SAR analysis provided the opportunity to directly test the impact of altering k-on and k-off rates, which revealed that slow k-off as far more important than fast k-on for degradation activity. 7/8

Comprehensive mutational analysis revealed the GEMIN3 degron by which dGEM3 mediates recognition by VHL. 6/8

We extensively characterize the properties of dGEM3 to show biochemical reconstitution of the VHL:dGEM3:GEMIN3 ternary complex and ubiquitination consistent with a molecular glue without pre-existing affinity between VHL and GEMIN3. 5/7

Proteomics analysis reveals dGEM3 is a highly selective degrader and the direct target is the SMN complex protein GEMIN3. 4/8

We discover dGEM3 as a VHL-based molecular glue causing significant gene expression changes. 3/8

We use an innovative gene expression read out approach to do target-agnostic screening of a focused 26,000 VHL ligand library for molecular glue degraders. 2/8

Detailed mechanistic studies revealed that EN1441 promotes AR-v7 insolubility leading to aggregation that precedes degradation. 5/7

Further studies revealed that EN1441 inhibits AR-v7 transcriptional activity, leading to gene expression changes consistent with AR pathway inhibition. Importantly, this inhibition is more complete compared to therapies that only inhibit full-length AR and not AR-v7. 4/7

EN1441 directly engages AR-v7 Cys125 and causes proteasome-dependent degradation. 3/7

AR is a heavily drugged target in prostate cancer. A splice variant AR-v7 the occurs during the natural progression of disease has been challenging to drug. We performed a screen for AR-v7 degraders using a covalent fragment library leading to discovery of EN1441. 2/7