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Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Thrombocytopenia'. | SARS-CoV-2 Infection (COVID-19) and Herpes Simplex Virus-1 Conjunctivitis: Concurrent Viral Infections or a Cause-Effect Result?
The pulmonary effects of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), the virus that causes coronavirus disease (COVID-19), are well documented; however, more evidence is needed to understand its effect on multiple organ systems. We present the case of a 69-year-old male with dyspnea for two weeks and bilateral conjunctivitis who tested positive for SARS-CoV-2. He was found to be hypoxic, requiring supplemental oxygen. On hospital day two, he complained of worsening left eye pain with the development of a left lower eyelid ulcer. He underwent a CT of facial bones, which showed findings consistent with pre-septal cellulitis and abscess. Samples from bilateral conjunctival secretions and left lower eyelid ulcer tested positive for herpes simplex virus-1 (HSV-1), and negative for SARS-CoV-2. He received supportive care, antibiotics, and famciclovir with almost complete resolution of his ocular complaints. This case illustrates an atypical COVID-19 presentation and raises concern as to how this virus modulates the immune system, allowing for concurrent viral infections.
Introduction
Coronavirus disease 2019 (COVID-19) is primarily a respiratory infection; however, it is now known to affect multiple organs, including the eye [1]. Due to the novelty of the virus, there are scarce data on COVID-19-related ocular infections in the medical literature. One study conducted in China noted that 31.6% of COVID-19-infected patients had ocular manifestations consistent with conjunctivitis [2]. However, the initial presentation of COVID-19 being an ocular manifestation seems rare. Other viruses such as herpes simplex are known to commonly present with ocular manifestations, with the latter having an annual incidence of 11.8 new cases for every 100,000 population [3]. In fact, the reactivation of latent herpes simplex virus (HSV) in the sensory ganglia may lead to initial or recurrent disease, typically monocular [3].
Case presentation
A 69-year-old Caucasian male with a medical history of type II diabetes mellitus, coronary artery disease, peripheral arterial disease, and stage III squamous cell lung cancer receiving weekly docetaxel presented to the ER in March 2020 with complaints of progressive dyspnea, cough, and scant white sputum for 14 days. He was a personal care aide who worked in a rehabilitation facility where multiple staff and patients had tested positive for COVID-19. Upon presentation, he was afebrile, normotensive, and hypoxemic, requiring 2 liters of oxygen via nasal cannula. He also reported bilateral eye pain, more significant in the left eye than the right, with associated bilateral conjunctival erythema, pruritus, chemosis, and foreign body sensation. He denied any blurry vision, changes in visual acuity, or previous ocular problems. Of note, the patient was enrolled in a double-blinded, randomized placebo-controlled trial with sarilumab on hospital day two (illness day 15) for the management of COVID-19. The patient electively self-withdrew from the trial three days post-infusion (drug versus placebo) administration due to personal reasons, unrelated to any safety concerns.
The patient tested positive for COVID-19 via a nasopharyngeal swab polymerase chain reaction (PCR). Laboratory workup revealed a normal white blood cell count, with normal neutrophil and lymphocyte counts. However, mild thrombocytopenia and vitamin D deficiency were noted. He presented with mild to moderate elevation in C-reactive protein (CRP), ferritin, D-dimer, and interleukin-6 level. His procalcitonin level, however, was mildly elevated (Table 1).
Table 1 Lab values at admission
Laboratory test Value at admission (reference range)
C-reactive protein 12.09 mg/dL (<0.5 mg/dL)
D-Dimer 2.0 mg/L (<0.5 mg/L)
Ferritin 824 ng/mL (21.81–274.66 ng/mL)
Hemoglobin A1c 8.3% (4.0–6.5%)
Interleukin 6 59 pg/mL (<5 pg/mL)
Lactic acid 2.5 mmol/L (0.5–2.2 mmol/L)
Lactic dehydrogenase 398 μ/l (125–220 μ/L)
Platelet count 103 103/μL (142.0–424.0 103/μL)
Procalcitonin 0.13 ng/ml (0.0–0.07 ng/mL)
Troponin 0.013 (0.000–0.023)
Vitamin D 23.2 ng/mL (>30 ng/mL)
White blood cell 4.38 103/μL (4.6–10.2 103/μL)
His chest X-ray at admission was unrevealing; however, a CT angiography (CTA) of his chest was negative for pulmonary embolism but revealed bilateral nodular and confluent sub-pleural and peri-bronchovascular ground-glass opacities. On the second day of admission (illness day 15), his left eye swelling and pain were noted to have worsened, with the development of a small ulcerative lesion anterior to the left lower eyelid (Figure 1).
Figure 1 Right and left eye (panel A and B, respectively) showing bilateral periorbital and conjunctival erythema, and a shallow ulcer in the left lower eyelid anteriorly (panel B)
A CT of facial bones showed a left-sided pre-septal and facial swelling consistent with cellulitis, and suggestive of a subdermal fluid collection measuring 1.5 x 0.4 x 0.9 cm, concerning for a superficial abscess (Figure 2). He was thus started on vancomycin and ceftriaxone.
Figure 2 CT facial bones with contrast showing left-sided pre-septal cellulitis and probable subdermal fluid collection (yellow arrow)
CT: computed tomography
His clinical presentation was most compatible with acute viral follicular conjunctivitis. For diagnostic purposes, conjunctival swabs were obtained from both eyes on hospital day seven (illness day 21), including a swab of the ulcer at the left lower eyelid. All the specimens were positive for HSV-1 PCR, and negative for severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), Adenovirus, and varicella-zoster virus (VZV) PCR.
His moderately severe SARS-CoV-2 infection was managed with a five-day course of hydroxychloroquine based on the limited evidence available at the time, in addition to supportive care. For the confirmed HSV-1 conjunctivitis, he was provided with a seven-day course of famciclovir and supportive care using artificial tears, cold compresses, and olopatadine hydrochloride ophthalmic solutions 0.1% every five to eight hours. The intravenous antimicrobial regimen targeting his pre-septal cellulitis was transitioned to oral cephalexin and doxycycline with instructions to complete 14 days of therapy.
During his inpatient stay, his oxygen requirements improved, along with his ocular symptoms. He was reached via telephone on post-discharge day six for continuity of care; he stated that he continued to notice some shortness of breath on exertion, though his ocular complaints had almost completely resolved.
Discussion
Coronaviruses had been previously reported to be associated with conjunctivitis in humans but deemed to be mild and rare [4,5]. However, ocular complications of SARS-CoV-2 have not been widely reported. A study performed in the province of Hubei in China by Wu et al. found that 31.6% of COVID-19 patients (n=39) had ocular manifestations consistent with conjunctivitis. A meta-analysis conducted by Loffredo et al., including three studies of 1,167 patients with COVID-19, found that only 3% of patients with severe disease and 0.7% of patients with non-severe disease had conjunctivitis. In SARS-CoV-2 conjunctivitis, patients can present with ocular redness, irritation, foreign body sensation, tearing, and chemosis [2,5,6]. Our patient presented with all of the above characteristics. In addition, our patient did not experience blurred vision, which is consistent with other studies [2,5].
Clinical examination findings of SARS-CoV-2 conjunctivitis are mostly consistent with mild follicular conjunctivitis as observed in several other studies [4,5,7]. Similarly, our patient's exam findings were consistent with acute viral follicular conjunctivitis, likely related to COVID-19 with superimposed HSV-1 reactivation. The potential false-negative COVID-19 PCR test of the conjunctival swabs can be explained by poor sample collection, scarcity of viral RNA in conjunctival fluid, or test error. Of note, Wu et al. reported that only 16.7% of their COVID-19-positive patients under study had positive results for SARS-CoV-2 on PCR from both conjunctivae. This implies that perhaps not all COVID-19-related conjunctivitis will have positive SARS-CoV-2 PCR upon testing of conjunctival swabs. Conjunctivitis can rarely present as an initial manifestation of COVID-19, and ocular symptoms more commonly affect patients with severe systemic diseases [2,5,8]. Moreover, patients with ocular symptoms have been reported to present with higher white blood cell and neutrophil counts, higher levels of procalcitonin, CRP, and LDH compared to patients without ocular symptoms [2]. Our patient did have neutrophilia, elevated CRP, and LDH, but a mild elevation of procalcitonin, and his COVID-19 was deemed moderate in severity. Even though there is a low prevalence of SARS-CoV-2 nucleotides in tears, just like SARS-CoV-1, it is possible to transmit via the eyes [2,5]. Furthermore, in order to demonstrate that the genome detected corresponds to the infectious virus, Colavita et al inoculated Vero-6 cells with ocular samples positive for SARS-CoV-2 RNA, and cytopathic effects were observed five days post-inoculum [1]. It has been suggested that viral loads in conjunctival specimens gradually decrease over time, with less potential for transmissibility accompanied by improvement of the ocular symptoms [4]. Several authors have reported outcomes ranging from improvement to resolution of symptoms by the third week of illness [1,4]. Significantly, ocular manifestations of COVID-19 conjunctivitis are thought to be self-limited, and there are currently no reports of sight-threatening manifestations.
In our case, the patient had several risk factors for immunosuppression, including age, multiple comorbidities, and the use of antineoplastic therapy. Furthermore, we believe that multiple factors led to his HSV-1 reactivation, including recent use of the cytotoxic agent for his underlying lung malignancy, and his underlying SARS-CoV-2 infection. In addition, our patient had bilateral ocular disease, whereas HSV-1 conjunctivitis is almost always unilateral [9]. One hypothesis is that the initial COVID-19 conjunctivitis led to repeated eye-rubbing and trauma resulting in HSV-1 reactivation complicated by superimposed bacterial infection and autoinoculation of the other eye. Corneal staining with fluorescein was not performed for the evaluation of herpetic keratitis.
HSV conjunctivitis has been deemed to be the second most common cause of viral conjunctivitis after Adenovirus infection, and also the most serious one [6,9]. In one study conducted in a Minnesota county, the annual incidence of herpes eye symptoms was found to be 11.8 new cases for every 100,000 population [3]. With HSV, vesicles may appear on the face or eyelids and vision may be affected [6]. Supportive care for both COVID-19 and HSV-1 conjunctivitis are indicated, with the use of preservative-free artificial tears, antihistamine eye drops, and cold compresses. A short course of topical antibiotics can be added to prevent or treat bacterial superinfection [5]. For HSV conjunctivitis, topical antivirals like acyclovir have been used, and most patients achieve resolution within 14-30 days [6]. For our immunosuppressed patient, a systemic option was preferred over a topical antiviral, in addition to supportive care, yielding excellent results. Although atypical manifestations of COVID-19 infections including ocular manifestations were not looked into during the initial phase of the pandemic, it has now become critical to consider the possibility of COVID-19 infection in multiple organ systems, as well as the risk of co-infections including reactivation of viruses, based on the evolving evidence. This would tremendously help us in taking good care of these patients as well as to timely put preventative measures in place to control its spread both in the healthcare settings and among the population at large.
Conclusions
Conjunctivitis is an atypical presentation of COVID-19 and can present with eye redness, ocular irritation, foreign body sensation, tearing, and chemosis. Detection of viral RNA in tears may not always be possible for diagnostic purposes. Nonetheless, COVID-19 conjunctivitis can in most instances be managed with a trial of frequent preservative-free artificial tears, cold compresses, and lubricating ophthalmic ointment. A short course of topical antibiotics can be added to prevent or treat bacterial superinfection. Lastly, HSV-1 should be on the differential diagnosis for any immunocompromised patient that presents with COVID-19 conjunctivitis.
The authors have declared that no competing interests exist.
Human Ethics
Consent was obtained by all participants in this study | DOCETAXEL | DrugsGivenReaction | CC BY | 33575153 | 18,984,996 | 2021-01-09 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Vitamin D deficiency'. | SARS-CoV-2 Infection (COVID-19) and Herpes Simplex Virus-1 Conjunctivitis: Concurrent Viral Infections or a Cause-Effect Result?
The pulmonary effects of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), the virus that causes coronavirus disease (COVID-19), are well documented; however, more evidence is needed to understand its effect on multiple organ systems. We present the case of a 69-year-old male with dyspnea for two weeks and bilateral conjunctivitis who tested positive for SARS-CoV-2. He was found to be hypoxic, requiring supplemental oxygen. On hospital day two, he complained of worsening left eye pain with the development of a left lower eyelid ulcer. He underwent a CT of facial bones, which showed findings consistent with pre-septal cellulitis and abscess. Samples from bilateral conjunctival secretions and left lower eyelid ulcer tested positive for herpes simplex virus-1 (HSV-1), and negative for SARS-CoV-2. He received supportive care, antibiotics, and famciclovir with almost complete resolution of his ocular complaints. This case illustrates an atypical COVID-19 presentation and raises concern as to how this virus modulates the immune system, allowing for concurrent viral infections.
Introduction
Coronavirus disease 2019 (COVID-19) is primarily a respiratory infection; however, it is now known to affect multiple organs, including the eye [1]. Due to the novelty of the virus, there are scarce data on COVID-19-related ocular infections in the medical literature. One study conducted in China noted that 31.6% of COVID-19-infected patients had ocular manifestations consistent with conjunctivitis [2]. However, the initial presentation of COVID-19 being an ocular manifestation seems rare. Other viruses such as herpes simplex are known to commonly present with ocular manifestations, with the latter having an annual incidence of 11.8 new cases for every 100,000 population [3]. In fact, the reactivation of latent herpes simplex virus (HSV) in the sensory ganglia may lead to initial or recurrent disease, typically monocular [3].
Case presentation
A 69-year-old Caucasian male with a medical history of type II diabetes mellitus, coronary artery disease, peripheral arterial disease, and stage III squamous cell lung cancer receiving weekly docetaxel presented to the ER in March 2020 with complaints of progressive dyspnea, cough, and scant white sputum for 14 days. He was a personal care aide who worked in a rehabilitation facility where multiple staff and patients had tested positive for COVID-19. Upon presentation, he was afebrile, normotensive, and hypoxemic, requiring 2 liters of oxygen via nasal cannula. He also reported bilateral eye pain, more significant in the left eye than the right, with associated bilateral conjunctival erythema, pruritus, chemosis, and foreign body sensation. He denied any blurry vision, changes in visual acuity, or previous ocular problems. Of note, the patient was enrolled in a double-blinded, randomized placebo-controlled trial with sarilumab on hospital day two (illness day 15) for the management of COVID-19. The patient electively self-withdrew from the trial three days post-infusion (drug versus placebo) administration due to personal reasons, unrelated to any safety concerns.
The patient tested positive for COVID-19 via a nasopharyngeal swab polymerase chain reaction (PCR). Laboratory workup revealed a normal white blood cell count, with normal neutrophil and lymphocyte counts. However, mild thrombocytopenia and vitamin D deficiency were noted. He presented with mild to moderate elevation in C-reactive protein (CRP), ferritin, D-dimer, and interleukin-6 level. His procalcitonin level, however, was mildly elevated (Table 1).
Table 1 Lab values at admission
Laboratory test Value at admission (reference range)
C-reactive protein 12.09 mg/dL (<0.5 mg/dL)
D-Dimer 2.0 mg/L (<0.5 mg/L)
Ferritin 824 ng/mL (21.81–274.66 ng/mL)
Hemoglobin A1c 8.3% (4.0–6.5%)
Interleukin 6 59 pg/mL (<5 pg/mL)
Lactic acid 2.5 mmol/L (0.5–2.2 mmol/L)
Lactic dehydrogenase 398 μ/l (125–220 μ/L)
Platelet count 103 103/μL (142.0–424.0 103/μL)
Procalcitonin 0.13 ng/ml (0.0–0.07 ng/mL)
Troponin 0.013 (0.000–0.023)
Vitamin D 23.2 ng/mL (>30 ng/mL)
White blood cell 4.38 103/μL (4.6–10.2 103/μL)
His chest X-ray at admission was unrevealing; however, a CT angiography (CTA) of his chest was negative for pulmonary embolism but revealed bilateral nodular and confluent sub-pleural and peri-bronchovascular ground-glass opacities. On the second day of admission (illness day 15), his left eye swelling and pain were noted to have worsened, with the development of a small ulcerative lesion anterior to the left lower eyelid (Figure 1).
Figure 1 Right and left eye (panel A and B, respectively) showing bilateral periorbital and conjunctival erythema, and a shallow ulcer in the left lower eyelid anteriorly (panel B)
A CT of facial bones showed a left-sided pre-septal and facial swelling consistent with cellulitis, and suggestive of a subdermal fluid collection measuring 1.5 x 0.4 x 0.9 cm, concerning for a superficial abscess (Figure 2). He was thus started on vancomycin and ceftriaxone.
Figure 2 CT facial bones with contrast showing left-sided pre-septal cellulitis and probable subdermal fluid collection (yellow arrow)
CT: computed tomography
His clinical presentation was most compatible with acute viral follicular conjunctivitis. For diagnostic purposes, conjunctival swabs were obtained from both eyes on hospital day seven (illness day 21), including a swab of the ulcer at the left lower eyelid. All the specimens were positive for HSV-1 PCR, and negative for severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), Adenovirus, and varicella-zoster virus (VZV) PCR.
His moderately severe SARS-CoV-2 infection was managed with a five-day course of hydroxychloroquine based on the limited evidence available at the time, in addition to supportive care. For the confirmed HSV-1 conjunctivitis, he was provided with a seven-day course of famciclovir and supportive care using artificial tears, cold compresses, and olopatadine hydrochloride ophthalmic solutions 0.1% every five to eight hours. The intravenous antimicrobial regimen targeting his pre-septal cellulitis was transitioned to oral cephalexin and doxycycline with instructions to complete 14 days of therapy.
During his inpatient stay, his oxygen requirements improved, along with his ocular symptoms. He was reached via telephone on post-discharge day six for continuity of care; he stated that he continued to notice some shortness of breath on exertion, though his ocular complaints had almost completely resolved.
Discussion
Coronaviruses had been previously reported to be associated with conjunctivitis in humans but deemed to be mild and rare [4,5]. However, ocular complications of SARS-CoV-2 have not been widely reported. A study performed in the province of Hubei in China by Wu et al. found that 31.6% of COVID-19 patients (n=39) had ocular manifestations consistent with conjunctivitis. A meta-analysis conducted by Loffredo et al., including three studies of 1,167 patients with COVID-19, found that only 3% of patients with severe disease and 0.7% of patients with non-severe disease had conjunctivitis. In SARS-CoV-2 conjunctivitis, patients can present with ocular redness, irritation, foreign body sensation, tearing, and chemosis [2,5,6]. Our patient presented with all of the above characteristics. In addition, our patient did not experience blurred vision, which is consistent with other studies [2,5].
Clinical examination findings of SARS-CoV-2 conjunctivitis are mostly consistent with mild follicular conjunctivitis as observed in several other studies [4,5,7]. Similarly, our patient's exam findings were consistent with acute viral follicular conjunctivitis, likely related to COVID-19 with superimposed HSV-1 reactivation. The potential false-negative COVID-19 PCR test of the conjunctival swabs can be explained by poor sample collection, scarcity of viral RNA in conjunctival fluid, or test error. Of note, Wu et al. reported that only 16.7% of their COVID-19-positive patients under study had positive results for SARS-CoV-2 on PCR from both conjunctivae. This implies that perhaps not all COVID-19-related conjunctivitis will have positive SARS-CoV-2 PCR upon testing of conjunctival swabs. Conjunctivitis can rarely present as an initial manifestation of COVID-19, and ocular symptoms more commonly affect patients with severe systemic diseases [2,5,8]. Moreover, patients with ocular symptoms have been reported to present with higher white blood cell and neutrophil counts, higher levels of procalcitonin, CRP, and LDH compared to patients without ocular symptoms [2]. Our patient did have neutrophilia, elevated CRP, and LDH, but a mild elevation of procalcitonin, and his COVID-19 was deemed moderate in severity. Even though there is a low prevalence of SARS-CoV-2 nucleotides in tears, just like SARS-CoV-1, it is possible to transmit via the eyes [2,5]. Furthermore, in order to demonstrate that the genome detected corresponds to the infectious virus, Colavita et al inoculated Vero-6 cells with ocular samples positive for SARS-CoV-2 RNA, and cytopathic effects were observed five days post-inoculum [1]. It has been suggested that viral loads in conjunctival specimens gradually decrease over time, with less potential for transmissibility accompanied by improvement of the ocular symptoms [4]. Several authors have reported outcomes ranging from improvement to resolution of symptoms by the third week of illness [1,4]. Significantly, ocular manifestations of COVID-19 conjunctivitis are thought to be self-limited, and there are currently no reports of sight-threatening manifestations.
In our case, the patient had several risk factors for immunosuppression, including age, multiple comorbidities, and the use of antineoplastic therapy. Furthermore, we believe that multiple factors led to his HSV-1 reactivation, including recent use of the cytotoxic agent for his underlying lung malignancy, and his underlying SARS-CoV-2 infection. In addition, our patient had bilateral ocular disease, whereas HSV-1 conjunctivitis is almost always unilateral [9]. One hypothesis is that the initial COVID-19 conjunctivitis led to repeated eye-rubbing and trauma resulting in HSV-1 reactivation complicated by superimposed bacterial infection and autoinoculation of the other eye. Corneal staining with fluorescein was not performed for the evaluation of herpetic keratitis.
HSV conjunctivitis has been deemed to be the second most common cause of viral conjunctivitis after Adenovirus infection, and also the most serious one [6,9]. In one study conducted in a Minnesota county, the annual incidence of herpes eye symptoms was found to be 11.8 new cases for every 100,000 population [3]. With HSV, vesicles may appear on the face or eyelids and vision may be affected [6]. Supportive care for both COVID-19 and HSV-1 conjunctivitis are indicated, with the use of preservative-free artificial tears, antihistamine eye drops, and cold compresses. A short course of topical antibiotics can be added to prevent or treat bacterial superinfection [5]. For HSV conjunctivitis, topical antivirals like acyclovir have been used, and most patients achieve resolution within 14-30 days [6]. For our immunosuppressed patient, a systemic option was preferred over a topical antiviral, in addition to supportive care, yielding excellent results. Although atypical manifestations of COVID-19 infections including ocular manifestations were not looked into during the initial phase of the pandemic, it has now become critical to consider the possibility of COVID-19 infection in multiple organ systems, as well as the risk of co-infections including reactivation of viruses, based on the evolving evidence. This would tremendously help us in taking good care of these patients as well as to timely put preventative measures in place to control its spread both in the healthcare settings and among the population at large.
Conclusions
Conjunctivitis is an atypical presentation of COVID-19 and can present with eye redness, ocular irritation, foreign body sensation, tearing, and chemosis. Detection of viral RNA in tears may not always be possible for diagnostic purposes. Nonetheless, COVID-19 conjunctivitis can in most instances be managed with a trial of frequent preservative-free artificial tears, cold compresses, and lubricating ophthalmic ointment. A short course of topical antibiotics can be added to prevent or treat bacterial superinfection. Lastly, HSV-1 should be on the differential diagnosis for any immunocompromised patient that presents with COVID-19 conjunctivitis.
The authors have declared that no competing interests exist.
Human Ethics
Consent was obtained by all participants in this study | DOCETAXEL | DrugsGivenReaction | CC BY | 33575153 | 18,984,996 | 2021-01-09 |
What was the outcome of reaction 'COVID-19'? | SARS-CoV-2 Infection (COVID-19) and Herpes Simplex Virus-1 Conjunctivitis: Concurrent Viral Infections or a Cause-Effect Result?
The pulmonary effects of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), the virus that causes coronavirus disease (COVID-19), are well documented; however, more evidence is needed to understand its effect on multiple organ systems. We present the case of a 69-year-old male with dyspnea for two weeks and bilateral conjunctivitis who tested positive for SARS-CoV-2. He was found to be hypoxic, requiring supplemental oxygen. On hospital day two, he complained of worsening left eye pain with the development of a left lower eyelid ulcer. He underwent a CT of facial bones, which showed findings consistent with pre-septal cellulitis and abscess. Samples from bilateral conjunctival secretions and left lower eyelid ulcer tested positive for herpes simplex virus-1 (HSV-1), and negative for SARS-CoV-2. He received supportive care, antibiotics, and famciclovir with almost complete resolution of his ocular complaints. This case illustrates an atypical COVID-19 presentation and raises concern as to how this virus modulates the immune system, allowing for concurrent viral infections.
Introduction
Coronavirus disease 2019 (COVID-19) is primarily a respiratory infection; however, it is now known to affect multiple organs, including the eye [1]. Due to the novelty of the virus, there are scarce data on COVID-19-related ocular infections in the medical literature. One study conducted in China noted that 31.6% of COVID-19-infected patients had ocular manifestations consistent with conjunctivitis [2]. However, the initial presentation of COVID-19 being an ocular manifestation seems rare. Other viruses such as herpes simplex are known to commonly present with ocular manifestations, with the latter having an annual incidence of 11.8 new cases for every 100,000 population [3]. In fact, the reactivation of latent herpes simplex virus (HSV) in the sensory ganglia may lead to initial or recurrent disease, typically monocular [3].
Case presentation
A 69-year-old Caucasian male with a medical history of type II diabetes mellitus, coronary artery disease, peripheral arterial disease, and stage III squamous cell lung cancer receiving weekly docetaxel presented to the ER in March 2020 with complaints of progressive dyspnea, cough, and scant white sputum for 14 days. He was a personal care aide who worked in a rehabilitation facility where multiple staff and patients had tested positive for COVID-19. Upon presentation, he was afebrile, normotensive, and hypoxemic, requiring 2 liters of oxygen via nasal cannula. He also reported bilateral eye pain, more significant in the left eye than the right, with associated bilateral conjunctival erythema, pruritus, chemosis, and foreign body sensation. He denied any blurry vision, changes in visual acuity, or previous ocular problems. Of note, the patient was enrolled in a double-blinded, randomized placebo-controlled trial with sarilumab on hospital day two (illness day 15) for the management of COVID-19. The patient electively self-withdrew from the trial three days post-infusion (drug versus placebo) administration due to personal reasons, unrelated to any safety concerns.
The patient tested positive for COVID-19 via a nasopharyngeal swab polymerase chain reaction (PCR). Laboratory workup revealed a normal white blood cell count, with normal neutrophil and lymphocyte counts. However, mild thrombocytopenia and vitamin D deficiency were noted. He presented with mild to moderate elevation in C-reactive protein (CRP), ferritin, D-dimer, and interleukin-6 level. His procalcitonin level, however, was mildly elevated (Table 1).
Table 1 Lab values at admission
Laboratory test Value at admission (reference range)
C-reactive protein 12.09 mg/dL (<0.5 mg/dL)
D-Dimer 2.0 mg/L (<0.5 mg/L)
Ferritin 824 ng/mL (21.81–274.66 ng/mL)
Hemoglobin A1c 8.3% (4.0–6.5%)
Interleukin 6 59 pg/mL (<5 pg/mL)
Lactic acid 2.5 mmol/L (0.5–2.2 mmol/L)
Lactic dehydrogenase 398 μ/l (125–220 μ/L)
Platelet count 103 103/μL (142.0–424.0 103/μL)
Procalcitonin 0.13 ng/ml (0.0–0.07 ng/mL)
Troponin 0.013 (0.000–0.023)
Vitamin D 23.2 ng/mL (>30 ng/mL)
White blood cell 4.38 103/μL (4.6–10.2 103/μL)
His chest X-ray at admission was unrevealing; however, a CT angiography (CTA) of his chest was negative for pulmonary embolism but revealed bilateral nodular and confluent sub-pleural and peri-bronchovascular ground-glass opacities. On the second day of admission (illness day 15), his left eye swelling and pain were noted to have worsened, with the development of a small ulcerative lesion anterior to the left lower eyelid (Figure 1).
Figure 1 Right and left eye (panel A and B, respectively) showing bilateral periorbital and conjunctival erythema, and a shallow ulcer in the left lower eyelid anteriorly (panel B)
A CT of facial bones showed a left-sided pre-septal and facial swelling consistent with cellulitis, and suggestive of a subdermal fluid collection measuring 1.5 x 0.4 x 0.9 cm, concerning for a superficial abscess (Figure 2). He was thus started on vancomycin and ceftriaxone.
Figure 2 CT facial bones with contrast showing left-sided pre-septal cellulitis and probable subdermal fluid collection (yellow arrow)
CT: computed tomography
His clinical presentation was most compatible with acute viral follicular conjunctivitis. For diagnostic purposes, conjunctival swabs were obtained from both eyes on hospital day seven (illness day 21), including a swab of the ulcer at the left lower eyelid. All the specimens were positive for HSV-1 PCR, and negative for severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), Adenovirus, and varicella-zoster virus (VZV) PCR.
His moderately severe SARS-CoV-2 infection was managed with a five-day course of hydroxychloroquine based on the limited evidence available at the time, in addition to supportive care. For the confirmed HSV-1 conjunctivitis, he was provided with a seven-day course of famciclovir and supportive care using artificial tears, cold compresses, and olopatadine hydrochloride ophthalmic solutions 0.1% every five to eight hours. The intravenous antimicrobial regimen targeting his pre-septal cellulitis was transitioned to oral cephalexin and doxycycline with instructions to complete 14 days of therapy.
During his inpatient stay, his oxygen requirements improved, along with his ocular symptoms. He was reached via telephone on post-discharge day six for continuity of care; he stated that he continued to notice some shortness of breath on exertion, though his ocular complaints had almost completely resolved.
Discussion
Coronaviruses had been previously reported to be associated with conjunctivitis in humans but deemed to be mild and rare [4,5]. However, ocular complications of SARS-CoV-2 have not been widely reported. A study performed in the province of Hubei in China by Wu et al. found that 31.6% of COVID-19 patients (n=39) had ocular manifestations consistent with conjunctivitis. A meta-analysis conducted by Loffredo et al., including three studies of 1,167 patients with COVID-19, found that only 3% of patients with severe disease and 0.7% of patients with non-severe disease had conjunctivitis. In SARS-CoV-2 conjunctivitis, patients can present with ocular redness, irritation, foreign body sensation, tearing, and chemosis [2,5,6]. Our patient presented with all of the above characteristics. In addition, our patient did not experience blurred vision, which is consistent with other studies [2,5].
Clinical examination findings of SARS-CoV-2 conjunctivitis are mostly consistent with mild follicular conjunctivitis as observed in several other studies [4,5,7]. Similarly, our patient's exam findings were consistent with acute viral follicular conjunctivitis, likely related to COVID-19 with superimposed HSV-1 reactivation. The potential false-negative COVID-19 PCR test of the conjunctival swabs can be explained by poor sample collection, scarcity of viral RNA in conjunctival fluid, or test error. Of note, Wu et al. reported that only 16.7% of their COVID-19-positive patients under study had positive results for SARS-CoV-2 on PCR from both conjunctivae. This implies that perhaps not all COVID-19-related conjunctivitis will have positive SARS-CoV-2 PCR upon testing of conjunctival swabs. Conjunctivitis can rarely present as an initial manifestation of COVID-19, and ocular symptoms more commonly affect patients with severe systemic diseases [2,5,8]. Moreover, patients with ocular symptoms have been reported to present with higher white blood cell and neutrophil counts, higher levels of procalcitonin, CRP, and LDH compared to patients without ocular symptoms [2]. Our patient did have neutrophilia, elevated CRP, and LDH, but a mild elevation of procalcitonin, and his COVID-19 was deemed moderate in severity. Even though there is a low prevalence of SARS-CoV-2 nucleotides in tears, just like SARS-CoV-1, it is possible to transmit via the eyes [2,5]. Furthermore, in order to demonstrate that the genome detected corresponds to the infectious virus, Colavita et al inoculated Vero-6 cells with ocular samples positive for SARS-CoV-2 RNA, and cytopathic effects were observed five days post-inoculum [1]. It has been suggested that viral loads in conjunctival specimens gradually decrease over time, with less potential for transmissibility accompanied by improvement of the ocular symptoms [4]. Several authors have reported outcomes ranging from improvement to resolution of symptoms by the third week of illness [1,4]. Significantly, ocular manifestations of COVID-19 conjunctivitis are thought to be self-limited, and there are currently no reports of sight-threatening manifestations.
In our case, the patient had several risk factors for immunosuppression, including age, multiple comorbidities, and the use of antineoplastic therapy. Furthermore, we believe that multiple factors led to his HSV-1 reactivation, including recent use of the cytotoxic agent for his underlying lung malignancy, and his underlying SARS-CoV-2 infection. In addition, our patient had bilateral ocular disease, whereas HSV-1 conjunctivitis is almost always unilateral [9]. One hypothesis is that the initial COVID-19 conjunctivitis led to repeated eye-rubbing and trauma resulting in HSV-1 reactivation complicated by superimposed bacterial infection and autoinoculation of the other eye. Corneal staining with fluorescein was not performed for the evaluation of herpetic keratitis.
HSV conjunctivitis has been deemed to be the second most common cause of viral conjunctivitis after Adenovirus infection, and also the most serious one [6,9]. In one study conducted in a Minnesota county, the annual incidence of herpes eye symptoms was found to be 11.8 new cases for every 100,000 population [3]. With HSV, vesicles may appear on the face or eyelids and vision may be affected [6]. Supportive care for both COVID-19 and HSV-1 conjunctivitis are indicated, with the use of preservative-free artificial tears, antihistamine eye drops, and cold compresses. A short course of topical antibiotics can be added to prevent or treat bacterial superinfection [5]. For HSV conjunctivitis, topical antivirals like acyclovir have been used, and most patients achieve resolution within 14-30 days [6]. For our immunosuppressed patient, a systemic option was preferred over a topical antiviral, in addition to supportive care, yielding excellent results. Although atypical manifestations of COVID-19 infections including ocular manifestations were not looked into during the initial phase of the pandemic, it has now become critical to consider the possibility of COVID-19 infection in multiple organ systems, as well as the risk of co-infections including reactivation of viruses, based on the evolving evidence. This would tremendously help us in taking good care of these patients as well as to timely put preventative measures in place to control its spread both in the healthcare settings and among the population at large.
Conclusions
Conjunctivitis is an atypical presentation of COVID-19 and can present with eye redness, ocular irritation, foreign body sensation, tearing, and chemosis. Detection of viral RNA in tears may not always be possible for diagnostic purposes. Nonetheless, COVID-19 conjunctivitis can in most instances be managed with a trial of frequent preservative-free artificial tears, cold compresses, and lubricating ophthalmic ointment. A short course of topical antibiotics can be added to prevent or treat bacterial superinfection. Lastly, HSV-1 should be on the differential diagnosis for any immunocompromised patient that presents with COVID-19 conjunctivitis.
The authors have declared that no competing interests exist.
Human Ethics
Consent was obtained by all participants in this study | Recovered | ReactionOutcome | CC BY | 33575153 | 18,984,996 | 2021-01-09 |
What was the outcome of reaction 'Ophthalmic herpes simplex'? | SARS-CoV-2 Infection (COVID-19) and Herpes Simplex Virus-1 Conjunctivitis: Concurrent Viral Infections or a Cause-Effect Result?
The pulmonary effects of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), the virus that causes coronavirus disease (COVID-19), are well documented; however, more evidence is needed to understand its effect on multiple organ systems. We present the case of a 69-year-old male with dyspnea for two weeks and bilateral conjunctivitis who tested positive for SARS-CoV-2. He was found to be hypoxic, requiring supplemental oxygen. On hospital day two, he complained of worsening left eye pain with the development of a left lower eyelid ulcer. He underwent a CT of facial bones, which showed findings consistent with pre-septal cellulitis and abscess. Samples from bilateral conjunctival secretions and left lower eyelid ulcer tested positive for herpes simplex virus-1 (HSV-1), and negative for SARS-CoV-2. He received supportive care, antibiotics, and famciclovir with almost complete resolution of his ocular complaints. This case illustrates an atypical COVID-19 presentation and raises concern as to how this virus modulates the immune system, allowing for concurrent viral infections.
Introduction
Coronavirus disease 2019 (COVID-19) is primarily a respiratory infection; however, it is now known to affect multiple organs, including the eye [1]. Due to the novelty of the virus, there are scarce data on COVID-19-related ocular infections in the medical literature. One study conducted in China noted that 31.6% of COVID-19-infected patients had ocular manifestations consistent with conjunctivitis [2]. However, the initial presentation of COVID-19 being an ocular manifestation seems rare. Other viruses such as herpes simplex are known to commonly present with ocular manifestations, with the latter having an annual incidence of 11.8 new cases for every 100,000 population [3]. In fact, the reactivation of latent herpes simplex virus (HSV) in the sensory ganglia may lead to initial or recurrent disease, typically monocular [3].
Case presentation
A 69-year-old Caucasian male with a medical history of type II diabetes mellitus, coronary artery disease, peripheral arterial disease, and stage III squamous cell lung cancer receiving weekly docetaxel presented to the ER in March 2020 with complaints of progressive dyspnea, cough, and scant white sputum for 14 days. He was a personal care aide who worked in a rehabilitation facility where multiple staff and patients had tested positive for COVID-19. Upon presentation, he was afebrile, normotensive, and hypoxemic, requiring 2 liters of oxygen via nasal cannula. He also reported bilateral eye pain, more significant in the left eye than the right, with associated bilateral conjunctival erythema, pruritus, chemosis, and foreign body sensation. He denied any blurry vision, changes in visual acuity, or previous ocular problems. Of note, the patient was enrolled in a double-blinded, randomized placebo-controlled trial with sarilumab on hospital day two (illness day 15) for the management of COVID-19. The patient electively self-withdrew from the trial three days post-infusion (drug versus placebo) administration due to personal reasons, unrelated to any safety concerns.
The patient tested positive for COVID-19 via a nasopharyngeal swab polymerase chain reaction (PCR). Laboratory workup revealed a normal white blood cell count, with normal neutrophil and lymphocyte counts. However, mild thrombocytopenia and vitamin D deficiency were noted. He presented with mild to moderate elevation in C-reactive protein (CRP), ferritin, D-dimer, and interleukin-6 level. His procalcitonin level, however, was mildly elevated (Table 1).
Table 1 Lab values at admission
Laboratory test Value at admission (reference range)
C-reactive protein 12.09 mg/dL (<0.5 mg/dL)
D-Dimer 2.0 mg/L (<0.5 mg/L)
Ferritin 824 ng/mL (21.81–274.66 ng/mL)
Hemoglobin A1c 8.3% (4.0–6.5%)
Interleukin 6 59 pg/mL (<5 pg/mL)
Lactic acid 2.5 mmol/L (0.5–2.2 mmol/L)
Lactic dehydrogenase 398 μ/l (125–220 μ/L)
Platelet count 103 103/μL (142.0–424.0 103/μL)
Procalcitonin 0.13 ng/ml (0.0–0.07 ng/mL)
Troponin 0.013 (0.000–0.023)
Vitamin D 23.2 ng/mL (>30 ng/mL)
White blood cell 4.38 103/μL (4.6–10.2 103/μL)
His chest X-ray at admission was unrevealing; however, a CT angiography (CTA) of his chest was negative for pulmonary embolism but revealed bilateral nodular and confluent sub-pleural and peri-bronchovascular ground-glass opacities. On the second day of admission (illness day 15), his left eye swelling and pain were noted to have worsened, with the development of a small ulcerative lesion anterior to the left lower eyelid (Figure 1).
Figure 1 Right and left eye (panel A and B, respectively) showing bilateral periorbital and conjunctival erythema, and a shallow ulcer in the left lower eyelid anteriorly (panel B)
A CT of facial bones showed a left-sided pre-septal and facial swelling consistent with cellulitis, and suggestive of a subdermal fluid collection measuring 1.5 x 0.4 x 0.9 cm, concerning for a superficial abscess (Figure 2). He was thus started on vancomycin and ceftriaxone.
Figure 2 CT facial bones with contrast showing left-sided pre-septal cellulitis and probable subdermal fluid collection (yellow arrow)
CT: computed tomography
His clinical presentation was most compatible with acute viral follicular conjunctivitis. For diagnostic purposes, conjunctival swabs were obtained from both eyes on hospital day seven (illness day 21), including a swab of the ulcer at the left lower eyelid. All the specimens were positive for HSV-1 PCR, and negative for severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), Adenovirus, and varicella-zoster virus (VZV) PCR.
His moderately severe SARS-CoV-2 infection was managed with a five-day course of hydroxychloroquine based on the limited evidence available at the time, in addition to supportive care. For the confirmed HSV-1 conjunctivitis, he was provided with a seven-day course of famciclovir and supportive care using artificial tears, cold compresses, and olopatadine hydrochloride ophthalmic solutions 0.1% every five to eight hours. The intravenous antimicrobial regimen targeting his pre-septal cellulitis was transitioned to oral cephalexin and doxycycline with instructions to complete 14 days of therapy.
During his inpatient stay, his oxygen requirements improved, along with his ocular symptoms. He was reached via telephone on post-discharge day six for continuity of care; he stated that he continued to notice some shortness of breath on exertion, though his ocular complaints had almost completely resolved.
Discussion
Coronaviruses had been previously reported to be associated with conjunctivitis in humans but deemed to be mild and rare [4,5]. However, ocular complications of SARS-CoV-2 have not been widely reported. A study performed in the province of Hubei in China by Wu et al. found that 31.6% of COVID-19 patients (n=39) had ocular manifestations consistent with conjunctivitis. A meta-analysis conducted by Loffredo et al., including three studies of 1,167 patients with COVID-19, found that only 3% of patients with severe disease and 0.7% of patients with non-severe disease had conjunctivitis. In SARS-CoV-2 conjunctivitis, patients can present with ocular redness, irritation, foreign body sensation, tearing, and chemosis [2,5,6]. Our patient presented with all of the above characteristics. In addition, our patient did not experience blurred vision, which is consistent with other studies [2,5].
Clinical examination findings of SARS-CoV-2 conjunctivitis are mostly consistent with mild follicular conjunctivitis as observed in several other studies [4,5,7]. Similarly, our patient's exam findings were consistent with acute viral follicular conjunctivitis, likely related to COVID-19 with superimposed HSV-1 reactivation. The potential false-negative COVID-19 PCR test of the conjunctival swabs can be explained by poor sample collection, scarcity of viral RNA in conjunctival fluid, or test error. Of note, Wu et al. reported that only 16.7% of their COVID-19-positive patients under study had positive results for SARS-CoV-2 on PCR from both conjunctivae. This implies that perhaps not all COVID-19-related conjunctivitis will have positive SARS-CoV-2 PCR upon testing of conjunctival swabs. Conjunctivitis can rarely present as an initial manifestation of COVID-19, and ocular symptoms more commonly affect patients with severe systemic diseases [2,5,8]. Moreover, patients with ocular symptoms have been reported to present with higher white blood cell and neutrophil counts, higher levels of procalcitonin, CRP, and LDH compared to patients without ocular symptoms [2]. Our patient did have neutrophilia, elevated CRP, and LDH, but a mild elevation of procalcitonin, and his COVID-19 was deemed moderate in severity. Even though there is a low prevalence of SARS-CoV-2 nucleotides in tears, just like SARS-CoV-1, it is possible to transmit via the eyes [2,5]. Furthermore, in order to demonstrate that the genome detected corresponds to the infectious virus, Colavita et al inoculated Vero-6 cells with ocular samples positive for SARS-CoV-2 RNA, and cytopathic effects were observed five days post-inoculum [1]. It has been suggested that viral loads in conjunctival specimens gradually decrease over time, with less potential for transmissibility accompanied by improvement of the ocular symptoms [4]. Several authors have reported outcomes ranging from improvement to resolution of symptoms by the third week of illness [1,4]. Significantly, ocular manifestations of COVID-19 conjunctivitis are thought to be self-limited, and there are currently no reports of sight-threatening manifestations.
In our case, the patient had several risk factors for immunosuppression, including age, multiple comorbidities, and the use of antineoplastic therapy. Furthermore, we believe that multiple factors led to his HSV-1 reactivation, including recent use of the cytotoxic agent for his underlying lung malignancy, and his underlying SARS-CoV-2 infection. In addition, our patient had bilateral ocular disease, whereas HSV-1 conjunctivitis is almost always unilateral [9]. One hypothesis is that the initial COVID-19 conjunctivitis led to repeated eye-rubbing and trauma resulting in HSV-1 reactivation complicated by superimposed bacterial infection and autoinoculation of the other eye. Corneal staining with fluorescein was not performed for the evaluation of herpetic keratitis.
HSV conjunctivitis has been deemed to be the second most common cause of viral conjunctivitis after Adenovirus infection, and also the most serious one [6,9]. In one study conducted in a Minnesota county, the annual incidence of herpes eye symptoms was found to be 11.8 new cases for every 100,000 population [3]. With HSV, vesicles may appear on the face or eyelids and vision may be affected [6]. Supportive care for both COVID-19 and HSV-1 conjunctivitis are indicated, with the use of preservative-free artificial tears, antihistamine eye drops, and cold compresses. A short course of topical antibiotics can be added to prevent or treat bacterial superinfection [5]. For HSV conjunctivitis, topical antivirals like acyclovir have been used, and most patients achieve resolution within 14-30 days [6]. For our immunosuppressed patient, a systemic option was preferred over a topical antiviral, in addition to supportive care, yielding excellent results. Although atypical manifestations of COVID-19 infections including ocular manifestations were not looked into during the initial phase of the pandemic, it has now become critical to consider the possibility of COVID-19 infection in multiple organ systems, as well as the risk of co-infections including reactivation of viruses, based on the evolving evidence. This would tremendously help us in taking good care of these patients as well as to timely put preventative measures in place to control its spread both in the healthcare settings and among the population at large.
Conclusions
Conjunctivitis is an atypical presentation of COVID-19 and can present with eye redness, ocular irritation, foreign body sensation, tearing, and chemosis. Detection of viral RNA in tears may not always be possible for diagnostic purposes. Nonetheless, COVID-19 conjunctivitis can in most instances be managed with a trial of frequent preservative-free artificial tears, cold compresses, and lubricating ophthalmic ointment. A short course of topical antibiotics can be added to prevent or treat bacterial superinfection. Lastly, HSV-1 should be on the differential diagnosis for any immunocompromised patient that presents with COVID-19 conjunctivitis.
The authors have declared that no competing interests exist.
Human Ethics
Consent was obtained by all participants in this study | Recovered | ReactionOutcome | CC BY | 33575153 | 18,984,996 | 2021-01-09 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'American trypanosomiasis'. | Reactivation of Chagas Disease in a Patient With an Autoimmune Rheumatic Disease: Case Report and Review of the Literature.
Reactivation of Chagas disease has been described in immunosuppressed patients, but there is a paucity of literature describing reactivation in patients on immunosuppressive therapies for the treatment of autoimmune rheumatic diseases. We describe a case of Chagas disease reactivation in a woman taking azathioprine and prednisone for limited cutaneous systemic sclerosis (lcSSc). Reactivation manifested as indurated and erythematous cutaneous nodules. Sequencing of a skin biopsy specimen confirmed the diagnosis of Chagas disease. She was treated with benznidazole with clinical improvement in the cutaneous lesions. However, her clinical course was complicated and included disseminated CMV disease and subsequent septic shock due to bacteremia. Our case and review of the literature highlight that screening for Chagas disease should be strongly considered for patients who will undergo immunosuppression for treatment of autoimmune disease if epidemiologically indicated.
PATIENT CASE
An 86-year-old woman developed painful subcutaneous nodules on her medial thighs and left upper extremity (Figure 1A, B). Over the ensuing 2 months, the nodules enlarged and became more indurated, erythematous, and painful. She also developed night sweats, fatigue, diarrhea, anorexia, and weight loss.
Figure 1. A, Skin lesions on left medial thigh. B, Skin lesions on right medial thigh.
The patient had a history of poorly controlled diabetes mellitus and limited cutaneous systemic sclerosis (lcSSc; a multisystem autoimmune rheumatic disease), manifested as pulmonary hypertension and Reynaud’s syndrome. Eight months before admission, she was diagnosed with tachy-brady syndrome, requiring a pacemaker. Echocardiogram at that time showed a dilated right ventricle with elevated pressures, preservation of left ventricle ejection fraction, and no wall motion abnormalities or apical aneurysms. For management of lcSSc, she had been on long-standing azathioprine (100 mg/d). Prednisone (15 mg/d) was added 3 months before the current presentation for management of a diagnosis of retinal vasculitis.
The patient was born and raised in a rural mountain village near Trujillo, Venezuela. As a child, she lived in a mud hut and cared for many animals including chickens, dogs, and cats. At 20 years of age, she emigrated to the United States. She traveled back to Venezuela only once at 70 years of age, and otherwise denied international travel.
She was admitted to the hospital, where she was initially afebrile and hemodynamically stable. Physical exam showed no mucosal abnormalities or regional lymphadenopathy. White blood cell count was 4.4 K/μL with profound lymphopenia (absolute lymphocytes 0.2 K/μL), hemoglobin 11.5 g/dL, and platelets 126 K/μL. Kidney and liver function were normal. An HIV antigen/antibody test was negative. Strongyloides serology was negative. Computed tomography scan showed scattered bilateral pulmonary nodules; normal caliber esophagus, small bowel, and large bowel; and inflammation in the ascending colon. Contrast-enhanced brain magnetic resonance imaging was normal.
Biopsy of the skin nodules (Figure 2A, B) showed a lymphohistiocytic infiltrate in the superficial and deep dermis with round intracellular organisms noted on hematoxylin/eosin stain. Periodic acid-Schiff-diastase, Gomori methenamine silver (GMS), Fite, and gram stains did not highlight the organisms. However, structures resembling kinetoplasts were minimally accentuated on GMS stain. Bacterial, fungal, and acid-fast bacillus cultures were ultimately negative. Serology for Trypanosoma cruzi (performed at Mayo Clinic Laboratories) was positive by both enzyme-linked immunosorbent assay and a lateral flow assay. A Giemsa-stained smear of peripheral blood (buffy coat) was negative, and real-time polymerase chain reaction (PCR) testing of a whole-blood sample for T. cruzi was also negative (performed at ARUP Laboratories). Tissue obtained from the cutaneous biopsy specimen identified T. cruzi by sequencing of the internal transcribed spacer 2 and D2 region of the 28S rRNA gene (performed at Stanford Health Care [1–3]).
Figure 2. A, Biopsy of skin nodules, 200× magnification, hematoxylin and eosin (H&E) stain. Histologic sections show an epidermis with mild spongiosis and an underlying lymphohistiocytic infiltrate. B, Biopsy of skin nodules, 400× magnification, H&E stain. Histologic sections show numerous parasitized histiocytes (demonstrated by arrows). The organisms are circular without a well-defined capsule.
Given the patient’s remote exposure to an area endemic for T. cruzi, she was diagnosed with reactivation of Chagas disease (CD). Immunosuppression was weaned, and oral benznidazole 150 mg twice daily (~5 mg/kg/d) was initiated.
Concurrent with the finding of CD, the patient was also found to have disseminated cytomegalovirus (CMV) disease. Plasma CMV viral load was >1.2 million copies/mL. Lung, colon, and skin biopsies all contained cells demonstrating cytopathic changes, which were positive by CMV immunostaining. Intravenous ganciclovir was started.
With treatment, the patient’s constitutional symptoms improved, and the skin lesions became less tender and erythematous. However, 1 month after the initiation of benznidazole, the patient developed Enterobacter cloacae bacteremia and septic shock. Shortly thereafter, she died.
Chagas Disease Epidemiology and Natural History
T. cruzi is primarily transmitted to humans via inoculation of wounds or mucosal membranes with infected feces of blood-sucking triatomine insects. CD is endemic throughout much of Latin America. However, as people have migrated from endemic areas, CD has become increasingly prevalent in traditionally nonendemic regions. In the United States, it is estimated that 1.3% of Latin American immigrants are infected with T. cruzi [4].
In acute CD, nearly all infected individuals have an effective host immune response that controls the parasitemia within 1–2 months of initial infection. However, in the absence of effective antiparasitic treatment, tissue infection persists for the life of the host. Chronic indeterminate CD occurs in persons without signs or symptoms of infection, with normal electrocardiogram, and normal radiographic appearance of the chest, esophagus, and colon. Indeterminate CD may persist for life or progress to determinate CD in 20%–30% of patients. Determinate CD results from tissue destruction related to persistent parasite replication and the immune response. It manifests with cardiac disease (conduction abnormalities, arrhythmias, dilated cardiomyopathy) and, less frequently, gastrointestinal disease (esophageal/colonic dysmotility and dilatation) [5, 6]. In some immunocompromised persons, chronic (indeterminate or determinate) CD can reactivate.
In conjunction with clinical symptoms, diagnosis of CD reactivation can be aided by laboratory testing. Serologic tests will be positive in most patients infected with T. cruzi. In chronic CD, patients can have transient parasitemia detected on microscopic examination of whole blood (or, preferably, buffy coat) smears [7]. With CD reactivation, patients often have more persistent parasitemia [8]. Positive T. cruzi blood PCR assays are suggestive but not diagnostic of reactivation. However, positive blood PCR results can herald the development of subsequent invasive Chagas disease reactivation in immunocompromised patients. Furthermore, rising parasite numbers demonstrated by quantitative PCR in serial specimens are highly suggestive of reactivated disease [8].
Reactivation of Chagas Disease
CD reactivation is best described in patients with advanced Chagas cardiomyopathy who undergo orthotopic heart transplantation [9–11]. The immunosuppressive drugs used to prevent transplant rejection predispose to CD reactivation. In these patients, reactivation most commonly manifests first as asymptomatic parasitemia or acute myocarditis [12]. Reactivation can also present as subcutaneous lesions [13], panniculitis [14], or, less commonly, meningoencephalitis [15]. Case series involving other solid organ transplant recipients (mostly renal transplant recipients) describe similar manifestations of CD reactivation [16–19]. CD reactivation is also known to occur in people with HIV/AIDS, most commonly presenting as meningoencephalitis [20] and/or brain abscesses (chagomas) [3, 21]. CD reactivation has also been described in patients receiving chemotherapy for hematologic and solid malignancies [22–25] and hematopoietic cell transplant (HCT) recipients [26–29].
Reactivation of Chagas Disease in Patients With Autoimmune Rheumatic Disease
There is a paucity of data regarding CD reactivation in patients receiving immunosuppressive therapy for autoimmune rheumatic diseases (ARDs; includes conditions such as systemic lupus erythematosus [SLE], rheumatoid arthritis, dermatomyositis, mixed connective tissue disease, and scleroderma). Compared with transplant recipients, patients with AIDS, and those receiving chemotherapy, patients undergoing treatment for ARD are often less immunosuppressed. However, it is unclear whether the dearth of literature in this cohort reflects infrequent reactivation or simply a lack of published case reports.
Among published English-language case reports that describe CD reactivation in patients with ARD on immunosuppressive regimens, the minority describe symptomatic CD reactivation (Table 1). Two patients developed brain chagomas—1 patient was successfully treated [30], and the other expired shortly after diagnosis [31]. Two other patients presented with Chagas skin lesions [30, 32], 1 of whom suffered recurrent disease after a course of benznidazole [30]. Other cases describe asymptomatic reactivation detected by T. cruzi PCR [33, 34], for which 1 patient was treated with off-label posaconazole after failing therapy with benznidazole [33]. The remaining patient reports describe possible reactivation detected through T. cruzi blood PCR positivity on a single test [30, 35]; given the absence of symptoms and serial testing, it is unclear if these patients had true CD reactivation or merely intermittently detectable parasites associated with chronic CD.
Table 1. Published Studies Describing Chagas Disease Reactivation (or Possible Reactivation) in Patients With Autoimmune Disease who Were Receiving Immunosuppressive Therapy
Study Patient Autoimmune Disease Immunosuppression Country of Origin Chagas Diagnosis Known Before Evaluation for Reactivation Evidence of Chagas Reactivation (or Possible Reactivation) Treatment Outcome
Current case 86 yo F Limited cutaneous systemic sclerosis Prednisone 15 mg/d, azathioprine 100 mg/d Venezuela No Skin lesions—erythematous, indurated, painful nodules on medial thighs and upper extremity Benznidazole Patient died 1 mo after initiation of benznidazole
Kaushal et al., 2019 [31] 88 yo F RA MTX Unknown No Brain chagoma Benznidazole Patient died shortly after diagnosis
German Sanchez et al., 2019 [30] 53 yo F SLE Prednisone 5 mg/d, MMF, CP All patients in study from Argentina No Brain chagoma and T. cruzi blood PCR pos Benznidazole PCR neg
68 yo F Psoriatic arthritis Prednisone 20 mg/d, HCQ, MTX Yes Panniculitis and T. cruzi blood PCR pos Benznidazole Relapse with recurrent skin lesions 1 y s/p treatment
7 of 13 patients with ARDs who had Chagas reactivation (or possible reactivation) 48 yo F RA MTX, adalimumab Yes Single T. Cruz blood PCR pos Benznidazole PCR neg
66 yo M RA Prednisone 5 mg/d, MTX, etanercept Yes Single T. Cruzi blood PCR pos Benznidazole PCR neg
Treatment durations for all patients were 1–2 mo 81 yo F RA Prednisone 5 mg/d, HCQ, MTX, leflunomide Yes Single T. Cruzi blood PCR pos Nifurtimox PCR neg
66 yo F Sjogren syndrome Prednisone 5 mg/d, HCQ Yes Single T. Cruzi blood PCR pos Benznidazole PCR neg
57 yo F Vasculitis Prednisone 5 mg/d, CP No Single T. cruzi blood PCR pos Benznidazole PCR neg
Vacas et al., 2017 [35] 57 yo M Psoriatic erythroderma Infliximab Argentina Yes Single T. cruzi blood PCR pos Benznidazole × 45 d PCR neg
Navarrete-Dechent et al., 2015 [34] 52 yo M Psoriasis Adalimumab Chile Yes—Chagas megacolon, received preemptive treatment with nifurtimox
T. cruzi blood PCR pos 8 months following preemptive nifurtimox Repeat course nifurtimox × 2 mo PCR neg
Burgos et al., 2012 [32] 44 yo F SLE Prednisone 50 mg/d, azathioprine 50 mg/d Paraguay No Parasitemia, skin lesions—erythematous, painful nodules that progressed to ulcer and eschar Benznidazole × 2 mo Clinically improved
Pinazo et al., 2010 [33] 44 yo F SLE Steroids, CP Argentina Yes—chronic indeterminate Chagas
T. cruzi blood PCR pos, treated with benznidazole with recurrent PCR pos Benznidazole × 2 mo, and then posaconazole × 3 mo for relapse Serial PCR neg after posaconazole
Abbreviations: CP, cyclophosphamide; HCQ, hydroxychloroquine; MMF, mycophenolate mofetil; MTX, methotrexate; NA, not applicable; neg, negative; PCR, polymerase chain reaction; pos, positive; RA, rheumatoid arthritis; SLE, systemic lupus erythematosus.
Although limited by small sample size and abbreviated longitudinal follow-up, other studies have attempted to investigate the risk for patients with ARD developing CD reactivation while on immunosuppression. One such study described 2 of 13 patients with ARD who developed symptomatic reactivation on immunosuppressive therapies over a 2-year period, and another 5 patients who developed possible CD reactivation detected by a single positive T. cruzi blood PCR test (Table 1) [30]. In other case series, the majority of immunosuppressed patients with ARD and CD were treated for chronic CD before the development of any evidence of CD reactivation, and only a small minority of these patients subsequently developed CD reactivation while receiving immunosuppression [25, 36, 37]. In 1 such study, 6 of 8 patients with ARD and CD were treated in this manner with varying regimens of benznidazole and nifurtimox. Three patients were treated for CD before a diagnosis of an ARD was made. Only 1 of the patients who received such treatment subsequently developed CD reactivation while receiving immunosuppression (further patient details not specified) [36]. In another study, 11 of 14 patients with chronic CD and ARD were treated for chronic CD with benznidazole for 60 days at the time of study enrollment, and none of the 14 patients experienced CD reactivation while receiving immunosuppression during follow-up [25]. Lastly, 3 patients with ARD and chronic CD were all treated at the time of CD diagnosis with benznidazole for 60 days; none of these patients developed subsequent CD reactivation during a 36-month follow-up period [37]. Though these preliminary data are intriguing, larger population studies, conducted over longer durations, are needed in order to draw more substantial conclusions about factors predisposing to CD reactivation in this cohort and the outcomes of prophylactic therapy.
DISCUSSION
Our patient is among the few in whom symptomatic CD reactivation was documented in the setting of immunosuppressive therapy for an ARD. Our patient likely had unrecognized determinate CD, clinically manifested by tachy-brady syndrome requiring a pacemaker [38]. CD reactivated after intensification of her immunosuppression regimen, ultimately manifesting as disseminated skin nodules, with sequencing of the skin biopsy revealing T. cruzi.
Our patient’s case was unique in that symptomatic CD reactivation occurred in the setting of a negative T. cruzi blood PCR. Typically, T. cruzi blood PCR positivity develops before symptomatic reactivation [11]. Though it is unclear why our patient’s PCR was negative, it is possibly related to blood volume collected or PCR test characteristics. Importantly, this demonstrates that PCR positivity should not be the sole diagnostic measure in the evaluation for CD reactivation; assessment must also include careful clinical evaluation.
CD reactivation in patients being treated for ARD additionally involves a different degree of immunosuppression compared with other better-studied cohorts. An effective host response against T. cruzi requires both cellular and humoral immunity [39]. Our patient was taking low-dose prednisone and azathioprine. Corticosteroids are nonspecific immune function inhibitors [40]. However, there are no reports that describe an increased incidence of CD reactivation in patients receiving corticosteroids as their sole form of immunosuppression. Azathioprine is an antimetabolite that decreases both T and B lymphocyte production [40]. However, some heart transplant recipients with a history of Chagas cardiomyopathy preferentially receive azathioprine over mycophenolate due to a 6-fold lower incidence of CD reactivation with azathioprine compared with mycophenolate [41]. Taken together, prednisone in doses <20 mg/d and azathioprine still seem to convey a relatively low risk for CD reactivation.
In our patient’s case, it is possible that more multifaceted immunosuppression led to her disease. Specifically, the synergistic effect of prednisone and azathioprine, in conjunction with her profound lymphopenia, advanced age, and uncontrolled diabetes, may have contributed to CD reactivation. However, these immunosuppressing factors still do not clearly explain our patient’s profound functional immunosuppression, further exhibited by concurrent disseminated CMV disease. Based on prior data [42–45], patients with rheumatoid arthritis and SLE may have an increased risk of infection independent of immunomodulatory therapy. Further research might better delineate the relationship between CD and immune function.
Screening for Chagas Disease in Patients With ARD
Currently, consensus guidelines recommend serologic screening for CD in transplant donors and recipients with epidemiologic risk factors. A positive serologic result should be confirmed by at least 2 distinct serologic methods. In the United States, CD treatment based solely on a positive serology result in either the donor or the recipient is generally not recommended given the toxicity of the therapeutic options [46, 47]. Alternatively, transplant recipients who are seropositive should be monitored for reactivation, especially during the times of most intense immunosuppression. Laboratory monitoring employs microscopy of blood/buffy coats and blood PCR. If monitoring reveals parasitemia and/or PCR positivity (especially increasing the parasite load on serial quantitative PCRs), patients are typically given preemptive CD treatment, as the development of detectable T. cruzi in this cohort often heralds the development of symptomatic reactivation [46–49].
Given that data regarding CD reactivation in patients with ARD are so scant, it is uncertain if similar screening and treatment guidelines should be applied to ARD patients in the face of immunosuppression. However, in light of our patient’s case and our review of the literature, our opinion is that strong consideration should be given to serologic screening for T. cruzi before immunosuppression for ARD in patients who have CD risk factors. (Note that serologic screening following immunosuppression may be falsely negative due to a blunted immune response.) Data are insufficient to comment on the risks versus benefits of CD treatment based solely on a positive serologic result in this cohort. However, similar to transplant recipients, for those who are seropositive, clinical assessment and serial blood microscopy/PCR monitoring should be employed during immunosuppressive therapy. There should be strong consideration of preemptive treatment of patients with parasitemia and/or PCR blood positivity (especially increasing parasite load on serial quantitative PCRs) even in the absence of symptoms; of course, evaluation/treatment for those with symptoms/signs concerning for CD reactivation. More research is needed to refine the screening and subsequent treatment approach for CD in patients with ARD.
CONCLUSIONS
This report describes CD reactivation in a patient with an ARD receiving immunosuppressive therapy. It highlights the need for more research regarding CD and reactivation in this patient population. It additionally suggests the need for broader serologic screening of patients with risk factors for CD before starting immunosuppression, and subsequent monitoring of at-risk patients while they receive such therapy.
Acknowledgments
Financial support. None
Potential conflicts of interest. All authors: no reported conflicts of interest. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed.
Disclaimer. The findings and conclusions in this report are those of the authors and do not necessarily represent the views of the Centers for Disease Control and Prevention.
Patient consent. Our study does not include factors necessitating patient consent. | AZATHIOPRINE, PREDNISONE | DrugsGivenReaction | CC BY-NC-ND | 33575423 | 19,507,537 | 2021-02 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Disease recurrence'. | Reactivation of Chagas Disease in a Patient With an Autoimmune Rheumatic Disease: Case Report and Review of the Literature.
Reactivation of Chagas disease has been described in immunosuppressed patients, but there is a paucity of literature describing reactivation in patients on immunosuppressive therapies for the treatment of autoimmune rheumatic diseases. We describe a case of Chagas disease reactivation in a woman taking azathioprine and prednisone for limited cutaneous systemic sclerosis (lcSSc). Reactivation manifested as indurated and erythematous cutaneous nodules. Sequencing of a skin biopsy specimen confirmed the diagnosis of Chagas disease. She was treated with benznidazole with clinical improvement in the cutaneous lesions. However, her clinical course was complicated and included disseminated CMV disease and subsequent septic shock due to bacteremia. Our case and review of the literature highlight that screening for Chagas disease should be strongly considered for patients who will undergo immunosuppression for treatment of autoimmune disease if epidemiologically indicated.
PATIENT CASE
An 86-year-old woman developed painful subcutaneous nodules on her medial thighs and left upper extremity (Figure 1A, B). Over the ensuing 2 months, the nodules enlarged and became more indurated, erythematous, and painful. She also developed night sweats, fatigue, diarrhea, anorexia, and weight loss.
Figure 1. A, Skin lesions on left medial thigh. B, Skin lesions on right medial thigh.
The patient had a history of poorly controlled diabetes mellitus and limited cutaneous systemic sclerosis (lcSSc; a multisystem autoimmune rheumatic disease), manifested as pulmonary hypertension and Reynaud’s syndrome. Eight months before admission, she was diagnosed with tachy-brady syndrome, requiring a pacemaker. Echocardiogram at that time showed a dilated right ventricle with elevated pressures, preservation of left ventricle ejection fraction, and no wall motion abnormalities or apical aneurysms. For management of lcSSc, she had been on long-standing azathioprine (100 mg/d). Prednisone (15 mg/d) was added 3 months before the current presentation for management of a diagnosis of retinal vasculitis.
The patient was born and raised in a rural mountain village near Trujillo, Venezuela. As a child, she lived in a mud hut and cared for many animals including chickens, dogs, and cats. At 20 years of age, she emigrated to the United States. She traveled back to Venezuela only once at 70 years of age, and otherwise denied international travel.
She was admitted to the hospital, where she was initially afebrile and hemodynamically stable. Physical exam showed no mucosal abnormalities or regional lymphadenopathy. White blood cell count was 4.4 K/μL with profound lymphopenia (absolute lymphocytes 0.2 K/μL), hemoglobin 11.5 g/dL, and platelets 126 K/μL. Kidney and liver function were normal. An HIV antigen/antibody test was negative. Strongyloides serology was negative. Computed tomography scan showed scattered bilateral pulmonary nodules; normal caliber esophagus, small bowel, and large bowel; and inflammation in the ascending colon. Contrast-enhanced brain magnetic resonance imaging was normal.
Biopsy of the skin nodules (Figure 2A, B) showed a lymphohistiocytic infiltrate in the superficial and deep dermis with round intracellular organisms noted on hematoxylin/eosin stain. Periodic acid-Schiff-diastase, Gomori methenamine silver (GMS), Fite, and gram stains did not highlight the organisms. However, structures resembling kinetoplasts were minimally accentuated on GMS stain. Bacterial, fungal, and acid-fast bacillus cultures were ultimately negative. Serology for Trypanosoma cruzi (performed at Mayo Clinic Laboratories) was positive by both enzyme-linked immunosorbent assay and a lateral flow assay. A Giemsa-stained smear of peripheral blood (buffy coat) was negative, and real-time polymerase chain reaction (PCR) testing of a whole-blood sample for T. cruzi was also negative (performed at ARUP Laboratories). Tissue obtained from the cutaneous biopsy specimen identified T. cruzi by sequencing of the internal transcribed spacer 2 and D2 region of the 28S rRNA gene (performed at Stanford Health Care [1–3]).
Figure 2. A, Biopsy of skin nodules, 200× magnification, hematoxylin and eosin (H&E) stain. Histologic sections show an epidermis with mild spongiosis and an underlying lymphohistiocytic infiltrate. B, Biopsy of skin nodules, 400× magnification, H&E stain. Histologic sections show numerous parasitized histiocytes (demonstrated by arrows). The organisms are circular without a well-defined capsule.
Given the patient’s remote exposure to an area endemic for T. cruzi, she was diagnosed with reactivation of Chagas disease (CD). Immunosuppression was weaned, and oral benznidazole 150 mg twice daily (~5 mg/kg/d) was initiated.
Concurrent with the finding of CD, the patient was also found to have disseminated cytomegalovirus (CMV) disease. Plasma CMV viral load was >1.2 million copies/mL. Lung, colon, and skin biopsies all contained cells demonstrating cytopathic changes, which were positive by CMV immunostaining. Intravenous ganciclovir was started.
With treatment, the patient’s constitutional symptoms improved, and the skin lesions became less tender and erythematous. However, 1 month after the initiation of benznidazole, the patient developed Enterobacter cloacae bacteremia and septic shock. Shortly thereafter, she died.
Chagas Disease Epidemiology and Natural History
T. cruzi is primarily transmitted to humans via inoculation of wounds or mucosal membranes with infected feces of blood-sucking triatomine insects. CD is endemic throughout much of Latin America. However, as people have migrated from endemic areas, CD has become increasingly prevalent in traditionally nonendemic regions. In the United States, it is estimated that 1.3% of Latin American immigrants are infected with T. cruzi [4].
In acute CD, nearly all infected individuals have an effective host immune response that controls the parasitemia within 1–2 months of initial infection. However, in the absence of effective antiparasitic treatment, tissue infection persists for the life of the host. Chronic indeterminate CD occurs in persons without signs or symptoms of infection, with normal electrocardiogram, and normal radiographic appearance of the chest, esophagus, and colon. Indeterminate CD may persist for life or progress to determinate CD in 20%–30% of patients. Determinate CD results from tissue destruction related to persistent parasite replication and the immune response. It manifests with cardiac disease (conduction abnormalities, arrhythmias, dilated cardiomyopathy) and, less frequently, gastrointestinal disease (esophageal/colonic dysmotility and dilatation) [5, 6]. In some immunocompromised persons, chronic (indeterminate or determinate) CD can reactivate.
In conjunction with clinical symptoms, diagnosis of CD reactivation can be aided by laboratory testing. Serologic tests will be positive in most patients infected with T. cruzi. In chronic CD, patients can have transient parasitemia detected on microscopic examination of whole blood (or, preferably, buffy coat) smears [7]. With CD reactivation, patients often have more persistent parasitemia [8]. Positive T. cruzi blood PCR assays are suggestive but not diagnostic of reactivation. However, positive blood PCR results can herald the development of subsequent invasive Chagas disease reactivation in immunocompromised patients. Furthermore, rising parasite numbers demonstrated by quantitative PCR in serial specimens are highly suggestive of reactivated disease [8].
Reactivation of Chagas Disease
CD reactivation is best described in patients with advanced Chagas cardiomyopathy who undergo orthotopic heart transplantation [9–11]. The immunosuppressive drugs used to prevent transplant rejection predispose to CD reactivation. In these patients, reactivation most commonly manifests first as asymptomatic parasitemia or acute myocarditis [12]. Reactivation can also present as subcutaneous lesions [13], panniculitis [14], or, less commonly, meningoencephalitis [15]. Case series involving other solid organ transplant recipients (mostly renal transplant recipients) describe similar manifestations of CD reactivation [16–19]. CD reactivation is also known to occur in people with HIV/AIDS, most commonly presenting as meningoencephalitis [20] and/or brain abscesses (chagomas) [3, 21]. CD reactivation has also been described in patients receiving chemotherapy for hematologic and solid malignancies [22–25] and hematopoietic cell transplant (HCT) recipients [26–29].
Reactivation of Chagas Disease in Patients With Autoimmune Rheumatic Disease
There is a paucity of data regarding CD reactivation in patients receiving immunosuppressive therapy for autoimmune rheumatic diseases (ARDs; includes conditions such as systemic lupus erythematosus [SLE], rheumatoid arthritis, dermatomyositis, mixed connective tissue disease, and scleroderma). Compared with transplant recipients, patients with AIDS, and those receiving chemotherapy, patients undergoing treatment for ARD are often less immunosuppressed. However, it is unclear whether the dearth of literature in this cohort reflects infrequent reactivation or simply a lack of published case reports.
Among published English-language case reports that describe CD reactivation in patients with ARD on immunosuppressive regimens, the minority describe symptomatic CD reactivation (Table 1). Two patients developed brain chagomas—1 patient was successfully treated [30], and the other expired shortly after diagnosis [31]. Two other patients presented with Chagas skin lesions [30, 32], 1 of whom suffered recurrent disease after a course of benznidazole [30]. Other cases describe asymptomatic reactivation detected by T. cruzi PCR [33, 34], for which 1 patient was treated with off-label posaconazole after failing therapy with benznidazole [33]. The remaining patient reports describe possible reactivation detected through T. cruzi blood PCR positivity on a single test [30, 35]; given the absence of symptoms and serial testing, it is unclear if these patients had true CD reactivation or merely intermittently detectable parasites associated with chronic CD.
Table 1. Published Studies Describing Chagas Disease Reactivation (or Possible Reactivation) in Patients With Autoimmune Disease who Were Receiving Immunosuppressive Therapy
Study Patient Autoimmune Disease Immunosuppression Country of Origin Chagas Diagnosis Known Before Evaluation for Reactivation Evidence of Chagas Reactivation (or Possible Reactivation) Treatment Outcome
Current case 86 yo F Limited cutaneous systemic sclerosis Prednisone 15 mg/d, azathioprine 100 mg/d Venezuela No Skin lesions—erythematous, indurated, painful nodules on medial thighs and upper extremity Benznidazole Patient died 1 mo after initiation of benznidazole
Kaushal et al., 2019 [31] 88 yo F RA MTX Unknown No Brain chagoma Benznidazole Patient died shortly after diagnosis
German Sanchez et al., 2019 [30] 53 yo F SLE Prednisone 5 mg/d, MMF, CP All patients in study from Argentina No Brain chagoma and T. cruzi blood PCR pos Benznidazole PCR neg
68 yo F Psoriatic arthritis Prednisone 20 mg/d, HCQ, MTX Yes Panniculitis and T. cruzi blood PCR pos Benznidazole Relapse with recurrent skin lesions 1 y s/p treatment
7 of 13 patients with ARDs who had Chagas reactivation (or possible reactivation) 48 yo F RA MTX, adalimumab Yes Single T. Cruz blood PCR pos Benznidazole PCR neg
66 yo M RA Prednisone 5 mg/d, MTX, etanercept Yes Single T. Cruzi blood PCR pos Benznidazole PCR neg
Treatment durations for all patients were 1–2 mo 81 yo F RA Prednisone 5 mg/d, HCQ, MTX, leflunomide Yes Single T. Cruzi blood PCR pos Nifurtimox PCR neg
66 yo F Sjogren syndrome Prednisone 5 mg/d, HCQ Yes Single T. Cruzi blood PCR pos Benznidazole PCR neg
57 yo F Vasculitis Prednisone 5 mg/d, CP No Single T. cruzi blood PCR pos Benznidazole PCR neg
Vacas et al., 2017 [35] 57 yo M Psoriatic erythroderma Infliximab Argentina Yes Single T. cruzi blood PCR pos Benznidazole × 45 d PCR neg
Navarrete-Dechent et al., 2015 [34] 52 yo M Psoriasis Adalimumab Chile Yes—Chagas megacolon, received preemptive treatment with nifurtimox
T. cruzi blood PCR pos 8 months following preemptive nifurtimox Repeat course nifurtimox × 2 mo PCR neg
Burgos et al., 2012 [32] 44 yo F SLE Prednisone 50 mg/d, azathioprine 50 mg/d Paraguay No Parasitemia, skin lesions—erythematous, painful nodules that progressed to ulcer and eschar Benznidazole × 2 mo Clinically improved
Pinazo et al., 2010 [33] 44 yo F SLE Steroids, CP Argentina Yes—chronic indeterminate Chagas
T. cruzi blood PCR pos, treated with benznidazole with recurrent PCR pos Benznidazole × 2 mo, and then posaconazole × 3 mo for relapse Serial PCR neg after posaconazole
Abbreviations: CP, cyclophosphamide; HCQ, hydroxychloroquine; MMF, mycophenolate mofetil; MTX, methotrexate; NA, not applicable; neg, negative; PCR, polymerase chain reaction; pos, positive; RA, rheumatoid arthritis; SLE, systemic lupus erythematosus.
Although limited by small sample size and abbreviated longitudinal follow-up, other studies have attempted to investigate the risk for patients with ARD developing CD reactivation while on immunosuppression. One such study described 2 of 13 patients with ARD who developed symptomatic reactivation on immunosuppressive therapies over a 2-year period, and another 5 patients who developed possible CD reactivation detected by a single positive T. cruzi blood PCR test (Table 1) [30]. In other case series, the majority of immunosuppressed patients with ARD and CD were treated for chronic CD before the development of any evidence of CD reactivation, and only a small minority of these patients subsequently developed CD reactivation while receiving immunosuppression [25, 36, 37]. In 1 such study, 6 of 8 patients with ARD and CD were treated in this manner with varying regimens of benznidazole and nifurtimox. Three patients were treated for CD before a diagnosis of an ARD was made. Only 1 of the patients who received such treatment subsequently developed CD reactivation while receiving immunosuppression (further patient details not specified) [36]. In another study, 11 of 14 patients with chronic CD and ARD were treated for chronic CD with benznidazole for 60 days at the time of study enrollment, and none of the 14 patients experienced CD reactivation while receiving immunosuppression during follow-up [25]. Lastly, 3 patients with ARD and chronic CD were all treated at the time of CD diagnosis with benznidazole for 60 days; none of these patients developed subsequent CD reactivation during a 36-month follow-up period [37]. Though these preliminary data are intriguing, larger population studies, conducted over longer durations, are needed in order to draw more substantial conclusions about factors predisposing to CD reactivation in this cohort and the outcomes of prophylactic therapy.
DISCUSSION
Our patient is among the few in whom symptomatic CD reactivation was documented in the setting of immunosuppressive therapy for an ARD. Our patient likely had unrecognized determinate CD, clinically manifested by tachy-brady syndrome requiring a pacemaker [38]. CD reactivated after intensification of her immunosuppression regimen, ultimately manifesting as disseminated skin nodules, with sequencing of the skin biopsy revealing T. cruzi.
Our patient’s case was unique in that symptomatic CD reactivation occurred in the setting of a negative T. cruzi blood PCR. Typically, T. cruzi blood PCR positivity develops before symptomatic reactivation [11]. Though it is unclear why our patient’s PCR was negative, it is possibly related to blood volume collected or PCR test characteristics. Importantly, this demonstrates that PCR positivity should not be the sole diagnostic measure in the evaluation for CD reactivation; assessment must also include careful clinical evaluation.
CD reactivation in patients being treated for ARD additionally involves a different degree of immunosuppression compared with other better-studied cohorts. An effective host response against T. cruzi requires both cellular and humoral immunity [39]. Our patient was taking low-dose prednisone and azathioprine. Corticosteroids are nonspecific immune function inhibitors [40]. However, there are no reports that describe an increased incidence of CD reactivation in patients receiving corticosteroids as their sole form of immunosuppression. Azathioprine is an antimetabolite that decreases both T and B lymphocyte production [40]. However, some heart transplant recipients with a history of Chagas cardiomyopathy preferentially receive azathioprine over mycophenolate due to a 6-fold lower incidence of CD reactivation with azathioprine compared with mycophenolate [41]. Taken together, prednisone in doses <20 mg/d and azathioprine still seem to convey a relatively low risk for CD reactivation.
In our patient’s case, it is possible that more multifaceted immunosuppression led to her disease. Specifically, the synergistic effect of prednisone and azathioprine, in conjunction with her profound lymphopenia, advanced age, and uncontrolled diabetes, may have contributed to CD reactivation. However, these immunosuppressing factors still do not clearly explain our patient’s profound functional immunosuppression, further exhibited by concurrent disseminated CMV disease. Based on prior data [42–45], patients with rheumatoid arthritis and SLE may have an increased risk of infection independent of immunomodulatory therapy. Further research might better delineate the relationship between CD and immune function.
Screening for Chagas Disease in Patients With ARD
Currently, consensus guidelines recommend serologic screening for CD in transplant donors and recipients with epidemiologic risk factors. A positive serologic result should be confirmed by at least 2 distinct serologic methods. In the United States, CD treatment based solely on a positive serology result in either the donor or the recipient is generally not recommended given the toxicity of the therapeutic options [46, 47]. Alternatively, transplant recipients who are seropositive should be monitored for reactivation, especially during the times of most intense immunosuppression. Laboratory monitoring employs microscopy of blood/buffy coats and blood PCR. If monitoring reveals parasitemia and/or PCR positivity (especially increasing the parasite load on serial quantitative PCRs), patients are typically given preemptive CD treatment, as the development of detectable T. cruzi in this cohort often heralds the development of symptomatic reactivation [46–49].
Given that data regarding CD reactivation in patients with ARD are so scant, it is uncertain if similar screening and treatment guidelines should be applied to ARD patients in the face of immunosuppression. However, in light of our patient’s case and our review of the literature, our opinion is that strong consideration should be given to serologic screening for T. cruzi before immunosuppression for ARD in patients who have CD risk factors. (Note that serologic screening following immunosuppression may be falsely negative due to a blunted immune response.) Data are insufficient to comment on the risks versus benefits of CD treatment based solely on a positive serologic result in this cohort. However, similar to transplant recipients, for those who are seropositive, clinical assessment and serial blood microscopy/PCR monitoring should be employed during immunosuppressive therapy. There should be strong consideration of preemptive treatment of patients with parasitemia and/or PCR blood positivity (especially increasing parasite load on serial quantitative PCRs) even in the absence of symptoms; of course, evaluation/treatment for those with symptoms/signs concerning for CD reactivation. More research is needed to refine the screening and subsequent treatment approach for CD in patients with ARD.
CONCLUSIONS
This report describes CD reactivation in a patient with an ARD receiving immunosuppressive therapy. It highlights the need for more research regarding CD and reactivation in this patient population. It additionally suggests the need for broader serologic screening of patients with risk factors for CD before starting immunosuppression, and subsequent monitoring of at-risk patients while they receive such therapy.
Acknowledgments
Financial support. None
Potential conflicts of interest. All authors: no reported conflicts of interest. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed.
Disclaimer. The findings and conclusions in this report are those of the authors and do not necessarily represent the views of the Centers for Disease Control and Prevention.
Patient consent. Our study does not include factors necessitating patient consent. | AZATHIOPRINE, PREDNISONE | DrugsGivenReaction | CC BY-NC-ND | 33575423 | 19,494,598 | 2021-02 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Disseminated cytomegaloviral infection'. | Reactivation of Chagas Disease in a Patient With an Autoimmune Rheumatic Disease: Case Report and Review of the Literature.
Reactivation of Chagas disease has been described in immunosuppressed patients, but there is a paucity of literature describing reactivation in patients on immunosuppressive therapies for the treatment of autoimmune rheumatic diseases. We describe a case of Chagas disease reactivation in a woman taking azathioprine and prednisone for limited cutaneous systemic sclerosis (lcSSc). Reactivation manifested as indurated and erythematous cutaneous nodules. Sequencing of a skin biopsy specimen confirmed the diagnosis of Chagas disease. She was treated with benznidazole with clinical improvement in the cutaneous lesions. However, her clinical course was complicated and included disseminated CMV disease and subsequent septic shock due to bacteremia. Our case and review of the literature highlight that screening for Chagas disease should be strongly considered for patients who will undergo immunosuppression for treatment of autoimmune disease if epidemiologically indicated.
PATIENT CASE
An 86-year-old woman developed painful subcutaneous nodules on her medial thighs and left upper extremity (Figure 1A, B). Over the ensuing 2 months, the nodules enlarged and became more indurated, erythematous, and painful. She also developed night sweats, fatigue, diarrhea, anorexia, and weight loss.
Figure 1. A, Skin lesions on left medial thigh. B, Skin lesions on right medial thigh.
The patient had a history of poorly controlled diabetes mellitus and limited cutaneous systemic sclerosis (lcSSc; a multisystem autoimmune rheumatic disease), manifested as pulmonary hypertension and Reynaud’s syndrome. Eight months before admission, she was diagnosed with tachy-brady syndrome, requiring a pacemaker. Echocardiogram at that time showed a dilated right ventricle with elevated pressures, preservation of left ventricle ejection fraction, and no wall motion abnormalities or apical aneurysms. For management of lcSSc, she had been on long-standing azathioprine (100 mg/d). Prednisone (15 mg/d) was added 3 months before the current presentation for management of a diagnosis of retinal vasculitis.
The patient was born and raised in a rural mountain village near Trujillo, Venezuela. As a child, she lived in a mud hut and cared for many animals including chickens, dogs, and cats. At 20 years of age, she emigrated to the United States. She traveled back to Venezuela only once at 70 years of age, and otherwise denied international travel.
She was admitted to the hospital, where she was initially afebrile and hemodynamically stable. Physical exam showed no mucosal abnormalities or regional lymphadenopathy. White blood cell count was 4.4 K/μL with profound lymphopenia (absolute lymphocytes 0.2 K/μL), hemoglobin 11.5 g/dL, and platelets 126 K/μL. Kidney and liver function were normal. An HIV antigen/antibody test was negative. Strongyloides serology was negative. Computed tomography scan showed scattered bilateral pulmonary nodules; normal caliber esophagus, small bowel, and large bowel; and inflammation in the ascending colon. Contrast-enhanced brain magnetic resonance imaging was normal.
Biopsy of the skin nodules (Figure 2A, B) showed a lymphohistiocytic infiltrate in the superficial and deep dermis with round intracellular organisms noted on hematoxylin/eosin stain. Periodic acid-Schiff-diastase, Gomori methenamine silver (GMS), Fite, and gram stains did not highlight the organisms. However, structures resembling kinetoplasts were minimally accentuated on GMS stain. Bacterial, fungal, and acid-fast bacillus cultures were ultimately negative. Serology for Trypanosoma cruzi (performed at Mayo Clinic Laboratories) was positive by both enzyme-linked immunosorbent assay and a lateral flow assay. A Giemsa-stained smear of peripheral blood (buffy coat) was negative, and real-time polymerase chain reaction (PCR) testing of a whole-blood sample for T. cruzi was also negative (performed at ARUP Laboratories). Tissue obtained from the cutaneous biopsy specimen identified T. cruzi by sequencing of the internal transcribed spacer 2 and D2 region of the 28S rRNA gene (performed at Stanford Health Care [1–3]).
Figure 2. A, Biopsy of skin nodules, 200× magnification, hematoxylin and eosin (H&E) stain. Histologic sections show an epidermis with mild spongiosis and an underlying lymphohistiocytic infiltrate. B, Biopsy of skin nodules, 400× magnification, H&E stain. Histologic sections show numerous parasitized histiocytes (demonstrated by arrows). The organisms are circular without a well-defined capsule.
Given the patient’s remote exposure to an area endemic for T. cruzi, she was diagnosed with reactivation of Chagas disease (CD). Immunosuppression was weaned, and oral benznidazole 150 mg twice daily (~5 mg/kg/d) was initiated.
Concurrent with the finding of CD, the patient was also found to have disseminated cytomegalovirus (CMV) disease. Plasma CMV viral load was >1.2 million copies/mL. Lung, colon, and skin biopsies all contained cells demonstrating cytopathic changes, which were positive by CMV immunostaining. Intravenous ganciclovir was started.
With treatment, the patient’s constitutional symptoms improved, and the skin lesions became less tender and erythematous. However, 1 month after the initiation of benznidazole, the patient developed Enterobacter cloacae bacteremia and septic shock. Shortly thereafter, she died.
Chagas Disease Epidemiology and Natural History
T. cruzi is primarily transmitted to humans via inoculation of wounds or mucosal membranes with infected feces of blood-sucking triatomine insects. CD is endemic throughout much of Latin America. However, as people have migrated from endemic areas, CD has become increasingly prevalent in traditionally nonendemic regions. In the United States, it is estimated that 1.3% of Latin American immigrants are infected with T. cruzi [4].
In acute CD, nearly all infected individuals have an effective host immune response that controls the parasitemia within 1–2 months of initial infection. However, in the absence of effective antiparasitic treatment, tissue infection persists for the life of the host. Chronic indeterminate CD occurs in persons without signs or symptoms of infection, with normal electrocardiogram, and normal radiographic appearance of the chest, esophagus, and colon. Indeterminate CD may persist for life or progress to determinate CD in 20%–30% of patients. Determinate CD results from tissue destruction related to persistent parasite replication and the immune response. It manifests with cardiac disease (conduction abnormalities, arrhythmias, dilated cardiomyopathy) and, less frequently, gastrointestinal disease (esophageal/colonic dysmotility and dilatation) [5, 6]. In some immunocompromised persons, chronic (indeterminate or determinate) CD can reactivate.
In conjunction with clinical symptoms, diagnosis of CD reactivation can be aided by laboratory testing. Serologic tests will be positive in most patients infected with T. cruzi. In chronic CD, patients can have transient parasitemia detected on microscopic examination of whole blood (or, preferably, buffy coat) smears [7]. With CD reactivation, patients often have more persistent parasitemia [8]. Positive T. cruzi blood PCR assays are suggestive but not diagnostic of reactivation. However, positive blood PCR results can herald the development of subsequent invasive Chagas disease reactivation in immunocompromised patients. Furthermore, rising parasite numbers demonstrated by quantitative PCR in serial specimens are highly suggestive of reactivated disease [8].
Reactivation of Chagas Disease
CD reactivation is best described in patients with advanced Chagas cardiomyopathy who undergo orthotopic heart transplantation [9–11]. The immunosuppressive drugs used to prevent transplant rejection predispose to CD reactivation. In these patients, reactivation most commonly manifests first as asymptomatic parasitemia or acute myocarditis [12]. Reactivation can also present as subcutaneous lesions [13], panniculitis [14], or, less commonly, meningoencephalitis [15]. Case series involving other solid organ transplant recipients (mostly renal transplant recipients) describe similar manifestations of CD reactivation [16–19]. CD reactivation is also known to occur in people with HIV/AIDS, most commonly presenting as meningoencephalitis [20] and/or brain abscesses (chagomas) [3, 21]. CD reactivation has also been described in patients receiving chemotherapy for hematologic and solid malignancies [22–25] and hematopoietic cell transplant (HCT) recipients [26–29].
Reactivation of Chagas Disease in Patients With Autoimmune Rheumatic Disease
There is a paucity of data regarding CD reactivation in patients receiving immunosuppressive therapy for autoimmune rheumatic diseases (ARDs; includes conditions such as systemic lupus erythematosus [SLE], rheumatoid arthritis, dermatomyositis, mixed connective tissue disease, and scleroderma). Compared with transplant recipients, patients with AIDS, and those receiving chemotherapy, patients undergoing treatment for ARD are often less immunosuppressed. However, it is unclear whether the dearth of literature in this cohort reflects infrequent reactivation or simply a lack of published case reports.
Among published English-language case reports that describe CD reactivation in patients with ARD on immunosuppressive regimens, the minority describe symptomatic CD reactivation (Table 1). Two patients developed brain chagomas—1 patient was successfully treated [30], and the other expired shortly after diagnosis [31]. Two other patients presented with Chagas skin lesions [30, 32], 1 of whom suffered recurrent disease after a course of benznidazole [30]. Other cases describe asymptomatic reactivation detected by T. cruzi PCR [33, 34], for which 1 patient was treated with off-label posaconazole after failing therapy with benznidazole [33]. The remaining patient reports describe possible reactivation detected through T. cruzi blood PCR positivity on a single test [30, 35]; given the absence of symptoms and serial testing, it is unclear if these patients had true CD reactivation or merely intermittently detectable parasites associated with chronic CD.
Table 1. Published Studies Describing Chagas Disease Reactivation (or Possible Reactivation) in Patients With Autoimmune Disease who Were Receiving Immunosuppressive Therapy
Study Patient Autoimmune Disease Immunosuppression Country of Origin Chagas Diagnosis Known Before Evaluation for Reactivation Evidence of Chagas Reactivation (or Possible Reactivation) Treatment Outcome
Current case 86 yo F Limited cutaneous systemic sclerosis Prednisone 15 mg/d, azathioprine 100 mg/d Venezuela No Skin lesions—erythematous, indurated, painful nodules on medial thighs and upper extremity Benznidazole Patient died 1 mo after initiation of benznidazole
Kaushal et al., 2019 [31] 88 yo F RA MTX Unknown No Brain chagoma Benznidazole Patient died shortly after diagnosis
German Sanchez et al., 2019 [30] 53 yo F SLE Prednisone 5 mg/d, MMF, CP All patients in study from Argentina No Brain chagoma and T. cruzi blood PCR pos Benznidazole PCR neg
68 yo F Psoriatic arthritis Prednisone 20 mg/d, HCQ, MTX Yes Panniculitis and T. cruzi blood PCR pos Benznidazole Relapse with recurrent skin lesions 1 y s/p treatment
7 of 13 patients with ARDs who had Chagas reactivation (or possible reactivation) 48 yo F RA MTX, adalimumab Yes Single T. Cruz blood PCR pos Benznidazole PCR neg
66 yo M RA Prednisone 5 mg/d, MTX, etanercept Yes Single T. Cruzi blood PCR pos Benznidazole PCR neg
Treatment durations for all patients were 1–2 mo 81 yo F RA Prednisone 5 mg/d, HCQ, MTX, leflunomide Yes Single T. Cruzi blood PCR pos Nifurtimox PCR neg
66 yo F Sjogren syndrome Prednisone 5 mg/d, HCQ Yes Single T. Cruzi blood PCR pos Benznidazole PCR neg
57 yo F Vasculitis Prednisone 5 mg/d, CP No Single T. cruzi blood PCR pos Benznidazole PCR neg
Vacas et al., 2017 [35] 57 yo M Psoriatic erythroderma Infliximab Argentina Yes Single T. cruzi blood PCR pos Benznidazole × 45 d PCR neg
Navarrete-Dechent et al., 2015 [34] 52 yo M Psoriasis Adalimumab Chile Yes—Chagas megacolon, received preemptive treatment with nifurtimox
T. cruzi blood PCR pos 8 months following preemptive nifurtimox Repeat course nifurtimox × 2 mo PCR neg
Burgos et al., 2012 [32] 44 yo F SLE Prednisone 50 mg/d, azathioprine 50 mg/d Paraguay No Parasitemia, skin lesions—erythematous, painful nodules that progressed to ulcer and eschar Benznidazole × 2 mo Clinically improved
Pinazo et al., 2010 [33] 44 yo F SLE Steroids, CP Argentina Yes—chronic indeterminate Chagas
T. cruzi blood PCR pos, treated with benznidazole with recurrent PCR pos Benznidazole × 2 mo, and then posaconazole × 3 mo for relapse Serial PCR neg after posaconazole
Abbreviations: CP, cyclophosphamide; HCQ, hydroxychloroquine; MMF, mycophenolate mofetil; MTX, methotrexate; NA, not applicable; neg, negative; PCR, polymerase chain reaction; pos, positive; RA, rheumatoid arthritis; SLE, systemic lupus erythematosus.
Although limited by small sample size and abbreviated longitudinal follow-up, other studies have attempted to investigate the risk for patients with ARD developing CD reactivation while on immunosuppression. One such study described 2 of 13 patients with ARD who developed symptomatic reactivation on immunosuppressive therapies over a 2-year period, and another 5 patients who developed possible CD reactivation detected by a single positive T. cruzi blood PCR test (Table 1) [30]. In other case series, the majority of immunosuppressed patients with ARD and CD were treated for chronic CD before the development of any evidence of CD reactivation, and only a small minority of these patients subsequently developed CD reactivation while receiving immunosuppression [25, 36, 37]. In 1 such study, 6 of 8 patients with ARD and CD were treated in this manner with varying regimens of benznidazole and nifurtimox. Three patients were treated for CD before a diagnosis of an ARD was made. Only 1 of the patients who received such treatment subsequently developed CD reactivation while receiving immunosuppression (further patient details not specified) [36]. In another study, 11 of 14 patients with chronic CD and ARD were treated for chronic CD with benznidazole for 60 days at the time of study enrollment, and none of the 14 patients experienced CD reactivation while receiving immunosuppression during follow-up [25]. Lastly, 3 patients with ARD and chronic CD were all treated at the time of CD diagnosis with benznidazole for 60 days; none of these patients developed subsequent CD reactivation during a 36-month follow-up period [37]. Though these preliminary data are intriguing, larger population studies, conducted over longer durations, are needed in order to draw more substantial conclusions about factors predisposing to CD reactivation in this cohort and the outcomes of prophylactic therapy.
DISCUSSION
Our patient is among the few in whom symptomatic CD reactivation was documented in the setting of immunosuppressive therapy for an ARD. Our patient likely had unrecognized determinate CD, clinically manifested by tachy-brady syndrome requiring a pacemaker [38]. CD reactivated after intensification of her immunosuppression regimen, ultimately manifesting as disseminated skin nodules, with sequencing of the skin biopsy revealing T. cruzi.
Our patient’s case was unique in that symptomatic CD reactivation occurred in the setting of a negative T. cruzi blood PCR. Typically, T. cruzi blood PCR positivity develops before symptomatic reactivation [11]. Though it is unclear why our patient’s PCR was negative, it is possibly related to blood volume collected or PCR test characteristics. Importantly, this demonstrates that PCR positivity should not be the sole diagnostic measure in the evaluation for CD reactivation; assessment must also include careful clinical evaluation.
CD reactivation in patients being treated for ARD additionally involves a different degree of immunosuppression compared with other better-studied cohorts. An effective host response against T. cruzi requires both cellular and humoral immunity [39]. Our patient was taking low-dose prednisone and azathioprine. Corticosteroids are nonspecific immune function inhibitors [40]. However, there are no reports that describe an increased incidence of CD reactivation in patients receiving corticosteroids as their sole form of immunosuppression. Azathioprine is an antimetabolite that decreases both T and B lymphocyte production [40]. However, some heart transplant recipients with a history of Chagas cardiomyopathy preferentially receive azathioprine over mycophenolate due to a 6-fold lower incidence of CD reactivation with azathioprine compared with mycophenolate [41]. Taken together, prednisone in doses <20 mg/d and azathioprine still seem to convey a relatively low risk for CD reactivation.
In our patient’s case, it is possible that more multifaceted immunosuppression led to her disease. Specifically, the synergistic effect of prednisone and azathioprine, in conjunction with her profound lymphopenia, advanced age, and uncontrolled diabetes, may have contributed to CD reactivation. However, these immunosuppressing factors still do not clearly explain our patient’s profound functional immunosuppression, further exhibited by concurrent disseminated CMV disease. Based on prior data [42–45], patients with rheumatoid arthritis and SLE may have an increased risk of infection independent of immunomodulatory therapy. Further research might better delineate the relationship between CD and immune function.
Screening for Chagas Disease in Patients With ARD
Currently, consensus guidelines recommend serologic screening for CD in transplant donors and recipients with epidemiologic risk factors. A positive serologic result should be confirmed by at least 2 distinct serologic methods. In the United States, CD treatment based solely on a positive serology result in either the donor or the recipient is generally not recommended given the toxicity of the therapeutic options [46, 47]. Alternatively, transplant recipients who are seropositive should be monitored for reactivation, especially during the times of most intense immunosuppression. Laboratory monitoring employs microscopy of blood/buffy coats and blood PCR. If monitoring reveals parasitemia and/or PCR positivity (especially increasing the parasite load on serial quantitative PCRs), patients are typically given preemptive CD treatment, as the development of detectable T. cruzi in this cohort often heralds the development of symptomatic reactivation [46–49].
Given that data regarding CD reactivation in patients with ARD are so scant, it is uncertain if similar screening and treatment guidelines should be applied to ARD patients in the face of immunosuppression. However, in light of our patient’s case and our review of the literature, our opinion is that strong consideration should be given to serologic screening for T. cruzi before immunosuppression for ARD in patients who have CD risk factors. (Note that serologic screening following immunosuppression may be falsely negative due to a blunted immune response.) Data are insufficient to comment on the risks versus benefits of CD treatment based solely on a positive serologic result in this cohort. However, similar to transplant recipients, for those who are seropositive, clinical assessment and serial blood microscopy/PCR monitoring should be employed during immunosuppressive therapy. There should be strong consideration of preemptive treatment of patients with parasitemia and/or PCR blood positivity (especially increasing parasite load on serial quantitative PCRs) even in the absence of symptoms; of course, evaluation/treatment for those with symptoms/signs concerning for CD reactivation. More research is needed to refine the screening and subsequent treatment approach for CD in patients with ARD.
CONCLUSIONS
This report describes CD reactivation in a patient with an ARD receiving immunosuppressive therapy. It highlights the need for more research regarding CD and reactivation in this patient population. It additionally suggests the need for broader serologic screening of patients with risk factors for CD before starting immunosuppression, and subsequent monitoring of at-risk patients while they receive such therapy.
Acknowledgments
Financial support. None
Potential conflicts of interest. All authors: no reported conflicts of interest. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed.
Disclaimer. The findings and conclusions in this report are those of the authors and do not necessarily represent the views of the Centers for Disease Control and Prevention.
Patient consent. Our study does not include factors necessitating patient consent. | AZATHIOPRINE, PREDNISONE | DrugsGivenReaction | CC BY-NC-ND | 33575423 | 19,494,598 | 2021-02 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Enterobacter infection'. | Reactivation of Chagas Disease in a Patient With an Autoimmune Rheumatic Disease: Case Report and Review of the Literature.
Reactivation of Chagas disease has been described in immunosuppressed patients, but there is a paucity of literature describing reactivation in patients on immunosuppressive therapies for the treatment of autoimmune rheumatic diseases. We describe a case of Chagas disease reactivation in a woman taking azathioprine and prednisone for limited cutaneous systemic sclerosis (lcSSc). Reactivation manifested as indurated and erythematous cutaneous nodules. Sequencing of a skin biopsy specimen confirmed the diagnosis of Chagas disease. She was treated with benznidazole with clinical improvement in the cutaneous lesions. However, her clinical course was complicated and included disseminated CMV disease and subsequent septic shock due to bacteremia. Our case and review of the literature highlight that screening for Chagas disease should be strongly considered for patients who will undergo immunosuppression for treatment of autoimmune disease if epidemiologically indicated.
PATIENT CASE
An 86-year-old woman developed painful subcutaneous nodules on her medial thighs and left upper extremity (Figure 1A, B). Over the ensuing 2 months, the nodules enlarged and became more indurated, erythematous, and painful. She also developed night sweats, fatigue, diarrhea, anorexia, and weight loss.
Figure 1. A, Skin lesions on left medial thigh. B, Skin lesions on right medial thigh.
The patient had a history of poorly controlled diabetes mellitus and limited cutaneous systemic sclerosis (lcSSc; a multisystem autoimmune rheumatic disease), manifested as pulmonary hypertension and Reynaud’s syndrome. Eight months before admission, she was diagnosed with tachy-brady syndrome, requiring a pacemaker. Echocardiogram at that time showed a dilated right ventricle with elevated pressures, preservation of left ventricle ejection fraction, and no wall motion abnormalities or apical aneurysms. For management of lcSSc, she had been on long-standing azathioprine (100 mg/d). Prednisone (15 mg/d) was added 3 months before the current presentation for management of a diagnosis of retinal vasculitis.
The patient was born and raised in a rural mountain village near Trujillo, Venezuela. As a child, she lived in a mud hut and cared for many animals including chickens, dogs, and cats. At 20 years of age, she emigrated to the United States. She traveled back to Venezuela only once at 70 years of age, and otherwise denied international travel.
She was admitted to the hospital, where she was initially afebrile and hemodynamically stable. Physical exam showed no mucosal abnormalities or regional lymphadenopathy. White blood cell count was 4.4 K/μL with profound lymphopenia (absolute lymphocytes 0.2 K/μL), hemoglobin 11.5 g/dL, and platelets 126 K/μL. Kidney and liver function were normal. An HIV antigen/antibody test was negative. Strongyloides serology was negative. Computed tomography scan showed scattered bilateral pulmonary nodules; normal caliber esophagus, small bowel, and large bowel; and inflammation in the ascending colon. Contrast-enhanced brain magnetic resonance imaging was normal.
Biopsy of the skin nodules (Figure 2A, B) showed a lymphohistiocytic infiltrate in the superficial and deep dermis with round intracellular organisms noted on hematoxylin/eosin stain. Periodic acid-Schiff-diastase, Gomori methenamine silver (GMS), Fite, and gram stains did not highlight the organisms. However, structures resembling kinetoplasts were minimally accentuated on GMS stain. Bacterial, fungal, and acid-fast bacillus cultures were ultimately negative. Serology for Trypanosoma cruzi (performed at Mayo Clinic Laboratories) was positive by both enzyme-linked immunosorbent assay and a lateral flow assay. A Giemsa-stained smear of peripheral blood (buffy coat) was negative, and real-time polymerase chain reaction (PCR) testing of a whole-blood sample for T. cruzi was also negative (performed at ARUP Laboratories). Tissue obtained from the cutaneous biopsy specimen identified T. cruzi by sequencing of the internal transcribed spacer 2 and D2 region of the 28S rRNA gene (performed at Stanford Health Care [1–3]).
Figure 2. A, Biopsy of skin nodules, 200× magnification, hematoxylin and eosin (H&E) stain. Histologic sections show an epidermis with mild spongiosis and an underlying lymphohistiocytic infiltrate. B, Biopsy of skin nodules, 400× magnification, H&E stain. Histologic sections show numerous parasitized histiocytes (demonstrated by arrows). The organisms are circular without a well-defined capsule.
Given the patient’s remote exposure to an area endemic for T. cruzi, she was diagnosed with reactivation of Chagas disease (CD). Immunosuppression was weaned, and oral benznidazole 150 mg twice daily (~5 mg/kg/d) was initiated.
Concurrent with the finding of CD, the patient was also found to have disseminated cytomegalovirus (CMV) disease. Plasma CMV viral load was >1.2 million copies/mL. Lung, colon, and skin biopsies all contained cells demonstrating cytopathic changes, which were positive by CMV immunostaining. Intravenous ganciclovir was started.
With treatment, the patient’s constitutional symptoms improved, and the skin lesions became less tender and erythematous. However, 1 month after the initiation of benznidazole, the patient developed Enterobacter cloacae bacteremia and septic shock. Shortly thereafter, she died.
Chagas Disease Epidemiology and Natural History
T. cruzi is primarily transmitted to humans via inoculation of wounds or mucosal membranes with infected feces of blood-sucking triatomine insects. CD is endemic throughout much of Latin America. However, as people have migrated from endemic areas, CD has become increasingly prevalent in traditionally nonendemic regions. In the United States, it is estimated that 1.3% of Latin American immigrants are infected with T. cruzi [4].
In acute CD, nearly all infected individuals have an effective host immune response that controls the parasitemia within 1–2 months of initial infection. However, in the absence of effective antiparasitic treatment, tissue infection persists for the life of the host. Chronic indeterminate CD occurs in persons without signs or symptoms of infection, with normal electrocardiogram, and normal radiographic appearance of the chest, esophagus, and colon. Indeterminate CD may persist for life or progress to determinate CD in 20%–30% of patients. Determinate CD results from tissue destruction related to persistent parasite replication and the immune response. It manifests with cardiac disease (conduction abnormalities, arrhythmias, dilated cardiomyopathy) and, less frequently, gastrointestinal disease (esophageal/colonic dysmotility and dilatation) [5, 6]. In some immunocompromised persons, chronic (indeterminate or determinate) CD can reactivate.
In conjunction with clinical symptoms, diagnosis of CD reactivation can be aided by laboratory testing. Serologic tests will be positive in most patients infected with T. cruzi. In chronic CD, patients can have transient parasitemia detected on microscopic examination of whole blood (or, preferably, buffy coat) smears [7]. With CD reactivation, patients often have more persistent parasitemia [8]. Positive T. cruzi blood PCR assays are suggestive but not diagnostic of reactivation. However, positive blood PCR results can herald the development of subsequent invasive Chagas disease reactivation in immunocompromised patients. Furthermore, rising parasite numbers demonstrated by quantitative PCR in serial specimens are highly suggestive of reactivated disease [8].
Reactivation of Chagas Disease
CD reactivation is best described in patients with advanced Chagas cardiomyopathy who undergo orthotopic heart transplantation [9–11]. The immunosuppressive drugs used to prevent transplant rejection predispose to CD reactivation. In these patients, reactivation most commonly manifests first as asymptomatic parasitemia or acute myocarditis [12]. Reactivation can also present as subcutaneous lesions [13], panniculitis [14], or, less commonly, meningoencephalitis [15]. Case series involving other solid organ transplant recipients (mostly renal transplant recipients) describe similar manifestations of CD reactivation [16–19]. CD reactivation is also known to occur in people with HIV/AIDS, most commonly presenting as meningoencephalitis [20] and/or brain abscesses (chagomas) [3, 21]. CD reactivation has also been described in patients receiving chemotherapy for hematologic and solid malignancies [22–25] and hematopoietic cell transplant (HCT) recipients [26–29].
Reactivation of Chagas Disease in Patients With Autoimmune Rheumatic Disease
There is a paucity of data regarding CD reactivation in patients receiving immunosuppressive therapy for autoimmune rheumatic diseases (ARDs; includes conditions such as systemic lupus erythematosus [SLE], rheumatoid arthritis, dermatomyositis, mixed connective tissue disease, and scleroderma). Compared with transplant recipients, patients with AIDS, and those receiving chemotherapy, patients undergoing treatment for ARD are often less immunosuppressed. However, it is unclear whether the dearth of literature in this cohort reflects infrequent reactivation or simply a lack of published case reports.
Among published English-language case reports that describe CD reactivation in patients with ARD on immunosuppressive regimens, the minority describe symptomatic CD reactivation (Table 1). Two patients developed brain chagomas—1 patient was successfully treated [30], and the other expired shortly after diagnosis [31]. Two other patients presented with Chagas skin lesions [30, 32], 1 of whom suffered recurrent disease after a course of benznidazole [30]. Other cases describe asymptomatic reactivation detected by T. cruzi PCR [33, 34], for which 1 patient was treated with off-label posaconazole after failing therapy with benznidazole [33]. The remaining patient reports describe possible reactivation detected through T. cruzi blood PCR positivity on a single test [30, 35]; given the absence of symptoms and serial testing, it is unclear if these patients had true CD reactivation or merely intermittently detectable parasites associated with chronic CD.
Table 1. Published Studies Describing Chagas Disease Reactivation (or Possible Reactivation) in Patients With Autoimmune Disease who Were Receiving Immunosuppressive Therapy
Study Patient Autoimmune Disease Immunosuppression Country of Origin Chagas Diagnosis Known Before Evaluation for Reactivation Evidence of Chagas Reactivation (or Possible Reactivation) Treatment Outcome
Current case 86 yo F Limited cutaneous systemic sclerosis Prednisone 15 mg/d, azathioprine 100 mg/d Venezuela No Skin lesions—erythematous, indurated, painful nodules on medial thighs and upper extremity Benznidazole Patient died 1 mo after initiation of benznidazole
Kaushal et al., 2019 [31] 88 yo F RA MTX Unknown No Brain chagoma Benznidazole Patient died shortly after diagnosis
German Sanchez et al., 2019 [30] 53 yo F SLE Prednisone 5 mg/d, MMF, CP All patients in study from Argentina No Brain chagoma and T. cruzi blood PCR pos Benznidazole PCR neg
68 yo F Psoriatic arthritis Prednisone 20 mg/d, HCQ, MTX Yes Panniculitis and T. cruzi blood PCR pos Benznidazole Relapse with recurrent skin lesions 1 y s/p treatment
7 of 13 patients with ARDs who had Chagas reactivation (or possible reactivation) 48 yo F RA MTX, adalimumab Yes Single T. Cruz blood PCR pos Benznidazole PCR neg
66 yo M RA Prednisone 5 mg/d, MTX, etanercept Yes Single T. Cruzi blood PCR pos Benznidazole PCR neg
Treatment durations for all patients were 1–2 mo 81 yo F RA Prednisone 5 mg/d, HCQ, MTX, leflunomide Yes Single T. Cruzi blood PCR pos Nifurtimox PCR neg
66 yo F Sjogren syndrome Prednisone 5 mg/d, HCQ Yes Single T. Cruzi blood PCR pos Benznidazole PCR neg
57 yo F Vasculitis Prednisone 5 mg/d, CP No Single T. cruzi blood PCR pos Benznidazole PCR neg
Vacas et al., 2017 [35] 57 yo M Psoriatic erythroderma Infliximab Argentina Yes Single T. cruzi blood PCR pos Benznidazole × 45 d PCR neg
Navarrete-Dechent et al., 2015 [34] 52 yo M Psoriasis Adalimumab Chile Yes—Chagas megacolon, received preemptive treatment with nifurtimox
T. cruzi blood PCR pos 8 months following preemptive nifurtimox Repeat course nifurtimox × 2 mo PCR neg
Burgos et al., 2012 [32] 44 yo F SLE Prednisone 50 mg/d, azathioprine 50 mg/d Paraguay No Parasitemia, skin lesions—erythematous, painful nodules that progressed to ulcer and eschar Benznidazole × 2 mo Clinically improved
Pinazo et al., 2010 [33] 44 yo F SLE Steroids, CP Argentina Yes—chronic indeterminate Chagas
T. cruzi blood PCR pos, treated with benznidazole with recurrent PCR pos Benznidazole × 2 mo, and then posaconazole × 3 mo for relapse Serial PCR neg after posaconazole
Abbreviations: CP, cyclophosphamide; HCQ, hydroxychloroquine; MMF, mycophenolate mofetil; MTX, methotrexate; NA, not applicable; neg, negative; PCR, polymerase chain reaction; pos, positive; RA, rheumatoid arthritis; SLE, systemic lupus erythematosus.
Although limited by small sample size and abbreviated longitudinal follow-up, other studies have attempted to investigate the risk for patients with ARD developing CD reactivation while on immunosuppression. One such study described 2 of 13 patients with ARD who developed symptomatic reactivation on immunosuppressive therapies over a 2-year period, and another 5 patients who developed possible CD reactivation detected by a single positive T. cruzi blood PCR test (Table 1) [30]. In other case series, the majority of immunosuppressed patients with ARD and CD were treated for chronic CD before the development of any evidence of CD reactivation, and only a small minority of these patients subsequently developed CD reactivation while receiving immunosuppression [25, 36, 37]. In 1 such study, 6 of 8 patients with ARD and CD were treated in this manner with varying regimens of benznidazole and nifurtimox. Three patients were treated for CD before a diagnosis of an ARD was made. Only 1 of the patients who received such treatment subsequently developed CD reactivation while receiving immunosuppression (further patient details not specified) [36]. In another study, 11 of 14 patients with chronic CD and ARD were treated for chronic CD with benznidazole for 60 days at the time of study enrollment, and none of the 14 patients experienced CD reactivation while receiving immunosuppression during follow-up [25]. Lastly, 3 patients with ARD and chronic CD were all treated at the time of CD diagnosis with benznidazole for 60 days; none of these patients developed subsequent CD reactivation during a 36-month follow-up period [37]. Though these preliminary data are intriguing, larger population studies, conducted over longer durations, are needed in order to draw more substantial conclusions about factors predisposing to CD reactivation in this cohort and the outcomes of prophylactic therapy.
DISCUSSION
Our patient is among the few in whom symptomatic CD reactivation was documented in the setting of immunosuppressive therapy for an ARD. Our patient likely had unrecognized determinate CD, clinically manifested by tachy-brady syndrome requiring a pacemaker [38]. CD reactivated after intensification of her immunosuppression regimen, ultimately manifesting as disseminated skin nodules, with sequencing of the skin biopsy revealing T. cruzi.
Our patient’s case was unique in that symptomatic CD reactivation occurred in the setting of a negative T. cruzi blood PCR. Typically, T. cruzi blood PCR positivity develops before symptomatic reactivation [11]. Though it is unclear why our patient’s PCR was negative, it is possibly related to blood volume collected or PCR test characteristics. Importantly, this demonstrates that PCR positivity should not be the sole diagnostic measure in the evaluation for CD reactivation; assessment must also include careful clinical evaluation.
CD reactivation in patients being treated for ARD additionally involves a different degree of immunosuppression compared with other better-studied cohorts. An effective host response against T. cruzi requires both cellular and humoral immunity [39]. Our patient was taking low-dose prednisone and azathioprine. Corticosteroids are nonspecific immune function inhibitors [40]. However, there are no reports that describe an increased incidence of CD reactivation in patients receiving corticosteroids as their sole form of immunosuppression. Azathioprine is an antimetabolite that decreases both T and B lymphocyte production [40]. However, some heart transplant recipients with a history of Chagas cardiomyopathy preferentially receive azathioprine over mycophenolate due to a 6-fold lower incidence of CD reactivation with azathioprine compared with mycophenolate [41]. Taken together, prednisone in doses <20 mg/d and azathioprine still seem to convey a relatively low risk for CD reactivation.
In our patient’s case, it is possible that more multifaceted immunosuppression led to her disease. Specifically, the synergistic effect of prednisone and azathioprine, in conjunction with her profound lymphopenia, advanced age, and uncontrolled diabetes, may have contributed to CD reactivation. However, these immunosuppressing factors still do not clearly explain our patient’s profound functional immunosuppression, further exhibited by concurrent disseminated CMV disease. Based on prior data [42–45], patients with rheumatoid arthritis and SLE may have an increased risk of infection independent of immunomodulatory therapy. Further research might better delineate the relationship between CD and immune function.
Screening for Chagas Disease in Patients With ARD
Currently, consensus guidelines recommend serologic screening for CD in transplant donors and recipients with epidemiologic risk factors. A positive serologic result should be confirmed by at least 2 distinct serologic methods. In the United States, CD treatment based solely on a positive serology result in either the donor or the recipient is generally not recommended given the toxicity of the therapeutic options [46, 47]. Alternatively, transplant recipients who are seropositive should be monitored for reactivation, especially during the times of most intense immunosuppression. Laboratory monitoring employs microscopy of blood/buffy coats and blood PCR. If monitoring reveals parasitemia and/or PCR positivity (especially increasing the parasite load on serial quantitative PCRs), patients are typically given preemptive CD treatment, as the development of detectable T. cruzi in this cohort often heralds the development of symptomatic reactivation [46–49].
Given that data regarding CD reactivation in patients with ARD are so scant, it is uncertain if similar screening and treatment guidelines should be applied to ARD patients in the face of immunosuppression. However, in light of our patient’s case and our review of the literature, our opinion is that strong consideration should be given to serologic screening for T. cruzi before immunosuppression for ARD in patients who have CD risk factors. (Note that serologic screening following immunosuppression may be falsely negative due to a blunted immune response.) Data are insufficient to comment on the risks versus benefits of CD treatment based solely on a positive serologic result in this cohort. However, similar to transplant recipients, for those who are seropositive, clinical assessment and serial blood microscopy/PCR monitoring should be employed during immunosuppressive therapy. There should be strong consideration of preemptive treatment of patients with parasitemia and/or PCR blood positivity (especially increasing parasite load on serial quantitative PCRs) even in the absence of symptoms; of course, evaluation/treatment for those with symptoms/signs concerning for CD reactivation. More research is needed to refine the screening and subsequent treatment approach for CD in patients with ARD.
CONCLUSIONS
This report describes CD reactivation in a patient with an ARD receiving immunosuppressive therapy. It highlights the need for more research regarding CD and reactivation in this patient population. It additionally suggests the need for broader serologic screening of patients with risk factors for CD before starting immunosuppression, and subsequent monitoring of at-risk patients while they receive such therapy.
Acknowledgments
Financial support. None
Potential conflicts of interest. All authors: no reported conflicts of interest. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed.
Disclaimer. The findings and conclusions in this report are those of the authors and do not necessarily represent the views of the Centers for Disease Control and Prevention.
Patient consent. Our study does not include factors necessitating patient consent. | AZATHIOPRINE, PREDNISONE | DrugsGivenReaction | CC BY-NC-ND | 33575423 | 19,494,598 | 2021-02 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Septic shock'. | Reactivation of Chagas Disease in a Patient With an Autoimmune Rheumatic Disease: Case Report and Review of the Literature.
Reactivation of Chagas disease has been described in immunosuppressed patients, but there is a paucity of literature describing reactivation in patients on immunosuppressive therapies for the treatment of autoimmune rheumatic diseases. We describe a case of Chagas disease reactivation in a woman taking azathioprine and prednisone for limited cutaneous systemic sclerosis (lcSSc). Reactivation manifested as indurated and erythematous cutaneous nodules. Sequencing of a skin biopsy specimen confirmed the diagnosis of Chagas disease. She was treated with benznidazole with clinical improvement in the cutaneous lesions. However, her clinical course was complicated and included disseminated CMV disease and subsequent septic shock due to bacteremia. Our case and review of the literature highlight that screening for Chagas disease should be strongly considered for patients who will undergo immunosuppression for treatment of autoimmune disease if epidemiologically indicated.
PATIENT CASE
An 86-year-old woman developed painful subcutaneous nodules on her medial thighs and left upper extremity (Figure 1A, B). Over the ensuing 2 months, the nodules enlarged and became more indurated, erythematous, and painful. She also developed night sweats, fatigue, diarrhea, anorexia, and weight loss.
Figure 1. A, Skin lesions on left medial thigh. B, Skin lesions on right medial thigh.
The patient had a history of poorly controlled diabetes mellitus and limited cutaneous systemic sclerosis (lcSSc; a multisystem autoimmune rheumatic disease), manifested as pulmonary hypertension and Reynaud’s syndrome. Eight months before admission, she was diagnosed with tachy-brady syndrome, requiring a pacemaker. Echocardiogram at that time showed a dilated right ventricle with elevated pressures, preservation of left ventricle ejection fraction, and no wall motion abnormalities or apical aneurysms. For management of lcSSc, she had been on long-standing azathioprine (100 mg/d). Prednisone (15 mg/d) was added 3 months before the current presentation for management of a diagnosis of retinal vasculitis.
The patient was born and raised in a rural mountain village near Trujillo, Venezuela. As a child, she lived in a mud hut and cared for many animals including chickens, dogs, and cats. At 20 years of age, she emigrated to the United States. She traveled back to Venezuela only once at 70 years of age, and otherwise denied international travel.
She was admitted to the hospital, where she was initially afebrile and hemodynamically stable. Physical exam showed no mucosal abnormalities or regional lymphadenopathy. White blood cell count was 4.4 K/μL with profound lymphopenia (absolute lymphocytes 0.2 K/μL), hemoglobin 11.5 g/dL, and platelets 126 K/μL. Kidney and liver function were normal. An HIV antigen/antibody test was negative. Strongyloides serology was negative. Computed tomography scan showed scattered bilateral pulmonary nodules; normal caliber esophagus, small bowel, and large bowel; and inflammation in the ascending colon. Contrast-enhanced brain magnetic resonance imaging was normal.
Biopsy of the skin nodules (Figure 2A, B) showed a lymphohistiocytic infiltrate in the superficial and deep dermis with round intracellular organisms noted on hematoxylin/eosin stain. Periodic acid-Schiff-diastase, Gomori methenamine silver (GMS), Fite, and gram stains did not highlight the organisms. However, structures resembling kinetoplasts were minimally accentuated on GMS stain. Bacterial, fungal, and acid-fast bacillus cultures were ultimately negative. Serology for Trypanosoma cruzi (performed at Mayo Clinic Laboratories) was positive by both enzyme-linked immunosorbent assay and a lateral flow assay. A Giemsa-stained smear of peripheral blood (buffy coat) was negative, and real-time polymerase chain reaction (PCR) testing of a whole-blood sample for T. cruzi was also negative (performed at ARUP Laboratories). Tissue obtained from the cutaneous biopsy specimen identified T. cruzi by sequencing of the internal transcribed spacer 2 and D2 region of the 28S rRNA gene (performed at Stanford Health Care [1–3]).
Figure 2. A, Biopsy of skin nodules, 200× magnification, hematoxylin and eosin (H&E) stain. Histologic sections show an epidermis with mild spongiosis and an underlying lymphohistiocytic infiltrate. B, Biopsy of skin nodules, 400× magnification, H&E stain. Histologic sections show numerous parasitized histiocytes (demonstrated by arrows). The organisms are circular without a well-defined capsule.
Given the patient’s remote exposure to an area endemic for T. cruzi, she was diagnosed with reactivation of Chagas disease (CD). Immunosuppression was weaned, and oral benznidazole 150 mg twice daily (~5 mg/kg/d) was initiated.
Concurrent with the finding of CD, the patient was also found to have disseminated cytomegalovirus (CMV) disease. Plasma CMV viral load was >1.2 million copies/mL. Lung, colon, and skin biopsies all contained cells demonstrating cytopathic changes, which were positive by CMV immunostaining. Intravenous ganciclovir was started.
With treatment, the patient’s constitutional symptoms improved, and the skin lesions became less tender and erythematous. However, 1 month after the initiation of benznidazole, the patient developed Enterobacter cloacae bacteremia and septic shock. Shortly thereafter, she died.
Chagas Disease Epidemiology and Natural History
T. cruzi is primarily transmitted to humans via inoculation of wounds or mucosal membranes with infected feces of blood-sucking triatomine insects. CD is endemic throughout much of Latin America. However, as people have migrated from endemic areas, CD has become increasingly prevalent in traditionally nonendemic regions. In the United States, it is estimated that 1.3% of Latin American immigrants are infected with T. cruzi [4].
In acute CD, nearly all infected individuals have an effective host immune response that controls the parasitemia within 1–2 months of initial infection. However, in the absence of effective antiparasitic treatment, tissue infection persists for the life of the host. Chronic indeterminate CD occurs in persons without signs or symptoms of infection, with normal electrocardiogram, and normal radiographic appearance of the chest, esophagus, and colon. Indeterminate CD may persist for life or progress to determinate CD in 20%–30% of patients. Determinate CD results from tissue destruction related to persistent parasite replication and the immune response. It manifests with cardiac disease (conduction abnormalities, arrhythmias, dilated cardiomyopathy) and, less frequently, gastrointestinal disease (esophageal/colonic dysmotility and dilatation) [5, 6]. In some immunocompromised persons, chronic (indeterminate or determinate) CD can reactivate.
In conjunction with clinical symptoms, diagnosis of CD reactivation can be aided by laboratory testing. Serologic tests will be positive in most patients infected with T. cruzi. In chronic CD, patients can have transient parasitemia detected on microscopic examination of whole blood (or, preferably, buffy coat) smears [7]. With CD reactivation, patients often have more persistent parasitemia [8]. Positive T. cruzi blood PCR assays are suggestive but not diagnostic of reactivation. However, positive blood PCR results can herald the development of subsequent invasive Chagas disease reactivation in immunocompromised patients. Furthermore, rising parasite numbers demonstrated by quantitative PCR in serial specimens are highly suggestive of reactivated disease [8].
Reactivation of Chagas Disease
CD reactivation is best described in patients with advanced Chagas cardiomyopathy who undergo orthotopic heart transplantation [9–11]. The immunosuppressive drugs used to prevent transplant rejection predispose to CD reactivation. In these patients, reactivation most commonly manifests first as asymptomatic parasitemia or acute myocarditis [12]. Reactivation can also present as subcutaneous lesions [13], panniculitis [14], or, less commonly, meningoencephalitis [15]. Case series involving other solid organ transplant recipients (mostly renal transplant recipients) describe similar manifestations of CD reactivation [16–19]. CD reactivation is also known to occur in people with HIV/AIDS, most commonly presenting as meningoencephalitis [20] and/or brain abscesses (chagomas) [3, 21]. CD reactivation has also been described in patients receiving chemotherapy for hematologic and solid malignancies [22–25] and hematopoietic cell transplant (HCT) recipients [26–29].
Reactivation of Chagas Disease in Patients With Autoimmune Rheumatic Disease
There is a paucity of data regarding CD reactivation in patients receiving immunosuppressive therapy for autoimmune rheumatic diseases (ARDs; includes conditions such as systemic lupus erythematosus [SLE], rheumatoid arthritis, dermatomyositis, mixed connective tissue disease, and scleroderma). Compared with transplant recipients, patients with AIDS, and those receiving chemotherapy, patients undergoing treatment for ARD are often less immunosuppressed. However, it is unclear whether the dearth of literature in this cohort reflects infrequent reactivation or simply a lack of published case reports.
Among published English-language case reports that describe CD reactivation in patients with ARD on immunosuppressive regimens, the minority describe symptomatic CD reactivation (Table 1). Two patients developed brain chagomas—1 patient was successfully treated [30], and the other expired shortly after diagnosis [31]. Two other patients presented with Chagas skin lesions [30, 32], 1 of whom suffered recurrent disease after a course of benznidazole [30]. Other cases describe asymptomatic reactivation detected by T. cruzi PCR [33, 34], for which 1 patient was treated with off-label posaconazole after failing therapy with benznidazole [33]. The remaining patient reports describe possible reactivation detected through T. cruzi blood PCR positivity on a single test [30, 35]; given the absence of symptoms and serial testing, it is unclear if these patients had true CD reactivation or merely intermittently detectable parasites associated with chronic CD.
Table 1. Published Studies Describing Chagas Disease Reactivation (or Possible Reactivation) in Patients With Autoimmune Disease who Were Receiving Immunosuppressive Therapy
Study Patient Autoimmune Disease Immunosuppression Country of Origin Chagas Diagnosis Known Before Evaluation for Reactivation Evidence of Chagas Reactivation (or Possible Reactivation) Treatment Outcome
Current case 86 yo F Limited cutaneous systemic sclerosis Prednisone 15 mg/d, azathioprine 100 mg/d Venezuela No Skin lesions—erythematous, indurated, painful nodules on medial thighs and upper extremity Benznidazole Patient died 1 mo after initiation of benznidazole
Kaushal et al., 2019 [31] 88 yo F RA MTX Unknown No Brain chagoma Benznidazole Patient died shortly after diagnosis
German Sanchez et al., 2019 [30] 53 yo F SLE Prednisone 5 mg/d, MMF, CP All patients in study from Argentina No Brain chagoma and T. cruzi blood PCR pos Benznidazole PCR neg
68 yo F Psoriatic arthritis Prednisone 20 mg/d, HCQ, MTX Yes Panniculitis and T. cruzi blood PCR pos Benznidazole Relapse with recurrent skin lesions 1 y s/p treatment
7 of 13 patients with ARDs who had Chagas reactivation (or possible reactivation) 48 yo F RA MTX, adalimumab Yes Single T. Cruz blood PCR pos Benznidazole PCR neg
66 yo M RA Prednisone 5 mg/d, MTX, etanercept Yes Single T. Cruzi blood PCR pos Benznidazole PCR neg
Treatment durations for all patients were 1–2 mo 81 yo F RA Prednisone 5 mg/d, HCQ, MTX, leflunomide Yes Single T. Cruzi blood PCR pos Nifurtimox PCR neg
66 yo F Sjogren syndrome Prednisone 5 mg/d, HCQ Yes Single T. Cruzi blood PCR pos Benznidazole PCR neg
57 yo F Vasculitis Prednisone 5 mg/d, CP No Single T. cruzi blood PCR pos Benznidazole PCR neg
Vacas et al., 2017 [35] 57 yo M Psoriatic erythroderma Infliximab Argentina Yes Single T. cruzi blood PCR pos Benznidazole × 45 d PCR neg
Navarrete-Dechent et al., 2015 [34] 52 yo M Psoriasis Adalimumab Chile Yes—Chagas megacolon, received preemptive treatment with nifurtimox
T. cruzi blood PCR pos 8 months following preemptive nifurtimox Repeat course nifurtimox × 2 mo PCR neg
Burgos et al., 2012 [32] 44 yo F SLE Prednisone 50 mg/d, azathioprine 50 mg/d Paraguay No Parasitemia, skin lesions—erythematous, painful nodules that progressed to ulcer and eschar Benznidazole × 2 mo Clinically improved
Pinazo et al., 2010 [33] 44 yo F SLE Steroids, CP Argentina Yes—chronic indeterminate Chagas
T. cruzi blood PCR pos, treated with benznidazole with recurrent PCR pos Benznidazole × 2 mo, and then posaconazole × 3 mo for relapse Serial PCR neg after posaconazole
Abbreviations: CP, cyclophosphamide; HCQ, hydroxychloroquine; MMF, mycophenolate mofetil; MTX, methotrexate; NA, not applicable; neg, negative; PCR, polymerase chain reaction; pos, positive; RA, rheumatoid arthritis; SLE, systemic lupus erythematosus.
Although limited by small sample size and abbreviated longitudinal follow-up, other studies have attempted to investigate the risk for patients with ARD developing CD reactivation while on immunosuppression. One such study described 2 of 13 patients with ARD who developed symptomatic reactivation on immunosuppressive therapies over a 2-year period, and another 5 patients who developed possible CD reactivation detected by a single positive T. cruzi blood PCR test (Table 1) [30]. In other case series, the majority of immunosuppressed patients with ARD and CD were treated for chronic CD before the development of any evidence of CD reactivation, and only a small minority of these patients subsequently developed CD reactivation while receiving immunosuppression [25, 36, 37]. In 1 such study, 6 of 8 patients with ARD and CD were treated in this manner with varying regimens of benznidazole and nifurtimox. Three patients were treated for CD before a diagnosis of an ARD was made. Only 1 of the patients who received such treatment subsequently developed CD reactivation while receiving immunosuppression (further patient details not specified) [36]. In another study, 11 of 14 patients with chronic CD and ARD were treated for chronic CD with benznidazole for 60 days at the time of study enrollment, and none of the 14 patients experienced CD reactivation while receiving immunosuppression during follow-up [25]. Lastly, 3 patients with ARD and chronic CD were all treated at the time of CD diagnosis with benznidazole for 60 days; none of these patients developed subsequent CD reactivation during a 36-month follow-up period [37]. Though these preliminary data are intriguing, larger population studies, conducted over longer durations, are needed in order to draw more substantial conclusions about factors predisposing to CD reactivation in this cohort and the outcomes of prophylactic therapy.
DISCUSSION
Our patient is among the few in whom symptomatic CD reactivation was documented in the setting of immunosuppressive therapy for an ARD. Our patient likely had unrecognized determinate CD, clinically manifested by tachy-brady syndrome requiring a pacemaker [38]. CD reactivated after intensification of her immunosuppression regimen, ultimately manifesting as disseminated skin nodules, with sequencing of the skin biopsy revealing T. cruzi.
Our patient’s case was unique in that symptomatic CD reactivation occurred in the setting of a negative T. cruzi blood PCR. Typically, T. cruzi blood PCR positivity develops before symptomatic reactivation [11]. Though it is unclear why our patient’s PCR was negative, it is possibly related to blood volume collected or PCR test characteristics. Importantly, this demonstrates that PCR positivity should not be the sole diagnostic measure in the evaluation for CD reactivation; assessment must also include careful clinical evaluation.
CD reactivation in patients being treated for ARD additionally involves a different degree of immunosuppression compared with other better-studied cohorts. An effective host response against T. cruzi requires both cellular and humoral immunity [39]. Our patient was taking low-dose prednisone and azathioprine. Corticosteroids are nonspecific immune function inhibitors [40]. However, there are no reports that describe an increased incidence of CD reactivation in patients receiving corticosteroids as their sole form of immunosuppression. Azathioprine is an antimetabolite that decreases both T and B lymphocyte production [40]. However, some heart transplant recipients with a history of Chagas cardiomyopathy preferentially receive azathioprine over mycophenolate due to a 6-fold lower incidence of CD reactivation with azathioprine compared with mycophenolate [41]. Taken together, prednisone in doses <20 mg/d and azathioprine still seem to convey a relatively low risk for CD reactivation.
In our patient’s case, it is possible that more multifaceted immunosuppression led to her disease. Specifically, the synergistic effect of prednisone and azathioprine, in conjunction with her profound lymphopenia, advanced age, and uncontrolled diabetes, may have contributed to CD reactivation. However, these immunosuppressing factors still do not clearly explain our patient’s profound functional immunosuppression, further exhibited by concurrent disseminated CMV disease. Based on prior data [42–45], patients with rheumatoid arthritis and SLE may have an increased risk of infection independent of immunomodulatory therapy. Further research might better delineate the relationship between CD and immune function.
Screening for Chagas Disease in Patients With ARD
Currently, consensus guidelines recommend serologic screening for CD in transplant donors and recipients with epidemiologic risk factors. A positive serologic result should be confirmed by at least 2 distinct serologic methods. In the United States, CD treatment based solely on a positive serology result in either the donor or the recipient is generally not recommended given the toxicity of the therapeutic options [46, 47]. Alternatively, transplant recipients who are seropositive should be monitored for reactivation, especially during the times of most intense immunosuppression. Laboratory monitoring employs microscopy of blood/buffy coats and blood PCR. If monitoring reveals parasitemia and/or PCR positivity (especially increasing the parasite load on serial quantitative PCRs), patients are typically given preemptive CD treatment, as the development of detectable T. cruzi in this cohort often heralds the development of symptomatic reactivation [46–49].
Given that data regarding CD reactivation in patients with ARD are so scant, it is uncertain if similar screening and treatment guidelines should be applied to ARD patients in the face of immunosuppression. However, in light of our patient’s case and our review of the literature, our opinion is that strong consideration should be given to serologic screening for T. cruzi before immunosuppression for ARD in patients who have CD risk factors. (Note that serologic screening following immunosuppression may be falsely negative due to a blunted immune response.) Data are insufficient to comment on the risks versus benefits of CD treatment based solely on a positive serologic result in this cohort. However, similar to transplant recipients, for those who are seropositive, clinical assessment and serial blood microscopy/PCR monitoring should be employed during immunosuppressive therapy. There should be strong consideration of preemptive treatment of patients with parasitemia and/or PCR blood positivity (especially increasing parasite load on serial quantitative PCRs) even in the absence of symptoms; of course, evaluation/treatment for those with symptoms/signs concerning for CD reactivation. More research is needed to refine the screening and subsequent treatment approach for CD in patients with ARD.
CONCLUSIONS
This report describes CD reactivation in a patient with an ARD receiving immunosuppressive therapy. It highlights the need for more research regarding CD and reactivation in this patient population. It additionally suggests the need for broader serologic screening of patients with risk factors for CD before starting immunosuppression, and subsequent monitoring of at-risk patients while they receive such therapy.
Acknowledgments
Financial support. None
Potential conflicts of interest. All authors: no reported conflicts of interest. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed.
Disclaimer. The findings and conclusions in this report are those of the authors and do not necessarily represent the views of the Centers for Disease Control and Prevention.
Patient consent. Our study does not include factors necessitating patient consent. | AZATHIOPRINE, PREDNISONE | DrugsGivenReaction | CC BY-NC-ND | 33575423 | 19,494,598 | 2021-02 |
What was the outcome of reaction 'American trypanosomiasis'? | Reactivation of Chagas Disease in a Patient With an Autoimmune Rheumatic Disease: Case Report and Review of the Literature.
Reactivation of Chagas disease has been described in immunosuppressed patients, but there is a paucity of literature describing reactivation in patients on immunosuppressive therapies for the treatment of autoimmune rheumatic diseases. We describe a case of Chagas disease reactivation in a woman taking azathioprine and prednisone for limited cutaneous systemic sclerosis (lcSSc). Reactivation manifested as indurated and erythematous cutaneous nodules. Sequencing of a skin biopsy specimen confirmed the diagnosis of Chagas disease. She was treated with benznidazole with clinical improvement in the cutaneous lesions. However, her clinical course was complicated and included disseminated CMV disease and subsequent septic shock due to bacteremia. Our case and review of the literature highlight that screening for Chagas disease should be strongly considered for patients who will undergo immunosuppression for treatment of autoimmune disease if epidemiologically indicated.
PATIENT CASE
An 86-year-old woman developed painful subcutaneous nodules on her medial thighs and left upper extremity (Figure 1A, B). Over the ensuing 2 months, the nodules enlarged and became more indurated, erythematous, and painful. She also developed night sweats, fatigue, diarrhea, anorexia, and weight loss.
Figure 1. A, Skin lesions on left medial thigh. B, Skin lesions on right medial thigh.
The patient had a history of poorly controlled diabetes mellitus and limited cutaneous systemic sclerosis (lcSSc; a multisystem autoimmune rheumatic disease), manifested as pulmonary hypertension and Reynaud’s syndrome. Eight months before admission, she was diagnosed with tachy-brady syndrome, requiring a pacemaker. Echocardiogram at that time showed a dilated right ventricle with elevated pressures, preservation of left ventricle ejection fraction, and no wall motion abnormalities or apical aneurysms. For management of lcSSc, she had been on long-standing azathioprine (100 mg/d). Prednisone (15 mg/d) was added 3 months before the current presentation for management of a diagnosis of retinal vasculitis.
The patient was born and raised in a rural mountain village near Trujillo, Venezuela. As a child, she lived in a mud hut and cared for many animals including chickens, dogs, and cats. At 20 years of age, she emigrated to the United States. She traveled back to Venezuela only once at 70 years of age, and otherwise denied international travel.
She was admitted to the hospital, where she was initially afebrile and hemodynamically stable. Physical exam showed no mucosal abnormalities or regional lymphadenopathy. White blood cell count was 4.4 K/μL with profound lymphopenia (absolute lymphocytes 0.2 K/μL), hemoglobin 11.5 g/dL, and platelets 126 K/μL. Kidney and liver function were normal. An HIV antigen/antibody test was negative. Strongyloides serology was negative. Computed tomography scan showed scattered bilateral pulmonary nodules; normal caliber esophagus, small bowel, and large bowel; and inflammation in the ascending colon. Contrast-enhanced brain magnetic resonance imaging was normal.
Biopsy of the skin nodules (Figure 2A, B) showed a lymphohistiocytic infiltrate in the superficial and deep dermis with round intracellular organisms noted on hematoxylin/eosin stain. Periodic acid-Schiff-diastase, Gomori methenamine silver (GMS), Fite, and gram stains did not highlight the organisms. However, structures resembling kinetoplasts were minimally accentuated on GMS stain. Bacterial, fungal, and acid-fast bacillus cultures were ultimately negative. Serology for Trypanosoma cruzi (performed at Mayo Clinic Laboratories) was positive by both enzyme-linked immunosorbent assay and a lateral flow assay. A Giemsa-stained smear of peripheral blood (buffy coat) was negative, and real-time polymerase chain reaction (PCR) testing of a whole-blood sample for T. cruzi was also negative (performed at ARUP Laboratories). Tissue obtained from the cutaneous biopsy specimen identified T. cruzi by sequencing of the internal transcribed spacer 2 and D2 region of the 28S rRNA gene (performed at Stanford Health Care [1–3]).
Figure 2. A, Biopsy of skin nodules, 200× magnification, hematoxylin and eosin (H&E) stain. Histologic sections show an epidermis with mild spongiosis and an underlying lymphohistiocytic infiltrate. B, Biopsy of skin nodules, 400× magnification, H&E stain. Histologic sections show numerous parasitized histiocytes (demonstrated by arrows). The organisms are circular without a well-defined capsule.
Given the patient’s remote exposure to an area endemic for T. cruzi, she was diagnosed with reactivation of Chagas disease (CD). Immunosuppression was weaned, and oral benznidazole 150 mg twice daily (~5 mg/kg/d) was initiated.
Concurrent with the finding of CD, the patient was also found to have disseminated cytomegalovirus (CMV) disease. Plasma CMV viral load was >1.2 million copies/mL. Lung, colon, and skin biopsies all contained cells demonstrating cytopathic changes, which were positive by CMV immunostaining. Intravenous ganciclovir was started.
With treatment, the patient’s constitutional symptoms improved, and the skin lesions became less tender and erythematous. However, 1 month after the initiation of benznidazole, the patient developed Enterobacter cloacae bacteremia and septic shock. Shortly thereafter, she died.
Chagas Disease Epidemiology and Natural History
T. cruzi is primarily transmitted to humans via inoculation of wounds or mucosal membranes with infected feces of blood-sucking triatomine insects. CD is endemic throughout much of Latin America. However, as people have migrated from endemic areas, CD has become increasingly prevalent in traditionally nonendemic regions. In the United States, it is estimated that 1.3% of Latin American immigrants are infected with T. cruzi [4].
In acute CD, nearly all infected individuals have an effective host immune response that controls the parasitemia within 1–2 months of initial infection. However, in the absence of effective antiparasitic treatment, tissue infection persists for the life of the host. Chronic indeterminate CD occurs in persons without signs or symptoms of infection, with normal electrocardiogram, and normal radiographic appearance of the chest, esophagus, and colon. Indeterminate CD may persist for life or progress to determinate CD in 20%–30% of patients. Determinate CD results from tissue destruction related to persistent parasite replication and the immune response. It manifests with cardiac disease (conduction abnormalities, arrhythmias, dilated cardiomyopathy) and, less frequently, gastrointestinal disease (esophageal/colonic dysmotility and dilatation) [5, 6]. In some immunocompromised persons, chronic (indeterminate or determinate) CD can reactivate.
In conjunction with clinical symptoms, diagnosis of CD reactivation can be aided by laboratory testing. Serologic tests will be positive in most patients infected with T. cruzi. In chronic CD, patients can have transient parasitemia detected on microscopic examination of whole blood (or, preferably, buffy coat) smears [7]. With CD reactivation, patients often have more persistent parasitemia [8]. Positive T. cruzi blood PCR assays are suggestive but not diagnostic of reactivation. However, positive blood PCR results can herald the development of subsequent invasive Chagas disease reactivation in immunocompromised patients. Furthermore, rising parasite numbers demonstrated by quantitative PCR in serial specimens are highly suggestive of reactivated disease [8].
Reactivation of Chagas Disease
CD reactivation is best described in patients with advanced Chagas cardiomyopathy who undergo orthotopic heart transplantation [9–11]. The immunosuppressive drugs used to prevent transplant rejection predispose to CD reactivation. In these patients, reactivation most commonly manifests first as asymptomatic parasitemia or acute myocarditis [12]. Reactivation can also present as subcutaneous lesions [13], panniculitis [14], or, less commonly, meningoencephalitis [15]. Case series involving other solid organ transplant recipients (mostly renal transplant recipients) describe similar manifestations of CD reactivation [16–19]. CD reactivation is also known to occur in people with HIV/AIDS, most commonly presenting as meningoencephalitis [20] and/or brain abscesses (chagomas) [3, 21]. CD reactivation has also been described in patients receiving chemotherapy for hematologic and solid malignancies [22–25] and hematopoietic cell transplant (HCT) recipients [26–29].
Reactivation of Chagas Disease in Patients With Autoimmune Rheumatic Disease
There is a paucity of data regarding CD reactivation in patients receiving immunosuppressive therapy for autoimmune rheumatic diseases (ARDs; includes conditions such as systemic lupus erythematosus [SLE], rheumatoid arthritis, dermatomyositis, mixed connective tissue disease, and scleroderma). Compared with transplant recipients, patients with AIDS, and those receiving chemotherapy, patients undergoing treatment for ARD are often less immunosuppressed. However, it is unclear whether the dearth of literature in this cohort reflects infrequent reactivation or simply a lack of published case reports.
Among published English-language case reports that describe CD reactivation in patients with ARD on immunosuppressive regimens, the minority describe symptomatic CD reactivation (Table 1). Two patients developed brain chagomas—1 patient was successfully treated [30], and the other expired shortly after diagnosis [31]. Two other patients presented with Chagas skin lesions [30, 32], 1 of whom suffered recurrent disease after a course of benznidazole [30]. Other cases describe asymptomatic reactivation detected by T. cruzi PCR [33, 34], for which 1 patient was treated with off-label posaconazole after failing therapy with benznidazole [33]. The remaining patient reports describe possible reactivation detected through T. cruzi blood PCR positivity on a single test [30, 35]; given the absence of symptoms and serial testing, it is unclear if these patients had true CD reactivation or merely intermittently detectable parasites associated with chronic CD.
Table 1. Published Studies Describing Chagas Disease Reactivation (or Possible Reactivation) in Patients With Autoimmune Disease who Were Receiving Immunosuppressive Therapy
Study Patient Autoimmune Disease Immunosuppression Country of Origin Chagas Diagnosis Known Before Evaluation for Reactivation Evidence of Chagas Reactivation (or Possible Reactivation) Treatment Outcome
Current case 86 yo F Limited cutaneous systemic sclerosis Prednisone 15 mg/d, azathioprine 100 mg/d Venezuela No Skin lesions—erythematous, indurated, painful nodules on medial thighs and upper extremity Benznidazole Patient died 1 mo after initiation of benznidazole
Kaushal et al., 2019 [31] 88 yo F RA MTX Unknown No Brain chagoma Benznidazole Patient died shortly after diagnosis
German Sanchez et al., 2019 [30] 53 yo F SLE Prednisone 5 mg/d, MMF, CP All patients in study from Argentina No Brain chagoma and T. cruzi blood PCR pos Benznidazole PCR neg
68 yo F Psoriatic arthritis Prednisone 20 mg/d, HCQ, MTX Yes Panniculitis and T. cruzi blood PCR pos Benznidazole Relapse with recurrent skin lesions 1 y s/p treatment
7 of 13 patients with ARDs who had Chagas reactivation (or possible reactivation) 48 yo F RA MTX, adalimumab Yes Single T. Cruz blood PCR pos Benznidazole PCR neg
66 yo M RA Prednisone 5 mg/d, MTX, etanercept Yes Single T. Cruzi blood PCR pos Benznidazole PCR neg
Treatment durations for all patients were 1–2 mo 81 yo F RA Prednisone 5 mg/d, HCQ, MTX, leflunomide Yes Single T. Cruzi blood PCR pos Nifurtimox PCR neg
66 yo F Sjogren syndrome Prednisone 5 mg/d, HCQ Yes Single T. Cruzi blood PCR pos Benznidazole PCR neg
57 yo F Vasculitis Prednisone 5 mg/d, CP No Single T. cruzi blood PCR pos Benznidazole PCR neg
Vacas et al., 2017 [35] 57 yo M Psoriatic erythroderma Infliximab Argentina Yes Single T. cruzi blood PCR pos Benznidazole × 45 d PCR neg
Navarrete-Dechent et al., 2015 [34] 52 yo M Psoriasis Adalimumab Chile Yes—Chagas megacolon, received preemptive treatment with nifurtimox
T. cruzi blood PCR pos 8 months following preemptive nifurtimox Repeat course nifurtimox × 2 mo PCR neg
Burgos et al., 2012 [32] 44 yo F SLE Prednisone 50 mg/d, azathioprine 50 mg/d Paraguay No Parasitemia, skin lesions—erythematous, painful nodules that progressed to ulcer and eschar Benznidazole × 2 mo Clinically improved
Pinazo et al., 2010 [33] 44 yo F SLE Steroids, CP Argentina Yes—chronic indeterminate Chagas
T. cruzi blood PCR pos, treated with benznidazole with recurrent PCR pos Benznidazole × 2 mo, and then posaconazole × 3 mo for relapse Serial PCR neg after posaconazole
Abbreviations: CP, cyclophosphamide; HCQ, hydroxychloroquine; MMF, mycophenolate mofetil; MTX, methotrexate; NA, not applicable; neg, negative; PCR, polymerase chain reaction; pos, positive; RA, rheumatoid arthritis; SLE, systemic lupus erythematosus.
Although limited by small sample size and abbreviated longitudinal follow-up, other studies have attempted to investigate the risk for patients with ARD developing CD reactivation while on immunosuppression. One such study described 2 of 13 patients with ARD who developed symptomatic reactivation on immunosuppressive therapies over a 2-year period, and another 5 patients who developed possible CD reactivation detected by a single positive T. cruzi blood PCR test (Table 1) [30]. In other case series, the majority of immunosuppressed patients with ARD and CD were treated for chronic CD before the development of any evidence of CD reactivation, and only a small minority of these patients subsequently developed CD reactivation while receiving immunosuppression [25, 36, 37]. In 1 such study, 6 of 8 patients with ARD and CD were treated in this manner with varying regimens of benznidazole and nifurtimox. Three patients were treated for CD before a diagnosis of an ARD was made. Only 1 of the patients who received such treatment subsequently developed CD reactivation while receiving immunosuppression (further patient details not specified) [36]. In another study, 11 of 14 patients with chronic CD and ARD were treated for chronic CD with benznidazole for 60 days at the time of study enrollment, and none of the 14 patients experienced CD reactivation while receiving immunosuppression during follow-up [25]. Lastly, 3 patients with ARD and chronic CD were all treated at the time of CD diagnosis with benznidazole for 60 days; none of these patients developed subsequent CD reactivation during a 36-month follow-up period [37]. Though these preliminary data are intriguing, larger population studies, conducted over longer durations, are needed in order to draw more substantial conclusions about factors predisposing to CD reactivation in this cohort and the outcomes of prophylactic therapy.
DISCUSSION
Our patient is among the few in whom symptomatic CD reactivation was documented in the setting of immunosuppressive therapy for an ARD. Our patient likely had unrecognized determinate CD, clinically manifested by tachy-brady syndrome requiring a pacemaker [38]. CD reactivated after intensification of her immunosuppression regimen, ultimately manifesting as disseminated skin nodules, with sequencing of the skin biopsy revealing T. cruzi.
Our patient’s case was unique in that symptomatic CD reactivation occurred in the setting of a negative T. cruzi blood PCR. Typically, T. cruzi blood PCR positivity develops before symptomatic reactivation [11]. Though it is unclear why our patient’s PCR was negative, it is possibly related to blood volume collected or PCR test characteristics. Importantly, this demonstrates that PCR positivity should not be the sole diagnostic measure in the evaluation for CD reactivation; assessment must also include careful clinical evaluation.
CD reactivation in patients being treated for ARD additionally involves a different degree of immunosuppression compared with other better-studied cohorts. An effective host response against T. cruzi requires both cellular and humoral immunity [39]. Our patient was taking low-dose prednisone and azathioprine. Corticosteroids are nonspecific immune function inhibitors [40]. However, there are no reports that describe an increased incidence of CD reactivation in patients receiving corticosteroids as their sole form of immunosuppression. Azathioprine is an antimetabolite that decreases both T and B lymphocyte production [40]. However, some heart transplant recipients with a history of Chagas cardiomyopathy preferentially receive azathioprine over mycophenolate due to a 6-fold lower incidence of CD reactivation with azathioprine compared with mycophenolate [41]. Taken together, prednisone in doses <20 mg/d and azathioprine still seem to convey a relatively low risk for CD reactivation.
In our patient’s case, it is possible that more multifaceted immunosuppression led to her disease. Specifically, the synergistic effect of prednisone and azathioprine, in conjunction with her profound lymphopenia, advanced age, and uncontrolled diabetes, may have contributed to CD reactivation. However, these immunosuppressing factors still do not clearly explain our patient’s profound functional immunosuppression, further exhibited by concurrent disseminated CMV disease. Based on prior data [42–45], patients with rheumatoid arthritis and SLE may have an increased risk of infection independent of immunomodulatory therapy. Further research might better delineate the relationship between CD and immune function.
Screening for Chagas Disease in Patients With ARD
Currently, consensus guidelines recommend serologic screening for CD in transplant donors and recipients with epidemiologic risk factors. A positive serologic result should be confirmed by at least 2 distinct serologic methods. In the United States, CD treatment based solely on a positive serology result in either the donor or the recipient is generally not recommended given the toxicity of the therapeutic options [46, 47]. Alternatively, transplant recipients who are seropositive should be monitored for reactivation, especially during the times of most intense immunosuppression. Laboratory monitoring employs microscopy of blood/buffy coats and blood PCR. If monitoring reveals parasitemia and/or PCR positivity (especially increasing the parasite load on serial quantitative PCRs), patients are typically given preemptive CD treatment, as the development of detectable T. cruzi in this cohort often heralds the development of symptomatic reactivation [46–49].
Given that data regarding CD reactivation in patients with ARD are so scant, it is uncertain if similar screening and treatment guidelines should be applied to ARD patients in the face of immunosuppression. However, in light of our patient’s case and our review of the literature, our opinion is that strong consideration should be given to serologic screening for T. cruzi before immunosuppression for ARD in patients who have CD risk factors. (Note that serologic screening following immunosuppression may be falsely negative due to a blunted immune response.) Data are insufficient to comment on the risks versus benefits of CD treatment based solely on a positive serologic result in this cohort. However, similar to transplant recipients, for those who are seropositive, clinical assessment and serial blood microscopy/PCR monitoring should be employed during immunosuppressive therapy. There should be strong consideration of preemptive treatment of patients with parasitemia and/or PCR blood positivity (especially increasing parasite load on serial quantitative PCRs) even in the absence of symptoms; of course, evaluation/treatment for those with symptoms/signs concerning for CD reactivation. More research is needed to refine the screening and subsequent treatment approach for CD in patients with ARD.
CONCLUSIONS
This report describes CD reactivation in a patient with an ARD receiving immunosuppressive therapy. It highlights the need for more research regarding CD and reactivation in this patient population. It additionally suggests the need for broader serologic screening of patients with risk factors for CD before starting immunosuppression, and subsequent monitoring of at-risk patients while they receive such therapy.
Acknowledgments
Financial support. None
Potential conflicts of interest. All authors: no reported conflicts of interest. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed.
Disclaimer. The findings and conclusions in this report are those of the authors and do not necessarily represent the views of the Centers for Disease Control and Prevention.
Patient consent. Our study does not include factors necessitating patient consent. | Recovering | ReactionOutcome | CC BY-NC-ND | 33575423 | 19,507,537 | 2021-02 |
What was the outcome of reaction 'Disseminated cytomegaloviral infection'? | Reactivation of Chagas Disease in a Patient With an Autoimmune Rheumatic Disease: Case Report and Review of the Literature.
Reactivation of Chagas disease has been described in immunosuppressed patients, but there is a paucity of literature describing reactivation in patients on immunosuppressive therapies for the treatment of autoimmune rheumatic diseases. We describe a case of Chagas disease reactivation in a woman taking azathioprine and prednisone for limited cutaneous systemic sclerosis (lcSSc). Reactivation manifested as indurated and erythematous cutaneous nodules. Sequencing of a skin biopsy specimen confirmed the diagnosis of Chagas disease. She was treated with benznidazole with clinical improvement in the cutaneous lesions. However, her clinical course was complicated and included disseminated CMV disease and subsequent septic shock due to bacteremia. Our case and review of the literature highlight that screening for Chagas disease should be strongly considered for patients who will undergo immunosuppression for treatment of autoimmune disease if epidemiologically indicated.
PATIENT CASE
An 86-year-old woman developed painful subcutaneous nodules on her medial thighs and left upper extremity (Figure 1A, B). Over the ensuing 2 months, the nodules enlarged and became more indurated, erythematous, and painful. She also developed night sweats, fatigue, diarrhea, anorexia, and weight loss.
Figure 1. A, Skin lesions on left medial thigh. B, Skin lesions on right medial thigh.
The patient had a history of poorly controlled diabetes mellitus and limited cutaneous systemic sclerosis (lcSSc; a multisystem autoimmune rheumatic disease), manifested as pulmonary hypertension and Reynaud’s syndrome. Eight months before admission, she was diagnosed with tachy-brady syndrome, requiring a pacemaker. Echocardiogram at that time showed a dilated right ventricle with elevated pressures, preservation of left ventricle ejection fraction, and no wall motion abnormalities or apical aneurysms. For management of lcSSc, she had been on long-standing azathioprine (100 mg/d). Prednisone (15 mg/d) was added 3 months before the current presentation for management of a diagnosis of retinal vasculitis.
The patient was born and raised in a rural mountain village near Trujillo, Venezuela. As a child, she lived in a mud hut and cared for many animals including chickens, dogs, and cats. At 20 years of age, she emigrated to the United States. She traveled back to Venezuela only once at 70 years of age, and otherwise denied international travel.
She was admitted to the hospital, where she was initially afebrile and hemodynamically stable. Physical exam showed no mucosal abnormalities or regional lymphadenopathy. White blood cell count was 4.4 K/μL with profound lymphopenia (absolute lymphocytes 0.2 K/μL), hemoglobin 11.5 g/dL, and platelets 126 K/μL. Kidney and liver function were normal. An HIV antigen/antibody test was negative. Strongyloides serology was negative. Computed tomography scan showed scattered bilateral pulmonary nodules; normal caliber esophagus, small bowel, and large bowel; and inflammation in the ascending colon. Contrast-enhanced brain magnetic resonance imaging was normal.
Biopsy of the skin nodules (Figure 2A, B) showed a lymphohistiocytic infiltrate in the superficial and deep dermis with round intracellular organisms noted on hematoxylin/eosin stain. Periodic acid-Schiff-diastase, Gomori methenamine silver (GMS), Fite, and gram stains did not highlight the organisms. However, structures resembling kinetoplasts were minimally accentuated on GMS stain. Bacterial, fungal, and acid-fast bacillus cultures were ultimately negative. Serology for Trypanosoma cruzi (performed at Mayo Clinic Laboratories) was positive by both enzyme-linked immunosorbent assay and a lateral flow assay. A Giemsa-stained smear of peripheral blood (buffy coat) was negative, and real-time polymerase chain reaction (PCR) testing of a whole-blood sample for T. cruzi was also negative (performed at ARUP Laboratories). Tissue obtained from the cutaneous biopsy specimen identified T. cruzi by sequencing of the internal transcribed spacer 2 and D2 region of the 28S rRNA gene (performed at Stanford Health Care [1–3]).
Figure 2. A, Biopsy of skin nodules, 200× magnification, hematoxylin and eosin (H&E) stain. Histologic sections show an epidermis with mild spongiosis and an underlying lymphohistiocytic infiltrate. B, Biopsy of skin nodules, 400× magnification, H&E stain. Histologic sections show numerous parasitized histiocytes (demonstrated by arrows). The organisms are circular without a well-defined capsule.
Given the patient’s remote exposure to an area endemic for T. cruzi, she was diagnosed with reactivation of Chagas disease (CD). Immunosuppression was weaned, and oral benznidazole 150 mg twice daily (~5 mg/kg/d) was initiated.
Concurrent with the finding of CD, the patient was also found to have disseminated cytomegalovirus (CMV) disease. Plasma CMV viral load was >1.2 million copies/mL. Lung, colon, and skin biopsies all contained cells demonstrating cytopathic changes, which were positive by CMV immunostaining. Intravenous ganciclovir was started.
With treatment, the patient’s constitutional symptoms improved, and the skin lesions became less tender and erythematous. However, 1 month after the initiation of benznidazole, the patient developed Enterobacter cloacae bacteremia and septic shock. Shortly thereafter, she died.
Chagas Disease Epidemiology and Natural History
T. cruzi is primarily transmitted to humans via inoculation of wounds or mucosal membranes with infected feces of blood-sucking triatomine insects. CD is endemic throughout much of Latin America. However, as people have migrated from endemic areas, CD has become increasingly prevalent in traditionally nonendemic regions. In the United States, it is estimated that 1.3% of Latin American immigrants are infected with T. cruzi [4].
In acute CD, nearly all infected individuals have an effective host immune response that controls the parasitemia within 1–2 months of initial infection. However, in the absence of effective antiparasitic treatment, tissue infection persists for the life of the host. Chronic indeterminate CD occurs in persons without signs or symptoms of infection, with normal electrocardiogram, and normal radiographic appearance of the chest, esophagus, and colon. Indeterminate CD may persist for life or progress to determinate CD in 20%–30% of patients. Determinate CD results from tissue destruction related to persistent parasite replication and the immune response. It manifests with cardiac disease (conduction abnormalities, arrhythmias, dilated cardiomyopathy) and, less frequently, gastrointestinal disease (esophageal/colonic dysmotility and dilatation) [5, 6]. In some immunocompromised persons, chronic (indeterminate or determinate) CD can reactivate.
In conjunction with clinical symptoms, diagnosis of CD reactivation can be aided by laboratory testing. Serologic tests will be positive in most patients infected with T. cruzi. In chronic CD, patients can have transient parasitemia detected on microscopic examination of whole blood (or, preferably, buffy coat) smears [7]. With CD reactivation, patients often have more persistent parasitemia [8]. Positive T. cruzi blood PCR assays are suggestive but not diagnostic of reactivation. However, positive blood PCR results can herald the development of subsequent invasive Chagas disease reactivation in immunocompromised patients. Furthermore, rising parasite numbers demonstrated by quantitative PCR in serial specimens are highly suggestive of reactivated disease [8].
Reactivation of Chagas Disease
CD reactivation is best described in patients with advanced Chagas cardiomyopathy who undergo orthotopic heart transplantation [9–11]. The immunosuppressive drugs used to prevent transplant rejection predispose to CD reactivation. In these patients, reactivation most commonly manifests first as asymptomatic parasitemia or acute myocarditis [12]. Reactivation can also present as subcutaneous lesions [13], panniculitis [14], or, less commonly, meningoencephalitis [15]. Case series involving other solid organ transplant recipients (mostly renal transplant recipients) describe similar manifestations of CD reactivation [16–19]. CD reactivation is also known to occur in people with HIV/AIDS, most commonly presenting as meningoencephalitis [20] and/or brain abscesses (chagomas) [3, 21]. CD reactivation has also been described in patients receiving chemotherapy for hematologic and solid malignancies [22–25] and hematopoietic cell transplant (HCT) recipients [26–29].
Reactivation of Chagas Disease in Patients With Autoimmune Rheumatic Disease
There is a paucity of data regarding CD reactivation in patients receiving immunosuppressive therapy for autoimmune rheumatic diseases (ARDs; includes conditions such as systemic lupus erythematosus [SLE], rheumatoid arthritis, dermatomyositis, mixed connective tissue disease, and scleroderma). Compared with transplant recipients, patients with AIDS, and those receiving chemotherapy, patients undergoing treatment for ARD are often less immunosuppressed. However, it is unclear whether the dearth of literature in this cohort reflects infrequent reactivation or simply a lack of published case reports.
Among published English-language case reports that describe CD reactivation in patients with ARD on immunosuppressive regimens, the minority describe symptomatic CD reactivation (Table 1). Two patients developed brain chagomas—1 patient was successfully treated [30], and the other expired shortly after diagnosis [31]. Two other patients presented with Chagas skin lesions [30, 32], 1 of whom suffered recurrent disease after a course of benznidazole [30]. Other cases describe asymptomatic reactivation detected by T. cruzi PCR [33, 34], for which 1 patient was treated with off-label posaconazole after failing therapy with benznidazole [33]. The remaining patient reports describe possible reactivation detected through T. cruzi blood PCR positivity on a single test [30, 35]; given the absence of symptoms and serial testing, it is unclear if these patients had true CD reactivation or merely intermittently detectable parasites associated with chronic CD.
Table 1. Published Studies Describing Chagas Disease Reactivation (or Possible Reactivation) in Patients With Autoimmune Disease who Were Receiving Immunosuppressive Therapy
Study Patient Autoimmune Disease Immunosuppression Country of Origin Chagas Diagnosis Known Before Evaluation for Reactivation Evidence of Chagas Reactivation (or Possible Reactivation) Treatment Outcome
Current case 86 yo F Limited cutaneous systemic sclerosis Prednisone 15 mg/d, azathioprine 100 mg/d Venezuela No Skin lesions—erythematous, indurated, painful nodules on medial thighs and upper extremity Benznidazole Patient died 1 mo after initiation of benznidazole
Kaushal et al., 2019 [31] 88 yo F RA MTX Unknown No Brain chagoma Benznidazole Patient died shortly after diagnosis
German Sanchez et al., 2019 [30] 53 yo F SLE Prednisone 5 mg/d, MMF, CP All patients in study from Argentina No Brain chagoma and T. cruzi blood PCR pos Benznidazole PCR neg
68 yo F Psoriatic arthritis Prednisone 20 mg/d, HCQ, MTX Yes Panniculitis and T. cruzi blood PCR pos Benznidazole Relapse with recurrent skin lesions 1 y s/p treatment
7 of 13 patients with ARDs who had Chagas reactivation (or possible reactivation) 48 yo F RA MTX, adalimumab Yes Single T. Cruz blood PCR pos Benznidazole PCR neg
66 yo M RA Prednisone 5 mg/d, MTX, etanercept Yes Single T. Cruzi blood PCR pos Benznidazole PCR neg
Treatment durations for all patients were 1–2 mo 81 yo F RA Prednisone 5 mg/d, HCQ, MTX, leflunomide Yes Single T. Cruzi blood PCR pos Nifurtimox PCR neg
66 yo F Sjogren syndrome Prednisone 5 mg/d, HCQ Yes Single T. Cruzi blood PCR pos Benznidazole PCR neg
57 yo F Vasculitis Prednisone 5 mg/d, CP No Single T. cruzi blood PCR pos Benznidazole PCR neg
Vacas et al., 2017 [35] 57 yo M Psoriatic erythroderma Infliximab Argentina Yes Single T. cruzi blood PCR pos Benznidazole × 45 d PCR neg
Navarrete-Dechent et al., 2015 [34] 52 yo M Psoriasis Adalimumab Chile Yes—Chagas megacolon, received preemptive treatment with nifurtimox
T. cruzi blood PCR pos 8 months following preemptive nifurtimox Repeat course nifurtimox × 2 mo PCR neg
Burgos et al., 2012 [32] 44 yo F SLE Prednisone 50 mg/d, azathioprine 50 mg/d Paraguay No Parasitemia, skin lesions—erythematous, painful nodules that progressed to ulcer and eschar Benznidazole × 2 mo Clinically improved
Pinazo et al., 2010 [33] 44 yo F SLE Steroids, CP Argentina Yes—chronic indeterminate Chagas
T. cruzi blood PCR pos, treated with benznidazole with recurrent PCR pos Benznidazole × 2 mo, and then posaconazole × 3 mo for relapse Serial PCR neg after posaconazole
Abbreviations: CP, cyclophosphamide; HCQ, hydroxychloroquine; MMF, mycophenolate mofetil; MTX, methotrexate; NA, not applicable; neg, negative; PCR, polymerase chain reaction; pos, positive; RA, rheumatoid arthritis; SLE, systemic lupus erythematosus.
Although limited by small sample size and abbreviated longitudinal follow-up, other studies have attempted to investigate the risk for patients with ARD developing CD reactivation while on immunosuppression. One such study described 2 of 13 patients with ARD who developed symptomatic reactivation on immunosuppressive therapies over a 2-year period, and another 5 patients who developed possible CD reactivation detected by a single positive T. cruzi blood PCR test (Table 1) [30]. In other case series, the majority of immunosuppressed patients with ARD and CD were treated for chronic CD before the development of any evidence of CD reactivation, and only a small minority of these patients subsequently developed CD reactivation while receiving immunosuppression [25, 36, 37]. In 1 such study, 6 of 8 patients with ARD and CD were treated in this manner with varying regimens of benznidazole and nifurtimox. Three patients were treated for CD before a diagnosis of an ARD was made. Only 1 of the patients who received such treatment subsequently developed CD reactivation while receiving immunosuppression (further patient details not specified) [36]. In another study, 11 of 14 patients with chronic CD and ARD were treated for chronic CD with benznidazole for 60 days at the time of study enrollment, and none of the 14 patients experienced CD reactivation while receiving immunosuppression during follow-up [25]. Lastly, 3 patients with ARD and chronic CD were all treated at the time of CD diagnosis with benznidazole for 60 days; none of these patients developed subsequent CD reactivation during a 36-month follow-up period [37]. Though these preliminary data are intriguing, larger population studies, conducted over longer durations, are needed in order to draw more substantial conclusions about factors predisposing to CD reactivation in this cohort and the outcomes of prophylactic therapy.
DISCUSSION
Our patient is among the few in whom symptomatic CD reactivation was documented in the setting of immunosuppressive therapy for an ARD. Our patient likely had unrecognized determinate CD, clinically manifested by tachy-brady syndrome requiring a pacemaker [38]. CD reactivated after intensification of her immunosuppression regimen, ultimately manifesting as disseminated skin nodules, with sequencing of the skin biopsy revealing T. cruzi.
Our patient’s case was unique in that symptomatic CD reactivation occurred in the setting of a negative T. cruzi blood PCR. Typically, T. cruzi blood PCR positivity develops before symptomatic reactivation [11]. Though it is unclear why our patient’s PCR was negative, it is possibly related to blood volume collected or PCR test characteristics. Importantly, this demonstrates that PCR positivity should not be the sole diagnostic measure in the evaluation for CD reactivation; assessment must also include careful clinical evaluation.
CD reactivation in patients being treated for ARD additionally involves a different degree of immunosuppression compared with other better-studied cohorts. An effective host response against T. cruzi requires both cellular and humoral immunity [39]. Our patient was taking low-dose prednisone and azathioprine. Corticosteroids are nonspecific immune function inhibitors [40]. However, there are no reports that describe an increased incidence of CD reactivation in patients receiving corticosteroids as their sole form of immunosuppression. Azathioprine is an antimetabolite that decreases both T and B lymphocyte production [40]. However, some heart transplant recipients with a history of Chagas cardiomyopathy preferentially receive azathioprine over mycophenolate due to a 6-fold lower incidence of CD reactivation with azathioprine compared with mycophenolate [41]. Taken together, prednisone in doses <20 mg/d and azathioprine still seem to convey a relatively low risk for CD reactivation.
In our patient’s case, it is possible that more multifaceted immunosuppression led to her disease. Specifically, the synergistic effect of prednisone and azathioprine, in conjunction with her profound lymphopenia, advanced age, and uncontrolled diabetes, may have contributed to CD reactivation. However, these immunosuppressing factors still do not clearly explain our patient’s profound functional immunosuppression, further exhibited by concurrent disseminated CMV disease. Based on prior data [42–45], patients with rheumatoid arthritis and SLE may have an increased risk of infection independent of immunomodulatory therapy. Further research might better delineate the relationship between CD and immune function.
Screening for Chagas Disease in Patients With ARD
Currently, consensus guidelines recommend serologic screening for CD in transplant donors and recipients with epidemiologic risk factors. A positive serologic result should be confirmed by at least 2 distinct serologic methods. In the United States, CD treatment based solely on a positive serology result in either the donor or the recipient is generally not recommended given the toxicity of the therapeutic options [46, 47]. Alternatively, transplant recipients who are seropositive should be monitored for reactivation, especially during the times of most intense immunosuppression. Laboratory monitoring employs microscopy of blood/buffy coats and blood PCR. If monitoring reveals parasitemia and/or PCR positivity (especially increasing the parasite load on serial quantitative PCRs), patients are typically given preemptive CD treatment, as the development of detectable T. cruzi in this cohort often heralds the development of symptomatic reactivation [46–49].
Given that data regarding CD reactivation in patients with ARD are so scant, it is uncertain if similar screening and treatment guidelines should be applied to ARD patients in the face of immunosuppression. However, in light of our patient’s case and our review of the literature, our opinion is that strong consideration should be given to serologic screening for T. cruzi before immunosuppression for ARD in patients who have CD risk factors. (Note that serologic screening following immunosuppression may be falsely negative due to a blunted immune response.) Data are insufficient to comment on the risks versus benefits of CD treatment based solely on a positive serologic result in this cohort. However, similar to transplant recipients, for those who are seropositive, clinical assessment and serial blood microscopy/PCR monitoring should be employed during immunosuppressive therapy. There should be strong consideration of preemptive treatment of patients with parasitemia and/or PCR blood positivity (especially increasing parasite load on serial quantitative PCRs) even in the absence of symptoms; of course, evaluation/treatment for those with symptoms/signs concerning for CD reactivation. More research is needed to refine the screening and subsequent treatment approach for CD in patients with ARD.
CONCLUSIONS
This report describes CD reactivation in a patient with an ARD receiving immunosuppressive therapy. It highlights the need for more research regarding CD and reactivation in this patient population. It additionally suggests the need for broader serologic screening of patients with risk factors for CD before starting immunosuppression, and subsequent monitoring of at-risk patients while they receive such therapy.
Acknowledgments
Financial support. None
Potential conflicts of interest. All authors: no reported conflicts of interest. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed.
Disclaimer. The findings and conclusions in this report are those of the authors and do not necessarily represent the views of the Centers for Disease Control and Prevention.
Patient consent. Our study does not include factors necessitating patient consent. | Recovering | ReactionOutcome | CC BY-NC-ND | 33575423 | 19,494,598 | 2021-02 |
What was the outcome of reaction 'Enterobacter infection'? | Reactivation of Chagas Disease in a Patient With an Autoimmune Rheumatic Disease: Case Report and Review of the Literature.
Reactivation of Chagas disease has been described in immunosuppressed patients, but there is a paucity of literature describing reactivation in patients on immunosuppressive therapies for the treatment of autoimmune rheumatic diseases. We describe a case of Chagas disease reactivation in a woman taking azathioprine and prednisone for limited cutaneous systemic sclerosis (lcSSc). Reactivation manifested as indurated and erythematous cutaneous nodules. Sequencing of a skin biopsy specimen confirmed the diagnosis of Chagas disease. She was treated with benznidazole with clinical improvement in the cutaneous lesions. However, her clinical course was complicated and included disseminated CMV disease and subsequent septic shock due to bacteremia. Our case and review of the literature highlight that screening for Chagas disease should be strongly considered for patients who will undergo immunosuppression for treatment of autoimmune disease if epidemiologically indicated.
PATIENT CASE
An 86-year-old woman developed painful subcutaneous nodules on her medial thighs and left upper extremity (Figure 1A, B). Over the ensuing 2 months, the nodules enlarged and became more indurated, erythematous, and painful. She also developed night sweats, fatigue, diarrhea, anorexia, and weight loss.
Figure 1. A, Skin lesions on left medial thigh. B, Skin lesions on right medial thigh.
The patient had a history of poorly controlled diabetes mellitus and limited cutaneous systemic sclerosis (lcSSc; a multisystem autoimmune rheumatic disease), manifested as pulmonary hypertension and Reynaud’s syndrome. Eight months before admission, she was diagnosed with tachy-brady syndrome, requiring a pacemaker. Echocardiogram at that time showed a dilated right ventricle with elevated pressures, preservation of left ventricle ejection fraction, and no wall motion abnormalities or apical aneurysms. For management of lcSSc, she had been on long-standing azathioprine (100 mg/d). Prednisone (15 mg/d) was added 3 months before the current presentation for management of a diagnosis of retinal vasculitis.
The patient was born and raised in a rural mountain village near Trujillo, Venezuela. As a child, she lived in a mud hut and cared for many animals including chickens, dogs, and cats. At 20 years of age, she emigrated to the United States. She traveled back to Venezuela only once at 70 years of age, and otherwise denied international travel.
She was admitted to the hospital, where she was initially afebrile and hemodynamically stable. Physical exam showed no mucosal abnormalities or regional lymphadenopathy. White blood cell count was 4.4 K/μL with profound lymphopenia (absolute lymphocytes 0.2 K/μL), hemoglobin 11.5 g/dL, and platelets 126 K/μL. Kidney and liver function were normal. An HIV antigen/antibody test was negative. Strongyloides serology was negative. Computed tomography scan showed scattered bilateral pulmonary nodules; normal caliber esophagus, small bowel, and large bowel; and inflammation in the ascending colon. Contrast-enhanced brain magnetic resonance imaging was normal.
Biopsy of the skin nodules (Figure 2A, B) showed a lymphohistiocytic infiltrate in the superficial and deep dermis with round intracellular organisms noted on hematoxylin/eosin stain. Periodic acid-Schiff-diastase, Gomori methenamine silver (GMS), Fite, and gram stains did not highlight the organisms. However, structures resembling kinetoplasts were minimally accentuated on GMS stain. Bacterial, fungal, and acid-fast bacillus cultures were ultimately negative. Serology for Trypanosoma cruzi (performed at Mayo Clinic Laboratories) was positive by both enzyme-linked immunosorbent assay and a lateral flow assay. A Giemsa-stained smear of peripheral blood (buffy coat) was negative, and real-time polymerase chain reaction (PCR) testing of a whole-blood sample for T. cruzi was also negative (performed at ARUP Laboratories). Tissue obtained from the cutaneous biopsy specimen identified T. cruzi by sequencing of the internal transcribed spacer 2 and D2 region of the 28S rRNA gene (performed at Stanford Health Care [1–3]).
Figure 2. A, Biopsy of skin nodules, 200× magnification, hematoxylin and eosin (H&E) stain. Histologic sections show an epidermis with mild spongiosis and an underlying lymphohistiocytic infiltrate. B, Biopsy of skin nodules, 400× magnification, H&E stain. Histologic sections show numerous parasitized histiocytes (demonstrated by arrows). The organisms are circular without a well-defined capsule.
Given the patient’s remote exposure to an area endemic for T. cruzi, she was diagnosed with reactivation of Chagas disease (CD). Immunosuppression was weaned, and oral benznidazole 150 mg twice daily (~5 mg/kg/d) was initiated.
Concurrent with the finding of CD, the patient was also found to have disseminated cytomegalovirus (CMV) disease. Plasma CMV viral load was >1.2 million copies/mL. Lung, colon, and skin biopsies all contained cells demonstrating cytopathic changes, which were positive by CMV immunostaining. Intravenous ganciclovir was started.
With treatment, the patient’s constitutional symptoms improved, and the skin lesions became less tender and erythematous. However, 1 month after the initiation of benznidazole, the patient developed Enterobacter cloacae bacteremia and septic shock. Shortly thereafter, she died.
Chagas Disease Epidemiology and Natural History
T. cruzi is primarily transmitted to humans via inoculation of wounds or mucosal membranes with infected feces of blood-sucking triatomine insects. CD is endemic throughout much of Latin America. However, as people have migrated from endemic areas, CD has become increasingly prevalent in traditionally nonendemic regions. In the United States, it is estimated that 1.3% of Latin American immigrants are infected with T. cruzi [4].
In acute CD, nearly all infected individuals have an effective host immune response that controls the parasitemia within 1–2 months of initial infection. However, in the absence of effective antiparasitic treatment, tissue infection persists for the life of the host. Chronic indeterminate CD occurs in persons without signs or symptoms of infection, with normal electrocardiogram, and normal radiographic appearance of the chest, esophagus, and colon. Indeterminate CD may persist for life or progress to determinate CD in 20%–30% of patients. Determinate CD results from tissue destruction related to persistent parasite replication and the immune response. It manifests with cardiac disease (conduction abnormalities, arrhythmias, dilated cardiomyopathy) and, less frequently, gastrointestinal disease (esophageal/colonic dysmotility and dilatation) [5, 6]. In some immunocompromised persons, chronic (indeterminate or determinate) CD can reactivate.
In conjunction with clinical symptoms, diagnosis of CD reactivation can be aided by laboratory testing. Serologic tests will be positive in most patients infected with T. cruzi. In chronic CD, patients can have transient parasitemia detected on microscopic examination of whole blood (or, preferably, buffy coat) smears [7]. With CD reactivation, patients often have more persistent parasitemia [8]. Positive T. cruzi blood PCR assays are suggestive but not diagnostic of reactivation. However, positive blood PCR results can herald the development of subsequent invasive Chagas disease reactivation in immunocompromised patients. Furthermore, rising parasite numbers demonstrated by quantitative PCR in serial specimens are highly suggestive of reactivated disease [8].
Reactivation of Chagas Disease
CD reactivation is best described in patients with advanced Chagas cardiomyopathy who undergo orthotopic heart transplantation [9–11]. The immunosuppressive drugs used to prevent transplant rejection predispose to CD reactivation. In these patients, reactivation most commonly manifests first as asymptomatic parasitemia or acute myocarditis [12]. Reactivation can also present as subcutaneous lesions [13], panniculitis [14], or, less commonly, meningoencephalitis [15]. Case series involving other solid organ transplant recipients (mostly renal transplant recipients) describe similar manifestations of CD reactivation [16–19]. CD reactivation is also known to occur in people with HIV/AIDS, most commonly presenting as meningoencephalitis [20] and/or brain abscesses (chagomas) [3, 21]. CD reactivation has also been described in patients receiving chemotherapy for hematologic and solid malignancies [22–25] and hematopoietic cell transplant (HCT) recipients [26–29].
Reactivation of Chagas Disease in Patients With Autoimmune Rheumatic Disease
There is a paucity of data regarding CD reactivation in patients receiving immunosuppressive therapy for autoimmune rheumatic diseases (ARDs; includes conditions such as systemic lupus erythematosus [SLE], rheumatoid arthritis, dermatomyositis, mixed connective tissue disease, and scleroderma). Compared with transplant recipients, patients with AIDS, and those receiving chemotherapy, patients undergoing treatment for ARD are often less immunosuppressed. However, it is unclear whether the dearth of literature in this cohort reflects infrequent reactivation or simply a lack of published case reports.
Among published English-language case reports that describe CD reactivation in patients with ARD on immunosuppressive regimens, the minority describe symptomatic CD reactivation (Table 1). Two patients developed brain chagomas—1 patient was successfully treated [30], and the other expired shortly after diagnosis [31]. Two other patients presented with Chagas skin lesions [30, 32], 1 of whom suffered recurrent disease after a course of benznidazole [30]. Other cases describe asymptomatic reactivation detected by T. cruzi PCR [33, 34], for which 1 patient was treated with off-label posaconazole after failing therapy with benznidazole [33]. The remaining patient reports describe possible reactivation detected through T. cruzi blood PCR positivity on a single test [30, 35]; given the absence of symptoms and serial testing, it is unclear if these patients had true CD reactivation or merely intermittently detectable parasites associated with chronic CD.
Table 1. Published Studies Describing Chagas Disease Reactivation (or Possible Reactivation) in Patients With Autoimmune Disease who Were Receiving Immunosuppressive Therapy
Study Patient Autoimmune Disease Immunosuppression Country of Origin Chagas Diagnosis Known Before Evaluation for Reactivation Evidence of Chagas Reactivation (or Possible Reactivation) Treatment Outcome
Current case 86 yo F Limited cutaneous systemic sclerosis Prednisone 15 mg/d, azathioprine 100 mg/d Venezuela No Skin lesions—erythematous, indurated, painful nodules on medial thighs and upper extremity Benznidazole Patient died 1 mo after initiation of benznidazole
Kaushal et al., 2019 [31] 88 yo F RA MTX Unknown No Brain chagoma Benznidazole Patient died shortly after diagnosis
German Sanchez et al., 2019 [30] 53 yo F SLE Prednisone 5 mg/d, MMF, CP All patients in study from Argentina No Brain chagoma and T. cruzi blood PCR pos Benznidazole PCR neg
68 yo F Psoriatic arthritis Prednisone 20 mg/d, HCQ, MTX Yes Panniculitis and T. cruzi blood PCR pos Benznidazole Relapse with recurrent skin lesions 1 y s/p treatment
7 of 13 patients with ARDs who had Chagas reactivation (or possible reactivation) 48 yo F RA MTX, adalimumab Yes Single T. Cruz blood PCR pos Benznidazole PCR neg
66 yo M RA Prednisone 5 mg/d, MTX, etanercept Yes Single T. Cruzi blood PCR pos Benznidazole PCR neg
Treatment durations for all patients were 1–2 mo 81 yo F RA Prednisone 5 mg/d, HCQ, MTX, leflunomide Yes Single T. Cruzi blood PCR pos Nifurtimox PCR neg
66 yo F Sjogren syndrome Prednisone 5 mg/d, HCQ Yes Single T. Cruzi blood PCR pos Benznidazole PCR neg
57 yo F Vasculitis Prednisone 5 mg/d, CP No Single T. cruzi blood PCR pos Benznidazole PCR neg
Vacas et al., 2017 [35] 57 yo M Psoriatic erythroderma Infliximab Argentina Yes Single T. cruzi blood PCR pos Benznidazole × 45 d PCR neg
Navarrete-Dechent et al., 2015 [34] 52 yo M Psoriasis Adalimumab Chile Yes—Chagas megacolon, received preemptive treatment with nifurtimox
T. cruzi blood PCR pos 8 months following preemptive nifurtimox Repeat course nifurtimox × 2 mo PCR neg
Burgos et al., 2012 [32] 44 yo F SLE Prednisone 50 mg/d, azathioprine 50 mg/d Paraguay No Parasitemia, skin lesions—erythematous, painful nodules that progressed to ulcer and eschar Benznidazole × 2 mo Clinically improved
Pinazo et al., 2010 [33] 44 yo F SLE Steroids, CP Argentina Yes—chronic indeterminate Chagas
T. cruzi blood PCR pos, treated with benznidazole with recurrent PCR pos Benznidazole × 2 mo, and then posaconazole × 3 mo for relapse Serial PCR neg after posaconazole
Abbreviations: CP, cyclophosphamide; HCQ, hydroxychloroquine; MMF, mycophenolate mofetil; MTX, methotrexate; NA, not applicable; neg, negative; PCR, polymerase chain reaction; pos, positive; RA, rheumatoid arthritis; SLE, systemic lupus erythematosus.
Although limited by small sample size and abbreviated longitudinal follow-up, other studies have attempted to investigate the risk for patients with ARD developing CD reactivation while on immunosuppression. One such study described 2 of 13 patients with ARD who developed symptomatic reactivation on immunosuppressive therapies over a 2-year period, and another 5 patients who developed possible CD reactivation detected by a single positive T. cruzi blood PCR test (Table 1) [30]. In other case series, the majority of immunosuppressed patients with ARD and CD were treated for chronic CD before the development of any evidence of CD reactivation, and only a small minority of these patients subsequently developed CD reactivation while receiving immunosuppression [25, 36, 37]. In 1 such study, 6 of 8 patients with ARD and CD were treated in this manner with varying regimens of benznidazole and nifurtimox. Three patients were treated for CD before a diagnosis of an ARD was made. Only 1 of the patients who received such treatment subsequently developed CD reactivation while receiving immunosuppression (further patient details not specified) [36]. In another study, 11 of 14 patients with chronic CD and ARD were treated for chronic CD with benznidazole for 60 days at the time of study enrollment, and none of the 14 patients experienced CD reactivation while receiving immunosuppression during follow-up [25]. Lastly, 3 patients with ARD and chronic CD were all treated at the time of CD diagnosis with benznidazole for 60 days; none of these patients developed subsequent CD reactivation during a 36-month follow-up period [37]. Though these preliminary data are intriguing, larger population studies, conducted over longer durations, are needed in order to draw more substantial conclusions about factors predisposing to CD reactivation in this cohort and the outcomes of prophylactic therapy.
DISCUSSION
Our patient is among the few in whom symptomatic CD reactivation was documented in the setting of immunosuppressive therapy for an ARD. Our patient likely had unrecognized determinate CD, clinically manifested by tachy-brady syndrome requiring a pacemaker [38]. CD reactivated after intensification of her immunosuppression regimen, ultimately manifesting as disseminated skin nodules, with sequencing of the skin biopsy revealing T. cruzi.
Our patient’s case was unique in that symptomatic CD reactivation occurred in the setting of a negative T. cruzi blood PCR. Typically, T. cruzi blood PCR positivity develops before symptomatic reactivation [11]. Though it is unclear why our patient’s PCR was negative, it is possibly related to blood volume collected or PCR test characteristics. Importantly, this demonstrates that PCR positivity should not be the sole diagnostic measure in the evaluation for CD reactivation; assessment must also include careful clinical evaluation.
CD reactivation in patients being treated for ARD additionally involves a different degree of immunosuppression compared with other better-studied cohorts. An effective host response against T. cruzi requires both cellular and humoral immunity [39]. Our patient was taking low-dose prednisone and azathioprine. Corticosteroids are nonspecific immune function inhibitors [40]. However, there are no reports that describe an increased incidence of CD reactivation in patients receiving corticosteroids as their sole form of immunosuppression. Azathioprine is an antimetabolite that decreases both T and B lymphocyte production [40]. However, some heart transplant recipients with a history of Chagas cardiomyopathy preferentially receive azathioprine over mycophenolate due to a 6-fold lower incidence of CD reactivation with azathioprine compared with mycophenolate [41]. Taken together, prednisone in doses <20 mg/d and azathioprine still seem to convey a relatively low risk for CD reactivation.
In our patient’s case, it is possible that more multifaceted immunosuppression led to her disease. Specifically, the synergistic effect of prednisone and azathioprine, in conjunction with her profound lymphopenia, advanced age, and uncontrolled diabetes, may have contributed to CD reactivation. However, these immunosuppressing factors still do not clearly explain our patient’s profound functional immunosuppression, further exhibited by concurrent disseminated CMV disease. Based on prior data [42–45], patients with rheumatoid arthritis and SLE may have an increased risk of infection independent of immunomodulatory therapy. Further research might better delineate the relationship between CD and immune function.
Screening for Chagas Disease in Patients With ARD
Currently, consensus guidelines recommend serologic screening for CD in transplant donors and recipients with epidemiologic risk factors. A positive serologic result should be confirmed by at least 2 distinct serologic methods. In the United States, CD treatment based solely on a positive serology result in either the donor or the recipient is generally not recommended given the toxicity of the therapeutic options [46, 47]. Alternatively, transplant recipients who are seropositive should be monitored for reactivation, especially during the times of most intense immunosuppression. Laboratory monitoring employs microscopy of blood/buffy coats and blood PCR. If monitoring reveals parasitemia and/or PCR positivity (especially increasing the parasite load on serial quantitative PCRs), patients are typically given preemptive CD treatment, as the development of detectable T. cruzi in this cohort often heralds the development of symptomatic reactivation [46–49].
Given that data regarding CD reactivation in patients with ARD are so scant, it is uncertain if similar screening and treatment guidelines should be applied to ARD patients in the face of immunosuppression. However, in light of our patient’s case and our review of the literature, our opinion is that strong consideration should be given to serologic screening for T. cruzi before immunosuppression for ARD in patients who have CD risk factors. (Note that serologic screening following immunosuppression may be falsely negative due to a blunted immune response.) Data are insufficient to comment on the risks versus benefits of CD treatment based solely on a positive serologic result in this cohort. However, similar to transplant recipients, for those who are seropositive, clinical assessment and serial blood microscopy/PCR monitoring should be employed during immunosuppressive therapy. There should be strong consideration of preemptive treatment of patients with parasitemia and/or PCR blood positivity (especially increasing parasite load on serial quantitative PCRs) even in the absence of symptoms; of course, evaluation/treatment for those with symptoms/signs concerning for CD reactivation. More research is needed to refine the screening and subsequent treatment approach for CD in patients with ARD.
CONCLUSIONS
This report describes CD reactivation in a patient with an ARD receiving immunosuppressive therapy. It highlights the need for more research regarding CD and reactivation in this patient population. It additionally suggests the need for broader serologic screening of patients with risk factors for CD before starting immunosuppression, and subsequent monitoring of at-risk patients while they receive such therapy.
Acknowledgments
Financial support. None
Potential conflicts of interest. All authors: no reported conflicts of interest. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed.
Disclaimer. The findings and conclusions in this report are those of the authors and do not necessarily represent the views of the Centers for Disease Control and Prevention.
Patient consent. Our study does not include factors necessitating patient consent. | Fatal | ReactionOutcome | CC BY-NC-ND | 33575423 | 19,494,598 | 2021-02 |
What was the outcome of reaction 'Septic shock'? | Reactivation of Chagas Disease in a Patient With an Autoimmune Rheumatic Disease: Case Report and Review of the Literature.
Reactivation of Chagas disease has been described in immunosuppressed patients, but there is a paucity of literature describing reactivation in patients on immunosuppressive therapies for the treatment of autoimmune rheumatic diseases. We describe a case of Chagas disease reactivation in a woman taking azathioprine and prednisone for limited cutaneous systemic sclerosis (lcSSc). Reactivation manifested as indurated and erythematous cutaneous nodules. Sequencing of a skin biopsy specimen confirmed the diagnosis of Chagas disease. She was treated with benznidazole with clinical improvement in the cutaneous lesions. However, her clinical course was complicated and included disseminated CMV disease and subsequent septic shock due to bacteremia. Our case and review of the literature highlight that screening for Chagas disease should be strongly considered for patients who will undergo immunosuppression for treatment of autoimmune disease if epidemiologically indicated.
PATIENT CASE
An 86-year-old woman developed painful subcutaneous nodules on her medial thighs and left upper extremity (Figure 1A, B). Over the ensuing 2 months, the nodules enlarged and became more indurated, erythematous, and painful. She also developed night sweats, fatigue, diarrhea, anorexia, and weight loss.
Figure 1. A, Skin lesions on left medial thigh. B, Skin lesions on right medial thigh.
The patient had a history of poorly controlled diabetes mellitus and limited cutaneous systemic sclerosis (lcSSc; a multisystem autoimmune rheumatic disease), manifested as pulmonary hypertension and Reynaud’s syndrome. Eight months before admission, she was diagnosed with tachy-brady syndrome, requiring a pacemaker. Echocardiogram at that time showed a dilated right ventricle with elevated pressures, preservation of left ventricle ejection fraction, and no wall motion abnormalities or apical aneurysms. For management of lcSSc, she had been on long-standing azathioprine (100 mg/d). Prednisone (15 mg/d) was added 3 months before the current presentation for management of a diagnosis of retinal vasculitis.
The patient was born and raised in a rural mountain village near Trujillo, Venezuela. As a child, she lived in a mud hut and cared for many animals including chickens, dogs, and cats. At 20 years of age, she emigrated to the United States. She traveled back to Venezuela only once at 70 years of age, and otherwise denied international travel.
She was admitted to the hospital, where she was initially afebrile and hemodynamically stable. Physical exam showed no mucosal abnormalities or regional lymphadenopathy. White blood cell count was 4.4 K/μL with profound lymphopenia (absolute lymphocytes 0.2 K/μL), hemoglobin 11.5 g/dL, and platelets 126 K/μL. Kidney and liver function were normal. An HIV antigen/antibody test was negative. Strongyloides serology was negative. Computed tomography scan showed scattered bilateral pulmonary nodules; normal caliber esophagus, small bowel, and large bowel; and inflammation in the ascending colon. Contrast-enhanced brain magnetic resonance imaging was normal.
Biopsy of the skin nodules (Figure 2A, B) showed a lymphohistiocytic infiltrate in the superficial and deep dermis with round intracellular organisms noted on hematoxylin/eosin stain. Periodic acid-Schiff-diastase, Gomori methenamine silver (GMS), Fite, and gram stains did not highlight the organisms. However, structures resembling kinetoplasts were minimally accentuated on GMS stain. Bacterial, fungal, and acid-fast bacillus cultures were ultimately negative. Serology for Trypanosoma cruzi (performed at Mayo Clinic Laboratories) was positive by both enzyme-linked immunosorbent assay and a lateral flow assay. A Giemsa-stained smear of peripheral blood (buffy coat) was negative, and real-time polymerase chain reaction (PCR) testing of a whole-blood sample for T. cruzi was also negative (performed at ARUP Laboratories). Tissue obtained from the cutaneous biopsy specimen identified T. cruzi by sequencing of the internal transcribed spacer 2 and D2 region of the 28S rRNA gene (performed at Stanford Health Care [1–3]).
Figure 2. A, Biopsy of skin nodules, 200× magnification, hematoxylin and eosin (H&E) stain. Histologic sections show an epidermis with mild spongiosis and an underlying lymphohistiocytic infiltrate. B, Biopsy of skin nodules, 400× magnification, H&E stain. Histologic sections show numerous parasitized histiocytes (demonstrated by arrows). The organisms are circular without a well-defined capsule.
Given the patient’s remote exposure to an area endemic for T. cruzi, she was diagnosed with reactivation of Chagas disease (CD). Immunosuppression was weaned, and oral benznidazole 150 mg twice daily (~5 mg/kg/d) was initiated.
Concurrent with the finding of CD, the patient was also found to have disseminated cytomegalovirus (CMV) disease. Plasma CMV viral load was >1.2 million copies/mL. Lung, colon, and skin biopsies all contained cells demonstrating cytopathic changes, which were positive by CMV immunostaining. Intravenous ganciclovir was started.
With treatment, the patient’s constitutional symptoms improved, and the skin lesions became less tender and erythematous. However, 1 month after the initiation of benznidazole, the patient developed Enterobacter cloacae bacteremia and septic shock. Shortly thereafter, she died.
Chagas Disease Epidemiology and Natural History
T. cruzi is primarily transmitted to humans via inoculation of wounds or mucosal membranes with infected feces of blood-sucking triatomine insects. CD is endemic throughout much of Latin America. However, as people have migrated from endemic areas, CD has become increasingly prevalent in traditionally nonendemic regions. In the United States, it is estimated that 1.3% of Latin American immigrants are infected with T. cruzi [4].
In acute CD, nearly all infected individuals have an effective host immune response that controls the parasitemia within 1–2 months of initial infection. However, in the absence of effective antiparasitic treatment, tissue infection persists for the life of the host. Chronic indeterminate CD occurs in persons without signs or symptoms of infection, with normal electrocardiogram, and normal radiographic appearance of the chest, esophagus, and colon. Indeterminate CD may persist for life or progress to determinate CD in 20%–30% of patients. Determinate CD results from tissue destruction related to persistent parasite replication and the immune response. It manifests with cardiac disease (conduction abnormalities, arrhythmias, dilated cardiomyopathy) and, less frequently, gastrointestinal disease (esophageal/colonic dysmotility and dilatation) [5, 6]. In some immunocompromised persons, chronic (indeterminate or determinate) CD can reactivate.
In conjunction with clinical symptoms, diagnosis of CD reactivation can be aided by laboratory testing. Serologic tests will be positive in most patients infected with T. cruzi. In chronic CD, patients can have transient parasitemia detected on microscopic examination of whole blood (or, preferably, buffy coat) smears [7]. With CD reactivation, patients often have more persistent parasitemia [8]. Positive T. cruzi blood PCR assays are suggestive but not diagnostic of reactivation. However, positive blood PCR results can herald the development of subsequent invasive Chagas disease reactivation in immunocompromised patients. Furthermore, rising parasite numbers demonstrated by quantitative PCR in serial specimens are highly suggestive of reactivated disease [8].
Reactivation of Chagas Disease
CD reactivation is best described in patients with advanced Chagas cardiomyopathy who undergo orthotopic heart transplantation [9–11]. The immunosuppressive drugs used to prevent transplant rejection predispose to CD reactivation. In these patients, reactivation most commonly manifests first as asymptomatic parasitemia or acute myocarditis [12]. Reactivation can also present as subcutaneous lesions [13], panniculitis [14], or, less commonly, meningoencephalitis [15]. Case series involving other solid organ transplant recipients (mostly renal transplant recipients) describe similar manifestations of CD reactivation [16–19]. CD reactivation is also known to occur in people with HIV/AIDS, most commonly presenting as meningoencephalitis [20] and/or brain abscesses (chagomas) [3, 21]. CD reactivation has also been described in patients receiving chemotherapy for hematologic and solid malignancies [22–25] and hematopoietic cell transplant (HCT) recipients [26–29].
Reactivation of Chagas Disease in Patients With Autoimmune Rheumatic Disease
There is a paucity of data regarding CD reactivation in patients receiving immunosuppressive therapy for autoimmune rheumatic diseases (ARDs; includes conditions such as systemic lupus erythematosus [SLE], rheumatoid arthritis, dermatomyositis, mixed connective tissue disease, and scleroderma). Compared with transplant recipients, patients with AIDS, and those receiving chemotherapy, patients undergoing treatment for ARD are often less immunosuppressed. However, it is unclear whether the dearth of literature in this cohort reflects infrequent reactivation or simply a lack of published case reports.
Among published English-language case reports that describe CD reactivation in patients with ARD on immunosuppressive regimens, the minority describe symptomatic CD reactivation (Table 1). Two patients developed brain chagomas—1 patient was successfully treated [30], and the other expired shortly after diagnosis [31]. Two other patients presented with Chagas skin lesions [30, 32], 1 of whom suffered recurrent disease after a course of benznidazole [30]. Other cases describe asymptomatic reactivation detected by T. cruzi PCR [33, 34], for which 1 patient was treated with off-label posaconazole after failing therapy with benznidazole [33]. The remaining patient reports describe possible reactivation detected through T. cruzi blood PCR positivity on a single test [30, 35]; given the absence of symptoms and serial testing, it is unclear if these patients had true CD reactivation or merely intermittently detectable parasites associated with chronic CD.
Table 1. Published Studies Describing Chagas Disease Reactivation (or Possible Reactivation) in Patients With Autoimmune Disease who Were Receiving Immunosuppressive Therapy
Study Patient Autoimmune Disease Immunosuppression Country of Origin Chagas Diagnosis Known Before Evaluation for Reactivation Evidence of Chagas Reactivation (or Possible Reactivation) Treatment Outcome
Current case 86 yo F Limited cutaneous systemic sclerosis Prednisone 15 mg/d, azathioprine 100 mg/d Venezuela No Skin lesions—erythematous, indurated, painful nodules on medial thighs and upper extremity Benznidazole Patient died 1 mo after initiation of benznidazole
Kaushal et al., 2019 [31] 88 yo F RA MTX Unknown No Brain chagoma Benznidazole Patient died shortly after diagnosis
German Sanchez et al., 2019 [30] 53 yo F SLE Prednisone 5 mg/d, MMF, CP All patients in study from Argentina No Brain chagoma and T. cruzi blood PCR pos Benznidazole PCR neg
68 yo F Psoriatic arthritis Prednisone 20 mg/d, HCQ, MTX Yes Panniculitis and T. cruzi blood PCR pos Benznidazole Relapse with recurrent skin lesions 1 y s/p treatment
7 of 13 patients with ARDs who had Chagas reactivation (or possible reactivation) 48 yo F RA MTX, adalimumab Yes Single T. Cruz blood PCR pos Benznidazole PCR neg
66 yo M RA Prednisone 5 mg/d, MTX, etanercept Yes Single T. Cruzi blood PCR pos Benznidazole PCR neg
Treatment durations for all patients were 1–2 mo 81 yo F RA Prednisone 5 mg/d, HCQ, MTX, leflunomide Yes Single T. Cruzi blood PCR pos Nifurtimox PCR neg
66 yo F Sjogren syndrome Prednisone 5 mg/d, HCQ Yes Single T. Cruzi blood PCR pos Benznidazole PCR neg
57 yo F Vasculitis Prednisone 5 mg/d, CP No Single T. cruzi blood PCR pos Benznidazole PCR neg
Vacas et al., 2017 [35] 57 yo M Psoriatic erythroderma Infliximab Argentina Yes Single T. cruzi blood PCR pos Benznidazole × 45 d PCR neg
Navarrete-Dechent et al., 2015 [34] 52 yo M Psoriasis Adalimumab Chile Yes—Chagas megacolon, received preemptive treatment with nifurtimox
T. cruzi blood PCR pos 8 months following preemptive nifurtimox Repeat course nifurtimox × 2 mo PCR neg
Burgos et al., 2012 [32] 44 yo F SLE Prednisone 50 mg/d, azathioprine 50 mg/d Paraguay No Parasitemia, skin lesions—erythematous, painful nodules that progressed to ulcer and eschar Benznidazole × 2 mo Clinically improved
Pinazo et al., 2010 [33] 44 yo F SLE Steroids, CP Argentina Yes—chronic indeterminate Chagas
T. cruzi blood PCR pos, treated with benznidazole with recurrent PCR pos Benznidazole × 2 mo, and then posaconazole × 3 mo for relapse Serial PCR neg after posaconazole
Abbreviations: CP, cyclophosphamide; HCQ, hydroxychloroquine; MMF, mycophenolate mofetil; MTX, methotrexate; NA, not applicable; neg, negative; PCR, polymerase chain reaction; pos, positive; RA, rheumatoid arthritis; SLE, systemic lupus erythematosus.
Although limited by small sample size and abbreviated longitudinal follow-up, other studies have attempted to investigate the risk for patients with ARD developing CD reactivation while on immunosuppression. One such study described 2 of 13 patients with ARD who developed symptomatic reactivation on immunosuppressive therapies over a 2-year period, and another 5 patients who developed possible CD reactivation detected by a single positive T. cruzi blood PCR test (Table 1) [30]. In other case series, the majority of immunosuppressed patients with ARD and CD were treated for chronic CD before the development of any evidence of CD reactivation, and only a small minority of these patients subsequently developed CD reactivation while receiving immunosuppression [25, 36, 37]. In 1 such study, 6 of 8 patients with ARD and CD were treated in this manner with varying regimens of benznidazole and nifurtimox. Three patients were treated for CD before a diagnosis of an ARD was made. Only 1 of the patients who received such treatment subsequently developed CD reactivation while receiving immunosuppression (further patient details not specified) [36]. In another study, 11 of 14 patients with chronic CD and ARD were treated for chronic CD with benznidazole for 60 days at the time of study enrollment, and none of the 14 patients experienced CD reactivation while receiving immunosuppression during follow-up [25]. Lastly, 3 patients with ARD and chronic CD were all treated at the time of CD diagnosis with benznidazole for 60 days; none of these patients developed subsequent CD reactivation during a 36-month follow-up period [37]. Though these preliminary data are intriguing, larger population studies, conducted over longer durations, are needed in order to draw more substantial conclusions about factors predisposing to CD reactivation in this cohort and the outcomes of prophylactic therapy.
DISCUSSION
Our patient is among the few in whom symptomatic CD reactivation was documented in the setting of immunosuppressive therapy for an ARD. Our patient likely had unrecognized determinate CD, clinically manifested by tachy-brady syndrome requiring a pacemaker [38]. CD reactivated after intensification of her immunosuppression regimen, ultimately manifesting as disseminated skin nodules, with sequencing of the skin biopsy revealing T. cruzi.
Our patient’s case was unique in that symptomatic CD reactivation occurred in the setting of a negative T. cruzi blood PCR. Typically, T. cruzi blood PCR positivity develops before symptomatic reactivation [11]. Though it is unclear why our patient’s PCR was negative, it is possibly related to blood volume collected or PCR test characteristics. Importantly, this demonstrates that PCR positivity should not be the sole diagnostic measure in the evaluation for CD reactivation; assessment must also include careful clinical evaluation.
CD reactivation in patients being treated for ARD additionally involves a different degree of immunosuppression compared with other better-studied cohorts. An effective host response against T. cruzi requires both cellular and humoral immunity [39]. Our patient was taking low-dose prednisone and azathioprine. Corticosteroids are nonspecific immune function inhibitors [40]. However, there are no reports that describe an increased incidence of CD reactivation in patients receiving corticosteroids as their sole form of immunosuppression. Azathioprine is an antimetabolite that decreases both T and B lymphocyte production [40]. However, some heart transplant recipients with a history of Chagas cardiomyopathy preferentially receive azathioprine over mycophenolate due to a 6-fold lower incidence of CD reactivation with azathioprine compared with mycophenolate [41]. Taken together, prednisone in doses <20 mg/d and azathioprine still seem to convey a relatively low risk for CD reactivation.
In our patient’s case, it is possible that more multifaceted immunosuppression led to her disease. Specifically, the synergistic effect of prednisone and azathioprine, in conjunction with her profound lymphopenia, advanced age, and uncontrolled diabetes, may have contributed to CD reactivation. However, these immunosuppressing factors still do not clearly explain our patient’s profound functional immunosuppression, further exhibited by concurrent disseminated CMV disease. Based on prior data [42–45], patients with rheumatoid arthritis and SLE may have an increased risk of infection independent of immunomodulatory therapy. Further research might better delineate the relationship between CD and immune function.
Screening for Chagas Disease in Patients With ARD
Currently, consensus guidelines recommend serologic screening for CD in transplant donors and recipients with epidemiologic risk factors. A positive serologic result should be confirmed by at least 2 distinct serologic methods. In the United States, CD treatment based solely on a positive serology result in either the donor or the recipient is generally not recommended given the toxicity of the therapeutic options [46, 47]. Alternatively, transplant recipients who are seropositive should be monitored for reactivation, especially during the times of most intense immunosuppression. Laboratory monitoring employs microscopy of blood/buffy coats and blood PCR. If monitoring reveals parasitemia and/or PCR positivity (especially increasing the parasite load on serial quantitative PCRs), patients are typically given preemptive CD treatment, as the development of detectable T. cruzi in this cohort often heralds the development of symptomatic reactivation [46–49].
Given that data regarding CD reactivation in patients with ARD are so scant, it is uncertain if similar screening and treatment guidelines should be applied to ARD patients in the face of immunosuppression. However, in light of our patient’s case and our review of the literature, our opinion is that strong consideration should be given to serologic screening for T. cruzi before immunosuppression for ARD in patients who have CD risk factors. (Note that serologic screening following immunosuppression may be falsely negative due to a blunted immune response.) Data are insufficient to comment on the risks versus benefits of CD treatment based solely on a positive serologic result in this cohort. However, similar to transplant recipients, for those who are seropositive, clinical assessment and serial blood microscopy/PCR monitoring should be employed during immunosuppressive therapy. There should be strong consideration of preemptive treatment of patients with parasitemia and/or PCR blood positivity (especially increasing parasite load on serial quantitative PCRs) even in the absence of symptoms; of course, evaluation/treatment for those with symptoms/signs concerning for CD reactivation. More research is needed to refine the screening and subsequent treatment approach for CD in patients with ARD.
CONCLUSIONS
This report describes CD reactivation in a patient with an ARD receiving immunosuppressive therapy. It highlights the need for more research regarding CD and reactivation in this patient population. It additionally suggests the need for broader serologic screening of patients with risk factors for CD before starting immunosuppression, and subsequent monitoring of at-risk patients while they receive such therapy.
Acknowledgments
Financial support. None
Potential conflicts of interest. All authors: no reported conflicts of interest. All authors have submitted the ICMJE Form for Disclosure of Potential Conflicts of Interest. Conflicts that the editors consider relevant to the content of the manuscript have been disclosed.
Disclaimer. The findings and conclusions in this report are those of the authors and do not necessarily represent the views of the Centers for Disease Control and Prevention.
Patient consent. Our study does not include factors necessitating patient consent. | Fatal | ReactionOutcome | CC BY-NC-ND | 33575423 | 19,494,598 | 2021-02 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Transplant rejection'. | Safety and effectiveness of everolimus in maintenance kidney transplant patients in the real-world setting: results from a 2-year post-marketing surveillance study in Japan.
BACKGROUND
Data on real-world use of everolimus (EVR) in Japanese maintenance kidney transplant (KTx) patients are limited. This post-marketing surveillance study was conducted to assess the safety and effectiveness of EVR, and identify factors affecting renal impairment.
METHODS
Adult maintenance KTx patients were enrolled within 14 days of initiating EVR. Patient medical data were collected using electronic data capture case report forms at 6 months, 1, and 2 years after initiating EVR, or at discontinuation.
RESULTS
All patients receiving EVR in Japan during the surveillance period were enrolled (N = 263). Mean time from transplantation to EVR initiation was 75.7 months. Decreased renal function (31.56%) was the primary reason for initiating EVR. In combination with EVR, the mean daily dose of tacrolimus and cyclosporine could be reduced to ~ 79 and ~ 64%, by 2 years, respectively. Incidences of serious adverse events and adverse drug reactions were 15.97 and 49.43%, respectively. Two-year graft survival rate was 95.82% and low in patients with baseline estimated glomerular filtration rate (eGFR; modification of diet in renal disease) < 30 mL/min/1.73 m2 (69.57%; P < 0.0001) and urinary protein/creatinine ratio (UPCR) ≥ 0.55 g/gCr (84.21%; P = 0.0206). Throughout the survey, mean eGFR values were stable (> 55 mL/min/1.73 m2). Renal impairment was influenced by patient and donor age, eGFR, and UPCR at baseline.
CONCLUSIONS
No new safety concerns for the use of EVR in adult maintenance KTx patients were identified. Early EVR initiation may be considered in these patients before renal function deterioration occurs.
Introduction
Current immunosuppressive protocols with calcineurin inhibitors (CNIs) provide good short-term efficacy but their long-term use is associated with chronic nephrotoxicity [1, 2], CNI arteriolopathy [3], diabetes [4, 5], and cardiovascular complications [6]. Thus, immunosuppressive strategies that can facilitate CNI minimization/elimination, while maintaining long-term anti-rejection efficacy are being developed [7–9].
Several studies have reported the efficacy and safety of everolimus (EVR) as a maintenance immunosuppressant in kidney transplant (KTx) patients [10–14]. The main reasons for switching to an EVR-based regimen were interstitial fibrosis and tubular atrophy, CNI-associated nephrotoxicity, cancer, viral infections, and generalized vascular disease [10, 11].
