Sensitivity Of Cancer Cells Research Articles

Development of Natural-Product-Inspired ABCB1 Inhibitors Through Regioselective Tryptophan C3-Benzylation.

The emergence of drug resistance in cancer cells eventually causing relapse is a serious threat that demands new advances. Upregulation of the ATP-dependent binding cassette (ABC) transporters, such as ABCB1, significantly contributes to the emergence of drug resistance in cancer. Despite more than 30 years of therapeutic discovery, and several generations of inhibitors against P-gp, the search for effective agents that minimize toxicity to human cells, while maintaining efflux pump inhibition is still underway. Leads derived from natural product scaffolds are well-known to be effective in various therapeutic approaches. Inspired by the biosynthetic pathway to Nocardioazine A, a marine alkaloid known to inhibit the P-gp efflux pump in cancer cells, we devised a regioselective pathway to create structurally unique indole-C3-benzyl cyclo-L-Trp-L-Trp diketopiperazines (DKPs). Using bat cells as a model to derive effective ABCB1 inhibitors for targeting human P-gp efflux pumps, we have recently identified exo-C3-N-Dbn-Trp2 (13) as a lead ABCB1 inhibitor. This C3-benzylated lead inhibited ABCB1 better than Verapamil.[21] Additionally, C3-N-Dbn-Trp2 restored chemotherapy sensitivity in drug-resistant human cancer cells and had no adverse effect on cell proliferation in cell cultures. For a clearer structure-activity relationship, we developed a broader screen to test C3-functionalized pyrroloindolines as ABCB1 inhibitors and observed that C3-benzylation is outperforming respective isoprenylated derivatives. Results arising from the molecular docking studies indicate that the interactions at the access tunnel between ABCB1 and the inhibitor result in a powerful predictor for the efficacy of the inhibitor. Based on fluorescence-based assays, we conclude that the most efficacious inhibitor is the p-cyano-derived exo-C3-N-Dbn-Trp2 (33 a), closely followed by the p-nitro substituted analogue. By combining assay results with molecular docking studies, we further correlate that the predictions based on the inhibitor interactions at the access tunnel provide clues about the design of improved ABCB1 inhibitors. As it has been well documented that ABCB1 itself is powerfully engaged in multi-drug resistance, this work lays the foundation for the design of a new class of inhibitors based on the endogenous amino acid-derived cyclo-L-Trp-L-Trp DKP scaffold.

Extracellular vesicles in cancer drug resistance: Mechanistic insights and therapeutic implications

AbstractDespite significant advancements in the treatment of cancer, therapeutic resistance remains a major hurdle for the achievement of full cures. Besides being typically driven by intratumoral heterogeneity, such acquired resistance also relies heavily on cell‐cell communication whereby extracellular vesicles (EVs) carrying resistance‐related components can be transferred from drug‐resistant cells or nontumorigenic members within tumor microenvironment (TME) to their sensitive neighbors. The cargo and membrane surface proteins of EVs derived from drug‐resistant cells and TME cells carry abundant biological information, transferring therapy‐resistant capacity to sensitive cancer cells. Hence, a deep understanding of the roles of EVs in cancer therapy resistance will facilitate identifying biomarkers and developing new approaches to restore therapy sensitivity. In this review, we summarize our current understanding regarding the causes and effects of EV‐mediated cell–cell communication in drug resistance, with a particular notice on how various kinds of cargoes derived from different cells within TME are linked to the resistant phenotype. We also discuss how this knowledge can contribute to improvements in clinical practice, that is, the opportunities and challenges of EVs in functioning as potential biomarkers in predicting therapeutic resistance and, by extension, EV‐based therapy in achieving deeper and longer‐lasting clinical responses.

Pericytes Modulate Third-Generation Tyrosine Kinase Inhibitor Sensitivity in EGFR-Mutated Lung Cancer Cells Through IL32-β5-Integrin Paracrine Signaling.

