Chemoresistant Epithelial Ovarian Cancer Cells Research Articles

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

ObjectiveOvarian 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. MethodsThe human EOC cell lines A2780, HeyA8, SKOV3ip1, A2780-CP20, HeyA8-MDR, and SKOV3-TR were treated with BNZ-111, and cell proliferation, apoptosis, and cell cycle were assessed. ResultsIt 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. ConclusionBNZ-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.

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.

Predicting chemoresponsiveness in epithelial ovarian cancer patients using circulating small extracellular vesicle-derived plasma gelsolin

BackgroundResistance to chemotherapy continues to be a challenge when treating epithelial ovarian cancer (EOC), contributing to low patient survival rates. While CA125, the conventional EOC biomarker, has been useful in monitoring patients’ response to therapy, there are no biomarkers used to predict treatment response prior to chemotherapy. Previous work in vitro showed that plasma gelsolin (pGSN) is highly expressed in chemoresistant EOC cell lines, where it is secreted in small extracellular vesicles (sEVs). Whether sEVs from tumour cells are secreted into the circulation of EOC patients and could be used to predict patient chemoresponsiveness is yet to be determined. This study aims to identify if sEV-pGSN in the circulation could be a predictive biomarker for chemoresistance in EOC.MethodsSandwich ELISA was used to measure pGSN concentrations from plasma samples of 96 EOC patients (primarily high grade serous EOC). sEVs were isolated using ExoQuick ULTRA and characterized using western blot, nanoparticle tracking analysis, and electron microscopy after which pGSN was measured from the sEVs. Patients were stratified as platinum sensitive or resistant groups based on first progression free interval (PFI) of 6 or 12 months.ResultsTotal circulating pGSN was significantly decreased and sEV-pGSN increased in patients with a PFI ≤ 12 months (chemoresistant) compared to those with a PFI > 12 months (chemosensitive). The ratio of total pGSN to sEV-pGSN further differentiated these groups and was a strong predictive marker for chemoresistance (sensitivity: 73.91%, specificity: 72.46%). Predetermined CA125 was not different between chemosensitive and chemoresistant groups and was not predictive of chemoresponsiveness prior to treatment. When CA125 was combined with the ratio of total pGSN/sEV-pGSN, it was a significant predictor of chemoresponsiveness, but the test performance was not as robust as the total pGSN/sEV-pGSN alone.ConclusionsTotal pGSN/sEV-pGSN was the best predictor of chemoresponsiveness prior to treatment, outperforming the individual biomarkers (CA125, total pGSN, and sEV-pGSN). This multianalyte predictor of chemoresponsiveness could help to inform physicians’ treatment and follow up plan at the time of EOC diagnosis, thus improving patients’ outcomes.

Anti-cancer effect of fenbendazole-incorporated PLGA nanoparticles in ovarian cancer.

Fenbendazole (FZ) has potential anti-cancer effects, but its poor water solubility limits its use for cancer therapy. In this study, we investigated the anti-cancer effect of FZ with different drug delivery methods on epithelial ovarian cancer (EOC) in both in vitro and in vivo models. EOC cell lines were treated with FZ and cell proliferation was assessed. The effect of FZ on tumor growth in cell line xenograft mouse model of EOC was examined according to the delivery route, including oral and intraperitoneal administration. To improve the systemic delivery of FZ by converting fat-soluble drugs to hydrophilic, we prepared FZ-encapsulated poly(D,L-lactide-co-glycolide) acid (PLGA) nanoparticles (FZ-PLGA-NPs). We investigated the preclinical efficacy of FZ-PLGA-NPs by analyzing cell proliferation, apoptosis, and in vivo models including cell lines and patient-derived xenograft (PDX) of EOC. FZ significantly decreased cell proliferation of both chemosensitive and chemoresistant EOC cells. However, in cell line xenograft mouse models, there was no effect of oral FZ treatment on tumor reduction. When administered intraperitoneally, FZ was not absorbed but aggregated in the intraperitoneal space. We synthesized FZ-PLGA-NPs to obtain water solubility and enhance drug absorption. FZ-PLGA-NPs significantly decreased cell proliferation in EOC cell lines. Intravenous injection of FZ-PLGA-NP in xenograft mouse models with HeyA8 and HeyA8-MDR significantly reduced tumor weight compared to the control group. FZ-PLGA-NPs showed anti-cancer effects in PDX model as well. FZ-incorporated PLGA nanoparticles exerted significant anti-cancer effects in EOC cells and xenograft models including PDX. These results warrant further investigation in clinical trials.