Although EVR has been approved in Japan for “inhibition of graft rejection in kidney transplantation” in 2011, data on clinical experience with EVR in Japanese maintenance KTx patients are limited. In a previous post-marketing surveillance (PMS) study in Japan, efficacy and safety of EVR in both de novo and maintenance KTx patients was reported up to 2 years. However, no data on renal function, influence of baseline characteristics on efficacy and safety were reported [14]. The current PMS study was planned to assess the safety and effectiveness of EVR in adult maintenance KTx patients and to identify factors responsible for subsequent renal impairment.
Patients and methods
Survey design and population
This was a 2-year, observational, non-interventional, multicenter, PMS study conducted between September 2014 and August 2018 by a central registration system. KTx patients aged ≥ 18 years receiving EVR in the maintenance period were enrolled from December 2014–15. Patients were enrolled within 14 days of initiating EVR and enrollment continued until December 31, 2015. Written informed consent was obtained from all patients before enrollment. Patients who had previously participated in other EVR studies or had exposure to EVR < 3 months prior to enrollment were excluded. Patient medical data were collected using electronic data capture case report forms (CRFs) at 6 months, 1 and 2 years after initiating EVR, or at discontinuation.
Survey objectives
The survey objective was to assess the safety and effectiveness of EVR in maintenance KTx patients and to identify baseline (at the time of EVR initiation) characteristics influencing renal impairment by observing changes in renal function before and after EVR use. Safety and effectiveness of EVR and changes in renal function from baseline were also assessed in elderly patients (≥ 65 years).
Sample size determination
A sample size of 200 patients was determined to explore factors influencing renal impairment after EVR initiation. Using findings from a previous retrospective survey in maintenance KTx patients [11], baseline renal function and donor age were considered as prediction factors, each with two categories (i.e., worse/better baseline renal function and younger/older donors). With 200 patients and a 5% alpha level using a chi-squared test, the probabilities of detecting a significant difference between the two categories each for baseline renal function and donor age were 78 and 90%, respectively.
Survey endpoints and assessments
Safety and effectiveness observation periods were defined as 2 years from EVR initiation until discontinuation or graft loss + 30 days. Safety endpoints included the assessment of serious adverse events (SAEs), adverse drug reactions (ADRs), death, and ADRs by baseline characteristics. SAEs were defined as life-threatening events or death, permanent or significant disability/impairment, congenital abnormality, in-patient hospitalization or prolongation of hospitalization, or medically significant event. Adverse events (AEs) for which a causal relationship with EVR was likely were treated as ADRs. Terminology for AEs was standardized using the Medical Dictionary for Regulatory Activities/Japanese edition, version 21.0. For all AEs, details of causality, action taken, and outcome at each visit until recovery or stabilization were recorded by the investigator. Effectiveness endpoints included the assessment of graft rejection, graft survival, and patient survival rates by baseline characteristics. Graft rejection was clinically diagnosed with/without biopsy and effectiveness rates were defined as the proportion of patients without rejection. ADRs and treatment effectiveness by baseline characteristics were also assessed in elderly patients.
Renal function after initiating EVR was assessed by change in estimated glomerular filtration rate (eGFR; using the Japanese equation [15], international formula [modification of diet in renal disease; MDRD], and serum cystatin C) over time.
Renal impairment at final assessment was defined as percentage decrease in renal function below the 25th percentile eGFR (MDRD) value from EVR initiation, and was assessed by baseline characteristics.
The target trough levels (C0) for EVR were determined as 3–8 ng/mL. Mean EVR C0 and the proportion of patients within the EVR C0 categories (< 3 ng/mL, 3–8 ng/mL, and > 8 ng/mL) were assessed at Months 1, 3, 6, 12, and 24, and at discontinuation.
Analysis sets
The safety and effectiveness analysis sets were defined as patients with at least one fixed CRF volume in whom none of the exclusion criteria (such as deviations in enrollment/unconfirmed enrollment, patients not receiving EVR, no visit following first dose, unfixed first CRF volume, < 6 months post-transplantation, off-label use, duplicate cases, outside of the contract period, and participation in clinical study of an unapproved drug) were applied.
Statistical analysis
To assess the influence of baseline characteristics on ADR or effectiveness, Fisher’s exact test (for nominal categorical baseline characteristics) or the Mann–Whitney U test (for ordinal categorical baseline characteristics with more than two levels) were performed, with a two-sided significance level of 5%. In testing, “unknown,” “not reported,” and “not evaluable” data were excluded. The Mantel–Haenszel test was performed to adjust the stratified effect of baseline characteristics for which a significant difference (P < 0.05) between factors was observed. A factor was suspected to influence ADR or effectiveness if the adjusted analysis showed a significant difference (P < 0.05) between factors. Missing values were not imputed and the value from the last assessment point was carried forward for the final assessment. The proportion and odds ratio (95% CI) of patients with renal impairment by baseline characteristics were evaluated in a descriptive manner.
Results
Patient demographics and baseline characteristics
The survey enrolled 263 patients from 34 medical institutions, and CRFs for all patients were fixed on August 31, 2018. All 263 patients were included in the safety and effectiveness analysis sets. Demographic and baseline characteristics are presented in Table 1. Mean patient and donor ages were 51.5 ± 13.10 years and 55.8 ± 11.69 years, respectively. Of the 263 patients, 56 (21.29%) were elderly (≥ 65 years). The mean time from transplantation to EVR initiation was 75.7 ± 63.17 months. Decreased renal function (31.56%) was the primary reason for initiating EVR. Baseline eGFR (Japanese equation) was ≤ 60 mL/min/1.73 m2 in ~ 80% of patients. EVR was discontinued in 65 (24.71%) patients; AEs being the primary reason for discontinuation in 46 (17.49%) patients.Table 1 Demographic and baseline characteristics (safety analysis set; N = 263)
Characteristics Number of patients, n (%) Characteristics Number of patients, n (%)
Sex HLA mismatches
Male 163 (61.98) < 3 79 (30.04)
Female 100 (38.02) ≥ 3 132 (50.19)
Age (years), mean ± SD 51.5 ± 13.10 Unknown 52 (19.77)
< 50 125 (47.53) Immunological risk at Tx
≥ 50 and < 65 82 (31.18) High risk: ABO-i or PRA( +) 61 (23.19)
≥ 65 56 (21.29) Normal risk 196 (74.52)
Donor age (years), mean ± SD 55.8 ± 11.69 Unknown 6 (2.28)
< 50 65 (24.71) Donor type
≥ 50 and < 65 118 (44.87) Living 225 (85.55)
≥ 65 55 (20.91) Cardiac arrest 25 (9.51)
Unknown 25 (9.51) Brain death 11 (4.18)
Time since Tx (months), mean ± SD 75.7 ± 63.17 Unknown 2 (0.76)
≥ 6 months and < 1 year 48 (18.25) Reason for initiating EVR
≥ 1 year and < 5 years 73 (27.76) Decreased renal function 83 (31.56)
≥ 5 years and < 10 years 88 (33.46) Malignant tumor 36 (13.69)
≥ 10 years 54 (20.53) Cardiovascular event 4 (1.52)
Body weight (kg), mean ± SD 58.8 ± 11.69 (n = 240) Arteriosclerosis 29 (11.03)
Height (cm), mean ± SD 163.7 ± 8.85 (n = 243) Cytomegalovirus infection 11 (4.18)
BMI (kg/m2), mean ± SD 21.9 ± 3.63 (n = 234) Antimetabolite-related AE 3 (1.14)
< 18.5 39 (14.83) MMF-related AE 11 (4.18)
≥ 18.5 and < 25.0 160 (60.84) Other 86 (32.70)
≥ 25.0 35 (13.31) eGFR (Japanese equation; mL/min/1.73 m2)c
Unknown 29 (11.03) < 30 51 (19.39)
Primary disease leading to KTxa ≥ 30 and ≤ 60 161 (61.22)
Chronic glomerulonephritis 80 (30.42) > 60 48 (18.25)
Focal glomerulosclerosis 10 (3.80) Unknown 3 (1.14)
IgA nephropathy 51 (19.39) eGFR (MDRD; mL/min/1.73 m2)c
Interstitial nephritis 2 (0.76) < 30 23 (8.75)
Polycystic kidney 20 (7.60) ≥ 30 and ≤ 60 121 (46.01)
Nephrosclerosis 14 (5.32) > 60 116 (44.11)
Hypoplastic/dysplastic kidney 4 (1.52) Unknown 3 (1.14)
Diabetic nephropathy 26 (9.89) eGFR (serum cystatin C; mL/min/1.73 m2)c
Other 60 (22.81) < 30 22 (8.37)
History of graft rejectionb ≥ 30 and ≤ 60 80 (30.42)
Cellular rejection 12 (4.56) > 60 26 (9.89)
ABMR 22 (8.37) Unknown 135 (51.33)
Relationship with donor UPCR (g/gCr)c
Blood relative 129 (49.05) < 0.55 147 (55.89)
Spouse 89 (33.84) ≥ 0.55 19 (7.22)
Other 45 (17.11) Unknown 97 (36.88)
Unknown 0 (0.0)
As the safety and effectiveness analysis sets are the same, the composition ratios remain the same for both sets
aPrimary disease leading to kidney transplantation allowed multiple selections
bWithin 6 months before the start of EVR treatment
cAt the start of EVR treatment
ABMR antibody-mediated rejection, ABO-i ABO incompatible, AE adverse event, BMI body mass index, eGFR estimated glomerular filtration rate, EVR everolimus, HLA human leukocyte antigen, IgA immunoglobulin A, KTx kidney transplantation, MDRD modification of diet in renal disease, MMF mycophenolate mofetil, PRA panel reactive antibody, SD standard deviation, Tx transplantation, UPCR urinary protein/creatinine ratio
Immunosuppression
The mean treatment and observation periods were 613.2 and 644.3 days, respectively. Most patients (72.62%) received EVR for at least 2 years with a mean daily dose of 1.3 mg. Mean EVR C0 was within the target range throughout the observation period and was 4.39 ± 2.23 ng/mL at the final assessment (Fig. 1a). Adherence to the target EVR C0 was seen in 58.02% of patients (Fig. 1b). During the safety observation period, 182 (69.20%) patients received concomitant tacrolimus and 82 (31.18%) patients received concomitant cyclosporine at least once. By 2 years, the mean daily dose of tacrolimus and cyclosporine was reduced to ~ 79 and ~ 64% of the dose, respectively. The majority of the patients also received mycophenolate mofetil (MMF) (n = 224 [85.17%]) and corticosteroids (n = 225 [85.55%]) during the survey.Fig. 1 Exposure of everolimus (safety analysis set): a Mean (SD) EVR C0 over time, b Proportion of patients with adherence to the EVR target range (3–8 ng/mL). The shaded box indicates the protocol-defined EVR target C0 range (3–8 ng/mL). *Final assessment means at the end of EVR treatment or at discontinuation. C0 trough level, EVR everolimus, M month, SD standard deviation
Safety
The overall incidence of SAEs was 15.97%. Kidney transplant rejection and renal impairment were the most common (1.14% each) SAEs. ADRs occurred in 49.43% of patients, most commonly reported were stomatitis (15.97%), proteinuria (9.89%), hyperlipidemia (5.32%), and peripheral edema (3.80%) (Table 2).Table 2 Incidence rates (≥ 1%) of ADRs by preferred term (safety analysis set; N = 263)
Incidence of ADRs n/N (%) 130/263 (49.43)
Type of ADR Incidence, n (%)
Nasopharyngitis 3 (1.14)
Kidney transplant rejection 3 (1.14)
Dyslipidemia 6 (2.28)
Lipid metabolism disorder 3 (1.14)
Hyperlipidemia 14 (5.32)
Diarrhea 3 (1.14)
Stomatitis 42 (15.97)
Rash 4 (1.52)
Proteinuria 26 (9.89)
Renal impairment 5 (1.90)
Concomitant disease aggravated 3 (1.14)
Peripheral edema 10 (3.80)
Albumin urine present 7 (2.66)
Blood creatinine increased 3 (1.14)
Protein urine 3 (1.14)
Protein urine present 5 (1.90)
ADR adverse drug reaction
Two deaths were reported during the survey. One patient (76 years) died from gastric cancer. Onset was seen on Day 28 post-EVR initiation, and the time from transplantation to EVR initiation was 4.2 years. Another patient (72 years) died from subarachnoid hemorrhage with an onset of 40 days after EVR initiation. The time post-transplantation to EVR initiation was 6.2 years. Both deaths were reported to be unrelated to EVR treatment.
While investigating the incidence of ADRs by baseline characteristics (Table 3), history of antibody-mediated rejection (ABMR), donor type, and reasons for initiating EVR were identified as contributing factors. The frequency of ADRs was higher in patients with versus without a history of ABMR (77.27 versus 47.08%; P = 0.0074). Most ADRs occurred in patients receiving an allograft from brain-death donors (63.64%), followed by living (51.11%) and cardiac arrest (28.0%) donors (P = 0.0494). However, adjusted analysis for ABMR and donor type categories showed no significant differences, suggesting that these findings may be due to confounders. The incidence of ADRs (P = 0.0010) by reasons for initiating EVR is provided in Table 3. As the adjusted analysis also found significant differences, reasons for initiating EVR should be considered as an influencing factor. The incidence of ADRs in elderly patients (41.07%) was numerically lower than in non-elderly patients (51.69%).Table 3 Incidence of ADRs by baseline characteristics (safety analysis set)
aShaded categories were not considered for tests
bAt the start of EVR treatment
ABMR antibody-mediated rejection, ABO-i ABO incompatible, ADR adverse drug reaction, AE adverse event, BMI body mass index, eGFR estimated glomerular filtration rate, EVR everolimus, HLA human leukocyte antigen, IgA immunoglobulin A, KTx kidney transplantation, MDRD modification of diet in renal disease, MMF mycophenolate mofetil, PRA panel reactive antibody, Tx transplantation, UPCR urinary protein/creatinine ratio
Effectiveness
Overall incidence of graft rejection was low (6.84%) in this population. In total, 19 events were reported in 18 patients (Table S1). The 2-year graft survival rate was 95.82% and the patient survival rate was 99.24%. Effectiveness analysis by baseline characteristics is presented in Table 4.Table 4 Effectiveness analysis by baseline characteristics (effectiveness analysis set)
aP values calculated using the Mann − Whitney U test. All other P values are calculated using Fisher’s exact test
bShaded categories were not considered for tests
cAt the start of EVR treatment
ABMR antibody-mediated rejection, ABO-i ABO incompatible, AE adverse event, BMI body mass index, eGFR estimated glomerular filtration rate, EVR everolimus, HLA human leukocyte antigen, IgA immunoglobulin A, KTx kidney transplantation, MDRD modification of diet in renal disease, MMF mycophenolate mofetil, PRA panel reactive antibody, Tx transplantation, UPCR urinary protein/creatinine ratio
Graft rejection
Baseline characteristics such as nephrosclerosis, history of cellular rejection and ABMR, and eGFR (Japanese equation) were found to influence graft rejection (Table 4). Patients with nephrosclerosis before transplantation showed a lower rate of effectiveness (71.43%) compared to patients without nephrosclerosis (94.38%; P = 0.0101). The rates of effectiveness were lower in patients with versus without a history of cellular rejection (66.67 versus 94.80%; P = 0.0044) and in patients with versus without a history of ABMR (77.27 versus 95.00%; P = 0.0083). As the adjusted analysis also found a significant difference, the influence of nephrosclerosis and history of both cellular rejection and ABMR could not be eliminated. The rates of effectiveness were higher in patients within the eGFR categories (Japanese equation) of ≥ 30 and ≤ 60 mL/min/1.73 m2, and > 60 mL/min/1.73 m2 (P = 0.0220). These differences by eGFR category may be due to confounding factors, as the adjusted analysis showed no significant differences. Effectiveness rates were comparable between elderly and non-elderly patients (96.43 versus 92.27%; P = 0.3786).
Graft survival
Graft survival rates were influenced by baseline eGFR (Japanese equation and MDRD) and urinary protein/creatinine ratio (UPCR). Patients with lower baseline eGFR showed lower graft survival rates compared to patients with higher eGFR (> 60 mL/min/1.73 m2; P = 0.0003 for eGFR [Japanese equation] and P < 0.0001 for eGFR [MDRD]). Baseline UPCR of < 0.55 g/gCr showed a higher 2-year graft survival rate (97.96%) versus UPCR ≥ 0.55 g/gCr (84.21%; P = 0.0206) (Table 4). As adjusted analysis failed to show significant differences, the influence of baseline eGFR (Japanese equation) and UPCR could be due to confounding factors. Graft survival rates were comparable between elderly and non-elderly patients (96.43 versus 95.65%; P = 1.0000).
Patient survival
Patient survival rates were associated with baseline characteristics such as patient and donor age. The survival rates were lower in elderly (96.43%) versus non-elderly (100.00%; P = 0.0447) patients (Table 4). Adjusted analysis for patient age also showed significant differences. The two deaths reported during the survey due to gastric cancer and subarachnoid hemorrhage, respectively, were seen in elderly patients. However, the causal relationship between these AEs leading to death and EVR was eliminated for each instance; therefore, an advanced age was not considered to directly affect survival in these patients. Patient survival rates by donor age categories (P = 0.0406) are provided in Table 4. The adjusted analysis showed that the influence of donor age on patient survival could be due to confounding factors.
Renal function
Baseline mean eGFR values were maintained until 2 years after the treatment or the final assessment (Fig. 2). For patients aged < 65 years and ≥ 65 years, mean eGFR (MDRD) at baseline and final assessment were comparable (Fig. 3a). Among patients with baseline UPCR ≥ 0.55 g/gCr, mean eGFR (MDRD) at the final assessment was numerically lower compared to baseline (Fig. 3b).Fig. 2 Mean (SD) change in eGFR over time (safety analysis set). *Final assessment means at the end of EVR treatment or at discontinuation. eGFR estimated glomerular filtration rate, EVR everolimus, M month, MDRD modification of diet in renal disease, SD standard deviation
Fig. 3 Mean (SD) change in eGFR (MDRD; safety analysis set): a By patient age (< 65 years vs ≥ 65 years), b By UPCR categories (< 0.55 g/gCr vs ≥ 0.55 g/gCr). *Final assessment means at the end of EVR treatment or at discontinuation; †Changes over time in the renal function data during the observation period were calculated for patients who had data at each measurement time point out of the 263 patients in the safety analysis set. eGFR estimated glomerular filtration rate, EVR everolimus, MDRD modification of diet in renal disease, SD standard deviation, UPCR urinary protein/creatinine ratio
The proportion of patients with renal impairment (eGFR [MDRD]) at the final assessment by baseline characteristics is presented in Table 5 (renal impairment assessments by eGFR [Japanese equation] and eGFR [serum cystatin C] are given in Table S2 and S3, respectively). Baseline factors such as patient and donor age, eGFR, and UPCR were found to influence renal impairment after EVR treatment. Incidence of renal impairment was lower in the patient age group of ≥ 65 years (16.98%) versus < 50 years (31.71%). In patients with donors aged ≥ 65 years, the incidence of renal impairment was higher (38.89%) versus donors aged < 50 years (20.31%). Irrespective of the formula used for measuring eGFR, more patients with baseline eGFR < 30 mL/min/1.73 m2 showed renal impairment at the final assessment. Of 23 patients with baseline eGFR (MDRD) < 30 mL/min/1.73 m2, 12 (52.17%) showed renal impairment at the final assessment. Similarly, incidence (52.63%) of renal impairment was higher in patients with baseline UPCR ≥ 0.55 g/gCr.Table 5 Proportion of patients with renal impairment by baseline characteristics: percentage decrease in renal function below the 25th percentile eGFR (MDRD)
aShaded categories were not considered for tests
bAt the start of EVR treatment
cAngiotensin II receptor antagonists, angiotensin-converting enzyme inhibitors, treatment drugs for dyslipidemia, and treatment drugs for diabetes mellitus including insulin
dPatients with CNI dose reduction were defined as those in whom the dosage of CNIs was reduced by ≥ 30% relative to the dose at the start of treatment at ≥ 2 time points out of all assessment points
AE adverse event, CI confidence interval, CNI calcineurin inhibitor, eGFR estimated glomerular filtration rate, EVR everolimus, HLA human leukocyte antigen, MDRD modification of diet in renal disease, MMF mycophenolate mofetil, OR odds ratio, Tx transplantation, UPCR urinary protein/creatinine ratio
Discussion
This large PMS study in Japan reported real-world use of EVR in 263 adults, maintenance KTx patients (with 225 living donor transplants), and identified baseline characteristics influencing the subsequent renal impairment. AEs were the primary reason for EVR discontinuation and the most common ADRs reported were stomatitis, proteinuria, hyperlipidemia, and peripheral edema. These safety findings are consistent with those previously reported in the literature for EVR [9–12]. In our survey, with EVR initiation, the mean daily dose of tacrolimus and cyclosporine was reduced to ~ 79 and ~ 64% by 2 years, respectively. Even though blood CNI concentrations were measured at various time points, mean CNI C0 levels were not reported due to a lack of data on the timing of blood sampling. However, post-hoc analysis (data not shown) showed an overall decline in blood CNI levels over the course of EVR treatment.
Late conversion (≥ 3 years after transplantation) to EVR has shown to impact renal function and graft survival [11]. Although the overall graft survival rate (95.82%) at 2 years was high in our survey, low rates were seen in patients with baseline eGFR (MDRD) < 30 mL/min/1.73 m2 and UPCR ≥ 0.55 g/gCr. Similar results were observed in the ASCERTAIN study (assessing the effect of late-conversion [mean 5.6 years post-transplant] to EVR with CNI elimination/minimization in maintenance KTx patients with renal impairment at baseline [defined as GFR 30–70 mL/min]), where overall, 93.3% of patients receiving EVR survived with a functioning graft at Month (M) 24 [12]. This suggests that longer follow-up (beyond 2 years) is needed to conclude the clinical benefits of EVR.
In the ASCERTAIN study, renal function was stable in all treatment groups at M24. Although no overall renal benefit was seen with EVR-based regimen, post-hoc analyses showed that the increase in measured glomerular filtration rate (mGFR) was higher in patients with baseline creatinine clearance (CrCl) > 50 mL/min in the CNI elimination group versus the control group (P = 0.017) [12]. In a retrospective study in KTx patients, improvement in renal function was statistically significant at 1-year in patients who were converted (median 5.8 years post-transplant) to EVR with baseline CrCl ≥ 40 mL/min and proteinuria < 550 mg/day (P = 0.005) [11]. Consistent with these findings, baseline mean eGFR values were maintained during our survey. However, renal impairment at final assessment was higher in patients with baseline eGFR (MDRD) < 30 mL/min/1.73 m2 and UPCR ≥ 0.55 g/gCr. This suggests that patients with good baseline renal function may benefit from an EVR-based regimen. In contrast, Nojima et al. reported significant improvement in renal function at 1-year in Japanese KTx patients, including patients with low baseline eGFR (< 30 mL/min) converted to EVR-based regimen at a mean 7.4 years post-transplant. This could be due to the low CNI C0 levels observed in this study [16].
The lower incidence of renal impairment in elderly patients versus patients aged < 50 years could be due to a high proportion of (i) elderly patients who received a graft from donors aged < 65 years and (ii) patients aged < 50 years who received a graft from donors aged ≥ 50 years. As renal function declines with age [11], graft function may have been impaired at transplantation in patients < 50 years who received a graft from elderly donors. However, adjusted analysis with confounders was not performed to confirm this finding.
The low graft rejection rate in our survey could be because most patients received EVR + CNI + MMF + corticosteroids therapy. Despite the significant difference in the effectiveness rates between patients with versus without nephrosclerosis, clinical factors potentially associated with the observed difference could not be identified in this survey setting due to a small number of patients with nephrosclerosis.
Randomized studies with early EVR initiation (< 6 months post-transplant) have shown clinical benefits of using EVR. In the ZEUS study, conversion to EVR with cyclosporine elimination showed significant improvement in renal function up to 5 years (P < 0.001), while maintaining efficacy and safety [17, 18]. Similarly, conversion to EVR with cyclosporine elimination was associated with a significant increase in mGFR (P = 0.012) at M12 in the CENTRAL study [19] and a significant increase in eGFR (P < 0.001) up to M24 in the ELEVATE study [20]. Results from the largest study in de novo KTx patients (TRANSFORM) with 50% living donor transplants showed comparable antirejection efficacy, stable renal function, and low incidence of de novo donor-specific antibodies (dnDSA) and viral infections with EVR-based regimen at M24 [9]. The clinical benefit of EVR was also evident in de novo Japanese KTx patients in the 12-month A1202 study, where EVR + reduced-exposure cyclosporine (EVR + rCsA) group showed numerically higher median eGFR values (58 mL/min/1.73 m2 versus 55.25 mL/min/1.73 m2; P = 0.063) and comparable safety versus MMF + standard-exposure cyclosporine (MMF + sCsA) group. The graft survival rate at M12 was 100% in both the treatment groups [21]. Moreover, when participants (N = 24) from this study were followed-up at 10 years, the graft survival rate was maintained in the EVR + rCsA (100%) group but was reduced in the MMF + sCsA (90.9%) group. In addition, dnDSA-free survival was significantly better in the EVR + rCsA group [22]. Although these results are from a small population, early EVR initiation showed better clinical outcomes. More robust clinical evidence is needed to conclude the long-term benefits of EVR in Japanese KTx patients.
The main limitations of this survey were lack of a comparator arm to conclude the clinical benefits of EVR and a limited follow-up period of 2 years. In addition, protocol-defined criteria for confirmation of rejection and graft survival were not applicable. However, the survey provided useful insights into the safety and effectiveness of EVR use in Japanese KTx patients in a real-world setting.
In conclusion, this survey showed that EVR initiation can facilitate the reduction of mean daily doses of tacrolimus and cyclosporine to ~ 79 and ~ 64% by 2 years, respectively. Although overall patient and graft survival rates at 2 years were high, graft survival rates were affected by baseline eGFR and UPCR values. Renal impairment was higher in patients with poor baseline eGFR and UPCR. Thus, early EVR initiation (< 6 months post-transplant) may be considered in maintenance KTx patients to prevent renal function deterioration. No new safety concerns for EVR use in Japanese maintenance KTx patients were identified during the survey.
Supplementary Information
Below is the link to the electronic supplementary material.Supplementary file1 (DOCX 30 KB)
Supplementary file2 (DOCX 35 KB)
Supplementary file3 (DOCX 35 KB)
Abbreviations
ABMR Antibody-mediated rejection
ADR Adverse drug reaction
AE Adverse event
C0 Trough level
CNI Calcineurin inhibitor
CRF Case report form
DSA Donor-specific antibodies
eGFR Estimated glomerular filtration rate
EVR Everolimus
MDRD Modification of diet in renal disease
mGFR Measured glomerular filtration rate
MMF Mycophenolate mofetil
rCsA Reduced-exposure cyclosporine
SAE Serious adverse event
SD Standard deviation
sCsA Standard-exposure cyclosporine
UPCR Urinary protein/creatinine ratio
Acknowledgements
The authors thank the patients and the investigators who participated in this survey, as well as Peter Bernhardt (Novartis Pharma AG) and Osamu Kamisawa (former employee, Novartis Pharma K.K.) for their contributions to the study design. The authors thank Ryosuke Imada and Asako Itakura (Novartis Pharma K.K.) as well as Moksha Shah and Aparajita Mandal (Novartis Healthcare Pvt. Ltd.) for medical writing and editorial assistance.
Author contributions
NH: Designed research/study and wrote the paper. MY: Performed research/study. SK: Analyzed data. YW: Interpretation of data for the study.
Funding
The study was funded by Novartis Pharma K.K., Japan.
Compliance with ethical standards
Conflict of interest
Employment: Naomi Hayase, Mariko Yamada, Shuhei Kaneko, and Yoko Watanabe (Novartis Pharma K.K.).
Ethical approval
In accordance with good post-marketing study practice (GPSP), the requirement of ethical approval by an institutional review board was not mandatory for this post-marketing surveillance study.
Informed consent
Written informed consent was obtained from all patients before enrollment.
Publisher's Note
Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. | EVEROLIMUS, MYCOPHENOLATE MOFETIL, TACROLIMUS | DrugsGivenReaction | CC BY | 33575935 | 19,794,642 | 2021-06 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Blood stem cell transplant failure'. | Post-HSCT graft failure due to refractory human cytomegalovirus successfully treated with haploidentical donor-derived immunoglobulins and stem cell graft infusion: A case report.
Human cytomegalovirus (HCMV) remains an important cause of transplant-related morbidity and mortality. The incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations as a result of a lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen.
We describe the case of an 8-year-old βE-thalassemic girl from Bangladesh who was seropositive for human cytomegalovirus (HCMV) and underwent hematopoietic stem cell transplantation from a HLA-matched, unrelated, HCMV-seronegative donor. Despite administering antiviral prophylaxis with commercial pooled anti-HCMV immunoglobulin (Ig) from day +1, the post-transplant course was complicated by prompt viral reactivation, and foscarnet therapy was initiated. The virus was refractory to treatment, leading rapidly to complete bone marrow failure, and targeted immunotherapy was proposed as a second-line therapy. Hypothesizing that the patient and her relatives may have been exposed to similar HCMV strains, we selected the patient's mother, who presented a high HCMV antibody titer, as the donor of virus strain-specific anti-HCMV Ig and T-lymphocytes. Complete viral clearance was achieved after two transfusions of the mother's plasma. Subsequently, the patient underwent a haploidentical rescue transplant, promptly reaching full hematological recovery.
These findings suggest that treatment with virus strain-specific Ig may offer a new therapeutic option for critically ill patients.
pmc1 Introduction
Human cytomegalovirus (HCMV) is the most significant cause of opportunistic viral infection in allogeneic hematopoietic stem cell transplantation (HSCT) and remains an important cause of transplant-related morbidity and mortality (Wagner-Drouet et al., 2019). Several studies have shown that the incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations (Ganepola et al., 2007; Zhou et al., 2009; Ozdemir et al., 2007). This is likely a consequence of delayed HCMV-specific immune reconstitution due to the lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen (Styczynski, 2018; Ljungman et al., 2014). HCMV infection can lead to pneumonia, gastroenteritis, retinitis, hepatitis, and encephalitis among HSCT recipients (Griffiths, 2012). In addition, HCMV infection of the bone marrow can result in impaired graft acceptance and, in the most extreme cases, complete graft failure (Steffens et al., 1998). Treatment of HCMV disease essentially consists of administering antiviral drugs, namely ganciclovir and foscarnet, and intravenous immunoglobulin. Although chemoprophylaxis is not generally recommended in HSCT, immunoglobulin, whether pooled or HCMV-specific, have shown some efficacy in preventing recurrent HCMV infections (Boeckh and Ljungman, 2009).
We describe a case of a pediatric HSCT recipient who had complications of HCMV recurrence and was treated successfully with targeted immunological therapy, using haploidentical donor-derived immunoglobulin (Ig).
2 Case presentation
An 8-year-old girl was diagnosed at the age of two in her country of origin (Bangladesh) as βE-thalassemic. She was treated with monthly blood transfusions and weekly iron chelation with deferoxamine, plus supplementation with folic acid. A suboptimal regimen of chelation therapy led to severe systemic siderosis, and her family travelled to Italy to undergo HSCT. Since a human leukocyte antigen (HLA)-matched related donor was unavailable at that time, an HLA-matched (10/10) unrelated donor was identified. The patient was HCMV-seropositive, whereas the donor was HCMV-seronegative, thus posing a high-risk serological combination (D-/R+) for post-transplant HCMV infection and associated complications.
The conditioning regimen consisted of fludarabine, busulfan, thiotepa, and anti-thymocyte globulin (ATG). The infused donor's bone marrow contained 3.79 × 108 total nuclear cells/kg recipient weight, and the vitality of the cells contained in the graft was 96.9%. Tacrolimus and mycophenolate mofetil were used for graft-versus-host disease (GVHD) prophylaxis. Anti-HCMV prophylaxis commenced at day +1 with commercial pooled anti-HCMV Ig (Megalotect, Biotest Pharma GmbH, Germany) at a dose of 100 U/kg twice a week.
The initial post-transplant period was uneventful. The first occurrence of HCMV viremia (600 viral genome copies/mL) was detected at day +14, and antiviral therapy with foscarnet was started immediately. At day +17, neutrophil engraftment was achieved, but not the platelet and the erythrocyte counterparts. At day +20, the patient presented fever without clinical symptoms and without an increase of C-reactive protein. Afterwards, a full blood count revealed a progressive drop in leukocytes and platelets, whereas HCMV viremia continued to increase despite antiviral therapy. The patient also developed a severe, HCMV-related hemorrhagic cystitis, which required frequent platelet transfusions, initially once daily for 5 days, and thereafter twice daily for 7 days.
As the infection appeared unresponsive to foscarnet, ganciclovir treatment was started at day +25. This switch in antiviral therapy followed an increase in viral load, reaching a maximum titer of 5100 viral genome copies/mL at day +30. Simultaneously, the clinical condition of the patient continued to deteriorate, and the leukocyte count decreased drastically. Therefore, the diagnosis of secondary graft failure due to HCMV infection was established. Because of the failure of anti-HCMV treatment, ganciclovir was withdrawn, and foscarnet therapy was resumed. The possible explanations for treatment failure include the presence of antiviral resistance mutations or an ineffective activity of the Ig administered. As the first option seemed improbable because resistance mutations usually arise later than 6 weeks after HSCT, we opted to address the latter option. Therefore, we considered using antibodies (Abs) that were more likely to inhibit the HCMV strains causing the infection in the patient. Although the diversity of circulating HCMV strains is known to be extensive (Puchhammer-Stöckl and Görzer, 2011), we hypothesized that the patient and her relatives may have been exposed to similar strains on a familial basis. As a result, we performed serological tests on the patient's mother, father and brother. We selected the patient's mother, who presented the highest anti-HCMV IgG titer, as the best donor of strain-specific HCMV Ab-rich plasma and virus-specific T-lymphocytes in this situation. At day +32, the patient's mother underwent leukapheresis. The lymphocytes were sent immediately to a cell factory to produce specific anti-HCMV T-lymphocytes. On the same day, the plasma enriched in HCMV strain-specific Abs was infused into the patient. At day +34, the viral load decreased for the first time, to 1100 viral genome copies/mL, and the patient began new conditioning for a haploidentical rescue transplant from her mother. In light of the concurrent bone marrow aplasia, the conditioning regimen was conducted solely with ATG. At day +38, the girl received a second plasma transfusion from her mother. On the day of the second transplant (day +40), complete viral clearance was documented, and the girl received 11.3 × 106 CD34 cells/kg of mother's graft. GVHD prophylaxis was performed with post-transplant cyclophosphamide, tacrolimus, and mycophenolate mofetil. The engraftment was rapid, with a neutrophil count of >500/μL achieved at day +15, and the final platelets and packed red blood cells were transfused at day +20. Since HCMV viremia remained negative during the entire post-transplant period, we discarded the use of the mother's virus-specific T-lymphocytes. The patient was discharged at day +27 in perfect clinical condition, with continuation of antiviral valganciclovir prophylaxis. Immunosuppressive treatment and antiviral prophylaxis were discontinued six months later, and the patient remains in perfect health 2 years after the second transplant.
Therapeutic interventions, HCMV load, and white blood cell count during the entire episode are summarized in Fig. 1. In addition, the HCMV strains present in the patient were characterized by high-throughput sequencing (HTS) of an archived (−80 °C) plasma sample from day +19. DNA was extracted from 800 μl of plasma using a QIAamp MinElute virus spin kit (QIAGEN, Crawley, UK). An aliquot of 50 μl of DNA was used to generate an HCMV-enriched sequencing library as described previously (Suárez et al., 2019). The library was loaded onto a NextSeq DNA sequencer (Illumina, San Diego, CA, USA), generating 150 bp paired-ended reads (Table 1). HCMV strain enumeration was performed by genotyping 12 hypervariable HCMV genes from the reads as described previously (Suárez et al., 2019). The detection of four genotypes of the RL13 gene indicated the presence of at least four HCMV strains (a strain being defined as a particular constellation of genotypes of the hypervariable genes analysed), each including one of these RL13 genotypes (Table 2). The data was also inspected for the presence of resistance mutations in the antiviral target genes (UL54 and UL97) using LoFreq (Wilm et al., 2012). Briefly, the reads were mapped to the genome of HCMV reference strain Merlin (GenBank accession no. AY446894.2), and non-synonymous variants known to confer resistance were identified. No resistance mutations were detected.Fig. 1 Information on viremia, white blood cell (WBC) count, total serum IgG and antiviral therapy. Data on HCMV viremia is expressed in viral genome copies/mL (right Y-axis). Application and duration of antiviral treatments are represented by colored bars (yellow, anti-HCMV-specific Ig; red, foscarnet; blue, ganciclovir/valganciclovir). Chronological data (X-axis) are represented in days after the first transplant. The plasma sample analysed by high-throughput sequencing (HTS) is represented by a filled blue circle.
Fig. 1
Table 1 Overview of HTS data.
Table 1Sample ID TRI-SCT1
Sample type Plasma
Days after transplant +19
HCMV load (genome copies/mL) 2100
Data deposition (ENA project no.)a PRJEB38333
Sequencing library (genome copies) 1440
Trimmed reads (no.) 5,033,480
HCMV reads (no.)b 936,773
HCMV reads; %b 19
Coverage depth (reads/nt)b 885
a The sequence dataset was purged of human reads and deposited in the European Nucleotide Archive (ENA).
b Reads mapped to the HCMV reference strain Merlin genome; HTS, high-throughput sequencing; nt, nucleotide.
Table 2 Genotyping of hypervariable HCMV genes.
Table 2
3 Discussion
Various factors are implicated in the outcome of HCMV infection in transplant patients, the first of which relates to the HCMV serological status of donor and recipient. The serological status of the first donor may have played an important role in this case, as it has been reported that using an HCMV-seronegative unrelated HSCT donor (D-) for a seropositive recipient (R+) results in reduced overall survival after myeloablative conditioning. In contrast, an HCMV seropositive donor is associated with improved event-free survival and reduced non-relapse mortality (Styczynski, 2018). Thus, the absence of an HCMV-specific immunological memory in the donor's cells may have placed the HCMV-positive recipient at a higher risk of HCMV reactivation and associated complications.
In relation to viral factors, the clinical course of the patient may have been influenced by two main factors: the presence of multiple HCMV strains or the presence of mutations conferring antiviral resistance. Infection with multiple HCMV strains is common after transplantation (Görzer et al., 2010), and is associated with delayed HCMV clearance during antiviral therapy, even in the absence of antiviral resistance mutations. It is also associated with worse clinical outcomes, including an increase in graft rejection and faster progression of disease. In principle, the mechanisms behind this increased pathogenicity can be viral, for example involving recombination and complementation of different strains within the host, or immunological, since control of multiple strains may be more challenging for the immune system. Overall, in clinically symptomatic immunocompromised hosts, the presence of multiple HCMV strains is more threatening than the presence of a single strain (Manuel et al., 2009).
In the present study, the initial failure of antiviral therapy could not be explained by the development of resistance mutations in the HCMV strains present in the patient. This finding is not surprising, as detection of resistance mutations is unusual during the first 6 weeks of antiviral therapy, whereas the sample analysed in this case was collected only 13 days after initiation of foscarnet treatment (Lurain and Chou, 2010; Springer et al., 2005).
In regard to the initial failure of the immunotherapy with commercial pooled Ig, this may have been due to the lack of particular Abs with neutralizing capacity against the HCMV strains present in the patient. Assuming that the donors providing the Ig had been exposed to strains circulating in Europe, the Abs may have been more suited for neutralizing HCMV strains commonly circulating in that region. However, it is possible that the strains present in the patient, who originated from Bangladesh, may have differed immunologically from strains in other regions of the world. The extent to which this is true is unknown, as current knowledge of HCMV genome variability is derived almost entirely from strains circulating in a few European countries. Therefore, it would be useful to assess the neutralizing capacity of commercial pooled Ig in order to avoid potential therapeutic failure, especially in patients from non-European countries. The clinical problem in this case was addressed successfully by using the Ab-rich plasma of the patient's mother in a kind of personalized immunotherapy, on the premise that both mother and daughter may have been exposed to the same pool of viral strains. In this HCMV-positive recipient with a severe viral reactivation, Ig obtained from a household donor afforded an efficacious treatment that led to complete viral clearance.
To the best of our knowledge, this is the first report of the successful use of HCMV strain-specific Ig therapy in an HSCT recipient. The efficacy of this approach as a means of prevention has been demonstrated previously only in mouse models involving murine cytomegalovirus (MCMV) reactivation after bone marrow transplantation (Martins et al., 2019). In this setting, mice received serum obtained either from latently infected (seropositive) donors or from seronegative donors. The transfer of immune serum protected mice from viral reactivation without affecting the development of GVHD. Remarkably, complete viral protection was obtained with a small volume (5 μL) of strain-specific immune serum. On the other hand, when specific donor antibodies were administered to mice infected with antigenically mismatched MCMV strains, they were not efficient in preventing viral reactivation.
Our case highlights the importance of monitoring HSCT recipients comprehensively, longitudinally and with high sensitivity for viral factors, including the presence of multiple strains and resistance mutations. Importantly, it also provides an affordable targeted immunological therapy in patients from countries with constrained access to alternative therapies.
4 Conclusion
HSCT is a complex and specific setting in which a recipient may rapidly develop viral reactivation and consequent life-threatening complications. Ab-rich plasma may represent a fast and feasible therapeutic option to overcome viral activity and allow successful engraftment of the donor's bone marrow.
Funding
No specific funding was received.
Consent for publication
The authors have obtained consent from the parents of the patient to publish individual patient data.
Declaration of competing interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgments
The sequencing work was supported by funding from the Medical Research Council (grant number MC_UU_12014/3 to AJD). | BUSULFAN, FLUDARABINE PHOSPHATE, LAPINE T-LYMPHOCYTE IMMUNE GLOBULIN, MYCOPHENOLATE MOFETIL, TACROLIMUS, THIOTEPA | DrugsGivenReaction | CC BY | 33577809 | 19,010,959 | 2021-04 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Drug ineffective'. | Post-HSCT graft failure due to refractory human cytomegalovirus successfully treated with haploidentical donor-derived immunoglobulins and stem cell graft infusion: A case report.
Human cytomegalovirus (HCMV) remains an important cause of transplant-related morbidity and mortality. The incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations as a result of a lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen.
We describe the case of an 8-year-old βE-thalassemic girl from Bangladesh who was seropositive for human cytomegalovirus (HCMV) and underwent hematopoietic stem cell transplantation from a HLA-matched, unrelated, HCMV-seronegative donor. Despite administering antiviral prophylaxis with commercial pooled anti-HCMV immunoglobulin (Ig) from day +1, the post-transplant course was complicated by prompt viral reactivation, and foscarnet therapy was initiated. The virus was refractory to treatment, leading rapidly to complete bone marrow failure, and targeted immunotherapy was proposed as a second-line therapy. Hypothesizing that the patient and her relatives may have been exposed to similar HCMV strains, we selected the patient's mother, who presented a high HCMV antibody titer, as the donor of virus strain-specific anti-HCMV Ig and T-lymphocytes. Complete viral clearance was achieved after two transfusions of the mother's plasma. Subsequently, the patient underwent a haploidentical rescue transplant, promptly reaching full hematological recovery.
These findings suggest that treatment with virus strain-specific Ig may offer a new therapeutic option for critically ill patients.
pmc1 Introduction
Human cytomegalovirus (HCMV) is the most significant cause of opportunistic viral infection in allogeneic hematopoietic stem cell transplantation (HSCT) and remains an important cause of transplant-related morbidity and mortality (Wagner-Drouet et al., 2019). Several studies have shown that the incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations (Ganepola et al., 2007; Zhou et al., 2009; Ozdemir et al., 2007). This is likely a consequence of delayed HCMV-specific immune reconstitution due to the lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen (Styczynski, 2018; Ljungman et al., 2014). HCMV infection can lead to pneumonia, gastroenteritis, retinitis, hepatitis, and encephalitis among HSCT recipients (Griffiths, 2012). In addition, HCMV infection of the bone marrow can result in impaired graft acceptance and, in the most extreme cases, complete graft failure (Steffens et al., 1998). Treatment of HCMV disease essentially consists of administering antiviral drugs, namely ganciclovir and foscarnet, and intravenous immunoglobulin. Although chemoprophylaxis is not generally recommended in HSCT, immunoglobulin, whether pooled or HCMV-specific, have shown some efficacy in preventing recurrent HCMV infections (Boeckh and Ljungman, 2009).
We describe a case of a pediatric HSCT recipient who had complications of HCMV recurrence and was treated successfully with targeted immunological therapy, using haploidentical donor-derived immunoglobulin (Ig).
2 Case presentation
An 8-year-old girl was diagnosed at the age of two in her country of origin (Bangladesh) as βE-thalassemic. She was treated with monthly blood transfusions and weekly iron chelation with deferoxamine, plus supplementation with folic acid. A suboptimal regimen of chelation therapy led to severe systemic siderosis, and her family travelled to Italy to undergo HSCT. Since a human leukocyte antigen (HLA)-matched related donor was unavailable at that time, an HLA-matched (10/10) unrelated donor was identified. The patient was HCMV-seropositive, whereas the donor was HCMV-seronegative, thus posing a high-risk serological combination (D-/R+) for post-transplant HCMV infection and associated complications.
The conditioning regimen consisted of fludarabine, busulfan, thiotepa, and anti-thymocyte globulin (ATG). The infused donor's bone marrow contained 3.79 × 108 total nuclear cells/kg recipient weight, and the vitality of the cells contained in the graft was 96.9%. Tacrolimus and mycophenolate mofetil were used for graft-versus-host disease (GVHD) prophylaxis. Anti-HCMV prophylaxis commenced at day +1 with commercial pooled anti-HCMV Ig (Megalotect, Biotest Pharma GmbH, Germany) at a dose of 100 U/kg twice a week.
The initial post-transplant period was uneventful. The first occurrence of HCMV viremia (600 viral genome copies/mL) was detected at day +14, and antiviral therapy with foscarnet was started immediately. At day +17, neutrophil engraftment was achieved, but not the platelet and the erythrocyte counterparts. At day +20, the patient presented fever without clinical symptoms and without an increase of C-reactive protein. Afterwards, a full blood count revealed a progressive drop in leukocytes and platelets, whereas HCMV viremia continued to increase despite antiviral therapy. The patient also developed a severe, HCMV-related hemorrhagic cystitis, which required frequent platelet transfusions, initially once daily for 5 days, and thereafter twice daily for 7 days.
As the infection appeared unresponsive to foscarnet, ganciclovir treatment was started at day +25. This switch in antiviral therapy followed an increase in viral load, reaching a maximum titer of 5100 viral genome copies/mL at day +30. Simultaneously, the clinical condition of the patient continued to deteriorate, and the leukocyte count decreased drastically. Therefore, the diagnosis of secondary graft failure due to HCMV infection was established. Because of the failure of anti-HCMV treatment, ganciclovir was withdrawn, and foscarnet therapy was resumed. The possible explanations for treatment failure include the presence of antiviral resistance mutations or an ineffective activity of the Ig administered. As the first option seemed improbable because resistance mutations usually arise later than 6 weeks after HSCT, we opted to address the latter option. Therefore, we considered using antibodies (Abs) that were more likely to inhibit the HCMV strains causing the infection in the patient. Although the diversity of circulating HCMV strains is known to be extensive (Puchhammer-Stöckl and Görzer, 2011), we hypothesized that the patient and her relatives may have been exposed to similar strains on a familial basis. As a result, we performed serological tests on the patient's mother, father and brother. We selected the patient's mother, who presented the highest anti-HCMV IgG titer, as the best donor of strain-specific HCMV Ab-rich plasma and virus-specific T-lymphocytes in this situation. At day +32, the patient's mother underwent leukapheresis. The lymphocytes were sent immediately to a cell factory to produce specific anti-HCMV T-lymphocytes. On the same day, the plasma enriched in HCMV strain-specific Abs was infused into the patient. At day +34, the viral load decreased for the first time, to 1100 viral genome copies/mL, and the patient began new conditioning for a haploidentical rescue transplant from her mother. In light of the concurrent bone marrow aplasia, the conditioning regimen was conducted solely with ATG. At day +38, the girl received a second plasma transfusion from her mother. On the day of the second transplant (day +40), complete viral clearance was documented, and the girl received 11.3 × 106 CD34 cells/kg of mother's graft. GVHD prophylaxis was performed with post-transplant cyclophosphamide, tacrolimus, and mycophenolate mofetil. The engraftment was rapid, with a neutrophil count of >500/μL achieved at day +15, and the final platelets and packed red blood cells were transfused at day +20. Since HCMV viremia remained negative during the entire post-transplant period, we discarded the use of the mother's virus-specific T-lymphocytes. The patient was discharged at day +27 in perfect clinical condition, with continuation of antiviral valganciclovir prophylaxis. Immunosuppressive treatment and antiviral prophylaxis were discontinued six months later, and the patient remains in perfect health 2 years after the second transplant.
Therapeutic interventions, HCMV load, and white blood cell count during the entire episode are summarized in Fig. 1. In addition, the HCMV strains present in the patient were characterized by high-throughput sequencing (HTS) of an archived (−80 °C) plasma sample from day +19. DNA was extracted from 800 μl of plasma using a QIAamp MinElute virus spin kit (QIAGEN, Crawley, UK). An aliquot of 50 μl of DNA was used to generate an HCMV-enriched sequencing library as described previously (Suárez et al., 2019). The library was loaded onto a NextSeq DNA sequencer (Illumina, San Diego, CA, USA), generating 150 bp paired-ended reads (Table 1). HCMV strain enumeration was performed by genotyping 12 hypervariable HCMV genes from the reads as described previously (Suárez et al., 2019). The detection of four genotypes of the RL13 gene indicated the presence of at least four HCMV strains (a strain being defined as a particular constellation of genotypes of the hypervariable genes analysed), each including one of these RL13 genotypes (Table 2). The data was also inspected for the presence of resistance mutations in the antiviral target genes (UL54 and UL97) using LoFreq (Wilm et al., 2012). Briefly, the reads were mapped to the genome of HCMV reference strain Merlin (GenBank accession no. AY446894.2), and non-synonymous variants known to confer resistance were identified. No resistance mutations were detected.Fig. 1 Information on viremia, white blood cell (WBC) count, total serum IgG and antiviral therapy. Data on HCMV viremia is expressed in viral genome copies/mL (right Y-axis). Application and duration of antiviral treatments are represented by colored bars (yellow, anti-HCMV-specific Ig; red, foscarnet; blue, ganciclovir/valganciclovir). Chronological data (X-axis) are represented in days after the first transplant. The plasma sample analysed by high-throughput sequencing (HTS) is represented by a filled blue circle.
Fig. 1
Table 1 Overview of HTS data.
Table 1Sample ID TRI-SCT1
Sample type Plasma
Days after transplant +19
HCMV load (genome copies/mL) 2100
Data deposition (ENA project no.)a PRJEB38333
Sequencing library (genome copies) 1440
Trimmed reads (no.) 5,033,480
HCMV reads (no.)b 936,773
HCMV reads; %b 19
Coverage depth (reads/nt)b 885
a The sequence dataset was purged of human reads and deposited in the European Nucleotide Archive (ENA).
b Reads mapped to the HCMV reference strain Merlin genome; HTS, high-throughput sequencing; nt, nucleotide.
Table 2 Genotyping of hypervariable HCMV genes.
Table 2
3 Discussion
Various factors are implicated in the outcome of HCMV infection in transplant patients, the first of which relates to the HCMV serological status of donor and recipient. The serological status of the first donor may have played an important role in this case, as it has been reported that using an HCMV-seronegative unrelated HSCT donor (D-) for a seropositive recipient (R+) results in reduced overall survival after myeloablative conditioning. In contrast, an HCMV seropositive donor is associated with improved event-free survival and reduced non-relapse mortality (Styczynski, 2018). Thus, the absence of an HCMV-specific immunological memory in the donor's cells may have placed the HCMV-positive recipient at a higher risk of HCMV reactivation and associated complications.
In relation to viral factors, the clinical course of the patient may have been influenced by two main factors: the presence of multiple HCMV strains or the presence of mutations conferring antiviral resistance. Infection with multiple HCMV strains is common after transplantation (Görzer et al., 2010), and is associated with delayed HCMV clearance during antiviral therapy, even in the absence of antiviral resistance mutations. It is also associated with worse clinical outcomes, including an increase in graft rejection and faster progression of disease. In principle, the mechanisms behind this increased pathogenicity can be viral, for example involving recombination and complementation of different strains within the host, or immunological, since control of multiple strains may be more challenging for the immune system. Overall, in clinically symptomatic immunocompromised hosts, the presence of multiple HCMV strains is more threatening than the presence of a single strain (Manuel et al., 2009).
In the present study, the initial failure of antiviral therapy could not be explained by the development of resistance mutations in the HCMV strains present in the patient. This finding is not surprising, as detection of resistance mutations is unusual during the first 6 weeks of antiviral therapy, whereas the sample analysed in this case was collected only 13 days after initiation of foscarnet treatment (Lurain and Chou, 2010; Springer et al., 2005).
In regard to the initial failure of the immunotherapy with commercial pooled Ig, this may have been due to the lack of particular Abs with neutralizing capacity against the HCMV strains present in the patient. Assuming that the donors providing the Ig had been exposed to strains circulating in Europe, the Abs may have been more suited for neutralizing HCMV strains commonly circulating in that region. However, it is possible that the strains present in the patient, who originated from Bangladesh, may have differed immunologically from strains in other regions of the world. The extent to which this is true is unknown, as current knowledge of HCMV genome variability is derived almost entirely from strains circulating in a few European countries. Therefore, it would be useful to assess the neutralizing capacity of commercial pooled Ig in order to avoid potential therapeutic failure, especially in patients from non-European countries. The clinical problem in this case was addressed successfully by using the Ab-rich plasma of the patient's mother in a kind of personalized immunotherapy, on the premise that both mother and daughter may have been exposed to the same pool of viral strains. In this HCMV-positive recipient with a severe viral reactivation, Ig obtained from a household donor afforded an efficacious treatment that led to complete viral clearance.
To the best of our knowledge, this is the first report of the successful use of HCMV strain-specific Ig therapy in an HSCT recipient. The efficacy of this approach as a means of prevention has been demonstrated previously only in mouse models involving murine cytomegalovirus (MCMV) reactivation after bone marrow transplantation (Martins et al., 2019). In this setting, mice received serum obtained either from latently infected (seropositive) donors or from seronegative donors. The transfer of immune serum protected mice from viral reactivation without affecting the development of GVHD. Remarkably, complete viral protection was obtained with a small volume (5 μL) of strain-specific immune serum. On the other hand, when specific donor antibodies were administered to mice infected with antigenically mismatched MCMV strains, they were not efficient in preventing viral reactivation.
Our case highlights the importance of monitoring HSCT recipients comprehensively, longitudinally and with high sensitivity for viral factors, including the presence of multiple strains and resistance mutations. Importantly, it also provides an affordable targeted immunological therapy in patients from countries with constrained access to alternative therapies.
4 Conclusion
HSCT is a complex and specific setting in which a recipient may rapidly develop viral reactivation and consequent life-threatening complications. Ab-rich plasma may represent a fast and feasible therapeutic option to overcome viral activity and allow successful engraftment of the donor's bone marrow.
Funding
No specific funding was received.
Consent for publication
The authors have obtained consent from the parents of the patient to publish individual patient data.
Declaration of competing interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgments
The sequencing work was supported by funding from the Medical Research Council (grant number MC_UU_12014/3 to AJD). | FOSCARNET, GANCICLOVIR | DrugsGivenReaction | CC BY | 33577809 | 19,082,624 | 2021-04 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Product use issue'. | Post-HSCT graft failure due to refractory human cytomegalovirus successfully treated with haploidentical donor-derived immunoglobulins and stem cell graft infusion: A case report.
Human cytomegalovirus (HCMV) remains an important cause of transplant-related morbidity and mortality. The incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations as a result of a lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen.
We describe the case of an 8-year-old βE-thalassemic girl from Bangladesh who was seropositive for human cytomegalovirus (HCMV) and underwent hematopoietic stem cell transplantation from a HLA-matched, unrelated, HCMV-seronegative donor. Despite administering antiviral prophylaxis with commercial pooled anti-HCMV immunoglobulin (Ig) from day +1, the post-transplant course was complicated by prompt viral reactivation, and foscarnet therapy was initiated. The virus was refractory to treatment, leading rapidly to complete bone marrow failure, and targeted immunotherapy was proposed as a second-line therapy. Hypothesizing that the patient and her relatives may have been exposed to similar HCMV strains, we selected the patient's mother, who presented a high HCMV antibody titer, as the donor of virus strain-specific anti-HCMV Ig and T-lymphocytes. Complete viral clearance was achieved after two transfusions of the mother's plasma. Subsequently, the patient underwent a haploidentical rescue transplant, promptly reaching full hematological recovery.
These findings suggest that treatment with virus strain-specific Ig may offer a new therapeutic option for critically ill patients.
pmc1 Introduction
Human cytomegalovirus (HCMV) is the most significant cause of opportunistic viral infection in allogeneic hematopoietic stem cell transplantation (HSCT) and remains an important cause of transplant-related morbidity and mortality (Wagner-Drouet et al., 2019). Several studies have shown that the incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations (Ganepola et al., 2007; Zhou et al., 2009; Ozdemir et al., 2007). This is likely a consequence of delayed HCMV-specific immune reconstitution due to the lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen (Styczynski, 2018; Ljungman et al., 2014). HCMV infection can lead to pneumonia, gastroenteritis, retinitis, hepatitis, and encephalitis among HSCT recipients (Griffiths, 2012). In addition, HCMV infection of the bone marrow can result in impaired graft acceptance and, in the most extreme cases, complete graft failure (Steffens et al., 1998). Treatment of HCMV disease essentially consists of administering antiviral drugs, namely ganciclovir and foscarnet, and intravenous immunoglobulin. Although chemoprophylaxis is not generally recommended in HSCT, immunoglobulin, whether pooled or HCMV-specific, have shown some efficacy in preventing recurrent HCMV infections (Boeckh and Ljungman, 2009).
We describe a case of a pediatric HSCT recipient who had complications of HCMV recurrence and was treated successfully with targeted immunological therapy, using haploidentical donor-derived immunoglobulin (Ig).
2 Case presentation
An 8-year-old girl was diagnosed at the age of two in her country of origin (Bangladesh) as βE-thalassemic. She was treated with monthly blood transfusions and weekly iron chelation with deferoxamine, plus supplementation with folic acid. A suboptimal regimen of chelation therapy led to severe systemic siderosis, and her family travelled to Italy to undergo HSCT. Since a human leukocyte antigen (HLA)-matched related donor was unavailable at that time, an HLA-matched (10/10) unrelated donor was identified. The patient was HCMV-seropositive, whereas the donor was HCMV-seronegative, thus posing a high-risk serological combination (D-/R+) for post-transplant HCMV infection and associated complications.
The conditioning regimen consisted of fludarabine, busulfan, thiotepa, and anti-thymocyte globulin (ATG). The infused donor's bone marrow contained 3.79 × 108 total nuclear cells/kg recipient weight, and the vitality of the cells contained in the graft was 96.9%. Tacrolimus and mycophenolate mofetil were used for graft-versus-host disease (GVHD) prophylaxis. Anti-HCMV prophylaxis commenced at day +1 with commercial pooled anti-HCMV Ig (Megalotect, Biotest Pharma GmbH, Germany) at a dose of 100 U/kg twice a week.
The initial post-transplant period was uneventful. The first occurrence of HCMV viremia (600 viral genome copies/mL) was detected at day +14, and antiviral therapy with foscarnet was started immediately. At day +17, neutrophil engraftment was achieved, but not the platelet and the erythrocyte counterparts. At day +20, the patient presented fever without clinical symptoms and without an increase of C-reactive protein. Afterwards, a full blood count revealed a progressive drop in leukocytes and platelets, whereas HCMV viremia continued to increase despite antiviral therapy. The patient also developed a severe, HCMV-related hemorrhagic cystitis, which required frequent platelet transfusions, initially once daily for 5 days, and thereafter twice daily for 7 days.
As the infection appeared unresponsive to foscarnet, ganciclovir treatment was started at day +25. This switch in antiviral therapy followed an increase in viral load, reaching a maximum titer of 5100 viral genome copies/mL at day +30. Simultaneously, the clinical condition of the patient continued to deteriorate, and the leukocyte count decreased drastically. Therefore, the diagnosis of secondary graft failure due to HCMV infection was established. Because of the failure of anti-HCMV treatment, ganciclovir was withdrawn, and foscarnet therapy was resumed. The possible explanations for treatment failure include the presence of antiviral resistance mutations or an ineffective activity of the Ig administered. As the first option seemed improbable because resistance mutations usually arise later than 6 weeks after HSCT, we opted to address the latter option. Therefore, we considered using antibodies (Abs) that were more likely to inhibit the HCMV strains causing the infection in the patient. Although the diversity of circulating HCMV strains is known to be extensive (Puchhammer-Stöckl and Görzer, 2011), we hypothesized that the patient and her relatives may have been exposed to similar strains on a familial basis. As a result, we performed serological tests on the patient's mother, father and brother. We selected the patient's mother, who presented the highest anti-HCMV IgG titer, as the best donor of strain-specific HCMV Ab-rich plasma and virus-specific T-lymphocytes in this situation. At day +32, the patient's mother underwent leukapheresis. The lymphocytes were sent immediately to a cell factory to produce specific anti-HCMV T-lymphocytes. On the same day, the plasma enriched in HCMV strain-specific Abs was infused into the patient. At day +34, the viral load decreased for the first time, to 1100 viral genome copies/mL, and the patient began new conditioning for a haploidentical rescue transplant from her mother. In light of the concurrent bone marrow aplasia, the conditioning regimen was conducted solely with ATG. At day +38, the girl received a second plasma transfusion from her mother. On the day of the second transplant (day +40), complete viral clearance was documented, and the girl received 11.3 × 106 CD34 cells/kg of mother's graft. GVHD prophylaxis was performed with post-transplant cyclophosphamide, tacrolimus, and mycophenolate mofetil. The engraftment was rapid, with a neutrophil count of >500/μL achieved at day +15, and the final platelets and packed red blood cells were transfused at day +20. Since HCMV viremia remained negative during the entire post-transplant period, we discarded the use of the mother's virus-specific T-lymphocytes. The patient was discharged at day +27 in perfect clinical condition, with continuation of antiviral valganciclovir prophylaxis. Immunosuppressive treatment and antiviral prophylaxis were discontinued six months later, and the patient remains in perfect health 2 years after the second transplant.
Therapeutic interventions, HCMV load, and white blood cell count during the entire episode are summarized in Fig. 1. In addition, the HCMV strains present in the patient were characterized by high-throughput sequencing (HTS) of an archived (−80 °C) plasma sample from day +19. DNA was extracted from 800 μl of plasma using a QIAamp MinElute virus spin kit (QIAGEN, Crawley, UK). An aliquot of 50 μl of DNA was used to generate an HCMV-enriched sequencing library as described previously (Suárez et al., 2019). The library was loaded onto a NextSeq DNA sequencer (Illumina, San Diego, CA, USA), generating 150 bp paired-ended reads (Table 1). HCMV strain enumeration was performed by genotyping 12 hypervariable HCMV genes from the reads as described previously (Suárez et al., 2019). The detection of four genotypes of the RL13 gene indicated the presence of at least four HCMV strains (a strain being defined as a particular constellation of genotypes of the hypervariable genes analysed), each including one of these RL13 genotypes (Table 2). The data was also inspected for the presence of resistance mutations in the antiviral target genes (UL54 and UL97) using LoFreq (Wilm et al., 2012). Briefly, the reads were mapped to the genome of HCMV reference strain Merlin (GenBank accession no. AY446894.2), and non-synonymous variants known to confer resistance were identified. No resistance mutations were detected.Fig. 1 Information on viremia, white blood cell (WBC) count, total serum IgG and antiviral therapy. Data on HCMV viremia is expressed in viral genome copies/mL (right Y-axis). Application and duration of antiviral treatments are represented by colored bars (yellow, anti-HCMV-specific Ig; red, foscarnet; blue, ganciclovir/valganciclovir). Chronological data (X-axis) are represented in days after the first transplant. The plasma sample analysed by high-throughput sequencing (HTS) is represented by a filled blue circle.
Fig. 1
Table 1 Overview of HTS data.
Table 1Sample ID TRI-SCT1
Sample type Plasma
Days after transplant +19
HCMV load (genome copies/mL) 2100
Data deposition (ENA project no.)a PRJEB38333
Sequencing library (genome copies) 1440
Trimmed reads (no.) 5,033,480
HCMV reads (no.)b 936,773
HCMV reads; %b 19
Coverage depth (reads/nt)b 885
a The sequence dataset was purged of human reads and deposited in the European Nucleotide Archive (ENA).
b Reads mapped to the HCMV reference strain Merlin genome; HTS, high-throughput sequencing; nt, nucleotide.
Table 2 Genotyping of hypervariable HCMV genes.
Table 2
3 Discussion
Various factors are implicated in the outcome of HCMV infection in transplant patients, the first of which relates to the HCMV serological status of donor and recipient. The serological status of the first donor may have played an important role in this case, as it has been reported that using an HCMV-seronegative unrelated HSCT donor (D-) for a seropositive recipient (R+) results in reduced overall survival after myeloablative conditioning. In contrast, an HCMV seropositive donor is associated with improved event-free survival and reduced non-relapse mortality (Styczynski, 2018). Thus, the absence of an HCMV-specific immunological memory in the donor's cells may have placed the HCMV-positive recipient at a higher risk of HCMV reactivation and associated complications.
In relation to viral factors, the clinical course of the patient may have been influenced by two main factors: the presence of multiple HCMV strains or the presence of mutations conferring antiviral resistance. Infection with multiple HCMV strains is common after transplantation (Görzer et al., 2010), and is associated with delayed HCMV clearance during antiviral therapy, even in the absence of antiviral resistance mutations. It is also associated with worse clinical outcomes, including an increase in graft rejection and faster progression of disease. In principle, the mechanisms behind this increased pathogenicity can be viral, for example involving recombination and complementation of different strains within the host, or immunological, since control of multiple strains may be more challenging for the immune system. Overall, in clinically symptomatic immunocompromised hosts, the presence of multiple HCMV strains is more threatening than the presence of a single strain (Manuel et al., 2009).
In the present study, the initial failure of antiviral therapy could not be explained by the development of resistance mutations in the HCMV strains present in the patient. This finding is not surprising, as detection of resistance mutations is unusual during the first 6 weeks of antiviral therapy, whereas the sample analysed in this case was collected only 13 days after initiation of foscarnet treatment (Lurain and Chou, 2010; Springer et al., 2005).
In regard to the initial failure of the immunotherapy with commercial pooled Ig, this may have been due to the lack of particular Abs with neutralizing capacity against the HCMV strains present in the patient. Assuming that the donors providing the Ig had been exposed to strains circulating in Europe, the Abs may have been more suited for neutralizing HCMV strains commonly circulating in that region. However, it is possible that the strains present in the patient, who originated from Bangladesh, may have differed immunologically from strains in other regions of the world. The extent to which this is true is unknown, as current knowledge of HCMV genome variability is derived almost entirely from strains circulating in a few European countries. Therefore, it would be useful to assess the neutralizing capacity of commercial pooled Ig in order to avoid potential therapeutic failure, especially in patients from non-European countries. The clinical problem in this case was addressed successfully by using the Ab-rich plasma of the patient's mother in a kind of personalized immunotherapy, on the premise that both mother and daughter may have been exposed to the same pool of viral strains. In this HCMV-positive recipient with a severe viral reactivation, Ig obtained from a household donor afforded an efficacious treatment that led to complete viral clearance.
To the best of our knowledge, this is the first report of the successful use of HCMV strain-specific Ig therapy in an HSCT recipient. The efficacy of this approach as a means of prevention has been demonstrated previously only in mouse models involving murine cytomegalovirus (MCMV) reactivation after bone marrow transplantation (Martins et al., 2019). In this setting, mice received serum obtained either from latently infected (seropositive) donors or from seronegative donors. The transfer of immune serum protected mice from viral reactivation without affecting the development of GVHD. Remarkably, complete viral protection was obtained with a small volume (5 μL) of strain-specific immune serum. On the other hand, when specific donor antibodies were administered to mice infected with antigenically mismatched MCMV strains, they were not efficient in preventing viral reactivation.
Our case highlights the importance of monitoring HSCT recipients comprehensively, longitudinally and with high sensitivity for viral factors, including the presence of multiple strains and resistance mutations. Importantly, it also provides an affordable targeted immunological therapy in patients from countries with constrained access to alternative therapies.
4 Conclusion
HSCT is a complex and specific setting in which a recipient may rapidly develop viral reactivation and consequent life-threatening complications. Ab-rich plasma may represent a fast and feasible therapeutic option to overcome viral activity and allow successful engraftment of the donor's bone marrow.
Funding
No specific funding was received.
Consent for publication
The authors have obtained consent from the parents of the patient to publish individual patient data.
Declaration of competing interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgments
The sequencing work was supported by funding from the Medical Research Council (grant number MC_UU_12014/3 to AJD). | FOSCARNET, GANCICLOVIR | DrugsGivenReaction | CC BY | 33577809 | 19,082,624 | 2021-04 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Treatment failure'. | Post-HSCT graft failure due to refractory human cytomegalovirus successfully treated with haploidentical donor-derived immunoglobulins and stem cell graft infusion: A case report.
Human cytomegalovirus (HCMV) remains an important cause of transplant-related morbidity and mortality. The incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations as a result of a lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen.
We describe the case of an 8-year-old βE-thalassemic girl from Bangladesh who was seropositive for human cytomegalovirus (HCMV) and underwent hematopoietic stem cell transplantation from a HLA-matched, unrelated, HCMV-seronegative donor. Despite administering antiviral prophylaxis with commercial pooled anti-HCMV immunoglobulin (Ig) from day +1, the post-transplant course was complicated by prompt viral reactivation, and foscarnet therapy was initiated. The virus was refractory to treatment, leading rapidly to complete bone marrow failure, and targeted immunotherapy was proposed as a second-line therapy. Hypothesizing that the patient and her relatives may have been exposed to similar HCMV strains, we selected the patient's mother, who presented a high HCMV antibody titer, as the donor of virus strain-specific anti-HCMV Ig and T-lymphocytes. Complete viral clearance was achieved after two transfusions of the mother's plasma. Subsequently, the patient underwent a haploidentical rescue transplant, promptly reaching full hematological recovery.
These findings suggest that treatment with virus strain-specific Ig may offer a new therapeutic option for critically ill patients.
pmc1 Introduction
Human cytomegalovirus (HCMV) is the most significant cause of opportunistic viral infection in allogeneic hematopoietic stem cell transplantation (HSCT) and remains an important cause of transplant-related morbidity and mortality (Wagner-Drouet et al., 2019). Several studies have shown that the incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations (Ganepola et al., 2007; Zhou et al., 2009; Ozdemir et al., 2007). This is likely a consequence of delayed HCMV-specific immune reconstitution due to the lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen (Styczynski, 2018; Ljungman et al., 2014). HCMV infection can lead to pneumonia, gastroenteritis, retinitis, hepatitis, and encephalitis among HSCT recipients (Griffiths, 2012). In addition, HCMV infection of the bone marrow can result in impaired graft acceptance and, in the most extreme cases, complete graft failure (Steffens et al., 1998). Treatment of HCMV disease essentially consists of administering antiviral drugs, namely ganciclovir and foscarnet, and intravenous immunoglobulin. Although chemoprophylaxis is not generally recommended in HSCT, immunoglobulin, whether pooled or HCMV-specific, have shown some efficacy in preventing recurrent HCMV infections (Boeckh and Ljungman, 2009).
We describe a case of a pediatric HSCT recipient who had complications of HCMV recurrence and was treated successfully with targeted immunological therapy, using haploidentical donor-derived immunoglobulin (Ig).
2 Case presentation
An 8-year-old girl was diagnosed at the age of two in her country of origin (Bangladesh) as βE-thalassemic. She was treated with monthly blood transfusions and weekly iron chelation with deferoxamine, plus supplementation with folic acid. A suboptimal regimen of chelation therapy led to severe systemic siderosis, and her family travelled to Italy to undergo HSCT. Since a human leukocyte antigen (HLA)-matched related donor was unavailable at that time, an HLA-matched (10/10) unrelated donor was identified. The patient was HCMV-seropositive, whereas the donor was HCMV-seronegative, thus posing a high-risk serological combination (D-/R+) for post-transplant HCMV infection and associated complications.
The conditioning regimen consisted of fludarabine, busulfan, thiotepa, and anti-thymocyte globulin (ATG). The infused donor's bone marrow contained 3.79 × 108 total nuclear cells/kg recipient weight, and the vitality of the cells contained in the graft was 96.9%. Tacrolimus and mycophenolate mofetil were used for graft-versus-host disease (GVHD) prophylaxis. Anti-HCMV prophylaxis commenced at day +1 with commercial pooled anti-HCMV Ig (Megalotect, Biotest Pharma GmbH, Germany) at a dose of 100 U/kg twice a week.
The initial post-transplant period was uneventful. The first occurrence of HCMV viremia (600 viral genome copies/mL) was detected at day +14, and antiviral therapy with foscarnet was started immediately. At day +17, neutrophil engraftment was achieved, but not the platelet and the erythrocyte counterparts. At day +20, the patient presented fever without clinical symptoms and without an increase of C-reactive protein. Afterwards, a full blood count revealed a progressive drop in leukocytes and platelets, whereas HCMV viremia continued to increase despite antiviral therapy. The patient also developed a severe, HCMV-related hemorrhagic cystitis, which required frequent platelet transfusions, initially once daily for 5 days, and thereafter twice daily for 7 days.
As the infection appeared unresponsive to foscarnet, ganciclovir treatment was started at day +25. This switch in antiviral therapy followed an increase in viral load, reaching a maximum titer of 5100 viral genome copies/mL at day +30. Simultaneously, the clinical condition of the patient continued to deteriorate, and the leukocyte count decreased drastically. Therefore, the diagnosis of secondary graft failure due to HCMV infection was established. Because of the failure of anti-HCMV treatment, ganciclovir was withdrawn, and foscarnet therapy was resumed. The possible explanations for treatment failure include the presence of antiviral resistance mutations or an ineffective activity of the Ig administered. As the first option seemed improbable because resistance mutations usually arise later than 6 weeks after HSCT, we opted to address the latter option. Therefore, we considered using antibodies (Abs) that were more likely to inhibit the HCMV strains causing the infection in the patient. Although the diversity of circulating HCMV strains is known to be extensive (Puchhammer-Stöckl and Görzer, 2011), we hypothesized that the patient and her relatives may have been exposed to similar strains on a familial basis. As a result, we performed serological tests on the patient's mother, father and brother. We selected the patient's mother, who presented the highest anti-HCMV IgG titer, as the best donor of strain-specific HCMV Ab-rich plasma and virus-specific T-lymphocytes in this situation. At day +32, the patient's mother underwent leukapheresis. The lymphocytes were sent immediately to a cell factory to produce specific anti-HCMV T-lymphocytes. On the same day, the plasma enriched in HCMV strain-specific Abs was infused into the patient. At day +34, the viral load decreased for the first time, to 1100 viral genome copies/mL, and the patient began new conditioning for a haploidentical rescue transplant from her mother. In light of the concurrent bone marrow aplasia, the conditioning regimen was conducted solely with ATG. At day +38, the girl received a second plasma transfusion from her mother. On the day of the second transplant (day +40), complete viral clearance was documented, and the girl received 11.3 × 106 CD34 cells/kg of mother's graft. GVHD prophylaxis was performed with post-transplant cyclophosphamide, tacrolimus, and mycophenolate mofetil. The engraftment was rapid, with a neutrophil count of >500/μL achieved at day +15, and the final platelets and packed red blood cells were transfused at day +20. Since HCMV viremia remained negative during the entire post-transplant period, we discarded the use of the mother's virus-specific T-lymphocytes. The patient was discharged at day +27 in perfect clinical condition, with continuation of antiviral valganciclovir prophylaxis. Immunosuppressive treatment and antiviral prophylaxis were discontinued six months later, and the patient remains in perfect health 2 years after the second transplant.
Therapeutic interventions, HCMV load, and white blood cell count during the entire episode are summarized in Fig. 1. In addition, the HCMV strains present in the patient were characterized by high-throughput sequencing (HTS) of an archived (−80 °C) plasma sample from day +19. DNA was extracted from 800 μl of plasma using a QIAamp MinElute virus spin kit (QIAGEN, Crawley, UK). An aliquot of 50 μl of DNA was used to generate an HCMV-enriched sequencing library as described previously (Suárez et al., 2019). The library was loaded onto a NextSeq DNA sequencer (Illumina, San Diego, CA, USA), generating 150 bp paired-ended reads (Table 1). HCMV strain enumeration was performed by genotyping 12 hypervariable HCMV genes from the reads as described previously (Suárez et al., 2019). The detection of four genotypes of the RL13 gene indicated the presence of at least four HCMV strains (a strain being defined as a particular constellation of genotypes of the hypervariable genes analysed), each including one of these RL13 genotypes (Table 2). The data was also inspected for the presence of resistance mutations in the antiviral target genes (UL54 and UL97) using LoFreq (Wilm et al., 2012). Briefly, the reads were mapped to the genome of HCMV reference strain Merlin (GenBank accession no. AY446894.2), and non-synonymous variants known to confer resistance were identified. No resistance mutations were detected.Fig. 1 Information on viremia, white blood cell (WBC) count, total serum IgG and antiviral therapy. Data on HCMV viremia is expressed in viral genome copies/mL (right Y-axis). Application and duration of antiviral treatments are represented by colored bars (yellow, anti-HCMV-specific Ig; red, foscarnet; blue, ganciclovir/valganciclovir). Chronological data (X-axis) are represented in days after the first transplant. The plasma sample analysed by high-throughput sequencing (HTS) is represented by a filled blue circle.
Fig. 1
Table 1 Overview of HTS data.
Table 1Sample ID TRI-SCT1
Sample type Plasma
Days after transplant +19
HCMV load (genome copies/mL) 2100
Data deposition (ENA project no.)a PRJEB38333
Sequencing library (genome copies) 1440
Trimmed reads (no.) 5,033,480
HCMV reads (no.)b 936,773
HCMV reads; %b 19
Coverage depth (reads/nt)b 885
a The sequence dataset was purged of human reads and deposited in the European Nucleotide Archive (ENA).
b Reads mapped to the HCMV reference strain Merlin genome; HTS, high-throughput sequencing; nt, nucleotide.
Table 2 Genotyping of hypervariable HCMV genes.
Table 2
3 Discussion
Various factors are implicated in the outcome of HCMV infection in transplant patients, the first of which relates to the HCMV serological status of donor and recipient. The serological status of the first donor may have played an important role in this case, as it has been reported that using an HCMV-seronegative unrelated HSCT donor (D-) for a seropositive recipient (R+) results in reduced overall survival after myeloablative conditioning. In contrast, an HCMV seropositive donor is associated with improved event-free survival and reduced non-relapse mortality (Styczynski, 2018). Thus, the absence of an HCMV-specific immunological memory in the donor's cells may have placed the HCMV-positive recipient at a higher risk of HCMV reactivation and associated complications.
In relation to viral factors, the clinical course of the patient may have been influenced by two main factors: the presence of multiple HCMV strains or the presence of mutations conferring antiviral resistance. Infection with multiple HCMV strains is common after transplantation (Görzer et al., 2010), and is associated with delayed HCMV clearance during antiviral therapy, even in the absence of antiviral resistance mutations. It is also associated with worse clinical outcomes, including an increase in graft rejection and faster progression of disease. In principle, the mechanisms behind this increased pathogenicity can be viral, for example involving recombination and complementation of different strains within the host, or immunological, since control of multiple strains may be more challenging for the immune system. Overall, in clinically symptomatic immunocompromised hosts, the presence of multiple HCMV strains is more threatening than the presence of a single strain (Manuel et al., 2009).
In the present study, the initial failure of antiviral therapy could not be explained by the development of resistance mutations in the HCMV strains present in the patient. This finding is not surprising, as detection of resistance mutations is unusual during the first 6 weeks of antiviral therapy, whereas the sample analysed in this case was collected only 13 days after initiation of foscarnet treatment (Lurain and Chou, 2010; Springer et al., 2005).
In regard to the initial failure of the immunotherapy with commercial pooled Ig, this may have been due to the lack of particular Abs with neutralizing capacity against the HCMV strains present in the patient. Assuming that the donors providing the Ig had been exposed to strains circulating in Europe, the Abs may have been more suited for neutralizing HCMV strains commonly circulating in that region. However, it is possible that the strains present in the patient, who originated from Bangladesh, may have differed immunologically from strains in other regions of the world. The extent to which this is true is unknown, as current knowledge of HCMV genome variability is derived almost entirely from strains circulating in a few European countries. Therefore, it would be useful to assess the neutralizing capacity of commercial pooled Ig in order to avoid potential therapeutic failure, especially in patients from non-European countries. The clinical problem in this case was addressed successfully by using the Ab-rich plasma of the patient's mother in a kind of personalized immunotherapy, on the premise that both mother and daughter may have been exposed to the same pool of viral strains. In this HCMV-positive recipient with a severe viral reactivation, Ig obtained from a household donor afforded an efficacious treatment that led to complete viral clearance.
To the best of our knowledge, this is the first report of the successful use of HCMV strain-specific Ig therapy in an HSCT recipient. The efficacy of this approach as a means of prevention has been demonstrated previously only in mouse models involving murine cytomegalovirus (MCMV) reactivation after bone marrow transplantation (Martins et al., 2019). In this setting, mice received serum obtained either from latently infected (seropositive) donors or from seronegative donors. The transfer of immune serum protected mice from viral reactivation without affecting the development of GVHD. Remarkably, complete viral protection was obtained with a small volume (5 μL) of strain-specific immune serum. On the other hand, when specific donor antibodies were administered to mice infected with antigenically mismatched MCMV strains, they were not efficient in preventing viral reactivation.
Our case highlights the importance of monitoring HSCT recipients comprehensively, longitudinally and with high sensitivity for viral factors, including the presence of multiple strains and resistance mutations. Importantly, it also provides an affordable targeted immunological therapy in patients from countries with constrained access to alternative therapies.
4 Conclusion
HSCT is a complex and specific setting in which a recipient may rapidly develop viral reactivation and consequent life-threatening complications. Ab-rich plasma may represent a fast and feasible therapeutic option to overcome viral activity and allow successful engraftment of the donor's bone marrow.
Funding
No specific funding was received.
Consent for publication
The authors have obtained consent from the parents of the patient to publish individual patient data.
Declaration of competing interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgments
The sequencing work was supported by funding from the Medical Research Council (grant number MC_UU_12014/3 to AJD). | BUSULFAN, FLUDARABINE PHOSPHATE, FOSCARNET, GANCICLOVIR, MYCOPHENOLATE MOFETIL, TACROLIMUS, THIOTEPA, THYMOCYTE IMMUNE GLOBULIN NOS | DrugsGivenReaction | CC BY | 33577809 | 19,141,715 | 2021-04 |
What was the dosage of drug 'BUSULFAN'? | Post-HSCT graft failure due to refractory human cytomegalovirus successfully treated with haploidentical donor-derived immunoglobulins and stem cell graft infusion: A case report.
Human cytomegalovirus (HCMV) remains an important cause of transplant-related morbidity and mortality. The incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations as a result of a lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen.
We describe the case of an 8-year-old βE-thalassemic girl from Bangladesh who was seropositive for human cytomegalovirus (HCMV) and underwent hematopoietic stem cell transplantation from a HLA-matched, unrelated, HCMV-seronegative donor. Despite administering antiviral prophylaxis with commercial pooled anti-HCMV immunoglobulin (Ig) from day +1, the post-transplant course was complicated by prompt viral reactivation, and foscarnet therapy was initiated. The virus was refractory to treatment, leading rapidly to complete bone marrow failure, and targeted immunotherapy was proposed as a second-line therapy. Hypothesizing that the patient and her relatives may have been exposed to similar HCMV strains, we selected the patient's mother, who presented a high HCMV antibody titer, as the donor of virus strain-specific anti-HCMV Ig and T-lymphocytes. Complete viral clearance was achieved after two transfusions of the mother's plasma. Subsequently, the patient underwent a haploidentical rescue transplant, promptly reaching full hematological recovery.
These findings suggest that treatment with virus strain-specific Ig may offer a new therapeutic option for critically ill patients.
pmc1 Introduction
Human cytomegalovirus (HCMV) is the most significant cause of opportunistic viral infection in allogeneic hematopoietic stem cell transplantation (HSCT) and remains an important cause of transplant-related morbidity and mortality (Wagner-Drouet et al., 2019). Several studies have shown that the incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations (Ganepola et al., 2007; Zhou et al., 2009; Ozdemir et al., 2007). This is likely a consequence of delayed HCMV-specific immune reconstitution due to the lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen (Styczynski, 2018; Ljungman et al., 2014). HCMV infection can lead to pneumonia, gastroenteritis, retinitis, hepatitis, and encephalitis among HSCT recipients (Griffiths, 2012). In addition, HCMV infection of the bone marrow can result in impaired graft acceptance and, in the most extreme cases, complete graft failure (Steffens et al., 1998). Treatment of HCMV disease essentially consists of administering antiviral drugs, namely ganciclovir and foscarnet, and intravenous immunoglobulin. Although chemoprophylaxis is not generally recommended in HSCT, immunoglobulin, whether pooled or HCMV-specific, have shown some efficacy in preventing recurrent HCMV infections (Boeckh and Ljungman, 2009).
We describe a case of a pediatric HSCT recipient who had complications of HCMV recurrence and was treated successfully with targeted immunological therapy, using haploidentical donor-derived immunoglobulin (Ig).
2 Case presentation
An 8-year-old girl was diagnosed at the age of two in her country of origin (Bangladesh) as βE-thalassemic. She was treated with monthly blood transfusions and weekly iron chelation with deferoxamine, plus supplementation with folic acid. A suboptimal regimen of chelation therapy led to severe systemic siderosis, and her family travelled to Italy to undergo HSCT. Since a human leukocyte antigen (HLA)-matched related donor was unavailable at that time, an HLA-matched (10/10) unrelated donor was identified. The patient was HCMV-seropositive, whereas the donor was HCMV-seronegative, thus posing a high-risk serological combination (D-/R+) for post-transplant HCMV infection and associated complications.
The conditioning regimen consisted of fludarabine, busulfan, thiotepa, and anti-thymocyte globulin (ATG). The infused donor's bone marrow contained 3.79 × 108 total nuclear cells/kg recipient weight, and the vitality of the cells contained in the graft was 96.9%. Tacrolimus and mycophenolate mofetil were used for graft-versus-host disease (GVHD) prophylaxis. Anti-HCMV prophylaxis commenced at day +1 with commercial pooled anti-HCMV Ig (Megalotect, Biotest Pharma GmbH, Germany) at a dose of 100 U/kg twice a week.
The initial post-transplant period was uneventful. The first occurrence of HCMV viremia (600 viral genome copies/mL) was detected at day +14, and antiviral therapy with foscarnet was started immediately. At day +17, neutrophil engraftment was achieved, but not the platelet and the erythrocyte counterparts. At day +20, the patient presented fever without clinical symptoms and without an increase of C-reactive protein. Afterwards, a full blood count revealed a progressive drop in leukocytes and platelets, whereas HCMV viremia continued to increase despite antiviral therapy. The patient also developed a severe, HCMV-related hemorrhagic cystitis, which required frequent platelet transfusions, initially once daily for 5 days, and thereafter twice daily for 7 days.
As the infection appeared unresponsive to foscarnet, ganciclovir treatment was started at day +25. This switch in antiviral therapy followed an increase in viral load, reaching a maximum titer of 5100 viral genome copies/mL at day +30. Simultaneously, the clinical condition of the patient continued to deteriorate, and the leukocyte count decreased drastically. Therefore, the diagnosis of secondary graft failure due to HCMV infection was established. Because of the failure of anti-HCMV treatment, ganciclovir was withdrawn, and foscarnet therapy was resumed. The possible explanations for treatment failure include the presence of antiviral resistance mutations or an ineffective activity of the Ig administered. As the first option seemed improbable because resistance mutations usually arise later than 6 weeks after HSCT, we opted to address the latter option. Therefore, we considered using antibodies (Abs) that were more likely to inhibit the HCMV strains causing the infection in the patient. Although the diversity of circulating HCMV strains is known to be extensive (Puchhammer-Stöckl and Görzer, 2011), we hypothesized that the patient and her relatives may have been exposed to similar strains on a familial basis. As a result, we performed serological tests on the patient's mother, father and brother. We selected the patient's mother, who presented the highest anti-HCMV IgG titer, as the best donor of strain-specific HCMV Ab-rich plasma and virus-specific T-lymphocytes in this situation. At day +32, the patient's mother underwent leukapheresis. The lymphocytes were sent immediately to a cell factory to produce specific anti-HCMV T-lymphocytes. On the same day, the plasma enriched in HCMV strain-specific Abs was infused into the patient. At day +34, the viral load decreased for the first time, to 1100 viral genome copies/mL, and the patient began new conditioning for a haploidentical rescue transplant from her mother. In light of the concurrent bone marrow aplasia, the conditioning regimen was conducted solely with ATG. At day +38, the girl received a second plasma transfusion from her mother. On the day of the second transplant (day +40), complete viral clearance was documented, and the girl received 11.3 × 106 CD34 cells/kg of mother's graft. GVHD prophylaxis was performed with post-transplant cyclophosphamide, tacrolimus, and mycophenolate mofetil. The engraftment was rapid, with a neutrophil count of >500/μL achieved at day +15, and the final platelets and packed red blood cells were transfused at day +20. Since HCMV viremia remained negative during the entire post-transplant period, we discarded the use of the mother's virus-specific T-lymphocytes. The patient was discharged at day +27 in perfect clinical condition, with continuation of antiviral valganciclovir prophylaxis. Immunosuppressive treatment and antiviral prophylaxis were discontinued six months later, and the patient remains in perfect health 2 years after the second transplant.
Therapeutic interventions, HCMV load, and white blood cell count during the entire episode are summarized in Fig. 1. In addition, the HCMV strains present in the patient were characterized by high-throughput sequencing (HTS) of an archived (−80 °C) plasma sample from day +19. DNA was extracted from 800 μl of plasma using a QIAamp MinElute virus spin kit (QIAGEN, Crawley, UK). An aliquot of 50 μl of DNA was used to generate an HCMV-enriched sequencing library as described previously (Suárez et al., 2019). The library was loaded onto a NextSeq DNA sequencer (Illumina, San Diego, CA, USA), generating 150 bp paired-ended reads (Table 1). HCMV strain enumeration was performed by genotyping 12 hypervariable HCMV genes from the reads as described previously (Suárez et al., 2019). The detection of four genotypes of the RL13 gene indicated the presence of at least four HCMV strains (a strain being defined as a particular constellation of genotypes of the hypervariable genes analysed), each including one of these RL13 genotypes (Table 2). The data was also inspected for the presence of resistance mutations in the antiviral target genes (UL54 and UL97) using LoFreq (Wilm et al., 2012). Briefly, the reads were mapped to the genome of HCMV reference strain Merlin (GenBank accession no. AY446894.2), and non-synonymous variants known to confer resistance were identified. No resistance mutations were detected.Fig. 1 Information on viremia, white blood cell (WBC) count, total serum IgG and antiviral therapy. Data on HCMV viremia is expressed in viral genome copies/mL (right Y-axis). Application and duration of antiviral treatments are represented by colored bars (yellow, anti-HCMV-specific Ig; red, foscarnet; blue, ganciclovir/valganciclovir). Chronological data (X-axis) are represented in days after the first transplant. The plasma sample analysed by high-throughput sequencing (HTS) is represented by a filled blue circle.
Fig. 1
Table 1 Overview of HTS data.
Table 1Sample ID TRI-SCT1
Sample type Plasma
Days after transplant +19
HCMV load (genome copies/mL) 2100
Data deposition (ENA project no.)a PRJEB38333
Sequencing library (genome copies) 1440
Trimmed reads (no.) 5,033,480
HCMV reads (no.)b 936,773
HCMV reads; %b 19
Coverage depth (reads/nt)b 885
a The sequence dataset was purged of human reads and deposited in the European Nucleotide Archive (ENA).
b Reads mapped to the HCMV reference strain Merlin genome; HTS, high-throughput sequencing; nt, nucleotide.
Table 2 Genotyping of hypervariable HCMV genes.
Table 2
3 Discussion
Various factors are implicated in the outcome of HCMV infection in transplant patients, the first of which relates to the HCMV serological status of donor and recipient. The serological status of the first donor may have played an important role in this case, as it has been reported that using an HCMV-seronegative unrelated HSCT donor (D-) for a seropositive recipient (R+) results in reduced overall survival after myeloablative conditioning. In contrast, an HCMV seropositive donor is associated with improved event-free survival and reduced non-relapse mortality (Styczynski, 2018). Thus, the absence of an HCMV-specific immunological memory in the donor's cells may have placed the HCMV-positive recipient at a higher risk of HCMV reactivation and associated complications.
In relation to viral factors, the clinical course of the patient may have been influenced by two main factors: the presence of multiple HCMV strains or the presence of mutations conferring antiviral resistance. Infection with multiple HCMV strains is common after transplantation (Görzer et al., 2010), and is associated with delayed HCMV clearance during antiviral therapy, even in the absence of antiviral resistance mutations. It is also associated with worse clinical outcomes, including an increase in graft rejection and faster progression of disease. In principle, the mechanisms behind this increased pathogenicity can be viral, for example involving recombination and complementation of different strains within the host, or immunological, since control of multiple strains may be more challenging for the immune system. Overall, in clinically symptomatic immunocompromised hosts, the presence of multiple HCMV strains is more threatening than the presence of a single strain (Manuel et al., 2009).
In the present study, the initial failure of antiviral therapy could not be explained by the development of resistance mutations in the HCMV strains present in the patient. This finding is not surprising, as detection of resistance mutations is unusual during the first 6 weeks of antiviral therapy, whereas the sample analysed in this case was collected only 13 days after initiation of foscarnet treatment (Lurain and Chou, 2010; Springer et al., 2005).
In regard to the initial failure of the immunotherapy with commercial pooled Ig, this may have been due to the lack of particular Abs with neutralizing capacity against the HCMV strains present in the patient. Assuming that the donors providing the Ig had been exposed to strains circulating in Europe, the Abs may have been more suited for neutralizing HCMV strains commonly circulating in that region. However, it is possible that the strains present in the patient, who originated from Bangladesh, may have differed immunologically from strains in other regions of the world. The extent to which this is true is unknown, as current knowledge of HCMV genome variability is derived almost entirely from strains circulating in a few European countries. Therefore, it would be useful to assess the neutralizing capacity of commercial pooled Ig in order to avoid potential therapeutic failure, especially in patients from non-European countries. The clinical problem in this case was addressed successfully by using the Ab-rich plasma of the patient's mother in a kind of personalized immunotherapy, on the premise that both mother and daughter may have been exposed to the same pool of viral strains. In this HCMV-positive recipient with a severe viral reactivation, Ig obtained from a household donor afforded an efficacious treatment that led to complete viral clearance.
To the best of our knowledge, this is the first report of the successful use of HCMV strain-specific Ig therapy in an HSCT recipient. The efficacy of this approach as a means of prevention has been demonstrated previously only in mouse models involving murine cytomegalovirus (MCMV) reactivation after bone marrow transplantation (Martins et al., 2019). In this setting, mice received serum obtained either from latently infected (seropositive) donors or from seronegative donors. The transfer of immune serum protected mice from viral reactivation without affecting the development of GVHD. Remarkably, complete viral protection was obtained with a small volume (5 μL) of strain-specific immune serum. On the other hand, when specific donor antibodies were administered to mice infected with antigenically mismatched MCMV strains, they were not efficient in preventing viral reactivation.
Our case highlights the importance of monitoring HSCT recipients comprehensively, longitudinally and with high sensitivity for viral factors, including the presence of multiple strains and resistance mutations. Importantly, it also provides an affordable targeted immunological therapy in patients from countries with constrained access to alternative therapies.
4 Conclusion
HSCT is a complex and specific setting in which a recipient may rapidly develop viral reactivation and consequent life-threatening complications. Ab-rich plasma may represent a fast and feasible therapeutic option to overcome viral activity and allow successful engraftment of the donor's bone marrow.
Funding
No specific funding was received.
Consent for publication
The authors have obtained consent from the parents of the patient to publish individual patient data.
Declaration of competing interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgments
The sequencing work was supported by funding from the Medical Research Council (grant number MC_UU_12014/3 to AJD). | UNK UNK, UNKNOWN FREQ. | DrugDosageText | CC BY | 33577809 | 19,010,959 | 2021-04 |
What was the dosage of drug 'FLUDARABINE PHOSPHATE'? | Post-HSCT graft failure due to refractory human cytomegalovirus successfully treated with haploidentical donor-derived immunoglobulins and stem cell graft infusion: A case report.
Human cytomegalovirus (HCMV) remains an important cause of transplant-related morbidity and mortality. The incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations as a result of a lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen.
We describe the case of an 8-year-old βE-thalassemic girl from Bangladesh who was seropositive for human cytomegalovirus (HCMV) and underwent hematopoietic stem cell transplantation from a HLA-matched, unrelated, HCMV-seronegative donor. Despite administering antiviral prophylaxis with commercial pooled anti-HCMV immunoglobulin (Ig) from day +1, the post-transplant course was complicated by prompt viral reactivation, and foscarnet therapy was initiated. The virus was refractory to treatment, leading rapidly to complete bone marrow failure, and targeted immunotherapy was proposed as a second-line therapy. Hypothesizing that the patient and her relatives may have been exposed to similar HCMV strains, we selected the patient's mother, who presented a high HCMV antibody titer, as the donor of virus strain-specific anti-HCMV Ig and T-lymphocytes. Complete viral clearance was achieved after two transfusions of the mother's plasma. Subsequently, the patient underwent a haploidentical rescue transplant, promptly reaching full hematological recovery.
These findings suggest that treatment with virus strain-specific Ig may offer a new therapeutic option for critically ill patients.
pmc1 Introduction
Human cytomegalovirus (HCMV) is the most significant cause of opportunistic viral infection in allogeneic hematopoietic stem cell transplantation (HSCT) and remains an important cause of transplant-related morbidity and mortality (Wagner-Drouet et al., 2019). Several studies have shown that the incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations (Ganepola et al., 2007; Zhou et al., 2009; Ozdemir et al., 2007). This is likely a consequence of delayed HCMV-specific immune reconstitution due to the lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen (Styczynski, 2018; Ljungman et al., 2014). HCMV infection can lead to pneumonia, gastroenteritis, retinitis, hepatitis, and encephalitis among HSCT recipients (Griffiths, 2012). In addition, HCMV infection of the bone marrow can result in impaired graft acceptance and, in the most extreme cases, complete graft failure (Steffens et al., 1998). Treatment of HCMV disease essentially consists of administering antiviral drugs, namely ganciclovir and foscarnet, and intravenous immunoglobulin. Although chemoprophylaxis is not generally recommended in HSCT, immunoglobulin, whether pooled or HCMV-specific, have shown some efficacy in preventing recurrent HCMV infections (Boeckh and Ljungman, 2009).
We describe a case of a pediatric HSCT recipient who had complications of HCMV recurrence and was treated successfully with targeted immunological therapy, using haploidentical donor-derived immunoglobulin (Ig).
2 Case presentation
An 8-year-old girl was diagnosed at the age of two in her country of origin (Bangladesh) as βE-thalassemic. She was treated with monthly blood transfusions and weekly iron chelation with deferoxamine, plus supplementation with folic acid. A suboptimal regimen of chelation therapy led to severe systemic siderosis, and her family travelled to Italy to undergo HSCT. Since a human leukocyte antigen (HLA)-matched related donor was unavailable at that time, an HLA-matched (10/10) unrelated donor was identified. The patient was HCMV-seropositive, whereas the donor was HCMV-seronegative, thus posing a high-risk serological combination (D-/R+) for post-transplant HCMV infection and associated complications.
The conditioning regimen consisted of fludarabine, busulfan, thiotepa, and anti-thymocyte globulin (ATG). The infused donor's bone marrow contained 3.79 × 108 total nuclear cells/kg recipient weight, and the vitality of the cells contained in the graft was 96.9%. Tacrolimus and mycophenolate mofetil were used for graft-versus-host disease (GVHD) prophylaxis. Anti-HCMV prophylaxis commenced at day +1 with commercial pooled anti-HCMV Ig (Megalotect, Biotest Pharma GmbH, Germany) at a dose of 100 U/kg twice a week.
The initial post-transplant period was uneventful. The first occurrence of HCMV viremia (600 viral genome copies/mL) was detected at day +14, and antiviral therapy with foscarnet was started immediately. At day +17, neutrophil engraftment was achieved, but not the platelet and the erythrocyte counterparts. At day +20, the patient presented fever without clinical symptoms and without an increase of C-reactive protein. Afterwards, a full blood count revealed a progressive drop in leukocytes and platelets, whereas HCMV viremia continued to increase despite antiviral therapy. The patient also developed a severe, HCMV-related hemorrhagic cystitis, which required frequent platelet transfusions, initially once daily for 5 days, and thereafter twice daily for 7 days.
As the infection appeared unresponsive to foscarnet, ganciclovir treatment was started at day +25. This switch in antiviral therapy followed an increase in viral load, reaching a maximum titer of 5100 viral genome copies/mL at day +30. Simultaneously, the clinical condition of the patient continued to deteriorate, and the leukocyte count decreased drastically. Therefore, the diagnosis of secondary graft failure due to HCMV infection was established. Because of the failure of anti-HCMV treatment, ganciclovir was withdrawn, and foscarnet therapy was resumed. The possible explanations for treatment failure include the presence of antiviral resistance mutations or an ineffective activity of the Ig administered. As the first option seemed improbable because resistance mutations usually arise later than 6 weeks after HSCT, we opted to address the latter option. Therefore, we considered using antibodies (Abs) that were more likely to inhibit the HCMV strains causing the infection in the patient. Although the diversity of circulating HCMV strains is known to be extensive (Puchhammer-Stöckl and Görzer, 2011), we hypothesized that the patient and her relatives may have been exposed to similar strains on a familial basis. As a result, we performed serological tests on the patient's mother, father and brother. We selected the patient's mother, who presented the highest anti-HCMV IgG titer, as the best donor of strain-specific HCMV Ab-rich plasma and virus-specific T-lymphocytes in this situation. At day +32, the patient's mother underwent leukapheresis. The lymphocytes were sent immediately to a cell factory to produce specific anti-HCMV T-lymphocytes. On the same day, the plasma enriched in HCMV strain-specific Abs was infused into the patient. At day +34, the viral load decreased for the first time, to 1100 viral genome copies/mL, and the patient began new conditioning for a haploidentical rescue transplant from her mother. In light of the concurrent bone marrow aplasia, the conditioning regimen was conducted solely with ATG. At day +38, the girl received a second plasma transfusion from her mother. On the day of the second transplant (day +40), complete viral clearance was documented, and the girl received 11.3 × 106 CD34 cells/kg of mother's graft. GVHD prophylaxis was performed with post-transplant cyclophosphamide, tacrolimus, and mycophenolate mofetil. The engraftment was rapid, with a neutrophil count of >500/μL achieved at day +15, and the final platelets and packed red blood cells were transfused at day +20. Since HCMV viremia remained negative during the entire post-transplant period, we discarded the use of the mother's virus-specific T-lymphocytes. The patient was discharged at day +27 in perfect clinical condition, with continuation of antiviral valganciclovir prophylaxis. Immunosuppressive treatment and antiviral prophylaxis were discontinued six months later, and the patient remains in perfect health 2 years after the second transplant.
Therapeutic interventions, HCMV load, and white blood cell count during the entire episode are summarized in Fig. 1. In addition, the HCMV strains present in the patient were characterized by high-throughput sequencing (HTS) of an archived (−80 °C) plasma sample from day +19. DNA was extracted from 800 μl of plasma using a QIAamp MinElute virus spin kit (QIAGEN, Crawley, UK). An aliquot of 50 μl of DNA was used to generate an HCMV-enriched sequencing library as described previously (Suárez et al., 2019). The library was loaded onto a NextSeq DNA sequencer (Illumina, San Diego, CA, USA), generating 150 bp paired-ended reads (Table 1). HCMV strain enumeration was performed by genotyping 12 hypervariable HCMV genes from the reads as described previously (Suárez et al., 2019). The detection of four genotypes of the RL13 gene indicated the presence of at least four HCMV strains (a strain being defined as a particular constellation of genotypes of the hypervariable genes analysed), each including one of these RL13 genotypes (Table 2). The data was also inspected for the presence of resistance mutations in the antiviral target genes (UL54 and UL97) using LoFreq (Wilm et al., 2012). Briefly, the reads were mapped to the genome of HCMV reference strain Merlin (GenBank accession no. AY446894.2), and non-synonymous variants known to confer resistance were identified. No resistance mutations were detected.Fig. 1 Information on viremia, white blood cell (WBC) count, total serum IgG and antiviral therapy. Data on HCMV viremia is expressed in viral genome copies/mL (right Y-axis). Application and duration of antiviral treatments are represented by colored bars (yellow, anti-HCMV-specific Ig; red, foscarnet; blue, ganciclovir/valganciclovir). Chronological data (X-axis) are represented in days after the first transplant. The plasma sample analysed by high-throughput sequencing (HTS) is represented by a filled blue circle.
Fig. 1
Table 1 Overview of HTS data.
Table 1Sample ID TRI-SCT1
Sample type Plasma
Days after transplant +19
HCMV load (genome copies/mL) 2100
Data deposition (ENA project no.)a PRJEB38333
Sequencing library (genome copies) 1440
Trimmed reads (no.) 5,033,480
HCMV reads (no.)b 936,773
HCMV reads; %b 19
Coverage depth (reads/nt)b 885
a The sequence dataset was purged of human reads and deposited in the European Nucleotide Archive (ENA).
b Reads mapped to the HCMV reference strain Merlin genome; HTS, high-throughput sequencing; nt, nucleotide.
Table 2 Genotyping of hypervariable HCMV genes.
Table 2
3 Discussion
Various factors are implicated in the outcome of HCMV infection in transplant patients, the first of which relates to the HCMV serological status of donor and recipient. The serological status of the first donor may have played an important role in this case, as it has been reported that using an HCMV-seronegative unrelated HSCT donor (D-) for a seropositive recipient (R+) results in reduced overall survival after myeloablative conditioning. In contrast, an HCMV seropositive donor is associated with improved event-free survival and reduced non-relapse mortality (Styczynski, 2018). Thus, the absence of an HCMV-specific immunological memory in the donor's cells may have placed the HCMV-positive recipient at a higher risk of HCMV reactivation and associated complications.
In relation to viral factors, the clinical course of the patient may have been influenced by two main factors: the presence of multiple HCMV strains or the presence of mutations conferring antiviral resistance. Infection with multiple HCMV strains is common after transplantation (Görzer et al., 2010), and is associated with delayed HCMV clearance during antiviral therapy, even in the absence of antiviral resistance mutations. It is also associated with worse clinical outcomes, including an increase in graft rejection and faster progression of disease. In principle, the mechanisms behind this increased pathogenicity can be viral, for example involving recombination and complementation of different strains within the host, or immunological, since control of multiple strains may be more challenging for the immune system. Overall, in clinically symptomatic immunocompromised hosts, the presence of multiple HCMV strains is more threatening than the presence of a single strain (Manuel et al., 2009).
In the present study, the initial failure of antiviral therapy could not be explained by the development of resistance mutations in the HCMV strains present in the patient. This finding is not surprising, as detection of resistance mutations is unusual during the first 6 weeks of antiviral therapy, whereas the sample analysed in this case was collected only 13 days after initiation of foscarnet treatment (Lurain and Chou, 2010; Springer et al., 2005).
In regard to the initial failure of the immunotherapy with commercial pooled Ig, this may have been due to the lack of particular Abs with neutralizing capacity against the HCMV strains present in the patient. Assuming that the donors providing the Ig had been exposed to strains circulating in Europe, the Abs may have been more suited for neutralizing HCMV strains commonly circulating in that region. However, it is possible that the strains present in the patient, who originated from Bangladesh, may have differed immunologically from strains in other regions of the world. The extent to which this is true is unknown, as current knowledge of HCMV genome variability is derived almost entirely from strains circulating in a few European countries. Therefore, it would be useful to assess the neutralizing capacity of commercial pooled Ig in order to avoid potential therapeutic failure, especially in patients from non-European countries. The clinical problem in this case was addressed successfully by using the Ab-rich plasma of the patient's mother in a kind of personalized immunotherapy, on the premise that both mother and daughter may have been exposed to the same pool of viral strains. In this HCMV-positive recipient with a severe viral reactivation, Ig obtained from a household donor afforded an efficacious treatment that led to complete viral clearance.
To the best of our knowledge, this is the first report of the successful use of HCMV strain-specific Ig therapy in an HSCT recipient. The efficacy of this approach as a means of prevention has been demonstrated previously only in mouse models involving murine cytomegalovirus (MCMV) reactivation after bone marrow transplantation (Martins et al., 2019). In this setting, mice received serum obtained either from latently infected (seropositive) donors or from seronegative donors. The transfer of immune serum protected mice from viral reactivation without affecting the development of GVHD. Remarkably, complete viral protection was obtained with a small volume (5 μL) of strain-specific immune serum. On the other hand, when specific donor antibodies were administered to mice infected with antigenically mismatched MCMV strains, they were not efficient in preventing viral reactivation.
Our case highlights the importance of monitoring HSCT recipients comprehensively, longitudinally and with high sensitivity for viral factors, including the presence of multiple strains and resistance mutations. Importantly, it also provides an affordable targeted immunological therapy in patients from countries with constrained access to alternative therapies.
4 Conclusion
HSCT is a complex and specific setting in which a recipient may rapidly develop viral reactivation and consequent life-threatening complications. Ab-rich plasma may represent a fast and feasible therapeutic option to overcome viral activity and allow successful engraftment of the donor's bone marrow.
Funding
No specific funding was received.
Consent for publication
The authors have obtained consent from the parents of the patient to publish individual patient data.
Declaration of competing interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgments
The sequencing work was supported by funding from the Medical Research Council (grant number MC_UU_12014/3 to AJD). | UNK UNK, UNKNOWN FREQ. | DrugDosageText | CC BY | 33577809 | 19,010,959 | 2021-04 |
What was the dosage of drug 'LAPINE T-LYMPHOCYTE IMMUNE GLOBULIN'? | Post-HSCT graft failure due to refractory human cytomegalovirus successfully treated with haploidentical donor-derived immunoglobulins and stem cell graft infusion: A case report.
Human cytomegalovirus (HCMV) remains an important cause of transplant-related morbidity and mortality. The incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations as a result of a lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen.
We describe the case of an 8-year-old βE-thalassemic girl from Bangladesh who was seropositive for human cytomegalovirus (HCMV) and underwent hematopoietic stem cell transplantation from a HLA-matched, unrelated, HCMV-seronegative donor. Despite administering antiviral prophylaxis with commercial pooled anti-HCMV immunoglobulin (Ig) from day +1, the post-transplant course was complicated by prompt viral reactivation, and foscarnet therapy was initiated. The virus was refractory to treatment, leading rapidly to complete bone marrow failure, and targeted immunotherapy was proposed as a second-line therapy. Hypothesizing that the patient and her relatives may have been exposed to similar HCMV strains, we selected the patient's mother, who presented a high HCMV antibody titer, as the donor of virus strain-specific anti-HCMV Ig and T-lymphocytes. Complete viral clearance was achieved after two transfusions of the mother's plasma. Subsequently, the patient underwent a haploidentical rescue transplant, promptly reaching full hematological recovery.
These findings suggest that treatment with virus strain-specific Ig may offer a new therapeutic option for critically ill patients.
pmc1 Introduction
Human cytomegalovirus (HCMV) is the most significant cause of opportunistic viral infection in allogeneic hematopoietic stem cell transplantation (HSCT) and remains an important cause of transplant-related morbidity and mortality (Wagner-Drouet et al., 2019). Several studies have shown that the incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations (Ganepola et al., 2007; Zhou et al., 2009; Ozdemir et al., 2007). This is likely a consequence of delayed HCMV-specific immune reconstitution due to the lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen (Styczynski, 2018; Ljungman et al., 2014). HCMV infection can lead to pneumonia, gastroenteritis, retinitis, hepatitis, and encephalitis among HSCT recipients (Griffiths, 2012). In addition, HCMV infection of the bone marrow can result in impaired graft acceptance and, in the most extreme cases, complete graft failure (Steffens et al., 1998). Treatment of HCMV disease essentially consists of administering antiviral drugs, namely ganciclovir and foscarnet, and intravenous immunoglobulin. Although chemoprophylaxis is not generally recommended in HSCT, immunoglobulin, whether pooled or HCMV-specific, have shown some efficacy in preventing recurrent HCMV infections (Boeckh and Ljungman, 2009).
We describe a case of a pediatric HSCT recipient who had complications of HCMV recurrence and was treated successfully with targeted immunological therapy, using haploidentical donor-derived immunoglobulin (Ig).
2 Case presentation
An 8-year-old girl was diagnosed at the age of two in her country of origin (Bangladesh) as βE-thalassemic. She was treated with monthly blood transfusions and weekly iron chelation with deferoxamine, plus supplementation with folic acid. A suboptimal regimen of chelation therapy led to severe systemic siderosis, and her family travelled to Italy to undergo HSCT. Since a human leukocyte antigen (HLA)-matched related donor was unavailable at that time, an HLA-matched (10/10) unrelated donor was identified. The patient was HCMV-seropositive, whereas the donor was HCMV-seronegative, thus posing a high-risk serological combination (D-/R+) for post-transplant HCMV infection and associated complications.
The conditioning regimen consisted of fludarabine, busulfan, thiotepa, and anti-thymocyte globulin (ATG). The infused donor's bone marrow contained 3.79 × 108 total nuclear cells/kg recipient weight, and the vitality of the cells contained in the graft was 96.9%. Tacrolimus and mycophenolate mofetil were used for graft-versus-host disease (GVHD) prophylaxis. Anti-HCMV prophylaxis commenced at day +1 with commercial pooled anti-HCMV Ig (Megalotect, Biotest Pharma GmbH, Germany) at a dose of 100 U/kg twice a week.
The initial post-transplant period was uneventful. The first occurrence of HCMV viremia (600 viral genome copies/mL) was detected at day +14, and antiviral therapy with foscarnet was started immediately. At day +17, neutrophil engraftment was achieved, but not the platelet and the erythrocyte counterparts. At day +20, the patient presented fever without clinical symptoms and without an increase of C-reactive protein. Afterwards, a full blood count revealed a progressive drop in leukocytes and platelets, whereas HCMV viremia continued to increase despite antiviral therapy. The patient also developed a severe, HCMV-related hemorrhagic cystitis, which required frequent platelet transfusions, initially once daily for 5 days, and thereafter twice daily for 7 days.
As the infection appeared unresponsive to foscarnet, ganciclovir treatment was started at day +25. This switch in antiviral therapy followed an increase in viral load, reaching a maximum titer of 5100 viral genome copies/mL at day +30. Simultaneously, the clinical condition of the patient continued to deteriorate, and the leukocyte count decreased drastically. Therefore, the diagnosis of secondary graft failure due to HCMV infection was established. Because of the failure of anti-HCMV treatment, ganciclovir was withdrawn, and foscarnet therapy was resumed. The possible explanations for treatment failure include the presence of antiviral resistance mutations or an ineffective activity of the Ig administered. As the first option seemed improbable because resistance mutations usually arise later than 6 weeks after HSCT, we opted to address the latter option. Therefore, we considered using antibodies (Abs) that were more likely to inhibit the HCMV strains causing the infection in the patient. Although the diversity of circulating HCMV strains is known to be extensive (Puchhammer-Stöckl and Görzer, 2011), we hypothesized that the patient and her relatives may have been exposed to similar strains on a familial basis. As a result, we performed serological tests on the patient's mother, father and brother. We selected the patient's mother, who presented the highest anti-HCMV IgG titer, as the best donor of strain-specific HCMV Ab-rich plasma and virus-specific T-lymphocytes in this situation. At day +32, the patient's mother underwent leukapheresis. The lymphocytes were sent immediately to a cell factory to produce specific anti-HCMV T-lymphocytes. On the same day, the plasma enriched in HCMV strain-specific Abs was infused into the patient. At day +34, the viral load decreased for the first time, to 1100 viral genome copies/mL, and the patient began new conditioning for a haploidentical rescue transplant from her mother. In light of the concurrent bone marrow aplasia, the conditioning regimen was conducted solely with ATG. At day +38, the girl received a second plasma transfusion from her mother. On the day of the second transplant (day +40), complete viral clearance was documented, and the girl received 11.3 × 106 CD34 cells/kg of mother's graft. GVHD prophylaxis was performed with post-transplant cyclophosphamide, tacrolimus, and mycophenolate mofetil. The engraftment was rapid, with a neutrophil count of >500/μL achieved at day +15, and the final platelets and packed red blood cells were transfused at day +20. Since HCMV viremia remained negative during the entire post-transplant period, we discarded the use of the mother's virus-specific T-lymphocytes. The patient was discharged at day +27 in perfect clinical condition, with continuation of antiviral valganciclovir prophylaxis. Immunosuppressive treatment and antiviral prophylaxis were discontinued six months later, and the patient remains in perfect health 2 years after the second transplant.
Therapeutic interventions, HCMV load, and white blood cell count during the entire episode are summarized in Fig. 1. In addition, the HCMV strains present in the patient were characterized by high-throughput sequencing (HTS) of an archived (−80 °C) plasma sample from day +19. DNA was extracted from 800 μl of plasma using a QIAamp MinElute virus spin kit (QIAGEN, Crawley, UK). An aliquot of 50 μl of DNA was used to generate an HCMV-enriched sequencing library as described previously (Suárez et al., 2019). The library was loaded onto a NextSeq DNA sequencer (Illumina, San Diego, CA, USA), generating 150 bp paired-ended reads (Table 1). HCMV strain enumeration was performed by genotyping 12 hypervariable HCMV genes from the reads as described previously (Suárez et al., 2019). The detection of four genotypes of the RL13 gene indicated the presence of at least four HCMV strains (a strain being defined as a particular constellation of genotypes of the hypervariable genes analysed), each including one of these RL13 genotypes (Table 2). The data was also inspected for the presence of resistance mutations in the antiviral target genes (UL54 and UL97) using LoFreq (Wilm et al., 2012). Briefly, the reads were mapped to the genome of HCMV reference strain Merlin (GenBank accession no. AY446894.2), and non-synonymous variants known to confer resistance were identified. No resistance mutations were detected.Fig. 1 Information on viremia, white blood cell (WBC) count, total serum IgG and antiviral therapy. Data on HCMV viremia is expressed in viral genome copies/mL (right Y-axis). Application and duration of antiviral treatments are represented by colored bars (yellow, anti-HCMV-specific Ig; red, foscarnet; blue, ganciclovir/valganciclovir). Chronological data (X-axis) are represented in days after the first transplant. The plasma sample analysed by high-throughput sequencing (HTS) is represented by a filled blue circle.
Fig. 1
Table 1 Overview of HTS data.
Table 1Sample ID TRI-SCT1
Sample type Plasma
Days after transplant +19
HCMV load (genome copies/mL) 2100
Data deposition (ENA project no.)a PRJEB38333
Sequencing library (genome copies) 1440
Trimmed reads (no.) 5,033,480
HCMV reads (no.)b 936,773
HCMV reads; %b 19
Coverage depth (reads/nt)b 885
a The sequence dataset was purged of human reads and deposited in the European Nucleotide Archive (ENA).
b Reads mapped to the HCMV reference strain Merlin genome; HTS, high-throughput sequencing; nt, nucleotide.
Table 2 Genotyping of hypervariable HCMV genes.
Table 2
3 Discussion
Various factors are implicated in the outcome of HCMV infection in transplant patients, the first of which relates to the HCMV serological status of donor and recipient. The serological status of the first donor may have played an important role in this case, as it has been reported that using an HCMV-seronegative unrelated HSCT donor (D-) for a seropositive recipient (R+) results in reduced overall survival after myeloablative conditioning. In contrast, an HCMV seropositive donor is associated with improved event-free survival and reduced non-relapse mortality (Styczynski, 2018). Thus, the absence of an HCMV-specific immunological memory in the donor's cells may have placed the HCMV-positive recipient at a higher risk of HCMV reactivation and associated complications.
In relation to viral factors, the clinical course of the patient may have been influenced by two main factors: the presence of multiple HCMV strains or the presence of mutations conferring antiviral resistance. Infection with multiple HCMV strains is common after transplantation (Görzer et al., 2010), and is associated with delayed HCMV clearance during antiviral therapy, even in the absence of antiviral resistance mutations. It is also associated with worse clinical outcomes, including an increase in graft rejection and faster progression of disease. In principle, the mechanisms behind this increased pathogenicity can be viral, for example involving recombination and complementation of different strains within the host, or immunological, since control of multiple strains may be more challenging for the immune system. Overall, in clinically symptomatic immunocompromised hosts, the presence of multiple HCMV strains is more threatening than the presence of a single strain (Manuel et al., 2009).
In the present study, the initial failure of antiviral therapy could not be explained by the development of resistance mutations in the HCMV strains present in the patient. This finding is not surprising, as detection of resistance mutations is unusual during the first 6 weeks of antiviral therapy, whereas the sample analysed in this case was collected only 13 days after initiation of foscarnet treatment (Lurain and Chou, 2010; Springer et al., 2005).
In regard to the initial failure of the immunotherapy with commercial pooled Ig, this may have been due to the lack of particular Abs with neutralizing capacity against the HCMV strains present in the patient. Assuming that the donors providing the Ig had been exposed to strains circulating in Europe, the Abs may have been more suited for neutralizing HCMV strains commonly circulating in that region. However, it is possible that the strains present in the patient, who originated from Bangladesh, may have differed immunologically from strains in other regions of the world. The extent to which this is true is unknown, as current knowledge of HCMV genome variability is derived almost entirely from strains circulating in a few European countries. Therefore, it would be useful to assess the neutralizing capacity of commercial pooled Ig in order to avoid potential therapeutic failure, especially in patients from non-European countries. The clinical problem in this case was addressed successfully by using the Ab-rich plasma of the patient's mother in a kind of personalized immunotherapy, on the premise that both mother and daughter may have been exposed to the same pool of viral strains. In this HCMV-positive recipient with a severe viral reactivation, Ig obtained from a household donor afforded an efficacious treatment that led to complete viral clearance.
To the best of our knowledge, this is the first report of the successful use of HCMV strain-specific Ig therapy in an HSCT recipient. The efficacy of this approach as a means of prevention has been demonstrated previously only in mouse models involving murine cytomegalovirus (MCMV) reactivation after bone marrow transplantation (Martins et al., 2019). In this setting, mice received serum obtained either from latently infected (seropositive) donors or from seronegative donors. The transfer of immune serum protected mice from viral reactivation without affecting the development of GVHD. Remarkably, complete viral protection was obtained with a small volume (5 μL) of strain-specific immune serum. On the other hand, when specific donor antibodies were administered to mice infected with antigenically mismatched MCMV strains, they were not efficient in preventing viral reactivation.
Our case highlights the importance of monitoring HSCT recipients comprehensively, longitudinally and with high sensitivity for viral factors, including the presence of multiple strains and resistance mutations. Importantly, it also provides an affordable targeted immunological therapy in patients from countries with constrained access to alternative therapies.
4 Conclusion
HSCT is a complex and specific setting in which a recipient may rapidly develop viral reactivation and consequent life-threatening complications. Ab-rich plasma may represent a fast and feasible therapeutic option to overcome viral activity and allow successful engraftment of the donor's bone marrow.
Funding
No specific funding was received.
Consent for publication
The authors have obtained consent from the parents of the patient to publish individual patient data.
Declaration of competing interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgments
The sequencing work was supported by funding from the Medical Research Council (grant number MC_UU_12014/3 to AJD). | UNK UNK, UNKNOWN FREQ. | DrugDosageText | CC BY | 33577809 | 19,010,959 | 2021-04 |
What was the dosage of drug 'MYCOPHENOLATE MOFETIL'? | Post-HSCT graft failure due to refractory human cytomegalovirus successfully treated with haploidentical donor-derived immunoglobulins and stem cell graft infusion: A case report.
Human cytomegalovirus (HCMV) remains an important cause of transplant-related morbidity and mortality. The incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations as a result of a lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen.
We describe the case of an 8-year-old βE-thalassemic girl from Bangladesh who was seropositive for human cytomegalovirus (HCMV) and underwent hematopoietic stem cell transplantation from a HLA-matched, unrelated, HCMV-seronegative donor. Despite administering antiviral prophylaxis with commercial pooled anti-HCMV immunoglobulin (Ig) from day +1, the post-transplant course was complicated by prompt viral reactivation, and foscarnet therapy was initiated. The virus was refractory to treatment, leading rapidly to complete bone marrow failure, and targeted immunotherapy was proposed as a second-line therapy. Hypothesizing that the patient and her relatives may have been exposed to similar HCMV strains, we selected the patient's mother, who presented a high HCMV antibody titer, as the donor of virus strain-specific anti-HCMV Ig and T-lymphocytes. Complete viral clearance was achieved after two transfusions of the mother's plasma. Subsequently, the patient underwent a haploidentical rescue transplant, promptly reaching full hematological recovery.
These findings suggest that treatment with virus strain-specific Ig may offer a new therapeutic option for critically ill patients.
pmc1 Introduction
Human cytomegalovirus (HCMV) is the most significant cause of opportunistic viral infection in allogeneic hematopoietic stem cell transplantation (HSCT) and remains an important cause of transplant-related morbidity and mortality (Wagner-Drouet et al., 2019). Several studies have shown that the incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations (Ganepola et al., 2007; Zhou et al., 2009; Ozdemir et al., 2007). This is likely a consequence of delayed HCMV-specific immune reconstitution due to the lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen (Styczynski, 2018; Ljungman et al., 2014). HCMV infection can lead to pneumonia, gastroenteritis, retinitis, hepatitis, and encephalitis among HSCT recipients (Griffiths, 2012). In addition, HCMV infection of the bone marrow can result in impaired graft acceptance and, in the most extreme cases, complete graft failure (Steffens et al., 1998). Treatment of HCMV disease essentially consists of administering antiviral drugs, namely ganciclovir and foscarnet, and intravenous immunoglobulin. Although chemoprophylaxis is not generally recommended in HSCT, immunoglobulin, whether pooled or HCMV-specific, have shown some efficacy in preventing recurrent HCMV infections (Boeckh and Ljungman, 2009).
We describe a case of a pediatric HSCT recipient who had complications of HCMV recurrence and was treated successfully with targeted immunological therapy, using haploidentical donor-derived immunoglobulin (Ig).
2 Case presentation
An 8-year-old girl was diagnosed at the age of two in her country of origin (Bangladesh) as βE-thalassemic. She was treated with monthly blood transfusions and weekly iron chelation with deferoxamine, plus supplementation with folic acid. A suboptimal regimen of chelation therapy led to severe systemic siderosis, and her family travelled to Italy to undergo HSCT. Since a human leukocyte antigen (HLA)-matched related donor was unavailable at that time, an HLA-matched (10/10) unrelated donor was identified. The patient was HCMV-seropositive, whereas the donor was HCMV-seronegative, thus posing a high-risk serological combination (D-/R+) for post-transplant HCMV infection and associated complications.
The conditioning regimen consisted of fludarabine, busulfan, thiotepa, and anti-thymocyte globulin (ATG). The infused donor's bone marrow contained 3.79 × 108 total nuclear cells/kg recipient weight, and the vitality of the cells contained in the graft was 96.9%. Tacrolimus and mycophenolate mofetil were used for graft-versus-host disease (GVHD) prophylaxis. Anti-HCMV prophylaxis commenced at day +1 with commercial pooled anti-HCMV Ig (Megalotect, Biotest Pharma GmbH, Germany) at a dose of 100 U/kg twice a week.
The initial post-transplant period was uneventful. The first occurrence of HCMV viremia (600 viral genome copies/mL) was detected at day +14, and antiviral therapy with foscarnet was started immediately. At day +17, neutrophil engraftment was achieved, but not the platelet and the erythrocyte counterparts. At day +20, the patient presented fever without clinical symptoms and without an increase of C-reactive protein. Afterwards, a full blood count revealed a progressive drop in leukocytes and platelets, whereas HCMV viremia continued to increase despite antiviral therapy. The patient also developed a severe, HCMV-related hemorrhagic cystitis, which required frequent platelet transfusions, initially once daily for 5 days, and thereafter twice daily for 7 days.
As the infection appeared unresponsive to foscarnet, ganciclovir treatment was started at day +25. This switch in antiviral therapy followed an increase in viral load, reaching a maximum titer of 5100 viral genome copies/mL at day +30. Simultaneously, the clinical condition of the patient continued to deteriorate, and the leukocyte count decreased drastically. Therefore, the diagnosis of secondary graft failure due to HCMV infection was established. Because of the failure of anti-HCMV treatment, ganciclovir was withdrawn, and foscarnet therapy was resumed. The possible explanations for treatment failure include the presence of antiviral resistance mutations or an ineffective activity of the Ig administered. As the first option seemed improbable because resistance mutations usually arise later than 6 weeks after HSCT, we opted to address the latter option. Therefore, we considered using antibodies (Abs) that were more likely to inhibit the HCMV strains causing the infection in the patient. Although the diversity of circulating HCMV strains is known to be extensive (Puchhammer-Stöckl and Görzer, 2011), we hypothesized that the patient and her relatives may have been exposed to similar strains on a familial basis. As a result, we performed serological tests on the patient's mother, father and brother. We selected the patient's mother, who presented the highest anti-HCMV IgG titer, as the best donor of strain-specific HCMV Ab-rich plasma and virus-specific T-lymphocytes in this situation. At day +32, the patient's mother underwent leukapheresis. The lymphocytes were sent immediately to a cell factory to produce specific anti-HCMV T-lymphocytes. On the same day, the plasma enriched in HCMV strain-specific Abs was infused into the patient. At day +34, the viral load decreased for the first time, to 1100 viral genome copies/mL, and the patient began new conditioning for a haploidentical rescue transplant from her mother. In light of the concurrent bone marrow aplasia, the conditioning regimen was conducted solely with ATG. At day +38, the girl received a second plasma transfusion from her mother. On the day of the second transplant (day +40), complete viral clearance was documented, and the girl received 11.3 × 106 CD34 cells/kg of mother's graft. GVHD prophylaxis was performed with post-transplant cyclophosphamide, tacrolimus, and mycophenolate mofetil. The engraftment was rapid, with a neutrophil count of >500/μL achieved at day +15, and the final platelets and packed red blood cells were transfused at day +20. Since HCMV viremia remained negative during the entire post-transplant period, we discarded the use of the mother's virus-specific T-lymphocytes. The patient was discharged at day +27 in perfect clinical condition, with continuation of antiviral valganciclovir prophylaxis. Immunosuppressive treatment and antiviral prophylaxis were discontinued six months later, and the patient remains in perfect health 2 years after the second transplant.
Therapeutic interventions, HCMV load, and white blood cell count during the entire episode are summarized in Fig. 1. In addition, the HCMV strains present in the patient were characterized by high-throughput sequencing (HTS) of an archived (−80 °C) plasma sample from day +19. DNA was extracted from 800 μl of plasma using a QIAamp MinElute virus spin kit (QIAGEN, Crawley, UK). An aliquot of 50 μl of DNA was used to generate an HCMV-enriched sequencing library as described previously (Suárez et al., 2019). The library was loaded onto a NextSeq DNA sequencer (Illumina, San Diego, CA, USA), generating 150 bp paired-ended reads (Table 1). HCMV strain enumeration was performed by genotyping 12 hypervariable HCMV genes from the reads as described previously (Suárez et al., 2019). The detection of four genotypes of the RL13 gene indicated the presence of at least four HCMV strains (a strain being defined as a particular constellation of genotypes of the hypervariable genes analysed), each including one of these RL13 genotypes (Table 2). The data was also inspected for the presence of resistance mutations in the antiviral target genes (UL54 and UL97) using LoFreq (Wilm et al., 2012). Briefly, the reads were mapped to the genome of HCMV reference strain Merlin (GenBank accession no. AY446894.2), and non-synonymous variants known to confer resistance were identified. No resistance mutations were detected.Fig. 1 Information on viremia, white blood cell (WBC) count, total serum IgG and antiviral therapy. Data on HCMV viremia is expressed in viral genome copies/mL (right Y-axis). Application and duration of antiviral treatments are represented by colored bars (yellow, anti-HCMV-specific Ig; red, foscarnet; blue, ganciclovir/valganciclovir). Chronological data (X-axis) are represented in days after the first transplant. The plasma sample analysed by high-throughput sequencing (HTS) is represented by a filled blue circle.
Fig. 1
Table 1 Overview of HTS data.
Table 1Sample ID TRI-SCT1
Sample type Plasma
Days after transplant +19
HCMV load (genome copies/mL) 2100
Data deposition (ENA project no.)a PRJEB38333
Sequencing library (genome copies) 1440
Trimmed reads (no.) 5,033,480
HCMV reads (no.)b 936,773
HCMV reads; %b 19
Coverage depth (reads/nt)b 885
a The sequence dataset was purged of human reads and deposited in the European Nucleotide Archive (ENA).
b Reads mapped to the HCMV reference strain Merlin genome; HTS, high-throughput sequencing; nt, nucleotide.
Table 2 Genotyping of hypervariable HCMV genes.
Table 2
3 Discussion
Various factors are implicated in the outcome of HCMV infection in transplant patients, the first of which relates to the HCMV serological status of donor and recipient. The serological status of the first donor may have played an important role in this case, as it has been reported that using an HCMV-seronegative unrelated HSCT donor (D-) for a seropositive recipient (R+) results in reduced overall survival after myeloablative conditioning. In contrast, an HCMV seropositive donor is associated with improved event-free survival and reduced non-relapse mortality (Styczynski, 2018). Thus, the absence of an HCMV-specific immunological memory in the donor's cells may have placed the HCMV-positive recipient at a higher risk of HCMV reactivation and associated complications.
In relation to viral factors, the clinical course of the patient may have been influenced by two main factors: the presence of multiple HCMV strains or the presence of mutations conferring antiviral resistance. Infection with multiple HCMV strains is common after transplantation (Görzer et al., 2010), and is associated with delayed HCMV clearance during antiviral therapy, even in the absence of antiviral resistance mutations. It is also associated with worse clinical outcomes, including an increase in graft rejection and faster progression of disease. In principle, the mechanisms behind this increased pathogenicity can be viral, for example involving recombination and complementation of different strains within the host, or immunological, since control of multiple strains may be more challenging for the immune system. Overall, in clinically symptomatic immunocompromised hosts, the presence of multiple HCMV strains is more threatening than the presence of a single strain (Manuel et al., 2009).
In the present study, the initial failure of antiviral therapy could not be explained by the development of resistance mutations in the HCMV strains present in the patient. This finding is not surprising, as detection of resistance mutations is unusual during the first 6 weeks of antiviral therapy, whereas the sample analysed in this case was collected only 13 days after initiation of foscarnet treatment (Lurain and Chou, 2010; Springer et al., 2005).
In regard to the initial failure of the immunotherapy with commercial pooled Ig, this may have been due to the lack of particular Abs with neutralizing capacity against the HCMV strains present in the patient. Assuming that the donors providing the Ig had been exposed to strains circulating in Europe, the Abs may have been more suited for neutralizing HCMV strains commonly circulating in that region. However, it is possible that the strains present in the patient, who originated from Bangladesh, may have differed immunologically from strains in other regions of the world. The extent to which this is true is unknown, as current knowledge of HCMV genome variability is derived almost entirely from strains circulating in a few European countries. Therefore, it would be useful to assess the neutralizing capacity of commercial pooled Ig in order to avoid potential therapeutic failure, especially in patients from non-European countries. The clinical problem in this case was addressed successfully by using the Ab-rich plasma of the patient's mother in a kind of personalized immunotherapy, on the premise that both mother and daughter may have been exposed to the same pool of viral strains. In this HCMV-positive recipient with a severe viral reactivation, Ig obtained from a household donor afforded an efficacious treatment that led to complete viral clearance.
To the best of our knowledge, this is the first report of the successful use of HCMV strain-specific Ig therapy in an HSCT recipient. The efficacy of this approach as a means of prevention has been demonstrated previously only in mouse models involving murine cytomegalovirus (MCMV) reactivation after bone marrow transplantation (Martins et al., 2019). In this setting, mice received serum obtained either from latently infected (seropositive) donors or from seronegative donors. The transfer of immune serum protected mice from viral reactivation without affecting the development of GVHD. Remarkably, complete viral protection was obtained with a small volume (5 μL) of strain-specific immune serum. On the other hand, when specific donor antibodies were administered to mice infected with antigenically mismatched MCMV strains, they were not efficient in preventing viral reactivation.
Our case highlights the importance of monitoring HSCT recipients comprehensively, longitudinally and with high sensitivity for viral factors, including the presence of multiple strains and resistance mutations. Importantly, it also provides an affordable targeted immunological therapy in patients from countries with constrained access to alternative therapies.
4 Conclusion
HSCT is a complex and specific setting in which a recipient may rapidly develop viral reactivation and consequent life-threatening complications. Ab-rich plasma may represent a fast and feasible therapeutic option to overcome viral activity and allow successful engraftment of the donor's bone marrow.
Funding
No specific funding was received.
Consent for publication
The authors have obtained consent from the parents of the patient to publish individual patient data.
Declaration of competing interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgments
The sequencing work was supported by funding from the Medical Research Council (grant number MC_UU_12014/3 to AJD). | UNK UNK, UNKNOWN FREQ. | DrugDosageText | CC BY | 33577809 | 19,010,959 | 2021-04 |
What was the dosage of drug 'TACROLIMUS'? | Post-HSCT graft failure due to refractory human cytomegalovirus successfully treated with haploidentical donor-derived immunoglobulins and stem cell graft infusion: A case report.
Human cytomegalovirus (HCMV) remains an important cause of transplant-related morbidity and mortality. The incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations as a result of a lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen.
We describe the case of an 8-year-old βE-thalassemic girl from Bangladesh who was seropositive for human cytomegalovirus (HCMV) and underwent hematopoietic stem cell transplantation from a HLA-matched, unrelated, HCMV-seronegative donor. Despite administering antiviral prophylaxis with commercial pooled anti-HCMV immunoglobulin (Ig) from day +1, the post-transplant course was complicated by prompt viral reactivation, and foscarnet therapy was initiated. The virus was refractory to treatment, leading rapidly to complete bone marrow failure, and targeted immunotherapy was proposed as a second-line therapy. Hypothesizing that the patient and her relatives may have been exposed to similar HCMV strains, we selected the patient's mother, who presented a high HCMV antibody titer, as the donor of virus strain-specific anti-HCMV Ig and T-lymphocytes. Complete viral clearance was achieved after two transfusions of the mother's plasma. Subsequently, the patient underwent a haploidentical rescue transplant, promptly reaching full hematological recovery.
These findings suggest that treatment with virus strain-specific Ig may offer a new therapeutic option for critically ill patients.
pmc1 Introduction
Human cytomegalovirus (HCMV) is the most significant cause of opportunistic viral infection in allogeneic hematopoietic stem cell transplantation (HSCT) and remains an important cause of transplant-related morbidity and mortality (Wagner-Drouet et al., 2019). Several studies have shown that the incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations (Ganepola et al., 2007; Zhou et al., 2009; Ozdemir et al., 2007). This is likely a consequence of delayed HCMV-specific immune reconstitution due to the lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen (Styczynski, 2018; Ljungman et al., 2014). HCMV infection can lead to pneumonia, gastroenteritis, retinitis, hepatitis, and encephalitis among HSCT recipients (Griffiths, 2012). In addition, HCMV infection of the bone marrow can result in impaired graft acceptance and, in the most extreme cases, complete graft failure (Steffens et al., 1998). Treatment of HCMV disease essentially consists of administering antiviral drugs, namely ganciclovir and foscarnet, and intravenous immunoglobulin. Although chemoprophylaxis is not generally recommended in HSCT, immunoglobulin, whether pooled or HCMV-specific, have shown some efficacy in preventing recurrent HCMV infections (Boeckh and Ljungman, 2009).
We describe a case of a pediatric HSCT recipient who had complications of HCMV recurrence and was treated successfully with targeted immunological therapy, using haploidentical donor-derived immunoglobulin (Ig).
2 Case presentation
An 8-year-old girl was diagnosed at the age of two in her country of origin (Bangladesh) as βE-thalassemic. She was treated with monthly blood transfusions and weekly iron chelation with deferoxamine, plus supplementation with folic acid. A suboptimal regimen of chelation therapy led to severe systemic siderosis, and her family travelled to Italy to undergo HSCT. Since a human leukocyte antigen (HLA)-matched related donor was unavailable at that time, an HLA-matched (10/10) unrelated donor was identified. The patient was HCMV-seropositive, whereas the donor was HCMV-seronegative, thus posing a high-risk serological combination (D-/R+) for post-transplant HCMV infection and associated complications.
The conditioning regimen consisted of fludarabine, busulfan, thiotepa, and anti-thymocyte globulin (ATG). The infused donor's bone marrow contained 3.79 × 108 total nuclear cells/kg recipient weight, and the vitality of the cells contained in the graft was 96.9%. Tacrolimus and mycophenolate mofetil were used for graft-versus-host disease (GVHD) prophylaxis. Anti-HCMV prophylaxis commenced at day +1 with commercial pooled anti-HCMV Ig (Megalotect, Biotest Pharma GmbH, Germany) at a dose of 100 U/kg twice a week.
The initial post-transplant period was uneventful. The first occurrence of HCMV viremia (600 viral genome copies/mL) was detected at day +14, and antiviral therapy with foscarnet was started immediately. At day +17, neutrophil engraftment was achieved, but not the platelet and the erythrocyte counterparts. At day +20, the patient presented fever without clinical symptoms and without an increase of C-reactive protein. Afterwards, a full blood count revealed a progressive drop in leukocytes and platelets, whereas HCMV viremia continued to increase despite antiviral therapy. The patient also developed a severe, HCMV-related hemorrhagic cystitis, which required frequent platelet transfusions, initially once daily for 5 days, and thereafter twice daily for 7 days.
As the infection appeared unresponsive to foscarnet, ganciclovir treatment was started at day +25. This switch in antiviral therapy followed an increase in viral load, reaching a maximum titer of 5100 viral genome copies/mL at day +30. Simultaneously, the clinical condition of the patient continued to deteriorate, and the leukocyte count decreased drastically. Therefore, the diagnosis of secondary graft failure due to HCMV infection was established. Because of the failure of anti-HCMV treatment, ganciclovir was withdrawn, and foscarnet therapy was resumed. The possible explanations for treatment failure include the presence of antiviral resistance mutations or an ineffective activity of the Ig administered. As the first option seemed improbable because resistance mutations usually arise later than 6 weeks after HSCT, we opted to address the latter option. Therefore, we considered using antibodies (Abs) that were more likely to inhibit the HCMV strains causing the infection in the patient. Although the diversity of circulating HCMV strains is known to be extensive (Puchhammer-Stöckl and Görzer, 2011), we hypothesized that the patient and her relatives may have been exposed to similar strains on a familial basis. As a result, we performed serological tests on the patient's mother, father and brother. We selected the patient's mother, who presented the highest anti-HCMV IgG titer, as the best donor of strain-specific HCMV Ab-rich plasma and virus-specific T-lymphocytes in this situation. At day +32, the patient's mother underwent leukapheresis. The lymphocytes were sent immediately to a cell factory to produce specific anti-HCMV T-lymphocytes. On the same day, the plasma enriched in HCMV strain-specific Abs was infused into the patient. At day +34, the viral load decreased for the first time, to 1100 viral genome copies/mL, and the patient began new conditioning for a haploidentical rescue transplant from her mother. In light of the concurrent bone marrow aplasia, the conditioning regimen was conducted solely with ATG. At day +38, the girl received a second plasma transfusion from her mother. On the day of the second transplant (day +40), complete viral clearance was documented, and the girl received 11.3 × 106 CD34 cells/kg of mother's graft. GVHD prophylaxis was performed with post-transplant cyclophosphamide, tacrolimus, and mycophenolate mofetil. The engraftment was rapid, with a neutrophil count of >500/μL achieved at day +15, and the final platelets and packed red blood cells were transfused at day +20. Since HCMV viremia remained negative during the entire post-transplant period, we discarded the use of the mother's virus-specific T-lymphocytes. The patient was discharged at day +27 in perfect clinical condition, with continuation of antiviral valganciclovir prophylaxis. Immunosuppressive treatment and antiviral prophylaxis were discontinued six months later, and the patient remains in perfect health 2 years after the second transplant.
Therapeutic interventions, HCMV load, and white blood cell count during the entire episode are summarized in Fig. 1. In addition, the HCMV strains present in the patient were characterized by high-throughput sequencing (HTS) of an archived (−80 °C) plasma sample from day +19. DNA was extracted from 800 μl of plasma using a QIAamp MinElute virus spin kit (QIAGEN, Crawley, UK). An aliquot of 50 μl of DNA was used to generate an HCMV-enriched sequencing library as described previously (Suárez et al., 2019). The library was loaded onto a NextSeq DNA sequencer (Illumina, San Diego, CA, USA), generating 150 bp paired-ended reads (Table 1). HCMV strain enumeration was performed by genotyping 12 hypervariable HCMV genes from the reads as described previously (Suárez et al., 2019). The detection of four genotypes of the RL13 gene indicated the presence of at least four HCMV strains (a strain being defined as a particular constellation of genotypes of the hypervariable genes analysed), each including one of these RL13 genotypes (Table 2). The data was also inspected for the presence of resistance mutations in the antiviral target genes (UL54 and UL97) using LoFreq (Wilm et al., 2012). Briefly, the reads were mapped to the genome of HCMV reference strain Merlin (GenBank accession no. AY446894.2), and non-synonymous variants known to confer resistance were identified. No resistance mutations were detected.Fig. 1 Information on viremia, white blood cell (WBC) count, total serum IgG and antiviral therapy. Data on HCMV viremia is expressed in viral genome copies/mL (right Y-axis). Application and duration of antiviral treatments are represented by colored bars (yellow, anti-HCMV-specific Ig; red, foscarnet; blue, ganciclovir/valganciclovir). Chronological data (X-axis) are represented in days after the first transplant. The plasma sample analysed by high-throughput sequencing (HTS) is represented by a filled blue circle.
Fig. 1
Table 1 Overview of HTS data.
Table 1Sample ID TRI-SCT1
Sample type Plasma
Days after transplant +19
HCMV load (genome copies/mL) 2100
Data deposition (ENA project no.)a PRJEB38333
Sequencing library (genome copies) 1440
Trimmed reads (no.) 5,033,480
HCMV reads (no.)b 936,773
HCMV reads; %b 19
Coverage depth (reads/nt)b 885
a The sequence dataset was purged of human reads and deposited in the European Nucleotide Archive (ENA).
b Reads mapped to the HCMV reference strain Merlin genome; HTS, high-throughput sequencing; nt, nucleotide.
Table 2 Genotyping of hypervariable HCMV genes.
Table 2
3 Discussion
Various factors are implicated in the outcome of HCMV infection in transplant patients, the first of which relates to the HCMV serological status of donor and recipient. The serological status of the first donor may have played an important role in this case, as it has been reported that using an HCMV-seronegative unrelated HSCT donor (D-) for a seropositive recipient (R+) results in reduced overall survival after myeloablative conditioning. In contrast, an HCMV seropositive donor is associated with improved event-free survival and reduced non-relapse mortality (Styczynski, 2018). Thus, the absence of an HCMV-specific immunological memory in the donor's cells may have placed the HCMV-positive recipient at a higher risk of HCMV reactivation and associated complications.
In relation to viral factors, the clinical course of the patient may have been influenced by two main factors: the presence of multiple HCMV strains or the presence of mutations conferring antiviral resistance. Infection with multiple HCMV strains is common after transplantation (Görzer et al., 2010), and is associated with delayed HCMV clearance during antiviral therapy, even in the absence of antiviral resistance mutations. It is also associated with worse clinical outcomes, including an increase in graft rejection and faster progression of disease. In principle, the mechanisms behind this increased pathogenicity can be viral, for example involving recombination and complementation of different strains within the host, or immunological, since control of multiple strains may be more challenging for the immune system. Overall, in clinically symptomatic immunocompromised hosts, the presence of multiple HCMV strains is more threatening than the presence of a single strain (Manuel et al., 2009).
In the present study, the initial failure of antiviral therapy could not be explained by the development of resistance mutations in the HCMV strains present in the patient. This finding is not surprising, as detection of resistance mutations is unusual during the first 6 weeks of antiviral therapy, whereas the sample analysed in this case was collected only 13 days after initiation of foscarnet treatment (Lurain and Chou, 2010; Springer et al., 2005).
In regard to the initial failure of the immunotherapy with commercial pooled Ig, this may have been due to the lack of particular Abs with neutralizing capacity against the HCMV strains present in the patient. Assuming that the donors providing the Ig had been exposed to strains circulating in Europe, the Abs may have been more suited for neutralizing HCMV strains commonly circulating in that region. However, it is possible that the strains present in the patient, who originated from Bangladesh, may have differed immunologically from strains in other regions of the world. The extent to which this is true is unknown, as current knowledge of HCMV genome variability is derived almost entirely from strains circulating in a few European countries. Therefore, it would be useful to assess the neutralizing capacity of commercial pooled Ig in order to avoid potential therapeutic failure, especially in patients from non-European countries. The clinical problem in this case was addressed successfully by using the Ab-rich plasma of the patient's mother in a kind of personalized immunotherapy, on the premise that both mother and daughter may have been exposed to the same pool of viral strains. In this HCMV-positive recipient with a severe viral reactivation, Ig obtained from a household donor afforded an efficacious treatment that led to complete viral clearance.
To the best of our knowledge, this is the first report of the successful use of HCMV strain-specific Ig therapy in an HSCT recipient. The efficacy of this approach as a means of prevention has been demonstrated previously only in mouse models involving murine cytomegalovirus (MCMV) reactivation after bone marrow transplantation (Martins et al., 2019). In this setting, mice received serum obtained either from latently infected (seropositive) donors or from seronegative donors. The transfer of immune serum protected mice from viral reactivation without affecting the development of GVHD. Remarkably, complete viral protection was obtained with a small volume (5 μL) of strain-specific immune serum. On the other hand, when specific donor antibodies were administered to mice infected with antigenically mismatched MCMV strains, they were not efficient in preventing viral reactivation.
Our case highlights the importance of monitoring HSCT recipients comprehensively, longitudinally and with high sensitivity for viral factors, including the presence of multiple strains and resistance mutations. Importantly, it also provides an affordable targeted immunological therapy in patients from countries with constrained access to alternative therapies.
4 Conclusion
HSCT is a complex and specific setting in which a recipient may rapidly develop viral reactivation and consequent life-threatening complications. Ab-rich plasma may represent a fast and feasible therapeutic option to overcome viral activity and allow successful engraftment of the donor's bone marrow.
Funding
No specific funding was received.
Consent for publication
The authors have obtained consent from the parents of the patient to publish individual patient data.
Declaration of competing interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgments
The sequencing work was supported by funding from the Medical Research Council (grant number MC_UU_12014/3 to AJD). | UNK UNK, UNKNOWN FREQ. | DrugDosageText | CC BY | 33577809 | 19,010,959 | 2021-04 |
What was the dosage of drug 'THIOTEPA'? | Post-HSCT graft failure due to refractory human cytomegalovirus successfully treated with haploidentical donor-derived immunoglobulins and stem cell graft infusion: A case report.
Human cytomegalovirus (HCMV) remains an important cause of transplant-related morbidity and mortality. The incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations as a result of a lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen.
We describe the case of an 8-year-old βE-thalassemic girl from Bangladesh who was seropositive for human cytomegalovirus (HCMV) and underwent hematopoietic stem cell transplantation from a HLA-matched, unrelated, HCMV-seronegative donor. Despite administering antiviral prophylaxis with commercial pooled anti-HCMV immunoglobulin (Ig) from day +1, the post-transplant course was complicated by prompt viral reactivation, and foscarnet therapy was initiated. The virus was refractory to treatment, leading rapidly to complete bone marrow failure, and targeted immunotherapy was proposed as a second-line therapy. Hypothesizing that the patient and her relatives may have been exposed to similar HCMV strains, we selected the patient's mother, who presented a high HCMV antibody titer, as the donor of virus strain-specific anti-HCMV Ig and T-lymphocytes. Complete viral clearance was achieved after two transfusions of the mother's plasma. Subsequently, the patient underwent a haploidentical rescue transplant, promptly reaching full hematological recovery.
These findings suggest that treatment with virus strain-specific Ig may offer a new therapeutic option for critically ill patients.
pmc1 Introduction
Human cytomegalovirus (HCMV) is the most significant cause of opportunistic viral infection in allogeneic hematopoietic stem cell transplantation (HSCT) and remains an important cause of transplant-related morbidity and mortality (Wagner-Drouet et al., 2019). Several studies have shown that the incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations (Ganepola et al., 2007; Zhou et al., 2009; Ozdemir et al., 2007). This is likely a consequence of delayed HCMV-specific immune reconstitution due to the lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen (Styczynski, 2018; Ljungman et al., 2014). HCMV infection can lead to pneumonia, gastroenteritis, retinitis, hepatitis, and encephalitis among HSCT recipients (Griffiths, 2012). In addition, HCMV infection of the bone marrow can result in impaired graft acceptance and, in the most extreme cases, complete graft failure (Steffens et al., 1998). Treatment of HCMV disease essentially consists of administering antiviral drugs, namely ganciclovir and foscarnet, and intravenous immunoglobulin. Although chemoprophylaxis is not generally recommended in HSCT, immunoglobulin, whether pooled or HCMV-specific, have shown some efficacy in preventing recurrent HCMV infections (Boeckh and Ljungman, 2009).
We describe a case of a pediatric HSCT recipient who had complications of HCMV recurrence and was treated successfully with targeted immunological therapy, using haploidentical donor-derived immunoglobulin (Ig).
2 Case presentation
An 8-year-old girl was diagnosed at the age of two in her country of origin (Bangladesh) as βE-thalassemic. She was treated with monthly blood transfusions and weekly iron chelation with deferoxamine, plus supplementation with folic acid. A suboptimal regimen of chelation therapy led to severe systemic siderosis, and her family travelled to Italy to undergo HSCT. Since a human leukocyte antigen (HLA)-matched related donor was unavailable at that time, an HLA-matched (10/10) unrelated donor was identified. The patient was HCMV-seropositive, whereas the donor was HCMV-seronegative, thus posing a high-risk serological combination (D-/R+) for post-transplant HCMV infection and associated complications.
The conditioning regimen consisted of fludarabine, busulfan, thiotepa, and anti-thymocyte globulin (ATG). The infused donor's bone marrow contained 3.79 × 108 total nuclear cells/kg recipient weight, and the vitality of the cells contained in the graft was 96.9%. Tacrolimus and mycophenolate mofetil were used for graft-versus-host disease (GVHD) prophylaxis. Anti-HCMV prophylaxis commenced at day +1 with commercial pooled anti-HCMV Ig (Megalotect, Biotest Pharma GmbH, Germany) at a dose of 100 U/kg twice a week.
The initial post-transplant period was uneventful. The first occurrence of HCMV viremia (600 viral genome copies/mL) was detected at day +14, and antiviral therapy with foscarnet was started immediately. At day +17, neutrophil engraftment was achieved, but not the platelet and the erythrocyte counterparts. At day +20, the patient presented fever without clinical symptoms and without an increase of C-reactive protein. Afterwards, a full blood count revealed a progressive drop in leukocytes and platelets, whereas HCMV viremia continued to increase despite antiviral therapy. The patient also developed a severe, HCMV-related hemorrhagic cystitis, which required frequent platelet transfusions, initially once daily for 5 days, and thereafter twice daily for 7 days.
As the infection appeared unresponsive to foscarnet, ganciclovir treatment was started at day +25. This switch in antiviral therapy followed an increase in viral load, reaching a maximum titer of 5100 viral genome copies/mL at day +30. Simultaneously, the clinical condition of the patient continued to deteriorate, and the leukocyte count decreased drastically. Therefore, the diagnosis of secondary graft failure due to HCMV infection was established. Because of the failure of anti-HCMV treatment, ganciclovir was withdrawn, and foscarnet therapy was resumed. The possible explanations for treatment failure include the presence of antiviral resistance mutations or an ineffective activity of the Ig administered. As the first option seemed improbable because resistance mutations usually arise later than 6 weeks after HSCT, we opted to address the latter option. Therefore, we considered using antibodies (Abs) that were more likely to inhibit the HCMV strains causing the infection in the patient. Although the diversity of circulating HCMV strains is known to be extensive (Puchhammer-Stöckl and Görzer, 2011), we hypothesized that the patient and her relatives may have been exposed to similar strains on a familial basis. As a result, we performed serological tests on the patient's mother, father and brother. We selected the patient's mother, who presented the highest anti-HCMV IgG titer, as the best donor of strain-specific HCMV Ab-rich plasma and virus-specific T-lymphocytes in this situation. At day +32, the patient's mother underwent leukapheresis. The lymphocytes were sent immediately to a cell factory to produce specific anti-HCMV T-lymphocytes. On the same day, the plasma enriched in HCMV strain-specific Abs was infused into the patient. At day +34, the viral load decreased for the first time, to 1100 viral genome copies/mL, and the patient began new conditioning for a haploidentical rescue transplant from her mother. In light of the concurrent bone marrow aplasia, the conditioning regimen was conducted solely with ATG. At day +38, the girl received a second plasma transfusion from her mother. On the day of the second transplant (day +40), complete viral clearance was documented, and the girl received 11.3 × 106 CD34 cells/kg of mother's graft. GVHD prophylaxis was performed with post-transplant cyclophosphamide, tacrolimus, and mycophenolate mofetil. The engraftment was rapid, with a neutrophil count of >500/μL achieved at day +15, and the final platelets and packed red blood cells were transfused at day +20. Since HCMV viremia remained negative during the entire post-transplant period, we discarded the use of the mother's virus-specific T-lymphocytes. The patient was discharged at day +27 in perfect clinical condition, with continuation of antiviral valganciclovir prophylaxis. Immunosuppressive treatment and antiviral prophylaxis were discontinued six months later, and the patient remains in perfect health 2 years after the second transplant.
Therapeutic interventions, HCMV load, and white blood cell count during the entire episode are summarized in Fig. 1. In addition, the HCMV strains present in the patient were characterized by high-throughput sequencing (HTS) of an archived (−80 °C) plasma sample from day +19. DNA was extracted from 800 μl of plasma using a QIAamp MinElute virus spin kit (QIAGEN, Crawley, UK). An aliquot of 50 μl of DNA was used to generate an HCMV-enriched sequencing library as described previously (Suárez et al., 2019). The library was loaded onto a NextSeq DNA sequencer (Illumina, San Diego, CA, USA), generating 150 bp paired-ended reads (Table 1). HCMV strain enumeration was performed by genotyping 12 hypervariable HCMV genes from the reads as described previously (Suárez et al., 2019). The detection of four genotypes of the RL13 gene indicated the presence of at least four HCMV strains (a strain being defined as a particular constellation of genotypes of the hypervariable genes analysed), each including one of these RL13 genotypes (Table 2). The data was also inspected for the presence of resistance mutations in the antiviral target genes (UL54 and UL97) using LoFreq (Wilm et al., 2012). Briefly, the reads were mapped to the genome of HCMV reference strain Merlin (GenBank accession no. AY446894.2), and non-synonymous variants known to confer resistance were identified. No resistance mutations were detected.Fig. 1 Information on viremia, white blood cell (WBC) count, total serum IgG and antiviral therapy. Data on HCMV viremia is expressed in viral genome copies/mL (right Y-axis). Application and duration of antiviral treatments are represented by colored bars (yellow, anti-HCMV-specific Ig; red, foscarnet; blue, ganciclovir/valganciclovir). Chronological data (X-axis) are represented in days after the first transplant. The plasma sample analysed by high-throughput sequencing (HTS) is represented by a filled blue circle.
Fig. 1
Table 1 Overview of HTS data.
Table 1Sample ID TRI-SCT1
Sample type Plasma
Days after transplant +19
HCMV load (genome copies/mL) 2100
Data deposition (ENA project no.)a PRJEB38333
Sequencing library (genome copies) 1440
Trimmed reads (no.) 5,033,480
HCMV reads (no.)b 936,773
HCMV reads; %b 19
Coverage depth (reads/nt)b 885
a The sequence dataset was purged of human reads and deposited in the European Nucleotide Archive (ENA).
b Reads mapped to the HCMV reference strain Merlin genome; HTS, high-throughput sequencing; nt, nucleotide.
Table 2 Genotyping of hypervariable HCMV genes.
Table 2
3 Discussion
Various factors are implicated in the outcome of HCMV infection in transplant patients, the first of which relates to the HCMV serological status of donor and recipient. The serological status of the first donor may have played an important role in this case, as it has been reported that using an HCMV-seronegative unrelated HSCT donor (D-) for a seropositive recipient (R+) results in reduced overall survival after myeloablative conditioning. In contrast, an HCMV seropositive donor is associated with improved event-free survival and reduced non-relapse mortality (Styczynski, 2018). Thus, the absence of an HCMV-specific immunological memory in the donor's cells may have placed the HCMV-positive recipient at a higher risk of HCMV reactivation and associated complications.
In relation to viral factors, the clinical course of the patient may have been influenced by two main factors: the presence of multiple HCMV strains or the presence of mutations conferring antiviral resistance. Infection with multiple HCMV strains is common after transplantation (Görzer et al., 2010), and is associated with delayed HCMV clearance during antiviral therapy, even in the absence of antiviral resistance mutations. It is also associated with worse clinical outcomes, including an increase in graft rejection and faster progression of disease. In principle, the mechanisms behind this increased pathogenicity can be viral, for example involving recombination and complementation of different strains within the host, or immunological, since control of multiple strains may be more challenging for the immune system. Overall, in clinically symptomatic immunocompromised hosts, the presence of multiple HCMV strains is more threatening than the presence of a single strain (Manuel et al., 2009).
In the present study, the initial failure of antiviral therapy could not be explained by the development of resistance mutations in the HCMV strains present in the patient. This finding is not surprising, as detection of resistance mutations is unusual during the first 6 weeks of antiviral therapy, whereas the sample analysed in this case was collected only 13 days after initiation of foscarnet treatment (Lurain and Chou, 2010; Springer et al., 2005).
In regard to the initial failure of the immunotherapy with commercial pooled Ig, this may have been due to the lack of particular Abs with neutralizing capacity against the HCMV strains present in the patient. Assuming that the donors providing the Ig had been exposed to strains circulating in Europe, the Abs may have been more suited for neutralizing HCMV strains commonly circulating in that region. However, it is possible that the strains present in the patient, who originated from Bangladesh, may have differed immunologically from strains in other regions of the world. The extent to which this is true is unknown, as current knowledge of HCMV genome variability is derived almost entirely from strains circulating in a few European countries. Therefore, it would be useful to assess the neutralizing capacity of commercial pooled Ig in order to avoid potential therapeutic failure, especially in patients from non-European countries. The clinical problem in this case was addressed successfully by using the Ab-rich plasma of the patient's mother in a kind of personalized immunotherapy, on the premise that both mother and daughter may have been exposed to the same pool of viral strains. In this HCMV-positive recipient with a severe viral reactivation, Ig obtained from a household donor afforded an efficacious treatment that led to complete viral clearance.
To the best of our knowledge, this is the first report of the successful use of HCMV strain-specific Ig therapy in an HSCT recipient. The efficacy of this approach as a means of prevention has been demonstrated previously only in mouse models involving murine cytomegalovirus (MCMV) reactivation after bone marrow transplantation (Martins et al., 2019). In this setting, mice received serum obtained either from latently infected (seropositive) donors or from seronegative donors. The transfer of immune serum protected mice from viral reactivation without affecting the development of GVHD. Remarkably, complete viral protection was obtained with a small volume (5 μL) of strain-specific immune serum. On the other hand, when specific donor antibodies were administered to mice infected with antigenically mismatched MCMV strains, they were not efficient in preventing viral reactivation.
Our case highlights the importance of monitoring HSCT recipients comprehensively, longitudinally and with high sensitivity for viral factors, including the presence of multiple strains and resistance mutations. Importantly, it also provides an affordable targeted immunological therapy in patients from countries with constrained access to alternative therapies.
4 Conclusion
HSCT is a complex and specific setting in which a recipient may rapidly develop viral reactivation and consequent life-threatening complications. Ab-rich plasma may represent a fast and feasible therapeutic option to overcome viral activity and allow successful engraftment of the donor's bone marrow.
Funding
No specific funding was received.
Consent for publication
The authors have obtained consent from the parents of the patient to publish individual patient data.
Declaration of competing interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgments
The sequencing work was supported by funding from the Medical Research Council (grant number MC_UU_12014/3 to AJD). | UNK UNK, UNKNOWN FREQ. | DrugDosageText | CC BY | 33577809 | 19,010,959 | 2021-04 |
What was the outcome of reaction 'Blood stem cell transplant failure'? | Post-HSCT graft failure due to refractory human cytomegalovirus successfully treated with haploidentical donor-derived immunoglobulins and stem cell graft infusion: A case report.
Human cytomegalovirus (HCMV) remains an important cause of transplant-related morbidity and mortality. The incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations as a result of a lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen.
We describe the case of an 8-year-old βE-thalassemic girl from Bangladesh who was seropositive for human cytomegalovirus (HCMV) and underwent hematopoietic stem cell transplantation from a HLA-matched, unrelated, HCMV-seronegative donor. Despite administering antiviral prophylaxis with commercial pooled anti-HCMV immunoglobulin (Ig) from day +1, the post-transplant course was complicated by prompt viral reactivation, and foscarnet therapy was initiated. The virus was refractory to treatment, leading rapidly to complete bone marrow failure, and targeted immunotherapy was proposed as a second-line therapy. Hypothesizing that the patient and her relatives may have been exposed to similar HCMV strains, we selected the patient's mother, who presented a high HCMV antibody titer, as the donor of virus strain-specific anti-HCMV Ig and T-lymphocytes. Complete viral clearance was achieved after two transfusions of the mother's plasma. Subsequently, the patient underwent a haploidentical rescue transplant, promptly reaching full hematological recovery.
These findings suggest that treatment with virus strain-specific Ig may offer a new therapeutic option for critically ill patients.
pmc1 Introduction
Human cytomegalovirus (HCMV) is the most significant cause of opportunistic viral infection in allogeneic hematopoietic stem cell transplantation (HSCT) and remains an important cause of transplant-related morbidity and mortality (Wagner-Drouet et al., 2019). Several studies have shown that the incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations (Ganepola et al., 2007; Zhou et al., 2009; Ozdemir et al., 2007). This is likely a consequence of delayed HCMV-specific immune reconstitution due to the lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen (Styczynski, 2018; Ljungman et al., 2014). HCMV infection can lead to pneumonia, gastroenteritis, retinitis, hepatitis, and encephalitis among HSCT recipients (Griffiths, 2012). In addition, HCMV infection of the bone marrow can result in impaired graft acceptance and, in the most extreme cases, complete graft failure (Steffens et al., 1998). Treatment of HCMV disease essentially consists of administering antiviral drugs, namely ganciclovir and foscarnet, and intravenous immunoglobulin. Although chemoprophylaxis is not generally recommended in HSCT, immunoglobulin, whether pooled or HCMV-specific, have shown some efficacy in preventing recurrent HCMV infections (Boeckh and Ljungman, 2009).
We describe a case of a pediatric HSCT recipient who had complications of HCMV recurrence and was treated successfully with targeted immunological therapy, using haploidentical donor-derived immunoglobulin (Ig).
2 Case presentation
An 8-year-old girl was diagnosed at the age of two in her country of origin (Bangladesh) as βE-thalassemic. She was treated with monthly blood transfusions and weekly iron chelation with deferoxamine, plus supplementation with folic acid. A suboptimal regimen of chelation therapy led to severe systemic siderosis, and her family travelled to Italy to undergo HSCT. Since a human leukocyte antigen (HLA)-matched related donor was unavailable at that time, an HLA-matched (10/10) unrelated donor was identified. The patient was HCMV-seropositive, whereas the donor was HCMV-seronegative, thus posing a high-risk serological combination (D-/R+) for post-transplant HCMV infection and associated complications.
The conditioning regimen consisted of fludarabine, busulfan, thiotepa, and anti-thymocyte globulin (ATG). The infused donor's bone marrow contained 3.79 × 108 total nuclear cells/kg recipient weight, and the vitality of the cells contained in the graft was 96.9%. Tacrolimus and mycophenolate mofetil were used for graft-versus-host disease (GVHD) prophylaxis. Anti-HCMV prophylaxis commenced at day +1 with commercial pooled anti-HCMV Ig (Megalotect, Biotest Pharma GmbH, Germany) at a dose of 100 U/kg twice a week.
The initial post-transplant period was uneventful. The first occurrence of HCMV viremia (600 viral genome copies/mL) was detected at day +14, and antiviral therapy with foscarnet was started immediately. At day +17, neutrophil engraftment was achieved, but not the platelet and the erythrocyte counterparts. At day +20, the patient presented fever without clinical symptoms and without an increase of C-reactive protein. Afterwards, a full blood count revealed a progressive drop in leukocytes and platelets, whereas HCMV viremia continued to increase despite antiviral therapy. The patient also developed a severe, HCMV-related hemorrhagic cystitis, which required frequent platelet transfusions, initially once daily for 5 days, and thereafter twice daily for 7 days.
As the infection appeared unresponsive to foscarnet, ganciclovir treatment was started at day +25. This switch in antiviral therapy followed an increase in viral load, reaching a maximum titer of 5100 viral genome copies/mL at day +30. Simultaneously, the clinical condition of the patient continued to deteriorate, and the leukocyte count decreased drastically. Therefore, the diagnosis of secondary graft failure due to HCMV infection was established. Because of the failure of anti-HCMV treatment, ganciclovir was withdrawn, and foscarnet therapy was resumed. The possible explanations for treatment failure include the presence of antiviral resistance mutations or an ineffective activity of the Ig administered. As the first option seemed improbable because resistance mutations usually arise later than 6 weeks after HSCT, we opted to address the latter option. Therefore, we considered using antibodies (Abs) that were more likely to inhibit the HCMV strains causing the infection in the patient. Although the diversity of circulating HCMV strains is known to be extensive (Puchhammer-Stöckl and Görzer, 2011), we hypothesized that the patient and her relatives may have been exposed to similar strains on a familial basis. As a result, we performed serological tests on the patient's mother, father and brother. We selected the patient's mother, who presented the highest anti-HCMV IgG titer, as the best donor of strain-specific HCMV Ab-rich plasma and virus-specific T-lymphocytes in this situation. At day +32, the patient's mother underwent leukapheresis. The lymphocytes were sent immediately to a cell factory to produce specific anti-HCMV T-lymphocytes. On the same day, the plasma enriched in HCMV strain-specific Abs was infused into the patient. At day +34, the viral load decreased for the first time, to 1100 viral genome copies/mL, and the patient began new conditioning for a haploidentical rescue transplant from her mother. In light of the concurrent bone marrow aplasia, the conditioning regimen was conducted solely with ATG. At day +38, the girl received a second plasma transfusion from her mother. On the day of the second transplant (day +40), complete viral clearance was documented, and the girl received 11.3 × 106 CD34 cells/kg of mother's graft. GVHD prophylaxis was performed with post-transplant cyclophosphamide, tacrolimus, and mycophenolate mofetil. The engraftment was rapid, with a neutrophil count of >500/μL achieved at day +15, and the final platelets and packed red blood cells were transfused at day +20. Since HCMV viremia remained negative during the entire post-transplant period, we discarded the use of the mother's virus-specific T-lymphocytes. The patient was discharged at day +27 in perfect clinical condition, with continuation of antiviral valganciclovir prophylaxis. Immunosuppressive treatment and antiviral prophylaxis were discontinued six months later, and the patient remains in perfect health 2 years after the second transplant.
Therapeutic interventions, HCMV load, and white blood cell count during the entire episode are summarized in Fig. 1. In addition, the HCMV strains present in the patient were characterized by high-throughput sequencing (HTS) of an archived (−80 °C) plasma sample from day +19. DNA was extracted from 800 μl of plasma using a QIAamp MinElute virus spin kit (QIAGEN, Crawley, UK). An aliquot of 50 μl of DNA was used to generate an HCMV-enriched sequencing library as described previously (Suárez et al., 2019). The library was loaded onto a NextSeq DNA sequencer (Illumina, San Diego, CA, USA), generating 150 bp paired-ended reads (Table 1). HCMV strain enumeration was performed by genotyping 12 hypervariable HCMV genes from the reads as described previously (Suárez et al., 2019). The detection of four genotypes of the RL13 gene indicated the presence of at least four HCMV strains (a strain being defined as a particular constellation of genotypes of the hypervariable genes analysed), each including one of these RL13 genotypes (Table 2). The data was also inspected for the presence of resistance mutations in the antiviral target genes (UL54 and UL97) using LoFreq (Wilm et al., 2012). Briefly, the reads were mapped to the genome of HCMV reference strain Merlin (GenBank accession no. AY446894.2), and non-synonymous variants known to confer resistance were identified. No resistance mutations were detected.Fig. 1 Information on viremia, white blood cell (WBC) count, total serum IgG and antiviral therapy. Data on HCMV viremia is expressed in viral genome copies/mL (right Y-axis). Application and duration of antiviral treatments are represented by colored bars (yellow, anti-HCMV-specific Ig; red, foscarnet; blue, ganciclovir/valganciclovir). Chronological data (X-axis) are represented in days after the first transplant. The plasma sample analysed by high-throughput sequencing (HTS) is represented by a filled blue circle.
Fig. 1
Table 1 Overview of HTS data.
Table 1Sample ID TRI-SCT1
Sample type Plasma
Days after transplant +19
HCMV load (genome copies/mL) 2100
Data deposition (ENA project no.)a PRJEB38333
Sequencing library (genome copies) 1440
Trimmed reads (no.) 5,033,480
HCMV reads (no.)b 936,773
HCMV reads; %b 19
Coverage depth (reads/nt)b 885
a The sequence dataset was purged of human reads and deposited in the European Nucleotide Archive (ENA).
b Reads mapped to the HCMV reference strain Merlin genome; HTS, high-throughput sequencing; nt, nucleotide.
Table 2 Genotyping of hypervariable HCMV genes.
Table 2
3 Discussion
Various factors are implicated in the outcome of HCMV infection in transplant patients, the first of which relates to the HCMV serological status of donor and recipient. The serological status of the first donor may have played an important role in this case, as it has been reported that using an HCMV-seronegative unrelated HSCT donor (D-) for a seropositive recipient (R+) results in reduced overall survival after myeloablative conditioning. In contrast, an HCMV seropositive donor is associated with improved event-free survival and reduced non-relapse mortality (Styczynski, 2018). Thus, the absence of an HCMV-specific immunological memory in the donor's cells may have placed the HCMV-positive recipient at a higher risk of HCMV reactivation and associated complications.
In relation to viral factors, the clinical course of the patient may have been influenced by two main factors: the presence of multiple HCMV strains or the presence of mutations conferring antiviral resistance. Infection with multiple HCMV strains is common after transplantation (Görzer et al., 2010), and is associated with delayed HCMV clearance during antiviral therapy, even in the absence of antiviral resistance mutations. It is also associated with worse clinical outcomes, including an increase in graft rejection and faster progression of disease. In principle, the mechanisms behind this increased pathogenicity can be viral, for example involving recombination and complementation of different strains within the host, or immunological, since control of multiple strains may be more challenging for the immune system. Overall, in clinically symptomatic immunocompromised hosts, the presence of multiple HCMV strains is more threatening than the presence of a single strain (Manuel et al., 2009).
In the present study, the initial failure of antiviral therapy could not be explained by the development of resistance mutations in the HCMV strains present in the patient. This finding is not surprising, as detection of resistance mutations is unusual during the first 6 weeks of antiviral therapy, whereas the sample analysed in this case was collected only 13 days after initiation of foscarnet treatment (Lurain and Chou, 2010; Springer et al., 2005).
In regard to the initial failure of the immunotherapy with commercial pooled Ig, this may have been due to the lack of particular Abs with neutralizing capacity against the HCMV strains present in the patient. Assuming that the donors providing the Ig had been exposed to strains circulating in Europe, the Abs may have been more suited for neutralizing HCMV strains commonly circulating in that region. However, it is possible that the strains present in the patient, who originated from Bangladesh, may have differed immunologically from strains in other regions of the world. The extent to which this is true is unknown, as current knowledge of HCMV genome variability is derived almost entirely from strains circulating in a few European countries. Therefore, it would be useful to assess the neutralizing capacity of commercial pooled Ig in order to avoid potential therapeutic failure, especially in patients from non-European countries. The clinical problem in this case was addressed successfully by using the Ab-rich plasma of the patient's mother in a kind of personalized immunotherapy, on the premise that both mother and daughter may have been exposed to the same pool of viral strains. In this HCMV-positive recipient with a severe viral reactivation, Ig obtained from a household donor afforded an efficacious treatment that led to complete viral clearance.
To the best of our knowledge, this is the first report of the successful use of HCMV strain-specific Ig therapy in an HSCT recipient. The efficacy of this approach as a means of prevention has been demonstrated previously only in mouse models involving murine cytomegalovirus (MCMV) reactivation after bone marrow transplantation (Martins et al., 2019). In this setting, mice received serum obtained either from latently infected (seropositive) donors or from seronegative donors. The transfer of immune serum protected mice from viral reactivation without affecting the development of GVHD. Remarkably, complete viral protection was obtained with a small volume (5 μL) of strain-specific immune serum. On the other hand, when specific donor antibodies were administered to mice infected with antigenically mismatched MCMV strains, they were not efficient in preventing viral reactivation.
Our case highlights the importance of monitoring HSCT recipients comprehensively, longitudinally and with high sensitivity for viral factors, including the presence of multiple strains and resistance mutations. Importantly, it also provides an affordable targeted immunological therapy in patients from countries with constrained access to alternative therapies.
4 Conclusion
HSCT is a complex and specific setting in which a recipient may rapidly develop viral reactivation and consequent life-threatening complications. Ab-rich plasma may represent a fast and feasible therapeutic option to overcome viral activity and allow successful engraftment of the donor's bone marrow.
Funding
No specific funding was received.
Consent for publication
The authors have obtained consent from the parents of the patient to publish individual patient data.
Declaration of competing interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgments
The sequencing work was supported by funding from the Medical Research Council (grant number MC_UU_12014/3 to AJD). | Recovered | ReactionOutcome | CC BY | 33577809 | 19,010,959 | 2021-04 |
What was the outcome of reaction 'Cystitis haemorrhagic'? | Post-HSCT graft failure due to refractory human cytomegalovirus successfully treated with haploidentical donor-derived immunoglobulins and stem cell graft infusion: A case report.
Human cytomegalovirus (HCMV) remains an important cause of transplant-related morbidity and mortality. The incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations as a result of a lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen.
We describe the case of an 8-year-old βE-thalassemic girl from Bangladesh who was seropositive for human cytomegalovirus (HCMV) and underwent hematopoietic stem cell transplantation from a HLA-matched, unrelated, HCMV-seronegative donor. Despite administering antiviral prophylaxis with commercial pooled anti-HCMV immunoglobulin (Ig) from day +1, the post-transplant course was complicated by prompt viral reactivation, and foscarnet therapy was initiated. The virus was refractory to treatment, leading rapidly to complete bone marrow failure, and targeted immunotherapy was proposed as a second-line therapy. Hypothesizing that the patient and her relatives may have been exposed to similar HCMV strains, we selected the patient's mother, who presented a high HCMV antibody titer, as the donor of virus strain-specific anti-HCMV Ig and T-lymphocytes. Complete viral clearance was achieved after two transfusions of the mother's plasma. Subsequently, the patient underwent a haploidentical rescue transplant, promptly reaching full hematological recovery.
These findings suggest that treatment with virus strain-specific Ig may offer a new therapeutic option for critically ill patients.
pmc1 Introduction
Human cytomegalovirus (HCMV) is the most significant cause of opportunistic viral infection in allogeneic hematopoietic stem cell transplantation (HSCT) and remains an important cause of transplant-related morbidity and mortality (Wagner-Drouet et al., 2019). Several studies have shown that the incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations (Ganepola et al., 2007; Zhou et al., 2009; Ozdemir et al., 2007). This is likely a consequence of delayed HCMV-specific immune reconstitution due to the lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen (Styczynski, 2018; Ljungman et al., 2014). HCMV infection can lead to pneumonia, gastroenteritis, retinitis, hepatitis, and encephalitis among HSCT recipients (Griffiths, 2012). In addition, HCMV infection of the bone marrow can result in impaired graft acceptance and, in the most extreme cases, complete graft failure (Steffens et al., 1998). Treatment of HCMV disease essentially consists of administering antiviral drugs, namely ganciclovir and foscarnet, and intravenous immunoglobulin. Although chemoprophylaxis is not generally recommended in HSCT, immunoglobulin, whether pooled or HCMV-specific, have shown some efficacy in preventing recurrent HCMV infections (Boeckh and Ljungman, 2009).
We describe a case of a pediatric HSCT recipient who had complications of HCMV recurrence and was treated successfully with targeted immunological therapy, using haploidentical donor-derived immunoglobulin (Ig).
2 Case presentation
An 8-year-old girl was diagnosed at the age of two in her country of origin (Bangladesh) as βE-thalassemic. She was treated with monthly blood transfusions and weekly iron chelation with deferoxamine, plus supplementation with folic acid. A suboptimal regimen of chelation therapy led to severe systemic siderosis, and her family travelled to Italy to undergo HSCT. Since a human leukocyte antigen (HLA)-matched related donor was unavailable at that time, an HLA-matched (10/10) unrelated donor was identified. The patient was HCMV-seropositive, whereas the donor was HCMV-seronegative, thus posing a high-risk serological combination (D-/R+) for post-transplant HCMV infection and associated complications.
The conditioning regimen consisted of fludarabine, busulfan, thiotepa, and anti-thymocyte globulin (ATG). The infused donor's bone marrow contained 3.79 × 108 total nuclear cells/kg recipient weight, and the vitality of the cells contained in the graft was 96.9%. Tacrolimus and mycophenolate mofetil were used for graft-versus-host disease (GVHD) prophylaxis. Anti-HCMV prophylaxis commenced at day +1 with commercial pooled anti-HCMV Ig (Megalotect, Biotest Pharma GmbH, Germany) at a dose of 100 U/kg twice a week.
The initial post-transplant period was uneventful. The first occurrence of HCMV viremia (600 viral genome copies/mL) was detected at day +14, and antiviral therapy with foscarnet was started immediately. At day +17, neutrophil engraftment was achieved, but not the platelet and the erythrocyte counterparts. At day +20, the patient presented fever without clinical symptoms and without an increase of C-reactive protein. Afterwards, a full blood count revealed a progressive drop in leukocytes and platelets, whereas HCMV viremia continued to increase despite antiviral therapy. The patient also developed a severe, HCMV-related hemorrhagic cystitis, which required frequent platelet transfusions, initially once daily for 5 days, and thereafter twice daily for 7 days.
As the infection appeared unresponsive to foscarnet, ganciclovir treatment was started at day +25. This switch in antiviral therapy followed an increase in viral load, reaching a maximum titer of 5100 viral genome copies/mL at day +30. Simultaneously, the clinical condition of the patient continued to deteriorate, and the leukocyte count decreased drastically. Therefore, the diagnosis of secondary graft failure due to HCMV infection was established. Because of the failure of anti-HCMV treatment, ganciclovir was withdrawn, and foscarnet therapy was resumed. The possible explanations for treatment failure include the presence of antiviral resistance mutations or an ineffective activity of the Ig administered. As the first option seemed improbable because resistance mutations usually arise later than 6 weeks after HSCT, we opted to address the latter option. Therefore, we considered using antibodies (Abs) that were more likely to inhibit the HCMV strains causing the infection in the patient. Although the diversity of circulating HCMV strains is known to be extensive (Puchhammer-Stöckl and Görzer, 2011), we hypothesized that the patient and her relatives may have been exposed to similar strains on a familial basis. As a result, we performed serological tests on the patient's mother, father and brother. We selected the patient's mother, who presented the highest anti-HCMV IgG titer, as the best donor of strain-specific HCMV Ab-rich plasma and virus-specific T-lymphocytes in this situation. At day +32, the patient's mother underwent leukapheresis. The lymphocytes were sent immediately to a cell factory to produce specific anti-HCMV T-lymphocytes. On the same day, the plasma enriched in HCMV strain-specific Abs was infused into the patient. At day +34, the viral load decreased for the first time, to 1100 viral genome copies/mL, and the patient began new conditioning for a haploidentical rescue transplant from her mother. In light of the concurrent bone marrow aplasia, the conditioning regimen was conducted solely with ATG. At day +38, the girl received a second plasma transfusion from her mother. On the day of the second transplant (day +40), complete viral clearance was documented, and the girl received 11.3 × 106 CD34 cells/kg of mother's graft. GVHD prophylaxis was performed with post-transplant cyclophosphamide, tacrolimus, and mycophenolate mofetil. The engraftment was rapid, with a neutrophil count of >500/μL achieved at day +15, and the final platelets and packed red blood cells were transfused at day +20. Since HCMV viremia remained negative during the entire post-transplant period, we discarded the use of the mother's virus-specific T-lymphocytes. The patient was discharged at day +27 in perfect clinical condition, with continuation of antiviral valganciclovir prophylaxis. Immunosuppressive treatment and antiviral prophylaxis were discontinued six months later, and the patient remains in perfect health 2 years after the second transplant.
Therapeutic interventions, HCMV load, and white blood cell count during the entire episode are summarized in Fig. 1. In addition, the HCMV strains present in the patient were characterized by high-throughput sequencing (HTS) of an archived (−80 °C) plasma sample from day +19. DNA was extracted from 800 μl of plasma using a QIAamp MinElute virus spin kit (QIAGEN, Crawley, UK). An aliquot of 50 μl of DNA was used to generate an HCMV-enriched sequencing library as described previously (Suárez et al., 2019). The library was loaded onto a NextSeq DNA sequencer (Illumina, San Diego, CA, USA), generating 150 bp paired-ended reads (Table 1). HCMV strain enumeration was performed by genotyping 12 hypervariable HCMV genes from the reads as described previously (Suárez et al., 2019). The detection of four genotypes of the RL13 gene indicated the presence of at least four HCMV strains (a strain being defined as a particular constellation of genotypes of the hypervariable genes analysed), each including one of these RL13 genotypes (Table 2). The data was also inspected for the presence of resistance mutations in the antiviral target genes (UL54 and UL97) using LoFreq (Wilm et al., 2012). Briefly, the reads were mapped to the genome of HCMV reference strain Merlin (GenBank accession no. AY446894.2), and non-synonymous variants known to confer resistance were identified. No resistance mutations were detected.Fig. 1 Information on viremia, white blood cell (WBC) count, total serum IgG and antiviral therapy. Data on HCMV viremia is expressed in viral genome copies/mL (right Y-axis). Application and duration of antiviral treatments are represented by colored bars (yellow, anti-HCMV-specific Ig; red, foscarnet; blue, ganciclovir/valganciclovir). Chronological data (X-axis) are represented in days after the first transplant. The plasma sample analysed by high-throughput sequencing (HTS) is represented by a filled blue circle.
Fig. 1
Table 1 Overview of HTS data.
Table 1Sample ID TRI-SCT1
Sample type Plasma
Days after transplant +19
HCMV load (genome copies/mL) 2100
Data deposition (ENA project no.)a PRJEB38333
Sequencing library (genome copies) 1440
Trimmed reads (no.) 5,033,480
HCMV reads (no.)b 936,773
HCMV reads; %b 19
Coverage depth (reads/nt)b 885
a The sequence dataset was purged of human reads and deposited in the European Nucleotide Archive (ENA).
b Reads mapped to the HCMV reference strain Merlin genome; HTS, high-throughput sequencing; nt, nucleotide.
Table 2 Genotyping of hypervariable HCMV genes.
Table 2
3 Discussion
Various factors are implicated in the outcome of HCMV infection in transplant patients, the first of which relates to the HCMV serological status of donor and recipient. The serological status of the first donor may have played an important role in this case, as it has been reported that using an HCMV-seronegative unrelated HSCT donor (D-) for a seropositive recipient (R+) results in reduced overall survival after myeloablative conditioning. In contrast, an HCMV seropositive donor is associated with improved event-free survival and reduced non-relapse mortality (Styczynski, 2018). Thus, the absence of an HCMV-specific immunological memory in the donor's cells may have placed the HCMV-positive recipient at a higher risk of HCMV reactivation and associated complications.
In relation to viral factors, the clinical course of the patient may have been influenced by two main factors: the presence of multiple HCMV strains or the presence of mutations conferring antiviral resistance. Infection with multiple HCMV strains is common after transplantation (Görzer et al., 2010), and is associated with delayed HCMV clearance during antiviral therapy, even in the absence of antiviral resistance mutations. It is also associated with worse clinical outcomes, including an increase in graft rejection and faster progression of disease. In principle, the mechanisms behind this increased pathogenicity can be viral, for example involving recombination and complementation of different strains within the host, or immunological, since control of multiple strains may be more challenging for the immune system. Overall, in clinically symptomatic immunocompromised hosts, the presence of multiple HCMV strains is more threatening than the presence of a single strain (Manuel et al., 2009).
In the present study, the initial failure of antiviral therapy could not be explained by the development of resistance mutations in the HCMV strains present in the patient. This finding is not surprising, as detection of resistance mutations is unusual during the first 6 weeks of antiviral therapy, whereas the sample analysed in this case was collected only 13 days after initiation of foscarnet treatment (Lurain and Chou, 2010; Springer et al., 2005).
In regard to the initial failure of the immunotherapy with commercial pooled Ig, this may have been due to the lack of particular Abs with neutralizing capacity against the HCMV strains present in the patient. Assuming that the donors providing the Ig had been exposed to strains circulating in Europe, the Abs may have been more suited for neutralizing HCMV strains commonly circulating in that region. However, it is possible that the strains present in the patient, who originated from Bangladesh, may have differed immunologically from strains in other regions of the world. The extent to which this is true is unknown, as current knowledge of HCMV genome variability is derived almost entirely from strains circulating in a few European countries. Therefore, it would be useful to assess the neutralizing capacity of commercial pooled Ig in order to avoid potential therapeutic failure, especially in patients from non-European countries. The clinical problem in this case was addressed successfully by using the Ab-rich plasma of the patient's mother in a kind of personalized immunotherapy, on the premise that both mother and daughter may have been exposed to the same pool of viral strains. In this HCMV-positive recipient with a severe viral reactivation, Ig obtained from a household donor afforded an efficacious treatment that led to complete viral clearance.
To the best of our knowledge, this is the first report of the successful use of HCMV strain-specific Ig therapy in an HSCT recipient. The efficacy of this approach as a means of prevention has been demonstrated previously only in mouse models involving murine cytomegalovirus (MCMV) reactivation after bone marrow transplantation (Martins et al., 2019). In this setting, mice received serum obtained either from latently infected (seropositive) donors or from seronegative donors. The transfer of immune serum protected mice from viral reactivation without affecting the development of GVHD. Remarkably, complete viral protection was obtained with a small volume (5 μL) of strain-specific immune serum. On the other hand, when specific donor antibodies were administered to mice infected with antigenically mismatched MCMV strains, they were not efficient in preventing viral reactivation.
Our case highlights the importance of monitoring HSCT recipients comprehensively, longitudinally and with high sensitivity for viral factors, including the presence of multiple strains and resistance mutations. Importantly, it also provides an affordable targeted immunological therapy in patients from countries with constrained access to alternative therapies.
4 Conclusion
HSCT is a complex and specific setting in which a recipient may rapidly develop viral reactivation and consequent life-threatening complications. Ab-rich plasma may represent a fast and feasible therapeutic option to overcome viral activity and allow successful engraftment of the donor's bone marrow.
Funding
No specific funding was received.
Consent for publication
The authors have obtained consent from the parents of the patient to publish individual patient data.
Declaration of competing interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgments
The sequencing work was supported by funding from the Medical Research Council (grant number MC_UU_12014/3 to AJD). | Recovered | ReactionOutcome | CC BY | 33577809 | 19,256,788 | 2021-04 |
What was the outcome of reaction 'Cytomegalovirus infection reactivation'? | Post-HSCT graft failure due to refractory human cytomegalovirus successfully treated with haploidentical donor-derived immunoglobulins and stem cell graft infusion: A case report.
Human cytomegalovirus (HCMV) remains an important cause of transplant-related morbidity and mortality. The incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations as a result of a lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen.
We describe the case of an 8-year-old βE-thalassemic girl from Bangladesh who was seropositive for human cytomegalovirus (HCMV) and underwent hematopoietic stem cell transplantation from a HLA-matched, unrelated, HCMV-seronegative donor. Despite administering antiviral prophylaxis with commercial pooled anti-HCMV immunoglobulin (Ig) from day +1, the post-transplant course was complicated by prompt viral reactivation, and foscarnet therapy was initiated. The virus was refractory to treatment, leading rapidly to complete bone marrow failure, and targeted immunotherapy was proposed as a second-line therapy. Hypothesizing that the patient and her relatives may have been exposed to similar HCMV strains, we selected the patient's mother, who presented a high HCMV antibody titer, as the donor of virus strain-specific anti-HCMV Ig and T-lymphocytes. Complete viral clearance was achieved after two transfusions of the mother's plasma. Subsequently, the patient underwent a haploidentical rescue transplant, promptly reaching full hematological recovery.
These findings suggest that treatment with virus strain-specific Ig may offer a new therapeutic option for critically ill patients.
pmc1 Introduction
Human cytomegalovirus (HCMV) is the most significant cause of opportunistic viral infection in allogeneic hematopoietic stem cell transplantation (HSCT) and remains an important cause of transplant-related morbidity and mortality (Wagner-Drouet et al., 2019). Several studies have shown that the incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations (Ganepola et al., 2007; Zhou et al., 2009; Ozdemir et al., 2007). This is likely a consequence of delayed HCMV-specific immune reconstitution due to the lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen (Styczynski, 2018; Ljungman et al., 2014). HCMV infection can lead to pneumonia, gastroenteritis, retinitis, hepatitis, and encephalitis among HSCT recipients (Griffiths, 2012). In addition, HCMV infection of the bone marrow can result in impaired graft acceptance and, in the most extreme cases, complete graft failure (Steffens et al., 1998). Treatment of HCMV disease essentially consists of administering antiviral drugs, namely ganciclovir and foscarnet, and intravenous immunoglobulin. Although chemoprophylaxis is not generally recommended in HSCT, immunoglobulin, whether pooled or HCMV-specific, have shown some efficacy in preventing recurrent HCMV infections (Boeckh and Ljungman, 2009).
We describe a case of a pediatric HSCT recipient who had complications of HCMV recurrence and was treated successfully with targeted immunological therapy, using haploidentical donor-derived immunoglobulin (Ig).
2 Case presentation
An 8-year-old girl was diagnosed at the age of two in her country of origin (Bangladesh) as βE-thalassemic. She was treated with monthly blood transfusions and weekly iron chelation with deferoxamine, plus supplementation with folic acid. A suboptimal regimen of chelation therapy led to severe systemic siderosis, and her family travelled to Italy to undergo HSCT. Since a human leukocyte antigen (HLA)-matched related donor was unavailable at that time, an HLA-matched (10/10) unrelated donor was identified. The patient was HCMV-seropositive, whereas the donor was HCMV-seronegative, thus posing a high-risk serological combination (D-/R+) for post-transplant HCMV infection and associated complications.
The conditioning regimen consisted of fludarabine, busulfan, thiotepa, and anti-thymocyte globulin (ATG). The infused donor's bone marrow contained 3.79 × 108 total nuclear cells/kg recipient weight, and the vitality of the cells contained in the graft was 96.9%. Tacrolimus and mycophenolate mofetil were used for graft-versus-host disease (GVHD) prophylaxis. Anti-HCMV prophylaxis commenced at day +1 with commercial pooled anti-HCMV Ig (Megalotect, Biotest Pharma GmbH, Germany) at a dose of 100 U/kg twice a week.
The initial post-transplant period was uneventful. The first occurrence of HCMV viremia (600 viral genome copies/mL) was detected at day +14, and antiviral therapy with foscarnet was started immediately. At day +17, neutrophil engraftment was achieved, but not the platelet and the erythrocyte counterparts. At day +20, the patient presented fever without clinical symptoms and without an increase of C-reactive protein. Afterwards, a full blood count revealed a progressive drop in leukocytes and platelets, whereas HCMV viremia continued to increase despite antiviral therapy. The patient also developed a severe, HCMV-related hemorrhagic cystitis, which required frequent platelet transfusions, initially once daily for 5 days, and thereafter twice daily for 7 days.
As the infection appeared unresponsive to foscarnet, ganciclovir treatment was started at day +25. This switch in antiviral therapy followed an increase in viral load, reaching a maximum titer of 5100 viral genome copies/mL at day +30. Simultaneously, the clinical condition of the patient continued to deteriorate, and the leukocyte count decreased drastically. Therefore, the diagnosis of secondary graft failure due to HCMV infection was established. Because of the failure of anti-HCMV treatment, ganciclovir was withdrawn, and foscarnet therapy was resumed. The possible explanations for treatment failure include the presence of antiviral resistance mutations or an ineffective activity of the Ig administered. As the first option seemed improbable because resistance mutations usually arise later than 6 weeks after HSCT, we opted to address the latter option. Therefore, we considered using antibodies (Abs) that were more likely to inhibit the HCMV strains causing the infection in the patient. Although the diversity of circulating HCMV strains is known to be extensive (Puchhammer-Stöckl and Görzer, 2011), we hypothesized that the patient and her relatives may have been exposed to similar strains on a familial basis. As a result, we performed serological tests on the patient's mother, father and brother. We selected the patient's mother, who presented the highest anti-HCMV IgG titer, as the best donor of strain-specific HCMV Ab-rich plasma and virus-specific T-lymphocytes in this situation. At day +32, the patient's mother underwent leukapheresis. The lymphocytes were sent immediately to a cell factory to produce specific anti-HCMV T-lymphocytes. On the same day, the plasma enriched in HCMV strain-specific Abs was infused into the patient. At day +34, the viral load decreased for the first time, to 1100 viral genome copies/mL, and the patient began new conditioning for a haploidentical rescue transplant from her mother. In light of the concurrent bone marrow aplasia, the conditioning regimen was conducted solely with ATG. At day +38, the girl received a second plasma transfusion from her mother. On the day of the second transplant (day +40), complete viral clearance was documented, and the girl received 11.3 × 106 CD34 cells/kg of mother's graft. GVHD prophylaxis was performed with post-transplant cyclophosphamide, tacrolimus, and mycophenolate mofetil. The engraftment was rapid, with a neutrophil count of >500/μL achieved at day +15, and the final platelets and packed red blood cells were transfused at day +20. Since HCMV viremia remained negative during the entire post-transplant period, we discarded the use of the mother's virus-specific T-lymphocytes. The patient was discharged at day +27 in perfect clinical condition, with continuation of antiviral valganciclovir prophylaxis. Immunosuppressive treatment and antiviral prophylaxis were discontinued six months later, and the patient remains in perfect health 2 years after the second transplant.
Therapeutic interventions, HCMV load, and white blood cell count during the entire episode are summarized in Fig. 1. In addition, the HCMV strains present in the patient were characterized by high-throughput sequencing (HTS) of an archived (−80 °C) plasma sample from day +19. DNA was extracted from 800 μl of plasma using a QIAamp MinElute virus spin kit (QIAGEN, Crawley, UK). An aliquot of 50 μl of DNA was used to generate an HCMV-enriched sequencing library as described previously (Suárez et al., 2019). The library was loaded onto a NextSeq DNA sequencer (Illumina, San Diego, CA, USA), generating 150 bp paired-ended reads (Table 1). HCMV strain enumeration was performed by genotyping 12 hypervariable HCMV genes from the reads as described previously (Suárez et al., 2019). The detection of four genotypes of the RL13 gene indicated the presence of at least four HCMV strains (a strain being defined as a particular constellation of genotypes of the hypervariable genes analysed), each including one of these RL13 genotypes (Table 2). The data was also inspected for the presence of resistance mutations in the antiviral target genes (UL54 and UL97) using LoFreq (Wilm et al., 2012). Briefly, the reads were mapped to the genome of HCMV reference strain Merlin (GenBank accession no. AY446894.2), and non-synonymous variants known to confer resistance were identified. No resistance mutations were detected.Fig. 1 Information on viremia, white blood cell (WBC) count, total serum IgG and antiviral therapy. Data on HCMV viremia is expressed in viral genome copies/mL (right Y-axis). Application and duration of antiviral treatments are represented by colored bars (yellow, anti-HCMV-specific Ig; red, foscarnet; blue, ganciclovir/valganciclovir). Chronological data (X-axis) are represented in days after the first transplant. The plasma sample analysed by high-throughput sequencing (HTS) is represented by a filled blue circle.
Fig. 1
Table 1 Overview of HTS data.
Table 1Sample ID TRI-SCT1
Sample type Plasma
Days after transplant +19
HCMV load (genome copies/mL) 2100
Data deposition (ENA project no.)a PRJEB38333
Sequencing library (genome copies) 1440
Trimmed reads (no.) 5,033,480
HCMV reads (no.)b 936,773
HCMV reads; %b 19
Coverage depth (reads/nt)b 885
a The sequence dataset was purged of human reads and deposited in the European Nucleotide Archive (ENA).
b Reads mapped to the HCMV reference strain Merlin genome; HTS, high-throughput sequencing; nt, nucleotide.
Table 2 Genotyping of hypervariable HCMV genes.
Table 2
3 Discussion
Various factors are implicated in the outcome of HCMV infection in transplant patients, the first of which relates to the HCMV serological status of donor and recipient. The serological status of the first donor may have played an important role in this case, as it has been reported that using an HCMV-seronegative unrelated HSCT donor (D-) for a seropositive recipient (R+) results in reduced overall survival after myeloablative conditioning. In contrast, an HCMV seropositive donor is associated with improved event-free survival and reduced non-relapse mortality (Styczynski, 2018). Thus, the absence of an HCMV-specific immunological memory in the donor's cells may have placed the HCMV-positive recipient at a higher risk of HCMV reactivation and associated complications.
In relation to viral factors, the clinical course of the patient may have been influenced by two main factors: the presence of multiple HCMV strains or the presence of mutations conferring antiviral resistance. Infection with multiple HCMV strains is common after transplantation (Görzer et al., 2010), and is associated with delayed HCMV clearance during antiviral therapy, even in the absence of antiviral resistance mutations. It is also associated with worse clinical outcomes, including an increase in graft rejection and faster progression of disease. In principle, the mechanisms behind this increased pathogenicity can be viral, for example involving recombination and complementation of different strains within the host, or immunological, since control of multiple strains may be more challenging for the immune system. Overall, in clinically symptomatic immunocompromised hosts, the presence of multiple HCMV strains is more threatening than the presence of a single strain (Manuel et al., 2009).
In the present study, the initial failure of antiviral therapy could not be explained by the development of resistance mutations in the HCMV strains present in the patient. This finding is not surprising, as detection of resistance mutations is unusual during the first 6 weeks of antiviral therapy, whereas the sample analysed in this case was collected only 13 days after initiation of foscarnet treatment (Lurain and Chou, 2010; Springer et al., 2005).
In regard to the initial failure of the immunotherapy with commercial pooled Ig, this may have been due to the lack of particular Abs with neutralizing capacity against the HCMV strains present in the patient. Assuming that the donors providing the Ig had been exposed to strains circulating in Europe, the Abs may have been more suited for neutralizing HCMV strains commonly circulating in that region. However, it is possible that the strains present in the patient, who originated from Bangladesh, may have differed immunologically from strains in other regions of the world. The extent to which this is true is unknown, as current knowledge of HCMV genome variability is derived almost entirely from strains circulating in a few European countries. Therefore, it would be useful to assess the neutralizing capacity of commercial pooled Ig in order to avoid potential therapeutic failure, especially in patients from non-European countries. The clinical problem in this case was addressed successfully by using the Ab-rich plasma of the patient's mother in a kind of personalized immunotherapy, on the premise that both mother and daughter may have been exposed to the same pool of viral strains. In this HCMV-positive recipient with a severe viral reactivation, Ig obtained from a household donor afforded an efficacious treatment that led to complete viral clearance.
To the best of our knowledge, this is the first report of the successful use of HCMV strain-specific Ig therapy in an HSCT recipient. The efficacy of this approach as a means of prevention has been demonstrated previously only in mouse models involving murine cytomegalovirus (MCMV) reactivation after bone marrow transplantation (Martins et al., 2019). In this setting, mice received serum obtained either from latently infected (seropositive) donors or from seronegative donors. The transfer of immune serum protected mice from viral reactivation without affecting the development of GVHD. Remarkably, complete viral protection was obtained with a small volume (5 μL) of strain-specific immune serum. On the other hand, when specific donor antibodies were administered to mice infected with antigenically mismatched MCMV strains, they were not efficient in preventing viral reactivation.
Our case highlights the importance of monitoring HSCT recipients comprehensively, longitudinally and with high sensitivity for viral factors, including the presence of multiple strains and resistance mutations. Importantly, it also provides an affordable targeted immunological therapy in patients from countries with constrained access to alternative therapies.
4 Conclusion
HSCT is a complex and specific setting in which a recipient may rapidly develop viral reactivation and consequent life-threatening complications. Ab-rich plasma may represent a fast and feasible therapeutic option to overcome viral activity and allow successful engraftment of the donor's bone marrow.
Funding
No specific funding was received.
Consent for publication
The authors have obtained consent from the parents of the patient to publish individual patient data.
Declaration of competing interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgments
The sequencing work was supported by funding from the Medical Research Council (grant number MC_UU_12014/3 to AJD). | Recovered | ReactionOutcome | CC BY | 33577809 | 19,256,788 | 2021-04 |
What was the outcome of reaction 'Cytomegalovirus infection'? | Post-HSCT graft failure due to refractory human cytomegalovirus successfully treated with haploidentical donor-derived immunoglobulins and stem cell graft infusion: A case report.
Human cytomegalovirus (HCMV) remains an important cause of transplant-related morbidity and mortality. The incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations as a result of a lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen.
We describe the case of an 8-year-old βE-thalassemic girl from Bangladesh who was seropositive for human cytomegalovirus (HCMV) and underwent hematopoietic stem cell transplantation from a HLA-matched, unrelated, HCMV-seronegative donor. Despite administering antiviral prophylaxis with commercial pooled anti-HCMV immunoglobulin (Ig) from day +1, the post-transplant course was complicated by prompt viral reactivation, and foscarnet therapy was initiated. The virus was refractory to treatment, leading rapidly to complete bone marrow failure, and targeted immunotherapy was proposed as a second-line therapy. Hypothesizing that the patient and her relatives may have been exposed to similar HCMV strains, we selected the patient's mother, who presented a high HCMV antibody titer, as the donor of virus strain-specific anti-HCMV Ig and T-lymphocytes. Complete viral clearance was achieved after two transfusions of the mother's plasma. Subsequently, the patient underwent a haploidentical rescue transplant, promptly reaching full hematological recovery.
These findings suggest that treatment with virus strain-specific Ig may offer a new therapeutic option for critically ill patients.
pmc1 Introduction
Human cytomegalovirus (HCMV) is the most significant cause of opportunistic viral infection in allogeneic hematopoietic stem cell transplantation (HSCT) and remains an important cause of transplant-related morbidity and mortality (Wagner-Drouet et al., 2019). Several studies have shown that the incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations (Ganepola et al., 2007; Zhou et al., 2009; Ozdemir et al., 2007). This is likely a consequence of delayed HCMV-specific immune reconstitution due to the lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen (Styczynski, 2018; Ljungman et al., 2014). HCMV infection can lead to pneumonia, gastroenteritis, retinitis, hepatitis, and encephalitis among HSCT recipients (Griffiths, 2012). In addition, HCMV infection of the bone marrow can result in impaired graft acceptance and, in the most extreme cases, complete graft failure (Steffens et al., 1998). Treatment of HCMV disease essentially consists of administering antiviral drugs, namely ganciclovir and foscarnet, and intravenous immunoglobulin. Although chemoprophylaxis is not generally recommended in HSCT, immunoglobulin, whether pooled or HCMV-specific, have shown some efficacy in preventing recurrent HCMV infections (Boeckh and Ljungman, 2009).
We describe a case of a pediatric HSCT recipient who had complications of HCMV recurrence and was treated successfully with targeted immunological therapy, using haploidentical donor-derived immunoglobulin (Ig).
2 Case presentation
An 8-year-old girl was diagnosed at the age of two in her country of origin (Bangladesh) as βE-thalassemic. She was treated with monthly blood transfusions and weekly iron chelation with deferoxamine, plus supplementation with folic acid. A suboptimal regimen of chelation therapy led to severe systemic siderosis, and her family travelled to Italy to undergo HSCT. Since a human leukocyte antigen (HLA)-matched related donor was unavailable at that time, an HLA-matched (10/10) unrelated donor was identified. The patient was HCMV-seropositive, whereas the donor was HCMV-seronegative, thus posing a high-risk serological combination (D-/R+) for post-transplant HCMV infection and associated complications.
The conditioning regimen consisted of fludarabine, busulfan, thiotepa, and anti-thymocyte globulin (ATG). The infused donor's bone marrow contained 3.79 × 108 total nuclear cells/kg recipient weight, and the vitality of the cells contained in the graft was 96.9%. Tacrolimus and mycophenolate mofetil were used for graft-versus-host disease (GVHD) prophylaxis. Anti-HCMV prophylaxis commenced at day +1 with commercial pooled anti-HCMV Ig (Megalotect, Biotest Pharma GmbH, Germany) at a dose of 100 U/kg twice a week.
The initial post-transplant period was uneventful. The first occurrence of HCMV viremia (600 viral genome copies/mL) was detected at day +14, and antiviral therapy with foscarnet was started immediately. At day +17, neutrophil engraftment was achieved, but not the platelet and the erythrocyte counterparts. At day +20, the patient presented fever without clinical symptoms and without an increase of C-reactive protein. Afterwards, a full blood count revealed a progressive drop in leukocytes and platelets, whereas HCMV viremia continued to increase despite antiviral therapy. The patient also developed a severe, HCMV-related hemorrhagic cystitis, which required frequent platelet transfusions, initially once daily for 5 days, and thereafter twice daily for 7 days.
As the infection appeared unresponsive to foscarnet, ganciclovir treatment was started at day +25. This switch in antiviral therapy followed an increase in viral load, reaching a maximum titer of 5100 viral genome copies/mL at day +30. Simultaneously, the clinical condition of the patient continued to deteriorate, and the leukocyte count decreased drastically. Therefore, the diagnosis of secondary graft failure due to HCMV infection was established. Because of the failure of anti-HCMV treatment, ganciclovir was withdrawn, and foscarnet therapy was resumed. The possible explanations for treatment failure include the presence of antiviral resistance mutations or an ineffective activity of the Ig administered. As the first option seemed improbable because resistance mutations usually arise later than 6 weeks after HSCT, we opted to address the latter option. Therefore, we considered using antibodies (Abs) that were more likely to inhibit the HCMV strains causing the infection in the patient. Although the diversity of circulating HCMV strains is known to be extensive (Puchhammer-Stöckl and Görzer, 2011), we hypothesized that the patient and her relatives may have been exposed to similar strains on a familial basis. As a result, we performed serological tests on the patient's mother, father and brother. We selected the patient's mother, who presented the highest anti-HCMV IgG titer, as the best donor of strain-specific HCMV Ab-rich plasma and virus-specific T-lymphocytes in this situation. At day +32, the patient's mother underwent leukapheresis. The lymphocytes were sent immediately to a cell factory to produce specific anti-HCMV T-lymphocytes. On the same day, the plasma enriched in HCMV strain-specific Abs was infused into the patient. At day +34, the viral load decreased for the first time, to 1100 viral genome copies/mL, and the patient began new conditioning for a haploidentical rescue transplant from her mother. In light of the concurrent bone marrow aplasia, the conditioning regimen was conducted solely with ATG. At day +38, the girl received a second plasma transfusion from her mother. On the day of the second transplant (day +40), complete viral clearance was documented, and the girl received 11.3 × 106 CD34 cells/kg of mother's graft. GVHD prophylaxis was performed with post-transplant cyclophosphamide, tacrolimus, and mycophenolate mofetil. The engraftment was rapid, with a neutrophil count of >500/μL achieved at day +15, and the final platelets and packed red blood cells were transfused at day +20. Since HCMV viremia remained negative during the entire post-transplant period, we discarded the use of the mother's virus-specific T-lymphocytes. The patient was discharged at day +27 in perfect clinical condition, with continuation of antiviral valganciclovir prophylaxis. Immunosuppressive treatment and antiviral prophylaxis were discontinued six months later, and the patient remains in perfect health 2 years after the second transplant.
Therapeutic interventions, HCMV load, and white blood cell count during the entire episode are summarized in Fig. 1. In addition, the HCMV strains present in the patient were characterized by high-throughput sequencing (HTS) of an archived (−80 °C) plasma sample from day +19. DNA was extracted from 800 μl of plasma using a QIAamp MinElute virus spin kit (QIAGEN, Crawley, UK). An aliquot of 50 μl of DNA was used to generate an HCMV-enriched sequencing library as described previously (Suárez et al., 2019). The library was loaded onto a NextSeq DNA sequencer (Illumina, San Diego, CA, USA), generating 150 bp paired-ended reads (Table 1). HCMV strain enumeration was performed by genotyping 12 hypervariable HCMV genes from the reads as described previously (Suárez et al., 2019). The detection of four genotypes of the RL13 gene indicated the presence of at least four HCMV strains (a strain being defined as a particular constellation of genotypes of the hypervariable genes analysed), each including one of these RL13 genotypes (Table 2). The data was also inspected for the presence of resistance mutations in the antiviral target genes (UL54 and UL97) using LoFreq (Wilm et al., 2012). Briefly, the reads were mapped to the genome of HCMV reference strain Merlin (GenBank accession no. AY446894.2), and non-synonymous variants known to confer resistance were identified. No resistance mutations were detected.Fig. 1 Information on viremia, white blood cell (WBC) count, total serum IgG and antiviral therapy. Data on HCMV viremia is expressed in viral genome copies/mL (right Y-axis). Application and duration of antiviral treatments are represented by colored bars (yellow, anti-HCMV-specific Ig; red, foscarnet; blue, ganciclovir/valganciclovir). Chronological data (X-axis) are represented in days after the first transplant. The plasma sample analysed by high-throughput sequencing (HTS) is represented by a filled blue circle.
Fig. 1
Table 1 Overview of HTS data.
Table 1Sample ID TRI-SCT1
Sample type Plasma
Days after transplant +19
HCMV load (genome copies/mL) 2100
Data deposition (ENA project no.)a PRJEB38333
Sequencing library (genome copies) 1440
Trimmed reads (no.) 5,033,480
HCMV reads (no.)b 936,773
HCMV reads; %b 19
Coverage depth (reads/nt)b 885
a The sequence dataset was purged of human reads and deposited in the European Nucleotide Archive (ENA).
b Reads mapped to the HCMV reference strain Merlin genome; HTS, high-throughput sequencing; nt, nucleotide.
Table 2 Genotyping of hypervariable HCMV genes.
Table 2
3 Discussion
Various factors are implicated in the outcome of HCMV infection in transplant patients, the first of which relates to the HCMV serological status of donor and recipient. The serological status of the first donor may have played an important role in this case, as it has been reported that using an HCMV-seronegative unrelated HSCT donor (D-) for a seropositive recipient (R+) results in reduced overall survival after myeloablative conditioning. In contrast, an HCMV seropositive donor is associated with improved event-free survival and reduced non-relapse mortality (Styczynski, 2018). Thus, the absence of an HCMV-specific immunological memory in the donor's cells may have placed the HCMV-positive recipient at a higher risk of HCMV reactivation and associated complications.
