Project Summary Targeted protein degradation (TPD) is an emerging therapeutic modality with the potential to overcome limitations of traditional pharmacological inhibition approaches. TPD uses small molecules (degraders) to hijack the cellular degradation machinery by recruiting E3 ubiquitin ligases to proteins of interest (POI), which would not otherwise be recognized as substrate. Despite the tremendous promise of TPD approach, major bottlenecks still exist, including: 1) ligand discovery against undruggable disease-causing proteins. 2) a dearth of available E3 ligase recruiters that can be exploited for TPD applications, as there are limited number of E3 ligases, with most of the published degraders using ligands for cereblon and VHL, despite the existence of >600 E3 ligases in the human genome. Therefore, it is a great deal of interest in the discovering of new E3 ligase ligands to expand the scope of new therapeutics. Recently, several research groups discovered cysteine-reactive covalent recruiters against previously untargeted E3 ligases that have been successfully used in TPD by using chemical proteomic approaches. While targeting cysteines serves as a useful starting point, there is a need to consider other types of chemistry to generate covalent E3 recruiters for targeting new E3 ligases to expand the scope of TPD. One such possibility is to target potential nucleophilic residue, such as lysine, tyrosine, and serine on E3 ligases, respectively. However, specific biocompatible electrophiles targeting lysines, tyrosine, and serine are largely unexplored for TPD application. The ultimate goal of this proposal is to design, synthesize and validate new E3 ligases ligands that enable the development of novel degraders. To achieve this, we will use a chemoproteomic strategy that leverages broadly reactive, lysine, tyrosine, and serine-directed electrophilic warheads coupled to selective ligands for intracellular proteins (JQ1 for BRD4 or SLF for FKBP12) to screen for heterobifunctional degrader compounds that operate by covalent adduction of E3 ligases. To this end, we have already succeeded in developing a series of compounds to promote ligand-induced protein degradation. These proof-of-concept degraders led to the induced ubiquitination and proteasomal degradation of target proteins. We will perform further chemoproteomic studies to identify the E3 ligase targets of these degraders responsible for the degradation activity. With our novel approaches, we expect to see the repertoire of E3 ligases recruited for targeted protein degradation to be further enriched, paving the way for the next generation of tissue- and disease- specific molecular degraders.

Project Narrative The research proposed herein aims to discover a new class of covalent E3 ligase recruiters by chemoproteomic approaches. This research will generate novel degraders with the potential to expand the repertoire of E3 ligases, as well as tissue- and disease-specific molecular degraders.