Binding of Intracellular Myeloperoxidase to αV/β1 Integrin Serves as a Mechanism of Survival in Epithelial Ovarian Cancer.

We were the first to report that epithelial ovarian cancer (EOC) cells and tissues express myeloperoxidase (MPO) that is known to play a role in immune surveillance and inflammation by myeloid cells. Additionally, we reported that MPO is colocalized with inducible nitric oxide synthase (iNOS), a key pro-oxidant enzyme, and plays a key role in regulating apoptosis in EOC cells. Whereas myeloid cells express MPO in a dimeric form, intriguingly, here we report the unique expression of only the monomeric form of MPO in EOC cells, tissues, and blood of an ovarian cancer patient. Additionally, we have identified a cell membrane receptor, αV/β1 integrin, that is uniquely expressed by both chemosensitive and chemoresistant EOC cells with significantly higher expression in chemoresistant EOC cells. More importantly, we have demonstrated that monoclonal antibodies against αV/β1 integrin induced cytotoxicity in EOC cells, but not in normal cells, that is also synergistic with conventional chemotherapies. Cytotoxicity of αV/β1 antibodies is due to conformational changes in αV/β1 integrin which prevents monomeric MPO binding to αV/β1 integrin inhibiting the activation of MPO, leading to increased apoptosis. Since normal epithelial cells and macrophages lack monomeric MPO and αV/β1 integrin system, targeting this unique MPO-dependent survival mechanism will selectively eliminate EOC cells and will be the target for developing specific ovarian cancer therapies.

SIAH1 reverses chemoresistance in epithelial ovarian cancer via ubiquitination of YBX-1

Chemoresistance is a severe outcome among patients with epithelial ovarian cancer (EOC) that leads to a poor prognosis. YBX-1 has been shown to cause treatment failure and cancer progression in EOC. However, strategies that directly target YBX-1 are not yet conceivable. Here, we identified that SIAH1 which was downregulated in chemoresistant EOC samples and cell lines functioned as novel E3 ligases to trigger degradation of YBX-1 at cytoplasm by RING finger domain. Mechanistic studies show that YBX-1 was ubiquitinated by SIAH1 at lys304 that lead to the instability of its target m5C-modified mRNAs, thus sensitized EOC cells to cDDP. Overexpression of SIAH1 enhanced the antitumor efficacy of cisplatin in vitro and in vivo, which were partially impaired by ectopic expression of YBX-1 or depletion of YBX-1 ubiquitination. In summary, our data identify the SIAH1/YBX-1 interaction as a therapeutic target for overcoming EOC chemoresistance.

An active RUNX1-ID1/ID3 axis governs differentiation and chemoresistance of cancer stem cell population in epithelial ovarian cancer cells

