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+## **Exploration of Targeted Anti-tumor Therapy**
+
+Open Access Review
+# **Early-stage triple negative breast cancer: the therapeutic role of ** **immunotherapy and the prognostic value of pathological complete ** **response**
+
+## **Authors**
+Pierluigi De Santis , Martina Perrone , Chiara Guarini, Anna Natalizia Santoro , Carmelo Laface, Daniela Carrozzo , Gaia Rachele Oliva , Palma Fedele
+
+1 Oncology Unit, Francavilla Fontana Ceglie Messapica Hospital District, 72021 Francavilla Fontana, Italy
+
+2 Department of Medicine and Translational Surgery, Università Cattolica del Sacro Cuore, 00168 Roma, Italy
+
+***Correspondence:** Palma Fedele, Oncology Unit, Francavilla Fontana Ceglie Messapica Hospital District, 72021 Francavilla
+
+[Fontana, Italy. minafedele@hotmail.com](mailto:minafedele@hotmail.com)
+
+**Academic Editor:** Laura Cerchia, Institute of Experimental Endocrinology and Oncology “G. Salvatore”-National Research
+
+Council (IEOS-CNR), Italy; Simona Camorani, Institute of Experimental Endocrinology and Oncology “G. Salvatore”-National
+
+Research Council (IEOS-CNR), Italy
+
+**Received:** May 26, 2023 **Accepted:** December 26, 2023 **Published:** February 28, 2024
+
+**Cite this article:** De Santis P, Perrone M, Guarini C, Santoro AN, Laface C, Carrozzo D, et al. Early-stage triple negative breast
+
+cancer: the therapeutic role of immunotherapy and the prognostic value of pathological complete response. Explor Target
+
+[Antitumor Ther. 2024;5:232–50. https://doi.org/10.37349/etat.2024.00215](https://doi.org/10.37349/etat.2024.00215)
+## **Abstract**
+
+Triple negative breast cancer (TNBC) represents an aggressive disease associated with a high risk of
+
+recurrence after curative treatment and a poor prognosis in the metastatic setting. Chemotherapy was for
+
+years the only treatment available in the early and metastatic setting, due to the lack of actionable targets.
+
+Clinical practice has changed following the results obtained with the addition of immunotherapy to
+
+standard chemotherapy, the development of novel drugs [i.e. antibody-drug conjugates (ADCs)], and the
+
+use of targeted treatments for patients carrying germline pathogenic breast cancer susceptibility genes
+
+( *BRCA* ) *1* or *BRCA 2* variants. The treatment of early-stage disease has had a shift in clinical practice since
+
+July 2021, after the Food and Drug Administration (FDA) approval of pembrolizumab in association with
+
+chemotherapy as neoadjuvant treatment for TNBC and as a single agent in the subsequent adjuvant setting.
+
+This intensive treatment based on the combination of a poly-chemotherapy and an immune checkpoint
+
+inhibitor (ICI) led to the improvement of short- and long-term outcomes, but it has highlighted some new
+
+unmet clinical needs in the treatment of early-stage TNBC: the selection of the most effective adjuvant
+
+therapy and the integration of pembrolizumab with other therapeutic strategies [capecitabine, poly(ADP
+ribose) polymerase (PARP) inhibitors] based on the achievement of pathologic complete response (pCR);
+
+the identification of predictive biomarkers to select patients who could most benefit from the addition of
+
+ICI, to minimize toxicities and to maximize outcomes; the possibility of de-escalating chemotherapy in favor
+
+of immune-combo or novel agents, such as ADCs; the role of immunotherapy in estrogen receptor (ER)-low
+
+patients. The advent of immunotherapy not only addresses current challenges in TNBC treatment but also
+
+holds the promise of a radical transformation in its therapeutic paradigm, enhancing significantly clinical
+
+outcomes and offering new perspectives for patients grappling with this aggressive form of breast cancer.
+
+**© The Author(s) 2024.** This is an Open Access article licensed under a Creative Commons Attribution 4.0 International
+
+License (https://creativecommons.org/licenses/by/4.0/), which permits unrestricted use, sharing, adaptation, distribution
+
+and reproduction in any medium or format, for any purpose, even commercially, as long as you give appropriate credit to the
+
+original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made.
+
+Explor Target Antitumor Ther. 2024;5:232–50 | https://doi.org/10.37349/etat.2024.00215 Page 232
+
+
+-----
+
+ **Keywords**
+
+Triple negative breast cancer, immunotherapy, pathological complete response, neoadjuvant combination
+
+treatment, adjuvant treatment 
+## **Introduction**
+
+Triple negative Breast Cancer (TNBC) is a histological subtype of breast cancer (BC) characterized by the
+immunohistochemical lack of expression (< 1%) of estrogen receptor (ER), progesterone receptor (PgR),
+and human epidermal growth factor receptor 2 (HER2). It accounts for approximately 10–20% of all BC,
+affecting mainly young, premenopausal women, and individuals with inherited gene alterations, such as BC
+susceptibility genes 1/2 ( *BRCA* *1/2* ) mutations [1–3]. It notably presents an aggressive biological behavior
+with a trend to have a higher grade and an often lymph node involvement at diagnosis, an inclination to
+metastasize after curative treatment, and a poorer prognosis in metastatic setting when compared with
+other BC subtypes [4, 5].
+
+For decades, treatment for early TNBC has been based on surgery and subsequent adjuvant
+chemotherapy (CHT) for the reduction of disease recurrence [6]. Therefore, conventional cytotoxic CHT has
+represented the backbone of systemic treatment in the early TNBC, including neoadjuvant treatment, which
+used to reduce tumor size in larger tumors increasing the chances of a breast-conserving surgery [7, 8]. In
+recent years the development of novel therapeutic approaches has been difficult, due to the heterogeneity
+of TNBC and lack of therapeutic targets [9, 10]. Nevertheless, immunotherapy and poly(ADP-ribose)
+polymerase (PARP) inhibitors have shown survival benefits in recent studies.
+
+Specifically, combinations of immune checkpoint inhibitors (ICIs) with CHT or other alternative
+therapeutic compounds could emerge as a successful therapeutic approach in the management of TNBC
+patients. Despite the progress in ICIs representing a notable milestone in TNBC treatment, additional
+investigations are necessary to tackle this issue comprehensively. A profound comprehension of tumor
+subtypes, alongside tumor microenvironment (TME) and in terms of molecular, genetic, and immune
+aspects, would amplify the potential for developing targeted immunotherapy to achieve superior
+therapeutic effectiveness, especially in TNBC [11].
+
+Therefore, in this review, we aimed to investigate the role of immunotherapy in early-stage TNBC, the
+prognostic value of pathologic complete response (pCR) with its therapeutic implications, and the future
+perspectives regarding the systemic treatment of early TNBC, including the discovery of new biomarkers. 
+## **The landscape of immunotherapy in TNBC**
+
+The immune system plays a crucial role in TNBC compared to the other molecular subtypes of BC. Although
+originally BC was considered non-immunogenic, TNBC has a high immunogenic potential, making it a
+promising candidate for immunotherapy, especially with ICIs [12, 13]. TNBC immunogenicity is related to
+intrinsic tumor cell signatures and tumoral surrounding microenvironment features.
+
+Over the last decades thanks to emerging technologies such as next-generation sequencing (NGS), the
+knowledge of the molecular and genetic background of TNBC improved, bringing to light its intertumoral
+and intratumoral heterogeneity.
+
+A first classification divided TNBC into six subtypes: basal-like 1 (BL1), basal-like 2 (BL2),
+mesenchymal (M), M stem-like (MSL), immunomodulatory (IM), and luminal androgen receptor (LAR) [14].
+
+Subsequently, analyzing RNA and DNA-based profiles of 198 TNBC tumors, a four-type classification of
+TNBC was shaped: basal-like immunosuppressed (BLIS), basal-like immune-activated (BLIA), M and LAR
+
+[15]. This classification was further revised with the identification of four specific TNBC subtypes: BL1, BL2,
+M, and LAR, omitting IM and MSL because of the dependence of these two subtypes on the TME features
+
+[14].
+
+Explor Target Antitumor Ther. 2024;5:232–50 | https://doi.org/10.37349/etat.2024.00215 Page 233
+
+
+-----
+
+In addition, TNBC could be classified into three microenvironment phenotypes or clusters:
+
+(1). Cluster 1: “immune-desert” with poor immune cell permeation, due to a high presence of *MYC*
+amplifications and, consequently, a lower recruitment of innate immune cells.
+
+(2). Cluster 2: “innate immune-inactivated” characterized by a hyper-activation of
+
+phosphatidylinositide 3-kinase/protein kinase B (PI3K-AKT) pathway in tumor cells, low tumor
+antigen burden and infiltration of deactivated innate immune cells, fibroblasts, and endothelial
+
+cells. Clusters 1 and 2 are therefore referred to as “cold tumors”.
+
+(3). Cluster 3: “immune-inflamed”, the so-called “hot tumor” that represents about 30% of TNBCs and
+
+is characterized by an abundant adaptive and innate immune cells infiltration and with a high
+expression of immune checkpoint molecules [16].
+
+The potential “hot” conversion of “cold” tumors could improve the efficacy of cancer immunotherapy.
+For example, local IM therapies can express a synergistic effect with immunotherapy by acting on
+components of the TME and immune system function, such as elevating the expression of tumor antigens
+and increasing the recruitment of activated immune cells in the TME [17].
+
+TNBC cancer cell immunological features include genomic instability and high tumor mutational
+burden (TMB), resulting in more somatic mutations and neoantigens [18].
+
+Moreover, approximately 10–20% of TNBC harbor *BRCA 1* or *BRCA 2* germinal mutations, with a
+consequent hereditary deficit in the DNA repair mechanism and strong genomic instability. Several studies
+have demonstrated that TNBC-carrying BRCA mutations are more sensitive to DNA-damaging drugs such as
+anthracyclines, but also platinum agents and PARP inhibitors [19–21]. Sensitivity to these drugs was also
+observed in tumors with alterations in other genes, sharing BRCA-mutant phenotype in the absence of a
+*BRCA 1/2* mutation, namely “BRCAness” [22, 23].
+
+Tumors with *BRCA 1/2* mutations or BRCAness TNBC are more immunogenic than TNBC without these
+genetic alterations [24–26].
+
+Compared to the other BC subtypes the immunogenic TME features in TNBC consist of higher levels of
+vascular endothelial growth factor (VEGF), that promote tumor cell growth and migration such as mitogenactivated protein kinases (MAPKs), tumor-associated macrophages (TAMs), and tumor-infiltrating
+lymphocytes (TILs), white blood cells that migrate towards the tumor, leading to an important
+immunogenic effect and consequently that are involved in killing cancer cells [27–29].
+
+TAMs regulate the interaction between the immune system and cancer cells. CD163+ M2 macrophages,
+which are associated with tumors characterized by higher proliferation and poorer differentiation [30], are
+more present in TNBC and basal-like BC [31]. A prosperous infiltration of TILs is found in TNBC tumors and
+the stroma surrounding them, with a recognized predictive and prognostic role, specifically for CD4+ CD8+
+T cells [32]. Several studies have reported better response to neoadjuvant CHT (NACT) [33] and better
+clinical outcomes in BC with high TIL infiltrate [34–39]. Based on this evidence, the international TILs
+working group started standardizing the evaluation of BC TILs to use it in clinical practice identifying those
+patients that may benefit from emerging immunotherapies with ICIs or combination therapies [40].
+
+All these TME elements contribute to TNBC immunogenicity which also appears to be closely related to
+the concept of TMB, depending on the ineffective DNA repair system with the consequent generation of
+high rates of neoantigens. The upregulated antigen presentation system leads to an increasing number of
+innate and adaptative immune cells and many cytokines interplaying with cancer cells. However, the exact
+relationship between TMB, neoantigens, and immune infiltration is not yet completely understood, and
+some studies have reported an inverse association between immune cells in TME and the rate of somatic
+copy number alterations [41, 42].
+
+Moreover, although TMB is comparable across the three clusters of TNBC, the “immune-inflated”
+phenotype is characterized by a higher degree of immune cells in the TME, but also a high expression of
+immune checkpoints by cancer cells [16]. The rate of TILs, indeed, has been positively related to
+
+Explor Target Antitumor Ther. 2024;5:232–50 | https://doi.org/10.37349/etat.2024.00215 Page 234
+
+
+-----
+
+programmed death ligand 1 (PD-L1) expression [43]. PD-L1 is an immune checkpoint that mediates local
+immune escape in many tumors inducing saturation of activated T cells. Even if PD-L1 prognostic role is yet
+controversial [44], however it results more overexpressed in TNBC compared with other BC and it can
+predict responsiveness to immunotherapy [16].
+
+Therefore, TNBC represents an aggressive BC subtype, associated with high mutational load, high
+tumor immunogenicity and TME diversity.
