Construction of Small‐Molecule Deubiquitinase‐Targeting Chimeras to Reactivate and Stabilize Mutant p53 Y220C in Vitro and in Vivo

Wild-type p53 is a tumor suppressor protein that is crucial in various pathological processes. The predominant strategy for p53 Y220C mutant involves binding reactivators to mutation-induced pocket to stabilize the protein thermally and restore wild-type structure. However, the corrected p53 Y220C remains vulnerable to ubiquitination, reducing protein levels and impairing function. Herein, we report the development of p53 Y220C-specific deubiquitinase-targeting chimeras (DUBTACs)-bifunctional small molecules concurrently recruiting both p53 Y220C and the deubiquitinase OTUB1. The most potent compound, A1, effectively restored wild-type conformation and potently removed ubiquitin. Furthermore, A1 demonstrated superior efficacy in inducing apoptosis in p53 Y220C-mutant Huh7 cells and achieved a 61.06% reduction in tumor weight in Huh7 xenograft model. Our results demonstrated the viability of the DUBTAC strategy for p53 Y220C, successfully re-establishing conformation and promoting deubiquitination. More importantly, this study provides the first proof-of-concept for DUBTAC application in vivo, establishing targeted deubiquitination as a therapeutic approach.