In relation to viral factors, the clinical course of the patient may have been influenced by two main factors: the presence of multiple HCMV strains or the presence of mutations conferring antiviral resistance. Infection with multiple HCMV strains is common after transplantation (Görzer et al., 2010), and is associated with delayed HCMV clearance during antiviral therapy, even in the absence of antiviral resistance mutations. It is also associated with worse clinical outcomes, including an increase in graft rejection and faster progression of disease. In principle, the mechanisms behind this increased pathogenicity can be viral, for example involving recombination and complementation of different strains within the host, or immunological, since control of multiple strains may be more challenging for the immune system. Overall, in clinically symptomatic immunocompromised hosts, the presence of multiple HCMV strains is more threatening than the presence of a single strain (Manuel et al., 2009).
In the present study, the initial failure of antiviral therapy could not be explained by the development of resistance mutations in the HCMV strains present in the patient. This finding is not surprising, as detection of resistance mutations is unusual during the first 6 weeks of antiviral therapy, whereas the sample analysed in this case was collected only 13 days after initiation of foscarnet treatment (Lurain and Chou, 2010; Springer et al., 2005).
In regard to the initial failure of the immunotherapy with commercial pooled Ig, this may have been due to the lack of particular Abs with neutralizing capacity against the HCMV strains present in the patient. Assuming that the donors providing the Ig had been exposed to strains circulating in Europe, the Abs may have been more suited for neutralizing HCMV strains commonly circulating in that region. However, it is possible that the strains present in the patient, who originated from Bangladesh, may have differed immunologically from strains in other regions of the world. The extent to which this is true is unknown, as current knowledge of HCMV genome variability is derived almost entirely from strains circulating in a few European countries. Therefore, it would be useful to assess the neutralizing capacity of commercial pooled Ig in order to avoid potential therapeutic failure, especially in patients from non-European countries. The clinical problem in this case was addressed successfully by using the Ab-rich plasma of the patient's mother in a kind of personalized immunotherapy, on the premise that both mother and daughter may have been exposed to the same pool of viral strains. In this HCMV-positive recipient with a severe viral reactivation, Ig obtained from a household donor afforded an efficacious treatment that led to complete viral clearance.
To the best of our knowledge, this is the first report of the successful use of HCMV strain-specific Ig therapy in an HSCT recipient. The efficacy of this approach as a means of prevention has been demonstrated previously only in mouse models involving murine cytomegalovirus (MCMV) reactivation after bone marrow transplantation (Martins et al., 2019). In this setting, mice received serum obtained either from latently infected (seropositive) donors or from seronegative donors. The transfer of immune serum protected mice from viral reactivation without affecting the development of GVHD. Remarkably, complete viral protection was obtained with a small volume (5 μL) of strain-specific immune serum. On the other hand, when specific donor antibodies were administered to mice infected with antigenically mismatched MCMV strains, they were not efficient in preventing viral reactivation.
Our case highlights the importance of monitoring HSCT recipients comprehensively, longitudinally and with high sensitivity for viral factors, including the presence of multiple strains and resistance mutations. Importantly, it also provides an affordable targeted immunological therapy in patients from countries with constrained access to alternative therapies.
4 Conclusion
HSCT is a complex and specific setting in which a recipient may rapidly develop viral reactivation and consequent life-threatening complications. Ab-rich plasma may represent a fast and feasible therapeutic option to overcome viral activity and allow successful engraftment of the donor's bone marrow.
Funding
No specific funding was received.
Consent for publication
The authors have obtained consent from the parents of the patient to publish individual patient data.
Declaration of competing interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgments
The sequencing work was supported by funding from the Medical Research Council (grant number MC_UU_12014/3 to AJD). | Recovered | ReactionOutcome | CC BY | 33577809 | 19,010,959 | 2021-04 |
What was the outcome of reaction 'Cytomegalovirus viraemia'? | Post-HSCT graft failure due to refractory human cytomegalovirus successfully treated with haploidentical donor-derived immunoglobulins and stem cell graft infusion: A case report.
Human cytomegalovirus (HCMV) remains an important cause of transplant-related morbidity and mortality. The incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations as a result of a lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen.
We describe the case of an 8-year-old βE-thalassemic girl from Bangladesh who was seropositive for human cytomegalovirus (HCMV) and underwent hematopoietic stem cell transplantation from a HLA-matched, unrelated, HCMV-seronegative donor. Despite administering antiviral prophylaxis with commercial pooled anti-HCMV immunoglobulin (Ig) from day +1, the post-transplant course was complicated by prompt viral reactivation, and foscarnet therapy was initiated. The virus was refractory to treatment, leading rapidly to complete bone marrow failure, and targeted immunotherapy was proposed as a second-line therapy. Hypothesizing that the patient and her relatives may have been exposed to similar HCMV strains, we selected the patient's mother, who presented a high HCMV antibody titer, as the donor of virus strain-specific anti-HCMV Ig and T-lymphocytes. Complete viral clearance was achieved after two transfusions of the mother's plasma. Subsequently, the patient underwent a haploidentical rescue transplant, promptly reaching full hematological recovery.
These findings suggest that treatment with virus strain-specific Ig may offer a new therapeutic option for critically ill patients.
pmc1 Introduction
Human cytomegalovirus (HCMV) is the most significant cause of opportunistic viral infection in allogeneic hematopoietic stem cell transplantation (HSCT) and remains an important cause of transplant-related morbidity and mortality (Wagner-Drouet et al., 2019). Several studies have shown that the incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations (Ganepola et al., 2007; Zhou et al., 2009; Ozdemir et al., 2007). This is likely a consequence of delayed HCMV-specific immune reconstitution due to the lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen (Styczynski, 2018; Ljungman et al., 2014). HCMV infection can lead to pneumonia, gastroenteritis, retinitis, hepatitis, and encephalitis among HSCT recipients (Griffiths, 2012). In addition, HCMV infection of the bone marrow can result in impaired graft acceptance and, in the most extreme cases, complete graft failure (Steffens et al., 1998). Treatment of HCMV disease essentially consists of administering antiviral drugs, namely ganciclovir and foscarnet, and intravenous immunoglobulin. Although chemoprophylaxis is not generally recommended in HSCT, immunoglobulin, whether pooled or HCMV-specific, have shown some efficacy in preventing recurrent HCMV infections (Boeckh and Ljungman, 2009).
We describe a case of a pediatric HSCT recipient who had complications of HCMV recurrence and was treated successfully with targeted immunological therapy, using haploidentical donor-derived immunoglobulin (Ig).
2 Case presentation
An 8-year-old girl was diagnosed at the age of two in her country of origin (Bangladesh) as βE-thalassemic. She was treated with monthly blood transfusions and weekly iron chelation with deferoxamine, plus supplementation with folic acid. A suboptimal regimen of chelation therapy led to severe systemic siderosis, and her family travelled to Italy to undergo HSCT. Since a human leukocyte antigen (HLA)-matched related donor was unavailable at that time, an HLA-matched (10/10) unrelated donor was identified. The patient was HCMV-seropositive, whereas the donor was HCMV-seronegative, thus posing a high-risk serological combination (D-/R+) for post-transplant HCMV infection and associated complications.
The conditioning regimen consisted of fludarabine, busulfan, thiotepa, and anti-thymocyte globulin (ATG). The infused donor's bone marrow contained 3.79 × 108 total nuclear cells/kg recipient weight, and the vitality of the cells contained in the graft was 96.9%. Tacrolimus and mycophenolate mofetil were used for graft-versus-host disease (GVHD) prophylaxis. Anti-HCMV prophylaxis commenced at day +1 with commercial pooled anti-HCMV Ig (Megalotect, Biotest Pharma GmbH, Germany) at a dose of 100 U/kg twice a week.
The initial post-transplant period was uneventful. The first occurrence of HCMV viremia (600 viral genome copies/mL) was detected at day +14, and antiviral therapy with foscarnet was started immediately. At day +17, neutrophil engraftment was achieved, but not the platelet and the erythrocyte counterparts. At day +20, the patient presented fever without clinical symptoms and without an increase of C-reactive protein. Afterwards, a full blood count revealed a progressive drop in leukocytes and platelets, whereas HCMV viremia continued to increase despite antiviral therapy. The patient also developed a severe, HCMV-related hemorrhagic cystitis, which required frequent platelet transfusions, initially once daily for 5 days, and thereafter twice daily for 7 days.
As the infection appeared unresponsive to foscarnet, ganciclovir treatment was started at day +25. This switch in antiviral therapy followed an increase in viral load, reaching a maximum titer of 5100 viral genome copies/mL at day +30. Simultaneously, the clinical condition of the patient continued to deteriorate, and the leukocyte count decreased drastically. Therefore, the diagnosis of secondary graft failure due to HCMV infection was established. Because of the failure of anti-HCMV treatment, ganciclovir was withdrawn, and foscarnet therapy was resumed. The possible explanations for treatment failure include the presence of antiviral resistance mutations or an ineffective activity of the Ig administered. As the first option seemed improbable because resistance mutations usually arise later than 6 weeks after HSCT, we opted to address the latter option. Therefore, we considered using antibodies (Abs) that were more likely to inhibit the HCMV strains causing the infection in the patient. Although the diversity of circulating HCMV strains is known to be extensive (Puchhammer-Stöckl and Görzer, 2011), we hypothesized that the patient and her relatives may have been exposed to similar strains on a familial basis. As a result, we performed serological tests on the patient's mother, father and brother. We selected the patient's mother, who presented the highest anti-HCMV IgG titer, as the best donor of strain-specific HCMV Ab-rich plasma and virus-specific T-lymphocytes in this situation. At day +32, the patient's mother underwent leukapheresis. The lymphocytes were sent immediately to a cell factory to produce specific anti-HCMV T-lymphocytes. On the same day, the plasma enriched in HCMV strain-specific Abs was infused into the patient. At day +34, the viral load decreased for the first time, to 1100 viral genome copies/mL, and the patient began new conditioning for a haploidentical rescue transplant from her mother. In light of the concurrent bone marrow aplasia, the conditioning regimen was conducted solely with ATG. At day +38, the girl received a second plasma transfusion from her mother. On the day of the second transplant (day +40), complete viral clearance was documented, and the girl received 11.3 × 106 CD34 cells/kg of mother's graft. GVHD prophylaxis was performed with post-transplant cyclophosphamide, tacrolimus, and mycophenolate mofetil. The engraftment was rapid, with a neutrophil count of >500/μL achieved at day +15, and the final platelets and packed red blood cells were transfused at day +20. Since HCMV viremia remained negative during the entire post-transplant period, we discarded the use of the mother's virus-specific T-lymphocytes. The patient was discharged at day +27 in perfect clinical condition, with continuation of antiviral valganciclovir prophylaxis. Immunosuppressive treatment and antiviral prophylaxis were discontinued six months later, and the patient remains in perfect health 2 years after the second transplant.
Therapeutic interventions, HCMV load, and white blood cell count during the entire episode are summarized in Fig. 1. In addition, the HCMV strains present in the patient were characterized by high-throughput sequencing (HTS) of an archived (−80 °C) plasma sample from day +19. DNA was extracted from 800 μl of plasma using a QIAamp MinElute virus spin kit (QIAGEN, Crawley, UK). An aliquot of 50 μl of DNA was used to generate an HCMV-enriched sequencing library as described previously (Suárez et al., 2019). The library was loaded onto a NextSeq DNA sequencer (Illumina, San Diego, CA, USA), generating 150 bp paired-ended reads (Table 1). HCMV strain enumeration was performed by genotyping 12 hypervariable HCMV genes from the reads as described previously (Suárez et al., 2019). The detection of four genotypes of the RL13 gene indicated the presence of at least four HCMV strains (a strain being defined as a particular constellation of genotypes of the hypervariable genes analysed), each including one of these RL13 genotypes (Table 2). The data was also inspected for the presence of resistance mutations in the antiviral target genes (UL54 and UL97) using LoFreq (Wilm et al., 2012). Briefly, the reads were mapped to the genome of HCMV reference strain Merlin (GenBank accession no. AY446894.2), and non-synonymous variants known to confer resistance were identified. No resistance mutations were detected.Fig. 1 Information on viremia, white blood cell (WBC) count, total serum IgG and antiviral therapy. Data on HCMV viremia is expressed in viral genome copies/mL (right Y-axis). Application and duration of antiviral treatments are represented by colored bars (yellow, anti-HCMV-specific Ig; red, foscarnet; blue, ganciclovir/valganciclovir). Chronological data (X-axis) are represented in days after the first transplant. The plasma sample analysed by high-throughput sequencing (HTS) is represented by a filled blue circle.
Fig. 1
Table 1 Overview of HTS data.
Table 1Sample ID TRI-SCT1
Sample type Plasma
Days after transplant +19
HCMV load (genome copies/mL) 2100
Data deposition (ENA project no.)a PRJEB38333
Sequencing library (genome copies) 1440
Trimmed reads (no.) 5,033,480
HCMV reads (no.)b 936,773
HCMV reads; %b 19
Coverage depth (reads/nt)b 885
a The sequence dataset was purged of human reads and deposited in the European Nucleotide Archive (ENA).
b Reads mapped to the HCMV reference strain Merlin genome; HTS, high-throughput sequencing; nt, nucleotide.
Table 2 Genotyping of hypervariable HCMV genes.
Table 2
3 Discussion
Various factors are implicated in the outcome of HCMV infection in transplant patients, the first of which relates to the HCMV serological status of donor and recipient. The serological status of the first donor may have played an important role in this case, as it has been reported that using an HCMV-seronegative unrelated HSCT donor (D-) for a seropositive recipient (R+) results in reduced overall survival after myeloablative conditioning. In contrast, an HCMV seropositive donor is associated with improved event-free survival and reduced non-relapse mortality (Styczynski, 2018). Thus, the absence of an HCMV-specific immunological memory in the donor's cells may have placed the HCMV-positive recipient at a higher risk of HCMV reactivation and associated complications.
In relation to viral factors, the clinical course of the patient may have been influenced by two main factors: the presence of multiple HCMV strains or the presence of mutations conferring antiviral resistance. Infection with multiple HCMV strains is common after transplantation (Görzer et al., 2010), and is associated with delayed HCMV clearance during antiviral therapy, even in the absence of antiviral resistance mutations. It is also associated with worse clinical outcomes, including an increase in graft rejection and faster progression of disease. In principle, the mechanisms behind this increased pathogenicity can be viral, for example involving recombination and complementation of different strains within the host, or immunological, since control of multiple strains may be more challenging for the immune system. Overall, in clinically symptomatic immunocompromised hosts, the presence of multiple HCMV strains is more threatening than the presence of a single strain (Manuel et al., 2009).
In the present study, the initial failure of antiviral therapy could not be explained by the development of resistance mutations in the HCMV strains present in the patient. This finding is not surprising, as detection of resistance mutations is unusual during the first 6 weeks of antiviral therapy, whereas the sample analysed in this case was collected only 13 days after initiation of foscarnet treatment (Lurain and Chou, 2010; Springer et al., 2005).
In regard to the initial failure of the immunotherapy with commercial pooled Ig, this may have been due to the lack of particular Abs with neutralizing capacity against the HCMV strains present in the patient. Assuming that the donors providing the Ig had been exposed to strains circulating in Europe, the Abs may have been more suited for neutralizing HCMV strains commonly circulating in that region. However, it is possible that the strains present in the patient, who originated from Bangladesh, may have differed immunologically from strains in other regions of the world. The extent to which this is true is unknown, as current knowledge of HCMV genome variability is derived almost entirely from strains circulating in a few European countries. Therefore, it would be useful to assess the neutralizing capacity of commercial pooled Ig in order to avoid potential therapeutic failure, especially in patients from non-European countries. The clinical problem in this case was addressed successfully by using the Ab-rich plasma of the patient's mother in a kind of personalized immunotherapy, on the premise that both mother and daughter may have been exposed to the same pool of viral strains. In this HCMV-positive recipient with a severe viral reactivation, Ig obtained from a household donor afforded an efficacious treatment that led to complete viral clearance.
To the best of our knowledge, this is the first report of the successful use of HCMV strain-specific Ig therapy in an HSCT recipient. The efficacy of this approach as a means of prevention has been demonstrated previously only in mouse models involving murine cytomegalovirus (MCMV) reactivation after bone marrow transplantation (Martins et al., 2019). In this setting, mice received serum obtained either from latently infected (seropositive) donors or from seronegative donors. The transfer of immune serum protected mice from viral reactivation without affecting the development of GVHD. Remarkably, complete viral protection was obtained with a small volume (5 μL) of strain-specific immune serum. On the other hand, when specific donor antibodies were administered to mice infected with antigenically mismatched MCMV strains, they were not efficient in preventing viral reactivation.
Our case highlights the importance of monitoring HSCT recipients comprehensively, longitudinally and with high sensitivity for viral factors, including the presence of multiple strains and resistance mutations. Importantly, it also provides an affordable targeted immunological therapy in patients from countries with constrained access to alternative therapies.
4 Conclusion
HSCT is a complex and specific setting in which a recipient may rapidly develop viral reactivation and consequent life-threatening complications. Ab-rich plasma may represent a fast and feasible therapeutic option to overcome viral activity and allow successful engraftment of the donor's bone marrow.
Funding
No specific funding was received.
Consent for publication
The authors have obtained consent from the parents of the patient to publish individual patient data.
Declaration of competing interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgments
The sequencing work was supported by funding from the Medical Research Council (grant number MC_UU_12014/3 to AJD). | Recovered | ReactionOutcome | CC BY | 33577809 | 19,009,259 | 2021-04 |
What was the outcome of reaction 'Pyrexia'? | Post-HSCT graft failure due to refractory human cytomegalovirus successfully treated with haploidentical donor-derived immunoglobulins and stem cell graft infusion: A case report.
Human cytomegalovirus (HCMV) remains an important cause of transplant-related morbidity and mortality. The incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations as a result of a lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen.
We describe the case of an 8-year-old βE-thalassemic girl from Bangladesh who was seropositive for human cytomegalovirus (HCMV) and underwent hematopoietic stem cell transplantation from a HLA-matched, unrelated, HCMV-seronegative donor. Despite administering antiviral prophylaxis with commercial pooled anti-HCMV immunoglobulin (Ig) from day +1, the post-transplant course was complicated by prompt viral reactivation, and foscarnet therapy was initiated. The virus was refractory to treatment, leading rapidly to complete bone marrow failure, and targeted immunotherapy was proposed as a second-line therapy. Hypothesizing that the patient and her relatives may have been exposed to similar HCMV strains, we selected the patient's mother, who presented a high HCMV antibody titer, as the donor of virus strain-specific anti-HCMV Ig and T-lymphocytes. Complete viral clearance was achieved after two transfusions of the mother's plasma. Subsequently, the patient underwent a haploidentical rescue transplant, promptly reaching full hematological recovery.
These findings suggest that treatment with virus strain-specific Ig may offer a new therapeutic option for critically ill patients.
pmc1 Introduction
Human cytomegalovirus (HCMV) is the most significant cause of opportunistic viral infection in allogeneic hematopoietic stem cell transplantation (HSCT) and remains an important cause of transplant-related morbidity and mortality (Wagner-Drouet et al., 2019). Several studies have shown that the incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations (Ganepola et al., 2007; Zhou et al., 2009; Ozdemir et al., 2007). This is likely a consequence of delayed HCMV-specific immune reconstitution due to the lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen (Styczynski, 2018; Ljungman et al., 2014). HCMV infection can lead to pneumonia, gastroenteritis, retinitis, hepatitis, and encephalitis among HSCT recipients (Griffiths, 2012). In addition, HCMV infection of the bone marrow can result in impaired graft acceptance and, in the most extreme cases, complete graft failure (Steffens et al., 1998). Treatment of HCMV disease essentially consists of administering antiviral drugs, namely ganciclovir and foscarnet, and intravenous immunoglobulin. Although chemoprophylaxis is not generally recommended in HSCT, immunoglobulin, whether pooled or HCMV-specific, have shown some efficacy in preventing recurrent HCMV infections (Boeckh and Ljungman, 2009).
We describe a case of a pediatric HSCT recipient who had complications of HCMV recurrence and was treated successfully with targeted immunological therapy, using haploidentical donor-derived immunoglobulin (Ig).
2 Case presentation
An 8-year-old girl was diagnosed at the age of two in her country of origin (Bangladesh) as βE-thalassemic. She was treated with monthly blood transfusions and weekly iron chelation with deferoxamine, plus supplementation with folic acid. A suboptimal regimen of chelation therapy led to severe systemic siderosis, and her family travelled to Italy to undergo HSCT. Since a human leukocyte antigen (HLA)-matched related donor was unavailable at that time, an HLA-matched (10/10) unrelated donor was identified. The patient was HCMV-seropositive, whereas the donor was HCMV-seronegative, thus posing a high-risk serological combination (D-/R+) for post-transplant HCMV infection and associated complications.
The conditioning regimen consisted of fludarabine, busulfan, thiotepa, and anti-thymocyte globulin (ATG). The infused donor's bone marrow contained 3.79 × 108 total nuclear cells/kg recipient weight, and the vitality of the cells contained in the graft was 96.9%. Tacrolimus and mycophenolate mofetil were used for graft-versus-host disease (GVHD) prophylaxis. Anti-HCMV prophylaxis commenced at day +1 with commercial pooled anti-HCMV Ig (Megalotect, Biotest Pharma GmbH, Germany) at a dose of 100 U/kg twice a week.
The initial post-transplant period was uneventful. The first occurrence of HCMV viremia (600 viral genome copies/mL) was detected at day +14, and antiviral therapy with foscarnet was started immediately. At day +17, neutrophil engraftment was achieved, but not the platelet and the erythrocyte counterparts. At day +20, the patient presented fever without clinical symptoms and without an increase of C-reactive protein. Afterwards, a full blood count revealed a progressive drop in leukocytes and platelets, whereas HCMV viremia continued to increase despite antiviral therapy. The patient also developed a severe, HCMV-related hemorrhagic cystitis, which required frequent platelet transfusions, initially once daily for 5 days, and thereafter twice daily for 7 days.
As the infection appeared unresponsive to foscarnet, ganciclovir treatment was started at day +25. This switch in antiviral therapy followed an increase in viral load, reaching a maximum titer of 5100 viral genome copies/mL at day +30. Simultaneously, the clinical condition of the patient continued to deteriorate, and the leukocyte count decreased drastically. Therefore, the diagnosis of secondary graft failure due to HCMV infection was established. Because of the failure of anti-HCMV treatment, ganciclovir was withdrawn, and foscarnet therapy was resumed. The possible explanations for treatment failure include the presence of antiviral resistance mutations or an ineffective activity of the Ig administered. As the first option seemed improbable because resistance mutations usually arise later than 6 weeks after HSCT, we opted to address the latter option. Therefore, we considered using antibodies (Abs) that were more likely to inhibit the HCMV strains causing the infection in the patient. Although the diversity of circulating HCMV strains is known to be extensive (Puchhammer-Stöckl and Görzer, 2011), we hypothesized that the patient and her relatives may have been exposed to similar strains on a familial basis. As a result, we performed serological tests on the patient's mother, father and brother. We selected the patient's mother, who presented the highest anti-HCMV IgG titer, as the best donor of strain-specific HCMV Ab-rich plasma and virus-specific T-lymphocytes in this situation. At day +32, the patient's mother underwent leukapheresis. The lymphocytes were sent immediately to a cell factory to produce specific anti-HCMV T-lymphocytes. On the same day, the plasma enriched in HCMV strain-specific Abs was infused into the patient. At day +34, the viral load decreased for the first time, to 1100 viral genome copies/mL, and the patient began new conditioning for a haploidentical rescue transplant from her mother. In light of the concurrent bone marrow aplasia, the conditioning regimen was conducted solely with ATG. At day +38, the girl received a second plasma transfusion from her mother. On the day of the second transplant (day +40), complete viral clearance was documented, and the girl received 11.3 × 106 CD34 cells/kg of mother's graft. GVHD prophylaxis was performed with post-transplant cyclophosphamide, tacrolimus, and mycophenolate mofetil. The engraftment was rapid, with a neutrophil count of >500/μL achieved at day +15, and the final platelets and packed red blood cells were transfused at day +20. Since HCMV viremia remained negative during the entire post-transplant period, we discarded the use of the mother's virus-specific T-lymphocytes. The patient was discharged at day +27 in perfect clinical condition, with continuation of antiviral valganciclovir prophylaxis. Immunosuppressive treatment and antiviral prophylaxis were discontinued six months later, and the patient remains in perfect health 2 years after the second transplant.
Therapeutic interventions, HCMV load, and white blood cell count during the entire episode are summarized in Fig. 1. In addition, the HCMV strains present in the patient were characterized by high-throughput sequencing (HTS) of an archived (−80 °C) plasma sample from day +19. DNA was extracted from 800 μl of plasma using a QIAamp MinElute virus spin kit (QIAGEN, Crawley, UK). An aliquot of 50 μl of DNA was used to generate an HCMV-enriched sequencing library as described previously (Suárez et al., 2019). The library was loaded onto a NextSeq DNA sequencer (Illumina, San Diego, CA, USA), generating 150 bp paired-ended reads (Table 1). HCMV strain enumeration was performed by genotyping 12 hypervariable HCMV genes from the reads as described previously (Suárez et al., 2019). The detection of four genotypes of the RL13 gene indicated the presence of at least four HCMV strains (a strain being defined as a particular constellation of genotypes of the hypervariable genes analysed), each including one of these RL13 genotypes (Table 2). The data was also inspected for the presence of resistance mutations in the antiviral target genes (UL54 and UL97) using LoFreq (Wilm et al., 2012). Briefly, the reads were mapped to the genome of HCMV reference strain Merlin (GenBank accession no. AY446894.2), and non-synonymous variants known to confer resistance were identified. No resistance mutations were detected.Fig. 1 Information on viremia, white blood cell (WBC) count, total serum IgG and antiviral therapy. Data on HCMV viremia is expressed in viral genome copies/mL (right Y-axis). Application and duration of antiviral treatments are represented by colored bars (yellow, anti-HCMV-specific Ig; red, foscarnet; blue, ganciclovir/valganciclovir). Chronological data (X-axis) are represented in days after the first transplant. The plasma sample analysed by high-throughput sequencing (HTS) is represented by a filled blue circle.
Fig. 1
Table 1 Overview of HTS data.
Table 1Sample ID TRI-SCT1
Sample type Plasma
Days after transplant +19
HCMV load (genome copies/mL) 2100
Data deposition (ENA project no.)a PRJEB38333
Sequencing library (genome copies) 1440
Trimmed reads (no.) 5,033,480
HCMV reads (no.)b 936,773
HCMV reads; %b 19
Coverage depth (reads/nt)b 885
a The sequence dataset was purged of human reads and deposited in the European Nucleotide Archive (ENA).
b Reads mapped to the HCMV reference strain Merlin genome; HTS, high-throughput sequencing; nt, nucleotide.
Table 2 Genotyping of hypervariable HCMV genes.
Table 2
3 Discussion
Various factors are implicated in the outcome of HCMV infection in transplant patients, the first of which relates to the HCMV serological status of donor and recipient. The serological status of the first donor may have played an important role in this case, as it has been reported that using an HCMV-seronegative unrelated HSCT donor (D-) for a seropositive recipient (R+) results in reduced overall survival after myeloablative conditioning. In contrast, an HCMV seropositive donor is associated with improved event-free survival and reduced non-relapse mortality (Styczynski, 2018). Thus, the absence of an HCMV-specific immunological memory in the donor's cells may have placed the HCMV-positive recipient at a higher risk of HCMV reactivation and associated complications.
In relation to viral factors, the clinical course of the patient may have been influenced by two main factors: the presence of multiple HCMV strains or the presence of mutations conferring antiviral resistance. Infection with multiple HCMV strains is common after transplantation (Görzer et al., 2010), and is associated with delayed HCMV clearance during antiviral therapy, even in the absence of antiviral resistance mutations. It is also associated with worse clinical outcomes, including an increase in graft rejection and faster progression of disease. In principle, the mechanisms behind this increased pathogenicity can be viral, for example involving recombination and complementation of different strains within the host, or immunological, since control of multiple strains may be more challenging for the immune system. Overall, in clinically symptomatic immunocompromised hosts, the presence of multiple HCMV strains is more threatening than the presence of a single strain (Manuel et al., 2009).
In the present study, the initial failure of antiviral therapy could not be explained by the development of resistance mutations in the HCMV strains present in the patient. This finding is not surprising, as detection of resistance mutations is unusual during the first 6 weeks of antiviral therapy, whereas the sample analysed in this case was collected only 13 days after initiation of foscarnet treatment (Lurain and Chou, 2010; Springer et al., 2005).
In regard to the initial failure of the immunotherapy with commercial pooled Ig, this may have been due to the lack of particular Abs with neutralizing capacity against the HCMV strains present in the patient. Assuming that the donors providing the Ig had been exposed to strains circulating in Europe, the Abs may have been more suited for neutralizing HCMV strains commonly circulating in that region. However, it is possible that the strains present in the patient, who originated from Bangladesh, may have differed immunologically from strains in other regions of the world. The extent to which this is true is unknown, as current knowledge of HCMV genome variability is derived almost entirely from strains circulating in a few European countries. Therefore, it would be useful to assess the neutralizing capacity of commercial pooled Ig in order to avoid potential therapeutic failure, especially in patients from non-European countries. The clinical problem in this case was addressed successfully by using the Ab-rich plasma of the patient's mother in a kind of personalized immunotherapy, on the premise that both mother and daughter may have been exposed to the same pool of viral strains. In this HCMV-positive recipient with a severe viral reactivation, Ig obtained from a household donor afforded an efficacious treatment that led to complete viral clearance.
To the best of our knowledge, this is the first report of the successful use of HCMV strain-specific Ig therapy in an HSCT recipient. The efficacy of this approach as a means of prevention has been demonstrated previously only in mouse models involving murine cytomegalovirus (MCMV) reactivation after bone marrow transplantation (Martins et al., 2019). In this setting, mice received serum obtained either from latently infected (seropositive) donors or from seronegative donors. The transfer of immune serum protected mice from viral reactivation without affecting the development of GVHD. Remarkably, complete viral protection was obtained with a small volume (5 μL) of strain-specific immune serum. On the other hand, when specific donor antibodies were administered to mice infected with antigenically mismatched MCMV strains, they were not efficient in preventing viral reactivation.
Our case highlights the importance of monitoring HSCT recipients comprehensively, longitudinally and with high sensitivity for viral factors, including the presence of multiple strains and resistance mutations. Importantly, it also provides an affordable targeted immunological therapy in patients from countries with constrained access to alternative therapies.
4 Conclusion
HSCT is a complex and specific setting in which a recipient may rapidly develop viral reactivation and consequent life-threatening complications. Ab-rich plasma may represent a fast and feasible therapeutic option to overcome viral activity and allow successful engraftment of the donor's bone marrow.
Funding
No specific funding was received.
Consent for publication
The authors have obtained consent from the parents of the patient to publish individual patient data.
Declaration of competing interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgments
The sequencing work was supported by funding from the Medical Research Council (grant number MC_UU_12014/3 to AJD). | Recovered | ReactionOutcome | CC BY | 33577809 | 19,090,951 | 2021-04 |
What was the outcome of reaction 'Transplant failure'? | Post-HSCT graft failure due to refractory human cytomegalovirus successfully treated with haploidentical donor-derived immunoglobulins and stem cell graft infusion: A case report.
Human cytomegalovirus (HCMV) remains an important cause of transplant-related morbidity and mortality. The incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations as a result of a lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen.
We describe the case of an 8-year-old βE-thalassemic girl from Bangladesh who was seropositive for human cytomegalovirus (HCMV) and underwent hematopoietic stem cell transplantation from a HLA-matched, unrelated, HCMV-seronegative donor. Despite administering antiviral prophylaxis with commercial pooled anti-HCMV immunoglobulin (Ig) from day +1, the post-transplant course was complicated by prompt viral reactivation, and foscarnet therapy was initiated. The virus was refractory to treatment, leading rapidly to complete bone marrow failure, and targeted immunotherapy was proposed as a second-line therapy. Hypothesizing that the patient and her relatives may have been exposed to similar HCMV strains, we selected the patient's mother, who presented a high HCMV antibody titer, as the donor of virus strain-specific anti-HCMV Ig and T-lymphocytes. Complete viral clearance was achieved after two transfusions of the mother's plasma. Subsequently, the patient underwent a haploidentical rescue transplant, promptly reaching full hematological recovery.
These findings suggest that treatment with virus strain-specific Ig may offer a new therapeutic option for critically ill patients.
pmc1 Introduction
Human cytomegalovirus (HCMV) is the most significant cause of opportunistic viral infection in allogeneic hematopoietic stem cell transplantation (HSCT) and remains an important cause of transplant-related morbidity and mortality (Wagner-Drouet et al., 2019). Several studies have shown that the incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations (Ganepola et al., 2007; Zhou et al., 2009; Ozdemir et al., 2007). This is likely a consequence of delayed HCMV-specific immune reconstitution due to the lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen (Styczynski, 2018; Ljungman et al., 2014). HCMV infection can lead to pneumonia, gastroenteritis, retinitis, hepatitis, and encephalitis among HSCT recipients (Griffiths, 2012). In addition, HCMV infection of the bone marrow can result in impaired graft acceptance and, in the most extreme cases, complete graft failure (Steffens et al., 1998). Treatment of HCMV disease essentially consists of administering antiviral drugs, namely ganciclovir and foscarnet, and intravenous immunoglobulin. Although chemoprophylaxis is not generally recommended in HSCT, immunoglobulin, whether pooled or HCMV-specific, have shown some efficacy in preventing recurrent HCMV infections (Boeckh and Ljungman, 2009).
We describe a case of a pediatric HSCT recipient who had complications of HCMV recurrence and was treated successfully with targeted immunological therapy, using haploidentical donor-derived immunoglobulin (Ig).
2 Case presentation
An 8-year-old girl was diagnosed at the age of two in her country of origin (Bangladesh) as βE-thalassemic. She was treated with monthly blood transfusions and weekly iron chelation with deferoxamine, plus supplementation with folic acid. A suboptimal regimen of chelation therapy led to severe systemic siderosis, and her family travelled to Italy to undergo HSCT. Since a human leukocyte antigen (HLA)-matched related donor was unavailable at that time, an HLA-matched (10/10) unrelated donor was identified. The patient was HCMV-seropositive, whereas the donor was HCMV-seronegative, thus posing a high-risk serological combination (D-/R+) for post-transplant HCMV infection and associated complications.
The conditioning regimen consisted of fludarabine, busulfan, thiotepa, and anti-thymocyte globulin (ATG). The infused donor's bone marrow contained 3.79 × 108 total nuclear cells/kg recipient weight, and the vitality of the cells contained in the graft was 96.9%. Tacrolimus and mycophenolate mofetil were used for graft-versus-host disease (GVHD) prophylaxis. Anti-HCMV prophylaxis commenced at day +1 with commercial pooled anti-HCMV Ig (Megalotect, Biotest Pharma GmbH, Germany) at a dose of 100 U/kg twice a week.
The initial post-transplant period was uneventful. The first occurrence of HCMV viremia (600 viral genome copies/mL) was detected at day +14, and antiviral therapy with foscarnet was started immediately. At day +17, neutrophil engraftment was achieved, but not the platelet and the erythrocyte counterparts. At day +20, the patient presented fever without clinical symptoms and without an increase of C-reactive protein. Afterwards, a full blood count revealed a progressive drop in leukocytes and platelets, whereas HCMV viremia continued to increase despite antiviral therapy. The patient also developed a severe, HCMV-related hemorrhagic cystitis, which required frequent platelet transfusions, initially once daily for 5 days, and thereafter twice daily for 7 days.
As the infection appeared unresponsive to foscarnet, ganciclovir treatment was started at day +25. This switch in antiviral therapy followed an increase in viral load, reaching a maximum titer of 5100 viral genome copies/mL at day +30. Simultaneously, the clinical condition of the patient continued to deteriorate, and the leukocyte count decreased drastically. Therefore, the diagnosis of secondary graft failure due to HCMV infection was established. Because of the failure of anti-HCMV treatment, ganciclovir was withdrawn, and foscarnet therapy was resumed. The possible explanations for treatment failure include the presence of antiviral resistance mutations or an ineffective activity of the Ig administered. As the first option seemed improbable because resistance mutations usually arise later than 6 weeks after HSCT, we opted to address the latter option. Therefore, we considered using antibodies (Abs) that were more likely to inhibit the HCMV strains causing the infection in the patient. Although the diversity of circulating HCMV strains is known to be extensive (Puchhammer-Stöckl and Görzer, 2011), we hypothesized that the patient and her relatives may have been exposed to similar strains on a familial basis. As a result, we performed serological tests on the patient's mother, father and brother. We selected the patient's mother, who presented the highest anti-HCMV IgG titer, as the best donor of strain-specific HCMV Ab-rich plasma and virus-specific T-lymphocytes in this situation. At day +32, the patient's mother underwent leukapheresis. The lymphocytes were sent immediately to a cell factory to produce specific anti-HCMV T-lymphocytes. On the same day, the plasma enriched in HCMV strain-specific Abs was infused into the patient. At day +34, the viral load decreased for the first time, to 1100 viral genome copies/mL, and the patient began new conditioning for a haploidentical rescue transplant from her mother. In light of the concurrent bone marrow aplasia, the conditioning regimen was conducted solely with ATG. At day +38, the girl received a second plasma transfusion from her mother. On the day of the second transplant (day +40), complete viral clearance was documented, and the girl received 11.3 × 106 CD34 cells/kg of mother's graft. GVHD prophylaxis was performed with post-transplant cyclophosphamide, tacrolimus, and mycophenolate mofetil. The engraftment was rapid, with a neutrophil count of >500/μL achieved at day +15, and the final platelets and packed red blood cells were transfused at day +20. Since HCMV viremia remained negative during the entire post-transplant period, we discarded the use of the mother's virus-specific T-lymphocytes. The patient was discharged at day +27 in perfect clinical condition, with continuation of antiviral valganciclovir prophylaxis. Immunosuppressive treatment and antiviral prophylaxis were discontinued six months later, and the patient remains in perfect health 2 years after the second transplant.
Therapeutic interventions, HCMV load, and white blood cell count during the entire episode are summarized in Fig. 1. In addition, the HCMV strains present in the patient were characterized by high-throughput sequencing (HTS) of an archived (−80 °C) plasma sample from day +19. DNA was extracted from 800 μl of plasma using a QIAamp MinElute virus spin kit (QIAGEN, Crawley, UK). An aliquot of 50 μl of DNA was used to generate an HCMV-enriched sequencing library as described previously (Suárez et al., 2019). The library was loaded onto a NextSeq DNA sequencer (Illumina, San Diego, CA, USA), generating 150 bp paired-ended reads (Table 1). HCMV strain enumeration was performed by genotyping 12 hypervariable HCMV genes from the reads as described previously (Suárez et al., 2019). The detection of four genotypes of the RL13 gene indicated the presence of at least four HCMV strains (a strain being defined as a particular constellation of genotypes of the hypervariable genes analysed), each including one of these RL13 genotypes (Table 2). The data was also inspected for the presence of resistance mutations in the antiviral target genes (UL54 and UL97) using LoFreq (Wilm et al., 2012). Briefly, the reads were mapped to the genome of HCMV reference strain Merlin (GenBank accession no. AY446894.2), and non-synonymous variants known to confer resistance were identified. No resistance mutations were detected.Fig. 1 Information on viremia, white blood cell (WBC) count, total serum IgG and antiviral therapy. Data on HCMV viremia is expressed in viral genome copies/mL (right Y-axis). Application and duration of antiviral treatments are represented by colored bars (yellow, anti-HCMV-specific Ig; red, foscarnet; blue, ganciclovir/valganciclovir). Chronological data (X-axis) are represented in days after the first transplant. The plasma sample analysed by high-throughput sequencing (HTS) is represented by a filled blue circle.
Fig. 1
Table 1 Overview of HTS data.
Table 1Sample ID TRI-SCT1
Sample type Plasma
Days after transplant +19
HCMV load (genome copies/mL) 2100
Data deposition (ENA project no.)a PRJEB38333
Sequencing library (genome copies) 1440
Trimmed reads (no.) 5,033,480
HCMV reads (no.)b 936,773
HCMV reads; %b 19
Coverage depth (reads/nt)b 885
a The sequence dataset was purged of human reads and deposited in the European Nucleotide Archive (ENA).
b Reads mapped to the HCMV reference strain Merlin genome; HTS, high-throughput sequencing; nt, nucleotide.
Table 2 Genotyping of hypervariable HCMV genes.
Table 2
3 Discussion
Various factors are implicated in the outcome of HCMV infection in transplant patients, the first of which relates to the HCMV serological status of donor and recipient. The serological status of the first donor may have played an important role in this case, as it has been reported that using an HCMV-seronegative unrelated HSCT donor (D-) for a seropositive recipient (R+) results in reduced overall survival after myeloablative conditioning. In contrast, an HCMV seropositive donor is associated with improved event-free survival and reduced non-relapse mortality (Styczynski, 2018). Thus, the absence of an HCMV-specific immunological memory in the donor's cells may have placed the HCMV-positive recipient at a higher risk of HCMV reactivation and associated complications.
In relation to viral factors, the clinical course of the patient may have been influenced by two main factors: the presence of multiple HCMV strains or the presence of mutations conferring antiviral resistance. Infection with multiple HCMV strains is common after transplantation (Görzer et al., 2010), and is associated with delayed HCMV clearance during antiviral therapy, even in the absence of antiviral resistance mutations. It is also associated with worse clinical outcomes, including an increase in graft rejection and faster progression of disease. In principle, the mechanisms behind this increased pathogenicity can be viral, for example involving recombination and complementation of different strains within the host, or immunological, since control of multiple strains may be more challenging for the immune system. Overall, in clinically symptomatic immunocompromised hosts, the presence of multiple HCMV strains is more threatening than the presence of a single strain (Manuel et al., 2009).
In the present study, the initial failure of antiviral therapy could not be explained by the development of resistance mutations in the HCMV strains present in the patient. This finding is not surprising, as detection of resistance mutations is unusual during the first 6 weeks of antiviral therapy, whereas the sample analysed in this case was collected only 13 days after initiation of foscarnet treatment (Lurain and Chou, 2010; Springer et al., 2005).
In regard to the initial failure of the immunotherapy with commercial pooled Ig, this may have been due to the lack of particular Abs with neutralizing capacity against the HCMV strains present in the patient. Assuming that the donors providing the Ig had been exposed to strains circulating in Europe, the Abs may have been more suited for neutralizing HCMV strains commonly circulating in that region. However, it is possible that the strains present in the patient, who originated from Bangladesh, may have differed immunologically from strains in other regions of the world. The extent to which this is true is unknown, as current knowledge of HCMV genome variability is derived almost entirely from strains circulating in a few European countries. Therefore, it would be useful to assess the neutralizing capacity of commercial pooled Ig in order to avoid potential therapeutic failure, especially in patients from non-European countries. The clinical problem in this case was addressed successfully by using the Ab-rich plasma of the patient's mother in a kind of personalized immunotherapy, on the premise that both mother and daughter may have been exposed to the same pool of viral strains. In this HCMV-positive recipient with a severe viral reactivation, Ig obtained from a household donor afforded an efficacious treatment that led to complete viral clearance.
To the best of our knowledge, this is the first report of the successful use of HCMV strain-specific Ig therapy in an HSCT recipient. The efficacy of this approach as a means of prevention has been demonstrated previously only in mouse models involving murine cytomegalovirus (MCMV) reactivation after bone marrow transplantation (Martins et al., 2019). In this setting, mice received serum obtained either from latently infected (seropositive) donors or from seronegative donors. The transfer of immune serum protected mice from viral reactivation without affecting the development of GVHD. Remarkably, complete viral protection was obtained with a small volume (5 μL) of strain-specific immune serum. On the other hand, when specific donor antibodies were administered to mice infected with antigenically mismatched MCMV strains, they were not efficient in preventing viral reactivation.
Our case highlights the importance of monitoring HSCT recipients comprehensively, longitudinally and with high sensitivity for viral factors, including the presence of multiple strains and resistance mutations. Importantly, it also provides an affordable targeted immunological therapy in patients from countries with constrained access to alternative therapies.
4 Conclusion
HSCT is a complex and specific setting in which a recipient may rapidly develop viral reactivation and consequent life-threatening complications. Ab-rich plasma may represent a fast and feasible therapeutic option to overcome viral activity and allow successful engraftment of the donor's bone marrow.
Funding
No specific funding was received.
Consent for publication
The authors have obtained consent from the parents of the patient to publish individual patient data.
Declaration of competing interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgments
The sequencing work was supported by funding from the Medical Research Council (grant number MC_UU_12014/3 to AJD). | Recovered | ReactionOutcome | CC BY | 33577809 | 19,256,788 | 2021-04 |
What was the outcome of reaction 'Viral haemorrhagic cystitis'? | Post-HSCT graft failure due to refractory human cytomegalovirus successfully treated with haploidentical donor-derived immunoglobulins and stem cell graft infusion: A case report.
Human cytomegalovirus (HCMV) remains an important cause of transplant-related morbidity and mortality. The incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations as a result of a lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen.
We describe the case of an 8-year-old βE-thalassemic girl from Bangladesh who was seropositive for human cytomegalovirus (HCMV) and underwent hematopoietic stem cell transplantation from a HLA-matched, unrelated, HCMV-seronegative donor. Despite administering antiviral prophylaxis with commercial pooled anti-HCMV immunoglobulin (Ig) from day +1, the post-transplant course was complicated by prompt viral reactivation, and foscarnet therapy was initiated. The virus was refractory to treatment, leading rapidly to complete bone marrow failure, and targeted immunotherapy was proposed as a second-line therapy. Hypothesizing that the patient and her relatives may have been exposed to similar HCMV strains, we selected the patient's mother, who presented a high HCMV antibody titer, as the donor of virus strain-specific anti-HCMV Ig and T-lymphocytes. Complete viral clearance was achieved after two transfusions of the mother's plasma. Subsequently, the patient underwent a haploidentical rescue transplant, promptly reaching full hematological recovery.
These findings suggest that treatment with virus strain-specific Ig may offer a new therapeutic option for critically ill patients.
pmc1 Introduction
Human cytomegalovirus (HCMV) is the most significant cause of opportunistic viral infection in allogeneic hematopoietic stem cell transplantation (HSCT) and remains an important cause of transplant-related morbidity and mortality (Wagner-Drouet et al., 2019). Several studies have shown that the incidence of HCMV recurrence in the donor seronegative (D-)/recipient seropositive (R+) group is significantly higher than in other serostatus combinations (Ganepola et al., 2007; Zhou et al., 2009; Ozdemir et al., 2007). This is likely a consequence of delayed HCMV-specific immune reconstitution due to the lack of pre-existing HCMV-specific memory T-lymphocytes in the donor, coupled with the eradication of the recipient's cellular immunity due to the conditioning regimen (Styczynski, 2018; Ljungman et al., 2014). HCMV infection can lead to pneumonia, gastroenteritis, retinitis, hepatitis, and encephalitis among HSCT recipients (Griffiths, 2012). In addition, HCMV infection of the bone marrow can result in impaired graft acceptance and, in the most extreme cases, complete graft failure (Steffens et al., 1998). Treatment of HCMV disease essentially consists of administering antiviral drugs, namely ganciclovir and foscarnet, and intravenous immunoglobulin. Although chemoprophylaxis is not generally recommended in HSCT, immunoglobulin, whether pooled or HCMV-specific, have shown some efficacy in preventing recurrent HCMV infections (Boeckh and Ljungman, 2009).
We describe a case of a pediatric HSCT recipient who had complications of HCMV recurrence and was treated successfully with targeted immunological therapy, using haploidentical donor-derived immunoglobulin (Ig).
2 Case presentation
An 8-year-old girl was diagnosed at the age of two in her country of origin (Bangladesh) as βE-thalassemic. She was treated with monthly blood transfusions and weekly iron chelation with deferoxamine, plus supplementation with folic acid. A suboptimal regimen of chelation therapy led to severe systemic siderosis, and her family travelled to Italy to undergo HSCT. Since a human leukocyte antigen (HLA)-matched related donor was unavailable at that time, an HLA-matched (10/10) unrelated donor was identified. The patient was HCMV-seropositive, whereas the donor was HCMV-seronegative, thus posing a high-risk serological combination (D-/R+) for post-transplant HCMV infection and associated complications.
The conditioning regimen consisted of fludarabine, busulfan, thiotepa, and anti-thymocyte globulin (ATG). The infused donor's bone marrow contained 3.79 × 108 total nuclear cells/kg recipient weight, and the vitality of the cells contained in the graft was 96.9%. Tacrolimus and mycophenolate mofetil were used for graft-versus-host disease (GVHD) prophylaxis. Anti-HCMV prophylaxis commenced at day +1 with commercial pooled anti-HCMV Ig (Megalotect, Biotest Pharma GmbH, Germany) at a dose of 100 U/kg twice a week.
The initial post-transplant period was uneventful. The first occurrence of HCMV viremia (600 viral genome copies/mL) was detected at day +14, and antiviral therapy with foscarnet was started immediately. At day +17, neutrophil engraftment was achieved, but not the platelet and the erythrocyte counterparts. At day +20, the patient presented fever without clinical symptoms and without an increase of C-reactive protein. Afterwards, a full blood count revealed a progressive drop in leukocytes and platelets, whereas HCMV viremia continued to increase despite antiviral therapy. The patient also developed a severe, HCMV-related hemorrhagic cystitis, which required frequent platelet transfusions, initially once daily for 5 days, and thereafter twice daily for 7 days.
As the infection appeared unresponsive to foscarnet, ganciclovir treatment was started at day +25. This switch in antiviral therapy followed an increase in viral load, reaching a maximum titer of 5100 viral genome copies/mL at day +30. Simultaneously, the clinical condition of the patient continued to deteriorate, and the leukocyte count decreased drastically. Therefore, the diagnosis of secondary graft failure due to HCMV infection was established. Because of the failure of anti-HCMV treatment, ganciclovir was withdrawn, and foscarnet therapy was resumed. The possible explanations for treatment failure include the presence of antiviral resistance mutations or an ineffective activity of the Ig administered. As the first option seemed improbable because resistance mutations usually arise later than 6 weeks after HSCT, we opted to address the latter option. Therefore, we considered using antibodies (Abs) that were more likely to inhibit the HCMV strains causing the infection in the patient. Although the diversity of circulating HCMV strains is known to be extensive (Puchhammer-Stöckl and Görzer, 2011), we hypothesized that the patient and her relatives may have been exposed to similar strains on a familial basis. As a result, we performed serological tests on the patient's mother, father and brother. We selected the patient's mother, who presented the highest anti-HCMV IgG titer, as the best donor of strain-specific HCMV Ab-rich plasma and virus-specific T-lymphocytes in this situation. At day +32, the patient's mother underwent leukapheresis. The lymphocytes were sent immediately to a cell factory to produce specific anti-HCMV T-lymphocytes. On the same day, the plasma enriched in HCMV strain-specific Abs was infused into the patient. At day +34, the viral load decreased for the first time, to 1100 viral genome copies/mL, and the patient began new conditioning for a haploidentical rescue transplant from her mother. In light of the concurrent bone marrow aplasia, the conditioning regimen was conducted solely with ATG. At day +38, the girl received a second plasma transfusion from her mother. On the day of the second transplant (day +40), complete viral clearance was documented, and the girl received 11.3 × 106 CD34 cells/kg of mother's graft. GVHD prophylaxis was performed with post-transplant cyclophosphamide, tacrolimus, and mycophenolate mofetil. The engraftment was rapid, with a neutrophil count of >500/μL achieved at day +15, and the final platelets and packed red blood cells were transfused at day +20. Since HCMV viremia remained negative during the entire post-transplant period, we discarded the use of the mother's virus-specific T-lymphocytes. The patient was discharged at day +27 in perfect clinical condition, with continuation of antiviral valganciclovir prophylaxis. Immunosuppressive treatment and antiviral prophylaxis were discontinued six months later, and the patient remains in perfect health 2 years after the second transplant.
Therapeutic interventions, HCMV load, and white blood cell count during the entire episode are summarized in Fig. 1. In addition, the HCMV strains present in the patient were characterized by high-throughput sequencing (HTS) of an archived (−80 °C) plasma sample from day +19. DNA was extracted from 800 μl of plasma using a QIAamp MinElute virus spin kit (QIAGEN, Crawley, UK). An aliquot of 50 μl of DNA was used to generate an HCMV-enriched sequencing library as described previously (Suárez et al., 2019). The library was loaded onto a NextSeq DNA sequencer (Illumina, San Diego, CA, USA), generating 150 bp paired-ended reads (Table 1). HCMV strain enumeration was performed by genotyping 12 hypervariable HCMV genes from the reads as described previously (Suárez et al., 2019). The detection of four genotypes of the RL13 gene indicated the presence of at least four HCMV strains (a strain being defined as a particular constellation of genotypes of the hypervariable genes analysed), each including one of these RL13 genotypes (Table 2). The data was also inspected for the presence of resistance mutations in the antiviral target genes (UL54 and UL97) using LoFreq (Wilm et al., 2012). Briefly, the reads were mapped to the genome of HCMV reference strain Merlin (GenBank accession no. AY446894.2), and non-synonymous variants known to confer resistance were identified. No resistance mutations were detected.Fig. 1 Information on viremia, white blood cell (WBC) count, total serum IgG and antiviral therapy. Data on HCMV viremia is expressed in viral genome copies/mL (right Y-axis). Application and duration of antiviral treatments are represented by colored bars (yellow, anti-HCMV-specific Ig; red, foscarnet; blue, ganciclovir/valganciclovir). Chronological data (X-axis) are represented in days after the first transplant. The plasma sample analysed by high-throughput sequencing (HTS) is represented by a filled blue circle.
Fig. 1
Table 1 Overview of HTS data.
Table 1Sample ID TRI-SCT1
Sample type Plasma
Days after transplant +19
HCMV load (genome copies/mL) 2100
Data deposition (ENA project no.)a PRJEB38333
Sequencing library (genome copies) 1440
Trimmed reads (no.) 5,033,480
HCMV reads (no.)b 936,773
HCMV reads; %b 19
Coverage depth (reads/nt)b 885
a The sequence dataset was purged of human reads and deposited in the European Nucleotide Archive (ENA).
b Reads mapped to the HCMV reference strain Merlin genome; HTS, high-throughput sequencing; nt, nucleotide.
Table 2 Genotyping of hypervariable HCMV genes.
Table 2
3 Discussion
Various factors are implicated in the outcome of HCMV infection in transplant patients, the first of which relates to the HCMV serological status of donor and recipient. The serological status of the first donor may have played an important role in this case, as it has been reported that using an HCMV-seronegative unrelated HSCT donor (D-) for a seropositive recipient (R+) results in reduced overall survival after myeloablative conditioning. In contrast, an HCMV seropositive donor is associated with improved event-free survival and reduced non-relapse mortality (Styczynski, 2018). Thus, the absence of an HCMV-specific immunological memory in the donor's cells may have placed the HCMV-positive recipient at a higher risk of HCMV reactivation and associated complications.
In relation to viral factors, the clinical course of the patient may have been influenced by two main factors: the presence of multiple HCMV strains or the presence of mutations conferring antiviral resistance. Infection with multiple HCMV strains is common after transplantation (Görzer et al., 2010), and is associated with delayed HCMV clearance during antiviral therapy, even in the absence of antiviral resistance mutations. It is also associated with worse clinical outcomes, including an increase in graft rejection and faster progression of disease. In principle, the mechanisms behind this increased pathogenicity can be viral, for example involving recombination and complementation of different strains within the host, or immunological, since control of multiple strains may be more challenging for the immune system. Overall, in clinically symptomatic immunocompromised hosts, the presence of multiple HCMV strains is more threatening than the presence of a single strain (Manuel et al., 2009).
In the present study, the initial failure of antiviral therapy could not be explained by the development of resistance mutations in the HCMV strains present in the patient. This finding is not surprising, as detection of resistance mutations is unusual during the first 6 weeks of antiviral therapy, whereas the sample analysed in this case was collected only 13 days after initiation of foscarnet treatment (Lurain and Chou, 2010; Springer et al., 2005).
In regard to the initial failure of the immunotherapy with commercial pooled Ig, this may have been due to the lack of particular Abs with neutralizing capacity against the HCMV strains present in the patient. Assuming that the donors providing the Ig had been exposed to strains circulating in Europe, the Abs may have been more suited for neutralizing HCMV strains commonly circulating in that region. However, it is possible that the strains present in the patient, who originated from Bangladesh, may have differed immunologically from strains in other regions of the world. The extent to which this is true is unknown, as current knowledge of HCMV genome variability is derived almost entirely from strains circulating in a few European countries. Therefore, it would be useful to assess the neutralizing capacity of commercial pooled Ig in order to avoid potential therapeutic failure, especially in patients from non-European countries. The clinical problem in this case was addressed successfully by using the Ab-rich plasma of the patient's mother in a kind of personalized immunotherapy, on the premise that both mother and daughter may have been exposed to the same pool of viral strains. In this HCMV-positive recipient with a severe viral reactivation, Ig obtained from a household donor afforded an efficacious treatment that led to complete viral clearance.
To the best of our knowledge, this is the first report of the successful use of HCMV strain-specific Ig therapy in an HSCT recipient. The efficacy of this approach as a means of prevention has been demonstrated previously only in mouse models involving murine cytomegalovirus (MCMV) reactivation after bone marrow transplantation (Martins et al., 2019). In this setting, mice received serum obtained either from latently infected (seropositive) donors or from seronegative donors. The transfer of immune serum protected mice from viral reactivation without affecting the development of GVHD. Remarkably, complete viral protection was obtained with a small volume (5 μL) of strain-specific immune serum. On the other hand, when specific donor antibodies were administered to mice infected with antigenically mismatched MCMV strains, they were not efficient in preventing viral reactivation.
Our case highlights the importance of monitoring HSCT recipients comprehensively, longitudinally and with high sensitivity for viral factors, including the presence of multiple strains and resistance mutations. Importantly, it also provides an affordable targeted immunological therapy in patients from countries with constrained access to alternative therapies.
4 Conclusion
HSCT is a complex and specific setting in which a recipient may rapidly develop viral reactivation and consequent life-threatening complications. Ab-rich plasma may represent a fast and feasible therapeutic option to overcome viral activity and allow successful engraftment of the donor's bone marrow.
Funding
No specific funding was received.
Consent for publication
The authors have obtained consent from the parents of the patient to publish individual patient data.
Declaration of competing interest
The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper.
Acknowledgments
The sequencing work was supported by funding from the Medical Research Council (grant number MC_UU_12014/3 to AJD). | Recovered | ReactionOutcome | CC BY | 33577809 | 19,010,959 | 2021-04 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Death'. | Efficacy of Denosumab Therapy Following Treatment with Bisphosphonates in Women with Osteoporosis: A Cohort Study.
Denosumab is a human monoclonal antibody that neutralizes RANKL, a cytokine able to interact with the RANK receptor on preosteoclasts and osteoclasts, decreasing their recruitment and differentiation, leading to a decreased bone resorption. The aim of this observational real-life study was to analyze adherence to denosumab therapy and assess its efficacy in increasing bone mineral density (BMD) and modulating biochemical skeletal markers following previous treatments with bisphosphonates in a group of post-menopausal women with osteoporosis. Women were recruited in the specialized center from March 2012 to September 2019. Biochemical markers were recorded at baseline and every six months prior to subsequent drug injection. Dual X-ray absorptiometry was requested at baseline and after 18/24 months. Comparing BMD at baseline and after denosumab therapy in naive patients and in those previously treated with bisphosphonates, a positive therapeutic effect was observed in both groups. The results of our real-life study demonstrate, as expected, that BMD values significantly increased upon denosumab treatment. Interestingly, denosumab showed an increased efficacy in patients previously treated with bisphosphonates. Moreover, biochemical markers data indicate that osteoporotic patients, without other concomitant unstable health conditions, could be evaluated once a year, decreasing the number of specialistic center access.
1. Introduction
During the last decades, in industrialized countries a significant increase of life expectancy with a gradual aging of the population has been observed. By 2060, America’s population of adults over the age of 65 is expected to double the current number [1]. In this scenario, the prolonged life expectancy, the inappropriate eating behaviors and lifestyle have contributed to the increased incidence of osteoporosis, a skeletal metabolic chronic disease characterized by compromised bone strength due to the reduction of both bone quantity and quality. The decreased bone strength leads the affected subjects to an increased risk of developing vertebral and non-vertebral fragility fractures, with increased morbidity, mortality, health service utilization, health care costs, and deterioration of quality of life [2,3,4,5,6,7,8].
All pharmacological treatments proposed in the last decades for osteoporotic patients aimed to reduce the risk of osteoporosis-related fractures by increasing bone mineral density and improving bone quality by modulating bone remodeling as demonstrated in controlled randomized clinical trial [9,10,11,12,13].
Denosumab, one of the latest pharmacological treatment developed as anti-osteoporotic treatment, is a fully human monoclonal antibody with high specificity to human RANKL, a receptor activator of NFκB ligand that is an essential factor in activating osteoclast differentiation and activity and, thus, activating bone resorption. This antiresorptive drug is administered by a subcutaneous injection every 6 months and it reversibly reduces osteoclast number and activity decreasing bone resorption. On the basis of this mechanism, denosumab has been demonstrated to significantly decrease new vertebral, nonvertebral, and hip fractures for up to 10 years of treatment in post-menopausal women affected by osteoporosis as demonstrated by the Fracture REduction Evaluation of Denosumab in Osteoporosis every 6 Months (FREEDOM) Extension trial [14,15,16].
Few studies, however, have evaluated adherence and efficacy of denosumab treatment in increasing BMD and potentially modulating biochemical skeletal markers in everyday life outpatients [17,18,19]. Thus, the aim of this observational real-life study was to analyze adherence to denosumab therapy and assess its efficacy in increasing bone mineral density (BMD) and modulating biochemical skeletal markers following previous treatments with bisphosphonates in a group of post-menopausal women with osteoporosis. In particular, it was evaluated and compared BMD at baseline and after denosumab therapy in either naive patients or patients previously treated with anti-resorptive drugs, since some previous studies suggested potential altered response to denosumab upon previous antiresorptive drugs [20,21,22].
2. Materials and Methods
2.1. Recruitment
Four hundred twenty-eight post-menopausal women (mean age = 71.7 ± 8.6 years) were recruited for this study. All subjects were affected by osteoporosis and referring to the specialized center for the management of osteoporosis in Policlinico Umberto I Hospital, Rome, from March 2012 to September 2019. Patients were evaluated every six months from the date of the initial prescription. The study respects the Declaration of Helsinki and was approved by the local Ethical Committees of the institution (prot 648/17) involved and met the guidelines of the responsible governmental agency.
2.2. Inclusion Criteria
Inclusion criteria were fixed on the basis of a previous work and AIFA’s advices for drug treatments: diagnosis of post-menopausal osteoporosis, possibility to receive a prescription during the study period according to the Italian reimbursement criteria for anti-osteoporotic pharmacological treatment [23,24].
Specifically, subjects with: (i) at least one prevalent hip or vertebral fracture (with contraindications to take oral pharmacological treatment), (ii) subjects aged ≥50 years with a femoral neck (FN) T-score ≤ −4 standard deviations (SD), (iii) subjects aged ≥50 years with a FN T-score ≤ −3 SD and at least one risk factor (family history of fragility vertebral or hip fractures, rheumatoid arthritis and other connective tissue diseases, previous wrist fragility fracture, early menopause, chronic corticosteroid therapy) were included.
The study was approved by the local University Institutional Review Board (prot n5061/18) and all participants gave their written informed consent before study participation.
2.3. Intervention
Before starting the new pharmacological treatment, the referring specialist or a resident fellow explained to the patients the mode of action and the method of taking the drug. Treatment consisted of a subcutaneous injection of denosumab (Prolia®, Amgen Inc. Thousand Oaks, CA 91320-1799, USA) 60 mg/every 6 months. Each patient was clinically evaluated before the first administration of denosumab (baseline) and then every 6 months, before the following injection, until the end of the observational study period. All patients continued taking all other concomitant therapies, including calcium and/or vitamin D supplementation. The specialist allowed all patients, or their relatives, to contact the clinical group via e-mail or telephone in case of doubts, questions or important communications. At the date of the initial prescription (baseline) the following data and information (y/n) were collected: age (years), age of menopause (years), weight (kg), height (m), body mass index (BMI), history of smoking; family history for osteoporosis; family history for fragility fractures; history for fragility vertebral and/or femoral and/or any other skeletal site fractures; previous hormonal replacement therapy (HRT), and/or chronic corticosteroid therapy; any previous and/or current supplementation with calcium and/or vitamin D and any previous antiresorptive therapy with bisphosphonates (BPs, including risedronate, alendronate, clodronate, zoledronic acid, ibandronate) or strontium ranelate or teriparatide. BMD [T-score at lumbar spine (LS), total proximal femur (F) and femoral neck (FN)] was evaluated by dual X-ray absorptiometry (DXA) measurement. In regards of the biochemical markers, serum parathyroid hormone (PTH), calcium (Ca), alkaline phosphatase (ALP), and 25(OH) vitamin D3 (VitD) were analyzed. Since this is a real-life study, blood tests were not centralized in our hospital center, but patients performed them in the morning between 07.00 a.m. and 09.00 a.m. after an overnight fast in other clinical centers which are required to use standardized methods (Table 1). As previously mentioned, some patients were under medications for other diseases, which however did not affect bone metabolism.
Weight (kg) and height (m) as anthropometric parameters were recorded to calculate BMI, according to the following formula: BMI = weight/height2 (kg/m2).
Osteoporosis was diagnosticated by DXA, based on the T-score for BMD less than or equal to −2.5 SD [25]. At baseline visit, lumbar, total proximal and femoral neck DXA were requested. Vertebrae affected by fractures, or other structural change or artifact, were excluded following the International Society for Clinical Densitometry—ISCD —official guidelines. Left hips affected by fractures, and underwent hip replacement surgery, were excluded and substituted by measurement of right hips. The following DXA was required after 18/24 months.
Biochemical markers were measured at baseline, approximately a week before drug administration, and at 6, 12, 18 and 24 months of denosumab treatment, in order to allow the patients to bring results to the visit.
For all subjects, bone density at lumbar spine, total proximal femur and femoral neck (LS T-score, F T-score and FN T-score) at baseline, were compared with values collected over time.
Of all participants who were prescribed denosumab, 43 dropped out (10.04%) before the end of the study. Reasons for drop-out are reported in the results section.
Since denosumab is also prescribed in post-menopausal women with hormone-responsive breast cancer under aromatase inhibitors (AI) adjuvant therapy, denosumab was prescribed to 53 women affected by breast cancer (BC group) [26]. In fact, as estrogens stimulate breast cancer cells, AI are aimed to inhibit the production of endogenous estrogens from aromatase in adipose tissue. On the other hand, low levels of estrogens can induce bone loss predisposing to osteoporosis and fractures [27,28,29]. BC group was excluded from the study for reasons that are reported in the results.
2.4. Statistical Analysis
Statistical analysis was performed on the following parameters: age, weight, height, BMI, age of menopause, BMD (LS T-score, F T-score, FN T-score), PTH, ALP, Ca and VitD. Differences between the means of above cited parameters were estimated using a two-sample t-test comparison between women with and without breast cancer. Pearson’s correlations among above cited parameters were estimated. The chi-square test was used to compare variables’ frequencies among groups (normal BMD, osteopenia, osteoporosis).
ANOVA_RM (repeated measures) were performed on BMD, PTH, ALP, Ca and VitD for different times. The time of the measurements were called T0 (baseline) and T1, T2, T3, T4, T5, T6, T7, T8 and T9 corresponding to 6, 12, 18, 24, 30, 36, 42, 48, 54 months, respectively. Statistical significance was defined as p ≤ 0.05. All statistical analysis was performed by SPSS version 24.0 software (SPSS Inc., Chicago, IL, USA).
3. Results
As previously described in the material and methods section, data were collected for each patient for a maximum of 8 years every six months from the beginning of the therapy, individually. Thus, we estimated and reported adherence of each patient, drop out and reason of it, densitometric analysis, biochemical markers, eventual missing data. To increase the power of the analysis, we considered the first 24 months of therapy for each patient in order to have a satisfactory number of patients to be considered.
3.1. Drop-Out
Of 428 enrolled subjects, 43 dropped out (10.04%) before the end of the study: in particular, the majority dropped within the first 12 months of therapy (87%), whereas the rest (13%) dropped out after 12 months. Thus, from our data it results that outpatients had an adherence to therapy of 89.6% up to the end of therapy. As depicted in Table 2, reasons of drop out were: death (6/43, 14%), refusal of therapy (13/43, 30.2%), unspecific disturbs (2/43, 4.6%), fear of collateral effects (4/43, 9.4%), fear of injective drugs (3/43, 6.9%), suggested by another specialist to change therapy (10/43, 23.2%), followed in another specialized center (5/43, 11.6%) (Table 2). Indeed, several studies have demonstrated that higher number of drop out occurs within the first 12 months of chronic pharmacological intervention [30,31,32].
3.2. Comparison between Groups of Women with or without Breast Cancer
Two-sample t-test comparison between the group of 53 women with breast cancer (BC) and the group of 332 women without breast cancer (NoBC) depicted significant differences in age (p < 0.01). Consequently, the two groups could not be studied as a single population and BC women in therapy con denosumab were not included in the statistical analysis of this study to address the aim of the study.
3.3. Partition between Naive Subjects and Subjects Treated with Previous Therapies
Of the 332 remaining patients (mean age = 72.8 ± 7.9 years), 237 women had at least one vertebral fragility fracture and 16 women had a previous femoral fracture.
From this group of 332 women, 126 subjects were considered naive (no previous anti-osteoporotic therapy), 122 were previously treated with BPs therapy, 43 either teriparatide or strontium ranelate and 41 had received two or more of the above-mentioned therapies, as shown in Table 3. In order to compare naive group (NG) with BPs previously treated group (BPG) data, the 84 subjects who received teriparatide or strontium ranelate prior to denosumab were not included in statistical analysis (ANOVA).
3.4. Pearson’s Correlations at Baseline
Pearson’s correlations among main parameters at baseline were estimated.
Interestingly, analysis showed that BMD femoral neck (FN T-score) was significantly negatively related with age (p < 0.01). Correlation values showed that menopause age was positively correlated with F T-score (p < 0.01) and FN T-score (p < 0.01), age was positively correlated with BMI (p < 0.01) and LS T-score (p < 0.05). VitD was inversely correlated with weight (p < 0.05) and Ca was negatively correlated only with PTH (p < 0.01). FN T-score was positively correlated with all parameters analyzed (p < 0.01) except Ca and VitD, while it was negatively correlated with PTH (p < 0.05).
3.5. Efficacy of Denosumab Therapy in Increasing BMD
We compared BMD values at lumbar spine, femoral and hip neck before (at baseline (T0) and after 24 (T4), 48 (T8) and 54 (T9) months of pharmacological treatment. In Figure 1, as expected, compared to baseline, BMD values significantly increased after 24 months of treatment. The chi-square test revealed that there was a significant change in BMD in patients in therapy with denosumab: evaluation showed that the number of subjects affected by a low BMD, compatible with an osteoporotic T-score was reduced after 48 months of denosumab therapy, whereas the percent of subjects with osteopenia was increased.
3.6. Previous Therapy
The interest was to evaluate whether previous therapies could have influenced the efficacy of denosumab therapy. Two different groups were considered: a group of subjects (n = 122) previously treated at least with one of the mentioned different bisphosphonates (BPG) and a naive group of subjects (n = 126) that have no taken any previous anti-osteoporotic medication (NG). BMD measurements (LS T-score, F T-score F and FN T-score) before (at baseline) and after denosumab therapy were compared by ANOVA_RM (Table 4).
A significant improvement (p < 0.01) was observed after denosumab therapy (Figure 2A–C). Differences between the two groups (previous therapy effect, see last line of Table 4) was not observed in Lumbar spine (Figure 2A) and Femoral (LS T-score, p = 0.91 and F T-score, p = 0.07) (Figure 2B) but was observed in Femoral Neck (FN T-score, p = 0.05) (Figure 2C).
3.7. Biochemical Markers
In order to study whether changes in biochemical marker values occurred during denosumab treatment in related variables, ANOVA (by ANOVA_RM) analysis were performed on PTH, ALP, Ca and VitD for different times.
It was observed that PTH levels significantly increased from T0 to T1 (N = 36, p < 0.001) but did not change substantially until 24 months. As expected by an anti-resorptive drug, ALP levels markedly decreased following treatment (N = 51, p < 0.001). In later time-points, biochemical markers data were available for a lower number of patients, which however demonstrated no further changes as compared to those that occurred within the first 24 months of therapy. Indeed, serum calcium levels remained unchanged upon treatment (ANOVA, N = 39, p > 0.05) while serum VitD levels significantly increased from T0 to T1 but did not change significantly after T3 (ANOVA, N = 79, p < 0.001) (data not showed).
4. Discussion
This Italian observational real-life study evaluated the effect of denosumab in a group of post-menopausal women. The results showed a good response in BMD after 2 years of denosumab treatment in all skeletal sites evaluated, as demonstrated by the decreased percent of BMD values compatible with osteoporosis and the increased percent of BMD values compatible with osteopenia. As expected, these data, regarding the improvements of BMD in a population of osteoporotic out-patients, confirm results already showed in randomized clinical trial [33,34,35].
Moreover, our findings, comparing BMD at baseline and over time in naive group (NG) with BPs previously treated group (BPG), showed a significant improvement of BMD in all skeletal sites evaluated (p < 0.01) after denosumab treatment in both groups of patients, suggesting that previous BP therapy does not lead to a decreased efficacy of denosumab. On the other hand, it is interesting to note that the increase of BMD at femoral neck was significantly higher (p = 0.05) in patients previously treated with BPs as previously showed by Sánchez et al., suggesting a therapy effect [17]. These data could be explained by the different mechanism of action of these anti-resorptive drugs: while the effect of denosumab on bone metabolism might stop upon discontinuation, BPs might maintain the antiresorptive effect even after discontinuation [36].
Pearson’s correlations at baseline among BMD regions confirm FN T-score as the most reliable measure, being negatively and positively correlated with age and menopause age, respectively. Indeed, our data further indicate that older patients have a lower FN T-score at baseline and that, as expected, an older age at menopause has a protective effect on the bone, likely due to circulating estrogens, leading to higher FN T-score. Further, considering other parameters, VitD was inversely correlated with body weight, as calculated by BMI, since in the present study body composition data were collected. Nevertheless, on the base of BMI of the patients, we can speculate that higher levels of fat mass could contribute to low VitD levels, since numerous studies, including the ones of our group, have shown that obese/overweight subjects have lower vitamin D levels, considering that the storage of this fat-soluble vitamin is mainly in the adipose tissue [37,38,39,40].
Interestingly, the measurement of bone biochemical markers after the first infusion of denosumab, showed a significant increase of serum PTH and VitD levels after 6 months of denosumab treatment (p < 0.001), confirming previous observation by Makras and colleagues [41]. But, unlike described in a more recent manuscript, PTH levels did not further change after the first six months of treatment [42]. The increase of circulating PTH after 6 months of treatment was not associated with lower levels of VitD and calcium, but this rise could be caused indirectly as result of an increase in bone anabolic activity in response to the strong decrease of osteoclasts (OCs) activity, as hypothesized by Nakamura et al. [43].
In regard of the analysis of ALP levels, we only considered the first 12 months of treatment during which, generally, bone formation markers are lowered upon anti-resorptive therapy [44].
Unfortunately, a lower number of patients had blood test analysis data, but in those patients, who have been followed for a longer period, no further changes were observed compared to those that occurred within the first 24 months of therapy.
The increase of VitD levels after 6 months is likely due to the supplementation leading to an optimization. In fact, only a small proportion of patients (38.8%) used to take supplementation of VitD before starting denosumab therapy, but most of them did not assume it regularly and the others did not take any VitD supplementation at all.
It is important to enlighten that, as previously explained, this is a real-life study and not a randomized clinical trial, thus data were collected for 8 years every six months, but for several different reasons (compliance of patients to prescription, economic reason) not all subjects could always afford biochemical tests. Although this might be a limitation, we have observed and reported this lack of data, which might often occur in a real-life approach of patients affected by osteoporosis. Nevertheless, BMD results confirmed that the first 24 months are relevant to test the efficacy of denosumab therapy. Moreover, our results show that denosumab therapy can influence PTH, ALP, VitD and Ca levels within the first 24 months of treatment, while no further significant modification can be observed after then assuming that, once stabilized, it is not essential to request blood tests every six months. Indeed, the outcomes of this study might be very important from a socio-economic point of view and suggest that, stable patients, without other concomitant unstable health condition, can be evaluated after 12 months and the blood tests control at the same time. In everyday life this clinical approach could be most useful and in line with the purpose of saving social and health costs, but also maintaining a good quality of life of these fragile subjects.
The study has some limitations. Due to the nature of the study perform on outpatients, we could not have centralized blood test evaluations, which would have allowed a more accurate analysis of all the bone markers, nevertheless, the examination of biochemical markers can be considered comparable within the limitations of a real life study. In addition, the number of patients in the longer therapy period are a smaller number as compared with the more recent enrolled patients.
5. Conclusions
The dropout was confirmed within 12 months. Stable patients, without other concomitant unstable health condition, can be evaluated after 12 months. Our results indicate that denosumab increases his efficacy in patient previously treated with bisphosphonates.
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Author Contributions
Conceptualization, S.M., G.I., C.M.; methodology, G.Z., C.M.; software, G.Z., C.B.; validation, G.Z., E.M., C.M.; formal analysis, G.Z., M.C.G., V.M.B.; investigation, C.M., E.M., R.F., S.M.; resources, C.B., A.L., S.M.; data curation, G.Z., C.M.; writing—original draft preparation. G.Z., C.M.; writing—review and editing G.Z., C.M., S.M., G.I., M.C.G.; visualization, S.M.; supervision, S.M., G.I.; project administration, S.M.; funding acquisition, S.M., C.B., A.L. All authors have read and agreed to the published version of the manuscript.
Funding
V.M.B. was supported by a fellowship by MIUR grant number 2017HBHA98 to Silvia Migliaccio.
Institutional Review Board Statement
The study was conducted according to the guidelines of the Declaration of Helsinki, and was approved by the local Sapienza University, Policlinico Umberto I Institutional Review Board (prot n. 5061/18).
Informed Consent Statement
All participants gave their written informed consent before study participation.
Data Availability Statement
The data presented in this study are available on request from the corresponding authors.
Conflicts of Interest
The authors declare no conflict of interest.
Abbreviations
RANKL receptor activator of nuclear factor kappa B ligand
BMD bone mineral density; PTH, parathyroid hormone
VitD 25-hydroxy-vitamin D3
NFkB nuclear factor kappa-light-chain-enhancer of activated B cells
FN femoral neck; SD, standard deviations
BMI body mass index
HRT hormonal replacement therapy
BPs bisphosphonates
DXA Dual X-ray Absorptiometry
LS lumbar spine
F total proximal femur
Ca calcium
ALP alkaline phosphatase
AI aromatase inhibitors
BC breast cancer
NoBC without breast cancer
BPG BPs previously treated group
NG naive group
OCs osteoclasts
Figure 1 Percentage of change as compared to baseline of subjects with osteoporosis, osteopenia and normal bone density during treatment (* p < 0.05).
Figure 2 Comparison between BMD T-score (mean value, standard error) in NG (blue line) and BPG (red line) group at baseline T0 and after T1 denosumab. (A) Lumbar spine T-score measurement; (B) Femoral T-score measurement; (C), Femoral Neck T-score measurement. * Represents statistical significant difference effect for time (p ≤ 0.05), while # indicates statistical significant difference effect for time therapy (p ≤ 0.05) as more extensively reported in Table 4.
ijerph-18-01728-t001_Table 1Table 1 Study population characteristics at baseline (before denosumab treatment).
Parameters Value
Age (years) 72.8 ± 7.9
Age of menopause (years) 48.7 ± 4.7
BMI (kg/m2) 24.8 ± 4.5
Habitual smokers (n, %) 21 (6.3%)
Family History of osteoporosis (n, %) 107 (32.3%)
Family History of fragility fractures (n, %) 47 (14.1%)
At least 1 vertebral fragility fracture (n, %) 237 (71.3%)
FN osteoporotic fracture (n, %) 16 (4.8%)
Osteoporotic fracture at any other site (n, %) 33 (9.9%)
Previous HRT (n, %) 32 (9.6%)
History of glucocorticoid use (n, %) 3 (0.9%)
Previous supplementation with Ca/VitD (n, %) 129 (38.8%)
Prior anti-osteoporotic treatment (n, %) 206 (62.0%)
LS T-score (SD) −2.7 ± 1.2
F T-score (SD) −2.1 ± 0.9
FN T-score (SD) −2.5 ± 0.7
Serum PTH (pg/mL) 50.6 ± 23.3
Serum calcium (mg/dL) 9.5 ± 0.6
Serum ALP (U/L) 94.5 ± 52
Serum 25(OH)vitamin D3 (ng/mL) 32.8 ± 15
ijerph-18-01728-t002_Table 2Table 2 Causes of dropouts (n = 43/428, 10.04%).
Causes (n, %)
Death 6 (14.0%)
Refusal of therapy 13 (30.2%)
Unspecified disturbances 2 (4.6%)
Fear of collateral effects 4 (9.4%)
Fear of injectable drugs 3 (6.9%)
Suggested to change therapy 1 10 (23.2%)
Followed in another center 5 (11.6%)
1 by another specialist.
ijerph-18-01728-t003_Table 3Table 3 Prior anti-osteoporotic treatment (n = 206/332, 62.04%).
Treatment (n, %)
Bisphosphonates 122 (59.2%)
Strontium ranelate or Teriparatide 43 (20.9%)
Multiple treatment 41 (19.9%)
ijerph-18-01728-t004_Table 4Table 4 BMD comparison between NG and BPG group.
Group LS T-Score F T-Score FN T-Score
Number of subjects NG (n) 29 28 30
BPG (n) 37 37 35
ANOVA results Time effect (p) 1 <0.01 <0.01 <0.01
Time × therapy (p) 2 0.91 0.07 0.05
1 Efficacy of denosumab; 2 differences in efficacy of denosumab between the groups; n, number of subjects; p, statistical probability. | CALCIUM, DENOSUMAB | DrugsGivenReaction | CC BY | 33579002 | 18,955,234 | 2021-02-10 |
What was the administration route of drug 'DENOSUMAB'? | Efficacy of Denosumab Therapy Following Treatment with Bisphosphonates in Women with Osteoporosis: A Cohort Study.
Denosumab is a human monoclonal antibody that neutralizes RANKL, a cytokine able to interact with the RANK receptor on preosteoclasts and osteoclasts, decreasing their recruitment and differentiation, leading to a decreased bone resorption. The aim of this observational real-life study was to analyze adherence to denosumab therapy and assess its efficacy in increasing bone mineral density (BMD) and modulating biochemical skeletal markers following previous treatments with bisphosphonates in a group of post-menopausal women with osteoporosis. Women were recruited in the specialized center from March 2012 to September 2019. Biochemical markers were recorded at baseline and every six months prior to subsequent drug injection. Dual X-ray absorptiometry was requested at baseline and after 18/24 months. Comparing BMD at baseline and after denosumab therapy in naive patients and in those previously treated with bisphosphonates, a positive therapeutic effect was observed in both groups. The results of our real-life study demonstrate, as expected, that BMD values significantly increased upon denosumab treatment. Interestingly, denosumab showed an increased efficacy in patients previously treated with bisphosphonates. Moreover, biochemical markers data indicate that osteoporotic patients, without other concomitant unstable health conditions, could be evaluated once a year, decreasing the number of specialistic center access.
1. Introduction
During the last decades, in industrialized countries a significant increase of life expectancy with a gradual aging of the population has been observed. By 2060, America’s population of adults over the age of 65 is expected to double the current number [1]. In this scenario, the prolonged life expectancy, the inappropriate eating behaviors and lifestyle have contributed to the increased incidence of osteoporosis, a skeletal metabolic chronic disease characterized by compromised bone strength due to the reduction of both bone quantity and quality. The decreased bone strength leads the affected subjects to an increased risk of developing vertebral and non-vertebral fragility fractures, with increased morbidity, mortality, health service utilization, health care costs, and deterioration of quality of life [2,3,4,5,6,7,8].
All pharmacological treatments proposed in the last decades for osteoporotic patients aimed to reduce the risk of osteoporosis-related fractures by increasing bone mineral density and improving bone quality by modulating bone remodeling as demonstrated in controlled randomized clinical trial [9,10,11,12,13].
Denosumab, one of the latest pharmacological treatment developed as anti-osteoporotic treatment, is a fully human monoclonal antibody with high specificity to human RANKL, a receptor activator of NFκB ligand that is an essential factor in activating osteoclast differentiation and activity and, thus, activating bone resorption. This antiresorptive drug is administered by a subcutaneous injection every 6 months and it reversibly reduces osteoclast number and activity decreasing bone resorption. On the basis of this mechanism, denosumab has been demonstrated to significantly decrease new vertebral, nonvertebral, and hip fractures for up to 10 years of treatment in post-menopausal women affected by osteoporosis as demonstrated by the Fracture REduction Evaluation of Denosumab in Osteoporosis every 6 Months (FREEDOM) Extension trial [14,15,16].
Few studies, however, have evaluated adherence and efficacy of denosumab treatment in increasing BMD and potentially modulating biochemical skeletal markers in everyday life outpatients [17,18,19]. Thus, the aim of this observational real-life study was to analyze adherence to denosumab therapy and assess its efficacy in increasing bone mineral density (BMD) and modulating biochemical skeletal markers following previous treatments with bisphosphonates in a group of post-menopausal women with osteoporosis. In particular, it was evaluated and compared BMD at baseline and after denosumab therapy in either naive patients or patients previously treated with anti-resorptive drugs, since some previous studies suggested potential altered response to denosumab upon previous antiresorptive drugs [20,21,22].
2. Materials and Methods
2.1. Recruitment
Four hundred twenty-eight post-menopausal women (mean age = 71.7 ± 8.6 years) were recruited for this study. All subjects were affected by osteoporosis and referring to the specialized center for the management of osteoporosis in Policlinico Umberto I Hospital, Rome, from March 2012 to September 2019. Patients were evaluated every six months from the date of the initial prescription. The study respects the Declaration of Helsinki and was approved by the local Ethical Committees of the institution (prot 648/17) involved and met the guidelines of the responsible governmental agency.
2.2. Inclusion Criteria
Inclusion criteria were fixed on the basis of a previous work and AIFA’s advices for drug treatments: diagnosis of post-menopausal osteoporosis, possibility to receive a prescription during the study period according to the Italian reimbursement criteria for anti-osteoporotic pharmacological treatment [23,24].
Specifically, subjects with: (i) at least one prevalent hip or vertebral fracture (with contraindications to take oral pharmacological treatment), (ii) subjects aged ≥50 years with a femoral neck (FN) T-score ≤ −4 standard deviations (SD), (iii) subjects aged ≥50 years with a FN T-score ≤ −3 SD and at least one risk factor (family history of fragility vertebral or hip fractures, rheumatoid arthritis and other connective tissue diseases, previous wrist fragility fracture, early menopause, chronic corticosteroid therapy) were included.
The study was approved by the local University Institutional Review Board (prot n5061/18) and all participants gave their written informed consent before study participation.
2.3. Intervention
Before starting the new pharmacological treatment, the referring specialist or a resident fellow explained to the patients the mode of action and the method of taking the drug. Treatment consisted of a subcutaneous injection of denosumab (Prolia®, Amgen Inc. Thousand Oaks, CA 91320-1799, USA) 60 mg/every 6 months. Each patient was clinically evaluated before the first administration of denosumab (baseline) and then every 6 months, before the following injection, until the end of the observational study period. All patients continued taking all other concomitant therapies, including calcium and/or vitamin D supplementation. The specialist allowed all patients, or their relatives, to contact the clinical group via e-mail or telephone in case of doubts, questions or important communications. At the date of the initial prescription (baseline) the following data and information (y/n) were collected: age (years), age of menopause (years), weight (kg), height (m), body mass index (BMI), history of smoking; family history for osteoporosis; family history for fragility fractures; history for fragility vertebral and/or femoral and/or any other skeletal site fractures; previous hormonal replacement therapy (HRT), and/or chronic corticosteroid therapy; any previous and/or current supplementation with calcium and/or vitamin D and any previous antiresorptive therapy with bisphosphonates (BPs, including risedronate, alendronate, clodronate, zoledronic acid, ibandronate) or strontium ranelate or teriparatide. BMD [T-score at lumbar spine (LS), total proximal femur (F) and femoral neck (FN)] was evaluated by dual X-ray absorptiometry (DXA) measurement. In regards of the biochemical markers, serum parathyroid hormone (PTH), calcium (Ca), alkaline phosphatase (ALP), and 25(OH) vitamin D3 (VitD) were analyzed. Since this is a real-life study, blood tests were not centralized in our hospital center, but patients performed them in the morning between 07.00 a.m. and 09.00 a.m. after an overnight fast in other clinical centers which are required to use standardized methods (Table 1). As previously mentioned, some patients were under medications for other diseases, which however did not affect bone metabolism.
Weight (kg) and height (m) as anthropometric parameters were recorded to calculate BMI, according to the following formula: BMI = weight/height2 (kg/m2).
Osteoporosis was diagnosticated by DXA, based on the T-score for BMD less than or equal to −2.5 SD [25]. At baseline visit, lumbar, total proximal and femoral neck DXA were requested. Vertebrae affected by fractures, or other structural change or artifact, were excluded following the International Society for Clinical Densitometry—ISCD —official guidelines. Left hips affected by fractures, and underwent hip replacement surgery, were excluded and substituted by measurement of right hips. The following DXA was required after 18/24 months.
Biochemical markers were measured at baseline, approximately a week before drug administration, and at 6, 12, 18 and 24 months of denosumab treatment, in order to allow the patients to bring results to the visit.
For all subjects, bone density at lumbar spine, total proximal femur and femoral neck (LS T-score, F T-score and FN T-score) at baseline, were compared with values collected over time.
Of all participants who were prescribed denosumab, 43 dropped out (10.04%) before the end of the study. Reasons for drop-out are reported in the results section.
Since denosumab is also prescribed in post-menopausal women with hormone-responsive breast cancer under aromatase inhibitors (AI) adjuvant therapy, denosumab was prescribed to 53 women affected by breast cancer (BC group) [26]. In fact, as estrogens stimulate breast cancer cells, AI are aimed to inhibit the production of endogenous estrogens from aromatase in adipose tissue. On the other hand, low levels of estrogens can induce bone loss predisposing to osteoporosis and fractures [27,28,29]. BC group was excluded from the study for reasons that are reported in the results.
2.4. Statistical Analysis
Statistical analysis was performed on the following parameters: age, weight, height, BMI, age of menopause, BMD (LS T-score, F T-score, FN T-score), PTH, ALP, Ca and VitD. Differences between the means of above cited parameters were estimated using a two-sample t-test comparison between women with and without breast cancer. Pearson’s correlations among above cited parameters were estimated. The chi-square test was used to compare variables’ frequencies among groups (normal BMD, osteopenia, osteoporosis).
ANOVA_RM (repeated measures) were performed on BMD, PTH, ALP, Ca and VitD for different times. The time of the measurements were called T0 (baseline) and T1, T2, T3, T4, T5, T6, T7, T8 and T9 corresponding to 6, 12, 18, 24, 30, 36, 42, 48, 54 months, respectively. Statistical significance was defined as p ≤ 0.05. All statistical analysis was performed by SPSS version 24.0 software (SPSS Inc., Chicago, IL, USA).
3. Results
As previously described in the material and methods section, data were collected for each patient for a maximum of 8 years every six months from the beginning of the therapy, individually. Thus, we estimated and reported adherence of each patient, drop out and reason of it, densitometric analysis, biochemical markers, eventual missing data. To increase the power of the analysis, we considered the first 24 months of therapy for each patient in order to have a satisfactory number of patients to be considered.
3.1. Drop-Out
Of 428 enrolled subjects, 43 dropped out (10.04%) before the end of the study: in particular, the majority dropped within the first 12 months of therapy (87%), whereas the rest (13%) dropped out after 12 months. Thus, from our data it results that outpatients had an adherence to therapy of 89.6% up to the end of therapy. As depicted in Table 2, reasons of drop out were: death (6/43, 14%), refusal of therapy (13/43, 30.2%), unspecific disturbs (2/43, 4.6%), fear of collateral effects (4/43, 9.4%), fear of injective drugs (3/43, 6.9%), suggested by another specialist to change therapy (10/43, 23.2%), followed in another specialized center (5/43, 11.6%) (Table 2). Indeed, several studies have demonstrated that higher number of drop out occurs within the first 12 months of chronic pharmacological intervention [30,31,32].
3.2. Comparison between Groups of Women with or without Breast Cancer
Two-sample t-test comparison between the group of 53 women with breast cancer (BC) and the group of 332 women without breast cancer (NoBC) depicted significant differences in age (p < 0.01). Consequently, the two groups could not be studied as a single population and BC women in therapy con denosumab were not included in the statistical analysis of this study to address the aim of the study.
3.3. Partition between Naive Subjects and Subjects Treated with Previous Therapies
Of the 332 remaining patients (mean age = 72.8 ± 7.9 years), 237 women had at least one vertebral fragility fracture and 16 women had a previous femoral fracture.
From this group of 332 women, 126 subjects were considered naive (no previous anti-osteoporotic therapy), 122 were previously treated with BPs therapy, 43 either teriparatide or strontium ranelate and 41 had received two or more of the above-mentioned therapies, as shown in Table 3. In order to compare naive group (NG) with BPs previously treated group (BPG) data, the 84 subjects who received teriparatide or strontium ranelate prior to denosumab were not included in statistical analysis (ANOVA).
3.4. Pearson’s Correlations at Baseline
Pearson’s correlations among main parameters at baseline were estimated.
Interestingly, analysis showed that BMD femoral neck (FN T-score) was significantly negatively related with age (p < 0.01). Correlation values showed that menopause age was positively correlated with F T-score (p < 0.01) and FN T-score (p < 0.01), age was positively correlated with BMI (p < 0.01) and LS T-score (p < 0.05). VitD was inversely correlated with weight (p < 0.05) and Ca was negatively correlated only with PTH (p < 0.01). FN T-score was positively correlated with all parameters analyzed (p < 0.01) except Ca and VitD, while it was negatively correlated with PTH (p < 0.05).
3.5. Efficacy of Denosumab Therapy in Increasing BMD
We compared BMD values at lumbar spine, femoral and hip neck before (at baseline (T0) and after 24 (T4), 48 (T8) and 54 (T9) months of pharmacological treatment. In Figure 1, as expected, compared to baseline, BMD values significantly increased after 24 months of treatment. The chi-square test revealed that there was a significant change in BMD in patients in therapy with denosumab: evaluation showed that the number of subjects affected by a low BMD, compatible with an osteoporotic T-score was reduced after 48 months of denosumab therapy, whereas the percent of subjects with osteopenia was increased.
3.6. Previous Therapy
The interest was to evaluate whether previous therapies could have influenced the efficacy of denosumab therapy. Two different groups were considered: a group of subjects (n = 122) previously treated at least with one of the mentioned different bisphosphonates (BPG) and a naive group of subjects (n = 126) that have no taken any previous anti-osteoporotic medication (NG). BMD measurements (LS T-score, F T-score F and FN T-score) before (at baseline) and after denosumab therapy were compared by ANOVA_RM (Table 4).
A significant improvement (p < 0.01) was observed after denosumab therapy (Figure 2A–C). Differences between the two groups (previous therapy effect, see last line of Table 4) was not observed in Lumbar spine (Figure 2A) and Femoral (LS T-score, p = 0.91 and F T-score, p = 0.07) (Figure 2B) but was observed in Femoral Neck (FN T-score, p = 0.05) (Figure 2C).
3.7. Biochemical Markers
In order to study whether changes in biochemical marker values occurred during denosumab treatment in related variables, ANOVA (by ANOVA_RM) analysis were performed on PTH, ALP, Ca and VitD for different times.
It was observed that PTH levels significantly increased from T0 to T1 (N = 36, p < 0.001) but did not change substantially until 24 months. As expected by an anti-resorptive drug, ALP levels markedly decreased following treatment (N = 51, p < 0.001). In later time-points, biochemical markers data were available for a lower number of patients, which however demonstrated no further changes as compared to those that occurred within the first 24 months of therapy. Indeed, serum calcium levels remained unchanged upon treatment (ANOVA, N = 39, p > 0.05) while serum VitD levels significantly increased from T0 to T1 but did not change significantly after T3 (ANOVA, N = 79, p < 0.001) (data not showed).
4. Discussion
This Italian observational real-life study evaluated the effect of denosumab in a group of post-menopausal women. The results showed a good response in BMD after 2 years of denosumab treatment in all skeletal sites evaluated, as demonstrated by the decreased percent of BMD values compatible with osteoporosis and the increased percent of BMD values compatible with osteopenia. As expected, these data, regarding the improvements of BMD in a population of osteoporotic out-patients, confirm results already showed in randomized clinical trial [33,34,35].
Moreover, our findings, comparing BMD at baseline and over time in naive group (NG) with BPs previously treated group (BPG), showed a significant improvement of BMD in all skeletal sites evaluated (p < 0.01) after denosumab treatment in both groups of patients, suggesting that previous BP therapy does not lead to a decreased efficacy of denosumab. On the other hand, it is interesting to note that the increase of BMD at femoral neck was significantly higher (p = 0.05) in patients previously treated with BPs as previously showed by Sánchez et al., suggesting a therapy effect [17]. These data could be explained by the different mechanism of action of these anti-resorptive drugs: while the effect of denosumab on bone metabolism might stop upon discontinuation, BPs might maintain the antiresorptive effect even after discontinuation [36].
Pearson’s correlations at baseline among BMD regions confirm FN T-score as the most reliable measure, being negatively and positively correlated with age and menopause age, respectively. Indeed, our data further indicate that older patients have a lower FN T-score at baseline and that, as expected, an older age at menopause has a protective effect on the bone, likely due to circulating estrogens, leading to higher FN T-score. Further, considering other parameters, VitD was inversely correlated with body weight, as calculated by BMI, since in the present study body composition data were collected. Nevertheless, on the base of BMI of the patients, we can speculate that higher levels of fat mass could contribute to low VitD levels, since numerous studies, including the ones of our group, have shown that obese/overweight subjects have lower vitamin D levels, considering that the storage of this fat-soluble vitamin is mainly in the adipose tissue [37,38,39,40].
Interestingly, the measurement of bone biochemical markers after the first infusion of denosumab, showed a significant increase of serum PTH and VitD levels after 6 months of denosumab treatment (p < 0.001), confirming previous observation by Makras and colleagues [41]. But, unlike described in a more recent manuscript, PTH levels did not further change after the first six months of treatment [42]. The increase of circulating PTH after 6 months of treatment was not associated with lower levels of VitD and calcium, but this rise could be caused indirectly as result of an increase in bone anabolic activity in response to the strong decrease of osteoclasts (OCs) activity, as hypothesized by Nakamura et al. [43].
In regard of the analysis of ALP levels, we only considered the first 12 months of treatment during which, generally, bone formation markers are lowered upon anti-resorptive therapy [44].
Unfortunately, a lower number of patients had blood test analysis data, but in those patients, who have been followed for a longer period, no further changes were observed compared to those that occurred within the first 24 months of therapy.
The increase of VitD levels after 6 months is likely due to the supplementation leading to an optimization. In fact, only a small proportion of patients (38.8%) used to take supplementation of VitD before starting denosumab therapy, but most of them did not assume it regularly and the others did not take any VitD supplementation at all.
It is important to enlighten that, as previously explained, this is a real-life study and not a randomized clinical trial, thus data were collected for 8 years every six months, but for several different reasons (compliance of patients to prescription, economic reason) not all subjects could always afford biochemical tests. Although this might be a limitation, we have observed and reported this lack of data, which might often occur in a real-life approach of patients affected by osteoporosis. Nevertheless, BMD results confirmed that the first 24 months are relevant to test the efficacy of denosumab therapy. Moreover, our results show that denosumab therapy can influence PTH, ALP, VitD and Ca levels within the first 24 months of treatment, while no further significant modification can be observed after then assuming that, once stabilized, it is not essential to request blood tests every six months. Indeed, the outcomes of this study might be very important from a socio-economic point of view and suggest that, stable patients, without other concomitant unstable health condition, can be evaluated after 12 months and the blood tests control at the same time. In everyday life this clinical approach could be most useful and in line with the purpose of saving social and health costs, but also maintaining a good quality of life of these fragile subjects.
The study has some limitations. Due to the nature of the study perform on outpatients, we could not have centralized blood test evaluations, which would have allowed a more accurate analysis of all the bone markers, nevertheless, the examination of biochemical markers can be considered comparable within the limitations of a real life study. In addition, the number of patients in the longer therapy period are a smaller number as compared with the more recent enrolled patients.
5. Conclusions
The dropout was confirmed within 12 months. Stable patients, without other concomitant unstable health condition, can be evaluated after 12 months. Our results indicate that denosumab increases his efficacy in patient previously treated with bisphosphonates.
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Author Contributions
Conceptualization, S.M., G.I., C.M.; methodology, G.Z., C.M.; software, G.Z., C.B.; validation, G.Z., E.M., C.M.; formal analysis, G.Z., M.C.G., V.M.B.; investigation, C.M., E.M., R.F., S.M.; resources, C.B., A.L., S.M.; data curation, G.Z., C.M.; writing—original draft preparation. G.Z., C.M.; writing—review and editing G.Z., C.M., S.M., G.I., M.C.G.; visualization, S.M.; supervision, S.M., G.I.; project administration, S.M.; funding acquisition, S.M., C.B., A.L. All authors have read and agreed to the published version of the manuscript.
Funding
V.M.B. was supported by a fellowship by MIUR grant number 2017HBHA98 to Silvia Migliaccio.
Institutional Review Board Statement
The study was conducted according to the guidelines of the Declaration of Helsinki, and was approved by the local Sapienza University, Policlinico Umberto I Institutional Review Board (prot n. 5061/18).
Informed Consent Statement
All participants gave their written informed consent before study participation.
Data Availability Statement
The data presented in this study are available on request from the corresponding authors.
Conflicts of Interest
The authors declare no conflict of interest.
Abbreviations
RANKL receptor activator of nuclear factor kappa B ligand
BMD bone mineral density; PTH, parathyroid hormone
VitD 25-hydroxy-vitamin D3
NFkB nuclear factor kappa-light-chain-enhancer of activated B cells
FN femoral neck; SD, standard deviations
BMI body mass index
HRT hormonal replacement therapy
BPs bisphosphonates
DXA Dual X-ray Absorptiometry
LS lumbar spine
F total proximal femur
Ca calcium
ALP alkaline phosphatase
AI aromatase inhibitors
BC breast cancer
NoBC without breast cancer
BPG BPs previously treated group
NG naive group
OCs osteoclasts
Figure 1 Percentage of change as compared to baseline of subjects with osteoporosis, osteopenia and normal bone density during treatment (* p < 0.05).
Figure 2 Comparison between BMD T-score (mean value, standard error) in NG (blue line) and BPG (red line) group at baseline T0 and after T1 denosumab. (A) Lumbar spine T-score measurement; (B) Femoral T-score measurement; (C), Femoral Neck T-score measurement. * Represents statistical significant difference effect for time (p ≤ 0.05), while # indicates statistical significant difference effect for time therapy (p ≤ 0.05) as more extensively reported in Table 4.
ijerph-18-01728-t001_Table 1Table 1 Study population characteristics at baseline (before denosumab treatment).
Parameters Value
Age (years) 72.8 ± 7.9
Age of menopause (years) 48.7 ± 4.7
BMI (kg/m2) 24.8 ± 4.5
Habitual smokers (n, %) 21 (6.3%)
Family History of osteoporosis (n, %) 107 (32.3%)
Family History of fragility fractures (n, %) 47 (14.1%)
At least 1 vertebral fragility fracture (n, %) 237 (71.3%)
FN osteoporotic fracture (n, %) 16 (4.8%)
Osteoporotic fracture at any other site (n, %) 33 (9.9%)
Previous HRT (n, %) 32 (9.6%)
History of glucocorticoid use (n, %) 3 (0.9%)
Previous supplementation with Ca/VitD (n, %) 129 (38.8%)
Prior anti-osteoporotic treatment (n, %) 206 (62.0%)
LS T-score (SD) −2.7 ± 1.2
F T-score (SD) −2.1 ± 0.9
FN T-score (SD) −2.5 ± 0.7
Serum PTH (pg/mL) 50.6 ± 23.3
Serum calcium (mg/dL) 9.5 ± 0.6
Serum ALP (U/L) 94.5 ± 52
Serum 25(OH)vitamin D3 (ng/mL) 32.8 ± 15
ijerph-18-01728-t002_Table 2Table 2 Causes of dropouts (n = 43/428, 10.04%).
Causes (n, %)
Death 6 (14.0%)
Refusal of therapy 13 (30.2%)
Unspecified disturbances 2 (4.6%)
Fear of collateral effects 4 (9.4%)
Fear of injectable drugs 3 (6.9%)
Suggested to change therapy 1 10 (23.2%)
Followed in another center 5 (11.6%)
1 by another specialist.
ijerph-18-01728-t003_Table 3Table 3 Prior anti-osteoporotic treatment (n = 206/332, 62.04%).
Treatment (n, %)
Bisphosphonates 122 (59.2%)
Strontium ranelate or Teriparatide 43 (20.9%)
Multiple treatment 41 (19.9%)
ijerph-18-01728-t004_Table 4Table 4 BMD comparison between NG and BPG group.
Group LS T-Score F T-Score FN T-Score
Number of subjects NG (n) 29 28 30
BPG (n) 37 37 35
ANOVA results Time effect (p) 1 <0.01 <0.01 <0.01
Time × therapy (p) 2 0.91 0.07 0.05
1 Efficacy of denosumab; 2 differences in efficacy of denosumab between the groups; n, number of subjects; p, statistical probability. | Subcutaneous | DrugAdministrationRoute | CC BY | 33579002 | 18,955,234 | 2021-02-10 |
What was the outcome of reaction 'Death'? | Efficacy of Denosumab Therapy Following Treatment with Bisphosphonates in Women with Osteoporosis: A Cohort Study.
Denosumab is a human monoclonal antibody that neutralizes RANKL, a cytokine able to interact with the RANK receptor on preosteoclasts and osteoclasts, decreasing their recruitment and differentiation, leading to a decreased bone resorption. The aim of this observational real-life study was to analyze adherence to denosumab therapy and assess its efficacy in increasing bone mineral density (BMD) and modulating biochemical skeletal markers following previous treatments with bisphosphonates in a group of post-menopausal women with osteoporosis. Women were recruited in the specialized center from March 2012 to September 2019. Biochemical markers were recorded at baseline and every six months prior to subsequent drug injection. Dual X-ray absorptiometry was requested at baseline and after 18/24 months. Comparing BMD at baseline and after denosumab therapy in naive patients and in those previously treated with bisphosphonates, a positive therapeutic effect was observed in both groups. The results of our real-life study demonstrate, as expected, that BMD values significantly increased upon denosumab treatment. Interestingly, denosumab showed an increased efficacy in patients previously treated with bisphosphonates. Moreover, biochemical markers data indicate that osteoporotic patients, without other concomitant unstable health conditions, could be evaluated once a year, decreasing the number of specialistic center access.
1. Introduction
During the last decades, in industrialized countries a significant increase of life expectancy with a gradual aging of the population has been observed. By 2060, America’s population of adults over the age of 65 is expected to double the current number [1]. In this scenario, the prolonged life expectancy, the inappropriate eating behaviors and lifestyle have contributed to the increased incidence of osteoporosis, a skeletal metabolic chronic disease characterized by compromised bone strength due to the reduction of both bone quantity and quality. The decreased bone strength leads the affected subjects to an increased risk of developing vertebral and non-vertebral fragility fractures, with increased morbidity, mortality, health service utilization, health care costs, and deterioration of quality of life [2,3,4,5,6,7,8].
All pharmacological treatments proposed in the last decades for osteoporotic patients aimed to reduce the risk of osteoporosis-related fractures by increasing bone mineral density and improving bone quality by modulating bone remodeling as demonstrated in controlled randomized clinical trial [9,10,11,12,13].
Denosumab, one of the latest pharmacological treatment developed as anti-osteoporotic treatment, is a fully human monoclonal antibody with high specificity to human RANKL, a receptor activator of NFκB ligand that is an essential factor in activating osteoclast differentiation and activity and, thus, activating bone resorption. This antiresorptive drug is administered by a subcutaneous injection every 6 months and it reversibly reduces osteoclast number and activity decreasing bone resorption. On the basis of this mechanism, denosumab has been demonstrated to significantly decrease new vertebral, nonvertebral, and hip fractures for up to 10 years of treatment in post-menopausal women affected by osteoporosis as demonstrated by the Fracture REduction Evaluation of Denosumab in Osteoporosis every 6 Months (FREEDOM) Extension trial [14,15,16].
Few studies, however, have evaluated adherence and efficacy of denosumab treatment in increasing BMD and potentially modulating biochemical skeletal markers in everyday life outpatients [17,18,19]. Thus, the aim of this observational real-life study was to analyze adherence to denosumab therapy and assess its efficacy in increasing bone mineral density (BMD) and modulating biochemical skeletal markers following previous treatments with bisphosphonates in a group of post-menopausal women with osteoporosis. In particular, it was evaluated and compared BMD at baseline and after denosumab therapy in either naive patients or patients previously treated with anti-resorptive drugs, since some previous studies suggested potential altered response to denosumab upon previous antiresorptive drugs [20,21,22].
2. Materials and Methods
2.1. Recruitment
Four hundred twenty-eight post-menopausal women (mean age = 71.7 ± 8.6 years) were recruited for this study. All subjects were affected by osteoporosis and referring to the specialized center for the management of osteoporosis in Policlinico Umberto I Hospital, Rome, from March 2012 to September 2019. Patients were evaluated every six months from the date of the initial prescription. The study respects the Declaration of Helsinki and was approved by the local Ethical Committees of the institution (prot 648/17) involved and met the guidelines of the responsible governmental agency.
2.2. Inclusion Criteria
Inclusion criteria were fixed on the basis of a previous work and AIFA’s advices for drug treatments: diagnosis of post-menopausal osteoporosis, possibility to receive a prescription during the study period according to the Italian reimbursement criteria for anti-osteoporotic pharmacological treatment [23,24].
Specifically, subjects with: (i) at least one prevalent hip or vertebral fracture (with contraindications to take oral pharmacological treatment), (ii) subjects aged ≥50 years with a femoral neck (FN) T-score ≤ −4 standard deviations (SD), (iii) subjects aged ≥50 years with a FN T-score ≤ −3 SD and at least one risk factor (family history of fragility vertebral or hip fractures, rheumatoid arthritis and other connective tissue diseases, previous wrist fragility fracture, early menopause, chronic corticosteroid therapy) were included.
The study was approved by the local University Institutional Review Board (prot n5061/18) and all participants gave their written informed consent before study participation.
2.3. Intervention
Before starting the new pharmacological treatment, the referring specialist or a resident fellow explained to the patients the mode of action and the method of taking the drug. Treatment consisted of a subcutaneous injection of denosumab (Prolia®, Amgen Inc. Thousand Oaks, CA 91320-1799, USA) 60 mg/every 6 months. Each patient was clinically evaluated before the first administration of denosumab (baseline) and then every 6 months, before the following injection, until the end of the observational study period. All patients continued taking all other concomitant therapies, including calcium and/or vitamin D supplementation. The specialist allowed all patients, or their relatives, to contact the clinical group via e-mail or telephone in case of doubts, questions or important communications. At the date of the initial prescription (baseline) the following data and information (y/n) were collected: age (years), age of menopause (years), weight (kg), height (m), body mass index (BMI), history of smoking; family history for osteoporosis; family history for fragility fractures; history for fragility vertebral and/or femoral and/or any other skeletal site fractures; previous hormonal replacement therapy (HRT), and/or chronic corticosteroid therapy; any previous and/or current supplementation with calcium and/or vitamin D and any previous antiresorptive therapy with bisphosphonates (BPs, including risedronate, alendronate, clodronate, zoledronic acid, ibandronate) or strontium ranelate or teriparatide. BMD [T-score at lumbar spine (LS), total proximal femur (F) and femoral neck (FN)] was evaluated by dual X-ray absorptiometry (DXA) measurement. In regards of the biochemical markers, serum parathyroid hormone (PTH), calcium (Ca), alkaline phosphatase (ALP), and 25(OH) vitamin D3 (VitD) were analyzed. Since this is a real-life study, blood tests were not centralized in our hospital center, but patients performed them in the morning between 07.00 a.m. and 09.00 a.m. after an overnight fast in other clinical centers which are required to use standardized methods (Table 1). As previously mentioned, some patients were under medications for other diseases, which however did not affect bone metabolism.
Weight (kg) and height (m) as anthropometric parameters were recorded to calculate BMI, according to the following formula: BMI = weight/height2 (kg/m2).
Osteoporosis was diagnosticated by DXA, based on the T-score for BMD less than or equal to −2.5 SD [25]. At baseline visit, lumbar, total proximal and femoral neck DXA were requested. Vertebrae affected by fractures, or other structural change or artifact, were excluded following the International Society for Clinical Densitometry—ISCD —official guidelines. Left hips affected by fractures, and underwent hip replacement surgery, were excluded and substituted by measurement of right hips. The following DXA was required after 18/24 months.
Biochemical markers were measured at baseline, approximately a week before drug administration, and at 6, 12, 18 and 24 months of denosumab treatment, in order to allow the patients to bring results to the visit.
For all subjects, bone density at lumbar spine, total proximal femur and femoral neck (LS T-score, F T-score and FN T-score) at baseline, were compared with values collected over time.
Of all participants who were prescribed denosumab, 43 dropped out (10.04%) before the end of the study. Reasons for drop-out are reported in the results section.
Since denosumab is also prescribed in post-menopausal women with hormone-responsive breast cancer under aromatase inhibitors (AI) adjuvant therapy, denosumab was prescribed to 53 women affected by breast cancer (BC group) [26]. In fact, as estrogens stimulate breast cancer cells, AI are aimed to inhibit the production of endogenous estrogens from aromatase in adipose tissue. On the other hand, low levels of estrogens can induce bone loss predisposing to osteoporosis and fractures [27,28,29]. BC group was excluded from the study for reasons that are reported in the results.
2.4. Statistical Analysis
Statistical analysis was performed on the following parameters: age, weight, height, BMI, age of menopause, BMD (LS T-score, F T-score, FN T-score), PTH, ALP, Ca and VitD. Differences between the means of above cited parameters were estimated using a two-sample t-test comparison between women with and without breast cancer. Pearson’s correlations among above cited parameters were estimated. The chi-square test was used to compare variables’ frequencies among groups (normal BMD, osteopenia, osteoporosis).
ANOVA_RM (repeated measures) were performed on BMD, PTH, ALP, Ca and VitD for different times. The time of the measurements were called T0 (baseline) and T1, T2, T3, T4, T5, T6, T7, T8 and T9 corresponding to 6, 12, 18, 24, 30, 36, 42, 48, 54 months, respectively. Statistical significance was defined as p ≤ 0.05. All statistical analysis was performed by SPSS version 24.0 software (SPSS Inc., Chicago, IL, USA).
3. Results
As previously described in the material and methods section, data were collected for each patient for a maximum of 8 years every six months from the beginning of the therapy, individually. Thus, we estimated and reported adherence of each patient, drop out and reason of it, densitometric analysis, biochemical markers, eventual missing data. To increase the power of the analysis, we considered the first 24 months of therapy for each patient in order to have a satisfactory number of patients to be considered.
3.1. Drop-Out
Of 428 enrolled subjects, 43 dropped out (10.04%) before the end of the study: in particular, the majority dropped within the first 12 months of therapy (87%), whereas the rest (13%) dropped out after 12 months. Thus, from our data it results that outpatients had an adherence to therapy of 89.6% up to the end of therapy. As depicted in Table 2, reasons of drop out were: death (6/43, 14%), refusal of therapy (13/43, 30.2%), unspecific disturbs (2/43, 4.6%), fear of collateral effects (4/43, 9.4%), fear of injective drugs (3/43, 6.9%), suggested by another specialist to change therapy (10/43, 23.2%), followed in another specialized center (5/43, 11.6%) (Table 2). Indeed, several studies have demonstrated that higher number of drop out occurs within the first 12 months of chronic pharmacological intervention [30,31,32].
3.2. Comparison between Groups of Women with or without Breast Cancer
Two-sample t-test comparison between the group of 53 women with breast cancer (BC) and the group of 332 women without breast cancer (NoBC) depicted significant differences in age (p < 0.01). Consequently, the two groups could not be studied as a single population and BC women in therapy con denosumab were not included in the statistical analysis of this study to address the aim of the study.
3.3. Partition between Naive Subjects and Subjects Treated with Previous Therapies
Of the 332 remaining patients (mean age = 72.8 ± 7.9 years), 237 women had at least one vertebral fragility fracture and 16 women had a previous femoral fracture.
From this group of 332 women, 126 subjects were considered naive (no previous anti-osteoporotic therapy), 122 were previously treated with BPs therapy, 43 either teriparatide or strontium ranelate and 41 had received two or more of the above-mentioned therapies, as shown in Table 3. In order to compare naive group (NG) with BPs previously treated group (BPG) data, the 84 subjects who received teriparatide or strontium ranelate prior to denosumab were not included in statistical analysis (ANOVA).
3.4. Pearson’s Correlations at Baseline
Pearson’s correlations among main parameters at baseline were estimated.
Interestingly, analysis showed that BMD femoral neck (FN T-score) was significantly negatively related with age (p < 0.01). Correlation values showed that menopause age was positively correlated with F T-score (p < 0.01) and FN T-score (p < 0.01), age was positively correlated with BMI (p < 0.01) and LS T-score (p < 0.05). VitD was inversely correlated with weight (p < 0.05) and Ca was negatively correlated only with PTH (p < 0.01). FN T-score was positively correlated with all parameters analyzed (p < 0.01) except Ca and VitD, while it was negatively correlated with PTH (p < 0.05).
3.5. Efficacy of Denosumab Therapy in Increasing BMD
We compared BMD values at lumbar spine, femoral and hip neck before (at baseline (T0) and after 24 (T4), 48 (T8) and 54 (T9) months of pharmacological treatment. In Figure 1, as expected, compared to baseline, BMD values significantly increased after 24 months of treatment. The chi-square test revealed that there was a significant change in BMD in patients in therapy with denosumab: evaluation showed that the number of subjects affected by a low BMD, compatible with an osteoporotic T-score was reduced after 48 months of denosumab therapy, whereas the percent of subjects with osteopenia was increased.
3.6. Previous Therapy
The interest was to evaluate whether previous therapies could have influenced the efficacy of denosumab therapy. Two different groups were considered: a group of subjects (n = 122) previously treated at least with one of the mentioned different bisphosphonates (BPG) and a naive group of subjects (n = 126) that have no taken any previous anti-osteoporotic medication (NG). BMD measurements (LS T-score, F T-score F and FN T-score) before (at baseline) and after denosumab therapy were compared by ANOVA_RM (Table 4).
A significant improvement (p < 0.01) was observed after denosumab therapy (Figure 2A–C). Differences between the two groups (previous therapy effect, see last line of Table 4) was not observed in Lumbar spine (Figure 2A) and Femoral (LS T-score, p = 0.91 and F T-score, p = 0.07) (Figure 2B) but was observed in Femoral Neck (FN T-score, p = 0.05) (Figure 2C).
3.7. Biochemical Markers
In order to study whether changes in biochemical marker values occurred during denosumab treatment in related variables, ANOVA (by ANOVA_RM) analysis were performed on PTH, ALP, Ca and VitD for different times.
It was observed that PTH levels significantly increased from T0 to T1 (N = 36, p < 0.001) but did not change substantially until 24 months. As expected by an anti-resorptive drug, ALP levels markedly decreased following treatment (N = 51, p < 0.001). In later time-points, biochemical markers data were available for a lower number of patients, which however demonstrated no further changes as compared to those that occurred within the first 24 months of therapy. Indeed, serum calcium levels remained unchanged upon treatment (ANOVA, N = 39, p > 0.05) while serum VitD levels significantly increased from T0 to T1 but did not change significantly after T3 (ANOVA, N = 79, p < 0.001) (data not showed).
4. Discussion
This Italian observational real-life study evaluated the effect of denosumab in a group of post-menopausal women. The results showed a good response in BMD after 2 years of denosumab treatment in all skeletal sites evaluated, as demonstrated by the decreased percent of BMD values compatible with osteoporosis and the increased percent of BMD values compatible with osteopenia. As expected, these data, regarding the improvements of BMD in a population of osteoporotic out-patients, confirm results already showed in randomized clinical trial [33,34,35].
Moreover, our findings, comparing BMD at baseline and over time in naive group (NG) with BPs previously treated group (BPG), showed a significant improvement of BMD in all skeletal sites evaluated (p < 0.01) after denosumab treatment in both groups of patients, suggesting that previous BP therapy does not lead to a decreased efficacy of denosumab. On the other hand, it is interesting to note that the increase of BMD at femoral neck was significantly higher (p = 0.05) in patients previously treated with BPs as previously showed by Sánchez et al., suggesting a therapy effect [17]. These data could be explained by the different mechanism of action of these anti-resorptive drugs: while the effect of denosumab on bone metabolism might stop upon discontinuation, BPs might maintain the antiresorptive effect even after discontinuation [36].
Pearson’s correlations at baseline among BMD regions confirm FN T-score as the most reliable measure, being negatively and positively correlated with age and menopause age, respectively. Indeed, our data further indicate that older patients have a lower FN T-score at baseline and that, as expected, an older age at menopause has a protective effect on the bone, likely due to circulating estrogens, leading to higher FN T-score. Further, considering other parameters, VitD was inversely correlated with body weight, as calculated by BMI, since in the present study body composition data were collected. Nevertheless, on the base of BMI of the patients, we can speculate that higher levels of fat mass could contribute to low VitD levels, since numerous studies, including the ones of our group, have shown that obese/overweight subjects have lower vitamin D levels, considering that the storage of this fat-soluble vitamin is mainly in the adipose tissue [37,38,39,40].
Interestingly, the measurement of bone biochemical markers after the first infusion of denosumab, showed a significant increase of serum PTH and VitD levels after 6 months of denosumab treatment (p < 0.001), confirming previous observation by Makras and colleagues [41]. But, unlike described in a more recent manuscript, PTH levels did not further change after the first six months of treatment [42]. The increase of circulating PTH after 6 months of treatment was not associated with lower levels of VitD and calcium, but this rise could be caused indirectly as result of an increase in bone anabolic activity in response to the strong decrease of osteoclasts (OCs) activity, as hypothesized by Nakamura et al. [43].
In regard of the analysis of ALP levels, we only considered the first 12 months of treatment during which, generally, bone formation markers are lowered upon anti-resorptive therapy [44].
Unfortunately, a lower number of patients had blood test analysis data, but in those patients, who have been followed for a longer period, no further changes were observed compared to those that occurred within the first 24 months of therapy.
The increase of VitD levels after 6 months is likely due to the supplementation leading to an optimization. In fact, only a small proportion of patients (38.8%) used to take supplementation of VitD before starting denosumab therapy, but most of them did not assume it regularly and the others did not take any VitD supplementation at all.
It is important to enlighten that, as previously explained, this is a real-life study and not a randomized clinical trial, thus data were collected for 8 years every six months, but for several different reasons (compliance of patients to prescription, economic reason) not all subjects could always afford biochemical tests. Although this might be a limitation, we have observed and reported this lack of data, which might often occur in a real-life approach of patients affected by osteoporosis. Nevertheless, BMD results confirmed that the first 24 months are relevant to test the efficacy of denosumab therapy. Moreover, our results show that denosumab therapy can influence PTH, ALP, VitD and Ca levels within the first 24 months of treatment, while no further significant modification can be observed after then assuming that, once stabilized, it is not essential to request blood tests every six months. Indeed, the outcomes of this study might be very important from a socio-economic point of view and suggest that, stable patients, without other concomitant unstable health condition, can be evaluated after 12 months and the blood tests control at the same time. In everyday life this clinical approach could be most useful and in line with the purpose of saving social and health costs, but also maintaining a good quality of life of these fragile subjects.
The study has some limitations. Due to the nature of the study perform on outpatients, we could not have centralized blood test evaluations, which would have allowed a more accurate analysis of all the bone markers, nevertheless, the examination of biochemical markers can be considered comparable within the limitations of a real life study. In addition, the number of patients in the longer therapy period are a smaller number as compared with the more recent enrolled patients.
5. Conclusions
The dropout was confirmed within 12 months. Stable patients, without other concomitant unstable health condition, can be evaluated after 12 months. Our results indicate that denosumab increases his efficacy in patient previously treated with bisphosphonates.
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Author Contributions
Conceptualization, S.M., G.I., C.M.; methodology, G.Z., C.M.; software, G.Z., C.B.; validation, G.Z., E.M., C.M.; formal analysis, G.Z., M.C.G., V.M.B.; investigation, C.M., E.M., R.F., S.M.; resources, C.B., A.L., S.M.; data curation, G.Z., C.M.; writing—original draft preparation. G.Z., C.M.; writing—review and editing G.Z., C.M., S.M., G.I., M.C.G.; visualization, S.M.; supervision, S.M., G.I.; project administration, S.M.; funding acquisition, S.M., C.B., A.L. All authors have read and agreed to the published version of the manuscript.
Funding
V.M.B. was supported by a fellowship by MIUR grant number 2017HBHA98 to Silvia Migliaccio.
Institutional Review Board Statement
The study was conducted according to the guidelines of the Declaration of Helsinki, and was approved by the local Sapienza University, Policlinico Umberto I Institutional Review Board (prot n. 5061/18).
Informed Consent Statement
All participants gave their written informed consent before study participation.
Data Availability Statement
The data presented in this study are available on request from the corresponding authors.
Conflicts of Interest
The authors declare no conflict of interest.
Abbreviations
RANKL receptor activator of nuclear factor kappa B ligand
BMD bone mineral density; PTH, parathyroid hormone
VitD 25-hydroxy-vitamin D3
NFkB nuclear factor kappa-light-chain-enhancer of activated B cells
FN femoral neck; SD, standard deviations
BMI body mass index
HRT hormonal replacement therapy
BPs bisphosphonates
DXA Dual X-ray Absorptiometry
LS lumbar spine
F total proximal femur
Ca calcium
ALP alkaline phosphatase
AI aromatase inhibitors
BC breast cancer
NoBC without breast cancer
BPG BPs previously treated group
NG naive group
OCs osteoclasts
Figure 1 Percentage of change as compared to baseline of subjects with osteoporosis, osteopenia and normal bone density during treatment (* p < 0.05).
Figure 2 Comparison between BMD T-score (mean value, standard error) in NG (blue line) and BPG (red line) group at baseline T0 and after T1 denosumab. (A) Lumbar spine T-score measurement; (B) Femoral T-score measurement; (C), Femoral Neck T-score measurement. * Represents statistical significant difference effect for time (p ≤ 0.05), while # indicates statistical significant difference effect for time therapy (p ≤ 0.05) as more extensively reported in Table 4.
ijerph-18-01728-t001_Table 1Table 1 Study population characteristics at baseline (before denosumab treatment).
Parameters Value
Age (years) 72.8 ± 7.9
Age of menopause (years) 48.7 ± 4.7
BMI (kg/m2) 24.8 ± 4.5
Habitual smokers (n, %) 21 (6.3%)
Family History of osteoporosis (n, %) 107 (32.3%)
Family History of fragility fractures (n, %) 47 (14.1%)
At least 1 vertebral fragility fracture (n, %) 237 (71.3%)
FN osteoporotic fracture (n, %) 16 (4.8%)
Osteoporotic fracture at any other site (n, %) 33 (9.9%)
Previous HRT (n, %) 32 (9.6%)
History of glucocorticoid use (n, %) 3 (0.9%)
Previous supplementation with Ca/VitD (n, %) 129 (38.8%)
Prior anti-osteoporotic treatment (n, %) 206 (62.0%)
LS T-score (SD) −2.7 ± 1.2
F T-score (SD) −2.1 ± 0.9
FN T-score (SD) −2.5 ± 0.7
Serum PTH (pg/mL) 50.6 ± 23.3
Serum calcium (mg/dL) 9.5 ± 0.6
Serum ALP (U/L) 94.5 ± 52
Serum 25(OH)vitamin D3 (ng/mL) 32.8 ± 15
ijerph-18-01728-t002_Table 2Table 2 Causes of dropouts (n = 43/428, 10.04%).
Causes (n, %)
Death 6 (14.0%)
Refusal of therapy 13 (30.2%)
Unspecified disturbances 2 (4.6%)
Fear of collateral effects 4 (9.4%)
Fear of injectable drugs 3 (6.9%)
Suggested to change therapy 1 10 (23.2%)
Followed in another center 5 (11.6%)
1 by another specialist.
ijerph-18-01728-t003_Table 3Table 3 Prior anti-osteoporotic treatment (n = 206/332, 62.04%).
Treatment (n, %)
Bisphosphonates 122 (59.2%)
Strontium ranelate or Teriparatide 43 (20.9%)
Multiple treatment 41 (19.9%)
ijerph-18-01728-t004_Table 4Table 4 BMD comparison between NG and BPG group.
Group LS T-Score F T-Score FN T-Score
Number of subjects NG (n) 29 28 30
BPG (n) 37 37 35
ANOVA results Time effect (p) 1 <0.01 <0.01 <0.01
Time × therapy (p) 2 0.91 0.07 0.05
1 Efficacy of denosumab; 2 differences in efficacy of denosumab between the groups; n, number of subjects; p, statistical probability. | Fatal | ReactionOutcome | CC BY | 33579002 | 18,955,234 | 2021-02-10 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'American trypanosomiasis'. | SARS-CoV-2 Infection and CMV Dissemination in Transplant Recipients as a Treatment for Chagas Cardiomyopathy: A Case Report.
Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 has infected over 90 million people worldwide, therefore it is considered a pandemic. SARS-CoV-2 infection can lead to severe pneumonia, acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure. Individuals receiving a heart transplantation (HT) may be at higher risk of adverse outcomes attributable to COVID-19 due to immunosuppressives, as well as concomitant infections that may also influence the prognoses. Herein, we describe the first report of two cases of HT recipients with concomitant infections by SARS-CoV-2, Trypanosoma cruzi, and cytomegalovirus (CMV) dissemination, from the first day of hospitalization due to COVID-19 in the intensive care unit (ICU) until the death of the patients.
1. Introduction
Coronavirus disease 2019 (COVID-19), an infectious respiratory disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spread on a pandemic scale since the first case was reported in Wuhan, China, in 2019 [1]. Most patients with the disease have mild-to-moderate symptoms; however, approximately 15% develop severe pneumonia, while approximately 5% develop acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure [2]. Lymphopenia is a recurrent feature in these patients, with a significant reduction in CD4+ T cells, CD8+ T cells, B cells, and natural killer (NK) cells [3], increasing the susceptibility of patients to severe illness and co-infection [4]. In fact, co-infections are associated with worsening of the clinical condition [5].
Chagas disease (CD) is a zoonosis whose etiologic agent is the protozoan Trypanosoma cruzi. It is estimated that 6–8 million people worldwide are infected [6]. CD has two distinct phases: an acute one, which is rare, with a strong production of type 1 cytokines, and a chronic phase, which develops in 30–40% of CD cases. The chronic phase of CD can be characterized by cardiomyopathy, arrhythmias, megaviscera, and, more rarely, polyneuropathy and stroke [7]. TCD4+ and TCD8+ lymphocytes are the main cells responsible for controlling parasitic infection. However, the immune response also contributes to tissue damage and pathology [8].
Chagasic cardiomyopathy represents the main cause of mortality from this disease, which can lead to heart failure, whose indication for treatment, especially in endemic countries, may include heart transplantation (HT) as a strategy to curb the evolution of this complication [6,9]. However, the immunosuppression and possible reactivation of the causative agent T. cruzi following HT require intensive clinical care and laboratory monitoring [10].
Other infections such as that caused by cytomegalovirus (CMV), a herpes virus that infects up to 60–100% of people in adulthood, are associated with transplant complications [11]. Primary CMV infection is generally asymptomatic in immunocompetent individuals, with the virus generating a latent infection. On the contrary, in immunocompromised and immunosuppressed populations, such as solid organ transplantation, hematopoietic stem cell transplantation, and HIV/AIDS patients, CMV reactivation is responsible for significant morbidity and mortality [12]. Although reactivation of infections such as CMV and CD are often described in transplant recipients, there are no reports of concomitant infection by T. cruzi, CMV, and SARS-CoV-2 in the context of HT in the literature to date.
Therefore, in this report, we investigated the progress of two patients who underwent HT at the Heart Institute of Hospital das Clínicas (Incor) and were subsequently transferred to the special intensive care unit (ICU) of the Hospital das Clínicas (Hospital das Clínicas, Faculty of Medicine, University of São Paulo-HCFMUSP) due to SARS-CoV-2 infection. These patients were diagnosed with COVID-19 by nasopharyngeal detection of SARS-CoV-2 RNA using reverse transcriptase polymerase chain reaction (RT-PCR). In addition, during hospitalization, CMV dissemination was evidenced by quantitative DNA detection in the blood. We describe herein the laboratory data from the first day of hospitalization due to COVID-19 until the death of the patients.
The baseline characteristics of these two patients, as well as their past clinical data, are summarized in Table 1.
2. Case Report
2.1. Patient 1
Female, 55 years old, presenting positive serology for CD since 2015, received HT on 2 May as a treatment for Chagas cardiomyopathy (Figure 1A) and underwent immunosuppressive therapy with methylprednisolone and azathioprine. Post-transplantation, she developed pneumonia, treated with meropenem and linezolid, and a remittent Candida tropicalis infection, treated with micafungin, with improvement. However, on 29 May, she developed dyspnea and desaturation, with a chest tomography suggestive of COVID-19. Her polymerase chain reaction (PCR) for SARS-CoV-2 was positive on 1 June.
On 3 June, she was transferred to the ICU specialized for treatment of severe SARS-CoV-2 infections. Laboratory analyses then showed that the patient had a reduced number of erythrocytes and hemoglobin level, and these reductions became more accentuated by day 33 in the ICU (Figure 2A,B). On this same day, the number of neutrophils peaked (Figure 2D). From the 4th to the 16th day in ICU, important lymphopenia developed (Figure 2F), resulting in an increase in the neutrophil-to-lymphocyte ratio in the same period (Figure 2H). Additionally, from the eighth day onward, she presented severe thrombocytopenia that persisted until death (Figure 2I).
Throughout the ICU hospitalization period, there were sustained high levels of creatinine, urea, D-dimer, C-reactive protein (CRP), and lactate dehydrogenase (Figure 2J,K,N,S,W). On the 32nd day, she presented a sharp increase in prothrombin time and activated partial thromboplastin time.
The cardiac function markers of creatine kinase myocardial band (CK-MB) and troponin remained elevated from the first days of ICU admission until death (Figure 2U,X). On the day of admission to the ICU, the N-terminal pro b-type natriuretic peptide (NT pro-BNP) was 70,000 pg/mL (reference value of <125 pg/mL) (Figure 2V). On the third day of hospitalization, NT pro-BNP (155,117 pg/mL) and troponin (0.28 ng/mL, reference value of <0.014 ng/mL) peaked, being related to the period in which she presented signs of graft rejection, which was treated with pulse methylprednisolone, thymoglobulin, and plasmapheresis. In addition, disseminated CMV infection was diagnosed (RT-PCR viral loads of 122 IU/mL on day 10 and 54 IU/mL on day 19 of hospitalization), for which she received ganciclovir.
After 47 days after diagnosis of SARS-CoV-2, the patient died (18 July) due to multiple organ dysfunctions associated with COVID-19.
2.2. Patient 2
Male, 62 years old, previously diagnosed with CD, received HT on 5 December 2019 as treatment for Chagas cardiomyopathy (Figure 1B). He underwent immunosuppressive therapy with cyclosporine, azathioprine, and prednisone. During postoperative hospitalization he presented Chagas reactivation characterized by skin biopsy and humoral graft rejection, being treated with plasmapheresis and methylprednisolone.
The patient was admitted to the ICU of the Hospital das Clínicas on 8 June, presenting cellulitis, deep vein thrombosis, and Chagas reactivation (a lower limb chagoma). The latter was treated with benznidazol. During hospitalization, he tested positive for PCR of SARS-CoV-2 on 29 June. He developed ARDS and septic and cardiogenic shock, which were the causes of his death.
Throughout the ICU hospitalization period, he maintained decreased levels of erythrocytes, hemoglobin, and lymphocytes (Figure 2A,B,F), as well as a high neutrophil/lymphocyte ratio (Figure 2H). From the 12th day, the number of platelets decreased and continued until the time of death (Figure 2I). The values of creatinine, urea, and glucose also remained high throughout the hospitalization period (Figure 2J,K,T). In addition, disseminated CMV (viral loads of 711 IU/mL detected on the 28th day and 1477 IU/mL on the 35th day of hospitalization) was diagnosed and treated with ganciclovir.
One day before death, the following laboratory parameters peaked: D-dimer (7612 ng/mL FEU, reference value of <500 ng/mL FEU), CRP (280.6 ng/mL, reference value of 0.300 ng/mL), CK-MB (8.38 ng/mL, reference value of 0.10–4.94 ng/mL), NT pro-BNP (55,393 pg/mL), lactate dehydrogenase (1161 U/L, reference value of 135–225 U/L), and troponin (0.701 ng/mL) (Figure 2N,S,U–X).
On 20 July, 36 days after admission to the ICU and 21 days after a positive COVID-19 diagnosis, the patient died.
3. Discussion
Herein, we described the first report of triple infection (SARS-CoV-2 infection, T. cruzi infection, and CMV dissemination) in HT recipients. Patients received HT as a form of treatment for Chagas cardiomyopathy.
We described the laboratory data from the first day of hospitalization in the ICU due to COVID-19 until the time of death. Both patients were admitted to a referral center for treatment for COVID-19 in the metropolitan region of São Paulo, a city in southeastern Brazil. We hypothesize that the triple infection by SARS-CoV-2 and CMV may have been an important cause of death and of the worsening in CD patients with HT. To date, there have been no similar reports of patients presenting these three concomitant infectious diseases and HT receptors.
COVID-19 infection among transplant recipients increases the potential for developing severe illness [13] and may vary between different organ transplants [14,15].
SARS-CoV-2 entry´s receptor angiotensin converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2) are expressed in different tissues, such as the lungs, heart, liver, kidneys, testicles, thyroid, and adipose tissue [16]. Some patients with COVID-19 develop severe disease characterized by respiratory distress syndrome and systemic manifestations. This condition has been associated with the dysregulated release of pro-inflammatory cytokines, termed “cytokine storm,” which may induce multi-organ failure [1,17].
The use of immunosuppressants and post-surgical opportunistic infections can also lead to damage of multisystem organs or even death [18]. There is a therapeutic paradox here, because while insufficient immunosuppression results in graft loss due to rejection, excessive immunosuppression can result in serious infection, including SARS-COV-2 [13], besides contributing to the reactivation of pathogens [19].
Long-term administration of immunosuppressants to solid organ transplant (SOT) receptors to reduce the risk of graft rejection may increase the risk of respiratory infections [20], although there is no clear clinical evidence of increased morbidity/mortality in SARS-CoV-2 infection [21].
Fernandez-Ruiz and collaborators [22] described a cohort of SOT receptors affected by COVID-19, 44% of whom were kidney transplant recipients, 33.3% liver transplant recipients, and 22.2% heart transplant recipients. The lethality rate was 27.8%, suggesting that SARS-CoV-2 infection had a severe course in SOT recipients.
Recently, we described the first patients with CD affected by SARS-CoV-2. The CD patients presented an increase in COVID-19 laboratory hallmarks and a rapid disease progression. Despite the efforts of the health staff, both patients died [3].
HT may be associated with the reactivation of pathogens, as reported in CD [10]. In a Brazilian cohort, the reactivation rate of Chagas disease after heart transplantation was reported to be 38.8% [23]. This fact made HT as a treatment for Chagas cardiomyopathy initially controversial. However, currently, especially in endemic countries, it is the most viable therapeutic option for patients with end-stage heart failure [24]. We hypothesize that, in the cases presented here, HT followed by reactivation of CD conferred an additional risk factor for the worsening of COVID-19.
Transplant recipients are also commonly affected by CMV reactivation, being associated with significant morbidity and mortality [12] that may worsen the infectious condition of COVID-19 [25]. In SOT, risk factors include the use of immunosuppressants for transplantation, advanced age, acute rejection, and other concomitant infections [11], with all these characteristics being present in both patients described herein. In the present report, we observed that immunosuppression may have contributed to the susceptibility to superinfections and more severe clinical manifestations in individuals undergoing HT.
Since overactivated immune responses can be one of the causes of organ damage, the anti-inflammatory effects of immunosuppression can be protective, reducing the cytokine storm related to complications in COVID-19 [26]. In this context, it has been described that immunosuppressive therapy with calcineurin inhibitors in patients with solid organ transplantation or systemic rheumatic diseases promotes a clinical course in SARS-CoV-2 infection, which is generally mild, and with an apparently low risk of superinfection [27]. In addition, immunosuppression has not been evaluated as a risk factor for SARS or MERS [28].
However, in the present case report, we observed that immunosuppression may have contributed to the susceptibility to SARS-CoV-2 infection, the reactivation of pathogens, and more serious clinical manifestations in individuals undergoing HT. Corroborating our findings, it was shown that patients with COVID-19 and cancer, due to their systemic immunosuppressive condition caused by malignancy and anticancer treatments, such as chemotherapy, had an increased risk of SARS-CoV-2 infection and a worse prognosis [29]. It has also been reported that chronic use of corticosteroids prior to SARS-CoV-2 contamination is associated with critical disease outcomes, including a high risk of death [30].
These contradictory observations show that knowledge about the relationship between COVID-19 and the patient’s immune condition is limited. Further studies are needed to elucidate the immune responses and prognosis of COVID-19.
In general, HT in patient 1 proved to be successful for the treatment of Chagas cardiomyopathy, with no reactivation of the pathogen. However, after SARS-CoV-2 infection, she presented graft rejection, treated with methylprednisolone, thymoglobulin, and plasmapheresis. There is a description in eye transplantation that COVID-19 infection can compromise the balance of immunoregulatory responses that allow graft survival, contributing to rejection in individuals infected with SARS-CoV-2 [31]. Thus, this change in the balance of attenuation of the immune response, such as a reduction in the number of regulatory T cells (Tregs) [32], may favor direct cardiotoxic action and multiple organ dysfunction.
During postoperative hospitalization, patient 2 presented Chagas reactivation, evidenced by skin biopsy, indicating that the CD was not completely controlled. In addition, it is important to consider that immunosuppressive therapy can contribute to the reactivation of CD [33]. After HT, the patient presented graft rejection and was treated with plasmapheresis and methylprednisolone. With this, before the SARS-CoV-2 infection, the patient was already weakened and died 21 days after the infection, in relation to patient 1 who died after 47 days of infection.
4. Conclusions
This report highlights the first case of an association between COVID-19, CD, and CMV dissemination in HT recipients. The patients had rapid disease progression to death. We believe that HT and the usage of immunosuppressive drugs, as well as immunosuppression generated by concomitant infections, may be an important risk factor for the development of severe COVID-19, especially in endemic areas with underreported CD infection.
Author Contributions
Conceptualization, S.C.G.-S., G.B., and R.W.A.; investigation, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., and S.M.; resources, M.N.S. and A.J.d.S.D.; writing—original draft preparation, S.C.G.-S., G.B., and R.W.A.; writing—review and editing, S.C.G.-S., G.B., and R.W.A.; visualization, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., S.M., A.J.d.S.D., and M.N.S.; supervision, M.N.S.; funding acquisition, A.J.d.S.D. and M.N.S. All authors have read and agreed to the published version of the manuscript.
Funding
This research was funded by Fundação de Amparo à Pesquisa (FAPESP), grant numbers 2019/22448-0, 19/02679-7, 2017/18199-9, 2018/18230-6, and 2019/26928-6. Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, CAPES:88887.503842/2020-00.
Institutional Review Board Statement
This study was approved by the local ethics committee (HCFMUSP nº 30800520.7.0000.0068-2020) and was carried out in accordance with the 2013 revision of the Declaration of Helsinki.
Informed Consent Statement
Informed consent was obtained from all of the subjects involved in the study.
Data Availability Statement
The laboratory data and imaging exams presented herein came from Hospital das Clinics, Faculty of Medicine, University of São Paulo (HCFMUSP) with prior approval for their use.
Conflicts of Interest
The authors declare no conflict of interest.
Figure 1 Chest X-ray of (a) patient 1 from May 2020 and (b) patient 2 from June 2015, taken just before heart transplantation, showing the Chagasic cardiomyopathy.
Figure 2 Daily clinical features of patients, from the first day of hospitalization in the ICU until death. Blood levels of (A) erythrocytes, (B) hemoglobin, (C) leukocytes, (D) neutrophils, (E) eosinophils, (F) lymphocytes, (G) monocytes, (H) neutrophil-to-lymphocyte ratio, (I) platelets, (J) creatinine, (K) urea, (L) prothrombin time, (M) activated partial thromboplastin time, (N) D-dimer, (O) pH, (P) pO2, (Q) pCO2, (R) oxygen peripheral, (S) CPR, (T) glucose, (U) CK-MB, (V) NT-proBNP, (W) lactate dehydrogenase, and (X) troponin T. Gray boxes represent the reference values.
tropicalmed-06-00022-t001_Table 1 Table 1 Patients’ characteristics, comorbidities, treatment, and complications.
Patient 1 Patient 2
Sex Female Male
Age 55 62
Positive SARS-CoV-2 RT-PCR date 1 June 2020 29 June 2020
Heart transplant date 2 May 2020 5 December 2019
Death date 30 days 207 days
Days between positive SARS-CoV-2 diagnosis and death 47 days 21 days
Complications during ICU stay ARDS due to COVID–19; heart transplant rejection, disseminated cytomegalovirus; aggravated chronic kidney disease and pressure ulcer ARDS due to COVID–19, disseminated cytomegalovirus and pancytopenia due to hemophagocytosis
Previous comorbidities Dilated cardiomyopathy of Chagas etiology, hypothyroidism by thyroidectomy by nodule 10 years ago Cardiomyopathy of Chagas etiology, disseminated cytomegalovirus, deep vein thrombosis, systemic arterial hypertension, diabetes, dyslipidemia, and chronic renal failure
Previous use of medications Carvedilol, losartan, furosemide, levothyroxine, isosorbide, and hydralazine Unknown
Medicines used during ICU stay Methylprednisolone, azathioprine, anti-thymocyte globulin, cyclosporine, tacrolimus, meropenem, linezolid, micafungin, vancomycin, polymyxin B, tigecycline, amikacin, fluconazole, ganciclovir, and hydrocortisone Meropenem, colistin, linezolid
fluconazole, amikacin, cyclosporine, azathioprine, and prednisone
ARD, acute respiratory distress syndrome; COVID-19, coronavirus disease 2019; ICU, intensive care unit; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. | AZATHIOPRINE, CYCLOSPORINE, METHYLPREDNISOLONE SODIUM SUCCINATE, PREDNISONE | DrugsGivenReaction | CC BY | 33579042 | 19,731,502 | 2021-02-10 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Dyspnoea'. | SARS-CoV-2 Infection and CMV Dissemination in Transplant Recipients as a Treatment for Chagas Cardiomyopathy: A Case Report.
Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 has infected over 90 million people worldwide, therefore it is considered a pandemic. SARS-CoV-2 infection can lead to severe pneumonia, acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure. Individuals receiving a heart transplantation (HT) may be at higher risk of adverse outcomes attributable to COVID-19 due to immunosuppressives, as well as concomitant infections that may also influence the prognoses. Herein, we describe the first report of two cases of HT recipients with concomitant infections by SARS-CoV-2, Trypanosoma cruzi, and cytomegalovirus (CMV) dissemination, from the first day of hospitalization due to COVID-19 in the intensive care unit (ICU) until the death of the patients.
1. Introduction
Coronavirus disease 2019 (COVID-19), an infectious respiratory disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spread on a pandemic scale since the first case was reported in Wuhan, China, in 2019 [1]. Most patients with the disease have mild-to-moderate symptoms; however, approximately 15% develop severe pneumonia, while approximately 5% develop acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure [2]. Lymphopenia is a recurrent feature in these patients, with a significant reduction in CD4+ T cells, CD8+ T cells, B cells, and natural killer (NK) cells [3], increasing the susceptibility of patients to severe illness and co-infection [4]. In fact, co-infections are associated with worsening of the clinical condition [5].
Chagas disease (CD) is a zoonosis whose etiologic agent is the protozoan Trypanosoma cruzi. It is estimated that 6–8 million people worldwide are infected [6]. CD has two distinct phases: an acute one, which is rare, with a strong production of type 1 cytokines, and a chronic phase, which develops in 30–40% of CD cases. The chronic phase of CD can be characterized by cardiomyopathy, arrhythmias, megaviscera, and, more rarely, polyneuropathy and stroke [7]. TCD4+ and TCD8+ lymphocytes are the main cells responsible for controlling parasitic infection. However, the immune response also contributes to tissue damage and pathology [8].
Chagasic cardiomyopathy represents the main cause of mortality from this disease, which can lead to heart failure, whose indication for treatment, especially in endemic countries, may include heart transplantation (HT) as a strategy to curb the evolution of this complication [6,9]. However, the immunosuppression and possible reactivation of the causative agent T. cruzi following HT require intensive clinical care and laboratory monitoring [10].
Other infections such as that caused by cytomegalovirus (CMV), a herpes virus that infects up to 60–100% of people in adulthood, are associated with transplant complications [11]. Primary CMV infection is generally asymptomatic in immunocompetent individuals, with the virus generating a latent infection. On the contrary, in immunocompromised and immunosuppressed populations, such as solid organ transplantation, hematopoietic stem cell transplantation, and HIV/AIDS patients, CMV reactivation is responsible for significant morbidity and mortality [12]. Although reactivation of infections such as CMV and CD are often described in transplant recipients, there are no reports of concomitant infection by T. cruzi, CMV, and SARS-CoV-2 in the context of HT in the literature to date.
Therefore, in this report, we investigated the progress of two patients who underwent HT at the Heart Institute of Hospital das Clínicas (Incor) and were subsequently transferred to the special intensive care unit (ICU) of the Hospital das Clínicas (Hospital das Clínicas, Faculty of Medicine, University of São Paulo-HCFMUSP) due to SARS-CoV-2 infection. These patients were diagnosed with COVID-19 by nasopharyngeal detection of SARS-CoV-2 RNA using reverse transcriptase polymerase chain reaction (RT-PCR). In addition, during hospitalization, CMV dissemination was evidenced by quantitative DNA detection in the blood. We describe herein the laboratory data from the first day of hospitalization due to COVID-19 until the death of the patients.
The baseline characteristics of these two patients, as well as their past clinical data, are summarized in Table 1.
2. Case Report
2.1. Patient 1
Female, 55 years old, presenting positive serology for CD since 2015, received HT on 2 May as a treatment for Chagas cardiomyopathy (Figure 1A) and underwent immunosuppressive therapy with methylprednisolone and azathioprine. Post-transplantation, she developed pneumonia, treated with meropenem and linezolid, and a remittent Candida tropicalis infection, treated with micafungin, with improvement. However, on 29 May, she developed dyspnea and desaturation, with a chest tomography suggestive of COVID-19. Her polymerase chain reaction (PCR) for SARS-CoV-2 was positive on 1 June.
On 3 June, she was transferred to the ICU specialized for treatment of severe SARS-CoV-2 infections. Laboratory analyses then showed that the patient had a reduced number of erythrocytes and hemoglobin level, and these reductions became more accentuated by day 33 in the ICU (Figure 2A,B). On this same day, the number of neutrophils peaked (Figure 2D). From the 4th to the 16th day in ICU, important lymphopenia developed (Figure 2F), resulting in an increase in the neutrophil-to-lymphocyte ratio in the same period (Figure 2H). Additionally, from the eighth day onward, she presented severe thrombocytopenia that persisted until death (Figure 2I).
Throughout the ICU hospitalization period, there were sustained high levels of creatinine, urea, D-dimer, C-reactive protein (CRP), and lactate dehydrogenase (Figure 2J,K,N,S,W). On the 32nd day, she presented a sharp increase in prothrombin time and activated partial thromboplastin time.
The cardiac function markers of creatine kinase myocardial band (CK-MB) and troponin remained elevated from the first days of ICU admission until death (Figure 2U,X). On the day of admission to the ICU, the N-terminal pro b-type natriuretic peptide (NT pro-BNP) was 70,000 pg/mL (reference value of <125 pg/mL) (Figure 2V). On the third day of hospitalization, NT pro-BNP (155,117 pg/mL) and troponin (0.28 ng/mL, reference value of <0.014 ng/mL) peaked, being related to the period in which she presented signs of graft rejection, which was treated with pulse methylprednisolone, thymoglobulin, and plasmapheresis. In addition, disseminated CMV infection was diagnosed (RT-PCR viral loads of 122 IU/mL on day 10 and 54 IU/mL on day 19 of hospitalization), for which she received ganciclovir.
After 47 days after diagnosis of SARS-CoV-2, the patient died (18 July) due to multiple organ dysfunctions associated with COVID-19.
2.2. Patient 2
Male, 62 years old, previously diagnosed with CD, received HT on 5 December 2019 as treatment for Chagas cardiomyopathy (Figure 1B). He underwent immunosuppressive therapy with cyclosporine, azathioprine, and prednisone. During postoperative hospitalization he presented Chagas reactivation characterized by skin biopsy and humoral graft rejection, being treated with plasmapheresis and methylprednisolone.
The patient was admitted to the ICU of the Hospital das Clínicas on 8 June, presenting cellulitis, deep vein thrombosis, and Chagas reactivation (a lower limb chagoma). The latter was treated with benznidazol. During hospitalization, he tested positive for PCR of SARS-CoV-2 on 29 June. He developed ARDS and septic and cardiogenic shock, which were the causes of his death.
Throughout the ICU hospitalization period, he maintained decreased levels of erythrocytes, hemoglobin, and lymphocytes (Figure 2A,B,F), as well as a high neutrophil/lymphocyte ratio (Figure 2H). From the 12th day, the number of platelets decreased and continued until the time of death (Figure 2I). The values of creatinine, urea, and glucose also remained high throughout the hospitalization period (Figure 2J,K,T). In addition, disseminated CMV (viral loads of 711 IU/mL detected on the 28th day and 1477 IU/mL on the 35th day of hospitalization) was diagnosed and treated with ganciclovir.
One day before death, the following laboratory parameters peaked: D-dimer (7612 ng/mL FEU, reference value of <500 ng/mL FEU), CRP (280.6 ng/mL, reference value of 0.300 ng/mL), CK-MB (8.38 ng/mL, reference value of 0.10–4.94 ng/mL), NT pro-BNP (55,393 pg/mL), lactate dehydrogenase (1161 U/L, reference value of 135–225 U/L), and troponin (0.701 ng/mL) (Figure 2N,S,U–X).
On 20 July, 36 days after admission to the ICU and 21 days after a positive COVID-19 diagnosis, the patient died.
3. Discussion
Herein, we described the first report of triple infection (SARS-CoV-2 infection, T. cruzi infection, and CMV dissemination) in HT recipients. Patients received HT as a form of treatment for Chagas cardiomyopathy.
We described the laboratory data from the first day of hospitalization in the ICU due to COVID-19 until the time of death. Both patients were admitted to a referral center for treatment for COVID-19 in the metropolitan region of São Paulo, a city in southeastern Brazil. We hypothesize that the triple infection by SARS-CoV-2 and CMV may have been an important cause of death and of the worsening in CD patients with HT. To date, there have been no similar reports of patients presenting these three concomitant infectious diseases and HT receptors.
COVID-19 infection among transplant recipients increases the potential for developing severe illness [13] and may vary between different organ transplants [14,15].
SARS-CoV-2 entry´s receptor angiotensin converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2) are expressed in different tissues, such as the lungs, heart, liver, kidneys, testicles, thyroid, and adipose tissue [16]. Some patients with COVID-19 develop severe disease characterized by respiratory distress syndrome and systemic manifestations. This condition has been associated with the dysregulated release of pro-inflammatory cytokines, termed “cytokine storm,” which may induce multi-organ failure [1,17].
The use of immunosuppressants and post-surgical opportunistic infections can also lead to damage of multisystem organs or even death [18]. There is a therapeutic paradox here, because while insufficient immunosuppression results in graft loss due to rejection, excessive immunosuppression can result in serious infection, including SARS-COV-2 [13], besides contributing to the reactivation of pathogens [19].
Long-term administration of immunosuppressants to solid organ transplant (SOT) receptors to reduce the risk of graft rejection may increase the risk of respiratory infections [20], although there is no clear clinical evidence of increased morbidity/mortality in SARS-CoV-2 infection [21].
Fernandez-Ruiz and collaborators [22] described a cohort of SOT receptors affected by COVID-19, 44% of whom were kidney transplant recipients, 33.3% liver transplant recipients, and 22.2% heart transplant recipients. The lethality rate was 27.8%, suggesting that SARS-CoV-2 infection had a severe course in SOT recipients.
Recently, we described the first patients with CD affected by SARS-CoV-2. The CD patients presented an increase in COVID-19 laboratory hallmarks and a rapid disease progression. Despite the efforts of the health staff, both patients died [3].
HT may be associated with the reactivation of pathogens, as reported in CD [10]. In a Brazilian cohort, the reactivation rate of Chagas disease after heart transplantation was reported to be 38.8% [23]. This fact made HT as a treatment for Chagas cardiomyopathy initially controversial. However, currently, especially in endemic countries, it is the most viable therapeutic option for patients with end-stage heart failure [24]. We hypothesize that, in the cases presented here, HT followed by reactivation of CD conferred an additional risk factor for the worsening of COVID-19.
Transplant recipients are also commonly affected by CMV reactivation, being associated with significant morbidity and mortality [12] that may worsen the infectious condition of COVID-19 [25]. In SOT, risk factors include the use of immunosuppressants for transplantation, advanced age, acute rejection, and other concomitant infections [11], with all these characteristics being present in both patients described herein. In the present report, we observed that immunosuppression may have contributed to the susceptibility to superinfections and more severe clinical manifestations in individuals undergoing HT.
Since overactivated immune responses can be one of the causes of organ damage, the anti-inflammatory effects of immunosuppression can be protective, reducing the cytokine storm related to complications in COVID-19 [26]. In this context, it has been described that immunosuppressive therapy with calcineurin inhibitors in patients with solid organ transplantation or systemic rheumatic diseases promotes a clinical course in SARS-CoV-2 infection, which is generally mild, and with an apparently low risk of superinfection [27]. In addition, immunosuppression has not been evaluated as a risk factor for SARS or MERS [28].
However, in the present case report, we observed that immunosuppression may have contributed to the susceptibility to SARS-CoV-2 infection, the reactivation of pathogens, and more serious clinical manifestations in individuals undergoing HT. Corroborating our findings, it was shown that patients with COVID-19 and cancer, due to their systemic immunosuppressive condition caused by malignancy and anticancer treatments, such as chemotherapy, had an increased risk of SARS-CoV-2 infection and a worse prognosis [29]. It has also been reported that chronic use of corticosteroids prior to SARS-CoV-2 contamination is associated with critical disease outcomes, including a high risk of death [30].
These contradictory observations show that knowledge about the relationship between COVID-19 and the patient’s immune condition is limited. Further studies are needed to elucidate the immune responses and prognosis of COVID-19.
In general, HT in patient 1 proved to be successful for the treatment of Chagas cardiomyopathy, with no reactivation of the pathogen. However, after SARS-CoV-2 infection, she presented graft rejection, treated with methylprednisolone, thymoglobulin, and plasmapheresis. There is a description in eye transplantation that COVID-19 infection can compromise the balance of immunoregulatory responses that allow graft survival, contributing to rejection in individuals infected with SARS-CoV-2 [31]. Thus, this change in the balance of attenuation of the immune response, such as a reduction in the number of regulatory T cells (Tregs) [32], may favor direct cardiotoxic action and multiple organ dysfunction.
During postoperative hospitalization, patient 2 presented Chagas reactivation, evidenced by skin biopsy, indicating that the CD was not completely controlled. In addition, it is important to consider that immunosuppressive therapy can contribute to the reactivation of CD [33]. After HT, the patient presented graft rejection and was treated with plasmapheresis and methylprednisolone. With this, before the SARS-CoV-2 infection, the patient was already weakened and died 21 days after the infection, in relation to patient 1 who died after 47 days of infection.
4. Conclusions
This report highlights the first case of an association between COVID-19, CD, and CMV dissemination in HT recipients. The patients had rapid disease progression to death. We believe that HT and the usage of immunosuppressive drugs, as well as immunosuppression generated by concomitant infections, may be an important risk factor for the development of severe COVID-19, especially in endemic areas with underreported CD infection.
Author Contributions
Conceptualization, S.C.G.-S., G.B., and R.W.A.; investigation, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., and S.M.; resources, M.N.S. and A.J.d.S.D.; writing—original draft preparation, S.C.G.-S., G.B., and R.W.A.; writing—review and editing, S.C.G.-S., G.B., and R.W.A.; visualization, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., S.M., A.J.d.S.D., and M.N.S.; supervision, M.N.S.; funding acquisition, A.J.d.S.D. and M.N.S. All authors have read and agreed to the published version of the manuscript.
Funding
This research was funded by Fundação de Amparo à Pesquisa (FAPESP), grant numbers 2019/22448-0, 19/02679-7, 2017/18199-9, 2018/18230-6, and 2019/26928-6. Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, CAPES:88887.503842/2020-00.
Institutional Review Board Statement
This study was approved by the local ethics committee (HCFMUSP nº 30800520.7.0000.0068-2020) and was carried out in accordance with the 2013 revision of the Declaration of Helsinki.
Informed Consent Statement
Informed consent was obtained from all of the subjects involved in the study.
Data Availability Statement
The laboratory data and imaging exams presented herein came from Hospital das Clinics, Faculty of Medicine, University of São Paulo (HCFMUSP) with prior approval for their use.
Conflicts of Interest
The authors declare no conflict of interest.
Figure 1 Chest X-ray of (a) patient 1 from May 2020 and (b) patient 2 from June 2015, taken just before heart transplantation, showing the Chagasic cardiomyopathy.
Figure 2 Daily clinical features of patients, from the first day of hospitalization in the ICU until death. Blood levels of (A) erythrocytes, (B) hemoglobin, (C) leukocytes, (D) neutrophils, (E) eosinophils, (F) lymphocytes, (G) monocytes, (H) neutrophil-to-lymphocyte ratio, (I) platelets, (J) creatinine, (K) urea, (L) prothrombin time, (M) activated partial thromboplastin time, (N) D-dimer, (O) pH, (P) pO2, (Q) pCO2, (R) oxygen peripheral, (S) CPR, (T) glucose, (U) CK-MB, (V) NT-proBNP, (W) lactate dehydrogenase, and (X) troponin T. Gray boxes represent the reference values.
tropicalmed-06-00022-t001_Table 1 Table 1 Patients’ characteristics, comorbidities, treatment, and complications.
Patient 1 Patient 2
Sex Female Male
Age 55 62
Positive SARS-CoV-2 RT-PCR date 1 June 2020 29 June 2020
Heart transplant date 2 May 2020 5 December 2019
Death date 30 days 207 days
Days between positive SARS-CoV-2 diagnosis and death 47 days 21 days
Complications during ICU stay ARDS due to COVID–19; heart transplant rejection, disseminated cytomegalovirus; aggravated chronic kidney disease and pressure ulcer ARDS due to COVID–19, disseminated cytomegalovirus and pancytopenia due to hemophagocytosis
Previous comorbidities Dilated cardiomyopathy of Chagas etiology, hypothyroidism by thyroidectomy by nodule 10 years ago Cardiomyopathy of Chagas etiology, disseminated cytomegalovirus, deep vein thrombosis, systemic arterial hypertension, diabetes, dyslipidemia, and chronic renal failure
Previous use of medications Carvedilol, losartan, furosemide, levothyroxine, isosorbide, and hydralazine Unknown
Medicines used during ICU stay Methylprednisolone, azathioprine, anti-thymocyte globulin, cyclosporine, tacrolimus, meropenem, linezolid, micafungin, vancomycin, polymyxin B, tigecycline, amikacin, fluconazole, ganciclovir, and hydrocortisone Meropenem, colistin, linezolid
fluconazole, amikacin, cyclosporine, azathioprine, and prednisone
ARD, acute respiratory distress syndrome; COVID-19, coronavirus disease 2019; ICU, intensive care unit; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. | AZATHIOPRINE, CARVEDILOL, CYCLOSPORINE, FUROSEMIDE, HYDRALAZINE HYDROCHLORIDE, HYDROCORTISONE, ISOSORBIDE, LAPINE T-LYMPHOCYTE IMMUNE GLOBULIN, LEVOTHYROXINE, LOSARTAN, METHYLPREDNISOLONE, TACROLIMUS | DrugsGivenReaction | CC BY | 33579042 | 19,681,859 | 2021-02-10 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Haemophagocytic lymphohistiocytosis'. | SARS-CoV-2 Infection and CMV Dissemination in Transplant Recipients as a Treatment for Chagas Cardiomyopathy: A Case Report.
Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 has infected over 90 million people worldwide, therefore it is considered a pandemic. SARS-CoV-2 infection can lead to severe pneumonia, acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure. Individuals receiving a heart transplantation (HT) may be at higher risk of adverse outcomes attributable to COVID-19 due to immunosuppressives, as well as concomitant infections that may also influence the prognoses. Herein, we describe the first report of two cases of HT recipients with concomitant infections by SARS-CoV-2, Trypanosoma cruzi, and cytomegalovirus (CMV) dissemination, from the first day of hospitalization due to COVID-19 in the intensive care unit (ICU) until the death of the patients.
1. Introduction
Coronavirus disease 2019 (COVID-19), an infectious respiratory disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spread on a pandemic scale since the first case was reported in Wuhan, China, in 2019 [1]. Most patients with the disease have mild-to-moderate symptoms; however, approximately 15% develop severe pneumonia, while approximately 5% develop acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure [2]. Lymphopenia is a recurrent feature in these patients, with a significant reduction in CD4+ T cells, CD8+ T cells, B cells, and natural killer (NK) cells [3], increasing the susceptibility of patients to severe illness and co-infection [4]. In fact, co-infections are associated with worsening of the clinical condition [5].
Chagas disease (CD) is a zoonosis whose etiologic agent is the protozoan Trypanosoma cruzi. It is estimated that 6–8 million people worldwide are infected [6]. CD has two distinct phases: an acute one, which is rare, with a strong production of type 1 cytokines, and a chronic phase, which develops in 30–40% of CD cases. The chronic phase of CD can be characterized by cardiomyopathy, arrhythmias, megaviscera, and, more rarely, polyneuropathy and stroke [7]. TCD4+ and TCD8+ lymphocytes are the main cells responsible for controlling parasitic infection. However, the immune response also contributes to tissue damage and pathology [8].
Chagasic cardiomyopathy represents the main cause of mortality from this disease, which can lead to heart failure, whose indication for treatment, especially in endemic countries, may include heart transplantation (HT) as a strategy to curb the evolution of this complication [6,9]. However, the immunosuppression and possible reactivation of the causative agent T. cruzi following HT require intensive clinical care and laboratory monitoring [10].
Other infections such as that caused by cytomegalovirus (CMV), a herpes virus that infects up to 60–100% of people in adulthood, are associated with transplant complications [11]. Primary CMV infection is generally asymptomatic in immunocompetent individuals, with the virus generating a latent infection. On the contrary, in immunocompromised and immunosuppressed populations, such as solid organ transplantation, hematopoietic stem cell transplantation, and HIV/AIDS patients, CMV reactivation is responsible for significant morbidity and mortality [12]. Although reactivation of infections such as CMV and CD are often described in transplant recipients, there are no reports of concomitant infection by T. cruzi, CMV, and SARS-CoV-2 in the context of HT in the literature to date.
Therefore, in this report, we investigated the progress of two patients who underwent HT at the Heart Institute of Hospital das Clínicas (Incor) and were subsequently transferred to the special intensive care unit (ICU) of the Hospital das Clínicas (Hospital das Clínicas, Faculty of Medicine, University of São Paulo-HCFMUSP) due to SARS-CoV-2 infection. These patients were diagnosed with COVID-19 by nasopharyngeal detection of SARS-CoV-2 RNA using reverse transcriptase polymerase chain reaction (RT-PCR). In addition, during hospitalization, CMV dissemination was evidenced by quantitative DNA detection in the blood. We describe herein the laboratory data from the first day of hospitalization due to COVID-19 until the death of the patients.
The baseline characteristics of these two patients, as well as their past clinical data, are summarized in Table 1.
2. Case Report
2.1. Patient 1
Female, 55 years old, presenting positive serology for CD since 2015, received HT on 2 May as a treatment for Chagas cardiomyopathy (Figure 1A) and underwent immunosuppressive therapy with methylprednisolone and azathioprine. Post-transplantation, she developed pneumonia, treated with meropenem and linezolid, and a remittent Candida tropicalis infection, treated with micafungin, with improvement. However, on 29 May, she developed dyspnea and desaturation, with a chest tomography suggestive of COVID-19. Her polymerase chain reaction (PCR) for SARS-CoV-2 was positive on 1 June.
On 3 June, she was transferred to the ICU specialized for treatment of severe SARS-CoV-2 infections. Laboratory analyses then showed that the patient had a reduced number of erythrocytes and hemoglobin level, and these reductions became more accentuated by day 33 in the ICU (Figure 2A,B). On this same day, the number of neutrophils peaked (Figure 2D). From the 4th to the 16th day in ICU, important lymphopenia developed (Figure 2F), resulting in an increase in the neutrophil-to-lymphocyte ratio in the same period (Figure 2H). Additionally, from the eighth day onward, she presented severe thrombocytopenia that persisted until death (Figure 2I).
Throughout the ICU hospitalization period, there were sustained high levels of creatinine, urea, D-dimer, C-reactive protein (CRP), and lactate dehydrogenase (Figure 2J,K,N,S,W). On the 32nd day, she presented a sharp increase in prothrombin time and activated partial thromboplastin time.
The cardiac function markers of creatine kinase myocardial band (CK-MB) and troponin remained elevated from the first days of ICU admission until death (Figure 2U,X). On the day of admission to the ICU, the N-terminal pro b-type natriuretic peptide (NT pro-BNP) was 70,000 pg/mL (reference value of <125 pg/mL) (Figure 2V). On the third day of hospitalization, NT pro-BNP (155,117 pg/mL) and troponin (0.28 ng/mL, reference value of <0.014 ng/mL) peaked, being related to the period in which she presented signs of graft rejection, which was treated with pulse methylprednisolone, thymoglobulin, and plasmapheresis. In addition, disseminated CMV infection was diagnosed (RT-PCR viral loads of 122 IU/mL on day 10 and 54 IU/mL on day 19 of hospitalization), for which she received ganciclovir.
After 47 days after diagnosis of SARS-CoV-2, the patient died (18 July) due to multiple organ dysfunctions associated with COVID-19.
2.2. Patient 2
Male, 62 years old, previously diagnosed with CD, received HT on 5 December 2019 as treatment for Chagas cardiomyopathy (Figure 1B). He underwent immunosuppressive therapy with cyclosporine, azathioprine, and prednisone. During postoperative hospitalization he presented Chagas reactivation characterized by skin biopsy and humoral graft rejection, being treated with plasmapheresis and methylprednisolone.
The patient was admitted to the ICU of the Hospital das Clínicas on 8 June, presenting cellulitis, deep vein thrombosis, and Chagas reactivation (a lower limb chagoma). The latter was treated with benznidazol. During hospitalization, he tested positive for PCR of SARS-CoV-2 on 29 June. He developed ARDS and septic and cardiogenic shock, which were the causes of his death.
Throughout the ICU hospitalization period, he maintained decreased levels of erythrocytes, hemoglobin, and lymphocytes (Figure 2A,B,F), as well as a high neutrophil/lymphocyte ratio (Figure 2H). From the 12th day, the number of platelets decreased and continued until the time of death (Figure 2I). The values of creatinine, urea, and glucose also remained high throughout the hospitalization period (Figure 2J,K,T). In addition, disseminated CMV (viral loads of 711 IU/mL detected on the 28th day and 1477 IU/mL on the 35th day of hospitalization) was diagnosed and treated with ganciclovir.
One day before death, the following laboratory parameters peaked: D-dimer (7612 ng/mL FEU, reference value of <500 ng/mL FEU), CRP (280.6 ng/mL, reference value of 0.300 ng/mL), CK-MB (8.38 ng/mL, reference value of 0.10–4.94 ng/mL), NT pro-BNP (55,393 pg/mL), lactate dehydrogenase (1161 U/L, reference value of 135–225 U/L), and troponin (0.701 ng/mL) (Figure 2N,S,U–X).
On 20 July, 36 days after admission to the ICU and 21 days after a positive COVID-19 diagnosis, the patient died.
3. Discussion
Herein, we described the first report of triple infection (SARS-CoV-2 infection, T. cruzi infection, and CMV dissemination) in HT recipients. Patients received HT as a form of treatment for Chagas cardiomyopathy.
We described the laboratory data from the first day of hospitalization in the ICU due to COVID-19 until the time of death. Both patients were admitted to a referral center for treatment for COVID-19 in the metropolitan region of São Paulo, a city in southeastern Brazil. We hypothesize that the triple infection by SARS-CoV-2 and CMV may have been an important cause of death and of the worsening in CD patients with HT. To date, there have been no similar reports of patients presenting these three concomitant infectious diseases and HT receptors.
COVID-19 infection among transplant recipients increases the potential for developing severe illness [13] and may vary between different organ transplants [14,15].
SARS-CoV-2 entry´s receptor angiotensin converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2) are expressed in different tissues, such as the lungs, heart, liver, kidneys, testicles, thyroid, and adipose tissue [16]. Some patients with COVID-19 develop severe disease characterized by respiratory distress syndrome and systemic manifestations. This condition has been associated with the dysregulated release of pro-inflammatory cytokines, termed “cytokine storm,” which may induce multi-organ failure [1,17].
The use of immunosuppressants and post-surgical opportunistic infections can also lead to damage of multisystem organs or even death [18]. There is a therapeutic paradox here, because while insufficient immunosuppression results in graft loss due to rejection, excessive immunosuppression can result in serious infection, including SARS-COV-2 [13], besides contributing to the reactivation of pathogens [19].
Long-term administration of immunosuppressants to solid organ transplant (SOT) receptors to reduce the risk of graft rejection may increase the risk of respiratory infections [20], although there is no clear clinical evidence of increased morbidity/mortality in SARS-CoV-2 infection [21].
Fernandez-Ruiz and collaborators [22] described a cohort of SOT receptors affected by COVID-19, 44% of whom were kidney transplant recipients, 33.3% liver transplant recipients, and 22.2% heart transplant recipients. The lethality rate was 27.8%, suggesting that SARS-CoV-2 infection had a severe course in SOT recipients.
Recently, we described the first patients with CD affected by SARS-CoV-2. The CD patients presented an increase in COVID-19 laboratory hallmarks and a rapid disease progression. Despite the efforts of the health staff, both patients died [3].
HT may be associated with the reactivation of pathogens, as reported in CD [10]. In a Brazilian cohort, the reactivation rate of Chagas disease after heart transplantation was reported to be 38.8% [23]. This fact made HT as a treatment for Chagas cardiomyopathy initially controversial. However, currently, especially in endemic countries, it is the most viable therapeutic option for patients with end-stage heart failure [24]. We hypothesize that, in the cases presented here, HT followed by reactivation of CD conferred an additional risk factor for the worsening of COVID-19.
Transplant recipients are also commonly affected by CMV reactivation, being associated with significant morbidity and mortality [12] that may worsen the infectious condition of COVID-19 [25]. In SOT, risk factors include the use of immunosuppressants for transplantation, advanced age, acute rejection, and other concomitant infections [11], with all these characteristics being present in both patients described herein. In the present report, we observed that immunosuppression may have contributed to the susceptibility to superinfections and more severe clinical manifestations in individuals undergoing HT.
Since overactivated immune responses can be one of the causes of organ damage, the anti-inflammatory effects of immunosuppression can be protective, reducing the cytokine storm related to complications in COVID-19 [26]. In this context, it has been described that immunosuppressive therapy with calcineurin inhibitors in patients with solid organ transplantation or systemic rheumatic diseases promotes a clinical course in SARS-CoV-2 infection, which is generally mild, and with an apparently low risk of superinfection [27]. In addition, immunosuppression has not been evaluated as a risk factor for SARS or MERS [28].
However, in the present case report, we observed that immunosuppression may have contributed to the susceptibility to SARS-CoV-2 infection, the reactivation of pathogens, and more serious clinical manifestations in individuals undergoing HT. Corroborating our findings, it was shown that patients with COVID-19 and cancer, due to their systemic immunosuppressive condition caused by malignancy and anticancer treatments, such as chemotherapy, had an increased risk of SARS-CoV-2 infection and a worse prognosis [29]. It has also been reported that chronic use of corticosteroids prior to SARS-CoV-2 contamination is associated with critical disease outcomes, including a high risk of death [30].
These contradictory observations show that knowledge about the relationship between COVID-19 and the patient’s immune condition is limited. Further studies are needed to elucidate the immune responses and prognosis of COVID-19.
In general, HT in patient 1 proved to be successful for the treatment of Chagas cardiomyopathy, with no reactivation of the pathogen. However, after SARS-CoV-2 infection, she presented graft rejection, treated with methylprednisolone, thymoglobulin, and plasmapheresis. There is a description in eye transplantation that COVID-19 infection can compromise the balance of immunoregulatory responses that allow graft survival, contributing to rejection in individuals infected with SARS-CoV-2 [31]. Thus, this change in the balance of attenuation of the immune response, such as a reduction in the number of regulatory T cells (Tregs) [32], may favor direct cardiotoxic action and multiple organ dysfunction.
During postoperative hospitalization, patient 2 presented Chagas reactivation, evidenced by skin biopsy, indicating that the CD was not completely controlled. In addition, it is important to consider that immunosuppressive therapy can contribute to the reactivation of CD [33]. After HT, the patient presented graft rejection and was treated with plasmapheresis and methylprednisolone. With this, before the SARS-CoV-2 infection, the patient was already weakened and died 21 days after the infection, in relation to patient 1 who died after 47 days of infection.
4. Conclusions
This report highlights the first case of an association between COVID-19, CD, and CMV dissemination in HT recipients. The patients had rapid disease progression to death. We believe that HT and the usage of immunosuppressive drugs, as well as immunosuppression generated by concomitant infections, may be an important risk factor for the development of severe COVID-19, especially in endemic areas with underreported CD infection.
Author Contributions
Conceptualization, S.C.G.-S., G.B., and R.W.A.; investigation, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., and S.M.; resources, M.N.S. and A.J.d.S.D.; writing—original draft preparation, S.C.G.-S., G.B., and R.W.A.; writing—review and editing, S.C.G.-S., G.B., and R.W.A.; visualization, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., S.M., A.J.d.S.D., and M.N.S.; supervision, M.N.S.; funding acquisition, A.J.d.S.D. and M.N.S. All authors have read and agreed to the published version of the manuscript.
Funding
This research was funded by Fundação de Amparo à Pesquisa (FAPESP), grant numbers 2019/22448-0, 19/02679-7, 2017/18199-9, 2018/18230-6, and 2019/26928-6. Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, CAPES:88887.503842/2020-00.
Institutional Review Board Statement
This study was approved by the local ethics committee (HCFMUSP nº 30800520.7.0000.0068-2020) and was carried out in accordance with the 2013 revision of the Declaration of Helsinki.
Informed Consent Statement
Informed consent was obtained from all of the subjects involved in the study.
Data Availability Statement
The laboratory data and imaging exams presented herein came from Hospital das Clinics, Faculty of Medicine, University of São Paulo (HCFMUSP) with prior approval for their use.
Conflicts of Interest
The authors declare no conflict of interest.
Figure 1 Chest X-ray of (a) patient 1 from May 2020 and (b) patient 2 from June 2015, taken just before heart transplantation, showing the Chagasic cardiomyopathy.
Figure 2 Daily clinical features of patients, from the first day of hospitalization in the ICU until death. Blood levels of (A) erythrocytes, (B) hemoglobin, (C) leukocytes, (D) neutrophils, (E) eosinophils, (F) lymphocytes, (G) monocytes, (H) neutrophil-to-lymphocyte ratio, (I) platelets, (J) creatinine, (K) urea, (L) prothrombin time, (M) activated partial thromboplastin time, (N) D-dimer, (O) pH, (P) pO2, (Q) pCO2, (R) oxygen peripheral, (S) CPR, (T) glucose, (U) CK-MB, (V) NT-proBNP, (W) lactate dehydrogenase, and (X) troponin T. Gray boxes represent the reference values.
tropicalmed-06-00022-t001_Table 1 Table 1 Patients’ characteristics, comorbidities, treatment, and complications.
Patient 1 Patient 2
Sex Female Male
Age 55 62
Positive SARS-CoV-2 RT-PCR date 1 June 2020 29 June 2020
Heart transplant date 2 May 2020 5 December 2019
Death date 30 days 207 days
Days between positive SARS-CoV-2 diagnosis and death 47 days 21 days
Complications during ICU stay ARDS due to COVID–19; heart transplant rejection, disseminated cytomegalovirus; aggravated chronic kidney disease and pressure ulcer ARDS due to COVID–19, disseminated cytomegalovirus and pancytopenia due to hemophagocytosis
Previous comorbidities Dilated cardiomyopathy of Chagas etiology, hypothyroidism by thyroidectomy by nodule 10 years ago Cardiomyopathy of Chagas etiology, disseminated cytomegalovirus, deep vein thrombosis, systemic arterial hypertension, diabetes, dyslipidemia, and chronic renal failure
Previous use of medications Carvedilol, losartan, furosemide, levothyroxine, isosorbide, and hydralazine Unknown
Medicines used during ICU stay Methylprednisolone, azathioprine, anti-thymocyte globulin, cyclosporine, tacrolimus, meropenem, linezolid, micafungin, vancomycin, polymyxin B, tigecycline, amikacin, fluconazole, ganciclovir, and hydrocortisone Meropenem, colistin, linezolid
fluconazole, amikacin, cyclosporine, azathioprine, and prednisone
ARD, acute respiratory distress syndrome; COVID-19, coronavirus disease 2019; ICU, intensive care unit; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. | AZATHIOPRINE, CYCLOSPORINE, METHYLPREDNISOLONE SODIUM SUCCINATE, PREDNISONE | DrugsGivenReaction | CC BY | 33579042 | 19,731,502 | 2021-02-10 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Infection reactivation'. | SARS-CoV-2 Infection and CMV Dissemination in Transplant Recipients as a Treatment for Chagas Cardiomyopathy: A Case Report.
Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 has infected over 90 million people worldwide, therefore it is considered a pandemic. SARS-CoV-2 infection can lead to severe pneumonia, acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure. Individuals receiving a heart transplantation (HT) may be at higher risk of adverse outcomes attributable to COVID-19 due to immunosuppressives, as well as concomitant infections that may also influence the prognoses. Herein, we describe the first report of two cases of HT recipients with concomitant infections by SARS-CoV-2, Trypanosoma cruzi, and cytomegalovirus (CMV) dissemination, from the first day of hospitalization due to COVID-19 in the intensive care unit (ICU) until the death of the patients.
1. Introduction
Coronavirus disease 2019 (COVID-19), an infectious respiratory disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spread on a pandemic scale since the first case was reported in Wuhan, China, in 2019 [1]. Most patients with the disease have mild-to-moderate symptoms; however, approximately 15% develop severe pneumonia, while approximately 5% develop acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure [2]. Lymphopenia is a recurrent feature in these patients, with a significant reduction in CD4+ T cells, CD8+ T cells, B cells, and natural killer (NK) cells [3], increasing the susceptibility of patients to severe illness and co-infection [4]. In fact, co-infections are associated with worsening of the clinical condition [5].
Chagas disease (CD) is a zoonosis whose etiologic agent is the protozoan Trypanosoma cruzi. It is estimated that 6–8 million people worldwide are infected [6]. CD has two distinct phases: an acute one, which is rare, with a strong production of type 1 cytokines, and a chronic phase, which develops in 30–40% of CD cases. The chronic phase of CD can be characterized by cardiomyopathy, arrhythmias, megaviscera, and, more rarely, polyneuropathy and stroke [7]. TCD4+ and TCD8+ lymphocytes are the main cells responsible for controlling parasitic infection. However, the immune response also contributes to tissue damage and pathology [8].
Chagasic cardiomyopathy represents the main cause of mortality from this disease, which can lead to heart failure, whose indication for treatment, especially in endemic countries, may include heart transplantation (HT) as a strategy to curb the evolution of this complication [6,9]. However, the immunosuppression and possible reactivation of the causative agent T. cruzi following HT require intensive clinical care and laboratory monitoring [10].
Other infections such as that caused by cytomegalovirus (CMV), a herpes virus that infects up to 60–100% of people in adulthood, are associated with transplant complications [11]. Primary CMV infection is generally asymptomatic in immunocompetent individuals, with the virus generating a latent infection. On the contrary, in immunocompromised and immunosuppressed populations, such as solid organ transplantation, hematopoietic stem cell transplantation, and HIV/AIDS patients, CMV reactivation is responsible for significant morbidity and mortality [12]. Although reactivation of infections such as CMV and CD are often described in transplant recipients, there are no reports of concomitant infection by T. cruzi, CMV, and SARS-CoV-2 in the context of HT in the literature to date.
Therefore, in this report, we investigated the progress of two patients who underwent HT at the Heart Institute of Hospital das Clínicas (Incor) and were subsequently transferred to the special intensive care unit (ICU) of the Hospital das Clínicas (Hospital das Clínicas, Faculty of Medicine, University of São Paulo-HCFMUSP) due to SARS-CoV-2 infection. These patients were diagnosed with COVID-19 by nasopharyngeal detection of SARS-CoV-2 RNA using reverse transcriptase polymerase chain reaction (RT-PCR). In addition, during hospitalization, CMV dissemination was evidenced by quantitative DNA detection in the blood. We describe herein the laboratory data from the first day of hospitalization due to COVID-19 until the death of the patients.
The baseline characteristics of these two patients, as well as their past clinical data, are summarized in Table 1.
2. Case Report
2.1. Patient 1
Female, 55 years old, presenting positive serology for CD since 2015, received HT on 2 May as a treatment for Chagas cardiomyopathy (Figure 1A) and underwent immunosuppressive therapy with methylprednisolone and azathioprine. Post-transplantation, she developed pneumonia, treated with meropenem and linezolid, and a remittent Candida tropicalis infection, treated with micafungin, with improvement. However, on 29 May, she developed dyspnea and desaturation, with a chest tomography suggestive of COVID-19. Her polymerase chain reaction (PCR) for SARS-CoV-2 was positive on 1 June.
On 3 June, she was transferred to the ICU specialized for treatment of severe SARS-CoV-2 infections. Laboratory analyses then showed that the patient had a reduced number of erythrocytes and hemoglobin level, and these reductions became more accentuated by day 33 in the ICU (Figure 2A,B). On this same day, the number of neutrophils peaked (Figure 2D). From the 4th to the 16th day in ICU, important lymphopenia developed (Figure 2F), resulting in an increase in the neutrophil-to-lymphocyte ratio in the same period (Figure 2H). Additionally, from the eighth day onward, she presented severe thrombocytopenia that persisted until death (Figure 2I).
Throughout the ICU hospitalization period, there were sustained high levels of creatinine, urea, D-dimer, C-reactive protein (CRP), and lactate dehydrogenase (Figure 2J,K,N,S,W). On the 32nd day, she presented a sharp increase in prothrombin time and activated partial thromboplastin time.
The cardiac function markers of creatine kinase myocardial band (CK-MB) and troponin remained elevated from the first days of ICU admission until death (Figure 2U,X). On the day of admission to the ICU, the N-terminal pro b-type natriuretic peptide (NT pro-BNP) was 70,000 pg/mL (reference value of <125 pg/mL) (Figure 2V). On the third day of hospitalization, NT pro-BNP (155,117 pg/mL) and troponin (0.28 ng/mL, reference value of <0.014 ng/mL) peaked, being related to the period in which she presented signs of graft rejection, which was treated with pulse methylprednisolone, thymoglobulin, and plasmapheresis. In addition, disseminated CMV infection was diagnosed (RT-PCR viral loads of 122 IU/mL on day 10 and 54 IU/mL on day 19 of hospitalization), for which she received ganciclovir.
After 47 days after diagnosis of SARS-CoV-2, the patient died (18 July) due to multiple organ dysfunctions associated with COVID-19.
2.2. Patient 2
Male, 62 years old, previously diagnosed with CD, received HT on 5 December 2019 as treatment for Chagas cardiomyopathy (Figure 1B). He underwent immunosuppressive therapy with cyclosporine, azathioprine, and prednisone. During postoperative hospitalization he presented Chagas reactivation characterized by skin biopsy and humoral graft rejection, being treated with plasmapheresis and methylprednisolone.
The patient was admitted to the ICU of the Hospital das Clínicas on 8 June, presenting cellulitis, deep vein thrombosis, and Chagas reactivation (a lower limb chagoma). The latter was treated with benznidazol. During hospitalization, he tested positive for PCR of SARS-CoV-2 on 29 June. He developed ARDS and septic and cardiogenic shock, which were the causes of his death.
Throughout the ICU hospitalization period, he maintained decreased levels of erythrocytes, hemoglobin, and lymphocytes (Figure 2A,B,F), as well as a high neutrophil/lymphocyte ratio (Figure 2H). From the 12th day, the number of platelets decreased and continued until the time of death (Figure 2I). The values of creatinine, urea, and glucose also remained high throughout the hospitalization period (Figure 2J,K,T). In addition, disseminated CMV (viral loads of 711 IU/mL detected on the 28th day and 1477 IU/mL on the 35th day of hospitalization) was diagnosed and treated with ganciclovir.
One day before death, the following laboratory parameters peaked: D-dimer (7612 ng/mL FEU, reference value of <500 ng/mL FEU), CRP (280.6 ng/mL, reference value of 0.300 ng/mL), CK-MB (8.38 ng/mL, reference value of 0.10–4.94 ng/mL), NT pro-BNP (55,393 pg/mL), lactate dehydrogenase (1161 U/L, reference value of 135–225 U/L), and troponin (0.701 ng/mL) (Figure 2N,S,U–X).
On 20 July, 36 days after admission to the ICU and 21 days after a positive COVID-19 diagnosis, the patient died.
3. Discussion
Herein, we described the first report of triple infection (SARS-CoV-2 infection, T. cruzi infection, and CMV dissemination) in HT recipients. Patients received HT as a form of treatment for Chagas cardiomyopathy.
We described the laboratory data from the first day of hospitalization in the ICU due to COVID-19 until the time of death. Both patients were admitted to a referral center for treatment for COVID-19 in the metropolitan region of São Paulo, a city in southeastern Brazil. We hypothesize that the triple infection by SARS-CoV-2 and CMV may have been an important cause of death and of the worsening in CD patients with HT. To date, there have been no similar reports of patients presenting these three concomitant infectious diseases and HT receptors.
COVID-19 infection among transplant recipients increases the potential for developing severe illness [13] and may vary between different organ transplants [14,15].
SARS-CoV-2 entry´s receptor angiotensin converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2) are expressed in different tissues, such as the lungs, heart, liver, kidneys, testicles, thyroid, and adipose tissue [16]. Some patients with COVID-19 develop severe disease characterized by respiratory distress syndrome and systemic manifestations. This condition has been associated with the dysregulated release of pro-inflammatory cytokines, termed “cytokine storm,” which may induce multi-organ failure [1,17].
The use of immunosuppressants and post-surgical opportunistic infections can also lead to damage of multisystem organs or even death [18]. There is a therapeutic paradox here, because while insufficient immunosuppression results in graft loss due to rejection, excessive immunosuppression can result in serious infection, including SARS-COV-2 [13], besides contributing to the reactivation of pathogens [19].
Long-term administration of immunosuppressants to solid organ transplant (SOT) receptors to reduce the risk of graft rejection may increase the risk of respiratory infections [20], although there is no clear clinical evidence of increased morbidity/mortality in SARS-CoV-2 infection [21].
Fernandez-Ruiz and collaborators [22] described a cohort of SOT receptors affected by COVID-19, 44% of whom were kidney transplant recipients, 33.3% liver transplant recipients, and 22.2% heart transplant recipients. The lethality rate was 27.8%, suggesting that SARS-CoV-2 infection had a severe course in SOT recipients.
Recently, we described the first patients with CD affected by SARS-CoV-2. The CD patients presented an increase in COVID-19 laboratory hallmarks and a rapid disease progression. Despite the efforts of the health staff, both patients died [3].
HT may be associated with the reactivation of pathogens, as reported in CD [10]. In a Brazilian cohort, the reactivation rate of Chagas disease after heart transplantation was reported to be 38.8% [23]. This fact made HT as a treatment for Chagas cardiomyopathy initially controversial. However, currently, especially in endemic countries, it is the most viable therapeutic option for patients with end-stage heart failure [24]. We hypothesize that, in the cases presented here, HT followed by reactivation of CD conferred an additional risk factor for the worsening of COVID-19.
Transplant recipients are also commonly affected by CMV reactivation, being associated with significant morbidity and mortality [12] that may worsen the infectious condition of COVID-19 [25]. In SOT, risk factors include the use of immunosuppressants for transplantation, advanced age, acute rejection, and other concomitant infections [11], with all these characteristics being present in both patients described herein. In the present report, we observed that immunosuppression may have contributed to the susceptibility to superinfections and more severe clinical manifestations in individuals undergoing HT.
Since overactivated immune responses can be one of the causes of organ damage, the anti-inflammatory effects of immunosuppression can be protective, reducing the cytokine storm related to complications in COVID-19 [26]. In this context, it has been described that immunosuppressive therapy with calcineurin inhibitors in patients with solid organ transplantation or systemic rheumatic diseases promotes a clinical course in SARS-CoV-2 infection, which is generally mild, and with an apparently low risk of superinfection [27]. In addition, immunosuppression has not been evaluated as a risk factor for SARS or MERS [28].
However, in the present case report, we observed that immunosuppression may have contributed to the susceptibility to SARS-CoV-2 infection, the reactivation of pathogens, and more serious clinical manifestations in individuals undergoing HT. Corroborating our findings, it was shown that patients with COVID-19 and cancer, due to their systemic immunosuppressive condition caused by malignancy and anticancer treatments, such as chemotherapy, had an increased risk of SARS-CoV-2 infection and a worse prognosis [29]. It has also been reported that chronic use of corticosteroids prior to SARS-CoV-2 contamination is associated with critical disease outcomes, including a high risk of death [30].
These contradictory observations show that knowledge about the relationship between COVID-19 and the patient’s immune condition is limited. Further studies are needed to elucidate the immune responses and prognosis of COVID-19.
In general, HT in patient 1 proved to be successful for the treatment of Chagas cardiomyopathy, with no reactivation of the pathogen. However, after SARS-CoV-2 infection, she presented graft rejection, treated with methylprednisolone, thymoglobulin, and plasmapheresis. There is a description in eye transplantation that COVID-19 infection can compromise the balance of immunoregulatory responses that allow graft survival, contributing to rejection in individuals infected with SARS-CoV-2 [31]. Thus, this change in the balance of attenuation of the immune response, such as a reduction in the number of regulatory T cells (Tregs) [32], may favor direct cardiotoxic action and multiple organ dysfunction.
During postoperative hospitalization, patient 2 presented Chagas reactivation, evidenced by skin biopsy, indicating that the CD was not completely controlled. In addition, it is important to consider that immunosuppressive therapy can contribute to the reactivation of CD [33]. After HT, the patient presented graft rejection and was treated with plasmapheresis and methylprednisolone. With this, before the SARS-CoV-2 infection, the patient was already weakened and died 21 days after the infection, in relation to patient 1 who died after 47 days of infection.
4. Conclusions
This report highlights the first case of an association between COVID-19, CD, and CMV dissemination in HT recipients. The patients had rapid disease progression to death. We believe that HT and the usage of immunosuppressive drugs, as well as immunosuppression generated by concomitant infections, may be an important risk factor for the development of severe COVID-19, especially in endemic areas with underreported CD infection.
Author Contributions
Conceptualization, S.C.G.-S., G.B., and R.W.A.; investigation, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., and S.M.; resources, M.N.S. and A.J.d.S.D.; writing—original draft preparation, S.C.G.-S., G.B., and R.W.A.; writing—review and editing, S.C.G.-S., G.B., and R.W.A.; visualization, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., S.M., A.J.d.S.D., and M.N.S.; supervision, M.N.S.; funding acquisition, A.J.d.S.D. and M.N.S. All authors have read and agreed to the published version of the manuscript.
Funding
This research was funded by Fundação de Amparo à Pesquisa (FAPESP), grant numbers 2019/22448-0, 19/02679-7, 2017/18199-9, 2018/18230-6, and 2019/26928-6. Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, CAPES:88887.503842/2020-00.
Institutional Review Board Statement
This study was approved by the local ethics committee (HCFMUSP nº 30800520.7.0000.0068-2020) and was carried out in accordance with the 2013 revision of the Declaration of Helsinki.
Informed Consent Statement
Informed consent was obtained from all of the subjects involved in the study.
Data Availability Statement
The laboratory data and imaging exams presented herein came from Hospital das Clinics, Faculty of Medicine, University of São Paulo (HCFMUSP) with prior approval for their use.
Conflicts of Interest
The authors declare no conflict of interest.
Figure 1 Chest X-ray of (a) patient 1 from May 2020 and (b) patient 2 from June 2015, taken just before heart transplantation, showing the Chagasic cardiomyopathy.
Figure 2 Daily clinical features of patients, from the first day of hospitalization in the ICU until death. Blood levels of (A) erythrocytes, (B) hemoglobin, (C) leukocytes, (D) neutrophils, (E) eosinophils, (F) lymphocytes, (G) monocytes, (H) neutrophil-to-lymphocyte ratio, (I) platelets, (J) creatinine, (K) urea, (L) prothrombin time, (M) activated partial thromboplastin time, (N) D-dimer, (O) pH, (P) pO2, (Q) pCO2, (R) oxygen peripheral, (S) CPR, (T) glucose, (U) CK-MB, (V) NT-proBNP, (W) lactate dehydrogenase, and (X) troponin T. Gray boxes represent the reference values.
tropicalmed-06-00022-t001_Table 1 Table 1 Patients’ characteristics, comorbidities, treatment, and complications.
Patient 1 Patient 2
Sex Female Male
Age 55 62
Positive SARS-CoV-2 RT-PCR date 1 June 2020 29 June 2020
Heart transplant date 2 May 2020 5 December 2019
Death date 30 days 207 days
Days between positive SARS-CoV-2 diagnosis and death 47 days 21 days
Complications during ICU stay ARDS due to COVID–19; heart transplant rejection, disseminated cytomegalovirus; aggravated chronic kidney disease and pressure ulcer ARDS due to COVID–19, disseminated cytomegalovirus and pancytopenia due to hemophagocytosis
Previous comorbidities Dilated cardiomyopathy of Chagas etiology, hypothyroidism by thyroidectomy by nodule 10 years ago Cardiomyopathy of Chagas etiology, disseminated cytomegalovirus, deep vein thrombosis, systemic arterial hypertension, diabetes, dyslipidemia, and chronic renal failure
Previous use of medications Carvedilol, losartan, furosemide, levothyroxine, isosorbide, and hydralazine Unknown
Medicines used during ICU stay Methylprednisolone, azathioprine, anti-thymocyte globulin, cyclosporine, tacrolimus, meropenem, linezolid, micafungin, vancomycin, polymyxin B, tigecycline, amikacin, fluconazole, ganciclovir, and hydrocortisone Meropenem, colistin, linezolid
fluconazole, amikacin, cyclosporine, azathioprine, and prednisone
ARD, acute respiratory distress syndrome; COVID-19, coronavirus disease 2019; ICU, intensive care unit; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. | AZATHIOPRINE, CYCLOSPORINE, METHYLPREDNISOLONE SODIUM SUCCINATE, PREDNISONE | DrugsGivenReaction | CC BY | 33579042 | 19,731,502 | 2021-02-10 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Oxygen saturation decreased'. | SARS-CoV-2 Infection and CMV Dissemination in Transplant Recipients as a Treatment for Chagas Cardiomyopathy: A Case Report.
Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 has infected over 90 million people worldwide, therefore it is considered a pandemic. SARS-CoV-2 infection can lead to severe pneumonia, acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure. Individuals receiving a heart transplantation (HT) may be at higher risk of adverse outcomes attributable to COVID-19 due to immunosuppressives, as well as concomitant infections that may also influence the prognoses. Herein, we describe the first report of two cases of HT recipients with concomitant infections by SARS-CoV-2, Trypanosoma cruzi, and cytomegalovirus (CMV) dissemination, from the first day of hospitalization due to COVID-19 in the intensive care unit (ICU) until the death of the patients.
1. Introduction
Coronavirus disease 2019 (COVID-19), an infectious respiratory disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spread on a pandemic scale since the first case was reported in Wuhan, China, in 2019 [1]. Most patients with the disease have mild-to-moderate symptoms; however, approximately 15% develop severe pneumonia, while approximately 5% develop acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure [2]. Lymphopenia is a recurrent feature in these patients, with a significant reduction in CD4+ T cells, CD8+ T cells, B cells, and natural killer (NK) cells [3], increasing the susceptibility of patients to severe illness and co-infection [4]. In fact, co-infections are associated with worsening of the clinical condition [5].
Chagas disease (CD) is a zoonosis whose etiologic agent is the protozoan Trypanosoma cruzi. It is estimated that 6–8 million people worldwide are infected [6]. CD has two distinct phases: an acute one, which is rare, with a strong production of type 1 cytokines, and a chronic phase, which develops in 30–40% of CD cases. The chronic phase of CD can be characterized by cardiomyopathy, arrhythmias, megaviscera, and, more rarely, polyneuropathy and stroke [7]. TCD4+ and TCD8+ lymphocytes are the main cells responsible for controlling parasitic infection. However, the immune response also contributes to tissue damage and pathology [8].
Chagasic cardiomyopathy represents the main cause of mortality from this disease, which can lead to heart failure, whose indication for treatment, especially in endemic countries, may include heart transplantation (HT) as a strategy to curb the evolution of this complication [6,9]. However, the immunosuppression and possible reactivation of the causative agent T. cruzi following HT require intensive clinical care and laboratory monitoring [10].
Other infections such as that caused by cytomegalovirus (CMV), a herpes virus that infects up to 60–100% of people in adulthood, are associated with transplant complications [11]. Primary CMV infection is generally asymptomatic in immunocompetent individuals, with the virus generating a latent infection. On the contrary, in immunocompromised and immunosuppressed populations, such as solid organ transplantation, hematopoietic stem cell transplantation, and HIV/AIDS patients, CMV reactivation is responsible for significant morbidity and mortality [12]. Although reactivation of infections such as CMV and CD are often described in transplant recipients, there are no reports of concomitant infection by T. cruzi, CMV, and SARS-CoV-2 in the context of HT in the literature to date.
Therefore, in this report, we investigated the progress of two patients who underwent HT at the Heart Institute of Hospital das Clínicas (Incor) and were subsequently transferred to the special intensive care unit (ICU) of the Hospital das Clínicas (Hospital das Clínicas, Faculty of Medicine, University of São Paulo-HCFMUSP) due to SARS-CoV-2 infection. These patients were diagnosed with COVID-19 by nasopharyngeal detection of SARS-CoV-2 RNA using reverse transcriptase polymerase chain reaction (RT-PCR). In addition, during hospitalization, CMV dissemination was evidenced by quantitative DNA detection in the blood. We describe herein the laboratory data from the first day of hospitalization due to COVID-19 until the death of the patients.
The baseline characteristics of these two patients, as well as their past clinical data, are summarized in Table 1.
2. Case Report
2.1. Patient 1
Female, 55 years old, presenting positive serology for CD since 2015, received HT on 2 May as a treatment for Chagas cardiomyopathy (Figure 1A) and underwent immunosuppressive therapy with methylprednisolone and azathioprine. Post-transplantation, she developed pneumonia, treated with meropenem and linezolid, and a remittent Candida tropicalis infection, treated with micafungin, with improvement. However, on 29 May, she developed dyspnea and desaturation, with a chest tomography suggestive of COVID-19. Her polymerase chain reaction (PCR) for SARS-CoV-2 was positive on 1 June.
On 3 June, she was transferred to the ICU specialized for treatment of severe SARS-CoV-2 infections. Laboratory analyses then showed that the patient had a reduced number of erythrocytes and hemoglobin level, and these reductions became more accentuated by day 33 in the ICU (Figure 2A,B). On this same day, the number of neutrophils peaked (Figure 2D). From the 4th to the 16th day in ICU, important lymphopenia developed (Figure 2F), resulting in an increase in the neutrophil-to-lymphocyte ratio in the same period (Figure 2H). Additionally, from the eighth day onward, she presented severe thrombocytopenia that persisted until death (Figure 2I).
Throughout the ICU hospitalization period, there were sustained high levels of creatinine, urea, D-dimer, C-reactive protein (CRP), and lactate dehydrogenase (Figure 2J,K,N,S,W). On the 32nd day, she presented a sharp increase in prothrombin time and activated partial thromboplastin time.
The cardiac function markers of creatine kinase myocardial band (CK-MB) and troponin remained elevated from the first days of ICU admission until death (Figure 2U,X). On the day of admission to the ICU, the N-terminal pro b-type natriuretic peptide (NT pro-BNP) was 70,000 pg/mL (reference value of <125 pg/mL) (Figure 2V). On the third day of hospitalization, NT pro-BNP (155,117 pg/mL) and troponin (0.28 ng/mL, reference value of <0.014 ng/mL) peaked, being related to the period in which she presented signs of graft rejection, which was treated with pulse methylprednisolone, thymoglobulin, and plasmapheresis. In addition, disseminated CMV infection was diagnosed (RT-PCR viral loads of 122 IU/mL on day 10 and 54 IU/mL on day 19 of hospitalization), for which she received ganciclovir.
After 47 days after diagnosis of SARS-CoV-2, the patient died (18 July) due to multiple organ dysfunctions associated with COVID-19.
2.2. Patient 2
Male, 62 years old, previously diagnosed with CD, received HT on 5 December 2019 as treatment for Chagas cardiomyopathy (Figure 1B). He underwent immunosuppressive therapy with cyclosporine, azathioprine, and prednisone. During postoperative hospitalization he presented Chagas reactivation characterized by skin biopsy and humoral graft rejection, being treated with plasmapheresis and methylprednisolone.
The patient was admitted to the ICU of the Hospital das Clínicas on 8 June, presenting cellulitis, deep vein thrombosis, and Chagas reactivation (a lower limb chagoma). The latter was treated with benznidazol. During hospitalization, he tested positive for PCR of SARS-CoV-2 on 29 June. He developed ARDS and septic and cardiogenic shock, which were the causes of his death.
Throughout the ICU hospitalization period, he maintained decreased levels of erythrocytes, hemoglobin, and lymphocytes (Figure 2A,B,F), as well as a high neutrophil/lymphocyte ratio (Figure 2H). From the 12th day, the number of platelets decreased and continued until the time of death (Figure 2I). The values of creatinine, urea, and glucose also remained high throughout the hospitalization period (Figure 2J,K,T). In addition, disseminated CMV (viral loads of 711 IU/mL detected on the 28th day and 1477 IU/mL on the 35th day of hospitalization) was diagnosed and treated with ganciclovir.
One day before death, the following laboratory parameters peaked: D-dimer (7612 ng/mL FEU, reference value of <500 ng/mL FEU), CRP (280.6 ng/mL, reference value of 0.300 ng/mL), CK-MB (8.38 ng/mL, reference value of 0.10–4.94 ng/mL), NT pro-BNP (55,393 pg/mL), lactate dehydrogenase (1161 U/L, reference value of 135–225 U/L), and troponin (0.701 ng/mL) (Figure 2N,S,U–X).
On 20 July, 36 days after admission to the ICU and 21 days after a positive COVID-19 diagnosis, the patient died.
3. Discussion
Herein, we described the first report of triple infection (SARS-CoV-2 infection, T. cruzi infection, and CMV dissemination) in HT recipients. Patients received HT as a form of treatment for Chagas cardiomyopathy.
We described the laboratory data from the first day of hospitalization in the ICU due to COVID-19 until the time of death. Both patients were admitted to a referral center for treatment for COVID-19 in the metropolitan region of São Paulo, a city in southeastern Brazil. We hypothesize that the triple infection by SARS-CoV-2 and CMV may have been an important cause of death and of the worsening in CD patients with HT. To date, there have been no similar reports of patients presenting these three concomitant infectious diseases and HT receptors.
COVID-19 infection among transplant recipients increases the potential for developing severe illness [13] and may vary between different organ transplants [14,15].
SARS-CoV-2 entry´s receptor angiotensin converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2) are expressed in different tissues, such as the lungs, heart, liver, kidneys, testicles, thyroid, and adipose tissue [16]. Some patients with COVID-19 develop severe disease characterized by respiratory distress syndrome and systemic manifestations. This condition has been associated with the dysregulated release of pro-inflammatory cytokines, termed “cytokine storm,” which may induce multi-organ failure [1,17].
The use of immunosuppressants and post-surgical opportunistic infections can also lead to damage of multisystem organs or even death [18]. There is a therapeutic paradox here, because while insufficient immunosuppression results in graft loss due to rejection, excessive immunosuppression can result in serious infection, including SARS-COV-2 [13], besides contributing to the reactivation of pathogens [19].
Long-term administration of immunosuppressants to solid organ transplant (SOT) receptors to reduce the risk of graft rejection may increase the risk of respiratory infections [20], although there is no clear clinical evidence of increased morbidity/mortality in SARS-CoV-2 infection [21].
Fernandez-Ruiz and collaborators [22] described a cohort of SOT receptors affected by COVID-19, 44% of whom were kidney transplant recipients, 33.3% liver transplant recipients, and 22.2% heart transplant recipients. The lethality rate was 27.8%, suggesting that SARS-CoV-2 infection had a severe course in SOT recipients.
Recently, we described the first patients with CD affected by SARS-CoV-2. The CD patients presented an increase in COVID-19 laboratory hallmarks and a rapid disease progression. Despite the efforts of the health staff, both patients died [3].
HT may be associated with the reactivation of pathogens, as reported in CD [10]. In a Brazilian cohort, the reactivation rate of Chagas disease after heart transplantation was reported to be 38.8% [23]. This fact made HT as a treatment for Chagas cardiomyopathy initially controversial. However, currently, especially in endemic countries, it is the most viable therapeutic option for patients with end-stage heart failure [24]. We hypothesize that, in the cases presented here, HT followed by reactivation of CD conferred an additional risk factor for the worsening of COVID-19.
Transplant recipients are also commonly affected by CMV reactivation, being associated with significant morbidity and mortality [12] that may worsen the infectious condition of COVID-19 [25]. In SOT, risk factors include the use of immunosuppressants for transplantation, advanced age, acute rejection, and other concomitant infections [11], with all these characteristics being present in both patients described herein. In the present report, we observed that immunosuppression may have contributed to the susceptibility to superinfections and more severe clinical manifestations in individuals undergoing HT.
Since overactivated immune responses can be one of the causes of organ damage, the anti-inflammatory effects of immunosuppression can be protective, reducing the cytokine storm related to complications in COVID-19 [26]. In this context, it has been described that immunosuppressive therapy with calcineurin inhibitors in patients with solid organ transplantation or systemic rheumatic diseases promotes a clinical course in SARS-CoV-2 infection, which is generally mild, and with an apparently low risk of superinfection [27]. In addition, immunosuppression has not been evaluated as a risk factor for SARS or MERS [28].
However, in the present case report, we observed that immunosuppression may have contributed to the susceptibility to SARS-CoV-2 infection, the reactivation of pathogens, and more serious clinical manifestations in individuals undergoing HT. Corroborating our findings, it was shown that patients with COVID-19 and cancer, due to their systemic immunosuppressive condition caused by malignancy and anticancer treatments, such as chemotherapy, had an increased risk of SARS-CoV-2 infection and a worse prognosis [29]. It has also been reported that chronic use of corticosteroids prior to SARS-CoV-2 contamination is associated with critical disease outcomes, including a high risk of death [30].
These contradictory observations show that knowledge about the relationship between COVID-19 and the patient’s immune condition is limited. Further studies are needed to elucidate the immune responses and prognosis of COVID-19.
In general, HT in patient 1 proved to be successful for the treatment of Chagas cardiomyopathy, with no reactivation of the pathogen. However, after SARS-CoV-2 infection, she presented graft rejection, treated with methylprednisolone, thymoglobulin, and plasmapheresis. There is a description in eye transplantation that COVID-19 infection can compromise the balance of immunoregulatory responses that allow graft survival, contributing to rejection in individuals infected with SARS-CoV-2 [31]. Thus, this change in the balance of attenuation of the immune response, such as a reduction in the number of regulatory T cells (Tregs) [32], may favor direct cardiotoxic action and multiple organ dysfunction.
During postoperative hospitalization, patient 2 presented Chagas reactivation, evidenced by skin biopsy, indicating that the CD was not completely controlled. In addition, it is important to consider that immunosuppressive therapy can contribute to the reactivation of CD [33]. After HT, the patient presented graft rejection and was treated with plasmapheresis and methylprednisolone. With this, before the SARS-CoV-2 infection, the patient was already weakened and died 21 days after the infection, in relation to patient 1 who died after 47 days of infection.
4. Conclusions
This report highlights the first case of an association between COVID-19, CD, and CMV dissemination in HT recipients. The patients had rapid disease progression to death. We believe that HT and the usage of immunosuppressive drugs, as well as immunosuppression generated by concomitant infections, may be an important risk factor for the development of severe COVID-19, especially in endemic areas with underreported CD infection.
Author Contributions
Conceptualization, S.C.G.-S., G.B., and R.W.A.; investigation, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., and S.M.; resources, M.N.S. and A.J.d.S.D.; writing—original draft preparation, S.C.G.-S., G.B., and R.W.A.; writing—review and editing, S.C.G.-S., G.B., and R.W.A.; visualization, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., S.M., A.J.d.S.D., and M.N.S.; supervision, M.N.S.; funding acquisition, A.J.d.S.D. and M.N.S. All authors have read and agreed to the published version of the manuscript.
Funding
This research was funded by Fundação de Amparo à Pesquisa (FAPESP), grant numbers 2019/22448-0, 19/02679-7, 2017/18199-9, 2018/18230-6, and 2019/26928-6. Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, CAPES:88887.503842/2020-00.
Institutional Review Board Statement
This study was approved by the local ethics committee (HCFMUSP nº 30800520.7.0000.0068-2020) and was carried out in accordance with the 2013 revision of the Declaration of Helsinki.
Informed Consent Statement
Informed consent was obtained from all of the subjects involved in the study.
Data Availability Statement
The laboratory data and imaging exams presented herein came from Hospital das Clinics, Faculty of Medicine, University of São Paulo (HCFMUSP) with prior approval for their use.
Conflicts of Interest
The authors declare no conflict of interest.
Figure 1 Chest X-ray of (a) patient 1 from May 2020 and (b) patient 2 from June 2015, taken just before heart transplantation, showing the Chagasic cardiomyopathy.
Figure 2 Daily clinical features of patients, from the first day of hospitalization in the ICU until death. Blood levels of (A) erythrocytes, (B) hemoglobin, (C) leukocytes, (D) neutrophils, (E) eosinophils, (F) lymphocytes, (G) monocytes, (H) neutrophil-to-lymphocyte ratio, (I) platelets, (J) creatinine, (K) urea, (L) prothrombin time, (M) activated partial thromboplastin time, (N) D-dimer, (O) pH, (P) pO2, (Q) pCO2, (R) oxygen peripheral, (S) CPR, (T) glucose, (U) CK-MB, (V) NT-proBNP, (W) lactate dehydrogenase, and (X) troponin T. Gray boxes represent the reference values.
tropicalmed-06-00022-t001_Table 1 Table 1 Patients’ characteristics, comorbidities, treatment, and complications.