+## **New paradigms in early TNBC: from CHT to immunotherapy**
+
+**CHT in adjuvant treatment for TNBC**
+
+In early TNBC patients, CHT represents the mainstay of adjuvant and neoadjuvant treatments. Adjuvant
+CHT is recommended for tumor sizes greater than 1.0 cm and patients with nodal involvement, regardless
+of tumor size. Therefore, it can be considered for tumor sizes between 0.6–1.0 cm [45]. A recent large metaanalysis demonstrated that adjuvant CHT with anthracyclines-containing regimens plus taxanes, compared
+with no CHT, can reduce BC mortality rates by about 40% during the first decade after diagnosis. Moreover,
+regimens with higher cumulative and dose-dense schedules of anthracycline (with granulocyte colonystimulating factor support) have shown better survival benefits and more reductions in recurrence [46].
+Three-weekly docetaxel and paclitaxel can be considered in adjuvant setting, but weekly paclitaxel, in a
+subgroup analysis, has shown improved outcomes and is preferred for TNBC [47]. In TNBC in frail patients
+with a known history of heart disease, to minimize the cardiotoxicity of adjuvant treatments, docetaxel
+combined with cyclophosphamide (TC) has proven to be a viable alternative to doxorubicin and
+cyclophosphamide (AC), demonstrating a favorable disease-free survival (DFS) [48]. Therefore, there is a
+broad spectrum of chemotherapeutic treatments for early TNBC that should be customised according to the
+patient and expected toxicities.
+
+**The role of platinum in adjuvant setting for TNBC**
+
+TNBC patients commonly harbor *BRCA 1/2* or BRCAness mutations with a homologous recombination
+deficiency (HRD) that makes them particularly susceptible to platinum agents due to their ability to hit
+cancer cells that have deficient DNA repair mechanisms [49–51]. Several retrospective single-center studies
+have explored the role of adjuvant platinum combined with standard anthracycline and taxane-based
+regimens, with controversial results not showing clear clinical benefits [52, 53]. Nevertheless, a recent
+phase III trials have demonstrated a longer 5-year DFS (86.5% *vs.* 80.3%) with similar results in distant DFS
+and relapse-free survival (RFS) of platinum-containing adjuvant regimens (paclitaxel-carboplatin)
+compared to a standard anthracyclines-containing regimen followed by taxane, however with no benefit in
+overall survival (OS) [54].
+
+Another important factor is platinum resistance. Platinum sensitivity may be affected by changes in the
+hazard ratio (HR) pathway or, in the case of patients with *BRCA 1/2* mutations, by the secondary
+appearance of new *BRCA 1 or 2* mutations that make cancer cells less sensitive to platinum [55, 56].
+
+Other mechanisms of resistance to platinum compounds are:
+
+(1). Modification of drug transport within the tumor cell, by determining decreased influx or increased
+
+efflux.
+
+(2). Increase of detoxification systems.
+
+(3). Decrease of cell apoptosis [57].
+
+Therefore, the benefit of adjuvant platinum-based regimens remains controversial and needs
+validation by prospective adjuvant ongoing trials.
+
+Explor Target Antitumor Ther. 2024;5:232–50 | https://doi.org/10.37349/etat.2024.00215 Page 235
+
+
+-----
+
+**Neoadjuvant treatments for TNBC**
+
+NACT
+
+Several treatment guidelines recommend NACT as the preferred option for stage II or III TNBC and for
+stage I with a tumor size greater than 1 cm. It can be considered in stage I TNBC with a tumor size from
+0.6 cm to 1 cm and/or in the case of tumors with nodal micrometastases. [6, 45]. There is no significant
+difference in survival benefits between patients receiving neoadjuvant or adjuvant CHT after surgical
+resection. However, neoadjuvant treatments can be useful for inoperable tumors rendering them operable
+and they can also downstage patients with operable BC promoting breast-conservation [58, 59]. The use of
+neoadjuvant treatments provides important prognostic information based on response to therapy.
+Achieving a pCR, defined as the lack of cancer cells in tissue samples of breast and axillary lymph nodes,
+after a neoadjuvant treatment, is associated with favorable disease-free and OS in early TNBC, as
+demonstrated in Collaborative Trials in Neoadjuvant Breast Cancer (CTNeoBC) pooled analysis. In this
+study, patients with early BC treated with NACT and followed by surgery who obtained pCR (ypT0 ypN0,
+ypT0/is ypN0) were associated with improved event-free survival (EFS) and OS, especially in TNBC (HR =
+0.24 and HR = 0.16, respectively) [33]. Like adjuvant treatment, traditional NACT is based on anthracyclines
+and taxanes, and a dose-dense regimen is preferred in neoadjuvant settings based on proven improved DFS
+and OS in a large meta-analysis [60].
+
+In recent years, the use of platinum-based combination regimens has been the focus of neoadjuvant
+treatment to increase the rate of pCR in TNBC. Three recent studies demonstrated that combining platinum
+with taxane and anthracycline led to an improvement in the pCR rate in TNBC, with a similar survival
+benefit [61–63]. In Brightness Trial patients with II–III stage TNBC were randomly assigned to receive
+paclitaxel alone, paclitaxel and carboplatin and this combination with a PARP inhibitor, veliparib followed
+by AC. Although the addition of veliparib and carboplatin was associated with an increase of patients who
+achieved a pCR compared to paclitaxel alone (53% *vs.* 31%, *P* < 0.0001), but not to paclitaxel and
+carboplatin, this benefit could be related to the addition of the carboplatin [63]. The initial rationale for
+using the combination of platinum in NACT was that sporadic TNBC can show BRCAness with a major
+response to platinum regimens [50, 51]. However, the greatest benefit was seen in patients who were
+germline BRCA wild type, and only a marginal benefit was observed in the germline BRCA mutant
+subgroup, as was shown in the recent GeparOLA trial. In this trial, patients were randomized to
+neoadjuvant therapy with paclitaxel and carboplatin *vs.* neoadjuvant therapy with paclitaxel and olaparib
+(PARP inhibitor). In both arms, the combination of epirubicin and cyclophosphamide was administered
+next. This study, although limited by a small number of patients enrolled, showed an advantage for the
+carboplatin arm in patients without BRCA mutation (germ or somatic) and high HRD. The 4-year invasive
+DFS (iDFS) rate with olaparib-paclitaxel was 81.2% *vs.* 93.4% with carboplatin-paclitaxel (CP) [HR = 3.03;
+95% confidence interval (CI) = 0.67–13.67; log-rank *P* = 0.1290]. The 4-year OS rate was 89.2% with the
+olaparib combination *vs.* 96.6% with carboplatin (HR = 3.27; 95% CI = 0.39–27.20; log-rank *P* = 0.2444).
+The trend of the iDFS curves was similar in the two treatment arms and independent of germline or somatic
+BRCA mutation [64].
+
+Platinum combinations are currently recommended for selected patients with TNBC who require
+adequate local control before surgical resection [45]. A more recent phase III trial presented at the San
+Antonio Breast Cancer Symposium evaluating the efficacy and safety of adding carboplatin to standard
+sequential taxane-anthracycline NACT in patients with TNBC who had no evidence of metastatic disease,
+has observed improvements in terms of DFS (5-year DFS were 70.6% and 64.5% respectively with a HR =
+0.79, 95% CI = 0.61–1.02, *P* = 0.073) and OS (5-year OS were 74.0% and 66.7% respectively with a HR =
+0.75, 95% CI = 0.57–0.98, *P* = 0.034) with the addition of carboplatin, but these benefits were limited to
+patients who were 50 years of age or younger. Therefore, the pCR in the intention-to-treat population was
+54.5% in the carboplatin arm and 40.3% in the control arm ( *P* < 0.001) [65].
+
+The inclusion of platinum agents as NACT for TNBC remains controversial. Long-term outcomes and
+new prospective studies are needed to clarify the role of platinum agents in this setting.
+
+Explor Target Antitumor Ther. 2024;5:232–50 | https://doi.org/10.37349/etat.2024.00215 Page 236
+
+
+-----
+
+Neoadjuvant immunotherapy
+
+The success of ICIs in metastatic TNBC led to expand their role in neoadjuvant settings. Pembrolizumab and
+atezolizumab have shown progression-free survival (PFS) benefits in phase III trials in advanced setting
+
+[66, 67]. In contrast to atezolizumab that showed conflicting results [68, 69], pembrolizumab consistently
+showed OS benefits in advanced TNBC [66, 70]. In early-stage TNBC two studies evaluated atezolizumab in
+neoadjuvant setting. In the NeoTRIPaPDL1 trial, no improvement in pCR was shown with the addition of
+atezolizumab to a non-anthracycline-containing CHT regimen [71]. More recent Impassion031 phase III
+study evaluating the association of atezolizumab to a standard NACT (nab-palclitaxel weekly for 12 weeks
+followed by 4 cycles of AC), has demonstrated a significant improvement of pCR rates in intention to treat
+(ITT) population (58% in atezolizumab arm *vs.* 41% in placebo arm, *P* = 0.0044), regardless of PD-L1 status,
+meeting the primary endpoint of the study [72]. Therefore, in early BC the combination of pembrolizumab
+with paclitaxel-carboplatin followed by anthracycline increased pCR rate and EFS rate in the KEYNOTE-522
+trial, representing a turning point for the role of immunotherapy in neoadjuvant therapy of TNBC and
+establishing pembrolizumab as a standard treatment during neoadjuvant treatment for stage II and III
+TNBC. The trial evaluated the combination of pembrolizumab (18 cycles, 200 mg every 3 weeks) combined
+with four cycles of paclitaxel (weekly or 3-weekly) and carboplatin (3-weekly), followed by 3-weekly AC for
+4 cycles, compared to placebo with CHT. Pembrolizumab arm showed a 13.6% improvement in pCR [64.8%
+(95% CI = 59.9–69.5%) *vs.* 51.2% (95% CI = 44.1–58.3%)] and in EFS rate [84.5% (95% CI = 81.7–86.9%)
+*vs.* 76.8% (95% CI = 72.2–80.7%)], meeting the primary endpoint of the study, regardless nodal
+involvement and PD-L1 status. The average duration of follow-up is still immature, but a trend of
+superiority in terms of OS in the pembrolizumab arm was nevertheless detected [73]. Limits of this study
+are the lack of biomarkers that predict what patient may benefit from the addition of pembrolizumab and
+the non-utilization of dose-dense schedule of AC which showed superior OS benefit in the neoadjuvant
+setting in TNBC [74].
+
+Moreover, the recent GeparNuevo trial showed that durvalumab (1,500 mg every 4 weeks) added to
+NACT consisting of nab-paclitaxel 125 mg/m [2] weekly for 12 weeks, followed by epirubicin/
+cyclophosphamide every 2 weeks, in early TNBC significantly improved iDFS (85.6% with durvalumab *vs.*
+77.2% with placebo HR = 0.48, 95% CI = 0.24–0.97, stratified log-rank *P* = 0.036) and OS (95.2% *vs.* 83.5%
+with a HR = 0.24, 95% CI = 0.08–0.72, *P* = 0.006), despite a modest pCR increase and no adjuvant
+component of durvalumab [75]. Future studies should aim to define the role of immunotherapy in the
+treatment of early TNBC, to define the ideal duration of these treatments, and should research new
+biomarkers to personalize treatments.
+## **Pathological complete response: prognostic role and therapeutic ** **implications**
+
+In clinical practice, the achievement of pCR after neoadjuvant treatment is correlated to the improvement of
+long-term benefits concerning EFS and OS. Its prognostic value is greatest in aggressive tumor subtypes,
+like in TNBC (EFS: HR = 0.24; OS: HR = 0.16) [33]. Patients who have residual invasive BC after the receipt
+of NACT have a high risk of relapse. Patients with TNBC who do not experience pCR have an estimated 5year EFS of 57% and OS of 47% (compared with 90% EFS and 84% OS, respectively, for patients with earlystage TNBC who demonstrate pCR) [76, 77].
