Discovery of Ubiquitin-Specific Protease 7 (USP7) Inhibitors with Novel Scaffold Structures by Virtual Screening, Molecular Dynamics Simulation, and Biological Evaluation.

USP7 has been regarded as a potential therapeutic target for cancer. In this study, virtual screening, molecular dynamics (MD) simulation, and biological evaluation have been applied for the discovery of novel USP7 inhibitors targeting the catalytic active site. Among the obtained compounds, compound 12 with a novel scaffold structure exhibited certain USP7 inhibitory activity (Ub-AMC assay IC ₅₀ = 18.40 ± 1.75 μM, Ub-Rho assay IC ₅₀ = 7.75 μM). The binding affinity between USP7 _CD (USP7 catalytic domain) and this hit compound was confirmed with a K _D value of 4.46 ± 0.86 μM. Preliminary in vitro studies disclosed its antiproliferative activity on human prostate cancer cell line LNCaP with an IC ₅₀ value of 15.43 ± 3.49 μM. MD simulation revealed the detailed differences of protein-ligand interactions between USP7 _CD and the ligands, including the reference compound ALM4 and compound 12 , providing some important information for improving the bioactivity of 12 . This hit compound will serve as a promising starting point for facilitating the further discovery of novel USP7 inhibitors.