Chemoresistant Ovarian Cancer Research Articles

Multicompartmentalized microvascularized tumor-on-a-chip to study tumor-stroma interactions and drug resistance in ovarian cancer.

The majority of ovarian cancer (OC) patients receiving standard of care chemotherapy develop chemoresistance within 5 years. The tumor microenvironment (TME) is a dynamic and influential player in disease progression and therapeutic response. However, there is a lack of models that allow us to elucidate the compartmentalized nature of TME in a controllable, yet physiologically relevant manner and its critical role in modulating drug resistance. We developed a 3D microvascularized multiniche tumor-on-a-chip formed by five chambers (central cancer chamber, flanked by two lateral stromal chambers and two external circulation chambers) to recapitulate OC-TME compartmentalization and study its influence on drug resistance. Stromal chambers included endothelial cells alone or cocultured with normal fibroblasts or cancer-associated fibroblasts (CAF). The tumor-on-a-chip recapitulated spatial TME compartmentalization including vessel-like structure, stromal-mediated extracellular matrix (ECM) remodeling, generation of oxygen gradients, and delayed drug diffusion/penetration from the circulation chamber towards the cancer chamber. The cancer chamber mimicked metastasis-like migration and increased drug resistance to carboplatin/paclitaxel treatment in the presence of CAF when compared to normal fibroblasts. CAF-mediated drug resistance was rescued by ECM targeted therapy. Critically, these results demonstrate that cellular crosstalk recreation and spatial organization through compartmentalization are essential to determining the effect of the compartmentalized OC-TME on drug resistance. Our results present a functionally characterized microvascularized multiniche tumor-on-a-chip able to recapitulate TME compartmentalization influencing drug resistance. This technology holds the potential to guide the design of more effective and targeted therapeutic strategies to overcome chemoresistance in OC.

Unraveling the role of ubiquitin C-terminal hydrolase L1 in high grade serous ovarian cancer pathogenesis.

e15078 Background: High grade serous ovarian cancer (HGSOC) is the most lethal of all gynecologic malignancies, due to the fact that the majority of patients are diagnosed at an advanced stage and eventually develop chemoresistant disease 12-18 months following completion of frontline therapy. Therefore, novel targeted therapies are urgently needed to combat chemoresistance and improve patient clinical outcomes. Ubiquitin C-terminal hydrolase L1 (UCHL1) is a deubiquitinating enzyme that plays a vital role in protein homeostasis and is highly overexpressed in HGSOC. It was previously uncovered that targeting UCHL1 with small molecule inhibition reduced cell viability in a chemoresistant ovarian cancer cell line. Methods: Comparative-label free proteomic analysis was used to uncover significant changes in HGSOC cell line OVCAR8WT following treatment with UCHL1 small molecule inhibitor, LDN-5744 or corresponding DMSO control. Significant pathway changes were determined by KEGG analysis. Matched chemosensitive and resistant HGSOC cells PEA1 and PEA2 were treated in combination with LDN-5744 and carboplatin, with cell viability was determined by an MTS assay. Furthermore, western blot analysis was employed to compare common cell growth pathway changes as a result of UCHL1 inhibition in HGSOC cells. Results: Proteomic analysis revealed a significant (p<0.05) upregulation in methyl phosphate capping enzyme (MEPCE) (4.9-fold), asparagine synthetase (ASNS) (2.5-fold), Phosphoserine aminotransferase 1 (PSAT1) (1.7-fold), and downregulation in centrosomal protein 55 (CEP55) (-7.2-fold), These identified proteomic changes were confirmed by western blot. KEGG pathways analysis using all significantly (p<0.05) differentially expressed proteins showcased significant (p<0.05) enrichment in amino acid biosynthesis, and metabolism, and chemical carcinogenesis-reactive oxygen species. Chemoresistant PEA2 cells treated in combination with LDN-5744 and carboplatin demonstrated a 59% reduction in cell viability, significantly (p<0.001) lower than the 13% and 22%, that was observed from UCHL1 and carboplatin treatment alone, respectively. Conversely, PEA1, the chemosensitive counterpart to PEA2 demonstrated a significant (p<0.001) 37% increase in cell viability upon combinatorial treatment compared with carboplatin alone, indicating that UCHL1 inhibition rescues the cells from chemotherapy induced cell death. Finally, LDN-5744 treatment in PEA2 led to a reduction in levels of phospho-(p-)AKT, p-STAT3, and p-ERK, while conversely in PEA1 cells levels of these proteins were increase or unchanged. Conclusions: Our findings demonstrate that targeting UCHL1 results in prominent metabolic changes in HGSOC cells and that the efficacy of UCHL1 inhibition is heavily dependent upon platinum status. Overall, this study highlights UCHL1’s novel therapeutic role in the recurrent HGSOC.