+
+After pre-operative CHT and surgical treatment, patients can receive postoperative radiation therapy
+(RT). Patients with hormone receptor-positive BC [hormone receptor-positive (HR+) BC] are candidates for
+adjuvant endocrine treatment. However, until recently, no adjuvant CHT was expected as standard in
+patients with TNBC. Only follow-up was recommended in those who have pCR or in those with residual
+invasive BC after the receipt of neoadjuvant regimens [78]. To address the unmet clinical need for optimal
+adjuvant treatment in the subgroup of patients with TNBC at high risk of recurrence (those who have not
+achieved the pCR after NACT containing anthracycline, taxane, or both), the Capecitabine for Residual
+Cancer as Adjuvant Therapy (CREATE-X) was designed. The trial did not include only patients with TNBC
+
+Explor Target Antitumor Ther. 2024;5:232–50 | https://doi.org/10.37349/etat.2024.00215 Page 237
+
+
+-----
+
+but also patients with HR+ HER2 negative BC [79]. The results of this phase III trial showed that the
+addition of adjuvant capecitabine (1,250 mg per square meter of body-surface area, twice per day, on days
+1 to 14, every 3 weeks for six or eight cycles) was safe and effective in prolonging DFS and OS among the
+ITT population. The study showed a superior DFS in the capecitabine group than in the control group
+(74.1% *vs.* 67.6% at 5 years; HR = 0.70; *P* = 0.01). Therefore, OS was longer in the experimental group:
+89.2% *vs.* 83.6% of the patients were alive at 5 years (HR = 0.59; *P* = 0.01). Thirty percent of the patients
+had triple negative (TN) disease, and they represent the subgroup with poor prognosis (approximately half
+the patients with TNBC who had a pCR did not have the recurrence of the disease) [33]. The benefit of
+capecitabine *vs.* control in DFS and OS was notable among this subgroup of patients (HR = 0.58 and HR =
+0.52, respectively) [79].
+
+The reflection in the treatment algorithm due to these results was significant.
+
+Some limits of this study are the exclusion of patients who had reached the pCR, for whom only followup was indicated, and the lack of efficacy results selected for residual cancer burden (RCB). The RCB
+quantifies the extent of residual disease after neoadjuvant treatment at the time of surgery. This score uses
+the diameter of residual disease, percentage of vital tumor cells, and diameter of the largest involved lymph
+node to calculate the amount of residual disease. It has been validated with distinct prognostic RCB classes
+in all BC subtypes, with the most significant discriminatory power in TN and Her-2 positive BC. It is
+categorized as RCB-0 (equivalent to a pCR), RCB-1, RCB-2, and RCB-3, reflecting increasingly larger residual
+cancer and respective poor prognoses (in terms of EFS) [80]. Finally, the CREATE-X trial did not examine
+capecitabine efficacy in patients with germline *BRCA 1* or *BRCA 2* pathogenic variants (less than 15% of
+those enrolled) [79].
+
+OlympiA is a phase III study designed to investigate how the PARP inhibitor olaparib might improve
+DFS and OS in patients with resected HR+ BC and TNBC with germline *BRCA 1* or *BRCA 2* mutation. It
+enrolled patients treated with CHT (containing anthracyclines, taxanes or the combination of both) in
+neoadjuvant or adjuvant setting and randomized them to receive olaparib (orally administered at the dose
+of 300 mg twice daily) *vs.* placebo for 1 year after surgical resection (and radiotherapy when indicated).
+Also in this trial, patients with TNBC who underwent NACT followed by surgery were required to have
+residual invasive BC in the breast and/or resected lymph nodes (non-pCR) [81]. Postneoadjuvant
+capecitabine was not foreseen in this trial. iDFS, the primary endpoint of the study, was significantly longer
+among patients assigned to receive olaparib than among those assigned placebo (HR = 0.58; *P* < 0.001). The
+percentage of patients alive and free of invasive disease at 3 years was 85.9% in the olaparib group and
+77.1% in the placebo group. The benefit of adjuvant olaparib was observed irrespective of the germline
+*BRCA* mutation (BRCA 1 *vs.* BRCA 2), the hormone-receptor status, or the timing of previous CHT
+(neoadjuvant *vs.* adjuvant) [81] 4-year iDFS for the olaparib group was 82.7% ( *vs.* 75.4% in placebo group)
+and 4-year distant DFS (DDFS) was 86.5% ( *vs.* 79.1%). Adjuvant olaparib improves OS, with an HR of 0.68
+and a *P* value of 0.009 at 3.5 years of median follow-up, meeting the significance threshold for OS at the
+second planned interim analysis. The OS benefit at 4 years in the olaparib arm compared with the placebo
+arm was reported (89.8% *vs.* 86.4%, respectively) [82].
+
+Both studies have defined the standard of adjuvant therapy post-NACT for patients with *BRCA* wild
+type (CREATE-X) and *BRCA* mutated (OlympiA) TNBC, that did not reach the pCR.
+
+The low percentage of *BRCA* mutated patients enrolled in the CREATE-X, the absence of pre-planned
+subgroup analyzes for this population do not allow for a description of the efficacy of capecitabine in this
+subgroup of patients.
+
+Moreover, there are no prospective randomized trials between capecitabine and olaparib to guide the
+clinical decision in this population, nor combination or sequence data between these two drugs.
+
+It would also be important to consider the potentially severe toxicity profile of such a combination,
+given their overlapping side effects (in particular, cytopenias).
+
+The treatment paradigm of early TNBC has had a real evolution since July 2021, with the introduction
+of immunotherapy following the Food and Drug Administration (FDA) approval of pembrolizumab for high
+Explor Target Antitumor Ther. 2024;5:232–50 | https://doi.org/10.37349/etat.2024.00215 Page 238
+
+
+-----
+
+risk TNBC (tumor size > 1 cm but ≤ 2 cm in diameter with nodal involvement or tumor size > 2 cm in
+diameter regardless of nodal involvement), regardless of tumor PD-L1 expression, in combination with CHT
+as neoadjuvant treatment, and then continued as a single agent as adjuvant treatment after surgery for a
+total duration of approximately 1 year [83].
+
+Results from the KEYNOTE-522 study were the basis for this approval, demonstrating a significantly
+higher rate of pCR at the time of definitive surgery among patients who received pembrolizumab plus NACT
+than among those who received placebo plus NACT and an improvement in long-term benefits [73, 84]. The
+aim of the trial was not to identify the contributions of the neoadjuvant and adjuvant treatment phases, so it
+is difficult to define if these long-term results are related to exposure to adjuvant pembrolizumab or a
+lesser RCB at the end of the neoadjuvant phase in the pembrolizumab–CHT group.
+
+An exploratory analysis of the study then provided data to further describe the prognosis related to the
+RCB after neoadjuvant experimental treatment (Figure 1) [85].
+
+**Figure 1.** The unmet need for the optimal adjuvant treatment according to RCB [85]
+
+The HR for recurrence event in subgroups RBO-0, RCB-1, RCB-2, and RCB-3 are respectively 0.70
+(rates: 5.2% *vs.* 7.3% in the pembrolizumab + CHT *vs.* placebo + CHT), 0.92 (rates: 17.4% *vs.* 20%), 0.52
+(rates: 25.5% *vs.* 44.3%), 1.24 (72.5% *vs.* 69.2%).
+
+The rate of recurrence was numerically lower in all RCB groups with pembrolizumab + CHT, except in
+the small RCB-3 subset (that is represented by 5% and 7% of the population in the study, respectively in
+the experimental and control group). Pembrolizumab shifted RCB to lower categories in most patients
+(RCB-0: 63% *vs.* 56% of patients in the experimental *vs.* the control arm; RCB-1: 9% *vs.* 11%; RCB 18% *vs.*
+20%).
+
+No patients in this trial received adjuvant capecitabine, and there are no randomized efficacy and
+safety data showing that multiagent therapy with pembrolizumab and capecitabine is superior to singleagent therapy in high-risk patients (stage II–III) who did not reach pCR.
+
+At the time, only results from phase II studies in metastatic TNBC demonstrated no new safety signals
+with this combination [86, 87].
+
+Pembrolizumab has also not been studied in combination with olaparib in the adjuvant setting, for the
+treatment of patients with *BRCA* mutations. No efficacy data are reported in the literature, even if some
+
+Explor Target Antitumor Ther. 2024;5:232–50 | https://doi.org/10.37349/etat.2024.00215 Page 239
+
+
+-----
+
+safety data are reported in the metastatic setting, in some early-phase studies that have evaluated the
+combination of PARPis and ICIs, not reporting unexpected toxicities [88, 89].
+
+Prospective trials would be needed to define what is the optimal adjuvant strategy according to RCB
+(single-agent CHT or poly-CHT), how the clinician should decide between olaparib, immunotherapy or
+capecitabine in the treatment of the population with *BRCA* mutations and whether these therapies can be
+administered in combination or sequence, with data in terms of efficacy and safety.
+
+Additional treatment strategies with new drugs are being studied as adjuvant treatment after NACT,
+with antibody-drug conjugates (ADCs) such as datopotamab deruxtecan (with or without durvalumab in
+TROPICS-Breast 03, ClinicalTrials.gov identifier: NCT05629585), and patritumab deruxtecan (HER3-DXd)
+which showed promising clinical response and biological changes in early TNBC [SOLTI TOT-HER3 window
+of opportunity trial part B, presented at European Society for Medical Oncology (ESMO) Breast 2023], or
+with ICIs (A-BRAVE trial, NCT02926196 and SWOG S1418/BR006 trial, NCT02954874).
+## **New biomarkers and frontiers in TNBC**
+
+Recent progress in integrating ICIs and novel agents has revolutionized the therapeutic approach for early
+TNBC. Treatment strategies now emphasize escalating chemotherapeutic agents based on standard
+neoadjuvant regimens. An example is a phase II trial (ACTRN12617000651381) presented at the San
+Antonio Breast Cancer Symposium 2022 evaluating in high-risk TNBC, the addition of ipilimumab and
+nivolumab to neoadjuvant paclitaxel following a suboptimal response to anthracycline-based CHT (< 50%
+tumor reduction) and resulting in promising objective response rate (ORR) (43.7%) and pCR (18.8%) rates,
+regardless of PD-L1 status.
+
+However, it is also crucial to identify subgroups of patients with favorable prognoses, where NACT
+could potentially be de-escalated. Therefore, discovering novel biomarkers to categorize patients with good
+prognoses and safely de-escalate NACT is essential.
+
+TILs show promise as a biomarker for selecting patients who may have favorable outcomes with
+treatment de-escalation. In recent trials, higher TILs levels were associated with a higher pCR rate [71, 75,
+90] and with a better response [75, 91]. Liquid biopsies, such as circulating tumor DNA (ctDNA), could
+serve as promising markers for identifying patients who might benefit from de-escalating or escalating
+neoadjuvant or adjuvant treatment. Rapid ctDNA clearance during NACT in early TNBC is linked to a high
+likelihood of achieving pCR [92]. Conversely, detecting ctDNA after completing NACT and surgery is
+associated with higher recurrence rates and poorer prognoses [93]. The use of dynamic biomarkers, such
+as ctDNA, to guide the choice of treatments in high-risk patients appears increasingly to be an important
+resource to be exploited in future studies.
+
+Furthermore, ADCs are emerging. Particularly, sacituzumab govitecan (SG) an ADC targeting Trop-2
+was approved in metastatic TNBC patients who received ≥ 2 prior systemic therapies in the light of the
+results of the phase III ASCENT study. In this trial patients were randomized (1:1) to receive sacituzumab
+govitecan 10 mg/kg via intravenous infusion on day 1 and day 8 of a 21-day treatment cycle or a treatment
+of physician’s choice (TPC) achieving the primary endpoint (PFS 4.8 *vs.* 1.7 months) and also demonstrating
+an advantage in terms of OS (11.8 months *vs.* 6.9 months) [94]. Another single-arm phase II trial is
+evaluating SG and atezolizumab in combination as adjuvant treatment for patients with TNBC who have
+residual invasive disease after neoadjuvant therapy and detectable ctDNA (ClinicalTrials.gov identifier
+NCT04434040).
+
+Finally, it is essential to redefine, with new dedicated trials, the role of ER-low (1–9%) BC which,
+biologically and prognostically very similar to TNBC, could potentially benefit from the addition of
+immunotherapy to CHT and the role of HER-2 low [score 1+ or 2+ not amplified in fluorescence *in situ*
+hybridization (FISH)] BC in the light of recent results of efficacy of trastuzumab deruxtecan in advanced BC
+HER-2 low. Therefore, future studies are likely to expand the armamentarium at our disposal in this setting.
+
+Explor Target Antitumor Ther. 2024;5:232–50 | https://doi.org/10.37349/etat.2024.00215 Page 240
+
+
+-----
+
+New frontiers in early TNBC are summarized in Figure 2.
+
+**Figure 2.** New frontiers in early TNBC
+## **Interpretation and clinical implications**
+
+TNBC has long been a challenging disease to treat due to its aggressive behavior and the lack of target
+therapies [95].
+
+Thanks to recent developments on TNBC, a series of therapeutic targets have been identified for the
+treatment of metastatic and early setting diseases. Especially for the radically operable disease, the chances
+of cure are increased with treatments aimed at reducing the odds of recurrence after tumor removal.
+
+Anthracycline and taxane-based poly-CHT remains the standard of treatment, most often administered
+preoperatively to assess tumor sensitivity. It aims to increase the rate of local control, making it useful to
+guide breast-conserving surgery and to ensure survival benefits by reaching the pCR.