Progression, relapse, and therapy resistance are the most challenging features of cancer therapy that have been postulated to be driven by Cancer Stem Cell (CSC) population. This enigmatic subpopulation of cancer cells has therefore emerged as promising therapeutic candidate. We earlier reported enrichment of CSC-like side population (SP) with increasing resistance towards Cisplatin and Paclitaxel either alone or in combination in epithelial ovarian cancer (EOC) cells. This SP population is a small proportion of the total population of cancer cells characterised with high expression of drug transporters, a unique feature of stem cells and thereby can be isolated through their efflux properties of DNA binding dyes. While the bulk non-SP (NSP) population of the cancer cells lack overexpression of the drug transporters and thus can be identified as the dye containing population. In this study, we show that increased expression of Runt related transcription factor 1 (RUNX1) maintains undifferentiated state of CSC-like SP cells through upregulation of inhibitors of DNA binding/differentiation genes (ID1 and ID3) in late cisplatin-paclitaxel resistant cells. Higher RUNX1 expression was found to correlate with decreased median overall survival and disease-free survival in The Cancer Genome Atlas (TCGA) data set of high grade serous ovarian cancer (HGSOC) patients. The protein-protein interaction network analysis of 397 upregulated genes in RUNX1-high samples of TCGA data show significant enrichment of pathways known to negatively regulate CSC differentiation. Intriguingly RUNX1 inhibition not only induces CSC differentiation but also downregulates anti-apoptotic protein BCL2 in both SP and NSP cells and potentiates cytotoxic effects of Cisplatin-Paclitaxel in chemoresistant EOC cells. Inhibition of BCL2 through Venetoclax treatment, a small molecule BH3 mimic, sensitized these cells to platinum taxol treatment. Altogether, our data reveal new regulatory roles by RUNX1 to modulate CSC differentiation via ID1 and ID3 and to promote chemoresistance through BCL2 upregulation.

IGF1R-α6 integrin-S100A4 network governs the organ-specific metastasis of chemoresistant epithelial ovarian cancer cells

Recurrent metastatic epithelial ovarian cancer (EOC) is challenging and associated with treatment limitations, as the mechanisms governing the metastatic behavior of chemoresistant EOC cells remain elusive. Using orthotopic xenograft mouse models of sensitive and acquired platinum-taxol-resistant A2780 EOC cells, we studied the mechanistic role of insulin like growth factor 1 receptor (IGF1R) signaling in the regulation of organ-specific metastasis of EOC cells undergoing acquirement of chemoresistance. Biochemical assays and organ-specific fibroblast-EOC cell co-culture were used to study the differential metastatic characteristics of sensitive vs. chemoresistant EOC cells, and the key molecule/s underlying the organ-specific homing of chemoresistant EOC cells were identified through subtractive LC/MS profiling of the co-culture secretome. The role of the identified molecule was validated through genetic/pharmacologic perturbation experiments. Acquired chemoresistance augmented organ-specific metastasis of EOC cells and enhanced lung homing, particularly for the late-stage chemoresistant cells, which was abrogated after IGF1R silencing. Escalation of chemoresistance (intrinsic and acquired) conferred EOC cells with higher adhesion toward primary lung fibroblasts, largely governed by the α6 integrin-IGF1R dual signaling axes. Subtractive analysis of the co-culture secretome revealed that interaction with lung fibroblasts induced the secretion of S100A4 from highly resistant EOC cells, which reciprocally activated lung fibroblasts. Genetic and pharmacologic inhibition of S100A4 significantly lowered distant metastases and completely abrogated lung-tropic nature of late-stage chemoresistant EOC cells. These results indicate that chemoresistance exacerbates organ-specific metastasis of EOC cells via the IGF1R-α6 integrin-S100A4 molecular network, of which S100A4 may serve as a potential target for the treatment of recurrent metastatic EOC.

Nuclear HKII-P-p53 (Ser15) Interaction is a Prognostic Biomarker for Chemoresponsiveness and Glycolytic Regulation in Epithelial Ovarian Cancer.