Plasma Gelsolin Inhibits Natural Killer Cell Function and Confers Chemoresistance in Epithelial Ovarian Cancer.

Plasma gelsolin (pGSN) overexpression in ovarian cancer (OVCA) disarms immune function, contributing to chemoresistance. The aim of this study was to investigate the immunoregulatory effects of pGSN expression on natural killer (NK) cell function in OVCA. OVCA tissues from primary surgeries underwent immunofluorescent staining of pGSN and the activated NK cell marker natural cytotoxicity triggering receptor 1 to analyze the prognostic impact of pGSN expression and activated NK cell infiltration. The immunoregulatory effects of pGSN on NK cells were assessed using apoptosis assay, cytokine secretion, immune checkpoint-receptor expression, and phosphorylation of STAT3. In OVCA tissue analyses, activated NK cell infiltration provided survival advantages to patients. However, high pGSN expression attenuated the survival benefits of activated NK cell infiltration. In the in vitro experiment, pGSN in OVCA cells induced NK cell death through cell-to-cell contact. pGSN increased T-cell immunoglobulin and mucin-domain-containing-3 expression (TIM-3) on activated NK cells. Further, it decreased interferon-γ production in activated TIM-3+ NK cells, attenuating their anti-tumor effects. Thus, increased pGSN expression suppresses the anti-tumor functions of NK cells. The study provides insights into why immunotherapy is rarely effective in patients with OVCA and suggests novel treatment strategies.

Single-cell transcriptome analysis of macrophage subpopulations contributing to chemotherapy resistance in ovarian cancer

BackgroundOvarian cancer, a fatal gynecological malignancy, is primarily managed through surgery and chemotherapy. However, a significant challenge arises as patients frequently experience relapse due to chemotherapy resistance. This study delves into the complex functions and underlying mechanisms of macrophages in chemotherapy resistance in ovarian cancer. MethodThe single-cell transcriptome sequencing data of ovarian cancer with or without chemotherapy were analyzed. Then, corresponding cell types were identified, and macrophages were extracted from all cells. Following the standardized single-cell analysis using the Seurat package, 15 distinct macrophage clusters were found and differentially expressed genes among them were analyzed. Moreover, their association with chemotherapy resistance was explored through cell proportions and gene expression. ResultIn the single-cell transcriptomic analysis of ovarian cancer tissues before and after chemotherapy, the cellular proportion of CXCL5+ macrophages, THBS1+ macrophages, and MMP9+ macrophages were significantly increased following chemotherapy. Further investigation revealed that these macrophage subpopulations upregulated the expression of multiple pro-tumorigenic angiogenic or invasive factors, in addition to CXCL5, THBS1, and MMP9, including CTSL, CXCL1, and CCL18. Finally, pathway enrichment analysis revealed the significant activation of signaling pathways, such as NOD-like receptor, MAPK, and TNF in these macrophage subpopulations, which provides direction for studying the mechanism of these subpopulations. ConclusionCXCL5+, THBS1+, and MMP9+ macrophage subpopulations exhibit an increased cellular prevalence post-chemotherapy and pro-tumorigenic molecular expression profiles, suggesting a close association with chemoresistance in ovarian cancer. These findings contribute to our understanding of the roles and mechanisms of macrophages in ovarian cancer chemoresistance, providing a theoretical basis and direction for the development of therapies targeting macrophages in overcoming ovarian cancer chemoresistance.

ATP6V1B1 regulates ovarian cancer progression and cisplatin sensitivity through the mTOR/autophagy pathway.

Early detection and effective chemotherapy for ovarian cancer, a serious gynecological malignancy, require further progress. This study aimed to investigate the molecular mechanism of ATPase H+-Transporting V1 Subunit B1 (ATP6V1B1) in ovarian cancer development and chemoresistance. Our data show that ATP6V1B1 is upregulated in ovarian cancer and correlated with decreased progression-free survival. Gain- and loss-of-function experiments demonstrated that ATP6V1B1 promotes the proliferation, migration, and invasion of ovarian cancer cells in vitro, while ATP6V1B1 knockout inhibits tumor growth in vivo. In addition, knocking down ATP6V1B1 increases the sensitivity of ovarian cancer cells to cisplatin. Mechanistic studies showed that ATP6V1B1 regulates the activation of the mTOR/autophagy pathway. Overall, our study confirmed the oncogenic role of ATP6V1B1 in ovarian cancer and revealed that ATP6V1B1 promotes ovarian cancer progression via the mTOR/autophagy axis.