+
+The introduction of immunotherapy in association with poly-CHT in the neoadjuvant setting has
+increased the rate of pCR, guaranteeing better results in terms of long-term benefits in the KEYNOTE-522,
+the pivotal trial that led to the approval in clinical practice of the use of the anti-PD1, pembrolizumab, in the
+early setting disease (neoadjuvant and adjuvant setting). These clinical findings were based on preclinical
+investigations that overturned the previous belief that BC was not an immunogenic disease [12].
+
+The actual need is to define the optimal adjuvant strategy after neoadjuvant chemo-immunotherapy,
+which must be affected by the patient’s risk of recurrence based on the histological prognostic and evidence
+after radical surgery, the individual’s tolerance of therapy-induced side effects (Figure 3).
+
+In patients with low RCB and a low overall risk of recurrence, pembrolizumab alone should be
+continued. In patients with poor prognostic features of high RCB, this strategy may not be the best choice.
+Patients with high RCB, BRCA wild type, could benefit from capecitabine alone, although it would be
+reasonable to use a combination of capecitabine and pembrolizumab. Patients with high RCB, germline
+*BRCA* mutations, could benefit from olaparib (according to the inclusion criteria of the OlympiA trial),
+although it would be reasonable to use olaparib and pembrolizumab in combination or sequentially.
+
+However, none of these strategies, in monotherapy and/or in combination, have evidence from specific
+randomized trials after the neoadjuvant immunotherapy. There are no data on efficacy and safety in this
+setting. Currently, the best schedule is not known, and new data are awaited on new adjuvant strategies.
+
+Explor Target Antitumor Ther. 2024;5:232–50 | https://doi.org/10.37349/etat.2024.00215 Page 241
+
+
+-----
+
+**Figure 3.** Current treatment algorithm for stage II–III TNBC
+
+Extensive efforts will also be required to investigate and expand access to immunotherapy to ER-low
+populations (ER 1–9%), not included in KEYNOTE-522. It represents a subgroup that does not formally
+meet the definition of TNBC, but shares biology, with nearly 90% of these tumors harboring a basal-like
+intrinsic subtype, and prognosis with TNBC and could share the same benefit from the addition of
+immunotherapy [96, 97].
+
+Furthermore, novel active agents are emerging for the treatment of TNBC and could provide an
+opportunity for a de-escalation of traditional CHT, the anti-trophoblast cell-surface antigen 2 (Trop2)
+sacituzumab govitecan that is currently being investigated in the early setting, including in combination
+with immunotherapy in the ASPRIA trial (ClinicalTrials.gov identifier NCT04434040).
+## **Conclusions**
+
+This review highlights the multitude of advances in the treatment of early-stage TNBC and the important
+
+issues raised.
+
+The management of triple-negative breast cancer (TNBC) has seen notable advancements with the
+identification of therapeutic targets and successful integration of immunotherapy in neoadjuvant
+treatment. However, the current challenge lies in determining the optimal adjuvant strategy post-chemoimmunotherapy, tailoring decisions to individual patient characteristics and prognostic factors. The
+uncertainty surrounding the efficacy and safety of these strategies necessitates further randomized studies,
+while ongoing research explores novel approaches, such as the potential use of innovative agents like
+sacituzumab govitecan in the context of de-escalating traditional CHT. The imperative to extend access to
+immunotherapy to subgroups, such as those with low ER expression, holds crucial promise, paving the way
+for a more personalized and targeted future direction in TNBC treatment.
+
+In the next few years, it will be necessary to design new prospective clinical trials and wait for the
+results of those in progress, for a better knowledge of the efficacy of combination therapies, therapeutic
+sequences and new target drugs for the treatment of a disease which up to a few years ago was considered
+“untargetable”. This should be accompanied by a commitment to biomarker discovery, which could help the
+oncologist make the best decision for patient care.
+
+Explor Target Antitumor Ther. 2024;5:232–50 | https://doi.org/10.37349/etat.2024.00215 Page 242
+
+
+-----
+
+## **Abbreviations**
+
+AC: doxorubicin and cyclophosphamide
+
+ADCs: antibody-drug conjugates
+
+BC: breast cancer
+
+BRCA: breast cancer susceptibility genes
+
+CHT: chemotherapy
+
+CI: confidence interval
+
+CREATE-X: Capecitabine for Residual Cancer as Adjuvant Therapy
+
+ctDNA: circulating tumor DNA
+
+DFS: disease-free survival
+
+EFS: event-free survival
+
+ER: estrogen receptor
+
+FDA: Food and Drug Administration
+
+HER2: human epidermal growth factor receptor 2
+
+HR: hazard ratio
+
+HR+: hormone receptor-positive
+
+ICI: immune checkpoint inhibitor
+
+iDFS: invasive disease-free survival
+
+IM: immunomodulatory
+
+LAR: luminal androgen receptor
+
+M: mesenchymal
+
+NACT: neoadjuvant chemotherapy
+
+OS: overall survival
+
+PARP: Poly(ADP-ribose) polymerase
+
+pCR: pathologic complete response
+
+PD-L1: programmed death ligand 1
+
+RCB: residual cancer burden
+
+TILs: tumor-infiltrating lymphocytes
+
+TMB: tumor mutational burden
+
+TME: tumor microenvironment
+
+TNBC: triple negative breast cancer **Declarations**
+
+**Author contributions**
+
+PDS: Conceptualization, Investigation, Writing—original draft, Writing—review & editing, Validation,
+Supervision. MP, CG, and GRO: Conceptualization, Investigation, Writing—original draft, Writing—review &
+editing. ANS, PF, and CL: Validation, Writing—review & editing, Supervision. DC: Investigation. All authors
+read and approved the submitted version.
+
+Explor Target Antitumor Ther. 2024;5:232–50 | https://doi.org/10.37349/etat.2024.00215 Page 243
+
+
+-----
+
+**Conflicts of interest**
+
+The authors declare that they have no conflicts of interest.
+
+**Ethical approval**
+
+Not applicable.
+
+**Consent to participate**
+
+Not applicable.
+
+**Consent to publication**
+
+Not applicable.
+
+**Availability of data and materials**
+
+Not applicable.
+
+**Funding**
+
+Not applicable.
+
+**Copyright**
+
+© The Author(s) 2024.
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+-----
+

ijo-57-06-1245.md

@@ -0,0 +1,1821 @@
+## **Authors**
+KWANG-AI WON, and CHARLES SPRUCK3
+ConsultantCA, Moraga, CA 94556; 
+Pin Pharmaceuticals, Inc., South San Francisco, CA 94080;
+Tumor Initiation and Maintenance Program, NCI-Designated Cancer Center,
+Sanford Burnham Prebys Medical Discovery Institute, La Jolla, CA 92037, USA
+Received July 14, 2020; Accepted September 9, 2020
+
+
+
+## **Abstract**
+
+Triple‑negative breast cancer (TNBC) accounts for
+10‑15% of all breast cancer cases. TNBCs lack estrogen and
+progesterone receptors and express low levels of HER2, and
+therefore do not respond to hormonal or anti‑HER2 therapies.
+TNBC is a particularly aggressive form of breast cancer that
+generally displays poorer prognosis compared to other breast
+cancer subtypes. TNBC is chemotherapy sensitive, and this
+treatment remains the standard of care despite its limited benefit.
+Recent advances with novel agents have been made for specific
+subgroups with PD‑L1 [+] tumors or germline *Brca* ‑mutated
+tumors. However, only a fraction of these patients responds to
+immune checkpoint or PARP inhibitors and even those who
+do respond often develop resistance and relapse. Various new
+agents and combination strategies have been explored to further
+understand molecular and immunological aspects of TNBC.
+In this review, we discuss clinical trials in the management of
+TNBC as well as perspectives for potential future treatments.
+
+## **Contents**
+
+1. Introduction
+2. Current treatment paradigm
+3. Investigational drugs
+4. New potential therapeutic strategies
+5. Conclusion
+
+## **1. Introduction**
+
+Breast cancer is characterized by heterogeneity at the
+molecular and clinical levels. Several biomarkers including
+
+*Correspondence to:* Dr Kwang‑Ai Won, ConsultantCA, 1988 Ascot
+Drive, Unit A, Moraga, CA 94556, USA
+E‑mail: wonk12pharm@yahoo.com
+
+*Key words:* triple‑negative breast cancer, clinical studies,
+immunotherapy, DNA‑damage response, targeted therapy,
+therapeutic strategy
+
+
+estrogen receptor α (ER α ), progesterone receptor (PR), and
+human epidermal growth factor receptor‑2 (ERBB2/HER2)
+have been established, and the main breast cancer subtypes are
+classified according to their molecular profile (1,2). Traditional
+staging of breast cancer is based on tumor size, lymph node
+involvement, and presence of metastasis, and recently biologic
+markers have been incorporated in the 8th edition of the
+American Joint Committee on Cancer (AJCC), improving the
+prognostic discrimination over anatomic staging alone (3).
+
+Triple‑negative breast cancer (TNBC) is characterized as
+having ≤1% cellular expression of ER and PR as determined
+by immunohistochemistry (IHC), and having HER2 expres­
+*in situ*
+sion of 0 to 1+ by IHC, or 2+ by IHC and fluorescence
+hybridization (FISH) negative (i.e. not an amplified gene
+copy number), according to American Society of Clinical
+Oncology/College of American Pathologists (ASCO/CAP)
+guidelines (4,5). TNBCs are comprised of at least four distinct
+transcriptional subtypes: Two basal subtypes, BL1 and BL2; a
+mesenchymal subtype M, which is devoid of immune cells; and
+a luminal androgen receptor (AR) subtype LAR (1,2). TNBC is
+also subdivided into 6 different subgroups based on molecular
+heterogeneity: Basal‑like; mesenchymal‑like; mesenchymal
+stem‑like; luminal AR expression; immunomodulatory; and
+unstable type (6). TNBC represents approximately 15‑20% of
+all newly diagnosed breast cancers and is generally a more
+aggressive disease with a poorer prognosis and higher grade
+than other types of breast cancer, accounting for 5% of all
+cancer‑related deaths annually. The median overall survival
+(OS) for the disease is 10.2 months with current therapies, with
+a 5‑year survival rate of ~65% for regional tumors and 11% for
+those that have spread to distant organs (7,8).
+
+In this review, we discuss current TNBC treatments and
+key examples of improved clinical benefit, as well as new
+therapeutic strategies with which to treat the disease.
+
+## **2. Current treatment paradigm**
+
+TNBC is chemotherapy sensitive, and this treatment remains
+the standard of care (SOC). Common chemotherapies
+include anthracycline (e.g., DNA intercalating agent and
+topoisomerase II blocker doxorubicin), alkylating agents (e.g.,
+cyclophosamide), an anti‑microtubule agent taxane, and an
+anti‑metabolite fluorouracil (5‑FU). The current SOC for newly
+
+
+-----
+
+1246 WON and SPRUCK: TNBC THERAPY: CURRENT AND FUTURE PERSPECTIVES
+
+
+diagnosed early TNBC consists of neoadjuvant chemotherapy,
+followed by surgery. For patients with relapsed/refractory
+TNBC, there is no standard chemotherapy regimen. Responses
+to treatment are usually short in duration and followed by
+rapid relapse, and visceral and brain metastases are common.
+Available therapies for patients with advanced TNBC include
+anti‑metabolites capecitabine and gemcitabine, non‑taxane
+microtubule inhibitor eribulin, and DNA cross‑linker
+platinums. The median progression‑free survival (PFS) with
+chemotherapy ranges from 1.7 to 3.7 months; the median OS
+from the onset of metastasis is 10 to 13 months. In clinical
+
+trials, patients with advanced TNBC treated with single‑agent
+taxane‑ or platinum‑based chemotherapy had a median PFS
+of 4 to 6 months and a median OS of 11 to 17 months (9‑11).
+
+New treatment options for patients with advanced TNBC
+have recently emerged, especially in cases where surgery is
+not an option.
+
+TNBC is more immunogenic than other breast cancer
+subtypes with tumor‑infiltrating lymphocytes (TILs) in its
+microenvironment. However, TNBC also displays a high level
+of programmed cell death‑ligand 1 (PD‑L1) expression (12,13).
