Abstract 3665: Insilco evaluation of multiple ubiquitin-specific protease 1 (USP1) inhibitors and selectivity profiling over other USPs using Alphafold

Abstract DNA damage triggers cell signaling cascades that mediate repair. This signaling is frequently dysregulated in cancers. The proteins that mediate this signaling are potential targets for therapeutic intervention. Ubiquitin-specific protease 1 (USP1) is one such target, with small-molecule inhibitors already in clinical trials. Here, we used USP1 crystal structures (PDB: 7ZH4 and 9FCJ) to study the comparison of multiple novel USP1 inhibitors alongside the clinical USP1 inhibitor KSQ-4279 and a well-established tool compound, ML323. Our findings indicate that each of these novel USP1 inhibitors binds to the same cryptic site on USP1 as ML323/KSQ4279, yet they induce subtle variations in the protein structure. Notably, one of the novel compounds exhibit additional interactions with D751 and D752 which we hypothesize may contribute to the unique structural disruptions associated with this novel inhibitor. This binding would likely enhance the thermal stability of USP1 significantly, potentially due to the inhibitors occupying a hydrophobic tunnel-like pocket within the USP1 structure. Furthermore, we have successfully employed AlphaFold structures of various USP proteins to demonstrate the selective binding of these novel compounds to USP1, setting them apart from all other known USP’s. Our results advance the in silico understanding of the binding modes of these novel ligands and underscore their selective potential. We aim to utilize this data for the design of new USP1 inhibitors. Citation Format: Prashant Kashinath Bhavar, Uday Kumar Surampudi, Appaji Baburao Mandhare, Rupesh Chikhale. Insilco evaluation of multiple ubiquitin-specific protease 1 (USP1) inhibitors and selectivity profiling over other USPs using Alphafold [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 3665.