OTUB2 silencing promotes ovarian cancer via mitochondrial metabolic reprogramming and can be synthetically targeted by CA9 inhibition

Ovarian cancer is an aggressive gynecological tumor characterized by a high relapse rate and chemoresistance. Ovarian cancer exhibits the cancer hallmark of elevated glycolysis, yet effective strategies targeting cancer cell metabolic reprogramming to overcome therapeutic resistance in ovarian cancer remain elusive. Here, we revealed that epigenetic silencing of Otubain 2 (OTUB2) is a driving force for mitochondrial metabolic reprogramming in ovarian cancer, which promotes tumorigenesis and chemoresistance. Mechanistically, OTUB2 silencing destabilizes sorting nexin 29 pseudogene 2 (SNX29P2), which subsequently prevents hypoxia-inducible factor-1 alpha (HIF-1α) from von Hippel-Lindau tumor suppressor-mediated degradation. Elevated HIF-1α activates the transcription of carbonic anhydrase 9 (CA9) and drives ovarian cancer progression and chemoresistance by promoting glycolysis. Importantly, pharmacological inhibition of CA9 substantially suppressed tumor growth and synergized with carboplatin in the treatment of OTUB2-silenced ovarian cancer. Thus, our study highlights the pivotal role of OTUB2/SNX29P2 in suppressing ovarian cancer development and proposes that targeting CA9-mediated glycolysis is an encouraging strategy for the treatment of ovarian cancer.

Platinum-induced upregulation of ITGA6 promotes chemoresistance and spreading in ovarian cancer

Platinum (PT)-resistant Epithelial Ovarian Cancer (EOC) grows as a metastatic disease, disseminating in the abdomen and pelvis. Very few options are available for PT-resistant EOC patients, and little is known about how the acquisition of PT-resistance mediates the increased spreading capabilities of EOC. Here, using isogenic PT-resistant cells, genetic and pharmacological approaches, and patient-derived models, we report that Integrin α6 (ITGA6) is overexpressed by PT-resistant cells and is necessary to sustain EOC metastatic ability and adhesion-dependent PT-resistance. Using in vitro approaches, we showed that PT induces a positive loop that, by stimulating ITGA6 transcription and secretion, contributes to the formation of a pre-metastatic niche enabling EOC cells to disseminate. At molecular level, ITGA6 engagement regulates the production and availability of insulin-like growth factors (IGFs), over-stimulating the IGF1R pathway and upregulating Snail expression. In vitro data were recapitulated using in vivo models in which the targeting of ITGA6 prevents PT-resistant EOC dissemination and improves PT-activity, supporting ITGA6 as a promising druggable target for EOC patients.

A Multi-Faceted Analysis Showing CRNDE Transcripts and a Recently Confirmed Micropeptide as Important Players in Ovarian Carcinogenesis.

CRNDE is considered an oncogene expressed as long non-coding RNA. Our previous paper is the only one reporting CRNDE as a micropeptide-coding gene. The amino acid sequence of this micropeptide (CRNDEP) has recently been confirmed by other researchers. This study aimed at providing a mass spectrometry (MS)-based validation of the CRNDEP sequence and an investigation of how the differential expression of CRNDE(P) influences the metabolism and chemoresistance of ovarian cancer (OvCa) cells. We also assessed cellular localization changes of CRNDEP, looked for its protein partners, and bioinformatically evaluated its RNA-binding capacities. Herein, we detected most of the CRNDEP sequence by MS. Moreover, our results corroborated the oncogenic role of CRNDE, portraying it as the gene impacting carcinogenesis at the stages of DNA transcription and replication, affecting the RNA metabolism, and stimulating the cell cycle progression and proliferation, with CRNDEP being detected in the centrosomes of dividing cells. We also showed that CRNDEP is located in nucleoli and revealed interactions of this micropeptide with p54, an RNA helicase. Additionally, we proved that high CRNDE(P) expression increases the resistance of OvCa cells to treatment with microtubule-targeted cytostatics. Furthermore, altered CRNDE(P) expression affected the activity of the microtubular cytoskeleton and the formation of focal adhesion plaques. Finally, according to our in silico analyses, CRNDEP is likely capable of RNA binding. All these results contribute to a better understanding of the CRNDE(P) role in OvCa biology, which may potentially improve the screening, diagnosis, and treatment of this disease.

