Abstract 6989: Advancing AOH1996: Enhanced anticancer activity of next-generation small molecule PCNA inhibitors

Abstract Proliferating cell nuclear antigen (PCNA) plays a critical role in at least nine distinct cellular pathways, including DNA replication and repair, transcription, cell cycle control, apoptosis induction, transcription-replication conflict resolution, and immune surveillance. Tumor cells express an acidic isoform of this protein (caPCNA), whereas non-cancer cells express only a basic isoform (nmPCNA). The absence of the caPCNA isoform in non-cancer cells makes it an ideal target for selectively killing cancer cells while preserving the viability of non-cancer cells. To this end, we developed a small molecule, AOH1996, that selectively inhibits the caPCNA isoform in cancer cells, inducing cell death in these cells while showing no detectable effects on non-cancer cells. AOH1996 directly binds to caPCNA, stabilizes its trimer structure, reduces chromatin association, and selectively inhibits tumor cell growth. Currently, in several phase I and phase Ib clinical trials, AOH1996 exhibits highly selective anti-cancer properties with essentially no off-target toxicities. This study describes our efforts to improve the clinical characteristics, potency, and stability of AOH analogues through a scaffold-hopping strategy that employs structure-activity relationship (SAR) studies and bioisostere replacement. SAR analysis and modification of four unique scaffolds related to AOH1996 led to the discovery of several additional analogues exhibiting enhanced potency, selectivity, and metabolic stability compared to the clinical lead. Over one hundred AOH analogues were designed, synthesized, and initially evaluated for effects on cell growth across eight tumor cell lines. Our results identify top analogues featuring unsymmetrical 3-methoxy-5-methyl groups on the terminal phenoxy moiety, combined with fluorine, nitrile, or chlorine substitutions at the #6 position of 1-naphthoic acid. Among these, AOH-3M5Me-6F and AOH-3M5Me-6CN emerged as the lead analogues, demonstrating up to nine- and seven-fold improved potency and 38% and 68% higher liver microsome stability, respectively. These findings position these analogues for further development as selective PCNA-targeting cancer therapies. Citation Format: Pouya Haratipour, Maryam Zangi, Mahshid Yaghoubi, Long Gu, Jennifer Jossart, John J. Perry, Linda H. Malkas, Robert J. Hickey. Advancing AOH1996: Enhanced anticancer activity of next-generation small molecule PCNA inhibitors [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 6989.