EGFR-mutated lung cancer patients sometimes display restricted responses to third-generation tyrosine kinase inhibitors (TKIs), potentially attributable to undervalued input from stromal cells, notably pericytes (PCs). The study shows that PCs isolated from EGFR-mutated patients have a unique secretome profile, notably secreting IL32 and affecting signaling pathways and biological processes linked to TKI sensitivity. Clinical evidence, supported by single-cell RNA sequencing and multiplex immunostaining of tumor tissues, confirms the presence of IL32-expressing pericytes closely interacting with β5-integrin-expressing cancer cells in EGFR-mutated patients, impacting therapeutic response and prognosis. Co-culture and conditioned medium experiments demonstrate that PCs reduce TKI effectiveness in EGFR-mutated cancer cells, a reversible phenomenon through silencing IL32 expression in PCs or depleting the IL32 receptor β5-integrin on cancer cells, thereby restoring cancer cell sensitivity. Mechanistically, it is shown that YY1 signaling upregulates IL32 secretion in PCs, subsequently activating the β5-integrin-Src-Akt pathway in EGFR-mutated cancer cells, contributing to their TKI sensitivity. In animal studies, co-injection of cancer cells with PCs compromises TKI effectiveness, independently of blood vessel functions, while inhibition of β5-integrin restores tumor cell sensitivity. Overall, the findings highlight direct crosstalk between cancer cells and pericytes, impacting TKI sensitivity via IL32-β5-integrin paracrine signaling, proposing an enhanced therapeutic approach for EGFR-mutated patients.

SiRNA-based strategies to combat drug resistance in gastric cancer.

Chemotherapy is a key treatment option for gastric cancer, but over 50% of patients develop either inherent or acquired resistance to these drugs, resulting in a 5-year survival rate of only about 20%. The primary treatment for advanced gastric cancer typically involves chemotherapy based on platinum or fluorouracil. Several factors can contribute to platinum resistance, including decreased drug uptake, increased drug efflux or metabolism, enhanced DNA repair, activation of pro-survival pathways, and inhibition of pro-apoptotic pathways. In recent years, there has been significant progress in biology aimed at finding innovative and more effective methods to overcome chemotherapy resistance. Small interfering RNAs (siRNAs) have emerged as a significant advancement in gene expression regulation, showing promise in enhancing the sensitivity of gastric cancer cells to chemotherapy drugs. However, siRNA therapies still face major challenges, particularly in terms of stability and efficient delivery in vivo. This article discusses the advances in siRNA therapy and its potential role in overcoming resistance to chemotherapeutic drugs such as cisplatin, 5-FU, doxorubicin, and paclitaxel in the treatment of gastric cancer.

LncRNA MAGI2-AS3 enhances cisplatin sensitivity of non-small cell lung cancer cells by regulating the miR-1269a/PTEN/AKT pathway

To investigate the mechanism mediating the regulatory effect of lncRNA MAGI2-AS3 on cisplatin (DDP) resistance in non-small cell lung cancer (NSCLC). MAGI2-AS3 and miR-1269a expression levels were detected by qRT-PCR in DDP-sensitive lung cancer cell lines (A549 and H1299) and their resistant counterparts (A549/DDP and H1299/DDP). In A549 and H1299 cells with MAGI2-AS3 silencing and A549/DDP and H1299/DDP cells overexpressing MAGI2-AS3, the effects of 20 μmol/L DDP on cell viability and apoptosis were examined with CCK-8 assay, colony formation assay, flow cytometry and Western blotting, and the changes in epithelial-mesenchymal transition (EMT) were assessed with wound healing and Transwell assays. The interaction between MAGI2-AS3, miR-1269a and PTEN was predicted using GEPIA, StarBase and miRDB and verified with luciferase reporter gene assay and radioimmunoprecipitation (RIP) assay. A miR-1269a mimic and pcDNA3.1-PTEN plasmid were used to perform the rescue assay. MAGI2-AS3 expression was significantly downregulated in lung cancer tissues (P < 0.05) in association with a poor prognosis (P < 0.05). In the two DDP-resistant lung cancer cell lines, MAGI2-AS3 expression was significantly lowered as compared with the sensitive cells. Silencing MAGI2-AS3 significantly enhanced cell viability and promoted EMT of A549 and H1299 cells irrespective of DDP treatment, and also decreased DDP-induced apoptosis of the cells. In A549/DDP and H1299/DDP cells, MAGI2-AS3 overexpression strongly repressed cell viability and EMT irrespective of DDP treatment and promoted DDP-induced cell apoptosis. Luciferase reporter gene and RIP assays confirmed the binding of MAGI2-AS3 with miR-1269a and the binding of miR-1269a with 3 '-UTR domain of PTEN. The rescue assay demonstrated that MAGI2-AS3 acted as a sponge for miR-1269a to promote PTEN expression and downregulate AKT phosphorylation, thus inhibiting EMT and promoting DDP-induced apoptosis of A549/DDP cells. MAGI2-AS3 enhances DDP sensitivity of NSCLC by targeted regulation of the miR-1269a/PTEN/AKT signaling axis.