+Thus, immunotherapies targeting the programmed cell
+death‑1 (PD‑1) receptor/PD‑L1 pathway that maintains immu­
+nosuppression in the tumor environment in TNBC have been
+explored and atezolizumab (anti‑PD‑L1 antibody) in combi­
+nation with nanoparticle albumin‑bound (nab)‑paclitaxel
+was approved as a first‑line therapy by the US Food and
+Drug Administration (FDA) based on the IMpassion130
+trial (NCT02425891) in 2019. This immuno‑chemotherapy
+became SOC for patients with PD‑L1 [+], unresectable, locally
+advanced or metastatic TNBC. Note that the survival benefit
+was exclusively in PD‑L1 [+] TNBC patients. The threshold is
+1% PD‑L1 expression on infiltrating immune cells by an
+approved companion diagnostic SP142 IHC assay and 41%
+of enrolled patients showed PD‑L1‑positive expression in
+the IMpassion130 trial. This is in contrast to studies in other
+types of cancer which showed benefit for checkpoint inhibitor
+therapy even in patients with negative PD‑L1 expression. In
+the first interim analysis of IMpassion130, the median PFS
+was 7.5 vs. 5.0 months with chemotherapy and the median OS
+was 25.0 vs. 15.5 months with chemotherapy among patients
+with PD‑L1 [+]
+tumors (14). In the pre‑specified second interim
+analysis (data cutoff January 2, 2019), the median OS was 25.0
+vs. 18.0 months with chemotherapy. Overall, the combination
+was well‑tolerated and immune‑related adverse events (AEs)
+included rash, hypothyroidism, and pneumonitis (15). Another
+immunotherapy, pembrolizumab (anti‑PD‑1 antibody), was
+approved in 2017 as a histology agnostic immunotherapy in all
+microsatellite instability‑high (MSI‑H) and/or mismatch repair
+deficient (dMMR) tumors. This is the first FDA‑approved
+cancer treatment based on a tumor biomarker without regard
+to the original location of the tumor. However, MSI‑H is rare
+in breast cancer (<2%) (16‑18).
+
+BRCA1 and BRCA2‑deficient tumors exhibit impaired
+homologous recombination repair (HRR) and synthetic lethality
+with poly(ADP‑ribose) polymerase (PARP) inhibitors (19,20).
+The FDA approved olaparib and talazoparib in 2018 to treat
+advanced‑stage HER2‑negative breast cancer in individuals
+with a *Brca1* or *Brca2* mutation. The FDA also approved the
+companion diagnostic test to identify germline *Brca* ‑mutated
+
+
+(gBRCAm) breast cancer patients. Approximately 5% of patients
+with breast cancer carry a gBRCAm. Olaparib approval was
+based on data from the OlympiAD Phase III (NCT02000622)
+trial comparing olaparib to physician's choice of chemotherapy
+(capecitabine, vinorelbine or eribulin). Olaparib was associated
+with a 42% increase in median PFS as compared to the control
+group (7 vs. 4 months) in gBRCAm HER2‑negative meta­
+static breast cancer patients with previous chemotherapy (21).
+There was no statistically significant improvement in OS with
+olaparib compared to the control group (19.3 vs. 17.1 months),
+but there was potential OS benefit among patients with no prior
+chemotherapy for metastatic breast cancer (HR 0.51, 95% CI
+0.29‑0.90) (22). Olaparib was generally well‑tolerated, with no
+evidence of cumulative toxicity including the risk of developing
+anemia during extended exposure. Talazoparib approval was
+based on data from the EMBRACA Phase III (NCT01945775)
+trial comparing talazoparib to gemcitabine or to the same
+physician choice of standard therapy as the OlympiAD trial.
+Talazoparib increased median PFS by 46% (8.6 vs. 5.6 months)
+in gBRCAm HER2‑negative locally advanced or metastatic
+breast cancer patients with previous chemotherapy including an
+anthracycline and/or taxane. Talazoparib presented with hema­
+tologic grade 3‑4 AEs (primarily anemia), which occurred in 55
+vs. 38% of the patients with standard therapy, and an improved
+side‑effect profile in patient‑reported outcomes (23).
+
+## **3. Investigational drugs**
+
+To improve therapeutic benefit in TNBC treatment,
+various agents have been explored in clinical studies. They
+include immuno‑ and targeted‑therapies in the networks of
+tumor‑stroma, DNA damage response (DDR), cell surface or
+intracellular receptors, and signaling pathways as well as cell
+surface markers for selective drug delivery, and antibody‑drug
+conjugates (ADCs) (Fig. 1). As of March 2020, 399 ongoing
+studies for TNBC have been listed on ClinicalTrials.gov and
+select Phase III studies are listed in Table I.
+
+*Immunotherapy: Immune checkpoint.* TILs are frequent
+in TNBC, correlate with increased pathologic complete
+response (pCR) to neoadjuvant chemotherapy, and are predic­
+tive of disease‑free survival (DFS) and OS in early‑stage
+TNBC (24‑26). Expression of immune regulatory checkpoints
+is an adaptive method of tumor resistance to infiltrating lympho­
+cytes within the tumor microenvironment. Multiple strategies
+have been used to enhance the response to PD‑1/PD‑L1
+blockade in pre‑clinical and early clinical studies, including
+several intratumoral immune modulators and targeted
+agents (27). The activity of immunotherapy, such as immune
+checkpoint inhibitors, can be enhanced by chemotherapeutic
+agents through the stimulation/release of antigens, thus leading
+to promotion of immunogenic cell death. Currently, clinical
+trials investigating the use of immune checkpoint inhibitors
+are ongoing either as a single agent or in various combinations
+with other agents beyond the metastatic setting and even in the
+first‑line setting (28).
+
+*Neoadjuvant treatment.* Studies determining benefit from
+neoadjuvant checkpoint inhibitor therapy have yielded mixed
+outcomes. Neoadjuvant chemotherapy with pembrolizumab
+
+
+-----
+
+INTERNATIONAL JOURNAL OF ONCOLOGY 57: 1245-1261, 2020 1247
+
+Figure 1. Immuno‑ and targeted‑therapies in key TNBC clinical studies. Various agents in the networks of TNBCs and immune cells have been explored,
+as well as tumor‑stroma interactions in the tumor microenvironment (TME). Targets and agents relevant to immune checkpoint, cell surface or intracel­
+lular receptors, signaling pathways, DNA damage response, and cell cycle checkpoint are shown. Various chemotherapy agents are listed in the box. AS,
+Adagloxad simolenin); LV, Ladiratuzumab vedotin; SG, Sacituzumab govitecan‑hziy; T‑DXd, tastuzumab deruxtecan; TNBC, triple‑negative breast cancer;
+A 2a R, adenosine 2A receptor; A 2b R, 2B receptor; PD‑1, programmed cell death‑1; PD‑L1, programmed cell death‑ligand 1; VEGF‑A, vascular endothelial
+growth factor A; RTKs, receptor tyrosine kinases; PARP, poly(ADP‑ribose) polymerase; CDK, cyclin‑dependent kinase; CD, cluster of differentiation; ATR,
+ataxia telangiectasia and Rad3‑related kinase; CHK1, checkpoint kinase 1; DNA‑PK, DNA‑dependent protein kinase; AR, androgen receptor; PI3K, phospha­
+tidylinositol 3‑kinase.
+
+
+have demonstrated manageable safety and promising anti­
+tumor activity for patients with early‑stage TNBC in the
+KEYNOTE‑173 Phase 1b (NCT02622074) (29) and I‑SPY2
+Phase II (NCT01042379) trials (30). The KEYNOTE‑522
+Phase III trial (NCT03036488) further explored neoadjuvant
+chemotherapy with or without pembrolizumab followed by
+surgery and pembrolizumab or placebo adjuvantly. The neoad­
+juvant combination showed a significantly higher pCR rate
+than the placebo‑chemotherapy group (65 vs. 51%). Note that
+a similar pCR benefit (~15%) in both the PD‑L‑positive and
+‑negative subgroups was observed, suggesting that neoadju­
+vant pembrolizumab may benefit patients regardless of PD‑L1
+levels. This is different from the advanced setting where only
+the PD‑L1‑positive patients benefit from atezolizumab. The
+toxicity profiles were as expected for each treatment, with
+similar rates (78 vs. 73%) of grade ≥3 treatment‑related AEs
+(TRAEs) (31).
+
+NeoTRIPaPDL1 Phase III (NCT02620280) trial also
+explored neoadjuvant chemotherapy with or without atezoli­
+zumab followed by surgery and four cycles of an anthracycline
+regimen. However, in this trial for patients with early‑stage
+high‑risk or locally advanced unilateral breast cancer there
+was no improvement in pCR with the combination therapy
+(44 vs. 41% with the control arm) (32). Note that the neoad­
+juvant chemo‑regimen was different from KEYNOTE‑522
+which included another round of chemotherapy following
+carboplatin and nab‑paclitaxel. The difference in the targets,
+PD‑1 for pembrolizumab vs. PD‑L1 for atezolizumab, may
+
+
+also have contributed to the different outcomes. Another
+
+Phase III (NCT03197935) trial, IMpassion031 study also
+explored atezolizumab in combination with chemotherapy
+(nab‑paclitaxel followed by doxorubicin and cyclophospha­
+mide) in comparison to placebo plus chemotherapy in the
+neoadjuvant setting. Treatment with atezolizumab continued
+adjuvantly for those in the combination arm of the study (33).
+The primary endpoint was pCR.
+
+*In the advanced setting.* As a first‑line treatment option for
+patients with locally recurrent, inoperable or metastatic TNBC,
+pembrolizumab was evaluated in combination with investiga­
+tor's choice of chemotherapy ( *i.e.* nab‑paclitaxel, paclitaxel or
+gemcitabine/carboplatin), compared to placebo plus chemo­
+therapy (KEYNOTE‑355 Phase III trial, NCT02819518).
+A significant PFS benefit with the pembrolizumab‑chemo
+combination in patients whose tumors expressed PD‑L1
+(CPS ���10) was reported (9.7 vs. 5.6 months for chemotherapy
+alone) (34). The study is currently in progress to evaluate OS,
+the other primary endpoint of the trial.
+
+In contrast to other studies of immunotherapy combined
+with SOC chemotherapy, the Tonic trial (NCT02499367)
+in metastatic TNBC was based on an adaptive trial design
+that explores a sequential treatment with anti‑PD‑1 antibody
+nivolumab after 2 weeks of chemotherapy or radiotherapy. The
+hypothesis is that short‑term treatment induces a more favor­
+able tumor microenvironment that would enhance sensitivity
+to immune checkpoint blockade in TNBC. The highest overall
+
+
+-----
+
+1248 WON and SPRUCK: TNBC THERAPY: CURRENT AND FUTURE PERSPECTIVES
+
+
+-----
+
+INTERNATIONAL JOURNAL OF ONCOLOGY 57: 1245-1261, 2020 1249
+
+
+-----
+
+1250 WON and SPRUCK: TNBC THERAPY: CURRENT AND FUTURE PERSPECTIVES
+
+
+-----
+
+INTERNATIONAL JOURNAL OF ONCOLOGY 57: 1245-1261, 2020 1251
+
+response rate (ORR) was observed with doxorubicin induc­
+tion (35%) followed by nivolumab/doxorubicin. Doxorubicin
+induction also upregulated immune‑related genes as well
+as inflammation, JAK‑STAT, and TNF‑ α signaling‑related
+genes, suggesting a more favorable tumor microenvironment
+induced by these chemotherapies (35). The InCITe Phase II
+trial (NCT03971409) also includes a two‑week induction of
+binimetinib (MEK inhibitor), utomilumab (4‑1BB agonist), or
+PF‑04518600 (anti‑OX40 antibody) which may help activate
+the immune system. The trial explores how well anti‑PD‑L1
+antibody avelumab might work with one of those agents after
+induction in stage IV or unresectable and recurrent TNBC.
+
+*Immunotherapy: Adenosine pathway.* Adenosine is catabo­
+lized from ATP and often overproduced and released by
+tumor cells. It is also converted from extracellular nucleotides
+
+by the plasma membrane protein, cluster of differentiation 73
+(CD73), which is upregulated in many cancer types (36,37).
+The excess adenosine in the tumor microenvironment
+
+activates the adenosine 2A receptor (A 2a R) and 2B receptor
+(A 2b R) (38,39) which are highly expressed on the cell surfaces
+of lymphocytes and myeloid cells, respectively, leading to
+immunosuppressive effects (Fig. 2). Targeting these receptors
+and enzymes could lead to reactivation of antitumor immunity
+by abrogating the inhibitory effect on the immune system
+and enhancing the cytotoxic T lymphocyte (CTL)‑mediated
+immune response (40,41).
+
+Combinations of adenosine pathway inhibitors and
+immune checkpoint inhibitors have been explored in clinical
+trials. NZV930 (SRF373) is an anti‑CD73 monoclonal
+antibody that binds to CD73 on tumor cells, leading to
+internalization of CD73, thereby preventing CD73‑mediated
+conversion of extracellular AMP to adenosine. A Phase I/Ib
+
+study (NCT03549000) is underway to evaluate NZV930 alone
+and in combination with PD‑1 inhibitor PDR001 and/or A 2a R
+antagonist NIR178 in patients with advanced malignancies
+including TNBC. NIR178 is an antagonist of A 2a R, blocking
+adenosine/A 2a R‑mediated inhibition of T lymphocytes. A
+Phase II study (NCT03207867) is underway for NIR178 in
+combination with PD‑1 inhibitor spartalizumab in multiple
+solid tumors and diffuse large B‑cell lymphoma (DLBCL)
+to assess if the addition of the adenosine antagonist improves
+the efficacy of PD‑1 inhibition. A dual adenosine A 2a R/A 2b R
+receptor antagonist, AB928, is currently being evaluated in a
+Phase I study (NCT03629756) in combination with the PD‑1
+inhibitor AB122 in patients with advanced malignancies. Early
+results show a favorable safety profile of AB928 combination
+therapy and predictable PK/PD correlation (42).
