Accelerating inhibitor discovery for deubiquitinating enzymes

Deubiquitinating enzymes (DUBs) are an emerging drug target class of similar to 100 proteases that cleave ubiquitin from protein substrates to regulate many cellular processes. A lack of selective chemical probes impedes pharmacologic interrogation of this important gene family. DUBs engage their cognate ligands through a myriad of interactions. We embrace this structural complexity to tailor a chemical diversification strategy for a DUB-focused covalent library. Pairing our library with activity-based protein profiling as a high-density primary screen, we identify selective hits against 23 endogenous DUBs spanning four subfamilies. Optimization of an azetidine hit yields a probe for the understudied DUB VCPIP1 with nanomolar potency and in-family selectivity. Our success in identifying good chemical starting points as well as structure-activity relationships across the gene family from a modest but purpose-build library challenges current paradigms that emphasize ultrahigh throughput in vitro or virtual screens against an ever-increasing scope of chemical space.

Automated summary

Deubiquitinating enzymes (DUBs) are a drug target class that regulate cellular processes by cleaving ubiquitin from protein substrates. This study used a diverse chemical library and activity-based protein profiling to identify selective hits against 23 DUBs. Optimization of a hit compound yielded a probe for the understudied DUB VCPIP1 with nanomolar potency and in-family selectivity.