Scaffolds functionalized with matrix metalloproteinase-responsive release of miRNA for synergistic magnetic hyperthermia and sensitizing chemotherapy of drug-tolerant breast cancer

Combining hyperthermia and chemotherapy for maximum anticancer efficacy remains a challenge because drug-tolerant cancer cells often evade this synergistic treatment due to drug resistance and asynchronous drug release. In this study, multifunctional scaffolds were designed to efficiently treat drug-tolerant breast cancer by improving the sensitization of breast cancer cells and synchronizing anticancer drug release with magnetic hyperthermia. The scaffolds contained microRNA-encapsulated matrix metalloproteinase-cleavable liposomes, doxorubicin-encapsulated thermoresponsive liposomes and Fe3O4 nanoparticles. The scaffolds could release microRNA specifically to improve the sensitization of breast cancer cells to anticancer drugs. The scaffolds also showed excellent hyperthermia effects under alternating magnetic field irradiation. Moreover, doxorubicin release was synchronized with magnetic hyperthermia. In vitro and in vivo studies demonstrated that the scaffolds effectively reduced drug resistance and eliminated doxorubicin-tolerant MDA-MB-231 cells through the synergistic effect of magnetic hyperthermia and sensitizing chemotherapy. Additionally, the scaffolds could support the proliferation and adipogenic differentiation of stem cells for adipose tissue regeneration after killing cancer cells at a late therapeutic stage. These composite scaffolds offer an innovative strategy for treating breast cancer, with synergistic anticancer effects and regenerative functions.

Co-delivery of Doxorubicin and VEGF siRNA using carcinoembryonic antigen-targeted chitosan nanoparticles for colorectal cancer

Developing a targeted nanocarrier-based therapeutic approach is highly effective in delivering therapeutics to specific cancer cells. Combining two or more therapeutic approaches with different mechanisms of action can effectively maintain their unique benefits while considerably improving cancer treatment. Doxorubicin (DOX) binds to DNA by intercalation and triggers a cascade of biochemical processes in tumor cells leading to death. Anti-VEGF medicines may sensitize cancer cells to chemotherapeutics by limiting blood supply and increasing penetration by disrupting tumor vasculature. In this study, DOX and vascular endothelial growth factor siRNA (VEGF siRNA) loaded chitosan nanoparticles (ChNPs) were prepared by using the ionic gelation method. The DOX-siRNA-ChNPs surface was then functionalized with carcinoembryonic antigen (CEA) targeted monoclonal antibody (DOX-siRNA-ChNPs-CMAb), thereby delivering therapeutic moieties into the cancer milieu. The cytotoxicity and cellular uptake of the developed nanocarrier were tested in-vitro on HT-29, which revealed that DOX-siRNA-ChNPs-CMAb possessed the highest cytotoxicity when compared with other prepared formulations and free DOX. The cellular uptake using fluorescence microscopy displayed a significantly higher uptake of DOX-siRNA-ChNPs-CMAb (targeted) than free DOX and DOX-siRNA-ChNPs (non-targeted). Apoptosis analysis in flow cytometry using annexin-V/PI staining demonstrated a substantially higher apoptosis rate of HT-29 cells using DOX-siRNA-ChNPs-CMAb than DOX-siRNA-ChNPs. Moreover, DOX-siRNA-ChNPs-CMAb were studied in CRC induced animal models, showing that DOX-siRNA-ChNPs-CMAb specifically targets CRC cells without showing organ toxicity. Thus, the novel approach of simultaneous delivery of DOX and siRNA using CMAb conjugated ChNPs demonstrated significant targeting in both in-vitro and in-vivo with minimal organ toxicity.

Extracellular vesicles from human breast cancer-resistant cells promote acquired drug resistance and pro-inflammatory macrophage response.