+
+*DNA‑damage response: PARP.* Approximately 60‑70% of
+breast cancer patients with an inherited *Brca1/2* mutation
+are TNBC subtype and 10‑30% of TNBC patients harbor
+a *Brca*
+pathogenic variant (43,44). A condition defined as
+‘BRCAness’ (45), which includes mutations in HRR genes
+through genetic or epigenetic inactivation, leads to suscep­
+tibility to both platinum and PARP inhibitors. Various
+PARP inhibitors (e.g. veliparib, niraparib, and rucaparib as
+well as olaparib and talazoparib) have been assessed in the
+neoadjuvant and adjuvant settings and in combination with
+other agents.
+
+
+-----
+
+1252 WON and SPRUCK: TNBC THERAPY: CURRENT AND FUTURE PERSPECTIVES
+
+(TOPACIO/KEYNOTE‑162 Phase II trial, NCT02657889)
+showed higher response rates in patients with tumor *Brca* muta­
+tions (tBRCAm): ORR of 28% in all (biomarker‑unselected)
+patients vs. 60% for tBRCAm patients. The combination
+therapy was safe with a tolerable safety profile (49).
+
+In MEDIOLA Phase I/II trial (NCT02734004) the combi­
+nation of olaparib and durvalumab showed ORR of 63% in a
+cohort of patients with gBRCAm metastatic breast cancer (50).
+In the I‑SPY 2 Phase II study (NCT01042379), adding the same
+combination to neoadjuvant paclitaxel led to improved pCR
+rates in patients with high‑risk, HER2‑negative stage II/III
+breast cancer compared with single‑agent paclitaxel. In those
+with TNBC, the pCR rate was 47 vs. 27% with paclitaxel alone.
+AEs were consistent with the known safety profiles of each
+agent alone (51). In metastatic TNBC, the efficacy of induction
+treatment of olaparib followed by the combination treatment of
+
+Figure 2. ATP‑adenosine pathway. Adenosine is generated from ATP by olaparib and durvalumab is being assessed in a Phase II study
+CD39 and CD73. It binds to A2 receptors on immune cells and blocks T cell (NCT03801369) (52). Patients with ≤2 prior chemotherapy
+priming, expansion, and activation, natural killer (NK) cell degranulation, regimens for metastatic breast cancer are eligible, but patients
+dendritic cell (DC) maturation and activation, and tumor‑associated macro­
+phage (TAM) M1 polarization, thus leading to immunosuppression. ATP, with gBRCAm TNBC are excluded. The primary end point is
+adenosine triphosphate; AMP, adenosine monophosphate; CD, cluster of ORR.
+differentiation. The DORA Phase II trial (NCT03167619) is evalu­
+
+ating olaparib as a maintenance therapy with or without
+durvalumab in patients with advanced TNBC who achieve at
+
+*Neoadjuvant and adjuvant settings.* A PARP inhibitor appears least stable disease after 3 cycles of platinum‑based chemo­
+to have efficacy for neoadjuvant treatment of patients with therapy. Another study of a PARP inhibitor as a maintenance
+gBRCAm TNBC. Talazoparib achieved encouraging pCR therapy, KEYLYNK‑009 Phase II/III trial (NCT04191135),
+in patients with gBRCAm breast cancer, including TNBC, is underway in metastatic TNBC to assess the efficacy of
+and HR [+] breast cancer, as a neoadjuvant single‑agent without olaparib plus pembrolizumab vs. chemotherapy plus pembro­
+the addition of chemotherapy (46). Currently a larger, multi­ lizumab after induction with first‑line chemotherapy plus
+center, neoadjuvant Phase II trial (NCT03499353) is ongoing. pembrolizumab (53).
+However, the addition of a PARP inhibitor to standard neoad­
+juvant chemotherapy was found to be not beneficial. In the *In combination with DDR‑HRR pathway inhibitors.*
+BrighTNess Phase III trial (NCT02032277) the addition of Resistance to PARP inhibitors can occur in certain cancer
+PARP inhibitor veliparib to carboplatin and paclitaxel followed contexts by various mechanisms, including increased HRR
+by doxorubicin and cyclophosphamide did not improve pCR capacity and decreased cell cycle progression and DNA
+whereas the addition of veliparib and carboplatin to paclitaxel replication stress. RAD51 overexpression has been observed
+did. Therefore, the addition of carboplatin but not veliparib to in a wide range of human cancers, particularly TNBCs and
+paclitaxel was proposed as a potential component of neoad­ serous ovarian cancers (54,55). Upregulation of RAD51 in
+juvant chemotherapy for patients with high‑risk TNBC (47). BRCA1‑defective cells is also associated with resistance to
+
+PARP inhibitors have also been studied as an adju­ PARP inhibitor (56,57). Inhibitors of key mediators of DNA
+vant single‑agent therapy. The OlympiA Phase III trial repair and replication, such as ataxia telangiectasia mutated
+(NCT02032823) was designed to assess olaparib in patients kinase (ATM), ataxia telangiectasia and Rad3‑related kinase
+with gBRCAm and high‑risk HER2‑negative breast cancer (ATR), checkpoint kinase 1 (CHK1) and checkpoint kinase
+who completed definitive local treatment and neoadjuvant or 2 (CHK2), DNA‑dependent protein kinase (DNA‑PK), and
+adjuvant chemotherapy. The primary outcome measure will be WEE1 kinase (Fig. 3) have been assessed to determine if they
+invasive DFS with a time frame of up to 10 years. can sensitize tumor cells to treatment with PARP inhibitors,
+
+as these inhibitors were found to prevent the accumulation of
+
+*In combination with immunotherapy.* A crosstalk exists RAD51 in TNBC (58).
+between PARP inhibition and the PD‑L1/PD‑1 immune The VIOLETTE Phase II study (NCT03330847) was
+checkpoint axis. PARP inhibitors upregulate PD‑L1 expres­ set up to assess the combinatory inhibition of PARP and a
+sion on tumor cells by inhibiting glycogen synthase kinase 3 component of the ATR‑CHK1‑WEE1 axis. Olaparib with
+beta (GSK3 β ) and activating the cGAS‑STING pathway (48). DDR kinase ATR inhibitor AZD6738 was compared to
+Thus, primary/acquired resistance to PARP inhibitors seems olaparib monotherapy in the second‑ or third‑line setting of
+to be associated with the development of immune evasion metastatic TNBC. Patients were stratified by *Brca* and HRR
+mechanisms. Multiple clinical studies are underway to assess gene mutation status and the primary endpoint was PFS (59).
+synergy between therapeutic strategies of PARP inhibition and The study also included a combination arm of olaparib with
+immune checkpoint blockers. the first‑in‑class WEE1 inhibitor adavosertib. WEE1 inhibitor
+
+In platinum‑resistant, advanced, or metastatic was found to potentiate the activity of DNA‑damaging agents
+TNBC, niraparib combined with pembrolizumab in preclinical TNBC models (60,61) and its potential clinical
+
+
+-----
+
+INTERNATIONAL JOURNAL OF ONCOLOGY 57: 1245-1261, 2020 1253
+
+Figure 3. DNA damage response pathways. Double‑strand breaks (DSB) or single‑strand breaks (SSB) activate DNA damage response (DDR) pathways,
+leading to cell cycle arrest and DNA repair or cell death depending on cell context. PARP1 senses DNA breaks and is involved in SSB repair. Oncogenic
+pathways including RAS, PI3K, AR, and MYC signaling can affect HR repair activity and contribute to resistance to PARP inhibitor treatment. MRN,
+MRE11-RAD50-NBS1 complex; ATRIP, ATP interacting protein; HR, homologous recombination; NHEJ, non­homologous end joining; H2AX, histone
+H2AX; XRCC4, X‑ray repair cross‑complementing protein 4; ATR, ataxia telangiectasia and Rad3‑related protein; CHK1/2, checkpoint kinase 1/2; CDK1/2,
+cyclin‑dependent kinase 1/2; DNA‑PK, DNA‑dependent protein kinase; AR, androgen receptor; PI3K, phosphatidylinositol 3‑kinase.
+
+
+value was observed in a Phase I study in patients with *Brca*
+mutations (62). However, the combination treatment arm of
+olaparib and adavosertib was discontinued in the VIOLETTE
+study and patients were offered the opportunity to continue
+treatment on olaparib monotherapy. The CHK1 inhibitor
+prexasertib in combination with olaparib was also explored in
+early clinical trials (63), but development of prexasertib was
+discontinued by the sponsor in 2019.
+
+*Intracellular signaling pathway targets*
+*PI3K/AKT pathway.* A wide range of malignancies including
+TNBC show dysregulated phosphatase and tensin homolog
+(PTEN)/phosphoinositide 3‑kinases (PI3K)/protein kinase B
+(AKT)/mammalian target of rapamycin (mTOR) signaling
+due to mutations in multiple signaling components. Loss
+of PTEN, a negative regulator of AKT, was found to be
+correlated with decreased T‑cell infiltration at tumor
+
+sites in patients, and inhibition of the PI3K‑AKT pathway
+re‑sensitized to T‑cell‑mediated immunotherapy (64). As the
+PI3K/AKT pathway has emerged as a potential mechanism
+of resistance to immunotherapy and chemotherapy, multiple
+clinical trials have assessed inhibitors of the various pathway
+
+components.
+
+Alpelisib is an oral PI3K inhibitor that selectively inhibits
+p110 α . It showed efficacy in targeting *Pik3ca* ‑mutated breast
+cancer (65) and was FDA approved in 2019 in combination
+with fulvestrant for postmenopausal women and men, with
+HR [+], HER2‑negative, *Pik3ca* ‑mutated, advanced or metastatic
+breast cancer following progression on or after an endo­
+crine‑based regimen. For patients with advanced TNBC, the
+EPIK‑B3 Phase III trial (NCT04251533) is planned with study
+start date of April 2020 to assess alpelisib in combination with
+nab‑paclitaxel. Patients have *Pik3ca* mutations or PTEN loss
+with ≤1 prior line of therapy for metastatic disease.
+
+IPI‑549 is a selective PI3K‑gamma inhibitor targeting
+immune‑suppressive tumor‑associated myeloid cells. The
+MARIO‑3 Phase II study (NCT03961698) was designed to
+explore the addition of IPI‑549 to the FDA approved regimen
+
+
+atezolizumab/nab‑paclitaxel in front‑line TNBC. Cohort A
+will be composed of patients with locally advanced, metastatic
+TNBC, which will include two sub‑cohorts based on PD‑L1
+IHC status. The primary objective is CR rate.
+
+Ipatasertib and capivasertib are pan‑AKT inhibitors that
+bind to all three isoforms of AKT. Both are now in Phase III
+trials evaluating the efficacy of combination with paclitaxel
+as first‑line therapy for locally advanced or metastatic TNBC.
+In the LOTUS Phase II trial, adding ipatasertib to first‑line
+paclitaxel improved PFS, particularly in patients with
+PTEN/PI3K/AKT‑altered tumors (HR, 0.44) (66). In this
+subgroup of patients, median OS was 23.1 vs. 16.2 months with
+placebo (HR, 0.65) (67). To confirm the findings from LOTUS,
+the IPATunity130 Phase III trial (NCT03337724) is evaluating
+ipatasertib + paclitaxel for PTEN/PI3K/AKT‑altered advanced
+TNBC or HR [+], HER2‑negative breast cancers. The primary
+endpoint is PFS (68). An independent trial also supported the
+potential benefit for addition of AKT inhibitor to chemotherapy.
+In the PAKT Phase II study (NCT02423603), addition of the
+oral AKT inhibitor capivasertib to first‑line paclitaxel resulted
+in significantly longer PFS and OS in patients with advanced
+TNBC, especially in patients with PTEN/PI3K/AKT‑altered
+tumors. The median PFS duration was 5.9 vs. 4.2 months with
+placebo, meeting the predefined significance level, and better
+benefit in patients with PTEN/PI3K/AKT‑altered tumors with
+median PFS of 9.3 months (HR, 0.30). The median OS was
+prolonged by 6.5 months with capivasertib (69). The most
+common AEs of grade ≥3 were diarrhea, infection, rash, and
+fatigue, similar to those observed with ipatasertib in the LOTUS
+trial. The CAPItello‑290 Phase III trial (NCT03997123) is
+underway and the primary endpoints are PFS and OS (70).
