Abstract LB-108: Functional genetic screen to identify genetic determinants of “BRCAness” in ovarian cancer

Abstract

Abstract Background: Ovarian cancer is the number one killer among gynecologic cancers in the US and the fifth leading cause of cancer death among American women. High-grade serous ovarian carcinoma (HGS-OC) is the most common subtype of epithelial ovarian cancer and accounts for up to 70% of all ovarian cancer cases. Although standard chemotherapeutic agents, such as platinum agents and taxanes are effective, their effect is not durable. Poly (ADP-ribose) polymerase (PARP) inhibitors represent a class of promising drugs for targeted cancer therapies. Cancer cells that show “BRCAness” are highly sensitive to PARP inhibitors, including those with deleterious mutations in BRCA1/2 or defects in other components that are important for homologous recombination (HR) repair pathway. However, the understanding of the regulation of HR function remains incomplete. Methods: To get a better understanding of how homologous recombination repair is regulated, we performed a genome-scale genetic screen to identify potential genetic determinants of “BRCAness” in ovarian cancer. We introduced the human Genome-scale CRISPR-Cas9 knockout (GeCKO) pooled library B (contains 58,031 unique guide sequences targeting 19,052 human genes) into PARP inhibitor-sensitive A2780 ovarian cancer cells that stably express CRISPR-associated protein 9, followed by one week of puromycin selection and treatment with vehicle or PARP inhibitor, olaparib (5μM and 10μM). Genomic DNA from surviving cells were extracted, and gRNA sequences were rescued by PCR and identified by Illumina sequencing. Subsequently, we used MAGeCK tool to identify gRNAs that were either enriched and dropped out after olaparib selection. Results: By comparing the vehicle- and olaparib-treated groups, we identified top 10 sgRNAs targeting genes including OR5J2, IL21, C12orf5 and NME2 are depleted after olaparib selection, indicating that they are potential candidates that may contribute to olaparib resistance in ovarian cancer cells. In contrast, top 10 sgRNAs targeting genes including PARP1, GGT6, PA2G4 and TNRC6C are enriched with olaparib selection, suggesting their potential role in determining “BRCAness” in ovarian cancer cells. Conclusion: In this genome-scale screen, we identified top sgRNA candidates that are enriched after olaparib selection, including PARP1, loss of which has been shown to cause PARP inhibitor resistance. Genes targeted by these sgRNAs would be the potential “BRCAness” inducers. We also identified top sgRNAs that are conferring more sensitivity after depletion. Further studies are ongoing to unveil the detailed mechanisms. By uncovering these potential genetic determinants of PARP inhibitor sensitivity, it will facilitate our understanding of PARP inhibitor resistance and provides us potential therapeutic targets in combination with PARP inhibitors to treat ovarian cancer. Citation Format: Pingping Fang, Jeremy Chien. Functional genetic screen to identify genetic determinants of “BRCAness” in ovarian cancer. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr LB-108.