Breast cancer is a significant public health problem around the world, ranking first in deaths due to cancer in females. The therapy to fight breast cancer involves different methods, including conventional chemotherapy. However, the acquired resistance that tumors develop during the treatment is still a central cause of cancer-associated deaths. One mechanism that induces drug resistance is cell communication via extracellular vesicles (EVs), which can carry efflux transporters and miRNA that increase sensitive cells' survivability to chemotherapy. Our study investigates the transcription changes modulated by EVs from tamoxifen- and doxorubicin-resistant breast cancer cells in sensitive cells and how these changes may induce acquired drug resistance, inhibit apoptosis, and increase survivability in the sensitive cells. Additionally, we exposed human macrophages to resistant EVs to understand the influence of EVs on immune responses. Our results suggest that the acquired drug resistance is associated with the ability of resistant EVs to upregulate several transporter classes, which are directly related to the increase of cell viability and survival of sensitive cells exposed to EVs before a low-dose drug treatment. In addition, we show evidence that resistant EVs may downregulate immune system factors to evade detection and block cell death by apoptosis in sensitive breast cancer cells. Our data also reveals that human macrophages in contact with resistant EVs trigger a pro-inflammatory cytokine secretion profile, an effect that may be helpful for future immunotherapy studies. These findings are the first transcriptome-wide analysis of cells exposed to resistant EVs, supporting that resistant EVs are associated with the acquired drug resistance process during chemotherapy by modulating different aspects of sensitive cancer cells that coffer the chemoresistance.

RON receptor tyrosine kinase regulates glycolysis through MAPK/CREB signaling to affect ferroptosis and chemotherapy sensitivity of thyroid cancer cells.

Anaplastic thyroid cancer (ATC) is one of the deadliest and most aggressive human malignancies for which there is currently no effective treatment. Tyrosine kinase receptor RON is highly expressed in various cancer types, including colon, pancreatic and thyroid cancer. However, its underlying role in ATC is not fully understood. The present study investigated the therapeutic potential and molecular mechanism of RON in ATC. RON expression in thyroid cancer cells was detected by western blotting. Glycolysis was assessed by measuring the extracellular acidification rate, glucose uptake, lactate concentration, and expression levels of glucose transporter 1, hexokinase 2 and pyruvate kinase M1/2. In addition, ferroptosis was assessed by detecting the levels of total iron, lipid peroxide and reactive oxygen species, and the expression levels of ferroptosis‑related proteins. Furthermore, mitochondrial function were assessed by JC‑1 staining and detection kits, respectively. The results demonstrated that RON was highly expressed in thyroid cancer cell lines. Furthermore, RON interference inhibited glycolysis, promoted ferroptosis, elevated cell sensitivity to chemotherapy and affected mitochondrial function in thyroid cancer cells. Further experiments demonstrated that RON interference affected the ferroptosis levels in thyroid cancer cells by inhibiting the glycolysis process. Mechanistically, the present results indicated that RON may affect ferroptosis, glycolysis and chemotherapy sensitivity by regulating MAPK/cAMP‑response element binding protein (CREB) signaling in thyroid cancer cells. In conclusion, the present study demonstrated that RON affected ferroptosis, glycolysis and chemotherapy sensitivity in thyroid cancer cells by regulating MAPK/CREB signaling, demonstrating its potential as a therapeutic target in thyroid cancer cells.

Chromatin accessibility is associated with therapeutic response in prostate cancer.

Treatment of advanced prostate cancer is challenging due to a lack of effective therapies. Therefore, it is important to understand the molecular mechanisms underlying therapeutic resistance in prostate cancer and to identify promising drug targets offering significant clinical advantages. Given the pivotal role of dysregulated transcriptional programs in the therapeutic response, it is essential to prioritize translational efforts targeting cancer-associated transcription factors (TFs). The present study investigated whether chromatin accessibility was associated with therapeutic resistance in prostate cancer using Assay for Transposase-Accessible Chromatin with sequencing (ATAC-seq) data. The bioinformatics analysis identified differences in chromatin accessibility between the drug response (Remission) and drug resistance (Disease) groups. Additionally, a significant association was observed between chromatin accessibility, transcriptional output and TF activity. Among TFs, forkhead box protein M1 (FOXM1) was identified as a TF with high activity and expression in the Disease group. Notably, the results of the computational analysis were validated by FOXM1 knockdown experiments, which resulted in suppressed cell proliferation and enhanced therapeutic sensitivity in prostate cancer cells. The present findings demonstrated that chromatin accessibility and TF activity may be associated with therapeutic resistance in prostate cancer. Additionally, these results provide the basis for future investigations aimed at understanding the molecular mechanisms of drug resistance and developing novel therapeutic approaches for prostate cancer.

The Tumor Suppressor TPD52-Governed Endoplasmic Reticulum Stress is Modulated by APCCdc20.