+
+Efficacy of immunotherapy was also found to be
+enhanced by AKT inhibitors as a first‑line therapy for locally
+advanced/metastatic TNBC. Phase Ib study (NCT03800836)
+was designed to evaluate the triplet combination of ipatasertib
+(I), atezolizumab (A), and paclitaxel or nab‑paclitaxel (P).
+Preliminary efficacy and safety data up to January 5, 2019
+showed that the triplet regimen had promising antitumor
+
+
+-----
+
+1254 WON and SPRUCK: TNBC THERAPY: CURRENT AND FUTURE PERSPECTIVES
+
+
+activity (73% confirmed ORR), irrespective of biomarker
+PD‑L1 status or PTEN/PI3K/AKT alteration status, and
+manageable toxicity (71). In Phase III trial (NCT04177108),
+patients were enrolled in two cohorts according to PD‑L1
+status: Cohort 1 for PD‑L1‑negative tumors and cohort 2 for
+PD‑L1‑positive tumors. Three arms, P + I + A vs. P + I vs. P,
+will be evaluated in cohort 1 and 2 arms, P + I + A vs. P + A,
+
+will be evaluated in cohort 2.
+
+*CDK4/6/Rb/E2F pathway.* The G 1 ‑S phase checkpoint of the
+cell cycle is regulated by CDK4/6 activity which is controlled
+by their binding partners D‑type cyclins and p16 INK4 inhib­
+itor. The active CDK4/6‑cyclin D complex phosphorylates the
+retinoblastoma (Rb) protein, thereby activating E2F function
+and transition from G 1 to S phase of the cell cycle (72). The
+FDA approved CDK4/6 inhibitors palbociclib, ribociclib, and
+abemaciclib based on improvements in PFS for the treatment
+of ER [+], HER2‑negative advanced or metastatic breast cancer
+in combination with an endocrine therapy. TNBCs with a Rb [+],
+p16 INK4‑negative profile might represent the subpopulation
+of TNBC suitable for treatment with CDK4/6 inhibitors.
+
+Preclinical combination studies of CDK4/6 inhibitors with
+
+chemotherapy suggest that the timing and sequence of drug
+exposure/drug delivery schedule might play a critical role in
+drug activity, and the evaluation of different schedules of treat­
+ment may represent a new approach (73,74). The hypothesis was
+that reversible G 1 arrest of palbociclib could synchronize tumor
+cells in the cell cycle and following their re‑entry later would
+ensure a higher fraction in mitosis (M) phase when exposed
+to paclitaxel. In the first combination trial for palbociclib and
+paclitaxel (NCT01320592) an alternative dosing schedule was
+feasible and safe, without evidence of additive toxicity in Rb [+]
+breast cancer regardless of subtype (75). Phase I follow‑up
+trial (NCT02599363) of ribocilcib and weekly paclitaxel is in
+progress in patients with Rb [+] advanced breast cancer. In this
+study, pharmacodynamic, histologic, and imaging biomarkers
+will be utilized to confirm synchronization and schedule and
+identify a patient population that benefits from this treatment
+approach.
+
+The standard chemotherapy regimen causes treat­
+ment‑limiting cumulative myelosuppression that may
+compromise antitumor efficacy in TNBC. CDK4/6 inhibitors
+induce transient G 1 arrest in immune cells and hematopoietic
+stem and progenitor cells, potentially helping to preserve
+T‑cell function and bone marrow. To test this hypothesis, an
+investigational CDK4/6 inhibitor trilaciclib in combination
+with gemcitabine and carboplatin was explored to evaluate
+benefit for patients with ≤2 prior chemotherapy regimens in
+metastatic TNBC. Phase II trial (NCT02978716) was negative
+for a safety‑related primary endpoint (i.e. no difference in the
+frequency or duration of severe grade 4 neutropenia). However,
+the median OS was improved by more than 60%, which was
+likely due to increased chemotherapy duration and exposure.
+Trilaciclib‑treated patients also had a higher number of acti­
+vated CD8 [+]
+T cells over the first 5 cycles of chemotherapy,
+which potentially enhanced antitumor immunity (76).
+
+*MYC and CDK.* Transcription factor c‑MYC triggers selective
+gene expression to promote cell growth and proliferation. It is
+amplified in several different cancer types including TNBC,
+
+
+functioning as a proto‑oncogene (77). c‑MYC compensates for
+BRCA loss by upregulating HRR through increased RAD51
+expression (55,78). TNBC patients with high c‑MYC and
+RAD51 expression exhibit poor prognosis and less favorable
+response to chemotherapy and PARP inhibitors (55,57,79).
+c‑MYC blockade in TNBC was found to be synthetic lethal
+with PARP inhibitors, independent of BRCA status (80).
+c‑MYC pathway activation in TNBC is also synthetic
+lethal with CDK inhibition (81). Dinaciclib is a pan‑CDK
+(CDK1/2/5/9) inhibitor and the combination with PARP1
+inhibitor veliparib is currently being pursued in patients with
+advanced solid tumors for which no curative therapy exists
+(Phase I trial, NCT01434316). Dinaciclib induced immuno­
+genic cell death (ICD) but also increased expression of PD1
+on tumor‑infiltrating T cells and expression of PD‑L1 on
+tumor cells, thus limiting its antitumor effect in preclinical
+studies. However, dinaciclib inhibits tumor growth in combi­
+nation with anti‑PD‑1 (82). Phase Ib trial (NCT01676753)
+was designed to evaluate the efficacy of combined dinaciclib
+and pembrolizumab in patients with metastatic or locally
+advanced and unresectable TNBC. Its clinical benefit rate was
+47% in preliminary efficacy analysis and high c‑MYC expres­
+sion correlated significantly with clinical response, warranting
+further validation of c‑MYC as a predicative biomarker of
+response to CDK/checkpoint inhibitors (83).
+
+*AR antagonists.* The androgen receptor (AR) is an intracellular
+steroid receptor that dimerizes and translocates to the nucleus
+after binding androgen ligands. In the nucleus, AR binds to
+androgen response elements to promote target gene transcrip­
+tion in a tissue‑specific manner. AR can also be activated
+in a ligand‑independent manner through crosstalk with key
+signaling pathways, including PI3K/AKT and ERK (84). AR
+is involved in cell cycle regulation and the epithelial‑to‑mesen­
+chymal transition (EMT) (85,86). AR has emerged as a new
+biomarker and a potential therapeutic target in TNBC. AR is
+expressed in ≥40% of TNBCs and its expression level varies
+considerably among TNBC molecular subtypes. It has been
+associated with favorable prognosis, with better DFS and
+higher OS in the LAR subtype (87,88). However, patients
+with AR [+] TNBCs have a decreased chance of achieving pCR
+to neoadjuvant chemotherapy and the LAR subtype has been
+linked to poorer response to chemotherapy compared to other
+TNBC patients (89‑91). Multiple selective AR inhibitors have
+been approved by the FDA for the treatment of prostate cancer
+and are currently part of standard care (92). The role of the AR
+in signaling pathways in TNBC is still not clear and clinical
+studies are underway to provide more insight into the role of
+the AR as well as to assess whether AR targeting is a valuable
+therapeutic strategy in TNBC.
+
+The first proof‑of‑concept trial of AR‑targeted treatment
+established activity of the first‑generation AR antagonist
+bicalutamide in patients with advanced AR [+] TNBC. The
+TBCRC 011 Phase II trial (NCT00468715) showed a modest
+clinical benefit rate (CBR) of 19% at 6 months and a median
+PFS duration of 12 weeks (93).
+
+AR [+] TNBC expresses a luminal profile with intact Rb
+protein, the target of CDK4/6 activity. Thus, CDK4/6 inhibi­
+tors may increase the efficacy of AR antagonists in metastatic
+AR [+] TNBC. The single group Phase I/II trial (NCT02605486)
+
+
+-----
+
+INTERNATIONAL JOURNAL OF ONCOLOGY 57: 1245-1261, 2020 1255
+
+
+was carried out to explore this hypothesis. The combination
+of palbociclib and bicalutamide was well‑tolerated with no
+unexpected toxicity (94). It also met its prespecified efficacy
+endpoint as measured by PFS with 11 patients (31 evaluable
+patients) at 6 months (95).
+
+As one of the second‑generation anti‑androgen thera­
+pies, abiraterone is a steroidal CYP17 inhibitor with potent
+hydroxylase activity, targeting androgen biosynthesis. The
+French Breast Cancer Intergroup (UCBG) 12‑1 Phase II trial
+(NCT01842321) was designed to evaluate abiraterone acetate
+(AA) with its requisite concomitant medication prednisone
+in AR [+] advanced or metastatic TNBC. Androgen deprivation
+by AA resulted in 20% of the 6‑month CBR. This treatment
+appeared to be beneficial for some patients with molecular
+apocrine tumors, a subtype that expresses AR but not ER α (96).
+Considering that prednisone stimulates the glucocorticoid
+receptor (GR), which is expressed in approximately 25% of
+TNBCs, GR activity might limit the efficacy of AA.
+
+Seviteronel is an investigational lyase‑selective
+non‑steroidal CYP17 inhibitor that targets androgen and
+estrogen production. The CLARITY‑01 Phase I/II trial
+(NCT02580448) was set up to evaluate seviteronel in locally
+advanced or metastatic TNBC or ER [+] breast cancer. It revealed
+
+that seviteronel was generally well‑tolerated and provided
+clinical benefit. A total of 26 and 11% of patients reached at
+least a CBR at 4 and 6 months, respectively. Levels of circu­
+lating tumor cells (CTCs) also decreased (97,98).
+
+A second‑generation AR antagonist enzalutamide not only
+competitively binds to the AR ligand‑binding domain, but also
+inhibits nuclear translocation of AR, DNA binding, and coacti­
+vator recruitment. Phase II single arm study (NCT01889238)
+assessed the efficacy of enzalutamide in patients with locally
+advanced or metastatic, AR [+] TNBC. The primary endpoint
+was CBR at 16 weeks, which was 25% in the intention‑to‑treat
+(ITT) population and 33% in the evaluable subgroup whose
+tumors expressed ≥10% nuclear AR. The only treatment‑related
+grade 3 or greater AE occurring in ≥2% of patients was
+fatigue (3.4%) (99). The randomized ENDEAR Phase III study
+(NCT02929576) comparing enzalutamide and paclitaxel to
+placebo and paclitaxel in advanced TNBC was in place (100)
+but withdrawn in 2018, citing that further understanding about
+the role of androgen signaling in TNBC was required. The
+TBCRC 032 Phase Ib/II trial (NCT02457910) investigated the
+safety and efficacy of enzalutamide alone or in combination with
+PI3K inhibitor taselisib in patients with metastatic AR [+] TNBC.
+Primary endpoint of CBR at 16 weeks was 36% and median PFS
+was 3.4 months. The trial was not completed due to termination of
+the development of taselisib. Although this study was exploratory
+due to sample size limitation, it revealed subtype‑specific treat­
+ment response (favorable trend for luminal over non‑luminal) and
+identified novel *Fgfr2* gene fusions that likely activate the PI3K
+pathway and AR splice variants that may contribute to enzalu­
+tamide resistance. Therefore, an AR IHC score of ≥10% alone
+may not identify patients with AR‑dependent tumors, and LAR
+subtype and AR splice variants may help identify patients likely
+to benefit from AR antagonists (101).
+
+*Cell surface targets*
+*Tumor‑associated carbohydrate antigens.* The Globo H
+antigen is a hexasaccharyl sphingolipid expressed on the
+
+
+surface of various cancer types and has been explored as a
+potential target for vaccine therapy. Adagloxad simolenin (AS)
+is an immune stimulant comprising the Globo H hexasac­
+charide epitope linked to the carrier protein keyhole limpet
+hemocyanin (KLH). KLH facilitates a more vigorous immune
+response given the weak antigen, Globo H. As a first‑in‑class
+active immunotherapy in development for metastatic breast
+cancer, AS with the saponin‑based adjuvant OBI‑821 induced
+antibodies reactive with Globo H [+] tumor cells that mediate
+
+antibody‑dependent cell‑mediated cytotoxicity (ADCC) and
+complement‑dependent cytotoxicity (CDC) (102). Phase II
+trial (NCT01516307) assessed low‑dose cyclophosphamide
+with or without active immunotherapy (AS + adjuvant) in
+post‑treated metastatic breast cancer subjects with stable
+disease or response to treatment. Although it did not meet its
+primary efficacy endpoint of PFS, patients who developed an
+immune response to the vaccine showed significantly improved
+PFS and OS (103). Based on these subgroup data, Phase III
+study (NCT03562637) of AS with adjuvant vs. placebo treat­
+ment is in progress for high‑risk early‑stage TNBC patients
+following neoadjuvant or adjuvant chemotherapy. Patients will
+be screened for Globo H expression (IHC H‑score ≥15) and
+the primary objective is improvement of invasive disease‑free
+survival (IDFS) in the time frame of 5 years.
