Abstract 4288: Discovery of a small molecule inhibitor of ADAR1 for cancer immunotherapy

Adenosine deaminase acting on RNA 1 (ADAR1) is a critical enzyme that catalyzes the conversion of adenosine to inosine in double-stranded RNA (dsRNA). By modifying endogenous dsRNA, ADAR1 prevents erroneous recognition by cytoplasmic dsRNA sensors, including MDA5, PKR, and ZBP1. Inhibition of ADAR1’s deaminase activity can activate MDA5-mediated innate immune responses and stimulate interferon production, while blocking ADAR1’s RNA-binding activity can induce PKR-mediated translational shutdown and cell death. Emerging evidence suggests that silencing ADAR1 enhances tumor responsiveness to immune checkpoint inhibitors, highlighting ADAR1 as a promising target in cancer immunotherapy. In this study, we aimed to develop small-molecule inhibitors of ADAR1 as potential therapeutic agents for ADAR1-dependent cancers. A primary RNA-editing assay was established to screen a library of 265,000 compounds, leading to the identification of 505 initial hits. These hits underwent triage validation involving target engagement studies, cell-based editing assays, and selectivity profiling. Among the validated compounds, RP04459 emerged as a lead candidate for further investigation. RP04459 effectively inhibited ADAR1’s RNA-editing activity in both biochemical and cellular assays. Treatment with RP04459 activated downstream interferon-β signaling in an MDA5-dependent manner, confirming its immunostimulatory potential. Detailed point mutation studies revealed the binding site of RP04459 on the ADAR1 protein, providing insight into its mechanism of action. The identification of RP04459 as a selective ADAR1 inhibitor underscores its potential to enhance immune-based therapies for cancers reliant on ADAR1 activity. Further preclinical evaluation of RP04459 may pave the way for novel therapeutic strategies in cancer immunotherapy Citation Format: Zijun Chen, Liqiang Shen, Qi Zhao, Jing Lu, Weiliang Li, Wen Jin, Xiang Ji, Mengchao Shi, Haowei Zhu, Lin Wang, Leduo Zhang, Xianqi Kong, Dawei Chen, Tianlun Zhou, Jiasheng Lu. Discovery of a small molecule inhibitor of ADAR1 for cancer immunotherapy [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 4288.