Aberrant regulation of unfolded protein response (UPR)/endoplasmic reticulum (ER) stress pathway is associated with cancer development, metastasis, and relapse, and the UPR signal transducer ATF6 has been proposed as a diagnostic and prognostic marker for many cancers. However, a causal molecular link between ATF6 activation and carcinogenesis is not established. Here, it is found that tumor protein D52 (TPD52) integrates ER stress and UPR signaling with the chaperone machinery by promoting S2P-mediated cleavage of ATF6. Although TPD52 has been generally considered as an oncogene, TPD52 is identified as a novel tumor suppressor in bladder cancer. Significantly, attenuation of the ER stress via depletion of TPD52 facilitated tumorigenesis in a subset of human carcinomas. Furthermore, the APCCdc20 E3 ligase is validated as the upstream regulator marking TPD52 for polyubiquitination-mediated proteolysis. In addition, inactivation of Cdc20 sensitized cancer cells to treatment with the ER stress inducer in a TPD52-dependent manner. Thus, the study suggests that TPD52 is a novel Cdc20 substrate that may modulate ER stress to prevent tumorigenesis.

APX2009 sensitizes hypoxic breast cancer cells to doxorubicin by increasing its accumulation and caspase-3/7-mediated apoptosis.

The association of targeted therapy with chemotherapy is encouraged to increase the treatment efficiency, especially in hypoxic triple-negative breast cancer. The APE1 redox activity has stood out as a potential tumor target. However, the effect of the association of the APE1 redox inhibitors with doxorubicin in hypoxia still needs to be evidenced. Therefore, our objective was to investigate the effect of the APX2009 (APE1 inhibitor) on the sensitization of breast cancer cells to doxorubicin in normoxia and hypoxia. The WST-1 assay was used to evaluate cell viability after APX2009 and doxorubicin application under normoxia and hypoxia conditions in the MCF-7 and MDA-MB-231 cells. Apoptosis was analyzed by annexin assay and detection of caspases-3/7 activity by luminescence-based assay. The clinical association between APE1 inhibition signature and doxorubicin sensitivity was evaluated by bioinformatics analyses. MDA-MB-231 and MCF-7 cell lines were more sensitive to APX2009 in normoxia than in hypoxia. Co-treatment with APX2009 and doxorubicin in hypoxia further decreased the viability of triple-negative MDA-MB-231 cells than treatment alone, which was accompanied by doxorubicin intracellular accumulation, and increase of apoptotic cells percentage, and caspases-3/7 activity. Moderate association was found between APE1 inhibition signature and doxorubicin sensitivity in the hypoxic basal subtype. The findings suggest that APX2009 sensitizes the MDA-MB-231 cells to doxorubicin in hypoxia by doxorubicin intracellular accumulation and caspases-3/7-mediated apoptosis.

Downregulation of circSTX6 suppresses tumor progression while facilitating radiosensitivity in cervical squamous cell carcinoma

Backgroundcervical squamous cell carcinoma (CSCC) is the second gynecological tumors that seriously threaten women's life quality. Circular RNA (circRNA) is related with cervical cancer carcinogenesis and radiosensitivity. AimTo investigate the performance of hsa_circ_0007905 (circSTX6) on regulating cellular activities and radiosensitivity in CSCC. MethodsThe relative expression of circSTX6 in different tissue samples was detected by RT-qPCR. The cellular activity influence of circSTX6 in cervical cancer cells was measured by CCK-8 and Transwell assays. The survival fractions of cancer cells were detected after the radiation treatment to explore the relationship between circSTX6 and radiosensitivity of cervical cancer. The downstream miRNAs were predicted and analyzed. Rescue experiments confirmed their targeting relationship. Bioinformatic analysis was performed to identify the potential targets of miR-203a-3p. ResultscircSTX6 was increased and miR-203a-3p was decreased in cervical cancer tissues and radio-resistant tissues. CircSTX6 expression was related to the patient's survival rates. CircSTX6 absence decreased cervical cancer cell proliferation and invasion while enhancing the sensitivity of cervical cancer cells to radiotherapy by regulating miR-203a-3p. RAB27B may be a target of miR-203a-3p. ConclusioncircSTX6 may be a clinical prognostic biomarker in CSCC. The absence of circSTX6 inhibits cellular behaviors and increases the sensitivity of cervical cancer cells to radiation by modulating miR-203a-3p/RAB27B axis.