Patient 1 Patient 2
Sex Female Male
Age 55 62
Positive SARS-CoV-2 RT-PCR date 1 June 2020 29 June 2020
Heart transplant date 2 May 2020 5 December 2019
Death date 30 days 207 days
Days between positive SARS-CoV-2 diagnosis and death 47 days 21 days
Complications during ICU stay ARDS due to COVID–19; heart transplant rejection, disseminated cytomegalovirus; aggravated chronic kidney disease and pressure ulcer ARDS due to COVID–19, disseminated cytomegalovirus and pancytopenia due to hemophagocytosis
Previous comorbidities Dilated cardiomyopathy of Chagas etiology, hypothyroidism by thyroidectomy by nodule 10 years ago Cardiomyopathy of Chagas etiology, disseminated cytomegalovirus, deep vein thrombosis, systemic arterial hypertension, diabetes, dyslipidemia, and chronic renal failure
Previous use of medications Carvedilol, losartan, furosemide, levothyroxine, isosorbide, and hydralazine Unknown
Medicines used during ICU stay Methylprednisolone, azathioprine, anti-thymocyte globulin, cyclosporine, tacrolimus, meropenem, linezolid, micafungin, vancomycin, polymyxin B, tigecycline, amikacin, fluconazole, ganciclovir, and hydrocortisone Meropenem, colistin, linezolid
fluconazole, amikacin, cyclosporine, azathioprine, and prednisone
ARD, acute respiratory distress syndrome; COVID-19, coronavirus disease 2019; ICU, intensive care unit; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. | AZATHIOPRINE, CARVEDILOL, CYCLOSPORINE, FUROSEMIDE, HYDRALAZINE HYDROCHLORIDE, HYDROCORTISONE, ISOSORBIDE, LAPINE T-LYMPHOCYTE IMMUNE GLOBULIN, LEVOTHYROXINE, LOSARTAN, METHYLPREDNISOLONE, TACROLIMUS | DrugsGivenReaction | CC BY | 33579042 | 19,681,859 | 2021-02-10 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Pancytopenia'. | SARS-CoV-2 Infection and CMV Dissemination in Transplant Recipients as a Treatment for Chagas Cardiomyopathy: A Case Report.
Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 has infected over 90 million people worldwide, therefore it is considered a pandemic. SARS-CoV-2 infection can lead to severe pneumonia, acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure. Individuals receiving a heart transplantation (HT) may be at higher risk of adverse outcomes attributable to COVID-19 due to immunosuppressives, as well as concomitant infections that may also influence the prognoses. Herein, we describe the first report of two cases of HT recipients with concomitant infections by SARS-CoV-2, Trypanosoma cruzi, and cytomegalovirus (CMV) dissemination, from the first day of hospitalization due to COVID-19 in the intensive care unit (ICU) until the death of the patients.
1. Introduction
Coronavirus disease 2019 (COVID-19), an infectious respiratory disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spread on a pandemic scale since the first case was reported in Wuhan, China, in 2019 [1]. Most patients with the disease have mild-to-moderate symptoms; however, approximately 15% develop severe pneumonia, while approximately 5% develop acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure [2]. Lymphopenia is a recurrent feature in these patients, with a significant reduction in CD4+ T cells, CD8+ T cells, B cells, and natural killer (NK) cells [3], increasing the susceptibility of patients to severe illness and co-infection [4]. In fact, co-infections are associated with worsening of the clinical condition [5].
Chagas disease (CD) is a zoonosis whose etiologic agent is the protozoan Trypanosoma cruzi. It is estimated that 6–8 million people worldwide are infected [6]. CD has two distinct phases: an acute one, which is rare, with a strong production of type 1 cytokines, and a chronic phase, which develops in 30–40% of CD cases. The chronic phase of CD can be characterized by cardiomyopathy, arrhythmias, megaviscera, and, more rarely, polyneuropathy and stroke [7]. TCD4+ and TCD8+ lymphocytes are the main cells responsible for controlling parasitic infection. However, the immune response also contributes to tissue damage and pathology [8].
Chagasic cardiomyopathy represents the main cause of mortality from this disease, which can lead to heart failure, whose indication for treatment, especially in endemic countries, may include heart transplantation (HT) as a strategy to curb the evolution of this complication [6,9]. However, the immunosuppression and possible reactivation of the causative agent T. cruzi following HT require intensive clinical care and laboratory monitoring [10].
Other infections such as that caused by cytomegalovirus (CMV), a herpes virus that infects up to 60–100% of people in adulthood, are associated with transplant complications [11]. Primary CMV infection is generally asymptomatic in immunocompetent individuals, with the virus generating a latent infection. On the contrary, in immunocompromised and immunosuppressed populations, such as solid organ transplantation, hematopoietic stem cell transplantation, and HIV/AIDS patients, CMV reactivation is responsible for significant morbidity and mortality [12]. Although reactivation of infections such as CMV and CD are often described in transplant recipients, there are no reports of concomitant infection by T. cruzi, CMV, and SARS-CoV-2 in the context of HT in the literature to date.
Therefore, in this report, we investigated the progress of two patients who underwent HT at the Heart Institute of Hospital das Clínicas (Incor) and were subsequently transferred to the special intensive care unit (ICU) of the Hospital das Clínicas (Hospital das Clínicas, Faculty of Medicine, University of São Paulo-HCFMUSP) due to SARS-CoV-2 infection. These patients were diagnosed with COVID-19 by nasopharyngeal detection of SARS-CoV-2 RNA using reverse transcriptase polymerase chain reaction (RT-PCR). In addition, during hospitalization, CMV dissemination was evidenced by quantitative DNA detection in the blood. We describe herein the laboratory data from the first day of hospitalization due to COVID-19 until the death of the patients.
The baseline characteristics of these two patients, as well as their past clinical data, are summarized in Table 1.
2. Case Report
2.1. Patient 1
Female, 55 years old, presenting positive serology for CD since 2015, received HT on 2 May as a treatment for Chagas cardiomyopathy (Figure 1A) and underwent immunosuppressive therapy with methylprednisolone and azathioprine. Post-transplantation, she developed pneumonia, treated with meropenem and linezolid, and a remittent Candida tropicalis infection, treated with micafungin, with improvement. However, on 29 May, she developed dyspnea and desaturation, with a chest tomography suggestive of COVID-19. Her polymerase chain reaction (PCR) for SARS-CoV-2 was positive on 1 June.
On 3 June, she was transferred to the ICU specialized for treatment of severe SARS-CoV-2 infections. Laboratory analyses then showed that the patient had a reduced number of erythrocytes and hemoglobin level, and these reductions became more accentuated by day 33 in the ICU (Figure 2A,B). On this same day, the number of neutrophils peaked (Figure 2D). From the 4th to the 16th day in ICU, important lymphopenia developed (Figure 2F), resulting in an increase in the neutrophil-to-lymphocyte ratio in the same period (Figure 2H). Additionally, from the eighth day onward, she presented severe thrombocytopenia that persisted until death (Figure 2I).
Throughout the ICU hospitalization period, there were sustained high levels of creatinine, urea, D-dimer, C-reactive protein (CRP), and lactate dehydrogenase (Figure 2J,K,N,S,W). On the 32nd day, she presented a sharp increase in prothrombin time and activated partial thromboplastin time.
The cardiac function markers of creatine kinase myocardial band (CK-MB) and troponin remained elevated from the first days of ICU admission until death (Figure 2U,X). On the day of admission to the ICU, the N-terminal pro b-type natriuretic peptide (NT pro-BNP) was 70,000 pg/mL (reference value of <125 pg/mL) (Figure 2V). On the third day of hospitalization, NT pro-BNP (155,117 pg/mL) and troponin (0.28 ng/mL, reference value of <0.014 ng/mL) peaked, being related to the period in which she presented signs of graft rejection, which was treated with pulse methylprednisolone, thymoglobulin, and plasmapheresis. In addition, disseminated CMV infection was diagnosed (RT-PCR viral loads of 122 IU/mL on day 10 and 54 IU/mL on day 19 of hospitalization), for which she received ganciclovir.
After 47 days after diagnosis of SARS-CoV-2, the patient died (18 July) due to multiple organ dysfunctions associated with COVID-19.
2.2. Patient 2
Male, 62 years old, previously diagnosed with CD, received HT on 5 December 2019 as treatment for Chagas cardiomyopathy (Figure 1B). He underwent immunosuppressive therapy with cyclosporine, azathioprine, and prednisone. During postoperative hospitalization he presented Chagas reactivation characterized by skin biopsy and humoral graft rejection, being treated with plasmapheresis and methylprednisolone.
The patient was admitted to the ICU of the Hospital das Clínicas on 8 June, presenting cellulitis, deep vein thrombosis, and Chagas reactivation (a lower limb chagoma). The latter was treated with benznidazol. During hospitalization, he tested positive for PCR of SARS-CoV-2 on 29 June. He developed ARDS and septic and cardiogenic shock, which were the causes of his death.
Throughout the ICU hospitalization period, he maintained decreased levels of erythrocytes, hemoglobin, and lymphocytes (Figure 2A,B,F), as well as a high neutrophil/lymphocyte ratio (Figure 2H). From the 12th day, the number of platelets decreased and continued until the time of death (Figure 2I). The values of creatinine, urea, and glucose also remained high throughout the hospitalization period (Figure 2J,K,T). In addition, disseminated CMV (viral loads of 711 IU/mL detected on the 28th day and 1477 IU/mL on the 35th day of hospitalization) was diagnosed and treated with ganciclovir.
One day before death, the following laboratory parameters peaked: D-dimer (7612 ng/mL FEU, reference value of <500 ng/mL FEU), CRP (280.6 ng/mL, reference value of 0.300 ng/mL), CK-MB (8.38 ng/mL, reference value of 0.10–4.94 ng/mL), NT pro-BNP (55,393 pg/mL), lactate dehydrogenase (1161 U/L, reference value of 135–225 U/L), and troponin (0.701 ng/mL) (Figure 2N,S,U–X).
On 20 July, 36 days after admission to the ICU and 21 days after a positive COVID-19 diagnosis, the patient died.
3. Discussion
Herein, we described the first report of triple infection (SARS-CoV-2 infection, T. cruzi infection, and CMV dissemination) in HT recipients. Patients received HT as a form of treatment for Chagas cardiomyopathy.
We described the laboratory data from the first day of hospitalization in the ICU due to COVID-19 until the time of death. Both patients were admitted to a referral center for treatment for COVID-19 in the metropolitan region of São Paulo, a city in southeastern Brazil. We hypothesize that the triple infection by SARS-CoV-2 and CMV may have been an important cause of death and of the worsening in CD patients with HT. To date, there have been no similar reports of patients presenting these three concomitant infectious diseases and HT receptors.
COVID-19 infection among transplant recipients increases the potential for developing severe illness [13] and may vary between different organ transplants [14,15].
SARS-CoV-2 entry´s receptor angiotensin converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2) are expressed in different tissues, such as the lungs, heart, liver, kidneys, testicles, thyroid, and adipose tissue [16]. Some patients with COVID-19 develop severe disease characterized by respiratory distress syndrome and systemic manifestations. This condition has been associated with the dysregulated release of pro-inflammatory cytokines, termed “cytokine storm,” which may induce multi-organ failure [1,17].
The use of immunosuppressants and post-surgical opportunistic infections can also lead to damage of multisystem organs or even death [18]. There is a therapeutic paradox here, because while insufficient immunosuppression results in graft loss due to rejection, excessive immunosuppression can result in serious infection, including SARS-COV-2 [13], besides contributing to the reactivation of pathogens [19].
Long-term administration of immunosuppressants to solid organ transplant (SOT) receptors to reduce the risk of graft rejection may increase the risk of respiratory infections [20], although there is no clear clinical evidence of increased morbidity/mortality in SARS-CoV-2 infection [21].
Fernandez-Ruiz and collaborators [22] described a cohort of SOT receptors affected by COVID-19, 44% of whom were kidney transplant recipients, 33.3% liver transplant recipients, and 22.2% heart transplant recipients. The lethality rate was 27.8%, suggesting that SARS-CoV-2 infection had a severe course in SOT recipients.
Recently, we described the first patients with CD affected by SARS-CoV-2. The CD patients presented an increase in COVID-19 laboratory hallmarks and a rapid disease progression. Despite the efforts of the health staff, both patients died [3].
HT may be associated with the reactivation of pathogens, as reported in CD [10]. In a Brazilian cohort, the reactivation rate of Chagas disease after heart transplantation was reported to be 38.8% [23]. This fact made HT as a treatment for Chagas cardiomyopathy initially controversial. However, currently, especially in endemic countries, it is the most viable therapeutic option for patients with end-stage heart failure [24]. We hypothesize that, in the cases presented here, HT followed by reactivation of CD conferred an additional risk factor for the worsening of COVID-19.
Transplant recipients are also commonly affected by CMV reactivation, being associated with significant morbidity and mortality [12] that may worsen the infectious condition of COVID-19 [25]. In SOT, risk factors include the use of immunosuppressants for transplantation, advanced age, acute rejection, and other concomitant infections [11], with all these characteristics being present in both patients described herein. In the present report, we observed that immunosuppression may have contributed to the susceptibility to superinfections and more severe clinical manifestations in individuals undergoing HT.
Since overactivated immune responses can be one of the causes of organ damage, the anti-inflammatory effects of immunosuppression can be protective, reducing the cytokine storm related to complications in COVID-19 [26]. In this context, it has been described that immunosuppressive therapy with calcineurin inhibitors in patients with solid organ transplantation or systemic rheumatic diseases promotes a clinical course in SARS-CoV-2 infection, which is generally mild, and with an apparently low risk of superinfection [27]. In addition, immunosuppression has not been evaluated as a risk factor for SARS or MERS [28].
However, in the present case report, we observed that immunosuppression may have contributed to the susceptibility to SARS-CoV-2 infection, the reactivation of pathogens, and more serious clinical manifestations in individuals undergoing HT. Corroborating our findings, it was shown that patients with COVID-19 and cancer, due to their systemic immunosuppressive condition caused by malignancy and anticancer treatments, such as chemotherapy, had an increased risk of SARS-CoV-2 infection and a worse prognosis [29]. It has also been reported that chronic use of corticosteroids prior to SARS-CoV-2 contamination is associated with critical disease outcomes, including a high risk of death [30].
These contradictory observations show that knowledge about the relationship between COVID-19 and the patient’s immune condition is limited. Further studies are needed to elucidate the immune responses and prognosis of COVID-19.
In general, HT in patient 1 proved to be successful for the treatment of Chagas cardiomyopathy, with no reactivation of the pathogen. However, after SARS-CoV-2 infection, she presented graft rejection, treated with methylprednisolone, thymoglobulin, and plasmapheresis. There is a description in eye transplantation that COVID-19 infection can compromise the balance of immunoregulatory responses that allow graft survival, contributing to rejection in individuals infected with SARS-CoV-2 [31]. Thus, this change in the balance of attenuation of the immune response, such as a reduction in the number of regulatory T cells (Tregs) [32], may favor direct cardiotoxic action and multiple organ dysfunction.
During postoperative hospitalization, patient 2 presented Chagas reactivation, evidenced by skin biopsy, indicating that the CD was not completely controlled. In addition, it is important to consider that immunosuppressive therapy can contribute to the reactivation of CD [33]. After HT, the patient presented graft rejection and was treated with plasmapheresis and methylprednisolone. With this, before the SARS-CoV-2 infection, the patient was already weakened and died 21 days after the infection, in relation to patient 1 who died after 47 days of infection.
4. Conclusions
This report highlights the first case of an association between COVID-19, CD, and CMV dissemination in HT recipients. The patients had rapid disease progression to death. We believe that HT and the usage of immunosuppressive drugs, as well as immunosuppression generated by concomitant infections, may be an important risk factor for the development of severe COVID-19, especially in endemic areas with underreported CD infection.
Author Contributions
Conceptualization, S.C.G.-S., G.B., and R.W.A.; investigation, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., and S.M.; resources, M.N.S. and A.J.d.S.D.; writing—original draft preparation, S.C.G.-S., G.B., and R.W.A.; writing—review and editing, S.C.G.-S., G.B., and R.W.A.; visualization, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., S.M., A.J.d.S.D., and M.N.S.; supervision, M.N.S.; funding acquisition, A.J.d.S.D. and M.N.S. All authors have read and agreed to the published version of the manuscript.
Funding
This research was funded by Fundação de Amparo à Pesquisa (FAPESP), grant numbers 2019/22448-0, 19/02679-7, 2017/18199-9, 2018/18230-6, and 2019/26928-6. Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, CAPES:88887.503842/2020-00.
Institutional Review Board Statement
This study was approved by the local ethics committee (HCFMUSP nº 30800520.7.0000.0068-2020) and was carried out in accordance with the 2013 revision of the Declaration of Helsinki.
Informed Consent Statement
Informed consent was obtained from all of the subjects involved in the study.
Data Availability Statement
The laboratory data and imaging exams presented herein came from Hospital das Clinics, Faculty of Medicine, University of São Paulo (HCFMUSP) with prior approval for their use.
Conflicts of Interest
The authors declare no conflict of interest.
Figure 1 Chest X-ray of (a) patient 1 from May 2020 and (b) patient 2 from June 2015, taken just before heart transplantation, showing the Chagasic cardiomyopathy.
Figure 2 Daily clinical features of patients, from the first day of hospitalization in the ICU until death. Blood levels of (A) erythrocytes, (B) hemoglobin, (C) leukocytes, (D) neutrophils, (E) eosinophils, (F) lymphocytes, (G) monocytes, (H) neutrophil-to-lymphocyte ratio, (I) platelets, (J) creatinine, (K) urea, (L) prothrombin time, (M) activated partial thromboplastin time, (N) D-dimer, (O) pH, (P) pO2, (Q) pCO2, (R) oxygen peripheral, (S) CPR, (T) glucose, (U) CK-MB, (V) NT-proBNP, (W) lactate dehydrogenase, and (X) troponin T. Gray boxes represent the reference values.
tropicalmed-06-00022-t001_Table 1 Table 1 Patients’ characteristics, comorbidities, treatment, and complications.
Patient 1 Patient 2
Sex Female Male
Age 55 62
Positive SARS-CoV-2 RT-PCR date 1 June 2020 29 June 2020
Heart transplant date 2 May 2020 5 December 2019
Death date 30 days 207 days
Days between positive SARS-CoV-2 diagnosis and death 47 days 21 days
Complications during ICU stay ARDS due to COVID–19; heart transplant rejection, disseminated cytomegalovirus; aggravated chronic kidney disease and pressure ulcer ARDS due to COVID–19, disseminated cytomegalovirus and pancytopenia due to hemophagocytosis
Previous comorbidities Dilated cardiomyopathy of Chagas etiology, hypothyroidism by thyroidectomy by nodule 10 years ago Cardiomyopathy of Chagas etiology, disseminated cytomegalovirus, deep vein thrombosis, systemic arterial hypertension, diabetes, dyslipidemia, and chronic renal failure
Previous use of medications Carvedilol, losartan, furosemide, levothyroxine, isosorbide, and hydralazine Unknown
Medicines used during ICU stay Methylprednisolone, azathioprine, anti-thymocyte globulin, cyclosporine, tacrolimus, meropenem, linezolid, micafungin, vancomycin, polymyxin B, tigecycline, amikacin, fluconazole, ganciclovir, and hydrocortisone Meropenem, colistin, linezolid
fluconazole, amikacin, cyclosporine, azathioprine, and prednisone
ARD, acute respiratory distress syndrome; COVID-19, coronavirus disease 2019; ICU, intensive care unit; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. | AZATHIOPRINE, CYCLOSPORINE, METHYLPREDNISOLONE SODIUM SUCCINATE, PREDNISONE | DrugsGivenReaction | CC BY | 33579042 | 19,731,502 | 2021-02-10 |
What was the dosage of drug 'METHYLPREDNISOLONE SODIUM SUCCINATE'? | SARS-CoV-2 Infection and CMV Dissemination in Transplant Recipients as a Treatment for Chagas Cardiomyopathy: A Case Report.
Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 has infected over 90 million people worldwide, therefore it is considered a pandemic. SARS-CoV-2 infection can lead to severe pneumonia, acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure. Individuals receiving a heart transplantation (HT) may be at higher risk of adverse outcomes attributable to COVID-19 due to immunosuppressives, as well as concomitant infections that may also influence the prognoses. Herein, we describe the first report of two cases of HT recipients with concomitant infections by SARS-CoV-2, Trypanosoma cruzi, and cytomegalovirus (CMV) dissemination, from the first day of hospitalization due to COVID-19 in the intensive care unit (ICU) until the death of the patients.
1. Introduction
Coronavirus disease 2019 (COVID-19), an infectious respiratory disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spread on a pandemic scale since the first case was reported in Wuhan, China, in 2019 [1]. Most patients with the disease have mild-to-moderate symptoms; however, approximately 15% develop severe pneumonia, while approximately 5% develop acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure [2]. Lymphopenia is a recurrent feature in these patients, with a significant reduction in CD4+ T cells, CD8+ T cells, B cells, and natural killer (NK) cells [3], increasing the susceptibility of patients to severe illness and co-infection [4]. In fact, co-infections are associated with worsening of the clinical condition [5].
Chagas disease (CD) is a zoonosis whose etiologic agent is the protozoan Trypanosoma cruzi. It is estimated that 6–8 million people worldwide are infected [6]. CD has two distinct phases: an acute one, which is rare, with a strong production of type 1 cytokines, and a chronic phase, which develops in 30–40% of CD cases. The chronic phase of CD can be characterized by cardiomyopathy, arrhythmias, megaviscera, and, more rarely, polyneuropathy and stroke [7]. TCD4+ and TCD8+ lymphocytes are the main cells responsible for controlling parasitic infection. However, the immune response also contributes to tissue damage and pathology [8].
Chagasic cardiomyopathy represents the main cause of mortality from this disease, which can lead to heart failure, whose indication for treatment, especially in endemic countries, may include heart transplantation (HT) as a strategy to curb the evolution of this complication [6,9]. However, the immunosuppression and possible reactivation of the causative agent T. cruzi following HT require intensive clinical care and laboratory monitoring [10].
Other infections such as that caused by cytomegalovirus (CMV), a herpes virus that infects up to 60–100% of people in adulthood, are associated with transplant complications [11]. Primary CMV infection is generally asymptomatic in immunocompetent individuals, with the virus generating a latent infection. On the contrary, in immunocompromised and immunosuppressed populations, such as solid organ transplantation, hematopoietic stem cell transplantation, and HIV/AIDS patients, CMV reactivation is responsible for significant morbidity and mortality [12]. Although reactivation of infections such as CMV and CD are often described in transplant recipients, there are no reports of concomitant infection by T. cruzi, CMV, and SARS-CoV-2 in the context of HT in the literature to date.
Therefore, in this report, we investigated the progress of two patients who underwent HT at the Heart Institute of Hospital das Clínicas (Incor) and were subsequently transferred to the special intensive care unit (ICU) of the Hospital das Clínicas (Hospital das Clínicas, Faculty of Medicine, University of São Paulo-HCFMUSP) due to SARS-CoV-2 infection. These patients were diagnosed with COVID-19 by nasopharyngeal detection of SARS-CoV-2 RNA using reverse transcriptase polymerase chain reaction (RT-PCR). In addition, during hospitalization, CMV dissemination was evidenced by quantitative DNA detection in the blood. We describe herein the laboratory data from the first day of hospitalization due to COVID-19 until the death of the patients.
The baseline characteristics of these two patients, as well as their past clinical data, are summarized in Table 1.
2. Case Report
2.1. Patient 1
Female, 55 years old, presenting positive serology for CD since 2015, received HT on 2 May as a treatment for Chagas cardiomyopathy (Figure 1A) and underwent immunosuppressive therapy with methylprednisolone and azathioprine. Post-transplantation, she developed pneumonia, treated with meropenem and linezolid, and a remittent Candida tropicalis infection, treated with micafungin, with improvement. However, on 29 May, she developed dyspnea and desaturation, with a chest tomography suggestive of COVID-19. Her polymerase chain reaction (PCR) for SARS-CoV-2 was positive on 1 June.
On 3 June, she was transferred to the ICU specialized for treatment of severe SARS-CoV-2 infections. Laboratory analyses then showed that the patient had a reduced number of erythrocytes and hemoglobin level, and these reductions became more accentuated by day 33 in the ICU (Figure 2A,B). On this same day, the number of neutrophils peaked (Figure 2D). From the 4th to the 16th day in ICU, important lymphopenia developed (Figure 2F), resulting in an increase in the neutrophil-to-lymphocyte ratio in the same period (Figure 2H). Additionally, from the eighth day onward, she presented severe thrombocytopenia that persisted until death (Figure 2I).
Throughout the ICU hospitalization period, there were sustained high levels of creatinine, urea, D-dimer, C-reactive protein (CRP), and lactate dehydrogenase (Figure 2J,K,N,S,W). On the 32nd day, she presented a sharp increase in prothrombin time and activated partial thromboplastin time.
The cardiac function markers of creatine kinase myocardial band (CK-MB) and troponin remained elevated from the first days of ICU admission until death (Figure 2U,X). On the day of admission to the ICU, the N-terminal pro b-type natriuretic peptide (NT pro-BNP) was 70,000 pg/mL (reference value of <125 pg/mL) (Figure 2V). On the third day of hospitalization, NT pro-BNP (155,117 pg/mL) and troponin (0.28 ng/mL, reference value of <0.014 ng/mL) peaked, being related to the period in which she presented signs of graft rejection, which was treated with pulse methylprednisolone, thymoglobulin, and plasmapheresis. In addition, disseminated CMV infection was diagnosed (RT-PCR viral loads of 122 IU/mL on day 10 and 54 IU/mL on day 19 of hospitalization), for which she received ganciclovir.
After 47 days after diagnosis of SARS-CoV-2, the patient died (18 July) due to multiple organ dysfunctions associated with COVID-19.
2.2. Patient 2
Male, 62 years old, previously diagnosed with CD, received HT on 5 December 2019 as treatment for Chagas cardiomyopathy (Figure 1B). He underwent immunosuppressive therapy with cyclosporine, azathioprine, and prednisone. During postoperative hospitalization he presented Chagas reactivation characterized by skin biopsy and humoral graft rejection, being treated with plasmapheresis and methylprednisolone.
The patient was admitted to the ICU of the Hospital das Clínicas on 8 June, presenting cellulitis, deep vein thrombosis, and Chagas reactivation (a lower limb chagoma). The latter was treated with benznidazol. During hospitalization, he tested positive for PCR of SARS-CoV-2 on 29 June. He developed ARDS and septic and cardiogenic shock, which were the causes of his death.
Throughout the ICU hospitalization period, he maintained decreased levels of erythrocytes, hemoglobin, and lymphocytes (Figure 2A,B,F), as well as a high neutrophil/lymphocyte ratio (Figure 2H). From the 12th day, the number of platelets decreased and continued until the time of death (Figure 2I). The values of creatinine, urea, and glucose also remained high throughout the hospitalization period (Figure 2J,K,T). In addition, disseminated CMV (viral loads of 711 IU/mL detected on the 28th day and 1477 IU/mL on the 35th day of hospitalization) was diagnosed and treated with ganciclovir.
One day before death, the following laboratory parameters peaked: D-dimer (7612 ng/mL FEU, reference value of <500 ng/mL FEU), CRP (280.6 ng/mL, reference value of 0.300 ng/mL), CK-MB (8.38 ng/mL, reference value of 0.10–4.94 ng/mL), NT pro-BNP (55,393 pg/mL), lactate dehydrogenase (1161 U/L, reference value of 135–225 U/L), and troponin (0.701 ng/mL) (Figure 2N,S,U–X).
On 20 July, 36 days after admission to the ICU and 21 days after a positive COVID-19 diagnosis, the patient died.
3. Discussion
Herein, we described the first report of triple infection (SARS-CoV-2 infection, T. cruzi infection, and CMV dissemination) in HT recipients. Patients received HT as a form of treatment for Chagas cardiomyopathy.
We described the laboratory data from the first day of hospitalization in the ICU due to COVID-19 until the time of death. Both patients were admitted to a referral center for treatment for COVID-19 in the metropolitan region of São Paulo, a city in southeastern Brazil. We hypothesize that the triple infection by SARS-CoV-2 and CMV may have been an important cause of death and of the worsening in CD patients with HT. To date, there have been no similar reports of patients presenting these three concomitant infectious diseases and HT receptors.
COVID-19 infection among transplant recipients increases the potential for developing severe illness [13] and may vary between different organ transplants [14,15].
SARS-CoV-2 entry´s receptor angiotensin converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2) are expressed in different tissues, such as the lungs, heart, liver, kidneys, testicles, thyroid, and adipose tissue [16]. Some patients with COVID-19 develop severe disease characterized by respiratory distress syndrome and systemic manifestations. This condition has been associated with the dysregulated release of pro-inflammatory cytokines, termed “cytokine storm,” which may induce multi-organ failure [1,17].
The use of immunosuppressants and post-surgical opportunistic infections can also lead to damage of multisystem organs or even death [18]. There is a therapeutic paradox here, because while insufficient immunosuppression results in graft loss due to rejection, excessive immunosuppression can result in serious infection, including SARS-COV-2 [13], besides contributing to the reactivation of pathogens [19].
Long-term administration of immunosuppressants to solid organ transplant (SOT) receptors to reduce the risk of graft rejection may increase the risk of respiratory infections [20], although there is no clear clinical evidence of increased morbidity/mortality in SARS-CoV-2 infection [21].
Fernandez-Ruiz and collaborators [22] described a cohort of SOT receptors affected by COVID-19, 44% of whom were kidney transplant recipients, 33.3% liver transplant recipients, and 22.2% heart transplant recipients. The lethality rate was 27.8%, suggesting that SARS-CoV-2 infection had a severe course in SOT recipients.
Recently, we described the first patients with CD affected by SARS-CoV-2. The CD patients presented an increase in COVID-19 laboratory hallmarks and a rapid disease progression. Despite the efforts of the health staff, both patients died [3].
HT may be associated with the reactivation of pathogens, as reported in CD [10]. In a Brazilian cohort, the reactivation rate of Chagas disease after heart transplantation was reported to be 38.8% [23]. This fact made HT as a treatment for Chagas cardiomyopathy initially controversial. However, currently, especially in endemic countries, it is the most viable therapeutic option for patients with end-stage heart failure [24]. We hypothesize that, in the cases presented here, HT followed by reactivation of CD conferred an additional risk factor for the worsening of COVID-19.
Transplant recipients are also commonly affected by CMV reactivation, being associated with significant morbidity and mortality [12] that may worsen the infectious condition of COVID-19 [25]. In SOT, risk factors include the use of immunosuppressants for transplantation, advanced age, acute rejection, and other concomitant infections [11], with all these characteristics being present in both patients described herein. In the present report, we observed that immunosuppression may have contributed to the susceptibility to superinfections and more severe clinical manifestations in individuals undergoing HT.
Since overactivated immune responses can be one of the causes of organ damage, the anti-inflammatory effects of immunosuppression can be protective, reducing the cytokine storm related to complications in COVID-19 [26]. In this context, it has been described that immunosuppressive therapy with calcineurin inhibitors in patients with solid organ transplantation or systemic rheumatic diseases promotes a clinical course in SARS-CoV-2 infection, which is generally mild, and with an apparently low risk of superinfection [27]. In addition, immunosuppression has not been evaluated as a risk factor for SARS or MERS [28].
However, in the present case report, we observed that immunosuppression may have contributed to the susceptibility to SARS-CoV-2 infection, the reactivation of pathogens, and more serious clinical manifestations in individuals undergoing HT. Corroborating our findings, it was shown that patients with COVID-19 and cancer, due to their systemic immunosuppressive condition caused by malignancy and anticancer treatments, such as chemotherapy, had an increased risk of SARS-CoV-2 infection and a worse prognosis [29]. It has also been reported that chronic use of corticosteroids prior to SARS-CoV-2 contamination is associated with critical disease outcomes, including a high risk of death [30].
These contradictory observations show that knowledge about the relationship between COVID-19 and the patient’s immune condition is limited. Further studies are needed to elucidate the immune responses and prognosis of COVID-19.
In general, HT in patient 1 proved to be successful for the treatment of Chagas cardiomyopathy, with no reactivation of the pathogen. However, after SARS-CoV-2 infection, she presented graft rejection, treated with methylprednisolone, thymoglobulin, and plasmapheresis. There is a description in eye transplantation that COVID-19 infection can compromise the balance of immunoregulatory responses that allow graft survival, contributing to rejection in individuals infected with SARS-CoV-2 [31]. Thus, this change in the balance of attenuation of the immune response, such as a reduction in the number of regulatory T cells (Tregs) [32], may favor direct cardiotoxic action and multiple organ dysfunction.
During postoperative hospitalization, patient 2 presented Chagas reactivation, evidenced by skin biopsy, indicating that the CD was not completely controlled. In addition, it is important to consider that immunosuppressive therapy can contribute to the reactivation of CD [33]. After HT, the patient presented graft rejection and was treated with plasmapheresis and methylprednisolone. With this, before the SARS-CoV-2 infection, the patient was already weakened and died 21 days after the infection, in relation to patient 1 who died after 47 days of infection.
4. Conclusions
This report highlights the first case of an association between COVID-19, CD, and CMV dissemination in HT recipients. The patients had rapid disease progression to death. We believe that HT and the usage of immunosuppressive drugs, as well as immunosuppression generated by concomitant infections, may be an important risk factor for the development of severe COVID-19, especially in endemic areas with underreported CD infection.
Author Contributions
Conceptualization, S.C.G.-S., G.B., and R.W.A.; investigation, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., and S.M.; resources, M.N.S. and A.J.d.S.D.; writing—original draft preparation, S.C.G.-S., G.B., and R.W.A.; writing—review and editing, S.C.G.-S., G.B., and R.W.A.; visualization, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., S.M., A.J.d.S.D., and M.N.S.; supervision, M.N.S.; funding acquisition, A.J.d.S.D. and M.N.S. All authors have read and agreed to the published version of the manuscript.
Funding
This research was funded by Fundação de Amparo à Pesquisa (FAPESP), grant numbers 2019/22448-0, 19/02679-7, 2017/18199-9, 2018/18230-6, and 2019/26928-6. Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, CAPES:88887.503842/2020-00.
Institutional Review Board Statement
This study was approved by the local ethics committee (HCFMUSP nº 30800520.7.0000.0068-2020) and was carried out in accordance with the 2013 revision of the Declaration of Helsinki.
Informed Consent Statement
Informed consent was obtained from all of the subjects involved in the study.
Data Availability Statement
The laboratory data and imaging exams presented herein came from Hospital das Clinics, Faculty of Medicine, University of São Paulo (HCFMUSP) with prior approval for their use.
Conflicts of Interest
The authors declare no conflict of interest.
Figure 1 Chest X-ray of (a) patient 1 from May 2020 and (b) patient 2 from June 2015, taken just before heart transplantation, showing the Chagasic cardiomyopathy.
Figure 2 Daily clinical features of patients, from the first day of hospitalization in the ICU until death. Blood levels of (A) erythrocytes, (B) hemoglobin, (C) leukocytes, (D) neutrophils, (E) eosinophils, (F) lymphocytes, (G) monocytes, (H) neutrophil-to-lymphocyte ratio, (I) platelets, (J) creatinine, (K) urea, (L) prothrombin time, (M) activated partial thromboplastin time, (N) D-dimer, (O) pH, (P) pO2, (Q) pCO2, (R) oxygen peripheral, (S) CPR, (T) glucose, (U) CK-MB, (V) NT-proBNP, (W) lactate dehydrogenase, and (X) troponin T. Gray boxes represent the reference values.
tropicalmed-06-00022-t001_Table 1 Table 1 Patients’ characteristics, comorbidities, treatment, and complications.
Patient 1 Patient 2
Sex Female Male
Age 55 62
Positive SARS-CoV-2 RT-PCR date 1 June 2020 29 June 2020
Heart transplant date 2 May 2020 5 December 2019
Death date 30 days 207 days
Days between positive SARS-CoV-2 diagnosis and death 47 days 21 days
Complications during ICU stay ARDS due to COVID–19; heart transplant rejection, disseminated cytomegalovirus; aggravated chronic kidney disease and pressure ulcer ARDS due to COVID–19, disseminated cytomegalovirus and pancytopenia due to hemophagocytosis
Previous comorbidities Dilated cardiomyopathy of Chagas etiology, hypothyroidism by thyroidectomy by nodule 10 years ago Cardiomyopathy of Chagas etiology, disseminated cytomegalovirus, deep vein thrombosis, systemic arterial hypertension, diabetes, dyslipidemia, and chronic renal failure
Previous use of medications Carvedilol, losartan, furosemide, levothyroxine, isosorbide, and hydralazine Unknown
Medicines used during ICU stay Methylprednisolone, azathioprine, anti-thymocyte globulin, cyclosporine, tacrolimus, meropenem, linezolid, micafungin, vancomycin, polymyxin B, tigecycline, amikacin, fluconazole, ganciclovir, and hydrocortisone Meropenem, colistin, linezolid
fluconazole, amikacin, cyclosporine, azathioprine, and prednisone
ARD, acute respiratory distress syndrome; COVID-19, coronavirus disease 2019; ICU, intensive care unit; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. | UNK (PULSE METHYLPREDNISOLONE) | DrugDosageText | CC BY | 33579042 | 19,731,501 | 2021-02-10 |
What was the dosage of drug 'METHYLPREDNISOLONE'? | SARS-CoV-2 Infection and CMV Dissemination in Transplant Recipients as a Treatment for Chagas Cardiomyopathy: A Case Report.
Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 has infected over 90 million people worldwide, therefore it is considered a pandemic. SARS-CoV-2 infection can lead to severe pneumonia, acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure. Individuals receiving a heart transplantation (HT) may be at higher risk of adverse outcomes attributable to COVID-19 due to immunosuppressives, as well as concomitant infections that may also influence the prognoses. Herein, we describe the first report of two cases of HT recipients with concomitant infections by SARS-CoV-2, Trypanosoma cruzi, and cytomegalovirus (CMV) dissemination, from the first day of hospitalization due to COVID-19 in the intensive care unit (ICU) until the death of the patients.
1. Introduction
Coronavirus disease 2019 (COVID-19), an infectious respiratory disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spread on a pandemic scale since the first case was reported in Wuhan, China, in 2019 [1]. Most patients with the disease have mild-to-moderate symptoms; however, approximately 15% develop severe pneumonia, while approximately 5% develop acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure [2]. Lymphopenia is a recurrent feature in these patients, with a significant reduction in CD4+ T cells, CD8+ T cells, B cells, and natural killer (NK) cells [3], increasing the susceptibility of patients to severe illness and co-infection [4]. In fact, co-infections are associated with worsening of the clinical condition [5].
Chagas disease (CD) is a zoonosis whose etiologic agent is the protozoan Trypanosoma cruzi. It is estimated that 6–8 million people worldwide are infected [6]. CD has two distinct phases: an acute one, which is rare, with a strong production of type 1 cytokines, and a chronic phase, which develops in 30–40% of CD cases. The chronic phase of CD can be characterized by cardiomyopathy, arrhythmias, megaviscera, and, more rarely, polyneuropathy and stroke [7]. TCD4+ and TCD8+ lymphocytes are the main cells responsible for controlling parasitic infection. However, the immune response also contributes to tissue damage and pathology [8].
Chagasic cardiomyopathy represents the main cause of mortality from this disease, which can lead to heart failure, whose indication for treatment, especially in endemic countries, may include heart transplantation (HT) as a strategy to curb the evolution of this complication [6,9]. However, the immunosuppression and possible reactivation of the causative agent T. cruzi following HT require intensive clinical care and laboratory monitoring [10].
Other infections such as that caused by cytomegalovirus (CMV), a herpes virus that infects up to 60–100% of people in adulthood, are associated with transplant complications [11]. Primary CMV infection is generally asymptomatic in immunocompetent individuals, with the virus generating a latent infection. On the contrary, in immunocompromised and immunosuppressed populations, such as solid organ transplantation, hematopoietic stem cell transplantation, and HIV/AIDS patients, CMV reactivation is responsible for significant morbidity and mortality [12]. Although reactivation of infections such as CMV and CD are often described in transplant recipients, there are no reports of concomitant infection by T. cruzi, CMV, and SARS-CoV-2 in the context of HT in the literature to date.
Therefore, in this report, we investigated the progress of two patients who underwent HT at the Heart Institute of Hospital das Clínicas (Incor) and were subsequently transferred to the special intensive care unit (ICU) of the Hospital das Clínicas (Hospital das Clínicas, Faculty of Medicine, University of São Paulo-HCFMUSP) due to SARS-CoV-2 infection. These patients were diagnosed with COVID-19 by nasopharyngeal detection of SARS-CoV-2 RNA using reverse transcriptase polymerase chain reaction (RT-PCR). In addition, during hospitalization, CMV dissemination was evidenced by quantitative DNA detection in the blood. We describe herein the laboratory data from the first day of hospitalization due to COVID-19 until the death of the patients.
The baseline characteristics of these two patients, as well as their past clinical data, are summarized in Table 1.
2. Case Report
2.1. Patient 1
Female, 55 years old, presenting positive serology for CD since 2015, received HT on 2 May as a treatment for Chagas cardiomyopathy (Figure 1A) and underwent immunosuppressive therapy with methylprednisolone and azathioprine. Post-transplantation, she developed pneumonia, treated with meropenem and linezolid, and a remittent Candida tropicalis infection, treated with micafungin, with improvement. However, on 29 May, she developed dyspnea and desaturation, with a chest tomography suggestive of COVID-19. Her polymerase chain reaction (PCR) for SARS-CoV-2 was positive on 1 June.
On 3 June, she was transferred to the ICU specialized for treatment of severe SARS-CoV-2 infections. Laboratory analyses then showed that the patient had a reduced number of erythrocytes and hemoglobin level, and these reductions became more accentuated by day 33 in the ICU (Figure 2A,B). On this same day, the number of neutrophils peaked (Figure 2D). From the 4th to the 16th day in ICU, important lymphopenia developed (Figure 2F), resulting in an increase in the neutrophil-to-lymphocyte ratio in the same period (Figure 2H). Additionally, from the eighth day onward, she presented severe thrombocytopenia that persisted until death (Figure 2I).
Throughout the ICU hospitalization period, there were sustained high levels of creatinine, urea, D-dimer, C-reactive protein (CRP), and lactate dehydrogenase (Figure 2J,K,N,S,W). On the 32nd day, she presented a sharp increase in prothrombin time and activated partial thromboplastin time.
The cardiac function markers of creatine kinase myocardial band (CK-MB) and troponin remained elevated from the first days of ICU admission until death (Figure 2U,X). On the day of admission to the ICU, the N-terminal pro b-type natriuretic peptide (NT pro-BNP) was 70,000 pg/mL (reference value of <125 pg/mL) (Figure 2V). On the third day of hospitalization, NT pro-BNP (155,117 pg/mL) and troponin (0.28 ng/mL, reference value of <0.014 ng/mL) peaked, being related to the period in which she presented signs of graft rejection, which was treated with pulse methylprednisolone, thymoglobulin, and plasmapheresis. In addition, disseminated CMV infection was diagnosed (RT-PCR viral loads of 122 IU/mL on day 10 and 54 IU/mL on day 19 of hospitalization), for which she received ganciclovir.
After 47 days after diagnosis of SARS-CoV-2, the patient died (18 July) due to multiple organ dysfunctions associated with COVID-19.
2.2. Patient 2
Male, 62 years old, previously diagnosed with CD, received HT on 5 December 2019 as treatment for Chagas cardiomyopathy (Figure 1B). He underwent immunosuppressive therapy with cyclosporine, azathioprine, and prednisone. During postoperative hospitalization he presented Chagas reactivation characterized by skin biopsy and humoral graft rejection, being treated with plasmapheresis and methylprednisolone.
The patient was admitted to the ICU of the Hospital das Clínicas on 8 June, presenting cellulitis, deep vein thrombosis, and Chagas reactivation (a lower limb chagoma). The latter was treated with benznidazol. During hospitalization, he tested positive for PCR of SARS-CoV-2 on 29 June. He developed ARDS and septic and cardiogenic shock, which were the causes of his death.
Throughout the ICU hospitalization period, he maintained decreased levels of erythrocytes, hemoglobin, and lymphocytes (Figure 2A,B,F), as well as a high neutrophil/lymphocyte ratio (Figure 2H). From the 12th day, the number of platelets decreased and continued until the time of death (Figure 2I). The values of creatinine, urea, and glucose also remained high throughout the hospitalization period (Figure 2J,K,T). In addition, disseminated CMV (viral loads of 711 IU/mL detected on the 28th day and 1477 IU/mL on the 35th day of hospitalization) was diagnosed and treated with ganciclovir.
One day before death, the following laboratory parameters peaked: D-dimer (7612 ng/mL FEU, reference value of <500 ng/mL FEU), CRP (280.6 ng/mL, reference value of 0.300 ng/mL), CK-MB (8.38 ng/mL, reference value of 0.10–4.94 ng/mL), NT pro-BNP (55,393 pg/mL), lactate dehydrogenase (1161 U/L, reference value of 135–225 U/L), and troponin (0.701 ng/mL) (Figure 2N,S,U–X).
On 20 July, 36 days after admission to the ICU and 21 days after a positive COVID-19 diagnosis, the patient died.
3. Discussion
Herein, we described the first report of triple infection (SARS-CoV-2 infection, T. cruzi infection, and CMV dissemination) in HT recipients. Patients received HT as a form of treatment for Chagas cardiomyopathy.
We described the laboratory data from the first day of hospitalization in the ICU due to COVID-19 until the time of death. Both patients were admitted to a referral center for treatment for COVID-19 in the metropolitan region of São Paulo, a city in southeastern Brazil. We hypothesize that the triple infection by SARS-CoV-2 and CMV may have been an important cause of death and of the worsening in CD patients with HT. To date, there have been no similar reports of patients presenting these three concomitant infectious diseases and HT receptors.
COVID-19 infection among transplant recipients increases the potential for developing severe illness [13] and may vary between different organ transplants [14,15].
SARS-CoV-2 entry´s receptor angiotensin converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2) are expressed in different tissues, such as the lungs, heart, liver, kidneys, testicles, thyroid, and adipose tissue [16]. Some patients with COVID-19 develop severe disease characterized by respiratory distress syndrome and systemic manifestations. This condition has been associated with the dysregulated release of pro-inflammatory cytokines, termed “cytokine storm,” which may induce multi-organ failure [1,17].
The use of immunosuppressants and post-surgical opportunistic infections can also lead to damage of multisystem organs or even death [18]. There is a therapeutic paradox here, because while insufficient immunosuppression results in graft loss due to rejection, excessive immunosuppression can result in serious infection, including SARS-COV-2 [13], besides contributing to the reactivation of pathogens [19].
Long-term administration of immunosuppressants to solid organ transplant (SOT) receptors to reduce the risk of graft rejection may increase the risk of respiratory infections [20], although there is no clear clinical evidence of increased morbidity/mortality in SARS-CoV-2 infection [21].
Fernandez-Ruiz and collaborators [22] described a cohort of SOT receptors affected by COVID-19, 44% of whom were kidney transplant recipients, 33.3% liver transplant recipients, and 22.2% heart transplant recipients. The lethality rate was 27.8%, suggesting that SARS-CoV-2 infection had a severe course in SOT recipients.
Recently, we described the first patients with CD affected by SARS-CoV-2. The CD patients presented an increase in COVID-19 laboratory hallmarks and a rapid disease progression. Despite the efforts of the health staff, both patients died [3].
HT may be associated with the reactivation of pathogens, as reported in CD [10]. In a Brazilian cohort, the reactivation rate of Chagas disease after heart transplantation was reported to be 38.8% [23]. This fact made HT as a treatment for Chagas cardiomyopathy initially controversial. However, currently, especially in endemic countries, it is the most viable therapeutic option for patients with end-stage heart failure [24]. We hypothesize that, in the cases presented here, HT followed by reactivation of CD conferred an additional risk factor for the worsening of COVID-19.
Transplant recipients are also commonly affected by CMV reactivation, being associated with significant morbidity and mortality [12] that may worsen the infectious condition of COVID-19 [25]. In SOT, risk factors include the use of immunosuppressants for transplantation, advanced age, acute rejection, and other concomitant infections [11], with all these characteristics being present in both patients described herein. In the present report, we observed that immunosuppression may have contributed to the susceptibility to superinfections and more severe clinical manifestations in individuals undergoing HT.
Since overactivated immune responses can be one of the causes of organ damage, the anti-inflammatory effects of immunosuppression can be protective, reducing the cytokine storm related to complications in COVID-19 [26]. In this context, it has been described that immunosuppressive therapy with calcineurin inhibitors in patients with solid organ transplantation or systemic rheumatic diseases promotes a clinical course in SARS-CoV-2 infection, which is generally mild, and with an apparently low risk of superinfection [27]. In addition, immunosuppression has not been evaluated as a risk factor for SARS or MERS [28].
However, in the present case report, we observed that immunosuppression may have contributed to the susceptibility to SARS-CoV-2 infection, the reactivation of pathogens, and more serious clinical manifestations in individuals undergoing HT. Corroborating our findings, it was shown that patients with COVID-19 and cancer, due to their systemic immunosuppressive condition caused by malignancy and anticancer treatments, such as chemotherapy, had an increased risk of SARS-CoV-2 infection and a worse prognosis [29]. It has also been reported that chronic use of corticosteroids prior to SARS-CoV-2 contamination is associated with critical disease outcomes, including a high risk of death [30].
These contradictory observations show that knowledge about the relationship between COVID-19 and the patient’s immune condition is limited. Further studies are needed to elucidate the immune responses and prognosis of COVID-19.
In general, HT in patient 1 proved to be successful for the treatment of Chagas cardiomyopathy, with no reactivation of the pathogen. However, after SARS-CoV-2 infection, she presented graft rejection, treated with methylprednisolone, thymoglobulin, and plasmapheresis. There is a description in eye transplantation that COVID-19 infection can compromise the balance of immunoregulatory responses that allow graft survival, contributing to rejection in individuals infected with SARS-CoV-2 [31]. Thus, this change in the balance of attenuation of the immune response, such as a reduction in the number of regulatory T cells (Tregs) [32], may favor direct cardiotoxic action and multiple organ dysfunction.
During postoperative hospitalization, patient 2 presented Chagas reactivation, evidenced by skin biopsy, indicating that the CD was not completely controlled. In addition, it is important to consider that immunosuppressive therapy can contribute to the reactivation of CD [33]. After HT, the patient presented graft rejection and was treated with plasmapheresis and methylprednisolone. With this, before the SARS-CoV-2 infection, the patient was already weakened and died 21 days after the infection, in relation to patient 1 who died after 47 days of infection.
4. Conclusions
This report highlights the first case of an association between COVID-19, CD, and CMV dissemination in HT recipients. The patients had rapid disease progression to death. We believe that HT and the usage of immunosuppressive drugs, as well as immunosuppression generated by concomitant infections, may be an important risk factor for the development of severe COVID-19, especially in endemic areas with underreported CD infection.
Author Contributions
Conceptualization, S.C.G.-S., G.B., and R.W.A.; investigation, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., and S.M.; resources, M.N.S. and A.J.d.S.D.; writing—original draft preparation, S.C.G.-S., G.B., and R.W.A.; writing—review and editing, S.C.G.-S., G.B., and R.W.A.; visualization, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., S.M., A.J.d.S.D., and M.N.S.; supervision, M.N.S.; funding acquisition, A.J.d.S.D. and M.N.S. All authors have read and agreed to the published version of the manuscript.
Funding
This research was funded by Fundação de Amparo à Pesquisa (FAPESP), grant numbers 2019/22448-0, 19/02679-7, 2017/18199-9, 2018/18230-6, and 2019/26928-6. Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, CAPES:88887.503842/2020-00.
Institutional Review Board Statement
This study was approved by the local ethics committee (HCFMUSP nº 30800520.7.0000.0068-2020) and was carried out in accordance with the 2013 revision of the Declaration of Helsinki.
Informed Consent Statement
Informed consent was obtained from all of the subjects involved in the study.
Data Availability Statement
The laboratory data and imaging exams presented herein came from Hospital das Clinics, Faculty of Medicine, University of São Paulo (HCFMUSP) with prior approval for their use.
Conflicts of Interest
The authors declare no conflict of interest.
Figure 1 Chest X-ray of (a) patient 1 from May 2020 and (b) patient 2 from June 2015, taken just before heart transplantation, showing the Chagasic cardiomyopathy.
Figure 2 Daily clinical features of patients, from the first day of hospitalization in the ICU until death. Blood levels of (A) erythrocytes, (B) hemoglobin, (C) leukocytes, (D) neutrophils, (E) eosinophils, (F) lymphocytes, (G) monocytes, (H) neutrophil-to-lymphocyte ratio, (I) platelets, (J) creatinine, (K) urea, (L) prothrombin time, (M) activated partial thromboplastin time, (N) D-dimer, (O) pH, (P) pO2, (Q) pCO2, (R) oxygen peripheral, (S) CPR, (T) glucose, (U) CK-MB, (V) NT-proBNP, (W) lactate dehydrogenase, and (X) troponin T. Gray boxes represent the reference values.
tropicalmed-06-00022-t001_Table 1 Table 1 Patients’ characteristics, comorbidities, treatment, and complications.
Patient 1 Patient 2
Sex Female Male
Age 55 62
Positive SARS-CoV-2 RT-PCR date 1 June 2020 29 June 2020
Heart transplant date 2 May 2020 5 December 2019
Death date 30 days 207 days
Days between positive SARS-CoV-2 diagnosis and death 47 days 21 days
Complications during ICU stay ARDS due to COVID–19; heart transplant rejection, disseminated cytomegalovirus; aggravated chronic kidney disease and pressure ulcer ARDS due to COVID–19, disseminated cytomegalovirus and pancytopenia due to hemophagocytosis
Previous comorbidities Dilated cardiomyopathy of Chagas etiology, hypothyroidism by thyroidectomy by nodule 10 years ago Cardiomyopathy of Chagas etiology, disseminated cytomegalovirus, deep vein thrombosis, systemic arterial hypertension, diabetes, dyslipidemia, and chronic renal failure
Previous use of medications Carvedilol, losartan, furosemide, levothyroxine, isosorbide, and hydralazine Unknown
Medicines used during ICU stay Methylprednisolone, azathioprine, anti-thymocyte globulin, cyclosporine, tacrolimus, meropenem, linezolid, micafungin, vancomycin, polymyxin B, tigecycline, amikacin, fluconazole, ganciclovir, and hydrocortisone Meropenem, colistin, linezolid
fluconazole, amikacin, cyclosporine, azathioprine, and prednisone
ARD, acute respiratory distress syndrome; COVID-19, coronavirus disease 2019; ICU, intensive care unit; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. | PULSE METHYLPREDNISOLONE | DrugDosageText | CC BY | 33579042 | 19,816,778 | 2021-02-10 |
What was the outcome of reaction 'Acute respiratory distress syndrome'? | SARS-CoV-2 Infection and CMV Dissemination in Transplant Recipients as a Treatment for Chagas Cardiomyopathy: A Case Report.
Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 has infected over 90 million people worldwide, therefore it is considered a pandemic. SARS-CoV-2 infection can lead to severe pneumonia, acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure. Individuals receiving a heart transplantation (HT) may be at higher risk of adverse outcomes attributable to COVID-19 due to immunosuppressives, as well as concomitant infections that may also influence the prognoses. Herein, we describe the first report of two cases of HT recipients with concomitant infections by SARS-CoV-2, Trypanosoma cruzi, and cytomegalovirus (CMV) dissemination, from the first day of hospitalization due to COVID-19 in the intensive care unit (ICU) until the death of the patients.
1. Introduction
Coronavirus disease 2019 (COVID-19), an infectious respiratory disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spread on a pandemic scale since the first case was reported in Wuhan, China, in 2019 [1]. Most patients with the disease have mild-to-moderate symptoms; however, approximately 15% develop severe pneumonia, while approximately 5% develop acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure [2]. Lymphopenia is a recurrent feature in these patients, with a significant reduction in CD4+ T cells, CD8+ T cells, B cells, and natural killer (NK) cells [3], increasing the susceptibility of patients to severe illness and co-infection [4]. In fact, co-infections are associated with worsening of the clinical condition [5].
Chagas disease (CD) is a zoonosis whose etiologic agent is the protozoan Trypanosoma cruzi. It is estimated that 6–8 million people worldwide are infected [6]. CD has two distinct phases: an acute one, which is rare, with a strong production of type 1 cytokines, and a chronic phase, which develops in 30–40% of CD cases. The chronic phase of CD can be characterized by cardiomyopathy, arrhythmias, megaviscera, and, more rarely, polyneuropathy and stroke [7]. TCD4+ and TCD8+ lymphocytes are the main cells responsible for controlling parasitic infection. However, the immune response also contributes to tissue damage and pathology [8].
Chagasic cardiomyopathy represents the main cause of mortality from this disease, which can lead to heart failure, whose indication for treatment, especially in endemic countries, may include heart transplantation (HT) as a strategy to curb the evolution of this complication [6,9]. However, the immunosuppression and possible reactivation of the causative agent T. cruzi following HT require intensive clinical care and laboratory monitoring [10].
Other infections such as that caused by cytomegalovirus (CMV), a herpes virus that infects up to 60–100% of people in adulthood, are associated with transplant complications [11]. Primary CMV infection is generally asymptomatic in immunocompetent individuals, with the virus generating a latent infection. On the contrary, in immunocompromised and immunosuppressed populations, such as solid organ transplantation, hematopoietic stem cell transplantation, and HIV/AIDS patients, CMV reactivation is responsible for significant morbidity and mortality [12]. Although reactivation of infections such as CMV and CD are often described in transplant recipients, there are no reports of concomitant infection by T. cruzi, CMV, and SARS-CoV-2 in the context of HT in the literature to date.
Therefore, in this report, we investigated the progress of two patients who underwent HT at the Heart Institute of Hospital das Clínicas (Incor) and were subsequently transferred to the special intensive care unit (ICU) of the Hospital das Clínicas (Hospital das Clínicas, Faculty of Medicine, University of São Paulo-HCFMUSP) due to SARS-CoV-2 infection. These patients were diagnosed with COVID-19 by nasopharyngeal detection of SARS-CoV-2 RNA using reverse transcriptase polymerase chain reaction (RT-PCR). In addition, during hospitalization, CMV dissemination was evidenced by quantitative DNA detection in the blood. We describe herein the laboratory data from the first day of hospitalization due to COVID-19 until the death of the patients.
The baseline characteristics of these two patients, as well as their past clinical data, are summarized in Table 1.
2. Case Report
2.1. Patient 1
Female, 55 years old, presenting positive serology for CD since 2015, received HT on 2 May as a treatment for Chagas cardiomyopathy (Figure 1A) and underwent immunosuppressive therapy with methylprednisolone and azathioprine. Post-transplantation, she developed pneumonia, treated with meropenem and linezolid, and a remittent Candida tropicalis infection, treated with micafungin, with improvement. However, on 29 May, she developed dyspnea and desaturation, with a chest tomography suggestive of COVID-19. Her polymerase chain reaction (PCR) for SARS-CoV-2 was positive on 1 June.
On 3 June, she was transferred to the ICU specialized for treatment of severe SARS-CoV-2 infections. Laboratory analyses then showed that the patient had a reduced number of erythrocytes and hemoglobin level, and these reductions became more accentuated by day 33 in the ICU (Figure 2A,B). On this same day, the number of neutrophils peaked (Figure 2D). From the 4th to the 16th day in ICU, important lymphopenia developed (Figure 2F), resulting in an increase in the neutrophil-to-lymphocyte ratio in the same period (Figure 2H). Additionally, from the eighth day onward, she presented severe thrombocytopenia that persisted until death (Figure 2I).
Throughout the ICU hospitalization period, there were sustained high levels of creatinine, urea, D-dimer, C-reactive protein (CRP), and lactate dehydrogenase (Figure 2J,K,N,S,W). On the 32nd day, she presented a sharp increase in prothrombin time and activated partial thromboplastin time.
The cardiac function markers of creatine kinase myocardial band (CK-MB) and troponin remained elevated from the first days of ICU admission until death (Figure 2U,X). On the day of admission to the ICU, the N-terminal pro b-type natriuretic peptide (NT pro-BNP) was 70,000 pg/mL (reference value of <125 pg/mL) (Figure 2V). On the third day of hospitalization, NT pro-BNP (155,117 pg/mL) and troponin (0.28 ng/mL, reference value of <0.014 ng/mL) peaked, being related to the period in which she presented signs of graft rejection, which was treated with pulse methylprednisolone, thymoglobulin, and plasmapheresis. In addition, disseminated CMV infection was diagnosed (RT-PCR viral loads of 122 IU/mL on day 10 and 54 IU/mL on day 19 of hospitalization), for which she received ganciclovir.
After 47 days after diagnosis of SARS-CoV-2, the patient died (18 July) due to multiple organ dysfunctions associated with COVID-19.
2.2. Patient 2
Male, 62 years old, previously diagnosed with CD, received HT on 5 December 2019 as treatment for Chagas cardiomyopathy (Figure 1B). He underwent immunosuppressive therapy with cyclosporine, azathioprine, and prednisone. During postoperative hospitalization he presented Chagas reactivation characterized by skin biopsy and humoral graft rejection, being treated with plasmapheresis and methylprednisolone.
The patient was admitted to the ICU of the Hospital das Clínicas on 8 June, presenting cellulitis, deep vein thrombosis, and Chagas reactivation (a lower limb chagoma). The latter was treated with benznidazol. During hospitalization, he tested positive for PCR of SARS-CoV-2 on 29 June. He developed ARDS and septic and cardiogenic shock, which were the causes of his death.
Throughout the ICU hospitalization period, he maintained decreased levels of erythrocytes, hemoglobin, and lymphocytes (Figure 2A,B,F), as well as a high neutrophil/lymphocyte ratio (Figure 2H). From the 12th day, the number of platelets decreased and continued until the time of death (Figure 2I). The values of creatinine, urea, and glucose also remained high throughout the hospitalization period (Figure 2J,K,T). In addition, disseminated CMV (viral loads of 711 IU/mL detected on the 28th day and 1477 IU/mL on the 35th day of hospitalization) was diagnosed and treated with ganciclovir.
One day before death, the following laboratory parameters peaked: D-dimer (7612 ng/mL FEU, reference value of <500 ng/mL FEU), CRP (280.6 ng/mL, reference value of 0.300 ng/mL), CK-MB (8.38 ng/mL, reference value of 0.10–4.94 ng/mL), NT pro-BNP (55,393 pg/mL), lactate dehydrogenase (1161 U/L, reference value of 135–225 U/L), and troponin (0.701 ng/mL) (Figure 2N,S,U–X).
On 20 July, 36 days after admission to the ICU and 21 days after a positive COVID-19 diagnosis, the patient died.
3. Discussion
Herein, we described the first report of triple infection (SARS-CoV-2 infection, T. cruzi infection, and CMV dissemination) in HT recipients. Patients received HT as a form of treatment for Chagas cardiomyopathy.
We described the laboratory data from the first day of hospitalization in the ICU due to COVID-19 until the time of death. Both patients were admitted to a referral center for treatment for COVID-19 in the metropolitan region of São Paulo, a city in southeastern Brazil. We hypothesize that the triple infection by SARS-CoV-2 and CMV may have been an important cause of death and of the worsening in CD patients with HT. To date, there have been no similar reports of patients presenting these three concomitant infectious diseases and HT receptors.
COVID-19 infection among transplant recipients increases the potential for developing severe illness [13] and may vary between different organ transplants [14,15].
SARS-CoV-2 entry´s receptor angiotensin converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2) are expressed in different tissues, such as the lungs, heart, liver, kidneys, testicles, thyroid, and adipose tissue [16]. Some patients with COVID-19 develop severe disease characterized by respiratory distress syndrome and systemic manifestations. This condition has been associated with the dysregulated release of pro-inflammatory cytokines, termed “cytokine storm,” which may induce multi-organ failure [1,17].
The use of immunosuppressants and post-surgical opportunistic infections can also lead to damage of multisystem organs or even death [18]. There is a therapeutic paradox here, because while insufficient immunosuppression results in graft loss due to rejection, excessive immunosuppression can result in serious infection, including SARS-COV-2 [13], besides contributing to the reactivation of pathogens [19].
Long-term administration of immunosuppressants to solid organ transplant (SOT) receptors to reduce the risk of graft rejection may increase the risk of respiratory infections [20], although there is no clear clinical evidence of increased morbidity/mortality in SARS-CoV-2 infection [21].
Fernandez-Ruiz and collaborators [22] described a cohort of SOT receptors affected by COVID-19, 44% of whom were kidney transplant recipients, 33.3% liver transplant recipients, and 22.2% heart transplant recipients. The lethality rate was 27.8%, suggesting that SARS-CoV-2 infection had a severe course in SOT recipients.
Recently, we described the first patients with CD affected by SARS-CoV-2. The CD patients presented an increase in COVID-19 laboratory hallmarks and a rapid disease progression. Despite the efforts of the health staff, both patients died [3].
HT may be associated with the reactivation of pathogens, as reported in CD [10]. In a Brazilian cohort, the reactivation rate of Chagas disease after heart transplantation was reported to be 38.8% [23]. This fact made HT as a treatment for Chagas cardiomyopathy initially controversial. However, currently, especially in endemic countries, it is the most viable therapeutic option for patients with end-stage heart failure [24]. We hypothesize that, in the cases presented here, HT followed by reactivation of CD conferred an additional risk factor for the worsening of COVID-19.
Transplant recipients are also commonly affected by CMV reactivation, being associated with significant morbidity and mortality [12] that may worsen the infectious condition of COVID-19 [25]. In SOT, risk factors include the use of immunosuppressants for transplantation, advanced age, acute rejection, and other concomitant infections [11], with all these characteristics being present in both patients described herein. In the present report, we observed that immunosuppression may have contributed to the susceptibility to superinfections and more severe clinical manifestations in individuals undergoing HT.
Since overactivated immune responses can be one of the causes of organ damage, the anti-inflammatory effects of immunosuppression can be protective, reducing the cytokine storm related to complications in COVID-19 [26]. In this context, it has been described that immunosuppressive therapy with calcineurin inhibitors in patients with solid organ transplantation or systemic rheumatic diseases promotes a clinical course in SARS-CoV-2 infection, which is generally mild, and with an apparently low risk of superinfection [27]. In addition, immunosuppression has not been evaluated as a risk factor for SARS or MERS [28].
However, in the present case report, we observed that immunosuppression may have contributed to the susceptibility to SARS-CoV-2 infection, the reactivation of pathogens, and more serious clinical manifestations in individuals undergoing HT. Corroborating our findings, it was shown that patients with COVID-19 and cancer, due to their systemic immunosuppressive condition caused by malignancy and anticancer treatments, such as chemotherapy, had an increased risk of SARS-CoV-2 infection and a worse prognosis [29]. It has also been reported that chronic use of corticosteroids prior to SARS-CoV-2 contamination is associated with critical disease outcomes, including a high risk of death [30].
These contradictory observations show that knowledge about the relationship between COVID-19 and the patient’s immune condition is limited. Further studies are needed to elucidate the immune responses and prognosis of COVID-19.
In general, HT in patient 1 proved to be successful for the treatment of Chagas cardiomyopathy, with no reactivation of the pathogen. However, after SARS-CoV-2 infection, she presented graft rejection, treated with methylprednisolone, thymoglobulin, and plasmapheresis. There is a description in eye transplantation that COVID-19 infection can compromise the balance of immunoregulatory responses that allow graft survival, contributing to rejection in individuals infected with SARS-CoV-2 [31]. Thus, this change in the balance of attenuation of the immune response, such as a reduction in the number of regulatory T cells (Tregs) [32], may favor direct cardiotoxic action and multiple organ dysfunction.
During postoperative hospitalization, patient 2 presented Chagas reactivation, evidenced by skin biopsy, indicating that the CD was not completely controlled. In addition, it is important to consider that immunosuppressive therapy can contribute to the reactivation of CD [33]. After HT, the patient presented graft rejection and was treated with plasmapheresis and methylprednisolone. With this, before the SARS-CoV-2 infection, the patient was already weakened and died 21 days after the infection, in relation to patient 1 who died after 47 days of infection.
4. Conclusions
This report highlights the first case of an association between COVID-19, CD, and CMV dissemination in HT recipients. The patients had rapid disease progression to death. We believe that HT and the usage of immunosuppressive drugs, as well as immunosuppression generated by concomitant infections, may be an important risk factor for the development of severe COVID-19, especially in endemic areas with underreported CD infection.
Author Contributions
Conceptualization, S.C.G.-S., G.B., and R.W.A.; investigation, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., and S.M.; resources, M.N.S. and A.J.d.S.D.; writing—original draft preparation, S.C.G.-S., G.B., and R.W.A.; writing—review and editing, S.C.G.-S., G.B., and R.W.A.; visualization, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., S.M., A.J.d.S.D., and M.N.S.; supervision, M.N.S.; funding acquisition, A.J.d.S.D. and M.N.S. All authors have read and agreed to the published version of the manuscript.
Funding
This research was funded by Fundação de Amparo à Pesquisa (FAPESP), grant numbers 2019/22448-0, 19/02679-7, 2017/18199-9, 2018/18230-6, and 2019/26928-6. Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, CAPES:88887.503842/2020-00.
Institutional Review Board Statement
This study was approved by the local ethics committee (HCFMUSP nº 30800520.7.0000.0068-2020) and was carried out in accordance with the 2013 revision of the Declaration of Helsinki.
Informed Consent Statement
Informed consent was obtained from all of the subjects involved in the study.
Data Availability Statement
The laboratory data and imaging exams presented herein came from Hospital das Clinics, Faculty of Medicine, University of São Paulo (HCFMUSP) with prior approval for their use.
Conflicts of Interest
The authors declare no conflict of interest.
Figure 1 Chest X-ray of (a) patient 1 from May 2020 and (b) patient 2 from June 2015, taken just before heart transplantation, showing the Chagasic cardiomyopathy.
Figure 2 Daily clinical features of patients, from the first day of hospitalization in the ICU until death. Blood levels of (A) erythrocytes, (B) hemoglobin, (C) leukocytes, (D) neutrophils, (E) eosinophils, (F) lymphocytes, (G) monocytes, (H) neutrophil-to-lymphocyte ratio, (I) platelets, (J) creatinine, (K) urea, (L) prothrombin time, (M) activated partial thromboplastin time, (N) D-dimer, (O) pH, (P) pO2, (Q) pCO2, (R) oxygen peripheral, (S) CPR, (T) glucose, (U) CK-MB, (V) NT-proBNP, (W) lactate dehydrogenase, and (X) troponin T. Gray boxes represent the reference values.
tropicalmed-06-00022-t001_Table 1 Table 1 Patients’ characteristics, comorbidities, treatment, and complications.
Patient 1 Patient 2
Sex Female Male
Age 55 62
Positive SARS-CoV-2 RT-PCR date 1 June 2020 29 June 2020
Heart transplant date 2 May 2020 5 December 2019
Death date 30 days 207 days
Days between positive SARS-CoV-2 diagnosis and death 47 days 21 days
Complications during ICU stay ARDS due to COVID–19; heart transplant rejection, disseminated cytomegalovirus; aggravated chronic kidney disease and pressure ulcer ARDS due to COVID–19, disseminated cytomegalovirus and pancytopenia due to hemophagocytosis
Previous comorbidities Dilated cardiomyopathy of Chagas etiology, hypothyroidism by thyroidectomy by nodule 10 years ago Cardiomyopathy of Chagas etiology, disseminated cytomegalovirus, deep vein thrombosis, systemic arterial hypertension, diabetes, dyslipidemia, and chronic renal failure
Previous use of medications Carvedilol, losartan, furosemide, levothyroxine, isosorbide, and hydralazine Unknown
Medicines used during ICU stay Methylprednisolone, azathioprine, anti-thymocyte globulin, cyclosporine, tacrolimus, meropenem, linezolid, micafungin, vancomycin, polymyxin B, tigecycline, amikacin, fluconazole, ganciclovir, and hydrocortisone Meropenem, colistin, linezolid
fluconazole, amikacin, cyclosporine, azathioprine, and prednisone
ARD, acute respiratory distress syndrome; COVID-19, coronavirus disease 2019; ICU, intensive care unit; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. | Fatal | ReactionOutcome | CC BY | 33579042 | 19,681,859 | 2021-02-10 |
What was the outcome of reaction 'American trypanosomiasis'? | SARS-CoV-2 Infection and CMV Dissemination in Transplant Recipients as a Treatment for Chagas Cardiomyopathy: A Case Report.
Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 has infected over 90 million people worldwide, therefore it is considered a pandemic. SARS-CoV-2 infection can lead to severe pneumonia, acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure. Individuals receiving a heart transplantation (HT) may be at higher risk of adverse outcomes attributable to COVID-19 due to immunosuppressives, as well as concomitant infections that may also influence the prognoses. Herein, we describe the first report of two cases of HT recipients with concomitant infections by SARS-CoV-2, Trypanosoma cruzi, and cytomegalovirus (CMV) dissemination, from the first day of hospitalization due to COVID-19 in the intensive care unit (ICU) until the death of the patients.
1. Introduction
Coronavirus disease 2019 (COVID-19), an infectious respiratory disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spread on a pandemic scale since the first case was reported in Wuhan, China, in 2019 [1]. Most patients with the disease have mild-to-moderate symptoms; however, approximately 15% develop severe pneumonia, while approximately 5% develop acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure [2]. Lymphopenia is a recurrent feature in these patients, with a significant reduction in CD4+ T cells, CD8+ T cells, B cells, and natural killer (NK) cells [3], increasing the susceptibility of patients to severe illness and co-infection [4]. In fact, co-infections are associated with worsening of the clinical condition [5].
Chagas disease (CD) is a zoonosis whose etiologic agent is the protozoan Trypanosoma cruzi. It is estimated that 6–8 million people worldwide are infected [6]. CD has two distinct phases: an acute one, which is rare, with a strong production of type 1 cytokines, and a chronic phase, which develops in 30–40% of CD cases. The chronic phase of CD can be characterized by cardiomyopathy, arrhythmias, megaviscera, and, more rarely, polyneuropathy and stroke [7]. TCD4+ and TCD8+ lymphocytes are the main cells responsible for controlling parasitic infection. However, the immune response also contributes to tissue damage and pathology [8].
Chagasic cardiomyopathy represents the main cause of mortality from this disease, which can lead to heart failure, whose indication for treatment, especially in endemic countries, may include heart transplantation (HT) as a strategy to curb the evolution of this complication [6,9]. However, the immunosuppression and possible reactivation of the causative agent T. cruzi following HT require intensive clinical care and laboratory monitoring [10].
Other infections such as that caused by cytomegalovirus (CMV), a herpes virus that infects up to 60–100% of people in adulthood, are associated with transplant complications [11]. Primary CMV infection is generally asymptomatic in immunocompetent individuals, with the virus generating a latent infection. On the contrary, in immunocompromised and immunosuppressed populations, such as solid organ transplantation, hematopoietic stem cell transplantation, and HIV/AIDS patients, CMV reactivation is responsible for significant morbidity and mortality [12]. Although reactivation of infections such as CMV and CD are often described in transplant recipients, there are no reports of concomitant infection by T. cruzi, CMV, and SARS-CoV-2 in the context of HT in the literature to date.
Therefore, in this report, we investigated the progress of two patients who underwent HT at the Heart Institute of Hospital das Clínicas (Incor) and were subsequently transferred to the special intensive care unit (ICU) of the Hospital das Clínicas (Hospital das Clínicas, Faculty of Medicine, University of São Paulo-HCFMUSP) due to SARS-CoV-2 infection. These patients were diagnosed with COVID-19 by nasopharyngeal detection of SARS-CoV-2 RNA using reverse transcriptase polymerase chain reaction (RT-PCR). In addition, during hospitalization, CMV dissemination was evidenced by quantitative DNA detection in the blood. We describe herein the laboratory data from the first day of hospitalization due to COVID-19 until the death of the patients.
The baseline characteristics of these two patients, as well as their past clinical data, are summarized in Table 1.
2. Case Report
2.1. Patient 1
Female, 55 years old, presenting positive serology for CD since 2015, received HT on 2 May as a treatment for Chagas cardiomyopathy (Figure 1A) and underwent immunosuppressive therapy with methylprednisolone and azathioprine. Post-transplantation, she developed pneumonia, treated with meropenem and linezolid, and a remittent Candida tropicalis infection, treated with micafungin, with improvement. However, on 29 May, she developed dyspnea and desaturation, with a chest tomography suggestive of COVID-19. Her polymerase chain reaction (PCR) for SARS-CoV-2 was positive on 1 June.
On 3 June, she was transferred to the ICU specialized for treatment of severe SARS-CoV-2 infections. Laboratory analyses then showed that the patient had a reduced number of erythrocytes and hemoglobin level, and these reductions became more accentuated by day 33 in the ICU (Figure 2A,B). On this same day, the number of neutrophils peaked (Figure 2D). From the 4th to the 16th day in ICU, important lymphopenia developed (Figure 2F), resulting in an increase in the neutrophil-to-lymphocyte ratio in the same period (Figure 2H). Additionally, from the eighth day onward, she presented severe thrombocytopenia that persisted until death (Figure 2I).
Throughout the ICU hospitalization period, there were sustained high levels of creatinine, urea, D-dimer, C-reactive protein (CRP), and lactate dehydrogenase (Figure 2J,K,N,S,W). On the 32nd day, she presented a sharp increase in prothrombin time and activated partial thromboplastin time.
The cardiac function markers of creatine kinase myocardial band (CK-MB) and troponin remained elevated from the first days of ICU admission until death (Figure 2U,X). On the day of admission to the ICU, the N-terminal pro b-type natriuretic peptide (NT pro-BNP) was 70,000 pg/mL (reference value of <125 pg/mL) (Figure 2V). On the third day of hospitalization, NT pro-BNP (155,117 pg/mL) and troponin (0.28 ng/mL, reference value of <0.014 ng/mL) peaked, being related to the period in which she presented signs of graft rejection, which was treated with pulse methylprednisolone, thymoglobulin, and plasmapheresis. In addition, disseminated CMV infection was diagnosed (RT-PCR viral loads of 122 IU/mL on day 10 and 54 IU/mL on day 19 of hospitalization), for which she received ganciclovir.
After 47 days after diagnosis of SARS-CoV-2, the patient died (18 July) due to multiple organ dysfunctions associated with COVID-19.
2.2. Patient 2
Male, 62 years old, previously diagnosed with CD, received HT on 5 December 2019 as treatment for Chagas cardiomyopathy (Figure 1B). He underwent immunosuppressive therapy with cyclosporine, azathioprine, and prednisone. During postoperative hospitalization he presented Chagas reactivation characterized by skin biopsy and humoral graft rejection, being treated with plasmapheresis and methylprednisolone.
The patient was admitted to the ICU of the Hospital das Clínicas on 8 June, presenting cellulitis, deep vein thrombosis, and Chagas reactivation (a lower limb chagoma). The latter was treated with benznidazol. During hospitalization, he tested positive for PCR of SARS-CoV-2 on 29 June. He developed ARDS and septic and cardiogenic shock, which were the causes of his death.
Throughout the ICU hospitalization period, he maintained decreased levels of erythrocytes, hemoglobin, and lymphocytes (Figure 2A,B,F), as well as a high neutrophil/lymphocyte ratio (Figure 2H). From the 12th day, the number of platelets decreased and continued until the time of death (Figure 2I). The values of creatinine, urea, and glucose also remained high throughout the hospitalization period (Figure 2J,K,T). In addition, disseminated CMV (viral loads of 711 IU/mL detected on the 28th day and 1477 IU/mL on the 35th day of hospitalization) was diagnosed and treated with ganciclovir.
One day before death, the following laboratory parameters peaked: D-dimer (7612 ng/mL FEU, reference value of <500 ng/mL FEU), CRP (280.6 ng/mL, reference value of 0.300 ng/mL), CK-MB (8.38 ng/mL, reference value of 0.10–4.94 ng/mL), NT pro-BNP (55,393 pg/mL), lactate dehydrogenase (1161 U/L, reference value of 135–225 U/L), and troponin (0.701 ng/mL) (Figure 2N,S,U–X).
On 20 July, 36 days after admission to the ICU and 21 days after a positive COVID-19 diagnosis, the patient died.
3. Discussion
Herein, we described the first report of triple infection (SARS-CoV-2 infection, T. cruzi infection, and CMV dissemination) in HT recipients. Patients received HT as a form of treatment for Chagas cardiomyopathy.
We described the laboratory data from the first day of hospitalization in the ICU due to COVID-19 until the time of death. Both patients were admitted to a referral center for treatment for COVID-19 in the metropolitan region of São Paulo, a city in southeastern Brazil. We hypothesize that the triple infection by SARS-CoV-2 and CMV may have been an important cause of death and of the worsening in CD patients with HT. To date, there have been no similar reports of patients presenting these three concomitant infectious diseases and HT receptors.
COVID-19 infection among transplant recipients increases the potential for developing severe illness [13] and may vary between different organ transplants [14,15].
SARS-CoV-2 entry´s receptor angiotensin converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2) are expressed in different tissues, such as the lungs, heart, liver, kidneys, testicles, thyroid, and adipose tissue [16]. Some patients with COVID-19 develop severe disease characterized by respiratory distress syndrome and systemic manifestations. This condition has been associated with the dysregulated release of pro-inflammatory cytokines, termed “cytokine storm,” which may induce multi-organ failure [1,17].
The use of immunosuppressants and post-surgical opportunistic infections can also lead to damage of multisystem organs or even death [18]. There is a therapeutic paradox here, because while insufficient immunosuppression results in graft loss due to rejection, excessive immunosuppression can result in serious infection, including SARS-COV-2 [13], besides contributing to the reactivation of pathogens [19].
Long-term administration of immunosuppressants to solid organ transplant (SOT) receptors to reduce the risk of graft rejection may increase the risk of respiratory infections [20], although there is no clear clinical evidence of increased morbidity/mortality in SARS-CoV-2 infection [21].
Fernandez-Ruiz and collaborators [22] described a cohort of SOT receptors affected by COVID-19, 44% of whom were kidney transplant recipients, 33.3% liver transplant recipients, and 22.2% heart transplant recipients. The lethality rate was 27.8%, suggesting that SARS-CoV-2 infection had a severe course in SOT recipients.
Recently, we described the first patients with CD affected by SARS-CoV-2. The CD patients presented an increase in COVID-19 laboratory hallmarks and a rapid disease progression. Despite the efforts of the health staff, both patients died [3].
HT may be associated with the reactivation of pathogens, as reported in CD [10]. In a Brazilian cohort, the reactivation rate of Chagas disease after heart transplantation was reported to be 38.8% [23]. This fact made HT as a treatment for Chagas cardiomyopathy initially controversial. However, currently, especially in endemic countries, it is the most viable therapeutic option for patients with end-stage heart failure [24]. We hypothesize that, in the cases presented here, HT followed by reactivation of CD conferred an additional risk factor for the worsening of COVID-19.
Transplant recipients are also commonly affected by CMV reactivation, being associated with significant morbidity and mortality [12] that may worsen the infectious condition of COVID-19 [25]. In SOT, risk factors include the use of immunosuppressants for transplantation, advanced age, acute rejection, and other concomitant infections [11], with all these characteristics being present in both patients described herein. In the present report, we observed that immunosuppression may have contributed to the susceptibility to superinfections and more severe clinical manifestations in individuals undergoing HT.
Since overactivated immune responses can be one of the causes of organ damage, the anti-inflammatory effects of immunosuppression can be protective, reducing the cytokine storm related to complications in COVID-19 [26]. In this context, it has been described that immunosuppressive therapy with calcineurin inhibitors in patients with solid organ transplantation or systemic rheumatic diseases promotes a clinical course in SARS-CoV-2 infection, which is generally mild, and with an apparently low risk of superinfection [27]. In addition, immunosuppression has not been evaluated as a risk factor for SARS or MERS [28].
However, in the present case report, we observed that immunosuppression may have contributed to the susceptibility to SARS-CoV-2 infection, the reactivation of pathogens, and more serious clinical manifestations in individuals undergoing HT. Corroborating our findings, it was shown that patients with COVID-19 and cancer, due to their systemic immunosuppressive condition caused by malignancy and anticancer treatments, such as chemotherapy, had an increased risk of SARS-CoV-2 infection and a worse prognosis [29]. It has also been reported that chronic use of corticosteroids prior to SARS-CoV-2 contamination is associated with critical disease outcomes, including a high risk of death [30].
These contradictory observations show that knowledge about the relationship between COVID-19 and the patient’s immune condition is limited. Further studies are needed to elucidate the immune responses and prognosis of COVID-19.
In general, HT in patient 1 proved to be successful for the treatment of Chagas cardiomyopathy, with no reactivation of the pathogen. However, after SARS-CoV-2 infection, she presented graft rejection, treated with methylprednisolone, thymoglobulin, and plasmapheresis. There is a description in eye transplantation that COVID-19 infection can compromise the balance of immunoregulatory responses that allow graft survival, contributing to rejection in individuals infected with SARS-CoV-2 [31]. Thus, this change in the balance of attenuation of the immune response, such as a reduction in the number of regulatory T cells (Tregs) [32], may favor direct cardiotoxic action and multiple organ dysfunction.
During postoperative hospitalization, patient 2 presented Chagas reactivation, evidenced by skin biopsy, indicating that the CD was not completely controlled. In addition, it is important to consider that immunosuppressive therapy can contribute to the reactivation of CD [33]. After HT, the patient presented graft rejection and was treated with plasmapheresis and methylprednisolone. With this, before the SARS-CoV-2 infection, the patient was already weakened and died 21 days after the infection, in relation to patient 1 who died after 47 days of infection.
4. Conclusions
This report highlights the first case of an association between COVID-19, CD, and CMV dissemination in HT recipients. The patients had rapid disease progression to death. We believe that HT and the usage of immunosuppressive drugs, as well as immunosuppression generated by concomitant infections, may be an important risk factor for the development of severe COVID-19, especially in endemic areas with underreported CD infection.
Author Contributions
Conceptualization, S.C.G.-S., G.B., and R.W.A.; investigation, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., and S.M.; resources, M.N.S. and A.J.d.S.D.; writing—original draft preparation, S.C.G.-S., G.B., and R.W.A.; writing—review and editing, S.C.G.-S., G.B., and R.W.A.; visualization, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., S.M., A.J.d.S.D., and M.N.S.; supervision, M.N.S.; funding acquisition, A.J.d.S.D. and M.N.S. All authors have read and agreed to the published version of the manuscript.
Funding
This research was funded by Fundação de Amparo à Pesquisa (FAPESP), grant numbers 2019/22448-0, 19/02679-7, 2017/18199-9, 2018/18230-6, and 2019/26928-6. Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, CAPES:88887.503842/2020-00.
Institutional Review Board Statement
This study was approved by the local ethics committee (HCFMUSP nº 30800520.7.0000.0068-2020) and was carried out in accordance with the 2013 revision of the Declaration of Helsinki.
Informed Consent Statement
Informed consent was obtained from all of the subjects involved in the study.
Data Availability Statement
The laboratory data and imaging exams presented herein came from Hospital das Clinics, Faculty of Medicine, University of São Paulo (HCFMUSP) with prior approval for their use.
Conflicts of Interest
The authors declare no conflict of interest.
Figure 1 Chest X-ray of (a) patient 1 from May 2020 and (b) patient 2 from June 2015, taken just before heart transplantation, showing the Chagasic cardiomyopathy.
Figure 2 Daily clinical features of patients, from the first day of hospitalization in the ICU until death. Blood levels of (A) erythrocytes, (B) hemoglobin, (C) leukocytes, (D) neutrophils, (E) eosinophils, (F) lymphocytes, (G) monocytes, (H) neutrophil-to-lymphocyte ratio, (I) platelets, (J) creatinine, (K) urea, (L) prothrombin time, (M) activated partial thromboplastin time, (N) D-dimer, (O) pH, (P) pO2, (Q) pCO2, (R) oxygen peripheral, (S) CPR, (T) glucose, (U) CK-MB, (V) NT-proBNP, (W) lactate dehydrogenase, and (X) troponin T. Gray boxes represent the reference values.
tropicalmed-06-00022-t001_Table 1 Table 1 Patients’ characteristics, comorbidities, treatment, and complications.
Patient 1 Patient 2
Sex Female Male
Age 55 62
Positive SARS-CoV-2 RT-PCR date 1 June 2020 29 June 2020
Heart transplant date 2 May 2020 5 December 2019
Death date 30 days 207 days
Days between positive SARS-CoV-2 diagnosis and death 47 days 21 days
Complications during ICU stay ARDS due to COVID–19; heart transplant rejection, disseminated cytomegalovirus; aggravated chronic kidney disease and pressure ulcer ARDS due to COVID–19, disseminated cytomegalovirus and pancytopenia due to hemophagocytosis
Previous comorbidities Dilated cardiomyopathy of Chagas etiology, hypothyroidism by thyroidectomy by nodule 10 years ago Cardiomyopathy of Chagas etiology, disseminated cytomegalovirus, deep vein thrombosis, systemic arterial hypertension, diabetes, dyslipidemia, and chronic renal failure
Previous use of medications Carvedilol, losartan, furosemide, levothyroxine, isosorbide, and hydralazine Unknown
Medicines used during ICU stay Methylprednisolone, azathioprine, anti-thymocyte globulin, cyclosporine, tacrolimus, meropenem, linezolid, micafungin, vancomycin, polymyxin B, tigecycline, amikacin, fluconazole, ganciclovir, and hydrocortisone Meropenem, colistin, linezolid
fluconazole, amikacin, cyclosporine, azathioprine, and prednisone
ARD, acute respiratory distress syndrome; COVID-19, coronavirus disease 2019; ICU, intensive care unit; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. | Fatal | ReactionOutcome | CC BY | 33579042 | 19,731,502 | 2021-02-10 |
What was the outcome of reaction 'COVID-19'? | SARS-CoV-2 Infection and CMV Dissemination in Transplant Recipients as a Treatment for Chagas Cardiomyopathy: A Case Report.
Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 has infected over 90 million people worldwide, therefore it is considered a pandemic. SARS-CoV-2 infection can lead to severe pneumonia, acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure. Individuals receiving a heart transplantation (HT) may be at higher risk of adverse outcomes attributable to COVID-19 due to immunosuppressives, as well as concomitant infections that may also influence the prognoses. Herein, we describe the first report of two cases of HT recipients with concomitant infections by SARS-CoV-2, Trypanosoma cruzi, and cytomegalovirus (CMV) dissemination, from the first day of hospitalization due to COVID-19 in the intensive care unit (ICU) until the death of the patients.
1. Introduction
Coronavirus disease 2019 (COVID-19), an infectious respiratory disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spread on a pandemic scale since the first case was reported in Wuhan, China, in 2019 [1]. Most patients with the disease have mild-to-moderate symptoms; however, approximately 15% develop severe pneumonia, while approximately 5% develop acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure [2]. Lymphopenia is a recurrent feature in these patients, with a significant reduction in CD4+ T cells, CD8+ T cells, B cells, and natural killer (NK) cells [3], increasing the susceptibility of patients to severe illness and co-infection [4]. In fact, co-infections are associated with worsening of the clinical condition [5].
Chagas disease (CD) is a zoonosis whose etiologic agent is the protozoan Trypanosoma cruzi. It is estimated that 6–8 million people worldwide are infected [6]. CD has two distinct phases: an acute one, which is rare, with a strong production of type 1 cytokines, and a chronic phase, which develops in 30–40% of CD cases. The chronic phase of CD can be characterized by cardiomyopathy, arrhythmias, megaviscera, and, more rarely, polyneuropathy and stroke [7]. TCD4+ and TCD8+ lymphocytes are the main cells responsible for controlling parasitic infection. However, the immune response also contributes to tissue damage and pathology [8].
Chagasic cardiomyopathy represents the main cause of mortality from this disease, which can lead to heart failure, whose indication for treatment, especially in endemic countries, may include heart transplantation (HT) as a strategy to curb the evolution of this complication [6,9]. However, the immunosuppression and possible reactivation of the causative agent T. cruzi following HT require intensive clinical care and laboratory monitoring [10].
Other infections such as that caused by cytomegalovirus (CMV), a herpes virus that infects up to 60–100% of people in adulthood, are associated with transplant complications [11]. Primary CMV infection is generally asymptomatic in immunocompetent individuals, with the virus generating a latent infection. On the contrary, in immunocompromised and immunosuppressed populations, such as solid organ transplantation, hematopoietic stem cell transplantation, and HIV/AIDS patients, CMV reactivation is responsible for significant morbidity and mortality [12]. Although reactivation of infections such as CMV and CD are often described in transplant recipients, there are no reports of concomitant infection by T. cruzi, CMV, and SARS-CoV-2 in the context of HT in the literature to date.
Therefore, in this report, we investigated the progress of two patients who underwent HT at the Heart Institute of Hospital das Clínicas (Incor) and were subsequently transferred to the special intensive care unit (ICU) of the Hospital das Clínicas (Hospital das Clínicas, Faculty of Medicine, University of São Paulo-HCFMUSP) due to SARS-CoV-2 infection. These patients were diagnosed with COVID-19 by nasopharyngeal detection of SARS-CoV-2 RNA using reverse transcriptase polymerase chain reaction (RT-PCR). In addition, during hospitalization, CMV dissemination was evidenced by quantitative DNA detection in the blood. We describe herein the laboratory data from the first day of hospitalization due to COVID-19 until the death of the patients.
The baseline characteristics of these two patients, as well as their past clinical data, are summarized in Table 1.
2. Case Report
2.1. Patient 1
Female, 55 years old, presenting positive serology for CD since 2015, received HT on 2 May as a treatment for Chagas cardiomyopathy (Figure 1A) and underwent immunosuppressive therapy with methylprednisolone and azathioprine. Post-transplantation, she developed pneumonia, treated with meropenem and linezolid, and a remittent Candida tropicalis infection, treated with micafungin, with improvement. However, on 29 May, she developed dyspnea and desaturation, with a chest tomography suggestive of COVID-19. Her polymerase chain reaction (PCR) for SARS-CoV-2 was positive on 1 June.
On 3 June, she was transferred to the ICU specialized for treatment of severe SARS-CoV-2 infections. Laboratory analyses then showed that the patient had a reduced number of erythrocytes and hemoglobin level, and these reductions became more accentuated by day 33 in the ICU (Figure 2A,B). On this same day, the number of neutrophils peaked (Figure 2D). From the 4th to the 16th day in ICU, important lymphopenia developed (Figure 2F), resulting in an increase in the neutrophil-to-lymphocyte ratio in the same period (Figure 2H). Additionally, from the eighth day onward, she presented severe thrombocytopenia that persisted until death (Figure 2I).
Throughout the ICU hospitalization period, there were sustained high levels of creatinine, urea, D-dimer, C-reactive protein (CRP), and lactate dehydrogenase (Figure 2J,K,N,S,W). On the 32nd day, she presented a sharp increase in prothrombin time and activated partial thromboplastin time.
The cardiac function markers of creatine kinase myocardial band (CK-MB) and troponin remained elevated from the first days of ICU admission until death (Figure 2U,X). On the day of admission to the ICU, the N-terminal pro b-type natriuretic peptide (NT pro-BNP) was 70,000 pg/mL (reference value of <125 pg/mL) (Figure 2V). On the third day of hospitalization, NT pro-BNP (155,117 pg/mL) and troponin (0.28 ng/mL, reference value of <0.014 ng/mL) peaked, being related to the period in which she presented signs of graft rejection, which was treated with pulse methylprednisolone, thymoglobulin, and plasmapheresis. In addition, disseminated CMV infection was diagnosed (RT-PCR viral loads of 122 IU/mL on day 10 and 54 IU/mL on day 19 of hospitalization), for which she received ganciclovir.
After 47 days after diagnosis of SARS-CoV-2, the patient died (18 July) due to multiple organ dysfunctions associated with COVID-19.
2.2. Patient 2
Male, 62 years old, previously diagnosed with CD, received HT on 5 December 2019 as treatment for Chagas cardiomyopathy (Figure 1B). He underwent immunosuppressive therapy with cyclosporine, azathioprine, and prednisone. During postoperative hospitalization he presented Chagas reactivation characterized by skin biopsy and humoral graft rejection, being treated with plasmapheresis and methylprednisolone.
The patient was admitted to the ICU of the Hospital das Clínicas on 8 June, presenting cellulitis, deep vein thrombosis, and Chagas reactivation (a lower limb chagoma). The latter was treated with benznidazol. During hospitalization, he tested positive for PCR of SARS-CoV-2 on 29 June. He developed ARDS and septic and cardiogenic shock, which were the causes of his death.
Throughout the ICU hospitalization period, he maintained decreased levels of erythrocytes, hemoglobin, and lymphocytes (Figure 2A,B,F), as well as a high neutrophil/lymphocyte ratio (Figure 2H). From the 12th day, the number of platelets decreased and continued until the time of death (Figure 2I). The values of creatinine, urea, and glucose also remained high throughout the hospitalization period (Figure 2J,K,T). In addition, disseminated CMV (viral loads of 711 IU/mL detected on the 28th day and 1477 IU/mL on the 35th day of hospitalization) was diagnosed and treated with ganciclovir.
One day before death, the following laboratory parameters peaked: D-dimer (7612 ng/mL FEU, reference value of <500 ng/mL FEU), CRP (280.6 ng/mL, reference value of 0.300 ng/mL), CK-MB (8.38 ng/mL, reference value of 0.10–4.94 ng/mL), NT pro-BNP (55,393 pg/mL), lactate dehydrogenase (1161 U/L, reference value of 135–225 U/L), and troponin (0.701 ng/mL) (Figure 2N,S,U–X).
On 20 July, 36 days after admission to the ICU and 21 days after a positive COVID-19 diagnosis, the patient died.
3. Discussion
Herein, we described the first report of triple infection (SARS-CoV-2 infection, T. cruzi infection, and CMV dissemination) in HT recipients. Patients received HT as a form of treatment for Chagas cardiomyopathy.
We described the laboratory data from the first day of hospitalization in the ICU due to COVID-19 until the time of death. Both patients were admitted to a referral center for treatment for COVID-19 in the metropolitan region of São Paulo, a city in southeastern Brazil. We hypothesize that the triple infection by SARS-CoV-2 and CMV may have been an important cause of death and of the worsening in CD patients with HT. To date, there have been no similar reports of patients presenting these three concomitant infectious diseases and HT receptors.
COVID-19 infection among transplant recipients increases the potential for developing severe illness [13] and may vary between different organ transplants [14,15].
SARS-CoV-2 entry´s receptor angiotensin converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2) are expressed in different tissues, such as the lungs, heart, liver, kidneys, testicles, thyroid, and adipose tissue [16]. Some patients with COVID-19 develop severe disease characterized by respiratory distress syndrome and systemic manifestations. This condition has been associated with the dysregulated release of pro-inflammatory cytokines, termed “cytokine storm,” which may induce multi-organ failure [1,17].
The use of immunosuppressants and post-surgical opportunistic infections can also lead to damage of multisystem organs or even death [18]. There is a therapeutic paradox here, because while insufficient immunosuppression results in graft loss due to rejection, excessive immunosuppression can result in serious infection, including SARS-COV-2 [13], besides contributing to the reactivation of pathogens [19].
Long-term administration of immunosuppressants to solid organ transplant (SOT) receptors to reduce the risk of graft rejection may increase the risk of respiratory infections [20], although there is no clear clinical evidence of increased morbidity/mortality in SARS-CoV-2 infection [21].
Fernandez-Ruiz and collaborators [22] described a cohort of SOT receptors affected by COVID-19, 44% of whom were kidney transplant recipients, 33.3% liver transplant recipients, and 22.2% heart transplant recipients. The lethality rate was 27.8%, suggesting that SARS-CoV-2 infection had a severe course in SOT recipients.
Recently, we described the first patients with CD affected by SARS-CoV-2. The CD patients presented an increase in COVID-19 laboratory hallmarks and a rapid disease progression. Despite the efforts of the health staff, both patients died [3].
HT may be associated with the reactivation of pathogens, as reported in CD [10]. In a Brazilian cohort, the reactivation rate of Chagas disease after heart transplantation was reported to be 38.8% [23]. This fact made HT as a treatment for Chagas cardiomyopathy initially controversial. However, currently, especially in endemic countries, it is the most viable therapeutic option for patients with end-stage heart failure [24]. We hypothesize that, in the cases presented here, HT followed by reactivation of CD conferred an additional risk factor for the worsening of COVID-19.
Transplant recipients are also commonly affected by CMV reactivation, being associated with significant morbidity and mortality [12] that may worsen the infectious condition of COVID-19 [25]. In SOT, risk factors include the use of immunosuppressants for transplantation, advanced age, acute rejection, and other concomitant infections [11], with all these characteristics being present in both patients described herein. In the present report, we observed that immunosuppression may have contributed to the susceptibility to superinfections and more severe clinical manifestations in individuals undergoing HT.
Since overactivated immune responses can be one of the causes of organ damage, the anti-inflammatory effects of immunosuppression can be protective, reducing the cytokine storm related to complications in COVID-19 [26]. In this context, it has been described that immunosuppressive therapy with calcineurin inhibitors in patients with solid organ transplantation or systemic rheumatic diseases promotes a clinical course in SARS-CoV-2 infection, which is generally mild, and with an apparently low risk of superinfection [27]. In addition, immunosuppression has not been evaluated as a risk factor for SARS or MERS [28].
However, in the present case report, we observed that immunosuppression may have contributed to the susceptibility to SARS-CoV-2 infection, the reactivation of pathogens, and more serious clinical manifestations in individuals undergoing HT. Corroborating our findings, it was shown that patients with COVID-19 and cancer, due to their systemic immunosuppressive condition caused by malignancy and anticancer treatments, such as chemotherapy, had an increased risk of SARS-CoV-2 infection and a worse prognosis [29]. It has also been reported that chronic use of corticosteroids prior to SARS-CoV-2 contamination is associated with critical disease outcomes, including a high risk of death [30].
These contradictory observations show that knowledge about the relationship between COVID-19 and the patient’s immune condition is limited. Further studies are needed to elucidate the immune responses and prognosis of COVID-19.
In general, HT in patient 1 proved to be successful for the treatment of Chagas cardiomyopathy, with no reactivation of the pathogen. However, after SARS-CoV-2 infection, she presented graft rejection, treated with methylprednisolone, thymoglobulin, and plasmapheresis. There is a description in eye transplantation that COVID-19 infection can compromise the balance of immunoregulatory responses that allow graft survival, contributing to rejection in individuals infected with SARS-CoV-2 [31]. Thus, this change in the balance of attenuation of the immune response, such as a reduction in the number of regulatory T cells (Tregs) [32], may favor direct cardiotoxic action and multiple organ dysfunction.
During postoperative hospitalization, patient 2 presented Chagas reactivation, evidenced by skin biopsy, indicating that the CD was not completely controlled. In addition, it is important to consider that immunosuppressive therapy can contribute to the reactivation of CD [33]. After HT, the patient presented graft rejection and was treated with plasmapheresis and methylprednisolone. With this, before the SARS-CoV-2 infection, the patient was already weakened and died 21 days after the infection, in relation to patient 1 who died after 47 days of infection.
4. Conclusions
This report highlights the first case of an association between COVID-19, CD, and CMV dissemination in HT recipients. The patients had rapid disease progression to death. We believe that HT and the usage of immunosuppressive drugs, as well as immunosuppression generated by concomitant infections, may be an important risk factor for the development of severe COVID-19, especially in endemic areas with underreported CD infection.
Author Contributions
Conceptualization, S.C.G.-S., G.B., and R.W.A.; investigation, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., and S.M.; resources, M.N.S. and A.J.d.S.D.; writing—original draft preparation, S.C.G.-S., G.B., and R.W.A.; writing—review and editing, S.C.G.-S., G.B., and R.W.A.; visualization, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., S.M., A.J.d.S.D., and M.N.S.; supervision, M.N.S.; funding acquisition, A.J.d.S.D. and M.N.S. All authors have read and agreed to the published version of the manuscript.
Funding
This research was funded by Fundação de Amparo à Pesquisa (FAPESP), grant numbers 2019/22448-0, 19/02679-7, 2017/18199-9, 2018/18230-6, and 2019/26928-6. Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, CAPES:88887.503842/2020-00.
Institutional Review Board Statement
This study was approved by the local ethics committee (HCFMUSP nº 30800520.7.0000.0068-2020) and was carried out in accordance with the 2013 revision of the Declaration of Helsinki.
Informed Consent Statement
Informed consent was obtained from all of the subjects involved in the study.
Data Availability Statement
The laboratory data and imaging exams presented herein came from Hospital das Clinics, Faculty of Medicine, University of São Paulo (HCFMUSP) with prior approval for their use.
Conflicts of Interest
The authors declare no conflict of interest.
Figure 1 Chest X-ray of (a) patient 1 from May 2020 and (b) patient 2 from June 2015, taken just before heart transplantation, showing the Chagasic cardiomyopathy.
Figure 2 Daily clinical features of patients, from the first day of hospitalization in the ICU until death. Blood levels of (A) erythrocytes, (B) hemoglobin, (C) leukocytes, (D) neutrophils, (E) eosinophils, (F) lymphocytes, (G) monocytes, (H) neutrophil-to-lymphocyte ratio, (I) platelets, (J) creatinine, (K) urea, (L) prothrombin time, (M) activated partial thromboplastin time, (N) D-dimer, (O) pH, (P) pO2, (Q) pCO2, (R) oxygen peripheral, (S) CPR, (T) glucose, (U) CK-MB, (V) NT-proBNP, (W) lactate dehydrogenase, and (X) troponin T. Gray boxes represent the reference values.
tropicalmed-06-00022-t001_Table 1 Table 1 Patients’ characteristics, comorbidities, treatment, and complications.
Patient 1 Patient 2
Sex Female Male
Age 55 62
Positive SARS-CoV-2 RT-PCR date 1 June 2020 29 June 2020
Heart transplant date 2 May 2020 5 December 2019
Death date 30 days 207 days
Days between positive SARS-CoV-2 diagnosis and death 47 days 21 days
Complications during ICU stay ARDS due to COVID–19; heart transplant rejection, disseminated cytomegalovirus; aggravated chronic kidney disease and pressure ulcer ARDS due to COVID–19, disseminated cytomegalovirus and pancytopenia due to hemophagocytosis
Previous comorbidities Dilated cardiomyopathy of Chagas etiology, hypothyroidism by thyroidectomy by nodule 10 years ago Cardiomyopathy of Chagas etiology, disseminated cytomegalovirus, deep vein thrombosis, systemic arterial hypertension, diabetes, dyslipidemia, and chronic renal failure
Previous use of medications Carvedilol, losartan, furosemide, levothyroxine, isosorbide, and hydralazine Unknown
Medicines used during ICU stay Methylprednisolone, azathioprine, anti-thymocyte globulin, cyclosporine, tacrolimus, meropenem, linezolid, micafungin, vancomycin, polymyxin B, tigecycline, amikacin, fluconazole, ganciclovir, and hydrocortisone Meropenem, colistin, linezolid
fluconazole, amikacin, cyclosporine, azathioprine, and prednisone
ARD, acute respiratory distress syndrome; COVID-19, coronavirus disease 2019; ICU, intensive care unit; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. | Fatal | ReactionOutcome | CC BY | 33579042 | 19,816,778 | 2021-02-10 |
What was the outcome of reaction 'Cardiogenic shock'? | SARS-CoV-2 Infection and CMV Dissemination in Transplant Recipients as a Treatment for Chagas Cardiomyopathy: A Case Report.
Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 has infected over 90 million people worldwide, therefore it is considered a pandemic. SARS-CoV-2 infection can lead to severe pneumonia, acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure. Individuals receiving a heart transplantation (HT) may be at higher risk of adverse outcomes attributable to COVID-19 due to immunosuppressives, as well as concomitant infections that may also influence the prognoses. Herein, we describe the first report of two cases of HT recipients with concomitant infections by SARS-CoV-2, Trypanosoma cruzi, and cytomegalovirus (CMV) dissemination, from the first day of hospitalization due to COVID-19 in the intensive care unit (ICU) until the death of the patients.
1. Introduction
Coronavirus disease 2019 (COVID-19), an infectious respiratory disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spread on a pandemic scale since the first case was reported in Wuhan, China, in 2019 [1]. Most patients with the disease have mild-to-moderate symptoms; however, approximately 15% develop severe pneumonia, while approximately 5% develop acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure [2]. Lymphopenia is a recurrent feature in these patients, with a significant reduction in CD4+ T cells, CD8+ T cells, B cells, and natural killer (NK) cells [3], increasing the susceptibility of patients to severe illness and co-infection [4]. In fact, co-infections are associated with worsening of the clinical condition [5].
Chagas disease (CD) is a zoonosis whose etiologic agent is the protozoan Trypanosoma cruzi. It is estimated that 6–8 million people worldwide are infected [6]. CD has two distinct phases: an acute one, which is rare, with a strong production of type 1 cytokines, and a chronic phase, which develops in 30–40% of CD cases. The chronic phase of CD can be characterized by cardiomyopathy, arrhythmias, megaviscera, and, more rarely, polyneuropathy and stroke [7]. TCD4+ and TCD8+ lymphocytes are the main cells responsible for controlling parasitic infection. However, the immune response also contributes to tissue damage and pathology [8].
Chagasic cardiomyopathy represents the main cause of mortality from this disease, which can lead to heart failure, whose indication for treatment, especially in endemic countries, may include heart transplantation (HT) as a strategy to curb the evolution of this complication [6,9]. However, the immunosuppression and possible reactivation of the causative agent T. cruzi following HT require intensive clinical care and laboratory monitoring [10].
Other infections such as that caused by cytomegalovirus (CMV), a herpes virus that infects up to 60–100% of people in adulthood, are associated with transplant complications [11]. Primary CMV infection is generally asymptomatic in immunocompetent individuals, with the virus generating a latent infection. On the contrary, in immunocompromised and immunosuppressed populations, such as solid organ transplantation, hematopoietic stem cell transplantation, and HIV/AIDS patients, CMV reactivation is responsible for significant morbidity and mortality [12]. Although reactivation of infections such as CMV and CD are often described in transplant recipients, there are no reports of concomitant infection by T. cruzi, CMV, and SARS-CoV-2 in the context of HT in the literature to date.
Therefore, in this report, we investigated the progress of two patients who underwent HT at the Heart Institute of Hospital das Clínicas (Incor) and were subsequently transferred to the special intensive care unit (ICU) of the Hospital das Clínicas (Hospital das Clínicas, Faculty of Medicine, University of São Paulo-HCFMUSP) due to SARS-CoV-2 infection. These patients were diagnosed with COVID-19 by nasopharyngeal detection of SARS-CoV-2 RNA using reverse transcriptase polymerase chain reaction (RT-PCR). In addition, during hospitalization, CMV dissemination was evidenced by quantitative DNA detection in the blood. We describe herein the laboratory data from the first day of hospitalization due to COVID-19 until the death of the patients.
The baseline characteristics of these two patients, as well as their past clinical data, are summarized in Table 1.
2. Case Report
2.1. Patient 1
Female, 55 years old, presenting positive serology for CD since 2015, received HT on 2 May as a treatment for Chagas cardiomyopathy (Figure 1A) and underwent immunosuppressive therapy with methylprednisolone and azathioprine. Post-transplantation, she developed pneumonia, treated with meropenem and linezolid, and a remittent Candida tropicalis infection, treated with micafungin, with improvement. However, on 29 May, she developed dyspnea and desaturation, with a chest tomography suggestive of COVID-19. Her polymerase chain reaction (PCR) for SARS-CoV-2 was positive on 1 June.
On 3 June, she was transferred to the ICU specialized for treatment of severe SARS-CoV-2 infections. Laboratory analyses then showed that the patient had a reduced number of erythrocytes and hemoglobin level, and these reductions became more accentuated by day 33 in the ICU (Figure 2A,B). On this same day, the number of neutrophils peaked (Figure 2D). From the 4th to the 16th day in ICU, important lymphopenia developed (Figure 2F), resulting in an increase in the neutrophil-to-lymphocyte ratio in the same period (Figure 2H). Additionally, from the eighth day onward, she presented severe thrombocytopenia that persisted until death (Figure 2I).
Throughout the ICU hospitalization period, there were sustained high levels of creatinine, urea, D-dimer, C-reactive protein (CRP), and lactate dehydrogenase (Figure 2J,K,N,S,W). On the 32nd day, she presented a sharp increase in prothrombin time and activated partial thromboplastin time.
The cardiac function markers of creatine kinase myocardial band (CK-MB) and troponin remained elevated from the first days of ICU admission until death (Figure 2U,X). On the day of admission to the ICU, the N-terminal pro b-type natriuretic peptide (NT pro-BNP) was 70,000 pg/mL (reference value of <125 pg/mL) (Figure 2V). On the third day of hospitalization, NT pro-BNP (155,117 pg/mL) and troponin (0.28 ng/mL, reference value of <0.014 ng/mL) peaked, being related to the period in which she presented signs of graft rejection, which was treated with pulse methylprednisolone, thymoglobulin, and plasmapheresis. In addition, disseminated CMV infection was diagnosed (RT-PCR viral loads of 122 IU/mL on day 10 and 54 IU/mL on day 19 of hospitalization), for which she received ganciclovir.
After 47 days after diagnosis of SARS-CoV-2, the patient died (18 July) due to multiple organ dysfunctions associated with COVID-19.
2.2. Patient 2
Male, 62 years old, previously diagnosed with CD, received HT on 5 December 2019 as treatment for Chagas cardiomyopathy (Figure 1B). He underwent immunosuppressive therapy with cyclosporine, azathioprine, and prednisone. During postoperative hospitalization he presented Chagas reactivation characterized by skin biopsy and humoral graft rejection, being treated with plasmapheresis and methylprednisolone.
The patient was admitted to the ICU of the Hospital das Clínicas on 8 June, presenting cellulitis, deep vein thrombosis, and Chagas reactivation (a lower limb chagoma). The latter was treated with benznidazol. During hospitalization, he tested positive for PCR of SARS-CoV-2 on 29 June. He developed ARDS and septic and cardiogenic shock, which were the causes of his death.
Throughout the ICU hospitalization period, he maintained decreased levels of erythrocytes, hemoglobin, and lymphocytes (Figure 2A,B,F), as well as a high neutrophil/lymphocyte ratio (Figure 2H). From the 12th day, the number of platelets decreased and continued until the time of death (Figure 2I). The values of creatinine, urea, and glucose also remained high throughout the hospitalization period (Figure 2J,K,T). In addition, disseminated CMV (viral loads of 711 IU/mL detected on the 28th day and 1477 IU/mL on the 35th day of hospitalization) was diagnosed and treated with ganciclovir.
One day before death, the following laboratory parameters peaked: D-dimer (7612 ng/mL FEU, reference value of <500 ng/mL FEU), CRP (280.6 ng/mL, reference value of 0.300 ng/mL), CK-MB (8.38 ng/mL, reference value of 0.10–4.94 ng/mL), NT pro-BNP (55,393 pg/mL), lactate dehydrogenase (1161 U/L, reference value of 135–225 U/L), and troponin (0.701 ng/mL) (Figure 2N,S,U–X).
On 20 July, 36 days after admission to the ICU and 21 days after a positive COVID-19 diagnosis, the patient died.
3. Discussion
Herein, we described the first report of triple infection (SARS-CoV-2 infection, T. cruzi infection, and CMV dissemination) in HT recipients. Patients received HT as a form of treatment for Chagas cardiomyopathy.
We described the laboratory data from the first day of hospitalization in the ICU due to COVID-19 until the time of death. Both patients were admitted to a referral center for treatment for COVID-19 in the metropolitan region of São Paulo, a city in southeastern Brazil. We hypothesize that the triple infection by SARS-CoV-2 and CMV may have been an important cause of death and of the worsening in CD patients with HT. To date, there have been no similar reports of patients presenting these three concomitant infectious diseases and HT receptors.
COVID-19 infection among transplant recipients increases the potential for developing severe illness [13] and may vary between different organ transplants [14,15].
SARS-CoV-2 entry´s receptor angiotensin converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2) are expressed in different tissues, such as the lungs, heart, liver, kidneys, testicles, thyroid, and adipose tissue [16]. Some patients with COVID-19 develop severe disease characterized by respiratory distress syndrome and systemic manifestations. This condition has been associated with the dysregulated release of pro-inflammatory cytokines, termed “cytokine storm,” which may induce multi-organ failure [1,17].
The use of immunosuppressants and post-surgical opportunistic infections can also lead to damage of multisystem organs or even death [18]. There is a therapeutic paradox here, because while insufficient immunosuppression results in graft loss due to rejection, excessive immunosuppression can result in serious infection, including SARS-COV-2 [13], besides contributing to the reactivation of pathogens [19].
Long-term administration of immunosuppressants to solid organ transplant (SOT) receptors to reduce the risk of graft rejection may increase the risk of respiratory infections [20], although there is no clear clinical evidence of increased morbidity/mortality in SARS-CoV-2 infection [21].
Fernandez-Ruiz and collaborators [22] described a cohort of SOT receptors affected by COVID-19, 44% of whom were kidney transplant recipients, 33.3% liver transplant recipients, and 22.2% heart transplant recipients. The lethality rate was 27.8%, suggesting that SARS-CoV-2 infection had a severe course in SOT recipients.
Recently, we described the first patients with CD affected by SARS-CoV-2. The CD patients presented an increase in COVID-19 laboratory hallmarks and a rapid disease progression. Despite the efforts of the health staff, both patients died [3].
HT may be associated with the reactivation of pathogens, as reported in CD [10]. In a Brazilian cohort, the reactivation rate of Chagas disease after heart transplantation was reported to be 38.8% [23]. This fact made HT as a treatment for Chagas cardiomyopathy initially controversial. However, currently, especially in endemic countries, it is the most viable therapeutic option for patients with end-stage heart failure [24]. We hypothesize that, in the cases presented here, HT followed by reactivation of CD conferred an additional risk factor for the worsening of COVID-19.
Transplant recipients are also commonly affected by CMV reactivation, being associated with significant morbidity and mortality [12] that may worsen the infectious condition of COVID-19 [25]. In SOT, risk factors include the use of immunosuppressants for transplantation, advanced age, acute rejection, and other concomitant infections [11], with all these characteristics being present in both patients described herein. In the present report, we observed that immunosuppression may have contributed to the susceptibility to superinfections and more severe clinical manifestations in individuals undergoing HT.
Since overactivated immune responses can be one of the causes of organ damage, the anti-inflammatory effects of immunosuppression can be protective, reducing the cytokine storm related to complications in COVID-19 [26]. In this context, it has been described that immunosuppressive therapy with calcineurin inhibitors in patients with solid organ transplantation or systemic rheumatic diseases promotes a clinical course in SARS-CoV-2 infection, which is generally mild, and with an apparently low risk of superinfection [27]. In addition, immunosuppression has not been evaluated as a risk factor for SARS or MERS [28].
However, in the present case report, we observed that immunosuppression may have contributed to the susceptibility to SARS-CoV-2 infection, the reactivation of pathogens, and more serious clinical manifestations in individuals undergoing HT. Corroborating our findings, it was shown that patients with COVID-19 and cancer, due to their systemic immunosuppressive condition caused by malignancy and anticancer treatments, such as chemotherapy, had an increased risk of SARS-CoV-2 infection and a worse prognosis [29]. It has also been reported that chronic use of corticosteroids prior to SARS-CoV-2 contamination is associated with critical disease outcomes, including a high risk of death [30].
These contradictory observations show that knowledge about the relationship between COVID-19 and the patient’s immune condition is limited. Further studies are needed to elucidate the immune responses and prognosis of COVID-19.
In general, HT in patient 1 proved to be successful for the treatment of Chagas cardiomyopathy, with no reactivation of the pathogen. However, after SARS-CoV-2 infection, she presented graft rejection, treated with methylprednisolone, thymoglobulin, and plasmapheresis. There is a description in eye transplantation that COVID-19 infection can compromise the balance of immunoregulatory responses that allow graft survival, contributing to rejection in individuals infected with SARS-CoV-2 [31]. Thus, this change in the balance of attenuation of the immune response, such as a reduction in the number of regulatory T cells (Tregs) [32], may favor direct cardiotoxic action and multiple organ dysfunction.
During postoperative hospitalization, patient 2 presented Chagas reactivation, evidenced by skin biopsy, indicating that the CD was not completely controlled. In addition, it is important to consider that immunosuppressive therapy can contribute to the reactivation of CD [33]. After HT, the patient presented graft rejection and was treated with plasmapheresis and methylprednisolone. With this, before the SARS-CoV-2 infection, the patient was already weakened and died 21 days after the infection, in relation to patient 1 who died after 47 days of infection.
4. Conclusions
This report highlights the first case of an association between COVID-19, CD, and CMV dissemination in HT recipients. The patients had rapid disease progression to death. We believe that HT and the usage of immunosuppressive drugs, as well as immunosuppression generated by concomitant infections, may be an important risk factor for the development of severe COVID-19, especially in endemic areas with underreported CD infection.
Author Contributions
Conceptualization, S.C.G.-S., G.B., and R.W.A.; investigation, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., and S.M.; resources, M.N.S. and A.J.d.S.D.; writing—original draft preparation, S.C.G.-S., G.B., and R.W.A.; writing—review and editing, S.C.G.-S., G.B., and R.W.A.; visualization, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., S.M., A.J.d.S.D., and M.N.S.; supervision, M.N.S.; funding acquisition, A.J.d.S.D. and M.N.S. All authors have read and agreed to the published version of the manuscript.
Funding
This research was funded by Fundação de Amparo à Pesquisa (FAPESP), grant numbers 2019/22448-0, 19/02679-7, 2017/18199-9, 2018/18230-6, and 2019/26928-6. Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, CAPES:88887.503842/2020-00.
Institutional Review Board Statement
This study was approved by the local ethics committee (HCFMUSP nº 30800520.7.0000.0068-2020) and was carried out in accordance with the 2013 revision of the Declaration of Helsinki.
Informed Consent Statement
Informed consent was obtained from all of the subjects involved in the study.
Data Availability Statement
The laboratory data and imaging exams presented herein came from Hospital das Clinics, Faculty of Medicine, University of São Paulo (HCFMUSP) with prior approval for their use.
Conflicts of Interest
The authors declare no conflict of interest.
Figure 1 Chest X-ray of (a) patient 1 from May 2020 and (b) patient 2 from June 2015, taken just before heart transplantation, showing the Chagasic cardiomyopathy.
Figure 2 Daily clinical features of patients, from the first day of hospitalization in the ICU until death. Blood levels of (A) erythrocytes, (B) hemoglobin, (C) leukocytes, (D) neutrophils, (E) eosinophils, (F) lymphocytes, (G) monocytes, (H) neutrophil-to-lymphocyte ratio, (I) platelets, (J) creatinine, (K) urea, (L) prothrombin time, (M) activated partial thromboplastin time, (N) D-dimer, (O) pH, (P) pO2, (Q) pCO2, (R) oxygen peripheral, (S) CPR, (T) glucose, (U) CK-MB, (V) NT-proBNP, (W) lactate dehydrogenase, and (X) troponin T. Gray boxes represent the reference values.
tropicalmed-06-00022-t001_Table 1 Table 1 Patients’ characteristics, comorbidities, treatment, and complications.
Patient 1 Patient 2
Sex Female Male
Age 55 62
Positive SARS-CoV-2 RT-PCR date 1 June 2020 29 June 2020
Heart transplant date 2 May 2020 5 December 2019
Death date 30 days 207 days
Days between positive SARS-CoV-2 diagnosis and death 47 days 21 days
Complications during ICU stay ARDS due to COVID–19; heart transplant rejection, disseminated cytomegalovirus; aggravated chronic kidney disease and pressure ulcer ARDS due to COVID–19, disseminated cytomegalovirus and pancytopenia due to hemophagocytosis
Previous comorbidities Dilated cardiomyopathy of Chagas etiology, hypothyroidism by thyroidectomy by nodule 10 years ago Cardiomyopathy of Chagas etiology, disseminated cytomegalovirus, deep vein thrombosis, systemic arterial hypertension, diabetes, dyslipidemia, and chronic renal failure
Previous use of medications Carvedilol, losartan, furosemide, levothyroxine, isosorbide, and hydralazine Unknown
Medicines used during ICU stay Methylprednisolone, azathioprine, anti-thymocyte globulin, cyclosporine, tacrolimus, meropenem, linezolid, micafungin, vancomycin, polymyxin B, tigecycline, amikacin, fluconazole, ganciclovir, and hydrocortisone Meropenem, colistin, linezolid
fluconazole, amikacin, cyclosporine, azathioprine, and prednisone
ARD, acute respiratory distress syndrome; COVID-19, coronavirus disease 2019; ICU, intensive care unit; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. | Fatal | ReactionOutcome | CC BY | 33579042 | 19,731,502 | 2021-02-10 |
What was the outcome of reaction 'Chronic kidney disease'? | SARS-CoV-2 Infection and CMV Dissemination in Transplant Recipients as a Treatment for Chagas Cardiomyopathy: A Case Report.
Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 has infected over 90 million people worldwide, therefore it is considered a pandemic. SARS-CoV-2 infection can lead to severe pneumonia, acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure. Individuals receiving a heart transplantation (HT) may be at higher risk of adverse outcomes attributable to COVID-19 due to immunosuppressives, as well as concomitant infections that may also influence the prognoses. Herein, we describe the first report of two cases of HT recipients with concomitant infections by SARS-CoV-2, Trypanosoma cruzi, and cytomegalovirus (CMV) dissemination, from the first day of hospitalization due to COVID-19 in the intensive care unit (ICU) until the death of the patients.
1. Introduction
Coronavirus disease 2019 (COVID-19), an infectious respiratory disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spread on a pandemic scale since the first case was reported in Wuhan, China, in 2019 [1]. Most patients with the disease have mild-to-moderate symptoms; however, approximately 15% develop severe pneumonia, while approximately 5% develop acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure [2]. Lymphopenia is a recurrent feature in these patients, with a significant reduction in CD4+ T cells, CD8+ T cells, B cells, and natural killer (NK) cells [3], increasing the susceptibility of patients to severe illness and co-infection [4]. In fact, co-infections are associated with worsening of the clinical condition [5].
Chagas disease (CD) is a zoonosis whose etiologic agent is the protozoan Trypanosoma cruzi. It is estimated that 6–8 million people worldwide are infected [6]. CD has two distinct phases: an acute one, which is rare, with a strong production of type 1 cytokines, and a chronic phase, which develops in 30–40% of CD cases. The chronic phase of CD can be characterized by cardiomyopathy, arrhythmias, megaviscera, and, more rarely, polyneuropathy and stroke [7]. TCD4+ and TCD8+ lymphocytes are the main cells responsible for controlling parasitic infection. However, the immune response also contributes to tissue damage and pathology [8].
Chagasic cardiomyopathy represents the main cause of mortality from this disease, which can lead to heart failure, whose indication for treatment, especially in endemic countries, may include heart transplantation (HT) as a strategy to curb the evolution of this complication [6,9]. However, the immunosuppression and possible reactivation of the causative agent T. cruzi following HT require intensive clinical care and laboratory monitoring [10].
Other infections such as that caused by cytomegalovirus (CMV), a herpes virus that infects up to 60–100% of people in adulthood, are associated with transplant complications [11]. Primary CMV infection is generally asymptomatic in immunocompetent individuals, with the virus generating a latent infection. On the contrary, in immunocompromised and immunosuppressed populations, such as solid organ transplantation, hematopoietic stem cell transplantation, and HIV/AIDS patients, CMV reactivation is responsible for significant morbidity and mortality [12]. Although reactivation of infections such as CMV and CD are often described in transplant recipients, there are no reports of concomitant infection by T. cruzi, CMV, and SARS-CoV-2 in the context of HT in the literature to date.
Therefore, in this report, we investigated the progress of two patients who underwent HT at the Heart Institute of Hospital das Clínicas (Incor) and were subsequently transferred to the special intensive care unit (ICU) of the Hospital das Clínicas (Hospital das Clínicas, Faculty of Medicine, University of São Paulo-HCFMUSP) due to SARS-CoV-2 infection. These patients were diagnosed with COVID-19 by nasopharyngeal detection of SARS-CoV-2 RNA using reverse transcriptase polymerase chain reaction (RT-PCR). In addition, during hospitalization, CMV dissemination was evidenced by quantitative DNA detection in the blood. We describe herein the laboratory data from the first day of hospitalization due to COVID-19 until the death of the patients.
The baseline characteristics of these two patients, as well as their past clinical data, are summarized in Table 1.
2. Case Report
2.1. Patient 1
Female, 55 years old, presenting positive serology for CD since 2015, received HT on 2 May as a treatment for Chagas cardiomyopathy (Figure 1A) and underwent immunosuppressive therapy with methylprednisolone and azathioprine. Post-transplantation, she developed pneumonia, treated with meropenem and linezolid, and a remittent Candida tropicalis infection, treated with micafungin, with improvement. However, on 29 May, she developed dyspnea and desaturation, with a chest tomography suggestive of COVID-19. Her polymerase chain reaction (PCR) for SARS-CoV-2 was positive on 1 June.
On 3 June, she was transferred to the ICU specialized for treatment of severe SARS-CoV-2 infections. Laboratory analyses then showed that the patient had a reduced number of erythrocytes and hemoglobin level, and these reductions became more accentuated by day 33 in the ICU (Figure 2A,B). On this same day, the number of neutrophils peaked (Figure 2D). From the 4th to the 16th day in ICU, important lymphopenia developed (Figure 2F), resulting in an increase in the neutrophil-to-lymphocyte ratio in the same period (Figure 2H). Additionally, from the eighth day onward, she presented severe thrombocytopenia that persisted until death (Figure 2I).
Throughout the ICU hospitalization period, there were sustained high levels of creatinine, urea, D-dimer, C-reactive protein (CRP), and lactate dehydrogenase (Figure 2J,K,N,S,W). On the 32nd day, she presented a sharp increase in prothrombin time and activated partial thromboplastin time.
The cardiac function markers of creatine kinase myocardial band (CK-MB) and troponin remained elevated from the first days of ICU admission until death (Figure 2U,X). On the day of admission to the ICU, the N-terminal pro b-type natriuretic peptide (NT pro-BNP) was 70,000 pg/mL (reference value of <125 pg/mL) (Figure 2V). On the third day of hospitalization, NT pro-BNP (155,117 pg/mL) and troponin (0.28 ng/mL, reference value of <0.014 ng/mL) peaked, being related to the period in which she presented signs of graft rejection, which was treated with pulse methylprednisolone, thymoglobulin, and plasmapheresis. In addition, disseminated CMV infection was diagnosed (RT-PCR viral loads of 122 IU/mL on day 10 and 54 IU/mL on day 19 of hospitalization), for which she received ganciclovir.
After 47 days after diagnosis of SARS-CoV-2, the patient died (18 July) due to multiple organ dysfunctions associated with COVID-19.
2.2. Patient 2
Male, 62 years old, previously diagnosed with CD, received HT on 5 December 2019 as treatment for Chagas cardiomyopathy (Figure 1B). He underwent immunosuppressive therapy with cyclosporine, azathioprine, and prednisone. During postoperative hospitalization he presented Chagas reactivation characterized by skin biopsy and humoral graft rejection, being treated with plasmapheresis and methylprednisolone.
The patient was admitted to the ICU of the Hospital das Clínicas on 8 June, presenting cellulitis, deep vein thrombosis, and Chagas reactivation (a lower limb chagoma). The latter was treated with benznidazol. During hospitalization, he tested positive for PCR of SARS-CoV-2 on 29 June. He developed ARDS and septic and cardiogenic shock, which were the causes of his death.
Throughout the ICU hospitalization period, he maintained decreased levels of erythrocytes, hemoglobin, and lymphocytes (Figure 2A,B,F), as well as a high neutrophil/lymphocyte ratio (Figure 2H). From the 12th day, the number of platelets decreased and continued until the time of death (Figure 2I). The values of creatinine, urea, and glucose also remained high throughout the hospitalization period (Figure 2J,K,T). In addition, disseminated CMV (viral loads of 711 IU/mL detected on the 28th day and 1477 IU/mL on the 35th day of hospitalization) was diagnosed and treated with ganciclovir.
One day before death, the following laboratory parameters peaked: D-dimer (7612 ng/mL FEU, reference value of <500 ng/mL FEU), CRP (280.6 ng/mL, reference value of 0.300 ng/mL), CK-MB (8.38 ng/mL, reference value of 0.10–4.94 ng/mL), NT pro-BNP (55,393 pg/mL), lactate dehydrogenase (1161 U/L, reference value of 135–225 U/L), and troponin (0.701 ng/mL) (Figure 2N,S,U–X).
On 20 July, 36 days after admission to the ICU and 21 days after a positive COVID-19 diagnosis, the patient died.
3. Discussion
Herein, we described the first report of triple infection (SARS-CoV-2 infection, T. cruzi infection, and CMV dissemination) in HT recipients. Patients received HT as a form of treatment for Chagas cardiomyopathy.
We described the laboratory data from the first day of hospitalization in the ICU due to COVID-19 until the time of death. Both patients were admitted to a referral center for treatment for COVID-19 in the metropolitan region of São Paulo, a city in southeastern Brazil. We hypothesize that the triple infection by SARS-CoV-2 and CMV may have been an important cause of death and of the worsening in CD patients with HT. To date, there have been no similar reports of patients presenting these three concomitant infectious diseases and HT receptors.
COVID-19 infection among transplant recipients increases the potential for developing severe illness [13] and may vary between different organ transplants [14,15].
SARS-CoV-2 entry´s receptor angiotensin converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2) are expressed in different tissues, such as the lungs, heart, liver, kidneys, testicles, thyroid, and adipose tissue [16]. Some patients with COVID-19 develop severe disease characterized by respiratory distress syndrome and systemic manifestations. This condition has been associated with the dysregulated release of pro-inflammatory cytokines, termed “cytokine storm,” which may induce multi-organ failure [1,17].
The use of immunosuppressants and post-surgical opportunistic infections can also lead to damage of multisystem organs or even death [18]. There is a therapeutic paradox here, because while insufficient immunosuppression results in graft loss due to rejection, excessive immunosuppression can result in serious infection, including SARS-COV-2 [13], besides contributing to the reactivation of pathogens [19].
Long-term administration of immunosuppressants to solid organ transplant (SOT) receptors to reduce the risk of graft rejection may increase the risk of respiratory infections [20], although there is no clear clinical evidence of increased morbidity/mortality in SARS-CoV-2 infection [21].
Fernandez-Ruiz and collaborators [22] described a cohort of SOT receptors affected by COVID-19, 44% of whom were kidney transplant recipients, 33.3% liver transplant recipients, and 22.2% heart transplant recipients. The lethality rate was 27.8%, suggesting that SARS-CoV-2 infection had a severe course in SOT recipients.
Recently, we described the first patients with CD affected by SARS-CoV-2. The CD patients presented an increase in COVID-19 laboratory hallmarks and a rapid disease progression. Despite the efforts of the health staff, both patients died [3].
HT may be associated with the reactivation of pathogens, as reported in CD [10]. In a Brazilian cohort, the reactivation rate of Chagas disease after heart transplantation was reported to be 38.8% [23]. This fact made HT as a treatment for Chagas cardiomyopathy initially controversial. However, currently, especially in endemic countries, it is the most viable therapeutic option for patients with end-stage heart failure [24]. We hypothesize that, in the cases presented here, HT followed by reactivation of CD conferred an additional risk factor for the worsening of COVID-19.
Transplant recipients are also commonly affected by CMV reactivation, being associated with significant morbidity and mortality [12] that may worsen the infectious condition of COVID-19 [25]. In SOT, risk factors include the use of immunosuppressants for transplantation, advanced age, acute rejection, and other concomitant infections [11], with all these characteristics being present in both patients described herein. In the present report, we observed that immunosuppression may have contributed to the susceptibility to superinfections and more severe clinical manifestations in individuals undergoing HT.
Since overactivated immune responses can be one of the causes of organ damage, the anti-inflammatory effects of immunosuppression can be protective, reducing the cytokine storm related to complications in COVID-19 [26]. In this context, it has been described that immunosuppressive therapy with calcineurin inhibitors in patients with solid organ transplantation or systemic rheumatic diseases promotes a clinical course in SARS-CoV-2 infection, which is generally mild, and with an apparently low risk of superinfection [27]. In addition, immunosuppression has not been evaluated as a risk factor for SARS or MERS [28].
However, in the present case report, we observed that immunosuppression may have contributed to the susceptibility to SARS-CoV-2 infection, the reactivation of pathogens, and more serious clinical manifestations in individuals undergoing HT. Corroborating our findings, it was shown that patients with COVID-19 and cancer, due to their systemic immunosuppressive condition caused by malignancy and anticancer treatments, such as chemotherapy, had an increased risk of SARS-CoV-2 infection and a worse prognosis [29]. It has also been reported that chronic use of corticosteroids prior to SARS-CoV-2 contamination is associated with critical disease outcomes, including a high risk of death [30].
These contradictory observations show that knowledge about the relationship between COVID-19 and the patient’s immune condition is limited. Further studies are needed to elucidate the immune responses and prognosis of COVID-19.
In general, HT in patient 1 proved to be successful for the treatment of Chagas cardiomyopathy, with no reactivation of the pathogen. However, after SARS-CoV-2 infection, she presented graft rejection, treated with methylprednisolone, thymoglobulin, and plasmapheresis. There is a description in eye transplantation that COVID-19 infection can compromise the balance of immunoregulatory responses that allow graft survival, contributing to rejection in individuals infected with SARS-CoV-2 [31]. Thus, this change in the balance of attenuation of the immune response, such as a reduction in the number of regulatory T cells (Tregs) [32], may favor direct cardiotoxic action and multiple organ dysfunction.
During postoperative hospitalization, patient 2 presented Chagas reactivation, evidenced by skin biopsy, indicating that the CD was not completely controlled. In addition, it is important to consider that immunosuppressive therapy can contribute to the reactivation of CD [33]. After HT, the patient presented graft rejection and was treated with plasmapheresis and methylprednisolone. With this, before the SARS-CoV-2 infection, the patient was already weakened and died 21 days after the infection, in relation to patient 1 who died after 47 days of infection.
4. Conclusions
This report highlights the first case of an association between COVID-19, CD, and CMV dissemination in HT recipients. The patients had rapid disease progression to death. We believe that HT and the usage of immunosuppressive drugs, as well as immunosuppression generated by concomitant infections, may be an important risk factor for the development of severe COVID-19, especially in endemic areas with underreported CD infection.
Author Contributions
Conceptualization, S.C.G.-S., G.B., and R.W.A.; investigation, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., and S.M.; resources, M.N.S. and A.J.d.S.D.; writing—original draft preparation, S.C.G.-S., G.B., and R.W.A.; writing—review and editing, S.C.G.-S., G.B., and R.W.A.; visualization, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., S.M., A.J.d.S.D., and M.N.S.; supervision, M.N.S.; funding acquisition, A.J.d.S.D. and M.N.S. All authors have read and agreed to the published version of the manuscript.
Funding
This research was funded by Fundação de Amparo à Pesquisa (FAPESP), grant numbers 2019/22448-0, 19/02679-7, 2017/18199-9, 2018/18230-6, and 2019/26928-6. Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, CAPES:88887.503842/2020-00.
Institutional Review Board Statement
This study was approved by the local ethics committee (HCFMUSP nº 30800520.7.0000.0068-2020) and was carried out in accordance with the 2013 revision of the Declaration of Helsinki.
Informed Consent Statement
Informed consent was obtained from all of the subjects involved in the study.
Data Availability Statement
The laboratory data and imaging exams presented herein came from Hospital das Clinics, Faculty of Medicine, University of São Paulo (HCFMUSP) with prior approval for their use.
Conflicts of Interest
The authors declare no conflict of interest.
Figure 1 Chest X-ray of (a) patient 1 from May 2020 and (b) patient 2 from June 2015, taken just before heart transplantation, showing the Chagasic cardiomyopathy.
Figure 2 Daily clinical features of patients, from the first day of hospitalization in the ICU until death. Blood levels of (A) erythrocytes, (B) hemoglobin, (C) leukocytes, (D) neutrophils, (E) eosinophils, (F) lymphocytes, (G) monocytes, (H) neutrophil-to-lymphocyte ratio, (I) platelets, (J) creatinine, (K) urea, (L) prothrombin time, (M) activated partial thromboplastin time, (N) D-dimer, (O) pH, (P) pO2, (Q) pCO2, (R) oxygen peripheral, (S) CPR, (T) glucose, (U) CK-MB, (V) NT-proBNP, (W) lactate dehydrogenase, and (X) troponin T. Gray boxes represent the reference values.
tropicalmed-06-00022-t001_Table 1 Table 1 Patients’ characteristics, comorbidities, treatment, and complications.
Patient 1 Patient 2
Sex Female Male
Age 55 62
Positive SARS-CoV-2 RT-PCR date 1 June 2020 29 June 2020
Heart transplant date 2 May 2020 5 December 2019
Death date 30 days 207 days
Days between positive SARS-CoV-2 diagnosis and death 47 days 21 days
Complications during ICU stay ARDS due to COVID–19; heart transplant rejection, disseminated cytomegalovirus; aggravated chronic kidney disease and pressure ulcer ARDS due to COVID–19, disseminated cytomegalovirus and pancytopenia due to hemophagocytosis
Previous comorbidities Dilated cardiomyopathy of Chagas etiology, hypothyroidism by thyroidectomy by nodule 10 years ago Cardiomyopathy of Chagas etiology, disseminated cytomegalovirus, deep vein thrombosis, systemic arterial hypertension, diabetes, dyslipidemia, and chronic renal failure
Previous use of medications Carvedilol, losartan, furosemide, levothyroxine, isosorbide, and hydralazine Unknown
Medicines used during ICU stay Methylprednisolone, azathioprine, anti-thymocyte globulin, cyclosporine, tacrolimus, meropenem, linezolid, micafungin, vancomycin, polymyxin B, tigecycline, amikacin, fluconazole, ganciclovir, and hydrocortisone Meropenem, colistin, linezolid
fluconazole, amikacin, cyclosporine, azathioprine, and prednisone
ARD, acute respiratory distress syndrome; COVID-19, coronavirus disease 2019; ICU, intensive care unit; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. | Fatal | ReactionOutcome | CC BY | 33579042 | 19,681,859 | 2021-02-10 |
What was the outcome of reaction 'Disseminated cytomegaloviral infection'? | SARS-CoV-2 Infection and CMV Dissemination in Transplant Recipients as a Treatment for Chagas Cardiomyopathy: A Case Report.
Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 has infected over 90 million people worldwide, therefore it is considered a pandemic. SARS-CoV-2 infection can lead to severe pneumonia, acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure. Individuals receiving a heart transplantation (HT) may be at higher risk of adverse outcomes attributable to COVID-19 due to immunosuppressives, as well as concomitant infections that may also influence the prognoses. Herein, we describe the first report of two cases of HT recipients with concomitant infections by SARS-CoV-2, Trypanosoma cruzi, and cytomegalovirus (CMV) dissemination, from the first day of hospitalization due to COVID-19 in the intensive care unit (ICU) until the death of the patients.
1. Introduction
Coronavirus disease 2019 (COVID-19), an infectious respiratory disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spread on a pandemic scale since the first case was reported in Wuhan, China, in 2019 [1]. Most patients with the disease have mild-to-moderate symptoms; however, approximately 15% develop severe pneumonia, while approximately 5% develop acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure [2]. Lymphopenia is a recurrent feature in these patients, with a significant reduction in CD4+ T cells, CD8+ T cells, B cells, and natural killer (NK) cells [3], increasing the susceptibility of patients to severe illness and co-infection [4]. In fact, co-infections are associated with worsening of the clinical condition [5].
Chagas disease (CD) is a zoonosis whose etiologic agent is the protozoan Trypanosoma cruzi. It is estimated that 6–8 million people worldwide are infected [6]. CD has two distinct phases: an acute one, which is rare, with a strong production of type 1 cytokines, and a chronic phase, which develops in 30–40% of CD cases. The chronic phase of CD can be characterized by cardiomyopathy, arrhythmias, megaviscera, and, more rarely, polyneuropathy and stroke [7]. TCD4+ and TCD8+ lymphocytes are the main cells responsible for controlling parasitic infection. However, the immune response also contributes to tissue damage and pathology [8].
Chagasic cardiomyopathy represents the main cause of mortality from this disease, which can lead to heart failure, whose indication for treatment, especially in endemic countries, may include heart transplantation (HT) as a strategy to curb the evolution of this complication [6,9]. However, the immunosuppression and possible reactivation of the causative agent T. cruzi following HT require intensive clinical care and laboratory monitoring [10].
Other infections such as that caused by cytomegalovirus (CMV), a herpes virus that infects up to 60–100% of people in adulthood, are associated with transplant complications [11]. Primary CMV infection is generally asymptomatic in immunocompetent individuals, with the virus generating a latent infection. On the contrary, in immunocompromised and immunosuppressed populations, such as solid organ transplantation, hematopoietic stem cell transplantation, and HIV/AIDS patients, CMV reactivation is responsible for significant morbidity and mortality [12]. Although reactivation of infections such as CMV and CD are often described in transplant recipients, there are no reports of concomitant infection by T. cruzi, CMV, and SARS-CoV-2 in the context of HT in the literature to date.
Therefore, in this report, we investigated the progress of two patients who underwent HT at the Heart Institute of Hospital das Clínicas (Incor) and were subsequently transferred to the special intensive care unit (ICU) of the Hospital das Clínicas (Hospital das Clínicas, Faculty of Medicine, University of São Paulo-HCFMUSP) due to SARS-CoV-2 infection. These patients were diagnosed with COVID-19 by nasopharyngeal detection of SARS-CoV-2 RNA using reverse transcriptase polymerase chain reaction (RT-PCR). In addition, during hospitalization, CMV dissemination was evidenced by quantitative DNA detection in the blood. We describe herein the laboratory data from the first day of hospitalization due to COVID-19 until the death of the patients.
The baseline characteristics of these two patients, as well as their past clinical data, are summarized in Table 1.
2. Case Report
2.1. Patient 1
Female, 55 years old, presenting positive serology for CD since 2015, received HT on 2 May as a treatment for Chagas cardiomyopathy (Figure 1A) and underwent immunosuppressive therapy with methylprednisolone and azathioprine. Post-transplantation, she developed pneumonia, treated with meropenem and linezolid, and a remittent Candida tropicalis infection, treated with micafungin, with improvement. However, on 29 May, she developed dyspnea and desaturation, with a chest tomography suggestive of COVID-19. Her polymerase chain reaction (PCR) for SARS-CoV-2 was positive on 1 June.
On 3 June, she was transferred to the ICU specialized for treatment of severe SARS-CoV-2 infections. Laboratory analyses then showed that the patient had a reduced number of erythrocytes and hemoglobin level, and these reductions became more accentuated by day 33 in the ICU (Figure 2A,B). On this same day, the number of neutrophils peaked (Figure 2D). From the 4th to the 16th day in ICU, important lymphopenia developed (Figure 2F), resulting in an increase in the neutrophil-to-lymphocyte ratio in the same period (Figure 2H). Additionally, from the eighth day onward, she presented severe thrombocytopenia that persisted until death (Figure 2I).
Throughout the ICU hospitalization period, there were sustained high levels of creatinine, urea, D-dimer, C-reactive protein (CRP), and lactate dehydrogenase (Figure 2J,K,N,S,W). On the 32nd day, she presented a sharp increase in prothrombin time and activated partial thromboplastin time.
The cardiac function markers of creatine kinase myocardial band (CK-MB) and troponin remained elevated from the first days of ICU admission until death (Figure 2U,X). On the day of admission to the ICU, the N-terminal pro b-type natriuretic peptide (NT pro-BNP) was 70,000 pg/mL (reference value of <125 pg/mL) (Figure 2V). On the third day of hospitalization, NT pro-BNP (155,117 pg/mL) and troponin (0.28 ng/mL, reference value of <0.014 ng/mL) peaked, being related to the period in which she presented signs of graft rejection, which was treated with pulse methylprednisolone, thymoglobulin, and plasmapheresis. In addition, disseminated CMV infection was diagnosed (RT-PCR viral loads of 122 IU/mL on day 10 and 54 IU/mL on day 19 of hospitalization), for which she received ganciclovir.
After 47 days after diagnosis of SARS-CoV-2, the patient died (18 July) due to multiple organ dysfunctions associated with COVID-19.
2.2. Patient 2
Male, 62 years old, previously diagnosed with CD, received HT on 5 December 2019 as treatment for Chagas cardiomyopathy (Figure 1B). He underwent immunosuppressive therapy with cyclosporine, azathioprine, and prednisone. During postoperative hospitalization he presented Chagas reactivation characterized by skin biopsy and humoral graft rejection, being treated with plasmapheresis and methylprednisolone.
The patient was admitted to the ICU of the Hospital das Clínicas on 8 June, presenting cellulitis, deep vein thrombosis, and Chagas reactivation (a lower limb chagoma). The latter was treated with benznidazol. During hospitalization, he tested positive for PCR of SARS-CoV-2 on 29 June. He developed ARDS and septic and cardiogenic shock, which were the causes of his death.
Throughout the ICU hospitalization period, he maintained decreased levels of erythrocytes, hemoglobin, and lymphocytes (Figure 2A,B,F), as well as a high neutrophil/lymphocyte ratio (Figure 2H). From the 12th day, the number of platelets decreased and continued until the time of death (Figure 2I). The values of creatinine, urea, and glucose also remained high throughout the hospitalization period (Figure 2J,K,T). In addition, disseminated CMV (viral loads of 711 IU/mL detected on the 28th day and 1477 IU/mL on the 35th day of hospitalization) was diagnosed and treated with ganciclovir.
One day before death, the following laboratory parameters peaked: D-dimer (7612 ng/mL FEU, reference value of <500 ng/mL FEU), CRP (280.6 ng/mL, reference value of 0.300 ng/mL), CK-MB (8.38 ng/mL, reference value of 0.10–4.94 ng/mL), NT pro-BNP (55,393 pg/mL), lactate dehydrogenase (1161 U/L, reference value of 135–225 U/L), and troponin (0.701 ng/mL) (Figure 2N,S,U–X).
On 20 July, 36 days after admission to the ICU and 21 days after a positive COVID-19 diagnosis, the patient died.
3. Discussion
Herein, we described the first report of triple infection (SARS-CoV-2 infection, T. cruzi infection, and CMV dissemination) in HT recipients. Patients received HT as a form of treatment for Chagas cardiomyopathy.
We described the laboratory data from the first day of hospitalization in the ICU due to COVID-19 until the time of death. Both patients were admitted to a referral center for treatment for COVID-19 in the metropolitan region of São Paulo, a city in southeastern Brazil. We hypothesize that the triple infection by SARS-CoV-2 and CMV may have been an important cause of death and of the worsening in CD patients with HT. To date, there have been no similar reports of patients presenting these three concomitant infectious diseases and HT receptors.
COVID-19 infection among transplant recipients increases the potential for developing severe illness [13] and may vary between different organ transplants [14,15].
SARS-CoV-2 entry´s receptor angiotensin converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2) are expressed in different tissues, such as the lungs, heart, liver, kidneys, testicles, thyroid, and adipose tissue [16]. Some patients with COVID-19 develop severe disease characterized by respiratory distress syndrome and systemic manifestations. This condition has been associated with the dysregulated release of pro-inflammatory cytokines, termed “cytokine storm,” which may induce multi-organ failure [1,17].
The use of immunosuppressants and post-surgical opportunistic infections can also lead to damage of multisystem organs or even death [18]. There is a therapeutic paradox here, because while insufficient immunosuppression results in graft loss due to rejection, excessive immunosuppression can result in serious infection, including SARS-COV-2 [13], besides contributing to the reactivation of pathogens [19].
Long-term administration of immunosuppressants to solid organ transplant (SOT) receptors to reduce the risk of graft rejection may increase the risk of respiratory infections [20], although there is no clear clinical evidence of increased morbidity/mortality in SARS-CoV-2 infection [21].
Fernandez-Ruiz and collaborators [22] described a cohort of SOT receptors affected by COVID-19, 44% of whom were kidney transplant recipients, 33.3% liver transplant recipients, and 22.2% heart transplant recipients. The lethality rate was 27.8%, suggesting that SARS-CoV-2 infection had a severe course in SOT recipients.
Recently, we described the first patients with CD affected by SARS-CoV-2. The CD patients presented an increase in COVID-19 laboratory hallmarks and a rapid disease progression. Despite the efforts of the health staff, both patients died [3].
HT may be associated with the reactivation of pathogens, as reported in CD [10]. In a Brazilian cohort, the reactivation rate of Chagas disease after heart transplantation was reported to be 38.8% [23]. This fact made HT as a treatment for Chagas cardiomyopathy initially controversial. However, currently, especially in endemic countries, it is the most viable therapeutic option for patients with end-stage heart failure [24]. We hypothesize that, in the cases presented here, HT followed by reactivation of CD conferred an additional risk factor for the worsening of COVID-19.
Transplant recipients are also commonly affected by CMV reactivation, being associated with significant morbidity and mortality [12] that may worsen the infectious condition of COVID-19 [25]. In SOT, risk factors include the use of immunosuppressants for transplantation, advanced age, acute rejection, and other concomitant infections [11], with all these characteristics being present in both patients described herein. In the present report, we observed that immunosuppression may have contributed to the susceptibility to superinfections and more severe clinical manifestations in individuals undergoing HT.
Since overactivated immune responses can be one of the causes of organ damage, the anti-inflammatory effects of immunosuppression can be protective, reducing the cytokine storm related to complications in COVID-19 [26]. In this context, it has been described that immunosuppressive therapy with calcineurin inhibitors in patients with solid organ transplantation or systemic rheumatic diseases promotes a clinical course in SARS-CoV-2 infection, which is generally mild, and with an apparently low risk of superinfection [27]. In addition, immunosuppression has not been evaluated as a risk factor for SARS or MERS [28].
However, in the present case report, we observed that immunosuppression may have contributed to the susceptibility to SARS-CoV-2 infection, the reactivation of pathogens, and more serious clinical manifestations in individuals undergoing HT. Corroborating our findings, it was shown that patients with COVID-19 and cancer, due to their systemic immunosuppressive condition caused by malignancy and anticancer treatments, such as chemotherapy, had an increased risk of SARS-CoV-2 infection and a worse prognosis [29]. It has also been reported that chronic use of corticosteroids prior to SARS-CoV-2 contamination is associated with critical disease outcomes, including a high risk of death [30].
These contradictory observations show that knowledge about the relationship between COVID-19 and the patient’s immune condition is limited. Further studies are needed to elucidate the immune responses and prognosis of COVID-19.
In general, HT in patient 1 proved to be successful for the treatment of Chagas cardiomyopathy, with no reactivation of the pathogen. However, after SARS-CoV-2 infection, she presented graft rejection, treated with methylprednisolone, thymoglobulin, and plasmapheresis. There is a description in eye transplantation that COVID-19 infection can compromise the balance of immunoregulatory responses that allow graft survival, contributing to rejection in individuals infected with SARS-CoV-2 [31]. Thus, this change in the balance of attenuation of the immune response, such as a reduction in the number of regulatory T cells (Tregs) [32], may favor direct cardiotoxic action and multiple organ dysfunction.
During postoperative hospitalization, patient 2 presented Chagas reactivation, evidenced by skin biopsy, indicating that the CD was not completely controlled. In addition, it is important to consider that immunosuppressive therapy can contribute to the reactivation of CD [33]. After HT, the patient presented graft rejection and was treated with plasmapheresis and methylprednisolone. With this, before the SARS-CoV-2 infection, the patient was already weakened and died 21 days after the infection, in relation to patient 1 who died after 47 days of infection.
4. Conclusions
This report highlights the first case of an association between COVID-19, CD, and CMV dissemination in HT recipients. The patients had rapid disease progression to death. We believe that HT and the usage of immunosuppressive drugs, as well as immunosuppression generated by concomitant infections, may be an important risk factor for the development of severe COVID-19, especially in endemic areas with underreported CD infection.
Author Contributions
Conceptualization, S.C.G.-S., G.B., and R.W.A.; investigation, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., and S.M.; resources, M.N.S. and A.J.d.S.D.; writing—original draft preparation, S.C.G.-S., G.B., and R.W.A.; writing—review and editing, S.C.G.-S., G.B., and R.W.A.; visualization, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., S.M., A.J.d.S.D., and M.N.S.; supervision, M.N.S.; funding acquisition, A.J.d.S.D. and M.N.S. All authors have read and agreed to the published version of the manuscript.
Funding
This research was funded by Fundação de Amparo à Pesquisa (FAPESP), grant numbers 2019/22448-0, 19/02679-7, 2017/18199-9, 2018/18230-6, and 2019/26928-6. Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, CAPES:88887.503842/2020-00.
Institutional Review Board Statement
This study was approved by the local ethics committee (HCFMUSP nº 30800520.7.0000.0068-2020) and was carried out in accordance with the 2013 revision of the Declaration of Helsinki.
Informed Consent Statement
Informed consent was obtained from all of the subjects involved in the study.
Data Availability Statement
The laboratory data and imaging exams presented herein came from Hospital das Clinics, Faculty of Medicine, University of São Paulo (HCFMUSP) with prior approval for their use.
Conflicts of Interest
The authors declare no conflict of interest.
Figure 1 Chest X-ray of (a) patient 1 from May 2020 and (b) patient 2 from June 2015, taken just before heart transplantation, showing the Chagasic cardiomyopathy.
Figure 2 Daily clinical features of patients, from the first day of hospitalization in the ICU until death. Blood levels of (A) erythrocytes, (B) hemoglobin, (C) leukocytes, (D) neutrophils, (E) eosinophils, (F) lymphocytes, (G) monocytes, (H) neutrophil-to-lymphocyte ratio, (I) platelets, (J) creatinine, (K) urea, (L) prothrombin time, (M) activated partial thromboplastin time, (N) D-dimer, (O) pH, (P) pO2, (Q) pCO2, (R) oxygen peripheral, (S) CPR, (T) glucose, (U) CK-MB, (V) NT-proBNP, (W) lactate dehydrogenase, and (X) troponin T. Gray boxes represent the reference values.
tropicalmed-06-00022-t001_Table 1 Table 1 Patients’ characteristics, comorbidities, treatment, and complications.
Patient 1 Patient 2
Sex Female Male
Age 55 62
Positive SARS-CoV-2 RT-PCR date 1 June 2020 29 June 2020
Heart transplant date 2 May 2020 5 December 2019
Death date 30 days 207 days
Days between positive SARS-CoV-2 diagnosis and death 47 days 21 days
Complications during ICU stay ARDS due to COVID–19; heart transplant rejection, disseminated cytomegalovirus; aggravated chronic kidney disease and pressure ulcer ARDS due to COVID–19, disseminated cytomegalovirus and pancytopenia due to hemophagocytosis
Previous comorbidities Dilated cardiomyopathy of Chagas etiology, hypothyroidism by thyroidectomy by nodule 10 years ago Cardiomyopathy of Chagas etiology, disseminated cytomegalovirus, deep vein thrombosis, systemic arterial hypertension, diabetes, dyslipidemia, and chronic renal failure
Previous use of medications Carvedilol, losartan, furosemide, levothyroxine, isosorbide, and hydralazine Unknown
Medicines used during ICU stay Methylprednisolone, azathioprine, anti-thymocyte globulin, cyclosporine, tacrolimus, meropenem, linezolid, micafungin, vancomycin, polymyxin B, tigecycline, amikacin, fluconazole, ganciclovir, and hydrocortisone Meropenem, colistin, linezolid
fluconazole, amikacin, cyclosporine, azathioprine, and prednisone
ARD, acute respiratory distress syndrome; COVID-19, coronavirus disease 2019; ICU, intensive care unit; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. | Fatal | ReactionOutcome | CC BY | 33579042 | 19,681,859 | 2021-02-10 |
What was the outcome of reaction 'Dyspnoea'? | SARS-CoV-2 Infection and CMV Dissemination in Transplant Recipients as a Treatment for Chagas Cardiomyopathy: A Case Report.
Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 has infected over 90 million people worldwide, therefore it is considered a pandemic. SARS-CoV-2 infection can lead to severe pneumonia, acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure. Individuals receiving a heart transplantation (HT) may be at higher risk of adverse outcomes attributable to COVID-19 due to immunosuppressives, as well as concomitant infections that may also influence the prognoses. Herein, we describe the first report of two cases of HT recipients with concomitant infections by SARS-CoV-2, Trypanosoma cruzi, and cytomegalovirus (CMV) dissemination, from the first day of hospitalization due to COVID-19 in the intensive care unit (ICU) until the death of the patients.
1. Introduction
Coronavirus disease 2019 (COVID-19), an infectious respiratory disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spread on a pandemic scale since the first case was reported in Wuhan, China, in 2019 [1]. Most patients with the disease have mild-to-moderate symptoms; however, approximately 15% develop severe pneumonia, while approximately 5% develop acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure [2]. Lymphopenia is a recurrent feature in these patients, with a significant reduction in CD4+ T cells, CD8+ T cells, B cells, and natural killer (NK) cells [3], increasing the susceptibility of patients to severe illness and co-infection [4]. In fact, co-infections are associated with worsening of the clinical condition [5].
Chagas disease (CD) is a zoonosis whose etiologic agent is the protozoan Trypanosoma cruzi. It is estimated that 6–8 million people worldwide are infected [6]. CD has two distinct phases: an acute one, which is rare, with a strong production of type 1 cytokines, and a chronic phase, which develops in 30–40% of CD cases. The chronic phase of CD can be characterized by cardiomyopathy, arrhythmias, megaviscera, and, more rarely, polyneuropathy and stroke [7]. TCD4+ and TCD8+ lymphocytes are the main cells responsible for controlling parasitic infection. However, the immune response also contributes to tissue damage and pathology [8].
Chagasic cardiomyopathy represents the main cause of mortality from this disease, which can lead to heart failure, whose indication for treatment, especially in endemic countries, may include heart transplantation (HT) as a strategy to curb the evolution of this complication [6,9]. However, the immunosuppression and possible reactivation of the causative agent T. cruzi following HT require intensive clinical care and laboratory monitoring [10].
Other infections such as that caused by cytomegalovirus (CMV), a herpes virus that infects up to 60–100% of people in adulthood, are associated with transplant complications [11]. Primary CMV infection is generally asymptomatic in immunocompetent individuals, with the virus generating a latent infection. On the contrary, in immunocompromised and immunosuppressed populations, such as solid organ transplantation, hematopoietic stem cell transplantation, and HIV/AIDS patients, CMV reactivation is responsible for significant morbidity and mortality [12]. Although reactivation of infections such as CMV and CD are often described in transplant recipients, there are no reports of concomitant infection by T. cruzi, CMV, and SARS-CoV-2 in the context of HT in the literature to date.
Therefore, in this report, we investigated the progress of two patients who underwent HT at the Heart Institute of Hospital das Clínicas (Incor) and were subsequently transferred to the special intensive care unit (ICU) of the Hospital das Clínicas (Hospital das Clínicas, Faculty of Medicine, University of São Paulo-HCFMUSP) due to SARS-CoV-2 infection. These patients were diagnosed with COVID-19 by nasopharyngeal detection of SARS-CoV-2 RNA using reverse transcriptase polymerase chain reaction (RT-PCR). In addition, during hospitalization, CMV dissemination was evidenced by quantitative DNA detection in the blood. We describe herein the laboratory data from the first day of hospitalization due to COVID-19 until the death of the patients.
The baseline characteristics of these two patients, as well as their past clinical data, are summarized in Table 1.
2. Case Report
2.1. Patient 1
Female, 55 years old, presenting positive serology for CD since 2015, received HT on 2 May as a treatment for Chagas cardiomyopathy (Figure 1A) and underwent immunosuppressive therapy with methylprednisolone and azathioprine. Post-transplantation, she developed pneumonia, treated with meropenem and linezolid, and a remittent Candida tropicalis infection, treated with micafungin, with improvement. However, on 29 May, she developed dyspnea and desaturation, with a chest tomography suggestive of COVID-19. Her polymerase chain reaction (PCR) for SARS-CoV-2 was positive on 1 June.
On 3 June, she was transferred to the ICU specialized for treatment of severe SARS-CoV-2 infections. Laboratory analyses then showed that the patient had a reduced number of erythrocytes and hemoglobin level, and these reductions became more accentuated by day 33 in the ICU (Figure 2A,B). On this same day, the number of neutrophils peaked (Figure 2D). From the 4th to the 16th day in ICU, important lymphopenia developed (Figure 2F), resulting in an increase in the neutrophil-to-lymphocyte ratio in the same period (Figure 2H). Additionally, from the eighth day onward, she presented severe thrombocytopenia that persisted until death (Figure 2I).
Throughout the ICU hospitalization period, there were sustained high levels of creatinine, urea, D-dimer, C-reactive protein (CRP), and lactate dehydrogenase (Figure 2J,K,N,S,W). On the 32nd day, she presented a sharp increase in prothrombin time and activated partial thromboplastin time.
The cardiac function markers of creatine kinase myocardial band (CK-MB) and troponin remained elevated from the first days of ICU admission until death (Figure 2U,X). On the day of admission to the ICU, the N-terminal pro b-type natriuretic peptide (NT pro-BNP) was 70,000 pg/mL (reference value of <125 pg/mL) (Figure 2V). On the third day of hospitalization, NT pro-BNP (155,117 pg/mL) and troponin (0.28 ng/mL, reference value of <0.014 ng/mL) peaked, being related to the period in which she presented signs of graft rejection, which was treated with pulse methylprednisolone, thymoglobulin, and plasmapheresis. In addition, disseminated CMV infection was diagnosed (RT-PCR viral loads of 122 IU/mL on day 10 and 54 IU/mL on day 19 of hospitalization), for which she received ganciclovir.
After 47 days after diagnosis of SARS-CoV-2, the patient died (18 July) due to multiple organ dysfunctions associated with COVID-19.
2.2. Patient 2
Male, 62 years old, previously diagnosed with CD, received HT on 5 December 2019 as treatment for Chagas cardiomyopathy (Figure 1B). He underwent immunosuppressive therapy with cyclosporine, azathioprine, and prednisone. During postoperative hospitalization he presented Chagas reactivation characterized by skin biopsy and humoral graft rejection, being treated with plasmapheresis and methylprednisolone.
The patient was admitted to the ICU of the Hospital das Clínicas on 8 June, presenting cellulitis, deep vein thrombosis, and Chagas reactivation (a lower limb chagoma). The latter was treated with benznidazol. During hospitalization, he tested positive for PCR of SARS-CoV-2 on 29 June. He developed ARDS and septic and cardiogenic shock, which were the causes of his death.
Throughout the ICU hospitalization period, he maintained decreased levels of erythrocytes, hemoglobin, and lymphocytes (Figure 2A,B,F), as well as a high neutrophil/lymphocyte ratio (Figure 2H). From the 12th day, the number of platelets decreased and continued until the time of death (Figure 2I). The values of creatinine, urea, and glucose also remained high throughout the hospitalization period (Figure 2J,K,T). In addition, disseminated CMV (viral loads of 711 IU/mL detected on the 28th day and 1477 IU/mL on the 35th day of hospitalization) was diagnosed and treated with ganciclovir.
One day before death, the following laboratory parameters peaked: D-dimer (7612 ng/mL FEU, reference value of <500 ng/mL FEU), CRP (280.6 ng/mL, reference value of 0.300 ng/mL), CK-MB (8.38 ng/mL, reference value of 0.10–4.94 ng/mL), NT pro-BNP (55,393 pg/mL), lactate dehydrogenase (1161 U/L, reference value of 135–225 U/L), and troponin (0.701 ng/mL) (Figure 2N,S,U–X).
On 20 July, 36 days after admission to the ICU and 21 days after a positive COVID-19 diagnosis, the patient died.
3. Discussion
Herein, we described the first report of triple infection (SARS-CoV-2 infection, T. cruzi infection, and CMV dissemination) in HT recipients. Patients received HT as a form of treatment for Chagas cardiomyopathy.
We described the laboratory data from the first day of hospitalization in the ICU due to COVID-19 until the time of death. Both patients were admitted to a referral center for treatment for COVID-19 in the metropolitan region of São Paulo, a city in southeastern Brazil. We hypothesize that the triple infection by SARS-CoV-2 and CMV may have been an important cause of death and of the worsening in CD patients with HT. To date, there have been no similar reports of patients presenting these three concomitant infectious diseases and HT receptors.
COVID-19 infection among transplant recipients increases the potential for developing severe illness [13] and may vary between different organ transplants [14,15].
SARS-CoV-2 entry´s receptor angiotensin converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2) are expressed in different tissues, such as the lungs, heart, liver, kidneys, testicles, thyroid, and adipose tissue [16]. Some patients with COVID-19 develop severe disease characterized by respiratory distress syndrome and systemic manifestations. This condition has been associated with the dysregulated release of pro-inflammatory cytokines, termed “cytokine storm,” which may induce multi-organ failure [1,17].
The use of immunosuppressants and post-surgical opportunistic infections can also lead to damage of multisystem organs or even death [18]. There is a therapeutic paradox here, because while insufficient immunosuppression results in graft loss due to rejection, excessive immunosuppression can result in serious infection, including SARS-COV-2 [13], besides contributing to the reactivation of pathogens [19].
Long-term administration of immunosuppressants to solid organ transplant (SOT) receptors to reduce the risk of graft rejection may increase the risk of respiratory infections [20], although there is no clear clinical evidence of increased morbidity/mortality in SARS-CoV-2 infection [21].
Fernandez-Ruiz and collaborators [22] described a cohort of SOT receptors affected by COVID-19, 44% of whom were kidney transplant recipients, 33.3% liver transplant recipients, and 22.2% heart transplant recipients. The lethality rate was 27.8%, suggesting that SARS-CoV-2 infection had a severe course in SOT recipients.
Recently, we described the first patients with CD affected by SARS-CoV-2. The CD patients presented an increase in COVID-19 laboratory hallmarks and a rapid disease progression. Despite the efforts of the health staff, both patients died [3].
HT may be associated with the reactivation of pathogens, as reported in CD [10]. In a Brazilian cohort, the reactivation rate of Chagas disease after heart transplantation was reported to be 38.8% [23]. This fact made HT as a treatment for Chagas cardiomyopathy initially controversial. However, currently, especially in endemic countries, it is the most viable therapeutic option for patients with end-stage heart failure [24]. We hypothesize that, in the cases presented here, HT followed by reactivation of CD conferred an additional risk factor for the worsening of COVID-19.
Transplant recipients are also commonly affected by CMV reactivation, being associated with significant morbidity and mortality [12] that may worsen the infectious condition of COVID-19 [25]. In SOT, risk factors include the use of immunosuppressants for transplantation, advanced age, acute rejection, and other concomitant infections [11], with all these characteristics being present in both patients described herein. In the present report, we observed that immunosuppression may have contributed to the susceptibility to superinfections and more severe clinical manifestations in individuals undergoing HT.
Since overactivated immune responses can be one of the causes of organ damage, the anti-inflammatory effects of immunosuppression can be protective, reducing the cytokine storm related to complications in COVID-19 [26]. In this context, it has been described that immunosuppressive therapy with calcineurin inhibitors in patients with solid organ transplantation or systemic rheumatic diseases promotes a clinical course in SARS-CoV-2 infection, which is generally mild, and with an apparently low risk of superinfection [27]. In addition, immunosuppression has not been evaluated as a risk factor for SARS or MERS [28].
However, in the present case report, we observed that immunosuppression may have contributed to the susceptibility to SARS-CoV-2 infection, the reactivation of pathogens, and more serious clinical manifestations in individuals undergoing HT. Corroborating our findings, it was shown that patients with COVID-19 and cancer, due to their systemic immunosuppressive condition caused by malignancy and anticancer treatments, such as chemotherapy, had an increased risk of SARS-CoV-2 infection and a worse prognosis [29]. It has also been reported that chronic use of corticosteroids prior to SARS-CoV-2 contamination is associated with critical disease outcomes, including a high risk of death [30].
These contradictory observations show that knowledge about the relationship between COVID-19 and the patient’s immune condition is limited. Further studies are needed to elucidate the immune responses and prognosis of COVID-19.
In general, HT in patient 1 proved to be successful for the treatment of Chagas cardiomyopathy, with no reactivation of the pathogen. However, after SARS-CoV-2 infection, she presented graft rejection, treated with methylprednisolone, thymoglobulin, and plasmapheresis. There is a description in eye transplantation that COVID-19 infection can compromise the balance of immunoregulatory responses that allow graft survival, contributing to rejection in individuals infected with SARS-CoV-2 [31]. Thus, this change in the balance of attenuation of the immune response, such as a reduction in the number of regulatory T cells (Tregs) [32], may favor direct cardiotoxic action and multiple organ dysfunction.
During postoperative hospitalization, patient 2 presented Chagas reactivation, evidenced by skin biopsy, indicating that the CD was not completely controlled. In addition, it is important to consider that immunosuppressive therapy can contribute to the reactivation of CD [33]. After HT, the patient presented graft rejection and was treated with plasmapheresis and methylprednisolone. With this, before the SARS-CoV-2 infection, the patient was already weakened and died 21 days after the infection, in relation to patient 1 who died after 47 days of infection.
4. Conclusions
This report highlights the first case of an association between COVID-19, CD, and CMV dissemination in HT recipients. The patients had rapid disease progression to death. We believe that HT and the usage of immunosuppressive drugs, as well as immunosuppression generated by concomitant infections, may be an important risk factor for the development of severe COVID-19, especially in endemic areas with underreported CD infection.
Author Contributions
Conceptualization, S.C.G.-S., G.B., and R.W.A.; investigation, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., and S.M.; resources, M.N.S. and A.J.d.S.D.; writing—original draft preparation, S.C.G.-S., G.B., and R.W.A.; writing—review and editing, S.C.G.-S., G.B., and R.W.A.; visualization, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., S.M., A.J.d.S.D., and M.N.S.; supervision, M.N.S.; funding acquisition, A.J.d.S.D. and M.N.S. All authors have read and agreed to the published version of the manuscript.
Funding
This research was funded by Fundação de Amparo à Pesquisa (FAPESP), grant numbers 2019/22448-0, 19/02679-7, 2017/18199-9, 2018/18230-6, and 2019/26928-6. Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, CAPES:88887.503842/2020-00.
Institutional Review Board Statement
This study was approved by the local ethics committee (HCFMUSP nº 30800520.7.0000.0068-2020) and was carried out in accordance with the 2013 revision of the Declaration of Helsinki.
Informed Consent Statement
Informed consent was obtained from all of the subjects involved in the study.
Data Availability Statement
The laboratory data and imaging exams presented herein came from Hospital das Clinics, Faculty of Medicine, University of São Paulo (HCFMUSP) with prior approval for their use.
Conflicts of Interest
The authors declare no conflict of interest.
Figure 1 Chest X-ray of (a) patient 1 from May 2020 and (b) patient 2 from June 2015, taken just before heart transplantation, showing the Chagasic cardiomyopathy.
Figure 2 Daily clinical features of patients, from the first day of hospitalization in the ICU until death. Blood levels of (A) erythrocytes, (B) hemoglobin, (C) leukocytes, (D) neutrophils, (E) eosinophils, (F) lymphocytes, (G) monocytes, (H) neutrophil-to-lymphocyte ratio, (I) platelets, (J) creatinine, (K) urea, (L) prothrombin time, (M) activated partial thromboplastin time, (N) D-dimer, (O) pH, (P) pO2, (Q) pCO2, (R) oxygen peripheral, (S) CPR, (T) glucose, (U) CK-MB, (V) NT-proBNP, (W) lactate dehydrogenase, and (X) troponin T. Gray boxes represent the reference values.
tropicalmed-06-00022-t001_Table 1 Table 1 Patients’ characteristics, comorbidities, treatment, and complications.
Patient 1 Patient 2
Sex Female Male
Age 55 62
Positive SARS-CoV-2 RT-PCR date 1 June 2020 29 June 2020
Heart transplant date 2 May 2020 5 December 2019
Death date 30 days 207 days
Days between positive SARS-CoV-2 diagnosis and death 47 days 21 days
Complications during ICU stay ARDS due to COVID–19; heart transplant rejection, disseminated cytomegalovirus; aggravated chronic kidney disease and pressure ulcer ARDS due to COVID–19, disseminated cytomegalovirus and pancytopenia due to hemophagocytosis
Previous comorbidities Dilated cardiomyopathy of Chagas etiology, hypothyroidism by thyroidectomy by nodule 10 years ago Cardiomyopathy of Chagas etiology, disseminated cytomegalovirus, deep vein thrombosis, systemic arterial hypertension, diabetes, dyslipidemia, and chronic renal failure
Previous use of medications Carvedilol, losartan, furosemide, levothyroxine, isosorbide, and hydralazine Unknown
Medicines used during ICU stay Methylprednisolone, azathioprine, anti-thymocyte globulin, cyclosporine, tacrolimus, meropenem, linezolid, micafungin, vancomycin, polymyxin B, tigecycline, amikacin, fluconazole, ganciclovir, and hydrocortisone Meropenem, colistin, linezolid
fluconazole, amikacin, cyclosporine, azathioprine, and prednisone
ARD, acute respiratory distress syndrome; COVID-19, coronavirus disease 2019; ICU, intensive care unit; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. | Fatal | ReactionOutcome | CC BY | 33579042 | 19,681,859 | 2021-02-10 |
What was the outcome of reaction 'Haemophagocytic lymphohistiocytosis'? | SARS-CoV-2 Infection and CMV Dissemination in Transplant Recipients as a Treatment for Chagas Cardiomyopathy: A Case Report.
Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 has infected over 90 million people worldwide, therefore it is considered a pandemic. SARS-CoV-2 infection can lead to severe pneumonia, acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure. Individuals receiving a heart transplantation (HT) may be at higher risk of adverse outcomes attributable to COVID-19 due to immunosuppressives, as well as concomitant infections that may also influence the prognoses. Herein, we describe the first report of two cases of HT recipients with concomitant infections by SARS-CoV-2, Trypanosoma cruzi, and cytomegalovirus (CMV) dissemination, from the first day of hospitalization due to COVID-19 in the intensive care unit (ICU) until the death of the patients.
1. Introduction
Coronavirus disease 2019 (COVID-19), an infectious respiratory disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spread on a pandemic scale since the first case was reported in Wuhan, China, in 2019 [1]. Most patients with the disease have mild-to-moderate symptoms; however, approximately 15% develop severe pneumonia, while approximately 5% develop acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure [2]. Lymphopenia is a recurrent feature in these patients, with a significant reduction in CD4+ T cells, CD8+ T cells, B cells, and natural killer (NK) cells [3], increasing the susceptibility of patients to severe illness and co-infection [4]. In fact, co-infections are associated with worsening of the clinical condition [5].
Chagas disease (CD) is a zoonosis whose etiologic agent is the protozoan Trypanosoma cruzi. It is estimated that 6–8 million people worldwide are infected [6]. CD has two distinct phases: an acute one, which is rare, with a strong production of type 1 cytokines, and a chronic phase, which develops in 30–40% of CD cases. The chronic phase of CD can be characterized by cardiomyopathy, arrhythmias, megaviscera, and, more rarely, polyneuropathy and stroke [7]. TCD4+ and TCD8+ lymphocytes are the main cells responsible for controlling parasitic infection. However, the immune response also contributes to tissue damage and pathology [8].
Chagasic cardiomyopathy represents the main cause of mortality from this disease, which can lead to heart failure, whose indication for treatment, especially in endemic countries, may include heart transplantation (HT) as a strategy to curb the evolution of this complication [6,9]. However, the immunosuppression and possible reactivation of the causative agent T. cruzi following HT require intensive clinical care and laboratory monitoring [10].
Other infections such as that caused by cytomegalovirus (CMV), a herpes virus that infects up to 60–100% of people in adulthood, are associated with transplant complications [11]. Primary CMV infection is generally asymptomatic in immunocompetent individuals, with the virus generating a latent infection. On the contrary, in immunocompromised and immunosuppressed populations, such as solid organ transplantation, hematopoietic stem cell transplantation, and HIV/AIDS patients, CMV reactivation is responsible for significant morbidity and mortality [12]. Although reactivation of infections such as CMV and CD are often described in transplant recipients, there are no reports of concomitant infection by T. cruzi, CMV, and SARS-CoV-2 in the context of HT in the literature to date.
Therefore, in this report, we investigated the progress of two patients who underwent HT at the Heart Institute of Hospital das Clínicas (Incor) and were subsequently transferred to the special intensive care unit (ICU) of the Hospital das Clínicas (Hospital das Clínicas, Faculty of Medicine, University of São Paulo-HCFMUSP) due to SARS-CoV-2 infection. These patients were diagnosed with COVID-19 by nasopharyngeal detection of SARS-CoV-2 RNA using reverse transcriptase polymerase chain reaction (RT-PCR). In addition, during hospitalization, CMV dissemination was evidenced by quantitative DNA detection in the blood. We describe herein the laboratory data from the first day of hospitalization due to COVID-19 until the death of the patients.
The baseline characteristics of these two patients, as well as their past clinical data, are summarized in Table 1.
2. Case Report
2.1. Patient 1
Female, 55 years old, presenting positive serology for CD since 2015, received HT on 2 May as a treatment for Chagas cardiomyopathy (Figure 1A) and underwent immunosuppressive therapy with methylprednisolone and azathioprine. Post-transplantation, she developed pneumonia, treated with meropenem and linezolid, and a remittent Candida tropicalis infection, treated with micafungin, with improvement. However, on 29 May, she developed dyspnea and desaturation, with a chest tomography suggestive of COVID-19. Her polymerase chain reaction (PCR) for SARS-CoV-2 was positive on 1 June.
On 3 June, she was transferred to the ICU specialized for treatment of severe SARS-CoV-2 infections. Laboratory analyses then showed that the patient had a reduced number of erythrocytes and hemoglobin level, and these reductions became more accentuated by day 33 in the ICU (Figure 2A,B). On this same day, the number of neutrophils peaked (Figure 2D). From the 4th to the 16th day in ICU, important lymphopenia developed (Figure 2F), resulting in an increase in the neutrophil-to-lymphocyte ratio in the same period (Figure 2H). Additionally, from the eighth day onward, she presented severe thrombocytopenia that persisted until death (Figure 2I).
Throughout the ICU hospitalization period, there were sustained high levels of creatinine, urea, D-dimer, C-reactive protein (CRP), and lactate dehydrogenase (Figure 2J,K,N,S,W). On the 32nd day, she presented a sharp increase in prothrombin time and activated partial thromboplastin time.
The cardiac function markers of creatine kinase myocardial band (CK-MB) and troponin remained elevated from the first days of ICU admission until death (Figure 2U,X). On the day of admission to the ICU, the N-terminal pro b-type natriuretic peptide (NT pro-BNP) was 70,000 pg/mL (reference value of <125 pg/mL) (Figure 2V). On the third day of hospitalization, NT pro-BNP (155,117 pg/mL) and troponin (0.28 ng/mL, reference value of <0.014 ng/mL) peaked, being related to the period in which she presented signs of graft rejection, which was treated with pulse methylprednisolone, thymoglobulin, and plasmapheresis. In addition, disseminated CMV infection was diagnosed (RT-PCR viral loads of 122 IU/mL on day 10 and 54 IU/mL on day 19 of hospitalization), for which she received ganciclovir.
After 47 days after diagnosis of SARS-CoV-2, the patient died (18 July) due to multiple organ dysfunctions associated with COVID-19.
2.2. Patient 2
Male, 62 years old, previously diagnosed with CD, received HT on 5 December 2019 as treatment for Chagas cardiomyopathy (Figure 1B). He underwent immunosuppressive therapy with cyclosporine, azathioprine, and prednisone. During postoperative hospitalization he presented Chagas reactivation characterized by skin biopsy and humoral graft rejection, being treated with plasmapheresis and methylprednisolone.
The patient was admitted to the ICU of the Hospital das Clínicas on 8 June, presenting cellulitis, deep vein thrombosis, and Chagas reactivation (a lower limb chagoma). The latter was treated with benznidazol. During hospitalization, he tested positive for PCR of SARS-CoV-2 on 29 June. He developed ARDS and septic and cardiogenic shock, which were the causes of his death.
Throughout the ICU hospitalization period, he maintained decreased levels of erythrocytes, hemoglobin, and lymphocytes (Figure 2A,B,F), as well as a high neutrophil/lymphocyte ratio (Figure 2H). From the 12th day, the number of platelets decreased and continued until the time of death (Figure 2I). The values of creatinine, urea, and glucose also remained high throughout the hospitalization period (Figure 2J,K,T). In addition, disseminated CMV (viral loads of 711 IU/mL detected on the 28th day and 1477 IU/mL on the 35th day of hospitalization) was diagnosed and treated with ganciclovir.
One day before death, the following laboratory parameters peaked: D-dimer (7612 ng/mL FEU, reference value of <500 ng/mL FEU), CRP (280.6 ng/mL, reference value of 0.300 ng/mL), CK-MB (8.38 ng/mL, reference value of 0.10–4.94 ng/mL), NT pro-BNP (55,393 pg/mL), lactate dehydrogenase (1161 U/L, reference value of 135–225 U/L), and troponin (0.701 ng/mL) (Figure 2N,S,U–X).
On 20 July, 36 days after admission to the ICU and 21 days after a positive COVID-19 diagnosis, the patient died.
3. Discussion
Herein, we described the first report of triple infection (SARS-CoV-2 infection, T. cruzi infection, and CMV dissemination) in HT recipients. Patients received HT as a form of treatment for Chagas cardiomyopathy.
We described the laboratory data from the first day of hospitalization in the ICU due to COVID-19 until the time of death. Both patients were admitted to a referral center for treatment for COVID-19 in the metropolitan region of São Paulo, a city in southeastern Brazil. We hypothesize that the triple infection by SARS-CoV-2 and CMV may have been an important cause of death and of the worsening in CD patients with HT. To date, there have been no similar reports of patients presenting these three concomitant infectious diseases and HT receptors.
COVID-19 infection among transplant recipients increases the potential for developing severe illness [13] and may vary between different organ transplants [14,15].
SARS-CoV-2 entry´s receptor angiotensin converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2) are expressed in different tissues, such as the lungs, heart, liver, kidneys, testicles, thyroid, and adipose tissue [16]. Some patients with COVID-19 develop severe disease characterized by respiratory distress syndrome and systemic manifestations. This condition has been associated with the dysregulated release of pro-inflammatory cytokines, termed “cytokine storm,” which may induce multi-organ failure [1,17].
The use of immunosuppressants and post-surgical opportunistic infections can also lead to damage of multisystem organs or even death [18]. There is a therapeutic paradox here, because while insufficient immunosuppression results in graft loss due to rejection, excessive immunosuppression can result in serious infection, including SARS-COV-2 [13], besides contributing to the reactivation of pathogens [19].
Long-term administration of immunosuppressants to solid organ transplant (SOT) receptors to reduce the risk of graft rejection may increase the risk of respiratory infections [20], although there is no clear clinical evidence of increased morbidity/mortality in SARS-CoV-2 infection [21].
Fernandez-Ruiz and collaborators [22] described a cohort of SOT receptors affected by COVID-19, 44% of whom were kidney transplant recipients, 33.3% liver transplant recipients, and 22.2% heart transplant recipients. The lethality rate was 27.8%, suggesting that SARS-CoV-2 infection had a severe course in SOT recipients.
Recently, we described the first patients with CD affected by SARS-CoV-2. The CD patients presented an increase in COVID-19 laboratory hallmarks and a rapid disease progression. Despite the efforts of the health staff, both patients died [3].
HT may be associated with the reactivation of pathogens, as reported in CD [10]. In a Brazilian cohort, the reactivation rate of Chagas disease after heart transplantation was reported to be 38.8% [23]. This fact made HT as a treatment for Chagas cardiomyopathy initially controversial. However, currently, especially in endemic countries, it is the most viable therapeutic option for patients with end-stage heart failure [24]. We hypothesize that, in the cases presented here, HT followed by reactivation of CD conferred an additional risk factor for the worsening of COVID-19.
Transplant recipients are also commonly affected by CMV reactivation, being associated with significant morbidity and mortality [12] that may worsen the infectious condition of COVID-19 [25]. In SOT, risk factors include the use of immunosuppressants for transplantation, advanced age, acute rejection, and other concomitant infections [11], with all these characteristics being present in both patients described herein. In the present report, we observed that immunosuppression may have contributed to the susceptibility to superinfections and more severe clinical manifestations in individuals undergoing HT.
Since overactivated immune responses can be one of the causes of organ damage, the anti-inflammatory effects of immunosuppression can be protective, reducing the cytokine storm related to complications in COVID-19 [26]. In this context, it has been described that immunosuppressive therapy with calcineurin inhibitors in patients with solid organ transplantation or systemic rheumatic diseases promotes a clinical course in SARS-CoV-2 infection, which is generally mild, and with an apparently low risk of superinfection [27]. In addition, immunosuppression has not been evaluated as a risk factor for SARS or MERS [28].
However, in the present case report, we observed that immunosuppression may have contributed to the susceptibility to SARS-CoV-2 infection, the reactivation of pathogens, and more serious clinical manifestations in individuals undergoing HT. Corroborating our findings, it was shown that patients with COVID-19 and cancer, due to their systemic immunosuppressive condition caused by malignancy and anticancer treatments, such as chemotherapy, had an increased risk of SARS-CoV-2 infection and a worse prognosis [29]. It has also been reported that chronic use of corticosteroids prior to SARS-CoV-2 contamination is associated with critical disease outcomes, including a high risk of death [30].
These contradictory observations show that knowledge about the relationship between COVID-19 and the patient’s immune condition is limited. Further studies are needed to elucidate the immune responses and prognosis of COVID-19.
In general, HT in patient 1 proved to be successful for the treatment of Chagas cardiomyopathy, with no reactivation of the pathogen. However, after SARS-CoV-2 infection, she presented graft rejection, treated with methylprednisolone, thymoglobulin, and plasmapheresis. There is a description in eye transplantation that COVID-19 infection can compromise the balance of immunoregulatory responses that allow graft survival, contributing to rejection in individuals infected with SARS-CoV-2 [31]. Thus, this change in the balance of attenuation of the immune response, such as a reduction in the number of regulatory T cells (Tregs) [32], may favor direct cardiotoxic action and multiple organ dysfunction.
During postoperative hospitalization, patient 2 presented Chagas reactivation, evidenced by skin biopsy, indicating that the CD was not completely controlled. In addition, it is important to consider that immunosuppressive therapy can contribute to the reactivation of CD [33]. After HT, the patient presented graft rejection and was treated with plasmapheresis and methylprednisolone. With this, before the SARS-CoV-2 infection, the patient was already weakened and died 21 days after the infection, in relation to patient 1 who died after 47 days of infection.
4. Conclusions
This report highlights the first case of an association between COVID-19, CD, and CMV dissemination in HT recipients. The patients had rapid disease progression to death. We believe that HT and the usage of immunosuppressive drugs, as well as immunosuppression generated by concomitant infections, may be an important risk factor for the development of severe COVID-19, especially in endemic areas with underreported CD infection.
Author Contributions
Conceptualization, S.C.G.-S., G.B., and R.W.A.; investigation, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., and S.M.; resources, M.N.S. and A.J.d.S.D.; writing—original draft preparation, S.C.G.-S., G.B., and R.W.A.; writing—review and editing, S.C.G.-S., G.B., and R.W.A.; visualization, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., S.M., A.J.d.S.D., and M.N.S.; supervision, M.N.S.; funding acquisition, A.J.d.S.D. and M.N.S. All authors have read and agreed to the published version of the manuscript.
Funding
This research was funded by Fundação de Amparo à Pesquisa (FAPESP), grant numbers 2019/22448-0, 19/02679-7, 2017/18199-9, 2018/18230-6, and 2019/26928-6. Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, CAPES:88887.503842/2020-00.
Institutional Review Board Statement
This study was approved by the local ethics committee (HCFMUSP nº 30800520.7.0000.0068-2020) and was carried out in accordance with the 2013 revision of the Declaration of Helsinki.
Informed Consent Statement
Informed consent was obtained from all of the subjects involved in the study.
Data Availability Statement
The laboratory data and imaging exams presented herein came from Hospital das Clinics, Faculty of Medicine, University of São Paulo (HCFMUSP) with prior approval for their use.
Conflicts of Interest
The authors declare no conflict of interest.
Figure 1 Chest X-ray of (a) patient 1 from May 2020 and (b) patient 2 from June 2015, taken just before heart transplantation, showing the Chagasic cardiomyopathy.
Figure 2 Daily clinical features of patients, from the first day of hospitalization in the ICU until death. Blood levels of (A) erythrocytes, (B) hemoglobin, (C) leukocytes, (D) neutrophils, (E) eosinophils, (F) lymphocytes, (G) monocytes, (H) neutrophil-to-lymphocyte ratio, (I) platelets, (J) creatinine, (K) urea, (L) prothrombin time, (M) activated partial thromboplastin time, (N) D-dimer, (O) pH, (P) pO2, (Q) pCO2, (R) oxygen peripheral, (S) CPR, (T) glucose, (U) CK-MB, (V) NT-proBNP, (W) lactate dehydrogenase, and (X) troponin T. Gray boxes represent the reference values.
tropicalmed-06-00022-t001_Table 1 Table 1 Patients’ characteristics, comorbidities, treatment, and complications.
Patient 1 Patient 2
Sex Female Male
Age 55 62
Positive SARS-CoV-2 RT-PCR date 1 June 2020 29 June 2020
Heart transplant date 2 May 2020 5 December 2019
Death date 30 days 207 days
Days between positive SARS-CoV-2 diagnosis and death 47 days 21 days
Complications during ICU stay ARDS due to COVID–19; heart transplant rejection, disseminated cytomegalovirus; aggravated chronic kidney disease and pressure ulcer ARDS due to COVID–19, disseminated cytomegalovirus and pancytopenia due to hemophagocytosis
Previous comorbidities Dilated cardiomyopathy of Chagas etiology, hypothyroidism by thyroidectomy by nodule 10 years ago Cardiomyopathy of Chagas etiology, disseminated cytomegalovirus, deep vein thrombosis, systemic arterial hypertension, diabetes, dyslipidemia, and chronic renal failure
Previous use of medications Carvedilol, losartan, furosemide, levothyroxine, isosorbide, and hydralazine Unknown
Medicines used during ICU stay Methylprednisolone, azathioprine, anti-thymocyte globulin, cyclosporine, tacrolimus, meropenem, linezolid, micafungin, vancomycin, polymyxin B, tigecycline, amikacin, fluconazole, ganciclovir, and hydrocortisone Meropenem, colistin, linezolid
fluconazole, amikacin, cyclosporine, azathioprine, and prednisone
ARD, acute respiratory distress syndrome; COVID-19, coronavirus disease 2019; ICU, intensive care unit; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. | Fatal | ReactionOutcome | CC BY | 33579042 | 19,731,502 | 2021-02-10 |
What was the outcome of reaction 'Infection reactivation'? | SARS-CoV-2 Infection and CMV Dissemination in Transplant Recipients as a Treatment for Chagas Cardiomyopathy: A Case Report.
Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 has infected over 90 million people worldwide, therefore it is considered a pandemic. SARS-CoV-2 infection can lead to severe pneumonia, acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure. Individuals receiving a heart transplantation (HT) may be at higher risk of adverse outcomes attributable to COVID-19 due to immunosuppressives, as well as concomitant infections that may also influence the prognoses. Herein, we describe the first report of two cases of HT recipients with concomitant infections by SARS-CoV-2, Trypanosoma cruzi, and cytomegalovirus (CMV) dissemination, from the first day of hospitalization due to COVID-19 in the intensive care unit (ICU) until the death of the patients.
1. Introduction
Coronavirus disease 2019 (COVID-19), an infectious respiratory disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spread on a pandemic scale since the first case was reported in Wuhan, China, in 2019 [1]. Most patients with the disease have mild-to-moderate symptoms; however, approximately 15% develop severe pneumonia, while approximately 5% develop acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure [2]. Lymphopenia is a recurrent feature in these patients, with a significant reduction in CD4+ T cells, CD8+ T cells, B cells, and natural killer (NK) cells [3], increasing the susceptibility of patients to severe illness and co-infection [4]. In fact, co-infections are associated with worsening of the clinical condition [5].
Chagas disease (CD) is a zoonosis whose etiologic agent is the protozoan Trypanosoma cruzi. It is estimated that 6–8 million people worldwide are infected [6]. CD has two distinct phases: an acute one, which is rare, with a strong production of type 1 cytokines, and a chronic phase, which develops in 30–40% of CD cases. The chronic phase of CD can be characterized by cardiomyopathy, arrhythmias, megaviscera, and, more rarely, polyneuropathy and stroke [7]. TCD4+ and TCD8+ lymphocytes are the main cells responsible for controlling parasitic infection. However, the immune response also contributes to tissue damage and pathology [8].
Chagasic cardiomyopathy represents the main cause of mortality from this disease, which can lead to heart failure, whose indication for treatment, especially in endemic countries, may include heart transplantation (HT) as a strategy to curb the evolution of this complication [6,9]. However, the immunosuppression and possible reactivation of the causative agent T. cruzi following HT require intensive clinical care and laboratory monitoring [10].
Other infections such as that caused by cytomegalovirus (CMV), a herpes virus that infects up to 60–100% of people in adulthood, are associated with transplant complications [11]. Primary CMV infection is generally asymptomatic in immunocompetent individuals, with the virus generating a latent infection. On the contrary, in immunocompromised and immunosuppressed populations, such as solid organ transplantation, hematopoietic stem cell transplantation, and HIV/AIDS patients, CMV reactivation is responsible for significant morbidity and mortality [12]. Although reactivation of infections such as CMV and CD are often described in transplant recipients, there are no reports of concomitant infection by T. cruzi, CMV, and SARS-CoV-2 in the context of HT in the literature to date.
Therefore, in this report, we investigated the progress of two patients who underwent HT at the Heart Institute of Hospital das Clínicas (Incor) and were subsequently transferred to the special intensive care unit (ICU) of the Hospital das Clínicas (Hospital das Clínicas, Faculty of Medicine, University of São Paulo-HCFMUSP) due to SARS-CoV-2 infection. These patients were diagnosed with COVID-19 by nasopharyngeal detection of SARS-CoV-2 RNA using reverse transcriptase polymerase chain reaction (RT-PCR). In addition, during hospitalization, CMV dissemination was evidenced by quantitative DNA detection in the blood. We describe herein the laboratory data from the first day of hospitalization due to COVID-19 until the death of the patients.
The baseline characteristics of these two patients, as well as their past clinical data, are summarized in Table 1.
2. Case Report
2.1. Patient 1
Female, 55 years old, presenting positive serology for CD since 2015, received HT on 2 May as a treatment for Chagas cardiomyopathy (Figure 1A) and underwent immunosuppressive therapy with methylprednisolone and azathioprine. Post-transplantation, she developed pneumonia, treated with meropenem and linezolid, and a remittent Candida tropicalis infection, treated with micafungin, with improvement. However, on 29 May, she developed dyspnea and desaturation, with a chest tomography suggestive of COVID-19. Her polymerase chain reaction (PCR) for SARS-CoV-2 was positive on 1 June.
On 3 June, she was transferred to the ICU specialized for treatment of severe SARS-CoV-2 infections. Laboratory analyses then showed that the patient had a reduced number of erythrocytes and hemoglobin level, and these reductions became more accentuated by day 33 in the ICU (Figure 2A,B). On this same day, the number of neutrophils peaked (Figure 2D). From the 4th to the 16th day in ICU, important lymphopenia developed (Figure 2F), resulting in an increase in the neutrophil-to-lymphocyte ratio in the same period (Figure 2H). Additionally, from the eighth day onward, she presented severe thrombocytopenia that persisted until death (Figure 2I).
Throughout the ICU hospitalization period, there were sustained high levels of creatinine, urea, D-dimer, C-reactive protein (CRP), and lactate dehydrogenase (Figure 2J,K,N,S,W). On the 32nd day, she presented a sharp increase in prothrombin time and activated partial thromboplastin time.
The cardiac function markers of creatine kinase myocardial band (CK-MB) and troponin remained elevated from the first days of ICU admission until death (Figure 2U,X). On the day of admission to the ICU, the N-terminal pro b-type natriuretic peptide (NT pro-BNP) was 70,000 pg/mL (reference value of <125 pg/mL) (Figure 2V). On the third day of hospitalization, NT pro-BNP (155,117 pg/mL) and troponin (0.28 ng/mL, reference value of <0.014 ng/mL) peaked, being related to the period in which she presented signs of graft rejection, which was treated with pulse methylprednisolone, thymoglobulin, and plasmapheresis. In addition, disseminated CMV infection was diagnosed (RT-PCR viral loads of 122 IU/mL on day 10 and 54 IU/mL on day 19 of hospitalization), for which she received ganciclovir.
After 47 days after diagnosis of SARS-CoV-2, the patient died (18 July) due to multiple organ dysfunctions associated with COVID-19.
2.2. Patient 2
Male, 62 years old, previously diagnosed with CD, received HT on 5 December 2019 as treatment for Chagas cardiomyopathy (Figure 1B). He underwent immunosuppressive therapy with cyclosporine, azathioprine, and prednisone. During postoperative hospitalization he presented Chagas reactivation characterized by skin biopsy and humoral graft rejection, being treated with plasmapheresis and methylprednisolone.
The patient was admitted to the ICU of the Hospital das Clínicas on 8 June, presenting cellulitis, deep vein thrombosis, and Chagas reactivation (a lower limb chagoma). The latter was treated with benznidazol. During hospitalization, he tested positive for PCR of SARS-CoV-2 on 29 June. He developed ARDS and septic and cardiogenic shock, which were the causes of his death.
Throughout the ICU hospitalization period, he maintained decreased levels of erythrocytes, hemoglobin, and lymphocytes (Figure 2A,B,F), as well as a high neutrophil/lymphocyte ratio (Figure 2H). From the 12th day, the number of platelets decreased and continued until the time of death (Figure 2I). The values of creatinine, urea, and glucose also remained high throughout the hospitalization period (Figure 2J,K,T). In addition, disseminated CMV (viral loads of 711 IU/mL detected on the 28th day and 1477 IU/mL on the 35th day of hospitalization) was diagnosed and treated with ganciclovir.
One day before death, the following laboratory parameters peaked: D-dimer (7612 ng/mL FEU, reference value of <500 ng/mL FEU), CRP (280.6 ng/mL, reference value of 0.300 ng/mL), CK-MB (8.38 ng/mL, reference value of 0.10–4.94 ng/mL), NT pro-BNP (55,393 pg/mL), lactate dehydrogenase (1161 U/L, reference value of 135–225 U/L), and troponin (0.701 ng/mL) (Figure 2N,S,U–X).
On 20 July, 36 days after admission to the ICU and 21 days after a positive COVID-19 diagnosis, the patient died.
3. Discussion
Herein, we described the first report of triple infection (SARS-CoV-2 infection, T. cruzi infection, and CMV dissemination) in HT recipients. Patients received HT as a form of treatment for Chagas cardiomyopathy.
We described the laboratory data from the first day of hospitalization in the ICU due to COVID-19 until the time of death. Both patients were admitted to a referral center for treatment for COVID-19 in the metropolitan region of São Paulo, a city in southeastern Brazil. We hypothesize that the triple infection by SARS-CoV-2 and CMV may have been an important cause of death and of the worsening in CD patients with HT. To date, there have been no similar reports of patients presenting these three concomitant infectious diseases and HT receptors.
COVID-19 infection among transplant recipients increases the potential for developing severe illness [13] and may vary between different organ transplants [14,15].
SARS-CoV-2 entry´s receptor angiotensin converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2) are expressed in different tissues, such as the lungs, heart, liver, kidneys, testicles, thyroid, and adipose tissue [16]. Some patients with COVID-19 develop severe disease characterized by respiratory distress syndrome and systemic manifestations. This condition has been associated with the dysregulated release of pro-inflammatory cytokines, termed “cytokine storm,” which may induce multi-organ failure [1,17].
The use of immunosuppressants and post-surgical opportunistic infections can also lead to damage of multisystem organs or even death [18]. There is a therapeutic paradox here, because while insufficient immunosuppression results in graft loss due to rejection, excessive immunosuppression can result in serious infection, including SARS-COV-2 [13], besides contributing to the reactivation of pathogens [19].
Long-term administration of immunosuppressants to solid organ transplant (SOT) receptors to reduce the risk of graft rejection may increase the risk of respiratory infections [20], although there is no clear clinical evidence of increased morbidity/mortality in SARS-CoV-2 infection [21].
Fernandez-Ruiz and collaborators [22] described a cohort of SOT receptors affected by COVID-19, 44% of whom were kidney transplant recipients, 33.3% liver transplant recipients, and 22.2% heart transplant recipients. The lethality rate was 27.8%, suggesting that SARS-CoV-2 infection had a severe course in SOT recipients.
Recently, we described the first patients with CD affected by SARS-CoV-2. The CD patients presented an increase in COVID-19 laboratory hallmarks and a rapid disease progression. Despite the efforts of the health staff, both patients died [3].
HT may be associated with the reactivation of pathogens, as reported in CD [10]. In a Brazilian cohort, the reactivation rate of Chagas disease after heart transplantation was reported to be 38.8% [23]. This fact made HT as a treatment for Chagas cardiomyopathy initially controversial. However, currently, especially in endemic countries, it is the most viable therapeutic option for patients with end-stage heart failure [24]. We hypothesize that, in the cases presented here, HT followed by reactivation of CD conferred an additional risk factor for the worsening of COVID-19.
Transplant recipients are also commonly affected by CMV reactivation, being associated with significant morbidity and mortality [12] that may worsen the infectious condition of COVID-19 [25]. In SOT, risk factors include the use of immunosuppressants for transplantation, advanced age, acute rejection, and other concomitant infections [11], with all these characteristics being present in both patients described herein. In the present report, we observed that immunosuppression may have contributed to the susceptibility to superinfections and more severe clinical manifestations in individuals undergoing HT.
Since overactivated immune responses can be one of the causes of organ damage, the anti-inflammatory effects of immunosuppression can be protective, reducing the cytokine storm related to complications in COVID-19 [26]. In this context, it has been described that immunosuppressive therapy with calcineurin inhibitors in patients with solid organ transplantation or systemic rheumatic diseases promotes a clinical course in SARS-CoV-2 infection, which is generally mild, and with an apparently low risk of superinfection [27]. In addition, immunosuppression has not been evaluated as a risk factor for SARS or MERS [28].
However, in the present case report, we observed that immunosuppression may have contributed to the susceptibility to SARS-CoV-2 infection, the reactivation of pathogens, and more serious clinical manifestations in individuals undergoing HT. Corroborating our findings, it was shown that patients with COVID-19 and cancer, due to their systemic immunosuppressive condition caused by malignancy and anticancer treatments, such as chemotherapy, had an increased risk of SARS-CoV-2 infection and a worse prognosis [29]. It has also been reported that chronic use of corticosteroids prior to SARS-CoV-2 contamination is associated with critical disease outcomes, including a high risk of death [30].
These contradictory observations show that knowledge about the relationship between COVID-19 and the patient’s immune condition is limited. Further studies are needed to elucidate the immune responses and prognosis of COVID-19.
In general, HT in patient 1 proved to be successful for the treatment of Chagas cardiomyopathy, with no reactivation of the pathogen. However, after SARS-CoV-2 infection, she presented graft rejection, treated with methylprednisolone, thymoglobulin, and plasmapheresis. There is a description in eye transplantation that COVID-19 infection can compromise the balance of immunoregulatory responses that allow graft survival, contributing to rejection in individuals infected with SARS-CoV-2 [31]. Thus, this change in the balance of attenuation of the immune response, such as a reduction in the number of regulatory T cells (Tregs) [32], may favor direct cardiotoxic action and multiple organ dysfunction.
During postoperative hospitalization, patient 2 presented Chagas reactivation, evidenced by skin biopsy, indicating that the CD was not completely controlled. In addition, it is important to consider that immunosuppressive therapy can contribute to the reactivation of CD [33]. After HT, the patient presented graft rejection and was treated with plasmapheresis and methylprednisolone. With this, before the SARS-CoV-2 infection, the patient was already weakened and died 21 days after the infection, in relation to patient 1 who died after 47 days of infection.
4. Conclusions
This report highlights the first case of an association between COVID-19, CD, and CMV dissemination in HT recipients. The patients had rapid disease progression to death. We believe that HT and the usage of immunosuppressive drugs, as well as immunosuppression generated by concomitant infections, may be an important risk factor for the development of severe COVID-19, especially in endemic areas with underreported CD infection.
Author Contributions
Conceptualization, S.C.G.-S., G.B., and R.W.A.; investigation, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., and S.M.; resources, M.N.S. and A.J.d.S.D.; writing—original draft preparation, S.C.G.-S., G.B., and R.W.A.; writing—review and editing, S.C.G.-S., G.B., and R.W.A.; visualization, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., S.M., A.J.d.S.D., and M.N.S.; supervision, M.N.S.; funding acquisition, A.J.d.S.D. and M.N.S. All authors have read and agreed to the published version of the manuscript.
Funding
This research was funded by Fundação de Amparo à Pesquisa (FAPESP), grant numbers 2019/22448-0, 19/02679-7, 2017/18199-9, 2018/18230-6, and 2019/26928-6. Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, CAPES:88887.503842/2020-00.
Institutional Review Board Statement
This study was approved by the local ethics committee (HCFMUSP nº 30800520.7.0000.0068-2020) and was carried out in accordance with the 2013 revision of the Declaration of Helsinki.
Informed Consent Statement
Informed consent was obtained from all of the subjects involved in the study.
Data Availability Statement
The laboratory data and imaging exams presented herein came from Hospital das Clinics, Faculty of Medicine, University of São Paulo (HCFMUSP) with prior approval for their use.
Conflicts of Interest
The authors declare no conflict of interest.
Figure 1 Chest X-ray of (a) patient 1 from May 2020 and (b) patient 2 from June 2015, taken just before heart transplantation, showing the Chagasic cardiomyopathy.
Figure 2 Daily clinical features of patients, from the first day of hospitalization in the ICU until death. Blood levels of (A) erythrocytes, (B) hemoglobin, (C) leukocytes, (D) neutrophils, (E) eosinophils, (F) lymphocytes, (G) monocytes, (H) neutrophil-to-lymphocyte ratio, (I) platelets, (J) creatinine, (K) urea, (L) prothrombin time, (M) activated partial thromboplastin time, (N) D-dimer, (O) pH, (P) pO2, (Q) pCO2, (R) oxygen peripheral, (S) CPR, (T) glucose, (U) CK-MB, (V) NT-proBNP, (W) lactate dehydrogenase, and (X) troponin T. Gray boxes represent the reference values.
tropicalmed-06-00022-t001_Table 1 Table 1 Patients’ characteristics, comorbidities, treatment, and complications.
Patient 1 Patient 2
Sex Female Male
Age 55 62
Positive SARS-CoV-2 RT-PCR date 1 June 2020 29 June 2020
Heart transplant date 2 May 2020 5 December 2019
Death date 30 days 207 days
Days between positive SARS-CoV-2 diagnosis and death 47 days 21 days
Complications during ICU stay ARDS due to COVID–19; heart transplant rejection, disseminated cytomegalovirus; aggravated chronic kidney disease and pressure ulcer ARDS due to COVID–19, disseminated cytomegalovirus and pancytopenia due to hemophagocytosis
Previous comorbidities Dilated cardiomyopathy of Chagas etiology, hypothyroidism by thyroidectomy by nodule 10 years ago Cardiomyopathy of Chagas etiology, disseminated cytomegalovirus, deep vein thrombosis, systemic arterial hypertension, diabetes, dyslipidemia, and chronic renal failure
Previous use of medications Carvedilol, losartan, furosemide, levothyroxine, isosorbide, and hydralazine Unknown
Medicines used during ICU stay Methylprednisolone, azathioprine, anti-thymocyte globulin, cyclosporine, tacrolimus, meropenem, linezolid, micafungin, vancomycin, polymyxin B, tigecycline, amikacin, fluconazole, ganciclovir, and hydrocortisone Meropenem, colistin, linezolid
fluconazole, amikacin, cyclosporine, azathioprine, and prednisone
ARD, acute respiratory distress syndrome; COVID-19, coronavirus disease 2019; ICU, intensive care unit; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. | Fatal | ReactionOutcome | CC BY | 33579042 | 19,731,502 | 2021-02-10 |
What was the outcome of reaction 'Lymphopenia'? | SARS-CoV-2 Infection and CMV Dissemination in Transplant Recipients as a Treatment for Chagas Cardiomyopathy: A Case Report.
Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 has infected over 90 million people worldwide, therefore it is considered a pandemic. SARS-CoV-2 infection can lead to severe pneumonia, acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure. Individuals receiving a heart transplantation (HT) may be at higher risk of adverse outcomes attributable to COVID-19 due to immunosuppressives, as well as concomitant infections that may also influence the prognoses. Herein, we describe the first report of two cases of HT recipients with concomitant infections by SARS-CoV-2, Trypanosoma cruzi, and cytomegalovirus (CMV) dissemination, from the first day of hospitalization due to COVID-19 in the intensive care unit (ICU) until the death of the patients.
1. Introduction
Coronavirus disease 2019 (COVID-19), an infectious respiratory disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spread on a pandemic scale since the first case was reported in Wuhan, China, in 2019 [1]. Most patients with the disease have mild-to-moderate symptoms; however, approximately 15% develop severe pneumonia, while approximately 5% develop acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure [2]. Lymphopenia is a recurrent feature in these patients, with a significant reduction in CD4+ T cells, CD8+ T cells, B cells, and natural killer (NK) cells [3], increasing the susceptibility of patients to severe illness and co-infection [4]. In fact, co-infections are associated with worsening of the clinical condition [5].
Chagas disease (CD) is a zoonosis whose etiologic agent is the protozoan Trypanosoma cruzi. It is estimated that 6–8 million people worldwide are infected [6]. CD has two distinct phases: an acute one, which is rare, with a strong production of type 1 cytokines, and a chronic phase, which develops in 30–40% of CD cases. The chronic phase of CD can be characterized by cardiomyopathy, arrhythmias, megaviscera, and, more rarely, polyneuropathy and stroke [7]. TCD4+ and TCD8+ lymphocytes are the main cells responsible for controlling parasitic infection. However, the immune response also contributes to tissue damage and pathology [8].
Chagasic cardiomyopathy represents the main cause of mortality from this disease, which can lead to heart failure, whose indication for treatment, especially in endemic countries, may include heart transplantation (HT) as a strategy to curb the evolution of this complication [6,9]. However, the immunosuppression and possible reactivation of the causative agent T. cruzi following HT require intensive clinical care and laboratory monitoring [10].
Other infections such as that caused by cytomegalovirus (CMV), a herpes virus that infects up to 60–100% of people in adulthood, are associated with transplant complications [11]. Primary CMV infection is generally asymptomatic in immunocompetent individuals, with the virus generating a latent infection. On the contrary, in immunocompromised and immunosuppressed populations, such as solid organ transplantation, hematopoietic stem cell transplantation, and HIV/AIDS patients, CMV reactivation is responsible for significant morbidity and mortality [12]. Although reactivation of infections such as CMV and CD are often described in transplant recipients, there are no reports of concomitant infection by T. cruzi, CMV, and SARS-CoV-2 in the context of HT in the literature to date.
Therefore, in this report, we investigated the progress of two patients who underwent HT at the Heart Institute of Hospital das Clínicas (Incor) and were subsequently transferred to the special intensive care unit (ICU) of the Hospital das Clínicas (Hospital das Clínicas, Faculty of Medicine, University of São Paulo-HCFMUSP) due to SARS-CoV-2 infection. These patients were diagnosed with COVID-19 by nasopharyngeal detection of SARS-CoV-2 RNA using reverse transcriptase polymerase chain reaction (RT-PCR). In addition, during hospitalization, CMV dissemination was evidenced by quantitative DNA detection in the blood. We describe herein the laboratory data from the first day of hospitalization due to COVID-19 until the death of the patients.
The baseline characteristics of these two patients, as well as their past clinical data, are summarized in Table 1.
2. Case Report
2.1. Patient 1
Female, 55 years old, presenting positive serology for CD since 2015, received HT on 2 May as a treatment for Chagas cardiomyopathy (Figure 1A) and underwent immunosuppressive therapy with methylprednisolone and azathioprine. Post-transplantation, she developed pneumonia, treated with meropenem and linezolid, and a remittent Candida tropicalis infection, treated with micafungin, with improvement. However, on 29 May, she developed dyspnea and desaturation, with a chest tomography suggestive of COVID-19. Her polymerase chain reaction (PCR) for SARS-CoV-2 was positive on 1 June.