RAD6 Overexpression and Ovarian Cancer Chemo-Resistance: Flow Cytometry and Immunohistochemistry

Objective: Ovarian cancer is one of the deadliest women’s cancers in the world. Nearly all the patients have poor prognosis and recurrence after standard therapy because of chemo-resistance. The standard therapy is cytoreductive debulking surgery followed by platinum-based chemotherapy. RAD6 has an important role in chemo-resistance. RAD6 is an E2 Ubiquitin-Conjugating Enzymes (UBE2) enzyme required for DNA repair, cell proliferation, and cell mutagenesis. Increased expression of RAD6 is believed to be associated with chemoresistance, recurrence, and poor prognosis of the disease. We aimed to study RAD6 relationship with ovarian cancer chemo-resistance and its ability to predict chemo-resistance. Methods: This study is an ambispective cohort study of 32 people in each group at the obstetrics-gynecology and pathology Department of Cipto Mangunkusumo, Tarakan, Dharmais, and Fatmawati Hospital. All patients will undergo standard cytoreductive debulking and histopathological examination followed by six series of chemotherapy followed by six months of observation. After the observation, we determine therapy response with the RECIST Criteria (Response Criteria in Solid Tumors). The chemo-resistance and chemo sensitive groups will be analyzed according to the therapy response. Our study is the first study examining RAD6 in ovarian cancer from flow cytometry blood test and directly from ovarian cancer tissue by double immunohistochemistry.

High-dimensional deconstruction of ovarian cancer at single-cell precision reveals HEBP2 that reshape the TIME and drive carboplatin resistance

BackgroundSingle-cell sequencing was employed to analyze the tumor immune microenvironment in ovarian cancer (OC) patients, exploring the evolutionary roles of various macrophage subgroups in OC progression and their correlation with fatty acid metabolism-related genes in contributing to drug resistance. MethodsThis study aimed to decipher the mechanisms underlying OC chemoresistance (OC-CR) and carboplatin resistance by integrating and analyzing multiple single-cell RNA sequencing datasets from OC patients. The tumor immune microenvironment in OC-CR patients exhibited notable alterations in cellular interactions and the proportions of different immune cell populations, in contrast to the cohort sensitive to OC chemotherapy. ResultsThe study demonstrates that the fatty acid-associated gene HEBP2 not only accelerates OC progression but also modifies the immune landscape of OC, driving the polarization from M0_TAM to M2_TAM. This shift results in a diminished efficacy of chemotherapy in OC. Furthermore, both in vitro and in vivo experiments underscored HEBP2′s role in boosting the proliferation of OC-resistant cell lines and suppressing apoptosis, thereby facilitating carboplatin resistance. ConclusionIn conclusion, the immune microenvironments of OC-CR significantly differ from those sensitive to chemotherapy, underscoring HEBP2′s role in fostering OC resistance. This establishes HEBP2 as a promising prognostic marker and a novel target for therapeutic strategies against OC resistance.

Frizzled class receptor 5 contributes to ovarian cancer chemoresistance through aldehyde dehydrogenase 1A1

BackgroundChemoresistance is associated with tumor relapse and unfavorable prognosis. Multiple mechanisms underlying chemoresistance have been elucidated, including stemness and DNA damage repair. Here, the involvement of the WNT receptor, FZD5, in ovarian cancer (OC) chemoresistance was investigated.MethodsOC cells were analyzed using in vitro techniques including cell transfection, western blot, immunofluorescence and phalloidin staining, CCK8 assay, colony formation, flowcytometry, real-time PCR, and tumorisphere formation. Pearson correlation analysis of the expression levels of relevant genes was conducted using data from the CCLE database. Further, the behavior of OC cells in vivo was assessed by generation of a mouse xenograft model.ResultsFunctional studies in OC cells showed that FZD5 contributes to epithelial phenotype maintenance, growth, stemness, HR repair, and chemoresistance. Mechanistically, FZD5 modulates the expression of ALDH1A1, a functional marker for cancer stem-like cells, in a β-catenin-dependent manner. ALDH1A1 activates Akt signaling, further upregulating RAD51 and BRCA1, to promote HR repair.ConclusionsTaken together, these findings demonstrate that the FZD5-ALDH1A1-Akt pathway is responsible for OC cell survival, and targeting this pathway can sensitize OC cells to DNA damage-based therapy.

