Targeted protein degradation using the endogenous Ubiquitin Proteasome System (UPS) represents a fundamentally new approach to drug discovery that potentially allows proteins that cannot be modulated by conventional small molecule inhibitors to be brought under therapeutic control. This provides the opportunity to develop therapeutics for traditionally undruggable targets including scaffolding proteins, protein-protein interactions and transcription factors. Traditional medicinal chemistry approaches have not been transferrable to drugging protein-protein interactions; consequently, prospective design of compounds to induce degradation is not well precedented, and to date has relied on empiricism and serendipity. Plexium has developed the DELPhe platform, which combines solid phase synthesis of DNA encoded libraries with high-throughput ultra-miniaturized cell-based assays, to screen large amounts of chemical space to identify monovalent or molecular glue degraders. Targeted libraries of drug-like molecules are created for each protein target or E3 ligase and are then assayed for their ability to cause degradation in intact cells. The DELPhe platform incorporates flexible readouts for identification of small molecule degraders by monitoring the loss of protein expression through immunofluorescence staining or through the generation of unique RNAseq signatures.
We describe here the use of Plexium’s DELPhe platform to sample extensive chemical space and discover small molecule monovalent degraders that demonstrate selective and sustained degradation of BRD4. Validation of screening hits identified monovalent degraders with rapid and potent degradation of BRD4 without any appreciable degradation of the highly homologous BRD2 and BRD3 proteins. Co-treatment with either proteosome or neddylation inhibitors blocked BRD4 degradation, demonstrating that loss of BRD4 protein was mediated by the ubiquitin proteosome system. Validated hits were further characterized in a ubiquitin ligase-focused CRISPR screen to identify the E3 ligase complex responsible for degradation. Collectively, these data demonstrate that Plexium’s DELPhe platform enables the rapid discovery of selective and potent monovalent degraders of BRD4.