High-Throughput Sequential Screening to Identify Modulators of ADAR Epitranscriptomic Enzymes
Adenosine-to-inosine (A-to-I) RNA editing, catalyzed by ADAR enzymes, is a key epitranscriptomic mechanism that regulates immunity, neurodevelopment, and tumorigenesis. While ADAR1 upregulation promotes cancer progression, reduced ADAR2 activity correlates with aggressive gliomas; yet no small-molecule modulators of ADAR enzymes have been identified. Here, we present a high-throughput screening platform for systematic discovery of chemical modulators of ADAR-dependent RNA editing. The assay enables quantitative detection of editing activity and incorporates orthogonal validation steps to assess specificity, enzymatic function, and downstream molecular programs. Using this platform on glioblastoma models, we find compounds that modify ADAR activity and inhibit tumor cell growth, uncovering new aspects of epitranscriptomic regulation. This work establishes a scalable strategy to uncover chemical modulators of A-to-I editing and highlights RNA-editing pathways as actionable therapeutic targets in glioblastoma.