Abstract 575: Identification of SORL1 as a key regulator of chemoresistance in ovarian cancer

Abstract This study aimed to understand the molecular mechanism of platinum chemoresistance in recurrent ovarian tumors with a focus on the SORL1-regulated pathways. SORL1 is an intracellular sorting receptor that binds to HER2 in breast cancer; this promotes HER2 recycling to the cell surface, which ultimately impacts the progression and treatment of breast cancer. The role of SORL1 in ovarian cancer has not been extensively studied. Method: RNA was extracted from 15 pairs of matched primary and recurrent high grade serous ovarian tumors collected from patients. Human transcriptomic microarray was used to identify the differentially expressed genes. The identified genes were validated by RT-QPCR and western blot in five ovarian cancer cell lines that were treated or untreated with carboplatin. To study the effects of identified target gene SORL1, overexpression and knockdown in ovarian cancer cell lines were established using SORL1 plasmid and shRNA. Proliferation and response to carboplatin were analyzed using celltiter Glo and caspase-3/7 activity assays respectively. EGFR and FGFR4 were identified as the interacting proteins of SORL1 in ovarian cancer cells through FGF1 and EGF1 treatment and co-immunoprecipitation. A proximity ligation assay was performed to validate their interactions. Finally, a xenograft mouse model was used to compare the tumor forming abilities and the response to carboplatin of SORL1-knockdown ovarian cancer cells and the control cancer cells in vivo. Protein expression of SORL1, EGFR, and FGFR4 in the collected tumors was determined via western blot. Result: Transcriptomic analysis of matched human primary and recurrent ovarian tumors identified SORL1 as an upregulated gene in recurrent tumors compared to primary samples. SORL1 expression was also upregulated in ovarian cancer cells surviving carboplatin treatment. Proliferation assays demonstrated that SORL1-overexpressing and knockdown cell lines had increased and inhibited proliferative capabilities respectively. Caspase-3 activity assays showed that SORL1-overexpressing and knockdown cell lines had reduced and increased sensitivity to carboplatin treatment respectively. SORL1 was found to bind to and promote the expression of EGFR and FGFR4. When injected subcutaneously into nude mice, SORL1-knockdown cells had reduced tumor forming capabilities and increased sensitivity to carboplatin. SORL1-knockdown caused the tumors to express lower levels of SORL1, EGFR, and FGFR4 proteins in comparison with the control tumors. Conclusion: This study demonstrates that SORL1 regulates the EGFR and FGF4 pathways to promote the growth and carboplatin resistance of ovarian cancer cells. Targeting this pathway has potential to aid in the treatment of platinum-resistant ovarian cancer. Citation Format: Miranda Mansolf, Ziyan Jian, Fangfang Bi, Tobias M. Hartwich, Yang Yang-Hartwich. Identification of SORL1 as a key regulator of chemoresistance in ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 575.

Abstract 5432: The role of nuclear factorκB-inducing kinase in ovarian cancer stem cells, chemoresistance, and relapse

Abstract Ovarian cancer is the most lethal gynecological cancer in the United States, largely due to its high relapse rates. Research suggests these high relapse rates are due to cancer stem-like cells (CSCs), a subpopulation of tumor cells that are resistant to chemotherapy and have enhanced tumor-initiation capacity. Post-treatment, CSCs self-renew to re-establish new tumors, leading to recurrent disease that is now resistant to chemotherapy. Given that over-expression of NF-κB has been linked to CSCs, cancer progression, and chemoresistance in various tumors, including ovarian, studies uncovering the mechanisms of this pathway are needed to develop novel therapeutics for ovarian cancer treatment. The upstream activator of the alternative NF-κB pathway, Nuclear Factor κB-inducing Kinase (NIK), could be an enticing target for eliminating CSCs, however its function in these cells is not established. In our current studies, we show that HGSOC cells grown as CSC-enriching 3D spheroids have increased NIK expression and activity relative to monolayer cultured cells. Moreover, cells expressing CSC markers (CD117+/CD133+), have increased NIK gene expression, further supporting an association between NIK and CSC maintenance. Upon treatment with chemotherapies carboplatin and paclitaxel, NIK gene expression increased in the surviving population, indicating a role in chemoresistance. Knockdown of NIK using siRNA led to changes in alternative NF-kB transcriptional activity and higher sensitivity to carboplatin and paclitaxel relative to vehicle controls. Current studies are evaluating the role of NIK in tumor-initiation capacity, asymmetric division, proliferation, and changes in expression of stemness genes. Lastly, an in vivo relapse model will test the feasibility of using a NIK inhibitor following chemotherapy to prevent relapse and prolong survival. Findings from this work will further define the role of NF-kB pathway components in facilitating CSCs survival and re-establishment of tumors and identify a new potential target for eliminating CSCs in ovarian cancer. Citation Format: Cassidy C. Lucht, Mikella Robinson, Samuel F. Gilbert, Omar Lujano Olazaba, Carrie D. House. The role of nuclear factorκB-inducing kinase in ovarian cancer stem cells, chemoresistance, and relapse [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 5432.