On 3 June, she was transferred to the ICU specialized for treatment of severe SARS-CoV-2 infections. Laboratory analyses then showed that the patient had a reduced number of erythrocytes and hemoglobin level, and these reductions became more accentuated by day 33 in the ICU (Figure 2A,B). On this same day, the number of neutrophils peaked (Figure 2D). From the 4th to the 16th day in ICU, important lymphopenia developed (Figure 2F), resulting in an increase in the neutrophil-to-lymphocyte ratio in the same period (Figure 2H). Additionally, from the eighth day onward, she presented severe thrombocytopenia that persisted until death (Figure 2I).
Throughout the ICU hospitalization period, there were sustained high levels of creatinine, urea, D-dimer, C-reactive protein (CRP), and lactate dehydrogenase (Figure 2J,K,N,S,W). On the 32nd day, she presented a sharp increase in prothrombin time and activated partial thromboplastin time.
The cardiac function markers of creatine kinase myocardial band (CK-MB) and troponin remained elevated from the first days of ICU admission until death (Figure 2U,X). On the day of admission to the ICU, the N-terminal pro b-type natriuretic peptide (NT pro-BNP) was 70,000 pg/mL (reference value of <125 pg/mL) (Figure 2V). On the third day of hospitalization, NT pro-BNP (155,117 pg/mL) and troponin (0.28 ng/mL, reference value of <0.014 ng/mL) peaked, being related to the period in which she presented signs of graft rejection, which was treated with pulse methylprednisolone, thymoglobulin, and plasmapheresis. In addition, disseminated CMV infection was diagnosed (RT-PCR viral loads of 122 IU/mL on day 10 and 54 IU/mL on day 19 of hospitalization), for which she received ganciclovir.
After 47 days after diagnosis of SARS-CoV-2, the patient died (18 July) due to multiple organ dysfunctions associated with COVID-19.
2.2. Patient 2
Male, 62 years old, previously diagnosed with CD, received HT on 5 December 2019 as treatment for Chagas cardiomyopathy (Figure 1B). He underwent immunosuppressive therapy with cyclosporine, azathioprine, and prednisone. During postoperative hospitalization he presented Chagas reactivation characterized by skin biopsy and humoral graft rejection, being treated with plasmapheresis and methylprednisolone.
The patient was admitted to the ICU of the Hospital das Clínicas on 8 June, presenting cellulitis, deep vein thrombosis, and Chagas reactivation (a lower limb chagoma). The latter was treated with benznidazol. During hospitalization, he tested positive for PCR of SARS-CoV-2 on 29 June. He developed ARDS and septic and cardiogenic shock, which were the causes of his death.
Throughout the ICU hospitalization period, he maintained decreased levels of erythrocytes, hemoglobin, and lymphocytes (Figure 2A,B,F), as well as a high neutrophil/lymphocyte ratio (Figure 2H). From the 12th day, the number of platelets decreased and continued until the time of death (Figure 2I). The values of creatinine, urea, and glucose also remained high throughout the hospitalization period (Figure 2J,K,T). In addition, disseminated CMV (viral loads of 711 IU/mL detected on the 28th day and 1477 IU/mL on the 35th day of hospitalization) was diagnosed and treated with ganciclovir.
One day before death, the following laboratory parameters peaked: D-dimer (7612 ng/mL FEU, reference value of <500 ng/mL FEU), CRP (280.6 ng/mL, reference value of 0.300 ng/mL), CK-MB (8.38 ng/mL, reference value of 0.10–4.94 ng/mL), NT pro-BNP (55,393 pg/mL), lactate dehydrogenase (1161 U/L, reference value of 135–225 U/L), and troponin (0.701 ng/mL) (Figure 2N,S,U–X).
On 20 July, 36 days after admission to the ICU and 21 days after a positive COVID-19 diagnosis, the patient died.
3. Discussion
Herein, we described the first report of triple infection (SARS-CoV-2 infection, T. cruzi infection, and CMV dissemination) in HT recipients. Patients received HT as a form of treatment for Chagas cardiomyopathy.
We described the laboratory data from the first day of hospitalization in the ICU due to COVID-19 until the time of death. Both patients were admitted to a referral center for treatment for COVID-19 in the metropolitan region of São Paulo, a city in southeastern Brazil. We hypothesize that the triple infection by SARS-CoV-2 and CMV may have been an important cause of death and of the worsening in CD patients with HT. To date, there have been no similar reports of patients presenting these three concomitant infectious diseases and HT receptors.
COVID-19 infection among transplant recipients increases the potential for developing severe illness [13] and may vary between different organ transplants [14,15].
SARS-CoV-2 entry´s receptor angiotensin converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2) are expressed in different tissues, such as the lungs, heart, liver, kidneys, testicles, thyroid, and adipose tissue [16]. Some patients with COVID-19 develop severe disease characterized by respiratory distress syndrome and systemic manifestations. This condition has been associated with the dysregulated release of pro-inflammatory cytokines, termed “cytokine storm,” which may induce multi-organ failure [1,17].
The use of immunosuppressants and post-surgical opportunistic infections can also lead to damage of multisystem organs or even death [18]. There is a therapeutic paradox here, because while insufficient immunosuppression results in graft loss due to rejection, excessive immunosuppression can result in serious infection, including SARS-COV-2 [13], besides contributing to the reactivation of pathogens [19].
Long-term administration of immunosuppressants to solid organ transplant (SOT) receptors to reduce the risk of graft rejection may increase the risk of respiratory infections [20], although there is no clear clinical evidence of increased morbidity/mortality in SARS-CoV-2 infection [21].
Fernandez-Ruiz and collaborators [22] described a cohort of SOT receptors affected by COVID-19, 44% of whom were kidney transplant recipients, 33.3% liver transplant recipients, and 22.2% heart transplant recipients. The lethality rate was 27.8%, suggesting that SARS-CoV-2 infection had a severe course in SOT recipients.
Recently, we described the first patients with CD affected by SARS-CoV-2. The CD patients presented an increase in COVID-19 laboratory hallmarks and a rapid disease progression. Despite the efforts of the health staff, both patients died [3].
HT may be associated with the reactivation of pathogens, as reported in CD [10]. In a Brazilian cohort, the reactivation rate of Chagas disease after heart transplantation was reported to be 38.8% [23]. This fact made HT as a treatment for Chagas cardiomyopathy initially controversial. However, currently, especially in endemic countries, it is the most viable therapeutic option for patients with end-stage heart failure [24]. We hypothesize that, in the cases presented here, HT followed by reactivation of CD conferred an additional risk factor for the worsening of COVID-19.
Transplant recipients are also commonly affected by CMV reactivation, being associated with significant morbidity and mortality [12] that may worsen the infectious condition of COVID-19 [25]. In SOT, risk factors include the use of immunosuppressants for transplantation, advanced age, acute rejection, and other concomitant infections [11], with all these characteristics being present in both patients described herein. In the present report, we observed that immunosuppression may have contributed to the susceptibility to superinfections and more severe clinical manifestations in individuals undergoing HT.
Since overactivated immune responses can be one of the causes of organ damage, the anti-inflammatory effects of immunosuppression can be protective, reducing the cytokine storm related to complications in COVID-19 [26]. In this context, it has been described that immunosuppressive therapy with calcineurin inhibitors in patients with solid organ transplantation or systemic rheumatic diseases promotes a clinical course in SARS-CoV-2 infection, which is generally mild, and with an apparently low risk of superinfection [27]. In addition, immunosuppression has not been evaluated as a risk factor for SARS or MERS [28].
However, in the present case report, we observed that immunosuppression may have contributed to the susceptibility to SARS-CoV-2 infection, the reactivation of pathogens, and more serious clinical manifestations in individuals undergoing HT. Corroborating our findings, it was shown that patients with COVID-19 and cancer, due to their systemic immunosuppressive condition caused by malignancy and anticancer treatments, such as chemotherapy, had an increased risk of SARS-CoV-2 infection and a worse prognosis [29]. It has also been reported that chronic use of corticosteroids prior to SARS-CoV-2 contamination is associated with critical disease outcomes, including a high risk of death [30].
These contradictory observations show that knowledge about the relationship between COVID-19 and the patient’s immune condition is limited. Further studies are needed to elucidate the immune responses and prognosis of COVID-19.
In general, HT in patient 1 proved to be successful for the treatment of Chagas cardiomyopathy, with no reactivation of the pathogen. However, after SARS-CoV-2 infection, she presented graft rejection, treated with methylprednisolone, thymoglobulin, and plasmapheresis. There is a description in eye transplantation that COVID-19 infection can compromise the balance of immunoregulatory responses that allow graft survival, contributing to rejection in individuals infected with SARS-CoV-2 [31]. Thus, this change in the balance of attenuation of the immune response, such as a reduction in the number of regulatory T cells (Tregs) [32], may favor direct cardiotoxic action and multiple organ dysfunction.
During postoperative hospitalization, patient 2 presented Chagas reactivation, evidenced by skin biopsy, indicating that the CD was not completely controlled. In addition, it is important to consider that immunosuppressive therapy can contribute to the reactivation of CD [33]. After HT, the patient presented graft rejection and was treated with plasmapheresis and methylprednisolone. With this, before the SARS-CoV-2 infection, the patient was already weakened and died 21 days after the infection, in relation to patient 1 who died after 47 days of infection.
4. Conclusions
This report highlights the first case of an association between COVID-19, CD, and CMV dissemination in HT recipients. The patients had rapid disease progression to death. We believe that HT and the usage of immunosuppressive drugs, as well as immunosuppression generated by concomitant infections, may be an important risk factor for the development of severe COVID-19, especially in endemic areas with underreported CD infection.
Author Contributions
Conceptualization, S.C.G.-S., G.B., and R.W.A.; investigation, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., and S.M.; resources, M.N.S. and A.J.d.S.D.; writing—original draft preparation, S.C.G.-S., G.B., and R.W.A.; writing—review and editing, S.C.G.-S., G.B., and R.W.A.; visualization, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., S.M., A.J.d.S.D., and M.N.S.; supervision, M.N.S.; funding acquisition, A.J.d.S.D. and M.N.S. All authors have read and agreed to the published version of the manuscript.
Funding
This research was funded by Fundação de Amparo à Pesquisa (FAPESP), grant numbers 2019/22448-0, 19/02679-7, 2017/18199-9, 2018/18230-6, and 2019/26928-6. Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, CAPES:88887.503842/2020-00.
Institutional Review Board Statement
This study was approved by the local ethics committee (HCFMUSP nº 30800520.7.0000.0068-2020) and was carried out in accordance with the 2013 revision of the Declaration of Helsinki.
Informed Consent Statement
Informed consent was obtained from all of the subjects involved in the study.
Data Availability Statement
The laboratory data and imaging exams presented herein came from Hospital das Clinics, Faculty of Medicine, University of São Paulo (HCFMUSP) with prior approval for their use.
Conflicts of Interest
The authors declare no conflict of interest.
Figure 1 Chest X-ray of (a) patient 1 from May 2020 and (b) patient 2 from June 2015, taken just before heart transplantation, showing the Chagasic cardiomyopathy.
Figure 2 Daily clinical features of patients, from the first day of hospitalization in the ICU until death. Blood levels of (A) erythrocytes, (B) hemoglobin, (C) leukocytes, (D) neutrophils, (E) eosinophils, (F) lymphocytes, (G) monocytes, (H) neutrophil-to-lymphocyte ratio, (I) platelets, (J) creatinine, (K) urea, (L) prothrombin time, (M) activated partial thromboplastin time, (N) D-dimer, (O) pH, (P) pO2, (Q) pCO2, (R) oxygen peripheral, (S) CPR, (T) glucose, (U) CK-MB, (V) NT-proBNP, (W) lactate dehydrogenase, and (X) troponin T. Gray boxes represent the reference values.
tropicalmed-06-00022-t001_Table 1 Table 1 Patients’ characteristics, comorbidities, treatment, and complications.
Patient 1 Patient 2
Sex Female Male
Age 55 62
Positive SARS-CoV-2 RT-PCR date 1 June 2020 29 June 2020
Heart transplant date 2 May 2020 5 December 2019
Death date 30 days 207 days
Days between positive SARS-CoV-2 diagnosis and death 47 days 21 days
Complications during ICU stay ARDS due to COVID–19; heart transplant rejection, disseminated cytomegalovirus; aggravated chronic kidney disease and pressure ulcer ARDS due to COVID–19, disseminated cytomegalovirus and pancytopenia due to hemophagocytosis
Previous comorbidities Dilated cardiomyopathy of Chagas etiology, hypothyroidism by thyroidectomy by nodule 10 years ago Cardiomyopathy of Chagas etiology, disseminated cytomegalovirus, deep vein thrombosis, systemic arterial hypertension, diabetes, dyslipidemia, and chronic renal failure
Previous use of medications Carvedilol, losartan, furosemide, levothyroxine, isosorbide, and hydralazine Unknown
Medicines used during ICU stay Methylprednisolone, azathioprine, anti-thymocyte globulin, cyclosporine, tacrolimus, meropenem, linezolid, micafungin, vancomycin, polymyxin B, tigecycline, amikacin, fluconazole, ganciclovir, and hydrocortisone Meropenem, colistin, linezolid
fluconazole, amikacin, cyclosporine, azathioprine, and prednisone
ARD, acute respiratory distress syndrome; COVID-19, coronavirus disease 2019; ICU, intensive care unit; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. | Fatal | ReactionOutcome | CC BY | 33579042 | 19,681,859 | 2021-02-10 |
What was the outcome of reaction 'Multiple organ dysfunction syndrome'? | SARS-CoV-2 Infection and CMV Dissemination in Transplant Recipients as a Treatment for Chagas Cardiomyopathy: A Case Report.
Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 has infected over 90 million people worldwide, therefore it is considered a pandemic. SARS-CoV-2 infection can lead to severe pneumonia, acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure. Individuals receiving a heart transplantation (HT) may be at higher risk of adverse outcomes attributable to COVID-19 due to immunosuppressives, as well as concomitant infections that may also influence the prognoses. Herein, we describe the first report of two cases of HT recipients with concomitant infections by SARS-CoV-2, Trypanosoma cruzi, and cytomegalovirus (CMV) dissemination, from the first day of hospitalization due to COVID-19 in the intensive care unit (ICU) until the death of the patients.
1. Introduction
Coronavirus disease 2019 (COVID-19), an infectious respiratory disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spread on a pandemic scale since the first case was reported in Wuhan, China, in 2019 [1]. Most patients with the disease have mild-to-moderate symptoms; however, approximately 15% develop severe pneumonia, while approximately 5% develop acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure [2]. Lymphopenia is a recurrent feature in these patients, with a significant reduction in CD4+ T cells, CD8+ T cells, B cells, and natural killer (NK) cells [3], increasing the susceptibility of patients to severe illness and co-infection [4]. In fact, co-infections are associated with worsening of the clinical condition [5].
Chagas disease (CD) is a zoonosis whose etiologic agent is the protozoan Trypanosoma cruzi. It is estimated that 6–8 million people worldwide are infected [6]. CD has two distinct phases: an acute one, which is rare, with a strong production of type 1 cytokines, and a chronic phase, which develops in 30–40% of CD cases. The chronic phase of CD can be characterized by cardiomyopathy, arrhythmias, megaviscera, and, more rarely, polyneuropathy and stroke [7]. TCD4+ and TCD8+ lymphocytes are the main cells responsible for controlling parasitic infection. However, the immune response also contributes to tissue damage and pathology [8].
Chagasic cardiomyopathy represents the main cause of mortality from this disease, which can lead to heart failure, whose indication for treatment, especially in endemic countries, may include heart transplantation (HT) as a strategy to curb the evolution of this complication [6,9]. However, the immunosuppression and possible reactivation of the causative agent T. cruzi following HT require intensive clinical care and laboratory monitoring [10].
Other infections such as that caused by cytomegalovirus (CMV), a herpes virus that infects up to 60–100% of people in adulthood, are associated with transplant complications [11]. Primary CMV infection is generally asymptomatic in immunocompetent individuals, with the virus generating a latent infection. On the contrary, in immunocompromised and immunosuppressed populations, such as solid organ transplantation, hematopoietic stem cell transplantation, and HIV/AIDS patients, CMV reactivation is responsible for significant morbidity and mortality [12]. Although reactivation of infections such as CMV and CD are often described in transplant recipients, there are no reports of concomitant infection by T. cruzi, CMV, and SARS-CoV-2 in the context of HT in the literature to date.
Therefore, in this report, we investigated the progress of two patients who underwent HT at the Heart Institute of Hospital das Clínicas (Incor) and were subsequently transferred to the special intensive care unit (ICU) of the Hospital das Clínicas (Hospital das Clínicas, Faculty of Medicine, University of São Paulo-HCFMUSP) due to SARS-CoV-2 infection. These patients were diagnosed with COVID-19 by nasopharyngeal detection of SARS-CoV-2 RNA using reverse transcriptase polymerase chain reaction (RT-PCR). In addition, during hospitalization, CMV dissemination was evidenced by quantitative DNA detection in the blood. We describe herein the laboratory data from the first day of hospitalization due to COVID-19 until the death of the patients.
The baseline characteristics of these two patients, as well as their past clinical data, are summarized in Table 1.
2. Case Report
2.1. Patient 1
Female, 55 years old, presenting positive serology for CD since 2015, received HT on 2 May as a treatment for Chagas cardiomyopathy (Figure 1A) and underwent immunosuppressive therapy with methylprednisolone and azathioprine. Post-transplantation, she developed pneumonia, treated with meropenem and linezolid, and a remittent Candida tropicalis infection, treated with micafungin, with improvement. However, on 29 May, she developed dyspnea and desaturation, with a chest tomography suggestive of COVID-19. Her polymerase chain reaction (PCR) for SARS-CoV-2 was positive on 1 June.
On 3 June, she was transferred to the ICU specialized for treatment of severe SARS-CoV-2 infections. Laboratory analyses then showed that the patient had a reduced number of erythrocytes and hemoglobin level, and these reductions became more accentuated by day 33 in the ICU (Figure 2A,B). On this same day, the number of neutrophils peaked (Figure 2D). From the 4th to the 16th day in ICU, important lymphopenia developed (Figure 2F), resulting in an increase in the neutrophil-to-lymphocyte ratio in the same period (Figure 2H). Additionally, from the eighth day onward, she presented severe thrombocytopenia that persisted until death (Figure 2I).
Throughout the ICU hospitalization period, there were sustained high levels of creatinine, urea, D-dimer, C-reactive protein (CRP), and lactate dehydrogenase (Figure 2J,K,N,S,W). On the 32nd day, she presented a sharp increase in prothrombin time and activated partial thromboplastin time.
The cardiac function markers of creatine kinase myocardial band (CK-MB) and troponin remained elevated from the first days of ICU admission until death (Figure 2U,X). On the day of admission to the ICU, the N-terminal pro b-type natriuretic peptide (NT pro-BNP) was 70,000 pg/mL (reference value of <125 pg/mL) (Figure 2V). On the third day of hospitalization, NT pro-BNP (155,117 pg/mL) and troponin (0.28 ng/mL, reference value of <0.014 ng/mL) peaked, being related to the period in which she presented signs of graft rejection, which was treated with pulse methylprednisolone, thymoglobulin, and plasmapheresis. In addition, disseminated CMV infection was diagnosed (RT-PCR viral loads of 122 IU/mL on day 10 and 54 IU/mL on day 19 of hospitalization), for which she received ganciclovir.
After 47 days after diagnosis of SARS-CoV-2, the patient died (18 July) due to multiple organ dysfunctions associated with COVID-19.
2.2. Patient 2
Male, 62 years old, previously diagnosed with CD, received HT on 5 December 2019 as treatment for Chagas cardiomyopathy (Figure 1B). He underwent immunosuppressive therapy with cyclosporine, azathioprine, and prednisone. During postoperative hospitalization he presented Chagas reactivation characterized by skin biopsy and humoral graft rejection, being treated with plasmapheresis and methylprednisolone.
The patient was admitted to the ICU of the Hospital das Clínicas on 8 June, presenting cellulitis, deep vein thrombosis, and Chagas reactivation (a lower limb chagoma). The latter was treated with benznidazol. During hospitalization, he tested positive for PCR of SARS-CoV-2 on 29 June. He developed ARDS and septic and cardiogenic shock, which were the causes of his death.
Throughout the ICU hospitalization period, he maintained decreased levels of erythrocytes, hemoglobin, and lymphocytes (Figure 2A,B,F), as well as a high neutrophil/lymphocyte ratio (Figure 2H). From the 12th day, the number of platelets decreased and continued until the time of death (Figure 2I). The values of creatinine, urea, and glucose also remained high throughout the hospitalization period (Figure 2J,K,T). In addition, disseminated CMV (viral loads of 711 IU/mL detected on the 28th day and 1477 IU/mL on the 35th day of hospitalization) was diagnosed and treated with ganciclovir.
One day before death, the following laboratory parameters peaked: D-dimer (7612 ng/mL FEU, reference value of <500 ng/mL FEU), CRP (280.6 ng/mL, reference value of 0.300 ng/mL), CK-MB (8.38 ng/mL, reference value of 0.10–4.94 ng/mL), NT pro-BNP (55,393 pg/mL), lactate dehydrogenase (1161 U/L, reference value of 135–225 U/L), and troponin (0.701 ng/mL) (Figure 2N,S,U–X).
On 20 July, 36 days after admission to the ICU and 21 days after a positive COVID-19 diagnosis, the patient died.
3. Discussion
Herein, we described the first report of triple infection (SARS-CoV-2 infection, T. cruzi infection, and CMV dissemination) in HT recipients. Patients received HT as a form of treatment for Chagas cardiomyopathy.
We described the laboratory data from the first day of hospitalization in the ICU due to COVID-19 until the time of death. Both patients were admitted to a referral center for treatment for COVID-19 in the metropolitan region of São Paulo, a city in southeastern Brazil. We hypothesize that the triple infection by SARS-CoV-2 and CMV may have been an important cause of death and of the worsening in CD patients with HT. To date, there have been no similar reports of patients presenting these three concomitant infectious diseases and HT receptors.
COVID-19 infection among transplant recipients increases the potential for developing severe illness [13] and may vary between different organ transplants [14,15].
SARS-CoV-2 entry´s receptor angiotensin converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2) are expressed in different tissues, such as the lungs, heart, liver, kidneys, testicles, thyroid, and adipose tissue [16]. Some patients with COVID-19 develop severe disease characterized by respiratory distress syndrome and systemic manifestations. This condition has been associated with the dysregulated release of pro-inflammatory cytokines, termed “cytokine storm,” which may induce multi-organ failure [1,17].
The use of immunosuppressants and post-surgical opportunistic infections can also lead to damage of multisystem organs or even death [18]. There is a therapeutic paradox here, because while insufficient immunosuppression results in graft loss due to rejection, excessive immunosuppression can result in serious infection, including SARS-COV-2 [13], besides contributing to the reactivation of pathogens [19].
Long-term administration of immunosuppressants to solid organ transplant (SOT) receptors to reduce the risk of graft rejection may increase the risk of respiratory infections [20], although there is no clear clinical evidence of increased morbidity/mortality in SARS-CoV-2 infection [21].
Fernandez-Ruiz and collaborators [22] described a cohort of SOT receptors affected by COVID-19, 44% of whom were kidney transplant recipients, 33.3% liver transplant recipients, and 22.2% heart transplant recipients. The lethality rate was 27.8%, suggesting that SARS-CoV-2 infection had a severe course in SOT recipients.
Recently, we described the first patients with CD affected by SARS-CoV-2. The CD patients presented an increase in COVID-19 laboratory hallmarks and a rapid disease progression. Despite the efforts of the health staff, both patients died [3].
HT may be associated with the reactivation of pathogens, as reported in CD [10]. In a Brazilian cohort, the reactivation rate of Chagas disease after heart transplantation was reported to be 38.8% [23]. This fact made HT as a treatment for Chagas cardiomyopathy initially controversial. However, currently, especially in endemic countries, it is the most viable therapeutic option for patients with end-stage heart failure [24]. We hypothesize that, in the cases presented here, HT followed by reactivation of CD conferred an additional risk factor for the worsening of COVID-19.
Transplant recipients are also commonly affected by CMV reactivation, being associated with significant morbidity and mortality [12] that may worsen the infectious condition of COVID-19 [25]. In SOT, risk factors include the use of immunosuppressants for transplantation, advanced age, acute rejection, and other concomitant infections [11], with all these characteristics being present in both patients described herein. In the present report, we observed that immunosuppression may have contributed to the susceptibility to superinfections and more severe clinical manifestations in individuals undergoing HT.
Since overactivated immune responses can be one of the causes of organ damage, the anti-inflammatory effects of immunosuppression can be protective, reducing the cytokine storm related to complications in COVID-19 [26]. In this context, it has been described that immunosuppressive therapy with calcineurin inhibitors in patients with solid organ transplantation or systemic rheumatic diseases promotes a clinical course in SARS-CoV-2 infection, which is generally mild, and with an apparently low risk of superinfection [27]. In addition, immunosuppression has not been evaluated as a risk factor for SARS or MERS [28].
However, in the present case report, we observed that immunosuppression may have contributed to the susceptibility to SARS-CoV-2 infection, the reactivation of pathogens, and more serious clinical manifestations in individuals undergoing HT. Corroborating our findings, it was shown that patients with COVID-19 and cancer, due to their systemic immunosuppressive condition caused by malignancy and anticancer treatments, such as chemotherapy, had an increased risk of SARS-CoV-2 infection and a worse prognosis [29]. It has also been reported that chronic use of corticosteroids prior to SARS-CoV-2 contamination is associated with critical disease outcomes, including a high risk of death [30].
These contradictory observations show that knowledge about the relationship between COVID-19 and the patient’s immune condition is limited. Further studies are needed to elucidate the immune responses and prognosis of COVID-19.
In general, HT in patient 1 proved to be successful for the treatment of Chagas cardiomyopathy, with no reactivation of the pathogen. However, after SARS-CoV-2 infection, she presented graft rejection, treated with methylprednisolone, thymoglobulin, and plasmapheresis. There is a description in eye transplantation that COVID-19 infection can compromise the balance of immunoregulatory responses that allow graft survival, contributing to rejection in individuals infected with SARS-CoV-2 [31]. Thus, this change in the balance of attenuation of the immune response, such as a reduction in the number of regulatory T cells (Tregs) [32], may favor direct cardiotoxic action and multiple organ dysfunction.
During postoperative hospitalization, patient 2 presented Chagas reactivation, evidenced by skin biopsy, indicating that the CD was not completely controlled. In addition, it is important to consider that immunosuppressive therapy can contribute to the reactivation of CD [33]. After HT, the patient presented graft rejection and was treated with plasmapheresis and methylprednisolone. With this, before the SARS-CoV-2 infection, the patient was already weakened and died 21 days after the infection, in relation to patient 1 who died after 47 days of infection.
4. Conclusions
This report highlights the first case of an association between COVID-19, CD, and CMV dissemination in HT recipients. The patients had rapid disease progression to death. We believe that HT and the usage of immunosuppressive drugs, as well as immunosuppression generated by concomitant infections, may be an important risk factor for the development of severe COVID-19, especially in endemic areas with underreported CD infection.
Author Contributions
Conceptualization, S.C.G.-S., G.B., and R.W.A.; investigation, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., and S.M.; resources, M.N.S. and A.J.d.S.D.; writing—original draft preparation, S.C.G.-S., G.B., and R.W.A.; writing—review and editing, S.C.G.-S., G.B., and R.W.A.; visualization, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., S.M., A.J.d.S.D., and M.N.S.; supervision, M.N.S.; funding acquisition, A.J.d.S.D. and M.N.S. All authors have read and agreed to the published version of the manuscript.
Funding
This research was funded by Fundação de Amparo à Pesquisa (FAPESP), grant numbers 2019/22448-0, 19/02679-7, 2017/18199-9, 2018/18230-6, and 2019/26928-6. Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, CAPES:88887.503842/2020-00.
Institutional Review Board Statement
This study was approved by the local ethics committee (HCFMUSP nº 30800520.7.0000.0068-2020) and was carried out in accordance with the 2013 revision of the Declaration of Helsinki.
Informed Consent Statement
Informed consent was obtained from all of the subjects involved in the study.
Data Availability Statement
The laboratory data and imaging exams presented herein came from Hospital das Clinics, Faculty of Medicine, University of São Paulo (HCFMUSP) with prior approval for their use.
Conflicts of Interest
The authors declare no conflict of interest.
Figure 1 Chest X-ray of (a) patient 1 from May 2020 and (b) patient 2 from June 2015, taken just before heart transplantation, showing the Chagasic cardiomyopathy.
Figure 2 Daily clinical features of patients, from the first day of hospitalization in the ICU until death. Blood levels of (A) erythrocytes, (B) hemoglobin, (C) leukocytes, (D) neutrophils, (E) eosinophils, (F) lymphocytes, (G) monocytes, (H) neutrophil-to-lymphocyte ratio, (I) platelets, (J) creatinine, (K) urea, (L) prothrombin time, (M) activated partial thromboplastin time, (N) D-dimer, (O) pH, (P) pO2, (Q) pCO2, (R) oxygen peripheral, (S) CPR, (T) glucose, (U) CK-MB, (V) NT-proBNP, (W) lactate dehydrogenase, and (X) troponin T. Gray boxes represent the reference values.
tropicalmed-06-00022-t001_Table 1 Table 1 Patients’ characteristics, comorbidities, treatment, and complications.
Patient 1 Patient 2
Sex Female Male
Age 55 62
Positive SARS-CoV-2 RT-PCR date 1 June 2020 29 June 2020
Heart transplant date 2 May 2020 5 December 2019
Death date 30 days 207 days
Days between positive SARS-CoV-2 diagnosis and death 47 days 21 days
Complications during ICU stay ARDS due to COVID–19; heart transplant rejection, disseminated cytomegalovirus; aggravated chronic kidney disease and pressure ulcer ARDS due to COVID–19, disseminated cytomegalovirus and pancytopenia due to hemophagocytosis
Previous comorbidities Dilated cardiomyopathy of Chagas etiology, hypothyroidism by thyroidectomy by nodule 10 years ago Cardiomyopathy of Chagas etiology, disseminated cytomegalovirus, deep vein thrombosis, systemic arterial hypertension, diabetes, dyslipidemia, and chronic renal failure
Previous use of medications Carvedilol, losartan, furosemide, levothyroxine, isosorbide, and hydralazine Unknown
Medicines used during ICU stay Methylprednisolone, azathioprine, anti-thymocyte globulin, cyclosporine, tacrolimus, meropenem, linezolid, micafungin, vancomycin, polymyxin B, tigecycline, amikacin, fluconazole, ganciclovir, and hydrocortisone Meropenem, colistin, linezolid
fluconazole, amikacin, cyclosporine, azathioprine, and prednisone
ARD, acute respiratory distress syndrome; COVID-19, coronavirus disease 2019; ICU, intensive care unit; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. | Fatal | ReactionOutcome | CC BY | 33579042 | 19,681,859 | 2021-02-10 |
What was the outcome of reaction 'Oxygen saturation decreased'? | SARS-CoV-2 Infection and CMV Dissemination in Transplant Recipients as a Treatment for Chagas Cardiomyopathy: A Case Report.
Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 has infected over 90 million people worldwide, therefore it is considered a pandemic. SARS-CoV-2 infection can lead to severe pneumonia, acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure. Individuals receiving a heart transplantation (HT) may be at higher risk of adverse outcomes attributable to COVID-19 due to immunosuppressives, as well as concomitant infections that may also influence the prognoses. Herein, we describe the first report of two cases of HT recipients with concomitant infections by SARS-CoV-2, Trypanosoma cruzi, and cytomegalovirus (CMV) dissemination, from the first day of hospitalization due to COVID-19 in the intensive care unit (ICU) until the death of the patients.
1. Introduction
Coronavirus disease 2019 (COVID-19), an infectious respiratory disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spread on a pandemic scale since the first case was reported in Wuhan, China, in 2019 [1]. Most patients with the disease have mild-to-moderate symptoms; however, approximately 15% develop severe pneumonia, while approximately 5% develop acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure [2]. Lymphopenia is a recurrent feature in these patients, with a significant reduction in CD4+ T cells, CD8+ T cells, B cells, and natural killer (NK) cells [3], increasing the susceptibility of patients to severe illness and co-infection [4]. In fact, co-infections are associated with worsening of the clinical condition [5].
Chagas disease (CD) is a zoonosis whose etiologic agent is the protozoan Trypanosoma cruzi. It is estimated that 6–8 million people worldwide are infected [6]. CD has two distinct phases: an acute one, which is rare, with a strong production of type 1 cytokines, and a chronic phase, which develops in 30–40% of CD cases. The chronic phase of CD can be characterized by cardiomyopathy, arrhythmias, megaviscera, and, more rarely, polyneuropathy and stroke [7]. TCD4+ and TCD8+ lymphocytes are the main cells responsible for controlling parasitic infection. However, the immune response also contributes to tissue damage and pathology [8].
Chagasic cardiomyopathy represents the main cause of mortality from this disease, which can lead to heart failure, whose indication for treatment, especially in endemic countries, may include heart transplantation (HT) as a strategy to curb the evolution of this complication [6,9]. However, the immunosuppression and possible reactivation of the causative agent T. cruzi following HT require intensive clinical care and laboratory monitoring [10].
Other infections such as that caused by cytomegalovirus (CMV), a herpes virus that infects up to 60–100% of people in adulthood, are associated with transplant complications [11]. Primary CMV infection is generally asymptomatic in immunocompetent individuals, with the virus generating a latent infection. On the contrary, in immunocompromised and immunosuppressed populations, such as solid organ transplantation, hematopoietic stem cell transplantation, and HIV/AIDS patients, CMV reactivation is responsible for significant morbidity and mortality [12]. Although reactivation of infections such as CMV and CD are often described in transplant recipients, there are no reports of concomitant infection by T. cruzi, CMV, and SARS-CoV-2 in the context of HT in the literature to date.
Therefore, in this report, we investigated the progress of two patients who underwent HT at the Heart Institute of Hospital das Clínicas (Incor) and were subsequently transferred to the special intensive care unit (ICU) of the Hospital das Clínicas (Hospital das Clínicas, Faculty of Medicine, University of São Paulo-HCFMUSP) due to SARS-CoV-2 infection. These patients were diagnosed with COVID-19 by nasopharyngeal detection of SARS-CoV-2 RNA using reverse transcriptase polymerase chain reaction (RT-PCR). In addition, during hospitalization, CMV dissemination was evidenced by quantitative DNA detection in the blood. We describe herein the laboratory data from the first day of hospitalization due to COVID-19 until the death of the patients.
The baseline characteristics of these two patients, as well as their past clinical data, are summarized in Table 1.
2. Case Report
2.1. Patient 1
Female, 55 years old, presenting positive serology for CD since 2015, received HT on 2 May as a treatment for Chagas cardiomyopathy (Figure 1A) and underwent immunosuppressive therapy with methylprednisolone and azathioprine. Post-transplantation, she developed pneumonia, treated with meropenem and linezolid, and a remittent Candida tropicalis infection, treated with micafungin, with improvement. However, on 29 May, she developed dyspnea and desaturation, with a chest tomography suggestive of COVID-19. Her polymerase chain reaction (PCR) for SARS-CoV-2 was positive on 1 June.
On 3 June, she was transferred to the ICU specialized for treatment of severe SARS-CoV-2 infections. Laboratory analyses then showed that the patient had a reduced number of erythrocytes and hemoglobin level, and these reductions became more accentuated by day 33 in the ICU (Figure 2A,B). On this same day, the number of neutrophils peaked (Figure 2D). From the 4th to the 16th day in ICU, important lymphopenia developed (Figure 2F), resulting in an increase in the neutrophil-to-lymphocyte ratio in the same period (Figure 2H). Additionally, from the eighth day onward, she presented severe thrombocytopenia that persisted until death (Figure 2I).
Throughout the ICU hospitalization period, there were sustained high levels of creatinine, urea, D-dimer, C-reactive protein (CRP), and lactate dehydrogenase (Figure 2J,K,N,S,W). On the 32nd day, she presented a sharp increase in prothrombin time and activated partial thromboplastin time.
The cardiac function markers of creatine kinase myocardial band (CK-MB) and troponin remained elevated from the first days of ICU admission until death (Figure 2U,X). On the day of admission to the ICU, the N-terminal pro b-type natriuretic peptide (NT pro-BNP) was 70,000 pg/mL (reference value of <125 pg/mL) (Figure 2V). On the third day of hospitalization, NT pro-BNP (155,117 pg/mL) and troponin (0.28 ng/mL, reference value of <0.014 ng/mL) peaked, being related to the period in which she presented signs of graft rejection, which was treated with pulse methylprednisolone, thymoglobulin, and plasmapheresis. In addition, disseminated CMV infection was diagnosed (RT-PCR viral loads of 122 IU/mL on day 10 and 54 IU/mL on day 19 of hospitalization), for which she received ganciclovir.
After 47 days after diagnosis of SARS-CoV-2, the patient died (18 July) due to multiple organ dysfunctions associated with COVID-19.
2.2. Patient 2
Male, 62 years old, previously diagnosed with CD, received HT on 5 December 2019 as treatment for Chagas cardiomyopathy (Figure 1B). He underwent immunosuppressive therapy with cyclosporine, azathioprine, and prednisone. During postoperative hospitalization he presented Chagas reactivation characterized by skin biopsy and humoral graft rejection, being treated with plasmapheresis and methylprednisolone.
The patient was admitted to the ICU of the Hospital das Clínicas on 8 June, presenting cellulitis, deep vein thrombosis, and Chagas reactivation (a lower limb chagoma). The latter was treated with benznidazol. During hospitalization, he tested positive for PCR of SARS-CoV-2 on 29 June. He developed ARDS and septic and cardiogenic shock, which were the causes of his death.
Throughout the ICU hospitalization period, he maintained decreased levels of erythrocytes, hemoglobin, and lymphocytes (Figure 2A,B,F), as well as a high neutrophil/lymphocyte ratio (Figure 2H). From the 12th day, the number of platelets decreased and continued until the time of death (Figure 2I). The values of creatinine, urea, and glucose also remained high throughout the hospitalization period (Figure 2J,K,T). In addition, disseminated CMV (viral loads of 711 IU/mL detected on the 28th day and 1477 IU/mL on the 35th day of hospitalization) was diagnosed and treated with ganciclovir.
One day before death, the following laboratory parameters peaked: D-dimer (7612 ng/mL FEU, reference value of <500 ng/mL FEU), CRP (280.6 ng/mL, reference value of 0.300 ng/mL), CK-MB (8.38 ng/mL, reference value of 0.10–4.94 ng/mL), NT pro-BNP (55,393 pg/mL), lactate dehydrogenase (1161 U/L, reference value of 135–225 U/L), and troponin (0.701 ng/mL) (Figure 2N,S,U–X).
On 20 July, 36 days after admission to the ICU and 21 days after a positive COVID-19 diagnosis, the patient died.
3. Discussion
Herein, we described the first report of triple infection (SARS-CoV-2 infection, T. cruzi infection, and CMV dissemination) in HT recipients. Patients received HT as a form of treatment for Chagas cardiomyopathy.
We described the laboratory data from the first day of hospitalization in the ICU due to COVID-19 until the time of death. Both patients were admitted to a referral center for treatment for COVID-19 in the metropolitan region of São Paulo, a city in southeastern Brazil. We hypothesize that the triple infection by SARS-CoV-2 and CMV may have been an important cause of death and of the worsening in CD patients with HT. To date, there have been no similar reports of patients presenting these three concomitant infectious diseases and HT receptors.
COVID-19 infection among transplant recipients increases the potential for developing severe illness [13] and may vary between different organ transplants [14,15].
SARS-CoV-2 entry´s receptor angiotensin converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2) are expressed in different tissues, such as the lungs, heart, liver, kidneys, testicles, thyroid, and adipose tissue [16]. Some patients with COVID-19 develop severe disease characterized by respiratory distress syndrome and systemic manifestations. This condition has been associated with the dysregulated release of pro-inflammatory cytokines, termed “cytokine storm,” which may induce multi-organ failure [1,17].
The use of immunosuppressants and post-surgical opportunistic infections can also lead to damage of multisystem organs or even death [18]. There is a therapeutic paradox here, because while insufficient immunosuppression results in graft loss due to rejection, excessive immunosuppression can result in serious infection, including SARS-COV-2 [13], besides contributing to the reactivation of pathogens [19].
Long-term administration of immunosuppressants to solid organ transplant (SOT) receptors to reduce the risk of graft rejection may increase the risk of respiratory infections [20], although there is no clear clinical evidence of increased morbidity/mortality in SARS-CoV-2 infection [21].
Fernandez-Ruiz and collaborators [22] described a cohort of SOT receptors affected by COVID-19, 44% of whom were kidney transplant recipients, 33.3% liver transplant recipients, and 22.2% heart transplant recipients. The lethality rate was 27.8%, suggesting that SARS-CoV-2 infection had a severe course in SOT recipients.
Recently, we described the first patients with CD affected by SARS-CoV-2. The CD patients presented an increase in COVID-19 laboratory hallmarks and a rapid disease progression. Despite the efforts of the health staff, both patients died [3].
HT may be associated with the reactivation of pathogens, as reported in CD [10]. In a Brazilian cohort, the reactivation rate of Chagas disease after heart transplantation was reported to be 38.8% [23]. This fact made HT as a treatment for Chagas cardiomyopathy initially controversial. However, currently, especially in endemic countries, it is the most viable therapeutic option for patients with end-stage heart failure [24]. We hypothesize that, in the cases presented here, HT followed by reactivation of CD conferred an additional risk factor for the worsening of COVID-19.
Transplant recipients are also commonly affected by CMV reactivation, being associated with significant morbidity and mortality [12] that may worsen the infectious condition of COVID-19 [25]. In SOT, risk factors include the use of immunosuppressants for transplantation, advanced age, acute rejection, and other concomitant infections [11], with all these characteristics being present in both patients described herein. In the present report, we observed that immunosuppression may have contributed to the susceptibility to superinfections and more severe clinical manifestations in individuals undergoing HT.
Since overactivated immune responses can be one of the causes of organ damage, the anti-inflammatory effects of immunosuppression can be protective, reducing the cytokine storm related to complications in COVID-19 [26]. In this context, it has been described that immunosuppressive therapy with calcineurin inhibitors in patients with solid organ transplantation or systemic rheumatic diseases promotes a clinical course in SARS-CoV-2 infection, which is generally mild, and with an apparently low risk of superinfection [27]. In addition, immunosuppression has not been evaluated as a risk factor for SARS or MERS [28].
However, in the present case report, we observed that immunosuppression may have contributed to the susceptibility to SARS-CoV-2 infection, the reactivation of pathogens, and more serious clinical manifestations in individuals undergoing HT. Corroborating our findings, it was shown that patients with COVID-19 and cancer, due to their systemic immunosuppressive condition caused by malignancy and anticancer treatments, such as chemotherapy, had an increased risk of SARS-CoV-2 infection and a worse prognosis [29]. It has also been reported that chronic use of corticosteroids prior to SARS-CoV-2 contamination is associated with critical disease outcomes, including a high risk of death [30].
These contradictory observations show that knowledge about the relationship between COVID-19 and the patient’s immune condition is limited. Further studies are needed to elucidate the immune responses and prognosis of COVID-19.
In general, HT in patient 1 proved to be successful for the treatment of Chagas cardiomyopathy, with no reactivation of the pathogen. However, after SARS-CoV-2 infection, she presented graft rejection, treated with methylprednisolone, thymoglobulin, and plasmapheresis. There is a description in eye transplantation that COVID-19 infection can compromise the balance of immunoregulatory responses that allow graft survival, contributing to rejection in individuals infected with SARS-CoV-2 [31]. Thus, this change in the balance of attenuation of the immune response, such as a reduction in the number of regulatory T cells (Tregs) [32], may favor direct cardiotoxic action and multiple organ dysfunction.
During postoperative hospitalization, patient 2 presented Chagas reactivation, evidenced by skin biopsy, indicating that the CD was not completely controlled. In addition, it is important to consider that immunosuppressive therapy can contribute to the reactivation of CD [33]. After HT, the patient presented graft rejection and was treated with plasmapheresis and methylprednisolone. With this, before the SARS-CoV-2 infection, the patient was already weakened and died 21 days after the infection, in relation to patient 1 who died after 47 days of infection.
4. Conclusions
This report highlights the first case of an association between COVID-19, CD, and CMV dissemination in HT recipients. The patients had rapid disease progression to death. We believe that HT and the usage of immunosuppressive drugs, as well as immunosuppression generated by concomitant infections, may be an important risk factor for the development of severe COVID-19, especially in endemic areas with underreported CD infection.
Author Contributions
Conceptualization, S.C.G.-S., G.B., and R.W.A.; investigation, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., and S.M.; resources, M.N.S. and A.J.d.S.D.; writing—original draft preparation, S.C.G.-S., G.B., and R.W.A.; writing—review and editing, S.C.G.-S., G.B., and R.W.A.; visualization, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., S.M., A.J.d.S.D., and M.N.S.; supervision, M.N.S.; funding acquisition, A.J.d.S.D. and M.N.S. All authors have read and agreed to the published version of the manuscript.
Funding
This research was funded by Fundação de Amparo à Pesquisa (FAPESP), grant numbers 2019/22448-0, 19/02679-7, 2017/18199-9, 2018/18230-6, and 2019/26928-6. Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, CAPES:88887.503842/2020-00.
Institutional Review Board Statement
This study was approved by the local ethics committee (HCFMUSP nº 30800520.7.0000.0068-2020) and was carried out in accordance with the 2013 revision of the Declaration of Helsinki.
Informed Consent Statement
Informed consent was obtained from all of the subjects involved in the study.
Data Availability Statement
The laboratory data and imaging exams presented herein came from Hospital das Clinics, Faculty of Medicine, University of São Paulo (HCFMUSP) with prior approval for their use.
Conflicts of Interest
The authors declare no conflict of interest.
Figure 1 Chest X-ray of (a) patient 1 from May 2020 and (b) patient 2 from June 2015, taken just before heart transplantation, showing the Chagasic cardiomyopathy.
Figure 2 Daily clinical features of patients, from the first day of hospitalization in the ICU until death. Blood levels of (A) erythrocytes, (B) hemoglobin, (C) leukocytes, (D) neutrophils, (E) eosinophils, (F) lymphocytes, (G) monocytes, (H) neutrophil-to-lymphocyte ratio, (I) platelets, (J) creatinine, (K) urea, (L) prothrombin time, (M) activated partial thromboplastin time, (N) D-dimer, (O) pH, (P) pO2, (Q) pCO2, (R) oxygen peripheral, (S) CPR, (T) glucose, (U) CK-MB, (V) NT-proBNP, (W) lactate dehydrogenase, and (X) troponin T. Gray boxes represent the reference values.
tropicalmed-06-00022-t001_Table 1 Table 1 Patients’ characteristics, comorbidities, treatment, and complications.
Patient 1 Patient 2
Sex Female Male
Age 55 62
Positive SARS-CoV-2 RT-PCR date 1 June 2020 29 June 2020
Heart transplant date 2 May 2020 5 December 2019
Death date 30 days 207 days
Days between positive SARS-CoV-2 diagnosis and death 47 days 21 days
Complications during ICU stay ARDS due to COVID–19; heart transplant rejection, disseminated cytomegalovirus; aggravated chronic kidney disease and pressure ulcer ARDS due to COVID–19, disseminated cytomegalovirus and pancytopenia due to hemophagocytosis
Previous comorbidities Dilated cardiomyopathy of Chagas etiology, hypothyroidism by thyroidectomy by nodule 10 years ago Cardiomyopathy of Chagas etiology, disseminated cytomegalovirus, deep vein thrombosis, systemic arterial hypertension, diabetes, dyslipidemia, and chronic renal failure
Previous use of medications Carvedilol, losartan, furosemide, levothyroxine, isosorbide, and hydralazine Unknown
Medicines used during ICU stay Methylprednisolone, azathioprine, anti-thymocyte globulin, cyclosporine, tacrolimus, meropenem, linezolid, micafungin, vancomycin, polymyxin B, tigecycline, amikacin, fluconazole, ganciclovir, and hydrocortisone Meropenem, colistin, linezolid
fluconazole, amikacin, cyclosporine, azathioprine, and prednisone
ARD, acute respiratory distress syndrome; COVID-19, coronavirus disease 2019; ICU, intensive care unit; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. | Fatal | ReactionOutcome | CC BY | 33579042 | 19,681,859 | 2021-02-10 |
What was the outcome of reaction 'Pancytopenia'? | SARS-CoV-2 Infection and CMV Dissemination in Transplant Recipients as a Treatment for Chagas Cardiomyopathy: A Case Report.
Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 has infected over 90 million people worldwide, therefore it is considered a pandemic. SARS-CoV-2 infection can lead to severe pneumonia, acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure. Individuals receiving a heart transplantation (HT) may be at higher risk of adverse outcomes attributable to COVID-19 due to immunosuppressives, as well as concomitant infections that may also influence the prognoses. Herein, we describe the first report of two cases of HT recipients with concomitant infections by SARS-CoV-2, Trypanosoma cruzi, and cytomegalovirus (CMV) dissemination, from the first day of hospitalization due to COVID-19 in the intensive care unit (ICU) until the death of the patients.
1. Introduction
Coronavirus disease 2019 (COVID-19), an infectious respiratory disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spread on a pandemic scale since the first case was reported in Wuhan, China, in 2019 [1]. Most patients with the disease have mild-to-moderate symptoms; however, approximately 15% develop severe pneumonia, while approximately 5% develop acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure [2]. Lymphopenia is a recurrent feature in these patients, with a significant reduction in CD4+ T cells, CD8+ T cells, B cells, and natural killer (NK) cells [3], increasing the susceptibility of patients to severe illness and co-infection [4]. In fact, co-infections are associated with worsening of the clinical condition [5].
Chagas disease (CD) is a zoonosis whose etiologic agent is the protozoan Trypanosoma cruzi. It is estimated that 6–8 million people worldwide are infected [6]. CD has two distinct phases: an acute one, which is rare, with a strong production of type 1 cytokines, and a chronic phase, which develops in 30–40% of CD cases. The chronic phase of CD can be characterized by cardiomyopathy, arrhythmias, megaviscera, and, more rarely, polyneuropathy and stroke [7]. TCD4+ and TCD8+ lymphocytes are the main cells responsible for controlling parasitic infection. However, the immune response also contributes to tissue damage and pathology [8].
Chagasic cardiomyopathy represents the main cause of mortality from this disease, which can lead to heart failure, whose indication for treatment, especially in endemic countries, may include heart transplantation (HT) as a strategy to curb the evolution of this complication [6,9]. However, the immunosuppression and possible reactivation of the causative agent T. cruzi following HT require intensive clinical care and laboratory monitoring [10].
Other infections such as that caused by cytomegalovirus (CMV), a herpes virus that infects up to 60–100% of people in adulthood, are associated with transplant complications [11]. Primary CMV infection is generally asymptomatic in immunocompetent individuals, with the virus generating a latent infection. On the contrary, in immunocompromised and immunosuppressed populations, such as solid organ transplantation, hematopoietic stem cell transplantation, and HIV/AIDS patients, CMV reactivation is responsible for significant morbidity and mortality [12]. Although reactivation of infections such as CMV and CD are often described in transplant recipients, there are no reports of concomitant infection by T. cruzi, CMV, and SARS-CoV-2 in the context of HT in the literature to date.
Therefore, in this report, we investigated the progress of two patients who underwent HT at the Heart Institute of Hospital das Clínicas (Incor) and were subsequently transferred to the special intensive care unit (ICU) of the Hospital das Clínicas (Hospital das Clínicas, Faculty of Medicine, University of São Paulo-HCFMUSP) due to SARS-CoV-2 infection. These patients were diagnosed with COVID-19 by nasopharyngeal detection of SARS-CoV-2 RNA using reverse transcriptase polymerase chain reaction (RT-PCR). In addition, during hospitalization, CMV dissemination was evidenced by quantitative DNA detection in the blood. We describe herein the laboratory data from the first day of hospitalization due to COVID-19 until the death of the patients.
The baseline characteristics of these two patients, as well as their past clinical data, are summarized in Table 1.
2. Case Report
2.1. Patient 1
Female, 55 years old, presenting positive serology for CD since 2015, received HT on 2 May as a treatment for Chagas cardiomyopathy (Figure 1A) and underwent immunosuppressive therapy with methylprednisolone and azathioprine. Post-transplantation, she developed pneumonia, treated with meropenem and linezolid, and a remittent Candida tropicalis infection, treated with micafungin, with improvement. However, on 29 May, she developed dyspnea and desaturation, with a chest tomography suggestive of COVID-19. Her polymerase chain reaction (PCR) for SARS-CoV-2 was positive on 1 June.
On 3 June, she was transferred to the ICU specialized for treatment of severe SARS-CoV-2 infections. Laboratory analyses then showed that the patient had a reduced number of erythrocytes and hemoglobin level, and these reductions became more accentuated by day 33 in the ICU (Figure 2A,B). On this same day, the number of neutrophils peaked (Figure 2D). From the 4th to the 16th day in ICU, important lymphopenia developed (Figure 2F), resulting in an increase in the neutrophil-to-lymphocyte ratio in the same period (Figure 2H). Additionally, from the eighth day onward, she presented severe thrombocytopenia that persisted until death (Figure 2I).
Throughout the ICU hospitalization period, there were sustained high levels of creatinine, urea, D-dimer, C-reactive protein (CRP), and lactate dehydrogenase (Figure 2J,K,N,S,W). On the 32nd day, she presented a sharp increase in prothrombin time and activated partial thromboplastin time.
The cardiac function markers of creatine kinase myocardial band (CK-MB) and troponin remained elevated from the first days of ICU admission until death (Figure 2U,X). On the day of admission to the ICU, the N-terminal pro b-type natriuretic peptide (NT pro-BNP) was 70,000 pg/mL (reference value of <125 pg/mL) (Figure 2V). On the third day of hospitalization, NT pro-BNP (155,117 pg/mL) and troponin (0.28 ng/mL, reference value of <0.014 ng/mL) peaked, being related to the period in which she presented signs of graft rejection, which was treated with pulse methylprednisolone, thymoglobulin, and plasmapheresis. In addition, disseminated CMV infection was diagnosed (RT-PCR viral loads of 122 IU/mL on day 10 and 54 IU/mL on day 19 of hospitalization), for which she received ganciclovir.
After 47 days after diagnosis of SARS-CoV-2, the patient died (18 July) due to multiple organ dysfunctions associated with COVID-19.
2.2. Patient 2
Male, 62 years old, previously diagnosed with CD, received HT on 5 December 2019 as treatment for Chagas cardiomyopathy (Figure 1B). He underwent immunosuppressive therapy with cyclosporine, azathioprine, and prednisone. During postoperative hospitalization he presented Chagas reactivation characterized by skin biopsy and humoral graft rejection, being treated with plasmapheresis and methylprednisolone.
The patient was admitted to the ICU of the Hospital das Clínicas on 8 June, presenting cellulitis, deep vein thrombosis, and Chagas reactivation (a lower limb chagoma). The latter was treated with benznidazol. During hospitalization, he tested positive for PCR of SARS-CoV-2 on 29 June. He developed ARDS and septic and cardiogenic shock, which were the causes of his death.
Throughout the ICU hospitalization period, he maintained decreased levels of erythrocytes, hemoglobin, and lymphocytes (Figure 2A,B,F), as well as a high neutrophil/lymphocyte ratio (Figure 2H). From the 12th day, the number of platelets decreased and continued until the time of death (Figure 2I). The values of creatinine, urea, and glucose also remained high throughout the hospitalization period (Figure 2J,K,T). In addition, disseminated CMV (viral loads of 711 IU/mL detected on the 28th day and 1477 IU/mL on the 35th day of hospitalization) was diagnosed and treated with ganciclovir.
One day before death, the following laboratory parameters peaked: D-dimer (7612 ng/mL FEU, reference value of <500 ng/mL FEU), CRP (280.6 ng/mL, reference value of 0.300 ng/mL), CK-MB (8.38 ng/mL, reference value of 0.10–4.94 ng/mL), NT pro-BNP (55,393 pg/mL), lactate dehydrogenase (1161 U/L, reference value of 135–225 U/L), and troponin (0.701 ng/mL) (Figure 2N,S,U–X).
On 20 July, 36 days after admission to the ICU and 21 days after a positive COVID-19 diagnosis, the patient died.
3. Discussion
Herein, we described the first report of triple infection (SARS-CoV-2 infection, T. cruzi infection, and CMV dissemination) in HT recipients. Patients received HT as a form of treatment for Chagas cardiomyopathy.
We described the laboratory data from the first day of hospitalization in the ICU due to COVID-19 until the time of death. Both patients were admitted to a referral center for treatment for COVID-19 in the metropolitan region of São Paulo, a city in southeastern Brazil. We hypothesize that the triple infection by SARS-CoV-2 and CMV may have been an important cause of death and of the worsening in CD patients with HT. To date, there have been no similar reports of patients presenting these three concomitant infectious diseases and HT receptors.
COVID-19 infection among transplant recipients increases the potential for developing severe illness [13] and may vary between different organ transplants [14,15].
SARS-CoV-2 entry´s receptor angiotensin converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2) are expressed in different tissues, such as the lungs, heart, liver, kidneys, testicles, thyroid, and adipose tissue [16]. Some patients with COVID-19 develop severe disease characterized by respiratory distress syndrome and systemic manifestations. This condition has been associated with the dysregulated release of pro-inflammatory cytokines, termed “cytokine storm,” which may induce multi-organ failure [1,17].
The use of immunosuppressants and post-surgical opportunistic infections can also lead to damage of multisystem organs or even death [18]. There is a therapeutic paradox here, because while insufficient immunosuppression results in graft loss due to rejection, excessive immunosuppression can result in serious infection, including SARS-COV-2 [13], besides contributing to the reactivation of pathogens [19].
Long-term administration of immunosuppressants to solid organ transplant (SOT) receptors to reduce the risk of graft rejection may increase the risk of respiratory infections [20], although there is no clear clinical evidence of increased morbidity/mortality in SARS-CoV-2 infection [21].
Fernandez-Ruiz and collaborators [22] described a cohort of SOT receptors affected by COVID-19, 44% of whom were kidney transplant recipients, 33.3% liver transplant recipients, and 22.2% heart transplant recipients. The lethality rate was 27.8%, suggesting that SARS-CoV-2 infection had a severe course in SOT recipients.
Recently, we described the first patients with CD affected by SARS-CoV-2. The CD patients presented an increase in COVID-19 laboratory hallmarks and a rapid disease progression. Despite the efforts of the health staff, both patients died [3].
HT may be associated with the reactivation of pathogens, as reported in CD [10]. In a Brazilian cohort, the reactivation rate of Chagas disease after heart transplantation was reported to be 38.8% [23]. This fact made HT as a treatment for Chagas cardiomyopathy initially controversial. However, currently, especially in endemic countries, it is the most viable therapeutic option for patients with end-stage heart failure [24]. We hypothesize that, in the cases presented here, HT followed by reactivation of CD conferred an additional risk factor for the worsening of COVID-19.
Transplant recipients are also commonly affected by CMV reactivation, being associated with significant morbidity and mortality [12] that may worsen the infectious condition of COVID-19 [25]. In SOT, risk factors include the use of immunosuppressants for transplantation, advanced age, acute rejection, and other concomitant infections [11], with all these characteristics being present in both patients described herein. In the present report, we observed that immunosuppression may have contributed to the susceptibility to superinfections and more severe clinical manifestations in individuals undergoing HT.
Since overactivated immune responses can be one of the causes of organ damage, the anti-inflammatory effects of immunosuppression can be protective, reducing the cytokine storm related to complications in COVID-19 [26]. In this context, it has been described that immunosuppressive therapy with calcineurin inhibitors in patients with solid organ transplantation or systemic rheumatic diseases promotes a clinical course in SARS-CoV-2 infection, which is generally mild, and with an apparently low risk of superinfection [27]. In addition, immunosuppression has not been evaluated as a risk factor for SARS or MERS [28].
However, in the present case report, we observed that immunosuppression may have contributed to the susceptibility to SARS-CoV-2 infection, the reactivation of pathogens, and more serious clinical manifestations in individuals undergoing HT. Corroborating our findings, it was shown that patients with COVID-19 and cancer, due to their systemic immunosuppressive condition caused by malignancy and anticancer treatments, such as chemotherapy, had an increased risk of SARS-CoV-2 infection and a worse prognosis [29]. It has also been reported that chronic use of corticosteroids prior to SARS-CoV-2 contamination is associated with critical disease outcomes, including a high risk of death [30].
These contradictory observations show that knowledge about the relationship between COVID-19 and the patient’s immune condition is limited. Further studies are needed to elucidate the immune responses and prognosis of COVID-19.
In general, HT in patient 1 proved to be successful for the treatment of Chagas cardiomyopathy, with no reactivation of the pathogen. However, after SARS-CoV-2 infection, she presented graft rejection, treated with methylprednisolone, thymoglobulin, and plasmapheresis. There is a description in eye transplantation that COVID-19 infection can compromise the balance of immunoregulatory responses that allow graft survival, contributing to rejection in individuals infected with SARS-CoV-2 [31]. Thus, this change in the balance of attenuation of the immune response, such as a reduction in the number of regulatory T cells (Tregs) [32], may favor direct cardiotoxic action and multiple organ dysfunction.
During postoperative hospitalization, patient 2 presented Chagas reactivation, evidenced by skin biopsy, indicating that the CD was not completely controlled. In addition, it is important to consider that immunosuppressive therapy can contribute to the reactivation of CD [33]. After HT, the patient presented graft rejection and was treated with plasmapheresis and methylprednisolone. With this, before the SARS-CoV-2 infection, the patient was already weakened and died 21 days after the infection, in relation to patient 1 who died after 47 days of infection.
4. Conclusions
This report highlights the first case of an association between COVID-19, CD, and CMV dissemination in HT recipients. The patients had rapid disease progression to death. We believe that HT and the usage of immunosuppressive drugs, as well as immunosuppression generated by concomitant infections, may be an important risk factor for the development of severe COVID-19, especially in endemic areas with underreported CD infection.
Author Contributions
Conceptualization, S.C.G.-S., G.B., and R.W.A.; investigation, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., and S.M.; resources, M.N.S. and A.J.d.S.D.; writing—original draft preparation, S.C.G.-S., G.B., and R.W.A.; writing—review and editing, S.C.G.-S., G.B., and R.W.A.; visualization, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., S.M., A.J.d.S.D., and M.N.S.; supervision, M.N.S.; funding acquisition, A.J.d.S.D. and M.N.S. All authors have read and agreed to the published version of the manuscript.
Funding
This research was funded by Fundação de Amparo à Pesquisa (FAPESP), grant numbers 2019/22448-0, 19/02679-7, 2017/18199-9, 2018/18230-6, and 2019/26928-6. Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, CAPES:88887.503842/2020-00.
Institutional Review Board Statement
This study was approved by the local ethics committee (HCFMUSP nº 30800520.7.0000.0068-2020) and was carried out in accordance with the 2013 revision of the Declaration of Helsinki.
Informed Consent Statement
Informed consent was obtained from all of the subjects involved in the study.
Data Availability Statement
The laboratory data and imaging exams presented herein came from Hospital das Clinics, Faculty of Medicine, University of São Paulo (HCFMUSP) with prior approval for their use.
Conflicts of Interest
The authors declare no conflict of interest.
Figure 1 Chest X-ray of (a) patient 1 from May 2020 and (b) patient 2 from June 2015, taken just before heart transplantation, showing the Chagasic cardiomyopathy.
Figure 2 Daily clinical features of patients, from the first day of hospitalization in the ICU until death. Blood levels of (A) erythrocytes, (B) hemoglobin, (C) leukocytes, (D) neutrophils, (E) eosinophils, (F) lymphocytes, (G) monocytes, (H) neutrophil-to-lymphocyte ratio, (I) platelets, (J) creatinine, (K) urea, (L) prothrombin time, (M) activated partial thromboplastin time, (N) D-dimer, (O) pH, (P) pO2, (Q) pCO2, (R) oxygen peripheral, (S) CPR, (T) glucose, (U) CK-MB, (V) NT-proBNP, (W) lactate dehydrogenase, and (X) troponin T. Gray boxes represent the reference values.
tropicalmed-06-00022-t001_Table 1 Table 1 Patients’ characteristics, comorbidities, treatment, and complications.
Patient 1 Patient 2
Sex Female Male
Age 55 62
Positive SARS-CoV-2 RT-PCR date 1 June 2020 29 June 2020
Heart transplant date 2 May 2020 5 December 2019
Death date 30 days 207 days
Days between positive SARS-CoV-2 diagnosis and death 47 days 21 days
Complications during ICU stay ARDS due to COVID–19; heart transplant rejection, disseminated cytomegalovirus; aggravated chronic kidney disease and pressure ulcer ARDS due to COVID–19, disseminated cytomegalovirus and pancytopenia due to hemophagocytosis
Previous comorbidities Dilated cardiomyopathy of Chagas etiology, hypothyroidism by thyroidectomy by nodule 10 years ago Cardiomyopathy of Chagas etiology, disseminated cytomegalovirus, deep vein thrombosis, systemic arterial hypertension, diabetes, dyslipidemia, and chronic renal failure
Previous use of medications Carvedilol, losartan, furosemide, levothyroxine, isosorbide, and hydralazine Unknown
Medicines used during ICU stay Methylprednisolone, azathioprine, anti-thymocyte globulin, cyclosporine, tacrolimus, meropenem, linezolid, micafungin, vancomycin, polymyxin B, tigecycline, amikacin, fluconazole, ganciclovir, and hydrocortisone Meropenem, colistin, linezolid
fluconazole, amikacin, cyclosporine, azathioprine, and prednisone
ARD, acute respiratory distress syndrome; COVID-19, coronavirus disease 2019; ICU, intensive care unit; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. | Fatal | ReactionOutcome | CC BY | 33579042 | 19,731,502 | 2021-02-10 |
What was the outcome of reaction 'Septic shock'? | SARS-CoV-2 Infection and CMV Dissemination in Transplant Recipients as a Treatment for Chagas Cardiomyopathy: A Case Report.
Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 has infected over 90 million people worldwide, therefore it is considered a pandemic. SARS-CoV-2 infection can lead to severe pneumonia, acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure. Individuals receiving a heart transplantation (HT) may be at higher risk of adverse outcomes attributable to COVID-19 due to immunosuppressives, as well as concomitant infections that may also influence the prognoses. Herein, we describe the first report of two cases of HT recipients with concomitant infections by SARS-CoV-2, Trypanosoma cruzi, and cytomegalovirus (CMV) dissemination, from the first day of hospitalization due to COVID-19 in the intensive care unit (ICU) until the death of the patients.
1. Introduction
Coronavirus disease 2019 (COVID-19), an infectious respiratory disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spread on a pandemic scale since the first case was reported in Wuhan, China, in 2019 [1]. Most patients with the disease have mild-to-moderate symptoms; however, approximately 15% develop severe pneumonia, while approximately 5% develop acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure [2]. Lymphopenia is a recurrent feature in these patients, with a significant reduction in CD4+ T cells, CD8+ T cells, B cells, and natural killer (NK) cells [3], increasing the susceptibility of patients to severe illness and co-infection [4]. In fact, co-infections are associated with worsening of the clinical condition [5].
Chagas disease (CD) is a zoonosis whose etiologic agent is the protozoan Trypanosoma cruzi. It is estimated that 6–8 million people worldwide are infected [6]. CD has two distinct phases: an acute one, which is rare, with a strong production of type 1 cytokines, and a chronic phase, which develops in 30–40% of CD cases. The chronic phase of CD can be characterized by cardiomyopathy, arrhythmias, megaviscera, and, more rarely, polyneuropathy and stroke [7]. TCD4+ and TCD8+ lymphocytes are the main cells responsible for controlling parasitic infection. However, the immune response also contributes to tissue damage and pathology [8].
Chagasic cardiomyopathy represents the main cause of mortality from this disease, which can lead to heart failure, whose indication for treatment, especially in endemic countries, may include heart transplantation (HT) as a strategy to curb the evolution of this complication [6,9]. However, the immunosuppression and possible reactivation of the causative agent T. cruzi following HT require intensive clinical care and laboratory monitoring [10].
Other infections such as that caused by cytomegalovirus (CMV), a herpes virus that infects up to 60–100% of people in adulthood, are associated with transplant complications [11]. Primary CMV infection is generally asymptomatic in immunocompetent individuals, with the virus generating a latent infection. On the contrary, in immunocompromised and immunosuppressed populations, such as solid organ transplantation, hematopoietic stem cell transplantation, and HIV/AIDS patients, CMV reactivation is responsible for significant morbidity and mortality [12]. Although reactivation of infections such as CMV and CD are often described in transplant recipients, there are no reports of concomitant infection by T. cruzi, CMV, and SARS-CoV-2 in the context of HT in the literature to date.
Therefore, in this report, we investigated the progress of two patients who underwent HT at the Heart Institute of Hospital das Clínicas (Incor) and were subsequently transferred to the special intensive care unit (ICU) of the Hospital das Clínicas (Hospital das Clínicas, Faculty of Medicine, University of São Paulo-HCFMUSP) due to SARS-CoV-2 infection. These patients were diagnosed with COVID-19 by nasopharyngeal detection of SARS-CoV-2 RNA using reverse transcriptase polymerase chain reaction (RT-PCR). In addition, during hospitalization, CMV dissemination was evidenced by quantitative DNA detection in the blood. We describe herein the laboratory data from the first day of hospitalization due to COVID-19 until the death of the patients.
The baseline characteristics of these two patients, as well as their past clinical data, are summarized in Table 1.
2. Case Report
2.1. Patient 1
Female, 55 years old, presenting positive serology for CD since 2015, received HT on 2 May as a treatment for Chagas cardiomyopathy (Figure 1A) and underwent immunosuppressive therapy with methylprednisolone and azathioprine. Post-transplantation, she developed pneumonia, treated with meropenem and linezolid, and a remittent Candida tropicalis infection, treated with micafungin, with improvement. However, on 29 May, she developed dyspnea and desaturation, with a chest tomography suggestive of COVID-19. Her polymerase chain reaction (PCR) for SARS-CoV-2 was positive on 1 June.
On 3 June, she was transferred to the ICU specialized for treatment of severe SARS-CoV-2 infections. Laboratory analyses then showed that the patient had a reduced number of erythrocytes and hemoglobin level, and these reductions became more accentuated by day 33 in the ICU (Figure 2A,B). On this same day, the number of neutrophils peaked (Figure 2D). From the 4th to the 16th day in ICU, important lymphopenia developed (Figure 2F), resulting in an increase in the neutrophil-to-lymphocyte ratio in the same period (Figure 2H). Additionally, from the eighth day onward, she presented severe thrombocytopenia that persisted until death (Figure 2I).
Throughout the ICU hospitalization period, there were sustained high levels of creatinine, urea, D-dimer, C-reactive protein (CRP), and lactate dehydrogenase (Figure 2J,K,N,S,W). On the 32nd day, she presented a sharp increase in prothrombin time and activated partial thromboplastin time.
The cardiac function markers of creatine kinase myocardial band (CK-MB) and troponin remained elevated from the first days of ICU admission until death (Figure 2U,X). On the day of admission to the ICU, the N-terminal pro b-type natriuretic peptide (NT pro-BNP) was 70,000 pg/mL (reference value of <125 pg/mL) (Figure 2V). On the third day of hospitalization, NT pro-BNP (155,117 pg/mL) and troponin (0.28 ng/mL, reference value of <0.014 ng/mL) peaked, being related to the period in which she presented signs of graft rejection, which was treated with pulse methylprednisolone, thymoglobulin, and plasmapheresis. In addition, disseminated CMV infection was diagnosed (RT-PCR viral loads of 122 IU/mL on day 10 and 54 IU/mL on day 19 of hospitalization), for which she received ganciclovir.
After 47 days after diagnosis of SARS-CoV-2, the patient died (18 July) due to multiple organ dysfunctions associated with COVID-19.
2.2. Patient 2
Male, 62 years old, previously diagnosed with CD, received HT on 5 December 2019 as treatment for Chagas cardiomyopathy (Figure 1B). He underwent immunosuppressive therapy with cyclosporine, azathioprine, and prednisone. During postoperative hospitalization he presented Chagas reactivation characterized by skin biopsy and humoral graft rejection, being treated with plasmapheresis and methylprednisolone.
The patient was admitted to the ICU of the Hospital das Clínicas on 8 June, presenting cellulitis, deep vein thrombosis, and Chagas reactivation (a lower limb chagoma). The latter was treated with benznidazol. During hospitalization, he tested positive for PCR of SARS-CoV-2 on 29 June. He developed ARDS and septic and cardiogenic shock, which were the causes of his death.
Throughout the ICU hospitalization period, he maintained decreased levels of erythrocytes, hemoglobin, and lymphocytes (Figure 2A,B,F), as well as a high neutrophil/lymphocyte ratio (Figure 2H). From the 12th day, the number of platelets decreased and continued until the time of death (Figure 2I). The values of creatinine, urea, and glucose also remained high throughout the hospitalization period (Figure 2J,K,T). In addition, disseminated CMV (viral loads of 711 IU/mL detected on the 28th day and 1477 IU/mL on the 35th day of hospitalization) was diagnosed and treated with ganciclovir.
One day before death, the following laboratory parameters peaked: D-dimer (7612 ng/mL FEU, reference value of <500 ng/mL FEU), CRP (280.6 ng/mL, reference value of 0.300 ng/mL), CK-MB (8.38 ng/mL, reference value of 0.10–4.94 ng/mL), NT pro-BNP (55,393 pg/mL), lactate dehydrogenase (1161 U/L, reference value of 135–225 U/L), and troponin (0.701 ng/mL) (Figure 2N,S,U–X).
On 20 July, 36 days after admission to the ICU and 21 days after a positive COVID-19 diagnosis, the patient died.
3. Discussion
Herein, we described the first report of triple infection (SARS-CoV-2 infection, T. cruzi infection, and CMV dissemination) in HT recipients. Patients received HT as a form of treatment for Chagas cardiomyopathy.
We described the laboratory data from the first day of hospitalization in the ICU due to COVID-19 until the time of death. Both patients were admitted to a referral center for treatment for COVID-19 in the metropolitan region of São Paulo, a city in southeastern Brazil. We hypothesize that the triple infection by SARS-CoV-2 and CMV may have been an important cause of death and of the worsening in CD patients with HT. To date, there have been no similar reports of patients presenting these three concomitant infectious diseases and HT receptors.
COVID-19 infection among transplant recipients increases the potential for developing severe illness [13] and may vary between different organ transplants [14,15].
SARS-CoV-2 entry´s receptor angiotensin converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2) are expressed in different tissues, such as the lungs, heart, liver, kidneys, testicles, thyroid, and adipose tissue [16]. Some patients with COVID-19 develop severe disease characterized by respiratory distress syndrome and systemic manifestations. This condition has been associated with the dysregulated release of pro-inflammatory cytokines, termed “cytokine storm,” which may induce multi-organ failure [1,17].
The use of immunosuppressants and post-surgical opportunistic infections can also lead to damage of multisystem organs or even death [18]. There is a therapeutic paradox here, because while insufficient immunosuppression results in graft loss due to rejection, excessive immunosuppression can result in serious infection, including SARS-COV-2 [13], besides contributing to the reactivation of pathogens [19].
Long-term administration of immunosuppressants to solid organ transplant (SOT) receptors to reduce the risk of graft rejection may increase the risk of respiratory infections [20], although there is no clear clinical evidence of increased morbidity/mortality in SARS-CoV-2 infection [21].
Fernandez-Ruiz and collaborators [22] described a cohort of SOT receptors affected by COVID-19, 44% of whom were kidney transplant recipients, 33.3% liver transplant recipients, and 22.2% heart transplant recipients. The lethality rate was 27.8%, suggesting that SARS-CoV-2 infection had a severe course in SOT recipients.
Recently, we described the first patients with CD affected by SARS-CoV-2. The CD patients presented an increase in COVID-19 laboratory hallmarks and a rapid disease progression. Despite the efforts of the health staff, both patients died [3].
HT may be associated with the reactivation of pathogens, as reported in CD [10]. In a Brazilian cohort, the reactivation rate of Chagas disease after heart transplantation was reported to be 38.8% [23]. This fact made HT as a treatment for Chagas cardiomyopathy initially controversial. However, currently, especially in endemic countries, it is the most viable therapeutic option for patients with end-stage heart failure [24]. We hypothesize that, in the cases presented here, HT followed by reactivation of CD conferred an additional risk factor for the worsening of COVID-19.
Transplant recipients are also commonly affected by CMV reactivation, being associated with significant morbidity and mortality [12] that may worsen the infectious condition of COVID-19 [25]. In SOT, risk factors include the use of immunosuppressants for transplantation, advanced age, acute rejection, and other concomitant infections [11], with all these characteristics being present in both patients described herein. In the present report, we observed that immunosuppression may have contributed to the susceptibility to superinfections and more severe clinical manifestations in individuals undergoing HT.
Since overactivated immune responses can be one of the causes of organ damage, the anti-inflammatory effects of immunosuppression can be protective, reducing the cytokine storm related to complications in COVID-19 [26]. In this context, it has been described that immunosuppressive therapy with calcineurin inhibitors in patients with solid organ transplantation or systemic rheumatic diseases promotes a clinical course in SARS-CoV-2 infection, which is generally mild, and with an apparently low risk of superinfection [27]. In addition, immunosuppression has not been evaluated as a risk factor for SARS or MERS [28].
However, in the present case report, we observed that immunosuppression may have contributed to the susceptibility to SARS-CoV-2 infection, the reactivation of pathogens, and more serious clinical manifestations in individuals undergoing HT. Corroborating our findings, it was shown that patients with COVID-19 and cancer, due to their systemic immunosuppressive condition caused by malignancy and anticancer treatments, such as chemotherapy, had an increased risk of SARS-CoV-2 infection and a worse prognosis [29]. It has also been reported that chronic use of corticosteroids prior to SARS-CoV-2 contamination is associated with critical disease outcomes, including a high risk of death [30].
These contradictory observations show that knowledge about the relationship between COVID-19 and the patient’s immune condition is limited. Further studies are needed to elucidate the immune responses and prognosis of COVID-19.
In general, HT in patient 1 proved to be successful for the treatment of Chagas cardiomyopathy, with no reactivation of the pathogen. However, after SARS-CoV-2 infection, she presented graft rejection, treated with methylprednisolone, thymoglobulin, and plasmapheresis. There is a description in eye transplantation that COVID-19 infection can compromise the balance of immunoregulatory responses that allow graft survival, contributing to rejection in individuals infected with SARS-CoV-2 [31]. Thus, this change in the balance of attenuation of the immune response, such as a reduction in the number of regulatory T cells (Tregs) [32], may favor direct cardiotoxic action and multiple organ dysfunction.
During postoperative hospitalization, patient 2 presented Chagas reactivation, evidenced by skin biopsy, indicating that the CD was not completely controlled. In addition, it is important to consider that immunosuppressive therapy can contribute to the reactivation of CD [33]. After HT, the patient presented graft rejection and was treated with plasmapheresis and methylprednisolone. With this, before the SARS-CoV-2 infection, the patient was already weakened and died 21 days after the infection, in relation to patient 1 who died after 47 days of infection.
4. Conclusions
This report highlights the first case of an association between COVID-19, CD, and CMV dissemination in HT recipients. The patients had rapid disease progression to death. We believe that HT and the usage of immunosuppressive drugs, as well as immunosuppression generated by concomitant infections, may be an important risk factor for the development of severe COVID-19, especially in endemic areas with underreported CD infection.
Author Contributions
Conceptualization, S.C.G.-S., G.B., and R.W.A.; investigation, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., and S.M.; resources, M.N.S. and A.J.d.S.D.; writing—original draft preparation, S.C.G.-S., G.B., and R.W.A.; writing—review and editing, S.C.G.-S., G.B., and R.W.A.; visualization, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., S.M., A.J.d.S.D., and M.N.S.; supervision, M.N.S.; funding acquisition, A.J.d.S.D. and M.N.S. All authors have read and agreed to the published version of the manuscript.
Funding
This research was funded by Fundação de Amparo à Pesquisa (FAPESP), grant numbers 2019/22448-0, 19/02679-7, 2017/18199-9, 2018/18230-6, and 2019/26928-6. Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, CAPES:88887.503842/2020-00.
Institutional Review Board Statement
This study was approved by the local ethics committee (HCFMUSP nº 30800520.7.0000.0068-2020) and was carried out in accordance with the 2013 revision of the Declaration of Helsinki.
Informed Consent Statement
Informed consent was obtained from all of the subjects involved in the study.
Data Availability Statement
The laboratory data and imaging exams presented herein came from Hospital das Clinics, Faculty of Medicine, University of São Paulo (HCFMUSP) with prior approval for their use.
Conflicts of Interest
The authors declare no conflict of interest.
Figure 1 Chest X-ray of (a) patient 1 from May 2020 and (b) patient 2 from June 2015, taken just before heart transplantation, showing the Chagasic cardiomyopathy.
Figure 2 Daily clinical features of patients, from the first day of hospitalization in the ICU until death. Blood levels of (A) erythrocytes, (B) hemoglobin, (C) leukocytes, (D) neutrophils, (E) eosinophils, (F) lymphocytes, (G) monocytes, (H) neutrophil-to-lymphocyte ratio, (I) platelets, (J) creatinine, (K) urea, (L) prothrombin time, (M) activated partial thromboplastin time, (N) D-dimer, (O) pH, (P) pO2, (Q) pCO2, (R) oxygen peripheral, (S) CPR, (T) glucose, (U) CK-MB, (V) NT-proBNP, (W) lactate dehydrogenase, and (X) troponin T. Gray boxes represent the reference values.
tropicalmed-06-00022-t001_Table 1 Table 1 Patients’ characteristics, comorbidities, treatment, and complications.
Patient 1 Patient 2
Sex Female Male
Age 55 62
Positive SARS-CoV-2 RT-PCR date 1 June 2020 29 June 2020
Heart transplant date 2 May 2020 5 December 2019
Death date 30 days 207 days
Days between positive SARS-CoV-2 diagnosis and death 47 days 21 days
Complications during ICU stay ARDS due to COVID–19; heart transplant rejection, disseminated cytomegalovirus; aggravated chronic kidney disease and pressure ulcer ARDS due to COVID–19, disseminated cytomegalovirus and pancytopenia due to hemophagocytosis
Previous comorbidities Dilated cardiomyopathy of Chagas etiology, hypothyroidism by thyroidectomy by nodule 10 years ago Cardiomyopathy of Chagas etiology, disseminated cytomegalovirus, deep vein thrombosis, systemic arterial hypertension, diabetes, dyslipidemia, and chronic renal failure
Previous use of medications Carvedilol, losartan, furosemide, levothyroxine, isosorbide, and hydralazine Unknown
Medicines used during ICU stay Methylprednisolone, azathioprine, anti-thymocyte globulin, cyclosporine, tacrolimus, meropenem, linezolid, micafungin, vancomycin, polymyxin B, tigecycline, amikacin, fluconazole, ganciclovir, and hydrocortisone Meropenem, colistin, linezolid
fluconazole, amikacin, cyclosporine, azathioprine, and prednisone
ARD, acute respiratory distress syndrome; COVID-19, coronavirus disease 2019; ICU, intensive care unit; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. | Fatal | ReactionOutcome | CC BY | 33579042 | 19,731,502 | 2021-02-10 |
What was the outcome of reaction 'Thrombocytopenia'? | SARS-CoV-2 Infection and CMV Dissemination in Transplant Recipients as a Treatment for Chagas Cardiomyopathy: A Case Report.
Coronavirus disease 2019 (COVID-19) is caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). COVID-19 has infected over 90 million people worldwide, therefore it is considered a pandemic. SARS-CoV-2 infection can lead to severe pneumonia, acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure. Individuals receiving a heart transplantation (HT) may be at higher risk of adverse outcomes attributable to COVID-19 due to immunosuppressives, as well as concomitant infections that may also influence the prognoses. Herein, we describe the first report of two cases of HT recipients with concomitant infections by SARS-CoV-2, Trypanosoma cruzi, and cytomegalovirus (CMV) dissemination, from the first day of hospitalization due to COVID-19 in the intensive care unit (ICU) until the death of the patients.
1. Introduction
Coronavirus disease 2019 (COVID-19), an infectious respiratory disease caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has spread on a pandemic scale since the first case was reported in Wuhan, China, in 2019 [1]. Most patients with the disease have mild-to-moderate symptoms; however, approximately 15% develop severe pneumonia, while approximately 5% develop acute respiratory distress syndrome (ARDS), septic shock, and/or organ failure [2]. Lymphopenia is a recurrent feature in these patients, with a significant reduction in CD4+ T cells, CD8+ T cells, B cells, and natural killer (NK) cells [3], increasing the susceptibility of patients to severe illness and co-infection [4]. In fact, co-infections are associated with worsening of the clinical condition [5].
Chagas disease (CD) is a zoonosis whose etiologic agent is the protozoan Trypanosoma cruzi. It is estimated that 6–8 million people worldwide are infected [6]. CD has two distinct phases: an acute one, which is rare, with a strong production of type 1 cytokines, and a chronic phase, which develops in 30–40% of CD cases. The chronic phase of CD can be characterized by cardiomyopathy, arrhythmias, megaviscera, and, more rarely, polyneuropathy and stroke [7]. TCD4+ and TCD8+ lymphocytes are the main cells responsible for controlling parasitic infection. However, the immune response also contributes to tissue damage and pathology [8].
Chagasic cardiomyopathy represents the main cause of mortality from this disease, which can lead to heart failure, whose indication for treatment, especially in endemic countries, may include heart transplantation (HT) as a strategy to curb the evolution of this complication [6,9]. However, the immunosuppression and possible reactivation of the causative agent T. cruzi following HT require intensive clinical care and laboratory monitoring [10].
Other infections such as that caused by cytomegalovirus (CMV), a herpes virus that infects up to 60–100% of people in adulthood, are associated with transplant complications [11]. Primary CMV infection is generally asymptomatic in immunocompetent individuals, with the virus generating a latent infection. On the contrary, in immunocompromised and immunosuppressed populations, such as solid organ transplantation, hematopoietic stem cell transplantation, and HIV/AIDS patients, CMV reactivation is responsible for significant morbidity and mortality [12]. Although reactivation of infections such as CMV and CD are often described in transplant recipients, there are no reports of concomitant infection by T. cruzi, CMV, and SARS-CoV-2 in the context of HT in the literature to date.
Therefore, in this report, we investigated the progress of two patients who underwent HT at the Heart Institute of Hospital das Clínicas (Incor) and were subsequently transferred to the special intensive care unit (ICU) of the Hospital das Clínicas (Hospital das Clínicas, Faculty of Medicine, University of São Paulo-HCFMUSP) due to SARS-CoV-2 infection. These patients were diagnosed with COVID-19 by nasopharyngeal detection of SARS-CoV-2 RNA using reverse transcriptase polymerase chain reaction (RT-PCR). In addition, during hospitalization, CMV dissemination was evidenced by quantitative DNA detection in the blood. We describe herein the laboratory data from the first day of hospitalization due to COVID-19 until the death of the patients.
The baseline characteristics of these two patients, as well as their past clinical data, are summarized in Table 1.
2. Case Report
2.1. Patient 1
Female, 55 years old, presenting positive serology for CD since 2015, received HT on 2 May as a treatment for Chagas cardiomyopathy (Figure 1A) and underwent immunosuppressive therapy with methylprednisolone and azathioprine. Post-transplantation, she developed pneumonia, treated with meropenem and linezolid, and a remittent Candida tropicalis infection, treated with micafungin, with improvement. However, on 29 May, she developed dyspnea and desaturation, with a chest tomography suggestive of COVID-19. Her polymerase chain reaction (PCR) for SARS-CoV-2 was positive on 1 June.
On 3 June, she was transferred to the ICU specialized for treatment of severe SARS-CoV-2 infections. Laboratory analyses then showed that the patient had a reduced number of erythrocytes and hemoglobin level, and these reductions became more accentuated by day 33 in the ICU (Figure 2A,B). On this same day, the number of neutrophils peaked (Figure 2D). From the 4th to the 16th day in ICU, important lymphopenia developed (Figure 2F), resulting in an increase in the neutrophil-to-lymphocyte ratio in the same period (Figure 2H). Additionally, from the eighth day onward, she presented severe thrombocytopenia that persisted until death (Figure 2I).
Throughout the ICU hospitalization period, there were sustained high levels of creatinine, urea, D-dimer, C-reactive protein (CRP), and lactate dehydrogenase (Figure 2J,K,N,S,W). On the 32nd day, she presented a sharp increase in prothrombin time and activated partial thromboplastin time.
The cardiac function markers of creatine kinase myocardial band (CK-MB) and troponin remained elevated from the first days of ICU admission until death (Figure 2U,X). On the day of admission to the ICU, the N-terminal pro b-type natriuretic peptide (NT pro-BNP) was 70,000 pg/mL (reference value of <125 pg/mL) (Figure 2V). On the third day of hospitalization, NT pro-BNP (155,117 pg/mL) and troponin (0.28 ng/mL, reference value of <0.014 ng/mL) peaked, being related to the period in which she presented signs of graft rejection, which was treated with pulse methylprednisolone, thymoglobulin, and plasmapheresis. In addition, disseminated CMV infection was diagnosed (RT-PCR viral loads of 122 IU/mL on day 10 and 54 IU/mL on day 19 of hospitalization), for which she received ganciclovir.
After 47 days after diagnosis of SARS-CoV-2, the patient died (18 July) due to multiple organ dysfunctions associated with COVID-19.
2.2. Patient 2
Male, 62 years old, previously diagnosed with CD, received HT on 5 December 2019 as treatment for Chagas cardiomyopathy (Figure 1B). He underwent immunosuppressive therapy with cyclosporine, azathioprine, and prednisone. During postoperative hospitalization he presented Chagas reactivation characterized by skin biopsy and humoral graft rejection, being treated with plasmapheresis and methylprednisolone.
The patient was admitted to the ICU of the Hospital das Clínicas on 8 June, presenting cellulitis, deep vein thrombosis, and Chagas reactivation (a lower limb chagoma). The latter was treated with benznidazol. During hospitalization, he tested positive for PCR of SARS-CoV-2 on 29 June. He developed ARDS and septic and cardiogenic shock, which were the causes of his death.
Throughout the ICU hospitalization period, he maintained decreased levels of erythrocytes, hemoglobin, and lymphocytes (Figure 2A,B,F), as well as a high neutrophil/lymphocyte ratio (Figure 2H). From the 12th day, the number of platelets decreased and continued until the time of death (Figure 2I). The values of creatinine, urea, and glucose also remained high throughout the hospitalization period (Figure 2J,K,T). In addition, disseminated CMV (viral loads of 711 IU/mL detected on the 28th day and 1477 IU/mL on the 35th day of hospitalization) was diagnosed and treated with ganciclovir.
One day before death, the following laboratory parameters peaked: D-dimer (7612 ng/mL FEU, reference value of <500 ng/mL FEU), CRP (280.6 ng/mL, reference value of 0.300 ng/mL), CK-MB (8.38 ng/mL, reference value of 0.10–4.94 ng/mL), NT pro-BNP (55,393 pg/mL), lactate dehydrogenase (1161 U/L, reference value of 135–225 U/L), and troponin (0.701 ng/mL) (Figure 2N,S,U–X).
On 20 July, 36 days after admission to the ICU and 21 days after a positive COVID-19 diagnosis, the patient died.
3. Discussion
Herein, we described the first report of triple infection (SARS-CoV-2 infection, T. cruzi infection, and CMV dissemination) in HT recipients. Patients received HT as a form of treatment for Chagas cardiomyopathy.
We described the laboratory data from the first day of hospitalization in the ICU due to COVID-19 until the time of death. Both patients were admitted to a referral center for treatment for COVID-19 in the metropolitan region of São Paulo, a city in southeastern Brazil. We hypothesize that the triple infection by SARS-CoV-2 and CMV may have been an important cause of death and of the worsening in CD patients with HT. To date, there have been no similar reports of patients presenting these three concomitant infectious diseases and HT receptors.
COVID-19 infection among transplant recipients increases the potential for developing severe illness [13] and may vary between different organ transplants [14,15].
SARS-CoV-2 entry´s receptor angiotensin converting enzyme 2 (ACE2) and transmembrane protease serine 2 (TMPRSS2) are expressed in different tissues, such as the lungs, heart, liver, kidneys, testicles, thyroid, and adipose tissue [16]. Some patients with COVID-19 develop severe disease characterized by respiratory distress syndrome and systemic manifestations. This condition has been associated with the dysregulated release of pro-inflammatory cytokines, termed “cytokine storm,” which may induce multi-organ failure [1,17].
The use of immunosuppressants and post-surgical opportunistic infections can also lead to damage of multisystem organs or even death [18]. There is a therapeutic paradox here, because while insufficient immunosuppression results in graft loss due to rejection, excessive immunosuppression can result in serious infection, including SARS-COV-2 [13], besides contributing to the reactivation of pathogens [19].
Long-term administration of immunosuppressants to solid organ transplant (SOT) receptors to reduce the risk of graft rejection may increase the risk of respiratory infections [20], although there is no clear clinical evidence of increased morbidity/mortality in SARS-CoV-2 infection [21].
Fernandez-Ruiz and collaborators [22] described a cohort of SOT receptors affected by COVID-19, 44% of whom were kidney transplant recipients, 33.3% liver transplant recipients, and 22.2% heart transplant recipients. The lethality rate was 27.8%, suggesting that SARS-CoV-2 infection had a severe course in SOT recipients.
Recently, we described the first patients with CD affected by SARS-CoV-2. The CD patients presented an increase in COVID-19 laboratory hallmarks and a rapid disease progression. Despite the efforts of the health staff, both patients died [3].
HT may be associated with the reactivation of pathogens, as reported in CD [10]. In a Brazilian cohort, the reactivation rate of Chagas disease after heart transplantation was reported to be 38.8% [23]. This fact made HT as a treatment for Chagas cardiomyopathy initially controversial. However, currently, especially in endemic countries, it is the most viable therapeutic option for patients with end-stage heart failure [24]. We hypothesize that, in the cases presented here, HT followed by reactivation of CD conferred an additional risk factor for the worsening of COVID-19.
Transplant recipients are also commonly affected by CMV reactivation, being associated with significant morbidity and mortality [12] that may worsen the infectious condition of COVID-19 [25]. In SOT, risk factors include the use of immunosuppressants for transplantation, advanced age, acute rejection, and other concomitant infections [11], with all these characteristics being present in both patients described herein. In the present report, we observed that immunosuppression may have contributed to the susceptibility to superinfections and more severe clinical manifestations in individuals undergoing HT.
Since overactivated immune responses can be one of the causes of organ damage, the anti-inflammatory effects of immunosuppression can be protective, reducing the cytokine storm related to complications in COVID-19 [26]. In this context, it has been described that immunosuppressive therapy with calcineurin inhibitors in patients with solid organ transplantation or systemic rheumatic diseases promotes a clinical course in SARS-CoV-2 infection, which is generally mild, and with an apparently low risk of superinfection [27]. In addition, immunosuppression has not been evaluated as a risk factor for SARS or MERS [28].
However, in the present case report, we observed that immunosuppression may have contributed to the susceptibility to SARS-CoV-2 infection, the reactivation of pathogens, and more serious clinical manifestations in individuals undergoing HT. Corroborating our findings, it was shown that patients with COVID-19 and cancer, due to their systemic immunosuppressive condition caused by malignancy and anticancer treatments, such as chemotherapy, had an increased risk of SARS-CoV-2 infection and a worse prognosis [29]. It has also been reported that chronic use of corticosteroids prior to SARS-CoV-2 contamination is associated with critical disease outcomes, including a high risk of death [30].
These contradictory observations show that knowledge about the relationship between COVID-19 and the patient’s immune condition is limited. Further studies are needed to elucidate the immune responses and prognosis of COVID-19.
In general, HT in patient 1 proved to be successful for the treatment of Chagas cardiomyopathy, with no reactivation of the pathogen. However, after SARS-CoV-2 infection, she presented graft rejection, treated with methylprednisolone, thymoglobulin, and plasmapheresis. There is a description in eye transplantation that COVID-19 infection can compromise the balance of immunoregulatory responses that allow graft survival, contributing to rejection in individuals infected with SARS-CoV-2 [31]. Thus, this change in the balance of attenuation of the immune response, such as a reduction in the number of regulatory T cells (Tregs) [32], may favor direct cardiotoxic action and multiple organ dysfunction.
During postoperative hospitalization, patient 2 presented Chagas reactivation, evidenced by skin biopsy, indicating that the CD was not completely controlled. In addition, it is important to consider that immunosuppressive therapy can contribute to the reactivation of CD [33]. After HT, the patient presented graft rejection and was treated with plasmapheresis and methylprednisolone. With this, before the SARS-CoV-2 infection, the patient was already weakened and died 21 days after the infection, in relation to patient 1 who died after 47 days of infection.
4. Conclusions
This report highlights the first case of an association between COVID-19, CD, and CMV dissemination in HT recipients. The patients had rapid disease progression to death. We believe that HT and the usage of immunosuppressive drugs, as well as immunosuppression generated by concomitant infections, may be an important risk factor for the development of severe COVID-19, especially in endemic areas with underreported CD infection.
Author Contributions
Conceptualization, S.C.G.-S., G.B., and R.W.A.; investigation, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., and S.M.; resources, M.N.S. and A.J.d.S.D.; writing—original draft preparation, S.C.G.-S., G.B., and R.W.A.; writing—review and editing, S.C.G.-S., G.B., and R.W.A.; visualization, S.C.G.-S., G.B., R.W.A., T.M.Y., F.M.E.T., L.d.M.O., D.R.B., A.J.P., E.A.d.O., A.C.C.C.B., M.M.d.S.A., I.G.F., N.Z.P., Y.Á.L.R., J.C.L., B.P., S.M., A.J.d.S.D., and M.N.S.; supervision, M.N.S.; funding acquisition, A.J.d.S.D. and M.N.S. All authors have read and agreed to the published version of the manuscript.
Funding
This research was funded by Fundação de Amparo à Pesquisa (FAPESP), grant numbers 2019/22448-0, 19/02679-7, 2017/18199-9, 2018/18230-6, and 2019/26928-6. Coordenação de Aperfeiçoamento de Pessoal de Nível Superior, CAPES:88887.503842/2020-00.
Institutional Review Board Statement
This study was approved by the local ethics committee (HCFMUSP nº 30800520.7.0000.0068-2020) and was carried out in accordance with the 2013 revision of the Declaration of Helsinki.
Informed Consent Statement
Informed consent was obtained from all of the subjects involved in the study.
Data Availability Statement
The laboratory data and imaging exams presented herein came from Hospital das Clinics, Faculty of Medicine, University of São Paulo (HCFMUSP) with prior approval for their use.
Conflicts of Interest
The authors declare no conflict of interest.
Figure 1 Chest X-ray of (a) patient 1 from May 2020 and (b) patient 2 from June 2015, taken just before heart transplantation, showing the Chagasic cardiomyopathy.
Figure 2 Daily clinical features of patients, from the first day of hospitalization in the ICU until death. Blood levels of (A) erythrocytes, (B) hemoglobin, (C) leukocytes, (D) neutrophils, (E) eosinophils, (F) lymphocytes, (G) monocytes, (H) neutrophil-to-lymphocyte ratio, (I) platelets, (J) creatinine, (K) urea, (L) prothrombin time, (M) activated partial thromboplastin time, (N) D-dimer, (O) pH, (P) pO2, (Q) pCO2, (R) oxygen peripheral, (S) CPR, (T) glucose, (U) CK-MB, (V) NT-proBNP, (W) lactate dehydrogenase, and (X) troponin T. Gray boxes represent the reference values.
tropicalmed-06-00022-t001_Table 1 Table 1 Patients’ characteristics, comorbidities, treatment, and complications.
Patient 1 Patient 2
Sex Female Male
Age 55 62
Positive SARS-CoV-2 RT-PCR date 1 June 2020 29 June 2020
Heart transplant date 2 May 2020 5 December 2019
Death date 30 days 207 days
Days between positive SARS-CoV-2 diagnosis and death 47 days 21 days
Complications during ICU stay ARDS due to COVID–19; heart transplant rejection, disseminated cytomegalovirus; aggravated chronic kidney disease and pressure ulcer ARDS due to COVID–19, disseminated cytomegalovirus and pancytopenia due to hemophagocytosis
Previous comorbidities Dilated cardiomyopathy of Chagas etiology, hypothyroidism by thyroidectomy by nodule 10 years ago Cardiomyopathy of Chagas etiology, disseminated cytomegalovirus, deep vein thrombosis, systemic arterial hypertension, diabetes, dyslipidemia, and chronic renal failure
Previous use of medications Carvedilol, losartan, furosemide, levothyroxine, isosorbide, and hydralazine Unknown
Medicines used during ICU stay Methylprednisolone, azathioprine, anti-thymocyte globulin, cyclosporine, tacrolimus, meropenem, linezolid, micafungin, vancomycin, polymyxin B, tigecycline, amikacin, fluconazole, ganciclovir, and hydrocortisone Meropenem, colistin, linezolid
fluconazole, amikacin, cyclosporine, azathioprine, and prednisone
ARD, acute respiratory distress syndrome; COVID-19, coronavirus disease 2019; ICU, intensive care unit; SARS-CoV-2, severe acute respiratory syndrome coronavirus 2.
Publisher’s Note: MDPI stays neutral with regard to jurisdictional claims in published maps and institutional affiliations. | Fatal | ReactionOutcome | CC BY | 33579042 | 19,681,859 | 2021-02-10 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Drug resistance'. | Hypermutated phenotype in gliosarcoma of the spinal cord.
Gliosarcoma is a variant of glioblastoma with equally poor prognosis and characterized by mixed glial and mesenchymal pathology. Metastasis is not uncommon but the involvement of the spinal cord is rare, and comprehensive genetic characterization of spinal gliosarcoma is lacking. We describe a patient initially diagnosed with a low-grade brain glioma via biopsy, followed by adjuvant radiation and temozolomide treatment. Nearly 2 years after diagnosis, she developed neurological deficits from an intradural, extramedullary tumor anterior to the spinal cord at T4, which was resected and diagnosed as gliosarcoma. Whole-exome sequencing (WES) of this tumor revealed a hypermutated phenotype, characterized by somatic mutations in key DNA mismatch repair (MMR) pathway genes, an abundance of C>T transitions within the identified somatic single nucleotide variations, and microsatellite stability, together consistent with temozolomide-mediated hypermutagenesis. This is the first report of a hypermutator phenotype in gliosarcoma, which may represent a novel genomic mechanism of progression from lower grade glioma.
Introduction
Gliosarcomas represent a variant of glioblastoma, characterized by mixed histopathological features of glial and mesenchymal origins1. Similar to glioblastoma, patients diagnosed with gliosarcoma exhibit poor prognosis and overall survival2. Metastasis is relatively common within the central nervous system (CNS), as well as systemic visceral sites1. Gliosarcomas involving the spinal cord are rare, making up approximately 1% of all malignant gliomas of the spine3 and may represent metastasis from intracranial tumors or less commonly, de novo formation4,5. To date, comprehensive genomic characterization of gliosarcomas remains vastly under-reported, compared to gliobastomas.
In this study, we report a patient with a spinal cord gliosarcoma with prior history of a low-grade brain glioma treated with adjuvant chemoradiation. Whole-exome sequencing (WES) of the resected spinal tumor revealed the first case of a hypermutator phenotype to be described in gliosarcoma.
Results
Case description
A 65-year-old female presented to an outside institution with several weeks of headaches and acute worsening of progressive numbness in her right lower extremity. Her past medical history was only significant for basal cell carcinoma of the left nasal ala, resected 5 years prior. Her family history was notable for breast cancer of middle-aged onset in her sister, mother, and grandmother but no prior diagnosis of a familial cancer predisposition syndrome. Her symptoms prompted magnetic resonance imaging (MRI) of the brain, which revealed a predominantly non-enhancing right parietal lesion with patchy enhancing foci extending to the splenium of the corpus callosum (Fig. 1a, b). Further imaging with computed tomography (CT) of the chest, abdomen, and pelvis did not reveal a primary systemic tumor to suggest a diagnosis of brain metastasis. She subsequently underwent stereotactic biopsy of her lesion with pathology demonstrating a glial neoplasm, consistent with World Health Organization (WHO) grade II astrocytoma (Fig. 1e–h). The Ki-67 index was 1–3% and immunohistochemical staining for TP53 was positive. Further molecular testing was negative for the EGFR variant III, IDH1 mutation, and MGMT promoter methylation. She was subsequently referred to our institution for further adjuvant treatment, where she underwent adjuvant radiation therapy and temozolomide treatment, as well as use of tumor-treating fields (TTF) with the Optune device (Novocure, Jersey, U.K.). A brain MRI obtained several months after completion of chemoradiation and TTF showed radiographic response with stable small foci of enhancement and drastic reduction in perilesional edema (Fig. 1c, d). Clinically, the patient was stable with moderate cognitive deficits and requirement of mild assistance with activities of daily living.Fig. 1 Imaging and histopathology of the low-grade glioma.
a Representative axial slices of T1-weighted MRI after contrast administration and (b) T2-weighted FLAIR obtained prior to stereotactic brain biopsy show a predominantly non-enhancing lesion within the deep right parietal lobe with small areas of patchy enhancement that is crossing the corpus callosum with associated edema. Repeat imaging obtained nearly 2 years after biopsy and completion of adjuvant chemoradation demonstrates (c) stable foci of small nodular enhancement on T1-weighted MRI after contrast administration with (d) minimal associated edema on T2-weighted FLAIR. e, f An infiltrating astrocytoma without necrosis or vascular proliferation is seen on hematoxylin & eosin staining. g Immunohistochemistry for GFAP-positive tumor cells is shown and (h) the Ki-67 proliferative index was ~3%.
Six months after completion of chemotherapy, she presented with increasing gait instability of two weeks duration and worsening thoracic back pain, localized between her shoulder blades. MRI of her thoracic and lumbar spine was obtained, demonstrating a homogenously enhancing dural-based, extramedullary lesion anterior to the spinal cord at T4 with moderate cord compression, causing edema extending from T2 to T7 (Fig. 2a–d). She was neurologically intact on examination other than brisk lower extremity reflexes, allodynia at the T4 level, and baseline right-sided hemibody numbness. Imaging findings were consistent with spinal meningioma, and the patient was admitted with plans for elective, inpatient surgery. However, two days after admission, she developed acute weakness in her bilateral lower extremities and subsequently was taken to the operating room for urgent resection of her tumor, which was removed in its entirety. Intra-operatively, the tumor was noted to be dural-based with a well-defined plane between tumor and the dura as well as the pia of the spinal cord. Motor evoked and somatosensory evoked potentials were improved at the end of the case, compared to baseline levels. Final pathology from the resected specimen was WHO IV gliosarcoma with biphasic tissue architecture, comprised of prominent GFAP-positive glial regions admixed with GFAP-negative spindled areas containing reticulin fibers (Fig. 2e–h). The Ki-67 index was elevated >10–15% and further routine molecular testing demonstrated MGMT promoter methylation but was otherwise negative for IDH1 mutation and ATRX loss.Fig. 2 Imaging and histopathology of the gliosarcoma.
a Representative sagittal and (b) axial slices from T1-weighted MRI after contrast administration show a homogeneously enhancing intradural, extramedullary lesion anterior to the spinal cord at the T4 level with associated cord compression seen on (c) sagittal and (d) axial slices on T2-weighted MRI. e Biphasic architecture is seen on hematoxylin & eosin staining, including (f) reticulin rich sarcomatous regions, interspersed with (g) GFAP-positive glial regions, and (h) an elevated Ki-67 proliferative index of ~10–15%.
The patient was subsequently discharged to rehabilitation and exhibited some improvement in lower extremity strength, albeit remained non-ambulatory and required significant assistance with activities of daily living. Further treatment with adjuvant chemoradiation was considered but given the unfavorable prognosis from new gliosarcoma diagnosis and poor performance status, she elected to undergo hospice care without further imaging or treatment and subsequently expired one month after surgery.
Genomic analysis
WES was performed on the gliosarcoma specimen, together with matching normal blood, to identify somatic single nucleotide variations (SNVs), insertions/deletions (INDEL), and copy number variations (CNV). The protocol for sequencing and analysis was in accordance with our previously described methods6. We detected 2372 somatic SNVs with a 96% C>T transition ratio (Fig. 3), consistent with a hypermutated phenotype. There were disease-associated alterations in key oncogenes, PTEN, PIK3R1, and ATM. Furthermore, there were key mutations in genes associated with the DNA mismatch repair (MMR) pathway, predicted to be deleterious based on scores from the following computational prediction tools: SIFT7, Polyphen2 HDIV Prediction8, Polyphen2 HVAR Prediction8, FATHMM9, MetaSVM10, and MutationTaster11. These included deleterious missense (p.P656L) and stop-gain (p.W105X) mutations in MSH6 and a stop-gain mutation in MSH3 (p.R727X). Additionally, there was a heterozygous germline variant in MSH2 (p.H46Q) of unknown significance. Somatic CNV analysis revealed amplifications of chromosomes 7, 20, and X, deletions on chromosomes 10, 13, 17, and focal CNV events on chromosomes 1 and 5. 27% of the genome was altered by CNV and/or loss-of-heterozygosity events.Fig. 3 Mutational signature plot of SNV in the resected spinal tumor specimen.
There was an overwhelming prevalence of C>T, with a SNV transition ratio of 96%, consistent with a hypermutated phenotype.
Given the relationship between microsatellite instability (MSI) and MMR deficiency in systemic cancers12, we performed further computational analysis with MSIsensor, an algorithm that predicts the microsatellite status of tumors from WES data13. The calculated MSI-score was 0.15, indicative of microsatellite stability per the tool guidelines, which uses a cut-off score of 3.5 for MSI.
Discussion
Gliosarcomas have a high predilection for metastasis to both visceral sites as well as the CNS, via hematogenous and leptomeningeal spread14. As such, most cases of gliosarcomas involving the spinal cord have been secondary to metastasis from intracranial origin4,15–19. In contrast, primary gliosarcomas of the spinal cord are less common5,20–22 and are vastly outnumbered by glioblastoma when analyzing primary malignant gliomas of the spinal cord3. Interestingly, these tumors may present as intramedullary15,23, as well as intradural, extramedullary lesions14,22, regardless of primary or metastatic etiology. Similar to their intracranial counterparts, spinal cord gliosarcomas demonstrate avid and typically homogenous enhancement, which in cases of intradural, extramedullary location, may be mistaken for a meningioma, as demonstrated in our patient. A prior history of a primary brain tumor must raise suspicion for an alternative diagnosis and highlights the need for surgical intervention to aid in definitive tissue diagnosis.
It remains unclear whether the gliosarcoma in our patient represented metastasis from her intracranial astrocytoma or de novo formation of a new primary tumor. This may have been addressed with additional WES data or targeted molecular testing from her original brain tumor, but unfortunately, inadequate tissue was available for further genomic studies. Additionally, further molecular testing of her brain tumor would have been helpful, as IDH-wildtype astrocytomas with grade II histology may exhibit survival outcomes akin to IDH-wildtype glioblastoma24,25. Based on recommendations from the Consortium to Inform Molecular and Practical Approaches to CNS Tumor Taxonomy (cIMPACT-NOW)26, molecular testing to assess for EGFR amplification, chromosome 7 gain/chromosome 10 loss, and TERT promoter mutations to diagnose a diffuse astrocytic glioma, IDH-wildtype, with molecular features of glioblastoma, WHO grade IV could have been performed to predict increased biological aggression and perhaps tailor clinical management27,28. The existing literature of metastatic gliosarcomas to the spinal cord have all been from primary intracranial gliosarcoma, so the notion that our patient’s gliosarcoma represented metastasis, despite the difference in WHO grade from her initial tumor, would be unique. There have been few reports of progression of low-grade gliomas into intracranial gliosarcomas, in instances with and without adjuvant chemoradiation29–31. These studies confirmed metastatic origin through single nucleotide polymorphism (SNP) arrays and fluorescence in-situ hybridization (FISH) for chromosomal analysis, but to date, comprehensive sequencing to delineate the genetic changes that may underlie either malignant progression into gliosarcomatous pathology or de novo spinal cord gliosarcoma formation have not been described.
In recent years, the prevalence of hypermutagenesis has been described across a variety of cancers, including gliomas32–34 and particularly after therapy-driven changes in tumor evolution at the time of recurrence. In particular, treatment with temozolomide may induce mutations in the MMR pathway, leading to therapeutic resistance and acquisition of the hypermutated phenotype35,36. Strikingly, we found a hypermutated phenotype in our patient’s specimen, including mutations in MMR genes, MSH3 and MSH6, which previously have not been associated with gliosarcoma pathology. Likewise, the abundance of C>T transitions detected within the somatic SNVs of our patient’s tumor was consistent with a MMR deficient hypermutated phenotype. We further explored the possibility of MSI, particularly given the strong correlation between MSI and MMR deficiency in systemic cancers that may serve as biomarkers for response to immunotherapies37–40. Utilizing the MSIsensor computational tool, we did not detect MSI despite the hypermutator phenotype of our patient’s tumor, a result consistent with prior studies failing to reveal an association between MSI and temozolomide-mediated hypermutagenesis in glioblastoma36,41. Taken together, these findings are consistent with a hypermutated phenotype secondary to temozolomide therapy in our patient’s gliosarcoma. Unfortunately, the WES data were not available in a timely enough manner to guide further treatment for our patient, such as the possibility of immunotherapy. Considering the clinical timeline of temozolomide treatment for her initial intracranial glioma but prior to spinal gliosarcoma diagnosis, this result also supports the notion of gliosarcomatous transformation from her lower grade glioma, rather than de novo tumor formation. We acknowledge that the inability to perform further genetic testing of her brain tumor is a key limitation of this study. Lastly, there are few reports of comprehensive genome sequencing of gliosarcomas, which have described key somatic mutations in known oncogenes including TP53, PTEN, RB1, and NF1 as well as amplification events of EGFR, CDK4/6, PDGFRA, MDM2, AKT1, and MET42–44. However, this case represents the first report of hypermutagenesis within gliosarcoma and raises the possibility that acquisition of this genotype may be a mechanism of gliosarcomatous transformation from lower grade tumors.
We considered whether our patient’s gliosarcoma arose de novo with a predisposition from her detected germline MSH2 variant. Sa et al identified a subset of treatment-naïve gliomas with de novo hypermutagenesis that had not been previously exposed to chemoradiation at the time of diagnosis45. Notably, these patients demonstrated germline mutations of MMR associated genes, including MSH2, MSH3, MSH6, and MLH3 and relevant family histories of various cancers, together suggestive of diagnosis of Lynch syndrome. Likewise, gliosarcoma has been described in Turcot syndrome46, a condition characterized by familial predisposition to colorectal and brain cancers, secondary to germline mutations of the MMR genes47. In contrast, our patient’s germline variant in MSH2 has been previously characterized as a benign variant, reported in individuals with colorectal, prostate, and ovarian cancers but not meeting clinical criteria for Lynch or Turcot syndrome or showing strong evidence for causality48,49. Furthermore, the lack of a second hit for MSH2 supports the notion of our patient’s variant as benign. As such, it remains unlikely that our patient’s germline MSH2 variant contributed to de novo tumor formation and/or acquisition of a hypermutator phenotype.
In summary, we describe the first case of a hypermutator phenotype associated with a gliosarcoma, characterized by somatic mutations in known MMR genes and consistent with temozolomide-mediated hypermutagenesis. Based on patterns of the described literature, it is more likely that our patient’s spinal cord gliosarcoma represents malignant progression and metastasis from her previous low-grade brain astrocytoma. As gliosarcoma remains a rare entity within primary malignant CNS tumors, even more so within primary spinal cord tumors, further comprehensive genomic characterization is needed to understand the genetic factors that can lead to malignant transformation from lower grade gliomas, as well as de novo formation of gliosarcomas.
Methods
Patient approval
This study was approved by Yale University’s Human Investigations Committee and Human Research Protection Program. Written informed consent was obtained from the parents of the patient. Specimens from resected tumors were evaluated microscopically by a board-certified neuropathologist.
WES and analysis
Genomic DNA from the resected tumor specimen and blood were isolated, and exome capture was performed with IDT xGen Exome Research Panel v1 with the additional spike-in regions of ~620 kb of RefGene coding regions. Sequencing of the captured regions was performed at Yale Center for Genome Analysis (YCGA) using Illumina NovaSeq6000 with 2 × 100 bp reads. Mean coverage of 152.4x and 230.6x was achieved for normal and tumor, respectively. Pre-processing of the raw data, including alignment, PCR duplicate identification, re-alignment around INDELs, and base quality score recalibration was performed “GATK Best Practices” guidelines (GATK v4.1.8.1). Somatic variant calling for SNV/INDELs together with CNV analysis and annotation was performed as previously described in reports from our institution50. Microsatellite status was determined with MSIsensor, a publically available computational algorithm designed to predict microsatellite stability based on WES data13.
Reporting summary
Further information on research design is available in the Nature Research Reporting Summary linked to this article.
Supplementary information
REPORTING SUMMARY
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Supplementary information
The online version contains supplementary material available at 10.1038/s41698-021-00143-w.
Author contributions
C.S.H., G.A.K., D.M., N.A.B., M.L.D., and E.Z.E. drafted the manuscript. C.S.H., G.A.K., A.J.K., A.A.E., A.B.K., D.M., N.A.B., and M.L.D. were involved in the clinical care of the patient and provided relevant material. C.S.H., E.Z.E., D.M., M.L.D., and E.Z.E. analyzed the data. All authors critically reviewed and revised the manuscript, including final approval of the completed version. C.S.H., M.L.D., and E.Z.E. conceived and supervised the study.
Data availability
The data generated and/or analyzed during the related study are described in the figshare metadata record: 10.6084/m9.figshare.1334755751. The whole-exome sequencing data of the resected spinal tumor are shared openly in the European Genotype-phenotype Archive under accession https://identifiers.org/ega.study:EGAS0000100486452.
Competing interests
The authors declare no competing interests. | TEMOZOLOMIDE | DrugsGivenReaction | CC BY | 33580181 | 19,179,073 | 2021-02-12 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Gliosarcoma'. | Hypermutated phenotype in gliosarcoma of the spinal cord.
Gliosarcoma is a variant of glioblastoma with equally poor prognosis and characterized by mixed glial and mesenchymal pathology. Metastasis is not uncommon but the involvement of the spinal cord is rare, and comprehensive genetic characterization of spinal gliosarcoma is lacking. We describe a patient initially diagnosed with a low-grade brain glioma via biopsy, followed by adjuvant radiation and temozolomide treatment. Nearly 2 years after diagnosis, she developed neurological deficits from an intradural, extramedullary tumor anterior to the spinal cord at T4, which was resected and diagnosed as gliosarcoma. Whole-exome sequencing (WES) of this tumor revealed a hypermutated phenotype, characterized by somatic mutations in key DNA mismatch repair (MMR) pathway genes, an abundance of C>T transitions within the identified somatic single nucleotide variations, and microsatellite stability, together consistent with temozolomide-mediated hypermutagenesis. This is the first report of a hypermutator phenotype in gliosarcoma, which may represent a novel genomic mechanism of progression from lower grade glioma.
Introduction
Gliosarcomas represent a variant of glioblastoma, characterized by mixed histopathological features of glial and mesenchymal origins1. Similar to glioblastoma, patients diagnosed with gliosarcoma exhibit poor prognosis and overall survival2. Metastasis is relatively common within the central nervous system (CNS), as well as systemic visceral sites1. Gliosarcomas involving the spinal cord are rare, making up approximately 1% of all malignant gliomas of the spine3 and may represent metastasis from intracranial tumors or less commonly, de novo formation4,5. To date, comprehensive genomic characterization of gliosarcomas remains vastly under-reported, compared to gliobastomas.
In this study, we report a patient with a spinal cord gliosarcoma with prior history of a low-grade brain glioma treated with adjuvant chemoradiation. Whole-exome sequencing (WES) of the resected spinal tumor revealed the first case of a hypermutator phenotype to be described in gliosarcoma.
Results
Case description
A 65-year-old female presented to an outside institution with several weeks of headaches and acute worsening of progressive numbness in her right lower extremity. Her past medical history was only significant for basal cell carcinoma of the left nasal ala, resected 5 years prior. Her family history was notable for breast cancer of middle-aged onset in her sister, mother, and grandmother but no prior diagnosis of a familial cancer predisposition syndrome. Her symptoms prompted magnetic resonance imaging (MRI) of the brain, which revealed a predominantly non-enhancing right parietal lesion with patchy enhancing foci extending to the splenium of the corpus callosum (Fig. 1a, b). Further imaging with computed tomography (CT) of the chest, abdomen, and pelvis did not reveal a primary systemic tumor to suggest a diagnosis of brain metastasis. She subsequently underwent stereotactic biopsy of her lesion with pathology demonstrating a glial neoplasm, consistent with World Health Organization (WHO) grade II astrocytoma (Fig. 1e–h). The Ki-67 index was 1–3% and immunohistochemical staining for TP53 was positive. Further molecular testing was negative for the EGFR variant III, IDH1 mutation, and MGMT promoter methylation. She was subsequently referred to our institution for further adjuvant treatment, where she underwent adjuvant radiation therapy and temozolomide treatment, as well as use of tumor-treating fields (TTF) with the Optune device (Novocure, Jersey, U.K.). A brain MRI obtained several months after completion of chemoradiation and TTF showed radiographic response with stable small foci of enhancement and drastic reduction in perilesional edema (Fig. 1c, d). Clinically, the patient was stable with moderate cognitive deficits and requirement of mild assistance with activities of daily living.Fig. 1 Imaging and histopathology of the low-grade glioma.
a Representative axial slices of T1-weighted MRI after contrast administration and (b) T2-weighted FLAIR obtained prior to stereotactic brain biopsy show a predominantly non-enhancing lesion within the deep right parietal lobe with small areas of patchy enhancement that is crossing the corpus callosum with associated edema. Repeat imaging obtained nearly 2 years after biopsy and completion of adjuvant chemoradation demonstrates (c) stable foci of small nodular enhancement on T1-weighted MRI after contrast administration with (d) minimal associated edema on T2-weighted FLAIR. e, f An infiltrating astrocytoma without necrosis or vascular proliferation is seen on hematoxylin & eosin staining. g Immunohistochemistry for GFAP-positive tumor cells is shown and (h) the Ki-67 proliferative index was ~3%.
Six months after completion of chemotherapy, she presented with increasing gait instability of two weeks duration and worsening thoracic back pain, localized between her shoulder blades. MRI of her thoracic and lumbar spine was obtained, demonstrating a homogenously enhancing dural-based, extramedullary lesion anterior to the spinal cord at T4 with moderate cord compression, causing edema extending from T2 to T7 (Fig. 2a–d). She was neurologically intact on examination other than brisk lower extremity reflexes, allodynia at the T4 level, and baseline right-sided hemibody numbness. Imaging findings were consistent with spinal meningioma, and the patient was admitted with plans for elective, inpatient surgery. However, two days after admission, she developed acute weakness in her bilateral lower extremities and subsequently was taken to the operating room for urgent resection of her tumor, which was removed in its entirety. Intra-operatively, the tumor was noted to be dural-based with a well-defined plane between tumor and the dura as well as the pia of the spinal cord. Motor evoked and somatosensory evoked potentials were improved at the end of the case, compared to baseline levels. Final pathology from the resected specimen was WHO IV gliosarcoma with biphasic tissue architecture, comprised of prominent GFAP-positive glial regions admixed with GFAP-negative spindled areas containing reticulin fibers (Fig. 2e–h). The Ki-67 index was elevated >10–15% and further routine molecular testing demonstrated MGMT promoter methylation but was otherwise negative for IDH1 mutation and ATRX loss.Fig. 2 Imaging and histopathology of the gliosarcoma.
a Representative sagittal and (b) axial slices from T1-weighted MRI after contrast administration show a homogeneously enhancing intradural, extramedullary lesion anterior to the spinal cord at the T4 level with associated cord compression seen on (c) sagittal and (d) axial slices on T2-weighted MRI. e Biphasic architecture is seen on hematoxylin & eosin staining, including (f) reticulin rich sarcomatous regions, interspersed with (g) GFAP-positive glial regions, and (h) an elevated Ki-67 proliferative index of ~10–15%.
The patient was subsequently discharged to rehabilitation and exhibited some improvement in lower extremity strength, albeit remained non-ambulatory and required significant assistance with activities of daily living. Further treatment with adjuvant chemoradiation was considered but given the unfavorable prognosis from new gliosarcoma diagnosis and poor performance status, she elected to undergo hospice care without further imaging or treatment and subsequently expired one month after surgery.
Genomic analysis
WES was performed on the gliosarcoma specimen, together with matching normal blood, to identify somatic single nucleotide variations (SNVs), insertions/deletions (INDEL), and copy number variations (CNV). The protocol for sequencing and analysis was in accordance with our previously described methods6. We detected 2372 somatic SNVs with a 96% C>T transition ratio (Fig. 3), consistent with a hypermutated phenotype. There were disease-associated alterations in key oncogenes, PTEN, PIK3R1, and ATM. Furthermore, there were key mutations in genes associated with the DNA mismatch repair (MMR) pathway, predicted to be deleterious based on scores from the following computational prediction tools: SIFT7, Polyphen2 HDIV Prediction8, Polyphen2 HVAR Prediction8, FATHMM9, MetaSVM10, and MutationTaster11. These included deleterious missense (p.P656L) and stop-gain (p.W105X) mutations in MSH6 and a stop-gain mutation in MSH3 (p.R727X). Additionally, there was a heterozygous germline variant in MSH2 (p.H46Q) of unknown significance. Somatic CNV analysis revealed amplifications of chromosomes 7, 20, and X, deletions on chromosomes 10, 13, 17, and focal CNV events on chromosomes 1 and 5. 27% of the genome was altered by CNV and/or loss-of-heterozygosity events.Fig. 3 Mutational signature plot of SNV in the resected spinal tumor specimen.
There was an overwhelming prevalence of C>T, with a SNV transition ratio of 96%, consistent with a hypermutated phenotype.
Given the relationship between microsatellite instability (MSI) and MMR deficiency in systemic cancers12, we performed further computational analysis with MSIsensor, an algorithm that predicts the microsatellite status of tumors from WES data13. The calculated MSI-score was 0.15, indicative of microsatellite stability per the tool guidelines, which uses a cut-off score of 3.5 for MSI.
Discussion
Gliosarcomas have a high predilection for metastasis to both visceral sites as well as the CNS, via hematogenous and leptomeningeal spread14. As such, most cases of gliosarcomas involving the spinal cord have been secondary to metastasis from intracranial origin4,15–19. In contrast, primary gliosarcomas of the spinal cord are less common5,20–22 and are vastly outnumbered by glioblastoma when analyzing primary malignant gliomas of the spinal cord3. Interestingly, these tumors may present as intramedullary15,23, as well as intradural, extramedullary lesions14,22, regardless of primary or metastatic etiology. Similar to their intracranial counterparts, spinal cord gliosarcomas demonstrate avid and typically homogenous enhancement, which in cases of intradural, extramedullary location, may be mistaken for a meningioma, as demonstrated in our patient. A prior history of a primary brain tumor must raise suspicion for an alternative diagnosis and highlights the need for surgical intervention to aid in definitive tissue diagnosis.
It remains unclear whether the gliosarcoma in our patient represented metastasis from her intracranial astrocytoma or de novo formation of a new primary tumor. This may have been addressed with additional WES data or targeted molecular testing from her original brain tumor, but unfortunately, inadequate tissue was available for further genomic studies. Additionally, further molecular testing of her brain tumor would have been helpful, as IDH-wildtype astrocytomas with grade II histology may exhibit survival outcomes akin to IDH-wildtype glioblastoma24,25. Based on recommendations from the Consortium to Inform Molecular and Practical Approaches to CNS Tumor Taxonomy (cIMPACT-NOW)26, molecular testing to assess for EGFR amplification, chromosome 7 gain/chromosome 10 loss, and TERT promoter mutations to diagnose a diffuse astrocytic glioma, IDH-wildtype, with molecular features of glioblastoma, WHO grade IV could have been performed to predict increased biological aggression and perhaps tailor clinical management27,28. The existing literature of metastatic gliosarcomas to the spinal cord have all been from primary intracranial gliosarcoma, so the notion that our patient’s gliosarcoma represented metastasis, despite the difference in WHO grade from her initial tumor, would be unique. There have been few reports of progression of low-grade gliomas into intracranial gliosarcomas, in instances with and without adjuvant chemoradiation29–31. These studies confirmed metastatic origin through single nucleotide polymorphism (SNP) arrays and fluorescence in-situ hybridization (FISH) for chromosomal analysis, but to date, comprehensive sequencing to delineate the genetic changes that may underlie either malignant progression into gliosarcomatous pathology or de novo spinal cord gliosarcoma formation have not been described.
In recent years, the prevalence of hypermutagenesis has been described across a variety of cancers, including gliomas32–34 and particularly after therapy-driven changes in tumor evolution at the time of recurrence. In particular, treatment with temozolomide may induce mutations in the MMR pathway, leading to therapeutic resistance and acquisition of the hypermutated phenotype35,36. Strikingly, we found a hypermutated phenotype in our patient’s specimen, including mutations in MMR genes, MSH3 and MSH6, which previously have not been associated with gliosarcoma pathology. Likewise, the abundance of C>T transitions detected within the somatic SNVs of our patient’s tumor was consistent with a MMR deficient hypermutated phenotype. We further explored the possibility of MSI, particularly given the strong correlation between MSI and MMR deficiency in systemic cancers that may serve as biomarkers for response to immunotherapies37–40. Utilizing the MSIsensor computational tool, we did not detect MSI despite the hypermutator phenotype of our patient’s tumor, a result consistent with prior studies failing to reveal an association between MSI and temozolomide-mediated hypermutagenesis in glioblastoma36,41. Taken together, these findings are consistent with a hypermutated phenotype secondary to temozolomide therapy in our patient’s gliosarcoma. Unfortunately, the WES data were not available in a timely enough manner to guide further treatment for our patient, such as the possibility of immunotherapy. Considering the clinical timeline of temozolomide treatment for her initial intracranial glioma but prior to spinal gliosarcoma diagnosis, this result also supports the notion of gliosarcomatous transformation from her lower grade glioma, rather than de novo tumor formation. We acknowledge that the inability to perform further genetic testing of her brain tumor is a key limitation of this study. Lastly, there are few reports of comprehensive genome sequencing of gliosarcomas, which have described key somatic mutations in known oncogenes including TP53, PTEN, RB1, and NF1 as well as amplification events of EGFR, CDK4/6, PDGFRA, MDM2, AKT1, and MET42–44. However, this case represents the first report of hypermutagenesis within gliosarcoma and raises the possibility that acquisition of this genotype may be a mechanism of gliosarcomatous transformation from lower grade tumors.
We considered whether our patient’s gliosarcoma arose de novo with a predisposition from her detected germline MSH2 variant. Sa et al identified a subset of treatment-naïve gliomas with de novo hypermutagenesis that had not been previously exposed to chemoradiation at the time of diagnosis45. Notably, these patients demonstrated germline mutations of MMR associated genes, including MSH2, MSH3, MSH6, and MLH3 and relevant family histories of various cancers, together suggestive of diagnosis of Lynch syndrome. Likewise, gliosarcoma has been described in Turcot syndrome46, a condition characterized by familial predisposition to colorectal and brain cancers, secondary to germline mutations of the MMR genes47. In contrast, our patient’s germline variant in MSH2 has been previously characterized as a benign variant, reported in individuals with colorectal, prostate, and ovarian cancers but not meeting clinical criteria for Lynch or Turcot syndrome or showing strong evidence for causality48,49. Furthermore, the lack of a second hit for MSH2 supports the notion of our patient’s variant as benign. As such, it remains unlikely that our patient’s germline MSH2 variant contributed to de novo tumor formation and/or acquisition of a hypermutator phenotype.
In summary, we describe the first case of a hypermutator phenotype associated with a gliosarcoma, characterized by somatic mutations in known MMR genes and consistent with temozolomide-mediated hypermutagenesis. Based on patterns of the described literature, it is more likely that our patient’s spinal cord gliosarcoma represents malignant progression and metastasis from her previous low-grade brain astrocytoma. As gliosarcoma remains a rare entity within primary malignant CNS tumors, even more so within primary spinal cord tumors, further comprehensive genomic characterization is needed to understand the genetic factors that can lead to malignant transformation from lower grade gliomas, as well as de novo formation of gliosarcomas.
Methods
Patient approval
This study was approved by Yale University’s Human Investigations Committee and Human Research Protection Program. Written informed consent was obtained from the parents of the patient. Specimens from resected tumors were evaluated microscopically by a board-certified neuropathologist.
WES and analysis
Genomic DNA from the resected tumor specimen and blood were isolated, and exome capture was performed with IDT xGen Exome Research Panel v1 with the additional spike-in regions of ~620 kb of RefGene coding regions. Sequencing of the captured regions was performed at Yale Center for Genome Analysis (YCGA) using Illumina NovaSeq6000 with 2 × 100 bp reads. Mean coverage of 152.4x and 230.6x was achieved for normal and tumor, respectively. Pre-processing of the raw data, including alignment, PCR duplicate identification, re-alignment around INDELs, and base quality score recalibration was performed “GATK Best Practices” guidelines (GATK v4.1.8.1). Somatic variant calling for SNV/INDELs together with CNV analysis and annotation was performed as previously described in reports from our institution50. Microsatellite status was determined with MSIsensor, a publically available computational algorithm designed to predict microsatellite stability based on WES data13.
Reporting summary
Further information on research design is available in the Nature Research Reporting Summary linked to this article.
Supplementary information
REPORTING SUMMARY
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Supplementary information
The online version contains supplementary material available at 10.1038/s41698-021-00143-w.
Author contributions
C.S.H., G.A.K., D.M., N.A.B., M.L.D., and E.Z.E. drafted the manuscript. C.S.H., G.A.K., A.J.K., A.A.E., A.B.K., D.M., N.A.B., and M.L.D. were involved in the clinical care of the patient and provided relevant material. C.S.H., E.Z.E., D.M., M.L.D., and E.Z.E. analyzed the data. All authors critically reviewed and revised the manuscript, including final approval of the completed version. C.S.H., M.L.D., and E.Z.E. conceived and supervised the study.
Data availability
The data generated and/or analyzed during the related study are described in the figshare metadata record: 10.6084/m9.figshare.1334755751. The whole-exome sequencing data of the resected spinal tumor are shared openly in the European Genotype-phenotype Archive under accession https://identifiers.org/ega.study:EGAS0000100486452.
Competing interests
The authors declare no competing interests. | TEMOZOLOMIDE | DrugsGivenReaction | CC BY | 33580181 | 18,945,561 | 2021-02-12 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Hypermutation'. | Hypermutated phenotype in gliosarcoma of the spinal cord.
Gliosarcoma is a variant of glioblastoma with equally poor prognosis and characterized by mixed glial and mesenchymal pathology. Metastasis is not uncommon but the involvement of the spinal cord is rare, and comprehensive genetic characterization of spinal gliosarcoma is lacking. We describe a patient initially diagnosed with a low-grade brain glioma via biopsy, followed by adjuvant radiation and temozolomide treatment. Nearly 2 years after diagnosis, she developed neurological deficits from an intradural, extramedullary tumor anterior to the spinal cord at T4, which was resected and diagnosed as gliosarcoma. Whole-exome sequencing (WES) of this tumor revealed a hypermutated phenotype, characterized by somatic mutations in key DNA mismatch repair (MMR) pathway genes, an abundance of C>T transitions within the identified somatic single nucleotide variations, and microsatellite stability, together consistent with temozolomide-mediated hypermutagenesis. This is the first report of a hypermutator phenotype in gliosarcoma, which may represent a novel genomic mechanism of progression from lower grade glioma.
Introduction
Gliosarcomas represent a variant of glioblastoma, characterized by mixed histopathological features of glial and mesenchymal origins1. Similar to glioblastoma, patients diagnosed with gliosarcoma exhibit poor prognosis and overall survival2. Metastasis is relatively common within the central nervous system (CNS), as well as systemic visceral sites1. Gliosarcomas involving the spinal cord are rare, making up approximately 1% of all malignant gliomas of the spine3 and may represent metastasis from intracranial tumors or less commonly, de novo formation4,5. To date, comprehensive genomic characterization of gliosarcomas remains vastly under-reported, compared to gliobastomas.
In this study, we report a patient with a spinal cord gliosarcoma with prior history of a low-grade brain glioma treated with adjuvant chemoradiation. Whole-exome sequencing (WES) of the resected spinal tumor revealed the first case of a hypermutator phenotype to be described in gliosarcoma.
Results
Case description
A 65-year-old female presented to an outside institution with several weeks of headaches and acute worsening of progressive numbness in her right lower extremity. Her past medical history was only significant for basal cell carcinoma of the left nasal ala, resected 5 years prior. Her family history was notable for breast cancer of middle-aged onset in her sister, mother, and grandmother but no prior diagnosis of a familial cancer predisposition syndrome. Her symptoms prompted magnetic resonance imaging (MRI) of the brain, which revealed a predominantly non-enhancing right parietal lesion with patchy enhancing foci extending to the splenium of the corpus callosum (Fig. 1a, b). Further imaging with computed tomography (CT) of the chest, abdomen, and pelvis did not reveal a primary systemic tumor to suggest a diagnosis of brain metastasis. She subsequently underwent stereotactic biopsy of her lesion with pathology demonstrating a glial neoplasm, consistent with World Health Organization (WHO) grade II astrocytoma (Fig. 1e–h). The Ki-67 index was 1–3% and immunohistochemical staining for TP53 was positive. Further molecular testing was negative for the EGFR variant III, IDH1 mutation, and MGMT promoter methylation. She was subsequently referred to our institution for further adjuvant treatment, where she underwent adjuvant radiation therapy and temozolomide treatment, as well as use of tumor-treating fields (TTF) with the Optune device (Novocure, Jersey, U.K.). A brain MRI obtained several months after completion of chemoradiation and TTF showed radiographic response with stable small foci of enhancement and drastic reduction in perilesional edema (Fig. 1c, d). Clinically, the patient was stable with moderate cognitive deficits and requirement of mild assistance with activities of daily living.Fig. 1 Imaging and histopathology of the low-grade glioma.
a Representative axial slices of T1-weighted MRI after contrast administration and (b) T2-weighted FLAIR obtained prior to stereotactic brain biopsy show a predominantly non-enhancing lesion within the deep right parietal lobe with small areas of patchy enhancement that is crossing the corpus callosum with associated edema. Repeat imaging obtained nearly 2 years after biopsy and completion of adjuvant chemoradation demonstrates (c) stable foci of small nodular enhancement on T1-weighted MRI after contrast administration with (d) minimal associated edema on T2-weighted FLAIR. e, f An infiltrating astrocytoma without necrosis or vascular proliferation is seen on hematoxylin & eosin staining. g Immunohistochemistry for GFAP-positive tumor cells is shown and (h) the Ki-67 proliferative index was ~3%.
Six months after completion of chemotherapy, she presented with increasing gait instability of two weeks duration and worsening thoracic back pain, localized between her shoulder blades. MRI of her thoracic and lumbar spine was obtained, demonstrating a homogenously enhancing dural-based, extramedullary lesion anterior to the spinal cord at T4 with moderate cord compression, causing edema extending from T2 to T7 (Fig. 2a–d). She was neurologically intact on examination other than brisk lower extremity reflexes, allodynia at the T4 level, and baseline right-sided hemibody numbness. Imaging findings were consistent with spinal meningioma, and the patient was admitted with plans for elective, inpatient surgery. However, two days after admission, she developed acute weakness in her bilateral lower extremities and subsequently was taken to the operating room for urgent resection of her tumor, which was removed in its entirety. Intra-operatively, the tumor was noted to be dural-based with a well-defined plane between tumor and the dura as well as the pia of the spinal cord. Motor evoked and somatosensory evoked potentials were improved at the end of the case, compared to baseline levels. Final pathology from the resected specimen was WHO IV gliosarcoma with biphasic tissue architecture, comprised of prominent GFAP-positive glial regions admixed with GFAP-negative spindled areas containing reticulin fibers (Fig. 2e–h). The Ki-67 index was elevated >10–15% and further routine molecular testing demonstrated MGMT promoter methylation but was otherwise negative for IDH1 mutation and ATRX loss.Fig. 2 Imaging and histopathology of the gliosarcoma.
a Representative sagittal and (b) axial slices from T1-weighted MRI after contrast administration show a homogeneously enhancing intradural, extramedullary lesion anterior to the spinal cord at the T4 level with associated cord compression seen on (c) sagittal and (d) axial slices on T2-weighted MRI. e Biphasic architecture is seen on hematoxylin & eosin staining, including (f) reticulin rich sarcomatous regions, interspersed with (g) GFAP-positive glial regions, and (h) an elevated Ki-67 proliferative index of ~10–15%.
The patient was subsequently discharged to rehabilitation and exhibited some improvement in lower extremity strength, albeit remained non-ambulatory and required significant assistance with activities of daily living. Further treatment with adjuvant chemoradiation was considered but given the unfavorable prognosis from new gliosarcoma diagnosis and poor performance status, she elected to undergo hospice care without further imaging or treatment and subsequently expired one month after surgery.
Genomic analysis
WES was performed on the gliosarcoma specimen, together with matching normal blood, to identify somatic single nucleotide variations (SNVs), insertions/deletions (INDEL), and copy number variations (CNV). The protocol for sequencing and analysis was in accordance with our previously described methods6. We detected 2372 somatic SNVs with a 96% C>T transition ratio (Fig. 3), consistent with a hypermutated phenotype. There were disease-associated alterations in key oncogenes, PTEN, PIK3R1, and ATM. Furthermore, there were key mutations in genes associated with the DNA mismatch repair (MMR) pathway, predicted to be deleterious based on scores from the following computational prediction tools: SIFT7, Polyphen2 HDIV Prediction8, Polyphen2 HVAR Prediction8, FATHMM9, MetaSVM10, and MutationTaster11. These included deleterious missense (p.P656L) and stop-gain (p.W105X) mutations in MSH6 and a stop-gain mutation in MSH3 (p.R727X). Additionally, there was a heterozygous germline variant in MSH2 (p.H46Q) of unknown significance. Somatic CNV analysis revealed amplifications of chromosomes 7, 20, and X, deletions on chromosomes 10, 13, 17, and focal CNV events on chromosomes 1 and 5. 27% of the genome was altered by CNV and/or loss-of-heterozygosity events.Fig. 3 Mutational signature plot of SNV in the resected spinal tumor specimen.
There was an overwhelming prevalence of C>T, with a SNV transition ratio of 96%, consistent with a hypermutated phenotype.
Given the relationship between microsatellite instability (MSI) and MMR deficiency in systemic cancers12, we performed further computational analysis with MSIsensor, an algorithm that predicts the microsatellite status of tumors from WES data13. The calculated MSI-score was 0.15, indicative of microsatellite stability per the tool guidelines, which uses a cut-off score of 3.5 for MSI.
Discussion
Gliosarcomas have a high predilection for metastasis to both visceral sites as well as the CNS, via hematogenous and leptomeningeal spread14. As such, most cases of gliosarcomas involving the spinal cord have been secondary to metastasis from intracranial origin4,15–19. In contrast, primary gliosarcomas of the spinal cord are less common5,20–22 and are vastly outnumbered by glioblastoma when analyzing primary malignant gliomas of the spinal cord3. Interestingly, these tumors may present as intramedullary15,23, as well as intradural, extramedullary lesions14,22, regardless of primary or metastatic etiology. Similar to their intracranial counterparts, spinal cord gliosarcomas demonstrate avid and typically homogenous enhancement, which in cases of intradural, extramedullary location, may be mistaken for a meningioma, as demonstrated in our patient. A prior history of a primary brain tumor must raise suspicion for an alternative diagnosis and highlights the need for surgical intervention to aid in definitive tissue diagnosis.
It remains unclear whether the gliosarcoma in our patient represented metastasis from her intracranial astrocytoma or de novo formation of a new primary tumor. This may have been addressed with additional WES data or targeted molecular testing from her original brain tumor, but unfortunately, inadequate tissue was available for further genomic studies. Additionally, further molecular testing of her brain tumor would have been helpful, as IDH-wildtype astrocytomas with grade II histology may exhibit survival outcomes akin to IDH-wildtype glioblastoma24,25. Based on recommendations from the Consortium to Inform Molecular and Practical Approaches to CNS Tumor Taxonomy (cIMPACT-NOW)26, molecular testing to assess for EGFR amplification, chromosome 7 gain/chromosome 10 loss, and TERT promoter mutations to diagnose a diffuse astrocytic glioma, IDH-wildtype, with molecular features of glioblastoma, WHO grade IV could have been performed to predict increased biological aggression and perhaps tailor clinical management27,28. The existing literature of metastatic gliosarcomas to the spinal cord have all been from primary intracranial gliosarcoma, so the notion that our patient’s gliosarcoma represented metastasis, despite the difference in WHO grade from her initial tumor, would be unique. There have been few reports of progression of low-grade gliomas into intracranial gliosarcomas, in instances with and without adjuvant chemoradiation29–31. These studies confirmed metastatic origin through single nucleotide polymorphism (SNP) arrays and fluorescence in-situ hybridization (FISH) for chromosomal analysis, but to date, comprehensive sequencing to delineate the genetic changes that may underlie either malignant progression into gliosarcomatous pathology or de novo spinal cord gliosarcoma formation have not been described.
In recent years, the prevalence of hypermutagenesis has been described across a variety of cancers, including gliomas32–34 and particularly after therapy-driven changes in tumor evolution at the time of recurrence. In particular, treatment with temozolomide may induce mutations in the MMR pathway, leading to therapeutic resistance and acquisition of the hypermutated phenotype35,36. Strikingly, we found a hypermutated phenotype in our patient’s specimen, including mutations in MMR genes, MSH3 and MSH6, which previously have not been associated with gliosarcoma pathology. Likewise, the abundance of C>T transitions detected within the somatic SNVs of our patient’s tumor was consistent with a MMR deficient hypermutated phenotype. We further explored the possibility of MSI, particularly given the strong correlation between MSI and MMR deficiency in systemic cancers that may serve as biomarkers for response to immunotherapies37–40. Utilizing the MSIsensor computational tool, we did not detect MSI despite the hypermutator phenotype of our patient’s tumor, a result consistent with prior studies failing to reveal an association between MSI and temozolomide-mediated hypermutagenesis in glioblastoma36,41. Taken together, these findings are consistent with a hypermutated phenotype secondary to temozolomide therapy in our patient’s gliosarcoma. Unfortunately, the WES data were not available in a timely enough manner to guide further treatment for our patient, such as the possibility of immunotherapy. Considering the clinical timeline of temozolomide treatment for her initial intracranial glioma but prior to spinal gliosarcoma diagnosis, this result also supports the notion of gliosarcomatous transformation from her lower grade glioma, rather than de novo tumor formation. We acknowledge that the inability to perform further genetic testing of her brain tumor is a key limitation of this study. Lastly, there are few reports of comprehensive genome sequencing of gliosarcomas, which have described key somatic mutations in known oncogenes including TP53, PTEN, RB1, and NF1 as well as amplification events of EGFR, CDK4/6, PDGFRA, MDM2, AKT1, and MET42–44. However, this case represents the first report of hypermutagenesis within gliosarcoma and raises the possibility that acquisition of this genotype may be a mechanism of gliosarcomatous transformation from lower grade tumors.
We considered whether our patient’s gliosarcoma arose de novo with a predisposition from her detected germline MSH2 variant. Sa et al identified a subset of treatment-naïve gliomas with de novo hypermutagenesis that had not been previously exposed to chemoradiation at the time of diagnosis45. Notably, these patients demonstrated germline mutations of MMR associated genes, including MSH2, MSH3, MSH6, and MLH3 and relevant family histories of various cancers, together suggestive of diagnosis of Lynch syndrome. Likewise, gliosarcoma has been described in Turcot syndrome46, a condition characterized by familial predisposition to colorectal and brain cancers, secondary to germline mutations of the MMR genes47. In contrast, our patient’s germline variant in MSH2 has been previously characterized as a benign variant, reported in individuals with colorectal, prostate, and ovarian cancers but not meeting clinical criteria for Lynch or Turcot syndrome or showing strong evidence for causality48,49. Furthermore, the lack of a second hit for MSH2 supports the notion of our patient’s variant as benign. As such, it remains unlikely that our patient’s germline MSH2 variant contributed to de novo tumor formation and/or acquisition of a hypermutator phenotype.
In summary, we describe the first case of a hypermutator phenotype associated with a gliosarcoma, characterized by somatic mutations in known MMR genes and consistent with temozolomide-mediated hypermutagenesis. Based on patterns of the described literature, it is more likely that our patient’s spinal cord gliosarcoma represents malignant progression and metastasis from her previous low-grade brain astrocytoma. As gliosarcoma remains a rare entity within primary malignant CNS tumors, even more so within primary spinal cord tumors, further comprehensive genomic characterization is needed to understand the genetic factors that can lead to malignant transformation from lower grade gliomas, as well as de novo formation of gliosarcomas.
Methods
Patient approval
This study was approved by Yale University’s Human Investigations Committee and Human Research Protection Program. Written informed consent was obtained from the parents of the patient. Specimens from resected tumors were evaluated microscopically by a board-certified neuropathologist.
WES and analysis
Genomic DNA from the resected tumor specimen and blood were isolated, and exome capture was performed with IDT xGen Exome Research Panel v1 with the additional spike-in regions of ~620 kb of RefGene coding regions. Sequencing of the captured regions was performed at Yale Center for Genome Analysis (YCGA) using Illumina NovaSeq6000 with 2 × 100 bp reads. Mean coverage of 152.4x and 230.6x was achieved for normal and tumor, respectively. Pre-processing of the raw data, including alignment, PCR duplicate identification, re-alignment around INDELs, and base quality score recalibration was performed “GATK Best Practices” guidelines (GATK v4.1.8.1). Somatic variant calling for SNV/INDELs together with CNV analysis and annotation was performed as previously described in reports from our institution50. Microsatellite status was determined with MSIsensor, a publically available computational algorithm designed to predict microsatellite stability based on WES data13.
Reporting summary
Further information on research design is available in the Nature Research Reporting Summary linked to this article.
Supplementary information
REPORTING SUMMARY
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Supplementary information
The online version contains supplementary material available at 10.1038/s41698-021-00143-w.
Author contributions
C.S.H., G.A.K., D.M., N.A.B., M.L.D., and E.Z.E. drafted the manuscript. C.S.H., G.A.K., A.J.K., A.A.E., A.B.K., D.M., N.A.B., and M.L.D. were involved in the clinical care of the patient and provided relevant material. C.S.H., E.Z.E., D.M., M.L.D., and E.Z.E. analyzed the data. All authors critically reviewed and revised the manuscript, including final approval of the completed version. C.S.H., M.L.D., and E.Z.E. conceived and supervised the study.
Data availability
The data generated and/or analyzed during the related study are described in the figshare metadata record: 10.6084/m9.figshare.1334755751. The whole-exome sequencing data of the resected spinal tumor are shared openly in the European Genotype-phenotype Archive under accession https://identifiers.org/ega.study:EGAS0000100486452.
Competing interests
The authors declare no competing interests. | TEMOZOLOMIDE | DrugsGivenReaction | CC BY | 33580181 | 19,179,073 | 2021-02-12 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Off label use'. | Hypermutated phenotype in gliosarcoma of the spinal cord.
Gliosarcoma is a variant of glioblastoma with equally poor prognosis and characterized by mixed glial and mesenchymal pathology. Metastasis is not uncommon but the involvement of the spinal cord is rare, and comprehensive genetic characterization of spinal gliosarcoma is lacking. We describe a patient initially diagnosed with a low-grade brain glioma via biopsy, followed by adjuvant radiation and temozolomide treatment. Nearly 2 years after diagnosis, she developed neurological deficits from an intradural, extramedullary tumor anterior to the spinal cord at T4, which was resected and diagnosed as gliosarcoma. Whole-exome sequencing (WES) of this tumor revealed a hypermutated phenotype, characterized by somatic mutations in key DNA mismatch repair (MMR) pathway genes, an abundance of C>T transitions within the identified somatic single nucleotide variations, and microsatellite stability, together consistent with temozolomide-mediated hypermutagenesis. This is the first report of a hypermutator phenotype in gliosarcoma, which may represent a novel genomic mechanism of progression from lower grade glioma.
Introduction
Gliosarcomas represent a variant of glioblastoma, characterized by mixed histopathological features of glial and mesenchymal origins1. Similar to glioblastoma, patients diagnosed with gliosarcoma exhibit poor prognosis and overall survival2. Metastasis is relatively common within the central nervous system (CNS), as well as systemic visceral sites1. Gliosarcomas involving the spinal cord are rare, making up approximately 1% of all malignant gliomas of the spine3 and may represent metastasis from intracranial tumors or less commonly, de novo formation4,5. To date, comprehensive genomic characterization of gliosarcomas remains vastly under-reported, compared to gliobastomas.
In this study, we report a patient with a spinal cord gliosarcoma with prior history of a low-grade brain glioma treated with adjuvant chemoradiation. Whole-exome sequencing (WES) of the resected spinal tumor revealed the first case of a hypermutator phenotype to be described in gliosarcoma.
Results
Case description
A 65-year-old female presented to an outside institution with several weeks of headaches and acute worsening of progressive numbness in her right lower extremity. Her past medical history was only significant for basal cell carcinoma of the left nasal ala, resected 5 years prior. Her family history was notable for breast cancer of middle-aged onset in her sister, mother, and grandmother but no prior diagnosis of a familial cancer predisposition syndrome. Her symptoms prompted magnetic resonance imaging (MRI) of the brain, which revealed a predominantly non-enhancing right parietal lesion with patchy enhancing foci extending to the splenium of the corpus callosum (Fig. 1a, b). Further imaging with computed tomography (CT) of the chest, abdomen, and pelvis did not reveal a primary systemic tumor to suggest a diagnosis of brain metastasis. She subsequently underwent stereotactic biopsy of her lesion with pathology demonstrating a glial neoplasm, consistent with World Health Organization (WHO) grade II astrocytoma (Fig. 1e–h). The Ki-67 index was 1–3% and immunohistochemical staining for TP53 was positive. Further molecular testing was negative for the EGFR variant III, IDH1 mutation, and MGMT promoter methylation. She was subsequently referred to our institution for further adjuvant treatment, where she underwent adjuvant radiation therapy and temozolomide treatment, as well as use of tumor-treating fields (TTF) with the Optune device (Novocure, Jersey, U.K.). A brain MRI obtained several months after completion of chemoradiation and TTF showed radiographic response with stable small foci of enhancement and drastic reduction in perilesional edema (Fig. 1c, d). Clinically, the patient was stable with moderate cognitive deficits and requirement of mild assistance with activities of daily living.Fig. 1 Imaging and histopathology of the low-grade glioma.
a Representative axial slices of T1-weighted MRI after contrast administration and (b) T2-weighted FLAIR obtained prior to stereotactic brain biopsy show a predominantly non-enhancing lesion within the deep right parietal lobe with small areas of patchy enhancement that is crossing the corpus callosum with associated edema. Repeat imaging obtained nearly 2 years after biopsy and completion of adjuvant chemoradation demonstrates (c) stable foci of small nodular enhancement on T1-weighted MRI after contrast administration with (d) minimal associated edema on T2-weighted FLAIR. e, f An infiltrating astrocytoma without necrosis or vascular proliferation is seen on hematoxylin & eosin staining. g Immunohistochemistry for GFAP-positive tumor cells is shown and (h) the Ki-67 proliferative index was ~3%.
Six months after completion of chemotherapy, she presented with increasing gait instability of two weeks duration and worsening thoracic back pain, localized between her shoulder blades. MRI of her thoracic and lumbar spine was obtained, demonstrating a homogenously enhancing dural-based, extramedullary lesion anterior to the spinal cord at T4 with moderate cord compression, causing edema extending from T2 to T7 (Fig. 2a–d). She was neurologically intact on examination other than brisk lower extremity reflexes, allodynia at the T4 level, and baseline right-sided hemibody numbness. Imaging findings were consistent with spinal meningioma, and the patient was admitted with plans for elective, inpatient surgery. However, two days after admission, she developed acute weakness in her bilateral lower extremities and subsequently was taken to the operating room for urgent resection of her tumor, which was removed in its entirety. Intra-operatively, the tumor was noted to be dural-based with a well-defined plane between tumor and the dura as well as the pia of the spinal cord. Motor evoked and somatosensory evoked potentials were improved at the end of the case, compared to baseline levels. Final pathology from the resected specimen was WHO IV gliosarcoma with biphasic tissue architecture, comprised of prominent GFAP-positive glial regions admixed with GFAP-negative spindled areas containing reticulin fibers (Fig. 2e–h). The Ki-67 index was elevated >10–15% and further routine molecular testing demonstrated MGMT promoter methylation but was otherwise negative for IDH1 mutation and ATRX loss.Fig. 2 Imaging and histopathology of the gliosarcoma.
a Representative sagittal and (b) axial slices from T1-weighted MRI after contrast administration show a homogeneously enhancing intradural, extramedullary lesion anterior to the spinal cord at the T4 level with associated cord compression seen on (c) sagittal and (d) axial slices on T2-weighted MRI. e Biphasic architecture is seen on hematoxylin & eosin staining, including (f) reticulin rich sarcomatous regions, interspersed with (g) GFAP-positive glial regions, and (h) an elevated Ki-67 proliferative index of ~10–15%.
The patient was subsequently discharged to rehabilitation and exhibited some improvement in lower extremity strength, albeit remained non-ambulatory and required significant assistance with activities of daily living. Further treatment with adjuvant chemoradiation was considered but given the unfavorable prognosis from new gliosarcoma diagnosis and poor performance status, she elected to undergo hospice care without further imaging or treatment and subsequently expired one month after surgery.
Genomic analysis
WES was performed on the gliosarcoma specimen, together with matching normal blood, to identify somatic single nucleotide variations (SNVs), insertions/deletions (INDEL), and copy number variations (CNV). The protocol for sequencing and analysis was in accordance with our previously described methods6. We detected 2372 somatic SNVs with a 96% C>T transition ratio (Fig. 3), consistent with a hypermutated phenotype. There were disease-associated alterations in key oncogenes, PTEN, PIK3R1, and ATM. Furthermore, there were key mutations in genes associated with the DNA mismatch repair (MMR) pathway, predicted to be deleterious based on scores from the following computational prediction tools: SIFT7, Polyphen2 HDIV Prediction8, Polyphen2 HVAR Prediction8, FATHMM9, MetaSVM10, and MutationTaster11. These included deleterious missense (p.P656L) and stop-gain (p.W105X) mutations in MSH6 and a stop-gain mutation in MSH3 (p.R727X). Additionally, there was a heterozygous germline variant in MSH2 (p.H46Q) of unknown significance. Somatic CNV analysis revealed amplifications of chromosomes 7, 20, and X, deletions on chromosomes 10, 13, 17, and focal CNV events on chromosomes 1 and 5. 27% of the genome was altered by CNV and/or loss-of-heterozygosity events.Fig. 3 Mutational signature plot of SNV in the resected spinal tumor specimen.
There was an overwhelming prevalence of C>T, with a SNV transition ratio of 96%, consistent with a hypermutated phenotype.
Given the relationship between microsatellite instability (MSI) and MMR deficiency in systemic cancers12, we performed further computational analysis with MSIsensor, an algorithm that predicts the microsatellite status of tumors from WES data13. The calculated MSI-score was 0.15, indicative of microsatellite stability per the tool guidelines, which uses a cut-off score of 3.5 for MSI.
Discussion
Gliosarcomas have a high predilection for metastasis to both visceral sites as well as the CNS, via hematogenous and leptomeningeal spread14. As such, most cases of gliosarcomas involving the spinal cord have been secondary to metastasis from intracranial origin4,15–19. In contrast, primary gliosarcomas of the spinal cord are less common5,20–22 and are vastly outnumbered by glioblastoma when analyzing primary malignant gliomas of the spinal cord3. Interestingly, these tumors may present as intramedullary15,23, as well as intradural, extramedullary lesions14,22, regardless of primary or metastatic etiology. Similar to their intracranial counterparts, spinal cord gliosarcomas demonstrate avid and typically homogenous enhancement, which in cases of intradural, extramedullary location, may be mistaken for a meningioma, as demonstrated in our patient. A prior history of a primary brain tumor must raise suspicion for an alternative diagnosis and highlights the need for surgical intervention to aid in definitive tissue diagnosis.
It remains unclear whether the gliosarcoma in our patient represented metastasis from her intracranial astrocytoma or de novo formation of a new primary tumor. This may have been addressed with additional WES data or targeted molecular testing from her original brain tumor, but unfortunately, inadequate tissue was available for further genomic studies. Additionally, further molecular testing of her brain tumor would have been helpful, as IDH-wildtype astrocytomas with grade II histology may exhibit survival outcomes akin to IDH-wildtype glioblastoma24,25. Based on recommendations from the Consortium to Inform Molecular and Practical Approaches to CNS Tumor Taxonomy (cIMPACT-NOW)26, molecular testing to assess for EGFR amplification, chromosome 7 gain/chromosome 10 loss, and TERT promoter mutations to diagnose a diffuse astrocytic glioma, IDH-wildtype, with molecular features of glioblastoma, WHO grade IV could have been performed to predict increased biological aggression and perhaps tailor clinical management27,28. The existing literature of metastatic gliosarcomas to the spinal cord have all been from primary intracranial gliosarcoma, so the notion that our patient’s gliosarcoma represented metastasis, despite the difference in WHO grade from her initial tumor, would be unique. There have been few reports of progression of low-grade gliomas into intracranial gliosarcomas, in instances with and without adjuvant chemoradiation29–31. These studies confirmed metastatic origin through single nucleotide polymorphism (SNP) arrays and fluorescence in-situ hybridization (FISH) for chromosomal analysis, but to date, comprehensive sequencing to delineate the genetic changes that may underlie either malignant progression into gliosarcomatous pathology or de novo spinal cord gliosarcoma formation have not been described.
In recent years, the prevalence of hypermutagenesis has been described across a variety of cancers, including gliomas32–34 and particularly after therapy-driven changes in tumor evolution at the time of recurrence. In particular, treatment with temozolomide may induce mutations in the MMR pathway, leading to therapeutic resistance and acquisition of the hypermutated phenotype35,36. Strikingly, we found a hypermutated phenotype in our patient’s specimen, including mutations in MMR genes, MSH3 and MSH6, which previously have not been associated with gliosarcoma pathology. Likewise, the abundance of C>T transitions detected within the somatic SNVs of our patient’s tumor was consistent with a MMR deficient hypermutated phenotype. We further explored the possibility of MSI, particularly given the strong correlation between MSI and MMR deficiency in systemic cancers that may serve as biomarkers for response to immunotherapies37–40. Utilizing the MSIsensor computational tool, we did not detect MSI despite the hypermutator phenotype of our patient’s tumor, a result consistent with prior studies failing to reveal an association between MSI and temozolomide-mediated hypermutagenesis in glioblastoma36,41. Taken together, these findings are consistent with a hypermutated phenotype secondary to temozolomide therapy in our patient’s gliosarcoma. Unfortunately, the WES data were not available in a timely enough manner to guide further treatment for our patient, such as the possibility of immunotherapy. Considering the clinical timeline of temozolomide treatment for her initial intracranial glioma but prior to spinal gliosarcoma diagnosis, this result also supports the notion of gliosarcomatous transformation from her lower grade glioma, rather than de novo tumor formation. We acknowledge that the inability to perform further genetic testing of her brain tumor is a key limitation of this study. Lastly, there are few reports of comprehensive genome sequencing of gliosarcomas, which have described key somatic mutations in known oncogenes including TP53, PTEN, RB1, and NF1 as well as amplification events of EGFR, CDK4/6, PDGFRA, MDM2, AKT1, and MET42–44. However, this case represents the first report of hypermutagenesis within gliosarcoma and raises the possibility that acquisition of this genotype may be a mechanism of gliosarcomatous transformation from lower grade tumors.
We considered whether our patient’s gliosarcoma arose de novo with a predisposition from her detected germline MSH2 variant. Sa et al identified a subset of treatment-naïve gliomas with de novo hypermutagenesis that had not been previously exposed to chemoradiation at the time of diagnosis45. Notably, these patients demonstrated germline mutations of MMR associated genes, including MSH2, MSH3, MSH6, and MLH3 and relevant family histories of various cancers, together suggestive of diagnosis of Lynch syndrome. Likewise, gliosarcoma has been described in Turcot syndrome46, a condition characterized by familial predisposition to colorectal and brain cancers, secondary to germline mutations of the MMR genes47. In contrast, our patient’s germline variant in MSH2 has been previously characterized as a benign variant, reported in individuals with colorectal, prostate, and ovarian cancers but not meeting clinical criteria for Lynch or Turcot syndrome or showing strong evidence for causality48,49. Furthermore, the lack of a second hit for MSH2 supports the notion of our patient’s variant as benign. As such, it remains unlikely that our patient’s germline MSH2 variant contributed to de novo tumor formation and/or acquisition of a hypermutator phenotype.
In summary, we describe the first case of a hypermutator phenotype associated with a gliosarcoma, characterized by somatic mutations in known MMR genes and consistent with temozolomide-mediated hypermutagenesis. Based on patterns of the described literature, it is more likely that our patient’s spinal cord gliosarcoma represents malignant progression and metastasis from her previous low-grade brain astrocytoma. As gliosarcoma remains a rare entity within primary malignant CNS tumors, even more so within primary spinal cord tumors, further comprehensive genomic characterization is needed to understand the genetic factors that can lead to malignant transformation from lower grade gliomas, as well as de novo formation of gliosarcomas.
Methods
Patient approval
This study was approved by Yale University’s Human Investigations Committee and Human Research Protection Program. Written informed consent was obtained from the parents of the patient. Specimens from resected tumors were evaluated microscopically by a board-certified neuropathologist.
WES and analysis
Genomic DNA from the resected tumor specimen and blood were isolated, and exome capture was performed with IDT xGen Exome Research Panel v1 with the additional spike-in regions of ~620 kb of RefGene coding regions. Sequencing of the captured regions was performed at Yale Center for Genome Analysis (YCGA) using Illumina NovaSeq6000 with 2 × 100 bp reads. Mean coverage of 152.4x and 230.6x was achieved for normal and tumor, respectively. Pre-processing of the raw data, including alignment, PCR duplicate identification, re-alignment around INDELs, and base quality score recalibration was performed “GATK Best Practices” guidelines (GATK v4.1.8.1). Somatic variant calling for SNV/INDELs together with CNV analysis and annotation was performed as previously described in reports from our institution50. Microsatellite status was determined with MSIsensor, a publically available computational algorithm designed to predict microsatellite stability based on WES data13.
Reporting summary
Further information on research design is available in the Nature Research Reporting Summary linked to this article.
Supplementary information
REPORTING SUMMARY
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Supplementary information
The online version contains supplementary material available at 10.1038/s41698-021-00143-w.
Author contributions
C.S.H., G.A.K., D.M., N.A.B., M.L.D., and E.Z.E. drafted the manuscript. C.S.H., G.A.K., A.J.K., A.A.E., A.B.K., D.M., N.A.B., and M.L.D. were involved in the clinical care of the patient and provided relevant material. C.S.H., E.Z.E., D.M., M.L.D., and E.Z.E. analyzed the data. All authors critically reviewed and revised the manuscript, including final approval of the completed version. C.S.H., M.L.D., and E.Z.E. conceived and supervised the study.
Data availability
The data generated and/or analyzed during the related study are described in the figshare metadata record: 10.6084/m9.figshare.1334755751. The whole-exome sequencing data of the resected spinal tumor are shared openly in the European Genotype-phenotype Archive under accession https://identifiers.org/ega.study:EGAS0000100486452.
Competing interests
The authors declare no competing interests. | TEMOZOLOMIDE | DrugsGivenReaction | CC BY | 33580181 | 18,945,561 | 2021-02-12 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Disease progression'. | Impressive response to dabrafenib, trametinib, and osimertinib in a metastatic EGFR-mutant/BRAF V600E lung adenocarcinoma patient.
The survival outcomes of the FLAURA trial support osimertinib as the new standard of care for untreated patients harboring activating mutations in the epidermal growth factor receptor (EGFR). Despite the initial response, disease progression invariably occurs. Although uncommon, BRAF V600E mutation arises as a unique mechanism of resistance, and thus far, no prospective studies are available to support concurrent EGFR/BRAF blockade. We report a case of impressive radiological and ctDNA response under dabrafenib, trametinib, and osimertinib in an advanced EGFR-mutant lung adenocarcinoma patient who developed BRAF V600E as one of the acquired resistance mechanisms to second-line osimertinib. Moreover, the patient experienced remarkable clinical improvement and good tolerance to combination therapy. The present case suggests the importance of prospective studies evaluating both efficacy and safety of the combination in later line settings and points towards the potential of ctDNA to monitor resistance mechanisms and treatment benefit in clinical practice.
Introduction
The use of osimertinib to target epidermal growth factor receptor (EGFR) has become the standard of care in untreated EGFR-mutant non–small cell lung cancer (NSCLC) patients. Although osimertinib can be highly active, showing more durable outcomes than first-generation tyrosine kinase inhibitors (TKI)1, most tumors invariably become resistant, limiting its long-term clinical benefit. The heterogeneity of resistance mechanisms to osimertinib, including EGFR C797S mutation, EGFR, and MET amplifications, off-target mutations in PIK3CA, KRAS, and HER2 as well as histologic transformation2–9, has stimulated routine performance of repeated biopsies to identify specific underlying mechanisms of resistance throughout the treatment course, and to guide the development of novel therapeutic strategies to overcome and prevent acquired resistance (AR)7,10–13.
Combined targeted therapy (TT) strategies have been increasingly addressed in prospective clinical trials14–17. A phase Ib/II trial reported 47% ORR with a combination of capmatinib (MET inhibitor) plus gefitinib in patients progressing to EGFR TKI and presenting MET dysregulation14. Likewise, SAVANNAH (NCT03778229)15 is an ongoing phase II trial designed to evaluate the efficacy of combination savolitinib and osimertinib in post-osimertinib progression/MET-positive patients. The biomarker-matched study ORCHARD (NCT03944772)16 is also underway to assess the efficacy of several osimertinib-based combinations following disease progression under frontline osimertinib.
BRAF mutations and fusions (i.e. AGK-BRAF, ESYT2-BRAF) have recently emerged as additional mechanisms of AR to third-generation EGFR TKI2,18–20. Studies demonstrating the efficacy of concurrent inhibition of EGFR and BRAF3 or MEK21 in pre-clinical models have raised clinicians’ expectations about overcoming AR by combining TT. Nevertheless, reports of successful combinations of TT for patients harboring BRAF-driven AR to osimertinib are very limited19,22–24 and no prospective data regarding efficacy and safety of BRAF/MEK/EGFR concurrent inhibition are available, with chemotherapy-based regimens remaining the treatment of choice in this unfavorable scenario. Similarly, PIK3CA mutations may also mediate AR to second-line osimertinib in 4–11%, but no clinical reports suggesting potential benefits of blocking these alterations in NSCLC are available6,9,25–27.
The detection of circulating tumor DNA (ctDNA) using liquid biopsies allows noninvasive real-time monitoring of treatment response and early detection of AR to TT, anticipating radiological response and treatment failure28,29. The widespread availability of highly specific and sensitive techniques to quantify ctDNA makes the longitudinal assessment of patients with NSCLC under TKI therapy very attractive. Nonetheless, for NSCLCs, ctDNA evaluation in routine clinical practice is currently limited to the detection of EGFR exon 19 deletion, EGFR L858R and T790M activating mutations29.
A man with metastatic lung adenocarcinoma harboring an EGFR mutation, who had progressed to erlotinib due to the emergence of the EGFR T790M mutation started osimertinib and remained on treatment for 15 months, then developing disease progression (PD). Sequential liquid biopsies were collected to monitor treatment response and disclosed the emergence of BRAF V600E and PIK3CA E545K resistance mutations 4 months before clinical progression. With this result and considering few available reports in the literature, our patient started a triple therapy with osimertinib, dabrafenib (BRAF inhibitor), and trametinib (MEK inhibitor).
Results
Case report
A 50-year-old non-smoker man was diagnosed with a tubule-papillary lung adenocarcinoma metastatic to bones and soft tissue (stage IVB - AJCC 8th edition) in July 2016. Several hypermetabolic bone lesions were observed on staging 18-fluorodeoxyglucose positron-emission tomography (18F-FDG PET-CT) scan at diagnosis. A baseline Next Generation Sequencing (NGS) assay (TruSightTumor™ - Illumina®) of a soft tissue metastasis revealed the presence of an activating EGFR exon 19 deletion (EGFR E746_A750del). No concurrent alterations in BRAF or in PIK3CA were identified at that time. ALK d5f3 immunohistochemistry and fluorescent in situ hybridization (FISH) for ROS1 and RET resulted negative.
In September 2016, the patient started on erlotinib 150 mg once daily (OD), achieving complete metabolic response in February 2017. We started to perform serial blood sample collections for ctDNA analysis in May 2017. In October 2017, oligo-progression (oligo-PD) was observed at the primary site and treated with stereotactic body radiation therapy (SBRT; 3 × 16 Gy). In November 2017, we identified the EGFR T790M and EGFR E746_A750del mutations in patient’s plasma using droplet digital PCR (ddPCR) and these results were confirmed in December 2017 with the Foundation ACT® ctDNA assay. Figure 1a shows all systemic and focal therapies of this patient since diagnosis.Fig. 1 Paired radiological and sequential blood-based ctDNA assessments throughout patient’s treatment demonstrating concordant results.
a Timeline displaying systemic and focal therapies since September 2016. b ctDNA levels in serial plasma samples based on fractional abundance of EGFR T790M, EGFR del19 (E746_A750 del), BRAF V600E, and PIK3CA E545K mutations. #2.4 CN results: EGFR del19 WT/RNAseP copy number ratio. c Paired 18F-FDG PET-CT scan maximum intensity projection (MIP) images displaying tumor burden variations. PD progression of disease, SBRT stereotatic body radiotherapy, Pembro pembrolizumab, CT chemotherapy, DTO dabrafenib, trametinib and osimertinib.
Erlotinib was kept until January 2018 when disease progression in the bones was observed and the fractional abundance (FA) of EGFR E746_A750del and EGFR T790M were 12.0 and 2.3%, respectively (Fig. 1b). The treatment was promptly switched to osimertinib 80 mg OD. Bone partial response (PR) was observed 2 months later, along with a significant drop in the FA of EGFR E746_A750del and EGFR T790M mutations in the plasma (Fig. 1b).
In July 2018, the patient developed oligo-PD in T3 vertebrae, which was treated with SBRT (1 × 20 Gy). At that point, ctDNA analysis revealed an increase in the FA of the EGFR E746_A750del in the plasma, but not in EGFR T790M, suggesting genetic heterogeneity between metastatic lesions. In December 2018, a new oligo-PD in the left iliac bone was detected by 18F-FDG PET-CT scan (Fig. 1c). At that time, ctDNA analysis revealed the emergence of BRAF V600E (FA: 0.4%) and PIK3CA E545K (FA: 0.9%) mutations, as well as a significant increase in the FA of the EGFR E746_A750del (FA: 7.5%) in the plasma. These mutations were also detected in a specimen from an iliac bone biopsy using a NGS assay (TruSightTumor™ - Illumina® - EGFR del19 allelic fraction 81.1%, EGFR del19 amplification – 12 copies, BRAF V600E allelic fraction 17.7% and PIK3CA E545K allelic fraction 32.7%). Even though the patient was treated with SBRT (1 × 18 Gy), he developed new bone metastases in March 2019 (Fig. 1b), consistent with the significant increase in the FA of EGFR E746_A750del, BRAF V600E and EGFR T790M mutations (Fig. 1b). At that specific timepoint and in April 2019, assessments of PIK3CA E545K plasma levels were not obtained due to limited amount of cfDNA.
Between April and September 2019, systemic treatment with carboplatin plus pemetrexed and pembrolizumab provided modest clinical benefit, reducing tumor burden and controlling the disease (Fig. 1b). Due to a new PD observed in October 2019, the treatment was switched to docetaxel monotherapy and, subsequently, to vinorelbine after progression under docetaxel; nonetheless, the patient experienced symptomatic PD in bones, lymph nodes, and lung in late January 2020 (Fig. 1c, 2a–d). He came to the clinic with a Karnofsky performance status of 70%, complaining of fatigue, appetite loss, and severe pain in the hips (despite regular use of 10 mg buprenorphine patch and oxycodone plus acetaminophen PO), which had been preventing him from performing his daily activities due to the inability to stay seated. At that point, high levels of all three resistance mutations were detected in the circulating DNA (PIK3CA E545K FA: 13.2%, BRAF V600E FA: 12.3%, and EGFR T790M FA: 5.3%; Fig. 1b). We also observed an exponential increase in the number of copies of EGFR E746_A750del, suggesting gene amplification (Fig. 1b).Fig. 2 18F-FDG PET-CT scan imaging depicting impressive overall tumor response under dabrafenib, trametinib, and osimertinib in three timepoints.
a–d baseline imaging (January/2020) showing: hypermetabolic spiculated mass (orange circle) in the left-superior lobe measuring 4.4 × 3.3 cm (SUVmax: 9.4); multiple hilar, mediastinal, retroperitoneal, and iliac hypermetabolic lymph nodes (red arrow) measuring up to 2.7 cm (SUVmax: 12.8); several hypermetabolic bone lesions throughout axial and appendicular skeleton (yellow circle); a left iliac bone lesion with signs of periosteal reaction and adjacent soft-tissue infiltration (SUVmax: 11.4). e–h first response evaluation imaging (April/2020) showing considerable partial response in the lung mass, measuring 3.8 × 2.5 cm (SUVmax: 2.7), as well as in several bone lesions, especially in the left-iliac bone (SUVmax: 4.8); complete response in lymph nodes. i–l third response evaluation imaging showing disease progression in the left ischium (SUVmax: 12.5; previous SUVmax: 5.9) (October/2020). SUVmax maximum standard uptake value.
In February 2020, after careful consideration due to patient’s good performance status, normal organ functions, severe pain, motivation and the emergence of BRAF V600E as an AR mechanism, he started on dabrafenib 75 mg twice daily (BID), trametinib 1 mg OD and osimertinib 80 mg OD. The use of a PI3K inhibitor was not considered an option due to the absence of reports in the literature suggesting efficacy of these drugs combined with osimertinib to treat advanced NSCLC patients. Within 2 weeks of treatment, the patient achieved complete resolution of the severe bone pain in the hips with no further need of opioid administration, as well as appetite gain and marked improvement in quality of life, which turned possible for him to resume his daily activities. As adverse events (AE), he experienced grade 1 fatigue, dysgeusia, fever, and nausea, all managed with symptomatic medication. Complete resolution of the fever occurred spontaneously within two weeks of treatment. An attempt to increase dosages of dabrafenib to 150 mg BID and trametinib 2 mg OD resulted unsuccessful due to persistent grade 2 fatigue. In April 2020, a new 18F-FDG PET-CT scan disclosed a complete response in lymph nodes and a dramatic PR in the lung and bones (PERCIST 1.1 evaluating 5 target lesions: Δ- 67%; Figs. 1c and 2e–h). Detection of EGFR T790M, BRAF V600E, and PIK3CA E545K mutations became negative in April 2020, while EGFR E746_A750del despite marginally positive, presented marked reduction (Fig. 1b) suggesting that combination dabrafenib, trametinib, and osimertinib might be an effective strategy to overcome PIK3CA E545K and BRAF V600E-driven resistance to osimertinib in advanced EGFR-mutant NSCLC patients. The patient remained in response until October 2020, when he developed asymptomatic bone PD in lumbar spine, left ischium, and right iliac bone, 8 months after starting this combination therapy (Figs. 1c and 2i–l). A FoundationOne®Liquid CDx plasma NGS disclosed the following alterations: CHCHD3-BRAF fusion, BRAF V600E, EGFR E746_750del, EGFR amplification, PIK3CA E545K, MAP2K2 (MEK2) C125S, MTAP rearrangement intron 5, TP53 V197M, and TP53 S241A. Owing to the considerable clinical benefit, good tolerance and lack of systemic treatment options, we decided to keep the patient on treatment and increase the doses of dabrafenib (150 mg BID) and trametinib (2 mg OD) every other day. He remains asymptomatic and tolerating well the proposed dose adjustment.
Discussion
Several studies have recently highlighted the importance of considering genes of interest within the context of commonly co-occurring mutations9,30. For example, as described by Blakely et al. through performing a cfDNA NGS analysis of 1,122 advanced stage EGFR-mutant NSCLCs, in ~93% of the patients, at least one more variables with known or likely known functional properties were present, disclosing the molecular complexity of this oncogenic driver and suggesting an association of co-occuring genomic alterations with TKI response and clinical outcomes9. In addition, Roper and colleagues reported the identification of at least two co-existing AR mechanisms in 73% of patients treated with osimertinib, as well as 6–23 different subclones per individual in a phylogenetic analysis performed in multiple metastatic sites of 15 individuals30. In the same publication, the authors also called the attention for a high incidence of acquired EGFR amplifications in post-osimertinib patients, which suggests maintenance of EGFR central role in the setting of progression, as previously reported by our group28 and also detected in the present case.
Although uncommon, BRAF V600E mediates AR in approximately 3% of the patients under second-line osimertinib6,20 and little is known about the efficacy of combined TT in this population3,21–23. Existing reports lack details regarding objective responses using standardized radiological criteria (i.e., RECIST 1.1; PERCIST) and the dynamics of resistance mechanisms through longitudinal ctDNA measurements22,23. Huang and colleagues22 described a case of an EGFRdel19/T790M + NSCLC patient who developed BRAF V600E-driven AR after second-line osimertinib and achieved stable disease under dabrafenib 150 mg BID, trametinib 1 mg OD and osimertinib 80 mg OD, with ongoing disease control 7.4 months after. Zhou and colleagues23 also reported their experience with dabrafenib 150 mg BID, trametinib 2 mg OD and osimertinib 80 mg OD leading to tumor reduction within 6 weeks of treatment, along with grade 2 rash and decreased appetite as AEs. Similarly, Meng et al. reported two cases treated with this triple regimen31. The first patient discontinued therapy after one month due to severe pneumonitis; the second one presented tumor response under dabrafenib 50 mg BID, trametinib 0.5 mg OD, and osimertinib 80 mg OD, with progression-free survival of 14 months. According to the authors, this reduced dose was prescribed owing to a grade 2 pyrexia, nausea, and vomiting under higher dabrafenib and trametinib doses. Dagogo-Jack and colleagues19 also described a successful case of combined EGFR/MAP kinase pathway blockade with osimertinib 80 mg OD and trametinib 1 mg OD; as treatment-related AEs, their patient experienced grade 2 diarrhea and fatigue, along with grade 1 rash and gastrointestinal bleeding. Nevertheless, it is worth highlighting the rapid clinical improvement, the remarkable radiologically confirmed objective response, as well as the good tolerance observed in this case even using only half standard dose of dabrafenib and trametinib approved for NSCLCs harboring BRAF V600E mutations. Since data regarding the efficacy of these combined approaches, the optimal drug association and dosing, as well as the toxicity profile are conflicting and largely unknown, further investigation into the mechanistic basis of this association represents an important priority. The Table 1 summarizes the toxicities arising under osimertinib plus BRAF/MEK inhibition and reported in the literature.Table 1 Summary of the toxicities arising under osimertinib plus BRAF/MEK inhibition and reported in the literature.
Reference Treatment Anorexia Nausea Vomiting Diarrhoea Fatigue Rash GI bleeding AST/ALT elevation Paroniquia Pyrexia Initial dose Dose reduction
Meng et al. D + T + O NR G2 G2 G2 NR NR NR NR NR G2 D (150 mg BID) + T (2 mg OD) + O (80 mg OD) D (50 mg BID) + T (0.5 mg/day) + O (80 mg/day)
Dagogo-Jack et al. T + O NR NR NR G2 G2 G1 G1 G1 NR NR T (1 mg OD) + O (80 mg OD) Not needed
Huang et al. D + T + O NR NR NR G1 NR NR NR NR G1 NR D (150 mg BID) + T (1 mg OD) + O (80 mg OD) Not needed
Zhou et al. D + T + O G1 NR NR NR NR G2 NR NR NR NR D (150 mg BID) + T (2 mg OD) + O (80 mg OD) Not needed
D dabrafenib, T trametinib, O osimertinib, NR not reported.
Liquid biopsy is emerging as an important diagnostic and predictive tool in the treatment of NSCLCs. The accurate identification of predictive genetic alterations is important for both patients’ management and the understanding of clonal evolution and AR to different therapies9,29,32. Also, circulating biomarkers from multiple disease sites better reflect systemic tumor burden, including alterations from genetically different metastatic lesions, which may be missed with single-site tissue biopsies8. Here, we were able to identify and monitor multiple concurrent mechanisms of resistance throughout the entire patient’s treatment. As shown in Fig. 1b, variations in blood ctDNA levels exhibited a positive correlation with imaging findings, even in situations involving focal treatments for oligo-PD. The observed concordance between radiological PD or response and ctDNA measurements also points towards its great potential to be incorporated into clinical practice to anticipate radiologic findings in a more effective manner.
Along with BRAF V600E mutation, the presence of PIK3CA E545K mutation is also associated with AR to osimertinib6,9,25,27. However, the clinical implication of concomitant targeting of EGFR and PI3K remains unclear. Whereas an alternative argument for the observed response could be a rechallenge following a long time upon off-osimertinib, in our opinion, this is an unlikely explanation33. Interestingly, the presence of a BRAF fusion along with a MEK2 mutation identified through plasma NGS in the setting of PD might represent a mechanism of resistance to this triple regimen34. Nonetheless, since pretreatment blood samples had not been analyzed using a similar methodology, this hypothesis deserves careful interpretation.
Increased understanding of the relationship of concurring genomic alterations in EGFR-mutant NSCLC may enable new therapeutic opportunities following disease progression to osimertinib. Here, we reported an impressive objective response to dabrafenib, trametinib, and osimertinib with concordant decrease in plasma ctDNA levels in a metastatic lung cancer patient harboring EGFR E746_A750del, BRAF V600E, and PIK3CA E545K activating mutations. This case report leads to a greater understanding of the currently limited literature regarding the management of EGFR-mutant NSCLC patients with acquired BRAFV600E mutation, since it reports a successful attempt to target both alterations concurrently while providing concordant and interesting data of serial ctDNA assessments throughout the entire treatment. Further investigation to optimize the efficacy and mitigate the toxicity profile of this drug association represents an important issue.
Methods
Patient
This study was approved by Hospital Sírio-Libanês Ethics Committee (HSL-RC 2020-16). The patient provided written informed consent for blood collection, ctDNA analysis and publication of this report.
Sample collection and plasma DNA extraction
Serial blood samples were collected between May 2017 and April 2020 (Fig. 1b). Peripheral blood (20 ml) was collected into tubes containing EDTA (BD, Franklin Lakes, New Jersey). Plasma was separated from the blood within 2 hours of blood collection, as previously described28. cfDNA was extracted using QIAamp MinElute Virus Vacuum Kit (Qiagen, Hilden, Germany) and stored at −80 °C.
ctDNA-ddPCR
Cell-free DNA (cfDNA) was quantified using the RNase P Copy Number Reference Assay (Life Technologies, Carlsbad, California). A total of 3000 genome-equivalents (~10 ng of cfDNA) were analyzed per assay to achieve a detection sensitivity of 0.2%. This detection limit has been assessed by using cell line-derived genomic DNA. A total of 10 ng of input DNA with varying proportions of mutant DNA was serially diluted into wild-type DNA to obtain samples with a mutant abundance of 1%, 0.5%, 0.1%, and 0.05% and subjected to droplet digital PCR (ddPCR). ddPCR was used to quantify the circulating levels of the EGFR activating mutation (EGFR E746_A750 del) and of the resistance mutations (EGFR T790M, PIK3CA E545K, and BRAF V600E). Probes and primers were obtained from BioRad (EGFR E746_A750 del #10041170, EGFR T790M #10040782, PIK3CA E545K #10041188, and BRAF V600E #10040779; Hercules, California). ddPCR was performed on the QX200 Droplet Digital PCR System, and data were analysed using QuantaSoft software (Bio-Rad). ctDNA quantification is presented as fractional abundance (FA–the proportion of the mutant allele in total cfDNA).
Reporting summary
Further information on research design is available in the Nature Research Reporting Summary linked to this article.
Supplementary information
Reporting Summary
Dataset 1
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
These authors contributed equally: Maurício Fernando Silva Almeida Ribeiro, Franciele Hinterholz Knebel.
Supplementary information
The online version contains supplementary material available at 10.1038/s41698-021-00149-4.
Acknowledgements
We thank the clinical research team at Hospital Sírio-Libanês for their assistance collecting samples. We thank Ernande Xavier dos Santos and Elisângela Monteiro for the outstanding technical support. We also thank Mrs. Dina Binzagr and Vivian Hannud for supporting the Translational Research Program at Hospital Sírio Libanês. This work was supported by Ludwig Cancer Research and Fundação de Amparo à Pesquisa do Estado de São Paulo – FAPESP. Lastly, we would like to thank the patient and his family for consenting to the present publication.
Author contributions
Conceptualization by M.F.S.A.R., F.H.K., J.V.M.A., A.A.C., and A.K.; Methodology by F.H.K. and A.A.C.; Resources br M.F.S.A.R., F.H.K., A.A.C., and J.V.M.A.; Writing of the first draft by M.F.S.A., F.H.K., and J.V.M.A.; Review, editing and approval of the final draft by all authors; Supervision by A.A.C. and A.K.; Funding acquisition by F.H.K. and A.A.C. M.F.S.A.R. and F.H.K. are co-first authors of the present work.
Data availability
The datasets that support the findings of this study are not publicly available in order to protect patient privacy. The data will be made available on reasonable request. For data access requests regarding the liquid biopsy (ctDNA quantification) data, please contact Dr. Franciele Knebel, email address: fhknebel@mochsl.org.br. For data access requests regarding the PET-CT high resolution images and PERCIST calculations, please contact Dr. José Marin, email address:jfgmarin@yahoo.com.br. For data access requests regarding the summary of the toxicities arising under osimertinib plus BRAF/MEK inhibition, please contact the corresponding author Dr. Maurício Ribeiro, email address: mauricio.fsaribeiro@hsl.org.br. The data generated and analysed during this study are described in the following metadata record: 10.6084/m9.figshare.1347594635.
Competing interests
The authors declare no competing interests. | ACETAMINOPHEN\OXYCODONE HYDROCHLORIDE, BUPRENORPHINE, DOCETAXEL | DrugsGivenReaction | CC BY | 33580193 | 19,460,717 | 2021-02-12 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Drug ineffective'. | Impressive response to dabrafenib, trametinib, and osimertinib in a metastatic EGFR-mutant/BRAF V600E lung adenocarcinoma patient.
The survival outcomes of the FLAURA trial support osimertinib as the new standard of care for untreated patients harboring activating mutations in the epidermal growth factor receptor (EGFR). Despite the initial response, disease progression invariably occurs. Although uncommon, BRAF V600E mutation arises as a unique mechanism of resistance, and thus far, no prospective studies are available to support concurrent EGFR/BRAF blockade. We report a case of impressive radiological and ctDNA response under dabrafenib, trametinib, and osimertinib in an advanced EGFR-mutant lung adenocarcinoma patient who developed BRAF V600E as one of the acquired resistance mechanisms to second-line osimertinib. Moreover, the patient experienced remarkable clinical improvement and good tolerance to combination therapy. The present case suggests the importance of prospective studies evaluating both efficacy and safety of the combination in later line settings and points towards the potential of ctDNA to monitor resistance mechanisms and treatment benefit in clinical practice.
Introduction
The use of osimertinib to target epidermal growth factor receptor (EGFR) has become the standard of care in untreated EGFR-mutant non–small cell lung cancer (NSCLC) patients. Although osimertinib can be highly active, showing more durable outcomes than first-generation tyrosine kinase inhibitors (TKI)1, most tumors invariably become resistant, limiting its long-term clinical benefit. The heterogeneity of resistance mechanisms to osimertinib, including EGFR C797S mutation, EGFR, and MET amplifications, off-target mutations in PIK3CA, KRAS, and HER2 as well as histologic transformation2–9, has stimulated routine performance of repeated biopsies to identify specific underlying mechanisms of resistance throughout the treatment course, and to guide the development of novel therapeutic strategies to overcome and prevent acquired resistance (AR)7,10–13.
Combined targeted therapy (TT) strategies have been increasingly addressed in prospective clinical trials14–17. A phase Ib/II trial reported 47% ORR with a combination of capmatinib (MET inhibitor) plus gefitinib in patients progressing to EGFR TKI and presenting MET dysregulation14. Likewise, SAVANNAH (NCT03778229)15 is an ongoing phase II trial designed to evaluate the efficacy of combination savolitinib and osimertinib in post-osimertinib progression/MET-positive patients. The biomarker-matched study ORCHARD (NCT03944772)16 is also underway to assess the efficacy of several osimertinib-based combinations following disease progression under frontline osimertinib.
BRAF mutations and fusions (i.e. AGK-BRAF, ESYT2-BRAF) have recently emerged as additional mechanisms of AR to third-generation EGFR TKI2,18–20. Studies demonstrating the efficacy of concurrent inhibition of EGFR and BRAF3 or MEK21 in pre-clinical models have raised clinicians’ expectations about overcoming AR by combining TT. Nevertheless, reports of successful combinations of TT for patients harboring BRAF-driven AR to osimertinib are very limited19,22–24 and no prospective data regarding efficacy and safety of BRAF/MEK/EGFR concurrent inhibition are available, with chemotherapy-based regimens remaining the treatment of choice in this unfavorable scenario. Similarly, PIK3CA mutations may also mediate AR to second-line osimertinib in 4–11%, but no clinical reports suggesting potential benefits of blocking these alterations in NSCLC are available6,9,25–27.
The detection of circulating tumor DNA (ctDNA) using liquid biopsies allows noninvasive real-time monitoring of treatment response and early detection of AR to TT, anticipating radiological response and treatment failure28,29. The widespread availability of highly specific and sensitive techniques to quantify ctDNA makes the longitudinal assessment of patients with NSCLC under TKI therapy very attractive. Nonetheless, for NSCLCs, ctDNA evaluation in routine clinical practice is currently limited to the detection of EGFR exon 19 deletion, EGFR L858R and T790M activating mutations29.
A man with metastatic lung adenocarcinoma harboring an EGFR mutation, who had progressed to erlotinib due to the emergence of the EGFR T790M mutation started osimertinib and remained on treatment for 15 months, then developing disease progression (PD). Sequential liquid biopsies were collected to monitor treatment response and disclosed the emergence of BRAF V600E and PIK3CA E545K resistance mutations 4 months before clinical progression. With this result and considering few available reports in the literature, our patient started a triple therapy with osimertinib, dabrafenib (BRAF inhibitor), and trametinib (MEK inhibitor).
Results
Case report
A 50-year-old non-smoker man was diagnosed with a tubule-papillary lung adenocarcinoma metastatic to bones and soft tissue (stage IVB - AJCC 8th edition) in July 2016. Several hypermetabolic bone lesions were observed on staging 18-fluorodeoxyglucose positron-emission tomography (18F-FDG PET-CT) scan at diagnosis. A baseline Next Generation Sequencing (NGS) assay (TruSightTumor™ - Illumina®) of a soft tissue metastasis revealed the presence of an activating EGFR exon 19 deletion (EGFR E746_A750del). No concurrent alterations in BRAF or in PIK3CA were identified at that time. ALK d5f3 immunohistochemistry and fluorescent in situ hybridization (FISH) for ROS1 and RET resulted negative.
In September 2016, the patient started on erlotinib 150 mg once daily (OD), achieving complete metabolic response in February 2017. We started to perform serial blood sample collections for ctDNA analysis in May 2017. In October 2017, oligo-progression (oligo-PD) was observed at the primary site and treated with stereotactic body radiation therapy (SBRT; 3 × 16 Gy). In November 2017, we identified the EGFR T790M and EGFR E746_A750del mutations in patient’s plasma using droplet digital PCR (ddPCR) and these results were confirmed in December 2017 with the Foundation ACT® ctDNA assay. Figure 1a shows all systemic and focal therapies of this patient since diagnosis.Fig. 1 Paired radiological and sequential blood-based ctDNA assessments throughout patient’s treatment demonstrating concordant results.
a Timeline displaying systemic and focal therapies since September 2016. b ctDNA levels in serial plasma samples based on fractional abundance of EGFR T790M, EGFR del19 (E746_A750 del), BRAF V600E, and PIK3CA E545K mutations. #2.4 CN results: EGFR del19 WT/RNAseP copy number ratio. c Paired 18F-FDG PET-CT scan maximum intensity projection (MIP) images displaying tumor burden variations. PD progression of disease, SBRT stereotatic body radiotherapy, Pembro pembrolizumab, CT chemotherapy, DTO dabrafenib, trametinib and osimertinib.
Erlotinib was kept until January 2018 when disease progression in the bones was observed and the fractional abundance (FA) of EGFR E746_A750del and EGFR T790M were 12.0 and 2.3%, respectively (Fig. 1b). The treatment was promptly switched to osimertinib 80 mg OD. Bone partial response (PR) was observed 2 months later, along with a significant drop in the FA of EGFR E746_A750del and EGFR T790M mutations in the plasma (Fig. 1b).
In July 2018, the patient developed oligo-PD in T3 vertebrae, which was treated with SBRT (1 × 20 Gy). At that point, ctDNA analysis revealed an increase in the FA of the EGFR E746_A750del in the plasma, but not in EGFR T790M, suggesting genetic heterogeneity between metastatic lesions. In December 2018, a new oligo-PD in the left iliac bone was detected by 18F-FDG PET-CT scan (Fig. 1c). At that time, ctDNA analysis revealed the emergence of BRAF V600E (FA: 0.4%) and PIK3CA E545K (FA: 0.9%) mutations, as well as a significant increase in the FA of the EGFR E746_A750del (FA: 7.5%) in the plasma. These mutations were also detected in a specimen from an iliac bone biopsy using a NGS assay (TruSightTumor™ - Illumina® - EGFR del19 allelic fraction 81.1%, EGFR del19 amplification – 12 copies, BRAF V600E allelic fraction 17.7% and PIK3CA E545K allelic fraction 32.7%). Even though the patient was treated with SBRT (1 × 18 Gy), he developed new bone metastases in March 2019 (Fig. 1b), consistent with the significant increase in the FA of EGFR E746_A750del, BRAF V600E and EGFR T790M mutations (Fig. 1b). At that specific timepoint and in April 2019, assessments of PIK3CA E545K plasma levels were not obtained due to limited amount of cfDNA.
Between April and September 2019, systemic treatment with carboplatin plus pemetrexed and pembrolizumab provided modest clinical benefit, reducing tumor burden and controlling the disease (Fig. 1b). Due to a new PD observed in October 2019, the treatment was switched to docetaxel monotherapy and, subsequently, to vinorelbine after progression under docetaxel; nonetheless, the patient experienced symptomatic PD in bones, lymph nodes, and lung in late January 2020 (Fig. 1c, 2a–d). He came to the clinic with a Karnofsky performance status of 70%, complaining of fatigue, appetite loss, and severe pain in the hips (despite regular use of 10 mg buprenorphine patch and oxycodone plus acetaminophen PO), which had been preventing him from performing his daily activities due to the inability to stay seated. At that point, high levels of all three resistance mutations were detected in the circulating DNA (PIK3CA E545K FA: 13.2%, BRAF V600E FA: 12.3%, and EGFR T790M FA: 5.3%; Fig. 1b). We also observed an exponential increase in the number of copies of EGFR E746_A750del, suggesting gene amplification (Fig. 1b).Fig. 2 18F-FDG PET-CT scan imaging depicting impressive overall tumor response under dabrafenib, trametinib, and osimertinib in three timepoints.
a–d baseline imaging (January/2020) showing: hypermetabolic spiculated mass (orange circle) in the left-superior lobe measuring 4.4 × 3.3 cm (SUVmax: 9.4); multiple hilar, mediastinal, retroperitoneal, and iliac hypermetabolic lymph nodes (red arrow) measuring up to 2.7 cm (SUVmax: 12.8); several hypermetabolic bone lesions throughout axial and appendicular skeleton (yellow circle); a left iliac bone lesion with signs of periosteal reaction and adjacent soft-tissue infiltration (SUVmax: 11.4). e–h first response evaluation imaging (April/2020) showing considerable partial response in the lung mass, measuring 3.8 × 2.5 cm (SUVmax: 2.7), as well as in several bone lesions, especially in the left-iliac bone (SUVmax: 4.8); complete response in lymph nodes. i–l third response evaluation imaging showing disease progression in the left ischium (SUVmax: 12.5; previous SUVmax: 5.9) (October/2020). SUVmax maximum standard uptake value.
In February 2020, after careful consideration due to patient’s good performance status, normal organ functions, severe pain, motivation and the emergence of BRAF V600E as an AR mechanism, he started on dabrafenib 75 mg twice daily (BID), trametinib 1 mg OD and osimertinib 80 mg OD. The use of a PI3K inhibitor was not considered an option due to the absence of reports in the literature suggesting efficacy of these drugs combined with osimertinib to treat advanced NSCLC patients. Within 2 weeks of treatment, the patient achieved complete resolution of the severe bone pain in the hips with no further need of opioid administration, as well as appetite gain and marked improvement in quality of life, which turned possible for him to resume his daily activities. As adverse events (AE), he experienced grade 1 fatigue, dysgeusia, fever, and nausea, all managed with symptomatic medication. Complete resolution of the fever occurred spontaneously within two weeks of treatment. An attempt to increase dosages of dabrafenib to 150 mg BID and trametinib 2 mg OD resulted unsuccessful due to persistent grade 2 fatigue. In April 2020, a new 18F-FDG PET-CT scan disclosed a complete response in lymph nodes and a dramatic PR in the lung and bones (PERCIST 1.1 evaluating 5 target lesions: Δ- 67%; Figs. 1c and 2e–h). Detection of EGFR T790M, BRAF V600E, and PIK3CA E545K mutations became negative in April 2020, while EGFR E746_A750del despite marginally positive, presented marked reduction (Fig. 1b) suggesting that combination dabrafenib, trametinib, and osimertinib might be an effective strategy to overcome PIK3CA E545K and BRAF V600E-driven resistance to osimertinib in advanced EGFR-mutant NSCLC patients. The patient remained in response until October 2020, when he developed asymptomatic bone PD in lumbar spine, left ischium, and right iliac bone, 8 months after starting this combination therapy (Figs. 1c and 2i–l). A FoundationOne®Liquid CDx plasma NGS disclosed the following alterations: CHCHD3-BRAF fusion, BRAF V600E, EGFR E746_750del, EGFR amplification, PIK3CA E545K, MAP2K2 (MEK2) C125S, MTAP rearrangement intron 5, TP53 V197M, and TP53 S241A. Owing to the considerable clinical benefit, good tolerance and lack of systemic treatment options, we decided to keep the patient on treatment and increase the doses of dabrafenib (150 mg BID) and trametinib (2 mg OD) every other day. He remains asymptomatic and tolerating well the proposed dose adjustment.
Discussion
Several studies have recently highlighted the importance of considering genes of interest within the context of commonly co-occurring mutations9,30. For example, as described by Blakely et al. through performing a cfDNA NGS analysis of 1,122 advanced stage EGFR-mutant NSCLCs, in ~93% of the patients, at least one more variables with known or likely known functional properties were present, disclosing the molecular complexity of this oncogenic driver and suggesting an association of co-occuring genomic alterations with TKI response and clinical outcomes9. In addition, Roper and colleagues reported the identification of at least two co-existing AR mechanisms in 73% of patients treated with osimertinib, as well as 6–23 different subclones per individual in a phylogenetic analysis performed in multiple metastatic sites of 15 individuals30. In the same publication, the authors also called the attention for a high incidence of acquired EGFR amplifications in post-osimertinib patients, which suggests maintenance of EGFR central role in the setting of progression, as previously reported by our group28 and also detected in the present case.
Although uncommon, BRAF V600E mediates AR in approximately 3% of the patients under second-line osimertinib6,20 and little is known about the efficacy of combined TT in this population3,21–23. Existing reports lack details regarding objective responses using standardized radiological criteria (i.e., RECIST 1.1; PERCIST) and the dynamics of resistance mechanisms through longitudinal ctDNA measurements22,23. Huang and colleagues22 described a case of an EGFRdel19/T790M + NSCLC patient who developed BRAF V600E-driven AR after second-line osimertinib and achieved stable disease under dabrafenib 150 mg BID, trametinib 1 mg OD and osimertinib 80 mg OD, with ongoing disease control 7.4 months after. Zhou and colleagues23 also reported their experience with dabrafenib 150 mg BID, trametinib 2 mg OD and osimertinib 80 mg OD leading to tumor reduction within 6 weeks of treatment, along with grade 2 rash and decreased appetite as AEs. Similarly, Meng et al. reported two cases treated with this triple regimen31. The first patient discontinued therapy after one month due to severe pneumonitis; the second one presented tumor response under dabrafenib 50 mg BID, trametinib 0.5 mg OD, and osimertinib 80 mg OD, with progression-free survival of 14 months. According to the authors, this reduced dose was prescribed owing to a grade 2 pyrexia, nausea, and vomiting under higher dabrafenib and trametinib doses. Dagogo-Jack and colleagues19 also described a successful case of combined EGFR/MAP kinase pathway blockade with osimertinib 80 mg OD and trametinib 1 mg OD; as treatment-related AEs, their patient experienced grade 2 diarrhea and fatigue, along with grade 1 rash and gastrointestinal bleeding. Nevertheless, it is worth highlighting the rapid clinical improvement, the remarkable radiologically confirmed objective response, as well as the good tolerance observed in this case even using only half standard dose of dabrafenib and trametinib approved for NSCLCs harboring BRAF V600E mutations. Since data regarding the efficacy of these combined approaches, the optimal drug association and dosing, as well as the toxicity profile are conflicting and largely unknown, further investigation into the mechanistic basis of this association represents an important priority. The Table 1 summarizes the toxicities arising under osimertinib plus BRAF/MEK inhibition and reported in the literature.Table 1 Summary of the toxicities arising under osimertinib plus BRAF/MEK inhibition and reported in the literature.
Reference Treatment Anorexia Nausea Vomiting Diarrhoea Fatigue Rash GI bleeding AST/ALT elevation Paroniquia Pyrexia Initial dose Dose reduction
Meng et al. D + T + O NR G2 G2 G2 NR NR NR NR NR G2 D (150 mg BID) + T (2 mg OD) + O (80 mg OD) D (50 mg BID) + T (0.5 mg/day) + O (80 mg/day)
Dagogo-Jack et al. T + O NR NR NR G2 G2 G1 G1 G1 NR NR T (1 mg OD) + O (80 mg OD) Not needed
Huang et al. D + T + O NR NR NR G1 NR NR NR NR G1 NR D (150 mg BID) + T (1 mg OD) + O (80 mg OD) Not needed
Zhou et al. D + T + O G1 NR NR NR NR G2 NR NR NR NR D (150 mg BID) + T (2 mg OD) + O (80 mg OD) Not needed
D dabrafenib, T trametinib, O osimertinib, NR not reported.
Liquid biopsy is emerging as an important diagnostic and predictive tool in the treatment of NSCLCs. The accurate identification of predictive genetic alterations is important for both patients’ management and the understanding of clonal evolution and AR to different therapies9,29,32. Also, circulating biomarkers from multiple disease sites better reflect systemic tumor burden, including alterations from genetically different metastatic lesions, which may be missed with single-site tissue biopsies8. Here, we were able to identify and monitor multiple concurrent mechanisms of resistance throughout the entire patient’s treatment. As shown in Fig. 1b, variations in blood ctDNA levels exhibited a positive correlation with imaging findings, even in situations involving focal treatments for oligo-PD. The observed concordance between radiological PD or response and ctDNA measurements also points towards its great potential to be incorporated into clinical practice to anticipate radiologic findings in a more effective manner.
Along with BRAF V600E mutation, the presence of PIK3CA E545K mutation is also associated with AR to osimertinib6,9,25,27. However, the clinical implication of concomitant targeting of EGFR and PI3K remains unclear. Whereas an alternative argument for the observed response could be a rechallenge following a long time upon off-osimertinib, in our opinion, this is an unlikely explanation33. Interestingly, the presence of a BRAF fusion along with a MEK2 mutation identified through plasma NGS in the setting of PD might represent a mechanism of resistance to this triple regimen34. Nonetheless, since pretreatment blood samples had not been analyzed using a similar methodology, this hypothesis deserves careful interpretation.
Increased understanding of the relationship of concurring genomic alterations in EGFR-mutant NSCLC may enable new therapeutic opportunities following disease progression to osimertinib. Here, we reported an impressive objective response to dabrafenib, trametinib, and osimertinib with concordant decrease in plasma ctDNA levels in a metastatic lung cancer patient harboring EGFR E746_A750del, BRAF V600E, and PIK3CA E545K activating mutations. This case report leads to a greater understanding of the currently limited literature regarding the management of EGFR-mutant NSCLC patients with acquired BRAFV600E mutation, since it reports a successful attempt to target both alterations concurrently while providing concordant and interesting data of serial ctDNA assessments throughout the entire treatment. Further investigation to optimize the efficacy and mitigate the toxicity profile of this drug association represents an important issue.
Methods
Patient
This study was approved by Hospital Sírio-Libanês Ethics Committee (HSL-RC 2020-16). The patient provided written informed consent for blood collection, ctDNA analysis and publication of this report.
Sample collection and plasma DNA extraction
Serial blood samples were collected between May 2017 and April 2020 (Fig. 1b). Peripheral blood (20 ml) was collected into tubes containing EDTA (BD, Franklin Lakes, New Jersey). Plasma was separated from the blood within 2 hours of blood collection, as previously described28. cfDNA was extracted using QIAamp MinElute Virus Vacuum Kit (Qiagen, Hilden, Germany) and stored at −80 °C.
ctDNA-ddPCR
Cell-free DNA (cfDNA) was quantified using the RNase P Copy Number Reference Assay (Life Technologies, Carlsbad, California). A total of 3000 genome-equivalents (~10 ng of cfDNA) were analyzed per assay to achieve a detection sensitivity of 0.2%. This detection limit has been assessed by using cell line-derived genomic DNA. A total of 10 ng of input DNA with varying proportions of mutant DNA was serially diluted into wild-type DNA to obtain samples with a mutant abundance of 1%, 0.5%, 0.1%, and 0.05% and subjected to droplet digital PCR (ddPCR). ddPCR was used to quantify the circulating levels of the EGFR activating mutation (EGFR E746_A750 del) and of the resistance mutations (EGFR T790M, PIK3CA E545K, and BRAF V600E). Probes and primers were obtained from BioRad (EGFR E746_A750 del #10041170, EGFR T790M #10040782, PIK3CA E545K #10041188, and BRAF V600E #10040779; Hercules, California). ddPCR was performed on the QX200 Droplet Digital PCR System, and data were analysed using QuantaSoft software (Bio-Rad). ctDNA quantification is presented as fractional abundance (FA–the proportion of the mutant allele in total cfDNA).
Reporting summary
Further information on research design is available in the Nature Research Reporting Summary linked to this article.
Supplementary information
Reporting Summary
Dataset 1
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
These authors contributed equally: Maurício Fernando Silva Almeida Ribeiro, Franciele Hinterholz Knebel.
Supplementary information
The online version contains supplementary material available at 10.1038/s41698-021-00149-4.
Acknowledgements
We thank the clinical research team at Hospital Sírio-Libanês for their assistance collecting samples. We thank Ernande Xavier dos Santos and Elisângela Monteiro for the outstanding technical support. We also thank Mrs. Dina Binzagr and Vivian Hannud for supporting the Translational Research Program at Hospital Sírio Libanês. This work was supported by Ludwig Cancer Research and Fundação de Amparo à Pesquisa do Estado de São Paulo – FAPESP. Lastly, we would like to thank the patient and his family for consenting to the present publication.
Author contributions
Conceptualization by M.F.S.A.R., F.H.K., J.V.M.A., A.A.C., and A.K.; Methodology by F.H.K. and A.A.C.; Resources br M.F.S.A.R., F.H.K., A.A.C., and J.V.M.A.; Writing of the first draft by M.F.S.A., F.H.K., and J.V.M.A.; Review, editing and approval of the final draft by all authors; Supervision by A.A.C. and A.K.; Funding acquisition by F.H.K. and A.A.C. M.F.S.A.R. and F.H.K. are co-first authors of the present work.
Data availability
The datasets that support the findings of this study are not publicly available in order to protect patient privacy. The data will be made available on reasonable request. For data access requests regarding the liquid biopsy (ctDNA quantification) data, please contact Dr. Franciele Knebel, email address: fhknebel@mochsl.org.br. For data access requests regarding the PET-CT high resolution images and PERCIST calculations, please contact Dr. José Marin, email address:jfgmarin@yahoo.com.br. For data access requests regarding the summary of the toxicities arising under osimertinib plus BRAF/MEK inhibition, please contact the corresponding author Dr. Maurício Ribeiro, email address: mauricio.fsaribeiro@hsl.org.br. The data generated and analysed during this study are described in the following metadata record: 10.6084/m9.figshare.1347594635.
Competing interests
The authors declare no competing interests. | ACETAMINOPHEN\OXYCODONE HYDROCHLORIDE, BUPRENORPHINE, DOCETAXEL | DrugsGivenReaction | CC BY | 33580193 | 19,460,717 | 2021-02-12 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Dysgeusia'. | Impressive response to dabrafenib, trametinib, and osimertinib in a metastatic EGFR-mutant/BRAF V600E lung adenocarcinoma patient.
The survival outcomes of the FLAURA trial support osimertinib as the new standard of care for untreated patients harboring activating mutations in the epidermal growth factor receptor (EGFR). Despite the initial response, disease progression invariably occurs. Although uncommon, BRAF V600E mutation arises as a unique mechanism of resistance, and thus far, no prospective studies are available to support concurrent EGFR/BRAF blockade. We report a case of impressive radiological and ctDNA response under dabrafenib, trametinib, and osimertinib in an advanced EGFR-mutant lung adenocarcinoma patient who developed BRAF V600E as one of the acquired resistance mechanisms to second-line osimertinib. Moreover, the patient experienced remarkable clinical improvement and good tolerance to combination therapy. The present case suggests the importance of prospective studies evaluating both efficacy and safety of the combination in later line settings and points towards the potential of ctDNA to monitor resistance mechanisms and treatment benefit in clinical practice.
Introduction
The use of osimertinib to target epidermal growth factor receptor (EGFR) has become the standard of care in untreated EGFR-mutant non–small cell lung cancer (NSCLC) patients. Although osimertinib can be highly active, showing more durable outcomes than first-generation tyrosine kinase inhibitors (TKI)1, most tumors invariably become resistant, limiting its long-term clinical benefit. The heterogeneity of resistance mechanisms to osimertinib, including EGFR C797S mutation, EGFR, and MET amplifications, off-target mutations in PIK3CA, KRAS, and HER2 as well as histologic transformation2–9, has stimulated routine performance of repeated biopsies to identify specific underlying mechanisms of resistance throughout the treatment course, and to guide the development of novel therapeutic strategies to overcome and prevent acquired resistance (AR)7,10–13.
Combined targeted therapy (TT) strategies have been increasingly addressed in prospective clinical trials14–17. A phase Ib/II trial reported 47% ORR with a combination of capmatinib (MET inhibitor) plus gefitinib in patients progressing to EGFR TKI and presenting MET dysregulation14. Likewise, SAVANNAH (NCT03778229)15 is an ongoing phase II trial designed to evaluate the efficacy of combination savolitinib and osimertinib in post-osimertinib progression/MET-positive patients. The biomarker-matched study ORCHARD (NCT03944772)16 is also underway to assess the efficacy of several osimertinib-based combinations following disease progression under frontline osimertinib.