Simple SummaryHexokinase II (HKII) is a key glycolysis enzyme associated with tumorigenesis, but its molecular mechanism and pathophysiological role in chemoresistant ovarian cancer remain elusive. In this study, we delineate the novel mechanism showing that activated phosphorylated-p53 (P-p53 Ser15) is required for the regulation of HKII intracellular trafficking and metabolic regulation in chemosensitive ovarian cancer, but not in chemoresistant ovarian cancer harboring p53 mutation. We have observed that increased nuclear HKII-P-p53 (Ser15) interaction is likely associated with chemosensitivity and better survival outcomes in epithelial ovarian cell lines, human primary epithelial ovarian cancer cells, and tumor sections. Nuclear HKII-P-p53 (Ser15) interaction may function as a promising prognostic biomarker, enabling prediction of patients with poor prognosis for deciding better clinical strategies.In epithelial ovarian cancer (EOC), carboplatin/cisplatin-induced chemoresistance is a major hurdle to successful treatment. Aerobic glycolysis is a common characteristic of cancer. However, the role of glycolytic metabolism in chemoresistance and its impact on clinical outcomes in EOC are not clear. Here, we show a functional interaction between the key glycolytic enzyme hexokinase II (HKII) and activated P-p53 (Ser15) in the regulation of bioenergetics and chemosensitivity. Using translational approaches with proximity ligation assessment in cancer cells and human EOC tumor sections, we showed that nuclear HKII-P-p53 (Ser15) interaction is increased after chemotherapy, and functions as a determinant of chemoresponsiveness as a prognostic biomarker. We also demonstrated that p53 is required for the intracellular nuclear HKII trafficking in the control of glycolysis in EOC, associated with chemosensitivity. Mechanistically, cisplatin-induced P-p53 (Ser15) recruits HKII and apoptosis-inducing factor (AIF) in chemosensitive EOC cells, enabling their translocation from the mitochondria to the nucleus, eliciting AIF-induced apoptosis. Conversely, in p53-defective chemoresistant EOC cells, HKII and AIF are strongly bound in the mitochondria and, therefore, apoptosis is suppressed. Collectively, our findings implicate nuclear HKII-P-p53(Ser15) interaction in chemosensitivity and could provide an effective clinical strategy as a promising biomarker during platinum-based therapy.

Abstract 1459: PTP4A3 phosphatase is a novel drug target for epithelial ovarian cancer

Abstract High grade serous ovarian cancer (HGSOC) is the deadliest epithelial ovarian cancer (EOC) variant, with a bleak outcome that kills >14,000 women annually in the US. In past four decades, first line therapies have changed only modestly. While dramatic advances in targeted- and immune-based therapies have been seen in some solid tumors, they have not been reported in patients with HGSOC. Recurrent and chemoresistant HGSOC is common. Therefore, it is imperative that new HGSOC drug targets be interrogated. The Protein Tyrosine Phosphatases of Regenerating Liver (PRL or PTP4A) family is significantly upregulated in a variety of tumors, including HGSOC. A bioinformatic analysis of the ovarian cancer using the KMplot Survival Database revealed that high PTP4A3 expression was associated with poor survival in EOC (p<0.0001, hazard ratio (HR)=1.35). In HGSOC early stage disease, PTP4A3 expression levels were also correlated with survival (stage I/II- p=0.0013, HR=3.2), with low expressing patients surviving twice as long as high expressing patients (upper quartile survival in low expression cohort = 81.2 months compared to high expression cohort = 44.3 months). This survival difference appeared independent of myc amplification and was more pronounced in advanced disease (stage III/IV). qPCR profiling of established HGSOC cell lines, including Kuramochi, COV362 and OVCAR4, showed 5-20 fold increases in PTP4A3 gene expression versus nonmalignant control cells. A potent, specific, allosteric small molecule inhibitor of PTP4A3, KVX-053, markedly reduced the growth and survival of chemoresistant and chemosensitive EOC cell lines independent of their BRCA or p53 status. Moreover, treatment of the HGSOC OVCAR4 cell line with KVX-053 (1 μM) inhibited the oncogenic Janus kinase-signal transducer and activator of transcription (JAK-STAT) signaling pathway through loss of phosphorylation of STAT3. Evaluation of a series of next-generation KVX-053-based analogs including chemodegraders yielded chemotypes with enhanced cytotoxic effects. These results demonstrate a clear path for the development of the PTP4A family of tyrosine phosphatases as a novel therapeutic target in a HGSOC. Citation Format: John Robert Cornelison, Ettore J. Rastelli, Duncan J. Hart, Anna J. Mendelson, Elizabeth R. Sharlow, Peter Wipf, John S. Lazo. PTP4A3 phosphatase is a novel drug target for epithelial ovarian cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1459.