Abstract 7139: Anti-cancer effects of benzimidazole derivative BNZ-111 on paclitaxel-resistant ovarian cancer

Abstract Ovarian cancer, a leading cause of cancer-related deaths in women, remains a formidable challenge, especially in the context of platinum-resistant disease. This study investigated the potential of the benzimidazole derivative BNZ-111 as a novel treatment strategy for platinum-resistant ovarian cancer. With improved pharmacokinetic parameters, BNZ-111 effectively disrupts tubulin polymerization through hydrogen bonds and hydrophobic interactions. It demonstrated strong cytotoxicity in both chemo-sensitive and chemo-resistant epithelial ovarian cancer cell lines, inducing apoptosis and G2/M cell cycle arrest. In vivo experiments using orthotopic and patient-derived xenograft models showed significant tumor growth inhibition without apparent toxicity to vital organs. Unlike paclitaxel, BNZ-111 proved effective in paclitaxel-resistant cells, potentially by bypassing interaction with MDR1 and modulating β-3 tubulin expression to suppress microtubule dynamics. This study suggests that BNZ-111, with favorable drug-like properties, holds promise as a therapeutic option for platinum-resistant ovarian cancer, addressing a critical clinical need in gynecologic oncology Citation Format: Jeong-Won Lee, Kwangho Lee, Byumseok Koh, Yoo-Young Lee, Duk-Soo Bae. Anti-cancer effects of benzimidazole derivative BNZ-111 on paclitaxel-resistant ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7139.

Abstract 7155: Dichloroacetate reverses cisplatin resistance in ovarian cancer through promoting ROS production

Abstract Chemotherapy resistance and the high rate of ovarian cancer relapse pose major challenges in the treatment of ovarian cancer. The development of chemoresistance contributes to poor overall prognosis associated with ovarian cancer and high cancer-related mortality in women worldwide. Herein, we found that Dichloroacetate (DCA), a pan-pyruvate dehydrogenase kinase (PDK) inhibitor, is a potent agent to overcome Cisplatin (CP) resistance in ovarian cancer treatment. Firstly, we elucidated that phosphorylation of pyruvate dehydrogenase (pPDH) was elevated in CP-resistant ovarian cancer cells. Then, blockage of pPDH by DCA treatment significantly inhibited cell migratory and invasive capacity in CP-resistant ovarian cancer cells through the transition of motile mesenchymal phenotypes into epithelial characteristics. DCA markedly decreased the number of sphere formations and attenuated the stem cell-like properties in CP-resistant ovarian cancer cells. In combination with CP treatment, DCA effectively inhibited cell proliferation in chemoresistant ovarian cancer cells by triggering apoptosis. Collectively, we suggested that DCA is a promising drug to reverse CP resistance for ovarian cancer treatment. We further tested the underlying mechanisms of DCA to restore CP sensitivity in ovarian cancer. We found that CP treatment promoted reactive oxygen species (ROS) production to induce cell death in CP-sensitive ovarian cancer. However, CP-resistant ovarian cancer cells exhibited higher basal levels of ROS compared with sensitive cancer cells, and CP treatment failed to generate ROS production. That means ovarian cancer cells steadily adapted to elevated ROS levels, then becoming ROS-adapted ovarian cancer cells to survive. Interestingly, DCA treatment results in the buildup of ROS production, causing cell death in CP-resistant ovarian cancer. Therefore, our study suggests that ROS production can be an attractive target in DCA treatment to restore CP sensitivity in ovarian cancer. Citation Format: Yen Thi Do, Seungmee Lee, Changmin Shin, Hyewon Chung, Jin Young Kim, Eunyoung Ha, Sojin Shin, Ji Hae Seo. Dichloroacetate reverses cisplatin resistance in ovarian cancer through promoting ROS production [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7155.

