PMCID string | Title string | Sentences string |
|---|---|---|
PMC9454178 | THZ1 targeting CDK7 suppresses c-KIT transcriptional activity in gastrointestinal stromal tumours | CDK4/6 inhibitors have gained FDA approval for the treatment of hormone receptor-positive breast cancer, and inhibitors targeting other cell cycle CDKs are currently in clinical trials for non-small cell lung cancer and other solid tumours . |
PMC9454178 | THZ1 targeting CDK7 suppresses c-KIT transcriptional activity in gastrointestinal stromal tumours | THZ1, a CDK7 inhibitor, exerts synergistic anticancer effects when combined with TKIs against neuroblastoma, glioma and non-small cell lung cancer [32, 37–39]. |
PMC9454178 | THZ1 targeting CDK7 suppresses c-KIT transcriptional activity in gastrointestinal stromal tumours | Therefore, targeting CDK7 may provide an alternative therapeutic option to block the reactivation of receptor tyrosine kinase pathways in RTK-driven neoplasms, especially for TKI-resistant cancer. |
PMC9454178 | THZ1 targeting CDK7 suppresses c-KIT transcriptional activity in gastrointestinal stromal tumours | Our research revealed that a combination of THZ1 and imatinib exerts synergistic antitumour effects in GIST imatinib-sensitive cells. |
PMC9454178 | THZ1 targeting CDK7 suppresses c-KIT transcriptional activity in gastrointestinal stromal tumours | To investigate the impact of CDK7 on c-KIT expression in GIST cells and the synergistic effect of THZ1 and imatinib, GIST-T1 and GIST-882 cells, which both harbour c-KIT gene mutations and are sensitive to imatinib, were used in our research. |
PMC9454178 | THZ1 targeting CDK7 suppresses c-KIT transcriptional activity in gastrointestinal stromal tumours | Both THZ1 and imatinib treatment led to c-KIT expression inhibition, and the combination treatment enhanced the inhibition of c-KIT expression and the downstream AKT and ERK signalling pathways. |
PMC9454178 | THZ1 targeting CDK7 suppresses c-KIT transcriptional activity in gastrointestinal stromal tumours | Taken together, our results indicate that THZ1 has effective antitumour activity against GISTs and may provide an additional therapeutic strategy for GIST patients with a poor response to imatinib. |
PMC9454178 | THZ1 targeting CDK7 suppresses c-KIT transcriptional activity in gastrointestinal stromal tumours | A superenhancer (SE) is a large cluster of genomic regulatory elements typically exhibiting an enrichment of histone H3 lysine 27 (H3K27ac) and densely bound by transcription factors and cofactors, playing critical roles in defining cell fate and identity . |
PMC9454178 | THZ1 targeting CDK7 suppresses c-KIT transcriptional activity in gastrointestinal stromal tumours | Interestingly, superenhancers frequently drive the expression of prominent oncogenes in cancer cells . |
PMC9454178 | THZ1 targeting CDK7 suppresses c-KIT transcriptional activity in gastrointestinal stromal tumours | Previous research has used H3K27ac chromatin immunoprecipitation with sequencing (ChIP-seq) of GIST tumour samples and cell lines and identified the SE clusters that drive c-KIT gene expression and are unique to GISTs . |
PMC9454178 | THZ1 targeting CDK7 suppresses c-KIT transcriptional activity in gastrointestinal stromal tumours | Subsequently, studies indicated that the SE domain was essential for c-KIT gene expression and tumorigenesis, including FOXF1, HAND1 and BARX1 . |
PMC9454178 | THZ1 targeting CDK7 suppresses c-KIT transcriptional activity in gastrointestinal stromal tumours | Disruption of the SE domain represents a therapeutic vulnerability in GIST . |
PMC9454178 | THZ1 targeting CDK7 suppresses c-KIT transcriptional activity in gastrointestinal stromal tumours | Among the genes screened out by ChIP-seq of GIST tumour samples and cell lines, OSR1 was hypothesized to bind to the c-KIT locus by ATAC sequencing . |
PMC9454178 | THZ1 targeting CDK7 suppresses c-KIT transcriptional activity in gastrointestinal stromal tumours | In our research, we found that CDK7 knockdown significantly inhibited the expression of c-KIT, and we identified the genes downregulated after THZ1 treatment by RNA-seq. |
PMC9454178 | THZ1 targeting CDK7 suppresses c-KIT transcriptional activity in gastrointestinal stromal tumours | Interestingly, we found that OSR1 was the predominantly downregulated gene. |
PMC9454178 | THZ1 targeting CDK7 suppresses c-KIT transcriptional activity in gastrointestinal stromal tumours | Therefore, we hypothesized that CDK7 knockdown inhibited the expression of c-KIT via OSR1. |
PMC9454178 | THZ1 targeting CDK7 suppresses c-KIT transcriptional activity in gastrointestinal stromal tumours | Subsequently, we proved that OSR1 expression was inhibited by THZ1 treatment in a dose-dependent manner and that OSR1 knockdown also significantly inhibited c-KIT expression. |