Theranostic potential of miRNAs in oral cancer: An emerging approach for precision oncology

AbstractMicroRNAs (miRNAs) have emerged as pivotal regulators in the pathogenesis of oral cancer, offering promising diagnostic and therapeutic opportunities in precision oncology. Dysregulated miRNA expression profiles are reliable biomarkers for oral cancer diagnosis and prognosis. Specific miRNAs, including miR‐21, miR‐31, miR‐375, miR‐let‐7a, miR‐125a, and miR‐200c, show altered expression patterns linked to oral cancer diagnosis, staging, and prognosis. Furthermore, these differentially expressed miRNAs can be detected in readily available biofluids such as saliva and plasma, improving their utility as non‐invasive diagnostic tools. Aberrant miRNA expression contributes to crucial oncogenic pathways in oral cancer progression, metastasis, and drug resistance. Using miRNA mimics or anti‐miRNAs to manipulate miRNA expression provides a mechanism for influencing downstream target genes and sensitizing cancer cells to conventional chemotherapy or targeted therapies. Preclinical studies have demonstrated the efficiency of miRNA‐based therapeutics in suppressing the development of oral cancer cells and triggering apoptosis. Furthermore, miRNAs have been linked to medication resistance, opening up new options for improving therapy response in patients with oral cancer. Further research and clinical validation are warranted to fully exploit the theranostic potential of miRNAs in oral cancer management and advance precision oncology.

Cyclin C promotes development and progression of B-cell acute lymphoblastic leukemia by counteracting p53-mediated stress responses.

Despite major therapeutic advances in the treatment of acute lymphoblastic leukemia (ALL), resistances and long-term toxicities still pose significant challenges. Cyclins and their associated cyclin-dependent kinases are one focus of cancer research when looking for targeted therapies. We discovered cyclin C as a key factor for B-ALL development and maintenance. While cyclin C is non-essential for normal hematopoiesis, CcncΔ/Δ BCR::ABL1+ B-ALL cells fail to elicit leukemia in mice. RNA sequencing experiments revealed a p53 pathway deregulation in CcncΔ/Δ BCR::ABL1+ cells resulting in the incapability of the leukemic cells to adequately respond to stress. A genome-wide CRISPR/Cas9 loss-of-function screen supplemented with additional knock-outs unveiled a dependency of human B-lymphoid cell lines on CCNC. High cyclin C levels in B-cell precursor (BCP) ALL patients were associated with poor event-free survival and increased risk of early disease recurrence after remission. Our findings highlight cyclin C as potential therapeutic target for B-ALL, particularly to enhance cancer cell sensitivity to stress and chemotherapy.

Epertinib counteracts multidrug resistance in cancer cells by antagonizing the drug efflux function of ABCB1 and ABCG2

A significant hurdle in cancer treatment arises from multidrug resistance (MDR), often due to overexpression of ATP-binding cassette (ABC) transporters like ABCB1 and/or ABCG2 in cancer cells. These transporters actively diminish the efficacy of cytotoxic drugs by facilitating ATP hydrolysis-dependent drug efflux and reducing intracellular drug accumulation in cancer cells. Addressing multidrug-resistant cancers poses a significant challenge due to the lack of approved treatments, prompting the exploration of alternative avenues like drug repurposing (also referred to as drug repositioning) of molecularly targeted agents to reverse MDR-mediated by ABCB1 and/or ABCG2 in multidrug-resistant cancer cells. Epertinib, a potent inhibitor of EGFR and HER2 currently in clinical trials for solid tumors, was investigated for its potential to resensitize ABCB1- and ABCG2-overexpressing multidrug-resistant cancer cells to chemotherapeutic agents. Our findings reveal that at sub-toxic, submicromolar concentrations, epertinib restores the sensitivity of multidrug-resistant cancer cells to cytotoxic drugs in a concentration-dependent manner. The results demonstrate that epertinib enhances drug-induced apoptosis in these cancer cells by impeding the drug-efflux function of ABCB1 and ABCG2 without altering their expression. ATPase activity and molecular docking were employed to reveal potential interaction sites between epertinib and the drug-binding pockets of ABCB1 and ABCG2. In summary, our study demonstrates an additional pharmacological capability of epertinib against the activity of ABCB1 and ABCG2. These findings suggest that incorporating epertinib into combination therapy could be advantageous for a specific patient subset with tumors exhibiting high levels of ABCB1 or ABCG2, warranting further exploration.