+
+*Antibody‑drug conjugates (ADCs).* An ADC is designed to
+be stable in plasma, target a tumor cell surface antigen with
+a high affinity and specificity, and is internalized, cleaved,
+and releases a payload drug which drives antitumor activity
+through direct cytotoxic cell killing and induces ICD.
+
+Sacituzumab govitecan‑hziy (SG) targets a glycoprotein,
+the human trophoblast cell‑surface antigen 2 (TROP‑2), that
+is expressed in more than 90% of TNBCs. Its payload is the
+active metabolite of irinotecan (SN‑38), which is conjugated
+to the anti‑TROP‑2 antibody by a cleavable linker. Phase I/II
+single group study (NCT01631552) included 108 patients with
+TNBC and 80% of patients had visceral metastases. The
+median number of prior regimens was 3 (range, 2‑10), which
+included chemotherapies and checkpoint inhibitors. Although
+it did not include biomarker selection of patients, 57 patients
+had moderate (2+) to strong (3+) and 5 patients had weak
+or absent TROP‑2 expression by IHC according to available
+data. The ORR was 33% and the median duration of response
+(DOR) was 7.7 months. The median PFS was 5.5 months and
+the median OS was 13.0 months. Myelotoxic effects were the
+main adverse reactions and grade 3 or 4 AEs included anemia
+and neutropenia (104). The confirmatory ASCENT Phase III
+study (NCT02574455) of SG in comparison with treatment
+of physician's choice for patients with metastatic TNBC
+was stopped due to compelling evidence of efficacy across
+multiple endpoints and SG was granted accelerated approval
+by the FDA based on the results of the IMMU‑132‑01 Phase II
+clinical trial for the treatment of adult patients with metastatic
+TNBC who have received ≥2 prior therapies for metastatic
+disease. It is the first ADC approved by the FDA specifically
+for relapsed or refractory metastatic TNBC as well as the first
+FDA‑approved anti‑TROP‑2 ADC.
+
+Ladiratuzumab vedotin (LV) targets LIV‑1, which is
+expressed in >90% of breast tumors with limited expression
+in normal tissues. LIV‑1 is a transmembrane protein with
+
+
+-----
+
+1256 WON and SPRUCK: TNBC THERAPY: CURRENT AND FUTURE PERSPECTIVES
+
+
+zinc transporter and metalloproteinase activity. The payload
+of LV is the microtubule disrupting agent monomethyl
+auristatin E (MMAE). Phase I study (NCT01969643) in
+patients with heavily pretreated metastatic TNBC showed
+25% ORR and medium PFS of 11 weeks. Treatment was
+
+generally well‑tolerated and related AEs were neutropenia,
+anemia, and neuropathy (105). LV was further explored
+in combination studies and in earlier lines of treatment.
+
+The SGNLVA‑002 Phase Ib/II trial (NCT03310957) was
+designed to assess whether combining LV and pembroli­
+zumab results in synergistic activity through LV‑induced
+ICD that creates a microenvironment favorable for
+enhanced anti‑PD‑L1 activity. It was for first‑line treatment
+of patients with unresectable locally advanced or metastatic
+TNBC. Initial dose‑finding studies revealed ORR of 35%
+with responses independent of PD‑L1 status and manage­
+able toxicity (106).
+
+ADC has also been explored for HER2‑low or nega­
+tive breast cancer. The rationale is based on the bystander
+effect, that is, the cleaved drug from an ADC may leak from
+the targeted tumor cell and affect cells in close proximity
+regardless of their target antigen expression status. Thus, an
+ADC having a high drug‑to‑antibody ratio and high‑potency
+payload would increase the killing of tumor cells even with
+low HER2 expression. Trastuzumab deruxtecan (T‑DXd)
+is the first HER2‑targeted agent to demonstrate promising
+clinical antitumor activity with a manageable safety profile
+in patients considered to be HER2‑negative. T‑DXd delivers
+a potent topoisomerase I inhibitor payload (an exatecan
+derivative) which is linked to a humanized anti‑HER2 anti­
+body. In Phase Ib (NCT02564900) trial of T‑DXd for heavily
+pretreated patients with advanced HER2‑low breast cancer,
+ORR was 37% with the median DOR being 10.4 months.
+Most toxicities were gastrointestinal or hematologic‑related,
+and interstitial lung disease (ILD) was an important
+identified risk (107). The DESTINY‑Breast04 Phase III
+(NCT03734029) was initiated to compare the efficacy and
+safety of T‑DXd to physician's choice (capecitabine, eribulin,
+gemcitabine, paclitaxel, or nab‑paclitaxel) in patients
+with HER2‑low, unresectable, and/or metastatic breast
+cancer (108).
+
+## **4. New potential therapeutic strategies**
+
+*Conversion of TNBC: Access to endocrine therapy.* Gene
+expression analysis and functional studies have revealed a high
+degree of plasticity and heterogeneity in luminal and basal‑like
+tumors. Expression of ER α, FOXA1 or GATA3 can result
+in transition from basal‑like breast cancer to luminal type
+whereas epigenetic reprogramming can result in a reverse tran­
+sition (109‑111). The CDK2‑EZH2 axis in tumors with TNBC
+phenotype (i.e *.* basal‑like breast cancer) has been explored for
+conversion to the ER α [+] subtype. Epigenetic enzyme EZH2, a
+histone‑lysine N‑methyltransferase that promotes histone H3
+lysine 27 mono‑, di‑ and tri‑methylation (H3K27me1/2/3),
+drives transcriptional repression (112,113). EZH2 can be
+phosphorylated at T416 (pT416‑EZH2) by cyclin E/CDK2 and
+>80% of TNBC patient specimens exhibit high pT416‑EZH2
+levels, which correlate with poorer survival (114). In preclinical
+studies, transgenic expression of a phospho‑mimicking mutant
+
+
+EZH2(T416D) in the mammary glands of mice reprogramed
+the committed luminal breast cancer cells into the basal‑like
+
+TNBC phenotype. In this setting inhibition of the CDK2‑EZH2
+axis by EZH2 inhibitors reactivated ER α expression and thus
+combination with tamoxifen suppressed tumor growth and
+improved the survival of mice bearing tumors with the TNBC
+phenotype (115). Therefore, inhibitors of CDK2 or EZH2
+combined with hormonal therapy may be a novel therapeutic
+strategy in TNBC with especially high pT416‑EZH2 levels.
+
+Another mechanism‑based therapy exploits the lack of
+ER expression due to hypermethylation of the ER α promoter.
+A combination epigenetic therapy of a DNA methyltrans­
+ferase (DNMT) inhibitor and a histone deacetylase (HDAC)
+inhibitor led to re‑expression of genes including ER α and
+restored tamoxifen sensitivity in ER‑negative breast cancer
+models (116,117). However, Phase II study (NCT01349959)
+in patients with advanced hormone‑resistant breast cancer
+or TNBC revealed that combination of DNMT inhibitor
+
+5‑azacitidine and HDAC inhibitor entinostat did not induce
+
+ER α expression and primary endpoint ORR was not met (118).
+ER α re‑expression induced by DNMT/HDAC inhibition might
+be attenuated by an active CDK2‑EZH2 axis, which affected
+outcomes in this study.
+
+The conversion of basal‑like breast cancer into ER α [+] is
+
+also under microenvironmental control. A paracrine signaling
+network involving platelet‑derived growth factor (PDGF)‑CC
+and PDGF receptor‑ α accelerated tumor growth through
+recruitment and activation of different subsets of cancer‑asso­
+ciated fibroblasts (119). In mouse models, impairing PDGF
+signaling was found to convert basal‑like breast cancers into
+ER α [+], and thus enhanced sensitivity to tamoxifen in previously
+resistant tumors (120). Therefore, PDGF inhibitors combined
+with endocrine therapy may be a novel therapeutic strategy in
+TNBC treatment.
+
+*Adaptive clinical studies: Molecular markers.* Under the
+master protocol framework, basket trials, where a targeted
+therapy is evaluated for multiple diseases that share common
+molecular alterations, and umbrella trials, where multiple
+targeted therapies are evaluated for a single disease that is
+stratified into multiple subgroups based on different molecular
+factors, have been developed (121). Recently there have been
+more adaptive, signal‑finding clinical trial designs coupled
+with correlative studies to investigate mechanisms of action.
+They also facilitate identifying active drug combinations as
+well as novel tumor indications. Patients are enrolled based on
+molecular markers from genetic profiling performed on their
+tumors. Some examples are listed below.
+
+In the OLAPCO Phase II trial (NCT02576444), PARP
+inhibitor olaparib was assessed in combination with various
+agents according to identified tumor mutations. It included
+AKT inhibitor capivasertib for tumors with mutations in
+the PI3K‑AKT pathway, WEE1 inhibitor adavosertib for
+tumors with *tp53* or/and *Kras* mutations, and ATR inhibitor
+ceralasertib for tumors with mutations in HRR genes. Primary
+outcome measure was ORR, and the trial also identified
+genetic determinants of response and resistance. Another
+Phase II trial (NCT03718091) evaluated ATR inhibitor M6620
+in selected solid tumors. Patients were enrolled in different
+
+cohorts based on tumor mutation status, including truncating
+
+
+-----
+
+INTERNATIONAL JOURNAL OF ONCOLOGY 57: 1245-1261, 2020 1257
+
+
+*Atm* mutations, germline *Brca* mutations, somatic *Brca* muta­
+tions or other HRR gene mutations, c‑MYC amplification,
+*Fbxw7* *Arid1a* muta­
+mutations, cyclin E amplification, and
+tions. Primary outcome measures included disease control rate
+(DCR) and changes in pCHK1 and γ H2AX levels. The I‑SPY
+2 Phase II trial (NCT01042379) was a neoadjuvant breast
+cancer trial using response‑adaptive randomization. It had
+multiple concurrent experimental arms with shared controls.
+Each biomarker signature was established at trial entry. A new
+regimen of combination with standard chemotherapy will be
+moved up to Phase III trial if it shows a high probability of
+improved pCR over standard chemotherapy.
+
+## **5. Conclusion**
+
+Developing novel treatments in both early and advanced
+TNBC settings remains a significant unmet need. Recent
+advances with novel agents have been made for specific
+subgroups with PD‑L1 [+] tumors or gBRCAm tumors. However,
+only a fraction of those patients respond to immune check­
+point or PARP inhibitors, and even those who do respond
+often develop resistance and relapse. In diverse tumor
+microenvironments, a given therapeutic agent shows vari­
+able responses, thus compromising the survival endpoints
+especially in an unselected TNBC population. Therefore,
+developing novel predictive biomarkers are crucial for
+selecting patients that will benefit the most from a given
+therapy. Single cell technologies will provide additional
+insight on tumor‑stroma interactions and facilitate compel­
+ling rationale for new treatments based on novel biomarkers.
+A non‑invasive testing of plasma circulating tumor DNA
+(ctDNA) and CTCs can potentially provide real‑time disease
+monitoring and even early therapy modification. However,
+their prognostic value needs further evaluation. With recent
+advances in multiomic analyses of cancers, there appears to
+be genomic and molecular similarities between TNBC and
+high‑grade serous ovarian carcinoma (HGSOC), suggesting
+that similar biological mechanisms drive some aspects of
+both cancer types. Therefore, treatment strategies for HGSOC
+can be explored in TNBC as well. The recent increase in the
+number of clinical trials investigating various new agents
+and combination strategies reflects further efforts to under­
+stand molecular and immunological aspects of TNBC. This
+may lead to more meaningful clinical benefits, including
+event‑free and overall survival.
+
+## **Acknowledgements**
+
+The authors would like to thank Professor Ian Collins of the
+
+Institute of Cancer Research, UK for valuable discussions on
+the DNA damage response pathways and checkpoint kinases.
+
+## **Funding**
+
+No funding was declared.
+
+## **Availability of data and materials**
+
+All information provided in this review is documented with
+relevant and current references.
+
+
+## **Authors' contributions**
+
+KAW was responsible for conceptualization, design, interpre­
+tation and visualization. KAW and CS were responsible for
+writing, reviewing and editing. Both authors approved the final
+manuscript.
+
+## **Ethics approval and consent to participate**
+
+Not applicable.
+
+## **Patient consent for publication**
+
+Not applicable.
+
+## **Competing interests**
+
+No competing interests are declared.
+
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+This work is licensed under a Creative Commons
+
+Attribution-NonCommercial-NoDerivatives 4.0
+
+International (CC BY-NC-ND 4.0) License.
+
+
+-----
+