Abstract 4671: Efficacy and mechanism of mebendazole for ovarian cancer therapy

Abstract Introduction: Ovarian cancer (OvCa) is the most lethal gynecological malignancy. Although most OvCa patients initially respond to frontline platinum-based therapies with approximately 80% of patients experiencing remission, approximately 75% of those women experience a recurrence with only approximately 50% of patients alive 5 years following diagnosis. There is a critical need to develop novel therapeutic agents or methods to overcome chemoresistance and toxicities experienced with conventional chemotherapy regimens used for OvCa patients. Repurposing drugs is a promising strategy to safely and more rapidly evaluate potentially effective treatment regimens. Over the past two decades, Mebendazole (MBZ), an anti-parasitic drug, has gained much attention in oncology due to its favorable biosafety profile and potent anti-cancerous activity seen in several human malignancies. Therefore, we examined the efficiency and mechanism of MBZ against OvCa. Methods: The in vitro efficacy of MBZ was evaluated in cisplatin-resistant (CPR) and parent (WT/sensitive) OVCAR-8 cell lines as well as in ascites samples collected from chemo-resistant OvCa patients by using 2D and/or 3D cell viability assay and invasion assay. The in vivo efficacy of MBZ was assessed in orthotopic animal models, utilizing CPR-OVCAR8 spheroids, and patient-derived xenograft (PDX) animal models with specimens from chemo-resistant ovarian cancer patients. Molecular mechanisms were investigated through western blot and immunofluorescence analysis. Result: Our results confirmed that MBZ efficiently decreased cell viability, spheroid size, and the invasion ability of OvCa cell lines and ascites samples. OvCa cells treated with MBZ demonstrated downregulation of proliferation markers and upregulation of apoptosis markers indicating inhibition of cell proliferation and induction of apoptosis. Mechanistically, MBZ inhibited the Wnt/β-catenin signaling pathway by downregulating the expression and nuclear localization of β-catenin and epithelial-mesenchymal transition (EMT) markers. Preclinical studies showed that MBZ at 50 mg/kg/day dose for 28 days significantly reduced tumor growth in PDXs and orthotopic tumor models without any evidence of toxicity. Conclusion: Collectively, our study strongly supports the therapeutic potential of MBZ against OvCa particularly, which warrants further clinical studies. Citation Format: Rajani Rai, Dhanamjai Penta, Debasish Dey, Doris M Benbrook, Lin Wang, Magdalena Bieniasz, Lauren E Dockery. Efficacy and mechanism of mebendazole for ovarian cancer therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4671.

Abstract 2603: Targeting chemo-resistant ovarian cancer with a first-in-class, first-in-human immune-oncology agents containing granzyme B

Abstract Ovarian cancer (OC) is the fifth leading cause of cancer-related mortality in women. The current standard of care consists of debulking surgery with platinum-taxane maintenance chemotherapy. However, OC will recur in most patients, resulting in a five-year survival rate of 45%. In the past decades, there have been several developments in front-line maintenance therapy including bevacizumab or PARP inhibitors. While these therapies have shown effectiveness in prolonging progression free survival, this has not translated into improvements in overall survival. It is, therefore, essential to develop more effective maintenance therapies and most of the current efforts are focused on targeted approaches and immune therapeutics. Though immunotherapeutic strategies have induced durable responses in patients with melanoma, lung cancer, and other malignancies, the OC response to immunotherapy is limited as there is a significant subset of patients who do not respond. Our laboratory has developed several fully human granzyme B (GrB) based fusion proteins against key OC targets including folate receptor 1 (GrB-Fc-mov018) and fibroblast growth factor-inducible 14 (GrB-Fc-IT4) which retain high specificity of the targeting agent, combined with the GrB specific activity, a pro-apoptotic mechanism of action, and a robust in vivo efficacy. We assessed the in vitro cytotoxicity of GrB-based constructs against a panel of OC cell lines sensitive (OVCAR8, SKOV3, A2780, HeyA8) or resistant (SKOV3-TX, A2780/Cl-16, HeyA8-MDR) to common chemotherapeutic agents. SKOV3-TX cells were >10 fold more resistant to Paclitaxel than the parental SKOV3 cells while A2780/Cl-16 were 11-fold more resistant to Cisplatin than the parental A2780 cells. In contrast, none of the chemo-resistant cell lines showed resistance to the GrB-based constructs and the IC50 values observed were similar to those of the parental cell lines in the low nanomolar range (<50 nM). Moreover, target expression analysis did not show significant differences between parental and chemo-resistant cell lines. A pilot in vivo efficacy study of GrB-Fc-IT4 and GrB-Fc-mov018 against a subcutaneous, established ovarian tumor (OVCAR8) xenograft showed promising growth inhibition of established tumors compared to vehicle controls. In vivo studies evaluating the response to different treatments of mice bearing chemo-resistant ovarian tumors are ongoing. Our results suggest that the mechanism of resistance developed to conventionally used chemotherapeutic agents such as Paclitaxel, Doxorubicin, Cisplatin or Mitomycin do not confer resistance to GrB-based constructs. Hence, we speculate that GrB-based constructs could be an effective therapy in OC patients with immune-excluded tumors that have developed resistance to conventional treatment. Research conducted, in part, by the Clayton Foundation for Research. Citation Format: Ana Alvarez-Cienfuegos, Lawrence H. Cheung, Khalid A. Mohamedali, Michael G. Rosenblum. Targeting chemo-resistant ovarian cancer with a first-in-class, first-in-human immune-oncology agents containing granzyme B [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2603.