PMC9454178 | THZ1 targeting CDK7 suppresses c-KIT transcriptional activity in gastrointestinal stromal tumours | Moreover, OSR1 overexpression reversed the inhibition of c-KIT expression induced by CDK7 knockdown. |
PMC9454178 | THZ1 targeting CDK7 suppresses c-KIT transcriptional activity in gastrointestinal stromal tumours | In summary, our research may have revealed the role of OSR1 in c-KIT expression attenuated by CDK7 inhibition. |
PMC9454178 | THZ1 targeting CDK7 suppresses c-KIT transcriptional activity in gastrointestinal stromal tumours | In summary, our results uncovered the positive correlations between CDK7 and the malignant potential of GISTs and indicated that targeting CDK7 with the selective inhibitor THZ1 may be a promising treatment for GIST patients. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Novel treatments in gastrointestinal stromal tumors (GISTs) are essential due to imatinib resistance and the modest results obtained with multi-target tyrosine kinase inhibitors. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | We investigated the possibility that the hydroalcoholic extract from the leaves of Arbutus unedo L. (AUN) could harbor novel chemotherapeutics. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | The bio-guided fractionation of AUN led to a subfraction, FR2-A, that affected the viability of both imatinib-sensitive and -resistant GIST cells. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Cells treated with FR2-A were positive for Annexin V staining, a marker of apoptosis. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | A rapid PARP-1 downregulation was observed, although without the traditional caspase-dependent cleavage. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | The fractionation of FR2-A produced nine further active subfractions (FRs), indicating that different molecules contributed to the effect promoted by FR2-A. NMR analysis revealed that pyrogallol-bearing compounds, such as gallic acid, gallic acid hexoside, gallocatechin, myricetin hexoside, and trigalloyl-glucose, are the main components of active FRs. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Notably, FRs similarly impaired the viability of GIST cells and peripheral blood mononuclear cells (PBMCs), suggesting a non-specific mechanism of action. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Nevertheless, despite the lack of specificity, the established FRs showed promising chemotherapeutic properties to broadly affect the viability of GIST cells, including those that are imatinib-resistant, encouraging further studies to investigate whether pyrogallol-bearing compounds could represent an alternative avenue in GISTs. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Plants have been an inestimable source of anticancer compounds, providing approximately 50% of the approved chemotherapeutics . |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | In fact, natural compounds (NPs) show promising features, such as scaffold diversity and structural complexity, that make them ideal for drug discovery . |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Arbutus unedo L. (A. unedo) is a Mediterranean plant used in traditional medicine to treat various illnesses, such as gastrointestinal, dermatological, cardio-vascular, and urological disorders, kidney diseases, or also used as a diuretic and antidiabetic, suggesting the presence of numerous NPs with promising pharmacological activities . |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Regarding potential uses in cancer, studies have reported that NPs extracted from the entire plant, fruits, honey, and leaves could promote cytotoxicity in cellular tumor models. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | In particular, it has been recently reported that the leaf extract of A. unedo reduced the viability of U2OS, a cellular model of osteosarcoma, without toxicity in human umbilical vein endothelial (HUVEC) cells, a healthy cellular model . |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | However, further investigations are required to identify the NPs responsible for the observed anticancer activity. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | GISTs are rare mesenchymal neoplasms caused by a gain of function mutation in KIT or PDGFRα, two tyrosine kinase receptors (TKRs). |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | For this reason, they are treated with imatinib, a tyrosine kinase inhibitor (TKI) . |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Although imatinib has significantly improved patient prognosis, providing remarkable amelioration of their life expectations, the therapy is not conclusive. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Indeed, imatinib mainly promotes partial response or stable disease in most patients, while complete response is only observed in 5% of patients . |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Moreover, imatinib treatment is frequently associated with drug resistance, mostly due to acquired secondary mutations in TKRs (50–60% of patients) , fostering the development and approvals of novel multi-target TKIs as further treatment lines . |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Unfortunately, resistance has not been effectively treated with a strategy exclusively based on TKIs, which have provided a modest increase in progressive free survival (PFS). |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Preclinical studies have reported that imatinib resistance could be more multifaceted than initially hypothesized, supporting the research of novel compounds that could broadly target imatinib-resistant GIST cells . |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Traditional chemotherapy could potentially represent an alternative strategy for treating GISTs. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Indeed, the lack of cell specificity, a limitation of traditional chemotherapy, could be an advantage, potentially targeting both TKI-sensitive and -resistant subclones through a non-specific mechanism of action. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | However, the available chemotherapeutics, including those effective in other soft tissue sarcomas, such as doxorubicin, had already been tested in GISTs before imatinib approval, showing no effect . |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Thus, no standard chemotherapy has been approved in GIST so far . |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Therefore, considering that identifying novel chemotherapeutics could be promising for treating GISTs and the interesting anticancer properties of A. unedo already reported in the literature, we hypothesized that A. unedo could harbor NPs with chemotherapeutic properties. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Therefore, we observed that an extract obtained from A. unedo leaves affected the viability of GIST cell lines characterized by different mutations and degrees of imatinib response. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Furthermore, a bio-guided assay fractionation procedure was employed to investigate the different compounds’ contribution to the crude extract’s activity. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | The pharmacological effect of the hydroalcoholic crude extract from A. unedo leaves (hereafter referred to as AUN only) was first assessed in the KIT-mutated imatinib-sensitive cell lines GIST-882 and GIST-T1. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | In detail, cells were treated with three serial 1:2 dilutions of AUN starting from 200 µg/mL for 72 h. AUN treatment significantly decreased the viability in GIST-882 (Figure 1a) and GIST-T1 (Figure 1b) with respect to the control. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | In GIST-882, a significant 75% and ~25% reduction was observed at 200 µg/mL and 100 µg/mL concentrations, while no effect was observed at 50 µg/mL. Similarly, cell viability was significantly impaired by about 75% at 200 µg/mL in GIST-T1, while a non-significant reduction was observed after 100 and 50 µg/mL treatments. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Considering the doubling time of GIST-882 and GIST-T1, experimentally calculated as approximately 48–72 h, the effects observed following AUN treatment could be due to both the inhibition of cell proliferation and cell death. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Interestingly, in the first gross investigation by flow cytometry, a further cell population with a lower cell size index and a slight increase in viability staining was observed in AUN-treated GIST-882 compared to the control (Supplementary Figure S1). |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Additionally, this population was characterized by reduced nucleus staining, supporting the hypothesis that these cells contain degraded chromosomal DNA. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | The two features—small size and degraded DNA—suggested that AUN could stimulate cell death by triggering apoptosis. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | This hypothesis was confirmed using the Annexin-V/7-AAD assay in both cell lines. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | In particular, in GIST-882, 24 h of AUN treatment promoted a significant increase in early (38%, p < 0.0001) and late (41%, p < 0.0001) apoptotic cells compared to the control (early 21% and late 13%, respectively; Figure 2—upper panel). |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Analogously, although with faster kinetics, an increase in early and late apoptotic cells was also observed in GIST-T1 after 6 h of treatment (Figure 2—lower panel). |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | In detail, we observed a significant increase in both early and late apoptosis compared to the control (31% vs. 15%; p < 0.0001 and 15% vs. 3%; p < 0.001, respectively). |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | AUN was first fractionated using a liquid/liquid partition to preliminary separate the compounds based on their polarity. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Hence, three fractions were obtained from AUN: FR1, which contained phytochemicals extracted by chloroform (yield about 0.33% w/w); FR2, which collected those that are instead extracted by ethyl acetate (yield about 8.8% w/w); and FR3, retaining the more hydrophilic phytochemicals in water. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Each fraction was then tested in GIST-882 and compared with AUN to identify the fraction where active phytochemicals were mainly distributed. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | As shown in Figure 3a, GIST-882 viability was impaired primarily by FR2, promoting a significant reduction of about 90% (p < 0.0001), which was even more intense than AUN. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Approximately a 30% viability reduction was observed in the FR3-treated sample (p < 0.05), while FR1 promoted no effect. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Overall, the data indicated that following liquid/liquid partition, active phytochemicals were mainly collected and enriched in FR2 with respect to FR1 and FR3. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Furthermore, the percentage of early apoptotic cells observed through the Annexin-V/7-AAD assay corroborated that the active proapoptotic phytochemicals were principally distributed in FR2 (Figure 3b). |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Similar results were observed in GIST-T1 cells, and in particular, cell viability was significantly reduced by 75% (p < 0.001) after 24 h of FR2 treatment (Supplementary Figure S2). |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Altogether, these data strengthen the fact that the active phytochemicals were extracted into FR2, and they can similarly target imatinib-sensitive cellular models characterized by different primary KIT mutations. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | The above data led us to consider FR2 as the fraction harboring the phytochemicals of interest and the starting material for a second step of bio-guided fractionation. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | In particular, it was subjected to reverse-phase medium-pressure liquid chromatography (MPLC), resulting in four FR2 subfractions (FR2-A, FR2-B, FR2-C, and FR2-D; Supplementary Figure S3) generated based on the chromatogram. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Afterwards, the activity of the four subfractions was tested in GIST-882, with cell viability evaluated after 24 h of treatment with two different concentrations (12.5 µg/mL and 50 µg/mL) compared to the control (Figure 4). |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | In total, 50 µg/mL of FR2-A significantly impaired cell viability (>95%; p < 0.0001), similarly to that observed after treatment with 200 µg/mL of FR2. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Moreover, at the same concentration, FR2-B and FR2-C promoted a significant 40% reduction in cell viability (p < 0.001). |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | At the lowest concentration (12.5 µg/mL), a significant reduction (p < 0.01) was observed after FR2-A treatment only; indeed, no effect of FR2 and the other FR2 subfractions was observed. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Overall, this result led us to identify FR2-A as the most active among the FR2 subfractions. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Consistent with this hypothesis, FR2-A also affected GIST-T1 viability, and the calculated IC50 was around 33 µg/mL in both cellular models (Supplementary Figure S4). |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Consequently, FR2-A was further investigated regarding its effect on imatinib-resistant cells and its mechanism of action. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | To further investigate FR2-A activity, GIST-48 and GIST-48b, recognized as imatinib-resistant cells , were treated with the FR2-A subfraction. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Notably, cell viability was impaired in both models, even at a higher IC50 compared to GIST-882 and GIST-T1 (Figure 5a,b). |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Interestingly, GIST-48b cells, unlike GIST-882, -T1, and -48 cells, are characterized by no detectable level of the oncoprotein KIT (Figure 5c), suggesting that the mechanism of action of FR2-A was KIT-independent and could provide an alternative treatment to TKIs. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | To further deepen the FR2-A mechanism of action, GIST-882 cells were treated in a time-course experiment of 48 h, with 33 and 66 µg/mL FR2-A as well as with the gold standard imatinib (Figure 6a). |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | In particular, 66 µg/mL of FR2-A reduces viability by 50% about 4 h after treatment, while 33 µg/mL of FR2-A requires about 6 h. In addition, FR2-A impaired almost entirely cell viability at 66 µg/mL, whereas the 33 µg/mL treatment did not reduce the viability by more than 50%. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Thus, it could be hypothesized that FR2-A affects GIST-882 viability in a dose- and time-dependent manner. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Regarding imatinib treatment, cell viability was significantly affected (p < 0.05), but only after 48 h, which may be due to reduced proliferation rather than reduced cell viability. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | In line with these observations, Annexin-V/7-AAD staining confirmed that imatinib promoted a slight increase (23.4% vs. 15.6%; p < 0.05) in early apoptotic cells compared to the control at 48 h (Figure 6b,c). |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | On the contrary, both concentrations of FR2-A gave rise to a remarkably higher increase in early apoptotic cells compared to the control, as previously shown with AUN and FR2. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | A total of 66 µg/mL of FR2-A led to a gradual and significant increase in the early apoptotic population starting from 2 h after treatment (p < 0.01), leading to a percentage of approximately 50–60% 48 h after treatment (p < 0.001). |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | The lower concentration, 33 µg/mL, required about 6 h to show a significant increase in early apoptotic cells, inducing the increase to about 40% at 48 h (Figure 6b,c). |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Considering that GIST-882 viability was quickly impaired 2 h after treatment, we focused on the mechanism of action of FR2-A at early time points. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Specifically, the activation of traditional apoptosis pathways based on the cleavage of caspase-3 or caspase-9 was monitored. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Although FR2-A promoted the rise of an Annexin V-positive and 7-AAD-negative population, commonly defined as early marks of apoptosis, no caspase activation was observed. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Failure to detect the cleavage of PARP-1, a well-recognized downstream target of caspase-3, confirmed the absence of traditional caspase-3 activation. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Nevertheless, treatment with 66 µg/mL of FR2-A determined a time-dependent downregulation of PARP-1. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | In detail, the level of PARP-1 was significantly reduced by about 50% after 2 h (p < 0.01), and even by about 85% after 3 h (p < 0.001), supporting the hypothesis that, at least in the late stage, the pharmacological effect could be strictly related to PARP-1 downregulation (Figure 7). |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Since FR2-A was associated with promising results, notably impairing the viability of GIST cells independently from KIT expression and mutational patterns, its chemical composition was analyzed by H NMR to identify the active compounds. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Intense spectral signals reveal that the most represented compound in FR2-A was arbutin in its β anomeric form (Supplementary Figure S5a). |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Therefore, GIST-882 cells were treated with pure β-arbutin at 200 µg/mL. However, no significant effect was detected on GIST-882 cell viability and apoptosis up to 72 h after treatment, indicating that β-arbutin was not the compound responsible for FR2-A activity (Supplementary Figure S5b). |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | In view of this result, FR2-A was further fractionated using size-exclusion chromatography (SEC), yielding 61 analyzable subfractions. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Among them, nine maintained the pharmacological activity in GIST-882 and GIST-T1, suggesting that more than one phytochemical was responsible for the FR2-A effect. |
PMC11085211 | Arbutus unedo L. Fractions Exhibit Chemotherapeutic Properties for the Treatment of Gastrointestinal Stromal Tumors | Specifically, 30 µg/mL of 2A-29, 2A-30, 2A-31, 2A-35, 2A-36, 2A-60, and 2A-61 significantly reduced the viability of both cell lines up to the limit of assay detection (p < 0.0001; Figure 8a); the same dose of 2A-48 and 2A-49 provided a significant viability reduction of no more than 60% (p < 0.0001). |
Subsets and Splits
No community queries yet
The top public SQL queries from the community will appear here once available.