LncRNA-H19 regulates chemoresistance to carboplatin in epithelial ovarian cancer through microRNA-29b-3p and STAT3.

Background: Platinum-based chemotherapy is part of current standard treatment for epithelial ovarian cancer (EOC). However, chemoresistance often rapidly developed, leading to chemotherapy failure and unfavored prognosis. Increasing evidence has demonstrated the important role of oncogenic long noncoding RNA H19 in various cancers, including EOC. No current study is available in exploring the role of lncRNA-H19 in carboplatin resistance of EOC and its underlying mechanism.Methods: Levels of lncRNA-H19, miR-29b-3p, and STAT3 mRNA were measured by qRT-PCR. The 50% inhibitory concentration value was detected with Cell Counting Kit-8 (CCK8). Colony-formation and CCK8 assays were employed to measure cell viability. Cell migration and invasion ability was evaluated with transwells. Western blot assay was utilized to measure P-gp, MRP1, LRP, and STAT3 protein levels. The targeting between lncRNA-H19 and miR-29b-3p, as well as miR-29b-3p and STAT3, was verified by dual-luciferase, RNA immunoprecipitation, and RNA pull-down experiments.Results: lncRNA-H19 and STAT3 were sharply increased, while miR-29b-3p was decreased in carboplatin-resistant EOC. Carboplatin efficacy was enhanced by lncRNA-H19 silencing in chemo-resistant EOC cells. lncRNA-H19 served as a competing endogenous RNA of miR-29b-3p, causing the derepression of miR-29b-3p downstream target STAT3, leading to chemoresistance in carboplatin-tolerated EOC.Conclusions: The lncRNA-H19/miR-29b-3p axis improved carboplatin resistance of EOC by targeting STAT3, indicating a possible approach to improving chemotherapy for EOC.

EGFR Blockade Reverses Cisplatin Resistance in Human Epithelial Ovarian Cancer Cells.

Background:EOC is one of the most lethal gynecological malignancy worldwide. Although the majority of EOC patients achieve clinical remission after induction therapy, over 80% relapse and succumb to the chemoresistant disease. Previous investigations have demonstrated the association of EGFR with resistance to cytotoxic chemotherapies, hormone therapy, and radiotherapy in the cancers. These studies have highlighted the role of EGFR as an attractive therapeutic target in cisplatin-resistant EOC cells. Methods:The human ovarian cell lines (SKOV3 and OVCAR3) were cultured according to ATCC recommendations. The MTT assay was used to determine the chemosensitivity of the cell lines in exposure to cisplatin and erlotinib. The qRT-PCR was applied to analyze the mRNA expression of the desired genes. Results:Erlotinib in combination with cisplatin reduced the cell proliferation in the chemoresistant EOC cells in comparison to monotherapy of the drugs (p < 0.05). Moreover, erlotinib/cisplatin combination synergistically decreased the expression of anti-apoptotic and also increased pro-apoptotic genes expression (p < 0.05). Cisplatin alone could increase the expression of MDR genes. The data suggested that EGFR and cisplatin drive chemoresistance in the EOC cells through MEKK signal transduction as well as through EGFR/MEKK pathways in the cells, respectively. Conclusion: Our findings propose that EGFR is an attractive therapeutic target in chemoresistant EOC to be exploited in translational oncology, and erlotinib/cisplatin combination treatment is a potential anti-cancer approach to overcome chemoresistance and inhibit the proliferation of the EOC cells.

Highly expressed STAT1 contributes to the suppression of stemness properties in human paclitaxel-resistant ovarian cancer cells.