BRAF mutations and fusions (i.e. AGK-BRAF, ESYT2-BRAF) have recently emerged as additional mechanisms of AR to third-generation EGFR TKI2,18–20. Studies demonstrating the efficacy of concurrent inhibition of EGFR and BRAF3 or MEK21 in pre-clinical models have raised clinicians’ expectations about overcoming AR by combining TT. Nevertheless, reports of successful combinations of TT for patients harboring BRAF-driven AR to osimertinib are very limited19,22–24 and no prospective data regarding efficacy and safety of BRAF/MEK/EGFR concurrent inhibition are available, with chemotherapy-based regimens remaining the treatment of choice in this unfavorable scenario. Similarly, PIK3CA mutations may also mediate AR to second-line osimertinib in 4–11%, but no clinical reports suggesting potential benefits of blocking these alterations in NSCLC are available6,9,25–27.
The detection of circulating tumor DNA (ctDNA) using liquid biopsies allows noninvasive real-time monitoring of treatment response and early detection of AR to TT, anticipating radiological response and treatment failure28,29. The widespread availability of highly specific and sensitive techniques to quantify ctDNA makes the longitudinal assessment of patients with NSCLC under TKI therapy very attractive. Nonetheless, for NSCLCs, ctDNA evaluation in routine clinical practice is currently limited to the detection of EGFR exon 19 deletion, EGFR L858R and T790M activating mutations29.
A man with metastatic lung adenocarcinoma harboring an EGFR mutation, who had progressed to erlotinib due to the emergence of the EGFR T790M mutation started osimertinib and remained on treatment for 15 months, then developing disease progression (PD). Sequential liquid biopsies were collected to monitor treatment response and disclosed the emergence of BRAF V600E and PIK3CA E545K resistance mutations 4 months before clinical progression. With this result and considering few available reports in the literature, our patient started a triple therapy with osimertinib, dabrafenib (BRAF inhibitor), and trametinib (MEK inhibitor).
Results
Case report
A 50-year-old non-smoker man was diagnosed with a tubule-papillary lung adenocarcinoma metastatic to bones and soft tissue (stage IVB - AJCC 8th edition) in July 2016. Several hypermetabolic bone lesions were observed on staging 18-fluorodeoxyglucose positron-emission tomography (18F-FDG PET-CT) scan at diagnosis. A baseline Next Generation Sequencing (NGS) assay (TruSightTumor™ - Illumina®) of a soft tissue metastasis revealed the presence of an activating EGFR exon 19 deletion (EGFR E746_A750del). No concurrent alterations in BRAF or in PIK3CA were identified at that time. ALK d5f3 immunohistochemistry and fluorescent in situ hybridization (FISH) for ROS1 and RET resulted negative.
In September 2016, the patient started on erlotinib 150 mg once daily (OD), achieving complete metabolic response in February 2017. We started to perform serial blood sample collections for ctDNA analysis in May 2017. In October 2017, oligo-progression (oligo-PD) was observed at the primary site and treated with stereotactic body radiation therapy (SBRT; 3 × 16 Gy). In November 2017, we identified the EGFR T790M and EGFR E746_A750del mutations in patient’s plasma using droplet digital PCR (ddPCR) and these results were confirmed in December 2017 with the Foundation ACT® ctDNA assay. Figure 1a shows all systemic and focal therapies of this patient since diagnosis.Fig. 1 Paired radiological and sequential blood-based ctDNA assessments throughout patient’s treatment demonstrating concordant results.
a Timeline displaying systemic and focal therapies since September 2016. b ctDNA levels in serial plasma samples based on fractional abundance of EGFR T790M, EGFR del19 (E746_A750 del), BRAF V600E, and PIK3CA E545K mutations. #2.4 CN results: EGFR del19 WT/RNAseP copy number ratio. c Paired 18F-FDG PET-CT scan maximum intensity projection (MIP) images displaying tumor burden variations. PD progression of disease, SBRT stereotatic body radiotherapy, Pembro pembrolizumab, CT chemotherapy, DTO dabrafenib, trametinib and osimertinib.
Erlotinib was kept until January 2018 when disease progression in the bones was observed and the fractional abundance (FA) of EGFR E746_A750del and EGFR T790M were 12.0 and 2.3%, respectively (Fig. 1b). The treatment was promptly switched to osimertinib 80 mg OD. Bone partial response (PR) was observed 2 months later, along with a significant drop in the FA of EGFR E746_A750del and EGFR T790M mutations in the plasma (Fig. 1b).
In July 2018, the patient developed oligo-PD in T3 vertebrae, which was treated with SBRT (1 × 20 Gy). At that point, ctDNA analysis revealed an increase in the FA of the EGFR E746_A750del in the plasma, but not in EGFR T790M, suggesting genetic heterogeneity between metastatic lesions. In December 2018, a new oligo-PD in the left iliac bone was detected by 18F-FDG PET-CT scan (Fig. 1c). At that time, ctDNA analysis revealed the emergence of BRAF V600E (FA: 0.4%) and PIK3CA E545K (FA: 0.9%) mutations, as well as a significant increase in the FA of the EGFR E746_A750del (FA: 7.5%) in the plasma. These mutations were also detected in a specimen from an iliac bone biopsy using a NGS assay (TruSightTumor™ - Illumina® - EGFR del19 allelic fraction 81.1%, EGFR del19 amplification – 12 copies, BRAF V600E allelic fraction 17.7% and PIK3CA E545K allelic fraction 32.7%). Even though the patient was treated with SBRT (1 × 18 Gy), he developed new bone metastases in March 2019 (Fig. 1b), consistent with the significant increase in the FA of EGFR E746_A750del, BRAF V600E and EGFR T790M mutations (Fig. 1b). At that specific timepoint and in April 2019, assessments of PIK3CA E545K plasma levels were not obtained due to limited amount of cfDNA.
Between April and September 2019, systemic treatment with carboplatin plus pemetrexed and pembrolizumab provided modest clinical benefit, reducing tumor burden and controlling the disease (Fig. 1b). Due to a new PD observed in October 2019, the treatment was switched to docetaxel monotherapy and, subsequently, to vinorelbine after progression under docetaxel; nonetheless, the patient experienced symptomatic PD in bones, lymph nodes, and lung in late January 2020 (Fig. 1c, 2a–d). He came to the clinic with a Karnofsky performance status of 70%, complaining of fatigue, appetite loss, and severe pain in the hips (despite regular use of 10 mg buprenorphine patch and oxycodone plus acetaminophen PO), which had been preventing him from performing his daily activities due to the inability to stay seated. At that point, high levels of all three resistance mutations were detected in the circulating DNA (PIK3CA E545K FA: 13.2%, BRAF V600E FA: 12.3%, and EGFR T790M FA: 5.3%; Fig. 1b). We also observed an exponential increase in the number of copies of EGFR E746_A750del, suggesting gene amplification (Fig. 1b).Fig. 2 18F-FDG PET-CT scan imaging depicting impressive overall tumor response under dabrafenib, trametinib, and osimertinib in three timepoints.
a–d baseline imaging (January/2020) showing: hypermetabolic spiculated mass (orange circle) in the left-superior lobe measuring 4.4 × 3.3 cm (SUVmax: 9.4); multiple hilar, mediastinal, retroperitoneal, and iliac hypermetabolic lymph nodes (red arrow) measuring up to 2.7 cm (SUVmax: 12.8); several hypermetabolic bone lesions throughout axial and appendicular skeleton (yellow circle); a left iliac bone lesion with signs of periosteal reaction and adjacent soft-tissue infiltration (SUVmax: 11.4). e–h first response evaluation imaging (April/2020) showing considerable partial response in the lung mass, measuring 3.8 × 2.5 cm (SUVmax: 2.7), as well as in several bone lesions, especially in the left-iliac bone (SUVmax: 4.8); complete response in lymph nodes. i–l third response evaluation imaging showing disease progression in the left ischium (SUVmax: 12.5; previous SUVmax: 5.9) (October/2020). SUVmax maximum standard uptake value.
In February 2020, after careful consideration due to patient’s good performance status, normal organ functions, severe pain, motivation and the emergence of BRAF V600E as an AR mechanism, he started on dabrafenib 75 mg twice daily (BID), trametinib 1 mg OD and osimertinib 80 mg OD. The use of a PI3K inhibitor was not considered an option due to the absence of reports in the literature suggesting efficacy of these drugs combined with osimertinib to treat advanced NSCLC patients. Within 2 weeks of treatment, the patient achieved complete resolution of the severe bone pain in the hips with no further need of opioid administration, as well as appetite gain and marked improvement in quality of life, which turned possible for him to resume his daily activities. As adverse events (AE), he experienced grade 1 fatigue, dysgeusia, fever, and nausea, all managed with symptomatic medication. Complete resolution of the fever occurred spontaneously within two weeks of treatment. An attempt to increase dosages of dabrafenib to 150 mg BID and trametinib 2 mg OD resulted unsuccessful due to persistent grade 2 fatigue. In April 2020, a new 18F-FDG PET-CT scan disclosed a complete response in lymph nodes and a dramatic PR in the lung and bones (PERCIST 1.1 evaluating 5 target lesions: Δ- 67%; Figs. 1c and 2e–h). Detection of EGFR T790M, BRAF V600E, and PIK3CA E545K mutations became negative in April 2020, while EGFR E746_A750del despite marginally positive, presented marked reduction (Fig. 1b) suggesting that combination dabrafenib, trametinib, and osimertinib might be an effective strategy to overcome PIK3CA E545K and BRAF V600E-driven resistance to osimertinib in advanced EGFR-mutant NSCLC patients. The patient remained in response until October 2020, when he developed asymptomatic bone PD in lumbar spine, left ischium, and right iliac bone, 8 months after starting this combination therapy (Figs. 1c and 2i–l). A FoundationOne®Liquid CDx plasma NGS disclosed the following alterations: CHCHD3-BRAF fusion, BRAF V600E, EGFR E746_750del, EGFR amplification, PIK3CA E545K, MAP2K2 (MEK2) C125S, MTAP rearrangement intron 5, TP53 V197M, and TP53 S241A. Owing to the considerable clinical benefit, good tolerance and lack of systemic treatment options, we decided to keep the patient on treatment and increase the doses of dabrafenib (150 mg BID) and trametinib (2 mg OD) every other day. He remains asymptomatic and tolerating well the proposed dose adjustment.
Discussion
Several studies have recently highlighted the importance of considering genes of interest within the context of commonly co-occurring mutations9,30. For example, as described by Blakely et al. through performing a cfDNA NGS analysis of 1,122 advanced stage EGFR-mutant NSCLCs, in ~93% of the patients, at least one more variables with known or likely known functional properties were present, disclosing the molecular complexity of this oncogenic driver and suggesting an association of co-occuring genomic alterations with TKI response and clinical outcomes9. In addition, Roper and colleagues reported the identification of at least two co-existing AR mechanisms in 73% of patients treated with osimertinib, as well as 6–23 different subclones per individual in a phylogenetic analysis performed in multiple metastatic sites of 15 individuals30. In the same publication, the authors also called the attention for a high incidence of acquired EGFR amplifications in post-osimertinib patients, which suggests maintenance of EGFR central role in the setting of progression, as previously reported by our group28 and also detected in the present case.
Although uncommon, BRAF V600E mediates AR in approximately 3% of the patients under second-line osimertinib6,20 and little is known about the efficacy of combined TT in this population3,21–23. Existing reports lack details regarding objective responses using standardized radiological criteria (i.e., RECIST 1.1; PERCIST) and the dynamics of resistance mechanisms through longitudinal ctDNA measurements22,23. Huang and colleagues22 described a case of an EGFRdel19/T790M + NSCLC patient who developed BRAF V600E-driven AR after second-line osimertinib and achieved stable disease under dabrafenib 150 mg BID, trametinib 1 mg OD and osimertinib 80 mg OD, with ongoing disease control 7.4 months after. Zhou and colleagues23 also reported their experience with dabrafenib 150 mg BID, trametinib 2 mg OD and osimertinib 80 mg OD leading to tumor reduction within 6 weeks of treatment, along with grade 2 rash and decreased appetite as AEs. Similarly, Meng et al. reported two cases treated with this triple regimen31. The first patient discontinued therapy after one month due to severe pneumonitis; the second one presented tumor response under dabrafenib 50 mg BID, trametinib 0.5 mg OD, and osimertinib 80 mg OD, with progression-free survival of 14 months. According to the authors, this reduced dose was prescribed owing to a grade 2 pyrexia, nausea, and vomiting under higher dabrafenib and trametinib doses. Dagogo-Jack and colleagues19 also described a successful case of combined EGFR/MAP kinase pathway blockade with osimertinib 80 mg OD and trametinib 1 mg OD; as treatment-related AEs, their patient experienced grade 2 diarrhea and fatigue, along with grade 1 rash and gastrointestinal bleeding. Nevertheless, it is worth highlighting the rapid clinical improvement, the remarkable radiologically confirmed objective response, as well as the good tolerance observed in this case even using only half standard dose of dabrafenib and trametinib approved for NSCLCs harboring BRAF V600E mutations. Since data regarding the efficacy of these combined approaches, the optimal drug association and dosing, as well as the toxicity profile are conflicting and largely unknown, further investigation into the mechanistic basis of this association represents an important priority. The Table 1 summarizes the toxicities arising under osimertinib plus BRAF/MEK inhibition and reported in the literature.Table 1 Summary of the toxicities arising under osimertinib plus BRAF/MEK inhibition and reported in the literature.
Reference Treatment Anorexia Nausea Vomiting Diarrhoea Fatigue Rash GI bleeding AST/ALT elevation Paroniquia Pyrexia Initial dose Dose reduction
Meng et al. D + T + O NR G2 G2 G2 NR NR NR NR NR G2 D (150 mg BID) + T (2 mg OD) + O (80 mg OD) D (50 mg BID) + T (0.5 mg/day) + O (80 mg/day)
Dagogo-Jack et al. T + O NR NR NR G2 G2 G1 G1 G1 NR NR T (1 mg OD) + O (80 mg OD) Not needed
Huang et al. D + T + O NR NR NR G1 NR NR NR NR G1 NR D (150 mg BID) + T (1 mg OD) + O (80 mg OD) Not needed
Zhou et al. D + T + O G1 NR NR NR NR G2 NR NR NR NR D (150 mg BID) + T (2 mg OD) + O (80 mg OD) Not needed
D dabrafenib, T trametinib, O osimertinib, NR not reported.
Liquid biopsy is emerging as an important diagnostic and predictive tool in the treatment of NSCLCs. The accurate identification of predictive genetic alterations is important for both patients’ management and the understanding of clonal evolution and AR to different therapies9,29,32. Also, circulating biomarkers from multiple disease sites better reflect systemic tumor burden, including alterations from genetically different metastatic lesions, which may be missed with single-site tissue biopsies8. Here, we were able to identify and monitor multiple concurrent mechanisms of resistance throughout the entire patient’s treatment. As shown in Fig. 1b, variations in blood ctDNA levels exhibited a positive correlation with imaging findings, even in situations involving focal treatments for oligo-PD. The observed concordance between radiological PD or response and ctDNA measurements also points towards its great potential to be incorporated into clinical practice to anticipate radiologic findings in a more effective manner.
Along with BRAF V600E mutation, the presence of PIK3CA E545K mutation is also associated with AR to osimertinib6,9,25,27. However, the clinical implication of concomitant targeting of EGFR and PI3K remains unclear. Whereas an alternative argument for the observed response could be a rechallenge following a long time upon off-osimertinib, in our opinion, this is an unlikely explanation33. Interestingly, the presence of a BRAF fusion along with a MEK2 mutation identified through plasma NGS in the setting of PD might represent a mechanism of resistance to this triple regimen34. Nonetheless, since pretreatment blood samples had not been analyzed using a similar methodology, this hypothesis deserves careful interpretation.
Increased understanding of the relationship of concurring genomic alterations in EGFR-mutant NSCLC may enable new therapeutic opportunities following disease progression to osimertinib. Here, we reported an impressive objective response to dabrafenib, trametinib, and osimertinib with concordant decrease in plasma ctDNA levels in a metastatic lung cancer patient harboring EGFR E746_A750del, BRAF V600E, and PIK3CA E545K activating mutations. This case report leads to a greater understanding of the currently limited literature regarding the management of EGFR-mutant NSCLC patients with acquired BRAFV600E mutation, since it reports a successful attempt to target both alterations concurrently while providing concordant and interesting data of serial ctDNA assessments throughout the entire treatment. Further investigation to optimize the efficacy and mitigate the toxicity profile of this drug association represents an important issue.
Methods
Patient
This study was approved by Hospital Sírio-Libanês Ethics Committee (HSL-RC 2020-16). The patient provided written informed consent for blood collection, ctDNA analysis and publication of this report.
Sample collection and plasma DNA extraction
Serial blood samples were collected between May 2017 and April 2020 (Fig. 1b). Peripheral blood (20 ml) was collected into tubes containing EDTA (BD, Franklin Lakes, New Jersey). Plasma was separated from the blood within 2 hours of blood collection, as previously described28. cfDNA was extracted using QIAamp MinElute Virus Vacuum Kit (Qiagen, Hilden, Germany) and stored at −80 °C.
ctDNA-ddPCR
Cell-free DNA (cfDNA) was quantified using the RNase P Copy Number Reference Assay (Life Technologies, Carlsbad, California). A total of 3000 genome-equivalents (~10 ng of cfDNA) were analyzed per assay to achieve a detection sensitivity of 0.2%. This detection limit has been assessed by using cell line-derived genomic DNA. A total of 10 ng of input DNA with varying proportions of mutant DNA was serially diluted into wild-type DNA to obtain samples with a mutant abundance of 1%, 0.5%, 0.1%, and 0.05% and subjected to droplet digital PCR (ddPCR). ddPCR was used to quantify the circulating levels of the EGFR activating mutation (EGFR E746_A750 del) and of the resistance mutations (EGFR T790M, PIK3CA E545K, and BRAF V600E). Probes and primers were obtained from BioRad (EGFR E746_A750 del #10041170, EGFR T790M #10040782, PIK3CA E545K #10041188, and BRAF V600E #10040779; Hercules, California). ddPCR was performed on the QX200 Droplet Digital PCR System, and data were analysed using QuantaSoft software (Bio-Rad). ctDNA quantification is presented as fractional abundance (FA–the proportion of the mutant allele in total cfDNA).
Reporting summary
Further information on research design is available in the Nature Research Reporting Summary linked to this article.
Supplementary information
Reporting Summary
Dataset 1
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
These authors contributed equally: Maurício Fernando Silva Almeida Ribeiro, Franciele Hinterholz Knebel.
Supplementary information
The online version contains supplementary material available at 10.1038/s41698-021-00149-4.
Acknowledgements
We thank the clinical research team at Hospital Sírio-Libanês for their assistance collecting samples. We thank Ernande Xavier dos Santos and Elisângela Monteiro for the outstanding technical support. We also thank Mrs. Dina Binzagr and Vivian Hannud for supporting the Translational Research Program at Hospital Sírio Libanês. This work was supported by Ludwig Cancer Research and Fundação de Amparo à Pesquisa do Estado de São Paulo – FAPESP. Lastly, we would like to thank the patient and his family for consenting to the present publication.
Author contributions
Conceptualization by M.F.S.A.R., F.H.K., J.V.M.A., A.A.C., and A.K.; Methodology by F.H.K. and A.A.C.; Resources br M.F.S.A.R., F.H.K., A.A.C., and J.V.M.A.; Writing of the first draft by M.F.S.A., F.H.K., and J.V.M.A.; Review, editing and approval of the final draft by all authors; Supervision by A.A.C. and A.K.; Funding acquisition by F.H.K. and A.A.C. M.F.S.A.R. and F.H.K. are co-first authors of the present work.
Data availability
The datasets that support the findings of this study are not publicly available in order to protect patient privacy. The data will be made available on reasonable request. For data access requests regarding the liquid biopsy (ctDNA quantification) data, please contact Dr. Franciele Knebel, email address: fhknebel@mochsl.org.br. For data access requests regarding the PET-CT high resolution images and PERCIST calculations, please contact Dr. José Marin, email address:jfgmarin@yahoo.com.br. For data access requests regarding the summary of the toxicities arising under osimertinib plus BRAF/MEK inhibition, please contact the corresponding author Dr. Maurício Ribeiro, email address: mauricio.fsaribeiro@hsl.org.br. The data generated and analysed during this study are described in the following metadata record: 10.6084/m9.figshare.1347594635.
Competing interests
The authors declare no competing interests. | DABRAFENIB, OSIMERTINIB, TRAMETINIB | DrugsGivenReaction | CC BY | 33580193 | 19,431,708 | 2021-02-12 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Fatigue'. | Impressive response to dabrafenib, trametinib, and osimertinib in a metastatic EGFR-mutant/BRAF V600E lung adenocarcinoma patient.
The survival outcomes of the FLAURA trial support osimertinib as the new standard of care for untreated patients harboring activating mutations in the epidermal growth factor receptor (EGFR). Despite the initial response, disease progression invariably occurs. Although uncommon, BRAF V600E mutation arises as a unique mechanism of resistance, and thus far, no prospective studies are available to support concurrent EGFR/BRAF blockade. We report a case of impressive radiological and ctDNA response under dabrafenib, trametinib, and osimertinib in an advanced EGFR-mutant lung adenocarcinoma patient who developed BRAF V600E as one of the acquired resistance mechanisms to second-line osimertinib. Moreover, the patient experienced remarkable clinical improvement and good tolerance to combination therapy. The present case suggests the importance of prospective studies evaluating both efficacy and safety of the combination in later line settings and points towards the potential of ctDNA to monitor resistance mechanisms and treatment benefit in clinical practice.
Introduction
The use of osimertinib to target epidermal growth factor receptor (EGFR) has become the standard of care in untreated EGFR-mutant non–small cell lung cancer (NSCLC) patients. Although osimertinib can be highly active, showing more durable outcomes than first-generation tyrosine kinase inhibitors (TKI)1, most tumors invariably become resistant, limiting its long-term clinical benefit. The heterogeneity of resistance mechanisms to osimertinib, including EGFR C797S mutation, EGFR, and MET amplifications, off-target mutations in PIK3CA, KRAS, and HER2 as well as histologic transformation2–9, has stimulated routine performance of repeated biopsies to identify specific underlying mechanisms of resistance throughout the treatment course, and to guide the development of novel therapeutic strategies to overcome and prevent acquired resistance (AR)7,10–13.
Combined targeted therapy (TT) strategies have been increasingly addressed in prospective clinical trials14–17. A phase Ib/II trial reported 47% ORR with a combination of capmatinib (MET inhibitor) plus gefitinib in patients progressing to EGFR TKI and presenting MET dysregulation14. Likewise, SAVANNAH (NCT03778229)15 is an ongoing phase II trial designed to evaluate the efficacy of combination savolitinib and osimertinib in post-osimertinib progression/MET-positive patients. The biomarker-matched study ORCHARD (NCT03944772)16 is also underway to assess the efficacy of several osimertinib-based combinations following disease progression under frontline osimertinib.
BRAF mutations and fusions (i.e. AGK-BRAF, ESYT2-BRAF) have recently emerged as additional mechanisms of AR to third-generation EGFR TKI2,18–20. Studies demonstrating the efficacy of concurrent inhibition of EGFR and BRAF3 or MEK21 in pre-clinical models have raised clinicians’ expectations about overcoming AR by combining TT. Nevertheless, reports of successful combinations of TT for patients harboring BRAF-driven AR to osimertinib are very limited19,22–24 and no prospective data regarding efficacy and safety of BRAF/MEK/EGFR concurrent inhibition are available, with chemotherapy-based regimens remaining the treatment of choice in this unfavorable scenario. Similarly, PIK3CA mutations may also mediate AR to second-line osimertinib in 4–11%, but no clinical reports suggesting potential benefits of blocking these alterations in NSCLC are available6,9,25–27.
The detection of circulating tumor DNA (ctDNA) using liquid biopsies allows noninvasive real-time monitoring of treatment response and early detection of AR to TT, anticipating radiological response and treatment failure28,29. The widespread availability of highly specific and sensitive techniques to quantify ctDNA makes the longitudinal assessment of patients with NSCLC under TKI therapy very attractive. Nonetheless, for NSCLCs, ctDNA evaluation in routine clinical practice is currently limited to the detection of EGFR exon 19 deletion, EGFR L858R and T790M activating mutations29.
A man with metastatic lung adenocarcinoma harboring an EGFR mutation, who had progressed to erlotinib due to the emergence of the EGFR T790M mutation started osimertinib and remained on treatment for 15 months, then developing disease progression (PD). Sequential liquid biopsies were collected to monitor treatment response and disclosed the emergence of BRAF V600E and PIK3CA E545K resistance mutations 4 months before clinical progression. With this result and considering few available reports in the literature, our patient started a triple therapy with osimertinib, dabrafenib (BRAF inhibitor), and trametinib (MEK inhibitor).
Results
Case report
A 50-year-old non-smoker man was diagnosed with a tubule-papillary lung adenocarcinoma metastatic to bones and soft tissue (stage IVB - AJCC 8th edition) in July 2016. Several hypermetabolic bone lesions were observed on staging 18-fluorodeoxyglucose positron-emission tomography (18F-FDG PET-CT) scan at diagnosis. A baseline Next Generation Sequencing (NGS) assay (TruSightTumor™ - Illumina®) of a soft tissue metastasis revealed the presence of an activating EGFR exon 19 deletion (EGFR E746_A750del). No concurrent alterations in BRAF or in PIK3CA were identified at that time. ALK d5f3 immunohistochemistry and fluorescent in situ hybridization (FISH) for ROS1 and RET resulted negative.
In September 2016, the patient started on erlotinib 150 mg once daily (OD), achieving complete metabolic response in February 2017. We started to perform serial blood sample collections for ctDNA analysis in May 2017. In October 2017, oligo-progression (oligo-PD) was observed at the primary site and treated with stereotactic body radiation therapy (SBRT; 3 × 16 Gy). In November 2017, we identified the EGFR T790M and EGFR E746_A750del mutations in patient’s plasma using droplet digital PCR (ddPCR) and these results were confirmed in December 2017 with the Foundation ACT® ctDNA assay. Figure 1a shows all systemic and focal therapies of this patient since diagnosis.Fig. 1 Paired radiological and sequential blood-based ctDNA assessments throughout patient’s treatment demonstrating concordant results.
a Timeline displaying systemic and focal therapies since September 2016. b ctDNA levels in serial plasma samples based on fractional abundance of EGFR T790M, EGFR del19 (E746_A750 del), BRAF V600E, and PIK3CA E545K mutations. #2.4 CN results: EGFR del19 WT/RNAseP copy number ratio. c Paired 18F-FDG PET-CT scan maximum intensity projection (MIP) images displaying tumor burden variations. PD progression of disease, SBRT stereotatic body radiotherapy, Pembro pembrolizumab, CT chemotherapy, DTO dabrafenib, trametinib and osimertinib.
Erlotinib was kept until January 2018 when disease progression in the bones was observed and the fractional abundance (FA) of EGFR E746_A750del and EGFR T790M were 12.0 and 2.3%, respectively (Fig. 1b). The treatment was promptly switched to osimertinib 80 mg OD. Bone partial response (PR) was observed 2 months later, along with a significant drop in the FA of EGFR E746_A750del and EGFR T790M mutations in the plasma (Fig. 1b).
In July 2018, the patient developed oligo-PD in T3 vertebrae, which was treated with SBRT (1 × 20 Gy). At that point, ctDNA analysis revealed an increase in the FA of the EGFR E746_A750del in the plasma, but not in EGFR T790M, suggesting genetic heterogeneity between metastatic lesions. In December 2018, a new oligo-PD in the left iliac bone was detected by 18F-FDG PET-CT scan (Fig. 1c). At that time, ctDNA analysis revealed the emergence of BRAF V600E (FA: 0.4%) and PIK3CA E545K (FA: 0.9%) mutations, as well as a significant increase in the FA of the EGFR E746_A750del (FA: 7.5%) in the plasma. These mutations were also detected in a specimen from an iliac bone biopsy using a NGS assay (TruSightTumor™ - Illumina® - EGFR del19 allelic fraction 81.1%, EGFR del19 amplification – 12 copies, BRAF V600E allelic fraction 17.7% and PIK3CA E545K allelic fraction 32.7%). Even though the patient was treated with SBRT (1 × 18 Gy), he developed new bone metastases in March 2019 (Fig. 1b), consistent with the significant increase in the FA of EGFR E746_A750del, BRAF V600E and EGFR T790M mutations (Fig. 1b). At that specific timepoint and in April 2019, assessments of PIK3CA E545K plasma levels were not obtained due to limited amount of cfDNA.
Between April and September 2019, systemic treatment with carboplatin plus pemetrexed and pembrolizumab provided modest clinical benefit, reducing tumor burden and controlling the disease (Fig. 1b). Due to a new PD observed in October 2019, the treatment was switched to docetaxel monotherapy and, subsequently, to vinorelbine after progression under docetaxel; nonetheless, the patient experienced symptomatic PD in bones, lymph nodes, and lung in late January 2020 (Fig. 1c, 2a–d). He came to the clinic with a Karnofsky performance status of 70%, complaining of fatigue, appetite loss, and severe pain in the hips (despite regular use of 10 mg buprenorphine patch and oxycodone plus acetaminophen PO), which had been preventing him from performing his daily activities due to the inability to stay seated. At that point, high levels of all three resistance mutations were detected in the circulating DNA (PIK3CA E545K FA: 13.2%, BRAF V600E FA: 12.3%, and EGFR T790M FA: 5.3%; Fig. 1b). We also observed an exponential increase in the number of copies of EGFR E746_A750del, suggesting gene amplification (Fig. 1b).Fig. 2 18F-FDG PET-CT scan imaging depicting impressive overall tumor response under dabrafenib, trametinib, and osimertinib in three timepoints.
a–d baseline imaging (January/2020) showing: hypermetabolic spiculated mass (orange circle) in the left-superior lobe measuring 4.4 × 3.3 cm (SUVmax: 9.4); multiple hilar, mediastinal, retroperitoneal, and iliac hypermetabolic lymph nodes (red arrow) measuring up to 2.7 cm (SUVmax: 12.8); several hypermetabolic bone lesions throughout axial and appendicular skeleton (yellow circle); a left iliac bone lesion with signs of periosteal reaction and adjacent soft-tissue infiltration (SUVmax: 11.4). e–h first response evaluation imaging (April/2020) showing considerable partial response in the lung mass, measuring 3.8 × 2.5 cm (SUVmax: 2.7), as well as in several bone lesions, especially in the left-iliac bone (SUVmax: 4.8); complete response in lymph nodes. i–l third response evaluation imaging showing disease progression in the left ischium (SUVmax: 12.5; previous SUVmax: 5.9) (October/2020). SUVmax maximum standard uptake value.
In February 2020, after careful consideration due to patient’s good performance status, normal organ functions, severe pain, motivation and the emergence of BRAF V600E as an AR mechanism, he started on dabrafenib 75 mg twice daily (BID), trametinib 1 mg OD and osimertinib 80 mg OD. The use of a PI3K inhibitor was not considered an option due to the absence of reports in the literature suggesting efficacy of these drugs combined with osimertinib to treat advanced NSCLC patients. Within 2 weeks of treatment, the patient achieved complete resolution of the severe bone pain in the hips with no further need of opioid administration, as well as appetite gain and marked improvement in quality of life, which turned possible for him to resume his daily activities. As adverse events (AE), he experienced grade 1 fatigue, dysgeusia, fever, and nausea, all managed with symptomatic medication. Complete resolution of the fever occurred spontaneously within two weeks of treatment. An attempt to increase dosages of dabrafenib to 150 mg BID and trametinib 2 mg OD resulted unsuccessful due to persistent grade 2 fatigue. In April 2020, a new 18F-FDG PET-CT scan disclosed a complete response in lymph nodes and a dramatic PR in the lung and bones (PERCIST 1.1 evaluating 5 target lesions: Δ- 67%; Figs. 1c and 2e–h). Detection of EGFR T790M, BRAF V600E, and PIK3CA E545K mutations became negative in April 2020, while EGFR E746_A750del despite marginally positive, presented marked reduction (Fig. 1b) suggesting that combination dabrafenib, trametinib, and osimertinib might be an effective strategy to overcome PIK3CA E545K and BRAF V600E-driven resistance to osimertinib in advanced EGFR-mutant NSCLC patients. The patient remained in response until October 2020, when he developed asymptomatic bone PD in lumbar spine, left ischium, and right iliac bone, 8 months after starting this combination therapy (Figs. 1c and 2i–l). A FoundationOne®Liquid CDx plasma NGS disclosed the following alterations: CHCHD3-BRAF fusion, BRAF V600E, EGFR E746_750del, EGFR amplification, PIK3CA E545K, MAP2K2 (MEK2) C125S, MTAP rearrangement intron 5, TP53 V197M, and TP53 S241A. Owing to the considerable clinical benefit, good tolerance and lack of systemic treatment options, we decided to keep the patient on treatment and increase the doses of dabrafenib (150 mg BID) and trametinib (2 mg OD) every other day. He remains asymptomatic and tolerating well the proposed dose adjustment.
Discussion
Several studies have recently highlighted the importance of considering genes of interest within the context of commonly co-occurring mutations9,30. For example, as described by Blakely et al. through performing a cfDNA NGS analysis of 1,122 advanced stage EGFR-mutant NSCLCs, in ~93% of the patients, at least one more variables with known or likely known functional properties were present, disclosing the molecular complexity of this oncogenic driver and suggesting an association of co-occuring genomic alterations with TKI response and clinical outcomes9. In addition, Roper and colleagues reported the identification of at least two co-existing AR mechanisms in 73% of patients treated with osimertinib, as well as 6–23 different subclones per individual in a phylogenetic analysis performed in multiple metastatic sites of 15 individuals30. In the same publication, the authors also called the attention for a high incidence of acquired EGFR amplifications in post-osimertinib patients, which suggests maintenance of EGFR central role in the setting of progression, as previously reported by our group28 and also detected in the present case.
Although uncommon, BRAF V600E mediates AR in approximately 3% of the patients under second-line osimertinib6,20 and little is known about the efficacy of combined TT in this population3,21–23. Existing reports lack details regarding objective responses using standardized radiological criteria (i.e., RECIST 1.1; PERCIST) and the dynamics of resistance mechanisms through longitudinal ctDNA measurements22,23. Huang and colleagues22 described a case of an EGFRdel19/T790M + NSCLC patient who developed BRAF V600E-driven AR after second-line osimertinib and achieved stable disease under dabrafenib 150 mg BID, trametinib 1 mg OD and osimertinib 80 mg OD, with ongoing disease control 7.4 months after. Zhou and colleagues23 also reported their experience with dabrafenib 150 mg BID, trametinib 2 mg OD and osimertinib 80 mg OD leading to tumor reduction within 6 weeks of treatment, along with grade 2 rash and decreased appetite as AEs. Similarly, Meng et al. reported two cases treated with this triple regimen31. The first patient discontinued therapy after one month due to severe pneumonitis; the second one presented tumor response under dabrafenib 50 mg BID, trametinib 0.5 mg OD, and osimertinib 80 mg OD, with progression-free survival of 14 months. According to the authors, this reduced dose was prescribed owing to a grade 2 pyrexia, nausea, and vomiting under higher dabrafenib and trametinib doses. Dagogo-Jack and colleagues19 also described a successful case of combined EGFR/MAP kinase pathway blockade with osimertinib 80 mg OD and trametinib 1 mg OD; as treatment-related AEs, their patient experienced grade 2 diarrhea and fatigue, along with grade 1 rash and gastrointestinal bleeding. Nevertheless, it is worth highlighting the rapid clinical improvement, the remarkable radiologically confirmed objective response, as well as the good tolerance observed in this case even using only half standard dose of dabrafenib and trametinib approved for NSCLCs harboring BRAF V600E mutations. Since data regarding the efficacy of these combined approaches, the optimal drug association and dosing, as well as the toxicity profile are conflicting and largely unknown, further investigation into the mechanistic basis of this association represents an important priority. The Table 1 summarizes the toxicities arising under osimertinib plus BRAF/MEK inhibition and reported in the literature.Table 1 Summary of the toxicities arising under osimertinib plus BRAF/MEK inhibition and reported in the literature.
Reference Treatment Anorexia Nausea Vomiting Diarrhoea Fatigue Rash GI bleeding AST/ALT elevation Paroniquia Pyrexia Initial dose Dose reduction
Meng et al. D + T + O NR G2 G2 G2 NR NR NR NR NR G2 D (150 mg BID) + T (2 mg OD) + O (80 mg OD) D (50 mg BID) + T (0.5 mg/day) + O (80 mg/day)
Dagogo-Jack et al. T + O NR NR NR G2 G2 G1 G1 G1 NR NR T (1 mg OD) + O (80 mg OD) Not needed
Huang et al. D + T + O NR NR NR G1 NR NR NR NR G1 NR D (150 mg BID) + T (1 mg OD) + O (80 mg OD) Not needed
Zhou et al. D + T + O G1 NR NR NR NR G2 NR NR NR NR D (150 mg BID) + T (2 mg OD) + O (80 mg OD) Not needed
D dabrafenib, T trametinib, O osimertinib, NR not reported.
Liquid biopsy is emerging as an important diagnostic and predictive tool in the treatment of NSCLCs. The accurate identification of predictive genetic alterations is important for both patients’ management and the understanding of clonal evolution and AR to different therapies9,29,32. Also, circulating biomarkers from multiple disease sites better reflect systemic tumor burden, including alterations from genetically different metastatic lesions, which may be missed with single-site tissue biopsies8. Here, we were able to identify and monitor multiple concurrent mechanisms of resistance throughout the entire patient’s treatment. As shown in Fig. 1b, variations in blood ctDNA levels exhibited a positive correlation with imaging findings, even in situations involving focal treatments for oligo-PD. The observed concordance between radiological PD or response and ctDNA measurements also points towards its great potential to be incorporated into clinical practice to anticipate radiologic findings in a more effective manner.
Along with BRAF V600E mutation, the presence of PIK3CA E545K mutation is also associated with AR to osimertinib6,9,25,27. However, the clinical implication of concomitant targeting of EGFR and PI3K remains unclear. Whereas an alternative argument for the observed response could be a rechallenge following a long time upon off-osimertinib, in our opinion, this is an unlikely explanation33. Interestingly, the presence of a BRAF fusion along with a MEK2 mutation identified through plasma NGS in the setting of PD might represent a mechanism of resistance to this triple regimen34. Nonetheless, since pretreatment blood samples had not been analyzed using a similar methodology, this hypothesis deserves careful interpretation.
Increased understanding of the relationship of concurring genomic alterations in EGFR-mutant NSCLC may enable new therapeutic opportunities following disease progression to osimertinib. Here, we reported an impressive objective response to dabrafenib, trametinib, and osimertinib with concordant decrease in plasma ctDNA levels in a metastatic lung cancer patient harboring EGFR E746_A750del, BRAF V600E, and PIK3CA E545K activating mutations. This case report leads to a greater understanding of the currently limited literature regarding the management of EGFR-mutant NSCLC patients with acquired BRAFV600E mutation, since it reports a successful attempt to target both alterations concurrently while providing concordant and interesting data of serial ctDNA assessments throughout the entire treatment. Further investigation to optimize the efficacy and mitigate the toxicity profile of this drug association represents an important issue.
Methods
Patient
This study was approved by Hospital Sírio-Libanês Ethics Committee (HSL-RC 2020-16). The patient provided written informed consent for blood collection, ctDNA analysis and publication of this report.
Sample collection and plasma DNA extraction
Serial blood samples were collected between May 2017 and April 2020 (Fig. 1b). Peripheral blood (20 ml) was collected into tubes containing EDTA (BD, Franklin Lakes, New Jersey). Plasma was separated from the blood within 2 hours of blood collection, as previously described28. cfDNA was extracted using QIAamp MinElute Virus Vacuum Kit (Qiagen, Hilden, Germany) and stored at −80 °C.
ctDNA-ddPCR
Cell-free DNA (cfDNA) was quantified using the RNase P Copy Number Reference Assay (Life Technologies, Carlsbad, California). A total of 3000 genome-equivalents (~10 ng of cfDNA) were analyzed per assay to achieve a detection sensitivity of 0.2%. This detection limit has been assessed by using cell line-derived genomic DNA. A total of 10 ng of input DNA with varying proportions of mutant DNA was serially diluted into wild-type DNA to obtain samples with a mutant abundance of 1%, 0.5%, 0.1%, and 0.05% and subjected to droplet digital PCR (ddPCR). ddPCR was used to quantify the circulating levels of the EGFR activating mutation (EGFR E746_A750 del) and of the resistance mutations (EGFR T790M, PIK3CA E545K, and BRAF V600E). Probes and primers were obtained from BioRad (EGFR E746_A750 del #10041170, EGFR T790M #10040782, PIK3CA E545K #10041188, and BRAF V600E #10040779; Hercules, California). ddPCR was performed on the QX200 Droplet Digital PCR System, and data were analysed using QuantaSoft software (Bio-Rad). ctDNA quantification is presented as fractional abundance (FA–the proportion of the mutant allele in total cfDNA).
Reporting summary
Further information on research design is available in the Nature Research Reporting Summary linked to this article.
Supplementary information
Reporting Summary
Dataset 1
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
These authors contributed equally: Maurício Fernando Silva Almeida Ribeiro, Franciele Hinterholz Knebel.
Supplementary information
The online version contains supplementary material available at 10.1038/s41698-021-00149-4.
Acknowledgements
We thank the clinical research team at Hospital Sírio-Libanês for their assistance collecting samples. We thank Ernande Xavier dos Santos and Elisângela Monteiro for the outstanding technical support. We also thank Mrs. Dina Binzagr and Vivian Hannud for supporting the Translational Research Program at Hospital Sírio Libanês. This work was supported by Ludwig Cancer Research and Fundação de Amparo à Pesquisa do Estado de São Paulo – FAPESP. Lastly, we would like to thank the patient and his family for consenting to the present publication.
Author contributions
Conceptualization by M.F.S.A.R., F.H.K., J.V.M.A., A.A.C., and A.K.; Methodology by F.H.K. and A.A.C.; Resources br M.F.S.A.R., F.H.K., A.A.C., and J.V.M.A.; Writing of the first draft by M.F.S.A., F.H.K., and J.V.M.A.; Review, editing and approval of the final draft by all authors; Supervision by A.A.C. and A.K.; Funding acquisition by F.H.K. and A.A.C. M.F.S.A.R. and F.H.K. are co-first authors of the present work.
Data availability
The datasets that support the findings of this study are not publicly available in order to protect patient privacy. The data will be made available on reasonable request. For data access requests regarding the liquid biopsy (ctDNA quantification) data, please contact Dr. Franciele Knebel, email address: fhknebel@mochsl.org.br. For data access requests regarding the PET-CT high resolution images and PERCIST calculations, please contact Dr. José Marin, email address:jfgmarin@yahoo.com.br. For data access requests regarding the summary of the toxicities arising under osimertinib plus BRAF/MEK inhibition, please contact the corresponding author Dr. Maurício Ribeiro, email address: mauricio.fsaribeiro@hsl.org.br. The data generated and analysed during this study are described in the following metadata record: 10.6084/m9.figshare.1347594635.
Competing interests
The authors declare no competing interests. | DABRAFENIB, OSIMERTINIB, TRAMETINIB | DrugsGivenReaction | CC BY | 33580193 | 19,431,708 | 2021-02-12 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Lung adenocarcinoma'. | Impressive response to dabrafenib, trametinib, and osimertinib in a metastatic EGFR-mutant/BRAF V600E lung adenocarcinoma patient.
The survival outcomes of the FLAURA trial support osimertinib as the new standard of care for untreated patients harboring activating mutations in the epidermal growth factor receptor (EGFR). Despite the initial response, disease progression invariably occurs. Although uncommon, BRAF V600E mutation arises as a unique mechanism of resistance, and thus far, no prospective studies are available to support concurrent EGFR/BRAF blockade. We report a case of impressive radiological and ctDNA response under dabrafenib, trametinib, and osimertinib in an advanced EGFR-mutant lung adenocarcinoma patient who developed BRAF V600E as one of the acquired resistance mechanisms to second-line osimertinib. Moreover, the patient experienced remarkable clinical improvement and good tolerance to combination therapy. The present case suggests the importance of prospective studies evaluating both efficacy and safety of the combination in later line settings and points towards the potential of ctDNA to monitor resistance mechanisms and treatment benefit in clinical practice.
Introduction
The use of osimertinib to target epidermal growth factor receptor (EGFR) has become the standard of care in untreated EGFR-mutant non–small cell lung cancer (NSCLC) patients. Although osimertinib can be highly active, showing more durable outcomes than first-generation tyrosine kinase inhibitors (TKI)1, most tumors invariably become resistant, limiting its long-term clinical benefit. The heterogeneity of resistance mechanisms to osimertinib, including EGFR C797S mutation, EGFR, and MET amplifications, off-target mutations in PIK3CA, KRAS, and HER2 as well as histologic transformation2–9, has stimulated routine performance of repeated biopsies to identify specific underlying mechanisms of resistance throughout the treatment course, and to guide the development of novel therapeutic strategies to overcome and prevent acquired resistance (AR)7,10–13.
Combined targeted therapy (TT) strategies have been increasingly addressed in prospective clinical trials14–17. A phase Ib/II trial reported 47% ORR with a combination of capmatinib (MET inhibitor) plus gefitinib in patients progressing to EGFR TKI and presenting MET dysregulation14. Likewise, SAVANNAH (NCT03778229)15 is an ongoing phase II trial designed to evaluate the efficacy of combination savolitinib and osimertinib in post-osimertinib progression/MET-positive patients. The biomarker-matched study ORCHARD (NCT03944772)16 is also underway to assess the efficacy of several osimertinib-based combinations following disease progression under frontline osimertinib.
BRAF mutations and fusions (i.e. AGK-BRAF, ESYT2-BRAF) have recently emerged as additional mechanisms of AR to third-generation EGFR TKI2,18–20. Studies demonstrating the efficacy of concurrent inhibition of EGFR and BRAF3 or MEK21 in pre-clinical models have raised clinicians’ expectations about overcoming AR by combining TT. Nevertheless, reports of successful combinations of TT for patients harboring BRAF-driven AR to osimertinib are very limited19,22–24 and no prospective data regarding efficacy and safety of BRAF/MEK/EGFR concurrent inhibition are available, with chemotherapy-based regimens remaining the treatment of choice in this unfavorable scenario. Similarly, PIK3CA mutations may also mediate AR to second-line osimertinib in 4–11%, but no clinical reports suggesting potential benefits of blocking these alterations in NSCLC are available6,9,25–27.
The detection of circulating tumor DNA (ctDNA) using liquid biopsies allows noninvasive real-time monitoring of treatment response and early detection of AR to TT, anticipating radiological response and treatment failure28,29. The widespread availability of highly specific and sensitive techniques to quantify ctDNA makes the longitudinal assessment of patients with NSCLC under TKI therapy very attractive. Nonetheless, for NSCLCs, ctDNA evaluation in routine clinical practice is currently limited to the detection of EGFR exon 19 deletion, EGFR L858R and T790M activating mutations29.
A man with metastatic lung adenocarcinoma harboring an EGFR mutation, who had progressed to erlotinib due to the emergence of the EGFR T790M mutation started osimertinib and remained on treatment for 15 months, then developing disease progression (PD). Sequential liquid biopsies were collected to monitor treatment response and disclosed the emergence of BRAF V600E and PIK3CA E545K resistance mutations 4 months before clinical progression. With this result and considering few available reports in the literature, our patient started a triple therapy with osimertinib, dabrafenib (BRAF inhibitor), and trametinib (MEK inhibitor).
Results
Case report
A 50-year-old non-smoker man was diagnosed with a tubule-papillary lung adenocarcinoma metastatic to bones and soft tissue (stage IVB - AJCC 8th edition) in July 2016. Several hypermetabolic bone lesions were observed on staging 18-fluorodeoxyglucose positron-emission tomography (18F-FDG PET-CT) scan at diagnosis. A baseline Next Generation Sequencing (NGS) assay (TruSightTumor™ - Illumina®) of a soft tissue metastasis revealed the presence of an activating EGFR exon 19 deletion (EGFR E746_A750del). No concurrent alterations in BRAF or in PIK3CA were identified at that time. ALK d5f3 immunohistochemistry and fluorescent in situ hybridization (FISH) for ROS1 and RET resulted negative.
In September 2016, the patient started on erlotinib 150 mg once daily (OD), achieving complete metabolic response in February 2017. We started to perform serial blood sample collections for ctDNA analysis in May 2017. In October 2017, oligo-progression (oligo-PD) was observed at the primary site and treated with stereotactic body radiation therapy (SBRT; 3 × 16 Gy). In November 2017, we identified the EGFR T790M and EGFR E746_A750del mutations in patient’s plasma using droplet digital PCR (ddPCR) and these results were confirmed in December 2017 with the Foundation ACT® ctDNA assay. Figure 1a shows all systemic and focal therapies of this patient since diagnosis.Fig. 1 Paired radiological and sequential blood-based ctDNA assessments throughout patient’s treatment demonstrating concordant results.
a Timeline displaying systemic and focal therapies since September 2016. b ctDNA levels in serial plasma samples based on fractional abundance of EGFR T790M, EGFR del19 (E746_A750 del), BRAF V600E, and PIK3CA E545K mutations. #2.4 CN results: EGFR del19 WT/RNAseP copy number ratio. c Paired 18F-FDG PET-CT scan maximum intensity projection (MIP) images displaying tumor burden variations. PD progression of disease, SBRT stereotatic body radiotherapy, Pembro pembrolizumab, CT chemotherapy, DTO dabrafenib, trametinib and osimertinib.
Erlotinib was kept until January 2018 when disease progression in the bones was observed and the fractional abundance (FA) of EGFR E746_A750del and EGFR T790M were 12.0 and 2.3%, respectively (Fig. 1b). The treatment was promptly switched to osimertinib 80 mg OD. Bone partial response (PR) was observed 2 months later, along with a significant drop in the FA of EGFR E746_A750del and EGFR T790M mutations in the plasma (Fig. 1b).
In July 2018, the patient developed oligo-PD in T3 vertebrae, which was treated with SBRT (1 × 20 Gy). At that point, ctDNA analysis revealed an increase in the FA of the EGFR E746_A750del in the plasma, but not in EGFR T790M, suggesting genetic heterogeneity between metastatic lesions. In December 2018, a new oligo-PD in the left iliac bone was detected by 18F-FDG PET-CT scan (Fig. 1c). At that time, ctDNA analysis revealed the emergence of BRAF V600E (FA: 0.4%) and PIK3CA E545K (FA: 0.9%) mutations, as well as a significant increase in the FA of the EGFR E746_A750del (FA: 7.5%) in the plasma. These mutations were also detected in a specimen from an iliac bone biopsy using a NGS assay (TruSightTumor™ - Illumina® - EGFR del19 allelic fraction 81.1%, EGFR del19 amplification – 12 copies, BRAF V600E allelic fraction 17.7% and PIK3CA E545K allelic fraction 32.7%). Even though the patient was treated with SBRT (1 × 18 Gy), he developed new bone metastases in March 2019 (Fig. 1b), consistent with the significant increase in the FA of EGFR E746_A750del, BRAF V600E and EGFR T790M mutations (Fig. 1b). At that specific timepoint and in April 2019, assessments of PIK3CA E545K plasma levels were not obtained due to limited amount of cfDNA.
Between April and September 2019, systemic treatment with carboplatin plus pemetrexed and pembrolizumab provided modest clinical benefit, reducing tumor burden and controlling the disease (Fig. 1b). Due to a new PD observed in October 2019, the treatment was switched to docetaxel monotherapy and, subsequently, to vinorelbine after progression under docetaxel; nonetheless, the patient experienced symptomatic PD in bones, lymph nodes, and lung in late January 2020 (Fig. 1c, 2a–d). He came to the clinic with a Karnofsky performance status of 70%, complaining of fatigue, appetite loss, and severe pain in the hips (despite regular use of 10 mg buprenorphine patch and oxycodone plus acetaminophen PO), which had been preventing him from performing his daily activities due to the inability to stay seated. At that point, high levels of all three resistance mutations were detected in the circulating DNA (PIK3CA E545K FA: 13.2%, BRAF V600E FA: 12.3%, and EGFR T790M FA: 5.3%; Fig. 1b). We also observed an exponential increase in the number of copies of EGFR E746_A750del, suggesting gene amplification (Fig. 1b).Fig. 2 18F-FDG PET-CT scan imaging depicting impressive overall tumor response under dabrafenib, trametinib, and osimertinib in three timepoints.
a–d baseline imaging (January/2020) showing: hypermetabolic spiculated mass (orange circle) in the left-superior lobe measuring 4.4 × 3.3 cm (SUVmax: 9.4); multiple hilar, mediastinal, retroperitoneal, and iliac hypermetabolic lymph nodes (red arrow) measuring up to 2.7 cm (SUVmax: 12.8); several hypermetabolic bone lesions throughout axial and appendicular skeleton (yellow circle); a left iliac bone lesion with signs of periosteal reaction and adjacent soft-tissue infiltration (SUVmax: 11.4). e–h first response evaluation imaging (April/2020) showing considerable partial response in the lung mass, measuring 3.8 × 2.5 cm (SUVmax: 2.7), as well as in several bone lesions, especially in the left-iliac bone (SUVmax: 4.8); complete response in lymph nodes. i–l third response evaluation imaging showing disease progression in the left ischium (SUVmax: 12.5; previous SUVmax: 5.9) (October/2020). SUVmax maximum standard uptake value.
In February 2020, after careful consideration due to patient’s good performance status, normal organ functions, severe pain, motivation and the emergence of BRAF V600E as an AR mechanism, he started on dabrafenib 75 mg twice daily (BID), trametinib 1 mg OD and osimertinib 80 mg OD. The use of a PI3K inhibitor was not considered an option due to the absence of reports in the literature suggesting efficacy of these drugs combined with osimertinib to treat advanced NSCLC patients. Within 2 weeks of treatment, the patient achieved complete resolution of the severe bone pain in the hips with no further need of opioid administration, as well as appetite gain and marked improvement in quality of life, which turned possible for him to resume his daily activities. As adverse events (AE), he experienced grade 1 fatigue, dysgeusia, fever, and nausea, all managed with symptomatic medication. Complete resolution of the fever occurred spontaneously within two weeks of treatment. An attempt to increase dosages of dabrafenib to 150 mg BID and trametinib 2 mg OD resulted unsuccessful due to persistent grade 2 fatigue. In April 2020, a new 18F-FDG PET-CT scan disclosed a complete response in lymph nodes and a dramatic PR in the lung and bones (PERCIST 1.1 evaluating 5 target lesions: Δ- 67%; Figs. 1c and 2e–h). Detection of EGFR T790M, BRAF V600E, and PIK3CA E545K mutations became negative in April 2020, while EGFR E746_A750del despite marginally positive, presented marked reduction (Fig. 1b) suggesting that combination dabrafenib, trametinib, and osimertinib might be an effective strategy to overcome PIK3CA E545K and BRAF V600E-driven resistance to osimertinib in advanced EGFR-mutant NSCLC patients. The patient remained in response until October 2020, when he developed asymptomatic bone PD in lumbar spine, left ischium, and right iliac bone, 8 months after starting this combination therapy (Figs. 1c and 2i–l). A FoundationOne®Liquid CDx plasma NGS disclosed the following alterations: CHCHD3-BRAF fusion, BRAF V600E, EGFR E746_750del, EGFR amplification, PIK3CA E545K, MAP2K2 (MEK2) C125S, MTAP rearrangement intron 5, TP53 V197M, and TP53 S241A. Owing to the considerable clinical benefit, good tolerance and lack of systemic treatment options, we decided to keep the patient on treatment and increase the doses of dabrafenib (150 mg BID) and trametinib (2 mg OD) every other day. He remains asymptomatic and tolerating well the proposed dose adjustment.
Discussion
Several studies have recently highlighted the importance of considering genes of interest within the context of commonly co-occurring mutations9,30. For example, as described by Blakely et al. through performing a cfDNA NGS analysis of 1,122 advanced stage EGFR-mutant NSCLCs, in ~93% of the patients, at least one more variables with known or likely known functional properties were present, disclosing the molecular complexity of this oncogenic driver and suggesting an association of co-occuring genomic alterations with TKI response and clinical outcomes9. In addition, Roper and colleagues reported the identification of at least two co-existing AR mechanisms in 73% of patients treated with osimertinib, as well as 6–23 different subclones per individual in a phylogenetic analysis performed in multiple metastatic sites of 15 individuals30. In the same publication, the authors also called the attention for a high incidence of acquired EGFR amplifications in post-osimertinib patients, which suggests maintenance of EGFR central role in the setting of progression, as previously reported by our group28 and also detected in the present case.
Although uncommon, BRAF V600E mediates AR in approximately 3% of the patients under second-line osimertinib6,20 and little is known about the efficacy of combined TT in this population3,21–23. Existing reports lack details regarding objective responses using standardized radiological criteria (i.e., RECIST 1.1; PERCIST) and the dynamics of resistance mechanisms through longitudinal ctDNA measurements22,23. Huang and colleagues22 described a case of an EGFRdel19/T790M + NSCLC patient who developed BRAF V600E-driven AR after second-line osimertinib and achieved stable disease under dabrafenib 150 mg BID, trametinib 1 mg OD and osimertinib 80 mg OD, with ongoing disease control 7.4 months after. Zhou and colleagues23 also reported their experience with dabrafenib 150 mg BID, trametinib 2 mg OD and osimertinib 80 mg OD leading to tumor reduction within 6 weeks of treatment, along with grade 2 rash and decreased appetite as AEs. Similarly, Meng et al. reported two cases treated with this triple regimen31. The first patient discontinued therapy after one month due to severe pneumonitis; the second one presented tumor response under dabrafenib 50 mg BID, trametinib 0.5 mg OD, and osimertinib 80 mg OD, with progression-free survival of 14 months. According to the authors, this reduced dose was prescribed owing to a grade 2 pyrexia, nausea, and vomiting under higher dabrafenib and trametinib doses. Dagogo-Jack and colleagues19 also described a successful case of combined EGFR/MAP kinase pathway blockade with osimertinib 80 mg OD and trametinib 1 mg OD; as treatment-related AEs, their patient experienced grade 2 diarrhea and fatigue, along with grade 1 rash and gastrointestinal bleeding. Nevertheless, it is worth highlighting the rapid clinical improvement, the remarkable radiologically confirmed objective response, as well as the good tolerance observed in this case even using only half standard dose of dabrafenib and trametinib approved for NSCLCs harboring BRAF V600E mutations. Since data regarding the efficacy of these combined approaches, the optimal drug association and dosing, as well as the toxicity profile are conflicting and largely unknown, further investigation into the mechanistic basis of this association represents an important priority. The Table 1 summarizes the toxicities arising under osimertinib plus BRAF/MEK inhibition and reported in the literature.Table 1 Summary of the toxicities arising under osimertinib plus BRAF/MEK inhibition and reported in the literature.
Reference Treatment Anorexia Nausea Vomiting Diarrhoea Fatigue Rash GI bleeding AST/ALT elevation Paroniquia Pyrexia Initial dose Dose reduction
Meng et al. D + T + O NR G2 G2 G2 NR NR NR NR NR G2 D (150 mg BID) + T (2 mg OD) + O (80 mg OD) D (50 mg BID) + T (0.5 mg/day) + O (80 mg/day)
Dagogo-Jack et al. T + O NR NR NR G2 G2 G1 G1 G1 NR NR T (1 mg OD) + O (80 mg OD) Not needed
Huang et al. D + T + O NR NR NR G1 NR NR NR NR G1 NR D (150 mg BID) + T (1 mg OD) + O (80 mg OD) Not needed
Zhou et al. D + T + O G1 NR NR NR NR G2 NR NR NR NR D (150 mg BID) + T (2 mg OD) + O (80 mg OD) Not needed
D dabrafenib, T trametinib, O osimertinib, NR not reported.
Liquid biopsy is emerging as an important diagnostic and predictive tool in the treatment of NSCLCs. The accurate identification of predictive genetic alterations is important for both patients’ management and the understanding of clonal evolution and AR to different therapies9,29,32. Also, circulating biomarkers from multiple disease sites better reflect systemic tumor burden, including alterations from genetically different metastatic lesions, which may be missed with single-site tissue biopsies8. Here, we were able to identify and monitor multiple concurrent mechanisms of resistance throughout the entire patient’s treatment. As shown in Fig. 1b, variations in blood ctDNA levels exhibited a positive correlation with imaging findings, even in situations involving focal treatments for oligo-PD. The observed concordance between radiological PD or response and ctDNA measurements also points towards its great potential to be incorporated into clinical practice to anticipate radiologic findings in a more effective manner.
Along with BRAF V600E mutation, the presence of PIK3CA E545K mutation is also associated with AR to osimertinib6,9,25,27. However, the clinical implication of concomitant targeting of EGFR and PI3K remains unclear. Whereas an alternative argument for the observed response could be a rechallenge following a long time upon off-osimertinib, in our opinion, this is an unlikely explanation33. Interestingly, the presence of a BRAF fusion along with a MEK2 mutation identified through plasma NGS in the setting of PD might represent a mechanism of resistance to this triple regimen34. Nonetheless, since pretreatment blood samples had not been analyzed using a similar methodology, this hypothesis deserves careful interpretation.
Increased understanding of the relationship of concurring genomic alterations in EGFR-mutant NSCLC may enable new therapeutic opportunities following disease progression to osimertinib. Here, we reported an impressive objective response to dabrafenib, trametinib, and osimertinib with concordant decrease in plasma ctDNA levels in a metastatic lung cancer patient harboring EGFR E746_A750del, BRAF V600E, and PIK3CA E545K activating mutations. This case report leads to a greater understanding of the currently limited literature regarding the management of EGFR-mutant NSCLC patients with acquired BRAFV600E mutation, since it reports a successful attempt to target both alterations concurrently while providing concordant and interesting data of serial ctDNA assessments throughout the entire treatment. Further investigation to optimize the efficacy and mitigate the toxicity profile of this drug association represents an important issue.
Methods
Patient
This study was approved by Hospital Sírio-Libanês Ethics Committee (HSL-RC 2020-16). The patient provided written informed consent for blood collection, ctDNA analysis and publication of this report.
Sample collection and plasma DNA extraction
Serial blood samples were collected between May 2017 and April 2020 (Fig. 1b). Peripheral blood (20 ml) was collected into tubes containing EDTA (BD, Franklin Lakes, New Jersey). Plasma was separated from the blood within 2 hours of blood collection, as previously described28. cfDNA was extracted using QIAamp MinElute Virus Vacuum Kit (Qiagen, Hilden, Germany) and stored at −80 °C.
ctDNA-ddPCR
Cell-free DNA (cfDNA) was quantified using the RNase P Copy Number Reference Assay (Life Technologies, Carlsbad, California). A total of 3000 genome-equivalents (~10 ng of cfDNA) were analyzed per assay to achieve a detection sensitivity of 0.2%. This detection limit has been assessed by using cell line-derived genomic DNA. A total of 10 ng of input DNA with varying proportions of mutant DNA was serially diluted into wild-type DNA to obtain samples with a mutant abundance of 1%, 0.5%, 0.1%, and 0.05% and subjected to droplet digital PCR (ddPCR). ddPCR was used to quantify the circulating levels of the EGFR activating mutation (EGFR E746_A750 del) and of the resistance mutations (EGFR T790M, PIK3CA E545K, and BRAF V600E). Probes and primers were obtained from BioRad (EGFR E746_A750 del #10041170, EGFR T790M #10040782, PIK3CA E545K #10041188, and BRAF V600E #10040779; Hercules, California). ddPCR was performed on the QX200 Droplet Digital PCR System, and data were analysed using QuantaSoft software (Bio-Rad). ctDNA quantification is presented as fractional abundance (FA–the proportion of the mutant allele in total cfDNA).
Reporting summary
Further information on research design is available in the Nature Research Reporting Summary linked to this article.
Supplementary information
Reporting Summary
Dataset 1
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
These authors contributed equally: Maurício Fernando Silva Almeida Ribeiro, Franciele Hinterholz Knebel.
Supplementary information
The online version contains supplementary material available at 10.1038/s41698-021-00149-4.
Acknowledgements
We thank the clinical research team at Hospital Sírio-Libanês for their assistance collecting samples. We thank Ernande Xavier dos Santos and Elisângela Monteiro for the outstanding technical support. We also thank Mrs. Dina Binzagr and Vivian Hannud for supporting the Translational Research Program at Hospital Sírio Libanês. This work was supported by Ludwig Cancer Research and Fundação de Amparo à Pesquisa do Estado de São Paulo – FAPESP. Lastly, we would like to thank the patient and his family for consenting to the present publication.
Author contributions
Conceptualization by M.F.S.A.R., F.H.K., J.V.M.A., A.A.C., and A.K.; Methodology by F.H.K. and A.A.C.; Resources br M.F.S.A.R., F.H.K., A.A.C., and J.V.M.A.; Writing of the first draft by M.F.S.A., F.H.K., and J.V.M.A.; Review, editing and approval of the final draft by all authors; Supervision by A.A.C. and A.K.; Funding acquisition by F.H.K. and A.A.C. M.F.S.A.R. and F.H.K. are co-first authors of the present work.
Data availability
The datasets that support the findings of this study are not publicly available in order to protect patient privacy. The data will be made available on reasonable request. For data access requests regarding the liquid biopsy (ctDNA quantification) data, please contact Dr. Franciele Knebel, email address: fhknebel@mochsl.org.br. For data access requests regarding the PET-CT high resolution images and PERCIST calculations, please contact Dr. José Marin, email address:jfgmarin@yahoo.com.br. For data access requests regarding the summary of the toxicities arising under osimertinib plus BRAF/MEK inhibition, please contact the corresponding author Dr. Maurício Ribeiro, email address: mauricio.fsaribeiro@hsl.org.br. The data generated and analysed during this study are described in the following metadata record: 10.6084/m9.figshare.1347594635.
Competing interests
The authors declare no competing interests. | DABRAFENIB, OSIMERTINIB, TRAMETINIB | DrugsGivenReaction | CC BY | 33580193 | 19,431,708 | 2021-02-12 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Malignant neoplasm progression'. | Impressive response to dabrafenib, trametinib, and osimertinib in a metastatic EGFR-mutant/BRAF V600E lung adenocarcinoma patient.
The survival outcomes of the FLAURA trial support osimertinib as the new standard of care for untreated patients harboring activating mutations in the epidermal growth factor receptor (EGFR). Despite the initial response, disease progression invariably occurs. Although uncommon, BRAF V600E mutation arises as a unique mechanism of resistance, and thus far, no prospective studies are available to support concurrent EGFR/BRAF blockade. We report a case of impressive radiological and ctDNA response under dabrafenib, trametinib, and osimertinib in an advanced EGFR-mutant lung adenocarcinoma patient who developed BRAF V600E as one of the acquired resistance mechanisms to second-line osimertinib. Moreover, the patient experienced remarkable clinical improvement and good tolerance to combination therapy. The present case suggests the importance of prospective studies evaluating both efficacy and safety of the combination in later line settings and points towards the potential of ctDNA to monitor resistance mechanisms and treatment benefit in clinical practice.
Introduction
The use of osimertinib to target epidermal growth factor receptor (EGFR) has become the standard of care in untreated EGFR-mutant non–small cell lung cancer (NSCLC) patients. Although osimertinib can be highly active, showing more durable outcomes than first-generation tyrosine kinase inhibitors (TKI)1, most tumors invariably become resistant, limiting its long-term clinical benefit. The heterogeneity of resistance mechanisms to osimertinib, including EGFR C797S mutation, EGFR, and MET amplifications, off-target mutations in PIK3CA, KRAS, and HER2 as well as histologic transformation2–9, has stimulated routine performance of repeated biopsies to identify specific underlying mechanisms of resistance throughout the treatment course, and to guide the development of novel therapeutic strategies to overcome and prevent acquired resistance (AR)7,10–13.
Combined targeted therapy (TT) strategies have been increasingly addressed in prospective clinical trials14–17. A phase Ib/II trial reported 47% ORR with a combination of capmatinib (MET inhibitor) plus gefitinib in patients progressing to EGFR TKI and presenting MET dysregulation14. Likewise, SAVANNAH (NCT03778229)15 is an ongoing phase II trial designed to evaluate the efficacy of combination savolitinib and osimertinib in post-osimertinib progression/MET-positive patients. The biomarker-matched study ORCHARD (NCT03944772)16 is also underway to assess the efficacy of several osimertinib-based combinations following disease progression under frontline osimertinib.
BRAF mutations and fusions (i.e. AGK-BRAF, ESYT2-BRAF) have recently emerged as additional mechanisms of AR to third-generation EGFR TKI2,18–20. Studies demonstrating the efficacy of concurrent inhibition of EGFR and BRAF3 or MEK21 in pre-clinical models have raised clinicians’ expectations about overcoming AR by combining TT. Nevertheless, reports of successful combinations of TT for patients harboring BRAF-driven AR to osimertinib are very limited19,22–24 and no prospective data regarding efficacy and safety of BRAF/MEK/EGFR concurrent inhibition are available, with chemotherapy-based regimens remaining the treatment of choice in this unfavorable scenario. Similarly, PIK3CA mutations may also mediate AR to second-line osimertinib in 4–11%, but no clinical reports suggesting potential benefits of blocking these alterations in NSCLC are available6,9,25–27.
The detection of circulating tumor DNA (ctDNA) using liquid biopsies allows noninvasive real-time monitoring of treatment response and early detection of AR to TT, anticipating radiological response and treatment failure28,29. The widespread availability of highly specific and sensitive techniques to quantify ctDNA makes the longitudinal assessment of patients with NSCLC under TKI therapy very attractive. Nonetheless, for NSCLCs, ctDNA evaluation in routine clinical practice is currently limited to the detection of EGFR exon 19 deletion, EGFR L858R and T790M activating mutations29.
A man with metastatic lung adenocarcinoma harboring an EGFR mutation, who had progressed to erlotinib due to the emergence of the EGFR T790M mutation started osimertinib and remained on treatment for 15 months, then developing disease progression (PD). Sequential liquid biopsies were collected to monitor treatment response and disclosed the emergence of BRAF V600E and PIK3CA E545K resistance mutations 4 months before clinical progression. With this result and considering few available reports in the literature, our patient started a triple therapy with osimertinib, dabrafenib (BRAF inhibitor), and trametinib (MEK inhibitor).
Results
Case report
A 50-year-old non-smoker man was diagnosed with a tubule-papillary lung adenocarcinoma metastatic to bones and soft tissue (stage IVB - AJCC 8th edition) in July 2016. Several hypermetabolic bone lesions were observed on staging 18-fluorodeoxyglucose positron-emission tomography (18F-FDG PET-CT) scan at diagnosis. A baseline Next Generation Sequencing (NGS) assay (TruSightTumor™ - Illumina®) of a soft tissue metastasis revealed the presence of an activating EGFR exon 19 deletion (EGFR E746_A750del). No concurrent alterations in BRAF or in PIK3CA were identified at that time. ALK d5f3 immunohistochemistry and fluorescent in situ hybridization (FISH) for ROS1 and RET resulted negative.
In September 2016, the patient started on erlotinib 150 mg once daily (OD), achieving complete metabolic response in February 2017. We started to perform serial blood sample collections for ctDNA analysis in May 2017. In October 2017, oligo-progression (oligo-PD) was observed at the primary site and treated with stereotactic body radiation therapy (SBRT; 3 × 16 Gy). In November 2017, we identified the EGFR T790M and EGFR E746_A750del mutations in patient’s plasma using droplet digital PCR (ddPCR) and these results were confirmed in December 2017 with the Foundation ACT® ctDNA assay. Figure 1a shows all systemic and focal therapies of this patient since diagnosis.Fig. 1 Paired radiological and sequential blood-based ctDNA assessments throughout patient’s treatment demonstrating concordant results.
a Timeline displaying systemic and focal therapies since September 2016. b ctDNA levels in serial plasma samples based on fractional abundance of EGFR T790M, EGFR del19 (E746_A750 del), BRAF V600E, and PIK3CA E545K mutations. #2.4 CN results: EGFR del19 WT/RNAseP copy number ratio. c Paired 18F-FDG PET-CT scan maximum intensity projection (MIP) images displaying tumor burden variations. PD progression of disease, SBRT stereotatic body radiotherapy, Pembro pembrolizumab, CT chemotherapy, DTO dabrafenib, trametinib and osimertinib.
Erlotinib was kept until January 2018 when disease progression in the bones was observed and the fractional abundance (FA) of EGFR E746_A750del and EGFR T790M were 12.0 and 2.3%, respectively (Fig. 1b). The treatment was promptly switched to osimertinib 80 mg OD. Bone partial response (PR) was observed 2 months later, along with a significant drop in the FA of EGFR E746_A750del and EGFR T790M mutations in the plasma (Fig. 1b).
In July 2018, the patient developed oligo-PD in T3 vertebrae, which was treated with SBRT (1 × 20 Gy). At that point, ctDNA analysis revealed an increase in the FA of the EGFR E746_A750del in the plasma, but not in EGFR T790M, suggesting genetic heterogeneity between metastatic lesions. In December 2018, a new oligo-PD in the left iliac bone was detected by 18F-FDG PET-CT scan (Fig. 1c). At that time, ctDNA analysis revealed the emergence of BRAF V600E (FA: 0.4%) and PIK3CA E545K (FA: 0.9%) mutations, as well as a significant increase in the FA of the EGFR E746_A750del (FA: 7.5%) in the plasma. These mutations were also detected in a specimen from an iliac bone biopsy using a NGS assay (TruSightTumor™ - Illumina® - EGFR del19 allelic fraction 81.1%, EGFR del19 amplification – 12 copies, BRAF V600E allelic fraction 17.7% and PIK3CA E545K allelic fraction 32.7%). Even though the patient was treated with SBRT (1 × 18 Gy), he developed new bone metastases in March 2019 (Fig. 1b), consistent with the significant increase in the FA of EGFR E746_A750del, BRAF V600E and EGFR T790M mutations (Fig. 1b). At that specific timepoint and in April 2019, assessments of PIK3CA E545K plasma levels were not obtained due to limited amount of cfDNA.
Between April and September 2019, systemic treatment with carboplatin plus pemetrexed and pembrolizumab provided modest clinical benefit, reducing tumor burden and controlling the disease (Fig. 1b). Due to a new PD observed in October 2019, the treatment was switched to docetaxel monotherapy and, subsequently, to vinorelbine after progression under docetaxel; nonetheless, the patient experienced symptomatic PD in bones, lymph nodes, and lung in late January 2020 (Fig. 1c, 2a–d). He came to the clinic with a Karnofsky performance status of 70%, complaining of fatigue, appetite loss, and severe pain in the hips (despite regular use of 10 mg buprenorphine patch and oxycodone plus acetaminophen PO), which had been preventing him from performing his daily activities due to the inability to stay seated. At that point, high levels of all three resistance mutations were detected in the circulating DNA (PIK3CA E545K FA: 13.2%, BRAF V600E FA: 12.3%, and EGFR T790M FA: 5.3%; Fig. 1b). We also observed an exponential increase in the number of copies of EGFR E746_A750del, suggesting gene amplification (Fig. 1b).Fig. 2 18F-FDG PET-CT scan imaging depicting impressive overall tumor response under dabrafenib, trametinib, and osimertinib in three timepoints.
a–d baseline imaging (January/2020) showing: hypermetabolic spiculated mass (orange circle) in the left-superior lobe measuring 4.4 × 3.3 cm (SUVmax: 9.4); multiple hilar, mediastinal, retroperitoneal, and iliac hypermetabolic lymph nodes (red arrow) measuring up to 2.7 cm (SUVmax: 12.8); several hypermetabolic bone lesions throughout axial and appendicular skeleton (yellow circle); a left iliac bone lesion with signs of periosteal reaction and adjacent soft-tissue infiltration (SUVmax: 11.4). e–h first response evaluation imaging (April/2020) showing considerable partial response in the lung mass, measuring 3.8 × 2.5 cm (SUVmax: 2.7), as well as in several bone lesions, especially in the left-iliac bone (SUVmax: 4.8); complete response in lymph nodes. i–l third response evaluation imaging showing disease progression in the left ischium (SUVmax: 12.5; previous SUVmax: 5.9) (October/2020). SUVmax maximum standard uptake value.
In February 2020, after careful consideration due to patient’s good performance status, normal organ functions, severe pain, motivation and the emergence of BRAF V600E as an AR mechanism, he started on dabrafenib 75 mg twice daily (BID), trametinib 1 mg OD and osimertinib 80 mg OD. The use of a PI3K inhibitor was not considered an option due to the absence of reports in the literature suggesting efficacy of these drugs combined with osimertinib to treat advanced NSCLC patients. Within 2 weeks of treatment, the patient achieved complete resolution of the severe bone pain in the hips with no further need of opioid administration, as well as appetite gain and marked improvement in quality of life, which turned possible for him to resume his daily activities. As adverse events (AE), he experienced grade 1 fatigue, dysgeusia, fever, and nausea, all managed with symptomatic medication. Complete resolution of the fever occurred spontaneously within two weeks of treatment. An attempt to increase dosages of dabrafenib to 150 mg BID and trametinib 2 mg OD resulted unsuccessful due to persistent grade 2 fatigue. In April 2020, a new 18F-FDG PET-CT scan disclosed a complete response in lymph nodes and a dramatic PR in the lung and bones (PERCIST 1.1 evaluating 5 target lesions: Δ- 67%; Figs. 1c and 2e–h). Detection of EGFR T790M, BRAF V600E, and PIK3CA E545K mutations became negative in April 2020, while EGFR E746_A750del despite marginally positive, presented marked reduction (Fig. 1b) suggesting that combination dabrafenib, trametinib, and osimertinib might be an effective strategy to overcome PIK3CA E545K and BRAF V600E-driven resistance to osimertinib in advanced EGFR-mutant NSCLC patients. The patient remained in response until October 2020, when he developed asymptomatic bone PD in lumbar spine, left ischium, and right iliac bone, 8 months after starting this combination therapy (Figs. 1c and 2i–l). A FoundationOne®Liquid CDx plasma NGS disclosed the following alterations: CHCHD3-BRAF fusion, BRAF V600E, EGFR E746_750del, EGFR amplification, PIK3CA E545K, MAP2K2 (MEK2) C125S, MTAP rearrangement intron 5, TP53 V197M, and TP53 S241A. Owing to the considerable clinical benefit, good tolerance and lack of systemic treatment options, we decided to keep the patient on treatment and increase the doses of dabrafenib (150 mg BID) and trametinib (2 mg OD) every other day. He remains asymptomatic and tolerating well the proposed dose adjustment.
Discussion
Several studies have recently highlighted the importance of considering genes of interest within the context of commonly co-occurring mutations9,30. For example, as described by Blakely et al. through performing a cfDNA NGS analysis of 1,122 advanced stage EGFR-mutant NSCLCs, in ~93% of the patients, at least one more variables with known or likely known functional properties were present, disclosing the molecular complexity of this oncogenic driver and suggesting an association of co-occuring genomic alterations with TKI response and clinical outcomes9. In addition, Roper and colleagues reported the identification of at least two co-existing AR mechanisms in 73% of patients treated with osimertinib, as well as 6–23 different subclones per individual in a phylogenetic analysis performed in multiple metastatic sites of 15 individuals30. In the same publication, the authors also called the attention for a high incidence of acquired EGFR amplifications in post-osimertinib patients, which suggests maintenance of EGFR central role in the setting of progression, as previously reported by our group28 and also detected in the present case.
Although uncommon, BRAF V600E mediates AR in approximately 3% of the patients under second-line osimertinib6,20 and little is known about the efficacy of combined TT in this population3,21–23. Existing reports lack details regarding objective responses using standardized radiological criteria (i.e., RECIST 1.1; PERCIST) and the dynamics of resistance mechanisms through longitudinal ctDNA measurements22,23. Huang and colleagues22 described a case of an EGFRdel19/T790M + NSCLC patient who developed BRAF V600E-driven AR after second-line osimertinib and achieved stable disease under dabrafenib 150 mg BID, trametinib 1 mg OD and osimertinib 80 mg OD, with ongoing disease control 7.4 months after. Zhou and colleagues23 also reported their experience with dabrafenib 150 mg BID, trametinib 2 mg OD and osimertinib 80 mg OD leading to tumor reduction within 6 weeks of treatment, along with grade 2 rash and decreased appetite as AEs. Similarly, Meng et al. reported two cases treated with this triple regimen31. The first patient discontinued therapy after one month due to severe pneumonitis; the second one presented tumor response under dabrafenib 50 mg BID, trametinib 0.5 mg OD, and osimertinib 80 mg OD, with progression-free survival of 14 months. According to the authors, this reduced dose was prescribed owing to a grade 2 pyrexia, nausea, and vomiting under higher dabrafenib and trametinib doses. Dagogo-Jack and colleagues19 also described a successful case of combined EGFR/MAP kinase pathway blockade with osimertinib 80 mg OD and trametinib 1 mg OD; as treatment-related AEs, their patient experienced grade 2 diarrhea and fatigue, along with grade 1 rash and gastrointestinal bleeding. Nevertheless, it is worth highlighting the rapid clinical improvement, the remarkable radiologically confirmed objective response, as well as the good tolerance observed in this case even using only half standard dose of dabrafenib and trametinib approved for NSCLCs harboring BRAF V600E mutations. Since data regarding the efficacy of these combined approaches, the optimal drug association and dosing, as well as the toxicity profile are conflicting and largely unknown, further investigation into the mechanistic basis of this association represents an important priority. The Table 1 summarizes the toxicities arising under osimertinib plus BRAF/MEK inhibition and reported in the literature.Table 1 Summary of the toxicities arising under osimertinib plus BRAF/MEK inhibition and reported in the literature.
Reference Treatment Anorexia Nausea Vomiting Diarrhoea Fatigue Rash GI bleeding AST/ALT elevation Paroniquia Pyrexia Initial dose Dose reduction
Meng et al. D + T + O NR G2 G2 G2 NR NR NR NR NR G2 D (150 mg BID) + T (2 mg OD) + O (80 mg OD) D (50 mg BID) + T (0.5 mg/day) + O (80 mg/day)
Dagogo-Jack et al. T + O NR NR NR G2 G2 G1 G1 G1 NR NR T (1 mg OD) + O (80 mg OD) Not needed
Huang et al. D + T + O NR NR NR G1 NR NR NR NR G1 NR D (150 mg BID) + T (1 mg OD) + O (80 mg OD) Not needed
Zhou et al. D + T + O G1 NR NR NR NR G2 NR NR NR NR D (150 mg BID) + T (2 mg OD) + O (80 mg OD) Not needed
D dabrafenib, T trametinib, O osimertinib, NR not reported.
Liquid biopsy is emerging as an important diagnostic and predictive tool in the treatment of NSCLCs. The accurate identification of predictive genetic alterations is important for both patients’ management and the understanding of clonal evolution and AR to different therapies9,29,32. Also, circulating biomarkers from multiple disease sites better reflect systemic tumor burden, including alterations from genetically different metastatic lesions, which may be missed with single-site tissue biopsies8. Here, we were able to identify and monitor multiple concurrent mechanisms of resistance throughout the entire patient’s treatment. As shown in Fig. 1b, variations in blood ctDNA levels exhibited a positive correlation with imaging findings, even in situations involving focal treatments for oligo-PD. The observed concordance between radiological PD or response and ctDNA measurements also points towards its great potential to be incorporated into clinical practice to anticipate radiologic findings in a more effective manner.
Along with BRAF V600E mutation, the presence of PIK3CA E545K mutation is also associated with AR to osimertinib6,9,25,27. However, the clinical implication of concomitant targeting of EGFR and PI3K remains unclear. Whereas an alternative argument for the observed response could be a rechallenge following a long time upon off-osimertinib, in our opinion, this is an unlikely explanation33. Interestingly, the presence of a BRAF fusion along with a MEK2 mutation identified through plasma NGS in the setting of PD might represent a mechanism of resistance to this triple regimen34. Nonetheless, since pretreatment blood samples had not been analyzed using a similar methodology, this hypothesis deserves careful interpretation.
Increased understanding of the relationship of concurring genomic alterations in EGFR-mutant NSCLC may enable new therapeutic opportunities following disease progression to osimertinib. Here, we reported an impressive objective response to dabrafenib, trametinib, and osimertinib with concordant decrease in plasma ctDNA levels in a metastatic lung cancer patient harboring EGFR E746_A750del, BRAF V600E, and PIK3CA E545K activating mutations. This case report leads to a greater understanding of the currently limited literature regarding the management of EGFR-mutant NSCLC patients with acquired BRAFV600E mutation, since it reports a successful attempt to target both alterations concurrently while providing concordant and interesting data of serial ctDNA assessments throughout the entire treatment. Further investigation to optimize the efficacy and mitigate the toxicity profile of this drug association represents an important issue.
Methods
Patient
This study was approved by Hospital Sírio-Libanês Ethics Committee (HSL-RC 2020-16). The patient provided written informed consent for blood collection, ctDNA analysis and publication of this report.
Sample collection and plasma DNA extraction
Serial blood samples were collected between May 2017 and April 2020 (Fig. 1b). Peripheral blood (20 ml) was collected into tubes containing EDTA (BD, Franklin Lakes, New Jersey). Plasma was separated from the blood within 2 hours of blood collection, as previously described28. cfDNA was extracted using QIAamp MinElute Virus Vacuum Kit (Qiagen, Hilden, Germany) and stored at −80 °C.
ctDNA-ddPCR
Cell-free DNA (cfDNA) was quantified using the RNase P Copy Number Reference Assay (Life Technologies, Carlsbad, California). A total of 3000 genome-equivalents (~10 ng of cfDNA) were analyzed per assay to achieve a detection sensitivity of 0.2%. This detection limit has been assessed by using cell line-derived genomic DNA. A total of 10 ng of input DNA with varying proportions of mutant DNA was serially diluted into wild-type DNA to obtain samples with a mutant abundance of 1%, 0.5%, 0.1%, and 0.05% and subjected to droplet digital PCR (ddPCR). ddPCR was used to quantify the circulating levels of the EGFR activating mutation (EGFR E746_A750 del) and of the resistance mutations (EGFR T790M, PIK3CA E545K, and BRAF V600E). Probes and primers were obtained from BioRad (EGFR E746_A750 del #10041170, EGFR T790M #10040782, PIK3CA E545K #10041188, and BRAF V600E #10040779; Hercules, California). ddPCR was performed on the QX200 Droplet Digital PCR System, and data were analysed using QuantaSoft software (Bio-Rad). ctDNA quantification is presented as fractional abundance (FA–the proportion of the mutant allele in total cfDNA).
Reporting summary
Further information on research design is available in the Nature Research Reporting Summary linked to this article.
Supplementary information
Reporting Summary
Dataset 1
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
These authors contributed equally: Maurício Fernando Silva Almeida Ribeiro, Franciele Hinterholz Knebel.
Supplementary information
The online version contains supplementary material available at 10.1038/s41698-021-00149-4.
Acknowledgements
We thank the clinical research team at Hospital Sírio-Libanês for their assistance collecting samples. We thank Ernande Xavier dos Santos and Elisângela Monteiro for the outstanding technical support. We also thank Mrs. Dina Binzagr and Vivian Hannud for supporting the Translational Research Program at Hospital Sírio Libanês. This work was supported by Ludwig Cancer Research and Fundação de Amparo à Pesquisa do Estado de São Paulo – FAPESP. Lastly, we would like to thank the patient and his family for consenting to the present publication.
Author contributions
Conceptualization by M.F.S.A.R., F.H.K., J.V.M.A., A.A.C., and A.K.; Methodology by F.H.K. and A.A.C.; Resources br M.F.S.A.R., F.H.K., A.A.C., and J.V.M.A.; Writing of the first draft by M.F.S.A., F.H.K., and J.V.M.A.; Review, editing and approval of the final draft by all authors; Supervision by A.A.C. and A.K.; Funding acquisition by F.H.K. and A.A.C. M.F.S.A.R. and F.H.K. are co-first authors of the present work.
Data availability
The datasets that support the findings of this study are not publicly available in order to protect patient privacy. The data will be made available on reasonable request. For data access requests regarding the liquid biopsy (ctDNA quantification) data, please contact Dr. Franciele Knebel, email address: fhknebel@mochsl.org.br. For data access requests regarding the PET-CT high resolution images and PERCIST calculations, please contact Dr. José Marin, email address:jfgmarin@yahoo.com.br. For data access requests regarding the summary of the toxicities arising under osimertinib plus BRAF/MEK inhibition, please contact the corresponding author Dr. Maurício Ribeiro, email address: mauricio.fsaribeiro@hsl.org.br. The data generated and analysed during this study are described in the following metadata record: 10.6084/m9.figshare.1347594635.
Competing interests
The authors declare no competing interests. | DABRAFENIB, OSIMERTINIB, TRAMETINIB | DrugsGivenReaction | CC BY | 33580193 | 19,431,708 | 2021-02-12 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Nausea'. | Impressive response to dabrafenib, trametinib, and osimertinib in a metastatic EGFR-mutant/BRAF V600E lung adenocarcinoma patient.
The survival outcomes of the FLAURA trial support osimertinib as the new standard of care for untreated patients harboring activating mutations in the epidermal growth factor receptor (EGFR). Despite the initial response, disease progression invariably occurs. Although uncommon, BRAF V600E mutation arises as a unique mechanism of resistance, and thus far, no prospective studies are available to support concurrent EGFR/BRAF blockade. We report a case of impressive radiological and ctDNA response under dabrafenib, trametinib, and osimertinib in an advanced EGFR-mutant lung adenocarcinoma patient who developed BRAF V600E as one of the acquired resistance mechanisms to second-line osimertinib. Moreover, the patient experienced remarkable clinical improvement and good tolerance to combination therapy. The present case suggests the importance of prospective studies evaluating both efficacy and safety of the combination in later line settings and points towards the potential of ctDNA to monitor resistance mechanisms and treatment benefit in clinical practice.
Introduction
The use of osimertinib to target epidermal growth factor receptor (EGFR) has become the standard of care in untreated EGFR-mutant non–small cell lung cancer (NSCLC) patients. Although osimertinib can be highly active, showing more durable outcomes than first-generation tyrosine kinase inhibitors (TKI)1, most tumors invariably become resistant, limiting its long-term clinical benefit. The heterogeneity of resistance mechanisms to osimertinib, including EGFR C797S mutation, EGFR, and MET amplifications, off-target mutations in PIK3CA, KRAS, and HER2 as well as histologic transformation2–9, has stimulated routine performance of repeated biopsies to identify specific underlying mechanisms of resistance throughout the treatment course, and to guide the development of novel therapeutic strategies to overcome and prevent acquired resistance (AR)7,10–13.
Combined targeted therapy (TT) strategies have been increasingly addressed in prospective clinical trials14–17. A phase Ib/II trial reported 47% ORR with a combination of capmatinib (MET inhibitor) plus gefitinib in patients progressing to EGFR TKI and presenting MET dysregulation14. Likewise, SAVANNAH (NCT03778229)15 is an ongoing phase II trial designed to evaluate the efficacy of combination savolitinib and osimertinib in post-osimertinib progression/MET-positive patients. The biomarker-matched study ORCHARD (NCT03944772)16 is also underway to assess the efficacy of several osimertinib-based combinations following disease progression under frontline osimertinib.
BRAF mutations and fusions (i.e. AGK-BRAF, ESYT2-BRAF) have recently emerged as additional mechanisms of AR to third-generation EGFR TKI2,18–20. Studies demonstrating the efficacy of concurrent inhibition of EGFR and BRAF3 or MEK21 in pre-clinical models have raised clinicians’ expectations about overcoming AR by combining TT. Nevertheless, reports of successful combinations of TT for patients harboring BRAF-driven AR to osimertinib are very limited19,22–24 and no prospective data regarding efficacy and safety of BRAF/MEK/EGFR concurrent inhibition are available, with chemotherapy-based regimens remaining the treatment of choice in this unfavorable scenario. Similarly, PIK3CA mutations may also mediate AR to second-line osimertinib in 4–11%, but no clinical reports suggesting potential benefits of blocking these alterations in NSCLC are available6,9,25–27.
The detection of circulating tumor DNA (ctDNA) using liquid biopsies allows noninvasive real-time monitoring of treatment response and early detection of AR to TT, anticipating radiological response and treatment failure28,29. The widespread availability of highly specific and sensitive techniques to quantify ctDNA makes the longitudinal assessment of patients with NSCLC under TKI therapy very attractive. Nonetheless, for NSCLCs, ctDNA evaluation in routine clinical practice is currently limited to the detection of EGFR exon 19 deletion, EGFR L858R and T790M activating mutations29.
A man with metastatic lung adenocarcinoma harboring an EGFR mutation, who had progressed to erlotinib due to the emergence of the EGFR T790M mutation started osimertinib and remained on treatment for 15 months, then developing disease progression (PD). Sequential liquid biopsies were collected to monitor treatment response and disclosed the emergence of BRAF V600E and PIK3CA E545K resistance mutations 4 months before clinical progression. With this result and considering few available reports in the literature, our patient started a triple therapy with osimertinib, dabrafenib (BRAF inhibitor), and trametinib (MEK inhibitor).
Results
Case report
A 50-year-old non-smoker man was diagnosed with a tubule-papillary lung adenocarcinoma metastatic to bones and soft tissue (stage IVB - AJCC 8th edition) in July 2016. Several hypermetabolic bone lesions were observed on staging 18-fluorodeoxyglucose positron-emission tomography (18F-FDG PET-CT) scan at diagnosis. A baseline Next Generation Sequencing (NGS) assay (TruSightTumor™ - Illumina®) of a soft tissue metastasis revealed the presence of an activating EGFR exon 19 deletion (EGFR E746_A750del). No concurrent alterations in BRAF or in PIK3CA were identified at that time. ALK d5f3 immunohistochemistry and fluorescent in situ hybridization (FISH) for ROS1 and RET resulted negative.
In September 2016, the patient started on erlotinib 150 mg once daily (OD), achieving complete metabolic response in February 2017. We started to perform serial blood sample collections for ctDNA analysis in May 2017. In October 2017, oligo-progression (oligo-PD) was observed at the primary site and treated with stereotactic body radiation therapy (SBRT; 3 × 16 Gy). In November 2017, we identified the EGFR T790M and EGFR E746_A750del mutations in patient’s plasma using droplet digital PCR (ddPCR) and these results were confirmed in December 2017 with the Foundation ACT® ctDNA assay. Figure 1a shows all systemic and focal therapies of this patient since diagnosis.Fig. 1 Paired radiological and sequential blood-based ctDNA assessments throughout patient’s treatment demonstrating concordant results.
a Timeline displaying systemic and focal therapies since September 2016. b ctDNA levels in serial plasma samples based on fractional abundance of EGFR T790M, EGFR del19 (E746_A750 del), BRAF V600E, and PIK3CA E545K mutations. #2.4 CN results: EGFR del19 WT/RNAseP copy number ratio. c Paired 18F-FDG PET-CT scan maximum intensity projection (MIP) images displaying tumor burden variations. PD progression of disease, SBRT stereotatic body radiotherapy, Pembro pembrolizumab, CT chemotherapy, DTO dabrafenib, trametinib and osimertinib.
Erlotinib was kept until January 2018 when disease progression in the bones was observed and the fractional abundance (FA) of EGFR E746_A750del and EGFR T790M were 12.0 and 2.3%, respectively (Fig. 1b). The treatment was promptly switched to osimertinib 80 mg OD. Bone partial response (PR) was observed 2 months later, along with a significant drop in the FA of EGFR E746_A750del and EGFR T790M mutations in the plasma (Fig. 1b).
In July 2018, the patient developed oligo-PD in T3 vertebrae, which was treated with SBRT (1 × 20 Gy). At that point, ctDNA analysis revealed an increase in the FA of the EGFR E746_A750del in the plasma, but not in EGFR T790M, suggesting genetic heterogeneity between metastatic lesions. In December 2018, a new oligo-PD in the left iliac bone was detected by 18F-FDG PET-CT scan (Fig. 1c). At that time, ctDNA analysis revealed the emergence of BRAF V600E (FA: 0.4%) and PIK3CA E545K (FA: 0.9%) mutations, as well as a significant increase in the FA of the EGFR E746_A750del (FA: 7.5%) in the plasma. These mutations were also detected in a specimen from an iliac bone biopsy using a NGS assay (TruSightTumor™ - Illumina® - EGFR del19 allelic fraction 81.1%, EGFR del19 amplification – 12 copies, BRAF V600E allelic fraction 17.7% and PIK3CA E545K allelic fraction 32.7%). Even though the patient was treated with SBRT (1 × 18 Gy), he developed new bone metastases in March 2019 (Fig. 1b), consistent with the significant increase in the FA of EGFR E746_A750del, BRAF V600E and EGFR T790M mutations (Fig. 1b). At that specific timepoint and in April 2019, assessments of PIK3CA E545K plasma levels were not obtained due to limited amount of cfDNA.
Between April and September 2019, systemic treatment with carboplatin plus pemetrexed and pembrolizumab provided modest clinical benefit, reducing tumor burden and controlling the disease (Fig. 1b). Due to a new PD observed in October 2019, the treatment was switched to docetaxel monotherapy and, subsequently, to vinorelbine after progression under docetaxel; nonetheless, the patient experienced symptomatic PD in bones, lymph nodes, and lung in late January 2020 (Fig. 1c, 2a–d). He came to the clinic with a Karnofsky performance status of 70%, complaining of fatigue, appetite loss, and severe pain in the hips (despite regular use of 10 mg buprenorphine patch and oxycodone plus acetaminophen PO), which had been preventing him from performing his daily activities due to the inability to stay seated. At that point, high levels of all three resistance mutations were detected in the circulating DNA (PIK3CA E545K FA: 13.2%, BRAF V600E FA: 12.3%, and EGFR T790M FA: 5.3%; Fig. 1b). We also observed an exponential increase in the number of copies of EGFR E746_A750del, suggesting gene amplification (Fig. 1b).Fig. 2 18F-FDG PET-CT scan imaging depicting impressive overall tumor response under dabrafenib, trametinib, and osimertinib in three timepoints.
a–d baseline imaging (January/2020) showing: hypermetabolic spiculated mass (orange circle) in the left-superior lobe measuring 4.4 × 3.3 cm (SUVmax: 9.4); multiple hilar, mediastinal, retroperitoneal, and iliac hypermetabolic lymph nodes (red arrow) measuring up to 2.7 cm (SUVmax: 12.8); several hypermetabolic bone lesions throughout axial and appendicular skeleton (yellow circle); a left iliac bone lesion with signs of periosteal reaction and adjacent soft-tissue infiltration (SUVmax: 11.4). e–h first response evaluation imaging (April/2020) showing considerable partial response in the lung mass, measuring 3.8 × 2.5 cm (SUVmax: 2.7), as well as in several bone lesions, especially in the left-iliac bone (SUVmax: 4.8); complete response in lymph nodes. i–l third response evaluation imaging showing disease progression in the left ischium (SUVmax: 12.5; previous SUVmax: 5.9) (October/2020). SUVmax maximum standard uptake value.
In February 2020, after careful consideration due to patient’s good performance status, normal organ functions, severe pain, motivation and the emergence of BRAF V600E as an AR mechanism, he started on dabrafenib 75 mg twice daily (BID), trametinib 1 mg OD and osimertinib 80 mg OD. The use of a PI3K inhibitor was not considered an option due to the absence of reports in the literature suggesting efficacy of these drugs combined with osimertinib to treat advanced NSCLC patients. Within 2 weeks of treatment, the patient achieved complete resolution of the severe bone pain in the hips with no further need of opioid administration, as well as appetite gain and marked improvement in quality of life, which turned possible for him to resume his daily activities. As adverse events (AE), he experienced grade 1 fatigue, dysgeusia, fever, and nausea, all managed with symptomatic medication. Complete resolution of the fever occurred spontaneously within two weeks of treatment. An attempt to increase dosages of dabrafenib to 150 mg BID and trametinib 2 mg OD resulted unsuccessful due to persistent grade 2 fatigue. In April 2020, a new 18F-FDG PET-CT scan disclosed a complete response in lymph nodes and a dramatic PR in the lung and bones (PERCIST 1.1 evaluating 5 target lesions: Δ- 67%; Figs. 1c and 2e–h). Detection of EGFR T790M, BRAF V600E, and PIK3CA E545K mutations became negative in April 2020, while EGFR E746_A750del despite marginally positive, presented marked reduction (Fig. 1b) suggesting that combination dabrafenib, trametinib, and osimertinib might be an effective strategy to overcome PIK3CA E545K and BRAF V600E-driven resistance to osimertinib in advanced EGFR-mutant NSCLC patients. The patient remained in response until October 2020, when he developed asymptomatic bone PD in lumbar spine, left ischium, and right iliac bone, 8 months after starting this combination therapy (Figs. 1c and 2i–l). A FoundationOne®Liquid CDx plasma NGS disclosed the following alterations: CHCHD3-BRAF fusion, BRAF V600E, EGFR E746_750del, EGFR amplification, PIK3CA E545K, MAP2K2 (MEK2) C125S, MTAP rearrangement intron 5, TP53 V197M, and TP53 S241A. Owing to the considerable clinical benefit, good tolerance and lack of systemic treatment options, we decided to keep the patient on treatment and increase the doses of dabrafenib (150 mg BID) and trametinib (2 mg OD) every other day. He remains asymptomatic and tolerating well the proposed dose adjustment.
Discussion
Several studies have recently highlighted the importance of considering genes of interest within the context of commonly co-occurring mutations9,30. For example, as described by Blakely et al. through performing a cfDNA NGS analysis of 1,122 advanced stage EGFR-mutant NSCLCs, in ~93% of the patients, at least one more variables with known or likely known functional properties were present, disclosing the molecular complexity of this oncogenic driver and suggesting an association of co-occuring genomic alterations with TKI response and clinical outcomes9. In addition, Roper and colleagues reported the identification of at least two co-existing AR mechanisms in 73% of patients treated with osimertinib, as well as 6–23 different subclones per individual in a phylogenetic analysis performed in multiple metastatic sites of 15 individuals30. In the same publication, the authors also called the attention for a high incidence of acquired EGFR amplifications in post-osimertinib patients, which suggests maintenance of EGFR central role in the setting of progression, as previously reported by our group28 and also detected in the present case.
Although uncommon, BRAF V600E mediates AR in approximately 3% of the patients under second-line osimertinib6,20 and little is known about the efficacy of combined TT in this population3,21–23. Existing reports lack details regarding objective responses using standardized radiological criteria (i.e., RECIST 1.1; PERCIST) and the dynamics of resistance mechanisms through longitudinal ctDNA measurements22,23. Huang and colleagues22 described a case of an EGFRdel19/T790M + NSCLC patient who developed BRAF V600E-driven AR after second-line osimertinib and achieved stable disease under dabrafenib 150 mg BID, trametinib 1 mg OD and osimertinib 80 mg OD, with ongoing disease control 7.4 months after. Zhou and colleagues23 also reported their experience with dabrafenib 150 mg BID, trametinib 2 mg OD and osimertinib 80 mg OD leading to tumor reduction within 6 weeks of treatment, along with grade 2 rash and decreased appetite as AEs. Similarly, Meng et al. reported two cases treated with this triple regimen31. The first patient discontinued therapy after one month due to severe pneumonitis; the second one presented tumor response under dabrafenib 50 mg BID, trametinib 0.5 mg OD, and osimertinib 80 mg OD, with progression-free survival of 14 months. According to the authors, this reduced dose was prescribed owing to a grade 2 pyrexia, nausea, and vomiting under higher dabrafenib and trametinib doses. Dagogo-Jack and colleagues19 also described a successful case of combined EGFR/MAP kinase pathway blockade with osimertinib 80 mg OD and trametinib 1 mg OD; as treatment-related AEs, their patient experienced grade 2 diarrhea and fatigue, along with grade 1 rash and gastrointestinal bleeding. Nevertheless, it is worth highlighting the rapid clinical improvement, the remarkable radiologically confirmed objective response, as well as the good tolerance observed in this case even using only half standard dose of dabrafenib and trametinib approved for NSCLCs harboring BRAF V600E mutations. Since data regarding the efficacy of these combined approaches, the optimal drug association and dosing, as well as the toxicity profile are conflicting and largely unknown, further investigation into the mechanistic basis of this association represents an important priority. The Table 1 summarizes the toxicities arising under osimertinib plus BRAF/MEK inhibition and reported in the literature.Table 1 Summary of the toxicities arising under osimertinib plus BRAF/MEK inhibition and reported in the literature.
Reference Treatment Anorexia Nausea Vomiting Diarrhoea Fatigue Rash GI bleeding AST/ALT elevation Paroniquia Pyrexia Initial dose Dose reduction
Meng et al. D + T + O NR G2 G2 G2 NR NR NR NR NR G2 D (150 mg BID) + T (2 mg OD) + O (80 mg OD) D (50 mg BID) + T (0.5 mg/day) + O (80 mg/day)
Dagogo-Jack et al. T + O NR NR NR G2 G2 G1 G1 G1 NR NR T (1 mg OD) + O (80 mg OD) Not needed
Huang et al. D + T + O NR NR NR G1 NR NR NR NR G1 NR D (150 mg BID) + T (1 mg OD) + O (80 mg OD) Not needed
Zhou et al. D + T + O G1 NR NR NR NR G2 NR NR NR NR D (150 mg BID) + T (2 mg OD) + O (80 mg OD) Not needed
D dabrafenib, T trametinib, O osimertinib, NR not reported.
Liquid biopsy is emerging as an important diagnostic and predictive tool in the treatment of NSCLCs. The accurate identification of predictive genetic alterations is important for both patients’ management and the understanding of clonal evolution and AR to different therapies9,29,32. Also, circulating biomarkers from multiple disease sites better reflect systemic tumor burden, including alterations from genetically different metastatic lesions, which may be missed with single-site tissue biopsies8. Here, we were able to identify and monitor multiple concurrent mechanisms of resistance throughout the entire patient’s treatment. As shown in Fig. 1b, variations in blood ctDNA levels exhibited a positive correlation with imaging findings, even in situations involving focal treatments for oligo-PD. The observed concordance between radiological PD or response and ctDNA measurements also points towards its great potential to be incorporated into clinical practice to anticipate radiologic findings in a more effective manner.
Along with BRAF V600E mutation, the presence of PIK3CA E545K mutation is also associated with AR to osimertinib6,9,25,27. However, the clinical implication of concomitant targeting of EGFR and PI3K remains unclear. Whereas an alternative argument for the observed response could be a rechallenge following a long time upon off-osimertinib, in our opinion, this is an unlikely explanation33. Interestingly, the presence of a BRAF fusion along with a MEK2 mutation identified through plasma NGS in the setting of PD might represent a mechanism of resistance to this triple regimen34. Nonetheless, since pretreatment blood samples had not been analyzed using a similar methodology, this hypothesis deserves careful interpretation.
Increased understanding of the relationship of concurring genomic alterations in EGFR-mutant NSCLC may enable new therapeutic opportunities following disease progression to osimertinib. Here, we reported an impressive objective response to dabrafenib, trametinib, and osimertinib with concordant decrease in plasma ctDNA levels in a metastatic lung cancer patient harboring EGFR E746_A750del, BRAF V600E, and PIK3CA E545K activating mutations. This case report leads to a greater understanding of the currently limited literature regarding the management of EGFR-mutant NSCLC patients with acquired BRAFV600E mutation, since it reports a successful attempt to target both alterations concurrently while providing concordant and interesting data of serial ctDNA assessments throughout the entire treatment. Further investigation to optimize the efficacy and mitigate the toxicity profile of this drug association represents an important issue.
Methods
Patient
This study was approved by Hospital Sírio-Libanês Ethics Committee (HSL-RC 2020-16). The patient provided written informed consent for blood collection, ctDNA analysis and publication of this report.
Sample collection and plasma DNA extraction
Serial blood samples were collected between May 2017 and April 2020 (Fig. 1b). Peripheral blood (20 ml) was collected into tubes containing EDTA (BD, Franklin Lakes, New Jersey). Plasma was separated from the blood within 2 hours of blood collection, as previously described28. cfDNA was extracted using QIAamp MinElute Virus Vacuum Kit (Qiagen, Hilden, Germany) and stored at −80 °C.
ctDNA-ddPCR
Cell-free DNA (cfDNA) was quantified using the RNase P Copy Number Reference Assay (Life Technologies, Carlsbad, California). A total of 3000 genome-equivalents (~10 ng of cfDNA) were analyzed per assay to achieve a detection sensitivity of 0.2%. This detection limit has been assessed by using cell line-derived genomic DNA. A total of 10 ng of input DNA with varying proportions of mutant DNA was serially diluted into wild-type DNA to obtain samples with a mutant abundance of 1%, 0.5%, 0.1%, and 0.05% and subjected to droplet digital PCR (ddPCR). ddPCR was used to quantify the circulating levels of the EGFR activating mutation (EGFR E746_A750 del) and of the resistance mutations (EGFR T790M, PIK3CA E545K, and BRAF V600E). Probes and primers were obtained from BioRad (EGFR E746_A750 del #10041170, EGFR T790M #10040782, PIK3CA E545K #10041188, and BRAF V600E #10040779; Hercules, California). ddPCR was performed on the QX200 Droplet Digital PCR System, and data were analysed using QuantaSoft software (Bio-Rad). ctDNA quantification is presented as fractional abundance (FA–the proportion of the mutant allele in total cfDNA).
Reporting summary
Further information on research design is available in the Nature Research Reporting Summary linked to this article.
Supplementary information
Reporting Summary
Dataset 1
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
These authors contributed equally: Maurício Fernando Silva Almeida Ribeiro, Franciele Hinterholz Knebel.
Supplementary information
The online version contains supplementary material available at 10.1038/s41698-021-00149-4.
Acknowledgements
We thank the clinical research team at Hospital Sírio-Libanês for their assistance collecting samples. We thank Ernande Xavier dos Santos and Elisângela Monteiro for the outstanding technical support. We also thank Mrs. Dina Binzagr and Vivian Hannud for supporting the Translational Research Program at Hospital Sírio Libanês. This work was supported by Ludwig Cancer Research and Fundação de Amparo à Pesquisa do Estado de São Paulo – FAPESP. Lastly, we would like to thank the patient and his family for consenting to the present publication.
Author contributions
Conceptualization by M.F.S.A.R., F.H.K., J.V.M.A., A.A.C., and A.K.; Methodology by F.H.K. and A.A.C.; Resources br M.F.S.A.R., F.H.K., A.A.C., and J.V.M.A.; Writing of the first draft by M.F.S.A., F.H.K., and J.V.M.A.; Review, editing and approval of the final draft by all authors; Supervision by A.A.C. and A.K.; Funding acquisition by F.H.K. and A.A.C. M.F.S.A.R. and F.H.K. are co-first authors of the present work.
Data availability
The datasets that support the findings of this study are not publicly available in order to protect patient privacy. The data will be made available on reasonable request. For data access requests regarding the liquid biopsy (ctDNA quantification) data, please contact Dr. Franciele Knebel, email address: fhknebel@mochsl.org.br. For data access requests regarding the PET-CT high resolution images and PERCIST calculations, please contact Dr. José Marin, email address:jfgmarin@yahoo.com.br. For data access requests regarding the summary of the toxicities arising under osimertinib plus BRAF/MEK inhibition, please contact the corresponding author Dr. Maurício Ribeiro, email address: mauricio.fsaribeiro@hsl.org.br. The data generated and analysed during this study are described in the following metadata record: 10.6084/m9.figshare.1347594635.
Competing interests
The authors declare no competing interests. | DABRAFENIB, OSIMERTINIB, TRAMETINIB | DrugsGivenReaction | CC BY | 33580193 | 19,431,708 | 2021-02-12 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Product use issue'. | Impressive response to dabrafenib, trametinib, and osimertinib in a metastatic EGFR-mutant/BRAF V600E lung adenocarcinoma patient.
The survival outcomes of the FLAURA trial support osimertinib as the new standard of care for untreated patients harboring activating mutations in the epidermal growth factor receptor (EGFR). Despite the initial response, disease progression invariably occurs. Although uncommon, BRAF V600E mutation arises as a unique mechanism of resistance, and thus far, no prospective studies are available to support concurrent EGFR/BRAF blockade. We report a case of impressive radiological and ctDNA response under dabrafenib, trametinib, and osimertinib in an advanced EGFR-mutant lung adenocarcinoma patient who developed BRAF V600E as one of the acquired resistance mechanisms to second-line osimertinib. Moreover, the patient experienced remarkable clinical improvement and good tolerance to combination therapy. The present case suggests the importance of prospective studies evaluating both efficacy and safety of the combination in later line settings and points towards the potential of ctDNA to monitor resistance mechanisms and treatment benefit in clinical practice.
Introduction
The use of osimertinib to target epidermal growth factor receptor (EGFR) has become the standard of care in untreated EGFR-mutant non–small cell lung cancer (NSCLC) patients. Although osimertinib can be highly active, showing more durable outcomes than first-generation tyrosine kinase inhibitors (TKI)1, most tumors invariably become resistant, limiting its long-term clinical benefit. The heterogeneity of resistance mechanisms to osimertinib, including EGFR C797S mutation, EGFR, and MET amplifications, off-target mutations in PIK3CA, KRAS, and HER2 as well as histologic transformation2–9, has stimulated routine performance of repeated biopsies to identify specific underlying mechanisms of resistance throughout the treatment course, and to guide the development of novel therapeutic strategies to overcome and prevent acquired resistance (AR)7,10–13.
Combined targeted therapy (TT) strategies have been increasingly addressed in prospective clinical trials14–17. A phase Ib/II trial reported 47% ORR with a combination of capmatinib (MET inhibitor) plus gefitinib in patients progressing to EGFR TKI and presenting MET dysregulation14. Likewise, SAVANNAH (NCT03778229)15 is an ongoing phase II trial designed to evaluate the efficacy of combination savolitinib and osimertinib in post-osimertinib progression/MET-positive patients. The biomarker-matched study ORCHARD (NCT03944772)16 is also underway to assess the efficacy of several osimertinib-based combinations following disease progression under frontline osimertinib.
BRAF mutations and fusions (i.e. AGK-BRAF, ESYT2-BRAF) have recently emerged as additional mechanisms of AR to third-generation EGFR TKI2,18–20. Studies demonstrating the efficacy of concurrent inhibition of EGFR and BRAF3 or MEK21 in pre-clinical models have raised clinicians’ expectations about overcoming AR by combining TT. Nevertheless, reports of successful combinations of TT for patients harboring BRAF-driven AR to osimertinib are very limited19,22–24 and no prospective data regarding efficacy and safety of BRAF/MEK/EGFR concurrent inhibition are available, with chemotherapy-based regimens remaining the treatment of choice in this unfavorable scenario. Similarly, PIK3CA mutations may also mediate AR to second-line osimertinib in 4–11%, but no clinical reports suggesting potential benefits of blocking these alterations in NSCLC are available6,9,25–27.
The detection of circulating tumor DNA (ctDNA) using liquid biopsies allows noninvasive real-time monitoring of treatment response and early detection of AR to TT, anticipating radiological response and treatment failure28,29. The widespread availability of highly specific and sensitive techniques to quantify ctDNA makes the longitudinal assessment of patients with NSCLC under TKI therapy very attractive. Nonetheless, for NSCLCs, ctDNA evaluation in routine clinical practice is currently limited to the detection of EGFR exon 19 deletion, EGFR L858R and T790M activating mutations29.
A man with metastatic lung adenocarcinoma harboring an EGFR mutation, who had progressed to erlotinib due to the emergence of the EGFR T790M mutation started osimertinib and remained on treatment for 15 months, then developing disease progression (PD). Sequential liquid biopsies were collected to monitor treatment response and disclosed the emergence of BRAF V600E and PIK3CA E545K resistance mutations 4 months before clinical progression. With this result and considering few available reports in the literature, our patient started a triple therapy with osimertinib, dabrafenib (BRAF inhibitor), and trametinib (MEK inhibitor).
Results
Case report
A 50-year-old non-smoker man was diagnosed with a tubule-papillary lung adenocarcinoma metastatic to bones and soft tissue (stage IVB - AJCC 8th edition) in July 2016. Several hypermetabolic bone lesions were observed on staging 18-fluorodeoxyglucose positron-emission tomography (18F-FDG PET-CT) scan at diagnosis. A baseline Next Generation Sequencing (NGS) assay (TruSightTumor™ - Illumina®) of a soft tissue metastasis revealed the presence of an activating EGFR exon 19 deletion (EGFR E746_A750del). No concurrent alterations in BRAF or in PIK3CA were identified at that time. ALK d5f3 immunohistochemistry and fluorescent in situ hybridization (FISH) for ROS1 and RET resulted negative.
In September 2016, the patient started on erlotinib 150 mg once daily (OD), achieving complete metabolic response in February 2017. We started to perform serial blood sample collections for ctDNA analysis in May 2017. In October 2017, oligo-progression (oligo-PD) was observed at the primary site and treated with stereotactic body radiation therapy (SBRT; 3 × 16 Gy). In November 2017, we identified the EGFR T790M and EGFR E746_A750del mutations in patient’s plasma using droplet digital PCR (ddPCR) and these results were confirmed in December 2017 with the Foundation ACT® ctDNA assay. Figure 1a shows all systemic and focal therapies of this patient since diagnosis.Fig. 1 Paired radiological and sequential blood-based ctDNA assessments throughout patient’s treatment demonstrating concordant results.
a Timeline displaying systemic and focal therapies since September 2016. b ctDNA levels in serial plasma samples based on fractional abundance of EGFR T790M, EGFR del19 (E746_A750 del), BRAF V600E, and PIK3CA E545K mutations. #2.4 CN results: EGFR del19 WT/RNAseP copy number ratio. c Paired 18F-FDG PET-CT scan maximum intensity projection (MIP) images displaying tumor burden variations. PD progression of disease, SBRT stereotatic body radiotherapy, Pembro pembrolizumab, CT chemotherapy, DTO dabrafenib, trametinib and osimertinib.
Erlotinib was kept until January 2018 when disease progression in the bones was observed and the fractional abundance (FA) of EGFR E746_A750del and EGFR T790M were 12.0 and 2.3%, respectively (Fig. 1b). The treatment was promptly switched to osimertinib 80 mg OD. Bone partial response (PR) was observed 2 months later, along with a significant drop in the FA of EGFR E746_A750del and EGFR T790M mutations in the plasma (Fig. 1b).
In July 2018, the patient developed oligo-PD in T3 vertebrae, which was treated with SBRT (1 × 20 Gy). At that point, ctDNA analysis revealed an increase in the FA of the EGFR E746_A750del in the plasma, but not in EGFR T790M, suggesting genetic heterogeneity between metastatic lesions. In December 2018, a new oligo-PD in the left iliac bone was detected by 18F-FDG PET-CT scan (Fig. 1c). At that time, ctDNA analysis revealed the emergence of BRAF V600E (FA: 0.4%) and PIK3CA E545K (FA: 0.9%) mutations, as well as a significant increase in the FA of the EGFR E746_A750del (FA: 7.5%) in the plasma. These mutations were also detected in a specimen from an iliac bone biopsy using a NGS assay (TruSightTumor™ - Illumina® - EGFR del19 allelic fraction 81.1%, EGFR del19 amplification – 12 copies, BRAF V600E allelic fraction 17.7% and PIK3CA E545K allelic fraction 32.7%). Even though the patient was treated with SBRT (1 × 18 Gy), he developed new bone metastases in March 2019 (Fig. 1b), consistent with the significant increase in the FA of EGFR E746_A750del, BRAF V600E and EGFR T790M mutations (Fig. 1b). At that specific timepoint and in April 2019, assessments of PIK3CA E545K plasma levels were not obtained due to limited amount of cfDNA.
Between April and September 2019, systemic treatment with carboplatin plus pemetrexed and pembrolizumab provided modest clinical benefit, reducing tumor burden and controlling the disease (Fig. 1b). Due to a new PD observed in October 2019, the treatment was switched to docetaxel monotherapy and, subsequently, to vinorelbine after progression under docetaxel; nonetheless, the patient experienced symptomatic PD in bones, lymph nodes, and lung in late January 2020 (Fig. 1c, 2a–d). He came to the clinic with a Karnofsky performance status of 70%, complaining of fatigue, appetite loss, and severe pain in the hips (despite regular use of 10 mg buprenorphine patch and oxycodone plus acetaminophen PO), which had been preventing him from performing his daily activities due to the inability to stay seated. At that point, high levels of all three resistance mutations were detected in the circulating DNA (PIK3CA E545K FA: 13.2%, BRAF V600E FA: 12.3%, and EGFR T790M FA: 5.3%; Fig. 1b). We also observed an exponential increase in the number of copies of EGFR E746_A750del, suggesting gene amplification (Fig. 1b).Fig. 2 18F-FDG PET-CT scan imaging depicting impressive overall tumor response under dabrafenib, trametinib, and osimertinib in three timepoints.
a–d baseline imaging (January/2020) showing: hypermetabolic spiculated mass (orange circle) in the left-superior lobe measuring 4.4 × 3.3 cm (SUVmax: 9.4); multiple hilar, mediastinal, retroperitoneal, and iliac hypermetabolic lymph nodes (red arrow) measuring up to 2.7 cm (SUVmax: 12.8); several hypermetabolic bone lesions throughout axial and appendicular skeleton (yellow circle); a left iliac bone lesion with signs of periosteal reaction and adjacent soft-tissue infiltration (SUVmax: 11.4). e–h first response evaluation imaging (April/2020) showing considerable partial response in the lung mass, measuring 3.8 × 2.5 cm (SUVmax: 2.7), as well as in several bone lesions, especially in the left-iliac bone (SUVmax: 4.8); complete response in lymph nodes. i–l third response evaluation imaging showing disease progression in the left ischium (SUVmax: 12.5; previous SUVmax: 5.9) (October/2020). SUVmax maximum standard uptake value.
In February 2020, after careful consideration due to patient’s good performance status, normal organ functions, severe pain, motivation and the emergence of BRAF V600E as an AR mechanism, he started on dabrafenib 75 mg twice daily (BID), trametinib 1 mg OD and osimertinib 80 mg OD. The use of a PI3K inhibitor was not considered an option due to the absence of reports in the literature suggesting efficacy of these drugs combined with osimertinib to treat advanced NSCLC patients. Within 2 weeks of treatment, the patient achieved complete resolution of the severe bone pain in the hips with no further need of opioid administration, as well as appetite gain and marked improvement in quality of life, which turned possible for him to resume his daily activities. As adverse events (AE), he experienced grade 1 fatigue, dysgeusia, fever, and nausea, all managed with symptomatic medication. Complete resolution of the fever occurred spontaneously within two weeks of treatment. An attempt to increase dosages of dabrafenib to 150 mg BID and trametinib 2 mg OD resulted unsuccessful due to persistent grade 2 fatigue. In April 2020, a new 18F-FDG PET-CT scan disclosed a complete response in lymph nodes and a dramatic PR in the lung and bones (PERCIST 1.1 evaluating 5 target lesions: Δ- 67%; Figs. 1c and 2e–h). Detection of EGFR T790M, BRAF V600E, and PIK3CA E545K mutations became negative in April 2020, while EGFR E746_A750del despite marginally positive, presented marked reduction (Fig. 1b) suggesting that combination dabrafenib, trametinib, and osimertinib might be an effective strategy to overcome PIK3CA E545K and BRAF V600E-driven resistance to osimertinib in advanced EGFR-mutant NSCLC patients. The patient remained in response until October 2020, when he developed asymptomatic bone PD in lumbar spine, left ischium, and right iliac bone, 8 months after starting this combination therapy (Figs. 1c and 2i–l). A FoundationOne®Liquid CDx plasma NGS disclosed the following alterations: CHCHD3-BRAF fusion, BRAF V600E, EGFR E746_750del, EGFR amplification, PIK3CA E545K, MAP2K2 (MEK2) C125S, MTAP rearrangement intron 5, TP53 V197M, and TP53 S241A. Owing to the considerable clinical benefit, good tolerance and lack of systemic treatment options, we decided to keep the patient on treatment and increase the doses of dabrafenib (150 mg BID) and trametinib (2 mg OD) every other day. He remains asymptomatic and tolerating well the proposed dose adjustment.
Discussion
Several studies have recently highlighted the importance of considering genes of interest within the context of commonly co-occurring mutations9,30. For example, as described by Blakely et al. through performing a cfDNA NGS analysis of 1,122 advanced stage EGFR-mutant NSCLCs, in ~93% of the patients, at least one more variables with known or likely known functional properties were present, disclosing the molecular complexity of this oncogenic driver and suggesting an association of co-occuring genomic alterations with TKI response and clinical outcomes9. In addition, Roper and colleagues reported the identification of at least two co-existing AR mechanisms in 73% of patients treated with osimertinib, as well as 6–23 different subclones per individual in a phylogenetic analysis performed in multiple metastatic sites of 15 individuals30. In the same publication, the authors also called the attention for a high incidence of acquired EGFR amplifications in post-osimertinib patients, which suggests maintenance of EGFR central role in the setting of progression, as previously reported by our group28 and also detected in the present case.
Although uncommon, BRAF V600E mediates AR in approximately 3% of the patients under second-line osimertinib6,20 and little is known about the efficacy of combined TT in this population3,21–23. Existing reports lack details regarding objective responses using standardized radiological criteria (i.e., RECIST 1.1; PERCIST) and the dynamics of resistance mechanisms through longitudinal ctDNA measurements22,23. Huang and colleagues22 described a case of an EGFRdel19/T790M + NSCLC patient who developed BRAF V600E-driven AR after second-line osimertinib and achieved stable disease under dabrafenib 150 mg BID, trametinib 1 mg OD and osimertinib 80 mg OD, with ongoing disease control 7.4 months after. Zhou and colleagues23 also reported their experience with dabrafenib 150 mg BID, trametinib 2 mg OD and osimertinib 80 mg OD leading to tumor reduction within 6 weeks of treatment, along with grade 2 rash and decreased appetite as AEs. Similarly, Meng et al. reported two cases treated with this triple regimen31. The first patient discontinued therapy after one month due to severe pneumonitis; the second one presented tumor response under dabrafenib 50 mg BID, trametinib 0.5 mg OD, and osimertinib 80 mg OD, with progression-free survival of 14 months. According to the authors, this reduced dose was prescribed owing to a grade 2 pyrexia, nausea, and vomiting under higher dabrafenib and trametinib doses. Dagogo-Jack and colleagues19 also described a successful case of combined EGFR/MAP kinase pathway blockade with osimertinib 80 mg OD and trametinib 1 mg OD; as treatment-related AEs, their patient experienced grade 2 diarrhea and fatigue, along with grade 1 rash and gastrointestinal bleeding. Nevertheless, it is worth highlighting the rapid clinical improvement, the remarkable radiologically confirmed objective response, as well as the good tolerance observed in this case even using only half standard dose of dabrafenib and trametinib approved for NSCLCs harboring BRAF V600E mutations. Since data regarding the efficacy of these combined approaches, the optimal drug association and dosing, as well as the toxicity profile are conflicting and largely unknown, further investigation into the mechanistic basis of this association represents an important priority. The Table 1 summarizes the toxicities arising under osimertinib plus BRAF/MEK inhibition and reported in the literature.Table 1 Summary of the toxicities arising under osimertinib plus BRAF/MEK inhibition and reported in the literature.
Reference Treatment Anorexia Nausea Vomiting Diarrhoea Fatigue Rash GI bleeding AST/ALT elevation Paroniquia Pyrexia Initial dose Dose reduction
Meng et al. D + T + O NR G2 G2 G2 NR NR NR NR NR G2 D (150 mg BID) + T (2 mg OD) + O (80 mg OD) D (50 mg BID) + T (0.5 mg/day) + O (80 mg/day)
Dagogo-Jack et al. T + O NR NR NR G2 G2 G1 G1 G1 NR NR T (1 mg OD) + O (80 mg OD) Not needed
Huang et al. D + T + O NR NR NR G1 NR NR NR NR G1 NR D (150 mg BID) + T (1 mg OD) + O (80 mg OD) Not needed
Zhou et al. D + T + O G1 NR NR NR NR G2 NR NR NR NR D (150 mg BID) + T (2 mg OD) + O (80 mg OD) Not needed
D dabrafenib, T trametinib, O osimertinib, NR not reported.
Liquid biopsy is emerging as an important diagnostic and predictive tool in the treatment of NSCLCs. The accurate identification of predictive genetic alterations is important for both patients’ management and the understanding of clonal evolution and AR to different therapies9,29,32. Also, circulating biomarkers from multiple disease sites better reflect systemic tumor burden, including alterations from genetically different metastatic lesions, which may be missed with single-site tissue biopsies8. Here, we were able to identify and monitor multiple concurrent mechanisms of resistance throughout the entire patient’s treatment. As shown in Fig. 1b, variations in blood ctDNA levels exhibited a positive correlation with imaging findings, even in situations involving focal treatments for oligo-PD. The observed concordance between radiological PD or response and ctDNA measurements also points towards its great potential to be incorporated into clinical practice to anticipate radiologic findings in a more effective manner.
Along with BRAF V600E mutation, the presence of PIK3CA E545K mutation is also associated with AR to osimertinib6,9,25,27. However, the clinical implication of concomitant targeting of EGFR and PI3K remains unclear. Whereas an alternative argument for the observed response could be a rechallenge following a long time upon off-osimertinib, in our opinion, this is an unlikely explanation33. Interestingly, the presence of a BRAF fusion along with a MEK2 mutation identified through plasma NGS in the setting of PD might represent a mechanism of resistance to this triple regimen34. Nonetheless, since pretreatment blood samples had not been analyzed using a similar methodology, this hypothesis deserves careful interpretation.
Increased understanding of the relationship of concurring genomic alterations in EGFR-mutant NSCLC may enable new therapeutic opportunities following disease progression to osimertinib. Here, we reported an impressive objective response to dabrafenib, trametinib, and osimertinib with concordant decrease in plasma ctDNA levels in a metastatic lung cancer patient harboring EGFR E746_A750del, BRAF V600E, and PIK3CA E545K activating mutations. This case report leads to a greater understanding of the currently limited literature regarding the management of EGFR-mutant NSCLC patients with acquired BRAFV600E mutation, since it reports a successful attempt to target both alterations concurrently while providing concordant and interesting data of serial ctDNA assessments throughout the entire treatment. Further investigation to optimize the efficacy and mitigate the toxicity profile of this drug association represents an important issue.
Methods
Patient
This study was approved by Hospital Sírio-Libanês Ethics Committee (HSL-RC 2020-16). The patient provided written informed consent for blood collection, ctDNA analysis and publication of this report.
Sample collection and plasma DNA extraction
Serial blood samples were collected between May 2017 and April 2020 (Fig. 1b). Peripheral blood (20 ml) was collected into tubes containing EDTA (BD, Franklin Lakes, New Jersey). Plasma was separated from the blood within 2 hours of blood collection, as previously described28. cfDNA was extracted using QIAamp MinElute Virus Vacuum Kit (Qiagen, Hilden, Germany) and stored at −80 °C.
ctDNA-ddPCR
Cell-free DNA (cfDNA) was quantified using the RNase P Copy Number Reference Assay (Life Technologies, Carlsbad, California). A total of 3000 genome-equivalents (~10 ng of cfDNA) were analyzed per assay to achieve a detection sensitivity of 0.2%. This detection limit has been assessed by using cell line-derived genomic DNA. A total of 10 ng of input DNA with varying proportions of mutant DNA was serially diluted into wild-type DNA to obtain samples with a mutant abundance of 1%, 0.5%, 0.1%, and 0.05% and subjected to droplet digital PCR (ddPCR). ddPCR was used to quantify the circulating levels of the EGFR activating mutation (EGFR E746_A750 del) and of the resistance mutations (EGFR T790M, PIK3CA E545K, and BRAF V600E). Probes and primers were obtained from BioRad (EGFR E746_A750 del #10041170, EGFR T790M #10040782, PIK3CA E545K #10041188, and BRAF V600E #10040779; Hercules, California). ddPCR was performed on the QX200 Droplet Digital PCR System, and data were analysed using QuantaSoft software (Bio-Rad). ctDNA quantification is presented as fractional abundance (FA–the proportion of the mutant allele in total cfDNA).
Reporting summary
Further information on research design is available in the Nature Research Reporting Summary linked to this article.
Supplementary information
Reporting Summary
Dataset 1
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
These authors contributed equally: Maurício Fernando Silva Almeida Ribeiro, Franciele Hinterholz Knebel.
Supplementary information
The online version contains supplementary material available at 10.1038/s41698-021-00149-4.
Acknowledgements
We thank the clinical research team at Hospital Sírio-Libanês for their assistance collecting samples. We thank Ernande Xavier dos Santos and Elisângela Monteiro for the outstanding technical support. We also thank Mrs. Dina Binzagr and Vivian Hannud for supporting the Translational Research Program at Hospital Sírio Libanês. This work was supported by Ludwig Cancer Research and Fundação de Amparo à Pesquisa do Estado de São Paulo – FAPESP. Lastly, we would like to thank the patient and his family for consenting to the present publication.
Author contributions
Conceptualization by M.F.S.A.R., F.H.K., J.V.M.A., A.A.C., and A.K.; Methodology by F.H.K. and A.A.C.; Resources br M.F.S.A.R., F.H.K., A.A.C., and J.V.M.A.; Writing of the first draft by M.F.S.A., F.H.K., and J.V.M.A.; Review, editing and approval of the final draft by all authors; Supervision by A.A.C. and A.K.; Funding acquisition by F.H.K. and A.A.C. M.F.S.A.R. and F.H.K. are co-first authors of the present work.
Data availability
The datasets that support the findings of this study are not publicly available in order to protect patient privacy. The data will be made available on reasonable request. For data access requests regarding the liquid biopsy (ctDNA quantification) data, please contact Dr. Franciele Knebel, email address: fhknebel@mochsl.org.br. For data access requests regarding the PET-CT high resolution images and PERCIST calculations, please contact Dr. José Marin, email address:jfgmarin@yahoo.com.br. For data access requests regarding the summary of the toxicities arising under osimertinib plus BRAF/MEK inhibition, please contact the corresponding author Dr. Maurício Ribeiro, email address: mauricio.fsaribeiro@hsl.org.br. The data generated and analysed during this study are described in the following metadata record: 10.6084/m9.figshare.1347594635.
Competing interests
The authors declare no competing interests. | DABRAFENIB, OSIMERTINIB, TRAMETINIB | DrugsGivenReaction | CC BY | 33580193 | 19,431,708 | 2021-02-12 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Pyrexia'. | Impressive response to dabrafenib, trametinib, and osimertinib in a metastatic EGFR-mutant/BRAF V600E lung adenocarcinoma patient.
The survival outcomes of the FLAURA trial support osimertinib as the new standard of care for untreated patients harboring activating mutations in the epidermal growth factor receptor (EGFR). Despite the initial response, disease progression invariably occurs. Although uncommon, BRAF V600E mutation arises as a unique mechanism of resistance, and thus far, no prospective studies are available to support concurrent EGFR/BRAF blockade. We report a case of impressive radiological and ctDNA response under dabrafenib, trametinib, and osimertinib in an advanced EGFR-mutant lung adenocarcinoma patient who developed BRAF V600E as one of the acquired resistance mechanisms to second-line osimertinib. Moreover, the patient experienced remarkable clinical improvement and good tolerance to combination therapy. The present case suggests the importance of prospective studies evaluating both efficacy and safety of the combination in later line settings and points towards the potential of ctDNA to monitor resistance mechanisms and treatment benefit in clinical practice.
Introduction
The use of osimertinib to target epidermal growth factor receptor (EGFR) has become the standard of care in untreated EGFR-mutant non–small cell lung cancer (NSCLC) patients. Although osimertinib can be highly active, showing more durable outcomes than first-generation tyrosine kinase inhibitors (TKI)1, most tumors invariably become resistant, limiting its long-term clinical benefit. The heterogeneity of resistance mechanisms to osimertinib, including EGFR C797S mutation, EGFR, and MET amplifications, off-target mutations in PIK3CA, KRAS, and HER2 as well as histologic transformation2–9, has stimulated routine performance of repeated biopsies to identify specific underlying mechanisms of resistance throughout the treatment course, and to guide the development of novel therapeutic strategies to overcome and prevent acquired resistance (AR)7,10–13.
Combined targeted therapy (TT) strategies have been increasingly addressed in prospective clinical trials14–17. A phase Ib/II trial reported 47% ORR with a combination of capmatinib (MET inhibitor) plus gefitinib in patients progressing to EGFR TKI and presenting MET dysregulation14. Likewise, SAVANNAH (NCT03778229)15 is an ongoing phase II trial designed to evaluate the efficacy of combination savolitinib and osimertinib in post-osimertinib progression/MET-positive patients. The biomarker-matched study ORCHARD (NCT03944772)16 is also underway to assess the efficacy of several osimertinib-based combinations following disease progression under frontline osimertinib.
BRAF mutations and fusions (i.e. AGK-BRAF, ESYT2-BRAF) have recently emerged as additional mechanisms of AR to third-generation EGFR TKI2,18–20. Studies demonstrating the efficacy of concurrent inhibition of EGFR and BRAF3 or MEK21 in pre-clinical models have raised clinicians’ expectations about overcoming AR by combining TT. Nevertheless, reports of successful combinations of TT for patients harboring BRAF-driven AR to osimertinib are very limited19,22–24 and no prospective data regarding efficacy and safety of BRAF/MEK/EGFR concurrent inhibition are available, with chemotherapy-based regimens remaining the treatment of choice in this unfavorable scenario. Similarly, PIK3CA mutations may also mediate AR to second-line osimertinib in 4–11%, but no clinical reports suggesting potential benefits of blocking these alterations in NSCLC are available6,9,25–27.
The detection of circulating tumor DNA (ctDNA) using liquid biopsies allows noninvasive real-time monitoring of treatment response and early detection of AR to TT, anticipating radiological response and treatment failure28,29. The widespread availability of highly specific and sensitive techniques to quantify ctDNA makes the longitudinal assessment of patients with NSCLC under TKI therapy very attractive. Nonetheless, for NSCLCs, ctDNA evaluation in routine clinical practice is currently limited to the detection of EGFR exon 19 deletion, EGFR L858R and T790M activating mutations29.
A man with metastatic lung adenocarcinoma harboring an EGFR mutation, who had progressed to erlotinib due to the emergence of the EGFR T790M mutation started osimertinib and remained on treatment for 15 months, then developing disease progression (PD). Sequential liquid biopsies were collected to monitor treatment response and disclosed the emergence of BRAF V600E and PIK3CA E545K resistance mutations 4 months before clinical progression. With this result and considering few available reports in the literature, our patient started a triple therapy with osimertinib, dabrafenib (BRAF inhibitor), and trametinib (MEK inhibitor).
Results
Case report
A 50-year-old non-smoker man was diagnosed with a tubule-papillary lung adenocarcinoma metastatic to bones and soft tissue (stage IVB - AJCC 8th edition) in July 2016. Several hypermetabolic bone lesions were observed on staging 18-fluorodeoxyglucose positron-emission tomography (18F-FDG PET-CT) scan at diagnosis. A baseline Next Generation Sequencing (NGS) assay (TruSightTumor™ - Illumina®) of a soft tissue metastasis revealed the presence of an activating EGFR exon 19 deletion (EGFR E746_A750del). No concurrent alterations in BRAF or in PIK3CA were identified at that time. ALK d5f3 immunohistochemistry and fluorescent in situ hybridization (FISH) for ROS1 and RET resulted negative.
In September 2016, the patient started on erlotinib 150 mg once daily (OD), achieving complete metabolic response in February 2017. We started to perform serial blood sample collections for ctDNA analysis in May 2017. In October 2017, oligo-progression (oligo-PD) was observed at the primary site and treated with stereotactic body radiation therapy (SBRT; 3 × 16 Gy). In November 2017, we identified the EGFR T790M and EGFR E746_A750del mutations in patient’s plasma using droplet digital PCR (ddPCR) and these results were confirmed in December 2017 with the Foundation ACT® ctDNA assay. Figure 1a shows all systemic and focal therapies of this patient since diagnosis.Fig. 1 Paired radiological and sequential blood-based ctDNA assessments throughout patient’s treatment demonstrating concordant results.
a Timeline displaying systemic and focal therapies since September 2016. b ctDNA levels in serial plasma samples based on fractional abundance of EGFR T790M, EGFR del19 (E746_A750 del), BRAF V600E, and PIK3CA E545K mutations. #2.4 CN results: EGFR del19 WT/RNAseP copy number ratio. c Paired 18F-FDG PET-CT scan maximum intensity projection (MIP) images displaying tumor burden variations. PD progression of disease, SBRT stereotatic body radiotherapy, Pembro pembrolizumab, CT chemotherapy, DTO dabrafenib, trametinib and osimertinib.
Erlotinib was kept until January 2018 when disease progression in the bones was observed and the fractional abundance (FA) of EGFR E746_A750del and EGFR T790M were 12.0 and 2.3%, respectively (Fig. 1b). The treatment was promptly switched to osimertinib 80 mg OD. Bone partial response (PR) was observed 2 months later, along with a significant drop in the FA of EGFR E746_A750del and EGFR T790M mutations in the plasma (Fig. 1b).
In July 2018, the patient developed oligo-PD in T3 vertebrae, which was treated with SBRT (1 × 20 Gy). At that point, ctDNA analysis revealed an increase in the FA of the EGFR E746_A750del in the plasma, but not in EGFR T790M, suggesting genetic heterogeneity between metastatic lesions. In December 2018, a new oligo-PD in the left iliac bone was detected by 18F-FDG PET-CT scan (Fig. 1c). At that time, ctDNA analysis revealed the emergence of BRAF V600E (FA: 0.4%) and PIK3CA E545K (FA: 0.9%) mutations, as well as a significant increase in the FA of the EGFR E746_A750del (FA: 7.5%) in the plasma. These mutations were also detected in a specimen from an iliac bone biopsy using a NGS assay (TruSightTumor™ - Illumina® - EGFR del19 allelic fraction 81.1%, EGFR del19 amplification – 12 copies, BRAF V600E allelic fraction 17.7% and PIK3CA E545K allelic fraction 32.7%). Even though the patient was treated with SBRT (1 × 18 Gy), he developed new bone metastases in March 2019 (Fig. 1b), consistent with the significant increase in the FA of EGFR E746_A750del, BRAF V600E and EGFR T790M mutations (Fig. 1b). At that specific timepoint and in April 2019, assessments of PIK3CA E545K plasma levels were not obtained due to limited amount of cfDNA.
Between April and September 2019, systemic treatment with carboplatin plus pemetrexed and pembrolizumab provided modest clinical benefit, reducing tumor burden and controlling the disease (Fig. 1b). Due to a new PD observed in October 2019, the treatment was switched to docetaxel monotherapy and, subsequently, to vinorelbine after progression under docetaxel; nonetheless, the patient experienced symptomatic PD in bones, lymph nodes, and lung in late January 2020 (Fig. 1c, 2a–d). He came to the clinic with a Karnofsky performance status of 70%, complaining of fatigue, appetite loss, and severe pain in the hips (despite regular use of 10 mg buprenorphine patch and oxycodone plus acetaminophen PO), which had been preventing him from performing his daily activities due to the inability to stay seated. At that point, high levels of all three resistance mutations were detected in the circulating DNA (PIK3CA E545K FA: 13.2%, BRAF V600E FA: 12.3%, and EGFR T790M FA: 5.3%; Fig. 1b). We also observed an exponential increase in the number of copies of EGFR E746_A750del, suggesting gene amplification (Fig. 1b).Fig. 2 18F-FDG PET-CT scan imaging depicting impressive overall tumor response under dabrafenib, trametinib, and osimertinib in three timepoints.
a–d baseline imaging (January/2020) showing: hypermetabolic spiculated mass (orange circle) in the left-superior lobe measuring 4.4 × 3.3 cm (SUVmax: 9.4); multiple hilar, mediastinal, retroperitoneal, and iliac hypermetabolic lymph nodes (red arrow) measuring up to 2.7 cm (SUVmax: 12.8); several hypermetabolic bone lesions throughout axial and appendicular skeleton (yellow circle); a left iliac bone lesion with signs of periosteal reaction and adjacent soft-tissue infiltration (SUVmax: 11.4). e–h first response evaluation imaging (April/2020) showing considerable partial response in the lung mass, measuring 3.8 × 2.5 cm (SUVmax: 2.7), as well as in several bone lesions, especially in the left-iliac bone (SUVmax: 4.8); complete response in lymph nodes. i–l third response evaluation imaging showing disease progression in the left ischium (SUVmax: 12.5; previous SUVmax: 5.9) (October/2020). SUVmax maximum standard uptake value.
In February 2020, after careful consideration due to patient’s good performance status, normal organ functions, severe pain, motivation and the emergence of BRAF V600E as an AR mechanism, he started on dabrafenib 75 mg twice daily (BID), trametinib 1 mg OD and osimertinib 80 mg OD. The use of a PI3K inhibitor was not considered an option due to the absence of reports in the literature suggesting efficacy of these drugs combined with osimertinib to treat advanced NSCLC patients. Within 2 weeks of treatment, the patient achieved complete resolution of the severe bone pain in the hips with no further need of opioid administration, as well as appetite gain and marked improvement in quality of life, which turned possible for him to resume his daily activities. As adverse events (AE), he experienced grade 1 fatigue, dysgeusia, fever, and nausea, all managed with symptomatic medication. Complete resolution of the fever occurred spontaneously within two weeks of treatment. An attempt to increase dosages of dabrafenib to 150 mg BID and trametinib 2 mg OD resulted unsuccessful due to persistent grade 2 fatigue. In April 2020, a new 18F-FDG PET-CT scan disclosed a complete response in lymph nodes and a dramatic PR in the lung and bones (PERCIST 1.1 evaluating 5 target lesions: Δ- 67%; Figs. 1c and 2e–h). Detection of EGFR T790M, BRAF V600E, and PIK3CA E545K mutations became negative in April 2020, while EGFR E746_A750del despite marginally positive, presented marked reduction (Fig. 1b) suggesting that combination dabrafenib, trametinib, and osimertinib might be an effective strategy to overcome PIK3CA E545K and BRAF V600E-driven resistance to osimertinib in advanced EGFR-mutant NSCLC patients. The patient remained in response until October 2020, when he developed asymptomatic bone PD in lumbar spine, left ischium, and right iliac bone, 8 months after starting this combination therapy (Figs. 1c and 2i–l). A FoundationOne®Liquid CDx plasma NGS disclosed the following alterations: CHCHD3-BRAF fusion, BRAF V600E, EGFR E746_750del, EGFR amplification, PIK3CA E545K, MAP2K2 (MEK2) C125S, MTAP rearrangement intron 5, TP53 V197M, and TP53 S241A. Owing to the considerable clinical benefit, good tolerance and lack of systemic treatment options, we decided to keep the patient on treatment and increase the doses of dabrafenib (150 mg BID) and trametinib (2 mg OD) every other day. He remains asymptomatic and tolerating well the proposed dose adjustment.
Discussion
Several studies have recently highlighted the importance of considering genes of interest within the context of commonly co-occurring mutations9,30. For example, as described by Blakely et al. through performing a cfDNA NGS analysis of 1,122 advanced stage EGFR-mutant NSCLCs, in ~93% of the patients, at least one more variables with known or likely known functional properties were present, disclosing the molecular complexity of this oncogenic driver and suggesting an association of co-occuring genomic alterations with TKI response and clinical outcomes9. In addition, Roper and colleagues reported the identification of at least two co-existing AR mechanisms in 73% of patients treated with osimertinib, as well as 6–23 different subclones per individual in a phylogenetic analysis performed in multiple metastatic sites of 15 individuals30. In the same publication, the authors also called the attention for a high incidence of acquired EGFR amplifications in post-osimertinib patients, which suggests maintenance of EGFR central role in the setting of progression, as previously reported by our group28 and also detected in the present case.
Although uncommon, BRAF V600E mediates AR in approximately 3% of the patients under second-line osimertinib6,20 and little is known about the efficacy of combined TT in this population3,21–23. Existing reports lack details regarding objective responses using standardized radiological criteria (i.e., RECIST 1.1; PERCIST) and the dynamics of resistance mechanisms through longitudinal ctDNA measurements22,23. Huang and colleagues22 described a case of an EGFRdel19/T790M + NSCLC patient who developed BRAF V600E-driven AR after second-line osimertinib and achieved stable disease under dabrafenib 150 mg BID, trametinib 1 mg OD and osimertinib 80 mg OD, with ongoing disease control 7.4 months after. Zhou and colleagues23 also reported their experience with dabrafenib 150 mg BID, trametinib 2 mg OD and osimertinib 80 mg OD leading to tumor reduction within 6 weeks of treatment, along with grade 2 rash and decreased appetite as AEs. Similarly, Meng et al. reported two cases treated with this triple regimen31. The first patient discontinued therapy after one month due to severe pneumonitis; the second one presented tumor response under dabrafenib 50 mg BID, trametinib 0.5 mg OD, and osimertinib 80 mg OD, with progression-free survival of 14 months. According to the authors, this reduced dose was prescribed owing to a grade 2 pyrexia, nausea, and vomiting under higher dabrafenib and trametinib doses. Dagogo-Jack and colleagues19 also described a successful case of combined EGFR/MAP kinase pathway blockade with osimertinib 80 mg OD and trametinib 1 mg OD; as treatment-related AEs, their patient experienced grade 2 diarrhea and fatigue, along with grade 1 rash and gastrointestinal bleeding. Nevertheless, it is worth highlighting the rapid clinical improvement, the remarkable radiologically confirmed objective response, as well as the good tolerance observed in this case even using only half standard dose of dabrafenib and trametinib approved for NSCLCs harboring BRAF V600E mutations. Since data regarding the efficacy of these combined approaches, the optimal drug association and dosing, as well as the toxicity profile are conflicting and largely unknown, further investigation into the mechanistic basis of this association represents an important priority. The Table 1 summarizes the toxicities arising under osimertinib plus BRAF/MEK inhibition and reported in the literature.Table 1 Summary of the toxicities arising under osimertinib plus BRAF/MEK inhibition and reported in the literature.
Reference Treatment Anorexia Nausea Vomiting Diarrhoea Fatigue Rash GI bleeding AST/ALT elevation Paroniquia Pyrexia Initial dose Dose reduction
Meng et al. D + T + O NR G2 G2 G2 NR NR NR NR NR G2 D (150 mg BID) + T (2 mg OD) + O (80 mg OD) D (50 mg BID) + T (0.5 mg/day) + O (80 mg/day)
Dagogo-Jack et al. T + O NR NR NR G2 G2 G1 G1 G1 NR NR T (1 mg OD) + O (80 mg OD) Not needed
Huang et al. D + T + O NR NR NR G1 NR NR NR NR G1 NR D (150 mg BID) + T (1 mg OD) + O (80 mg OD) Not needed
Zhou et al. D + T + O G1 NR NR NR NR G2 NR NR NR NR D (150 mg BID) + T (2 mg OD) + O (80 mg OD) Not needed
D dabrafenib, T trametinib, O osimertinib, NR not reported.
Liquid biopsy is emerging as an important diagnostic and predictive tool in the treatment of NSCLCs. The accurate identification of predictive genetic alterations is important for both patients’ management and the understanding of clonal evolution and AR to different therapies9,29,32. Also, circulating biomarkers from multiple disease sites better reflect systemic tumor burden, including alterations from genetically different metastatic lesions, which may be missed with single-site tissue biopsies8. Here, we were able to identify and monitor multiple concurrent mechanisms of resistance throughout the entire patient’s treatment. As shown in Fig. 1b, variations in blood ctDNA levels exhibited a positive correlation with imaging findings, even in situations involving focal treatments for oligo-PD. The observed concordance between radiological PD or response and ctDNA measurements also points towards its great potential to be incorporated into clinical practice to anticipate radiologic findings in a more effective manner.
Along with BRAF V600E mutation, the presence of PIK3CA E545K mutation is also associated with AR to osimertinib6,9,25,27. However, the clinical implication of concomitant targeting of EGFR and PI3K remains unclear. Whereas an alternative argument for the observed response could be a rechallenge following a long time upon off-osimertinib, in our opinion, this is an unlikely explanation33. Interestingly, the presence of a BRAF fusion along with a MEK2 mutation identified through plasma NGS in the setting of PD might represent a mechanism of resistance to this triple regimen34. Nonetheless, since pretreatment blood samples had not been analyzed using a similar methodology, this hypothesis deserves careful interpretation.
Increased understanding of the relationship of concurring genomic alterations in EGFR-mutant NSCLC may enable new therapeutic opportunities following disease progression to osimertinib. Here, we reported an impressive objective response to dabrafenib, trametinib, and osimertinib with concordant decrease in plasma ctDNA levels in a metastatic lung cancer patient harboring EGFR E746_A750del, BRAF V600E, and PIK3CA E545K activating mutations. This case report leads to a greater understanding of the currently limited literature regarding the management of EGFR-mutant NSCLC patients with acquired BRAFV600E mutation, since it reports a successful attempt to target both alterations concurrently while providing concordant and interesting data of serial ctDNA assessments throughout the entire treatment. Further investigation to optimize the efficacy and mitigate the toxicity profile of this drug association represents an important issue.
Methods
Patient
This study was approved by Hospital Sírio-Libanês Ethics Committee (HSL-RC 2020-16). The patient provided written informed consent for blood collection, ctDNA analysis and publication of this report.
Sample collection and plasma DNA extraction
Serial blood samples were collected between May 2017 and April 2020 (Fig. 1b). Peripheral blood (20 ml) was collected into tubes containing EDTA (BD, Franklin Lakes, New Jersey). Plasma was separated from the blood within 2 hours of blood collection, as previously described28. cfDNA was extracted using QIAamp MinElute Virus Vacuum Kit (Qiagen, Hilden, Germany) and stored at −80 °C.
ctDNA-ddPCR
Cell-free DNA (cfDNA) was quantified using the RNase P Copy Number Reference Assay (Life Technologies, Carlsbad, California). A total of 3000 genome-equivalents (~10 ng of cfDNA) were analyzed per assay to achieve a detection sensitivity of 0.2%. This detection limit has been assessed by using cell line-derived genomic DNA. A total of 10 ng of input DNA with varying proportions of mutant DNA was serially diluted into wild-type DNA to obtain samples with a mutant abundance of 1%, 0.5%, 0.1%, and 0.05% and subjected to droplet digital PCR (ddPCR). ddPCR was used to quantify the circulating levels of the EGFR activating mutation (EGFR E746_A750 del) and of the resistance mutations (EGFR T790M, PIK3CA E545K, and BRAF V600E). Probes and primers were obtained from BioRad (EGFR E746_A750 del #10041170, EGFR T790M #10040782, PIK3CA E545K #10041188, and BRAF V600E #10040779; Hercules, California). ddPCR was performed on the QX200 Droplet Digital PCR System, and data were analysed using QuantaSoft software (Bio-Rad). ctDNA quantification is presented as fractional abundance (FA–the proportion of the mutant allele in total cfDNA).
Reporting summary
Further information on research design is available in the Nature Research Reporting Summary linked to this article.
Supplementary information
Reporting Summary
Dataset 1
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
These authors contributed equally: Maurício Fernando Silva Almeida Ribeiro, Franciele Hinterholz Knebel.
Supplementary information
The online version contains supplementary material available at 10.1038/s41698-021-00149-4.
Acknowledgements
We thank the clinical research team at Hospital Sírio-Libanês for their assistance collecting samples. We thank Ernande Xavier dos Santos and Elisângela Monteiro for the outstanding technical support. We also thank Mrs. Dina Binzagr and Vivian Hannud for supporting the Translational Research Program at Hospital Sírio Libanês. This work was supported by Ludwig Cancer Research and Fundação de Amparo à Pesquisa do Estado de São Paulo – FAPESP. Lastly, we would like to thank the patient and his family for consenting to the present publication.
Author contributions
Conceptualization by M.F.S.A.R., F.H.K., J.V.M.A., A.A.C., and A.K.; Methodology by F.H.K. and A.A.C.; Resources br M.F.S.A.R., F.H.K., A.A.C., and J.V.M.A.; Writing of the first draft by M.F.S.A., F.H.K., and J.V.M.A.; Review, editing and approval of the final draft by all authors; Supervision by A.A.C. and A.K.; Funding acquisition by F.H.K. and A.A.C. M.F.S.A.R. and F.H.K. are co-first authors of the present work.
Data availability
The datasets that support the findings of this study are not publicly available in order to protect patient privacy. The data will be made available on reasonable request. For data access requests regarding the liquid biopsy (ctDNA quantification) data, please contact Dr. Franciele Knebel, email address: fhknebel@mochsl.org.br. For data access requests regarding the PET-CT high resolution images and PERCIST calculations, please contact Dr. José Marin, email address:jfgmarin@yahoo.com.br. For data access requests regarding the summary of the toxicities arising under osimertinib plus BRAF/MEK inhibition, please contact the corresponding author Dr. Maurício Ribeiro, email address: mauricio.fsaribeiro@hsl.org.br. The data generated and analysed during this study are described in the following metadata record: 10.6084/m9.figshare.1347594635.
Competing interests
The authors declare no competing interests. | DABRAFENIB, OSIMERTINIB, TRAMETINIB | DrugsGivenReaction | CC BY | 33580193 | 19,431,708 | 2021-02-12 |
What was the administration route of drug 'ACETAMINOPHEN\OXYCODONE HYDROCHLORIDE'? | Impressive response to dabrafenib, trametinib, and osimertinib in a metastatic EGFR-mutant/BRAF V600E lung adenocarcinoma patient.
The survival outcomes of the FLAURA trial support osimertinib as the new standard of care for untreated patients harboring activating mutations in the epidermal growth factor receptor (EGFR). Despite the initial response, disease progression invariably occurs. Although uncommon, BRAF V600E mutation arises as a unique mechanism of resistance, and thus far, no prospective studies are available to support concurrent EGFR/BRAF blockade. We report a case of impressive radiological and ctDNA response under dabrafenib, trametinib, and osimertinib in an advanced EGFR-mutant lung adenocarcinoma patient who developed BRAF V600E as one of the acquired resistance mechanisms to second-line osimertinib. Moreover, the patient experienced remarkable clinical improvement and good tolerance to combination therapy. The present case suggests the importance of prospective studies evaluating both efficacy and safety of the combination in later line settings and points towards the potential of ctDNA to monitor resistance mechanisms and treatment benefit in clinical practice.
Introduction
The use of osimertinib to target epidermal growth factor receptor (EGFR) has become the standard of care in untreated EGFR-mutant non–small cell lung cancer (NSCLC) patients. Although osimertinib can be highly active, showing more durable outcomes than first-generation tyrosine kinase inhibitors (TKI)1, most tumors invariably become resistant, limiting its long-term clinical benefit. The heterogeneity of resistance mechanisms to osimertinib, including EGFR C797S mutation, EGFR, and MET amplifications, off-target mutations in PIK3CA, KRAS, and HER2 as well as histologic transformation2–9, has stimulated routine performance of repeated biopsies to identify specific underlying mechanisms of resistance throughout the treatment course, and to guide the development of novel therapeutic strategies to overcome and prevent acquired resistance (AR)7,10–13.
Combined targeted therapy (TT) strategies have been increasingly addressed in prospective clinical trials14–17. A phase Ib/II trial reported 47% ORR with a combination of capmatinib (MET inhibitor) plus gefitinib in patients progressing to EGFR TKI and presenting MET dysregulation14. Likewise, SAVANNAH (NCT03778229)15 is an ongoing phase II trial designed to evaluate the efficacy of combination savolitinib and osimertinib in post-osimertinib progression/MET-positive patients. The biomarker-matched study ORCHARD (NCT03944772)16 is also underway to assess the efficacy of several osimertinib-based combinations following disease progression under frontline osimertinib.
BRAF mutations and fusions (i.e. AGK-BRAF, ESYT2-BRAF) have recently emerged as additional mechanisms of AR to third-generation EGFR TKI2,18–20. Studies demonstrating the efficacy of concurrent inhibition of EGFR and BRAF3 or MEK21 in pre-clinical models have raised clinicians’ expectations about overcoming AR by combining TT. Nevertheless, reports of successful combinations of TT for patients harboring BRAF-driven AR to osimertinib are very limited19,22–24 and no prospective data regarding efficacy and safety of BRAF/MEK/EGFR concurrent inhibition are available, with chemotherapy-based regimens remaining the treatment of choice in this unfavorable scenario. Similarly, PIK3CA mutations may also mediate AR to second-line osimertinib in 4–11%, but no clinical reports suggesting potential benefits of blocking these alterations in NSCLC are available6,9,25–27.
The detection of circulating tumor DNA (ctDNA) using liquid biopsies allows noninvasive real-time monitoring of treatment response and early detection of AR to TT, anticipating radiological response and treatment failure28,29. The widespread availability of highly specific and sensitive techniques to quantify ctDNA makes the longitudinal assessment of patients with NSCLC under TKI therapy very attractive. Nonetheless, for NSCLCs, ctDNA evaluation in routine clinical practice is currently limited to the detection of EGFR exon 19 deletion, EGFR L858R and T790M activating mutations29.
A man with metastatic lung adenocarcinoma harboring an EGFR mutation, who had progressed to erlotinib due to the emergence of the EGFR T790M mutation started osimertinib and remained on treatment for 15 months, then developing disease progression (PD). Sequential liquid biopsies were collected to monitor treatment response and disclosed the emergence of BRAF V600E and PIK3CA E545K resistance mutations 4 months before clinical progression. With this result and considering few available reports in the literature, our patient started a triple therapy with osimertinib, dabrafenib (BRAF inhibitor), and trametinib (MEK inhibitor).
Results
Case report
A 50-year-old non-smoker man was diagnosed with a tubule-papillary lung adenocarcinoma metastatic to bones and soft tissue (stage IVB - AJCC 8th edition) in July 2016. Several hypermetabolic bone lesions were observed on staging 18-fluorodeoxyglucose positron-emission tomography (18F-FDG PET-CT) scan at diagnosis. A baseline Next Generation Sequencing (NGS) assay (TruSightTumor™ - Illumina®) of a soft tissue metastasis revealed the presence of an activating EGFR exon 19 deletion (EGFR E746_A750del). No concurrent alterations in BRAF or in PIK3CA were identified at that time. ALK d5f3 immunohistochemistry and fluorescent in situ hybridization (FISH) for ROS1 and RET resulted negative.
In September 2016, the patient started on erlotinib 150 mg once daily (OD), achieving complete metabolic response in February 2017. We started to perform serial blood sample collections for ctDNA analysis in May 2017. In October 2017, oligo-progression (oligo-PD) was observed at the primary site and treated with stereotactic body radiation therapy (SBRT; 3 × 16 Gy). In November 2017, we identified the EGFR T790M and EGFR E746_A750del mutations in patient’s plasma using droplet digital PCR (ddPCR) and these results were confirmed in December 2017 with the Foundation ACT® ctDNA assay. Figure 1a shows all systemic and focal therapies of this patient since diagnosis.Fig. 1 Paired radiological and sequential blood-based ctDNA assessments throughout patient’s treatment demonstrating concordant results.
a Timeline displaying systemic and focal therapies since September 2016. b ctDNA levels in serial plasma samples based on fractional abundance of EGFR T790M, EGFR del19 (E746_A750 del), BRAF V600E, and PIK3CA E545K mutations. #2.4 CN results: EGFR del19 WT/RNAseP copy number ratio. c Paired 18F-FDG PET-CT scan maximum intensity projection (MIP) images displaying tumor burden variations. PD progression of disease, SBRT stereotatic body radiotherapy, Pembro pembrolizumab, CT chemotherapy, DTO dabrafenib, trametinib and osimertinib.
Erlotinib was kept until January 2018 when disease progression in the bones was observed and the fractional abundance (FA) of EGFR E746_A750del and EGFR T790M were 12.0 and 2.3%, respectively (Fig. 1b). The treatment was promptly switched to osimertinib 80 mg OD. Bone partial response (PR) was observed 2 months later, along with a significant drop in the FA of EGFR E746_A750del and EGFR T790M mutations in the plasma (Fig. 1b).
In July 2018, the patient developed oligo-PD in T3 vertebrae, which was treated with SBRT (1 × 20 Gy). At that point, ctDNA analysis revealed an increase in the FA of the EGFR E746_A750del in the plasma, but not in EGFR T790M, suggesting genetic heterogeneity between metastatic lesions. In December 2018, a new oligo-PD in the left iliac bone was detected by 18F-FDG PET-CT scan (Fig. 1c). At that time, ctDNA analysis revealed the emergence of BRAF V600E (FA: 0.4%) and PIK3CA E545K (FA: 0.9%) mutations, as well as a significant increase in the FA of the EGFR E746_A750del (FA: 7.5%) in the plasma. These mutations were also detected in a specimen from an iliac bone biopsy using a NGS assay (TruSightTumor™ - Illumina® - EGFR del19 allelic fraction 81.1%, EGFR del19 amplification – 12 copies, BRAF V600E allelic fraction 17.7% and PIK3CA E545K allelic fraction 32.7%). Even though the patient was treated with SBRT (1 × 18 Gy), he developed new bone metastases in March 2019 (Fig. 1b), consistent with the significant increase in the FA of EGFR E746_A750del, BRAF V600E and EGFR T790M mutations (Fig. 1b). At that specific timepoint and in April 2019, assessments of PIK3CA E545K plasma levels were not obtained due to limited amount of cfDNA.
Between April and September 2019, systemic treatment with carboplatin plus pemetrexed and pembrolizumab provided modest clinical benefit, reducing tumor burden and controlling the disease (Fig. 1b). Due to a new PD observed in October 2019, the treatment was switched to docetaxel monotherapy and, subsequently, to vinorelbine after progression under docetaxel; nonetheless, the patient experienced symptomatic PD in bones, lymph nodes, and lung in late January 2020 (Fig. 1c, 2a–d). He came to the clinic with a Karnofsky performance status of 70%, complaining of fatigue, appetite loss, and severe pain in the hips (despite regular use of 10 mg buprenorphine patch and oxycodone plus acetaminophen PO), which had been preventing him from performing his daily activities due to the inability to stay seated. At that point, high levels of all three resistance mutations were detected in the circulating DNA (PIK3CA E545K FA: 13.2%, BRAF V600E FA: 12.3%, and EGFR T790M FA: 5.3%; Fig. 1b). We also observed an exponential increase in the number of copies of EGFR E746_A750del, suggesting gene amplification (Fig. 1b).Fig. 2 18F-FDG PET-CT scan imaging depicting impressive overall tumor response under dabrafenib, trametinib, and osimertinib in three timepoints.
a–d baseline imaging (January/2020) showing: hypermetabolic spiculated mass (orange circle) in the left-superior lobe measuring 4.4 × 3.3 cm (SUVmax: 9.4); multiple hilar, mediastinal, retroperitoneal, and iliac hypermetabolic lymph nodes (red arrow) measuring up to 2.7 cm (SUVmax: 12.8); several hypermetabolic bone lesions throughout axial and appendicular skeleton (yellow circle); a left iliac bone lesion with signs of periosteal reaction and adjacent soft-tissue infiltration (SUVmax: 11.4). e–h first response evaluation imaging (April/2020) showing considerable partial response in the lung mass, measuring 3.8 × 2.5 cm (SUVmax: 2.7), as well as in several bone lesions, especially in the left-iliac bone (SUVmax: 4.8); complete response in lymph nodes. i–l third response evaluation imaging showing disease progression in the left ischium (SUVmax: 12.5; previous SUVmax: 5.9) (October/2020). SUVmax maximum standard uptake value.
In February 2020, after careful consideration due to patient’s good performance status, normal organ functions, severe pain, motivation and the emergence of BRAF V600E as an AR mechanism, he started on dabrafenib 75 mg twice daily (BID), trametinib 1 mg OD and osimertinib 80 mg OD. The use of a PI3K inhibitor was not considered an option due to the absence of reports in the literature suggesting efficacy of these drugs combined with osimertinib to treat advanced NSCLC patients. Within 2 weeks of treatment, the patient achieved complete resolution of the severe bone pain in the hips with no further need of opioid administration, as well as appetite gain and marked improvement in quality of life, which turned possible for him to resume his daily activities. As adverse events (AE), he experienced grade 1 fatigue, dysgeusia, fever, and nausea, all managed with symptomatic medication. Complete resolution of the fever occurred spontaneously within two weeks of treatment. An attempt to increase dosages of dabrafenib to 150 mg BID and trametinib 2 mg OD resulted unsuccessful due to persistent grade 2 fatigue. In April 2020, a new 18F-FDG PET-CT scan disclosed a complete response in lymph nodes and a dramatic PR in the lung and bones (PERCIST 1.1 evaluating 5 target lesions: Δ- 67%; Figs. 1c and 2e–h). Detection of EGFR T790M, BRAF V600E, and PIK3CA E545K mutations became negative in April 2020, while EGFR E746_A750del despite marginally positive, presented marked reduction (Fig. 1b) suggesting that combination dabrafenib, trametinib, and osimertinib might be an effective strategy to overcome PIK3CA E545K and BRAF V600E-driven resistance to osimertinib in advanced EGFR-mutant NSCLC patients. The patient remained in response until October 2020, when he developed asymptomatic bone PD in lumbar spine, left ischium, and right iliac bone, 8 months after starting this combination therapy (Figs. 1c and 2i–l). A FoundationOne®Liquid CDx plasma NGS disclosed the following alterations: CHCHD3-BRAF fusion, BRAF V600E, EGFR E746_750del, EGFR amplification, PIK3CA E545K, MAP2K2 (MEK2) C125S, MTAP rearrangement intron 5, TP53 V197M, and TP53 S241A. Owing to the considerable clinical benefit, good tolerance and lack of systemic treatment options, we decided to keep the patient on treatment and increase the doses of dabrafenib (150 mg BID) and trametinib (2 mg OD) every other day. He remains asymptomatic and tolerating well the proposed dose adjustment.
Discussion
Several studies have recently highlighted the importance of considering genes of interest within the context of commonly co-occurring mutations9,30. For example, as described by Blakely et al. through performing a cfDNA NGS analysis of 1,122 advanced stage EGFR-mutant NSCLCs, in ~93% of the patients, at least one more variables with known or likely known functional properties were present, disclosing the molecular complexity of this oncogenic driver and suggesting an association of co-occuring genomic alterations with TKI response and clinical outcomes9. In addition, Roper and colleagues reported the identification of at least two co-existing AR mechanisms in 73% of patients treated with osimertinib, as well as 6–23 different subclones per individual in a phylogenetic analysis performed in multiple metastatic sites of 15 individuals30. In the same publication, the authors also called the attention for a high incidence of acquired EGFR amplifications in post-osimertinib patients, which suggests maintenance of EGFR central role in the setting of progression, as previously reported by our group28 and also detected in the present case.
Although uncommon, BRAF V600E mediates AR in approximately 3% of the patients under second-line osimertinib6,20 and little is known about the efficacy of combined TT in this population3,21–23. Existing reports lack details regarding objective responses using standardized radiological criteria (i.e., RECIST 1.1; PERCIST) and the dynamics of resistance mechanisms through longitudinal ctDNA measurements22,23. Huang and colleagues22 described a case of an EGFRdel19/T790M + NSCLC patient who developed BRAF V600E-driven AR after second-line osimertinib and achieved stable disease under dabrafenib 150 mg BID, trametinib 1 mg OD and osimertinib 80 mg OD, with ongoing disease control 7.4 months after. Zhou and colleagues23 also reported their experience with dabrafenib 150 mg BID, trametinib 2 mg OD and osimertinib 80 mg OD leading to tumor reduction within 6 weeks of treatment, along with grade 2 rash and decreased appetite as AEs. Similarly, Meng et al. reported two cases treated with this triple regimen31. The first patient discontinued therapy after one month due to severe pneumonitis; the second one presented tumor response under dabrafenib 50 mg BID, trametinib 0.5 mg OD, and osimertinib 80 mg OD, with progression-free survival of 14 months. According to the authors, this reduced dose was prescribed owing to a grade 2 pyrexia, nausea, and vomiting under higher dabrafenib and trametinib doses. Dagogo-Jack and colleagues19 also described a successful case of combined EGFR/MAP kinase pathway blockade with osimertinib 80 mg OD and trametinib 1 mg OD; as treatment-related AEs, their patient experienced grade 2 diarrhea and fatigue, along with grade 1 rash and gastrointestinal bleeding. Nevertheless, it is worth highlighting the rapid clinical improvement, the remarkable radiologically confirmed objective response, as well as the good tolerance observed in this case even using only half standard dose of dabrafenib and trametinib approved for NSCLCs harboring BRAF V600E mutations. Since data regarding the efficacy of these combined approaches, the optimal drug association and dosing, as well as the toxicity profile are conflicting and largely unknown, further investigation into the mechanistic basis of this association represents an important priority. The Table 1 summarizes the toxicities arising under osimertinib plus BRAF/MEK inhibition and reported in the literature.Table 1 Summary of the toxicities arising under osimertinib plus BRAF/MEK inhibition and reported in the literature.
Reference Treatment Anorexia Nausea Vomiting Diarrhoea Fatigue Rash GI bleeding AST/ALT elevation Paroniquia Pyrexia Initial dose Dose reduction
Meng et al. D + T + O NR G2 G2 G2 NR NR NR NR NR G2 D (150 mg BID) + T (2 mg OD) + O (80 mg OD) D (50 mg BID) + T (0.5 mg/day) + O (80 mg/day)
Dagogo-Jack et al. T + O NR NR NR G2 G2 G1 G1 G1 NR NR T (1 mg OD) + O (80 mg OD) Not needed
Huang et al. D + T + O NR NR NR G1 NR NR NR NR G1 NR D (150 mg BID) + T (1 mg OD) + O (80 mg OD) Not needed
Zhou et al. D + T + O G1 NR NR NR NR G2 NR NR NR NR D (150 mg BID) + T (2 mg OD) + O (80 mg OD) Not needed
D dabrafenib, T trametinib, O osimertinib, NR not reported.
Liquid biopsy is emerging as an important diagnostic and predictive tool in the treatment of NSCLCs. The accurate identification of predictive genetic alterations is important for both patients’ management and the understanding of clonal evolution and AR to different therapies9,29,32. Also, circulating biomarkers from multiple disease sites better reflect systemic tumor burden, including alterations from genetically different metastatic lesions, which may be missed with single-site tissue biopsies8. Here, we were able to identify and monitor multiple concurrent mechanisms of resistance throughout the entire patient’s treatment. As shown in Fig. 1b, variations in blood ctDNA levels exhibited a positive correlation with imaging findings, even in situations involving focal treatments for oligo-PD. The observed concordance between radiological PD or response and ctDNA measurements also points towards its great potential to be incorporated into clinical practice to anticipate radiologic findings in a more effective manner.
Along with BRAF V600E mutation, the presence of PIK3CA E545K mutation is also associated with AR to osimertinib6,9,25,27. However, the clinical implication of concomitant targeting of EGFR and PI3K remains unclear. Whereas an alternative argument for the observed response could be a rechallenge following a long time upon off-osimertinib, in our opinion, this is an unlikely explanation33. Interestingly, the presence of a BRAF fusion along with a MEK2 mutation identified through plasma NGS in the setting of PD might represent a mechanism of resistance to this triple regimen34. Nonetheless, since pretreatment blood samples had not been analyzed using a similar methodology, this hypothesis deserves careful interpretation.
Increased understanding of the relationship of concurring genomic alterations in EGFR-mutant NSCLC may enable new therapeutic opportunities following disease progression to osimertinib. Here, we reported an impressive objective response to dabrafenib, trametinib, and osimertinib with concordant decrease in plasma ctDNA levels in a metastatic lung cancer patient harboring EGFR E746_A750del, BRAF V600E, and PIK3CA E545K activating mutations. This case report leads to a greater understanding of the currently limited literature regarding the management of EGFR-mutant NSCLC patients with acquired BRAFV600E mutation, since it reports a successful attempt to target both alterations concurrently while providing concordant and interesting data of serial ctDNA assessments throughout the entire treatment. Further investigation to optimize the efficacy and mitigate the toxicity profile of this drug association represents an important issue.
Methods
Patient
This study was approved by Hospital Sírio-Libanês Ethics Committee (HSL-RC 2020-16). The patient provided written informed consent for blood collection, ctDNA analysis and publication of this report.
Sample collection and plasma DNA extraction
Serial blood samples were collected between May 2017 and April 2020 (Fig. 1b). Peripheral blood (20 ml) was collected into tubes containing EDTA (BD, Franklin Lakes, New Jersey). Plasma was separated from the blood within 2 hours of blood collection, as previously described28. cfDNA was extracted using QIAamp MinElute Virus Vacuum Kit (Qiagen, Hilden, Germany) and stored at −80 °C.
ctDNA-ddPCR
Cell-free DNA (cfDNA) was quantified using the RNase P Copy Number Reference Assay (Life Technologies, Carlsbad, California). A total of 3000 genome-equivalents (~10 ng of cfDNA) were analyzed per assay to achieve a detection sensitivity of 0.2%. This detection limit has been assessed by using cell line-derived genomic DNA. A total of 10 ng of input DNA with varying proportions of mutant DNA was serially diluted into wild-type DNA to obtain samples with a mutant abundance of 1%, 0.5%, 0.1%, and 0.05% and subjected to droplet digital PCR (ddPCR). ddPCR was used to quantify the circulating levels of the EGFR activating mutation (EGFR E746_A750 del) and of the resistance mutations (EGFR T790M, PIK3CA E545K, and BRAF V600E). Probes and primers were obtained from BioRad (EGFR E746_A750 del #10041170, EGFR T790M #10040782, PIK3CA E545K #10041188, and BRAF V600E #10040779; Hercules, California). ddPCR was performed on the QX200 Droplet Digital PCR System, and data were analysed using QuantaSoft software (Bio-Rad). ctDNA quantification is presented as fractional abundance (FA–the proportion of the mutant allele in total cfDNA).
Reporting summary
Further information on research design is available in the Nature Research Reporting Summary linked to this article.
Supplementary information
Reporting Summary
Dataset 1
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
These authors contributed equally: Maurício Fernando Silva Almeida Ribeiro, Franciele Hinterholz Knebel.
Supplementary information
The online version contains supplementary material available at 10.1038/s41698-021-00149-4.
Acknowledgements
We thank the clinical research team at Hospital Sírio-Libanês for their assistance collecting samples. We thank Ernande Xavier dos Santos and Elisângela Monteiro for the outstanding technical support. We also thank Mrs. Dina Binzagr and Vivian Hannud for supporting the Translational Research Program at Hospital Sírio Libanês. This work was supported by Ludwig Cancer Research and Fundação de Amparo à Pesquisa do Estado de São Paulo – FAPESP. Lastly, we would like to thank the patient and his family for consenting to the present publication.
Author contributions
Conceptualization by M.F.S.A.R., F.H.K., J.V.M.A., A.A.C., and A.K.; Methodology by F.H.K. and A.A.C.; Resources br M.F.S.A.R., F.H.K., A.A.C., and J.V.M.A.; Writing of the first draft by M.F.S.A., F.H.K., and J.V.M.A.; Review, editing and approval of the final draft by all authors; Supervision by A.A.C. and A.K.; Funding acquisition by F.H.K. and A.A.C. M.F.S.A.R. and F.H.K. are co-first authors of the present work.
Data availability
The datasets that support the findings of this study are not publicly available in order to protect patient privacy. The data will be made available on reasonable request. For data access requests regarding the liquid biopsy (ctDNA quantification) data, please contact Dr. Franciele Knebel, email address: fhknebel@mochsl.org.br. For data access requests regarding the PET-CT high resolution images and PERCIST calculations, please contact Dr. José Marin, email address:jfgmarin@yahoo.com.br. For data access requests regarding the summary of the toxicities arising under osimertinib plus BRAF/MEK inhibition, please contact the corresponding author Dr. Maurício Ribeiro, email address: mauricio.fsaribeiro@hsl.org.br. The data generated and analysed during this study are described in the following metadata record: 10.6084/m9.figshare.1347594635.
Competing interests
The authors declare no competing interests. | Oral | DrugAdministrationRoute | CC BY | 33580193 | 19,460,717 | 2021-02-12 |
What was the outcome of reaction 'Pyrexia'? | Impressive response to dabrafenib, trametinib, and osimertinib in a metastatic EGFR-mutant/BRAF V600E lung adenocarcinoma patient.
The survival outcomes of the FLAURA trial support osimertinib as the new standard of care for untreated patients harboring activating mutations in the epidermal growth factor receptor (EGFR). Despite the initial response, disease progression invariably occurs. Although uncommon, BRAF V600E mutation arises as a unique mechanism of resistance, and thus far, no prospective studies are available to support concurrent EGFR/BRAF blockade. We report a case of impressive radiological and ctDNA response under dabrafenib, trametinib, and osimertinib in an advanced EGFR-mutant lung adenocarcinoma patient who developed BRAF V600E as one of the acquired resistance mechanisms to second-line osimertinib. Moreover, the patient experienced remarkable clinical improvement and good tolerance to combination therapy. The present case suggests the importance of prospective studies evaluating both efficacy and safety of the combination in later line settings and points towards the potential of ctDNA to monitor resistance mechanisms and treatment benefit in clinical practice.
Introduction
The use of osimertinib to target epidermal growth factor receptor (EGFR) has become the standard of care in untreated EGFR-mutant non–small cell lung cancer (NSCLC) patients. Although osimertinib can be highly active, showing more durable outcomes than first-generation tyrosine kinase inhibitors (TKI)1, most tumors invariably become resistant, limiting its long-term clinical benefit. The heterogeneity of resistance mechanisms to osimertinib, including EGFR C797S mutation, EGFR, and MET amplifications, off-target mutations in PIK3CA, KRAS, and HER2 as well as histologic transformation2–9, has stimulated routine performance of repeated biopsies to identify specific underlying mechanisms of resistance throughout the treatment course, and to guide the development of novel therapeutic strategies to overcome and prevent acquired resistance (AR)7,10–13.
Combined targeted therapy (TT) strategies have been increasingly addressed in prospective clinical trials14–17. A phase Ib/II trial reported 47% ORR with a combination of capmatinib (MET inhibitor) plus gefitinib in patients progressing to EGFR TKI and presenting MET dysregulation14. Likewise, SAVANNAH (NCT03778229)15 is an ongoing phase II trial designed to evaluate the efficacy of combination savolitinib and osimertinib in post-osimertinib progression/MET-positive patients. The biomarker-matched study ORCHARD (NCT03944772)16 is also underway to assess the efficacy of several osimertinib-based combinations following disease progression under frontline osimertinib.
BRAF mutations and fusions (i.e. AGK-BRAF, ESYT2-BRAF) have recently emerged as additional mechanisms of AR to third-generation EGFR TKI2,18–20. Studies demonstrating the efficacy of concurrent inhibition of EGFR and BRAF3 or MEK21 in pre-clinical models have raised clinicians’ expectations about overcoming AR by combining TT. Nevertheless, reports of successful combinations of TT for patients harboring BRAF-driven AR to osimertinib are very limited19,22–24 and no prospective data regarding efficacy and safety of BRAF/MEK/EGFR concurrent inhibition are available, with chemotherapy-based regimens remaining the treatment of choice in this unfavorable scenario. Similarly, PIK3CA mutations may also mediate AR to second-line osimertinib in 4–11%, but no clinical reports suggesting potential benefits of blocking these alterations in NSCLC are available6,9,25–27.
The detection of circulating tumor DNA (ctDNA) using liquid biopsies allows noninvasive real-time monitoring of treatment response and early detection of AR to TT, anticipating radiological response and treatment failure28,29. The widespread availability of highly specific and sensitive techniques to quantify ctDNA makes the longitudinal assessment of patients with NSCLC under TKI therapy very attractive. Nonetheless, for NSCLCs, ctDNA evaluation in routine clinical practice is currently limited to the detection of EGFR exon 19 deletion, EGFR L858R and T790M activating mutations29.
A man with metastatic lung adenocarcinoma harboring an EGFR mutation, who had progressed to erlotinib due to the emergence of the EGFR T790M mutation started osimertinib and remained on treatment for 15 months, then developing disease progression (PD). Sequential liquid biopsies were collected to monitor treatment response and disclosed the emergence of BRAF V600E and PIK3CA E545K resistance mutations 4 months before clinical progression. With this result and considering few available reports in the literature, our patient started a triple therapy with osimertinib, dabrafenib (BRAF inhibitor), and trametinib (MEK inhibitor).
Results
Case report
A 50-year-old non-smoker man was diagnosed with a tubule-papillary lung adenocarcinoma metastatic to bones and soft tissue (stage IVB - AJCC 8th edition) in July 2016. Several hypermetabolic bone lesions were observed on staging 18-fluorodeoxyglucose positron-emission tomography (18F-FDG PET-CT) scan at diagnosis. A baseline Next Generation Sequencing (NGS) assay (TruSightTumor™ - Illumina®) of a soft tissue metastasis revealed the presence of an activating EGFR exon 19 deletion (EGFR E746_A750del). No concurrent alterations in BRAF or in PIK3CA were identified at that time. ALK d5f3 immunohistochemistry and fluorescent in situ hybridization (FISH) for ROS1 and RET resulted negative.
In September 2016, the patient started on erlotinib 150 mg once daily (OD), achieving complete metabolic response in February 2017. We started to perform serial blood sample collections for ctDNA analysis in May 2017. In October 2017, oligo-progression (oligo-PD) was observed at the primary site and treated with stereotactic body radiation therapy (SBRT; 3 × 16 Gy). In November 2017, we identified the EGFR T790M and EGFR E746_A750del mutations in patient’s plasma using droplet digital PCR (ddPCR) and these results were confirmed in December 2017 with the Foundation ACT® ctDNA assay. Figure 1a shows all systemic and focal therapies of this patient since diagnosis.Fig. 1 Paired radiological and sequential blood-based ctDNA assessments throughout patient’s treatment demonstrating concordant results.
a Timeline displaying systemic and focal therapies since September 2016. b ctDNA levels in serial plasma samples based on fractional abundance of EGFR T790M, EGFR del19 (E746_A750 del), BRAF V600E, and PIK3CA E545K mutations. #2.4 CN results: EGFR del19 WT/RNAseP copy number ratio. c Paired 18F-FDG PET-CT scan maximum intensity projection (MIP) images displaying tumor burden variations. PD progression of disease, SBRT stereotatic body radiotherapy, Pembro pembrolizumab, CT chemotherapy, DTO dabrafenib, trametinib and osimertinib.
Erlotinib was kept until January 2018 when disease progression in the bones was observed and the fractional abundance (FA) of EGFR E746_A750del and EGFR T790M were 12.0 and 2.3%, respectively (Fig. 1b). The treatment was promptly switched to osimertinib 80 mg OD. Bone partial response (PR) was observed 2 months later, along with a significant drop in the FA of EGFR E746_A750del and EGFR T790M mutations in the plasma (Fig. 1b).
In July 2018, the patient developed oligo-PD in T3 vertebrae, which was treated with SBRT (1 × 20 Gy). At that point, ctDNA analysis revealed an increase in the FA of the EGFR E746_A750del in the plasma, but not in EGFR T790M, suggesting genetic heterogeneity between metastatic lesions. In December 2018, a new oligo-PD in the left iliac bone was detected by 18F-FDG PET-CT scan (Fig. 1c). At that time, ctDNA analysis revealed the emergence of BRAF V600E (FA: 0.4%) and PIK3CA E545K (FA: 0.9%) mutations, as well as a significant increase in the FA of the EGFR E746_A750del (FA: 7.5%) in the plasma. These mutations were also detected in a specimen from an iliac bone biopsy using a NGS assay (TruSightTumor™ - Illumina® - EGFR del19 allelic fraction 81.1%, EGFR del19 amplification – 12 copies, BRAF V600E allelic fraction 17.7% and PIK3CA E545K allelic fraction 32.7%). Even though the patient was treated with SBRT (1 × 18 Gy), he developed new bone metastases in March 2019 (Fig. 1b), consistent with the significant increase in the FA of EGFR E746_A750del, BRAF V600E and EGFR T790M mutations (Fig. 1b). At that specific timepoint and in April 2019, assessments of PIK3CA E545K plasma levels were not obtained due to limited amount of cfDNA.
Between April and September 2019, systemic treatment with carboplatin plus pemetrexed and pembrolizumab provided modest clinical benefit, reducing tumor burden and controlling the disease (Fig. 1b). Due to a new PD observed in October 2019, the treatment was switched to docetaxel monotherapy and, subsequently, to vinorelbine after progression under docetaxel; nonetheless, the patient experienced symptomatic PD in bones, lymph nodes, and lung in late January 2020 (Fig. 1c, 2a–d). He came to the clinic with a Karnofsky performance status of 70%, complaining of fatigue, appetite loss, and severe pain in the hips (despite regular use of 10 mg buprenorphine patch and oxycodone plus acetaminophen PO), which had been preventing him from performing his daily activities due to the inability to stay seated. At that point, high levels of all three resistance mutations were detected in the circulating DNA (PIK3CA E545K FA: 13.2%, BRAF V600E FA: 12.3%, and EGFR T790M FA: 5.3%; Fig. 1b). We also observed an exponential increase in the number of copies of EGFR E746_A750del, suggesting gene amplification (Fig. 1b).Fig. 2 18F-FDG PET-CT scan imaging depicting impressive overall tumor response under dabrafenib, trametinib, and osimertinib in three timepoints.
a–d baseline imaging (January/2020) showing: hypermetabolic spiculated mass (orange circle) in the left-superior lobe measuring 4.4 × 3.3 cm (SUVmax: 9.4); multiple hilar, mediastinal, retroperitoneal, and iliac hypermetabolic lymph nodes (red arrow) measuring up to 2.7 cm (SUVmax: 12.8); several hypermetabolic bone lesions throughout axial and appendicular skeleton (yellow circle); a left iliac bone lesion with signs of periosteal reaction and adjacent soft-tissue infiltration (SUVmax: 11.4). e–h first response evaluation imaging (April/2020) showing considerable partial response in the lung mass, measuring 3.8 × 2.5 cm (SUVmax: 2.7), as well as in several bone lesions, especially in the left-iliac bone (SUVmax: 4.8); complete response in lymph nodes. i–l third response evaluation imaging showing disease progression in the left ischium (SUVmax: 12.5; previous SUVmax: 5.9) (October/2020). SUVmax maximum standard uptake value.
In February 2020, after careful consideration due to patient’s good performance status, normal organ functions, severe pain, motivation and the emergence of BRAF V600E as an AR mechanism, he started on dabrafenib 75 mg twice daily (BID), trametinib 1 mg OD and osimertinib 80 mg OD. The use of a PI3K inhibitor was not considered an option due to the absence of reports in the literature suggesting efficacy of these drugs combined with osimertinib to treat advanced NSCLC patients. Within 2 weeks of treatment, the patient achieved complete resolution of the severe bone pain in the hips with no further need of opioid administration, as well as appetite gain and marked improvement in quality of life, which turned possible for him to resume his daily activities. As adverse events (AE), he experienced grade 1 fatigue, dysgeusia, fever, and nausea, all managed with symptomatic medication. Complete resolution of the fever occurred spontaneously within two weeks of treatment. An attempt to increase dosages of dabrafenib to 150 mg BID and trametinib 2 mg OD resulted unsuccessful due to persistent grade 2 fatigue. In April 2020, a new 18F-FDG PET-CT scan disclosed a complete response in lymph nodes and a dramatic PR in the lung and bones (PERCIST 1.1 evaluating 5 target lesions: Δ- 67%; Figs. 1c and 2e–h). Detection of EGFR T790M, BRAF V600E, and PIK3CA E545K mutations became negative in April 2020, while EGFR E746_A750del despite marginally positive, presented marked reduction (Fig. 1b) suggesting that combination dabrafenib, trametinib, and osimertinib might be an effective strategy to overcome PIK3CA E545K and BRAF V600E-driven resistance to osimertinib in advanced EGFR-mutant NSCLC patients. The patient remained in response until October 2020, when he developed asymptomatic bone PD in lumbar spine, left ischium, and right iliac bone, 8 months after starting this combination therapy (Figs. 1c and 2i–l). A FoundationOne®Liquid CDx plasma NGS disclosed the following alterations: CHCHD3-BRAF fusion, BRAF V600E, EGFR E746_750del, EGFR amplification, PIK3CA E545K, MAP2K2 (MEK2) C125S, MTAP rearrangement intron 5, TP53 V197M, and TP53 S241A. Owing to the considerable clinical benefit, good tolerance and lack of systemic treatment options, we decided to keep the patient on treatment and increase the doses of dabrafenib (150 mg BID) and trametinib (2 mg OD) every other day. He remains asymptomatic and tolerating well the proposed dose adjustment.
Discussion
Several studies have recently highlighted the importance of considering genes of interest within the context of commonly co-occurring mutations9,30. For example, as described by Blakely et al. through performing a cfDNA NGS analysis of 1,122 advanced stage EGFR-mutant NSCLCs, in ~93% of the patients, at least one more variables with known or likely known functional properties were present, disclosing the molecular complexity of this oncogenic driver and suggesting an association of co-occuring genomic alterations with TKI response and clinical outcomes9. In addition, Roper and colleagues reported the identification of at least two co-existing AR mechanisms in 73% of patients treated with osimertinib, as well as 6–23 different subclones per individual in a phylogenetic analysis performed in multiple metastatic sites of 15 individuals30. In the same publication, the authors also called the attention for a high incidence of acquired EGFR amplifications in post-osimertinib patients, which suggests maintenance of EGFR central role in the setting of progression, as previously reported by our group28 and also detected in the present case.
Although uncommon, BRAF V600E mediates AR in approximately 3% of the patients under second-line osimertinib6,20 and little is known about the efficacy of combined TT in this population3,21–23. Existing reports lack details regarding objective responses using standardized radiological criteria (i.e., RECIST 1.1; PERCIST) and the dynamics of resistance mechanisms through longitudinal ctDNA measurements22,23. Huang and colleagues22 described a case of an EGFRdel19/T790M + NSCLC patient who developed BRAF V600E-driven AR after second-line osimertinib and achieved stable disease under dabrafenib 150 mg BID, trametinib 1 mg OD and osimertinib 80 mg OD, with ongoing disease control 7.4 months after. Zhou and colleagues23 also reported their experience with dabrafenib 150 mg BID, trametinib 2 mg OD and osimertinib 80 mg OD leading to tumor reduction within 6 weeks of treatment, along with grade 2 rash and decreased appetite as AEs. Similarly, Meng et al. reported two cases treated with this triple regimen31. The first patient discontinued therapy after one month due to severe pneumonitis; the second one presented tumor response under dabrafenib 50 mg BID, trametinib 0.5 mg OD, and osimertinib 80 mg OD, with progression-free survival of 14 months. According to the authors, this reduced dose was prescribed owing to a grade 2 pyrexia, nausea, and vomiting under higher dabrafenib and trametinib doses. Dagogo-Jack and colleagues19 also described a successful case of combined EGFR/MAP kinase pathway blockade with osimertinib 80 mg OD and trametinib 1 mg OD; as treatment-related AEs, their patient experienced grade 2 diarrhea and fatigue, along with grade 1 rash and gastrointestinal bleeding. Nevertheless, it is worth highlighting the rapid clinical improvement, the remarkable radiologically confirmed objective response, as well as the good tolerance observed in this case even using only half standard dose of dabrafenib and trametinib approved for NSCLCs harboring BRAF V600E mutations. Since data regarding the efficacy of these combined approaches, the optimal drug association and dosing, as well as the toxicity profile are conflicting and largely unknown, further investigation into the mechanistic basis of this association represents an important priority. The Table 1 summarizes the toxicities arising under osimertinib plus BRAF/MEK inhibition and reported in the literature.Table 1 Summary of the toxicities arising under osimertinib plus BRAF/MEK inhibition and reported in the literature.
Reference Treatment Anorexia Nausea Vomiting Diarrhoea Fatigue Rash GI bleeding AST/ALT elevation Paroniquia Pyrexia Initial dose Dose reduction
Meng et al. D + T + O NR G2 G2 G2 NR NR NR NR NR G2 D (150 mg BID) + T (2 mg OD) + O (80 mg OD) D (50 mg BID) + T (0.5 mg/day) + O (80 mg/day)
Dagogo-Jack et al. T + O NR NR NR G2 G2 G1 G1 G1 NR NR T (1 mg OD) + O (80 mg OD) Not needed
Huang et al. D + T + O NR NR NR G1 NR NR NR NR G1 NR D (150 mg BID) + T (1 mg OD) + O (80 mg OD) Not needed
Zhou et al. D + T + O G1 NR NR NR NR G2 NR NR NR NR D (150 mg BID) + T (2 mg OD) + O (80 mg OD) Not needed
D dabrafenib, T trametinib, O osimertinib, NR not reported.
Liquid biopsy is emerging as an important diagnostic and predictive tool in the treatment of NSCLCs. The accurate identification of predictive genetic alterations is important for both patients’ management and the understanding of clonal evolution and AR to different therapies9,29,32. Also, circulating biomarkers from multiple disease sites better reflect systemic tumor burden, including alterations from genetically different metastatic lesions, which may be missed with single-site tissue biopsies8. Here, we were able to identify and monitor multiple concurrent mechanisms of resistance throughout the entire patient’s treatment. As shown in Fig. 1b, variations in blood ctDNA levels exhibited a positive correlation with imaging findings, even in situations involving focal treatments for oligo-PD. The observed concordance between radiological PD or response and ctDNA measurements also points towards its great potential to be incorporated into clinical practice to anticipate radiologic findings in a more effective manner.
Along with BRAF V600E mutation, the presence of PIK3CA E545K mutation is also associated with AR to osimertinib6,9,25,27. However, the clinical implication of concomitant targeting of EGFR and PI3K remains unclear. Whereas an alternative argument for the observed response could be a rechallenge following a long time upon off-osimertinib, in our opinion, this is an unlikely explanation33. Interestingly, the presence of a BRAF fusion along with a MEK2 mutation identified through plasma NGS in the setting of PD might represent a mechanism of resistance to this triple regimen34. Nonetheless, since pretreatment blood samples had not been analyzed using a similar methodology, this hypothesis deserves careful interpretation.
Increased understanding of the relationship of concurring genomic alterations in EGFR-mutant NSCLC may enable new therapeutic opportunities following disease progression to osimertinib. Here, we reported an impressive objective response to dabrafenib, trametinib, and osimertinib with concordant decrease in plasma ctDNA levels in a metastatic lung cancer patient harboring EGFR E746_A750del, BRAF V600E, and PIK3CA E545K activating mutations. This case report leads to a greater understanding of the currently limited literature regarding the management of EGFR-mutant NSCLC patients with acquired BRAFV600E mutation, since it reports a successful attempt to target both alterations concurrently while providing concordant and interesting data of serial ctDNA assessments throughout the entire treatment. Further investigation to optimize the efficacy and mitigate the toxicity profile of this drug association represents an important issue.
Methods
Patient
This study was approved by Hospital Sírio-Libanês Ethics Committee (HSL-RC 2020-16). The patient provided written informed consent for blood collection, ctDNA analysis and publication of this report.
Sample collection and plasma DNA extraction
Serial blood samples were collected between May 2017 and April 2020 (Fig. 1b). Peripheral blood (20 ml) was collected into tubes containing EDTA (BD, Franklin Lakes, New Jersey). Plasma was separated from the blood within 2 hours of blood collection, as previously described28. cfDNA was extracted using QIAamp MinElute Virus Vacuum Kit (Qiagen, Hilden, Germany) and stored at −80 °C.
ctDNA-ddPCR
Cell-free DNA (cfDNA) was quantified using the RNase P Copy Number Reference Assay (Life Technologies, Carlsbad, California). A total of 3000 genome-equivalents (~10 ng of cfDNA) were analyzed per assay to achieve a detection sensitivity of 0.2%. This detection limit has been assessed by using cell line-derived genomic DNA. A total of 10 ng of input DNA with varying proportions of mutant DNA was serially diluted into wild-type DNA to obtain samples with a mutant abundance of 1%, 0.5%, 0.1%, and 0.05% and subjected to droplet digital PCR (ddPCR). ddPCR was used to quantify the circulating levels of the EGFR activating mutation (EGFR E746_A750 del) and of the resistance mutations (EGFR T790M, PIK3CA E545K, and BRAF V600E). Probes and primers were obtained from BioRad (EGFR E746_A750 del #10041170, EGFR T790M #10040782, PIK3CA E545K #10041188, and BRAF V600E #10040779; Hercules, California). ddPCR was performed on the QX200 Droplet Digital PCR System, and data were analysed using QuantaSoft software (Bio-Rad). ctDNA quantification is presented as fractional abundance (FA–the proportion of the mutant allele in total cfDNA).
Reporting summary
Further information on research design is available in the Nature Research Reporting Summary linked to this article.
Supplementary information
Reporting Summary
Dataset 1
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
These authors contributed equally: Maurício Fernando Silva Almeida Ribeiro, Franciele Hinterholz Knebel.
Supplementary information
The online version contains supplementary material available at 10.1038/s41698-021-00149-4.
Acknowledgements
We thank the clinical research team at Hospital Sírio-Libanês for their assistance collecting samples. We thank Ernande Xavier dos Santos and Elisângela Monteiro for the outstanding technical support. We also thank Mrs. Dina Binzagr and Vivian Hannud for supporting the Translational Research Program at Hospital Sírio Libanês. This work was supported by Ludwig Cancer Research and Fundação de Amparo à Pesquisa do Estado de São Paulo – FAPESP. Lastly, we would like to thank the patient and his family for consenting to the present publication.
Author contributions
Conceptualization by M.F.S.A.R., F.H.K., J.V.M.A., A.A.C., and A.K.; Methodology by F.H.K. and A.A.C.; Resources br M.F.S.A.R., F.H.K., A.A.C., and J.V.M.A.; Writing of the first draft by M.F.S.A., F.H.K., and J.V.M.A.; Review, editing and approval of the final draft by all authors; Supervision by A.A.C. and A.K.; Funding acquisition by F.H.K. and A.A.C. M.F.S.A.R. and F.H.K. are co-first authors of the present work.
Data availability
The datasets that support the findings of this study are not publicly available in order to protect patient privacy. The data will be made available on reasonable request. For data access requests regarding the liquid biopsy (ctDNA quantification) data, please contact Dr. Franciele Knebel, email address: fhknebel@mochsl.org.br. For data access requests regarding the PET-CT high resolution images and PERCIST calculations, please contact Dr. José Marin, email address:jfgmarin@yahoo.com.br. For data access requests regarding the summary of the toxicities arising under osimertinib plus BRAF/MEK inhibition, please contact the corresponding author Dr. Maurício Ribeiro, email address: mauricio.fsaribeiro@hsl.org.br. The data generated and analysed during this study are described in the following metadata record: 10.6084/m9.figshare.1347594635.
Competing interests
The authors declare no competing interests. | Recovered | ReactionOutcome | CC BY | 33580193 | 19,431,708 | 2021-02-12 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Disease progression'. | Case Report: A 64-Year-Old Man with 10-Year History of Eosinophilic Granulomatosis with Polyangiitis with Bronchiectasis and Severe Klebsiella pneumonia.
BACKGROUND Eosinophilic granulomatosis with polyangiitis (EGPA) is a rare and severe progressive disease with multiple clinical manifestations and organ damage. Usually, it requires long-term monitoring of the state of many organs due to the gradual character of its manifestations. CASE REPORT We report a case of a long-term follow-up of a patient with eosinophilic granulomatosis with polyangiitis with emphasis on specific clinical features in this patient. A 64-year-old man was being followed up for 10 years. The initial diagnosis was allergic bronchial asthma; however, as new clinical signs and symptoms developed, the diagnosis of EGPA became obvious. A positive treatment response was seen, mainly manifested as reduced polyneuropathy. Meanwhile, bronchial asthma remained uncontrolled and bronchiectasis and Klebsiella pneumoniae colonization developed despite the combination treatment with prednisolone and methotrexate. Furthermore, the patient suffered a cerebral ischemic infarction. During the last hospital admission, severe uncontrolled bronchial asthma complicated with pneumonia resulted in the patient's death. CONCLUSIONS This clinical case shows the gradual development of EGPA with multiple-organ involvement, including respiratory manifestations and peripheral and central nervous system damage. Immunosuppressive treatment combined with complications of EGPA could have contributed to severe pneumonia development and death of the patient.
Background
Systemic vasculitides are difficult to diagnose due to the heterogenous clinical manifestations requiring a wide differential diagnostic search. One of the most uncommon vasculitis types is eosinophilic granulomatosis with polyangiitis (EGPA), or Churg-Strauss syndrome [1,2]. This vasculitis is associated with the presence of antineutrophilic cytoplasmic antibodies (ANCA) and is characterized by necrotizing inflammatory lesions of mainly small vessels, as well as eosinophilic and granulomatous inflammation in various organs. Bronchial asthma and eosinophilia are the most common clinical manifestations of EGPA [1–3]. In 2012, according to the current classification of systemic vasculitides, Churg-Strauss syndrome was renamed “Eosinophilic Granulomatosis with Polyangiitis” (EGPA) and classified into the ANCA-associated vasculitides group [3].
EGPA is a rare ANCA-associated vasculitis. The incidence rate is 0.5–6.8 new cases/year per million populations in Europe [2]. Prevalence is higher in northern latitudes compared to southern Europe [4]. The disease occurs most commonly in people 40–60 years of age [2,5]. No sex predominance has been clearly demonstrated [4].
The pathogenesis of the disease is still not completely clear, but the key role is attributed to autoimmune inflammation with the participation of eosinophils, CD8+, and CD4+lymphocytes (predominantly Th-2) and their proinflammatory cytokines: interleukin (IL)4, IL5, and IL13 [2,5]. Elevation of immunoglobulin (Ig)E, IgG4, and eotaxin-3 is common in EGPA patients [2,5]. The frequent (in 30–40% of cases) detection of ANCA (mainly antibodies to myeloperoxidase) also suggests the autoimmune nature of the disease. ANCA-positive patients are more commonly diagnosed with glomerulonephritis and mononeuritis, whereas the ANCA-negative ones more frequently demonstrate endocardial, myocardial, and pulmonary eosinophilic infiltrates [1,2,5,6].
There are 3 phases in the development of EGPA (these phases partially overlap and may not appear in sequence):
the allergic phase, distinguished by allergic upper respiratory tract inflammation (rhinosinusitis, allergic rhinitis) and bronchial asthma;
the eosinophilic phase, manifested with severe eosinophilia in combination with multiple eosinophilic infiltrates in various organs (most often in the lungs and gastrointestinal tract);
the vasculitis phase, with small- and medium-vessel inflammation of different organs, which can often be life-threatening [5,6].
Respiratory manifestations of EGPA include allergic rhinitis, sinusitis, nasal polyposis and, in more than 95% of patients, poorly controlled bronchial asthma [5]. Chest radiography findings include transient and migratory opacities, ground-glass opacity, focal consolidation, and small peribronchial and centrilobular nodules or masses [7]. Computed tomography reveals bronchial abnormalities, most commonly thickened wall and dilatation with secondary bronchiectasis formation, leading to frequent infections [7,8]. Pleural effusions and alveolar hemorrhages are uncommon manifestations of EGPA [1,5,7,8].
A characteristic neurological manifestation seen in 75% of patients with EGPA is peripheral neuropathy (mononeuritis multiplex) [1,5]. Upper-limb symptoms due to neuropathy, as well as vasculitis in epineural vessels, were more frequently reported in the ANCA-positive group [9]. Eosinophil-associated tissue damage and vascular occlusion leading to ischemia are more specific in ANCA-negative patients [9]. Also, the tissue damage caused by eosinophils may be associated with IL-5 concentration [10]. Brain hemorrhage, subarachnoid hemorrhage, and ischemic brain infarction are rare [1,5,6].
The main drugs for the treatment of EGPA are glucocorticoids. In organ- or life-threatening manifestations, the recommended starting dose of prednisone is 1 mg/kg/day with a maximal daily dose of 80 mg for 2–3 weeks, with tapering to a minimal supporting dose selected individually to control the symptoms and minimize the adverse effects of treatment. Methylprednisolone (7.5–15 mg/kg/day) can be used as a pulse therapy for life-threatening manifestations [11,12]. Most patients need prolonged or lifelong steroids. In the case of organ- or life-threatening manifestations, glucocorticoids in combination with cyclophosphamide are prescribed [11,12]. The combination of glucocorticoids and methotrexate or mycophenolate is required to induce remission in mild or moderate disease [11,12]. Mepolizumab (a humanized monoclonal antibody targeting IL-5) has been used for EGPA treatment [11]. For ANCA-positive patients with renal involvement or refractory disease, rituximab can be considered [3, 11].
Due to the rare occurrence of EGPA and the variety of its clinical manifestations, the diagnosis and treatment of this disease can be challenging. Currently, reports of long-term observations with the detailed patient follow-up are of great interest. The aim of our case report is to present rare manifestations of EGPA, such as bronchiectasis and stroke. We also focused on the infectious complications of immunosuppressive treatments. Thus, we would like to present a 10-year observation of a patient with EGPA.
Case Report
The male patient, born in 1953, was first hospitalized in the Pulmonary Department of our clinic in 2007 due to exacerbated asthma. The patient’s history was remarkable for multiple allergies (to house dust, cat and dog epidermis, and plant pollen) as well as nonsteroidal anti-inflammatory drugs intolerance (manifested as bronchospasm). The patient denied unhealthy habits and occupational hazards (he worked as a driver). His family history was not remarkable. The patient has had allergic rhinitis since adolescence. The first asthma symptoms appeared at the age of 18 while serving in the army. At the age of 22, an allergist diagnosed him with bronchial asthma. The patient had annual exacerbations (1–2 times a year) due to cold or allergen exposure, and acute respiratory viral infections. At the age of 42, polypoid rhinosinusitis was found, and a polypectomy was performed. Since 2006, the patient had been on controller treatment with salmeterol/fluticasone inhaler (25/250 mg, respectively) 2 doses 2 times a day.
In late 2009, extremely elevated eosinophils (53%) and total IgE (1000 IU/L) were found for the first time. Also, since 2009, the patient has had transient lung opacities, which usually disappeared after treatment with prednisolone 30 mg per day.
In August 2010, the patient noticed numbness on the back surface of the left foot, which spread to the entire left lower leg in 1 month, as well as numbness of the right foot and lower leg. Later, the patient started having tingling, twisting, and burning sensations aggravated by touching in the feet and lower legs. Also, the weakness in the distal parts of the legs developed, and he started having difficulties moving around on his own and falling while walking, and he was subsequently hospitalized in our clinic due to exacerbated asthma. On laboratory testing, high eosinophils (53%) and gamma-globulins (27.9%) were found in November 2010. The chest radiography showed emphysema and consolidation in the middle lobe of the right lung. A Neurology consult confirmed asymmetrical distal sensory-motor polyneuropathy demonstrated by lower distal asymmetrical paraparesis with decreased muscle strength in the foot flexors and extensors, more severe on the left side, low tone of the shin muscles, hypoactive Achilles reflexes, and reduced leg sensitivity starting from the middle of the shin and below, more severe in the peroneal nerve inner-vation area. The patient also demonstrated abnormal coordination (heel-to-knee, Romberg) test results and bilateral steppage gait with knee overextension. Electroneuromyography showed axonal lesion of all distal leg nerves and secondary myelinopathy. Thus, EGPA was suspected. The ANCA (anti-myeloperoxidase) titer was 30 U/ml (normal reference below 20 U/ml).
Allergic bronchopulmonary aspergillosis and other causes of hypereosinophilia were consistently ruled out. IgE and IgG for Aspergillus spp. and Aspergillus spp. in sputum were not found.
Thus, the presence of severe bronchial asthma, rhinosinusitis, high and persistent eosinophilia, transient pulmonary opacities, and polyneuropathy suggested EGPA (5 out of 6 clinical criteria of American College of Rheumatology present). Based on the clinical presentation and increased ANCA titer, the patient was diagnosed with EGPA. Systemic steroid treatment (prednisolone 300 mg intravenously and 30 mg orally) was prescribed, and 5 plasmapheresis sessions were performed. Later, prolonged combination immunosuppression (prednisolone 10 mg/day and methotrexate 25 mg/week) and controller asthma treatment (fluticasone propionate/salmeterol 250/25 µg 4 doses/ day, tiotropium bromide 18 µg per day) were prescribed. After 1 month on prednisolone and methotrexate, there was a significant neurological improvement: the motor and sensory disturbances decreased, and the patient was able to move around on his own. Further on, polyneuropathy completely resolved on the combination treatment. His asthma remained uncontrolled despite the combination treatment: he had frequent (3 times a year) severe exacerbations requiring hospitalization. In 2015, newly-emerged copious purulent sputum associated with frequent infection-related asthma exacerbations was noted. In January 2016, a patient underwent chest computed tomography (CT), which showed generalized bronchiectasis. Stenosis of the proximal bronchi was ruled out also by fibrobronchos-copy. The bronchial secretion microbiology revealed Klebsiella pneumoniae growth (107 colony-forming units).
In November 2016, during the asthma exacerbation, the patient suffered a cerebral ischemic infarction involving the right occipital lobe, confirmed by CT (Figures 1, 2).
On March 2, 2017, the patient was admitted to our hospital due to severe respiratory failure caused by pneumonia in the left upper lobe and severe asthma exacerbation.
The complete blood count showed high WBCs (15.2×109/l) with left shift. The IgE level was 144.0 IU/l, C-reactive protein was 48.8 mg/dL. Clinical chemistry was remarkable for urea nitrogen 18.4 mmol/l and creatinine 1.55 mg/dL. The sputum microscopy demonstrated 100 WBCs and 160 RBCs per vision field, and no eosinophils were revealed. A chest CT revealed multiple bronchiectasis and upper lobar pneumonia of the left lung (Figure 2). Klebsiella was again detected in the sputum culture.
The patient was prescribed intravenous antibiotics (ceftriaxone 2 g/day and ciprofloxacin 800 mg/day replaced by cefopera-zone 4 g/day and amikacin 1 g/day after 3 days due to low efficacy), bronchodilator and anti-inflammatory therapy (nebulized ipratropium bromide, fenoterol, and budesonide 2000 µg per day, prednisolone intravenously 150 mg and orally 30 mg). Despite the aggressive treatment, the respiratory failure progressed, and on day 6 of hospitalization the patient was transferred to the Intensive Care Unit and intubated. On day 7, the patient died. On autopsy, the EGPA diagnosis was confirmed, signs of ischemic infarction in the occipital lobe of the brain were found, and left upper lobe pneumonia with abscess formation and generalized bilateral bronchiectases were revealed.
Discussion
Based on the long follow-up period and careful interpretation of signs and symptoms, typical laboratory abnormalities, the diagnosis was clear. Among the most commonly used diagnostic criteria of EGPA are the American College of Rheumatology (ACR) classification criteria, Lanham’s criteria, and the Chapel Hill Consensus Conference criteria. The presence of 4 or more ACR classification criteria, including bronchial asthma, eosinophilia (more than 10%), paranasal sinus abnormalities, migratory or transient eosinophilic pulmonary infiltrates, extravascular eosinophil infiltration on biopsy, and mono- or polyneuropathy, has a sensitivity of 85% and a specificity of 100% [2,5,6]. We identified 5 out of 6 American College of Rheumatology clinical criteria present in our patient.
This clinical case shows gradual development of EGPA with multiple-organ involvement (see Figure 3). The disease onset was characterized by respiratory manifestations and suspicion of EGPA appeared only after the development of polyneuropathy (after 39 years) with hypereosinophilia. Polyneuropathy was noted at the age of 57, which is slightly later than the average age of onset reported in the literature [13]. Notably, the patient was ANCA-positive and his long-term neurological manifestation (mononeuritis) was quite typical for ANCA vasculitis [5,6,8,13]. The diagnosis became obvious after comprehensive assessment of the disease history with its gradually evolving signs and symptoms and laboratory abnormalities. Almost all manifestations of the disease seen in the patient were typical except for stroke, which is an exceptional feature for this case [1,5,6]. Neurological involvement manifested as ischemic stroke is an uncommon complication of EGPA (seen only in 5.2% of all patients) [1]. Thus, the patient demonstrated both peripheral and central nervous system involvement, which is unusual and life-threatening in EGPA.
This clinical case of EGPA also attracted our attention due to severe respiratory manifestations. While typical respiratory manifestations such as rhinosinusitis, asthma, and transient lung infiltrates are well documented in the literature, EGPA-associated bronchiectasis is rarely mentioned [14]. Bronchiectasis in the patient was detected by CT at the age of 63. The patient was a nonsmoker, and no other cause of bronchial abnormalities and airflow obstruction could be identified. Stenosis of the proximal bronchi was ruled out by fibrobronchoscopy and CT. The diagnosis of allergic bronchopulmonary aspergillosis was not confirmed. Klebsiella pneumoniae colonization was found during the last 2 years of the patient’s life. Klebsiella pneumoniae is a typical pathogen causing bronchiectasis [15]. It should be noted that aspergillosis and mycobacteriosis are common complications of bronchiectasis [15], but neither was identified in our patient.
