Cereblon target validation using a covalent inhibitor of neosubstrate recruitment

Abstract

Small molecule ligands of cereblon (CRBN), a component of an E3 ubiquitin ligase complex, such as immunomodulatory drugs (IMiDs) or proteolysis targeting chimeras (PROTACs), induce new interactions between the E3 and a target protein that is subsequently polyubiquitinated and proteasomally degraded. The development of new degraders requires validation of CRBN-dependence and existing methods include the use of engineered CRBN knockout cell lines, or PROTACs directed to CRBN itself. Technical limitations of these approaches necessitate a simple and rapid pharmacological method of CRBN inhibition. We developed a sulfonyl fluoride covalent CRBN ligand based on the IMiD EM12 called EM12-SO₂F that was designed to engage His353 on the surface of the IMiD binding site. EM12-SO₂F does not act as a molecular glue degrader like other IMiDs, and instead serves as an inhibitor of such function by blocking the degrader binding site. We demonstrate utility of EM12-SO₂F by inhibiting the degradation of the zinc-finger transcription factor and CRBN neosubstrate IKZF1 by the molecular glue degrader lenalidomide. Increasingly, libraries of degrader molecules are being screened phenotypically to identify starting points for hit elaboration, that simultaneously reveals new therapeutic targets amenable to degradation. Indeed, targeted protein degradation has become an exciting new therapeutic modality and EM12-SO₂F augments the chemical biology toolbox that will advance this area of drug discovery research.

Keywords: Cereblon; Histidine targeting; Molecular glue; PROTAC; Phenotypic screening; Sulfonyl fluoride; Target validation; Targeted protein degradation.