Expression of Concern: HER2 decreases drug sensitivity of ovarian cancer cells via inducing stem cell-like property in a NFκB-dependent way.

Expression of Concern: HER2 decreases drug sensitivity of ovarian cancer cells via inducing stem cell-like property in a NFκB-dependent way.

8038 Targeting Breast Cancer Stem Cells With Oncolytic Mammalian Orthoreovirus (MRV)

Abstract Disclosure: N.I. Roman Ortiz: None. J. Davydova: None. P. Danthi: None. J.H. Ostrander: None. Breast cancer is the most frequently diagnosed malignant cancer in women and is the second leading cause of cancer deaths among women in the United States. Patients with ER+ breast cancer can experience recurrence for more than 10 years after initial diagnosis. Breast cancer stem-like cells (BCSC) are slowly proliferative and exist as a minority sub-population (0-5%) within the tumor. Existing treatments target rapidly dividing cells, and BCSC are unresponsive to these therapies. Due to their self-renewal capabilities, BCSCs can fuel recurrence, metastasis, and resistance to therapies. Notably, no FDA-approved therapeutics selectively target BCSCs. Mammalian orthoreovirus (MRV) is an oncolytic virus tested in clinical trials for many cancer types including metastatic breast cancer. MRV was deemed safe but lacked efficacy as standalone treatment. We tested various laboratory MRV strains (T1L, R2) with T3D, a similar strain to the one evaluated in clinical trials. Our studies suggest that T3D is less effective in killing ER+ breast cancer cells and BCSCs compared to T1L and R2. To understand how T1L and R2 are more effective at killing ER+ breast cancer we aim to test the stages of MRV infection and how MRV induces cell death in ER+ therapy sensitive and resistant cells. MRV can enter cells by receptor mediated endocytosis through attachment with Junctional Adhesion Molecule A (JAM-A) and sialic acids as co-receptors. We have found that T1L attaches to cells in a JAM-A-dependent manner in ER+ sensitive cancer cells, while ER+ therapy resistant cells have increased resistance to JAM-A mediated endocytosis. Previous work has shown that apoptosis is involved in MRV-induced cell death. We found that apoptosis inhibitors in MRV-treated cells increase cell viability, suggesting that MRV-induces apoptotic cell death in ER+ breast cancer cells. Our current studies involve using RNA sequencing as a tool to assess gene expression of ER+ therapy sensitive and resistant cells infected with MRV at early and late stages of infection to determine key genes and pathways associated with cell death.We also aim to generate novel MRV strains with enhanced BCSC killing. We used a forward genetic approach and serially passaged MRV strains in BCSC enriched tumorsphere cultures of MCF-7 paclitaxel resistant (TaxR) cells to generate a selective oncolytic virus that can infect and kill BCSCs. Preliminary data has shown that serially passaged (SP) MRV strains (T1L SP, R2 SP) are more effective in decreasing cell viability and inhibiting tumorsphere formation compared to the parental strains (T1L and R2). Current studies are focused on sequencing SP clones to identify mutations that enhance BCSC death. We expect these studies to enhance MRV virotherapeutics for combined use with clinically relevant inhibitors (i.e. CDK4/6 inhibitors), anti-estrogens, or immunotherapy to prevent and treat metastatic ER+ breast cancer. Presentation: 6/2/2024