The patient was treated for a long time with a combination of prednisolone and methotrexate. There was a significant positive effect on polyneuropathy, but uncontrolled bronchial asthma persisted. This treatment regimen complies with current recommendations [3,11]. Patients with life-threatening and/or organ-threatening manifestations (including lesions of the central nervous system and severe peripheral neuropathy) require therapy with a combination of a glucocorticoid and a cytostatic [3,11].
The use of immunosuppressive drugs in patients with ANCA-associated vasculitis significantly increases the risk of infection and mortality [16–18]. Prednisone and methotrexate use were found to be significant predictors of hospitalization for pneumonia. Frequent infections (more than 1 episode) are reported in patients with ANCA-associated vasculitis during the first year of diagnosis and initiation of therapy [18]. We assume that bronchiectasis complicated by bacterial colonization and bronchial obstruction in combination with immuno-suppression led to severe pneumonia and death of the patient.
Conclusions
The present case report demonstrates a long-term follow-up of a patient with EGPA. Both common and rare manifestations of EGPA have been described in the patient, such as central nervous system damage and bronchiectasis. Immunosuppressive treatment probably contributed to Klebsiella infection development. Thus, long-term complications of EGPA in combination with adverse effects of medications could have caused the severe pneumonia and death of the patient.
Figure 1. Brain multi-spiral computed tomography performed in 2016 showing ischemic brain damage in the right occipital lobe.
Figure 2. (A–C) Chest computed tomography of patient showing large left upper lobe pneumonia and bronchiectasis in lover lobes.
Figure 3. Timeline from first symptoms to death. | AMIKACIN SULFATE, BUDESONIDE, CEFOPERAZONE, CEFTRIAXONE, CIPROFLOXACIN, FENOTEROL, FLUTICASONE PROPIONATE\SALMETEROL XINAFOATE, IPRATROPIUM BROMIDE, METHOTREXATE, PREDNISOLONE, TIOTROPIUM BROMIDE | DrugsGivenReaction | CC BY-NC-ND | 33582702 | 18,962,812 | 2021-02-14 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Klebsiella infection'. | Case Report: A 64-Year-Old Man with 10-Year History of Eosinophilic Granulomatosis with Polyangiitis with Bronchiectasis and Severe Klebsiella pneumonia.
BACKGROUND Eosinophilic granulomatosis with polyangiitis (EGPA) is a rare and severe progressive disease with multiple clinical manifestations and organ damage. Usually, it requires long-term monitoring of the state of many organs due to the gradual character of its manifestations. CASE REPORT We report a case of a long-term follow-up of a patient with eosinophilic granulomatosis with polyangiitis with emphasis on specific clinical features in this patient. A 64-year-old man was being followed up for 10 years. The initial diagnosis was allergic bronchial asthma; however, as new clinical signs and symptoms developed, the diagnosis of EGPA became obvious. A positive treatment response was seen, mainly manifested as reduced polyneuropathy. Meanwhile, bronchial asthma remained uncontrolled and bronchiectasis and Klebsiella pneumoniae colonization developed despite the combination treatment with prednisolone and methotrexate. Furthermore, the patient suffered a cerebral ischemic infarction. During the last hospital admission, severe uncontrolled bronchial asthma complicated with pneumonia resulted in the patient's death. CONCLUSIONS This clinical case shows the gradual development of EGPA with multiple-organ involvement, including respiratory manifestations and peripheral and central nervous system damage. Immunosuppressive treatment combined with complications of EGPA could have contributed to severe pneumonia development and death of the patient.
Background
Systemic vasculitides are difficult to diagnose due to the heterogenous clinical manifestations requiring a wide differential diagnostic search. One of the most uncommon vasculitis types is eosinophilic granulomatosis with polyangiitis (EGPA), or Churg-Strauss syndrome [1,2]. This vasculitis is associated with the presence of antineutrophilic cytoplasmic antibodies (ANCA) and is characterized by necrotizing inflammatory lesions of mainly small vessels, as well as eosinophilic and granulomatous inflammation in various organs. Bronchial asthma and eosinophilia are the most common clinical manifestations of EGPA [1–3]. In 2012, according to the current classification of systemic vasculitides, Churg-Strauss syndrome was renamed “Eosinophilic Granulomatosis with Polyangiitis” (EGPA) and classified into the ANCA-associated vasculitides group [3].
EGPA is a rare ANCA-associated vasculitis. The incidence rate is 0.5–6.8 new cases/year per million populations in Europe [2]. Prevalence is higher in northern latitudes compared to southern Europe [4]. The disease occurs most commonly in people 40–60 years of age [2,5]. No sex predominance has been clearly demonstrated [4].
The pathogenesis of the disease is still not completely clear, but the key role is attributed to autoimmune inflammation with the participation of eosinophils, CD8+, and CD4+lymphocytes (predominantly Th-2) and their proinflammatory cytokines: interleukin (IL)4, IL5, and IL13 [2,5]. Elevation of immunoglobulin (Ig)E, IgG4, and eotaxin-3 is common in EGPA patients [2,5]. The frequent (in 30–40% of cases) detection of ANCA (mainly antibodies to myeloperoxidase) also suggests the autoimmune nature of the disease. ANCA-positive patients are more commonly diagnosed with glomerulonephritis and mononeuritis, whereas the ANCA-negative ones more frequently demonstrate endocardial, myocardial, and pulmonary eosinophilic infiltrates [1,2,5,6].
There are 3 phases in the development of EGPA (these phases partially overlap and may not appear in sequence):
the allergic phase, distinguished by allergic upper respiratory tract inflammation (rhinosinusitis, allergic rhinitis) and bronchial asthma;
the eosinophilic phase, manifested with severe eosinophilia in combination with multiple eosinophilic infiltrates in various organs (most often in the lungs and gastrointestinal tract);
the vasculitis phase, with small- and medium-vessel inflammation of different organs, which can often be life-threatening [5,6].
Respiratory manifestations of EGPA include allergic rhinitis, sinusitis, nasal polyposis and, in more than 95% of patients, poorly controlled bronchial asthma [5]. Chest radiography findings include transient and migratory opacities, ground-glass opacity, focal consolidation, and small peribronchial and centrilobular nodules or masses [7]. Computed tomography reveals bronchial abnormalities, most commonly thickened wall and dilatation with secondary bronchiectasis formation, leading to frequent infections [7,8]. Pleural effusions and alveolar hemorrhages are uncommon manifestations of EGPA [1,5,7,8].
A characteristic neurological manifestation seen in 75% of patients with EGPA is peripheral neuropathy (mononeuritis multiplex) [1,5]. Upper-limb symptoms due to neuropathy, as well as vasculitis in epineural vessels, were more frequently reported in the ANCA-positive group [9]. Eosinophil-associated tissue damage and vascular occlusion leading to ischemia are more specific in ANCA-negative patients [9]. Also, the tissue damage caused by eosinophils may be associated with IL-5 concentration [10]. Brain hemorrhage, subarachnoid hemorrhage, and ischemic brain infarction are rare [1,5,6].
The main drugs for the treatment of EGPA are glucocorticoids. In organ- or life-threatening manifestations, the recommended starting dose of prednisone is 1 mg/kg/day with a maximal daily dose of 80 mg for 2–3 weeks, with tapering to a minimal supporting dose selected individually to control the symptoms and minimize the adverse effects of treatment. Methylprednisolone (7.5–15 mg/kg/day) can be used as a pulse therapy for life-threatening manifestations [11,12]. Most patients need prolonged or lifelong steroids. In the case of organ- or life-threatening manifestations, glucocorticoids in combination with cyclophosphamide are prescribed [11,12]. The combination of glucocorticoids and methotrexate or mycophenolate is required to induce remission in mild or moderate disease [11,12]. Mepolizumab (a humanized monoclonal antibody targeting IL-5) has been used for EGPA treatment [11]. For ANCA-positive patients with renal involvement or refractory disease, rituximab can be considered [3, 11].
Due to the rare occurrence of EGPA and the variety of its clinical manifestations, the diagnosis and treatment of this disease can be challenging. Currently, reports of long-term observations with the detailed patient follow-up are of great interest. The aim of our case report is to present rare manifestations of EGPA, such as bronchiectasis and stroke. We also focused on the infectious complications of immunosuppressive treatments. Thus, we would like to present a 10-year observation of a patient with EGPA.
Case Report
The male patient, born in 1953, was first hospitalized in the Pulmonary Department of our clinic in 2007 due to exacerbated asthma. The patient’s history was remarkable for multiple allergies (to house dust, cat and dog epidermis, and plant pollen) as well as nonsteroidal anti-inflammatory drugs intolerance (manifested as bronchospasm). The patient denied unhealthy habits and occupational hazards (he worked as a driver). His family history was not remarkable. The patient has had allergic rhinitis since adolescence. The first asthma symptoms appeared at the age of 18 while serving in the army. At the age of 22, an allergist diagnosed him with bronchial asthma. The patient had annual exacerbations (1–2 times a year) due to cold or allergen exposure, and acute respiratory viral infections. At the age of 42, polypoid rhinosinusitis was found, and a polypectomy was performed. Since 2006, the patient had been on controller treatment with salmeterol/fluticasone inhaler (25/250 mg, respectively) 2 doses 2 times a day.
In late 2009, extremely elevated eosinophils (53%) and total IgE (1000 IU/L) were found for the first time. Also, since 2009, the patient has had transient lung opacities, which usually disappeared after treatment with prednisolone 30 mg per day.
In August 2010, the patient noticed numbness on the back surface of the left foot, which spread to the entire left lower leg in 1 month, as well as numbness of the right foot and lower leg. Later, the patient started having tingling, twisting, and burning sensations aggravated by touching in the feet and lower legs. Also, the weakness in the distal parts of the legs developed, and he started having difficulties moving around on his own and falling while walking, and he was subsequently hospitalized in our clinic due to exacerbated asthma. On laboratory testing, high eosinophils (53%) and gamma-globulins (27.9%) were found in November 2010. The chest radiography showed emphysema and consolidation in the middle lobe of the right lung. A Neurology consult confirmed asymmetrical distal sensory-motor polyneuropathy demonstrated by lower distal asymmetrical paraparesis with decreased muscle strength in the foot flexors and extensors, more severe on the left side, low tone of the shin muscles, hypoactive Achilles reflexes, and reduced leg sensitivity starting from the middle of the shin and below, more severe in the peroneal nerve inner-vation area. The patient also demonstrated abnormal coordination (heel-to-knee, Romberg) test results and bilateral steppage gait with knee overextension. Electroneuromyography showed axonal lesion of all distal leg nerves and secondary myelinopathy. Thus, EGPA was suspected. The ANCA (anti-myeloperoxidase) titer was 30 U/ml (normal reference below 20 U/ml).
Allergic bronchopulmonary aspergillosis and other causes of hypereosinophilia were consistently ruled out. IgE and IgG for Aspergillus spp. and Aspergillus spp. in sputum were not found.
Thus, the presence of severe bronchial asthma, rhinosinusitis, high and persistent eosinophilia, transient pulmonary opacities, and polyneuropathy suggested EGPA (5 out of 6 clinical criteria of American College of Rheumatology present). Based on the clinical presentation and increased ANCA titer, the patient was diagnosed with EGPA. Systemic steroid treatment (prednisolone 300 mg intravenously and 30 mg orally) was prescribed, and 5 plasmapheresis sessions were performed. Later, prolonged combination immunosuppression (prednisolone 10 mg/day and methotrexate 25 mg/week) and controller asthma treatment (fluticasone propionate/salmeterol 250/25 µg 4 doses/ day, tiotropium bromide 18 µg per day) were prescribed. After 1 month on prednisolone and methotrexate, there was a significant neurological improvement: the motor and sensory disturbances decreased, and the patient was able to move around on his own. Further on, polyneuropathy completely resolved on the combination treatment. His asthma remained uncontrolled despite the combination treatment: he had frequent (3 times a year) severe exacerbations requiring hospitalization. In 2015, newly-emerged copious purulent sputum associated with frequent infection-related asthma exacerbations was noted. In January 2016, a patient underwent chest computed tomography (CT), which showed generalized bronchiectasis. Stenosis of the proximal bronchi was ruled out also by fibrobronchos-copy. The bronchial secretion microbiology revealed Klebsiella pneumoniae growth (107 colony-forming units).
In November 2016, during the asthma exacerbation, the patient suffered a cerebral ischemic infarction involving the right occipital lobe, confirmed by CT (Figures 1, 2).
On March 2, 2017, the patient was admitted to our hospital due to severe respiratory failure caused by pneumonia in the left upper lobe and severe asthma exacerbation.
The complete blood count showed high WBCs (15.2×109/l) with left shift. The IgE level was 144.0 IU/l, C-reactive protein was 48.8 mg/dL. Clinical chemistry was remarkable for urea nitrogen 18.4 mmol/l and creatinine 1.55 mg/dL. The sputum microscopy demonstrated 100 WBCs and 160 RBCs per vision field, and no eosinophils were revealed. A chest CT revealed multiple bronchiectasis and upper lobar pneumonia of the left lung (Figure 2). Klebsiella was again detected in the sputum culture.
The patient was prescribed intravenous antibiotics (ceftriaxone 2 g/day and ciprofloxacin 800 mg/day replaced by cefopera-zone 4 g/day and amikacin 1 g/day after 3 days due to low efficacy), bronchodilator and anti-inflammatory therapy (nebulized ipratropium bromide, fenoterol, and budesonide 2000 µg per day, prednisolone intravenously 150 mg and orally 30 mg). Despite the aggressive treatment, the respiratory failure progressed, and on day 6 of hospitalization the patient was transferred to the Intensive Care Unit and intubated. On day 7, the patient died. On autopsy, the EGPA diagnosis was confirmed, signs of ischemic infarction in the occipital lobe of the brain were found, and left upper lobe pneumonia with abscess formation and generalized bilateral bronchiectases were revealed.
Discussion
Based on the long follow-up period and careful interpretation of signs and symptoms, typical laboratory abnormalities, the diagnosis was clear. Among the most commonly used diagnostic criteria of EGPA are the American College of Rheumatology (ACR) classification criteria, Lanham’s criteria, and the Chapel Hill Consensus Conference criteria. The presence of 4 or more ACR classification criteria, including bronchial asthma, eosinophilia (more than 10%), paranasal sinus abnormalities, migratory or transient eosinophilic pulmonary infiltrates, extravascular eosinophil infiltration on biopsy, and mono- or polyneuropathy, has a sensitivity of 85% and a specificity of 100% [2,5,6]. We identified 5 out of 6 American College of Rheumatology clinical criteria present in our patient.
This clinical case shows gradual development of EGPA with multiple-organ involvement (see Figure 3). The disease onset was characterized by respiratory manifestations and suspicion of EGPA appeared only after the development of polyneuropathy (after 39 years) with hypereosinophilia. Polyneuropathy was noted at the age of 57, which is slightly later than the average age of onset reported in the literature [13]. Notably, the patient was ANCA-positive and his long-term neurological manifestation (mononeuritis) was quite typical for ANCA vasculitis [5,6,8,13]. The diagnosis became obvious after comprehensive assessment of the disease history with its gradually evolving signs and symptoms and laboratory abnormalities. Almost all manifestations of the disease seen in the patient were typical except for stroke, which is an exceptional feature for this case [1,5,6]. Neurological involvement manifested as ischemic stroke is an uncommon complication of EGPA (seen only in 5.2% of all patients) [1]. Thus, the patient demonstrated both peripheral and central nervous system involvement, which is unusual and life-threatening in EGPA.
This clinical case of EGPA also attracted our attention due to severe respiratory manifestations. While typical respiratory manifestations such as rhinosinusitis, asthma, and transient lung infiltrates are well documented in the literature, EGPA-associated bronchiectasis is rarely mentioned [14]. Bronchiectasis in the patient was detected by CT at the age of 63. The patient was a nonsmoker, and no other cause of bronchial abnormalities and airflow obstruction could be identified. Stenosis of the proximal bronchi was ruled out by fibrobronchoscopy and CT. The diagnosis of allergic bronchopulmonary aspergillosis was not confirmed. Klebsiella pneumoniae colonization was found during the last 2 years of the patient’s life. Klebsiella pneumoniae is a typical pathogen causing bronchiectasis [15]. It should be noted that aspergillosis and mycobacteriosis are common complications of bronchiectasis [15], but neither was identified in our patient.
The patient was treated for a long time with a combination of prednisolone and methotrexate. There was a significant positive effect on polyneuropathy, but uncontrolled bronchial asthma persisted. This treatment regimen complies with current recommendations [3,11]. Patients with life-threatening and/or organ-threatening manifestations (including lesions of the central nervous system and severe peripheral neuropathy) require therapy with a combination of a glucocorticoid and a cytostatic [3,11].
The use of immunosuppressive drugs in patients with ANCA-associated vasculitis significantly increases the risk of infection and mortality [16–18]. Prednisone and methotrexate use were found to be significant predictors of hospitalization for pneumonia. Frequent infections (more than 1 episode) are reported in patients with ANCA-associated vasculitis during the first year of diagnosis and initiation of therapy [18]. We assume that bronchiectasis complicated by bacterial colonization and bronchial obstruction in combination with immuno-suppression led to severe pneumonia and death of the patient.
Conclusions
The present case report demonstrates a long-term follow-up of a patient with EGPA. Both common and rare manifestations of EGPA have been described in the patient, such as central nervous system damage and bronchiectasis. Immunosuppressive treatment probably contributed to Klebsiella infection development. Thus, long-term complications of EGPA in combination with adverse effects of medications could have caused the severe pneumonia and death of the patient.
Figure 1. Brain multi-spiral computed tomography performed in 2016 showing ischemic brain damage in the right occipital lobe.
Figure 2. (A–C) Chest computed tomography of patient showing large left upper lobe pneumonia and bronchiectasis in lover lobes.
Figure 3. Timeline from first symptoms to death. | BUDESONIDE, FENOTEROL, FLUTICASONE PROPIONATE, IPRATROPIUM BROMIDE, METHOTREXATE, PREDNISOLONE, SALMETEROL, TIOTROPIUM BROMIDE | DrugsGivenReaction | CC BY-NC-ND | 33582702 | 18,975,017 | 2021-02-14 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Product use in unapproved indication'. | Case Report: A 64-Year-Old Man with 10-Year History of Eosinophilic Granulomatosis with Polyangiitis with Bronchiectasis and Severe Klebsiella pneumonia.
BACKGROUND Eosinophilic granulomatosis with polyangiitis (EGPA) is a rare and severe progressive disease with multiple clinical manifestations and organ damage. Usually, it requires long-term monitoring of the state of many organs due to the gradual character of its manifestations. CASE REPORT We report a case of a long-term follow-up of a patient with eosinophilic granulomatosis with polyangiitis with emphasis on specific clinical features in this patient. A 64-year-old man was being followed up for 10 years. The initial diagnosis was allergic bronchial asthma; however, as new clinical signs and symptoms developed, the diagnosis of EGPA became obvious. A positive treatment response was seen, mainly manifested as reduced polyneuropathy. Meanwhile, bronchial asthma remained uncontrolled and bronchiectasis and Klebsiella pneumoniae colonization developed despite the combination treatment with prednisolone and methotrexate. Furthermore, the patient suffered a cerebral ischemic infarction. During the last hospital admission, severe uncontrolled bronchial asthma complicated with pneumonia resulted in the patient's death. CONCLUSIONS This clinical case shows the gradual development of EGPA with multiple-organ involvement, including respiratory manifestations and peripheral and central nervous system damage. Immunosuppressive treatment combined with complications of EGPA could have contributed to severe pneumonia development and death of the patient.
Background
Systemic vasculitides are difficult to diagnose due to the heterogenous clinical manifestations requiring a wide differential diagnostic search. One of the most uncommon vasculitis types is eosinophilic granulomatosis with polyangiitis (EGPA), or Churg-Strauss syndrome [1,2]. This vasculitis is associated with the presence of antineutrophilic cytoplasmic antibodies (ANCA) and is characterized by necrotizing inflammatory lesions of mainly small vessels, as well as eosinophilic and granulomatous inflammation in various organs. Bronchial asthma and eosinophilia are the most common clinical manifestations of EGPA [1–3]. In 2012, according to the current classification of systemic vasculitides, Churg-Strauss syndrome was renamed “Eosinophilic Granulomatosis with Polyangiitis” (EGPA) and classified into the ANCA-associated vasculitides group [3].
EGPA is a rare ANCA-associated vasculitis. The incidence rate is 0.5–6.8 new cases/year per million populations in Europe [2]. Prevalence is higher in northern latitudes compared to southern Europe [4]. The disease occurs most commonly in people 40–60 years of age [2,5]. No sex predominance has been clearly demonstrated [4].
The pathogenesis of the disease is still not completely clear, but the key role is attributed to autoimmune inflammation with the participation of eosinophils, CD8+, and CD4+lymphocytes (predominantly Th-2) and their proinflammatory cytokines: interleukin (IL)4, IL5, and IL13 [2,5]. Elevation of immunoglobulin (Ig)E, IgG4, and eotaxin-3 is common in EGPA patients [2,5]. The frequent (in 30–40% of cases) detection of ANCA (mainly antibodies to myeloperoxidase) also suggests the autoimmune nature of the disease. ANCA-positive patients are more commonly diagnosed with glomerulonephritis and mononeuritis, whereas the ANCA-negative ones more frequently demonstrate endocardial, myocardial, and pulmonary eosinophilic infiltrates [1,2,5,6].
There are 3 phases in the development of EGPA (these phases partially overlap and may not appear in sequence):
the allergic phase, distinguished by allergic upper respiratory tract inflammation (rhinosinusitis, allergic rhinitis) and bronchial asthma;
the eosinophilic phase, manifested with severe eosinophilia in combination with multiple eosinophilic infiltrates in various organs (most often in the lungs and gastrointestinal tract);
the vasculitis phase, with small- and medium-vessel inflammation of different organs, which can often be life-threatening [5,6].
Respiratory manifestations of EGPA include allergic rhinitis, sinusitis, nasal polyposis and, in more than 95% of patients, poorly controlled bronchial asthma [5]. Chest radiography findings include transient and migratory opacities, ground-glass opacity, focal consolidation, and small peribronchial and centrilobular nodules or masses [7]. Computed tomography reveals bronchial abnormalities, most commonly thickened wall and dilatation with secondary bronchiectasis formation, leading to frequent infections [7,8]. Pleural effusions and alveolar hemorrhages are uncommon manifestations of EGPA [1,5,7,8].
A characteristic neurological manifestation seen in 75% of patients with EGPA is peripheral neuropathy (mononeuritis multiplex) [1,5]. Upper-limb symptoms due to neuropathy, as well as vasculitis in epineural vessels, were more frequently reported in the ANCA-positive group [9]. Eosinophil-associated tissue damage and vascular occlusion leading to ischemia are more specific in ANCA-negative patients [9]. Also, the tissue damage caused by eosinophils may be associated with IL-5 concentration [10]. Brain hemorrhage, subarachnoid hemorrhage, and ischemic brain infarction are rare [1,5,6].
The main drugs for the treatment of EGPA are glucocorticoids. In organ- or life-threatening manifestations, the recommended starting dose of prednisone is 1 mg/kg/day with a maximal daily dose of 80 mg for 2–3 weeks, with tapering to a minimal supporting dose selected individually to control the symptoms and minimize the adverse effects of treatment. Methylprednisolone (7.5–15 mg/kg/day) can be used as a pulse therapy for life-threatening manifestations [11,12]. Most patients need prolonged or lifelong steroids. In the case of organ- or life-threatening manifestations, glucocorticoids in combination with cyclophosphamide are prescribed [11,12]. The combination of glucocorticoids and methotrexate or mycophenolate is required to induce remission in mild or moderate disease [11,12]. Mepolizumab (a humanized monoclonal antibody targeting IL-5) has been used for EGPA treatment [11]. For ANCA-positive patients with renal involvement or refractory disease, rituximab can be considered [3, 11].
Due to the rare occurrence of EGPA and the variety of its clinical manifestations, the diagnosis and treatment of this disease can be challenging. Currently, reports of long-term observations with the detailed patient follow-up are of great interest. The aim of our case report is to present rare manifestations of EGPA, such as bronchiectasis and stroke. We also focused on the infectious complications of immunosuppressive treatments. Thus, we would like to present a 10-year observation of a patient with EGPA.
Case Report
The male patient, born in 1953, was first hospitalized in the Pulmonary Department of our clinic in 2007 due to exacerbated asthma. The patient’s history was remarkable for multiple allergies (to house dust, cat and dog epidermis, and plant pollen) as well as nonsteroidal anti-inflammatory drugs intolerance (manifested as bronchospasm). The patient denied unhealthy habits and occupational hazards (he worked as a driver). His family history was not remarkable. The patient has had allergic rhinitis since adolescence. The first asthma symptoms appeared at the age of 18 while serving in the army. At the age of 22, an allergist diagnosed him with bronchial asthma. The patient had annual exacerbations (1–2 times a year) due to cold or allergen exposure, and acute respiratory viral infections. At the age of 42, polypoid rhinosinusitis was found, and a polypectomy was performed. Since 2006, the patient had been on controller treatment with salmeterol/fluticasone inhaler (25/250 mg, respectively) 2 doses 2 times a day.
In late 2009, extremely elevated eosinophils (53%) and total IgE (1000 IU/L) were found for the first time. Also, since 2009, the patient has had transient lung opacities, which usually disappeared after treatment with prednisolone 30 mg per day.
In August 2010, the patient noticed numbness on the back surface of the left foot, which spread to the entire left lower leg in 1 month, as well as numbness of the right foot and lower leg. Later, the patient started having tingling, twisting, and burning sensations aggravated by touching in the feet and lower legs. Also, the weakness in the distal parts of the legs developed, and he started having difficulties moving around on his own and falling while walking, and he was subsequently hospitalized in our clinic due to exacerbated asthma. On laboratory testing, high eosinophils (53%) and gamma-globulins (27.9%) were found in November 2010. The chest radiography showed emphysema and consolidation in the middle lobe of the right lung. A Neurology consult confirmed asymmetrical distal sensory-motor polyneuropathy demonstrated by lower distal asymmetrical paraparesis with decreased muscle strength in the foot flexors and extensors, more severe on the left side, low tone of the shin muscles, hypoactive Achilles reflexes, and reduced leg sensitivity starting from the middle of the shin and below, more severe in the peroneal nerve inner-vation area. The patient also demonstrated abnormal coordination (heel-to-knee, Romberg) test results and bilateral steppage gait with knee overextension. Electroneuromyography showed axonal lesion of all distal leg nerves and secondary myelinopathy. Thus, EGPA was suspected. The ANCA (anti-myeloperoxidase) titer was 30 U/ml (normal reference below 20 U/ml).
Allergic bronchopulmonary aspergillosis and other causes of hypereosinophilia were consistently ruled out. IgE and IgG for Aspergillus spp. and Aspergillus spp. in sputum were not found.
Thus, the presence of severe bronchial asthma, rhinosinusitis, high and persistent eosinophilia, transient pulmonary opacities, and polyneuropathy suggested EGPA (5 out of 6 clinical criteria of American College of Rheumatology present). Based on the clinical presentation and increased ANCA titer, the patient was diagnosed with EGPA. Systemic steroid treatment (prednisolone 300 mg intravenously and 30 mg orally) was prescribed, and 5 plasmapheresis sessions were performed. Later, prolonged combination immunosuppression (prednisolone 10 mg/day and methotrexate 25 mg/week) and controller asthma treatment (fluticasone propionate/salmeterol 250/25 µg 4 doses/ day, tiotropium bromide 18 µg per day) were prescribed. After 1 month on prednisolone and methotrexate, there was a significant neurological improvement: the motor and sensory disturbances decreased, and the patient was able to move around on his own. Further on, polyneuropathy completely resolved on the combination treatment. His asthma remained uncontrolled despite the combination treatment: he had frequent (3 times a year) severe exacerbations requiring hospitalization. In 2015, newly-emerged copious purulent sputum associated with frequent infection-related asthma exacerbations was noted. In January 2016, a patient underwent chest computed tomography (CT), which showed generalized bronchiectasis. Stenosis of the proximal bronchi was ruled out also by fibrobronchos-copy. The bronchial secretion microbiology revealed Klebsiella pneumoniae growth (107 colony-forming units).
In November 2016, during the asthma exacerbation, the patient suffered a cerebral ischemic infarction involving the right occipital lobe, confirmed by CT (Figures 1, 2).
On March 2, 2017, the patient was admitted to our hospital due to severe respiratory failure caused by pneumonia in the left upper lobe and severe asthma exacerbation.
The complete blood count showed high WBCs (15.2×109/l) with left shift. The IgE level was 144.0 IU/l, C-reactive protein was 48.8 mg/dL. Clinical chemistry was remarkable for urea nitrogen 18.4 mmol/l and creatinine 1.55 mg/dL. The sputum microscopy demonstrated 100 WBCs and 160 RBCs per vision field, and no eosinophils were revealed. A chest CT revealed multiple bronchiectasis and upper lobar pneumonia of the left lung (Figure 2). Klebsiella was again detected in the sputum culture.
The patient was prescribed intravenous antibiotics (ceftriaxone 2 g/day and ciprofloxacin 800 mg/day replaced by cefopera-zone 4 g/day and amikacin 1 g/day after 3 days due to low efficacy), bronchodilator and anti-inflammatory therapy (nebulized ipratropium bromide, fenoterol, and budesonide 2000 µg per day, prednisolone intravenously 150 mg and orally 30 mg). Despite the aggressive treatment, the respiratory failure progressed, and on day 6 of hospitalization the patient was transferred to the Intensive Care Unit and intubated. On day 7, the patient died. On autopsy, the EGPA diagnosis was confirmed, signs of ischemic infarction in the occipital lobe of the brain were found, and left upper lobe pneumonia with abscess formation and generalized bilateral bronchiectases were revealed.
Discussion
Based on the long follow-up period and careful interpretation of signs and symptoms, typical laboratory abnormalities, the diagnosis was clear. Among the most commonly used diagnostic criteria of EGPA are the American College of Rheumatology (ACR) classification criteria, Lanham’s criteria, and the Chapel Hill Consensus Conference criteria. The presence of 4 or more ACR classification criteria, including bronchial asthma, eosinophilia (more than 10%), paranasal sinus abnormalities, migratory or transient eosinophilic pulmonary infiltrates, extravascular eosinophil infiltration on biopsy, and mono- or polyneuropathy, has a sensitivity of 85% and a specificity of 100% [2,5,6]. We identified 5 out of 6 American College of Rheumatology clinical criteria present in our patient.
This clinical case shows gradual development of EGPA with multiple-organ involvement (see Figure 3). The disease onset was characterized by respiratory manifestations and suspicion of EGPA appeared only after the development of polyneuropathy (after 39 years) with hypereosinophilia. Polyneuropathy was noted at the age of 57, which is slightly later than the average age of onset reported in the literature [13]. Notably, the patient was ANCA-positive and his long-term neurological manifestation (mononeuritis) was quite typical for ANCA vasculitis [5,6,8,13]. The diagnosis became obvious after comprehensive assessment of the disease history with its gradually evolving signs and symptoms and laboratory abnormalities. Almost all manifestations of the disease seen in the patient were typical except for stroke, which is an exceptional feature for this case [1,5,6]. Neurological involvement manifested as ischemic stroke is an uncommon complication of EGPA (seen only in 5.2% of all patients) [1]. Thus, the patient demonstrated both peripheral and central nervous system involvement, which is unusual and life-threatening in EGPA.
This clinical case of EGPA also attracted our attention due to severe respiratory manifestations. While typical respiratory manifestations such as rhinosinusitis, asthma, and transient lung infiltrates are well documented in the literature, EGPA-associated bronchiectasis is rarely mentioned [14]. Bronchiectasis in the patient was detected by CT at the age of 63. The patient was a nonsmoker, and no other cause of bronchial abnormalities and airflow obstruction could be identified. Stenosis of the proximal bronchi was ruled out by fibrobronchoscopy and CT. The diagnosis of allergic bronchopulmonary aspergillosis was not confirmed. Klebsiella pneumoniae colonization was found during the last 2 years of the patient’s life. Klebsiella pneumoniae is a typical pathogen causing bronchiectasis [15]. It should be noted that aspergillosis and mycobacteriosis are common complications of bronchiectasis [15], but neither was identified in our patient.
The patient was treated for a long time with a combination of prednisolone and methotrexate. There was a significant positive effect on polyneuropathy, but uncontrolled bronchial asthma persisted. This treatment regimen complies with current recommendations [3,11]. Patients with life-threatening and/or organ-threatening manifestations (including lesions of the central nervous system and severe peripheral neuropathy) require therapy with a combination of a glucocorticoid and a cytostatic [3,11].
The use of immunosuppressive drugs in patients with ANCA-associated vasculitis significantly increases the risk of infection and mortality [16–18]. Prednisone and methotrexate use were found to be significant predictors of hospitalization for pneumonia. Frequent infections (more than 1 episode) are reported in patients with ANCA-associated vasculitis during the first year of diagnosis and initiation of therapy [18]. We assume that bronchiectasis complicated by bacterial colonization and bronchial obstruction in combination with immuno-suppression led to severe pneumonia and death of the patient.
Conclusions
The present case report demonstrates a long-term follow-up of a patient with EGPA. Both common and rare manifestations of EGPA have been described in the patient, such as central nervous system damage and bronchiectasis. Immunosuppressive treatment probably contributed to Klebsiella infection development. Thus, long-term complications of EGPA in combination with adverse effects of medications could have caused the severe pneumonia and death of the patient.
Figure 1. Brain multi-spiral computed tomography performed in 2016 showing ischemic brain damage in the right occipital lobe.
Figure 2. (A–C) Chest computed tomography of patient showing large left upper lobe pneumonia and bronchiectasis in lover lobes.
Figure 3. Timeline from first symptoms to death. | BUDESONIDE, FENOTEROL, FLUTICASONE PROPIONATE, IPRATROPIUM BROMIDE, METHOTREXATE, PREDNISOLONE, SALMETEROL, TIOTROPIUM BROMIDE | DrugsGivenReaction | CC BY-NC-ND | 33582702 | 18,975,017 | 2021-02-14 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Sputum purulent'. | Case Report: A 64-Year-Old Man with 10-Year History of Eosinophilic Granulomatosis with Polyangiitis with Bronchiectasis and Severe Klebsiella pneumonia.
BACKGROUND Eosinophilic granulomatosis with polyangiitis (EGPA) is a rare and severe progressive disease with multiple clinical manifestations and organ damage. Usually, it requires long-term monitoring of the state of many organs due to the gradual character of its manifestations. CASE REPORT We report a case of a long-term follow-up of a patient with eosinophilic granulomatosis with polyangiitis with emphasis on specific clinical features in this patient. A 64-year-old man was being followed up for 10 years. The initial diagnosis was allergic bronchial asthma; however, as new clinical signs and symptoms developed, the diagnosis of EGPA became obvious. A positive treatment response was seen, mainly manifested as reduced polyneuropathy. Meanwhile, bronchial asthma remained uncontrolled and bronchiectasis and Klebsiella pneumoniae colonization developed despite the combination treatment with prednisolone and methotrexate. Furthermore, the patient suffered a cerebral ischemic infarction. During the last hospital admission, severe uncontrolled bronchial asthma complicated with pneumonia resulted in the patient's death. CONCLUSIONS This clinical case shows the gradual development of EGPA with multiple-organ involvement, including respiratory manifestations and peripheral and central nervous system damage. Immunosuppressive treatment combined with complications of EGPA could have contributed to severe pneumonia development and death of the patient.
Background
Systemic vasculitides are difficult to diagnose due to the heterogenous clinical manifestations requiring a wide differential diagnostic search. One of the most uncommon vasculitis types is eosinophilic granulomatosis with polyangiitis (EGPA), or Churg-Strauss syndrome [1,2]. This vasculitis is associated with the presence of antineutrophilic cytoplasmic antibodies (ANCA) and is characterized by necrotizing inflammatory lesions of mainly small vessels, as well as eosinophilic and granulomatous inflammation in various organs. Bronchial asthma and eosinophilia are the most common clinical manifestations of EGPA [1–3]. In 2012, according to the current classification of systemic vasculitides, Churg-Strauss syndrome was renamed “Eosinophilic Granulomatosis with Polyangiitis” (EGPA) and classified into the ANCA-associated vasculitides group [3].
EGPA is a rare ANCA-associated vasculitis. The incidence rate is 0.5–6.8 new cases/year per million populations in Europe [2]. Prevalence is higher in northern latitudes compared to southern Europe [4]. The disease occurs most commonly in people 40–60 years of age [2,5]. No sex predominance has been clearly demonstrated [4].
The pathogenesis of the disease is still not completely clear, but the key role is attributed to autoimmune inflammation with the participation of eosinophils, CD8+, and CD4+lymphocytes (predominantly Th-2) and their proinflammatory cytokines: interleukin (IL)4, IL5, and IL13 [2,5]. Elevation of immunoglobulin (Ig)E, IgG4, and eotaxin-3 is common in EGPA patients [2,5]. The frequent (in 30–40% of cases) detection of ANCA (mainly antibodies to myeloperoxidase) also suggests the autoimmune nature of the disease. ANCA-positive patients are more commonly diagnosed with glomerulonephritis and mononeuritis, whereas the ANCA-negative ones more frequently demonstrate endocardial, myocardial, and pulmonary eosinophilic infiltrates [1,2,5,6].
There are 3 phases in the development of EGPA (these phases partially overlap and may not appear in sequence):
the allergic phase, distinguished by allergic upper respiratory tract inflammation (rhinosinusitis, allergic rhinitis) and bronchial asthma;
the eosinophilic phase, manifested with severe eosinophilia in combination with multiple eosinophilic infiltrates in various organs (most often in the lungs and gastrointestinal tract);
the vasculitis phase, with small- and medium-vessel inflammation of different organs, which can often be life-threatening [5,6].
Respiratory manifestations of EGPA include allergic rhinitis, sinusitis, nasal polyposis and, in more than 95% of patients, poorly controlled bronchial asthma [5]. Chest radiography findings include transient and migratory opacities, ground-glass opacity, focal consolidation, and small peribronchial and centrilobular nodules or masses [7]. Computed tomography reveals bronchial abnormalities, most commonly thickened wall and dilatation with secondary bronchiectasis formation, leading to frequent infections [7,8]. Pleural effusions and alveolar hemorrhages are uncommon manifestations of EGPA [1,5,7,8].
A characteristic neurological manifestation seen in 75% of patients with EGPA is peripheral neuropathy (mononeuritis multiplex) [1,5]. Upper-limb symptoms due to neuropathy, as well as vasculitis in epineural vessels, were more frequently reported in the ANCA-positive group [9]. Eosinophil-associated tissue damage and vascular occlusion leading to ischemia are more specific in ANCA-negative patients [9]. Also, the tissue damage caused by eosinophils may be associated with IL-5 concentration [10]. Brain hemorrhage, subarachnoid hemorrhage, and ischemic brain infarction are rare [1,5,6].
The main drugs for the treatment of EGPA are glucocorticoids. In organ- or life-threatening manifestations, the recommended starting dose of prednisone is 1 mg/kg/day with a maximal daily dose of 80 mg for 2–3 weeks, with tapering to a minimal supporting dose selected individually to control the symptoms and minimize the adverse effects of treatment. Methylprednisolone (7.5–15 mg/kg/day) can be used as a pulse therapy for life-threatening manifestations [11,12]. Most patients need prolonged or lifelong steroids. In the case of organ- or life-threatening manifestations, glucocorticoids in combination with cyclophosphamide are prescribed [11,12]. The combination of glucocorticoids and methotrexate or mycophenolate is required to induce remission in mild or moderate disease [11,12]. Mepolizumab (a humanized monoclonal antibody targeting IL-5) has been used for EGPA treatment [11]. For ANCA-positive patients with renal involvement or refractory disease, rituximab can be considered [3, 11].
Due to the rare occurrence of EGPA and the variety of its clinical manifestations, the diagnosis and treatment of this disease can be challenging. Currently, reports of long-term observations with the detailed patient follow-up are of great interest. The aim of our case report is to present rare manifestations of EGPA, such as bronchiectasis and stroke. We also focused on the infectious complications of immunosuppressive treatments. Thus, we would like to present a 10-year observation of a patient with EGPA.
Case Report
The male patient, born in 1953, was first hospitalized in the Pulmonary Department of our clinic in 2007 due to exacerbated asthma. The patient’s history was remarkable for multiple allergies (to house dust, cat and dog epidermis, and plant pollen) as well as nonsteroidal anti-inflammatory drugs intolerance (manifested as bronchospasm). The patient denied unhealthy habits and occupational hazards (he worked as a driver). His family history was not remarkable. The patient has had allergic rhinitis since adolescence. The first asthma symptoms appeared at the age of 18 while serving in the army. At the age of 22, an allergist diagnosed him with bronchial asthma. The patient had annual exacerbations (1–2 times a year) due to cold or allergen exposure, and acute respiratory viral infections. At the age of 42, polypoid rhinosinusitis was found, and a polypectomy was performed. Since 2006, the patient had been on controller treatment with salmeterol/fluticasone inhaler (25/250 mg, respectively) 2 doses 2 times a day.
In late 2009, extremely elevated eosinophils (53%) and total IgE (1000 IU/L) were found for the first time. Also, since 2009, the patient has had transient lung opacities, which usually disappeared after treatment with prednisolone 30 mg per day.
In August 2010, the patient noticed numbness on the back surface of the left foot, which spread to the entire left lower leg in 1 month, as well as numbness of the right foot and lower leg. Later, the patient started having tingling, twisting, and burning sensations aggravated by touching in the feet and lower legs. Also, the weakness in the distal parts of the legs developed, and he started having difficulties moving around on his own and falling while walking, and he was subsequently hospitalized in our clinic due to exacerbated asthma. On laboratory testing, high eosinophils (53%) and gamma-globulins (27.9%) were found in November 2010. The chest radiography showed emphysema and consolidation in the middle lobe of the right lung. A Neurology consult confirmed asymmetrical distal sensory-motor polyneuropathy demonstrated by lower distal asymmetrical paraparesis with decreased muscle strength in the foot flexors and extensors, more severe on the left side, low tone of the shin muscles, hypoactive Achilles reflexes, and reduced leg sensitivity starting from the middle of the shin and below, more severe in the peroneal nerve inner-vation area. The patient also demonstrated abnormal coordination (heel-to-knee, Romberg) test results and bilateral steppage gait with knee overextension. Electroneuromyography showed axonal lesion of all distal leg nerves and secondary myelinopathy. Thus, EGPA was suspected. The ANCA (anti-myeloperoxidase) titer was 30 U/ml (normal reference below 20 U/ml).
Allergic bronchopulmonary aspergillosis and other causes of hypereosinophilia were consistently ruled out. IgE and IgG for Aspergillus spp. and Aspergillus spp. in sputum were not found.
Thus, the presence of severe bronchial asthma, rhinosinusitis, high and persistent eosinophilia, transient pulmonary opacities, and polyneuropathy suggested EGPA (5 out of 6 clinical criteria of American College of Rheumatology present). Based on the clinical presentation and increased ANCA titer, the patient was diagnosed with EGPA. Systemic steroid treatment (prednisolone 300 mg intravenously and 30 mg orally) was prescribed, and 5 plasmapheresis sessions were performed. Later, prolonged combination immunosuppression (prednisolone 10 mg/day and methotrexate 25 mg/week) and controller asthma treatment (fluticasone propionate/salmeterol 250/25 µg 4 doses/ day, tiotropium bromide 18 µg per day) were prescribed. After 1 month on prednisolone and methotrexate, there was a significant neurological improvement: the motor and sensory disturbances decreased, and the patient was able to move around on his own. Further on, polyneuropathy completely resolved on the combination treatment. His asthma remained uncontrolled despite the combination treatment: he had frequent (3 times a year) severe exacerbations requiring hospitalization. In 2015, newly-emerged copious purulent sputum associated with frequent infection-related asthma exacerbations was noted. In January 2016, a patient underwent chest computed tomography (CT), which showed generalized bronchiectasis. Stenosis of the proximal bronchi was ruled out also by fibrobronchos-copy. The bronchial secretion microbiology revealed Klebsiella pneumoniae growth (107 colony-forming units).
In November 2016, during the asthma exacerbation, the patient suffered a cerebral ischemic infarction involving the right occipital lobe, confirmed by CT (Figures 1, 2).
On March 2, 2017, the patient was admitted to our hospital due to severe respiratory failure caused by pneumonia in the left upper lobe and severe asthma exacerbation.
The complete blood count showed high WBCs (15.2×109/l) with left shift. The IgE level was 144.0 IU/l, C-reactive protein was 48.8 mg/dL. Clinical chemistry was remarkable for urea nitrogen 18.4 mmol/l and creatinine 1.55 mg/dL. The sputum microscopy demonstrated 100 WBCs and 160 RBCs per vision field, and no eosinophils were revealed. A chest CT revealed multiple bronchiectasis and upper lobar pneumonia of the left lung (Figure 2). Klebsiella was again detected in the sputum culture.
The patient was prescribed intravenous antibiotics (ceftriaxone 2 g/day and ciprofloxacin 800 mg/day replaced by cefopera-zone 4 g/day and amikacin 1 g/day after 3 days due to low efficacy), bronchodilator and anti-inflammatory therapy (nebulized ipratropium bromide, fenoterol, and budesonide 2000 µg per day, prednisolone intravenously 150 mg and orally 30 mg). Despite the aggressive treatment, the respiratory failure progressed, and on day 6 of hospitalization the patient was transferred to the Intensive Care Unit and intubated. On day 7, the patient died. On autopsy, the EGPA diagnosis was confirmed, signs of ischemic infarction in the occipital lobe of the brain were found, and left upper lobe pneumonia with abscess formation and generalized bilateral bronchiectases were revealed.
Discussion
Based on the long follow-up period and careful interpretation of signs and symptoms, typical laboratory abnormalities, the diagnosis was clear. Among the most commonly used diagnostic criteria of EGPA are the American College of Rheumatology (ACR) classification criteria, Lanham’s criteria, and the Chapel Hill Consensus Conference criteria. The presence of 4 or more ACR classification criteria, including bronchial asthma, eosinophilia (more than 10%), paranasal sinus abnormalities, migratory or transient eosinophilic pulmonary infiltrates, extravascular eosinophil infiltration on biopsy, and mono- or polyneuropathy, has a sensitivity of 85% and a specificity of 100% [2,5,6]. We identified 5 out of 6 American College of Rheumatology clinical criteria present in our patient.
This clinical case shows gradual development of EGPA with multiple-organ involvement (see Figure 3). The disease onset was characterized by respiratory manifestations and suspicion of EGPA appeared only after the development of polyneuropathy (after 39 years) with hypereosinophilia. Polyneuropathy was noted at the age of 57, which is slightly later than the average age of onset reported in the literature [13]. Notably, the patient was ANCA-positive and his long-term neurological manifestation (mononeuritis) was quite typical for ANCA vasculitis [5,6,8,13]. The diagnosis became obvious after comprehensive assessment of the disease history with its gradually evolving signs and symptoms and laboratory abnormalities. Almost all manifestations of the disease seen in the patient were typical except for stroke, which is an exceptional feature for this case [1,5,6]. Neurological involvement manifested as ischemic stroke is an uncommon complication of EGPA (seen only in 5.2% of all patients) [1]. Thus, the patient demonstrated both peripheral and central nervous system involvement, which is unusual and life-threatening in EGPA.
This clinical case of EGPA also attracted our attention due to severe respiratory manifestations. While typical respiratory manifestations such as rhinosinusitis, asthma, and transient lung infiltrates are well documented in the literature, EGPA-associated bronchiectasis is rarely mentioned [14]. Bronchiectasis in the patient was detected by CT at the age of 63. The patient was a nonsmoker, and no other cause of bronchial abnormalities and airflow obstruction could be identified. Stenosis of the proximal bronchi was ruled out by fibrobronchoscopy and CT. The diagnosis of allergic bronchopulmonary aspergillosis was not confirmed. Klebsiella pneumoniae colonization was found during the last 2 years of the patient’s life. Klebsiella pneumoniae is a typical pathogen causing bronchiectasis [15]. It should be noted that aspergillosis and mycobacteriosis are common complications of bronchiectasis [15], but neither was identified in our patient.
The patient was treated for a long time with a combination of prednisolone and methotrexate. There was a significant positive effect on polyneuropathy, but uncontrolled bronchial asthma persisted. This treatment regimen complies with current recommendations [3,11]. Patients with life-threatening and/or organ-threatening manifestations (including lesions of the central nervous system and severe peripheral neuropathy) require therapy with a combination of a glucocorticoid and a cytostatic [3,11].
The use of immunosuppressive drugs in patients with ANCA-associated vasculitis significantly increases the risk of infection and mortality [16–18]. Prednisone and methotrexate use were found to be significant predictors of hospitalization for pneumonia. Frequent infections (more than 1 episode) are reported in patients with ANCA-associated vasculitis during the first year of diagnosis and initiation of therapy [18]. We assume that bronchiectasis complicated by bacterial colonization and bronchial obstruction in combination with immuno-suppression led to severe pneumonia and death of the patient.
Conclusions
The present case report demonstrates a long-term follow-up of a patient with EGPA. Both common and rare manifestations of EGPA have been described in the patient, such as central nervous system damage and bronchiectasis. Immunosuppressive treatment probably contributed to Klebsiella infection development. Thus, long-term complications of EGPA in combination with adverse effects of medications could have caused the severe pneumonia and death of the patient.
Figure 1. Brain multi-spiral computed tomography performed in 2016 showing ischemic brain damage in the right occipital lobe.
Figure 2. (A–C) Chest computed tomography of patient showing large left upper lobe pneumonia and bronchiectasis in lover lobes.
Figure 3. Timeline from first symptoms to death. | AMIKACIN SULFATE, BUDESONIDE, CEFOPERAZONE, CEFTRIAXONE, CIPROFLOXACIN, FENOTEROL, FLUTICASONE PROPIONATE\SALMETEROL XINAFOATE, IPRATROPIUM BROMIDE, METHOTREXATE, PREDNISOLONE, TIOTROPIUM BROMIDE | DrugsGivenReaction | CC BY-NC-ND | 33582702 | 18,962,812 | 2021-02-14 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Therapeutic product effect incomplete'. | Case Report: A 64-Year-Old Man with 10-Year History of Eosinophilic Granulomatosis with Polyangiitis with Bronchiectasis and Severe Klebsiella pneumonia.
BACKGROUND Eosinophilic granulomatosis with polyangiitis (EGPA) is a rare and severe progressive disease with multiple clinical manifestations and organ damage. Usually, it requires long-term monitoring of the state of many organs due to the gradual character of its manifestations. CASE REPORT We report a case of a long-term follow-up of a patient with eosinophilic granulomatosis with polyangiitis with emphasis on specific clinical features in this patient. A 64-year-old man was being followed up for 10 years. The initial diagnosis was allergic bronchial asthma; however, as new clinical signs and symptoms developed, the diagnosis of EGPA became obvious. A positive treatment response was seen, mainly manifested as reduced polyneuropathy. Meanwhile, bronchial asthma remained uncontrolled and bronchiectasis and Klebsiella pneumoniae colonization developed despite the combination treatment with prednisolone and methotrexate. Furthermore, the patient suffered a cerebral ischemic infarction. During the last hospital admission, severe uncontrolled bronchial asthma complicated with pneumonia resulted in the patient's death. CONCLUSIONS This clinical case shows the gradual development of EGPA with multiple-organ involvement, including respiratory manifestations and peripheral and central nervous system damage. Immunosuppressive treatment combined with complications of EGPA could have contributed to severe pneumonia development and death of the patient.
Background
Systemic vasculitides are difficult to diagnose due to the heterogenous clinical manifestations requiring a wide differential diagnostic search. One of the most uncommon vasculitis types is eosinophilic granulomatosis with polyangiitis (EGPA), or Churg-Strauss syndrome [1,2]. This vasculitis is associated with the presence of antineutrophilic cytoplasmic antibodies (ANCA) and is characterized by necrotizing inflammatory lesions of mainly small vessels, as well as eosinophilic and granulomatous inflammation in various organs. Bronchial asthma and eosinophilia are the most common clinical manifestations of EGPA [1–3]. In 2012, according to the current classification of systemic vasculitides, Churg-Strauss syndrome was renamed “Eosinophilic Granulomatosis with Polyangiitis” (EGPA) and classified into the ANCA-associated vasculitides group [3].
EGPA is a rare ANCA-associated vasculitis. The incidence rate is 0.5–6.8 new cases/year per million populations in Europe [2]. Prevalence is higher in northern latitudes compared to southern Europe [4]. The disease occurs most commonly in people 40–60 years of age [2,5]. No sex predominance has been clearly demonstrated [4].
The pathogenesis of the disease is still not completely clear, but the key role is attributed to autoimmune inflammation with the participation of eosinophils, CD8+, and CD4+lymphocytes (predominantly Th-2) and their proinflammatory cytokines: interleukin (IL)4, IL5, and IL13 [2,5]. Elevation of immunoglobulin (Ig)E, IgG4, and eotaxin-3 is common in EGPA patients [2,5]. The frequent (in 30–40% of cases) detection of ANCA (mainly antibodies to myeloperoxidase) also suggests the autoimmune nature of the disease. ANCA-positive patients are more commonly diagnosed with glomerulonephritis and mononeuritis, whereas the ANCA-negative ones more frequently demonstrate endocardial, myocardial, and pulmonary eosinophilic infiltrates [1,2,5,6].
There are 3 phases in the development of EGPA (these phases partially overlap and may not appear in sequence):
the allergic phase, distinguished by allergic upper respiratory tract inflammation (rhinosinusitis, allergic rhinitis) and bronchial asthma;
the eosinophilic phase, manifested with severe eosinophilia in combination with multiple eosinophilic infiltrates in various organs (most often in the lungs and gastrointestinal tract);
the vasculitis phase, with small- and medium-vessel inflammation of different organs, which can often be life-threatening [5,6].
Respiratory manifestations of EGPA include allergic rhinitis, sinusitis, nasal polyposis and, in more than 95% of patients, poorly controlled bronchial asthma [5]. Chest radiography findings include transient and migratory opacities, ground-glass opacity, focal consolidation, and small peribronchial and centrilobular nodules or masses [7]. Computed tomography reveals bronchial abnormalities, most commonly thickened wall and dilatation with secondary bronchiectasis formation, leading to frequent infections [7,8]. Pleural effusions and alveolar hemorrhages are uncommon manifestations of EGPA [1,5,7,8].
A characteristic neurological manifestation seen in 75% of patients with EGPA is peripheral neuropathy (mononeuritis multiplex) [1,5]. Upper-limb symptoms due to neuropathy, as well as vasculitis in epineural vessels, were more frequently reported in the ANCA-positive group [9]. Eosinophil-associated tissue damage and vascular occlusion leading to ischemia are more specific in ANCA-negative patients [9]. Also, the tissue damage caused by eosinophils may be associated with IL-5 concentration [10]. Brain hemorrhage, subarachnoid hemorrhage, and ischemic brain infarction are rare [1,5,6].
The main drugs for the treatment of EGPA are glucocorticoids. In organ- or life-threatening manifestations, the recommended starting dose of prednisone is 1 mg/kg/day with a maximal daily dose of 80 mg for 2–3 weeks, with tapering to a minimal supporting dose selected individually to control the symptoms and minimize the adverse effects of treatment. Methylprednisolone (7.5–15 mg/kg/day) can be used as a pulse therapy for life-threatening manifestations [11,12]. Most patients need prolonged or lifelong steroids. In the case of organ- or life-threatening manifestations, glucocorticoids in combination with cyclophosphamide are prescribed [11,12]. The combination of glucocorticoids and methotrexate or mycophenolate is required to induce remission in mild or moderate disease [11,12]. Mepolizumab (a humanized monoclonal antibody targeting IL-5) has been used for EGPA treatment [11]. For ANCA-positive patients with renal involvement or refractory disease, rituximab can be considered [3, 11].
Due to the rare occurrence of EGPA and the variety of its clinical manifestations, the diagnosis and treatment of this disease can be challenging. Currently, reports of long-term observations with the detailed patient follow-up are of great interest. The aim of our case report is to present rare manifestations of EGPA, such as bronchiectasis and stroke. We also focused on the infectious complications of immunosuppressive treatments. Thus, we would like to present a 10-year observation of a patient with EGPA.
Case Report
The male patient, born in 1953, was first hospitalized in the Pulmonary Department of our clinic in 2007 due to exacerbated asthma. The patient’s history was remarkable for multiple allergies (to house dust, cat and dog epidermis, and plant pollen) as well as nonsteroidal anti-inflammatory drugs intolerance (manifested as bronchospasm). The patient denied unhealthy habits and occupational hazards (he worked as a driver). His family history was not remarkable. The patient has had allergic rhinitis since adolescence. The first asthma symptoms appeared at the age of 18 while serving in the army. At the age of 22, an allergist diagnosed him with bronchial asthma. The patient had annual exacerbations (1–2 times a year) due to cold or allergen exposure, and acute respiratory viral infections. At the age of 42, polypoid rhinosinusitis was found, and a polypectomy was performed. Since 2006, the patient had been on controller treatment with salmeterol/fluticasone inhaler (25/250 mg, respectively) 2 doses 2 times a day.
In late 2009, extremely elevated eosinophils (53%) and total IgE (1000 IU/L) were found for the first time. Also, since 2009, the patient has had transient lung opacities, which usually disappeared after treatment with prednisolone 30 mg per day.
In August 2010, the patient noticed numbness on the back surface of the left foot, which spread to the entire left lower leg in 1 month, as well as numbness of the right foot and lower leg. Later, the patient started having tingling, twisting, and burning sensations aggravated by touching in the feet and lower legs. Also, the weakness in the distal parts of the legs developed, and he started having difficulties moving around on his own and falling while walking, and he was subsequently hospitalized in our clinic due to exacerbated asthma. On laboratory testing, high eosinophils (53%) and gamma-globulins (27.9%) were found in November 2010. The chest radiography showed emphysema and consolidation in the middle lobe of the right lung. A Neurology consult confirmed asymmetrical distal sensory-motor polyneuropathy demonstrated by lower distal asymmetrical paraparesis with decreased muscle strength in the foot flexors and extensors, more severe on the left side, low tone of the shin muscles, hypoactive Achilles reflexes, and reduced leg sensitivity starting from the middle of the shin and below, more severe in the peroneal nerve inner-vation area. The patient also demonstrated abnormal coordination (heel-to-knee, Romberg) test results and bilateral steppage gait with knee overextension. Electroneuromyography showed axonal lesion of all distal leg nerves and secondary myelinopathy. Thus, EGPA was suspected. The ANCA (anti-myeloperoxidase) titer was 30 U/ml (normal reference below 20 U/ml).
Allergic bronchopulmonary aspergillosis and other causes of hypereosinophilia were consistently ruled out. IgE and IgG for Aspergillus spp. and Aspergillus spp. in sputum were not found.
Thus, the presence of severe bronchial asthma, rhinosinusitis, high and persistent eosinophilia, transient pulmonary opacities, and polyneuropathy suggested EGPA (5 out of 6 clinical criteria of American College of Rheumatology present). Based on the clinical presentation and increased ANCA titer, the patient was diagnosed with EGPA. Systemic steroid treatment (prednisolone 300 mg intravenously and 30 mg orally) was prescribed, and 5 plasmapheresis sessions were performed. Later, prolonged combination immunosuppression (prednisolone 10 mg/day and methotrexate 25 mg/week) and controller asthma treatment (fluticasone propionate/salmeterol 250/25 µg 4 doses/ day, tiotropium bromide 18 µg per day) were prescribed. After 1 month on prednisolone and methotrexate, there was a significant neurological improvement: the motor and sensory disturbances decreased, and the patient was able to move around on his own. Further on, polyneuropathy completely resolved on the combination treatment. His asthma remained uncontrolled despite the combination treatment: he had frequent (3 times a year) severe exacerbations requiring hospitalization. In 2015, newly-emerged copious purulent sputum associated with frequent infection-related asthma exacerbations was noted. In January 2016, a patient underwent chest computed tomography (CT), which showed generalized bronchiectasis. Stenosis of the proximal bronchi was ruled out also by fibrobronchos-copy. The bronchial secretion microbiology revealed Klebsiella pneumoniae growth (107 colony-forming units).
In November 2016, during the asthma exacerbation, the patient suffered a cerebral ischemic infarction involving the right occipital lobe, confirmed by CT (Figures 1, 2).
On March 2, 2017, the patient was admitted to our hospital due to severe respiratory failure caused by pneumonia in the left upper lobe and severe asthma exacerbation.
The complete blood count showed high WBCs (15.2×109/l) with left shift. The IgE level was 144.0 IU/l, C-reactive protein was 48.8 mg/dL. Clinical chemistry was remarkable for urea nitrogen 18.4 mmol/l and creatinine 1.55 mg/dL. The sputum microscopy demonstrated 100 WBCs and 160 RBCs per vision field, and no eosinophils were revealed. A chest CT revealed multiple bronchiectasis and upper lobar pneumonia of the left lung (Figure 2). Klebsiella was again detected in the sputum culture.
The patient was prescribed intravenous antibiotics (ceftriaxone 2 g/day and ciprofloxacin 800 mg/day replaced by cefopera-zone 4 g/day and amikacin 1 g/day after 3 days due to low efficacy), bronchodilator and anti-inflammatory therapy (nebulized ipratropium bromide, fenoterol, and budesonide 2000 µg per day, prednisolone intravenously 150 mg and orally 30 mg). Despite the aggressive treatment, the respiratory failure progressed, and on day 6 of hospitalization the patient was transferred to the Intensive Care Unit and intubated. On day 7, the patient died. On autopsy, the EGPA diagnosis was confirmed, signs of ischemic infarction in the occipital lobe of the brain were found, and left upper lobe pneumonia with abscess formation and generalized bilateral bronchiectases were revealed.
Discussion
Based on the long follow-up period and careful interpretation of signs and symptoms, typical laboratory abnormalities, the diagnosis was clear. Among the most commonly used diagnostic criteria of EGPA are the American College of Rheumatology (ACR) classification criteria, Lanham’s criteria, and the Chapel Hill Consensus Conference criteria. The presence of 4 or more ACR classification criteria, including bronchial asthma, eosinophilia (more than 10%), paranasal sinus abnormalities, migratory or transient eosinophilic pulmonary infiltrates, extravascular eosinophil infiltration on biopsy, and mono- or polyneuropathy, has a sensitivity of 85% and a specificity of 100% [2,5,6]. We identified 5 out of 6 American College of Rheumatology clinical criteria present in our patient.
This clinical case shows gradual development of EGPA with multiple-organ involvement (see Figure 3). The disease onset was characterized by respiratory manifestations and suspicion of EGPA appeared only after the development of polyneuropathy (after 39 years) with hypereosinophilia. Polyneuropathy was noted at the age of 57, which is slightly later than the average age of onset reported in the literature [13]. Notably, the patient was ANCA-positive and his long-term neurological manifestation (mononeuritis) was quite typical for ANCA vasculitis [5,6,8,13]. The diagnosis became obvious after comprehensive assessment of the disease history with its gradually evolving signs and symptoms and laboratory abnormalities. Almost all manifestations of the disease seen in the patient were typical except for stroke, which is an exceptional feature for this case [1,5,6]. Neurological involvement manifested as ischemic stroke is an uncommon complication of EGPA (seen only in 5.2% of all patients) [1]. Thus, the patient demonstrated both peripheral and central nervous system involvement, which is unusual and life-threatening in EGPA.
This clinical case of EGPA also attracted our attention due to severe respiratory manifestations. While typical respiratory manifestations such as rhinosinusitis, asthma, and transient lung infiltrates are well documented in the literature, EGPA-associated bronchiectasis is rarely mentioned [14]. Bronchiectasis in the patient was detected by CT at the age of 63. The patient was a nonsmoker, and no other cause of bronchial abnormalities and airflow obstruction could be identified. Stenosis of the proximal bronchi was ruled out by fibrobronchoscopy and CT. The diagnosis of allergic bronchopulmonary aspergillosis was not confirmed. Klebsiella pneumoniae colonization was found during the last 2 years of the patient’s life. Klebsiella pneumoniae is a typical pathogen causing bronchiectasis [15]. It should be noted that aspergillosis and mycobacteriosis are common complications of bronchiectasis [15], but neither was identified in our patient.
The patient was treated for a long time with a combination of prednisolone and methotrexate. There was a significant positive effect on polyneuropathy, but uncontrolled bronchial asthma persisted. This treatment regimen complies with current recommendations [3,11]. Patients with life-threatening and/or organ-threatening manifestations (including lesions of the central nervous system and severe peripheral neuropathy) require therapy with a combination of a glucocorticoid and a cytostatic [3,11].
The use of immunosuppressive drugs in patients with ANCA-associated vasculitis significantly increases the risk of infection and mortality [16–18]. Prednisone and methotrexate use were found to be significant predictors of hospitalization for pneumonia. Frequent infections (more than 1 episode) are reported in patients with ANCA-associated vasculitis during the first year of diagnosis and initiation of therapy [18]. We assume that bronchiectasis complicated by bacterial colonization and bronchial obstruction in combination with immuno-suppression led to severe pneumonia and death of the patient.
Conclusions
The present case report demonstrates a long-term follow-up of a patient with EGPA. Both common and rare manifestations of EGPA have been described in the patient, such as central nervous system damage and bronchiectasis. Immunosuppressive treatment probably contributed to Klebsiella infection development. Thus, long-term complications of EGPA in combination with adverse effects of medications could have caused the severe pneumonia and death of the patient.
Figure 1. Brain multi-spiral computed tomography performed in 2016 showing ischemic brain damage in the right occipital lobe.
Figure 2. (A–C) Chest computed tomography of patient showing large left upper lobe pneumonia and bronchiectasis in lover lobes.
Figure 3. Timeline from first symptoms to death. | AMIKACIN SULFATE, BUDESONIDE, CEFOPERAZONE, CEFTRIAXONE, CIPROFLOXACIN, FENOTEROL, FLUTICASONE PROPIONATE\SALMETEROL XINAFOATE, IPRATROPIUM BROMIDE, METHOTREXATE, PREDNISOLONE, TIOTROPIUM BROMIDE | DrugsGivenReaction | CC BY-NC-ND | 33582702 | 18,962,812 | 2021-02-14 |
What is the weight of the patient? | Case Report: A 64-Year-Old Man with 10-Year History of Eosinophilic Granulomatosis with Polyangiitis with Bronchiectasis and Severe Klebsiella pneumonia.
BACKGROUND Eosinophilic granulomatosis with polyangiitis (EGPA) is a rare and severe progressive disease with multiple clinical manifestations and organ damage. Usually, it requires long-term monitoring of the state of many organs due to the gradual character of its manifestations. CASE REPORT We report a case of a long-term follow-up of a patient with eosinophilic granulomatosis with polyangiitis with emphasis on specific clinical features in this patient. A 64-year-old man was being followed up for 10 years. The initial diagnosis was allergic bronchial asthma; however, as new clinical signs and symptoms developed, the diagnosis of EGPA became obvious. A positive treatment response was seen, mainly manifested as reduced polyneuropathy. Meanwhile, bronchial asthma remained uncontrolled and bronchiectasis and Klebsiella pneumoniae colonization developed despite the combination treatment with prednisolone and methotrexate. Furthermore, the patient suffered a cerebral ischemic infarction. During the last hospital admission, severe uncontrolled bronchial asthma complicated with pneumonia resulted in the patient's death. CONCLUSIONS This clinical case shows the gradual development of EGPA with multiple-organ involvement, including respiratory manifestations and peripheral and central nervous system damage. Immunosuppressive treatment combined with complications of EGPA could have contributed to severe pneumonia development and death of the patient.
Background
Systemic vasculitides are difficult to diagnose due to the heterogenous clinical manifestations requiring a wide differential diagnostic search. One of the most uncommon vasculitis types is eosinophilic granulomatosis with polyangiitis (EGPA), or Churg-Strauss syndrome [1,2]. This vasculitis is associated with the presence of antineutrophilic cytoplasmic antibodies (ANCA) and is characterized by necrotizing inflammatory lesions of mainly small vessels, as well as eosinophilic and granulomatous inflammation in various organs. Bronchial asthma and eosinophilia are the most common clinical manifestations of EGPA [1–3]. In 2012, according to the current classification of systemic vasculitides, Churg-Strauss syndrome was renamed “Eosinophilic Granulomatosis with Polyangiitis” (EGPA) and classified into the ANCA-associated vasculitides group [3].
EGPA is a rare ANCA-associated vasculitis. The incidence rate is 0.5–6.8 new cases/year per million populations in Europe [2]. Prevalence is higher in northern latitudes compared to southern Europe [4]. The disease occurs most commonly in people 40–60 years of age [2,5]. No sex predominance has been clearly demonstrated [4].
The pathogenesis of the disease is still not completely clear, but the key role is attributed to autoimmune inflammation with the participation of eosinophils, CD8+, and CD4+lymphocytes (predominantly Th-2) and their proinflammatory cytokines: interleukin (IL)4, IL5, and IL13 [2,5]. Elevation of immunoglobulin (Ig)E, IgG4, and eotaxin-3 is common in EGPA patients [2,5]. The frequent (in 30–40% of cases) detection of ANCA (mainly antibodies to myeloperoxidase) also suggests the autoimmune nature of the disease. ANCA-positive patients are more commonly diagnosed with glomerulonephritis and mononeuritis, whereas the ANCA-negative ones more frequently demonstrate endocardial, myocardial, and pulmonary eosinophilic infiltrates [1,2,5,6].
There are 3 phases in the development of EGPA (these phases partially overlap and may not appear in sequence):
the allergic phase, distinguished by allergic upper respiratory tract inflammation (rhinosinusitis, allergic rhinitis) and bronchial asthma;
the eosinophilic phase, manifested with severe eosinophilia in combination with multiple eosinophilic infiltrates in various organs (most often in the lungs and gastrointestinal tract);
the vasculitis phase, with small- and medium-vessel inflammation of different organs, which can often be life-threatening [5,6].
Respiratory manifestations of EGPA include allergic rhinitis, sinusitis, nasal polyposis and, in more than 95% of patients, poorly controlled bronchial asthma [5]. Chest radiography findings include transient and migratory opacities, ground-glass opacity, focal consolidation, and small peribronchial and centrilobular nodules or masses [7]. Computed tomography reveals bronchial abnormalities, most commonly thickened wall and dilatation with secondary bronchiectasis formation, leading to frequent infections [7,8]. Pleural effusions and alveolar hemorrhages are uncommon manifestations of EGPA [1,5,7,8].
A characteristic neurological manifestation seen in 75% of patients with EGPA is peripheral neuropathy (mononeuritis multiplex) [1,5]. Upper-limb symptoms due to neuropathy, as well as vasculitis in epineural vessels, were more frequently reported in the ANCA-positive group [9]. Eosinophil-associated tissue damage and vascular occlusion leading to ischemia are more specific in ANCA-negative patients [9]. Also, the tissue damage caused by eosinophils may be associated with IL-5 concentration [10]. Brain hemorrhage, subarachnoid hemorrhage, and ischemic brain infarction are rare [1,5,6].
The main drugs for the treatment of EGPA are glucocorticoids. In organ- or life-threatening manifestations, the recommended starting dose of prednisone is 1 mg/kg/day with a maximal daily dose of 80 mg for 2–3 weeks, with tapering to a minimal supporting dose selected individually to control the symptoms and minimize the adverse effects of treatment. Methylprednisolone (7.5–15 mg/kg/day) can be used as a pulse therapy for life-threatening manifestations [11,12]. Most patients need prolonged or lifelong steroids. In the case of organ- or life-threatening manifestations, glucocorticoids in combination with cyclophosphamide are prescribed [11,12]. The combination of glucocorticoids and methotrexate or mycophenolate is required to induce remission in mild or moderate disease [11,12]. Mepolizumab (a humanized monoclonal antibody targeting IL-5) has been used for EGPA treatment [11]. For ANCA-positive patients with renal involvement or refractory disease, rituximab can be considered [3, 11].
Due to the rare occurrence of EGPA and the variety of its clinical manifestations, the diagnosis and treatment of this disease can be challenging. Currently, reports of long-term observations with the detailed patient follow-up are of great interest. The aim of our case report is to present rare manifestations of EGPA, such as bronchiectasis and stroke. We also focused on the infectious complications of immunosuppressive treatments. Thus, we would like to present a 10-year observation of a patient with EGPA.
Case Report
The male patient, born in 1953, was first hospitalized in the Pulmonary Department of our clinic in 2007 due to exacerbated asthma. The patient’s history was remarkable for multiple allergies (to house dust, cat and dog epidermis, and plant pollen) as well as nonsteroidal anti-inflammatory drugs intolerance (manifested as bronchospasm). The patient denied unhealthy habits and occupational hazards (he worked as a driver). His family history was not remarkable. The patient has had allergic rhinitis since adolescence. The first asthma symptoms appeared at the age of 18 while serving in the army. At the age of 22, an allergist diagnosed him with bronchial asthma. The patient had annual exacerbations (1–2 times a year) due to cold or allergen exposure, and acute respiratory viral infections. At the age of 42, polypoid rhinosinusitis was found, and a polypectomy was performed. Since 2006, the patient had been on controller treatment with salmeterol/fluticasone inhaler (25/250 mg, respectively) 2 doses 2 times a day.
In late 2009, extremely elevated eosinophils (53%) and total IgE (1000 IU/L) were found for the first time. Also, since 2009, the patient has had transient lung opacities, which usually disappeared after treatment with prednisolone 30 mg per day.
In August 2010, the patient noticed numbness on the back surface of the left foot, which spread to the entire left lower leg in 1 month, as well as numbness of the right foot and lower leg. Later, the patient started having tingling, twisting, and burning sensations aggravated by touching in the feet and lower legs. Also, the weakness in the distal parts of the legs developed, and he started having difficulties moving around on his own and falling while walking, and he was subsequently hospitalized in our clinic due to exacerbated asthma. On laboratory testing, high eosinophils (53%) and gamma-globulins (27.9%) were found in November 2010. The chest radiography showed emphysema and consolidation in the middle lobe of the right lung. A Neurology consult confirmed asymmetrical distal sensory-motor polyneuropathy demonstrated by lower distal asymmetrical paraparesis with decreased muscle strength in the foot flexors and extensors, more severe on the left side, low tone of the shin muscles, hypoactive Achilles reflexes, and reduced leg sensitivity starting from the middle of the shin and below, more severe in the peroneal nerve inner-vation area. The patient also demonstrated abnormal coordination (heel-to-knee, Romberg) test results and bilateral steppage gait with knee overextension. Electroneuromyography showed axonal lesion of all distal leg nerves and secondary myelinopathy. Thus, EGPA was suspected. The ANCA (anti-myeloperoxidase) titer was 30 U/ml (normal reference below 20 U/ml).
Allergic bronchopulmonary aspergillosis and other causes of hypereosinophilia were consistently ruled out. IgE and IgG for Aspergillus spp. and Aspergillus spp. in sputum were not found.
Thus, the presence of severe bronchial asthma, rhinosinusitis, high and persistent eosinophilia, transient pulmonary opacities, and polyneuropathy suggested EGPA (5 out of 6 clinical criteria of American College of Rheumatology present). Based on the clinical presentation and increased ANCA titer, the patient was diagnosed with EGPA. Systemic steroid treatment (prednisolone 300 mg intravenously and 30 mg orally) was prescribed, and 5 plasmapheresis sessions were performed. Later, prolonged combination immunosuppression (prednisolone 10 mg/day and methotrexate 25 mg/week) and controller asthma treatment (fluticasone propionate/salmeterol 250/25 µg 4 doses/ day, tiotropium bromide 18 µg per day) were prescribed. After 1 month on prednisolone and methotrexate, there was a significant neurological improvement: the motor and sensory disturbances decreased, and the patient was able to move around on his own. Further on, polyneuropathy completely resolved on the combination treatment. His asthma remained uncontrolled despite the combination treatment: he had frequent (3 times a year) severe exacerbations requiring hospitalization. In 2015, newly-emerged copious purulent sputum associated with frequent infection-related asthma exacerbations was noted. In January 2016, a patient underwent chest computed tomography (CT), which showed generalized bronchiectasis. Stenosis of the proximal bronchi was ruled out also by fibrobronchos-copy. The bronchial secretion microbiology revealed Klebsiella pneumoniae growth (107 colony-forming units).
In November 2016, during the asthma exacerbation, the patient suffered a cerebral ischemic infarction involving the right occipital lobe, confirmed by CT (Figures 1, 2).
On March 2, 2017, the patient was admitted to our hospital due to severe respiratory failure caused by pneumonia in the left upper lobe and severe asthma exacerbation.
The complete blood count showed high WBCs (15.2×109/l) with left shift. The IgE level was 144.0 IU/l, C-reactive protein was 48.8 mg/dL. Clinical chemistry was remarkable for urea nitrogen 18.4 mmol/l and creatinine 1.55 mg/dL. The sputum microscopy demonstrated 100 WBCs and 160 RBCs per vision field, and no eosinophils were revealed. A chest CT revealed multiple bronchiectasis and upper lobar pneumonia of the left lung (Figure 2). Klebsiella was again detected in the sputum culture.
The patient was prescribed intravenous antibiotics (ceftriaxone 2 g/day and ciprofloxacin 800 mg/day replaced by cefopera-zone 4 g/day and amikacin 1 g/day after 3 days due to low efficacy), bronchodilator and anti-inflammatory therapy (nebulized ipratropium bromide, fenoterol, and budesonide 2000 µg per day, prednisolone intravenously 150 mg and orally 30 mg). Despite the aggressive treatment, the respiratory failure progressed, and on day 6 of hospitalization the patient was transferred to the Intensive Care Unit and intubated. On day 7, the patient died. On autopsy, the EGPA diagnosis was confirmed, signs of ischemic infarction in the occipital lobe of the brain were found, and left upper lobe pneumonia with abscess formation and generalized bilateral bronchiectases were revealed.
Discussion
Based on the long follow-up period and careful interpretation of signs and symptoms, typical laboratory abnormalities, the diagnosis was clear. Among the most commonly used diagnostic criteria of EGPA are the American College of Rheumatology (ACR) classification criteria, Lanham’s criteria, and the Chapel Hill Consensus Conference criteria. The presence of 4 or more ACR classification criteria, including bronchial asthma, eosinophilia (more than 10%), paranasal sinus abnormalities, migratory or transient eosinophilic pulmonary infiltrates, extravascular eosinophil infiltration on biopsy, and mono- or polyneuropathy, has a sensitivity of 85% and a specificity of 100% [2,5,6]. We identified 5 out of 6 American College of Rheumatology clinical criteria present in our patient.
This clinical case shows gradual development of EGPA with multiple-organ involvement (see Figure 3). The disease onset was characterized by respiratory manifestations and suspicion of EGPA appeared only after the development of polyneuropathy (after 39 years) with hypereosinophilia. Polyneuropathy was noted at the age of 57, which is slightly later than the average age of onset reported in the literature [13]. Notably, the patient was ANCA-positive and his long-term neurological manifestation (mononeuritis) was quite typical for ANCA vasculitis [5,6,8,13]. The diagnosis became obvious after comprehensive assessment of the disease history with its gradually evolving signs and symptoms and laboratory abnormalities. Almost all manifestations of the disease seen in the patient were typical except for stroke, which is an exceptional feature for this case [1,5,6]. Neurological involvement manifested as ischemic stroke is an uncommon complication of EGPA (seen only in 5.2% of all patients) [1]. Thus, the patient demonstrated both peripheral and central nervous system involvement, which is unusual and life-threatening in EGPA.
This clinical case of EGPA also attracted our attention due to severe respiratory manifestations. While typical respiratory manifestations such as rhinosinusitis, asthma, and transient lung infiltrates are well documented in the literature, EGPA-associated bronchiectasis is rarely mentioned [14]. Bronchiectasis in the patient was detected by CT at the age of 63. The patient was a nonsmoker, and no other cause of bronchial abnormalities and airflow obstruction could be identified. Stenosis of the proximal bronchi was ruled out by fibrobronchoscopy and CT. The diagnosis of allergic bronchopulmonary aspergillosis was not confirmed. Klebsiella pneumoniae colonization was found during the last 2 years of the patient’s life. Klebsiella pneumoniae is a typical pathogen causing bronchiectasis [15]. It should be noted that aspergillosis and mycobacteriosis are common complications of bronchiectasis [15], but neither was identified in our patient.
The patient was treated for a long time with a combination of prednisolone and methotrexate. There was a significant positive effect on polyneuropathy, but uncontrolled bronchial asthma persisted. This treatment regimen complies with current recommendations [3,11]. Patients with life-threatening and/or organ-threatening manifestations (including lesions of the central nervous system and severe peripheral neuropathy) require therapy with a combination of a glucocorticoid and a cytostatic [3,11].
The use of immunosuppressive drugs in patients with ANCA-associated vasculitis significantly increases the risk of infection and mortality [16–18]. Prednisone and methotrexate use were found to be significant predictors of hospitalization for pneumonia. Frequent infections (more than 1 episode) are reported in patients with ANCA-associated vasculitis during the first year of diagnosis and initiation of therapy [18]. We assume that bronchiectasis complicated by bacterial colonization and bronchial obstruction in combination with immuno-suppression led to severe pneumonia and death of the patient.
Conclusions
The present case report demonstrates a long-term follow-up of a patient with EGPA. Both common and rare manifestations of EGPA have been described in the patient, such as central nervous system damage and bronchiectasis. Immunosuppressive treatment probably contributed to Klebsiella infection development. Thus, long-term complications of EGPA in combination with adverse effects of medications could have caused the severe pneumonia and death of the patient.
Figure 1. Brain multi-spiral computed tomography performed in 2016 showing ischemic brain damage in the right occipital lobe.
Figure 2. (A–C) Chest computed tomography of patient showing large left upper lobe pneumonia and bronchiectasis in lover lobes.
Figure 3. Timeline from first symptoms to death. | 82 kg. | Weight | CC BY-NC-ND | 33582702 | 18,962,204 | 2021-02-14 |
What was the administration route of drug 'AMIKACIN'? | Case Report: A 64-Year-Old Man with 10-Year History of Eosinophilic Granulomatosis with Polyangiitis with Bronchiectasis and Severe Klebsiella pneumonia.
BACKGROUND Eosinophilic granulomatosis with polyangiitis (EGPA) is a rare and severe progressive disease with multiple clinical manifestations and organ damage. Usually, it requires long-term monitoring of the state of many organs due to the gradual character of its manifestations. CASE REPORT We report a case of a long-term follow-up of a patient with eosinophilic granulomatosis with polyangiitis with emphasis on specific clinical features in this patient. A 64-year-old man was being followed up for 10 years. The initial diagnosis was allergic bronchial asthma; however, as new clinical signs and symptoms developed, the diagnosis of EGPA became obvious. A positive treatment response was seen, mainly manifested as reduced polyneuropathy. Meanwhile, bronchial asthma remained uncontrolled and bronchiectasis and Klebsiella pneumoniae colonization developed despite the combination treatment with prednisolone and methotrexate. Furthermore, the patient suffered a cerebral ischemic infarction. During the last hospital admission, severe uncontrolled bronchial asthma complicated with pneumonia resulted in the patient's death. CONCLUSIONS This clinical case shows the gradual development of EGPA with multiple-organ involvement, including respiratory manifestations and peripheral and central nervous system damage. Immunosuppressive treatment combined with complications of EGPA could have contributed to severe pneumonia development and death of the patient.
Background
Systemic vasculitides are difficult to diagnose due to the heterogenous clinical manifestations requiring a wide differential diagnostic search. One of the most uncommon vasculitis types is eosinophilic granulomatosis with polyangiitis (EGPA), or Churg-Strauss syndrome [1,2]. This vasculitis is associated with the presence of antineutrophilic cytoplasmic antibodies (ANCA) and is characterized by necrotizing inflammatory lesions of mainly small vessels, as well as eosinophilic and granulomatous inflammation in various organs. Bronchial asthma and eosinophilia are the most common clinical manifestations of EGPA [1–3]. In 2012, according to the current classification of systemic vasculitides, Churg-Strauss syndrome was renamed “Eosinophilic Granulomatosis with Polyangiitis” (EGPA) and classified into the ANCA-associated vasculitides group [3].
EGPA is a rare ANCA-associated vasculitis. The incidence rate is 0.5–6.8 new cases/year per million populations in Europe [2]. Prevalence is higher in northern latitudes compared to southern Europe [4]. The disease occurs most commonly in people 40–60 years of age [2,5]. No sex predominance has been clearly demonstrated [4].
The pathogenesis of the disease is still not completely clear, but the key role is attributed to autoimmune inflammation with the participation of eosinophils, CD8+, and CD4+lymphocytes (predominantly Th-2) and their proinflammatory cytokines: interleukin (IL)4, IL5, and IL13 [2,5]. Elevation of immunoglobulin (Ig)E, IgG4, and eotaxin-3 is common in EGPA patients [2,5]. The frequent (in 30–40% of cases) detection of ANCA (mainly antibodies to myeloperoxidase) also suggests the autoimmune nature of the disease. ANCA-positive patients are more commonly diagnosed with glomerulonephritis and mononeuritis, whereas the ANCA-negative ones more frequently demonstrate endocardial, myocardial, and pulmonary eosinophilic infiltrates [1,2,5,6].
There are 3 phases in the development of EGPA (these phases partially overlap and may not appear in sequence):
the allergic phase, distinguished by allergic upper respiratory tract inflammation (rhinosinusitis, allergic rhinitis) and bronchial asthma;
the eosinophilic phase, manifested with severe eosinophilia in combination with multiple eosinophilic infiltrates in various organs (most often in the lungs and gastrointestinal tract);
the vasculitis phase, with small- and medium-vessel inflammation of different organs, which can often be life-threatening [5,6].
Respiratory manifestations of EGPA include allergic rhinitis, sinusitis, nasal polyposis and, in more than 95% of patients, poorly controlled bronchial asthma [5]. Chest radiography findings include transient and migratory opacities, ground-glass opacity, focal consolidation, and small peribronchial and centrilobular nodules or masses [7]. Computed tomography reveals bronchial abnormalities, most commonly thickened wall and dilatation with secondary bronchiectasis formation, leading to frequent infections [7,8]. Pleural effusions and alveolar hemorrhages are uncommon manifestations of EGPA [1,5,7,8].
A characteristic neurological manifestation seen in 75% of patients with EGPA is peripheral neuropathy (mononeuritis multiplex) [1,5]. Upper-limb symptoms due to neuropathy, as well as vasculitis in epineural vessels, were more frequently reported in the ANCA-positive group [9]. Eosinophil-associated tissue damage and vascular occlusion leading to ischemia are more specific in ANCA-negative patients [9]. Also, the tissue damage caused by eosinophils may be associated with IL-5 concentration [10]. Brain hemorrhage, subarachnoid hemorrhage, and ischemic brain infarction are rare [1,5,6].
The main drugs for the treatment of EGPA are glucocorticoids. In organ- or life-threatening manifestations, the recommended starting dose of prednisone is 1 mg/kg/day with a maximal daily dose of 80 mg for 2–3 weeks, with tapering to a minimal supporting dose selected individually to control the symptoms and minimize the adverse effects of treatment. Methylprednisolone (7.5–15 mg/kg/day) can be used as a pulse therapy for life-threatening manifestations [11,12]. Most patients need prolonged or lifelong steroids. In the case of organ- or life-threatening manifestations, glucocorticoids in combination with cyclophosphamide are prescribed [11,12]. The combination of glucocorticoids and methotrexate or mycophenolate is required to induce remission in mild or moderate disease [11,12]. Mepolizumab (a humanized monoclonal antibody targeting IL-5) has been used for EGPA treatment [11]. For ANCA-positive patients with renal involvement or refractory disease, rituximab can be considered [3, 11].
Due to the rare occurrence of EGPA and the variety of its clinical manifestations, the diagnosis and treatment of this disease can be challenging. Currently, reports of long-term observations with the detailed patient follow-up are of great interest. The aim of our case report is to present rare manifestations of EGPA, such as bronchiectasis and stroke. We also focused on the infectious complications of immunosuppressive treatments. Thus, we would like to present a 10-year observation of a patient with EGPA.
Case Report
The male patient, born in 1953, was first hospitalized in the Pulmonary Department of our clinic in 2007 due to exacerbated asthma. The patient’s history was remarkable for multiple allergies (to house dust, cat and dog epidermis, and plant pollen) as well as nonsteroidal anti-inflammatory drugs intolerance (manifested as bronchospasm). The patient denied unhealthy habits and occupational hazards (he worked as a driver). His family history was not remarkable. The patient has had allergic rhinitis since adolescence. The first asthma symptoms appeared at the age of 18 while serving in the army. At the age of 22, an allergist diagnosed him with bronchial asthma. The patient had annual exacerbations (1–2 times a year) due to cold or allergen exposure, and acute respiratory viral infections. At the age of 42, polypoid rhinosinusitis was found, and a polypectomy was performed. Since 2006, the patient had been on controller treatment with salmeterol/fluticasone inhaler (25/250 mg, respectively) 2 doses 2 times a day.
In late 2009, extremely elevated eosinophils (53%) and total IgE (1000 IU/L) were found for the first time. Also, since 2009, the patient has had transient lung opacities, which usually disappeared after treatment with prednisolone 30 mg per day.
In August 2010, the patient noticed numbness on the back surface of the left foot, which spread to the entire left lower leg in 1 month, as well as numbness of the right foot and lower leg. Later, the patient started having tingling, twisting, and burning sensations aggravated by touching in the feet and lower legs. Also, the weakness in the distal parts of the legs developed, and he started having difficulties moving around on his own and falling while walking, and he was subsequently hospitalized in our clinic due to exacerbated asthma. On laboratory testing, high eosinophils (53%) and gamma-globulins (27.9%) were found in November 2010. The chest radiography showed emphysema and consolidation in the middle lobe of the right lung. A Neurology consult confirmed asymmetrical distal sensory-motor polyneuropathy demonstrated by lower distal asymmetrical paraparesis with decreased muscle strength in the foot flexors and extensors, more severe on the left side, low tone of the shin muscles, hypoactive Achilles reflexes, and reduced leg sensitivity starting from the middle of the shin and below, more severe in the peroneal nerve inner-vation area. The patient also demonstrated abnormal coordination (heel-to-knee, Romberg) test results and bilateral steppage gait with knee overextension. Electroneuromyography showed axonal lesion of all distal leg nerves and secondary myelinopathy. Thus, EGPA was suspected. The ANCA (anti-myeloperoxidase) titer was 30 U/ml (normal reference below 20 U/ml).
Allergic bronchopulmonary aspergillosis and other causes of hypereosinophilia were consistently ruled out. IgE and IgG for Aspergillus spp. and Aspergillus spp. in sputum were not found.
Thus, the presence of severe bronchial asthma, rhinosinusitis, high and persistent eosinophilia, transient pulmonary opacities, and polyneuropathy suggested EGPA (5 out of 6 clinical criteria of American College of Rheumatology present). Based on the clinical presentation and increased ANCA titer, the patient was diagnosed with EGPA. Systemic steroid treatment (prednisolone 300 mg intravenously and 30 mg orally) was prescribed, and 5 plasmapheresis sessions were performed. Later, prolonged combination immunosuppression (prednisolone 10 mg/day and methotrexate 25 mg/week) and controller asthma treatment (fluticasone propionate/salmeterol 250/25 µg 4 doses/ day, tiotropium bromide 18 µg per day) were prescribed. After 1 month on prednisolone and methotrexate, there was a significant neurological improvement: the motor and sensory disturbances decreased, and the patient was able to move around on his own. Further on, polyneuropathy completely resolved on the combination treatment. His asthma remained uncontrolled despite the combination treatment: he had frequent (3 times a year) severe exacerbations requiring hospitalization. In 2015, newly-emerged copious purulent sputum associated with frequent infection-related asthma exacerbations was noted. In January 2016, a patient underwent chest computed tomography (CT), which showed generalized bronchiectasis. Stenosis of the proximal bronchi was ruled out also by fibrobronchos-copy. The bronchial secretion microbiology revealed Klebsiella pneumoniae growth (107 colony-forming units).
In November 2016, during the asthma exacerbation, the patient suffered a cerebral ischemic infarction involving the right occipital lobe, confirmed by CT (Figures 1, 2).
On March 2, 2017, the patient was admitted to our hospital due to severe respiratory failure caused by pneumonia in the left upper lobe and severe asthma exacerbation.
The complete blood count showed high WBCs (15.2×109/l) with left shift. The IgE level was 144.0 IU/l, C-reactive protein was 48.8 mg/dL. Clinical chemistry was remarkable for urea nitrogen 18.4 mmol/l and creatinine 1.55 mg/dL. The sputum microscopy demonstrated 100 WBCs and 160 RBCs per vision field, and no eosinophils were revealed. A chest CT revealed multiple bronchiectasis and upper lobar pneumonia of the left lung (Figure 2). Klebsiella was again detected in the sputum culture.
The patient was prescribed intravenous antibiotics (ceftriaxone 2 g/day and ciprofloxacin 800 mg/day replaced by cefopera-zone 4 g/day and amikacin 1 g/day after 3 days due to low efficacy), bronchodilator and anti-inflammatory therapy (nebulized ipratropium bromide, fenoterol, and budesonide 2000 µg per day, prednisolone intravenously 150 mg and orally 30 mg). Despite the aggressive treatment, the respiratory failure progressed, and on day 6 of hospitalization the patient was transferred to the Intensive Care Unit and intubated. On day 7, the patient died. On autopsy, the EGPA diagnosis was confirmed, signs of ischemic infarction in the occipital lobe of the brain were found, and left upper lobe pneumonia with abscess formation and generalized bilateral bronchiectases were revealed.
Discussion
Based on the long follow-up period and careful interpretation of signs and symptoms, typical laboratory abnormalities, the diagnosis was clear. Among the most commonly used diagnostic criteria of EGPA are the American College of Rheumatology (ACR) classification criteria, Lanham’s criteria, and the Chapel Hill Consensus Conference criteria. The presence of 4 or more ACR classification criteria, including bronchial asthma, eosinophilia (more than 10%), paranasal sinus abnormalities, migratory or transient eosinophilic pulmonary infiltrates, extravascular eosinophil infiltration on biopsy, and mono- or polyneuropathy, has a sensitivity of 85% and a specificity of 100% [2,5,6]. We identified 5 out of 6 American College of Rheumatology clinical criteria present in our patient.
This clinical case shows gradual development of EGPA with multiple-organ involvement (see Figure 3). The disease onset was characterized by respiratory manifestations and suspicion of EGPA appeared only after the development of polyneuropathy (after 39 years) with hypereosinophilia. Polyneuropathy was noted at the age of 57, which is slightly later than the average age of onset reported in the literature [13]. Notably, the patient was ANCA-positive and his long-term neurological manifestation (mononeuritis) was quite typical for ANCA vasculitis [5,6,8,13]. The diagnosis became obvious after comprehensive assessment of the disease history with its gradually evolving signs and symptoms and laboratory abnormalities. Almost all manifestations of the disease seen in the patient were typical except for stroke, which is an exceptional feature for this case [1,5,6]. Neurological involvement manifested as ischemic stroke is an uncommon complication of EGPA (seen only in 5.2% of all patients) [1]. Thus, the patient demonstrated both peripheral and central nervous system involvement, which is unusual and life-threatening in EGPA.
This clinical case of EGPA also attracted our attention due to severe respiratory manifestations. While typical respiratory manifestations such as rhinosinusitis, asthma, and transient lung infiltrates are well documented in the literature, EGPA-associated bronchiectasis is rarely mentioned [14]. Bronchiectasis in the patient was detected by CT at the age of 63. The patient was a nonsmoker, and no other cause of bronchial abnormalities and airflow obstruction could be identified. Stenosis of the proximal bronchi was ruled out by fibrobronchoscopy and CT. The diagnosis of allergic bronchopulmonary aspergillosis was not confirmed. Klebsiella pneumoniae colonization was found during the last 2 years of the patient’s life. Klebsiella pneumoniae is a typical pathogen causing bronchiectasis [15]. It should be noted that aspergillosis and mycobacteriosis are common complications of bronchiectasis [15], but neither was identified in our patient.
The patient was treated for a long time with a combination of prednisolone and methotrexate. There was a significant positive effect on polyneuropathy, but uncontrolled bronchial asthma persisted. This treatment regimen complies with current recommendations [3,11]. Patients with life-threatening and/or organ-threatening manifestations (including lesions of the central nervous system and severe peripheral neuropathy) require therapy with a combination of a glucocorticoid and a cytostatic [3,11].
The use of immunosuppressive drugs in patients with ANCA-associated vasculitis significantly increases the risk of infection and mortality [16–18]. Prednisone and methotrexate use were found to be significant predictors of hospitalization for pneumonia. Frequent infections (more than 1 episode) are reported in patients with ANCA-associated vasculitis during the first year of diagnosis and initiation of therapy [18]. We assume that bronchiectasis complicated by bacterial colonization and bronchial obstruction in combination with immuno-suppression led to severe pneumonia and death of the patient.
Conclusions
The present case report demonstrates a long-term follow-up of a patient with EGPA. Both common and rare manifestations of EGPA have been described in the patient, such as central nervous system damage and bronchiectasis. Immunosuppressive treatment probably contributed to Klebsiella infection development. Thus, long-term complications of EGPA in combination with adverse effects of medications could have caused the severe pneumonia and death of the patient.
Figure 1. Brain multi-spiral computed tomography performed in 2016 showing ischemic brain damage in the right occipital lobe.
Figure 2. (A–C) Chest computed tomography of patient showing large left upper lobe pneumonia and bronchiectasis in lover lobes.
Figure 3. Timeline from first symptoms to death. | Intravenous (not otherwise specified) | DrugAdministrationRoute | CC BY-NC-ND | 33582702 | 18,986,567 | 2021-02-14 |
What was the administration route of drug 'BUDESONIDE'? | Case Report: A 64-Year-Old Man with 10-Year History of Eosinophilic Granulomatosis with Polyangiitis with Bronchiectasis and Severe Klebsiella pneumonia.
BACKGROUND Eosinophilic granulomatosis with polyangiitis (EGPA) is a rare and severe progressive disease with multiple clinical manifestations and organ damage. Usually, it requires long-term monitoring of the state of many organs due to the gradual character of its manifestations. CASE REPORT We report a case of a long-term follow-up of a patient with eosinophilic granulomatosis with polyangiitis with emphasis on specific clinical features in this patient. A 64-year-old man was being followed up for 10 years. The initial diagnosis was allergic bronchial asthma; however, as new clinical signs and symptoms developed, the diagnosis of EGPA became obvious. A positive treatment response was seen, mainly manifested as reduced polyneuropathy. Meanwhile, bronchial asthma remained uncontrolled and bronchiectasis and Klebsiella pneumoniae colonization developed despite the combination treatment with prednisolone and methotrexate. Furthermore, the patient suffered a cerebral ischemic infarction. During the last hospital admission, severe uncontrolled bronchial asthma complicated with pneumonia resulted in the patient's death. CONCLUSIONS This clinical case shows the gradual development of EGPA with multiple-organ involvement, including respiratory manifestations and peripheral and central nervous system damage. Immunosuppressive treatment combined with complications of EGPA could have contributed to severe pneumonia development and death of the patient.
Background
Systemic vasculitides are difficult to diagnose due to the heterogenous clinical manifestations requiring a wide differential diagnostic search. One of the most uncommon vasculitis types is eosinophilic granulomatosis with polyangiitis (EGPA), or Churg-Strauss syndrome [1,2]. This vasculitis is associated with the presence of antineutrophilic cytoplasmic antibodies (ANCA) and is characterized by necrotizing inflammatory lesions of mainly small vessels, as well as eosinophilic and granulomatous inflammation in various organs. Bronchial asthma and eosinophilia are the most common clinical manifestations of EGPA [1–3]. In 2012, according to the current classification of systemic vasculitides, Churg-Strauss syndrome was renamed “Eosinophilic Granulomatosis with Polyangiitis” (EGPA) and classified into the ANCA-associated vasculitides group [3].
EGPA is a rare ANCA-associated vasculitis. The incidence rate is 0.5–6.8 new cases/year per million populations in Europe [2]. Prevalence is higher in northern latitudes compared to southern Europe [4]. The disease occurs most commonly in people 40–60 years of age [2,5]. No sex predominance has been clearly demonstrated [4].
The pathogenesis of the disease is still not completely clear, but the key role is attributed to autoimmune inflammation with the participation of eosinophils, CD8+, and CD4+lymphocytes (predominantly Th-2) and their proinflammatory cytokines: interleukin (IL)4, IL5, and IL13 [2,5]. Elevation of immunoglobulin (Ig)E, IgG4, and eotaxin-3 is common in EGPA patients [2,5]. The frequent (in 30–40% of cases) detection of ANCA (mainly antibodies to myeloperoxidase) also suggests the autoimmune nature of the disease. ANCA-positive patients are more commonly diagnosed with glomerulonephritis and mononeuritis, whereas the ANCA-negative ones more frequently demonstrate endocardial, myocardial, and pulmonary eosinophilic infiltrates [1,2,5,6].
There are 3 phases in the development of EGPA (these phases partially overlap and may not appear in sequence):
the allergic phase, distinguished by allergic upper respiratory tract inflammation (rhinosinusitis, allergic rhinitis) and bronchial asthma;
the eosinophilic phase, manifested with severe eosinophilia in combination with multiple eosinophilic infiltrates in various organs (most often in the lungs and gastrointestinal tract);
the vasculitis phase, with small- and medium-vessel inflammation of different organs, which can often be life-threatening [5,6].
Respiratory manifestations of EGPA include allergic rhinitis, sinusitis, nasal polyposis and, in more than 95% of patients, poorly controlled bronchial asthma [5]. Chest radiography findings include transient and migratory opacities, ground-glass opacity, focal consolidation, and small peribronchial and centrilobular nodules or masses [7]. Computed tomography reveals bronchial abnormalities, most commonly thickened wall and dilatation with secondary bronchiectasis formation, leading to frequent infections [7,8]. Pleural effusions and alveolar hemorrhages are uncommon manifestations of EGPA [1,5,7,8].
A characteristic neurological manifestation seen in 75% of patients with EGPA is peripheral neuropathy (mononeuritis multiplex) [1,5]. Upper-limb symptoms due to neuropathy, as well as vasculitis in epineural vessels, were more frequently reported in the ANCA-positive group [9]. Eosinophil-associated tissue damage and vascular occlusion leading to ischemia are more specific in ANCA-negative patients [9]. Also, the tissue damage caused by eosinophils may be associated with IL-5 concentration [10]. Brain hemorrhage, subarachnoid hemorrhage, and ischemic brain infarction are rare [1,5,6].
The main drugs for the treatment of EGPA are glucocorticoids. In organ- or life-threatening manifestations, the recommended starting dose of prednisone is 1 mg/kg/day with a maximal daily dose of 80 mg for 2–3 weeks, with tapering to a minimal supporting dose selected individually to control the symptoms and minimize the adverse effects of treatment. Methylprednisolone (7.5–15 mg/kg/day) can be used as a pulse therapy for life-threatening manifestations [11,12]. Most patients need prolonged or lifelong steroids. In the case of organ- or life-threatening manifestations, glucocorticoids in combination with cyclophosphamide are prescribed [11,12]. The combination of glucocorticoids and methotrexate or mycophenolate is required to induce remission in mild or moderate disease [11,12]. Mepolizumab (a humanized monoclonal antibody targeting IL-5) has been used for EGPA treatment [11]. For ANCA-positive patients with renal involvement or refractory disease, rituximab can be considered [3, 11].
Due to the rare occurrence of EGPA and the variety of its clinical manifestations, the diagnosis and treatment of this disease can be challenging. Currently, reports of long-term observations with the detailed patient follow-up are of great interest. The aim of our case report is to present rare manifestations of EGPA, such as bronchiectasis and stroke. We also focused on the infectious complications of immunosuppressive treatments. Thus, we would like to present a 10-year observation of a patient with EGPA.
Case Report
The male patient, born in 1953, was first hospitalized in the Pulmonary Department of our clinic in 2007 due to exacerbated asthma. The patient’s history was remarkable for multiple allergies (to house dust, cat and dog epidermis, and plant pollen) as well as nonsteroidal anti-inflammatory drugs intolerance (manifested as bronchospasm). The patient denied unhealthy habits and occupational hazards (he worked as a driver). His family history was not remarkable. The patient has had allergic rhinitis since adolescence. The first asthma symptoms appeared at the age of 18 while serving in the army. At the age of 22, an allergist diagnosed him with bronchial asthma. The patient had annual exacerbations (1–2 times a year) due to cold or allergen exposure, and acute respiratory viral infections. At the age of 42, polypoid rhinosinusitis was found, and a polypectomy was performed. Since 2006, the patient had been on controller treatment with salmeterol/fluticasone inhaler (25/250 mg, respectively) 2 doses 2 times a day.
In late 2009, extremely elevated eosinophils (53%) and total IgE (1000 IU/L) were found for the first time. Also, since 2009, the patient has had transient lung opacities, which usually disappeared after treatment with prednisolone 30 mg per day.
In August 2010, the patient noticed numbness on the back surface of the left foot, which spread to the entire left lower leg in 1 month, as well as numbness of the right foot and lower leg. Later, the patient started having tingling, twisting, and burning sensations aggravated by touching in the feet and lower legs. Also, the weakness in the distal parts of the legs developed, and he started having difficulties moving around on his own and falling while walking, and he was subsequently hospitalized in our clinic due to exacerbated asthma. On laboratory testing, high eosinophils (53%) and gamma-globulins (27.9%) were found in November 2010. The chest radiography showed emphysema and consolidation in the middle lobe of the right lung. A Neurology consult confirmed asymmetrical distal sensory-motor polyneuropathy demonstrated by lower distal asymmetrical paraparesis with decreased muscle strength in the foot flexors and extensors, more severe on the left side, low tone of the shin muscles, hypoactive Achilles reflexes, and reduced leg sensitivity starting from the middle of the shin and below, more severe in the peroneal nerve inner-vation area. The patient also demonstrated abnormal coordination (heel-to-knee, Romberg) test results and bilateral steppage gait with knee overextension. Electroneuromyography showed axonal lesion of all distal leg nerves and secondary myelinopathy. Thus, EGPA was suspected. The ANCA (anti-myeloperoxidase) titer was 30 U/ml (normal reference below 20 U/ml).
Allergic bronchopulmonary aspergillosis and other causes of hypereosinophilia were consistently ruled out. IgE and IgG for Aspergillus spp. and Aspergillus spp. in sputum were not found.
Thus, the presence of severe bronchial asthma, rhinosinusitis, high and persistent eosinophilia, transient pulmonary opacities, and polyneuropathy suggested EGPA (5 out of 6 clinical criteria of American College of Rheumatology present). Based on the clinical presentation and increased ANCA titer, the patient was diagnosed with EGPA. Systemic steroid treatment (prednisolone 300 mg intravenously and 30 mg orally) was prescribed, and 5 plasmapheresis sessions were performed. Later, prolonged combination immunosuppression (prednisolone 10 mg/day and methotrexate 25 mg/week) and controller asthma treatment (fluticasone propionate/salmeterol 250/25 µg 4 doses/ day, tiotropium bromide 18 µg per day) were prescribed. After 1 month on prednisolone and methotrexate, there was a significant neurological improvement: the motor and sensory disturbances decreased, and the patient was able to move around on his own. Further on, polyneuropathy completely resolved on the combination treatment. His asthma remained uncontrolled despite the combination treatment: he had frequent (3 times a year) severe exacerbations requiring hospitalization. In 2015, newly-emerged copious purulent sputum associated with frequent infection-related asthma exacerbations was noted. In January 2016, a patient underwent chest computed tomography (CT), which showed generalized bronchiectasis. Stenosis of the proximal bronchi was ruled out also by fibrobronchos-copy. The bronchial secretion microbiology revealed Klebsiella pneumoniae growth (107 colony-forming units).
In November 2016, during the asthma exacerbation, the patient suffered a cerebral ischemic infarction involving the right occipital lobe, confirmed by CT (Figures 1, 2).
On March 2, 2017, the patient was admitted to our hospital due to severe respiratory failure caused by pneumonia in the left upper lobe and severe asthma exacerbation.
The complete blood count showed high WBCs (15.2×109/l) with left shift. The IgE level was 144.0 IU/l, C-reactive protein was 48.8 mg/dL. Clinical chemistry was remarkable for urea nitrogen 18.4 mmol/l and creatinine 1.55 mg/dL. The sputum microscopy demonstrated 100 WBCs and 160 RBCs per vision field, and no eosinophils were revealed. A chest CT revealed multiple bronchiectasis and upper lobar pneumonia of the left lung (Figure 2). Klebsiella was again detected in the sputum culture.
The patient was prescribed intravenous antibiotics (ceftriaxone 2 g/day and ciprofloxacin 800 mg/day replaced by cefopera-zone 4 g/day and amikacin 1 g/day after 3 days due to low efficacy), bronchodilator and anti-inflammatory therapy (nebulized ipratropium bromide, fenoterol, and budesonide 2000 µg per day, prednisolone intravenously 150 mg and orally 30 mg). Despite the aggressive treatment, the respiratory failure progressed, and on day 6 of hospitalization the patient was transferred to the Intensive Care Unit and intubated. On day 7, the patient died. On autopsy, the EGPA diagnosis was confirmed, signs of ischemic infarction in the occipital lobe of the brain were found, and left upper lobe pneumonia with abscess formation and generalized bilateral bronchiectases were revealed.
Discussion
Based on the long follow-up period and careful interpretation of signs and symptoms, typical laboratory abnormalities, the diagnosis was clear. Among the most commonly used diagnostic criteria of EGPA are the American College of Rheumatology (ACR) classification criteria, Lanham’s criteria, and the Chapel Hill Consensus Conference criteria. The presence of 4 or more ACR classification criteria, including bronchial asthma, eosinophilia (more than 10%), paranasal sinus abnormalities, migratory or transient eosinophilic pulmonary infiltrates, extravascular eosinophil infiltration on biopsy, and mono- or polyneuropathy, has a sensitivity of 85% and a specificity of 100% [2,5,6]. We identified 5 out of 6 American College of Rheumatology clinical criteria present in our patient.
This clinical case shows gradual development of EGPA with multiple-organ involvement (see Figure 3). The disease onset was characterized by respiratory manifestations and suspicion of EGPA appeared only after the development of polyneuropathy (after 39 years) with hypereosinophilia. Polyneuropathy was noted at the age of 57, which is slightly later than the average age of onset reported in the literature [13]. Notably, the patient was ANCA-positive and his long-term neurological manifestation (mononeuritis) was quite typical for ANCA vasculitis [5,6,8,13]. The diagnosis became obvious after comprehensive assessment of the disease history with its gradually evolving signs and symptoms and laboratory abnormalities. Almost all manifestations of the disease seen in the patient were typical except for stroke, which is an exceptional feature for this case [1,5,6]. Neurological involvement manifested as ischemic stroke is an uncommon complication of EGPA (seen only in 5.2% of all patients) [1]. Thus, the patient demonstrated both peripheral and central nervous system involvement, which is unusual and life-threatening in EGPA.
This clinical case of EGPA also attracted our attention due to severe respiratory manifestations. While typical respiratory manifestations such as rhinosinusitis, asthma, and transient lung infiltrates are well documented in the literature, EGPA-associated bronchiectasis is rarely mentioned [14]. Bronchiectasis in the patient was detected by CT at the age of 63. The patient was a nonsmoker, and no other cause of bronchial abnormalities and airflow obstruction could be identified. Stenosis of the proximal bronchi was ruled out by fibrobronchoscopy and CT. The diagnosis of allergic bronchopulmonary aspergillosis was not confirmed. Klebsiella pneumoniae colonization was found during the last 2 years of the patient’s life. Klebsiella pneumoniae is a typical pathogen causing bronchiectasis [15]. It should be noted that aspergillosis and mycobacteriosis are common complications of bronchiectasis [15], but neither was identified in our patient.
The patient was treated for a long time with a combination of prednisolone and methotrexate. There was a significant positive effect on polyneuropathy, but uncontrolled bronchial asthma persisted. This treatment regimen complies with current recommendations [3,11]. Patients with life-threatening and/or organ-threatening manifestations (including lesions of the central nervous system and severe peripheral neuropathy) require therapy with a combination of a glucocorticoid and a cytostatic [3,11].
The use of immunosuppressive drugs in patients with ANCA-associated vasculitis significantly increases the risk of infection and mortality [16–18]. Prednisone and methotrexate use were found to be significant predictors of hospitalization for pneumonia. Frequent infections (more than 1 episode) are reported in patients with ANCA-associated vasculitis during the first year of diagnosis and initiation of therapy [18]. We assume that bronchiectasis complicated by bacterial colonization and bronchial obstruction in combination with immuno-suppression led to severe pneumonia and death of the patient.
Conclusions
The present case report demonstrates a long-term follow-up of a patient with EGPA. Both common and rare manifestations of EGPA have been described in the patient, such as central nervous system damage and bronchiectasis. Immunosuppressive treatment probably contributed to Klebsiella infection development. Thus, long-term complications of EGPA in combination with adverse effects of medications could have caused the severe pneumonia and death of the patient.
Figure 1. Brain multi-spiral computed tomography performed in 2016 showing ischemic brain damage in the right occipital lobe.
Figure 2. (A–C) Chest computed tomography of patient showing large left upper lobe pneumonia and bronchiectasis in lover lobes.
Figure 3. Timeline from first symptoms to death. | Intravenous (not otherwise specified) | DrugAdministrationRoute | CC BY-NC-ND | 33582702 | 19,021,629 | 2021-02-14 |
What was the administration route of drug 'CEFOPERAZONE'? | Case Report: A 64-Year-Old Man with 10-Year History of Eosinophilic Granulomatosis with Polyangiitis with Bronchiectasis and Severe Klebsiella pneumonia.
BACKGROUND Eosinophilic granulomatosis with polyangiitis (EGPA) is a rare and severe progressive disease with multiple clinical manifestations and organ damage. Usually, it requires long-term monitoring of the state of many organs due to the gradual character of its manifestations. CASE REPORT We report a case of a long-term follow-up of a patient with eosinophilic granulomatosis with polyangiitis with emphasis on specific clinical features in this patient. A 64-year-old man was being followed up for 10 years. The initial diagnosis was allergic bronchial asthma; however, as new clinical signs and symptoms developed, the diagnosis of EGPA became obvious. A positive treatment response was seen, mainly manifested as reduced polyneuropathy. Meanwhile, bronchial asthma remained uncontrolled and bronchiectasis and Klebsiella pneumoniae colonization developed despite the combination treatment with prednisolone and methotrexate. Furthermore, the patient suffered a cerebral ischemic infarction. During the last hospital admission, severe uncontrolled bronchial asthma complicated with pneumonia resulted in the patient's death. CONCLUSIONS This clinical case shows the gradual development of EGPA with multiple-organ involvement, including respiratory manifestations and peripheral and central nervous system damage. Immunosuppressive treatment combined with complications of EGPA could have contributed to severe pneumonia development and death of the patient.
Background
Systemic vasculitides are difficult to diagnose due to the heterogenous clinical manifestations requiring a wide differential diagnostic search. One of the most uncommon vasculitis types is eosinophilic granulomatosis with polyangiitis (EGPA), or Churg-Strauss syndrome [1,2]. This vasculitis is associated with the presence of antineutrophilic cytoplasmic antibodies (ANCA) and is characterized by necrotizing inflammatory lesions of mainly small vessels, as well as eosinophilic and granulomatous inflammation in various organs. Bronchial asthma and eosinophilia are the most common clinical manifestations of EGPA [1–3]. In 2012, according to the current classification of systemic vasculitides, Churg-Strauss syndrome was renamed “Eosinophilic Granulomatosis with Polyangiitis” (EGPA) and classified into the ANCA-associated vasculitides group [3].
EGPA is a rare ANCA-associated vasculitis. The incidence rate is 0.5–6.8 new cases/year per million populations in Europe [2]. Prevalence is higher in northern latitudes compared to southern Europe [4]. The disease occurs most commonly in people 40–60 years of age [2,5]. No sex predominance has been clearly demonstrated [4].
The pathogenesis of the disease is still not completely clear, but the key role is attributed to autoimmune inflammation with the participation of eosinophils, CD8+, and CD4+lymphocytes (predominantly Th-2) and their proinflammatory cytokines: interleukin (IL)4, IL5, and IL13 [2,5]. Elevation of immunoglobulin (Ig)E, IgG4, and eotaxin-3 is common in EGPA patients [2,5]. The frequent (in 30–40% of cases) detection of ANCA (mainly antibodies to myeloperoxidase) also suggests the autoimmune nature of the disease. ANCA-positive patients are more commonly diagnosed with glomerulonephritis and mononeuritis, whereas the ANCA-negative ones more frequently demonstrate endocardial, myocardial, and pulmonary eosinophilic infiltrates [1,2,5,6].
There are 3 phases in the development of EGPA (these phases partially overlap and may not appear in sequence):
the allergic phase, distinguished by allergic upper respiratory tract inflammation (rhinosinusitis, allergic rhinitis) and bronchial asthma;
the eosinophilic phase, manifested with severe eosinophilia in combination with multiple eosinophilic infiltrates in various organs (most often in the lungs and gastrointestinal tract);
the vasculitis phase, with small- and medium-vessel inflammation of different organs, which can often be life-threatening [5,6].
Respiratory manifestations of EGPA include allergic rhinitis, sinusitis, nasal polyposis and, in more than 95% of patients, poorly controlled bronchial asthma [5]. Chest radiography findings include transient and migratory opacities, ground-glass opacity, focal consolidation, and small peribronchial and centrilobular nodules or masses [7]. Computed tomography reveals bronchial abnormalities, most commonly thickened wall and dilatation with secondary bronchiectasis formation, leading to frequent infections [7,8]. Pleural effusions and alveolar hemorrhages are uncommon manifestations of EGPA [1,5,7,8].
A characteristic neurological manifestation seen in 75% of patients with EGPA is peripheral neuropathy (mononeuritis multiplex) [1,5]. Upper-limb symptoms due to neuropathy, as well as vasculitis in epineural vessels, were more frequently reported in the ANCA-positive group [9]. Eosinophil-associated tissue damage and vascular occlusion leading to ischemia are more specific in ANCA-negative patients [9]. Also, the tissue damage caused by eosinophils may be associated with IL-5 concentration [10]. Brain hemorrhage, subarachnoid hemorrhage, and ischemic brain infarction are rare [1,5,6].
The main drugs for the treatment of EGPA are glucocorticoids. In organ- or life-threatening manifestations, the recommended starting dose of prednisone is 1 mg/kg/day with a maximal daily dose of 80 mg for 2–3 weeks, with tapering to a minimal supporting dose selected individually to control the symptoms and minimize the adverse effects of treatment. Methylprednisolone (7.5–15 mg/kg/day) can be used as a pulse therapy for life-threatening manifestations [11,12]. Most patients need prolonged or lifelong steroids. In the case of organ- or life-threatening manifestations, glucocorticoids in combination with cyclophosphamide are prescribed [11,12]. The combination of glucocorticoids and methotrexate or mycophenolate is required to induce remission in mild or moderate disease [11,12]. Mepolizumab (a humanized monoclonal antibody targeting IL-5) has been used for EGPA treatment [11]. For ANCA-positive patients with renal involvement or refractory disease, rituximab can be considered [3, 11].
Due to the rare occurrence of EGPA and the variety of its clinical manifestations, the diagnosis and treatment of this disease can be challenging. Currently, reports of long-term observations with the detailed patient follow-up are of great interest. The aim of our case report is to present rare manifestations of EGPA, such as bronchiectasis and stroke. We also focused on the infectious complications of immunosuppressive treatments. Thus, we would like to present a 10-year observation of a patient with EGPA.
Case Report
The male patient, born in 1953, was first hospitalized in the Pulmonary Department of our clinic in 2007 due to exacerbated asthma. The patient’s history was remarkable for multiple allergies (to house dust, cat and dog epidermis, and plant pollen) as well as nonsteroidal anti-inflammatory drugs intolerance (manifested as bronchospasm). The patient denied unhealthy habits and occupational hazards (he worked as a driver). His family history was not remarkable. The patient has had allergic rhinitis since adolescence. The first asthma symptoms appeared at the age of 18 while serving in the army. At the age of 22, an allergist diagnosed him with bronchial asthma. The patient had annual exacerbations (1–2 times a year) due to cold or allergen exposure, and acute respiratory viral infections. At the age of 42, polypoid rhinosinusitis was found, and a polypectomy was performed. Since 2006, the patient had been on controller treatment with salmeterol/fluticasone inhaler (25/250 mg, respectively) 2 doses 2 times a day.
In late 2009, extremely elevated eosinophils (53%) and total IgE (1000 IU/L) were found for the first time. Also, since 2009, the patient has had transient lung opacities, which usually disappeared after treatment with prednisolone 30 mg per day.
In August 2010, the patient noticed numbness on the back surface of the left foot, which spread to the entire left lower leg in 1 month, as well as numbness of the right foot and lower leg. Later, the patient started having tingling, twisting, and burning sensations aggravated by touching in the feet and lower legs. Also, the weakness in the distal parts of the legs developed, and he started having difficulties moving around on his own and falling while walking, and he was subsequently hospitalized in our clinic due to exacerbated asthma. On laboratory testing, high eosinophils (53%) and gamma-globulins (27.9%) were found in November 2010. The chest radiography showed emphysema and consolidation in the middle lobe of the right lung. A Neurology consult confirmed asymmetrical distal sensory-motor polyneuropathy demonstrated by lower distal asymmetrical paraparesis with decreased muscle strength in the foot flexors and extensors, more severe on the left side, low tone of the shin muscles, hypoactive Achilles reflexes, and reduced leg sensitivity starting from the middle of the shin and below, more severe in the peroneal nerve inner-vation area. The patient also demonstrated abnormal coordination (heel-to-knee, Romberg) test results and bilateral steppage gait with knee overextension. Electroneuromyography showed axonal lesion of all distal leg nerves and secondary myelinopathy. Thus, EGPA was suspected. The ANCA (anti-myeloperoxidase) titer was 30 U/ml (normal reference below 20 U/ml).
Allergic bronchopulmonary aspergillosis and other causes of hypereosinophilia were consistently ruled out. IgE and IgG for Aspergillus spp. and Aspergillus spp. in sputum were not found.
Thus, the presence of severe bronchial asthma, rhinosinusitis, high and persistent eosinophilia, transient pulmonary opacities, and polyneuropathy suggested EGPA (5 out of 6 clinical criteria of American College of Rheumatology present). Based on the clinical presentation and increased ANCA titer, the patient was diagnosed with EGPA. Systemic steroid treatment (prednisolone 300 mg intravenously and 30 mg orally) was prescribed, and 5 plasmapheresis sessions were performed. Later, prolonged combination immunosuppression (prednisolone 10 mg/day and methotrexate 25 mg/week) and controller asthma treatment (fluticasone propionate/salmeterol 250/25 µg 4 doses/ day, tiotropium bromide 18 µg per day) were prescribed. After 1 month on prednisolone and methotrexate, there was a significant neurological improvement: the motor and sensory disturbances decreased, and the patient was able to move around on his own. Further on, polyneuropathy completely resolved on the combination treatment. His asthma remained uncontrolled despite the combination treatment: he had frequent (3 times a year) severe exacerbations requiring hospitalization. In 2015, newly-emerged copious purulent sputum associated with frequent infection-related asthma exacerbations was noted. In January 2016, a patient underwent chest computed tomography (CT), which showed generalized bronchiectasis. Stenosis of the proximal bronchi was ruled out also by fibrobronchos-copy. The bronchial secretion microbiology revealed Klebsiella pneumoniae growth (107 colony-forming units).
In November 2016, during the asthma exacerbation, the patient suffered a cerebral ischemic infarction involving the right occipital lobe, confirmed by CT (Figures 1, 2).
On March 2, 2017, the patient was admitted to our hospital due to severe respiratory failure caused by pneumonia in the left upper lobe and severe asthma exacerbation.
The complete blood count showed high WBCs (15.2×109/l) with left shift. The IgE level was 144.0 IU/l, C-reactive protein was 48.8 mg/dL. Clinical chemistry was remarkable for urea nitrogen 18.4 mmol/l and creatinine 1.55 mg/dL. The sputum microscopy demonstrated 100 WBCs and 160 RBCs per vision field, and no eosinophils were revealed. A chest CT revealed multiple bronchiectasis and upper lobar pneumonia of the left lung (Figure 2). Klebsiella was again detected in the sputum culture.
The patient was prescribed intravenous antibiotics (ceftriaxone 2 g/day and ciprofloxacin 800 mg/day replaced by cefopera-zone 4 g/day and amikacin 1 g/day after 3 days due to low efficacy), bronchodilator and anti-inflammatory therapy (nebulized ipratropium bromide, fenoterol, and budesonide 2000 µg per day, prednisolone intravenously 150 mg and orally 30 mg). Despite the aggressive treatment, the respiratory failure progressed, and on day 6 of hospitalization the patient was transferred to the Intensive Care Unit and intubated. On day 7, the patient died. On autopsy, the EGPA diagnosis was confirmed, signs of ischemic infarction in the occipital lobe of the brain were found, and left upper lobe pneumonia with abscess formation and generalized bilateral bronchiectases were revealed.
Discussion
Based on the long follow-up period and careful interpretation of signs and symptoms, typical laboratory abnormalities, the diagnosis was clear. Among the most commonly used diagnostic criteria of EGPA are the American College of Rheumatology (ACR) classification criteria, Lanham’s criteria, and the Chapel Hill Consensus Conference criteria. The presence of 4 or more ACR classification criteria, including bronchial asthma, eosinophilia (more than 10%), paranasal sinus abnormalities, migratory or transient eosinophilic pulmonary infiltrates, extravascular eosinophil infiltration on biopsy, and mono- or polyneuropathy, has a sensitivity of 85% and a specificity of 100% [2,5,6]. We identified 5 out of 6 American College of Rheumatology clinical criteria present in our patient.
This clinical case shows gradual development of EGPA with multiple-organ involvement (see Figure 3). The disease onset was characterized by respiratory manifestations and suspicion of EGPA appeared only after the development of polyneuropathy (after 39 years) with hypereosinophilia. Polyneuropathy was noted at the age of 57, which is slightly later than the average age of onset reported in the literature [13]. Notably, the patient was ANCA-positive and his long-term neurological manifestation (mononeuritis) was quite typical for ANCA vasculitis [5,6,8,13]. The diagnosis became obvious after comprehensive assessment of the disease history with its gradually evolving signs and symptoms and laboratory abnormalities. Almost all manifestations of the disease seen in the patient were typical except for stroke, which is an exceptional feature for this case [1,5,6]. Neurological involvement manifested as ischemic stroke is an uncommon complication of EGPA (seen only in 5.2% of all patients) [1]. Thus, the patient demonstrated both peripheral and central nervous system involvement, which is unusual and life-threatening in EGPA.
This clinical case of EGPA also attracted our attention due to severe respiratory manifestations. While typical respiratory manifestations such as rhinosinusitis, asthma, and transient lung infiltrates are well documented in the literature, EGPA-associated bronchiectasis is rarely mentioned [14]. Bronchiectasis in the patient was detected by CT at the age of 63. The patient was a nonsmoker, and no other cause of bronchial abnormalities and airflow obstruction could be identified. Stenosis of the proximal bronchi was ruled out by fibrobronchoscopy and CT. The diagnosis of allergic bronchopulmonary aspergillosis was not confirmed. Klebsiella pneumoniae colonization was found during the last 2 years of the patient’s life. Klebsiella pneumoniae is a typical pathogen causing bronchiectasis [15]. It should be noted that aspergillosis and mycobacteriosis are common complications of bronchiectasis [15], but neither was identified in our patient.
The patient was treated for a long time with a combination of prednisolone and methotrexate. There was a significant positive effect on polyneuropathy, but uncontrolled bronchial asthma persisted. This treatment regimen complies with current recommendations [3,11]. Patients with life-threatening and/or organ-threatening manifestations (including lesions of the central nervous system and severe peripheral neuropathy) require therapy with a combination of a glucocorticoid and a cytostatic [3,11].
The use of immunosuppressive drugs in patients with ANCA-associated vasculitis significantly increases the risk of infection and mortality [16–18]. Prednisone and methotrexate use were found to be significant predictors of hospitalization for pneumonia. Frequent infections (more than 1 episode) are reported in patients with ANCA-associated vasculitis during the first year of diagnosis and initiation of therapy [18]. We assume that bronchiectasis complicated by bacterial colonization and bronchial obstruction in combination with immuno-suppression led to severe pneumonia and death of the patient.
Conclusions
The present case report demonstrates a long-term follow-up of a patient with EGPA. Both common and rare manifestations of EGPA have been described in the patient, such as central nervous system damage and bronchiectasis. Immunosuppressive treatment probably contributed to Klebsiella infection development. Thus, long-term complications of EGPA in combination with adverse effects of medications could have caused the severe pneumonia and death of the patient.
Figure 1. Brain multi-spiral computed tomography performed in 2016 showing ischemic brain damage in the right occipital lobe.
Figure 2. (A–C) Chest computed tomography of patient showing large left upper lobe pneumonia and bronchiectasis in lover lobes.
Figure 3. Timeline from first symptoms to death. | Intravenous (not otherwise specified) | DrugAdministrationRoute | CC BY-NC-ND | 33582702 | 18,986,567 | 2021-02-14 |
What was the administration route of drug 'CEFTRIAXONE'? | Case Report: A 64-Year-Old Man with 10-Year History of Eosinophilic Granulomatosis with Polyangiitis with Bronchiectasis and Severe Klebsiella pneumonia.
BACKGROUND Eosinophilic granulomatosis with polyangiitis (EGPA) is a rare and severe progressive disease with multiple clinical manifestations and organ damage. Usually, it requires long-term monitoring of the state of many organs due to the gradual character of its manifestations. CASE REPORT We report a case of a long-term follow-up of a patient with eosinophilic granulomatosis with polyangiitis with emphasis on specific clinical features in this patient. A 64-year-old man was being followed up for 10 years. The initial diagnosis was allergic bronchial asthma; however, as new clinical signs and symptoms developed, the diagnosis of EGPA became obvious. A positive treatment response was seen, mainly manifested as reduced polyneuropathy. Meanwhile, bronchial asthma remained uncontrolled and bronchiectasis and Klebsiella pneumoniae colonization developed despite the combination treatment with prednisolone and methotrexate. Furthermore, the patient suffered a cerebral ischemic infarction. During the last hospital admission, severe uncontrolled bronchial asthma complicated with pneumonia resulted in the patient's death. CONCLUSIONS This clinical case shows the gradual development of EGPA with multiple-organ involvement, including respiratory manifestations and peripheral and central nervous system damage. Immunosuppressive treatment combined with complications of EGPA could have contributed to severe pneumonia development and death of the patient.
Background
Systemic vasculitides are difficult to diagnose due to the heterogenous clinical manifestations requiring a wide differential diagnostic search. One of the most uncommon vasculitis types is eosinophilic granulomatosis with polyangiitis (EGPA), or Churg-Strauss syndrome [1,2]. This vasculitis is associated with the presence of antineutrophilic cytoplasmic antibodies (ANCA) and is characterized by necrotizing inflammatory lesions of mainly small vessels, as well as eosinophilic and granulomatous inflammation in various organs. Bronchial asthma and eosinophilia are the most common clinical manifestations of EGPA [1–3]. In 2012, according to the current classification of systemic vasculitides, Churg-Strauss syndrome was renamed “Eosinophilic Granulomatosis with Polyangiitis” (EGPA) and classified into the ANCA-associated vasculitides group [3].
EGPA is a rare ANCA-associated vasculitis. The incidence rate is 0.5–6.8 new cases/year per million populations in Europe [2]. Prevalence is higher in northern latitudes compared to southern Europe [4]. The disease occurs most commonly in people 40–60 years of age [2,5]. No sex predominance has been clearly demonstrated [4].
The pathogenesis of the disease is still not completely clear, but the key role is attributed to autoimmune inflammation with the participation of eosinophils, CD8+, and CD4+lymphocytes (predominantly Th-2) and their proinflammatory cytokines: interleukin (IL)4, IL5, and IL13 [2,5]. Elevation of immunoglobulin (Ig)E, IgG4, and eotaxin-3 is common in EGPA patients [2,5]. The frequent (in 30–40% of cases) detection of ANCA (mainly antibodies to myeloperoxidase) also suggests the autoimmune nature of the disease. ANCA-positive patients are more commonly diagnosed with glomerulonephritis and mononeuritis, whereas the ANCA-negative ones more frequently demonstrate endocardial, myocardial, and pulmonary eosinophilic infiltrates [1,2,5,6].
There are 3 phases in the development of EGPA (these phases partially overlap and may not appear in sequence):
the allergic phase, distinguished by allergic upper respiratory tract inflammation (rhinosinusitis, allergic rhinitis) and bronchial asthma;
the eosinophilic phase, manifested with severe eosinophilia in combination with multiple eosinophilic infiltrates in various organs (most often in the lungs and gastrointestinal tract);
the vasculitis phase, with small- and medium-vessel inflammation of different organs, which can often be life-threatening [5,6].
Respiratory manifestations of EGPA include allergic rhinitis, sinusitis, nasal polyposis and, in more than 95% of patients, poorly controlled bronchial asthma [5]. Chest radiography findings include transient and migratory opacities, ground-glass opacity, focal consolidation, and small peribronchial and centrilobular nodules or masses [7]. Computed tomography reveals bronchial abnormalities, most commonly thickened wall and dilatation with secondary bronchiectasis formation, leading to frequent infections [7,8]. Pleural effusions and alveolar hemorrhages are uncommon manifestations of EGPA [1,5,7,8].
A characteristic neurological manifestation seen in 75% of patients with EGPA is peripheral neuropathy (mononeuritis multiplex) [1,5]. Upper-limb symptoms due to neuropathy, as well as vasculitis in epineural vessels, were more frequently reported in the ANCA-positive group [9]. Eosinophil-associated tissue damage and vascular occlusion leading to ischemia are more specific in ANCA-negative patients [9]. Also, the tissue damage caused by eosinophils may be associated with IL-5 concentration [10]. Brain hemorrhage, subarachnoid hemorrhage, and ischemic brain infarction are rare [1,5,6].
The main drugs for the treatment of EGPA are glucocorticoids. In organ- or life-threatening manifestations, the recommended starting dose of prednisone is 1 mg/kg/day with a maximal daily dose of 80 mg for 2–3 weeks, with tapering to a minimal supporting dose selected individually to control the symptoms and minimize the adverse effects of treatment. Methylprednisolone (7.5–15 mg/kg/day) can be used as a pulse therapy for life-threatening manifestations [11,12]. Most patients need prolonged or lifelong steroids. In the case of organ- or life-threatening manifestations, glucocorticoids in combination with cyclophosphamide are prescribed [11,12]. The combination of glucocorticoids and methotrexate or mycophenolate is required to induce remission in mild or moderate disease [11,12]. Mepolizumab (a humanized monoclonal antibody targeting IL-5) has been used for EGPA treatment [11]. For ANCA-positive patients with renal involvement or refractory disease, rituximab can be considered [3, 11].
Due to the rare occurrence of EGPA and the variety of its clinical manifestations, the diagnosis and treatment of this disease can be challenging. Currently, reports of long-term observations with the detailed patient follow-up are of great interest. The aim of our case report is to present rare manifestations of EGPA, such as bronchiectasis and stroke. We also focused on the infectious complications of immunosuppressive treatments. Thus, we would like to present a 10-year observation of a patient with EGPA.
Case Report
The male patient, born in 1953, was first hospitalized in the Pulmonary Department of our clinic in 2007 due to exacerbated asthma. The patient’s history was remarkable for multiple allergies (to house dust, cat and dog epidermis, and plant pollen) as well as nonsteroidal anti-inflammatory drugs intolerance (manifested as bronchospasm). The patient denied unhealthy habits and occupational hazards (he worked as a driver). His family history was not remarkable. The patient has had allergic rhinitis since adolescence. The first asthma symptoms appeared at the age of 18 while serving in the army. At the age of 22, an allergist diagnosed him with bronchial asthma. The patient had annual exacerbations (1–2 times a year) due to cold or allergen exposure, and acute respiratory viral infections. At the age of 42, polypoid rhinosinusitis was found, and a polypectomy was performed. Since 2006, the patient had been on controller treatment with salmeterol/fluticasone inhaler (25/250 mg, respectively) 2 doses 2 times a day.
In late 2009, extremely elevated eosinophils (53%) and total IgE (1000 IU/L) were found for the first time. Also, since 2009, the patient has had transient lung opacities, which usually disappeared after treatment with prednisolone 30 mg per day.
In August 2010, the patient noticed numbness on the back surface of the left foot, which spread to the entire left lower leg in 1 month, as well as numbness of the right foot and lower leg. Later, the patient started having tingling, twisting, and burning sensations aggravated by touching in the feet and lower legs. Also, the weakness in the distal parts of the legs developed, and he started having difficulties moving around on his own and falling while walking, and he was subsequently hospitalized in our clinic due to exacerbated asthma. On laboratory testing, high eosinophils (53%) and gamma-globulins (27.9%) were found in November 2010. The chest radiography showed emphysema and consolidation in the middle lobe of the right lung. A Neurology consult confirmed asymmetrical distal sensory-motor polyneuropathy demonstrated by lower distal asymmetrical paraparesis with decreased muscle strength in the foot flexors and extensors, more severe on the left side, low tone of the shin muscles, hypoactive Achilles reflexes, and reduced leg sensitivity starting from the middle of the shin and below, more severe in the peroneal nerve inner-vation area. The patient also demonstrated abnormal coordination (heel-to-knee, Romberg) test results and bilateral steppage gait with knee overextension. Electroneuromyography showed axonal lesion of all distal leg nerves and secondary myelinopathy. Thus, EGPA was suspected. The ANCA (anti-myeloperoxidase) titer was 30 U/ml (normal reference below 20 U/ml).
Allergic bronchopulmonary aspergillosis and other causes of hypereosinophilia were consistently ruled out. IgE and IgG for Aspergillus spp. and Aspergillus spp. in sputum were not found.
Thus, the presence of severe bronchial asthma, rhinosinusitis, high and persistent eosinophilia, transient pulmonary opacities, and polyneuropathy suggested EGPA (5 out of 6 clinical criteria of American College of Rheumatology present). Based on the clinical presentation and increased ANCA titer, the patient was diagnosed with EGPA. Systemic steroid treatment (prednisolone 300 mg intravenously and 30 mg orally) was prescribed, and 5 plasmapheresis sessions were performed. Later, prolonged combination immunosuppression (prednisolone 10 mg/day and methotrexate 25 mg/week) and controller asthma treatment (fluticasone propionate/salmeterol 250/25 µg 4 doses/ day, tiotropium bromide 18 µg per day) were prescribed. After 1 month on prednisolone and methotrexate, there was a significant neurological improvement: the motor and sensory disturbances decreased, and the patient was able to move around on his own. Further on, polyneuropathy completely resolved on the combination treatment. His asthma remained uncontrolled despite the combination treatment: he had frequent (3 times a year) severe exacerbations requiring hospitalization. In 2015, newly-emerged copious purulent sputum associated with frequent infection-related asthma exacerbations was noted. In January 2016, a patient underwent chest computed tomography (CT), which showed generalized bronchiectasis. Stenosis of the proximal bronchi was ruled out also by fibrobronchos-copy. The bronchial secretion microbiology revealed Klebsiella pneumoniae growth (107 colony-forming units).
In November 2016, during the asthma exacerbation, the patient suffered a cerebral ischemic infarction involving the right occipital lobe, confirmed by CT (Figures 1, 2).
On March 2, 2017, the patient was admitted to our hospital due to severe respiratory failure caused by pneumonia in the left upper lobe and severe asthma exacerbation.
The complete blood count showed high WBCs (15.2×109/l) with left shift. The IgE level was 144.0 IU/l, C-reactive protein was 48.8 mg/dL. Clinical chemistry was remarkable for urea nitrogen 18.4 mmol/l and creatinine 1.55 mg/dL. The sputum microscopy demonstrated 100 WBCs and 160 RBCs per vision field, and no eosinophils were revealed. A chest CT revealed multiple bronchiectasis and upper lobar pneumonia of the left lung (Figure 2). Klebsiella was again detected in the sputum culture.
The patient was prescribed intravenous antibiotics (ceftriaxone 2 g/day and ciprofloxacin 800 mg/day replaced by cefopera-zone 4 g/day and amikacin 1 g/day after 3 days due to low efficacy), bronchodilator and anti-inflammatory therapy (nebulized ipratropium bromide, fenoterol, and budesonide 2000 µg per day, prednisolone intravenously 150 mg and orally 30 mg). Despite the aggressive treatment, the respiratory failure progressed, and on day 6 of hospitalization the patient was transferred to the Intensive Care Unit and intubated. On day 7, the patient died. On autopsy, the EGPA diagnosis was confirmed, signs of ischemic infarction in the occipital lobe of the brain were found, and left upper lobe pneumonia with abscess formation and generalized bilateral bronchiectases were revealed.
Discussion
Based on the long follow-up period and careful interpretation of signs and symptoms, typical laboratory abnormalities, the diagnosis was clear. Among the most commonly used diagnostic criteria of EGPA are the American College of Rheumatology (ACR) classification criteria, Lanham’s criteria, and the Chapel Hill Consensus Conference criteria. The presence of 4 or more ACR classification criteria, including bronchial asthma, eosinophilia (more than 10%), paranasal sinus abnormalities, migratory or transient eosinophilic pulmonary infiltrates, extravascular eosinophil infiltration on biopsy, and mono- or polyneuropathy, has a sensitivity of 85% and a specificity of 100% [2,5,6]. We identified 5 out of 6 American College of Rheumatology clinical criteria present in our patient.
This clinical case shows gradual development of EGPA with multiple-organ involvement (see Figure 3). The disease onset was characterized by respiratory manifestations and suspicion of EGPA appeared only after the development of polyneuropathy (after 39 years) with hypereosinophilia. Polyneuropathy was noted at the age of 57, which is slightly later than the average age of onset reported in the literature [13]. Notably, the patient was ANCA-positive and his long-term neurological manifestation (mononeuritis) was quite typical for ANCA vasculitis [5,6,8,13]. The diagnosis became obvious after comprehensive assessment of the disease history with its gradually evolving signs and symptoms and laboratory abnormalities. Almost all manifestations of the disease seen in the patient were typical except for stroke, which is an exceptional feature for this case [1,5,6]. Neurological involvement manifested as ischemic stroke is an uncommon complication of EGPA (seen only in 5.2% of all patients) [1]. Thus, the patient demonstrated both peripheral and central nervous system involvement, which is unusual and life-threatening in EGPA.
This clinical case of EGPA also attracted our attention due to severe respiratory manifestations. While typical respiratory manifestations such as rhinosinusitis, asthma, and transient lung infiltrates are well documented in the literature, EGPA-associated bronchiectasis is rarely mentioned [14]. Bronchiectasis in the patient was detected by CT at the age of 63. The patient was a nonsmoker, and no other cause of bronchial abnormalities and airflow obstruction could be identified. Stenosis of the proximal bronchi was ruled out by fibrobronchoscopy and CT. The diagnosis of allergic bronchopulmonary aspergillosis was not confirmed. Klebsiella pneumoniae colonization was found during the last 2 years of the patient’s life. Klebsiella pneumoniae is a typical pathogen causing bronchiectasis [15]. It should be noted that aspergillosis and mycobacteriosis are common complications of bronchiectasis [15], but neither was identified in our patient.
The patient was treated for a long time with a combination of prednisolone and methotrexate. There was a significant positive effect on polyneuropathy, but uncontrolled bronchial asthma persisted. This treatment regimen complies with current recommendations [3,11]. Patients with life-threatening and/or organ-threatening manifestations (including lesions of the central nervous system and severe peripheral neuropathy) require therapy with a combination of a glucocorticoid and a cytostatic [3,11].
The use of immunosuppressive drugs in patients with ANCA-associated vasculitis significantly increases the risk of infection and mortality [16–18]. Prednisone and methotrexate use were found to be significant predictors of hospitalization for pneumonia. Frequent infections (more than 1 episode) are reported in patients with ANCA-associated vasculitis during the first year of diagnosis and initiation of therapy [18]. We assume that bronchiectasis complicated by bacterial colonization and bronchial obstruction in combination with immuno-suppression led to severe pneumonia and death of the patient.
Conclusions
The present case report demonstrates a long-term follow-up of a patient with EGPA. Both common and rare manifestations of EGPA have been described in the patient, such as central nervous system damage and bronchiectasis. Immunosuppressive treatment probably contributed to Klebsiella infection development. Thus, long-term complications of EGPA in combination with adverse effects of medications could have caused the severe pneumonia and death of the patient.
Figure 1. Brain multi-spiral computed tomography performed in 2016 showing ischemic brain damage in the right occipital lobe.
Figure 2. (A–C) Chest computed tomography of patient showing large left upper lobe pneumonia and bronchiectasis in lover lobes.
Figure 3. Timeline from first symptoms to death. | Intravenous (not otherwise specified) | DrugAdministrationRoute | CC BY-NC-ND | 33582702 | 18,986,567 | 2021-02-14 |
What was the administration route of drug 'CIPROFLOXACIN'? | Case Report: A 64-Year-Old Man with 10-Year History of Eosinophilic Granulomatosis with Polyangiitis with Bronchiectasis and Severe Klebsiella pneumonia.
BACKGROUND Eosinophilic granulomatosis with polyangiitis (EGPA) is a rare and severe progressive disease with multiple clinical manifestations and organ damage. Usually, it requires long-term monitoring of the state of many organs due to the gradual character of its manifestations. CASE REPORT We report a case of a long-term follow-up of a patient with eosinophilic granulomatosis with polyangiitis with emphasis on specific clinical features in this patient. A 64-year-old man was being followed up for 10 years. The initial diagnosis was allergic bronchial asthma; however, as new clinical signs and symptoms developed, the diagnosis of EGPA became obvious. A positive treatment response was seen, mainly manifested as reduced polyneuropathy. Meanwhile, bronchial asthma remained uncontrolled and bronchiectasis and Klebsiella pneumoniae colonization developed despite the combination treatment with prednisolone and methotrexate. Furthermore, the patient suffered a cerebral ischemic infarction. During the last hospital admission, severe uncontrolled bronchial asthma complicated with pneumonia resulted in the patient's death. CONCLUSIONS This clinical case shows the gradual development of EGPA with multiple-organ involvement, including respiratory manifestations and peripheral and central nervous system damage. Immunosuppressive treatment combined with complications of EGPA could have contributed to severe pneumonia development and death of the patient.
Background
Systemic vasculitides are difficult to diagnose due to the heterogenous clinical manifestations requiring a wide differential diagnostic search. One of the most uncommon vasculitis types is eosinophilic granulomatosis with polyangiitis (EGPA), or Churg-Strauss syndrome [1,2]. This vasculitis is associated with the presence of antineutrophilic cytoplasmic antibodies (ANCA) and is characterized by necrotizing inflammatory lesions of mainly small vessels, as well as eosinophilic and granulomatous inflammation in various organs. Bronchial asthma and eosinophilia are the most common clinical manifestations of EGPA [1–3]. In 2012, according to the current classification of systemic vasculitides, Churg-Strauss syndrome was renamed “Eosinophilic Granulomatosis with Polyangiitis” (EGPA) and classified into the ANCA-associated vasculitides group [3].
EGPA is a rare ANCA-associated vasculitis. The incidence rate is 0.5–6.8 new cases/year per million populations in Europe [2]. Prevalence is higher in northern latitudes compared to southern Europe [4]. The disease occurs most commonly in people 40–60 years of age [2,5]. No sex predominance has been clearly demonstrated [4].
The pathogenesis of the disease is still not completely clear, but the key role is attributed to autoimmune inflammation with the participation of eosinophils, CD8+, and CD4+lymphocytes (predominantly Th-2) and their proinflammatory cytokines: interleukin (IL)4, IL5, and IL13 [2,5]. Elevation of immunoglobulin (Ig)E, IgG4, and eotaxin-3 is common in EGPA patients [2,5]. The frequent (in 30–40% of cases) detection of ANCA (mainly antibodies to myeloperoxidase) also suggests the autoimmune nature of the disease. ANCA-positive patients are more commonly diagnosed with glomerulonephritis and mononeuritis, whereas the ANCA-negative ones more frequently demonstrate endocardial, myocardial, and pulmonary eosinophilic infiltrates [1,2,5,6].
There are 3 phases in the development of EGPA (these phases partially overlap and may not appear in sequence):
the allergic phase, distinguished by allergic upper respiratory tract inflammation (rhinosinusitis, allergic rhinitis) and bronchial asthma;
the eosinophilic phase, manifested with severe eosinophilia in combination with multiple eosinophilic infiltrates in various organs (most often in the lungs and gastrointestinal tract);
the vasculitis phase, with small- and medium-vessel inflammation of different organs, which can often be life-threatening [5,6].
Respiratory manifestations of EGPA include allergic rhinitis, sinusitis, nasal polyposis and, in more than 95% of patients, poorly controlled bronchial asthma [5]. Chest radiography findings include transient and migratory opacities, ground-glass opacity, focal consolidation, and small peribronchial and centrilobular nodules or masses [7]. Computed tomography reveals bronchial abnormalities, most commonly thickened wall and dilatation with secondary bronchiectasis formation, leading to frequent infections [7,8]. Pleural effusions and alveolar hemorrhages are uncommon manifestations of EGPA [1,5,7,8].
A characteristic neurological manifestation seen in 75% of patients with EGPA is peripheral neuropathy (mononeuritis multiplex) [1,5]. Upper-limb symptoms due to neuropathy, as well as vasculitis in epineural vessels, were more frequently reported in the ANCA-positive group [9]. Eosinophil-associated tissue damage and vascular occlusion leading to ischemia are more specific in ANCA-negative patients [9]. Also, the tissue damage caused by eosinophils may be associated with IL-5 concentration [10]. Brain hemorrhage, subarachnoid hemorrhage, and ischemic brain infarction are rare [1,5,6].
The main drugs for the treatment of EGPA are glucocorticoids. In organ- or life-threatening manifestations, the recommended starting dose of prednisone is 1 mg/kg/day with a maximal daily dose of 80 mg for 2–3 weeks, with tapering to a minimal supporting dose selected individually to control the symptoms and minimize the adverse effects of treatment. Methylprednisolone (7.5–15 mg/kg/day) can be used as a pulse therapy for life-threatening manifestations [11,12]. Most patients need prolonged or lifelong steroids. In the case of organ- or life-threatening manifestations, glucocorticoids in combination with cyclophosphamide are prescribed [11,12]. The combination of glucocorticoids and methotrexate or mycophenolate is required to induce remission in mild or moderate disease [11,12]. Mepolizumab (a humanized monoclonal antibody targeting IL-5) has been used for EGPA treatment [11]. For ANCA-positive patients with renal involvement or refractory disease, rituximab can be considered [3, 11].
Due to the rare occurrence of EGPA and the variety of its clinical manifestations, the diagnosis and treatment of this disease can be challenging. Currently, reports of long-term observations with the detailed patient follow-up are of great interest. The aim of our case report is to present rare manifestations of EGPA, such as bronchiectasis and stroke. We also focused on the infectious complications of immunosuppressive treatments. Thus, we would like to present a 10-year observation of a patient with EGPA.
Case Report
The male patient, born in 1953, was first hospitalized in the Pulmonary Department of our clinic in 2007 due to exacerbated asthma. The patient’s history was remarkable for multiple allergies (to house dust, cat and dog epidermis, and plant pollen) as well as nonsteroidal anti-inflammatory drugs intolerance (manifested as bronchospasm). The patient denied unhealthy habits and occupational hazards (he worked as a driver). His family history was not remarkable. The patient has had allergic rhinitis since adolescence. The first asthma symptoms appeared at the age of 18 while serving in the army. At the age of 22, an allergist diagnosed him with bronchial asthma. The patient had annual exacerbations (1–2 times a year) due to cold or allergen exposure, and acute respiratory viral infections. At the age of 42, polypoid rhinosinusitis was found, and a polypectomy was performed. Since 2006, the patient had been on controller treatment with salmeterol/fluticasone inhaler (25/250 mg, respectively) 2 doses 2 times a day.
In late 2009, extremely elevated eosinophils (53%) and total IgE (1000 IU/L) were found for the first time. Also, since 2009, the patient has had transient lung opacities, which usually disappeared after treatment with prednisolone 30 mg per day.
In August 2010, the patient noticed numbness on the back surface of the left foot, which spread to the entire left lower leg in 1 month, as well as numbness of the right foot and lower leg. Later, the patient started having tingling, twisting, and burning sensations aggravated by touching in the feet and lower legs. Also, the weakness in the distal parts of the legs developed, and he started having difficulties moving around on his own and falling while walking, and he was subsequently hospitalized in our clinic due to exacerbated asthma. On laboratory testing, high eosinophils (53%) and gamma-globulins (27.9%) were found in November 2010. The chest radiography showed emphysema and consolidation in the middle lobe of the right lung. A Neurology consult confirmed asymmetrical distal sensory-motor polyneuropathy demonstrated by lower distal asymmetrical paraparesis with decreased muscle strength in the foot flexors and extensors, more severe on the left side, low tone of the shin muscles, hypoactive Achilles reflexes, and reduced leg sensitivity starting from the middle of the shin and below, more severe in the peroneal nerve inner-vation area. The patient also demonstrated abnormal coordination (heel-to-knee, Romberg) test results and bilateral steppage gait with knee overextension. Electroneuromyography showed axonal lesion of all distal leg nerves and secondary myelinopathy. Thus, EGPA was suspected. The ANCA (anti-myeloperoxidase) titer was 30 U/ml (normal reference below 20 U/ml).
Allergic bronchopulmonary aspergillosis and other causes of hypereosinophilia were consistently ruled out. IgE and IgG for Aspergillus spp. and Aspergillus spp. in sputum were not found.
Thus, the presence of severe bronchial asthma, rhinosinusitis, high and persistent eosinophilia, transient pulmonary opacities, and polyneuropathy suggested EGPA (5 out of 6 clinical criteria of American College of Rheumatology present). Based on the clinical presentation and increased ANCA titer, the patient was diagnosed with EGPA. Systemic steroid treatment (prednisolone 300 mg intravenously and 30 mg orally) was prescribed, and 5 plasmapheresis sessions were performed. Later, prolonged combination immunosuppression (prednisolone 10 mg/day and methotrexate 25 mg/week) and controller asthma treatment (fluticasone propionate/salmeterol 250/25 µg 4 doses/ day, tiotropium bromide 18 µg per day) were prescribed. After 1 month on prednisolone and methotrexate, there was a significant neurological improvement: the motor and sensory disturbances decreased, and the patient was able to move around on his own. Further on, polyneuropathy completely resolved on the combination treatment. His asthma remained uncontrolled despite the combination treatment: he had frequent (3 times a year) severe exacerbations requiring hospitalization. In 2015, newly-emerged copious purulent sputum associated with frequent infection-related asthma exacerbations was noted. In January 2016, a patient underwent chest computed tomography (CT), which showed generalized bronchiectasis. Stenosis of the proximal bronchi was ruled out also by fibrobronchos-copy. The bronchial secretion microbiology revealed Klebsiella pneumoniae growth (107 colony-forming units).
In November 2016, during the asthma exacerbation, the patient suffered a cerebral ischemic infarction involving the right occipital lobe, confirmed by CT (Figures 1, 2).
On March 2, 2017, the patient was admitted to our hospital due to severe respiratory failure caused by pneumonia in the left upper lobe and severe asthma exacerbation.
The complete blood count showed high WBCs (15.2×109/l) with left shift. The IgE level was 144.0 IU/l, C-reactive protein was 48.8 mg/dL. Clinical chemistry was remarkable for urea nitrogen 18.4 mmol/l and creatinine 1.55 mg/dL. The sputum microscopy demonstrated 100 WBCs and 160 RBCs per vision field, and no eosinophils were revealed. A chest CT revealed multiple bronchiectasis and upper lobar pneumonia of the left lung (Figure 2). Klebsiella was again detected in the sputum culture.
The patient was prescribed intravenous antibiotics (ceftriaxone 2 g/day and ciprofloxacin 800 mg/day replaced by cefopera-zone 4 g/day and amikacin 1 g/day after 3 days due to low efficacy), bronchodilator and anti-inflammatory therapy (nebulized ipratropium bromide, fenoterol, and budesonide 2000 µg per day, prednisolone intravenously 150 mg and orally 30 mg). Despite the aggressive treatment, the respiratory failure progressed, and on day 6 of hospitalization the patient was transferred to the Intensive Care Unit and intubated. On day 7, the patient died. On autopsy, the EGPA diagnosis was confirmed, signs of ischemic infarction in the occipital lobe of the brain were found, and left upper lobe pneumonia with abscess formation and generalized bilateral bronchiectases were revealed.
Discussion
Based on the long follow-up period and careful interpretation of signs and symptoms, typical laboratory abnormalities, the diagnosis was clear. Among the most commonly used diagnostic criteria of EGPA are the American College of Rheumatology (ACR) classification criteria, Lanham’s criteria, and the Chapel Hill Consensus Conference criteria. The presence of 4 or more ACR classification criteria, including bronchial asthma, eosinophilia (more than 10%), paranasal sinus abnormalities, migratory or transient eosinophilic pulmonary infiltrates, extravascular eosinophil infiltration on biopsy, and mono- or polyneuropathy, has a sensitivity of 85% and a specificity of 100% [2,5,6]. We identified 5 out of 6 American College of Rheumatology clinical criteria present in our patient.
This clinical case shows gradual development of EGPA with multiple-organ involvement (see Figure 3). The disease onset was characterized by respiratory manifestations and suspicion of EGPA appeared only after the development of polyneuropathy (after 39 years) with hypereosinophilia. Polyneuropathy was noted at the age of 57, which is slightly later than the average age of onset reported in the literature [13]. Notably, the patient was ANCA-positive and his long-term neurological manifestation (mononeuritis) was quite typical for ANCA vasculitis [5,6,8,13]. The diagnosis became obvious after comprehensive assessment of the disease history with its gradually evolving signs and symptoms and laboratory abnormalities. Almost all manifestations of the disease seen in the patient were typical except for stroke, which is an exceptional feature for this case [1,5,6]. Neurological involvement manifested as ischemic stroke is an uncommon complication of EGPA (seen only in 5.2% of all patients) [1]. Thus, the patient demonstrated both peripheral and central nervous system involvement, which is unusual and life-threatening in EGPA.
This clinical case of EGPA also attracted our attention due to severe respiratory manifestations. While typical respiratory manifestations such as rhinosinusitis, asthma, and transient lung infiltrates are well documented in the literature, EGPA-associated bronchiectasis is rarely mentioned [14]. Bronchiectasis in the patient was detected by CT at the age of 63. The patient was a nonsmoker, and no other cause of bronchial abnormalities and airflow obstruction could be identified. Stenosis of the proximal bronchi was ruled out by fibrobronchoscopy and CT. The diagnosis of allergic bronchopulmonary aspergillosis was not confirmed. Klebsiella pneumoniae colonization was found during the last 2 years of the patient’s life. Klebsiella pneumoniae is a typical pathogen causing bronchiectasis [15]. It should be noted that aspergillosis and mycobacteriosis are common complications of bronchiectasis [15], but neither was identified in our patient.
The patient was treated for a long time with a combination of prednisolone and methotrexate. There was a significant positive effect on polyneuropathy, but uncontrolled bronchial asthma persisted. This treatment regimen complies with current recommendations [3,11]. Patients with life-threatening and/or organ-threatening manifestations (including lesions of the central nervous system and severe peripheral neuropathy) require therapy with a combination of a glucocorticoid and a cytostatic [3,11].
The use of immunosuppressive drugs in patients with ANCA-associated vasculitis significantly increases the risk of infection and mortality [16–18]. Prednisone and methotrexate use were found to be significant predictors of hospitalization for pneumonia. Frequent infections (more than 1 episode) are reported in patients with ANCA-associated vasculitis during the first year of diagnosis and initiation of therapy [18]. We assume that bronchiectasis complicated by bacterial colonization and bronchial obstruction in combination with immuno-suppression led to severe pneumonia and death of the patient.
Conclusions
The present case report demonstrates a long-term follow-up of a patient with EGPA. Both common and rare manifestations of EGPA have been described in the patient, such as central nervous system damage and bronchiectasis. Immunosuppressive treatment probably contributed to Klebsiella infection development. Thus, long-term complications of EGPA in combination with adverse effects of medications could have caused the severe pneumonia and death of the patient.
Figure 1. Brain multi-spiral computed tomography performed in 2016 showing ischemic brain damage in the right occipital lobe.
Figure 2. (A–C) Chest computed tomography of patient showing large left upper lobe pneumonia and bronchiectasis in lover lobes.
Figure 3. Timeline from first symptoms to death. | Intravenous (not otherwise specified) | DrugAdministrationRoute | CC BY-NC-ND | 33582702 | 18,986,567 | 2021-02-14 |
What was the administration route of drug 'FENOTEROL'? | Case Report: A 64-Year-Old Man with 10-Year History of Eosinophilic Granulomatosis with Polyangiitis with Bronchiectasis and Severe Klebsiella pneumonia.
BACKGROUND Eosinophilic granulomatosis with polyangiitis (EGPA) is a rare and severe progressive disease with multiple clinical manifestations and organ damage. Usually, it requires long-term monitoring of the state of many organs due to the gradual character of its manifestations. CASE REPORT We report a case of a long-term follow-up of a patient with eosinophilic granulomatosis with polyangiitis with emphasis on specific clinical features in this patient. A 64-year-old man was being followed up for 10 years. The initial diagnosis was allergic bronchial asthma; however, as new clinical signs and symptoms developed, the diagnosis of EGPA became obvious. A positive treatment response was seen, mainly manifested as reduced polyneuropathy. Meanwhile, bronchial asthma remained uncontrolled and bronchiectasis and Klebsiella pneumoniae colonization developed despite the combination treatment with prednisolone and methotrexate. Furthermore, the patient suffered a cerebral ischemic infarction. During the last hospital admission, severe uncontrolled bronchial asthma complicated with pneumonia resulted in the patient's death. CONCLUSIONS This clinical case shows the gradual development of EGPA with multiple-organ involvement, including respiratory manifestations and peripheral and central nervous system damage. Immunosuppressive treatment combined with complications of EGPA could have contributed to severe pneumonia development and death of the patient.
Background
Systemic vasculitides are difficult to diagnose due to the heterogenous clinical manifestations requiring a wide differential diagnostic search. One of the most uncommon vasculitis types is eosinophilic granulomatosis with polyangiitis (EGPA), or Churg-Strauss syndrome [1,2]. This vasculitis is associated with the presence of antineutrophilic cytoplasmic antibodies (ANCA) and is characterized by necrotizing inflammatory lesions of mainly small vessels, as well as eosinophilic and granulomatous inflammation in various organs. Bronchial asthma and eosinophilia are the most common clinical manifestations of EGPA [1–3]. In 2012, according to the current classification of systemic vasculitides, Churg-Strauss syndrome was renamed “Eosinophilic Granulomatosis with Polyangiitis” (EGPA) and classified into the ANCA-associated vasculitides group [3].
EGPA is a rare ANCA-associated vasculitis. The incidence rate is 0.5–6.8 new cases/year per million populations in Europe [2]. Prevalence is higher in northern latitudes compared to southern Europe [4]. The disease occurs most commonly in people 40–60 years of age [2,5]. No sex predominance has been clearly demonstrated [4].
The pathogenesis of the disease is still not completely clear, but the key role is attributed to autoimmune inflammation with the participation of eosinophils, CD8+, and CD4+lymphocytes (predominantly Th-2) and their proinflammatory cytokines: interleukin (IL)4, IL5, and IL13 [2,5]. Elevation of immunoglobulin (Ig)E, IgG4, and eotaxin-3 is common in EGPA patients [2,5]. The frequent (in 30–40% of cases) detection of ANCA (mainly antibodies to myeloperoxidase) also suggests the autoimmune nature of the disease. ANCA-positive patients are more commonly diagnosed with glomerulonephritis and mononeuritis, whereas the ANCA-negative ones more frequently demonstrate endocardial, myocardial, and pulmonary eosinophilic infiltrates [1,2,5,6].
There are 3 phases in the development of EGPA (these phases partially overlap and may not appear in sequence):
the allergic phase, distinguished by allergic upper respiratory tract inflammation (rhinosinusitis, allergic rhinitis) and bronchial asthma;
the eosinophilic phase, manifested with severe eosinophilia in combination with multiple eosinophilic infiltrates in various organs (most often in the lungs and gastrointestinal tract);
the vasculitis phase, with small- and medium-vessel inflammation of different organs, which can often be life-threatening [5,6].
Respiratory manifestations of EGPA include allergic rhinitis, sinusitis, nasal polyposis and, in more than 95% of patients, poorly controlled bronchial asthma [5]. Chest radiography findings include transient and migratory opacities, ground-glass opacity, focal consolidation, and small peribronchial and centrilobular nodules or masses [7]. Computed tomography reveals bronchial abnormalities, most commonly thickened wall and dilatation with secondary bronchiectasis formation, leading to frequent infections [7,8]. Pleural effusions and alveolar hemorrhages are uncommon manifestations of EGPA [1,5,7,8].
A characteristic neurological manifestation seen in 75% of patients with EGPA is peripheral neuropathy (mononeuritis multiplex) [1,5]. Upper-limb symptoms due to neuropathy, as well as vasculitis in epineural vessels, were more frequently reported in the ANCA-positive group [9]. Eosinophil-associated tissue damage and vascular occlusion leading to ischemia are more specific in ANCA-negative patients [9]. Also, the tissue damage caused by eosinophils may be associated with IL-5 concentration [10]. Brain hemorrhage, subarachnoid hemorrhage, and ischemic brain infarction are rare [1,5,6].
The main drugs for the treatment of EGPA are glucocorticoids. In organ- or life-threatening manifestations, the recommended starting dose of prednisone is 1 mg/kg/day with a maximal daily dose of 80 mg for 2–3 weeks, with tapering to a minimal supporting dose selected individually to control the symptoms and minimize the adverse effects of treatment. Methylprednisolone (7.5–15 mg/kg/day) can be used as a pulse therapy for life-threatening manifestations [11,12]. Most patients need prolonged or lifelong steroids. In the case of organ- or life-threatening manifestations, glucocorticoids in combination with cyclophosphamide are prescribed [11,12]. The combination of glucocorticoids and methotrexate or mycophenolate is required to induce remission in mild or moderate disease [11,12]. Mepolizumab (a humanized monoclonal antibody targeting IL-5) has been used for EGPA treatment [11]. For ANCA-positive patients with renal involvement or refractory disease, rituximab can be considered [3, 11].
Due to the rare occurrence of EGPA and the variety of its clinical manifestations, the diagnosis and treatment of this disease can be challenging. Currently, reports of long-term observations with the detailed patient follow-up are of great interest. The aim of our case report is to present rare manifestations of EGPA, such as bronchiectasis and stroke. We also focused on the infectious complications of immunosuppressive treatments. Thus, we would like to present a 10-year observation of a patient with EGPA.
Case Report
The male patient, born in 1953, was first hospitalized in the Pulmonary Department of our clinic in 2007 due to exacerbated asthma. The patient’s history was remarkable for multiple allergies (to house dust, cat and dog epidermis, and plant pollen) as well as nonsteroidal anti-inflammatory drugs intolerance (manifested as bronchospasm). The patient denied unhealthy habits and occupational hazards (he worked as a driver). His family history was not remarkable. The patient has had allergic rhinitis since adolescence. The first asthma symptoms appeared at the age of 18 while serving in the army. At the age of 22, an allergist diagnosed him with bronchial asthma. The patient had annual exacerbations (1–2 times a year) due to cold or allergen exposure, and acute respiratory viral infections. At the age of 42, polypoid rhinosinusitis was found, and a polypectomy was performed. Since 2006, the patient had been on controller treatment with salmeterol/fluticasone inhaler (25/250 mg, respectively) 2 doses 2 times a day.
In late 2009, extremely elevated eosinophils (53%) and total IgE (1000 IU/L) were found for the first time. Also, since 2009, the patient has had transient lung opacities, which usually disappeared after treatment with prednisolone 30 mg per day.
In August 2010, the patient noticed numbness on the back surface of the left foot, which spread to the entire left lower leg in 1 month, as well as numbness of the right foot and lower leg. Later, the patient started having tingling, twisting, and burning sensations aggravated by touching in the feet and lower legs. Also, the weakness in the distal parts of the legs developed, and he started having difficulties moving around on his own and falling while walking, and he was subsequently hospitalized in our clinic due to exacerbated asthma. On laboratory testing, high eosinophils (53%) and gamma-globulins (27.9%) were found in November 2010. The chest radiography showed emphysema and consolidation in the middle lobe of the right lung. A Neurology consult confirmed asymmetrical distal sensory-motor polyneuropathy demonstrated by lower distal asymmetrical paraparesis with decreased muscle strength in the foot flexors and extensors, more severe on the left side, low tone of the shin muscles, hypoactive Achilles reflexes, and reduced leg sensitivity starting from the middle of the shin and below, more severe in the peroneal nerve inner-vation area. The patient also demonstrated abnormal coordination (heel-to-knee, Romberg) test results and bilateral steppage gait with knee overextension. Electroneuromyography showed axonal lesion of all distal leg nerves and secondary myelinopathy. Thus, EGPA was suspected. The ANCA (anti-myeloperoxidase) titer was 30 U/ml (normal reference below 20 U/ml).
Allergic bronchopulmonary aspergillosis and other causes of hypereosinophilia were consistently ruled out. IgE and IgG for Aspergillus spp. and Aspergillus spp. in sputum were not found.
Thus, the presence of severe bronchial asthma, rhinosinusitis, high and persistent eosinophilia, transient pulmonary opacities, and polyneuropathy suggested EGPA (5 out of 6 clinical criteria of American College of Rheumatology present). Based on the clinical presentation and increased ANCA titer, the patient was diagnosed with EGPA. Systemic steroid treatment (prednisolone 300 mg intravenously and 30 mg orally) was prescribed, and 5 plasmapheresis sessions were performed. Later, prolonged combination immunosuppression (prednisolone 10 mg/day and methotrexate 25 mg/week) and controller asthma treatment (fluticasone propionate/salmeterol 250/25 µg 4 doses/ day, tiotropium bromide 18 µg per day) were prescribed. After 1 month on prednisolone and methotrexate, there was a significant neurological improvement: the motor and sensory disturbances decreased, and the patient was able to move around on his own. Further on, polyneuropathy completely resolved on the combination treatment. His asthma remained uncontrolled despite the combination treatment: he had frequent (3 times a year) severe exacerbations requiring hospitalization. In 2015, newly-emerged copious purulent sputum associated with frequent infection-related asthma exacerbations was noted. In January 2016, a patient underwent chest computed tomography (CT), which showed generalized bronchiectasis. Stenosis of the proximal bronchi was ruled out also by fibrobronchos-copy. The bronchial secretion microbiology revealed Klebsiella pneumoniae growth (107 colony-forming units).
In November 2016, during the asthma exacerbation, the patient suffered a cerebral ischemic infarction involving the right occipital lobe, confirmed by CT (Figures 1, 2).
On March 2, 2017, the patient was admitted to our hospital due to severe respiratory failure caused by pneumonia in the left upper lobe and severe asthma exacerbation.
The complete blood count showed high WBCs (15.2×109/l) with left shift. The IgE level was 144.0 IU/l, C-reactive protein was 48.8 mg/dL. Clinical chemistry was remarkable for urea nitrogen 18.4 mmol/l and creatinine 1.55 mg/dL. The sputum microscopy demonstrated 100 WBCs and 160 RBCs per vision field, and no eosinophils were revealed. A chest CT revealed multiple bronchiectasis and upper lobar pneumonia of the left lung (Figure 2). Klebsiella was again detected in the sputum culture.
The patient was prescribed intravenous antibiotics (ceftriaxone 2 g/day and ciprofloxacin 800 mg/day replaced by cefopera-zone 4 g/day and amikacin 1 g/day after 3 days due to low efficacy), bronchodilator and anti-inflammatory therapy (nebulized ipratropium bromide, fenoterol, and budesonide 2000 µg per day, prednisolone intravenously 150 mg and orally 30 mg). Despite the aggressive treatment, the respiratory failure progressed, and on day 6 of hospitalization the patient was transferred to the Intensive Care Unit and intubated. On day 7, the patient died. On autopsy, the EGPA diagnosis was confirmed, signs of ischemic infarction in the occipital lobe of the brain were found, and left upper lobe pneumonia with abscess formation and generalized bilateral bronchiectases were revealed.
Discussion
Based on the long follow-up period and careful interpretation of signs and symptoms, typical laboratory abnormalities, the diagnosis was clear. Among the most commonly used diagnostic criteria of EGPA are the American College of Rheumatology (ACR) classification criteria, Lanham’s criteria, and the Chapel Hill Consensus Conference criteria. The presence of 4 or more ACR classification criteria, including bronchial asthma, eosinophilia (more than 10%), paranasal sinus abnormalities, migratory or transient eosinophilic pulmonary infiltrates, extravascular eosinophil infiltration on biopsy, and mono- or polyneuropathy, has a sensitivity of 85% and a specificity of 100% [2,5,6]. We identified 5 out of 6 American College of Rheumatology clinical criteria present in our patient.
This clinical case shows gradual development of EGPA with multiple-organ involvement (see Figure 3). The disease onset was characterized by respiratory manifestations and suspicion of EGPA appeared only after the development of polyneuropathy (after 39 years) with hypereosinophilia. Polyneuropathy was noted at the age of 57, which is slightly later than the average age of onset reported in the literature [13]. Notably, the patient was ANCA-positive and his long-term neurological manifestation (mononeuritis) was quite typical for ANCA vasculitis [5,6,8,13]. The diagnosis became obvious after comprehensive assessment of the disease history with its gradually evolving signs and symptoms and laboratory abnormalities. Almost all manifestations of the disease seen in the patient were typical except for stroke, which is an exceptional feature for this case [1,5,6]. Neurological involvement manifested as ischemic stroke is an uncommon complication of EGPA (seen only in 5.2% of all patients) [1]. Thus, the patient demonstrated both peripheral and central nervous system involvement, which is unusual and life-threatening in EGPA.
This clinical case of EGPA also attracted our attention due to severe respiratory manifestations. While typical respiratory manifestations such as rhinosinusitis, asthma, and transient lung infiltrates are well documented in the literature, EGPA-associated bronchiectasis is rarely mentioned [14]. Bronchiectasis in the patient was detected by CT at the age of 63. The patient was a nonsmoker, and no other cause of bronchial abnormalities and airflow obstruction could be identified. Stenosis of the proximal bronchi was ruled out by fibrobronchoscopy and CT. The diagnosis of allergic bronchopulmonary aspergillosis was not confirmed. Klebsiella pneumoniae colonization was found during the last 2 years of the patient’s life. Klebsiella pneumoniae is a typical pathogen causing bronchiectasis [15]. It should be noted that aspergillosis and mycobacteriosis are common complications of bronchiectasis [15], but neither was identified in our patient.
The patient was treated for a long time with a combination of prednisolone and methotrexate. There was a significant positive effect on polyneuropathy, but uncontrolled bronchial asthma persisted. This treatment regimen complies with current recommendations [3,11]. Patients with life-threatening and/or organ-threatening manifestations (including lesions of the central nervous system and severe peripheral neuropathy) require therapy with a combination of a glucocorticoid and a cytostatic [3,11].
The use of immunosuppressive drugs in patients with ANCA-associated vasculitis significantly increases the risk of infection and mortality [16–18]. Prednisone and methotrexate use were found to be significant predictors of hospitalization for pneumonia. Frequent infections (more than 1 episode) are reported in patients with ANCA-associated vasculitis during the first year of diagnosis and initiation of therapy [18]. We assume that bronchiectasis complicated by bacterial colonization and bronchial obstruction in combination with immuno-suppression led to severe pneumonia and death of the patient.
Conclusions
The present case report demonstrates a long-term follow-up of a patient with EGPA. Both common and rare manifestations of EGPA have been described in the patient, such as central nervous system damage and bronchiectasis. Immunosuppressive treatment probably contributed to Klebsiella infection development. Thus, long-term complications of EGPA in combination with adverse effects of medications could have caused the severe pneumonia and death of the patient.
Figure 1. Brain multi-spiral computed tomography performed in 2016 showing ischemic brain damage in the right occipital lobe.
Figure 2. (A–C) Chest computed tomography of patient showing large left upper lobe pneumonia and bronchiectasis in lover lobes.
Figure 3. Timeline from first symptoms to death. | Intravenous (not otherwise specified) | DrugAdministrationRoute | CC BY-NC-ND | 33582702 | 19,021,629 | 2021-02-14 |
What was the administration route of drug 'FLUTICASONE PROPIONATE\SALMETEROL XINAFOATE'? | Case Report: A 64-Year-Old Man with 10-Year History of Eosinophilic Granulomatosis with Polyangiitis with Bronchiectasis and Severe Klebsiella pneumonia.
BACKGROUND Eosinophilic granulomatosis with polyangiitis (EGPA) is a rare and severe progressive disease with multiple clinical manifestations and organ damage. Usually, it requires long-term monitoring of the state of many organs due to the gradual character of its manifestations. CASE REPORT We report a case of a long-term follow-up of a patient with eosinophilic granulomatosis with polyangiitis with emphasis on specific clinical features in this patient. A 64-year-old man was being followed up for 10 years. The initial diagnosis was allergic bronchial asthma; however, as new clinical signs and symptoms developed, the diagnosis of EGPA became obvious. A positive treatment response was seen, mainly manifested as reduced polyneuropathy. Meanwhile, bronchial asthma remained uncontrolled and bronchiectasis and Klebsiella pneumoniae colonization developed despite the combination treatment with prednisolone and methotrexate. Furthermore, the patient suffered a cerebral ischemic infarction. During the last hospital admission, severe uncontrolled bronchial asthma complicated with pneumonia resulted in the patient's death. CONCLUSIONS This clinical case shows the gradual development of EGPA with multiple-organ involvement, including respiratory manifestations and peripheral and central nervous system damage. Immunosuppressive treatment combined with complications of EGPA could have contributed to severe pneumonia development and death of the patient.
Background
Systemic vasculitides are difficult to diagnose due to the heterogenous clinical manifestations requiring a wide differential diagnostic search. One of the most uncommon vasculitis types is eosinophilic granulomatosis with polyangiitis (EGPA), or Churg-Strauss syndrome [1,2]. This vasculitis is associated with the presence of antineutrophilic cytoplasmic antibodies (ANCA) and is characterized by necrotizing inflammatory lesions of mainly small vessels, as well as eosinophilic and granulomatous inflammation in various organs. Bronchial asthma and eosinophilia are the most common clinical manifestations of EGPA [1–3]. In 2012, according to the current classification of systemic vasculitides, Churg-Strauss syndrome was renamed “Eosinophilic Granulomatosis with Polyangiitis” (EGPA) and classified into the ANCA-associated vasculitides group [3].
EGPA is a rare ANCA-associated vasculitis. The incidence rate is 0.5–6.8 new cases/year per million populations in Europe [2]. Prevalence is higher in northern latitudes compared to southern Europe [4]. The disease occurs most commonly in people 40–60 years of age [2,5]. No sex predominance has been clearly demonstrated [4].
The pathogenesis of the disease is still not completely clear, but the key role is attributed to autoimmune inflammation with the participation of eosinophils, CD8+, and CD4+lymphocytes (predominantly Th-2) and their proinflammatory cytokines: interleukin (IL)4, IL5, and IL13 [2,5]. Elevation of immunoglobulin (Ig)E, IgG4, and eotaxin-3 is common in EGPA patients [2,5]. The frequent (in 30–40% of cases) detection of ANCA (mainly antibodies to myeloperoxidase) also suggests the autoimmune nature of the disease. ANCA-positive patients are more commonly diagnosed with glomerulonephritis and mononeuritis, whereas the ANCA-negative ones more frequently demonstrate endocardial, myocardial, and pulmonary eosinophilic infiltrates [1,2,5,6].
There are 3 phases in the development of EGPA (these phases partially overlap and may not appear in sequence):
the allergic phase, distinguished by allergic upper respiratory tract inflammation (rhinosinusitis, allergic rhinitis) and bronchial asthma;
the eosinophilic phase, manifested with severe eosinophilia in combination with multiple eosinophilic infiltrates in various organs (most often in the lungs and gastrointestinal tract);
the vasculitis phase, with small- and medium-vessel inflammation of different organs, which can often be life-threatening [5,6].
Respiratory manifestations of EGPA include allergic rhinitis, sinusitis, nasal polyposis and, in more than 95% of patients, poorly controlled bronchial asthma [5]. Chest radiography findings include transient and migratory opacities, ground-glass opacity, focal consolidation, and small peribronchial and centrilobular nodules or masses [7]. Computed tomography reveals bronchial abnormalities, most commonly thickened wall and dilatation with secondary bronchiectasis formation, leading to frequent infections [7,8]. Pleural effusions and alveolar hemorrhages are uncommon manifestations of EGPA [1,5,7,8].
A characteristic neurological manifestation seen in 75% of patients with EGPA is peripheral neuropathy (mononeuritis multiplex) [1,5]. Upper-limb symptoms due to neuropathy, as well as vasculitis in epineural vessels, were more frequently reported in the ANCA-positive group [9]. Eosinophil-associated tissue damage and vascular occlusion leading to ischemia are more specific in ANCA-negative patients [9]. Also, the tissue damage caused by eosinophils may be associated with IL-5 concentration [10]. Brain hemorrhage, subarachnoid hemorrhage, and ischemic brain infarction are rare [1,5,6].
The main drugs for the treatment of EGPA are glucocorticoids. In organ- or life-threatening manifestations, the recommended starting dose of prednisone is 1 mg/kg/day with a maximal daily dose of 80 mg for 2–3 weeks, with tapering to a minimal supporting dose selected individually to control the symptoms and minimize the adverse effects of treatment. Methylprednisolone (7.5–15 mg/kg/day) can be used as a pulse therapy for life-threatening manifestations [11,12]. Most patients need prolonged or lifelong steroids. In the case of organ- or life-threatening manifestations, glucocorticoids in combination with cyclophosphamide are prescribed [11,12]. The combination of glucocorticoids and methotrexate or mycophenolate is required to induce remission in mild or moderate disease [11,12]. Mepolizumab (a humanized monoclonal antibody targeting IL-5) has been used for EGPA treatment [11]. For ANCA-positive patients with renal involvement or refractory disease, rituximab can be considered [3, 11].
Due to the rare occurrence of EGPA and the variety of its clinical manifestations, the diagnosis and treatment of this disease can be challenging. Currently, reports of long-term observations with the detailed patient follow-up are of great interest. The aim of our case report is to present rare manifestations of EGPA, such as bronchiectasis and stroke. We also focused on the infectious complications of immunosuppressive treatments. Thus, we would like to present a 10-year observation of a patient with EGPA.
Case Report
The male patient, born in 1953, was first hospitalized in the Pulmonary Department of our clinic in 2007 due to exacerbated asthma. The patient’s history was remarkable for multiple allergies (to house dust, cat and dog epidermis, and plant pollen) as well as nonsteroidal anti-inflammatory drugs intolerance (manifested as bronchospasm). The patient denied unhealthy habits and occupational hazards (he worked as a driver). His family history was not remarkable. The patient has had allergic rhinitis since adolescence. The first asthma symptoms appeared at the age of 18 while serving in the army. At the age of 22, an allergist diagnosed him with bronchial asthma. The patient had annual exacerbations (1–2 times a year) due to cold or allergen exposure, and acute respiratory viral infections. At the age of 42, polypoid rhinosinusitis was found, and a polypectomy was performed. Since 2006, the patient had been on controller treatment with salmeterol/fluticasone inhaler (25/250 mg, respectively) 2 doses 2 times a day.
In late 2009, extremely elevated eosinophils (53%) and total IgE (1000 IU/L) were found for the first time. Also, since 2009, the patient has had transient lung opacities, which usually disappeared after treatment with prednisolone 30 mg per day.
In August 2010, the patient noticed numbness on the back surface of the left foot, which spread to the entire left lower leg in 1 month, as well as numbness of the right foot and lower leg. Later, the patient started having tingling, twisting, and burning sensations aggravated by touching in the feet and lower legs. Also, the weakness in the distal parts of the legs developed, and he started having difficulties moving around on his own and falling while walking, and he was subsequently hospitalized in our clinic due to exacerbated asthma. On laboratory testing, high eosinophils (53%) and gamma-globulins (27.9%) were found in November 2010. The chest radiography showed emphysema and consolidation in the middle lobe of the right lung. A Neurology consult confirmed asymmetrical distal sensory-motor polyneuropathy demonstrated by lower distal asymmetrical paraparesis with decreased muscle strength in the foot flexors and extensors, more severe on the left side, low tone of the shin muscles, hypoactive Achilles reflexes, and reduced leg sensitivity starting from the middle of the shin and below, more severe in the peroneal nerve inner-vation area. The patient also demonstrated abnormal coordination (heel-to-knee, Romberg) test results and bilateral steppage gait with knee overextension. Electroneuromyography showed axonal lesion of all distal leg nerves and secondary myelinopathy. Thus, EGPA was suspected. The ANCA (anti-myeloperoxidase) titer was 30 U/ml (normal reference below 20 U/ml).
Allergic bronchopulmonary aspergillosis and other causes of hypereosinophilia were consistently ruled out. IgE and IgG for Aspergillus spp. and Aspergillus spp. in sputum were not found.
Thus, the presence of severe bronchial asthma, rhinosinusitis, high and persistent eosinophilia, transient pulmonary opacities, and polyneuropathy suggested EGPA (5 out of 6 clinical criteria of American College of Rheumatology present). Based on the clinical presentation and increased ANCA titer, the patient was diagnosed with EGPA. Systemic steroid treatment (prednisolone 300 mg intravenously and 30 mg orally) was prescribed, and 5 plasmapheresis sessions were performed. Later, prolonged combination immunosuppression (prednisolone 10 mg/day and methotrexate 25 mg/week) and controller asthma treatment (fluticasone propionate/salmeterol 250/25 µg 4 doses/ day, tiotropium bromide 18 µg per day) were prescribed. After 1 month on prednisolone and methotrexate, there was a significant neurological improvement: the motor and sensory disturbances decreased, and the patient was able to move around on his own. Further on, polyneuropathy completely resolved on the combination treatment. His asthma remained uncontrolled despite the combination treatment: he had frequent (3 times a year) severe exacerbations requiring hospitalization. In 2015, newly-emerged copious purulent sputum associated with frequent infection-related asthma exacerbations was noted. In January 2016, a patient underwent chest computed tomography (CT), which showed generalized bronchiectasis. Stenosis of the proximal bronchi was ruled out also by fibrobronchos-copy. The bronchial secretion microbiology revealed Klebsiella pneumoniae growth (107 colony-forming units).
In November 2016, during the asthma exacerbation, the patient suffered a cerebral ischemic infarction involving the right occipital lobe, confirmed by CT (Figures 1, 2).
On March 2, 2017, the patient was admitted to our hospital due to severe respiratory failure caused by pneumonia in the left upper lobe and severe asthma exacerbation.
The complete blood count showed high WBCs (15.2×109/l) with left shift. The IgE level was 144.0 IU/l, C-reactive protein was 48.8 mg/dL. Clinical chemistry was remarkable for urea nitrogen 18.4 mmol/l and creatinine 1.55 mg/dL. The sputum microscopy demonstrated 100 WBCs and 160 RBCs per vision field, and no eosinophils were revealed. A chest CT revealed multiple bronchiectasis and upper lobar pneumonia of the left lung (Figure 2). Klebsiella was again detected in the sputum culture.
The patient was prescribed intravenous antibiotics (ceftriaxone 2 g/day and ciprofloxacin 800 mg/day replaced by cefopera-zone 4 g/day and amikacin 1 g/day after 3 days due to low efficacy), bronchodilator and anti-inflammatory therapy (nebulized ipratropium bromide, fenoterol, and budesonide 2000 µg per day, prednisolone intravenously 150 mg and orally 30 mg). Despite the aggressive treatment, the respiratory failure progressed, and on day 6 of hospitalization the patient was transferred to the Intensive Care Unit and intubated. On day 7, the patient died. On autopsy, the EGPA diagnosis was confirmed, signs of ischemic infarction in the occipital lobe of the brain were found, and left upper lobe pneumonia with abscess formation and generalized bilateral bronchiectases were revealed.
Discussion
Based on the long follow-up period and careful interpretation of signs and symptoms, typical laboratory abnormalities, the diagnosis was clear. Among the most commonly used diagnostic criteria of EGPA are the American College of Rheumatology (ACR) classification criteria, Lanham’s criteria, and the Chapel Hill Consensus Conference criteria. The presence of 4 or more ACR classification criteria, including bronchial asthma, eosinophilia (more than 10%), paranasal sinus abnormalities, migratory or transient eosinophilic pulmonary infiltrates, extravascular eosinophil infiltration on biopsy, and mono- or polyneuropathy, has a sensitivity of 85% and a specificity of 100% [2,5,6]. We identified 5 out of 6 American College of Rheumatology clinical criteria present in our patient.
This clinical case shows gradual development of EGPA with multiple-organ involvement (see Figure 3). The disease onset was characterized by respiratory manifestations and suspicion of EGPA appeared only after the development of polyneuropathy (after 39 years) with hypereosinophilia. Polyneuropathy was noted at the age of 57, which is slightly later than the average age of onset reported in the literature [13]. Notably, the patient was ANCA-positive and his long-term neurological manifestation (mononeuritis) was quite typical for ANCA vasculitis [5,6,8,13]. The diagnosis became obvious after comprehensive assessment of the disease history with its gradually evolving signs and symptoms and laboratory abnormalities. Almost all manifestations of the disease seen in the patient were typical except for stroke, which is an exceptional feature for this case [1,5,6]. Neurological involvement manifested as ischemic stroke is an uncommon complication of EGPA (seen only in 5.2% of all patients) [1]. Thus, the patient demonstrated both peripheral and central nervous system involvement, which is unusual and life-threatening in EGPA.
This clinical case of EGPA also attracted our attention due to severe respiratory manifestations. While typical respiratory manifestations such as rhinosinusitis, asthma, and transient lung infiltrates are well documented in the literature, EGPA-associated bronchiectasis is rarely mentioned [14]. Bronchiectasis in the patient was detected by CT at the age of 63. The patient was a nonsmoker, and no other cause of bronchial abnormalities and airflow obstruction could be identified. Stenosis of the proximal bronchi was ruled out by fibrobronchoscopy and CT. The diagnosis of allergic bronchopulmonary aspergillosis was not confirmed. Klebsiella pneumoniae colonization was found during the last 2 years of the patient’s life. Klebsiella pneumoniae is a typical pathogen causing bronchiectasis [15]. It should be noted that aspergillosis and mycobacteriosis are common complications of bronchiectasis [15], but neither was identified in our patient.
The patient was treated for a long time with a combination of prednisolone and methotrexate. There was a significant positive effect on polyneuropathy, but uncontrolled bronchial asthma persisted. This treatment regimen complies with current recommendations [3,11]. Patients with life-threatening and/or organ-threatening manifestations (including lesions of the central nervous system and severe peripheral neuropathy) require therapy with a combination of a glucocorticoid and a cytostatic [3,11].
The use of immunosuppressive drugs in patients with ANCA-associated vasculitis significantly increases the risk of infection and mortality [16–18]. Prednisone and methotrexate use were found to be significant predictors of hospitalization for pneumonia. Frequent infections (more than 1 episode) are reported in patients with ANCA-associated vasculitis during the first year of diagnosis and initiation of therapy [18]. We assume that bronchiectasis complicated by bacterial colonization and bronchial obstruction in combination with immuno-suppression led to severe pneumonia and death of the patient.
Conclusions
The present case report demonstrates a long-term follow-up of a patient with EGPA. Both common and rare manifestations of EGPA have been described in the patient, such as central nervous system damage and bronchiectasis. Immunosuppressive treatment probably contributed to Klebsiella infection development. Thus, long-term complications of EGPA in combination with adverse effects of medications could have caused the severe pneumonia and death of the patient.
Figure 1. Brain multi-spiral computed tomography performed in 2016 showing ischemic brain damage in the right occipital lobe.
Figure 2. (A–C) Chest computed tomography of patient showing large left upper lobe pneumonia and bronchiectasis in lover lobes.
Figure 3. Timeline from first symptoms to death. | Respiratory (inhalation) | DrugAdministrationRoute | CC BY-NC-ND | 33582702 | 18,986,567 | 2021-02-14 |
What was the administration route of drug 'IPRATROPIUM BROMIDE'? | Case Report: A 64-Year-Old Man with 10-Year History of Eosinophilic Granulomatosis with Polyangiitis with Bronchiectasis and Severe Klebsiella pneumonia.
BACKGROUND Eosinophilic granulomatosis with polyangiitis (EGPA) is a rare and severe progressive disease with multiple clinical manifestations and organ damage. Usually, it requires long-term monitoring of the state of many organs due to the gradual character of its manifestations. CASE REPORT We report a case of a long-term follow-up of a patient with eosinophilic granulomatosis with polyangiitis with emphasis on specific clinical features in this patient. A 64-year-old man was being followed up for 10 years. The initial diagnosis was allergic bronchial asthma; however, as new clinical signs and symptoms developed, the diagnosis of EGPA became obvious. A positive treatment response was seen, mainly manifested as reduced polyneuropathy. Meanwhile, bronchial asthma remained uncontrolled and bronchiectasis and Klebsiella pneumoniae colonization developed despite the combination treatment with prednisolone and methotrexate. Furthermore, the patient suffered a cerebral ischemic infarction. During the last hospital admission, severe uncontrolled bronchial asthma complicated with pneumonia resulted in the patient's death. CONCLUSIONS This clinical case shows the gradual development of EGPA with multiple-organ involvement, including respiratory manifestations and peripheral and central nervous system damage. Immunosuppressive treatment combined with complications of EGPA could have contributed to severe pneumonia development and death of the patient.
Background
Systemic vasculitides are difficult to diagnose due to the heterogenous clinical manifestations requiring a wide differential diagnostic search. One of the most uncommon vasculitis types is eosinophilic granulomatosis with polyangiitis (EGPA), or Churg-Strauss syndrome [1,2]. This vasculitis is associated with the presence of antineutrophilic cytoplasmic antibodies (ANCA) and is characterized by necrotizing inflammatory lesions of mainly small vessels, as well as eosinophilic and granulomatous inflammation in various organs. Bronchial asthma and eosinophilia are the most common clinical manifestations of EGPA [1–3]. In 2012, according to the current classification of systemic vasculitides, Churg-Strauss syndrome was renamed “Eosinophilic Granulomatosis with Polyangiitis” (EGPA) and classified into the ANCA-associated vasculitides group [3].
EGPA is a rare ANCA-associated vasculitis. The incidence rate is 0.5–6.8 new cases/year per million populations in Europe [2]. Prevalence is higher in northern latitudes compared to southern Europe [4]. The disease occurs most commonly in people 40–60 years of age [2,5]. No sex predominance has been clearly demonstrated [4].
The pathogenesis of the disease is still not completely clear, but the key role is attributed to autoimmune inflammation with the participation of eosinophils, CD8+, and CD4+lymphocytes (predominantly Th-2) and their proinflammatory cytokines: interleukin (IL)4, IL5, and IL13 [2,5]. Elevation of immunoglobulin (Ig)E, IgG4, and eotaxin-3 is common in EGPA patients [2,5]. The frequent (in 30–40% of cases) detection of ANCA (mainly antibodies to myeloperoxidase) also suggests the autoimmune nature of the disease. ANCA-positive patients are more commonly diagnosed with glomerulonephritis and mononeuritis, whereas the ANCA-negative ones more frequently demonstrate endocardial, myocardial, and pulmonary eosinophilic infiltrates [1,2,5,6].
There are 3 phases in the development of EGPA (these phases partially overlap and may not appear in sequence):
the allergic phase, distinguished by allergic upper respiratory tract inflammation (rhinosinusitis, allergic rhinitis) and bronchial asthma;
the eosinophilic phase, manifested with severe eosinophilia in combination with multiple eosinophilic infiltrates in various organs (most often in the lungs and gastrointestinal tract);
the vasculitis phase, with small- and medium-vessel inflammation of different organs, which can often be life-threatening [5,6].
Respiratory manifestations of EGPA include allergic rhinitis, sinusitis, nasal polyposis and, in more than 95% of patients, poorly controlled bronchial asthma [5]. Chest radiography findings include transient and migratory opacities, ground-glass opacity, focal consolidation, and small peribronchial and centrilobular nodules or masses [7]. Computed tomography reveals bronchial abnormalities, most commonly thickened wall and dilatation with secondary bronchiectasis formation, leading to frequent infections [7,8]. Pleural effusions and alveolar hemorrhages are uncommon manifestations of EGPA [1,5,7,8].
A characteristic neurological manifestation seen in 75% of patients with EGPA is peripheral neuropathy (mononeuritis multiplex) [1,5]. Upper-limb symptoms due to neuropathy, as well as vasculitis in epineural vessels, were more frequently reported in the ANCA-positive group [9]. Eosinophil-associated tissue damage and vascular occlusion leading to ischemia are more specific in ANCA-negative patients [9]. Also, the tissue damage caused by eosinophils may be associated with IL-5 concentration [10]. Brain hemorrhage, subarachnoid hemorrhage, and ischemic brain infarction are rare [1,5,6].
The main drugs for the treatment of EGPA are glucocorticoids. In organ- or life-threatening manifestations, the recommended starting dose of prednisone is 1 mg/kg/day with a maximal daily dose of 80 mg for 2–3 weeks, with tapering to a minimal supporting dose selected individually to control the symptoms and minimize the adverse effects of treatment. Methylprednisolone (7.5–15 mg/kg/day) can be used as a pulse therapy for life-threatening manifestations [11,12]. Most patients need prolonged or lifelong steroids. In the case of organ- or life-threatening manifestations, glucocorticoids in combination with cyclophosphamide are prescribed [11,12]. The combination of glucocorticoids and methotrexate or mycophenolate is required to induce remission in mild or moderate disease [11,12]. Mepolizumab (a humanized monoclonal antibody targeting IL-5) has been used for EGPA treatment [11]. For ANCA-positive patients with renal involvement or refractory disease, rituximab can be considered [3, 11].
Due to the rare occurrence of EGPA and the variety of its clinical manifestations, the diagnosis and treatment of this disease can be challenging. Currently, reports of long-term observations with the detailed patient follow-up are of great interest. The aim of our case report is to present rare manifestations of EGPA, such as bronchiectasis and stroke. We also focused on the infectious complications of immunosuppressive treatments. Thus, we would like to present a 10-year observation of a patient with EGPA.
Case Report
The male patient, born in 1953, was first hospitalized in the Pulmonary Department of our clinic in 2007 due to exacerbated asthma. The patient’s history was remarkable for multiple allergies (to house dust, cat and dog epidermis, and plant pollen) as well as nonsteroidal anti-inflammatory drugs intolerance (manifested as bronchospasm). The patient denied unhealthy habits and occupational hazards (he worked as a driver). His family history was not remarkable. The patient has had allergic rhinitis since adolescence. The first asthma symptoms appeared at the age of 18 while serving in the army. At the age of 22, an allergist diagnosed him with bronchial asthma. The patient had annual exacerbations (1–2 times a year) due to cold or allergen exposure, and acute respiratory viral infections. At the age of 42, polypoid rhinosinusitis was found, and a polypectomy was performed. Since 2006, the patient had been on controller treatment with salmeterol/fluticasone inhaler (25/250 mg, respectively) 2 doses 2 times a day.
In late 2009, extremely elevated eosinophils (53%) and total IgE (1000 IU/L) were found for the first time. Also, since 2009, the patient has had transient lung opacities, which usually disappeared after treatment with prednisolone 30 mg per day.
In August 2010, the patient noticed numbness on the back surface of the left foot, which spread to the entire left lower leg in 1 month, as well as numbness of the right foot and lower leg. Later, the patient started having tingling, twisting, and burning sensations aggravated by touching in the feet and lower legs. Also, the weakness in the distal parts of the legs developed, and he started having difficulties moving around on his own and falling while walking, and he was subsequently hospitalized in our clinic due to exacerbated asthma. On laboratory testing, high eosinophils (53%) and gamma-globulins (27.9%) were found in November 2010. The chest radiography showed emphysema and consolidation in the middle lobe of the right lung. A Neurology consult confirmed asymmetrical distal sensory-motor polyneuropathy demonstrated by lower distal asymmetrical paraparesis with decreased muscle strength in the foot flexors and extensors, more severe on the left side, low tone of the shin muscles, hypoactive Achilles reflexes, and reduced leg sensitivity starting from the middle of the shin and below, more severe in the peroneal nerve inner-vation area. The patient also demonstrated abnormal coordination (heel-to-knee, Romberg) test results and bilateral steppage gait with knee overextension. Electroneuromyography showed axonal lesion of all distal leg nerves and secondary myelinopathy. Thus, EGPA was suspected. The ANCA (anti-myeloperoxidase) titer was 30 U/ml (normal reference below 20 U/ml).
Allergic bronchopulmonary aspergillosis and other causes of hypereosinophilia were consistently ruled out. IgE and IgG for Aspergillus spp. and Aspergillus spp. in sputum were not found.
Thus, the presence of severe bronchial asthma, rhinosinusitis, high and persistent eosinophilia, transient pulmonary opacities, and polyneuropathy suggested EGPA (5 out of 6 clinical criteria of American College of Rheumatology present). Based on the clinical presentation and increased ANCA titer, the patient was diagnosed with EGPA. Systemic steroid treatment (prednisolone 300 mg intravenously and 30 mg orally) was prescribed, and 5 plasmapheresis sessions were performed. Later, prolonged combination immunosuppression (prednisolone 10 mg/day and methotrexate 25 mg/week) and controller asthma treatment (fluticasone propionate/salmeterol 250/25 µg 4 doses/ day, tiotropium bromide 18 µg per day) were prescribed. After 1 month on prednisolone and methotrexate, there was a significant neurological improvement: the motor and sensory disturbances decreased, and the patient was able to move around on his own. Further on, polyneuropathy completely resolved on the combination treatment. His asthma remained uncontrolled despite the combination treatment: he had frequent (3 times a year) severe exacerbations requiring hospitalization. In 2015, newly-emerged copious purulent sputum associated with frequent infection-related asthma exacerbations was noted. In January 2016, a patient underwent chest computed tomography (CT), which showed generalized bronchiectasis. Stenosis of the proximal bronchi was ruled out also by fibrobronchos-copy. The bronchial secretion microbiology revealed Klebsiella pneumoniae growth (107 colony-forming units).
In November 2016, during the asthma exacerbation, the patient suffered a cerebral ischemic infarction involving the right occipital lobe, confirmed by CT (Figures 1, 2).
On March 2, 2017, the patient was admitted to our hospital due to severe respiratory failure caused by pneumonia in the left upper lobe and severe asthma exacerbation.
The complete blood count showed high WBCs (15.2×109/l) with left shift. The IgE level was 144.0 IU/l, C-reactive protein was 48.8 mg/dL. Clinical chemistry was remarkable for urea nitrogen 18.4 mmol/l and creatinine 1.55 mg/dL. The sputum microscopy demonstrated 100 WBCs and 160 RBCs per vision field, and no eosinophils were revealed. A chest CT revealed multiple bronchiectasis and upper lobar pneumonia of the left lung (Figure 2). Klebsiella was again detected in the sputum culture.
The patient was prescribed intravenous antibiotics (ceftriaxone 2 g/day and ciprofloxacin 800 mg/day replaced by cefopera-zone 4 g/day and amikacin 1 g/day after 3 days due to low efficacy), bronchodilator and anti-inflammatory therapy (nebulized ipratropium bromide, fenoterol, and budesonide 2000 µg per day, prednisolone intravenously 150 mg and orally 30 mg). Despite the aggressive treatment, the respiratory failure progressed, and on day 6 of hospitalization the patient was transferred to the Intensive Care Unit and intubated. On day 7, the patient died. On autopsy, the EGPA diagnosis was confirmed, signs of ischemic infarction in the occipital lobe of the brain were found, and left upper lobe pneumonia with abscess formation and generalized bilateral bronchiectases were revealed.
Discussion
Based on the long follow-up period and careful interpretation of signs and symptoms, typical laboratory abnormalities, the diagnosis was clear. Among the most commonly used diagnostic criteria of EGPA are the American College of Rheumatology (ACR) classification criteria, Lanham’s criteria, and the Chapel Hill Consensus Conference criteria. The presence of 4 or more ACR classification criteria, including bronchial asthma, eosinophilia (more than 10%), paranasal sinus abnormalities, migratory or transient eosinophilic pulmonary infiltrates, extravascular eosinophil infiltration on biopsy, and mono- or polyneuropathy, has a sensitivity of 85% and a specificity of 100% [2,5,6]. We identified 5 out of 6 American College of Rheumatology clinical criteria present in our patient.
This clinical case shows gradual development of EGPA with multiple-organ involvement (see Figure 3). The disease onset was characterized by respiratory manifestations and suspicion of EGPA appeared only after the development of polyneuropathy (after 39 years) with hypereosinophilia. Polyneuropathy was noted at the age of 57, which is slightly later than the average age of onset reported in the literature [13]. Notably, the patient was ANCA-positive and his long-term neurological manifestation (mononeuritis) was quite typical for ANCA vasculitis [5,6,8,13]. The diagnosis became obvious after comprehensive assessment of the disease history with its gradually evolving signs and symptoms and laboratory abnormalities. Almost all manifestations of the disease seen in the patient were typical except for stroke, which is an exceptional feature for this case [1,5,6]. Neurological involvement manifested as ischemic stroke is an uncommon complication of EGPA (seen only in 5.2% of all patients) [1]. Thus, the patient demonstrated both peripheral and central nervous system involvement, which is unusual and life-threatening in EGPA.
This clinical case of EGPA also attracted our attention due to severe respiratory manifestations. While typical respiratory manifestations such as rhinosinusitis, asthma, and transient lung infiltrates are well documented in the literature, EGPA-associated bronchiectasis is rarely mentioned [14]. Bronchiectasis in the patient was detected by CT at the age of 63. The patient was a nonsmoker, and no other cause of bronchial abnormalities and airflow obstruction could be identified. Stenosis of the proximal bronchi was ruled out by fibrobronchoscopy and CT. The diagnosis of allergic bronchopulmonary aspergillosis was not confirmed. Klebsiella pneumoniae colonization was found during the last 2 years of the patient’s life. Klebsiella pneumoniae is a typical pathogen causing bronchiectasis [15]. It should be noted that aspergillosis and mycobacteriosis are common complications of bronchiectasis [15], but neither was identified in our patient.
The patient was treated for a long time with a combination of prednisolone and methotrexate. There was a significant positive effect on polyneuropathy, but uncontrolled bronchial asthma persisted. This treatment regimen complies with current recommendations [3,11]. Patients with life-threatening and/or organ-threatening manifestations (including lesions of the central nervous system and severe peripheral neuropathy) require therapy with a combination of a glucocorticoid and a cytostatic [3,11].
The use of immunosuppressive drugs in patients with ANCA-associated vasculitis significantly increases the risk of infection and mortality [16–18]. Prednisone and methotrexate use were found to be significant predictors of hospitalization for pneumonia. Frequent infections (more than 1 episode) are reported in patients with ANCA-associated vasculitis during the first year of diagnosis and initiation of therapy [18]. We assume that bronchiectasis complicated by bacterial colonization and bronchial obstruction in combination with immuno-suppression led to severe pneumonia and death of the patient.
Conclusions
The present case report demonstrates a long-term follow-up of a patient with EGPA. Both common and rare manifestations of EGPA have been described in the patient, such as central nervous system damage and bronchiectasis. Immunosuppressive treatment probably contributed to Klebsiella infection development. Thus, long-term complications of EGPA in combination with adverse effects of medications could have caused the severe pneumonia and death of the patient.
Figure 1. Brain multi-spiral computed tomography performed in 2016 showing ischemic brain damage in the right occipital lobe.
Figure 2. (A–C) Chest computed tomography of patient showing large left upper lobe pneumonia and bronchiectasis in lover lobes.
Figure 3. Timeline from first symptoms to death. | Respiratory (inhalation) | DrugAdministrationRoute | CC BY-NC-ND | 33582702 | 18,986,567 | 2021-02-14 |
What was the dosage of drug 'AMIKACIN SULFATE'? | Case Report: A 64-Year-Old Man with 10-Year History of Eosinophilic Granulomatosis with Polyangiitis with Bronchiectasis and Severe Klebsiella pneumonia.
BACKGROUND Eosinophilic granulomatosis with polyangiitis (EGPA) is a rare and severe progressive disease with multiple clinical manifestations and organ damage. Usually, it requires long-term monitoring of the state of many organs due to the gradual character of its manifestations. CASE REPORT We report a case of a long-term follow-up of a patient with eosinophilic granulomatosis with polyangiitis with emphasis on specific clinical features in this patient. A 64-year-old man was being followed up for 10 years. The initial diagnosis was allergic bronchial asthma; however, as new clinical signs and symptoms developed, the diagnosis of EGPA became obvious. A positive treatment response was seen, mainly manifested as reduced polyneuropathy. Meanwhile, bronchial asthma remained uncontrolled and bronchiectasis and Klebsiella pneumoniae colonization developed despite the combination treatment with prednisolone and methotrexate. Furthermore, the patient suffered a cerebral ischemic infarction. During the last hospital admission, severe uncontrolled bronchial asthma complicated with pneumonia resulted in the patient's death. CONCLUSIONS This clinical case shows the gradual development of EGPA with multiple-organ involvement, including respiratory manifestations and peripheral and central nervous system damage. Immunosuppressive treatment combined with complications of EGPA could have contributed to severe pneumonia development and death of the patient.
Background
Systemic vasculitides are difficult to diagnose due to the heterogenous clinical manifestations requiring a wide differential diagnostic search. One of the most uncommon vasculitis types is eosinophilic granulomatosis with polyangiitis (EGPA), or Churg-Strauss syndrome [1,2]. This vasculitis is associated with the presence of antineutrophilic cytoplasmic antibodies (ANCA) and is characterized by necrotizing inflammatory lesions of mainly small vessels, as well as eosinophilic and granulomatous inflammation in various organs. Bronchial asthma and eosinophilia are the most common clinical manifestations of EGPA [1–3]. In 2012, according to the current classification of systemic vasculitides, Churg-Strauss syndrome was renamed “Eosinophilic Granulomatosis with Polyangiitis” (EGPA) and classified into the ANCA-associated vasculitides group [3].
EGPA is a rare ANCA-associated vasculitis. The incidence rate is 0.5–6.8 new cases/year per million populations in Europe [2]. Prevalence is higher in northern latitudes compared to southern Europe [4]. The disease occurs most commonly in people 40–60 years of age [2,5]. No sex predominance has been clearly demonstrated [4].
The pathogenesis of the disease is still not completely clear, but the key role is attributed to autoimmune inflammation with the participation of eosinophils, CD8+, and CD4+lymphocytes (predominantly Th-2) and their proinflammatory cytokines: interleukin (IL)4, IL5, and IL13 [2,5]. Elevation of immunoglobulin (Ig)E, IgG4, and eotaxin-3 is common in EGPA patients [2,5]. The frequent (in 30–40% of cases) detection of ANCA (mainly antibodies to myeloperoxidase) also suggests the autoimmune nature of the disease. ANCA-positive patients are more commonly diagnosed with glomerulonephritis and mononeuritis, whereas the ANCA-negative ones more frequently demonstrate endocardial, myocardial, and pulmonary eosinophilic infiltrates [1,2,5,6].
There are 3 phases in the development of EGPA (these phases partially overlap and may not appear in sequence):
the allergic phase, distinguished by allergic upper respiratory tract inflammation (rhinosinusitis, allergic rhinitis) and bronchial asthma;
the eosinophilic phase, manifested with severe eosinophilia in combination with multiple eosinophilic infiltrates in various organs (most often in the lungs and gastrointestinal tract);
the vasculitis phase, with small- and medium-vessel inflammation of different organs, which can often be life-threatening [5,6].
Respiratory manifestations of EGPA include allergic rhinitis, sinusitis, nasal polyposis and, in more than 95% of patients, poorly controlled bronchial asthma [5]. Chest radiography findings include transient and migratory opacities, ground-glass opacity, focal consolidation, and small peribronchial and centrilobular nodules or masses [7]. Computed tomography reveals bronchial abnormalities, most commonly thickened wall and dilatation with secondary bronchiectasis formation, leading to frequent infections [7,8]. Pleural effusions and alveolar hemorrhages are uncommon manifestations of EGPA [1,5,7,8].
A characteristic neurological manifestation seen in 75% of patients with EGPA is peripheral neuropathy (mononeuritis multiplex) [1,5]. Upper-limb symptoms due to neuropathy, as well as vasculitis in epineural vessels, were more frequently reported in the ANCA-positive group [9]. Eosinophil-associated tissue damage and vascular occlusion leading to ischemia are more specific in ANCA-negative patients [9]. Also, the tissue damage caused by eosinophils may be associated with IL-5 concentration [10]. Brain hemorrhage, subarachnoid hemorrhage, and ischemic brain infarction are rare [1,5,6].
The main drugs for the treatment of EGPA are glucocorticoids. In organ- or life-threatening manifestations, the recommended starting dose of prednisone is 1 mg/kg/day with a maximal daily dose of 80 mg for 2–3 weeks, with tapering to a minimal supporting dose selected individually to control the symptoms and minimize the adverse effects of treatment. Methylprednisolone (7.5–15 mg/kg/day) can be used as a pulse therapy for life-threatening manifestations [11,12]. Most patients need prolonged or lifelong steroids. In the case of organ- or life-threatening manifestations, glucocorticoids in combination with cyclophosphamide are prescribed [11,12]. The combination of glucocorticoids and methotrexate or mycophenolate is required to induce remission in mild or moderate disease [11,12]. Mepolizumab (a humanized monoclonal antibody targeting IL-5) has been used for EGPA treatment [11]. For ANCA-positive patients with renal involvement or refractory disease, rituximab can be considered [3, 11].
Due to the rare occurrence of EGPA and the variety of its clinical manifestations, the diagnosis and treatment of this disease can be challenging. Currently, reports of long-term observations with the detailed patient follow-up are of great interest. The aim of our case report is to present rare manifestations of EGPA, such as bronchiectasis and stroke. We also focused on the infectious complications of immunosuppressive treatments. Thus, we would like to present a 10-year observation of a patient with EGPA.
Case Report
The male patient, born in 1953, was first hospitalized in the Pulmonary Department of our clinic in 2007 due to exacerbated asthma. The patient’s history was remarkable for multiple allergies (to house dust, cat and dog epidermis, and plant pollen) as well as nonsteroidal anti-inflammatory drugs intolerance (manifested as bronchospasm). The patient denied unhealthy habits and occupational hazards (he worked as a driver). His family history was not remarkable. The patient has had allergic rhinitis since adolescence. The first asthma symptoms appeared at the age of 18 while serving in the army. At the age of 22, an allergist diagnosed him with bronchial asthma. The patient had annual exacerbations (1–2 times a year) due to cold or allergen exposure, and acute respiratory viral infections. At the age of 42, polypoid rhinosinusitis was found, and a polypectomy was performed. Since 2006, the patient had been on controller treatment with salmeterol/fluticasone inhaler (25/250 mg, respectively) 2 doses 2 times a day.
In late 2009, extremely elevated eosinophils (53%) and total IgE (1000 IU/L) were found for the first time. Also, since 2009, the patient has had transient lung opacities, which usually disappeared after treatment with prednisolone 30 mg per day.
In August 2010, the patient noticed numbness on the back surface of the left foot, which spread to the entire left lower leg in 1 month, as well as numbness of the right foot and lower leg. Later, the patient started having tingling, twisting, and burning sensations aggravated by touching in the feet and lower legs. Also, the weakness in the distal parts of the legs developed, and he started having difficulties moving around on his own and falling while walking, and he was subsequently hospitalized in our clinic due to exacerbated asthma. On laboratory testing, high eosinophils (53%) and gamma-globulins (27.9%) were found in November 2010. The chest radiography showed emphysema and consolidation in the middle lobe of the right lung. A Neurology consult confirmed asymmetrical distal sensory-motor polyneuropathy demonstrated by lower distal asymmetrical paraparesis with decreased muscle strength in the foot flexors and extensors, more severe on the left side, low tone of the shin muscles, hypoactive Achilles reflexes, and reduced leg sensitivity starting from the middle of the shin and below, more severe in the peroneal nerve inner-vation area. The patient also demonstrated abnormal coordination (heel-to-knee, Romberg) test results and bilateral steppage gait with knee overextension. Electroneuromyography showed axonal lesion of all distal leg nerves and secondary myelinopathy. Thus, EGPA was suspected. The ANCA (anti-myeloperoxidase) titer was 30 U/ml (normal reference below 20 U/ml).