Abstract 2922: Sensory innervation as a novel driver of ovarian cancer progression

Abstract One of the main challenges currently faced in ovarian cancer treatment is that the primary platinum/taxane therapy used to treat patients does not meaningfully extend survival. The overall survival for ovarian cancer patients with advanced disease is less than 30% with most patients recurring within 5 years of initial therapy. Once resistant disease develops there are few second line therapy options, which inevitably leads to disease progression. Thus, there is a critical need to identify new therapies that can overcome ovarian cancer chemoresistance and produce meaningful increases in patient survival. In recent years, new efforts have focused on targeting the tumor microenvironment (TME), the complex mixture of non-cancer cells that surround and support the cancer cells within the tumor. Different strategies have evolved to target the TME, such as immune checkpoint inhibitors or angiogenesis inhibitors. However, these TME targeting therapies have limited effect when used to treat ovarian cancer patients. Most ovarian tumors have a microenvironment that is highly immunosuppressive and do not respond or are resistant to angiogenesis inhibitors. Thus, new TME targeting therapies must be developed that can induce a therapeutic effect in ovarian tumors and can be used to treat most ovarian cancer patients. A novel TME component ripe for therapeutic targeting are the nerve fibers that infiltrate tumors in a process termed tumor innervation. Recent studies in several cancers have shown that tumor innervation can promote tumor growth/metastasis. However, in ovarian cancer the role of innervation in promoting cancer progression remains undefined. Here we show that Transient Receptor Potential cation channel subfamily V member 1 (TRPV1)+ sensory innervation plays a significant role in driving ovarian cancer growth and metastasis. Analysis of patient samples shows that sensory nerve fiber innervation is much higher in ovarian tumors vs benign reproductive tissue. In addition, ablation of TRPV1 sensory nerve fibers in vivo causes reduced tumor burden and prolongs survival in a syngeneic mouse model of ovarian cancer metastasis. Taken together, our results establish TRPV1+ sensory innervation as a novel driver of ovarian cancer growth/metastasis and a potential therapeutic target for ovarian cancer treatment. Citation Format: Matthew Knarr, Katherine Cummins, Dusan Racordon, Hunter Reavis, Timothy Lippert, Ryan Hausler, Priyanka Rawat, Suyeon Ryu, Jamie Moon, Dave Hoon, Roger Greenberg, Paola Vermeer, Ronny Drapkin. Sensory innervation as a novel driver of ovarian cancer progression [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 2922.

PIK3R1 fusion drives chemoresistance in ovarian cancer by activating ERK1/2 and inducing rod and ring-like structures

Gene fusions are common in high-grade serous ovarian cancer (HGSC). Such genetic lesions may promote tumorigenesis, but the pathogenic mechanisms are currently poorly understood. Here, we investigated the role of a PIK3R1-CCDC178 fusion identified from a patient with advanced HGSC. We show that the fusion induces HGSC cell migration by regulating ERK1/2 and increases resistance to platinum treatment. Platinum resistance was associated with rod and ring-like cellular structure formation. These structures contained, in addition to the fusion protein, CIN85, a key regulator of PI3K-AKT-mTOR signaling. Our data suggest that the fusion-driven structure formation induces a previously unrecognized cell survival and resistance mechanism, which depends on ERK1/2-activation.

Impact of Obesity and Lysosomal Dysfunction on Chemoresistance in Ovarian Cancer.

Obesity is recognized as a significant risk factor for ovarian cancer, with accumulating evidence highlighting its impact on disease progression and chemoresistance. This review synthesizes current research elucidating the link between obesity-induced lysosomal dysfunction and ovarian cancer chemoresistance. Epidemiological studies consistently demonstrate a positive correlation between body mass index (BMI) and ovarian cancer risk, attributed in part to the predilection of epithelial ovarian cancer cells for adipose tissue, particularly the omentum. Adipokines released from the omentum contribute to cancer-associated characteristics, including energy supply to cancer cells. Moreover, obesity-induced alterations in lysosomal function have been implicated in systemic inflammation and lipid metabolism dysregulation, further exacerbating cancer progression. Lysosomes play a crucial role in drug resistance, as evidenced by studies demonstrating their involvement in mediating resistance to chemotherapy in ovarian cancer cells. Recent findings suggest that pharmacological inhibition of lysosomal calcium channels sensitizes drug-resistant ovarian cancer cells to cisplatin treatment, highlighting the therapeutic potential of targeting lysosomal dysfunction in obesity-related chemoresistance. This review underscores the importance of understanding the multifaceted roles of lysosomes in obesity-related drug resistance and their implications for the development of targeted therapeutic interventions in ovarian cancer management.