E3-Specific Degrader Discovery by Dynamic Tracing of Substrate Receptor Abundance

Targeted protein degradation (TPD) is a new pharmacology
based
on small-molecule degraders that induce proximity between a protein
of interest (POI) and an E3 ubiquitin ligase. Of the approximately
600 E3s encoded in the human genome, only around 2% can be co-opted
with degraders. This underrepresentation is caused by a paucity of
discovery approaches to identify degraders for defined E3s. This hampers
a rational expansion of the druggable proteome and stymies critical
advancements in the field, such as tissue- and cell-specific degradation.
Here, we focus on dynamic NEDD8 conjugation, a post-translational,
regulatory circuit that controls the activity of 250 cullin RING E3
ligases (CRLs). Leveraging this regulatory layer enabled us to develop
a scalable assay to identify compounds that alter the interactome
of an E3 of interest by tracing their abundance after pharmacologically
induced auto-degradation. Initial validation studies are performed
for CRBN and VHL, but proteomics studies indicate broad applicability
for many CRLs. Among amenable ligases, we select CRLDCAF15 for a proof-of-concept screen, leading to the identification of
a novel DCAF15-dependent molecular glue degrader inducing the degradation
of RBM23 and RBM39. Together, this strategy empowers the scalable
identification of degraders specific to a ligase of interest.