Abstract 2112: Identification of biomarkers associated with sensitivity to a novel PCNA inhibitor (AOH1996) by CRISPRi screening

Abstract Proliferating cell nuclear antigen (PCNA) plays an essential role in regulating DNA synthesis and repair and is essential for cancer cell growth and survival. We discovered a cancer-associated isoform of PCNA (caPCNA), which is ubiquitously and highly expressed in a broad range of cancer cells and tumor tissues but is not significantly expressed in non-malignant cells. Further studies of this novel caPCNA isoform as an anti-cancer drug target led to the identification of a cancer distinct region of PCNA which is partly delineated by the L126-Y133 peptide sequence and the discovery of a small molecule (AOH1996), which binds to this cancer distinct region as shown by crystallization studies. By targeting caPCNA, AOH1996 selectively kills cancer cells by interfering with the resolution of transcription-replication conflicts (TRC). Importantly, it causes no discernable toxicity at 6 times of its effective dose in mice. AOH1996 is currently in a phase 1 clinical trial for adult solid tumors at City of Hope (IND 138273). To identify genes that may be used as biomarkers for patient selection or potential targets for combination therapies, we screened a CRISPR interference (CRISPRi) library, which targets genes that are involved in cell signaling and cell cycle regulation. After culturing transduced cells in the presence of DMSO or 250 nM AOH1996 (corresponding to IC20) in triplicates for 14 generations, we measured the relative abundance of each guide RNA (gRNA) construct. Significant changes in the abundance of the gRNA constructs in AOH1996-treated cells relative to that in DMSO-treated cells were determined by the MAGeCK method. The target genes of the depleted or enriched gRNAs are predominantly involved in DNA repair, cell cycle regulation, and transcription regulation, which is consistent with the action mechanism of AOH1996. We validated the effect of these genes on sensitivity to AOH1996 and identified a panel of target proteins whose up or down-regulation is associated with sensitivity to AOH1996. Some of these proteins are targets of known chemotherapeutic drugs, which may be used in combination with AOH1996 in the clinic. Citation Format: Long Gu, Ming li, Chun-Wei Chen, Robert J. Hickey, Linda H. Malkas. Identification of biomarkers associated with sensitivity to a novel PCNA inhibitor (AOH1996) by CRISPRi screening [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 2112.