8038 Targeting Breast Cancer Stem Cells with Oncolytic Mammalian Orthoreovirus (MRV)

Abstract Disclosure: N.I. Roman Ortiz: None. J. Davydova: None. P. Danthi: None. J.H. Ostrander: None. Breast cancer is the most frequently diagnosed malignant cancer in women and is the second leading cause of cancer deaths among women in the United States. Patients with ER+ breast cancer can experience recurrence for more than 10 years after initial diagnosis. Breast cancer stem-like cells (BCSC) are slowly proliferative and exist as a minority sub-population (0-5%) within the tumor. Existing treatments target rapidly dividing cells, and BCSC are unresponsive to these therapies. Due to their self-renewal capabilities, BCSCs can fuel recurrence, metastasis, and resistance to therapies. Notably, no FDA-approved therapeutics selectively target BCSCs. Mammalian orthoreovirus (MRV) is an oncolytic virus tested in clinical trials for many cancer types including metastatic breast cancer. MRV was deemed safe but lacked efficacy as standalone treatment. We tested various laboratory MRV strains (T1L, R2) with T3D, a similar strain to the one evaluated in clinical trials. Our studies suggest that T3D is less effective in killing ER+ breast cancer cells and BCSCs compared to T1L and R2. To understand how T1L and R2 are more effective at killing ER+ breast cancer we aim to test the stages of MRV infection and how MRV induces cell death in ER+ therapy sensitive and resistant cells. MRV can enter cells by receptor mediated endocytosis through attachment with Junctional Adhesion Molecule A (JAM-A) and sialic acids as co-receptors. We have found that T1L attaches to cells in a JAM-A-dependent manner in ER+ sensitive cancer cells, while ER+ therapy resistant cells have increased resistance to JAM-A mediated endocytosis. Previous work has shown that apoptosis is involved in MRV-induced cell death. We found that apoptosis inhibitors in MRV-treated cells increase cell viability, suggesting that MRV-induces apoptotic cell death in ER+ breast cancer cells. Our current studies involve using RNA sequencing as a tool to assess gene expression of ER+ therapy sensitive and resistant cells infected with MRV at early and late stages of infection to determine key genes and pathways associated with cell death.We also aim to generate novel MRV strains with enhanced BCSC killing. We used a forward genetic approach and serially passaged MRV strains in BCSC enriched tumorsphere cultures of MCF-7 paclitaxel resistant (TaxR) cells to generate a selective oncolytic virus that can infect and kill BCSCs. Preliminary data has shown that serially passaged (SP) MRV strains (T1L SP, R2 SP) are more effective in decreasing cell viability and inhibiting tumorsphere formation compared to the parental strains (T1L and R2). Current studies are focused on sequencing SP clones to identify mutations that enhance BCSC death. We expect these studies to enhance MRV virotherapeutics for combined use with clinically relevant inhibitors (i.e. CDK4/6 inhibitors), anti-estrogens, or immunotherapy to prevent and treat metastatic ER+ breast cancer. Presentation: 6/2/2024

Non-cytotoxic levels of resveratrol enhance the anticancer effects of cisplatin by increasing the methyltransferase activity of CARM1 in human cancer cells

BackgroundResveratrol (RSVL) is a plant-derived polyhydroxyphenolic compound with excellent anticancer properties, alone or in combination with other chemotherapeutic drugs. However, the anticancer mechanism of RSVL is diverse and high concentrations are often required for RSVL to exert its anticancer effect, which would also adversely affect normal cells. PurposeThe main objective of this study is to investigate the molecular mechanism of how non-cytotoxic concentrations of RSVL enhance the anticancer effect of cisplatin involving a newly identified RSVL-binding protein. MethodsCell viability of cell lines from three cancer types exposed to RSVL and/or cisplatin was measured by NBB staining assay. RSVL-binding proteins were identified using RSVL-bound CNBr-activated Sepharose 4B beads coupled with LC-MS/MS, and the binding between RSVL and novel RSVL-binding protein was further confirmed with an in vitro pull-down assay. The expression of proteins was examined by immunoblot analysis, and the activity of methyltransferase was evaluated by in vitro methylation assay. The methylation level of H3R17 in the gene promoter was investigated using ChIP-qPCR. Bioinformatics analysis was conducted to identify pathway enrichment of genes, predict drug sensitivity, and analyze the survival of cancer patients. ResultsLow doses of RSVL might promote cancer cell growth whereas high doses of RSVL showed cytotoxic effects on normal cells. When co-treated with a lower cisplatin dose, non-cytotoxic RSVL levels showed synergistic anticancer effects. Here, coactivator-associated arginine methyltransferase 1 (CARM1) was identified as a novel RSVL-binding protein, and we showed that the upregulation of CARM1 increased the sensitivity of cancer cells to RSVL. Interestingly, we found that CARM1 was essential in the RSVL-induced sensitivity of cisplatin. Further molecular mechanistic studies revealed that RSVL could stabilize CARM1 protein, resulting in the upregulation and increased methyltransferase activity of CARM1. Additionally, we showed that the methylation levels of H3R17 in the promoter of p21, a downstream gene of CARM1 involving cell cycle arrest, were significantly increased after RSVL treatment. Finally, data from our bioinformatics analysis suggested that CARM1 could be utilized as a potential biomarker for chemotherapeutic drug sensitivity and prognosis in cancers. ConclusionsThis study identified CARM1 as a RSVL-binding protein for the first time and elucidated the potential roles of CARM1 in enhancing the efficacy of cisplatin by low doses of RSVL, which could have important clinical implications.