Signal transducer and activator of transcription-1 (STAT1) is an important factor in various cellular processes. The cancer stem cell (CSC) is considered as a tumor-initiating cell that drives the inner hierarchy in many cancers including epithelial ovarian cancer (EOC). Here, we explored for the first time the regulation of STAT1 on stemness properties in chemoresistant EOC cells. The paclitaxel (PTX)-resistant EOC cell line (OV3R-PTX) was derived from PTX-sensitive OVCAR-3 cells treated by the PTX regimen. A single cell clone OV3R-PTX-B4 was selected by fluorescence-activated cell sorting. PTX-resistant cells grew slowly in conventional 2D and 3D cultures, but tumor xenograft with PTX-resistant cells grew fast in nude mice. Interestingly, OV3R-PTX-B4 cells shared the characteristics of CSCs and stemness properties were found to be increased in the non-adherent spheroid culture system. The PTX-resistant cells had a high expression of CSC-related markers and low expression of STAT1 that had a high methylation level of CpG in its promoter region. Overexpressed STAT1 suppressed stemness properties, cell proliferation, and colony formation and favored the overall survival of patients with EOC. In summary, these data indicate a regulatory mechanism of STAT1 underlying drug resistance and provide a potential therapeutic application for EOC patients with PTX resistance.

Heat Shock Protein 60 (HSP60) Serves as a Potential Target for the Sensitization of Chemoresistant Ovarian Cancer Cells.

HSP60 is a mitochondrial chaperone protein that is associated with decreased overall survival of ovarian cancer patients. We determined whether targeting HSP60 with its monoclonal antibody would induce cytotoxicity in sensitive and chemoresistant ovarian cancer cells and whether it is synergistic when combined with chemotherapeutic drugs. Epithelial ovarian cancer (EOC) cells and their docetaxel- or cisplatin-resistant counterparts were utilized. HSP60 mRNA levels were determined by real-time RT-PCR. Cytotoxicity of HSP60 antibody (0.5 or 1.5μg/ml) alone and in combination with chemotherapy were assessed by MTT Cell Proliferation Assay. Unpaired t tests were used to compare groups for real-time RT-PCR. One-way ANOVA followed by Tukey's post hoc tests with Bonferroni correction was performed for cytotoxicity comparisons. Significant synergistic effects of the antibody combined with chemotherapy were determined by the CompuSyn Software. Basal HSP60 mRNA levels were increased in chemoresistant EOC cells as compared with their sensitive counterparts (p < 0.05). There was no significant difference in cytotoxicity between EOC cell types; however, treatment with the HSP60 antibody for 24h showed a dose response (0.5 and 1.5μg/ml) cytotoxic effect to both sensitive and chemoresistant EOC cells as compared with the isotype control (p< 0.05). Importantly, treatment with both doses of HSP60 antibody was not cytotoxic to normal macrophages. Combination of the HSP60 antibody with docetaxel or cisplatin was significantly synergistic in both sensitive and chemoresistant EOC cells. Here, we identify a novel target that may serve not only for ovarian cancer treatment but also for sensitization of patients to chemotherapy. The cytotoxic effect of HSP60 monoclonal antibody and its synergism with chemotherapeutic agents highlight HSP60 as a promising target for therapy and chemosensitization in ovarian cancer treatment.

Abstract 4754: CHTOP is a novel therapeutic target for chemoresistant epithelial ovarian cancer therapy