Allergic bronchopulmonary aspergillosis and other causes of hypereosinophilia were consistently ruled out. IgE and IgG for Aspergillus spp. and Aspergillus spp. in sputum were not found.
Thus, the presence of severe bronchial asthma, rhinosinusitis, high and persistent eosinophilia, transient pulmonary opacities, and polyneuropathy suggested EGPA (5 out of 6 clinical criteria of American College of Rheumatology present). Based on the clinical presentation and increased ANCA titer, the patient was diagnosed with EGPA. Systemic steroid treatment (prednisolone 300 mg intravenously and 30 mg orally) was prescribed, and 5 plasmapheresis sessions were performed. Later, prolonged combination immunosuppression (prednisolone 10 mg/day and methotrexate 25 mg/week) and controller asthma treatment (fluticasone propionate/salmeterol 250/25 µg 4 doses/ day, tiotropium bromide 18 µg per day) were prescribed. After 1 month on prednisolone and methotrexate, there was a significant neurological improvement: the motor and sensory disturbances decreased, and the patient was able to move around on his own. Further on, polyneuropathy completely resolved on the combination treatment. His asthma remained uncontrolled despite the combination treatment: he had frequent (3 times a year) severe exacerbations requiring hospitalization. In 2015, newly-emerged copious purulent sputum associated with frequent infection-related asthma exacerbations was noted. In January 2016, a patient underwent chest computed tomography (CT), which showed generalized bronchiectasis. Stenosis of the proximal bronchi was ruled out also by fibrobronchos-copy. The bronchial secretion microbiology revealed Klebsiella pneumoniae growth (107 colony-forming units).
In November 2016, during the asthma exacerbation, the patient suffered a cerebral ischemic infarction involving the right occipital lobe, confirmed by CT (Figures 1, 2).
On March 2, 2017, the patient was admitted to our hospital due to severe respiratory failure caused by pneumonia in the left upper lobe and severe asthma exacerbation.
The complete blood count showed high WBCs (15.2×109/l) with left shift. The IgE level was 144.0 IU/l, C-reactive protein was 48.8 mg/dL. Clinical chemistry was remarkable for urea nitrogen 18.4 mmol/l and creatinine 1.55 mg/dL. The sputum microscopy demonstrated 100 WBCs and 160 RBCs per vision field, and no eosinophils were revealed. A chest CT revealed multiple bronchiectasis and upper lobar pneumonia of the left lung (Figure 2). Klebsiella was again detected in the sputum culture.
The patient was prescribed intravenous antibiotics (ceftriaxone 2 g/day and ciprofloxacin 800 mg/day replaced by cefopera-zone 4 g/day and amikacin 1 g/day after 3 days due to low efficacy), bronchodilator and anti-inflammatory therapy (nebulized ipratropium bromide, fenoterol, and budesonide 2000 µg per day, prednisolone intravenously 150 mg and orally 30 mg). Despite the aggressive treatment, the respiratory failure progressed, and on day 6 of hospitalization the patient was transferred to the Intensive Care Unit and intubated. On day 7, the patient died. On autopsy, the EGPA diagnosis was confirmed, signs of ischemic infarction in the occipital lobe of the brain were found, and left upper lobe pneumonia with abscess formation and generalized bilateral bronchiectases were revealed.
Discussion
Based on the long follow-up period and careful interpretation of signs and symptoms, typical laboratory abnormalities, the diagnosis was clear. Among the most commonly used diagnostic criteria of EGPA are the American College of Rheumatology (ACR) classification criteria, Lanham’s criteria, and the Chapel Hill Consensus Conference criteria. The presence of 4 or more ACR classification criteria, including bronchial asthma, eosinophilia (more than 10%), paranasal sinus abnormalities, migratory or transient eosinophilic pulmonary infiltrates, extravascular eosinophil infiltration on biopsy, and mono- or polyneuropathy, has a sensitivity of 85% and a specificity of 100% [2,5,6]. We identified 5 out of 6 American College of Rheumatology clinical criteria present in our patient.
This clinical case shows gradual development of EGPA with multiple-organ involvement (see Figure 3). The disease onset was characterized by respiratory manifestations and suspicion of EGPA appeared only after the development of polyneuropathy (after 39 years) with hypereosinophilia. Polyneuropathy was noted at the age of 57, which is slightly later than the average age of onset reported in the literature [13]. Notably, the patient was ANCA-positive and his long-term neurological manifestation (mononeuritis) was quite typical for ANCA vasculitis [5,6,8,13]. The diagnosis became obvious after comprehensive assessment of the disease history with its gradually evolving signs and symptoms and laboratory abnormalities. Almost all manifestations of the disease seen in the patient were typical except for stroke, which is an exceptional feature for this case [1,5,6]. Neurological involvement manifested as ischemic stroke is an uncommon complication of EGPA (seen only in 5.2% of all patients) [1]. Thus, the patient demonstrated both peripheral and central nervous system involvement, which is unusual and life-threatening in EGPA.
This clinical case of EGPA also attracted our attention due to severe respiratory manifestations. While typical respiratory manifestations such as rhinosinusitis, asthma, and transient lung infiltrates are well documented in the literature, EGPA-associated bronchiectasis is rarely mentioned [14]. Bronchiectasis in the patient was detected by CT at the age of 63. The patient was a nonsmoker, and no other cause of bronchial abnormalities and airflow obstruction could be identified. Stenosis of the proximal bronchi was ruled out by fibrobronchoscopy and CT. The diagnosis of allergic bronchopulmonary aspergillosis was not confirmed. Klebsiella pneumoniae colonization was found during the last 2 years of the patient’s life. Klebsiella pneumoniae is a typical pathogen causing bronchiectasis [15]. It should be noted that aspergillosis and mycobacteriosis are common complications of bronchiectasis [15], but neither was identified in our patient.
The patient was treated for a long time with a combination of prednisolone and methotrexate. There was a significant positive effect on polyneuropathy, but uncontrolled bronchial asthma persisted. This treatment regimen complies with current recommendations [3,11]. Patients with life-threatening and/or organ-threatening manifestations (including lesions of the central nervous system and severe peripheral neuropathy) require therapy with a combination of a glucocorticoid and a cytostatic [3,11].
The use of immunosuppressive drugs in patients with ANCA-associated vasculitis significantly increases the risk of infection and mortality [16–18]. Prednisone and methotrexate use were found to be significant predictors of hospitalization for pneumonia. Frequent infections (more than 1 episode) are reported in patients with ANCA-associated vasculitis during the first year of diagnosis and initiation of therapy [18]. We assume that bronchiectasis complicated by bacterial colonization and bronchial obstruction in combination with immuno-suppression led to severe pneumonia and death of the patient.
Conclusions
The present case report demonstrates a long-term follow-up of a patient with EGPA. Both common and rare manifestations of EGPA have been described in the patient, such as central nervous system damage and bronchiectasis. Immunosuppressive treatment probably contributed to Klebsiella infection development. Thus, long-term complications of EGPA in combination with adverse effects of medications could have caused the severe pneumonia and death of the patient.
Figure 1. Brain multi-spiral computed tomography performed in 2016 showing ischemic brain damage in the right occipital lobe.
Figure 2. (A–C) Chest computed tomography of patient showing large left upper lobe pneumonia and bronchiectasis in lover lobes.
Figure 3. Timeline from first symptoms to death. | 1 G, PER DAY | DrugDosageText | CC BY-NC-ND | 33582702 | 18,962,812 | 2021-02-14 |
What was the dosage of drug 'FENOTEROL'? | Case Report: A 64-Year-Old Man with 10-Year History of Eosinophilic Granulomatosis with Polyangiitis with Bronchiectasis and Severe Klebsiella pneumonia.
BACKGROUND Eosinophilic granulomatosis with polyangiitis (EGPA) is a rare and severe progressive disease with multiple clinical manifestations and organ damage. Usually, it requires long-term monitoring of the state of many organs due to the gradual character of its manifestations. CASE REPORT We report a case of a long-term follow-up of a patient with eosinophilic granulomatosis with polyangiitis with emphasis on specific clinical features in this patient. A 64-year-old man was being followed up for 10 years. The initial diagnosis was allergic bronchial asthma; however, as new clinical signs and symptoms developed, the diagnosis of EGPA became obvious. A positive treatment response was seen, mainly manifested as reduced polyneuropathy. Meanwhile, bronchial asthma remained uncontrolled and bronchiectasis and Klebsiella pneumoniae colonization developed despite the combination treatment with prednisolone and methotrexate. Furthermore, the patient suffered a cerebral ischemic infarction. During the last hospital admission, severe uncontrolled bronchial asthma complicated with pneumonia resulted in the patient's death. CONCLUSIONS This clinical case shows the gradual development of EGPA with multiple-organ involvement, including respiratory manifestations and peripheral and central nervous system damage. Immunosuppressive treatment combined with complications of EGPA could have contributed to severe pneumonia development and death of the patient.
Background
Systemic vasculitides are difficult to diagnose due to the heterogenous clinical manifestations requiring a wide differential diagnostic search. One of the most uncommon vasculitis types is eosinophilic granulomatosis with polyangiitis (EGPA), or Churg-Strauss syndrome [1,2]. This vasculitis is associated with the presence of antineutrophilic cytoplasmic antibodies (ANCA) and is characterized by necrotizing inflammatory lesions of mainly small vessels, as well as eosinophilic and granulomatous inflammation in various organs. Bronchial asthma and eosinophilia are the most common clinical manifestations of EGPA [1–3]. In 2012, according to the current classification of systemic vasculitides, Churg-Strauss syndrome was renamed “Eosinophilic Granulomatosis with Polyangiitis” (EGPA) and classified into the ANCA-associated vasculitides group [3].
EGPA is a rare ANCA-associated vasculitis. The incidence rate is 0.5–6.8 new cases/year per million populations in Europe [2]. Prevalence is higher in northern latitudes compared to southern Europe [4]. The disease occurs most commonly in people 40–60 years of age [2,5]. No sex predominance has been clearly demonstrated [4].
The pathogenesis of the disease is still not completely clear, but the key role is attributed to autoimmune inflammation with the participation of eosinophils, CD8+, and CD4+lymphocytes (predominantly Th-2) and their proinflammatory cytokines: interleukin (IL)4, IL5, and IL13 [2,5]. Elevation of immunoglobulin (Ig)E, IgG4, and eotaxin-3 is common in EGPA patients [2,5]. The frequent (in 30–40% of cases) detection of ANCA (mainly antibodies to myeloperoxidase) also suggests the autoimmune nature of the disease. ANCA-positive patients are more commonly diagnosed with glomerulonephritis and mononeuritis, whereas the ANCA-negative ones more frequently demonstrate endocardial, myocardial, and pulmonary eosinophilic infiltrates [1,2,5,6].
There are 3 phases in the development of EGPA (these phases partially overlap and may not appear in sequence):
the allergic phase, distinguished by allergic upper respiratory tract inflammation (rhinosinusitis, allergic rhinitis) and bronchial asthma;
the eosinophilic phase, manifested with severe eosinophilia in combination with multiple eosinophilic infiltrates in various organs (most often in the lungs and gastrointestinal tract);
the vasculitis phase, with small- and medium-vessel inflammation of different organs, which can often be life-threatening [5,6].
Respiratory manifestations of EGPA include allergic rhinitis, sinusitis, nasal polyposis and, in more than 95% of patients, poorly controlled bronchial asthma [5]. Chest radiography findings include transient and migratory opacities, ground-glass opacity, focal consolidation, and small peribronchial and centrilobular nodules or masses [7]. Computed tomography reveals bronchial abnormalities, most commonly thickened wall and dilatation with secondary bronchiectasis formation, leading to frequent infections [7,8]. Pleural effusions and alveolar hemorrhages are uncommon manifestations of EGPA [1,5,7,8].
A characteristic neurological manifestation seen in 75% of patients with EGPA is peripheral neuropathy (mononeuritis multiplex) [1,5]. Upper-limb symptoms due to neuropathy, as well as vasculitis in epineural vessels, were more frequently reported in the ANCA-positive group [9]. Eosinophil-associated tissue damage and vascular occlusion leading to ischemia are more specific in ANCA-negative patients [9]. Also, the tissue damage caused by eosinophils may be associated with IL-5 concentration [10]. Brain hemorrhage, subarachnoid hemorrhage, and ischemic brain infarction are rare [1,5,6].
The main drugs for the treatment of EGPA are glucocorticoids. In organ- or life-threatening manifestations, the recommended starting dose of prednisone is 1 mg/kg/day with a maximal daily dose of 80 mg for 2–3 weeks, with tapering to a minimal supporting dose selected individually to control the symptoms and minimize the adverse effects of treatment. Methylprednisolone (7.5–15 mg/kg/day) can be used as a pulse therapy for life-threatening manifestations [11,12]. Most patients need prolonged or lifelong steroids. In the case of organ- or life-threatening manifestations, glucocorticoids in combination with cyclophosphamide are prescribed [11,12]. The combination of glucocorticoids and methotrexate or mycophenolate is required to induce remission in mild or moderate disease [11,12]. Mepolizumab (a humanized monoclonal antibody targeting IL-5) has been used for EGPA treatment [11]. For ANCA-positive patients with renal involvement or refractory disease, rituximab can be considered [3, 11].
Due to the rare occurrence of EGPA and the variety of its clinical manifestations, the diagnosis and treatment of this disease can be challenging. Currently, reports of long-term observations with the detailed patient follow-up are of great interest. The aim of our case report is to present rare manifestations of EGPA, such as bronchiectasis and stroke. We also focused on the infectious complications of immunosuppressive treatments. Thus, we would like to present a 10-year observation of a patient with EGPA.
Case Report
The male patient, born in 1953, was first hospitalized in the Pulmonary Department of our clinic in 2007 due to exacerbated asthma. The patient’s history was remarkable for multiple allergies (to house dust, cat and dog epidermis, and plant pollen) as well as nonsteroidal anti-inflammatory drugs intolerance (manifested as bronchospasm). The patient denied unhealthy habits and occupational hazards (he worked as a driver). His family history was not remarkable. The patient has had allergic rhinitis since adolescence. The first asthma symptoms appeared at the age of 18 while serving in the army. At the age of 22, an allergist diagnosed him with bronchial asthma. The patient had annual exacerbations (1–2 times a year) due to cold or allergen exposure, and acute respiratory viral infections. At the age of 42, polypoid rhinosinusitis was found, and a polypectomy was performed. Since 2006, the patient had been on controller treatment with salmeterol/fluticasone inhaler (25/250 mg, respectively) 2 doses 2 times a day.
In late 2009, extremely elevated eosinophils (53%) and total IgE (1000 IU/L) were found for the first time. Also, since 2009, the patient has had transient lung opacities, which usually disappeared after treatment with prednisolone 30 mg per day.
In August 2010, the patient noticed numbness on the back surface of the left foot, which spread to the entire left lower leg in 1 month, as well as numbness of the right foot and lower leg. Later, the patient started having tingling, twisting, and burning sensations aggravated by touching in the feet and lower legs. Also, the weakness in the distal parts of the legs developed, and he started having difficulties moving around on his own and falling while walking, and he was subsequently hospitalized in our clinic due to exacerbated asthma. On laboratory testing, high eosinophils (53%) and gamma-globulins (27.9%) were found in November 2010. The chest radiography showed emphysema and consolidation in the middle lobe of the right lung. A Neurology consult confirmed asymmetrical distal sensory-motor polyneuropathy demonstrated by lower distal asymmetrical paraparesis with decreased muscle strength in the foot flexors and extensors, more severe on the left side, low tone of the shin muscles, hypoactive Achilles reflexes, and reduced leg sensitivity starting from the middle of the shin and below, more severe in the peroneal nerve inner-vation area. The patient also demonstrated abnormal coordination (heel-to-knee, Romberg) test results and bilateral steppage gait with knee overextension. Electroneuromyography showed axonal lesion of all distal leg nerves and secondary myelinopathy. Thus, EGPA was suspected. The ANCA (anti-myeloperoxidase) titer was 30 U/ml (normal reference below 20 U/ml).
Allergic bronchopulmonary aspergillosis and other causes of hypereosinophilia were consistently ruled out. IgE and IgG for Aspergillus spp. and Aspergillus spp. in sputum were not found.
Thus, the presence of severe bronchial asthma, rhinosinusitis, high and persistent eosinophilia, transient pulmonary opacities, and polyneuropathy suggested EGPA (5 out of 6 clinical criteria of American College of Rheumatology present). Based on the clinical presentation and increased ANCA titer, the patient was diagnosed with EGPA. Systemic steroid treatment (prednisolone 300 mg intravenously and 30 mg orally) was prescribed, and 5 plasmapheresis sessions were performed. Later, prolonged combination immunosuppression (prednisolone 10 mg/day and methotrexate 25 mg/week) and controller asthma treatment (fluticasone propionate/salmeterol 250/25 µg 4 doses/ day, tiotropium bromide 18 µg per day) were prescribed. After 1 month on prednisolone and methotrexate, there was a significant neurological improvement: the motor and sensory disturbances decreased, and the patient was able to move around on his own. Further on, polyneuropathy completely resolved on the combination treatment. His asthma remained uncontrolled despite the combination treatment: he had frequent (3 times a year) severe exacerbations requiring hospitalization. In 2015, newly-emerged copious purulent sputum associated with frequent infection-related asthma exacerbations was noted. In January 2016, a patient underwent chest computed tomography (CT), which showed generalized bronchiectasis. Stenosis of the proximal bronchi was ruled out also by fibrobronchos-copy. The bronchial secretion microbiology revealed Klebsiella pneumoniae growth (107 colony-forming units).
In November 2016, during the asthma exacerbation, the patient suffered a cerebral ischemic infarction involving the right occipital lobe, confirmed by CT (Figures 1, 2).
On March 2, 2017, the patient was admitted to our hospital due to severe respiratory failure caused by pneumonia in the left upper lobe and severe asthma exacerbation.
The complete blood count showed high WBCs (15.2×109/l) with left shift. The IgE level was 144.0 IU/l, C-reactive protein was 48.8 mg/dL. Clinical chemistry was remarkable for urea nitrogen 18.4 mmol/l and creatinine 1.55 mg/dL. The sputum microscopy demonstrated 100 WBCs and 160 RBCs per vision field, and no eosinophils were revealed. A chest CT revealed multiple bronchiectasis and upper lobar pneumonia of the left lung (Figure 2). Klebsiella was again detected in the sputum culture.
The patient was prescribed intravenous antibiotics (ceftriaxone 2 g/day and ciprofloxacin 800 mg/day replaced by cefopera-zone 4 g/day and amikacin 1 g/day after 3 days due to low efficacy), bronchodilator and anti-inflammatory therapy (nebulized ipratropium bromide, fenoterol, and budesonide 2000 µg per day, prednisolone intravenously 150 mg and orally 30 mg). Despite the aggressive treatment, the respiratory failure progressed, and on day 6 of hospitalization the patient was transferred to the Intensive Care Unit and intubated. On day 7, the patient died. On autopsy, the EGPA diagnosis was confirmed, signs of ischemic infarction in the occipital lobe of the brain were found, and left upper lobe pneumonia with abscess formation and generalized bilateral bronchiectases were revealed.
Discussion
Based on the long follow-up period and careful interpretation of signs and symptoms, typical laboratory abnormalities, the diagnosis was clear. Among the most commonly used diagnostic criteria of EGPA are the American College of Rheumatology (ACR) classification criteria, Lanham’s criteria, and the Chapel Hill Consensus Conference criteria. The presence of 4 or more ACR classification criteria, including bronchial asthma, eosinophilia (more than 10%), paranasal sinus abnormalities, migratory or transient eosinophilic pulmonary infiltrates, extravascular eosinophil infiltration on biopsy, and mono- or polyneuropathy, has a sensitivity of 85% and a specificity of 100% [2,5,6]. We identified 5 out of 6 American College of Rheumatology clinical criteria present in our patient.
This clinical case shows gradual development of EGPA with multiple-organ involvement (see Figure 3). The disease onset was characterized by respiratory manifestations and suspicion of EGPA appeared only after the development of polyneuropathy (after 39 years) with hypereosinophilia. Polyneuropathy was noted at the age of 57, which is slightly later than the average age of onset reported in the literature [13]. Notably, the patient was ANCA-positive and his long-term neurological manifestation (mononeuritis) was quite typical for ANCA vasculitis [5,6,8,13]. The diagnosis became obvious after comprehensive assessment of the disease history with its gradually evolving signs and symptoms and laboratory abnormalities. Almost all manifestations of the disease seen in the patient were typical except for stroke, which is an exceptional feature for this case [1,5,6]. Neurological involvement manifested as ischemic stroke is an uncommon complication of EGPA (seen only in 5.2% of all patients) [1]. Thus, the patient demonstrated both peripheral and central nervous system involvement, which is unusual and life-threatening in EGPA.
This clinical case of EGPA also attracted our attention due to severe respiratory manifestations. While typical respiratory manifestations such as rhinosinusitis, asthma, and transient lung infiltrates are well documented in the literature, EGPA-associated bronchiectasis is rarely mentioned [14]. Bronchiectasis in the patient was detected by CT at the age of 63. The patient was a nonsmoker, and no other cause of bronchial abnormalities and airflow obstruction could be identified. Stenosis of the proximal bronchi was ruled out by fibrobronchoscopy and CT. The diagnosis of allergic bronchopulmonary aspergillosis was not confirmed. Klebsiella pneumoniae colonization was found during the last 2 years of the patient’s life. Klebsiella pneumoniae is a typical pathogen causing bronchiectasis [15]. It should be noted that aspergillosis and mycobacteriosis are common complications of bronchiectasis [15], but neither was identified in our patient.
The patient was treated for a long time with a combination of prednisolone and methotrexate. There was a significant positive effect on polyneuropathy, but uncontrolled bronchial asthma persisted. This treatment regimen complies with current recommendations [3,11]. Patients with life-threatening and/or organ-threatening manifestations (including lesions of the central nervous system and severe peripheral neuropathy) require therapy with a combination of a glucocorticoid and a cytostatic [3,11].
The use of immunosuppressive drugs in patients with ANCA-associated vasculitis significantly increases the risk of infection and mortality [16–18]. Prednisone and methotrexate use were found to be significant predictors of hospitalization for pneumonia. Frequent infections (more than 1 episode) are reported in patients with ANCA-associated vasculitis during the first year of diagnosis and initiation of therapy [18]. We assume that bronchiectasis complicated by bacterial colonization and bronchial obstruction in combination with immuno-suppression led to severe pneumonia and death of the patient.
Conclusions
The present case report demonstrates a long-term follow-up of a patient with EGPA. Both common and rare manifestations of EGPA have been described in the patient, such as central nervous system damage and bronchiectasis. Immunosuppressive treatment probably contributed to Klebsiella infection development. Thus, long-term complications of EGPA in combination with adverse effects of medications could have caused the severe pneumonia and death of the patient.
Figure 1. Brain multi-spiral computed tomography performed in 2016 showing ischemic brain damage in the right occipital lobe.
Figure 2. (A–C) Chest computed tomography of patient showing large left upper lobe pneumonia and bronchiectasis in lover lobes.
Figure 3. Timeline from first symptoms to death. | NEBULIZED 2000 UG PER DAY | DrugDosageText | CC BY-NC-ND | 33582702 | 18,975,017 | 2021-02-14 |
What was the dosage of drug 'METHOTREXATE SODIUM'? | Case Report: A 64-Year-Old Man with 10-Year History of Eosinophilic Granulomatosis with Polyangiitis with Bronchiectasis and Severe Klebsiella pneumonia.
BACKGROUND Eosinophilic granulomatosis with polyangiitis (EGPA) is a rare and severe progressive disease with multiple clinical manifestations and organ damage. Usually, it requires long-term monitoring of the state of many organs due to the gradual character of its manifestations. CASE REPORT We report a case of a long-term follow-up of a patient with eosinophilic granulomatosis with polyangiitis with emphasis on specific clinical features in this patient. A 64-year-old man was being followed up for 10 years. The initial diagnosis was allergic bronchial asthma; however, as new clinical signs and symptoms developed, the diagnosis of EGPA became obvious. A positive treatment response was seen, mainly manifested as reduced polyneuropathy. Meanwhile, bronchial asthma remained uncontrolled and bronchiectasis and Klebsiella pneumoniae colonization developed despite the combination treatment with prednisolone and methotrexate. Furthermore, the patient suffered a cerebral ischemic infarction. During the last hospital admission, severe uncontrolled bronchial asthma complicated with pneumonia resulted in the patient's death. CONCLUSIONS This clinical case shows the gradual development of EGPA with multiple-organ involvement, including respiratory manifestations and peripheral and central nervous system damage. Immunosuppressive treatment combined with complications of EGPA could have contributed to severe pneumonia development and death of the patient.
Background
Systemic vasculitides are difficult to diagnose due to the heterogenous clinical manifestations requiring a wide differential diagnostic search. One of the most uncommon vasculitis types is eosinophilic granulomatosis with polyangiitis (EGPA), or Churg-Strauss syndrome [1,2]. This vasculitis is associated with the presence of antineutrophilic cytoplasmic antibodies (ANCA) and is characterized by necrotizing inflammatory lesions of mainly small vessels, as well as eosinophilic and granulomatous inflammation in various organs. Bronchial asthma and eosinophilia are the most common clinical manifestations of EGPA [1–3]. In 2012, according to the current classification of systemic vasculitides, Churg-Strauss syndrome was renamed “Eosinophilic Granulomatosis with Polyangiitis” (EGPA) and classified into the ANCA-associated vasculitides group [3].
EGPA is a rare ANCA-associated vasculitis. The incidence rate is 0.5–6.8 new cases/year per million populations in Europe [2]. Prevalence is higher in northern latitudes compared to southern Europe [4]. The disease occurs most commonly in people 40–60 years of age [2,5]. No sex predominance has been clearly demonstrated [4].
The pathogenesis of the disease is still not completely clear, but the key role is attributed to autoimmune inflammation with the participation of eosinophils, CD8+, and CD4+lymphocytes (predominantly Th-2) and their proinflammatory cytokines: interleukin (IL)4, IL5, and IL13 [2,5]. Elevation of immunoglobulin (Ig)E, IgG4, and eotaxin-3 is common in EGPA patients [2,5]. The frequent (in 30–40% of cases) detection of ANCA (mainly antibodies to myeloperoxidase) also suggests the autoimmune nature of the disease. ANCA-positive patients are more commonly diagnosed with glomerulonephritis and mononeuritis, whereas the ANCA-negative ones more frequently demonstrate endocardial, myocardial, and pulmonary eosinophilic infiltrates [1,2,5,6].
There are 3 phases in the development of EGPA (these phases partially overlap and may not appear in sequence):
the allergic phase, distinguished by allergic upper respiratory tract inflammation (rhinosinusitis, allergic rhinitis) and bronchial asthma;
the eosinophilic phase, manifested with severe eosinophilia in combination with multiple eosinophilic infiltrates in various organs (most often in the lungs and gastrointestinal tract);
the vasculitis phase, with small- and medium-vessel inflammation of different organs, which can often be life-threatening [5,6].
Respiratory manifestations of EGPA include allergic rhinitis, sinusitis, nasal polyposis and, in more than 95% of patients, poorly controlled bronchial asthma [5]. Chest radiography findings include transient and migratory opacities, ground-glass opacity, focal consolidation, and small peribronchial and centrilobular nodules or masses [7]. Computed tomography reveals bronchial abnormalities, most commonly thickened wall and dilatation with secondary bronchiectasis formation, leading to frequent infections [7,8]. Pleural effusions and alveolar hemorrhages are uncommon manifestations of EGPA [1,5,7,8].
A characteristic neurological manifestation seen in 75% of patients with EGPA is peripheral neuropathy (mononeuritis multiplex) [1,5]. Upper-limb symptoms due to neuropathy, as well as vasculitis in epineural vessels, were more frequently reported in the ANCA-positive group [9]. Eosinophil-associated tissue damage and vascular occlusion leading to ischemia are more specific in ANCA-negative patients [9]. Also, the tissue damage caused by eosinophils may be associated with IL-5 concentration [10]. Brain hemorrhage, subarachnoid hemorrhage, and ischemic brain infarction are rare [1,5,6].
The main drugs for the treatment of EGPA are glucocorticoids. In organ- or life-threatening manifestations, the recommended starting dose of prednisone is 1 mg/kg/day with a maximal daily dose of 80 mg for 2–3 weeks, with tapering to a minimal supporting dose selected individually to control the symptoms and minimize the adverse effects of treatment. Methylprednisolone (7.5–15 mg/kg/day) can be used as a pulse therapy for life-threatening manifestations [11,12]. Most patients need prolonged or lifelong steroids. In the case of organ- or life-threatening manifestations, glucocorticoids in combination with cyclophosphamide are prescribed [11,12]. The combination of glucocorticoids and methotrexate or mycophenolate is required to induce remission in mild or moderate disease [11,12]. Mepolizumab (a humanized monoclonal antibody targeting IL-5) has been used for EGPA treatment [11]. For ANCA-positive patients with renal involvement or refractory disease, rituximab can be considered [3, 11].
Due to the rare occurrence of EGPA and the variety of its clinical manifestations, the diagnosis and treatment of this disease can be challenging. Currently, reports of long-term observations with the detailed patient follow-up are of great interest. The aim of our case report is to present rare manifestations of EGPA, such as bronchiectasis and stroke. We also focused on the infectious complications of immunosuppressive treatments. Thus, we would like to present a 10-year observation of a patient with EGPA.
Case Report
The male patient, born in 1953, was first hospitalized in the Pulmonary Department of our clinic in 2007 due to exacerbated asthma. The patient’s history was remarkable for multiple allergies (to house dust, cat and dog epidermis, and plant pollen) as well as nonsteroidal anti-inflammatory drugs intolerance (manifested as bronchospasm). The patient denied unhealthy habits and occupational hazards (he worked as a driver). His family history was not remarkable. The patient has had allergic rhinitis since adolescence. The first asthma symptoms appeared at the age of 18 while serving in the army. At the age of 22, an allergist diagnosed him with bronchial asthma. The patient had annual exacerbations (1–2 times a year) due to cold or allergen exposure, and acute respiratory viral infections. At the age of 42, polypoid rhinosinusitis was found, and a polypectomy was performed. Since 2006, the patient had been on controller treatment with salmeterol/fluticasone inhaler (25/250 mg, respectively) 2 doses 2 times a day.
In late 2009, extremely elevated eosinophils (53%) and total IgE (1000 IU/L) were found for the first time. Also, since 2009, the patient has had transient lung opacities, which usually disappeared after treatment with prednisolone 30 mg per day.
In August 2010, the patient noticed numbness on the back surface of the left foot, which spread to the entire left lower leg in 1 month, as well as numbness of the right foot and lower leg. Later, the patient started having tingling, twisting, and burning sensations aggravated by touching in the feet and lower legs. Also, the weakness in the distal parts of the legs developed, and he started having difficulties moving around on his own and falling while walking, and he was subsequently hospitalized in our clinic due to exacerbated asthma. On laboratory testing, high eosinophils (53%) and gamma-globulins (27.9%) were found in November 2010. The chest radiography showed emphysema and consolidation in the middle lobe of the right lung. A Neurology consult confirmed asymmetrical distal sensory-motor polyneuropathy demonstrated by lower distal asymmetrical paraparesis with decreased muscle strength in the foot flexors and extensors, more severe on the left side, low tone of the shin muscles, hypoactive Achilles reflexes, and reduced leg sensitivity starting from the middle of the shin and below, more severe in the peroneal nerve inner-vation area. The patient also demonstrated abnormal coordination (heel-to-knee, Romberg) test results and bilateral steppage gait with knee overextension. Electroneuromyography showed axonal lesion of all distal leg nerves and secondary myelinopathy. Thus, EGPA was suspected. The ANCA (anti-myeloperoxidase) titer was 30 U/ml (normal reference below 20 U/ml).
Allergic bronchopulmonary aspergillosis and other causes of hypereosinophilia were consistently ruled out. IgE and IgG for Aspergillus spp. and Aspergillus spp. in sputum were not found.
Thus, the presence of severe bronchial asthma, rhinosinusitis, high and persistent eosinophilia, transient pulmonary opacities, and polyneuropathy suggested EGPA (5 out of 6 clinical criteria of American College of Rheumatology present). Based on the clinical presentation and increased ANCA titer, the patient was diagnosed with EGPA. Systemic steroid treatment (prednisolone 300 mg intravenously and 30 mg orally) was prescribed, and 5 plasmapheresis sessions were performed. Later, prolonged combination immunosuppression (prednisolone 10 mg/day and methotrexate 25 mg/week) and controller asthma treatment (fluticasone propionate/salmeterol 250/25 µg 4 doses/ day, tiotropium bromide 18 µg per day) were prescribed. After 1 month on prednisolone and methotrexate, there was a significant neurological improvement: the motor and sensory disturbances decreased, and the patient was able to move around on his own. Further on, polyneuropathy completely resolved on the combination treatment. His asthma remained uncontrolled despite the combination treatment: he had frequent (3 times a year) severe exacerbations requiring hospitalization. In 2015, newly-emerged copious purulent sputum associated with frequent infection-related asthma exacerbations was noted. In January 2016, a patient underwent chest computed tomography (CT), which showed generalized bronchiectasis. Stenosis of the proximal bronchi was ruled out also by fibrobronchos-copy. The bronchial secretion microbiology revealed Klebsiella pneumoniae growth (107 colony-forming units).
In November 2016, during the asthma exacerbation, the patient suffered a cerebral ischemic infarction involving the right occipital lobe, confirmed by CT (Figures 1, 2).
On March 2, 2017, the patient was admitted to our hospital due to severe respiratory failure caused by pneumonia in the left upper lobe and severe asthma exacerbation.
The complete blood count showed high WBCs (15.2×109/l) with left shift. The IgE level was 144.0 IU/l, C-reactive protein was 48.8 mg/dL. Clinical chemistry was remarkable for urea nitrogen 18.4 mmol/l and creatinine 1.55 mg/dL. The sputum microscopy demonstrated 100 WBCs and 160 RBCs per vision field, and no eosinophils were revealed. A chest CT revealed multiple bronchiectasis and upper lobar pneumonia of the left lung (Figure 2). Klebsiella was again detected in the sputum culture.
The patient was prescribed intravenous antibiotics (ceftriaxone 2 g/day and ciprofloxacin 800 mg/day replaced by cefopera-zone 4 g/day and amikacin 1 g/day after 3 days due to low efficacy), bronchodilator and anti-inflammatory therapy (nebulized ipratropium bromide, fenoterol, and budesonide 2000 µg per day, prednisolone intravenously 150 mg and orally 30 mg). Despite the aggressive treatment, the respiratory failure progressed, and on day 6 of hospitalization the patient was transferred to the Intensive Care Unit and intubated. On day 7, the patient died. On autopsy, the EGPA diagnosis was confirmed, signs of ischemic infarction in the occipital lobe of the brain were found, and left upper lobe pneumonia with abscess formation and generalized bilateral bronchiectases were revealed.
Discussion
Based on the long follow-up period and careful interpretation of signs and symptoms, typical laboratory abnormalities, the diagnosis was clear. Among the most commonly used diagnostic criteria of EGPA are the American College of Rheumatology (ACR) classification criteria, Lanham’s criteria, and the Chapel Hill Consensus Conference criteria. The presence of 4 or more ACR classification criteria, including bronchial asthma, eosinophilia (more than 10%), paranasal sinus abnormalities, migratory or transient eosinophilic pulmonary infiltrates, extravascular eosinophil infiltration on biopsy, and mono- or polyneuropathy, has a sensitivity of 85% and a specificity of 100% [2,5,6]. We identified 5 out of 6 American College of Rheumatology clinical criteria present in our patient.
This clinical case shows gradual development of EGPA with multiple-organ involvement (see Figure 3). The disease onset was characterized by respiratory manifestations and suspicion of EGPA appeared only after the development of polyneuropathy (after 39 years) with hypereosinophilia. Polyneuropathy was noted at the age of 57, which is slightly later than the average age of onset reported in the literature [13]. Notably, the patient was ANCA-positive and his long-term neurological manifestation (mononeuritis) was quite typical for ANCA vasculitis [5,6,8,13]. The diagnosis became obvious after comprehensive assessment of the disease history with its gradually evolving signs and symptoms and laboratory abnormalities. Almost all manifestations of the disease seen in the patient were typical except for stroke, which is an exceptional feature for this case [1,5,6]. Neurological involvement manifested as ischemic stroke is an uncommon complication of EGPA (seen only in 5.2% of all patients) [1]. Thus, the patient demonstrated both peripheral and central nervous system involvement, which is unusual and life-threatening in EGPA.
This clinical case of EGPA also attracted our attention due to severe respiratory manifestations. While typical respiratory manifestations such as rhinosinusitis, asthma, and transient lung infiltrates are well documented in the literature, EGPA-associated bronchiectasis is rarely mentioned [14]. Bronchiectasis in the patient was detected by CT at the age of 63. The patient was a nonsmoker, and no other cause of bronchial abnormalities and airflow obstruction could be identified. Stenosis of the proximal bronchi was ruled out by fibrobronchoscopy and CT. The diagnosis of allergic bronchopulmonary aspergillosis was not confirmed. Klebsiella pneumoniae colonization was found during the last 2 years of the patient’s life. Klebsiella pneumoniae is a typical pathogen causing bronchiectasis [15]. It should be noted that aspergillosis and mycobacteriosis are common complications of bronchiectasis [15], but neither was identified in our patient.
The patient was treated for a long time with a combination of prednisolone and methotrexate. There was a significant positive effect on polyneuropathy, but uncontrolled bronchial asthma persisted. This treatment regimen complies with current recommendations [3,11]. Patients with life-threatening and/or organ-threatening manifestations (including lesions of the central nervous system and severe peripheral neuropathy) require therapy with a combination of a glucocorticoid and a cytostatic [3,11].
The use of immunosuppressive drugs in patients with ANCA-associated vasculitis significantly increases the risk of infection and mortality [16–18]. Prednisone and methotrexate use were found to be significant predictors of hospitalization for pneumonia. Frequent infections (more than 1 episode) are reported in patients with ANCA-associated vasculitis during the first year of diagnosis and initiation of therapy [18]. We assume that bronchiectasis complicated by bacterial colonization and bronchial obstruction in combination with immuno-suppression led to severe pneumonia and death of the patient.
Conclusions
The present case report demonstrates a long-term follow-up of a patient with EGPA. Both common and rare manifestations of EGPA have been described in the patient, such as central nervous system damage and bronchiectasis. Immunosuppressive treatment probably contributed to Klebsiella infection development. Thus, long-term complications of EGPA in combination with adverse effects of medications could have caused the severe pneumonia and death of the patient.
Figure 1. Brain multi-spiral computed tomography performed in 2016 showing ischemic brain damage in the right occipital lobe.
Figure 2. (A–C) Chest computed tomography of patient showing large left upper lobe pneumonia and bronchiectasis in lover lobes.
Figure 3. Timeline from first symptoms to death. | 25 MG, WEEKLY | DrugDosageText | CC BY-NC-ND | 33582702 | 18,986,917 | 2021-02-14 |
What was the dosage of drug 'TIOTROPIUM'? | Case Report: A 64-Year-Old Man with 10-Year History of Eosinophilic Granulomatosis with Polyangiitis with Bronchiectasis and Severe Klebsiella pneumonia.
BACKGROUND Eosinophilic granulomatosis with polyangiitis (EGPA) is a rare and severe progressive disease with multiple clinical manifestations and organ damage. Usually, it requires long-term monitoring of the state of many organs due to the gradual character of its manifestations. CASE REPORT We report a case of a long-term follow-up of a patient with eosinophilic granulomatosis with polyangiitis with emphasis on specific clinical features in this patient. A 64-year-old man was being followed up for 10 years. The initial diagnosis was allergic bronchial asthma; however, as new clinical signs and symptoms developed, the diagnosis of EGPA became obvious. A positive treatment response was seen, mainly manifested as reduced polyneuropathy. Meanwhile, bronchial asthma remained uncontrolled and bronchiectasis and Klebsiella pneumoniae colonization developed despite the combination treatment with prednisolone and methotrexate. Furthermore, the patient suffered a cerebral ischemic infarction. During the last hospital admission, severe uncontrolled bronchial asthma complicated with pneumonia resulted in the patient's death. CONCLUSIONS This clinical case shows the gradual development of EGPA with multiple-organ involvement, including respiratory manifestations and peripheral and central nervous system damage. Immunosuppressive treatment combined with complications of EGPA could have contributed to severe pneumonia development and death of the patient.
Background
Systemic vasculitides are difficult to diagnose due to the heterogenous clinical manifestations requiring a wide differential diagnostic search. One of the most uncommon vasculitis types is eosinophilic granulomatosis with polyangiitis (EGPA), or Churg-Strauss syndrome [1,2]. This vasculitis is associated with the presence of antineutrophilic cytoplasmic antibodies (ANCA) and is characterized by necrotizing inflammatory lesions of mainly small vessels, as well as eosinophilic and granulomatous inflammation in various organs. Bronchial asthma and eosinophilia are the most common clinical manifestations of EGPA [1–3]. In 2012, according to the current classification of systemic vasculitides, Churg-Strauss syndrome was renamed “Eosinophilic Granulomatosis with Polyangiitis” (EGPA) and classified into the ANCA-associated vasculitides group [3].
EGPA is a rare ANCA-associated vasculitis. The incidence rate is 0.5–6.8 new cases/year per million populations in Europe [2]. Prevalence is higher in northern latitudes compared to southern Europe [4]. The disease occurs most commonly in people 40–60 years of age [2,5]. No sex predominance has been clearly demonstrated [4].
The pathogenesis of the disease is still not completely clear, but the key role is attributed to autoimmune inflammation with the participation of eosinophils, CD8+, and CD4+lymphocytes (predominantly Th-2) and their proinflammatory cytokines: interleukin (IL)4, IL5, and IL13 [2,5]. Elevation of immunoglobulin (Ig)E, IgG4, and eotaxin-3 is common in EGPA patients [2,5]. The frequent (in 30–40% of cases) detection of ANCA (mainly antibodies to myeloperoxidase) also suggests the autoimmune nature of the disease. ANCA-positive patients are more commonly diagnosed with glomerulonephritis and mononeuritis, whereas the ANCA-negative ones more frequently demonstrate endocardial, myocardial, and pulmonary eosinophilic infiltrates [1,2,5,6].
There are 3 phases in the development of EGPA (these phases partially overlap and may not appear in sequence):
the allergic phase, distinguished by allergic upper respiratory tract inflammation (rhinosinusitis, allergic rhinitis) and bronchial asthma;
the eosinophilic phase, manifested with severe eosinophilia in combination with multiple eosinophilic infiltrates in various organs (most often in the lungs and gastrointestinal tract);
the vasculitis phase, with small- and medium-vessel inflammation of different organs, which can often be life-threatening [5,6].
Respiratory manifestations of EGPA include allergic rhinitis, sinusitis, nasal polyposis and, in more than 95% of patients, poorly controlled bronchial asthma [5]. Chest radiography findings include transient and migratory opacities, ground-glass opacity, focal consolidation, and small peribronchial and centrilobular nodules or masses [7]. Computed tomography reveals bronchial abnormalities, most commonly thickened wall and dilatation with secondary bronchiectasis formation, leading to frequent infections [7,8]. Pleural effusions and alveolar hemorrhages are uncommon manifestations of EGPA [1,5,7,8].
A characteristic neurological manifestation seen in 75% of patients with EGPA is peripheral neuropathy (mononeuritis multiplex) [1,5]. Upper-limb symptoms due to neuropathy, as well as vasculitis in epineural vessels, were more frequently reported in the ANCA-positive group [9]. Eosinophil-associated tissue damage and vascular occlusion leading to ischemia are more specific in ANCA-negative patients [9]. Also, the tissue damage caused by eosinophils may be associated with IL-5 concentration [10]. Brain hemorrhage, subarachnoid hemorrhage, and ischemic brain infarction are rare [1,5,6].
The main drugs for the treatment of EGPA are glucocorticoids. In organ- or life-threatening manifestations, the recommended starting dose of prednisone is 1 mg/kg/day with a maximal daily dose of 80 mg for 2–3 weeks, with tapering to a minimal supporting dose selected individually to control the symptoms and minimize the adverse effects of treatment. Methylprednisolone (7.5–15 mg/kg/day) can be used as a pulse therapy for life-threatening manifestations [11,12]. Most patients need prolonged or lifelong steroids. In the case of organ- or life-threatening manifestations, glucocorticoids in combination with cyclophosphamide are prescribed [11,12]. The combination of glucocorticoids and methotrexate or mycophenolate is required to induce remission in mild or moderate disease [11,12]. Mepolizumab (a humanized monoclonal antibody targeting IL-5) has been used for EGPA treatment [11]. For ANCA-positive patients with renal involvement or refractory disease, rituximab can be considered [3, 11].
Due to the rare occurrence of EGPA and the variety of its clinical manifestations, the diagnosis and treatment of this disease can be challenging. Currently, reports of long-term observations with the detailed patient follow-up are of great interest. The aim of our case report is to present rare manifestations of EGPA, such as bronchiectasis and stroke. We also focused on the infectious complications of immunosuppressive treatments. Thus, we would like to present a 10-year observation of a patient with EGPA.
Case Report
The male patient, born in 1953, was first hospitalized in the Pulmonary Department of our clinic in 2007 due to exacerbated asthma. The patient’s history was remarkable for multiple allergies (to house dust, cat and dog epidermis, and plant pollen) as well as nonsteroidal anti-inflammatory drugs intolerance (manifested as bronchospasm). The patient denied unhealthy habits and occupational hazards (he worked as a driver). His family history was not remarkable. The patient has had allergic rhinitis since adolescence. The first asthma symptoms appeared at the age of 18 while serving in the army. At the age of 22, an allergist diagnosed him with bronchial asthma. The patient had annual exacerbations (1–2 times a year) due to cold or allergen exposure, and acute respiratory viral infections. At the age of 42, polypoid rhinosinusitis was found, and a polypectomy was performed. Since 2006, the patient had been on controller treatment with salmeterol/fluticasone inhaler (25/250 mg, respectively) 2 doses 2 times a day.
In late 2009, extremely elevated eosinophils (53%) and total IgE (1000 IU/L) were found for the first time. Also, since 2009, the patient has had transient lung opacities, which usually disappeared after treatment with prednisolone 30 mg per day.
In August 2010, the patient noticed numbness on the back surface of the left foot, which spread to the entire left lower leg in 1 month, as well as numbness of the right foot and lower leg. Later, the patient started having tingling, twisting, and burning sensations aggravated by touching in the feet and lower legs. Also, the weakness in the distal parts of the legs developed, and he started having difficulties moving around on his own and falling while walking, and he was subsequently hospitalized in our clinic due to exacerbated asthma. On laboratory testing, high eosinophils (53%) and gamma-globulins (27.9%) were found in November 2010. The chest radiography showed emphysema and consolidation in the middle lobe of the right lung. A Neurology consult confirmed asymmetrical distal sensory-motor polyneuropathy demonstrated by lower distal asymmetrical paraparesis with decreased muscle strength in the foot flexors and extensors, more severe on the left side, low tone of the shin muscles, hypoactive Achilles reflexes, and reduced leg sensitivity starting from the middle of the shin and below, more severe in the peroneal nerve inner-vation area. The patient also demonstrated abnormal coordination (heel-to-knee, Romberg) test results and bilateral steppage gait with knee overextension. Electroneuromyography showed axonal lesion of all distal leg nerves and secondary myelinopathy. Thus, EGPA was suspected. The ANCA (anti-myeloperoxidase) titer was 30 U/ml (normal reference below 20 U/ml).
Allergic bronchopulmonary aspergillosis and other causes of hypereosinophilia were consistently ruled out. IgE and IgG for Aspergillus spp. and Aspergillus spp. in sputum were not found.
Thus, the presence of severe bronchial asthma, rhinosinusitis, high and persistent eosinophilia, transient pulmonary opacities, and polyneuropathy suggested EGPA (5 out of 6 clinical criteria of American College of Rheumatology present). Based on the clinical presentation and increased ANCA titer, the patient was diagnosed with EGPA. Systemic steroid treatment (prednisolone 300 mg intravenously and 30 mg orally) was prescribed, and 5 plasmapheresis sessions were performed. Later, prolonged combination immunosuppression (prednisolone 10 mg/day and methotrexate 25 mg/week) and controller asthma treatment (fluticasone propionate/salmeterol 250/25 µg 4 doses/ day, tiotropium bromide 18 µg per day) were prescribed. After 1 month on prednisolone and methotrexate, there was a significant neurological improvement: the motor and sensory disturbances decreased, and the patient was able to move around on his own. Further on, polyneuropathy completely resolved on the combination treatment. His asthma remained uncontrolled despite the combination treatment: he had frequent (3 times a year) severe exacerbations requiring hospitalization. In 2015, newly-emerged copious purulent sputum associated with frequent infection-related asthma exacerbations was noted. In January 2016, a patient underwent chest computed tomography (CT), which showed generalized bronchiectasis. Stenosis of the proximal bronchi was ruled out also by fibrobronchos-copy. The bronchial secretion microbiology revealed Klebsiella pneumoniae growth (107 colony-forming units).
In November 2016, during the asthma exacerbation, the patient suffered a cerebral ischemic infarction involving the right occipital lobe, confirmed by CT (Figures 1, 2).
On March 2, 2017, the patient was admitted to our hospital due to severe respiratory failure caused by pneumonia in the left upper lobe and severe asthma exacerbation.
The complete blood count showed high WBCs (15.2×109/l) with left shift. The IgE level was 144.0 IU/l, C-reactive protein was 48.8 mg/dL. Clinical chemistry was remarkable for urea nitrogen 18.4 mmol/l and creatinine 1.55 mg/dL. The sputum microscopy demonstrated 100 WBCs and 160 RBCs per vision field, and no eosinophils were revealed. A chest CT revealed multiple bronchiectasis and upper lobar pneumonia of the left lung (Figure 2). Klebsiella was again detected in the sputum culture.
The patient was prescribed intravenous antibiotics (ceftriaxone 2 g/day and ciprofloxacin 800 mg/day replaced by cefopera-zone 4 g/day and amikacin 1 g/day after 3 days due to low efficacy), bronchodilator and anti-inflammatory therapy (nebulized ipratropium bromide, fenoterol, and budesonide 2000 µg per day, prednisolone intravenously 150 mg and orally 30 mg). Despite the aggressive treatment, the respiratory failure progressed, and on day 6 of hospitalization the patient was transferred to the Intensive Care Unit and intubated. On day 7, the patient died. On autopsy, the EGPA diagnosis was confirmed, signs of ischemic infarction in the occipital lobe of the brain were found, and left upper lobe pneumonia with abscess formation and generalized bilateral bronchiectases were revealed.
Discussion
Based on the long follow-up period and careful interpretation of signs and symptoms, typical laboratory abnormalities, the diagnosis was clear. Among the most commonly used diagnostic criteria of EGPA are the American College of Rheumatology (ACR) classification criteria, Lanham’s criteria, and the Chapel Hill Consensus Conference criteria. The presence of 4 or more ACR classification criteria, including bronchial asthma, eosinophilia (more than 10%), paranasal sinus abnormalities, migratory or transient eosinophilic pulmonary infiltrates, extravascular eosinophil infiltration on biopsy, and mono- or polyneuropathy, has a sensitivity of 85% and a specificity of 100% [2,5,6]. We identified 5 out of 6 American College of Rheumatology clinical criteria present in our patient.
This clinical case shows gradual development of EGPA with multiple-organ involvement (see Figure 3). The disease onset was characterized by respiratory manifestations and suspicion of EGPA appeared only after the development of polyneuropathy (after 39 years) with hypereosinophilia. Polyneuropathy was noted at the age of 57, which is slightly later than the average age of onset reported in the literature [13]. Notably, the patient was ANCA-positive and his long-term neurological manifestation (mononeuritis) was quite typical for ANCA vasculitis [5,6,8,13]. The diagnosis became obvious after comprehensive assessment of the disease history with its gradually evolving signs and symptoms and laboratory abnormalities. Almost all manifestations of the disease seen in the patient were typical except for stroke, which is an exceptional feature for this case [1,5,6]. Neurological involvement manifested as ischemic stroke is an uncommon complication of EGPA (seen only in 5.2% of all patients) [1]. Thus, the patient demonstrated both peripheral and central nervous system involvement, which is unusual and life-threatening in EGPA.
This clinical case of EGPA also attracted our attention due to severe respiratory manifestations. While typical respiratory manifestations such as rhinosinusitis, asthma, and transient lung infiltrates are well documented in the literature, EGPA-associated bronchiectasis is rarely mentioned [14]. Bronchiectasis in the patient was detected by CT at the age of 63. The patient was a nonsmoker, and no other cause of bronchial abnormalities and airflow obstruction could be identified. Stenosis of the proximal bronchi was ruled out by fibrobronchoscopy and CT. The diagnosis of allergic bronchopulmonary aspergillosis was not confirmed. Klebsiella pneumoniae colonization was found during the last 2 years of the patient’s life. Klebsiella pneumoniae is a typical pathogen causing bronchiectasis [15]. It should be noted that aspergillosis and mycobacteriosis are common complications of bronchiectasis [15], but neither was identified in our patient.
The patient was treated for a long time with a combination of prednisolone and methotrexate. There was a significant positive effect on polyneuropathy, but uncontrolled bronchial asthma persisted. This treatment regimen complies with current recommendations [3,11]. Patients with life-threatening and/or organ-threatening manifestations (including lesions of the central nervous system and severe peripheral neuropathy) require therapy with a combination of a glucocorticoid and a cytostatic [3,11].
The use of immunosuppressive drugs in patients with ANCA-associated vasculitis significantly increases the risk of infection and mortality [16–18]. Prednisone and methotrexate use were found to be significant predictors of hospitalization for pneumonia. Frequent infections (more than 1 episode) are reported in patients with ANCA-associated vasculitis during the first year of diagnosis and initiation of therapy [18]. We assume that bronchiectasis complicated by bacterial colonization and bronchial obstruction in combination with immuno-suppression led to severe pneumonia and death of the patient.
Conclusions
The present case report demonstrates a long-term follow-up of a patient with EGPA. Both common and rare manifestations of EGPA have been described in the patient, such as central nervous system damage and bronchiectasis. Immunosuppressive treatment probably contributed to Klebsiella infection development. Thus, long-term complications of EGPA in combination with adverse effects of medications could have caused the severe pneumonia and death of the patient.
Figure 1. Brain multi-spiral computed tomography performed in 2016 showing ischemic brain damage in the right occipital lobe.
Figure 2. (A–C) Chest computed tomography of patient showing large left upper lobe pneumonia and bronchiectasis in lover lobes.
Figure 3. Timeline from first symptoms to death. | 18 µg (micrograms). | DrugDosage | CC BY-NC-ND | 33582702 | 18,962,204 | 2021-02-14 |
What was the outcome of reaction 'Asthma'? | Case Report: A 64-Year-Old Man with 10-Year History of Eosinophilic Granulomatosis with Polyangiitis with Bronchiectasis and Severe Klebsiella pneumonia.
BACKGROUND Eosinophilic granulomatosis with polyangiitis (EGPA) is a rare and severe progressive disease with multiple clinical manifestations and organ damage. Usually, it requires long-term monitoring of the state of many organs due to the gradual character of its manifestations. CASE REPORT We report a case of a long-term follow-up of a patient with eosinophilic granulomatosis with polyangiitis with emphasis on specific clinical features in this patient. A 64-year-old man was being followed up for 10 years. The initial diagnosis was allergic bronchial asthma; however, as new clinical signs and symptoms developed, the diagnosis of EGPA became obvious. A positive treatment response was seen, mainly manifested as reduced polyneuropathy. Meanwhile, bronchial asthma remained uncontrolled and bronchiectasis and Klebsiella pneumoniae colonization developed despite the combination treatment with prednisolone and methotrexate. Furthermore, the patient suffered a cerebral ischemic infarction. During the last hospital admission, severe uncontrolled bronchial asthma complicated with pneumonia resulted in the patient's death. CONCLUSIONS This clinical case shows the gradual development of EGPA with multiple-organ involvement, including respiratory manifestations and peripheral and central nervous system damage. Immunosuppressive treatment combined with complications of EGPA could have contributed to severe pneumonia development and death of the patient.
Background
Systemic vasculitides are difficult to diagnose due to the heterogenous clinical manifestations requiring a wide differential diagnostic search. One of the most uncommon vasculitis types is eosinophilic granulomatosis with polyangiitis (EGPA), or Churg-Strauss syndrome [1,2]. This vasculitis is associated with the presence of antineutrophilic cytoplasmic antibodies (ANCA) and is characterized by necrotizing inflammatory lesions of mainly small vessels, as well as eosinophilic and granulomatous inflammation in various organs. Bronchial asthma and eosinophilia are the most common clinical manifestations of EGPA [1–3]. In 2012, according to the current classification of systemic vasculitides, Churg-Strauss syndrome was renamed “Eosinophilic Granulomatosis with Polyangiitis” (EGPA) and classified into the ANCA-associated vasculitides group [3].
EGPA is a rare ANCA-associated vasculitis. The incidence rate is 0.5–6.8 new cases/year per million populations in Europe [2]. Prevalence is higher in northern latitudes compared to southern Europe [4]. The disease occurs most commonly in people 40–60 years of age [2,5]. No sex predominance has been clearly demonstrated [4].
The pathogenesis of the disease is still not completely clear, but the key role is attributed to autoimmune inflammation with the participation of eosinophils, CD8+, and CD4+lymphocytes (predominantly Th-2) and their proinflammatory cytokines: interleukin (IL)4, IL5, and IL13 [2,5]. Elevation of immunoglobulin (Ig)E, IgG4, and eotaxin-3 is common in EGPA patients [2,5]. The frequent (in 30–40% of cases) detection of ANCA (mainly antibodies to myeloperoxidase) also suggests the autoimmune nature of the disease. ANCA-positive patients are more commonly diagnosed with glomerulonephritis and mononeuritis, whereas the ANCA-negative ones more frequently demonstrate endocardial, myocardial, and pulmonary eosinophilic infiltrates [1,2,5,6].
There are 3 phases in the development of EGPA (these phases partially overlap and may not appear in sequence):
the allergic phase, distinguished by allergic upper respiratory tract inflammation (rhinosinusitis, allergic rhinitis) and bronchial asthma;
the eosinophilic phase, manifested with severe eosinophilia in combination with multiple eosinophilic infiltrates in various organs (most often in the lungs and gastrointestinal tract);
the vasculitis phase, with small- and medium-vessel inflammation of different organs, which can often be life-threatening [5,6].
Respiratory manifestations of EGPA include allergic rhinitis, sinusitis, nasal polyposis and, in more than 95% of patients, poorly controlled bronchial asthma [5]. Chest radiography findings include transient and migratory opacities, ground-glass opacity, focal consolidation, and small peribronchial and centrilobular nodules or masses [7]. Computed tomography reveals bronchial abnormalities, most commonly thickened wall and dilatation with secondary bronchiectasis formation, leading to frequent infections [7,8]. Pleural effusions and alveolar hemorrhages are uncommon manifestations of EGPA [1,5,7,8].
A characteristic neurological manifestation seen in 75% of patients with EGPA is peripheral neuropathy (mononeuritis multiplex) [1,5]. Upper-limb symptoms due to neuropathy, as well as vasculitis in epineural vessels, were more frequently reported in the ANCA-positive group [9]. Eosinophil-associated tissue damage and vascular occlusion leading to ischemia are more specific in ANCA-negative patients [9]. Also, the tissue damage caused by eosinophils may be associated with IL-5 concentration [10]. Brain hemorrhage, subarachnoid hemorrhage, and ischemic brain infarction are rare [1,5,6].
The main drugs for the treatment of EGPA are glucocorticoids. In organ- or life-threatening manifestations, the recommended starting dose of prednisone is 1 mg/kg/day with a maximal daily dose of 80 mg for 2–3 weeks, with tapering to a minimal supporting dose selected individually to control the symptoms and minimize the adverse effects of treatment. Methylprednisolone (7.5–15 mg/kg/day) can be used as a pulse therapy for life-threatening manifestations [11,12]. Most patients need prolonged or lifelong steroids. In the case of organ- or life-threatening manifestations, glucocorticoids in combination with cyclophosphamide are prescribed [11,12]. The combination of glucocorticoids and methotrexate or mycophenolate is required to induce remission in mild or moderate disease [11,12]. Mepolizumab (a humanized monoclonal antibody targeting IL-5) has been used for EGPA treatment [11]. For ANCA-positive patients with renal involvement or refractory disease, rituximab can be considered [3, 11].
Due to the rare occurrence of EGPA and the variety of its clinical manifestations, the diagnosis and treatment of this disease can be challenging. Currently, reports of long-term observations with the detailed patient follow-up are of great interest. The aim of our case report is to present rare manifestations of EGPA, such as bronchiectasis and stroke. We also focused on the infectious complications of immunosuppressive treatments. Thus, we would like to present a 10-year observation of a patient with EGPA.
Case Report
The male patient, born in 1953, was first hospitalized in the Pulmonary Department of our clinic in 2007 due to exacerbated asthma. The patient’s history was remarkable for multiple allergies (to house dust, cat and dog epidermis, and plant pollen) as well as nonsteroidal anti-inflammatory drugs intolerance (manifested as bronchospasm). The patient denied unhealthy habits and occupational hazards (he worked as a driver). His family history was not remarkable. The patient has had allergic rhinitis since adolescence. The first asthma symptoms appeared at the age of 18 while serving in the army. At the age of 22, an allergist diagnosed him with bronchial asthma. The patient had annual exacerbations (1–2 times a year) due to cold or allergen exposure, and acute respiratory viral infections. At the age of 42, polypoid rhinosinusitis was found, and a polypectomy was performed. Since 2006, the patient had been on controller treatment with salmeterol/fluticasone inhaler (25/250 mg, respectively) 2 doses 2 times a day.
In late 2009, extremely elevated eosinophils (53%) and total IgE (1000 IU/L) were found for the first time. Also, since 2009, the patient has had transient lung opacities, which usually disappeared after treatment with prednisolone 30 mg per day.
In August 2010, the patient noticed numbness on the back surface of the left foot, which spread to the entire left lower leg in 1 month, as well as numbness of the right foot and lower leg. Later, the patient started having tingling, twisting, and burning sensations aggravated by touching in the feet and lower legs. Also, the weakness in the distal parts of the legs developed, and he started having difficulties moving around on his own and falling while walking, and he was subsequently hospitalized in our clinic due to exacerbated asthma. On laboratory testing, high eosinophils (53%) and gamma-globulins (27.9%) were found in November 2010. The chest radiography showed emphysema and consolidation in the middle lobe of the right lung. A Neurology consult confirmed asymmetrical distal sensory-motor polyneuropathy demonstrated by lower distal asymmetrical paraparesis with decreased muscle strength in the foot flexors and extensors, more severe on the left side, low tone of the shin muscles, hypoactive Achilles reflexes, and reduced leg sensitivity starting from the middle of the shin and below, more severe in the peroneal nerve inner-vation area. The patient also demonstrated abnormal coordination (heel-to-knee, Romberg) test results and bilateral steppage gait with knee overextension. Electroneuromyography showed axonal lesion of all distal leg nerves and secondary myelinopathy. Thus, EGPA was suspected. The ANCA (anti-myeloperoxidase) titer was 30 U/ml (normal reference below 20 U/ml).
Allergic bronchopulmonary aspergillosis and other causes of hypereosinophilia were consistently ruled out. IgE and IgG for Aspergillus spp. and Aspergillus spp. in sputum were not found.
Thus, the presence of severe bronchial asthma, rhinosinusitis, high and persistent eosinophilia, transient pulmonary opacities, and polyneuropathy suggested EGPA (5 out of 6 clinical criteria of American College of Rheumatology present). Based on the clinical presentation and increased ANCA titer, the patient was diagnosed with EGPA. Systemic steroid treatment (prednisolone 300 mg intravenously and 30 mg orally) was prescribed, and 5 plasmapheresis sessions were performed. Later, prolonged combination immunosuppression (prednisolone 10 mg/day and methotrexate 25 mg/week) and controller asthma treatment (fluticasone propionate/salmeterol 250/25 µg 4 doses/ day, tiotropium bromide 18 µg per day) were prescribed. After 1 month on prednisolone and methotrexate, there was a significant neurological improvement: the motor and sensory disturbances decreased, and the patient was able to move around on his own. Further on, polyneuropathy completely resolved on the combination treatment. His asthma remained uncontrolled despite the combination treatment: he had frequent (3 times a year) severe exacerbations requiring hospitalization. In 2015, newly-emerged copious purulent sputum associated with frequent infection-related asthma exacerbations was noted. In January 2016, a patient underwent chest computed tomography (CT), which showed generalized bronchiectasis. Stenosis of the proximal bronchi was ruled out also by fibrobronchos-copy. The bronchial secretion microbiology revealed Klebsiella pneumoniae growth (107 colony-forming units).
In November 2016, during the asthma exacerbation, the patient suffered a cerebral ischemic infarction involving the right occipital lobe, confirmed by CT (Figures 1, 2).
On March 2, 2017, the patient was admitted to our hospital due to severe respiratory failure caused by pneumonia in the left upper lobe and severe asthma exacerbation.
The complete blood count showed high WBCs (15.2×109/l) with left shift. The IgE level was 144.0 IU/l, C-reactive protein was 48.8 mg/dL. Clinical chemistry was remarkable for urea nitrogen 18.4 mmol/l and creatinine 1.55 mg/dL. The sputum microscopy demonstrated 100 WBCs and 160 RBCs per vision field, and no eosinophils were revealed. A chest CT revealed multiple bronchiectasis and upper lobar pneumonia of the left lung (Figure 2). Klebsiella was again detected in the sputum culture.
The patient was prescribed intravenous antibiotics (ceftriaxone 2 g/day and ciprofloxacin 800 mg/day replaced by cefopera-zone 4 g/day and amikacin 1 g/day after 3 days due to low efficacy), bronchodilator and anti-inflammatory therapy (nebulized ipratropium bromide, fenoterol, and budesonide 2000 µg per day, prednisolone intravenously 150 mg and orally 30 mg). Despite the aggressive treatment, the respiratory failure progressed, and on day 6 of hospitalization the patient was transferred to the Intensive Care Unit and intubated. On day 7, the patient died. On autopsy, the EGPA diagnosis was confirmed, signs of ischemic infarction in the occipital lobe of the brain were found, and left upper lobe pneumonia with abscess formation and generalized bilateral bronchiectases were revealed.
Discussion
Based on the long follow-up period and careful interpretation of signs and symptoms, typical laboratory abnormalities, the diagnosis was clear. Among the most commonly used diagnostic criteria of EGPA are the American College of Rheumatology (ACR) classification criteria, Lanham’s criteria, and the Chapel Hill Consensus Conference criteria. The presence of 4 or more ACR classification criteria, including bronchial asthma, eosinophilia (more than 10%), paranasal sinus abnormalities, migratory or transient eosinophilic pulmonary infiltrates, extravascular eosinophil infiltration on biopsy, and mono- or polyneuropathy, has a sensitivity of 85% and a specificity of 100% [2,5,6]. We identified 5 out of 6 American College of Rheumatology clinical criteria present in our patient.
This clinical case shows gradual development of EGPA with multiple-organ involvement (see Figure 3). The disease onset was characterized by respiratory manifestations and suspicion of EGPA appeared only after the development of polyneuropathy (after 39 years) with hypereosinophilia. Polyneuropathy was noted at the age of 57, which is slightly later than the average age of onset reported in the literature [13]. Notably, the patient was ANCA-positive and his long-term neurological manifestation (mononeuritis) was quite typical for ANCA vasculitis [5,6,8,13]. The diagnosis became obvious after comprehensive assessment of the disease history with its gradually evolving signs and symptoms and laboratory abnormalities. Almost all manifestations of the disease seen in the patient were typical except for stroke, which is an exceptional feature for this case [1,5,6]. Neurological involvement manifested as ischemic stroke is an uncommon complication of EGPA (seen only in 5.2% of all patients) [1]. Thus, the patient demonstrated both peripheral and central nervous system involvement, which is unusual and life-threatening in EGPA.
This clinical case of EGPA also attracted our attention due to severe respiratory manifestations. While typical respiratory manifestations such as rhinosinusitis, asthma, and transient lung infiltrates are well documented in the literature, EGPA-associated bronchiectasis is rarely mentioned [14]. Bronchiectasis in the patient was detected by CT at the age of 63. The patient was a nonsmoker, and no other cause of bronchial abnormalities and airflow obstruction could be identified. Stenosis of the proximal bronchi was ruled out by fibrobronchoscopy and CT. The diagnosis of allergic bronchopulmonary aspergillosis was not confirmed. Klebsiella pneumoniae colonization was found during the last 2 years of the patient’s life. Klebsiella pneumoniae is a typical pathogen causing bronchiectasis [15]. It should be noted that aspergillosis and mycobacteriosis are common complications of bronchiectasis [15], but neither was identified in our patient.
The patient was treated for a long time with a combination of prednisolone and methotrexate. There was a significant positive effect on polyneuropathy, but uncontrolled bronchial asthma persisted. This treatment regimen complies with current recommendations [3,11]. Patients with life-threatening and/or organ-threatening manifestations (including lesions of the central nervous system and severe peripheral neuropathy) require therapy with a combination of a glucocorticoid and a cytostatic [3,11].
The use of immunosuppressive drugs in patients with ANCA-associated vasculitis significantly increases the risk of infection and mortality [16–18]. Prednisone and methotrexate use were found to be significant predictors of hospitalization for pneumonia. Frequent infections (more than 1 episode) are reported in patients with ANCA-associated vasculitis during the first year of diagnosis and initiation of therapy [18]. We assume that bronchiectasis complicated by bacterial colonization and bronchial obstruction in combination with immuno-suppression led to severe pneumonia and death of the patient.
Conclusions
The present case report demonstrates a long-term follow-up of a patient with EGPA. Both common and rare manifestations of EGPA have been described in the patient, such as central nervous system damage and bronchiectasis. Immunosuppressive treatment probably contributed to Klebsiella infection development. Thus, long-term complications of EGPA in combination with adverse effects of medications could have caused the severe pneumonia and death of the patient.
Figure 1. Brain multi-spiral computed tomography performed in 2016 showing ischemic brain damage in the right occipital lobe.
Figure 2. (A–C) Chest computed tomography of patient showing large left upper lobe pneumonia and bronchiectasis in lover lobes.
Figure 3. Timeline from first symptoms to death. | Fatal | ReactionOutcome | CC BY-NC-ND | 33582702 | 18,962,812 | 2021-02-14 |
What was the outcome of reaction 'Disease progression'? | Case Report: A 64-Year-Old Man with 10-Year History of Eosinophilic Granulomatosis with Polyangiitis with Bronchiectasis and Severe Klebsiella pneumonia.
BACKGROUND Eosinophilic granulomatosis with polyangiitis (EGPA) is a rare and severe progressive disease with multiple clinical manifestations and organ damage. Usually, it requires long-term monitoring of the state of many organs due to the gradual character of its manifestations. CASE REPORT We report a case of a long-term follow-up of a patient with eosinophilic granulomatosis with polyangiitis with emphasis on specific clinical features in this patient. A 64-year-old man was being followed up for 10 years. The initial diagnosis was allergic bronchial asthma; however, as new clinical signs and symptoms developed, the diagnosis of EGPA became obvious. A positive treatment response was seen, mainly manifested as reduced polyneuropathy. Meanwhile, bronchial asthma remained uncontrolled and bronchiectasis and Klebsiella pneumoniae colonization developed despite the combination treatment with prednisolone and methotrexate. Furthermore, the patient suffered a cerebral ischemic infarction. During the last hospital admission, severe uncontrolled bronchial asthma complicated with pneumonia resulted in the patient's death. CONCLUSIONS This clinical case shows the gradual development of EGPA with multiple-organ involvement, including respiratory manifestations and peripheral and central nervous system damage. Immunosuppressive treatment combined with complications of EGPA could have contributed to severe pneumonia development and death of the patient.
Background
Systemic vasculitides are difficult to diagnose due to the heterogenous clinical manifestations requiring a wide differential diagnostic search. One of the most uncommon vasculitis types is eosinophilic granulomatosis with polyangiitis (EGPA), or Churg-Strauss syndrome [1,2]. This vasculitis is associated with the presence of antineutrophilic cytoplasmic antibodies (ANCA) and is characterized by necrotizing inflammatory lesions of mainly small vessels, as well as eosinophilic and granulomatous inflammation in various organs. Bronchial asthma and eosinophilia are the most common clinical manifestations of EGPA [1–3]. In 2012, according to the current classification of systemic vasculitides, Churg-Strauss syndrome was renamed “Eosinophilic Granulomatosis with Polyangiitis” (EGPA) and classified into the ANCA-associated vasculitides group [3].
EGPA is a rare ANCA-associated vasculitis. The incidence rate is 0.5–6.8 new cases/year per million populations in Europe [2]. Prevalence is higher in northern latitudes compared to southern Europe [4]. The disease occurs most commonly in people 40–60 years of age [2,5]. No sex predominance has been clearly demonstrated [4].
The pathogenesis of the disease is still not completely clear, but the key role is attributed to autoimmune inflammation with the participation of eosinophils, CD8+, and CD4+lymphocytes (predominantly Th-2) and their proinflammatory cytokines: interleukin (IL)4, IL5, and IL13 [2,5]. Elevation of immunoglobulin (Ig)E, IgG4, and eotaxin-3 is common in EGPA patients [2,5]. The frequent (in 30–40% of cases) detection of ANCA (mainly antibodies to myeloperoxidase) also suggests the autoimmune nature of the disease. ANCA-positive patients are more commonly diagnosed with glomerulonephritis and mononeuritis, whereas the ANCA-negative ones more frequently demonstrate endocardial, myocardial, and pulmonary eosinophilic infiltrates [1,2,5,6].
There are 3 phases in the development of EGPA (these phases partially overlap and may not appear in sequence):
the allergic phase, distinguished by allergic upper respiratory tract inflammation (rhinosinusitis, allergic rhinitis) and bronchial asthma;
the eosinophilic phase, manifested with severe eosinophilia in combination with multiple eosinophilic infiltrates in various organs (most often in the lungs and gastrointestinal tract);
the vasculitis phase, with small- and medium-vessel inflammation of different organs, which can often be life-threatening [5,6].
Respiratory manifestations of EGPA include allergic rhinitis, sinusitis, nasal polyposis and, in more than 95% of patients, poorly controlled bronchial asthma [5]. Chest radiography findings include transient and migratory opacities, ground-glass opacity, focal consolidation, and small peribronchial and centrilobular nodules or masses [7]. Computed tomography reveals bronchial abnormalities, most commonly thickened wall and dilatation with secondary bronchiectasis formation, leading to frequent infections [7,8]. Pleural effusions and alveolar hemorrhages are uncommon manifestations of EGPA [1,5,7,8].
A characteristic neurological manifestation seen in 75% of patients with EGPA is peripheral neuropathy (mononeuritis multiplex) [1,5]. Upper-limb symptoms due to neuropathy, as well as vasculitis in epineural vessels, were more frequently reported in the ANCA-positive group [9]. Eosinophil-associated tissue damage and vascular occlusion leading to ischemia are more specific in ANCA-negative patients [9]. Also, the tissue damage caused by eosinophils may be associated with IL-5 concentration [10]. Brain hemorrhage, subarachnoid hemorrhage, and ischemic brain infarction are rare [1,5,6].
The main drugs for the treatment of EGPA are glucocorticoids. In organ- or life-threatening manifestations, the recommended starting dose of prednisone is 1 mg/kg/day with a maximal daily dose of 80 mg for 2–3 weeks, with tapering to a minimal supporting dose selected individually to control the symptoms and minimize the adverse effects of treatment. Methylprednisolone (7.5–15 mg/kg/day) can be used as a pulse therapy for life-threatening manifestations [11,12]. Most patients need prolonged or lifelong steroids. In the case of organ- or life-threatening manifestations, glucocorticoids in combination with cyclophosphamide are prescribed [11,12]. The combination of glucocorticoids and methotrexate or mycophenolate is required to induce remission in mild or moderate disease [11,12]. Mepolizumab (a humanized monoclonal antibody targeting IL-5) has been used for EGPA treatment [11]. For ANCA-positive patients with renal involvement or refractory disease, rituximab can be considered [3, 11].
Due to the rare occurrence of EGPA and the variety of its clinical manifestations, the diagnosis and treatment of this disease can be challenging. Currently, reports of long-term observations with the detailed patient follow-up are of great interest. The aim of our case report is to present rare manifestations of EGPA, such as bronchiectasis and stroke. We also focused on the infectious complications of immunosuppressive treatments. Thus, we would like to present a 10-year observation of a patient with EGPA.
Case Report
The male patient, born in 1953, was first hospitalized in the Pulmonary Department of our clinic in 2007 due to exacerbated asthma. The patient’s history was remarkable for multiple allergies (to house dust, cat and dog epidermis, and plant pollen) as well as nonsteroidal anti-inflammatory drugs intolerance (manifested as bronchospasm). The patient denied unhealthy habits and occupational hazards (he worked as a driver). His family history was not remarkable. The patient has had allergic rhinitis since adolescence. The first asthma symptoms appeared at the age of 18 while serving in the army. At the age of 22, an allergist diagnosed him with bronchial asthma. The patient had annual exacerbations (1–2 times a year) due to cold or allergen exposure, and acute respiratory viral infections. At the age of 42, polypoid rhinosinusitis was found, and a polypectomy was performed. Since 2006, the patient had been on controller treatment with salmeterol/fluticasone inhaler (25/250 mg, respectively) 2 doses 2 times a day.
In late 2009, extremely elevated eosinophils (53%) and total IgE (1000 IU/L) were found for the first time. Also, since 2009, the patient has had transient lung opacities, which usually disappeared after treatment with prednisolone 30 mg per day.
In August 2010, the patient noticed numbness on the back surface of the left foot, which spread to the entire left lower leg in 1 month, as well as numbness of the right foot and lower leg. Later, the patient started having tingling, twisting, and burning sensations aggravated by touching in the feet and lower legs. Also, the weakness in the distal parts of the legs developed, and he started having difficulties moving around on his own and falling while walking, and he was subsequently hospitalized in our clinic due to exacerbated asthma. On laboratory testing, high eosinophils (53%) and gamma-globulins (27.9%) were found in November 2010. The chest radiography showed emphysema and consolidation in the middle lobe of the right lung. A Neurology consult confirmed asymmetrical distal sensory-motor polyneuropathy demonstrated by lower distal asymmetrical paraparesis with decreased muscle strength in the foot flexors and extensors, more severe on the left side, low tone of the shin muscles, hypoactive Achilles reflexes, and reduced leg sensitivity starting from the middle of the shin and below, more severe in the peroneal nerve inner-vation area. The patient also demonstrated abnormal coordination (heel-to-knee, Romberg) test results and bilateral steppage gait with knee overextension. Electroneuromyography showed axonal lesion of all distal leg nerves and secondary myelinopathy. Thus, EGPA was suspected. The ANCA (anti-myeloperoxidase) titer was 30 U/ml (normal reference below 20 U/ml).
Allergic bronchopulmonary aspergillosis and other causes of hypereosinophilia were consistently ruled out. IgE and IgG for Aspergillus spp. and Aspergillus spp. in sputum were not found.
Thus, the presence of severe bronchial asthma, rhinosinusitis, high and persistent eosinophilia, transient pulmonary opacities, and polyneuropathy suggested EGPA (5 out of 6 clinical criteria of American College of Rheumatology present). Based on the clinical presentation and increased ANCA titer, the patient was diagnosed with EGPA. Systemic steroid treatment (prednisolone 300 mg intravenously and 30 mg orally) was prescribed, and 5 plasmapheresis sessions were performed. Later, prolonged combination immunosuppression (prednisolone 10 mg/day and methotrexate 25 mg/week) and controller asthma treatment (fluticasone propionate/salmeterol 250/25 µg 4 doses/ day, tiotropium bromide 18 µg per day) were prescribed. After 1 month on prednisolone and methotrexate, there was a significant neurological improvement: the motor and sensory disturbances decreased, and the patient was able to move around on his own. Further on, polyneuropathy completely resolved on the combination treatment. His asthma remained uncontrolled despite the combination treatment: he had frequent (3 times a year) severe exacerbations requiring hospitalization. In 2015, newly-emerged copious purulent sputum associated with frequent infection-related asthma exacerbations was noted. In January 2016, a patient underwent chest computed tomography (CT), which showed generalized bronchiectasis. Stenosis of the proximal bronchi was ruled out also by fibrobronchos-copy. The bronchial secretion microbiology revealed Klebsiella pneumoniae growth (107 colony-forming units).
In November 2016, during the asthma exacerbation, the patient suffered a cerebral ischemic infarction involving the right occipital lobe, confirmed by CT (Figures 1, 2).
On March 2, 2017, the patient was admitted to our hospital due to severe respiratory failure caused by pneumonia in the left upper lobe and severe asthma exacerbation.
The complete blood count showed high WBCs (15.2×109/l) with left shift. The IgE level was 144.0 IU/l, C-reactive protein was 48.8 mg/dL. Clinical chemistry was remarkable for urea nitrogen 18.4 mmol/l and creatinine 1.55 mg/dL. The sputum microscopy demonstrated 100 WBCs and 160 RBCs per vision field, and no eosinophils were revealed. A chest CT revealed multiple bronchiectasis and upper lobar pneumonia of the left lung (Figure 2). Klebsiella was again detected in the sputum culture.
The patient was prescribed intravenous antibiotics (ceftriaxone 2 g/day and ciprofloxacin 800 mg/day replaced by cefopera-zone 4 g/day and amikacin 1 g/day after 3 days due to low efficacy), bronchodilator and anti-inflammatory therapy (nebulized ipratropium bromide, fenoterol, and budesonide 2000 µg per day, prednisolone intravenously 150 mg and orally 30 mg). Despite the aggressive treatment, the respiratory failure progressed, and on day 6 of hospitalization the patient was transferred to the Intensive Care Unit and intubated. On day 7, the patient died. On autopsy, the EGPA diagnosis was confirmed, signs of ischemic infarction in the occipital lobe of the brain were found, and left upper lobe pneumonia with abscess formation and generalized bilateral bronchiectases were revealed.
Discussion
Based on the long follow-up period and careful interpretation of signs and symptoms, typical laboratory abnormalities, the diagnosis was clear. Among the most commonly used diagnostic criteria of EGPA are the American College of Rheumatology (ACR) classification criteria, Lanham’s criteria, and the Chapel Hill Consensus Conference criteria. The presence of 4 or more ACR classification criteria, including bronchial asthma, eosinophilia (more than 10%), paranasal sinus abnormalities, migratory or transient eosinophilic pulmonary infiltrates, extravascular eosinophil infiltration on biopsy, and mono- or polyneuropathy, has a sensitivity of 85% and a specificity of 100% [2,5,6]. We identified 5 out of 6 American College of Rheumatology clinical criteria present in our patient.
This clinical case shows gradual development of EGPA with multiple-organ involvement (see Figure 3). The disease onset was characterized by respiratory manifestations and suspicion of EGPA appeared only after the development of polyneuropathy (after 39 years) with hypereosinophilia. Polyneuropathy was noted at the age of 57, which is slightly later than the average age of onset reported in the literature [13]. Notably, the patient was ANCA-positive and his long-term neurological manifestation (mononeuritis) was quite typical for ANCA vasculitis [5,6,8,13]. The diagnosis became obvious after comprehensive assessment of the disease history with its gradually evolving signs and symptoms and laboratory abnormalities. Almost all manifestations of the disease seen in the patient were typical except for stroke, which is an exceptional feature for this case [1,5,6]. Neurological involvement manifested as ischemic stroke is an uncommon complication of EGPA (seen only in 5.2% of all patients) [1]. Thus, the patient demonstrated both peripheral and central nervous system involvement, which is unusual and life-threatening in EGPA.
This clinical case of EGPA also attracted our attention due to severe respiratory manifestations. While typical respiratory manifestations such as rhinosinusitis, asthma, and transient lung infiltrates are well documented in the literature, EGPA-associated bronchiectasis is rarely mentioned [14]. Bronchiectasis in the patient was detected by CT at the age of 63. The patient was a nonsmoker, and no other cause of bronchial abnormalities and airflow obstruction could be identified. Stenosis of the proximal bronchi was ruled out by fibrobronchoscopy and CT. The diagnosis of allergic bronchopulmonary aspergillosis was not confirmed. Klebsiella pneumoniae colonization was found during the last 2 years of the patient’s life. Klebsiella pneumoniae is a typical pathogen causing bronchiectasis [15]. It should be noted that aspergillosis and mycobacteriosis are common complications of bronchiectasis [15], but neither was identified in our patient.
The patient was treated for a long time with a combination of prednisolone and methotrexate. There was a significant positive effect on polyneuropathy, but uncontrolled bronchial asthma persisted. This treatment regimen complies with current recommendations [3,11]. Patients with life-threatening and/or organ-threatening manifestations (including lesions of the central nervous system and severe peripheral neuropathy) require therapy with a combination of a glucocorticoid and a cytostatic [3,11].
The use of immunosuppressive drugs in patients with ANCA-associated vasculitis significantly increases the risk of infection and mortality [16–18]. Prednisone and methotrexate use were found to be significant predictors of hospitalization for pneumonia. Frequent infections (more than 1 episode) are reported in patients with ANCA-associated vasculitis during the first year of diagnosis and initiation of therapy [18]. We assume that bronchiectasis complicated by bacterial colonization and bronchial obstruction in combination with immuno-suppression led to severe pneumonia and death of the patient.
Conclusions
The present case report demonstrates a long-term follow-up of a patient with EGPA. Both common and rare manifestations of EGPA have been described in the patient, such as central nervous system damage and bronchiectasis. Immunosuppressive treatment probably contributed to Klebsiella infection development. Thus, long-term complications of EGPA in combination with adverse effects of medications could have caused the severe pneumonia and death of the patient.
Figure 1. Brain multi-spiral computed tomography performed in 2016 showing ischemic brain damage in the right occipital lobe.
Figure 2. (A–C) Chest computed tomography of patient showing large left upper lobe pneumonia and bronchiectasis in lover lobes.
Figure 3. Timeline from first symptoms to death. | Fatal | ReactionOutcome | CC BY-NC-ND | 33582702 | 18,962,812 | 2021-02-14 |
What was the outcome of reaction 'Drug ineffective'? | Case Report: A 64-Year-Old Man with 10-Year History of Eosinophilic Granulomatosis with Polyangiitis with Bronchiectasis and Severe Klebsiella pneumonia.
BACKGROUND Eosinophilic granulomatosis with polyangiitis (EGPA) is a rare and severe progressive disease with multiple clinical manifestations and organ damage. Usually, it requires long-term monitoring of the state of many organs due to the gradual character of its manifestations. CASE REPORT We report a case of a long-term follow-up of a patient with eosinophilic granulomatosis with polyangiitis with emphasis on specific clinical features in this patient. A 64-year-old man was being followed up for 10 years. The initial diagnosis was allergic bronchial asthma; however, as new clinical signs and symptoms developed, the diagnosis of EGPA became obvious. A positive treatment response was seen, mainly manifested as reduced polyneuropathy. Meanwhile, bronchial asthma remained uncontrolled and bronchiectasis and Klebsiella pneumoniae colonization developed despite the combination treatment with prednisolone and methotrexate. Furthermore, the patient suffered a cerebral ischemic infarction. During the last hospital admission, severe uncontrolled bronchial asthma complicated with pneumonia resulted in the patient's death. CONCLUSIONS This clinical case shows the gradual development of EGPA with multiple-organ involvement, including respiratory manifestations and peripheral and central nervous system damage. Immunosuppressive treatment combined with complications of EGPA could have contributed to severe pneumonia development and death of the patient.
Background
Systemic vasculitides are difficult to diagnose due to the heterogenous clinical manifestations requiring a wide differential diagnostic search. One of the most uncommon vasculitis types is eosinophilic granulomatosis with polyangiitis (EGPA), or Churg-Strauss syndrome [1,2]. This vasculitis is associated with the presence of antineutrophilic cytoplasmic antibodies (ANCA) and is characterized by necrotizing inflammatory lesions of mainly small vessels, as well as eosinophilic and granulomatous inflammation in various organs. Bronchial asthma and eosinophilia are the most common clinical manifestations of EGPA [1–3]. In 2012, according to the current classification of systemic vasculitides, Churg-Strauss syndrome was renamed “Eosinophilic Granulomatosis with Polyangiitis” (EGPA) and classified into the ANCA-associated vasculitides group [3].
EGPA is a rare ANCA-associated vasculitis. The incidence rate is 0.5–6.8 new cases/year per million populations in Europe [2]. Prevalence is higher in northern latitudes compared to southern Europe [4]. The disease occurs most commonly in people 40–60 years of age [2,5]. No sex predominance has been clearly demonstrated [4].
The pathogenesis of the disease is still not completely clear, but the key role is attributed to autoimmune inflammation with the participation of eosinophils, CD8+, and CD4+lymphocytes (predominantly Th-2) and their proinflammatory cytokines: interleukin (IL)4, IL5, and IL13 [2,5]. Elevation of immunoglobulin (Ig)E, IgG4, and eotaxin-3 is common in EGPA patients [2,5]. The frequent (in 30–40% of cases) detection of ANCA (mainly antibodies to myeloperoxidase) also suggests the autoimmune nature of the disease. ANCA-positive patients are more commonly diagnosed with glomerulonephritis and mononeuritis, whereas the ANCA-negative ones more frequently demonstrate endocardial, myocardial, and pulmonary eosinophilic infiltrates [1,2,5,6].
There are 3 phases in the development of EGPA (these phases partially overlap and may not appear in sequence):
the allergic phase, distinguished by allergic upper respiratory tract inflammation (rhinosinusitis, allergic rhinitis) and bronchial asthma;
the eosinophilic phase, manifested with severe eosinophilia in combination with multiple eosinophilic infiltrates in various organs (most often in the lungs and gastrointestinal tract);
the vasculitis phase, with small- and medium-vessel inflammation of different organs, which can often be life-threatening [5,6].
Respiratory manifestations of EGPA include allergic rhinitis, sinusitis, nasal polyposis and, in more than 95% of patients, poorly controlled bronchial asthma [5]. Chest radiography findings include transient and migratory opacities, ground-glass opacity, focal consolidation, and small peribronchial and centrilobular nodules or masses [7]. Computed tomography reveals bronchial abnormalities, most commonly thickened wall and dilatation with secondary bronchiectasis formation, leading to frequent infections [7,8]. Pleural effusions and alveolar hemorrhages are uncommon manifestations of EGPA [1,5,7,8].
A characteristic neurological manifestation seen in 75% of patients with EGPA is peripheral neuropathy (mononeuritis multiplex) [1,5]. Upper-limb symptoms due to neuropathy, as well as vasculitis in epineural vessels, were more frequently reported in the ANCA-positive group [9]. Eosinophil-associated tissue damage and vascular occlusion leading to ischemia are more specific in ANCA-negative patients [9]. Also, the tissue damage caused by eosinophils may be associated with IL-5 concentration [10]. Brain hemorrhage, subarachnoid hemorrhage, and ischemic brain infarction are rare [1,5,6].
The main drugs for the treatment of EGPA are glucocorticoids. In organ- or life-threatening manifestations, the recommended starting dose of prednisone is 1 mg/kg/day with a maximal daily dose of 80 mg for 2–3 weeks, with tapering to a minimal supporting dose selected individually to control the symptoms and minimize the adverse effects of treatment. Methylprednisolone (7.5–15 mg/kg/day) can be used as a pulse therapy for life-threatening manifestations [11,12]. Most patients need prolonged or lifelong steroids. In the case of organ- or life-threatening manifestations, glucocorticoids in combination with cyclophosphamide are prescribed [11,12]. The combination of glucocorticoids and methotrexate or mycophenolate is required to induce remission in mild or moderate disease [11,12]. Mepolizumab (a humanized monoclonal antibody targeting IL-5) has been used for EGPA treatment [11]. For ANCA-positive patients with renal involvement or refractory disease, rituximab can be considered [3, 11].
Due to the rare occurrence of EGPA and the variety of its clinical manifestations, the diagnosis and treatment of this disease can be challenging. Currently, reports of long-term observations with the detailed patient follow-up are of great interest. The aim of our case report is to present rare manifestations of EGPA, such as bronchiectasis and stroke. We also focused on the infectious complications of immunosuppressive treatments. Thus, we would like to present a 10-year observation of a patient with EGPA.
Case Report
The male patient, born in 1953, was first hospitalized in the Pulmonary Department of our clinic in 2007 due to exacerbated asthma. The patient’s history was remarkable for multiple allergies (to house dust, cat and dog epidermis, and plant pollen) as well as nonsteroidal anti-inflammatory drugs intolerance (manifested as bronchospasm). The patient denied unhealthy habits and occupational hazards (he worked as a driver). His family history was not remarkable. The patient has had allergic rhinitis since adolescence. The first asthma symptoms appeared at the age of 18 while serving in the army. At the age of 22, an allergist diagnosed him with bronchial asthma. The patient had annual exacerbations (1–2 times a year) due to cold or allergen exposure, and acute respiratory viral infections. At the age of 42, polypoid rhinosinusitis was found, and a polypectomy was performed. Since 2006, the patient had been on controller treatment with salmeterol/fluticasone inhaler (25/250 mg, respectively) 2 doses 2 times a day.
In late 2009, extremely elevated eosinophils (53%) and total IgE (1000 IU/L) were found for the first time. Also, since 2009, the patient has had transient lung opacities, which usually disappeared after treatment with prednisolone 30 mg per day.
In August 2010, the patient noticed numbness on the back surface of the left foot, which spread to the entire left lower leg in 1 month, as well as numbness of the right foot and lower leg. Later, the patient started having tingling, twisting, and burning sensations aggravated by touching in the feet and lower legs. Also, the weakness in the distal parts of the legs developed, and he started having difficulties moving around on his own and falling while walking, and he was subsequently hospitalized in our clinic due to exacerbated asthma. On laboratory testing, high eosinophils (53%) and gamma-globulins (27.9%) were found in November 2010. The chest radiography showed emphysema and consolidation in the middle lobe of the right lung. A Neurology consult confirmed asymmetrical distal sensory-motor polyneuropathy demonstrated by lower distal asymmetrical paraparesis with decreased muscle strength in the foot flexors and extensors, more severe on the left side, low tone of the shin muscles, hypoactive Achilles reflexes, and reduced leg sensitivity starting from the middle of the shin and below, more severe in the peroneal nerve inner-vation area. The patient also demonstrated abnormal coordination (heel-to-knee, Romberg) test results and bilateral steppage gait with knee overextension. Electroneuromyography showed axonal lesion of all distal leg nerves and secondary myelinopathy. Thus, EGPA was suspected. The ANCA (anti-myeloperoxidase) titer was 30 U/ml (normal reference below 20 U/ml).
Allergic bronchopulmonary aspergillosis and other causes of hypereosinophilia were consistently ruled out. IgE and IgG for Aspergillus spp. and Aspergillus spp. in sputum were not found.
Thus, the presence of severe bronchial asthma, rhinosinusitis, high and persistent eosinophilia, transient pulmonary opacities, and polyneuropathy suggested EGPA (5 out of 6 clinical criteria of American College of Rheumatology present). Based on the clinical presentation and increased ANCA titer, the patient was diagnosed with EGPA. Systemic steroid treatment (prednisolone 300 mg intravenously and 30 mg orally) was prescribed, and 5 plasmapheresis sessions were performed. Later, prolonged combination immunosuppression (prednisolone 10 mg/day and methotrexate 25 mg/week) and controller asthma treatment (fluticasone propionate/salmeterol 250/25 µg 4 doses/ day, tiotropium bromide 18 µg per day) were prescribed. After 1 month on prednisolone and methotrexate, there was a significant neurological improvement: the motor and sensory disturbances decreased, and the patient was able to move around on his own. Further on, polyneuropathy completely resolved on the combination treatment. His asthma remained uncontrolled despite the combination treatment: he had frequent (3 times a year) severe exacerbations requiring hospitalization. In 2015, newly-emerged copious purulent sputum associated with frequent infection-related asthma exacerbations was noted. In January 2016, a patient underwent chest computed tomography (CT), which showed generalized bronchiectasis. Stenosis of the proximal bronchi was ruled out also by fibrobronchos-copy. The bronchial secretion microbiology revealed Klebsiella pneumoniae growth (107 colony-forming units).
In November 2016, during the asthma exacerbation, the patient suffered a cerebral ischemic infarction involving the right occipital lobe, confirmed by CT (Figures 1, 2).
On March 2, 2017, the patient was admitted to our hospital due to severe respiratory failure caused by pneumonia in the left upper lobe and severe asthma exacerbation.
The complete blood count showed high WBCs (15.2×109/l) with left shift. The IgE level was 144.0 IU/l, C-reactive protein was 48.8 mg/dL. Clinical chemistry was remarkable for urea nitrogen 18.4 mmol/l and creatinine 1.55 mg/dL. The sputum microscopy demonstrated 100 WBCs and 160 RBCs per vision field, and no eosinophils were revealed. A chest CT revealed multiple bronchiectasis and upper lobar pneumonia of the left lung (Figure 2). Klebsiella was again detected in the sputum culture.
The patient was prescribed intravenous antibiotics (ceftriaxone 2 g/day and ciprofloxacin 800 mg/day replaced by cefopera-zone 4 g/day and amikacin 1 g/day after 3 days due to low efficacy), bronchodilator and anti-inflammatory therapy (nebulized ipratropium bromide, fenoterol, and budesonide 2000 µg per day, prednisolone intravenously 150 mg and orally 30 mg). Despite the aggressive treatment, the respiratory failure progressed, and on day 6 of hospitalization the patient was transferred to the Intensive Care Unit and intubated. On day 7, the patient died. On autopsy, the EGPA diagnosis was confirmed, signs of ischemic infarction in the occipital lobe of the brain were found, and left upper lobe pneumonia with abscess formation and generalized bilateral bronchiectases were revealed.
Discussion
Based on the long follow-up period and careful interpretation of signs and symptoms, typical laboratory abnormalities, the diagnosis was clear. Among the most commonly used diagnostic criteria of EGPA are the American College of Rheumatology (ACR) classification criteria, Lanham’s criteria, and the Chapel Hill Consensus Conference criteria. The presence of 4 or more ACR classification criteria, including bronchial asthma, eosinophilia (more than 10%), paranasal sinus abnormalities, migratory or transient eosinophilic pulmonary infiltrates, extravascular eosinophil infiltration on biopsy, and mono- or polyneuropathy, has a sensitivity of 85% and a specificity of 100% [2,5,6]. We identified 5 out of 6 American College of Rheumatology clinical criteria present in our patient.
This clinical case shows gradual development of EGPA with multiple-organ involvement (see Figure 3). The disease onset was characterized by respiratory manifestations and suspicion of EGPA appeared only after the development of polyneuropathy (after 39 years) with hypereosinophilia. Polyneuropathy was noted at the age of 57, which is slightly later than the average age of onset reported in the literature [13]. Notably, the patient was ANCA-positive and his long-term neurological manifestation (mononeuritis) was quite typical for ANCA vasculitis [5,6,8,13]. The diagnosis became obvious after comprehensive assessment of the disease history with its gradually evolving signs and symptoms and laboratory abnormalities. Almost all manifestations of the disease seen in the patient were typical except for stroke, which is an exceptional feature for this case [1,5,6]. Neurological involvement manifested as ischemic stroke is an uncommon complication of EGPA (seen only in 5.2% of all patients) [1]. Thus, the patient demonstrated both peripheral and central nervous system involvement, which is unusual and life-threatening in EGPA.
This clinical case of EGPA also attracted our attention due to severe respiratory manifestations. While typical respiratory manifestations such as rhinosinusitis, asthma, and transient lung infiltrates are well documented in the literature, EGPA-associated bronchiectasis is rarely mentioned [14]. Bronchiectasis in the patient was detected by CT at the age of 63. The patient was a nonsmoker, and no other cause of bronchial abnormalities and airflow obstruction could be identified. Stenosis of the proximal bronchi was ruled out by fibrobronchoscopy and CT. The diagnosis of allergic bronchopulmonary aspergillosis was not confirmed. Klebsiella pneumoniae colonization was found during the last 2 years of the patient’s life. Klebsiella pneumoniae is a typical pathogen causing bronchiectasis [15]. It should be noted that aspergillosis and mycobacteriosis are common complications of bronchiectasis [15], but neither was identified in our patient.
The patient was treated for a long time with a combination of prednisolone and methotrexate. There was a significant positive effect on polyneuropathy, but uncontrolled bronchial asthma persisted. This treatment regimen complies with current recommendations [3,11]. Patients with life-threatening and/or organ-threatening manifestations (including lesions of the central nervous system and severe peripheral neuropathy) require therapy with a combination of a glucocorticoid and a cytostatic [3,11].
The use of immunosuppressive drugs in patients with ANCA-associated vasculitis significantly increases the risk of infection and mortality [16–18]. Prednisone and methotrexate use were found to be significant predictors of hospitalization for pneumonia. Frequent infections (more than 1 episode) are reported in patients with ANCA-associated vasculitis during the first year of diagnosis and initiation of therapy [18]. We assume that bronchiectasis complicated by bacterial colonization and bronchial obstruction in combination with immuno-suppression led to severe pneumonia and death of the patient.
Conclusions
The present case report demonstrates a long-term follow-up of a patient with EGPA. Both common and rare manifestations of EGPA have been described in the patient, such as central nervous system damage and bronchiectasis. Immunosuppressive treatment probably contributed to Klebsiella infection development. Thus, long-term complications of EGPA in combination with adverse effects of medications could have caused the severe pneumonia and death of the patient.
Figure 1. Brain multi-spiral computed tomography performed in 2016 showing ischemic brain damage in the right occipital lobe.
Figure 2. (A–C) Chest computed tomography of patient showing large left upper lobe pneumonia and bronchiectasis in lover lobes.
Figure 3. Timeline from first symptoms to death. | Fatal | ReactionOutcome | CC BY-NC-ND | 33582702 | 18,986,567 | 2021-02-14 |
What was the outcome of reaction 'Klebsiella infection'? | Case Report: A 64-Year-Old Man with 10-Year History of Eosinophilic Granulomatosis with Polyangiitis with Bronchiectasis and Severe Klebsiella pneumonia.
BACKGROUND Eosinophilic granulomatosis with polyangiitis (EGPA) is a rare and severe progressive disease with multiple clinical manifestations and organ damage. Usually, it requires long-term monitoring of the state of many organs due to the gradual character of its manifestations. CASE REPORT We report a case of a long-term follow-up of a patient with eosinophilic granulomatosis with polyangiitis with emphasis on specific clinical features in this patient. A 64-year-old man was being followed up for 10 years. The initial diagnosis was allergic bronchial asthma; however, as new clinical signs and symptoms developed, the diagnosis of EGPA became obvious. A positive treatment response was seen, mainly manifested as reduced polyneuropathy. Meanwhile, bronchial asthma remained uncontrolled and bronchiectasis and Klebsiella pneumoniae colonization developed despite the combination treatment with prednisolone and methotrexate. Furthermore, the patient suffered a cerebral ischemic infarction. During the last hospital admission, severe uncontrolled bronchial asthma complicated with pneumonia resulted in the patient's death. CONCLUSIONS This clinical case shows the gradual development of EGPA with multiple-organ involvement, including respiratory manifestations and peripheral and central nervous system damage. Immunosuppressive treatment combined with complications of EGPA could have contributed to severe pneumonia development and death of the patient.
Background
Systemic vasculitides are difficult to diagnose due to the heterogenous clinical manifestations requiring a wide differential diagnostic search. One of the most uncommon vasculitis types is eosinophilic granulomatosis with polyangiitis (EGPA), or Churg-Strauss syndrome [1,2]. This vasculitis is associated with the presence of antineutrophilic cytoplasmic antibodies (ANCA) and is characterized by necrotizing inflammatory lesions of mainly small vessels, as well as eosinophilic and granulomatous inflammation in various organs. Bronchial asthma and eosinophilia are the most common clinical manifestations of EGPA [1–3]. In 2012, according to the current classification of systemic vasculitides, Churg-Strauss syndrome was renamed “Eosinophilic Granulomatosis with Polyangiitis” (EGPA) and classified into the ANCA-associated vasculitides group [3].
EGPA is a rare ANCA-associated vasculitis. The incidence rate is 0.5–6.8 new cases/year per million populations in Europe [2]. Prevalence is higher in northern latitudes compared to southern Europe [4]. The disease occurs most commonly in people 40–60 years of age [2,5]. No sex predominance has been clearly demonstrated [4].
The pathogenesis of the disease is still not completely clear, but the key role is attributed to autoimmune inflammation with the participation of eosinophils, CD8+, and CD4+lymphocytes (predominantly Th-2) and their proinflammatory cytokines: interleukin (IL)4, IL5, and IL13 [2,5]. Elevation of immunoglobulin (Ig)E, IgG4, and eotaxin-3 is common in EGPA patients [2,5]. The frequent (in 30–40% of cases) detection of ANCA (mainly antibodies to myeloperoxidase) also suggests the autoimmune nature of the disease. ANCA-positive patients are more commonly diagnosed with glomerulonephritis and mononeuritis, whereas the ANCA-negative ones more frequently demonstrate endocardial, myocardial, and pulmonary eosinophilic infiltrates [1,2,5,6].
There are 3 phases in the development of EGPA (these phases partially overlap and may not appear in sequence):
the allergic phase, distinguished by allergic upper respiratory tract inflammation (rhinosinusitis, allergic rhinitis) and bronchial asthma;
the eosinophilic phase, manifested with severe eosinophilia in combination with multiple eosinophilic infiltrates in various organs (most often in the lungs and gastrointestinal tract);
the vasculitis phase, with small- and medium-vessel inflammation of different organs, which can often be life-threatening [5,6].
Respiratory manifestations of EGPA include allergic rhinitis, sinusitis, nasal polyposis and, in more than 95% of patients, poorly controlled bronchial asthma [5]. Chest radiography findings include transient and migratory opacities, ground-glass opacity, focal consolidation, and small peribronchial and centrilobular nodules or masses [7]. Computed tomography reveals bronchial abnormalities, most commonly thickened wall and dilatation with secondary bronchiectasis formation, leading to frequent infections [7,8]. Pleural effusions and alveolar hemorrhages are uncommon manifestations of EGPA [1,5,7,8].
A characteristic neurological manifestation seen in 75% of patients with EGPA is peripheral neuropathy (mononeuritis multiplex) [1,5]. Upper-limb symptoms due to neuropathy, as well as vasculitis in epineural vessels, were more frequently reported in the ANCA-positive group [9]. Eosinophil-associated tissue damage and vascular occlusion leading to ischemia are more specific in ANCA-negative patients [9]. Also, the tissue damage caused by eosinophils may be associated with IL-5 concentration [10]. Brain hemorrhage, subarachnoid hemorrhage, and ischemic brain infarction are rare [1,5,6].
The main drugs for the treatment of EGPA are glucocorticoids. In organ- or life-threatening manifestations, the recommended starting dose of prednisone is 1 mg/kg/day with a maximal daily dose of 80 mg for 2–3 weeks, with tapering to a minimal supporting dose selected individually to control the symptoms and minimize the adverse effects of treatment. Methylprednisolone (7.5–15 mg/kg/day) can be used as a pulse therapy for life-threatening manifestations [11,12]. Most patients need prolonged or lifelong steroids. In the case of organ- or life-threatening manifestations, glucocorticoids in combination with cyclophosphamide are prescribed [11,12]. The combination of glucocorticoids and methotrexate or mycophenolate is required to induce remission in mild or moderate disease [11,12]. Mepolizumab (a humanized monoclonal antibody targeting IL-5) has been used for EGPA treatment [11]. For ANCA-positive patients with renal involvement or refractory disease, rituximab can be considered [3, 11].
Due to the rare occurrence of EGPA and the variety of its clinical manifestations, the diagnosis and treatment of this disease can be challenging. Currently, reports of long-term observations with the detailed patient follow-up are of great interest. The aim of our case report is to present rare manifestations of EGPA, such as bronchiectasis and stroke. We also focused on the infectious complications of immunosuppressive treatments. Thus, we would like to present a 10-year observation of a patient with EGPA.
Case Report
The male patient, born in 1953, was first hospitalized in the Pulmonary Department of our clinic in 2007 due to exacerbated asthma. The patient’s history was remarkable for multiple allergies (to house dust, cat and dog epidermis, and plant pollen) as well as nonsteroidal anti-inflammatory drugs intolerance (manifested as bronchospasm). The patient denied unhealthy habits and occupational hazards (he worked as a driver). His family history was not remarkable. The patient has had allergic rhinitis since adolescence. The first asthma symptoms appeared at the age of 18 while serving in the army. At the age of 22, an allergist diagnosed him with bronchial asthma. The patient had annual exacerbations (1–2 times a year) due to cold or allergen exposure, and acute respiratory viral infections. At the age of 42, polypoid rhinosinusitis was found, and a polypectomy was performed. Since 2006, the patient had been on controller treatment with salmeterol/fluticasone inhaler (25/250 mg, respectively) 2 doses 2 times a day.
In late 2009, extremely elevated eosinophils (53%) and total IgE (1000 IU/L) were found for the first time. Also, since 2009, the patient has had transient lung opacities, which usually disappeared after treatment with prednisolone 30 mg per day.
In August 2010, the patient noticed numbness on the back surface of the left foot, which spread to the entire left lower leg in 1 month, as well as numbness of the right foot and lower leg. Later, the patient started having tingling, twisting, and burning sensations aggravated by touching in the feet and lower legs. Also, the weakness in the distal parts of the legs developed, and he started having difficulties moving around on his own and falling while walking, and he was subsequently hospitalized in our clinic due to exacerbated asthma. On laboratory testing, high eosinophils (53%) and gamma-globulins (27.9%) were found in November 2010. The chest radiography showed emphysema and consolidation in the middle lobe of the right lung. A Neurology consult confirmed asymmetrical distal sensory-motor polyneuropathy demonstrated by lower distal asymmetrical paraparesis with decreased muscle strength in the foot flexors and extensors, more severe on the left side, low tone of the shin muscles, hypoactive Achilles reflexes, and reduced leg sensitivity starting from the middle of the shin and below, more severe in the peroneal nerve inner-vation area. The patient also demonstrated abnormal coordination (heel-to-knee, Romberg) test results and bilateral steppage gait with knee overextension. Electroneuromyography showed axonal lesion of all distal leg nerves and secondary myelinopathy. Thus, EGPA was suspected. The ANCA (anti-myeloperoxidase) titer was 30 U/ml (normal reference below 20 U/ml).
Allergic bronchopulmonary aspergillosis and other causes of hypereosinophilia were consistently ruled out. IgE and IgG for Aspergillus spp. and Aspergillus spp. in sputum were not found.
Thus, the presence of severe bronchial asthma, rhinosinusitis, high and persistent eosinophilia, transient pulmonary opacities, and polyneuropathy suggested EGPA (5 out of 6 clinical criteria of American College of Rheumatology present). Based on the clinical presentation and increased ANCA titer, the patient was diagnosed with EGPA. Systemic steroid treatment (prednisolone 300 mg intravenously and 30 mg orally) was prescribed, and 5 plasmapheresis sessions were performed. Later, prolonged combination immunosuppression (prednisolone 10 mg/day and methotrexate 25 mg/week) and controller asthma treatment (fluticasone propionate/salmeterol 250/25 µg 4 doses/ day, tiotropium bromide 18 µg per day) were prescribed. After 1 month on prednisolone and methotrexate, there was a significant neurological improvement: the motor and sensory disturbances decreased, and the patient was able to move around on his own. Further on, polyneuropathy completely resolved on the combination treatment. His asthma remained uncontrolled despite the combination treatment: he had frequent (3 times a year) severe exacerbations requiring hospitalization. In 2015, newly-emerged copious purulent sputum associated with frequent infection-related asthma exacerbations was noted. In January 2016, a patient underwent chest computed tomography (CT), which showed generalized bronchiectasis. Stenosis of the proximal bronchi was ruled out also by fibrobronchos-copy. The bronchial secretion microbiology revealed Klebsiella pneumoniae growth (107 colony-forming units).
In November 2016, during the asthma exacerbation, the patient suffered a cerebral ischemic infarction involving the right occipital lobe, confirmed by CT (Figures 1, 2).
On March 2, 2017, the patient was admitted to our hospital due to severe respiratory failure caused by pneumonia in the left upper lobe and severe asthma exacerbation.
The complete blood count showed high WBCs (15.2×109/l) with left shift. The IgE level was 144.0 IU/l, C-reactive protein was 48.8 mg/dL. Clinical chemistry was remarkable for urea nitrogen 18.4 mmol/l and creatinine 1.55 mg/dL. The sputum microscopy demonstrated 100 WBCs and 160 RBCs per vision field, and no eosinophils were revealed. A chest CT revealed multiple bronchiectasis and upper lobar pneumonia of the left lung (Figure 2). Klebsiella was again detected in the sputum culture.
The patient was prescribed intravenous antibiotics (ceftriaxone 2 g/day and ciprofloxacin 800 mg/day replaced by cefopera-zone 4 g/day and amikacin 1 g/day after 3 days due to low efficacy), bronchodilator and anti-inflammatory therapy (nebulized ipratropium bromide, fenoterol, and budesonide 2000 µg per day, prednisolone intravenously 150 mg and orally 30 mg). Despite the aggressive treatment, the respiratory failure progressed, and on day 6 of hospitalization the patient was transferred to the Intensive Care Unit and intubated. On day 7, the patient died. On autopsy, the EGPA diagnosis was confirmed, signs of ischemic infarction in the occipital lobe of the brain were found, and left upper lobe pneumonia with abscess formation and generalized bilateral bronchiectases were revealed.
Discussion
Based on the long follow-up period and careful interpretation of signs and symptoms, typical laboratory abnormalities, the diagnosis was clear. Among the most commonly used diagnostic criteria of EGPA are the American College of Rheumatology (ACR) classification criteria, Lanham’s criteria, and the Chapel Hill Consensus Conference criteria. The presence of 4 or more ACR classification criteria, including bronchial asthma, eosinophilia (more than 10%), paranasal sinus abnormalities, migratory or transient eosinophilic pulmonary infiltrates, extravascular eosinophil infiltration on biopsy, and mono- or polyneuropathy, has a sensitivity of 85% and a specificity of 100% [2,5,6]. We identified 5 out of 6 American College of Rheumatology clinical criteria present in our patient.
This clinical case shows gradual development of EGPA with multiple-organ involvement (see Figure 3). The disease onset was characterized by respiratory manifestations and suspicion of EGPA appeared only after the development of polyneuropathy (after 39 years) with hypereosinophilia. Polyneuropathy was noted at the age of 57, which is slightly later than the average age of onset reported in the literature [13]. Notably, the patient was ANCA-positive and his long-term neurological manifestation (mononeuritis) was quite typical for ANCA vasculitis [5,6,8,13]. The diagnosis became obvious after comprehensive assessment of the disease history with its gradually evolving signs and symptoms and laboratory abnormalities. Almost all manifestations of the disease seen in the patient were typical except for stroke, which is an exceptional feature for this case [1,5,6]. Neurological involvement manifested as ischemic stroke is an uncommon complication of EGPA (seen only in 5.2% of all patients) [1]. Thus, the patient demonstrated both peripheral and central nervous system involvement, which is unusual and life-threatening in EGPA.
This clinical case of EGPA also attracted our attention due to severe respiratory manifestations. While typical respiratory manifestations such as rhinosinusitis, asthma, and transient lung infiltrates are well documented in the literature, EGPA-associated bronchiectasis is rarely mentioned [14]. Bronchiectasis in the patient was detected by CT at the age of 63. The patient was a nonsmoker, and no other cause of bronchial abnormalities and airflow obstruction could be identified. Stenosis of the proximal bronchi was ruled out by fibrobronchoscopy and CT. The diagnosis of allergic bronchopulmonary aspergillosis was not confirmed. Klebsiella pneumoniae colonization was found during the last 2 years of the patient’s life. Klebsiella pneumoniae is a typical pathogen causing bronchiectasis [15]. It should be noted that aspergillosis and mycobacteriosis are common complications of bronchiectasis [15], but neither was identified in our patient.
The patient was treated for a long time with a combination of prednisolone and methotrexate. There was a significant positive effect on polyneuropathy, but uncontrolled bronchial asthma persisted. This treatment regimen complies with current recommendations [3,11]. Patients with life-threatening and/or organ-threatening manifestations (including lesions of the central nervous system and severe peripheral neuropathy) require therapy with a combination of a glucocorticoid and a cytostatic [3,11].
The use of immunosuppressive drugs in patients with ANCA-associated vasculitis significantly increases the risk of infection and mortality [16–18]. Prednisone and methotrexate use were found to be significant predictors of hospitalization for pneumonia. Frequent infections (more than 1 episode) are reported in patients with ANCA-associated vasculitis during the first year of diagnosis and initiation of therapy [18]. We assume that bronchiectasis complicated by bacterial colonization and bronchial obstruction in combination with immuno-suppression led to severe pneumonia and death of the patient.
Conclusions
The present case report demonstrates a long-term follow-up of a patient with EGPA. Both common and rare manifestations of EGPA have been described in the patient, such as central nervous system damage and bronchiectasis. Immunosuppressive treatment probably contributed to Klebsiella infection development. Thus, long-term complications of EGPA in combination with adverse effects of medications could have caused the severe pneumonia and death of the patient.
Figure 1. Brain multi-spiral computed tomography performed in 2016 showing ischemic brain damage in the right occipital lobe.
Figure 2. (A–C) Chest computed tomography of patient showing large left upper lobe pneumonia and bronchiectasis in lover lobes.
Figure 3. Timeline from first symptoms to death. | Fatal | ReactionOutcome | CC BY-NC-ND | 33582702 | 18,975,017 | 2021-02-14 |
What was the outcome of reaction 'Pneumonia klebsiella'? | Case Report: A 64-Year-Old Man with 10-Year History of Eosinophilic Granulomatosis with Polyangiitis with Bronchiectasis and Severe Klebsiella pneumonia.
BACKGROUND Eosinophilic granulomatosis with polyangiitis (EGPA) is a rare and severe progressive disease with multiple clinical manifestations and organ damage. Usually, it requires long-term monitoring of the state of many organs due to the gradual character of its manifestations. CASE REPORT We report a case of a long-term follow-up of a patient with eosinophilic granulomatosis with polyangiitis with emphasis on specific clinical features in this patient. A 64-year-old man was being followed up for 10 years. The initial diagnosis was allergic bronchial asthma; however, as new clinical signs and symptoms developed, the diagnosis of EGPA became obvious. A positive treatment response was seen, mainly manifested as reduced polyneuropathy. Meanwhile, bronchial asthma remained uncontrolled and bronchiectasis and Klebsiella pneumoniae colonization developed despite the combination treatment with prednisolone and methotrexate. Furthermore, the patient suffered a cerebral ischemic infarction. During the last hospital admission, severe uncontrolled bronchial asthma complicated with pneumonia resulted in the patient's death. CONCLUSIONS This clinical case shows the gradual development of EGPA with multiple-organ involvement, including respiratory manifestations and peripheral and central nervous system damage. Immunosuppressive treatment combined with complications of EGPA could have contributed to severe pneumonia development and death of the patient.
Background
Systemic vasculitides are difficult to diagnose due to the heterogenous clinical manifestations requiring a wide differential diagnostic search. One of the most uncommon vasculitis types is eosinophilic granulomatosis with polyangiitis (EGPA), or Churg-Strauss syndrome [1,2]. This vasculitis is associated with the presence of antineutrophilic cytoplasmic antibodies (ANCA) and is characterized by necrotizing inflammatory lesions of mainly small vessels, as well as eosinophilic and granulomatous inflammation in various organs. Bronchial asthma and eosinophilia are the most common clinical manifestations of EGPA [1–3]. In 2012, according to the current classification of systemic vasculitides, Churg-Strauss syndrome was renamed “Eosinophilic Granulomatosis with Polyangiitis” (EGPA) and classified into the ANCA-associated vasculitides group [3].
EGPA is a rare ANCA-associated vasculitis. The incidence rate is 0.5–6.8 new cases/year per million populations in Europe [2]. Prevalence is higher in northern latitudes compared to southern Europe [4]. The disease occurs most commonly in people 40–60 years of age [2,5]. No sex predominance has been clearly demonstrated [4].
The pathogenesis of the disease is still not completely clear, but the key role is attributed to autoimmune inflammation with the participation of eosinophils, CD8+, and CD4+lymphocytes (predominantly Th-2) and their proinflammatory cytokines: interleukin (IL)4, IL5, and IL13 [2,5]. Elevation of immunoglobulin (Ig)E, IgG4, and eotaxin-3 is common in EGPA patients [2,5]. The frequent (in 30–40% of cases) detection of ANCA (mainly antibodies to myeloperoxidase) also suggests the autoimmune nature of the disease. ANCA-positive patients are more commonly diagnosed with glomerulonephritis and mononeuritis, whereas the ANCA-negative ones more frequently demonstrate endocardial, myocardial, and pulmonary eosinophilic infiltrates [1,2,5,6].
There are 3 phases in the development of EGPA (these phases partially overlap and may not appear in sequence):
the allergic phase, distinguished by allergic upper respiratory tract inflammation (rhinosinusitis, allergic rhinitis) and bronchial asthma;
the eosinophilic phase, manifested with severe eosinophilia in combination with multiple eosinophilic infiltrates in various organs (most often in the lungs and gastrointestinal tract);
the vasculitis phase, with small- and medium-vessel inflammation of different organs, which can often be life-threatening [5,6].
Respiratory manifestations of EGPA include allergic rhinitis, sinusitis, nasal polyposis and, in more than 95% of patients, poorly controlled bronchial asthma [5]. Chest radiography findings include transient and migratory opacities, ground-glass opacity, focal consolidation, and small peribronchial and centrilobular nodules or masses [7]. Computed tomography reveals bronchial abnormalities, most commonly thickened wall and dilatation with secondary bronchiectasis formation, leading to frequent infections [7,8]. Pleural effusions and alveolar hemorrhages are uncommon manifestations of EGPA [1,5,7,8].
A characteristic neurological manifestation seen in 75% of patients with EGPA is peripheral neuropathy (mononeuritis multiplex) [1,5]. Upper-limb symptoms due to neuropathy, as well as vasculitis in epineural vessels, were more frequently reported in the ANCA-positive group [9]. Eosinophil-associated tissue damage and vascular occlusion leading to ischemia are more specific in ANCA-negative patients [9]. Also, the tissue damage caused by eosinophils may be associated with IL-5 concentration [10]. Brain hemorrhage, subarachnoid hemorrhage, and ischemic brain infarction are rare [1,5,6].
The main drugs for the treatment of EGPA are glucocorticoids. In organ- or life-threatening manifestations, the recommended starting dose of prednisone is 1 mg/kg/day with a maximal daily dose of 80 mg for 2–3 weeks, with tapering to a minimal supporting dose selected individually to control the symptoms and minimize the adverse effects of treatment. Methylprednisolone (7.5–15 mg/kg/day) can be used as a pulse therapy for life-threatening manifestations [11,12]. Most patients need prolonged or lifelong steroids. In the case of organ- or life-threatening manifestations, glucocorticoids in combination with cyclophosphamide are prescribed [11,12]. The combination of glucocorticoids and methotrexate or mycophenolate is required to induce remission in mild or moderate disease [11,12]. Mepolizumab (a humanized monoclonal antibody targeting IL-5) has been used for EGPA treatment [11]. For ANCA-positive patients with renal involvement or refractory disease, rituximab can be considered [3, 11].
Due to the rare occurrence of EGPA and the variety of its clinical manifestations, the diagnosis and treatment of this disease can be challenging. Currently, reports of long-term observations with the detailed patient follow-up are of great interest. The aim of our case report is to present rare manifestations of EGPA, such as bronchiectasis and stroke. We also focused on the infectious complications of immunosuppressive treatments. Thus, we would like to present a 10-year observation of a patient with EGPA.
Case Report
The male patient, born in 1953, was first hospitalized in the Pulmonary Department of our clinic in 2007 due to exacerbated asthma. The patient’s history was remarkable for multiple allergies (to house dust, cat and dog epidermis, and plant pollen) as well as nonsteroidal anti-inflammatory drugs intolerance (manifested as bronchospasm). The patient denied unhealthy habits and occupational hazards (he worked as a driver). His family history was not remarkable. The patient has had allergic rhinitis since adolescence. The first asthma symptoms appeared at the age of 18 while serving in the army. At the age of 22, an allergist diagnosed him with bronchial asthma. The patient had annual exacerbations (1–2 times a year) due to cold or allergen exposure, and acute respiratory viral infections. At the age of 42, polypoid rhinosinusitis was found, and a polypectomy was performed. Since 2006, the patient had been on controller treatment with salmeterol/fluticasone inhaler (25/250 mg, respectively) 2 doses 2 times a day.
In late 2009, extremely elevated eosinophils (53%) and total IgE (1000 IU/L) were found for the first time. Also, since 2009, the patient has had transient lung opacities, which usually disappeared after treatment with prednisolone 30 mg per day.
In August 2010, the patient noticed numbness on the back surface of the left foot, which spread to the entire left lower leg in 1 month, as well as numbness of the right foot and lower leg. Later, the patient started having tingling, twisting, and burning sensations aggravated by touching in the feet and lower legs. Also, the weakness in the distal parts of the legs developed, and he started having difficulties moving around on his own and falling while walking, and he was subsequently hospitalized in our clinic due to exacerbated asthma. On laboratory testing, high eosinophils (53%) and gamma-globulins (27.9%) were found in November 2010. The chest radiography showed emphysema and consolidation in the middle lobe of the right lung. A Neurology consult confirmed asymmetrical distal sensory-motor polyneuropathy demonstrated by lower distal asymmetrical paraparesis with decreased muscle strength in the foot flexors and extensors, more severe on the left side, low tone of the shin muscles, hypoactive Achilles reflexes, and reduced leg sensitivity starting from the middle of the shin and below, more severe in the peroneal nerve inner-vation area. The patient also demonstrated abnormal coordination (heel-to-knee, Romberg) test results and bilateral steppage gait with knee overextension. Electroneuromyography showed axonal lesion of all distal leg nerves and secondary myelinopathy. Thus, EGPA was suspected. The ANCA (anti-myeloperoxidase) titer was 30 U/ml (normal reference below 20 U/ml).
Allergic bronchopulmonary aspergillosis and other causes of hypereosinophilia were consistently ruled out. IgE and IgG for Aspergillus spp. and Aspergillus spp. in sputum were not found.
Thus, the presence of severe bronchial asthma, rhinosinusitis, high and persistent eosinophilia, transient pulmonary opacities, and polyneuropathy suggested EGPA (5 out of 6 clinical criteria of American College of Rheumatology present). Based on the clinical presentation and increased ANCA titer, the patient was diagnosed with EGPA. Systemic steroid treatment (prednisolone 300 mg intravenously and 30 mg orally) was prescribed, and 5 plasmapheresis sessions were performed. Later, prolonged combination immunosuppression (prednisolone 10 mg/day and methotrexate 25 mg/week) and controller asthma treatment (fluticasone propionate/salmeterol 250/25 µg 4 doses/ day, tiotropium bromide 18 µg per day) were prescribed. After 1 month on prednisolone and methotrexate, there was a significant neurological improvement: the motor and sensory disturbances decreased, and the patient was able to move around on his own. Further on, polyneuropathy completely resolved on the combination treatment. His asthma remained uncontrolled despite the combination treatment: he had frequent (3 times a year) severe exacerbations requiring hospitalization. In 2015, newly-emerged copious purulent sputum associated with frequent infection-related asthma exacerbations was noted. In January 2016, a patient underwent chest computed tomography (CT), which showed generalized bronchiectasis. Stenosis of the proximal bronchi was ruled out also by fibrobronchos-copy. The bronchial secretion microbiology revealed Klebsiella pneumoniae growth (107 colony-forming units).
In November 2016, during the asthma exacerbation, the patient suffered a cerebral ischemic infarction involving the right occipital lobe, confirmed by CT (Figures 1, 2).
On March 2, 2017, the patient was admitted to our hospital due to severe respiratory failure caused by pneumonia in the left upper lobe and severe asthma exacerbation.
The complete blood count showed high WBCs (15.2×109/l) with left shift. The IgE level was 144.0 IU/l, C-reactive protein was 48.8 mg/dL. Clinical chemistry was remarkable for urea nitrogen 18.4 mmol/l and creatinine 1.55 mg/dL. The sputum microscopy demonstrated 100 WBCs and 160 RBCs per vision field, and no eosinophils were revealed. A chest CT revealed multiple bronchiectasis and upper lobar pneumonia of the left lung (Figure 2). Klebsiella was again detected in the sputum culture.
The patient was prescribed intravenous antibiotics (ceftriaxone 2 g/day and ciprofloxacin 800 mg/day replaced by cefopera-zone 4 g/day and amikacin 1 g/day after 3 days due to low efficacy), bronchodilator and anti-inflammatory therapy (nebulized ipratropium bromide, fenoterol, and budesonide 2000 µg per day, prednisolone intravenously 150 mg and orally 30 mg). Despite the aggressive treatment, the respiratory failure progressed, and on day 6 of hospitalization the patient was transferred to the Intensive Care Unit and intubated. On day 7, the patient died. On autopsy, the EGPA diagnosis was confirmed, signs of ischemic infarction in the occipital lobe of the brain were found, and left upper lobe pneumonia with abscess formation and generalized bilateral bronchiectases were revealed.
Discussion
Based on the long follow-up period and careful interpretation of signs and symptoms, typical laboratory abnormalities, the diagnosis was clear. Among the most commonly used diagnostic criteria of EGPA are the American College of Rheumatology (ACR) classification criteria, Lanham’s criteria, and the Chapel Hill Consensus Conference criteria. The presence of 4 or more ACR classification criteria, including bronchial asthma, eosinophilia (more than 10%), paranasal sinus abnormalities, migratory or transient eosinophilic pulmonary infiltrates, extravascular eosinophil infiltration on biopsy, and mono- or polyneuropathy, has a sensitivity of 85% and a specificity of 100% [2,5,6]. We identified 5 out of 6 American College of Rheumatology clinical criteria present in our patient.
This clinical case shows gradual development of EGPA with multiple-organ involvement (see Figure 3). The disease onset was characterized by respiratory manifestations and suspicion of EGPA appeared only after the development of polyneuropathy (after 39 years) with hypereosinophilia. Polyneuropathy was noted at the age of 57, which is slightly later than the average age of onset reported in the literature [13]. Notably, the patient was ANCA-positive and his long-term neurological manifestation (mononeuritis) was quite typical for ANCA vasculitis [5,6,8,13]. The diagnosis became obvious after comprehensive assessment of the disease history with its gradually evolving signs and symptoms and laboratory abnormalities. Almost all manifestations of the disease seen in the patient were typical except for stroke, which is an exceptional feature for this case [1,5,6]. Neurological involvement manifested as ischemic stroke is an uncommon complication of EGPA (seen only in 5.2% of all patients) [1]. Thus, the patient demonstrated both peripheral and central nervous system involvement, which is unusual and life-threatening in EGPA.
This clinical case of EGPA also attracted our attention due to severe respiratory manifestations. While typical respiratory manifestations such as rhinosinusitis, asthma, and transient lung infiltrates are well documented in the literature, EGPA-associated bronchiectasis is rarely mentioned [14]. Bronchiectasis in the patient was detected by CT at the age of 63. The patient was a nonsmoker, and no other cause of bronchial abnormalities and airflow obstruction could be identified. Stenosis of the proximal bronchi was ruled out by fibrobronchoscopy and CT. The diagnosis of allergic bronchopulmonary aspergillosis was not confirmed. Klebsiella pneumoniae colonization was found during the last 2 years of the patient’s life. Klebsiella pneumoniae is a typical pathogen causing bronchiectasis [15]. It should be noted that aspergillosis and mycobacteriosis are common complications of bronchiectasis [15], but neither was identified in our patient.
The patient was treated for a long time with a combination of prednisolone and methotrexate. There was a significant positive effect on polyneuropathy, but uncontrolled bronchial asthma persisted. This treatment regimen complies with current recommendations [3,11]. Patients with life-threatening and/or organ-threatening manifestations (including lesions of the central nervous system and severe peripheral neuropathy) require therapy with a combination of a glucocorticoid and a cytostatic [3,11].
The use of immunosuppressive drugs in patients with ANCA-associated vasculitis significantly increases the risk of infection and mortality [16–18]. Prednisone and methotrexate use were found to be significant predictors of hospitalization for pneumonia. Frequent infections (more than 1 episode) are reported in patients with ANCA-associated vasculitis during the first year of diagnosis and initiation of therapy [18]. We assume that bronchiectasis complicated by bacterial colonization and bronchial obstruction in combination with immuno-suppression led to severe pneumonia and death of the patient.
Conclusions
The present case report demonstrates a long-term follow-up of a patient with EGPA. Both common and rare manifestations of EGPA have been described in the patient, such as central nervous system damage and bronchiectasis. Immunosuppressive treatment probably contributed to Klebsiella infection development. Thus, long-term complications of EGPA in combination with adverse effects of medications could have caused the severe pneumonia and death of the patient.
Figure 1. Brain multi-spiral computed tomography performed in 2016 showing ischemic brain damage in the right occipital lobe.
Figure 2. (A–C) Chest computed tomography of patient showing large left upper lobe pneumonia and bronchiectasis in lover lobes.
Figure 3. Timeline from first symptoms to death. | Fatal | ReactionOutcome | CC BY-NC-ND | 33582702 | 18,962,204 | 2021-02-14 |
What was the outcome of reaction 'Pneumonia'? | Case Report: A 64-Year-Old Man with 10-Year History of Eosinophilic Granulomatosis with Polyangiitis with Bronchiectasis and Severe Klebsiella pneumonia.
BACKGROUND Eosinophilic granulomatosis with polyangiitis (EGPA) is a rare and severe progressive disease with multiple clinical manifestations and organ damage. Usually, it requires long-term monitoring of the state of many organs due to the gradual character of its manifestations. CASE REPORT We report a case of a long-term follow-up of a patient with eosinophilic granulomatosis with polyangiitis with emphasis on specific clinical features in this patient. A 64-year-old man was being followed up for 10 years. The initial diagnosis was allergic bronchial asthma; however, as new clinical signs and symptoms developed, the diagnosis of EGPA became obvious. A positive treatment response was seen, mainly manifested as reduced polyneuropathy. Meanwhile, bronchial asthma remained uncontrolled and bronchiectasis and Klebsiella pneumoniae colonization developed despite the combination treatment with prednisolone and methotrexate. Furthermore, the patient suffered a cerebral ischemic infarction. During the last hospital admission, severe uncontrolled bronchial asthma complicated with pneumonia resulted in the patient's death. CONCLUSIONS This clinical case shows the gradual development of EGPA with multiple-organ involvement, including respiratory manifestations and peripheral and central nervous system damage. Immunosuppressive treatment combined with complications of EGPA could have contributed to severe pneumonia development and death of the patient.
Background
Systemic vasculitides are difficult to diagnose due to the heterogenous clinical manifestations requiring a wide differential diagnostic search. One of the most uncommon vasculitis types is eosinophilic granulomatosis with polyangiitis (EGPA), or Churg-Strauss syndrome [1,2]. This vasculitis is associated with the presence of antineutrophilic cytoplasmic antibodies (ANCA) and is characterized by necrotizing inflammatory lesions of mainly small vessels, as well as eosinophilic and granulomatous inflammation in various organs. Bronchial asthma and eosinophilia are the most common clinical manifestations of EGPA [1–3]. In 2012, according to the current classification of systemic vasculitides, Churg-Strauss syndrome was renamed “Eosinophilic Granulomatosis with Polyangiitis” (EGPA) and classified into the ANCA-associated vasculitides group [3].
EGPA is a rare ANCA-associated vasculitis. The incidence rate is 0.5–6.8 new cases/year per million populations in Europe [2]. Prevalence is higher in northern latitudes compared to southern Europe [4]. The disease occurs most commonly in people 40–60 years of age [2,5]. No sex predominance has been clearly demonstrated [4].
The pathogenesis of the disease is still not completely clear, but the key role is attributed to autoimmune inflammation with the participation of eosinophils, CD8+, and CD4+lymphocytes (predominantly Th-2) and their proinflammatory cytokines: interleukin (IL)4, IL5, and IL13 [2,5]. Elevation of immunoglobulin (Ig)E, IgG4, and eotaxin-3 is common in EGPA patients [2,5]. The frequent (in 30–40% of cases) detection of ANCA (mainly antibodies to myeloperoxidase) also suggests the autoimmune nature of the disease. ANCA-positive patients are more commonly diagnosed with glomerulonephritis and mononeuritis, whereas the ANCA-negative ones more frequently demonstrate endocardial, myocardial, and pulmonary eosinophilic infiltrates [1,2,5,6].
There are 3 phases in the development of EGPA (these phases partially overlap and may not appear in sequence):
the allergic phase, distinguished by allergic upper respiratory tract inflammation (rhinosinusitis, allergic rhinitis) and bronchial asthma;
the eosinophilic phase, manifested with severe eosinophilia in combination with multiple eosinophilic infiltrates in various organs (most often in the lungs and gastrointestinal tract);
the vasculitis phase, with small- and medium-vessel inflammation of different organs, which can often be life-threatening [5,6].
Respiratory manifestations of EGPA include allergic rhinitis, sinusitis, nasal polyposis and, in more than 95% of patients, poorly controlled bronchial asthma [5]. Chest radiography findings include transient and migratory opacities, ground-glass opacity, focal consolidation, and small peribronchial and centrilobular nodules or masses [7]. Computed tomography reveals bronchial abnormalities, most commonly thickened wall and dilatation with secondary bronchiectasis formation, leading to frequent infections [7,8]. Pleural effusions and alveolar hemorrhages are uncommon manifestations of EGPA [1,5,7,8].
A characteristic neurological manifestation seen in 75% of patients with EGPA is peripheral neuropathy (mononeuritis multiplex) [1,5]. Upper-limb symptoms due to neuropathy, as well as vasculitis in epineural vessels, were more frequently reported in the ANCA-positive group [9]. Eosinophil-associated tissue damage and vascular occlusion leading to ischemia are more specific in ANCA-negative patients [9]. Also, the tissue damage caused by eosinophils may be associated with IL-5 concentration [10]. Brain hemorrhage, subarachnoid hemorrhage, and ischemic brain infarction are rare [1,5,6].
The main drugs for the treatment of EGPA are glucocorticoids. In organ- or life-threatening manifestations, the recommended starting dose of prednisone is 1 mg/kg/day with a maximal daily dose of 80 mg for 2–3 weeks, with tapering to a minimal supporting dose selected individually to control the symptoms and minimize the adverse effects of treatment. Methylprednisolone (7.5–15 mg/kg/day) can be used as a pulse therapy for life-threatening manifestations [11,12]. Most patients need prolonged or lifelong steroids. In the case of organ- or life-threatening manifestations, glucocorticoids in combination with cyclophosphamide are prescribed [11,12]. The combination of glucocorticoids and methotrexate or mycophenolate is required to induce remission in mild or moderate disease [11,12]. Mepolizumab (a humanized monoclonal antibody targeting IL-5) has been used for EGPA treatment [11]. For ANCA-positive patients with renal involvement or refractory disease, rituximab can be considered [3, 11].
Due to the rare occurrence of EGPA and the variety of its clinical manifestations, the diagnosis and treatment of this disease can be challenging. Currently, reports of long-term observations with the detailed patient follow-up are of great interest. The aim of our case report is to present rare manifestations of EGPA, such as bronchiectasis and stroke. We also focused on the infectious complications of immunosuppressive treatments. Thus, we would like to present a 10-year observation of a patient with EGPA.
Case Report
The male patient, born in 1953, was first hospitalized in the Pulmonary Department of our clinic in 2007 due to exacerbated asthma. The patient’s history was remarkable for multiple allergies (to house dust, cat and dog epidermis, and plant pollen) as well as nonsteroidal anti-inflammatory drugs intolerance (manifested as bronchospasm). The patient denied unhealthy habits and occupational hazards (he worked as a driver). His family history was not remarkable. The patient has had allergic rhinitis since adolescence. The first asthma symptoms appeared at the age of 18 while serving in the army. At the age of 22, an allergist diagnosed him with bronchial asthma. The patient had annual exacerbations (1–2 times a year) due to cold or allergen exposure, and acute respiratory viral infections. At the age of 42, polypoid rhinosinusitis was found, and a polypectomy was performed. Since 2006, the patient had been on controller treatment with salmeterol/fluticasone inhaler (25/250 mg, respectively) 2 doses 2 times a day.
In late 2009, extremely elevated eosinophils (53%) and total IgE (1000 IU/L) were found for the first time. Also, since 2009, the patient has had transient lung opacities, which usually disappeared after treatment with prednisolone 30 mg per day.
In August 2010, the patient noticed numbness on the back surface of the left foot, which spread to the entire left lower leg in 1 month, as well as numbness of the right foot and lower leg. Later, the patient started having tingling, twisting, and burning sensations aggravated by touching in the feet and lower legs. Also, the weakness in the distal parts of the legs developed, and he started having difficulties moving around on his own and falling while walking, and he was subsequently hospitalized in our clinic due to exacerbated asthma. On laboratory testing, high eosinophils (53%) and gamma-globulins (27.9%) were found in November 2010. The chest radiography showed emphysema and consolidation in the middle lobe of the right lung. A Neurology consult confirmed asymmetrical distal sensory-motor polyneuropathy demonstrated by lower distal asymmetrical paraparesis with decreased muscle strength in the foot flexors and extensors, more severe on the left side, low tone of the shin muscles, hypoactive Achilles reflexes, and reduced leg sensitivity starting from the middle of the shin and below, more severe in the peroneal nerve inner-vation area. The patient also demonstrated abnormal coordination (heel-to-knee, Romberg) test results and bilateral steppage gait with knee overextension. Electroneuromyography showed axonal lesion of all distal leg nerves and secondary myelinopathy. Thus, EGPA was suspected. The ANCA (anti-myeloperoxidase) titer was 30 U/ml (normal reference below 20 U/ml).
Allergic bronchopulmonary aspergillosis and other causes of hypereosinophilia were consistently ruled out. IgE and IgG for Aspergillus spp. and Aspergillus spp. in sputum were not found.
Thus, the presence of severe bronchial asthma, rhinosinusitis, high and persistent eosinophilia, transient pulmonary opacities, and polyneuropathy suggested EGPA (5 out of 6 clinical criteria of American College of Rheumatology present). Based on the clinical presentation and increased ANCA titer, the patient was diagnosed with EGPA. Systemic steroid treatment (prednisolone 300 mg intravenously and 30 mg orally) was prescribed, and 5 plasmapheresis sessions were performed. Later, prolonged combination immunosuppression (prednisolone 10 mg/day and methotrexate 25 mg/week) and controller asthma treatment (fluticasone propionate/salmeterol 250/25 µg 4 doses/ day, tiotropium bromide 18 µg per day) were prescribed. After 1 month on prednisolone and methotrexate, there was a significant neurological improvement: the motor and sensory disturbances decreased, and the patient was able to move around on his own. Further on, polyneuropathy completely resolved on the combination treatment. His asthma remained uncontrolled despite the combination treatment: he had frequent (3 times a year) severe exacerbations requiring hospitalization. In 2015, newly-emerged copious purulent sputum associated with frequent infection-related asthma exacerbations was noted. In January 2016, a patient underwent chest computed tomography (CT), which showed generalized bronchiectasis. Stenosis of the proximal bronchi was ruled out also by fibrobronchos-copy. The bronchial secretion microbiology revealed Klebsiella pneumoniae growth (107 colony-forming units).
In November 2016, during the asthma exacerbation, the patient suffered a cerebral ischemic infarction involving the right occipital lobe, confirmed by CT (Figures 1, 2).
On March 2, 2017, the patient was admitted to our hospital due to severe respiratory failure caused by pneumonia in the left upper lobe and severe asthma exacerbation.
The complete blood count showed high WBCs (15.2×109/l) with left shift. The IgE level was 144.0 IU/l, C-reactive protein was 48.8 mg/dL. Clinical chemistry was remarkable for urea nitrogen 18.4 mmol/l and creatinine 1.55 mg/dL. The sputum microscopy demonstrated 100 WBCs and 160 RBCs per vision field, and no eosinophils were revealed. A chest CT revealed multiple bronchiectasis and upper lobar pneumonia of the left lung (Figure 2). Klebsiella was again detected in the sputum culture.
The patient was prescribed intravenous antibiotics (ceftriaxone 2 g/day and ciprofloxacin 800 mg/day replaced by cefopera-zone 4 g/day and amikacin 1 g/day after 3 days due to low efficacy), bronchodilator and anti-inflammatory therapy (nebulized ipratropium bromide, fenoterol, and budesonide 2000 µg per day, prednisolone intravenously 150 mg and orally 30 mg). Despite the aggressive treatment, the respiratory failure progressed, and on day 6 of hospitalization the patient was transferred to the Intensive Care Unit and intubated. On day 7, the patient died. On autopsy, the EGPA diagnosis was confirmed, signs of ischemic infarction in the occipital lobe of the brain were found, and left upper lobe pneumonia with abscess formation and generalized bilateral bronchiectases were revealed.
Discussion
Based on the long follow-up period and careful interpretation of signs and symptoms, typical laboratory abnormalities, the diagnosis was clear. Among the most commonly used diagnostic criteria of EGPA are the American College of Rheumatology (ACR) classification criteria, Lanham’s criteria, and the Chapel Hill Consensus Conference criteria. The presence of 4 or more ACR classification criteria, including bronchial asthma, eosinophilia (more than 10%), paranasal sinus abnormalities, migratory or transient eosinophilic pulmonary infiltrates, extravascular eosinophil infiltration on biopsy, and mono- or polyneuropathy, has a sensitivity of 85% and a specificity of 100% [2,5,6]. We identified 5 out of 6 American College of Rheumatology clinical criteria present in our patient.
This clinical case shows gradual development of EGPA with multiple-organ involvement (see Figure 3). The disease onset was characterized by respiratory manifestations and suspicion of EGPA appeared only after the development of polyneuropathy (after 39 years) with hypereosinophilia. Polyneuropathy was noted at the age of 57, which is slightly later than the average age of onset reported in the literature [13]. Notably, the patient was ANCA-positive and his long-term neurological manifestation (mononeuritis) was quite typical for ANCA vasculitis [5,6,8,13]. The diagnosis became obvious after comprehensive assessment of the disease history with its gradually evolving signs and symptoms and laboratory abnormalities. Almost all manifestations of the disease seen in the patient were typical except for stroke, which is an exceptional feature for this case [1,5,6]. Neurological involvement manifested as ischemic stroke is an uncommon complication of EGPA (seen only in 5.2% of all patients) [1]. Thus, the patient demonstrated both peripheral and central nervous system involvement, which is unusual and life-threatening in EGPA.
This clinical case of EGPA also attracted our attention due to severe respiratory manifestations. While typical respiratory manifestations such as rhinosinusitis, asthma, and transient lung infiltrates are well documented in the literature, EGPA-associated bronchiectasis is rarely mentioned [14]. Bronchiectasis in the patient was detected by CT at the age of 63. The patient was a nonsmoker, and no other cause of bronchial abnormalities and airflow obstruction could be identified. Stenosis of the proximal bronchi was ruled out by fibrobronchoscopy and CT. The diagnosis of allergic bronchopulmonary aspergillosis was not confirmed. Klebsiella pneumoniae colonization was found during the last 2 years of the patient’s life. Klebsiella pneumoniae is a typical pathogen causing bronchiectasis [15]. It should be noted that aspergillosis and mycobacteriosis are common complications of bronchiectasis [15], but neither was identified in our patient.
The patient was treated for a long time with a combination of prednisolone and methotrexate. There was a significant positive effect on polyneuropathy, but uncontrolled bronchial asthma persisted. This treatment regimen complies with current recommendations [3,11]. Patients with life-threatening and/or organ-threatening manifestations (including lesions of the central nervous system and severe peripheral neuropathy) require therapy with a combination of a glucocorticoid and a cytostatic [3,11].
The use of immunosuppressive drugs in patients with ANCA-associated vasculitis significantly increases the risk of infection and mortality [16–18]. Prednisone and methotrexate use were found to be significant predictors of hospitalization for pneumonia. Frequent infections (more than 1 episode) are reported in patients with ANCA-associated vasculitis during the first year of diagnosis and initiation of therapy [18]. We assume that bronchiectasis complicated by bacterial colonization and bronchial obstruction in combination with immuno-suppression led to severe pneumonia and death of the patient.
Conclusions
The present case report demonstrates a long-term follow-up of a patient with EGPA. Both common and rare manifestations of EGPA have been described in the patient, such as central nervous system damage and bronchiectasis. Immunosuppressive treatment probably contributed to Klebsiella infection development. Thus, long-term complications of EGPA in combination with adverse effects of medications could have caused the severe pneumonia and death of the patient.
Figure 1. Brain multi-spiral computed tomography performed in 2016 showing ischemic brain damage in the right occipital lobe.
Figure 2. (A–C) Chest computed tomography of patient showing large left upper lobe pneumonia and bronchiectasis in lover lobes.
Figure 3. Timeline from first symptoms to death. | Fatal | ReactionOutcome | CC BY-NC-ND | 33582702 | 18,962,812 | 2021-02-14 |
What was the outcome of reaction 'Respiratory failure'? | Case Report: A 64-Year-Old Man with 10-Year History of Eosinophilic Granulomatosis with Polyangiitis with Bronchiectasis and Severe Klebsiella pneumonia.
BACKGROUND Eosinophilic granulomatosis with polyangiitis (EGPA) is a rare and severe progressive disease with multiple clinical manifestations and organ damage. Usually, it requires long-term monitoring of the state of many organs due to the gradual character of its manifestations. CASE REPORT We report a case of a long-term follow-up of a patient with eosinophilic granulomatosis with polyangiitis with emphasis on specific clinical features in this patient. A 64-year-old man was being followed up for 10 years. The initial diagnosis was allergic bronchial asthma; however, as new clinical signs and symptoms developed, the diagnosis of EGPA became obvious. A positive treatment response was seen, mainly manifested as reduced polyneuropathy. Meanwhile, bronchial asthma remained uncontrolled and bronchiectasis and Klebsiella pneumoniae colonization developed despite the combination treatment with prednisolone and methotrexate. Furthermore, the patient suffered a cerebral ischemic infarction. During the last hospital admission, severe uncontrolled bronchial asthma complicated with pneumonia resulted in the patient's death. CONCLUSIONS This clinical case shows the gradual development of EGPA with multiple-organ involvement, including respiratory manifestations and peripheral and central nervous system damage. Immunosuppressive treatment combined with complications of EGPA could have contributed to severe pneumonia development and death of the patient.
Background
Systemic vasculitides are difficult to diagnose due to the heterogenous clinical manifestations requiring a wide differential diagnostic search. One of the most uncommon vasculitis types is eosinophilic granulomatosis with polyangiitis (EGPA), or Churg-Strauss syndrome [1,2]. This vasculitis is associated with the presence of antineutrophilic cytoplasmic antibodies (ANCA) and is characterized by necrotizing inflammatory lesions of mainly small vessels, as well as eosinophilic and granulomatous inflammation in various organs. Bronchial asthma and eosinophilia are the most common clinical manifestations of EGPA [1–3]. In 2012, according to the current classification of systemic vasculitides, Churg-Strauss syndrome was renamed “Eosinophilic Granulomatosis with Polyangiitis” (EGPA) and classified into the ANCA-associated vasculitides group [3].
EGPA is a rare ANCA-associated vasculitis. The incidence rate is 0.5–6.8 new cases/year per million populations in Europe [2]. Prevalence is higher in northern latitudes compared to southern Europe [4]. The disease occurs most commonly in people 40–60 years of age [2,5]. No sex predominance has been clearly demonstrated [4].
The pathogenesis of the disease is still not completely clear, but the key role is attributed to autoimmune inflammation with the participation of eosinophils, CD8+, and CD4+lymphocytes (predominantly Th-2) and their proinflammatory cytokines: interleukin (IL)4, IL5, and IL13 [2,5]. Elevation of immunoglobulin (Ig)E, IgG4, and eotaxin-3 is common in EGPA patients [2,5]. The frequent (in 30–40% of cases) detection of ANCA (mainly antibodies to myeloperoxidase) also suggests the autoimmune nature of the disease. ANCA-positive patients are more commonly diagnosed with glomerulonephritis and mononeuritis, whereas the ANCA-negative ones more frequently demonstrate endocardial, myocardial, and pulmonary eosinophilic infiltrates [1,2,5,6].
There are 3 phases in the development of EGPA (these phases partially overlap and may not appear in sequence):
the allergic phase, distinguished by allergic upper respiratory tract inflammation (rhinosinusitis, allergic rhinitis) and bronchial asthma;
the eosinophilic phase, manifested with severe eosinophilia in combination with multiple eosinophilic infiltrates in various organs (most often in the lungs and gastrointestinal tract);
the vasculitis phase, with small- and medium-vessel inflammation of different organs, which can often be life-threatening [5,6].
Respiratory manifestations of EGPA include allergic rhinitis, sinusitis, nasal polyposis and, in more than 95% of patients, poorly controlled bronchial asthma [5]. Chest radiography findings include transient and migratory opacities, ground-glass opacity, focal consolidation, and small peribronchial and centrilobular nodules or masses [7]. Computed tomography reveals bronchial abnormalities, most commonly thickened wall and dilatation with secondary bronchiectasis formation, leading to frequent infections [7,8]. Pleural effusions and alveolar hemorrhages are uncommon manifestations of EGPA [1,5,7,8].
A characteristic neurological manifestation seen in 75% of patients with EGPA is peripheral neuropathy (mononeuritis multiplex) [1,5]. Upper-limb symptoms due to neuropathy, as well as vasculitis in epineural vessels, were more frequently reported in the ANCA-positive group [9]. Eosinophil-associated tissue damage and vascular occlusion leading to ischemia are more specific in ANCA-negative patients [9]. Also, the tissue damage caused by eosinophils may be associated with IL-5 concentration [10]. Brain hemorrhage, subarachnoid hemorrhage, and ischemic brain infarction are rare [1,5,6].
The main drugs for the treatment of EGPA are glucocorticoids. In organ- or life-threatening manifestations, the recommended starting dose of prednisone is 1 mg/kg/day with a maximal daily dose of 80 mg for 2–3 weeks, with tapering to a minimal supporting dose selected individually to control the symptoms and minimize the adverse effects of treatment. Methylprednisolone (7.5–15 mg/kg/day) can be used as a pulse therapy for life-threatening manifestations [11,12]. Most patients need prolonged or lifelong steroids. In the case of organ- or life-threatening manifestations, glucocorticoids in combination with cyclophosphamide are prescribed [11,12]. The combination of glucocorticoids and methotrexate or mycophenolate is required to induce remission in mild or moderate disease [11,12]. Mepolizumab (a humanized monoclonal antibody targeting IL-5) has been used for EGPA treatment [11]. For ANCA-positive patients with renal involvement or refractory disease, rituximab can be considered [3, 11].
Due to the rare occurrence of EGPA and the variety of its clinical manifestations, the diagnosis and treatment of this disease can be challenging. Currently, reports of long-term observations with the detailed patient follow-up are of great interest. The aim of our case report is to present rare manifestations of EGPA, such as bronchiectasis and stroke. We also focused on the infectious complications of immunosuppressive treatments. Thus, we would like to present a 10-year observation of a patient with EGPA.
Case Report
The male patient, born in 1953, was first hospitalized in the Pulmonary Department of our clinic in 2007 due to exacerbated asthma. The patient’s history was remarkable for multiple allergies (to house dust, cat and dog epidermis, and plant pollen) as well as nonsteroidal anti-inflammatory drugs intolerance (manifested as bronchospasm). The patient denied unhealthy habits and occupational hazards (he worked as a driver). His family history was not remarkable. The patient has had allergic rhinitis since adolescence. The first asthma symptoms appeared at the age of 18 while serving in the army. At the age of 22, an allergist diagnosed him with bronchial asthma. The patient had annual exacerbations (1–2 times a year) due to cold or allergen exposure, and acute respiratory viral infections. At the age of 42, polypoid rhinosinusitis was found, and a polypectomy was performed. Since 2006, the patient had been on controller treatment with salmeterol/fluticasone inhaler (25/250 mg, respectively) 2 doses 2 times a day.
In late 2009, extremely elevated eosinophils (53%) and total IgE (1000 IU/L) were found for the first time. Also, since 2009, the patient has had transient lung opacities, which usually disappeared after treatment with prednisolone 30 mg per day.
In August 2010, the patient noticed numbness on the back surface of the left foot, which spread to the entire left lower leg in 1 month, as well as numbness of the right foot and lower leg. Later, the patient started having tingling, twisting, and burning sensations aggravated by touching in the feet and lower legs. Also, the weakness in the distal parts of the legs developed, and he started having difficulties moving around on his own and falling while walking, and he was subsequently hospitalized in our clinic due to exacerbated asthma. On laboratory testing, high eosinophils (53%) and gamma-globulins (27.9%) were found in November 2010. The chest radiography showed emphysema and consolidation in the middle lobe of the right lung. A Neurology consult confirmed asymmetrical distal sensory-motor polyneuropathy demonstrated by lower distal asymmetrical paraparesis with decreased muscle strength in the foot flexors and extensors, more severe on the left side, low tone of the shin muscles, hypoactive Achilles reflexes, and reduced leg sensitivity starting from the middle of the shin and below, more severe in the peroneal nerve inner-vation area. The patient also demonstrated abnormal coordination (heel-to-knee, Romberg) test results and bilateral steppage gait with knee overextension. Electroneuromyography showed axonal lesion of all distal leg nerves and secondary myelinopathy. Thus, EGPA was suspected. The ANCA (anti-myeloperoxidase) titer was 30 U/ml (normal reference below 20 U/ml).
Allergic bronchopulmonary aspergillosis and other causes of hypereosinophilia were consistently ruled out. IgE and IgG for Aspergillus spp. and Aspergillus spp. in sputum were not found.
Thus, the presence of severe bronchial asthma, rhinosinusitis, high and persistent eosinophilia, transient pulmonary opacities, and polyneuropathy suggested EGPA (5 out of 6 clinical criteria of American College of Rheumatology present). Based on the clinical presentation and increased ANCA titer, the patient was diagnosed with EGPA. Systemic steroid treatment (prednisolone 300 mg intravenously and 30 mg orally) was prescribed, and 5 plasmapheresis sessions were performed. Later, prolonged combination immunosuppression (prednisolone 10 mg/day and methotrexate 25 mg/week) and controller asthma treatment (fluticasone propionate/salmeterol 250/25 µg 4 doses/ day, tiotropium bromide 18 µg per day) were prescribed. After 1 month on prednisolone and methotrexate, there was a significant neurological improvement: the motor and sensory disturbances decreased, and the patient was able to move around on his own. Further on, polyneuropathy completely resolved on the combination treatment. His asthma remained uncontrolled despite the combination treatment: he had frequent (3 times a year) severe exacerbations requiring hospitalization. In 2015, newly-emerged copious purulent sputum associated with frequent infection-related asthma exacerbations was noted. In January 2016, a patient underwent chest computed tomography (CT), which showed generalized bronchiectasis. Stenosis of the proximal bronchi was ruled out also by fibrobronchos-copy. The bronchial secretion microbiology revealed Klebsiella pneumoniae growth (107 colony-forming units).
In November 2016, during the asthma exacerbation, the patient suffered a cerebral ischemic infarction involving the right occipital lobe, confirmed by CT (Figures 1, 2).
On March 2, 2017, the patient was admitted to our hospital due to severe respiratory failure caused by pneumonia in the left upper lobe and severe asthma exacerbation.
The complete blood count showed high WBCs (15.2×109/l) with left shift. The IgE level was 144.0 IU/l, C-reactive protein was 48.8 mg/dL. Clinical chemistry was remarkable for urea nitrogen 18.4 mmol/l and creatinine 1.55 mg/dL. The sputum microscopy demonstrated 100 WBCs and 160 RBCs per vision field, and no eosinophils were revealed. A chest CT revealed multiple bronchiectasis and upper lobar pneumonia of the left lung (Figure 2). Klebsiella was again detected in the sputum culture.
The patient was prescribed intravenous antibiotics (ceftriaxone 2 g/day and ciprofloxacin 800 mg/day replaced by cefopera-zone 4 g/day and amikacin 1 g/day after 3 days due to low efficacy), bronchodilator and anti-inflammatory therapy (nebulized ipratropium bromide, fenoterol, and budesonide 2000 µg per day, prednisolone intravenously 150 mg and orally 30 mg). Despite the aggressive treatment, the respiratory failure progressed, and on day 6 of hospitalization the patient was transferred to the Intensive Care Unit and intubated. On day 7, the patient died. On autopsy, the EGPA diagnosis was confirmed, signs of ischemic infarction in the occipital lobe of the brain were found, and left upper lobe pneumonia with abscess formation and generalized bilateral bronchiectases were revealed.
Discussion
Based on the long follow-up period and careful interpretation of signs and symptoms, typical laboratory abnormalities, the diagnosis was clear. Among the most commonly used diagnostic criteria of EGPA are the American College of Rheumatology (ACR) classification criteria, Lanham’s criteria, and the Chapel Hill Consensus Conference criteria. The presence of 4 or more ACR classification criteria, including bronchial asthma, eosinophilia (more than 10%), paranasal sinus abnormalities, migratory or transient eosinophilic pulmonary infiltrates, extravascular eosinophil infiltration on biopsy, and mono- or polyneuropathy, has a sensitivity of 85% and a specificity of 100% [2,5,6]. We identified 5 out of 6 American College of Rheumatology clinical criteria present in our patient.
This clinical case shows gradual development of EGPA with multiple-organ involvement (see Figure 3). The disease onset was characterized by respiratory manifestations and suspicion of EGPA appeared only after the development of polyneuropathy (after 39 years) with hypereosinophilia. Polyneuropathy was noted at the age of 57, which is slightly later than the average age of onset reported in the literature [13]. Notably, the patient was ANCA-positive and his long-term neurological manifestation (mononeuritis) was quite typical for ANCA vasculitis [5,6,8,13]. The diagnosis became obvious after comprehensive assessment of the disease history with its gradually evolving signs and symptoms and laboratory abnormalities. Almost all manifestations of the disease seen in the patient were typical except for stroke, which is an exceptional feature for this case [1,5,6]. Neurological involvement manifested as ischemic stroke is an uncommon complication of EGPA (seen only in 5.2% of all patients) [1]. Thus, the patient demonstrated both peripheral and central nervous system involvement, which is unusual and life-threatening in EGPA.
This clinical case of EGPA also attracted our attention due to severe respiratory manifestations. While typical respiratory manifestations such as rhinosinusitis, asthma, and transient lung infiltrates are well documented in the literature, EGPA-associated bronchiectasis is rarely mentioned [14]. Bronchiectasis in the patient was detected by CT at the age of 63. The patient was a nonsmoker, and no other cause of bronchial abnormalities and airflow obstruction could be identified. Stenosis of the proximal bronchi was ruled out by fibrobronchoscopy and CT. The diagnosis of allergic bronchopulmonary aspergillosis was not confirmed. Klebsiella pneumoniae colonization was found during the last 2 years of the patient’s life. Klebsiella pneumoniae is a typical pathogen causing bronchiectasis [15]. It should be noted that aspergillosis and mycobacteriosis are common complications of bronchiectasis [15], but neither was identified in our patient.
The patient was treated for a long time with a combination of prednisolone and methotrexate. There was a significant positive effect on polyneuropathy, but uncontrolled bronchial asthma persisted. This treatment regimen complies with current recommendations [3,11]. Patients with life-threatening and/or organ-threatening manifestations (including lesions of the central nervous system and severe peripheral neuropathy) require therapy with a combination of a glucocorticoid and a cytostatic [3,11].
The use of immunosuppressive drugs in patients with ANCA-associated vasculitis significantly increases the risk of infection and mortality [16–18]. Prednisone and methotrexate use were found to be significant predictors of hospitalization for pneumonia. Frequent infections (more than 1 episode) are reported in patients with ANCA-associated vasculitis during the first year of diagnosis and initiation of therapy [18]. We assume that bronchiectasis complicated by bacterial colonization and bronchial obstruction in combination with immuno-suppression led to severe pneumonia and death of the patient.
Conclusions
The present case report demonstrates a long-term follow-up of a patient with EGPA. Both common and rare manifestations of EGPA have been described in the patient, such as central nervous system damage and bronchiectasis. Immunosuppressive treatment probably contributed to Klebsiella infection development. Thus, long-term complications of EGPA in combination with adverse effects of medications could have caused the severe pneumonia and death of the patient.
Figure 1. Brain multi-spiral computed tomography performed in 2016 showing ischemic brain damage in the right occipital lobe.
Figure 2. (A–C) Chest computed tomography of patient showing large left upper lobe pneumonia and bronchiectasis in lover lobes.
Figure 3. Timeline from first symptoms to death. | Fatal | ReactionOutcome | CC BY-NC-ND | 33582702 | 18,986,917 | 2021-02-14 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Asthenia'. | Practical Issues in the Use of Atezolizumab for Patients with Non-Small Cell Lung Cancer: Case Reports and Literature Review.
Atezolizumab is a monoclonal antibody targeting the programmed death ligand 1 (PD-L1) that was approved in 2017 in the USA and Europe for the second-line treatment of advanced or metastatic non-small cell lung cancer (NSCLC). This review article describes the practical clinical issues associated with atezolizumab treatment in NSCLC using a combination of four illustrative cases and a narrative literature review. The first two cases highlight the importance of tumor mutational status when making treatment decisions. A 62-year-old man with epidermal growth factor receptor (EGFR)-mutated, PD-L1-positive, stage IV lung adenocarcinoma received treatment with second-line atezolizumab + bevacizumab, carboplatin, and paclitaxel (BCP) after first-line osimertinib. In the second case, a 63-year-old man with stage IVb lung adenocarcinoma with anaplastic lymphoma kinase (ALK) translocation received sixth-line treatment with atezolizumab + BCP. The two final cases both had extensive metastases. A 55-year-old woman with EGFR-mutated lung adenocarcinoma received second-line treatment with atezolizumab + BCP after development of multiple metastases, followed by atezolizumab + bevacizumab until last follow-up. A 42-year-old man with PD-L1-positive pulmonary adenocarcinoma (negative for EGFR mutations) developed liver and brain metastases after several lines of therapy. He underwent holocranial radiation and received atezolizumab + BCP, which resulted in a decrease in all measurable and evaluable tumoral lesions. These illustrative cases indicate that the type and number of mutations may influence treatment response to atezolizumab, and that atezolizumab may provide clinical benefit in patients with high disease burden.
Key Summary Points
Why carry out this study?
Targeted immunotherapies, such as atezolizumab, have improved overall survival in patients with advanced or metastatic non-small cell lung cancer (NSCLC) in clinical trials.
Understanding how to treat patients on the basis of their targetable oncogenic mutations or those usually excluded from clinical trials (e.g., with treated brain metastases or high disease burden) is important for oncologists in the real-world clinical setting.
Four cases of treatment with atezolizumab plus bevacizumab, carboplatin, and paclitaxel in patients with NSCLC after at least one previous line of targeted therapy are described to illustrate these clinical issues.
What was learned from the study?
The presented cases suggest that both the number and type of mutations may be relevant for guiding treatment decisions in NSCLC, and that patients with high disease burden or brain metastases may benefit from atezolizumab-containing therapy.
Digital Features
This article is published with digital features, including a summary slide, to facilitate understanding of the article. To view digital features for this article go to https://doi.org/10.6084/m9.figshare.13521968.
Introduction
Non-small cell lung cancer (NSCLC) is the most common type of lung cancer and has a high rate of mortality [1, 2]. According to global estimates, lung cancer was the most commonly diagnosed cancer in 2018 (11.6% of all new cases) and the leading cause of cancer-related mortality (18.4% of all cancer deaths) [3]. In a US analysis of patients with NSCLC, the median overall survival (OS) time after diagnosis was only 13 months, and the estimated 4-year survival rate was between 22% and 28% [1]. However, OS has been improving over recent decades as a result of novel treatments [1, 2], including targeted therapies and immunotherapies.
One such novel treatment is atezolizumab, a monoclonal antibody targeting the programmed death ligand 1 (PD-L1) [4]. Atezolizumab was approved in the USA and Europe for the second-line treatment of advanced or metastatic NSCLC in 2017, based on the results of the phase II POPLAR study [5] and the phase III OAK study [6], in which atezolizumab was associated with significantly better survival compared with docetaxel. Since its approval, the IMpower150 study has demonstrated that atezolizumab in combination with bevacizumab, carboplatin, and paclitaxel (BCP) was more effective than BCP alone as first-line therapy in previously untreated patients with metastatic non-squamous NSCLC [7].
The current review describes the practical clinical issues associated with atezolizumab + BCP in patients with NSCLC, using a combination of illustrative case presentations and a narrative literature review.
Cases Illustrating Role of Mutational Status
Patient 1
A 62-year-old man with no relevant medical or surgical history was diagnosed with stage IIIC lung adenocarcinoma (cT3cN3cM0) in January 2018. He was a former smoker with a 15 pack-year history. Molecular testing showed an L858R point mutation in epidermal growth factor receptor (EGFR) exon 21 and a primary T790M mutation in EGFR exon 20. The tumor was PD-L1 positive, with a tumor proportion score (TPS) of 80% on immunohistochemistry (pharmDx 28–8; DAKO, Glostrup, Denmark).
On February 1, 2018, he began first-line treatment with osimertinib; the 12-week assessment showed a partial response by Response Evaluation Criteria In Solid Tumors (RECIST) 1.1 criteria. Response continued until March 2019, when the disease progressed to stage IV with mediastinal and retroperitoneal involvement (Fig. 1a). At this time, the patient was asymptomatic and started second-line treatment with atezolizumab in combination with BCP, based on the results of the IMpower150 clinical trial [7]. After two cycles, a computed tomography (CT) scan showed partial response by RECIST 1.1 criteria (Fig. 1b), and the patient was able to receive six cycles without significant adverse events.Fig. 1 Patient 1: Computed tomography scan of chest on a March 25, 2019, showing mediastinal and retroperitoneal involvement, b June 6, 2019 after two cycles of atezolizumab + BCP, c October 5, 2019 indicating a maintained radiologic response, and d January 30, 2020 showing tumor progression after 13 weeks of maintenance atezolizumab + bevacizumab. BCP bevacizumab, carboplatin, and paclitaxel
After confirming that the radiologic response was maintained (Fig. 1c), he continued maintenance treatment with atezolizumab + bevacizumab. In February 2020, after 13 cycles, tumor progression was noted (Fig. 1d) and treatment was discontinued. Biopsy showed amplification and overexpression of c-MET, so the patient initiated third-line treatment with telisotuzumab vedotin as part of a clinical trial (NCT03539536).
Patient 2
A 63-year-old man was diagnosed with a stage IVb lung adenocarcinoma with a brain metastasis (T2N3M1b) in February 2013. He was a former smoker (20 pack-year history) with occupational exposure to oil and its derivatives.
On June 13, 2013, he started induction chemotherapy with cisplatin + pemetrexed, undergoing radiosurgery for the brain lesion after the first cycle. He showed a partial radiologic and metabolic response after two cycles of cisplatin + pemetrexed, and a brain magnetic resonance imaging (MRI) revealed a reduction in the size of the brain metastasis.
He received two cycles of cisplatin + pemetrexed from June 13 to June 24, 2013, followed by radical-intent chemoradiation between June 28 and September 6, 2013, consisting of a 60 Gy dose and two cycles of weekly paclitaxel + carboplatin; however, he was unable to continue this treatment because of sustained leukopenia.
A follow-up assessment on October 6, 2014, found no signs of thoracoabdominal progression. Lesions consistent with metastases were identified in the cerebellar vermis and the right centrum semiovale; radiosurgery was administered using 20.7 Gy and 20.9 Gy, respectively, at these sites.
A CT scan on January 12, 2015, showed hilar-mediastinal progression, and a biopsy of the left hilar adenopathy showed that the tumor had wild-type KRAS, was EGFR- and BRAF-mutation negative, and had an anaplastic lymphoma kinase (ALK) translocation. In February 2015, he started second-line treatment with crizotinib and had a partial response. A brain MRI performed on November 3, 2015, showed oligoprogression, for which he received radiosurgery.
On April 4, 2016, he showed pulmonary progression, and 2 weeks later, he started third-line treatment with ceritinib as part of a clinical trial, with a best response of stable disease.
A CT scan on January 30, 2018, showed an increase in the size of the space-occupying lesions in the brain. On March 6 of the same year, he initiated fourth-line treatment with alectinib for metastases, with a best response of stable disease. A further CT scan on July 23, 2018, showed left hilar adenopathy progression and left upper lobe atelectasis. He started fifth-line therapy for metastases with brigatinib on September 5, 2018, and stable disease was his best response.
The patient developed left hemiparesis on January 16, 2019. A brain MRI on January 21 showed signs of radionecrosis, but an acute or subacute ischemic event could not be ruled out. Brain progression, with an increase in the size and number of lesions, was detected in March 2019 (Fig. 2a), and he received hippocampal-sparing whole brain radiation with focal boost (45 Gy dose) on the lesions.Fig. 2 Patient 2: brain magnetic resonance imaging scan results on a March 11, 2019, showing an increase in the size and number of brain lesions, and b March 22, 2020, after 4 cycles of atezolizumab + BCP followed by 13 cycles of maintenance atezolizumab + bevacizumab, and chest computed tomography scan results on c March 11, 2019, prior to, and d March 22, 2020, after atezolizumab-based treatment
On May 8, 2019, he began sixth-line treatment with atezolizumab + BCP. After four cycles (ending on August 8, 2019), he started maintenance treatment with atezolizumab + bevacizumab, achieving a partial brain response. In March 2020, he had received 13 cycles of maintenance treatment, and he showed stable disease as assessed by brain MRI (Fig. 2b) and CT scans of the chest (Fig. 2c, d), abdomen, and pelvis.
CASES Illustrating Management of High Disease Burden
Patient 3
A 55-year-old non-smoking woman without known morbidity was diagnosed with a stage IIIB lung adenocarcinoma (cT4N2) in June 2016. She received treatment with external radiation therapy (dose administered, 66 Gy) concomitantly with cisplatin and vinorelbine (three cycles). A follow-up CT scan showed partial response and the multidisciplinary Committee on Thoracic Tumors recommended surgical intervention. She underwent video-assisted thoracic surgery consisting of an upper right pulmonary lobectomy with systematic hilar and mediastinal lymph node dissection on October 5, 2016. Molecular analysis of the excised tumor showed EGFR exon 19 deletion.
In March 2018, disease progression was detected in lung, pleura, and bone, and in subcutaneous tissue and the lymphatic system. On March 27, she received palliative and decompressive radiation therapy of the lumbar spine (L5) at 8 Gy. Subsequently, in April 2018, she started treatment with afatinib at 40 mg/day, and had a partial response. Afatinib treatment continued until July 2019, when imaging identified disease progression in the liver and bone, and a sacral soft tissue mass (Fig. 3a). Analysis of liquid and sacral mass biopsy did not detect a resistant T790M mutation on EGFR exon 20.Fig. 3 Patient 3: Computed tomography of lumbosacral region on a July 4, 2019, before and b October 8, 2019, after three cycles of atezolizumab + BCP, showing partial response in the sacral soft tissue mass. BCP bevacizumab, carboplatin, and paclitaxel
At this time, the patient’s Eastern Cooperative Oncology Group (ECOG) performance status was 2, and she was negative for PD-L1 (TPS 0%). On August 7, 2019, on the basis of the results of the IMpower150 clinical trial [7], she began second-line treatment with atezolizumab + BCP. After three cycles she showed a partial response (Fig. 3b), and was able to complete six cycles of treatment. However, the patient developed febrile neutropenia and sepsis due to central catheter-related infections in the second cycle leading to hospital admission; grade 2 sensory neuropathy that reverted to grade 1 after discontinuing paclitaxel and carboplatin; and grade 1 asthenia. She then continued treatment with atezolizumab + bevacizumab. By June 2020, she had completed 14 cycles of treatment without relevant toxicities, but treatment was interrupted at that time because a CT scan showed progression of the liver and bone metastases.
Patient 4
A 42-year-old man was diagnosed with stage IIIB pulmonary adenocarcinoma (cT4N2M0) in June 2018; at this time, he had an ECOG performance status of 0. Molecular analysis showed the tumor was negative for EGFR mutations and ALK fusions, with PD-L1 expression on 30% of cancer cells. He was a smoker at the time of diagnosis, smoking one pack daily for 20 years (20 pack-years).
On August 22, 2018, treatment with cisplatin, gemcitabine, and paclitaxel was started, for a total of four 21-day cycles. Treatment was well tolerated, with the exception of nausea and asthenia (both grade 1). Repeat CT scans performed in September 2018 after two treatment cycles, and again in November after four treatment cycles, revealed decreasing tumor volume, with mediastinal contact still present; at this time, results of a brain MRI were normal.
In December 2018, the man underwent a right lower lobectomy; the right lower pulmonary lobe was found to have well-differentiated adenocarcinoma with an acinar pattern, without evidence of vascular invasion, and no visceral pleural infiltration. Surgical resection margins were not affected. TNM staging at this point was ypT1cN0M0. He subsequently received consolidative radiation therapy on the mediastinal bed; he had a performance status of 0 and no complications.
In October 2019, a follow-up examination revealed a de novo pulmonary micronodule in the control CT scan, but this was not seen on a positron emission tomography–CT scan. In February 2020, repeat CT scans revealed a growth on the left perihilar pulmonary nodule and a new right-hand micronodule. Metastases were seen in several liver segments. As a result of mild dizziness, a central nervous system (CNS) MRI scan was performed in January 2020, revealing the presence of brain metastases (Fig. 4a).Fig. 4 Patient 4: Brain magnetic resonance imaging scan results in a January 2020 showing brain metastases and b April 2020 showing reduced lesion size after atezolizumab + BCP treatment. BCP bevacizumab, carboplatin, and paclitaxel
Holocranial radiotherapy was administered, finishing at the beginning of February 2020. After 4 weeks, a repeat MRI did not show any significant changes in the size of the brain lesions. One month after discontinuing holocranial radiotherapy, in the second week of March 2020, he began treatment with four 21-day cycles of atezolizumab + BCP, followed by atezolizumab + bevacizumab until disease progression or toxicity. After two cycles, there were no notable toxicities, and an assessment performed at the end of April 2020 revealed a decrease in all measurable and evaluable tumoral lesions (Fig. 4b). By December 2020, 40 weeks after the start of atezolizumab + BCP, the patient was still in partial response and was being managed on chemotherapy-free maintenance treatment with atezolizumab + bevacizumab.
Compliance with Ethics Guidelines
Data on these patients were collected in accordance with the Helsinki Declaration of 1964 and its later amendments concerning human and animal rights. All patients provided written informed consent to all the diagnostic and therapeutic procedures, for the use of their medical images, and for inclusion in this manuscript. Ethical committee approval was not required, as per Spanish law.
Discussion
EGFR mutations are present in 10–20% of Caucasian patients and about 50% of Asian patients with NSCLC [8]. The L858R point mutation in EGFR exon 21 identified in patient 1 at diagnosis is one of the most common “classical” EGFR mutations, along with a deletion in exon 19 [8]. Together, these mutations represent 85% of EGFR mutations. These activating mutations make the tumor sensitive to tyrosine kinase inhibitors (TKIs) and are an indication for choosing a TKI agent as first-line therapy [9], as occurred in patient 1. Indeed, current NSCLC guidelines note the importance of molecular tumor analysis in determining first-line therapy [9, 10]. In Europe, testing for mutations or rearrangements of ALK, c-ros oncogene 1 (ROS1), and EGFR is considered mandatory, and testing for BRAF V600E mutations is important in countries where BRAF/MEK inhibitors are approved. Other tests include those for human epidermal growth factor receptor 2 (HER2) and MET exon mutations and for fusion genes in RET and NTRK1, which are all are considered evolving biomarkers [10]. The US National Comprehensive Cancer Network guidelines recommend the biomarker test panel for non-squamous NSCLC should, at a minimum, include EGFR mutations, BRAF mutations, ALK fusions, ROS1 fusions, and PD-L1 expression [9].
A primary T790M mutation in EGFR exon 20 is a resistance mutation and more likely to coexist with an L858R mutation than with an exon 19 deletion [11]. While osimertinib has been shown to be effective in patients with a primary T790M mutation, as shown in patient 1, most patients with such mutations progress within 1 year of starting osimertinib [11]. Patient 1 also had high expression of PD-L1 as shown by a TPS of 80%. This suggests that this patient is a good candidate for immunotherapy, but guidelines recommend the use of targeted therapy first-line (before immunotherapy) in patients with sensitizing mutations because the response rate is likely to be higher [9]. Once targeted therapy fails, immunotherapy is indicated.
ALK rearrangements are the driving mutations responsible for the development of NSCLC in 3–7% of patients, and, as observed in patient 2, these patients have a higher risk of developing brain metastases than those with other NSCLC subtypes [12]. The first-generation ALK inhibitor crizotinib, used for second-line treatment in patient 2, has antitumor advantages over chemotherapy in this subtype of patients; however, all patients eventually progress because of drug resistance [10]. Furthermore, the amount of crizotinib that crosses the blood–brain barrier is negligible, which limits its use in patients with brain metastases [10]. In patients with crizotinib-resistant ALK-rearranged NSCLC, one strategy is sequential treatment with next-generation ALK inhibitors, such as ceritinib, alectinib, or brigatinib [12], as used in patient 2.
In the IMpower150 study, most patients had wild-type EGFR and ALK, but 10% of patients were EGFR-mutation positive and 8.5% were ALK-rearrangement positive [7]. IMpower150 is the only trial that has shown positive results with immunotherapy in patients with EGFR- or ALK-positive NSCLC [10]. OS in the population that included patients with these mutations receiving the atezolizumab + BCP regimen was 19.8 months, which was similar to that in the population excluding patients with these mutations (19.5 months), and significantly longer than in the BCP group (15.0 months for the whole population and 14.7 months for the EGFR and ALK wild-type population) [13]. In patients with EGFR mutations, OS was 29.4 months in the atezolizumab + BCP group and 18.1 months in the BCP group [13]. Median progression-free survival (PFS) in patients with EGFR mutations receiving the atezolizumab + BCP regimen was 9.7 months, similar to the PFS in patients with wild-type EGFR (8.3 months), and significantly longer than in the BCP group irrespective of EGFR mutation status (6.1 months and 6.8 months, respectively, in the EGFR mutation and wild-type groups receiving BCP) [7]. Similarly, median PFS was significantly longer in the atezolizumab + BCP group than in the placebo + BCP group in patients with KRAS mutations (8.1 vs. 5.8 months, respectively), as well as in those with wild-type KRAS (9.7 vs. 5.8 months) [7].
The number of mutations, as well as the type, may be relevant to treatment response to atezolizumab. Data suggest that the survival (PFS and OS) benefit of PD-L1 or PD-1 inhibitors may be more marked in patients with a high tumor mutation burden (TMB) than in those with low TMB [14–16], although this result was not seen in some trials [17, 18]. Recent data from randomized trials show that a blood-based assay for TMB can be a useful and valid biomarker for atezolizumab [19].
The cases of patients 3 and 4 illustrated the use of atezolizumab in patients with high disease burden, which was characterized by extensive metastases, including in the liver. In the IMPower150 study, 13% of patients had liver metastases, and atezolizumab + BCP significantly prolonged PFS and OS compared with placebo + BCP in patients with or without liver metastases [7, 20]. However, the difference in PFS and OS between the atezolizumab + BCP group and the BCP group was more marked in the cohort with liver metastases [7]. In the group with liver metastases, the PFS hazard ratio was 0.42 (median PFS 7.4 vs. 4.9 months for atezolizumab + BCP vs. placebo + BCP), whereas in the group without liver metastases the PFS hazard ratio was 0.63 (median PFS 8.3 vs. 7.0 months, respectively) [7]; the OS hazard ratio was 0.52 in the group with liver metastases (median OS 13.3 vs. 9.4 months for atezolizumab + BCP vs. placebo + BCP), and in the group without liver metastases the OS hazard ratio was 0.82 (median OS 20.4 vs. 17.0 months, respectively) [20].
In addition to liver metastases, patient 3 also had bone metastases, which are associated with significantly reduced survival in patients with NSCLC [21, 22]. Moreover, bone metastases are associated with significant pain, fatigue, and disturbed sleep [23], which may negatively affect the patient’s performance status. Indeed, this patient had an ECOG performance status of 2. The IMpower150 study excluded patients with ECOG performance status of 2 or higher [7]; however, data indicate that between one-third and one-half of patients with NSCLC have poor performance status (e.g., ECOG PS of 2 or higher) [24]. It is encouraging that patient 3, who had multiple metastatic sites including in the appendicular skeleton, was able to benefit from treatment with atezolizumab over a prolonged period before disease progression.
The last case presented (patient 4) had both liver and brain metastases at the time of starting treatment with atezolizumab. Adenocarcinomas are the most common tumor type to metastasize to the CNS, and 30–64% of patients with NSCLC have CNS metastases [9, 10]. Historically, patients with brain metastases have had a particularly poor prognosis [25], although this is starting to change with the increasing availability of novel agents. However, patients with untreated brain metastases are usually excluded from clinical trials, many of which also exclude patients with treated brain metastases. Indeed, both the POPLAR study and IMPower150 excluded patients with untreated CNS metastases [5, 7], with neither study reporting on patients with treated brain metastases.
Patient 4 described here received treatment of brain metastases with holocranial radiotherapy, and subsequently derived benefit from atezolizumab treatment. The positive CNS responses in this patient is consistent with findings reported in the OAK trial [6]. In this trial, which specified inclusion of patients with treated, asymptomatic supratentorial CNS metastases, subgroup analysis demonstrated a survival benefit with atezolizumab compared with docetaxel in patients with treated CNS metastases at baseline, with a hazard ratio of 0.54 [95% confidence interval (CI) 0.31–0.94] [6].
These findings for atezolizumab in patients with brain metastases are also consistent with those of a phase II study of pembrolizumab in patients with NSCLC (n = 18) or melanoma and untreated brain metastases, in which pembrolizumab demonstrated activity in brain metastases, with a response rate for brain metastasis of 33% among patients with NSCLC [26].
Although this case series provides valuable information regarding the practical clinical issues associated with atezolizumab + BCP treatment, more well-designed studies are needed to develop evidence-based recommendations on immunotherapy to guide oncologists, immunologists, and other specialists in the management of patients with mutated NSCLC.
Conclusion
As more becomes known about the importance of the various genetic mutations, rearrangements, and expression profiles in NSCLC, testing for targetable oncogenic alterations and immuno-oncology therapy biomarkers is becoming more and more essential for treatment decisions [10]. The first two patients discussed here suggest that the number and type of mutations may be relevant for guiding treatment decisions in NSCLC. To our knowledge, atezolizumab + BCP is the only immunotherapy combination that has demonstrated efficacy in patients with mutated NSCLC [7, 27], and these cases show that the results of these studies have been transferred to clinical practice. Another factor that should be considered when deciding upon a course of therapy is a patient’s performance status. However, clinical trials often exclude patients with poor performance status; hence, the second two cases described here of patients with NSCLC and high disease burden (patients 3 and 4), who benefited from atezolizumab-containing therapy over a prolonged period, are particularly encouraging. Finally, the CNS responses reported here suggest that atezolizumab + bevacizumab may be an important option for the significant unmet need of treating brain metastases in NSCLC. These observations provide valuable insights into the practical clinical issues associated with atezolizumab + BCP treatment in patients with advanced or metastatic NSCLC. These illustrative clinical cases may also help clinicians to identify patients with NSCLC who will potentially gain the most benefit from the use of atezolizumab + BCP in routine clinical practice.
We thank the patients who allowed us to report their clinical details.
Funding
Medical writing assistance and the Rapid Service Fee for this manuscript were funded by Roche.
Medical Writing Assistance
We would like to thank Catherine Rees of Springer Healthcare Communications, and Marie Cheeseman, on behalf of Springer Healthcare Communications, who wrote the outline and first draft, respectively. This medical writing assistance was funded by Roche.
Authorship
All named authors meet the International Committee of Medical Journal Editors (ICMJE) criteria for authorship for this article, take responsibility for the integrity of the work as a whole, and have given their approval for this version to be published.
Compliance with Ethics Guidelines
Data from these patients were collected in accordance with the Helsinki Declaration of 1964 and its later amendments, concerning human and animal rights. All patients provided written informed consent to all the diagnostic-therapeutic procedures, for the use of their medical images, and for inclusion in this manuscript. Ethical committee approval was not required, as per Spanish law.
Disclosures
Dr Cobo reports receiving advisory or consultancy fees from Roche, Bristol Myers Squibb, AstraZeneca, Pfizer, and Boehringer, and travel and accommodation funding from Roche, Bristol Myers Squibb, and AstraZeneca outside of the submitted work. Dr Rodríguez-Abreu reports receiving advisory or consultancy fees from Roche, Merck, Bristol Myers Squibb, AstraZeneca, Pfizer, Boehringer, and Takeda, speaker’s fees from Roche, Merck, Bristol Myers Squibb, AstraZeneca, Boehringer, and Takeda, and travel and accommodation funding from Roche, Merck, Bristol Myers Squibb, and AstraZeneca outside of the submitted work. Diego Pérez Parente is an employee of Roche Farma, Spain. Pedro Ruiz Gracia is an employee of Roche Farma, Spain. Dr González reports receiving advisory or consultancy fees from Roche, Bristol Myers Squibb, AstraZeneca, Boehringer, and Sanofi, speaker’s fees from Roche, Merck, Bristol Myers Squibb, AstraZeneca, Lilly, and Ipsen, and travel and accommodation funding from Roche, Merck, Bristol Myers Squibb, and Lilly outside of the submitted work.
Data Availability
Qualified researchers may request access to individual patient level data through the clinical study data request platform (https://vivli.org/). Further details on Roche's criteria for eligible studies are available here (https://vivli.org/members/ourmembers/). For further details on Roche's Global Policy on the Sharing of Clinical Information and how to request access to related clinical study documents, see here (https://www.roche.com/research_and_development/who_we_are_how_we_work/clinical_trials/our_commitment_to_data_sharing.htm). | ATEZOLIZUMAB, BEVACIZUMAB, CARBOPLATIN, PACLITAXEL | DrugsGivenReaction | CC BY-NC | 33582978 | 18,969,138 | 2021-06 |
Give an alphabetized list of all active substances of drugs taken by the patient who experienced 'Device related infection'. | Practical Issues in the Use of Atezolizumab for Patients with Non-Small Cell Lung Cancer: Case Reports and Literature Review.
Atezolizumab is a monoclonal antibody targeting the programmed death ligand 1 (PD-L1) that was approved in 2017 in the USA and Europe for the second-line treatment of advanced or metastatic non-small cell lung cancer (NSCLC). This review article describes the practical clinical issues associated with atezolizumab treatment in NSCLC using a combination of four illustrative cases and a narrative literature review. The first two cases highlight the importance of tumor mutational status when making treatment decisions. A 62-year-old man with epidermal growth factor receptor (EGFR)-mutated, PD-L1-positive, stage IV lung adenocarcinoma received treatment with second-line atezolizumab + bevacizumab, carboplatin, and paclitaxel (BCP) after first-line osimertinib. In the second case, a 63-year-old man with stage IVb lung adenocarcinoma with anaplastic lymphoma kinase (ALK) translocation received sixth-line treatment with atezolizumab + BCP. The two final cases both had extensive metastases. A 55-year-old woman with EGFR-mutated lung adenocarcinoma received second-line treatment with atezolizumab + BCP after development of multiple metastases, followed by atezolizumab + bevacizumab until last follow-up. A 42-year-old man with PD-L1-positive pulmonary adenocarcinoma (negative for EGFR mutations) developed liver and brain metastases after several lines of therapy. He underwent holocranial radiation and received atezolizumab + BCP, which resulted in a decrease in all measurable and evaluable tumoral lesions. These illustrative cases indicate that the type and number of mutations may influence treatment response to atezolizumab, and that atezolizumab may provide clinical benefit in patients with high disease burden.
Key Summary Points
Why carry out this study?
Targeted immunotherapies, such as atezolizumab, have improved overall survival in patients with advanced or metastatic non-small cell lung cancer (NSCLC) in clinical trials.
Understanding how to treat patients on the basis of their targetable oncogenic mutations or those usually excluded from clinical trials (e.g., with treated brain metastases or high disease burden) is important for oncologists in the real-world clinical setting.
Four cases of treatment with atezolizumab plus bevacizumab, carboplatin, and paclitaxel in patients with NSCLC after at least one previous line of targeted therapy are described to illustrate these clinical issues.
What was learned from the study?
The presented cases suggest that both the number and type of mutations may be relevant for guiding treatment decisions in NSCLC, and that patients with high disease burden or brain metastases may benefit from atezolizumab-containing therapy.
Digital Features
This article is published with digital features, including a summary slide, to facilitate understanding of the article. To view digital features for this article go to https://doi.org/10.6084/m9.figshare.13521968.
Introduction
Non-small cell lung cancer (NSCLC) is the most common type of lung cancer and has a high rate of mortality [1, 2]. According to global estimates, lung cancer was the most commonly diagnosed cancer in 2018 (11.6% of all new cases) and the leading cause of cancer-related mortality (18.4% of all cancer deaths) [3]. In a US analysis of patients with NSCLC, the median overall survival (OS) time after diagnosis was only 13 months, and the estimated 4-year survival rate was between 22% and 28% [1]. However, OS has been improving over recent decades as a result of novel treatments [1, 2], including targeted therapies and immunotherapies.
One such novel treatment is atezolizumab, a monoclonal antibody targeting the programmed death ligand 1 (PD-L1) [4]. Atezolizumab was approved in the USA and Europe for the second-line treatment of advanced or metastatic NSCLC in 2017, based on the results of the phase II POPLAR study [5] and the phase III OAK study [6], in which atezolizumab was associated with significantly better survival compared with docetaxel. Since its approval, the IMpower150 study has demonstrated that atezolizumab in combination with bevacizumab, carboplatin, and paclitaxel (BCP) was more effective than BCP alone as first-line therapy in previously untreated patients with metastatic non-squamous NSCLC [7].
The current review describes the practical clinical issues associated with atezolizumab + BCP in patients with NSCLC, using a combination of illustrative case presentations and a narrative literature review.
Cases Illustrating Role of Mutational Status
Patient 1
A 62-year-old man with no relevant medical or surgical history was diagnosed with stage IIIC lung adenocarcinoma (cT3cN3cM0) in January 2018. He was a former smoker with a 15 pack-year history. Molecular testing showed an L858R point mutation in epidermal growth factor receptor (EGFR) exon 21 and a primary T790M mutation in EGFR exon 20. The tumor was PD-L1 positive, with a tumor proportion score (TPS) of 80% on immunohistochemistry (pharmDx 28–8; DAKO, Glostrup, Denmark).
On February 1, 2018, he began first-line treatment with osimertinib; the 12-week assessment showed a partial response by Response Evaluation Criteria In Solid Tumors (RECIST) 1.1 criteria. Response continued until March 2019, when the disease progressed to stage IV with mediastinal and retroperitoneal involvement (Fig. 1a). At this time, the patient was asymptomatic and started second-line treatment with atezolizumab in combination with BCP, based on the results of the IMpower150 clinical trial [7]. After two cycles, a computed tomography (CT) scan showed partial response by RECIST 1.1 criteria (Fig. 1b), and the patient was able to receive six cycles without significant adverse events.Fig. 1 Patient 1: Computed tomography scan of chest on a March 25, 2019, showing mediastinal and retroperitoneal involvement, b June 6, 2019 after two cycles of atezolizumab + BCP, c October 5, 2019 indicating a maintained radiologic response, and d January 30, 2020 showing tumor progression after 13 weeks of maintenance atezolizumab + bevacizumab. BCP bevacizumab, carboplatin, and paclitaxel
After confirming that the radiologic response was maintained (Fig. 1c), he continued maintenance treatment with atezolizumab + bevacizumab. In February 2020, after 13 cycles, tumor progression was noted (Fig. 1d) and treatment was discontinued. Biopsy showed amplification and overexpression of c-MET, so the patient initiated third-line treatment with telisotuzumab vedotin as part of a clinical trial (NCT03539536).
Patient 2
A 63-year-old man was diagnosed with a stage IVb lung adenocarcinoma with a brain metastasis (T2N3M1b) in February 2013. He was a former smoker (20 pack-year history) with occupational exposure to oil and its derivatives.
On June 13, 2013, he started induction chemotherapy with cisplatin + pemetrexed, undergoing radiosurgery for the brain lesion after the first cycle. He showed a partial radiologic and metabolic response after two cycles of cisplatin + pemetrexed, and a brain magnetic resonance imaging (MRI) revealed a reduction in the size of the brain metastasis.
He received two cycles of cisplatin + pemetrexed from June 13 to June 24, 2013, followed by radical-intent chemoradiation between June 28 and September 6, 2013, consisting of a 60 Gy dose and two cycles of weekly paclitaxel + carboplatin; however, he was unable to continue this treatment because of sustained leukopenia.
A follow-up assessment on October 6, 2014, found no signs of thoracoabdominal progression. Lesions consistent with metastases were identified in the cerebellar vermis and the right centrum semiovale; radiosurgery was administered using 20.7 Gy and 20.9 Gy, respectively, at these sites.
A CT scan on January 12, 2015, showed hilar-mediastinal progression, and a biopsy of the left hilar adenopathy showed that the tumor had wild-type KRAS, was EGFR- and BRAF-mutation negative, and had an anaplastic lymphoma kinase (ALK) translocation. In February 2015, he started second-line treatment with crizotinib and had a partial response. A brain MRI performed on November 3, 2015, showed oligoprogression, for which he received radiosurgery.
On April 4, 2016, he showed pulmonary progression, and 2 weeks later, he started third-line treatment with ceritinib as part of a clinical trial, with a best response of stable disease.
A CT scan on January 30, 2018, showed an increase in the size of the space-occupying lesions in the brain. On March 6 of the same year, he initiated fourth-line treatment with alectinib for metastases, with a best response of stable disease. A further CT scan on July 23, 2018, showed left hilar adenopathy progression and left upper lobe atelectasis. He started fifth-line therapy for metastases with brigatinib on September 5, 2018, and stable disease was his best response.
The patient developed left hemiparesis on January 16, 2019. A brain MRI on January 21 showed signs of radionecrosis, but an acute or subacute ischemic event could not be ruled out. Brain progression, with an increase in the size and number of lesions, was detected in March 2019 (Fig. 2a), and he received hippocampal-sparing whole brain radiation with focal boost (45 Gy dose) on the lesions.Fig. 2 Patient 2: brain magnetic resonance imaging scan results on a March 11, 2019, showing an increase in the size and number of brain lesions, and b March 22, 2020, after 4 cycles of atezolizumab + BCP followed by 13 cycles of maintenance atezolizumab + bevacizumab, and chest computed tomography scan results on c March 11, 2019, prior to, and d March 22, 2020, after atezolizumab-based treatment
On May 8, 2019, he began sixth-line treatment with atezolizumab + BCP. After four cycles (ending on August 8, 2019), he started maintenance treatment with atezolizumab + bevacizumab, achieving a partial brain response. In March 2020, he had received 13 cycles of maintenance treatment, and he showed stable disease as assessed by brain MRI (Fig. 2b) and CT scans of the chest (Fig. 2c, d), abdomen, and pelvis.
CASES Illustrating Management of High Disease Burden
Patient 3
A 55-year-old non-smoking woman without known morbidity was diagnosed with a stage IIIB lung adenocarcinoma (cT4N2) in June 2016. She received treatment with external radiation therapy (dose administered, 66 Gy) concomitantly with cisplatin and vinorelbine (three cycles). A follow-up CT scan showed partial response and the multidisciplinary Committee on Thoracic Tumors recommended surgical intervention. She underwent video-assisted thoracic surgery consisting of an upper right pulmonary lobectomy with systematic hilar and mediastinal lymph node dissection on October 5, 2016. Molecular analysis of the excised tumor showed EGFR exon 19 deletion.
In March 2018, disease progression was detected in lung, pleura, and bone, and in subcutaneous tissue and the lymphatic system. On March 27, she received palliative and decompressive radiation therapy of the lumbar spine (L5) at 8 Gy. Subsequently, in April 2018, she started treatment with afatinib at 40 mg/day, and had a partial response. Afatinib treatment continued until July 2019, when imaging identified disease progression in the liver and bone, and a sacral soft tissue mass (Fig. 3a). Analysis of liquid and sacral mass biopsy did not detect a resistant T790M mutation on EGFR exon 20.Fig. 3 Patient 3: Computed tomography of lumbosacral region on a July 4, 2019, before and b October 8, 2019, after three cycles of atezolizumab + BCP, showing partial response in the sacral soft tissue mass. BCP bevacizumab, carboplatin, and paclitaxel
At this time, the patient’s Eastern Cooperative Oncology Group (ECOG) performance status was 2, and she was negative for PD-L1 (TPS 0%). On August 7, 2019, on the basis of the results of the IMpower150 clinical trial [7], she began second-line treatment with atezolizumab + BCP. After three cycles she showed a partial response (Fig. 3b), and was able to complete six cycles of treatment. However, the patient developed febrile neutropenia and sepsis due to central catheter-related infections in the second cycle leading to hospital admission; grade 2 sensory neuropathy that reverted to grade 1 after discontinuing paclitaxel and carboplatin; and grade 1 asthenia. She then continued treatment with atezolizumab + bevacizumab. By June 2020, she had completed 14 cycles of treatment without relevant toxicities, but treatment was interrupted at that time because a CT scan showed progression of the liver and bone metastases.
Patient 4
A 42-year-old man was diagnosed with stage IIIB pulmonary adenocarcinoma (cT4N2M0) in June 2018; at this time, he had an ECOG performance status of 0. Molecular analysis showed the tumor was negative for EGFR mutations and ALK fusions, with PD-L1 expression on 30% of cancer cells. He was a smoker at the time of diagnosis, smoking one pack daily for 20 years (20 pack-years).
On August 22, 2018, treatment with cisplatin, gemcitabine, and paclitaxel was started, for a total of four 21-day cycles. Treatment was well tolerated, with the exception of nausea and asthenia (both grade 1). Repeat CT scans performed in September 2018 after two treatment cycles, and again in November after four treatment cycles, revealed decreasing tumor volume, with mediastinal contact still present; at this time, results of a brain MRI were normal.
In December 2018, the man underwent a right lower lobectomy; the right lower pulmonary lobe was found to have well-differentiated adenocarcinoma with an acinar pattern, without evidence of vascular invasion, and no visceral pleural infiltration. Surgical resection margins were not affected. TNM staging at this point was ypT1cN0M0. He subsequently received consolidative radiation therapy on the mediastinal bed; he had a performance status of 0 and no complications.
In October 2019, a follow-up examination revealed a de novo pulmonary micronodule in the control CT scan, but this was not seen on a positron emission tomography–CT scan. In February 2020, repeat CT scans revealed a growth on the left perihilar pulmonary nodule and a new right-hand micronodule. Metastases were seen in several liver segments. As a result of mild dizziness, a central nervous system (CNS) MRI scan was performed in January 2020, revealing the presence of brain metastases (Fig. 4a).Fig. 4 Patient 4: Brain magnetic resonance imaging scan results in a January 2020 showing brain metastases and b April 2020 showing reduced lesion size after atezolizumab + BCP treatment. BCP bevacizumab, carboplatin, and paclitaxel
Holocranial radiotherapy was administered, finishing at the beginning of February 2020. After 4 weeks, a repeat MRI did not show any significant changes in the size of the brain lesions. One month after discontinuing holocranial radiotherapy, in the second week of March 2020, he began treatment with four 21-day cycles of atezolizumab + BCP, followed by atezolizumab + bevacizumab until disease progression or toxicity. After two cycles, there were no notable toxicities, and an assessment performed at the end of April 2020 revealed a decrease in all measurable and evaluable tumoral lesions (Fig. 4b). By December 2020, 40 weeks after the start of atezolizumab + BCP, the patient was still in partial response and was being managed on chemotherapy-free maintenance treatment with atezolizumab + bevacizumab.
Compliance with Ethics Guidelines
Data on these patients were collected in accordance with the Helsinki Declaration of 1964 and its later amendments concerning human and animal rights. All patients provided written informed consent to all the diagnostic and therapeutic procedures, for the use of their medical images, and for inclusion in this manuscript. Ethical committee approval was not required, as per Spanish law.
Discussion
EGFR mutations are present in 10–20% of Caucasian patients and about 50% of Asian patients with NSCLC [8]. The L858R point mutation in EGFR exon 21 identified in patient 1 at diagnosis is one of the most common “classical” EGFR mutations, along with a deletion in exon 19 [8]. Together, these mutations represent 85% of EGFR mutations. These activating mutations make the tumor sensitive to tyrosine kinase inhibitors (TKIs) and are an indication for choosing a TKI agent as first-line therapy [9], as occurred in patient 1. Indeed, current NSCLC guidelines note the importance of molecular tumor analysis in determining first-line therapy [9, 10]. In Europe, testing for mutations or rearrangements of ALK, c-ros oncogene 1 (ROS1), and EGFR is considered mandatory, and testing for BRAF V600E mutations is important in countries where BRAF/MEK inhibitors are approved. Other tests include those for human epidermal growth factor receptor 2 (HER2) and MET exon mutations and for fusion genes in RET and NTRK1, which are all are considered evolving biomarkers [10]. The US National Comprehensive Cancer Network guidelines recommend the biomarker test panel for non-squamous NSCLC should, at a minimum, include EGFR mutations, BRAF mutations, ALK fusions, ROS1 fusions, and PD-L1 expression [9].
A primary T790M mutation in EGFR exon 20 is a resistance mutation and more likely to coexist with an L858R mutation than with an exon 19 deletion [11]. While osimertinib has been shown to be effective in patients with a primary T790M mutation, as shown in patient 1, most patients with such mutations progress within 1 year of starting osimertinib [11]. Patient 1 also had high expression of PD-L1 as shown by a TPS of 80%. This suggests that this patient is a good candidate for immunotherapy, but guidelines recommend the use of targeted therapy first-line (before immunotherapy) in patients with sensitizing mutations because the response rate is likely to be higher [9]. Once targeted therapy fails, immunotherapy is indicated.
ALK rearrangements are the driving mutations responsible for the development of NSCLC in 3–7% of patients, and, as observed in patient 2, these patients have a higher risk of developing brain metastases than those with other NSCLC subtypes [12]. The first-generation ALK inhibitor crizotinib, used for second-line treatment in patient 2, has antitumor advantages over chemotherapy in this subtype of patients; however, all patients eventually progress because of drug resistance [10]. Furthermore, the amount of crizotinib that crosses the blood–brain barrier is negligible, which limits its use in patients with brain metastases [10]. In patients with crizotinib-resistant ALK-rearranged NSCLC, one strategy is sequential treatment with next-generation ALK inhibitors, such as ceritinib, alectinib, or brigatinib [12], as used in patient 2.
In the IMpower150 study, most patients had wild-type EGFR and ALK, but 10% of patients were EGFR-mutation positive and 8.5% were ALK-rearrangement positive [7]. IMpower150 is the only trial that has shown positive results with immunotherapy in patients with EGFR- or ALK-positive NSCLC [10]. OS in the population that included patients with these mutations receiving the atezolizumab + BCP regimen was 19.8 months, which was similar to that in the population excluding patients with these mutations (19.5 months), and significantly longer than in the BCP group (15.0 months for the whole population and 14.7 months for the EGFR and ALK wild-type population) [13]. In patients with EGFR mutations, OS was 29.4 months in the atezolizumab + BCP group and 18.1 months in the BCP group [13]. Median progression-free survival (PFS) in patients with EGFR mutations receiving the atezolizumab + BCP regimen was 9.7 months, similar to the PFS in patients with wild-type EGFR (8.3 months), and significantly longer than in the BCP group irrespective of EGFR mutation status (6.1 months and 6.8 months, respectively, in the EGFR mutation and wild-type groups receiving BCP) [7]. Similarly, median PFS was significantly longer in the atezolizumab + BCP group than in the placebo + BCP group in patients with KRAS mutations (8.1 vs. 5.8 months, respectively), as well as in those with wild-type KRAS (9.7 vs. 5.8 months) [7].
The number of mutations, as well as the type, may be relevant to treatment response to atezolizumab. Data suggest that the survival (PFS and OS) benefit of PD-L1 or PD-1 inhibitors may be more marked in patients with a high tumor mutation burden (TMB) than in those with low TMB [14–16], although this result was not seen in some trials [17, 18]. Recent data from randomized trials show that a blood-based assay for TMB can be a useful and valid biomarker for atezolizumab [19].
The cases of patients 3 and 4 illustrated the use of atezolizumab in patients with high disease burden, which was characterized by extensive metastases, including in the liver. In the IMPower150 study, 13% of patients had liver metastases, and atezolizumab + BCP significantly prolonged PFS and OS compared with placebo + BCP in patients with or without liver metastases [7, 20]. However, the difference in PFS and OS between the atezolizumab + BCP group and the BCP group was more marked in the cohort with liver metastases [7]. In the group with liver metastases, the PFS hazard ratio was 0.42 (median PFS 7.4 vs. 4.9 months for atezolizumab + BCP vs. placebo + BCP), whereas in the group without liver metastases the PFS hazard ratio was 0.63 (median PFS 8.3 vs. 7.0 months, respectively) [7]; the OS hazard ratio was 0.52 in the group with liver metastases (median OS 13.3 vs. 9.4 months for atezolizumab + BCP vs. placebo + BCP), and in the group without liver metastases the OS hazard ratio was 0.82 (median OS 20.4 vs. 17.0 months, respectively) [20].
In addition to liver metastases, patient 3 also had bone metastases, which are associated with significantly reduced survival in patients with NSCLC [21, 22]. Moreover, bone metastases are associated with significant pain, fatigue, and disturbed sleep [23], which may negatively affect the patient’s performance status. Indeed, this patient had an ECOG performance status of 2. The IMpower150 study excluded patients with ECOG performance status of 2 or higher [7]; however, data indicate that between one-third and one-half of patients with NSCLC have poor performance status (e.g., ECOG PS of 2 or higher) [24]. It is encouraging that patient 3, who had multiple metastatic sites including in the appendicular skeleton, was able to benefit from treatment with atezolizumab over a prolonged period before disease progression.
The last case presented (patient 4) had both liver and brain metastases at the time of starting treatment with atezolizumab. Adenocarcinomas are the most common tumor type to metastasize to the CNS, and 30–64% of patients with NSCLC have CNS metastases [9, 10]. Historically, patients with brain metastases have had a particularly poor prognosis [25], although this is starting to change with the increasing availability of novel agents. However, patients with untreated brain metastases are usually excluded from clinical trials, many of which also exclude patients with treated brain metastases. Indeed, both the POPLAR study and IMPower150 excluded patients with untreated CNS metastases [5, 7], with neither study reporting on patients with treated brain metastases.
Patient 4 described here received treatment of brain metastases with holocranial radiotherapy, and subsequently derived benefit from atezolizumab treatment. The positive CNS responses in this patient is consistent with findings reported in the OAK trial [6]. In this trial, which specified inclusion of patients with treated, asymptomatic supratentorial CNS metastases, subgroup analysis demonstrated a survival benefit with atezolizumab compared with docetaxel in patients with treated CNS metastases at baseline, with a hazard ratio of 0.54 [95% confidence interval (CI) 0.31–0.94] [6].
These findings for atezolizumab in patients with brain metastases are also consistent with those of a phase II study of pembrolizumab in patients with NSCLC (n = 18) or melanoma and untreated brain metastases, in which pembrolizumab demonstrated activity in brain metastases, with a response rate for brain metastasis of 33% among patients with NSCLC [26].
Although this case series provides valuable information regarding the practical clinical issues associated with atezolizumab + BCP treatment, more well-designed studies are needed to develop evidence-based recommendations on immunotherapy to guide oncologists, immunologists, and other specialists in the management of patients with mutated NSCLC.
Conclusion
As more becomes known about the importance of the various genetic mutations, rearrangements, and expression profiles in NSCLC, testing for targetable oncogenic alterations and immuno-oncology therapy biomarkers is becoming more and more essential for treatment decisions [10]. The first two patients discussed here suggest that the number and type of mutations may be relevant for guiding treatment decisions in NSCLC. To our knowledge, atezolizumab + BCP is the only immunotherapy combination that has demonstrated efficacy in patients with mutated NSCLC [7, 27], and these cases show that the results of these studies have been transferred to clinical practice. Another factor that should be considered when deciding upon a course of therapy is a patient’s performance status. However, clinical trials often exclude patients with poor performance status; hence, the second two cases described here of patients with NSCLC and high disease burden (patients 3 and 4), who benefited from atezolizumab-containing therapy over a prolonged period, are particularly encouraging. Finally, the CNS responses reported here suggest that atezolizumab + bevacizumab may be an important option for the significant unmet need of treating brain metastases in NSCLC. These observations provide valuable insights into the practical clinical issues associated with atezolizumab + BCP treatment in patients with advanced or metastatic NSCLC. These illustrative clinical cases may also help clinicians to identify patients with NSCLC who will potentially gain the most benefit from the use of atezolizumab + BCP in routine clinical practice.
We thank the patients who allowed us to report their clinical details.
Funding
Medical writing assistance and the Rapid Service Fee for this manuscript were funded by Roche.
Medical Writing Assistance
We would like to thank Catherine Rees of Springer Healthcare Communications, and Marie Cheeseman, on behalf of Springer Healthcare Communications, who wrote the outline and first draft, respectively. This medical writing assistance was funded by Roche.
Authorship
All named authors meet the International Committee of Medical Journal Editors (ICMJE) criteria for authorship for this article, take responsibility for the integrity of the work as a whole, and have given their approval for this version to be published.
Compliance with Ethics Guidelines
Data from these patients were collected in accordance with the Helsinki Declaration of 1964 and its later amendments, concerning human and animal rights. All patients provided written informed consent to all the diagnostic-therapeutic procedures, for the use of their medical images, and for inclusion in this manuscript. Ethical committee approval was not required, as per Spanish law.
Disclosures
Dr Cobo reports receiving advisory or consultancy fees from Roche, Bristol Myers Squibb, AstraZeneca, Pfizer, and Boehringer, and travel and accommodation funding from Roche, Bristol Myers Squibb, and AstraZeneca outside of the submitted work. Dr Rodríguez-Abreu reports receiving advisory or consultancy fees from Roche, Merck, Bristol Myers Squibb, AstraZeneca, Pfizer, Boehringer, and Takeda, speaker’s fees from Roche, Merck, Bristol Myers Squibb, AstraZeneca, Boehringer, and Takeda, and travel and accommodation funding from Roche, Merck, Bristol Myers Squibb, and AstraZeneca outside of the submitted work. Diego Pérez Parente is an employee of Roche Farma, Spain. Pedro Ruiz Gracia is an employee of Roche Farma, Spain. Dr González reports receiving advisory or consultancy fees from Roche, Bristol Myers Squibb, AstraZeneca, Boehringer, and Sanofi, speaker’s fees from Roche, Merck, Bristol Myers Squibb, AstraZeneca, Lilly, and Ipsen, and travel and accommodation funding from Roche, Merck, Bristol Myers Squibb, and Lilly outside of the submitted work.
Data Availability
Qualified researchers may request access to individual patient level data through the clinical study data request platform (https://vivli.org/). Further details on Roche's criteria for eligible studies are available here (https://vivli.org/members/ourmembers/). For further details on Roche's Global Policy on the Sharing of Clinical Information and how to request access to related clinical study documents, see here (https://www.roche.com/research_and_development/who_we_are_how_we_work/clinical_trials/our_commitment_to_data_sharing.htm). | ATEZOLIZUMAB, BEVACIZUMAB, CARBOPLATIN, PACLITAXEL | DrugsGivenReaction | CC BY-NC | 33582978 | 18,610,802 | 2021-06 |
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