Abstract 115: S-adenosyl-methionine (SAM) synthesis protects ovarian cancer from chemotherapy induced DNA damage and contributes to cancer stem cell enrichment

Abstract High-grade serous ovarian cancer (HGSOC) is a deadly gynecological malignancy characterized by tumor recurrence and chemoresistance to first-line therapies, including platinum (Pt)-based chemotherapy, and Pt-induced increase in OC stem cells (OCSCs). Methionine (Met) has been shown to maintain CSC phenotypes, but its role in Pt-enriched OCSCs remains unclear. HGSOC cell lines were sorted using FACS into OCSCs and non-CSCs. Expression of methionine adenosyltransferase 2A (MAT2A), the enzyme converting Met to S-adenosyl-methionine (SAM), was higher (p<0.01) in OCSCs compared to non-CSCs. Supplementation of extracellular SAM (50μM) increased (p<0.05) the percentage of OCSCs, whereas inhibiting MAT2A by siRNA knockdown or the activity inhibitor prevented (p<0.05) the cisplatin (IC50, 16hr) induced increase in both OCSCs population and spheroid formation, demonstrating the importance of SAM biosynthesis in OCSC enrichment. The combination of cisplatin and siMAT2A also enhanced (p<0.01) DNA damage and apoptotic cell death compared to cisplatin. Mechanistically, siMAT2A inhibited (p<0.05) proteins involved in the DNA damage response and repair, including yH2Ax, ATM, ATR, Chk1 and FANCD2. MAT2A knockdown also suppressed (p<0.05) the cisplatin-induced upregulation of cyclin E/CDK2 which increased the percentage of cells by-passing the cell cycle checkpoint for DNA repair. Furthermore, siMAT2A increased (p<0.05) the binding of SAM sensor protein BMT2 on GATOR1, the negative regulatory complex of mTORC1, attenuating p70-S6 kinase and potentially contributing to downregulate cyclin E. Notably, the effects of siMAT2A were independent of the status of Met salvage enzyme methylthioadenosine phosphorylase. As SAM is a universal methyl donor for various methylation processes and given the established link between OCSC enrichment and aberrant DNA methylation, we used Infinium MethylationEPIC array v2.0 BeadChip to investigate genome-wide changes in DNA methylation using HGSOC cell line OVCAR3 treated with vehicle (control), cisplatin (15μM, 16hr) and/or siMAT2A (48hr). DNA methylation analysis identified differential methylated regions (DMRs) between three comparisons. Cisplatin alone increased (p<0.05) hypermethylated DMRs compared to the control. The combination of cisplatin and siMAT2A resulted in an increased (p<0.05) hypomethylated DMRs compared to cisplatin only, demonstrating a critical role of SAM synthesis in cisplatin-altered DNA methylation patterns. Enrichment analysis identified genes associated with stemness, differentiation, and cell death in the combination. Collectively, our study demonstrates a new role of SAM synthesis in Pt-induced DNA damage repair and OCSCs enrichment, providing a potential therapeutic target for improving chemotherapy outcomes and preventing OCSC enrichment in ovarian cancer. Citation Format: Shu Zhang, Tara X. Metcalfe, Zhen Fu, Christiane Hassel, Heather M. O'Hagan, Kenneth P. Nephew. S-adenosyl-methionine (SAM) synthesis protects ovarian cancer from chemotherapy induced DNA damage and contributes to cancer stem cell enrichment [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 115.