Abstract Background: Chemotherapy is the mainstay treatment for ovarian cancer (OC). Chemoresistance is a major challenge in epithelial ovarian cancer (EOC) therapy. CHTOP was identified as a potential chemoresistant biomarker in chemoresistant EOC cell lines using label-free LC-MS/MS proteomic technique. However, the role of CHTOP in EOC chemoresistance is still unclear. Aim: In this study, we aimed to investigate whether CHTOP can be used as a therapeutic target in chemoresistant EOC cells and to reveal the mechanism underlying chemosensitization. Methods: The expression difference of CHTOP was detected in chemoresistant and metastatic EOC cell lines by immunofluorescence (IF) and Western blot (WB). The expression of CHTOP in human EOC tissues was examined using immunohistochemistry (IHC). The effect of CHTOP knockdown (KD) on metastasis was examined using the Transwell® matrigel invasion and wound healing assays. Flow cytometry and TUNEL assay were employed to determine the association of CHTOP with apoptosis, while mammary sphere formation assay and IF were used to evaluate its regulation on EOC-cis cell stemness. Results: The higher expression of CHTOP was found in EOC-cis (A2780-cis and IGROV-1-cis) and metastatic EOC (SKOV-3 and OV-90) cells as compared to normal epithelial ovarian cells (HOSE) by IF and WB. Also, high expression of CHTOP was found in human EOC tissues and associated with poor prognosis in patients. In contrast, CHTOP KD significantly reduced the metastatic potential of EOC-cis cells and increased their apoptosis at the presence of cisplatin. Furthermore, CHTOP KD decreased the stemness of EOC-cis cells. Conclusion: Our findings suggest that CHTOP is associated with metastasis, apoptosis, and stemness in chemoresistant EOC cells, the survival in EOC patients, and might be a promising therapeutic target to overcome chemoresistance in EOC treatment. Keywords: ovarian cancer, chemoresistance, CHTOP, metastasis, apoptosis, stemness Citation Format: Xupeng Bai, Xiaojie Feng, Jie Ni, Julia Beretov, Junli Deng, Ying Zhu, Peter Graham, Yong Li. CHTOP is a novel therapeutic target for chemoresistant epithelial ovarian cancer therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4754.

CHTOP in Chemoresistant Epithelial Ovarian Cancer: A Novel and Potential Therapeutic Target.

Objective: Chemoresistance is a major challenge in epithelial ovarian cancer (EOC) treatment. Chromatin target of protein arginine methyltransferase (CHTOP) was identified as a potential biomarker in chemoresistant EOC cell lines using label-free LC-MS/MS quantitative proteomics. Thus, the aim of this study is to investigate the role of CHTOP in chemoresistant EOC and the underlying mechanism.Methods: The expression of CHTOP in human ovarian cancer cells and tissues was detected using immunofluorescence (IF), western blot (WB), and immunohistochemistry (IHC), respectively. Flow cytometry and TUNEL assay were employed to detect the effect of CHTOP knockdown (KD) in chemoresistant EOC cell apoptosis, while colony and sphere formation assays were used to evaluate its effect on cell stemness. The association of CHTOP with cell metastasis was determined using Matrigel invasion and wound-healing assays.Results: The higher level expression of CHTOP protein was found in chemoresistant EOC cells as compared to their sensitive parental cells or normal epithelial ovarian cells. Results from IHC and bioinformatic analysis showed CHTOP was highly expressed in human ovarian cancer tissues and associated with a poor progression-free survival in patients. In addition, CHTOP KD significantly enhanced cisplatin-induced apoptosis, reduced the stemness of chemoresistant EOC cells, and decreased their metastatic potential.Conclusion: Our findings suggest that CHTOP is associated with apoptosis, stemness, and metastasis in chemoresistant EOC cells and might be a promising target to overcome chemoresistance in EOC treatment.

Inhibition of PI3K/Akt/mTOR signaling pathway alleviates ovarian cancer chemoresistance through reversing epithelial-mesenchymal transition and decreasing cancer stem cell marker expression

