Identification of a privileged scaffold to accelerate DUB drug discovery

Abstract

The ubiquitin/proteasome (UPS) is a highly regulated protein modification pathway, where the covalent attachment of one or more ubiquitin proteins promotes proteolysis, or other non-proteolytic fates, of substrate proteins. Three distinct enzymes, E1, E2, and E3 ligases, are required to conjugate the C-terminus of ubiquitin to the target protein. Substrates that are destined for degradation are proteolyzed in a ubiquitin-dependent manner. Deubiquinating enzymes (DUBs) oppose the activity of ligases and remove ubiquitin from target proteins. Improper function of the UPS has been linked to several human pathologies, including neurodegeneration, inflammation, and cancer. Due to their central role in UPS regulation, DUBs are increasingly recognized as attractive targets for therapeutic intervention in these diseases. The clinical success of the proteosome inhibitor, Bortezomib, has validated the ubiquitin system as a target for cancer therapy. However, despite extensive research into the basic function and disease relevance of the ubiquitin system, high quality chemical probes and clinical agents have been developed for only a small fraction of proteins that write (E1-E2-E3), read, and erase (DUB) ubiquitin PTMs. In this work, we embarked on a campaign to identify reversible small molecules that target recently described DUB pockets. To generate chemical starting points for inhibitors that target these new pockets, we screened a chemically diverse small molecule library across a set of eight DUBs. We subsequently used our established suite of DUB assays and computational analysis to credential new chemotypes as “leads” for a noncovalent library targeting non-catalytic pockets conserved across most DUBs. Importantly, we identified analogs from these chemotypes that inhibit DUBs relevant to cancer therapy. We leveraged the additional sequence and structural diversity represented in these pockets to develop selective inhibitors across the gene family. Our new chemical tools will accelerate the development of selective probes for promising DUB targets and enable detailed mechanism studies to decipher the function of DUBs in oncology.