Design and Application of Cereblon-Recruiting Prodegraders

Abstract

Cereblon (CRBN) is one of the most widely co-opted E3 ligase substrate receptor proteins in targeted protein degradation (TPD), and several CRBN-recruiting degraders are currently in late-stage preclinical evaluation. While the diversity of CRBN-recruiting moieties has rapidly expanded, the glutarimide ring has remained largely conserved in these ligands. Glutarimides can form during protein cleavage events via the intramolecular cyclization of glutamine within the protein backbone. Inspired by this biological mechanism, we developed CRBN-recruiting prodegraders by replacing the glutarimide in parent degraders with uncyclized glutamine analogs. Prodegraders derived from a potent, cytotoxic GSPT1 degrader exhibited in cellulo cyclization, resulting in GSPT1 degradation. Optimization of the prodegrader scaffold revealed that glutarimide cyclization rates─and consequently, degradation kinetics─were tunable, with the most optimized prodegrader displaying degradation efficacy and cytotoxicity comparable to the parent, glutarimide-containing GSPT1 degrader. Furthermore, this prodegrader strategy can be readily applied to other known CRBN-based molecular glues and PROTACs. In contrast to conventional glutarimide-containing degraders, the amide of the prodegrader scaffold provides an accessible conjugation handle for stimulus-sensitive groups. We show that prodegraders can be conjugated to and released in vitro from a photolabile protecting group and a commonly used cathepsin-cleavable degrader–antibody conjugate (DAC) linker. Therefore, these prodegrader scaffolds introduce a generalizable conjugation strategy for CRBN-recruiting degraders to DAC linkers, eliminating the need for extensive degrader modifications to incorporate a conjugation handle. Overall, these findings establish the feasibility of utilizing glutamine analogs as a CRBN-recruiting prodegrader strategy and highlight their potential application in targeted drug delivery systems.