BackgroundOvarian cancer is the most common malignant tumor of the female reproductive tract. Chemoresistance is a major challenge for current ovarian cancer therapy. However, the mechanism underlying epithelial ovarian cancer (EOC) chemoresistance is not completely uncovered. The phosphatidylinositol-3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) signaling is an important intracellular pathway in regulating cell cycle, quiescence, and proliferation. The aim of this study is to investigate the role of PI3K/Akt/mTOR signaling pathway and its association with epithelial-mesenchymal transition (EMT) and cancer stem cell (CSC) marker expression in EOC chemoresistance.MethodsThe expressions of EMT and CSC markers were detected by immunofluorescence, western blot, and quantitative real-time PCR. BEZ235, a dual PI3K/mTOR inhibitor, was employed to investigate the role of PI3K/Akt/ mTOR signaling in regulating EMT and CSC marker expression. Students’ t test and one-way ANOVA with Tukey’s post-hoc test were used to compare the data from different groups.ResultsWe found that EMT and CSC marker expression were significantly enhanced in chemoresistant EOC cells, which was accompanied by the activation of PI3K/Akt/mTOR signaling. Compared with single cisplatin treatment, combined treatment with BEZ235 and cisplatin significantly disrupted the colony formation ability, induced higher ROS level and more apoptosis in chemoresistant EOC cells. Furthermore, the combination approach effectively inhibited PI3K/Akt/mTOR signaling pathway, reversed EMT, and decreased CSC marker expression in chemoresistant EOC cells compared with cisplatin mono-treatment.ConclusionsOur results first demonstrate that EMT and enhanced CSC marker expression triggered by activated PI3K/Akt/mTOR signaling are involved in the chemoresistance of EOC, and BEZ235 in combination with cisplatin might be a promising treatment option to reverse EOC chemoresistance.

156 - Modulation of mitochondrial fission activity maintains ovarian tumor initiating cells dependent on mitochondrial energetics

Self-renewing, therapy resistant ovarian tumor initiating cells (ovTICs) clearly contribute to development and recurrence of epithelial ovarian cancer (EOC), and recurrent EOC is incurable. Depending on the genetic and physiological context of a patient ovTICs can be dependent on mitochondrial energetics, specifically ATP production and redox homeostasis. Mitochondrial energetics can be repressed through fragmentation of mitochondria through elevated activity of mitochondrial fission protein Dynamin-related protein 1 (Drp1). Conversely, repression of Drp1 activity can augment mitochondrial energetics. Drp1 has also been implicated in maintenance of some stem cell lineages. However, Drp1 regulation of the stem-like ovTICs and the impact of fission activity on the mitochondrial energetics of these cells have not been well characterized. Hypothesis Modulation of mitochondrial energetics by fission, likely mediated by cyclic changes in Drp1 activity, maintains undifferentiated ovTICs. We studied mitochondrial properties in a derived chemoresistant EOC cell line previously shown to be enriched for ovTICs compared to its parental line. We found that the chemoresistant line has boosted mitochondrial energetics and repressed Drp1 activity compared to its parental line. We then FACS-sorted the chemoresistant line based on a TIC marker (Aldh) and mitochondrial membrane potential (ΔΨ) into 4 populations (Aldh-ΔΨlo, Aldh-ΔΨhi, Aldh+ΔΨlo, and Aldh+ΔΨhi). The sorted Aldh+ populations are dependent on mitochondrial energetics and cycle between ΔΨhi and ΔΨlo energetic states as they form in vitro tumorspheres, unlike the Aldh- populations. These energetic states have characteristic mitochondrial fission activity. Importantly, regulation of ATP production or redox state by fission is mutually exclusive for each energetic state. Understanding how fission can modulate the mitochondrial energetics of ovTICs that depend on mitochondria for ATP production and/or redox homeostasis will allow us to better identify and directly target TICs.

Exosomal expression of toll-like receptor 8-activating mir-146a-3p is upregulated by paclitaxel in chemoresistant epithelial ovarian cancer cells

Exosomal expression of toll-like receptor 8-activating mir-146a-3p is upregulated by paclitaxel in chemoresistant epithelial ovarian cancer cells

Targeting myeloperoxidase enhances apoptosis in chemoresistant epithelial ovarian cancer cells by reversing s-nitrosylation of caspase-3

Targeting myeloperoxidase enhances apoptosis in chemoresistant epithelial ovarian cancer cells by reversing s-nitrosylation of caspase-3