Metabolic Dependency on De Novo Pyrimidine Synthesis in Platinum Resistant Ovarian Cancer

Recurrent high grade serous ovarian cancer (OC) is mostly lethal due to development of resistance to platinum-based chemotherapy. We used cisplatin resistant (Cis-R) cells generated by repeat cisplatin treatment of sensitive (Cis-S) OC cell lines, and metabolomics to identify critical factors associated with platinum resistance. Pathway analysis of differentially abundant metabolites in Cis-R vs. Cis-S OVCAR5 and COV362 cells identified enrichment of pyrimidine metabolism (FDR<0.05) produced by increased de novo pyrimidine synthesis which was subsequently confirmed by flux analysis. Inhibition of this pathway using the DHODH inhibitor brequinar (BRQ) decreased viability, and produced expression changes in genes of the mitochondrial electron transport in Cis-R compared with Cis-S cells. Oxygen consumption rate (OCR) measured with a Seahorse assay indicated lower (P<0.05) OCR (baseline, ATP-related, and maximal) and spare respiratory capacity (SRC) in Cis-R vs. Cis-S OVCAR5 cells. In addition, BRQ had dose-related inhibitory effects on OCR across assay conditions in both Cis-R and Cis-S cells, SRC however, was inhibited by BRQ only in Cis-R cells. In vivo, BRQ attenuated growth of intraperitoneal (ip) Cis-R tumor xenografts but did not alter Cis-S tumors. Similarly, tumor growth was inhibited (p<0.01) by BRQ in a Cis-R PDX model. BRQ plus carboplatin enhanced (p<0.05) the inhibitory effects of carboplatin alone on total tumor weight and numbers of Cis-R ip tumor xenografts. Results indicate that Cis-R cells develop mechanisms that enhance de novo pyrimidine synthesis and maintain relatively low levels of mitochondrial oxidation, however these changes make Cis-R cells vulnerable to inhibitors of these processes. Overall design: To determine the relevance of de novo pyrimidine synthesis in resistance to cisplatin in ovarian cancer cells, we performed RNA-seq analysis of cisplatin resistant vs. cisplatin sensitive OVCAR5 cells treated with vehicle (control) or the DHODH inhibitor, brequinar (n=3 samples per group).