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Abstract Targeted protein degradation (TPD) is a powerful strategy for targeting and eliminating disease-causing proteins. While heterobifunctional Proteolysis-Targeting Chimeras (PROTACs) are more modular, the rational design of monovalent or molecular glue degraders remains challenging. In this study, we generated a small library of BET-domain inhibitor JQ1 analogs bearing elaborated electrophilic handles to identify permissive covalent degradative handles and E3 ligase pairs. We identified an elaborated fumaramide handle that, when appended onto JQ1, led to the proteasome-dependent degradation of BRD4. Further characterization revealed that the E3 ubiquitin ligase CUL4DCAF16—a common E3 ligase target of electrophilic degraders—was responsible for BRD4 loss by covalently targeting C173 on DCAF16. While this original fumaramide handle, when appended onto other protein-targeting ligands, did not accommodate the degradation of other neo-substrates, a truncated version of this handle attached to JQ1 was still capable of degrading BRD4, now through targeting both C173 and C178. This truncated fumaramide handle, when appended on various protein targeting ligands, and was also more permissive in degrading other neo-substrates, including CDK4/6, SMARCA2/4, and the androgen receptor (AR). We further demonstrated that this optimized truncated fumaramide handle, when transplanted onto an AR DNA binding domain-targeting ligand, could degrade both AR and the undruggable truncation variant of AR, AR-V7, in androgen-independent prostate cancer cells in a DCAF16-dependent manner. Overall, we have identified a unique DCAF16-targeting covalent degradative handle that can be transplanted across several protein-targeting ligands to induce the degradation of their respective targets for the modular design of monovalent or bifunctional degraders.
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This content will become publicly available on January 27, 2026
A Kinetic Scout Approach Accelerates Targeted Protein Degrader Development
Bifunctional molecules such as targeted protein degraders induce proximity to promote gain‐of‐function pharmacology. These powerful approaches have gained broad traction across academia and the pharmaceutical industry, leading to an intensive focus on strategies that can accelerate their identification and optimization. We and others have previously used chemical proteomics to map degradable target space, and these datasets have been used to develop and train multiparameter models to extend degradability predictions across the proteome. In this study, we now turn our attention to develop generalizable chemistry strategies to accelerate the development of new bifunctional degraders. We implement lysine‐targeted reversible‐covalent chemistry to rationally tune the binding kinetics at the protein‐of‐interest across a set of 25 targets. We define an unbiased workflow consisting of global proteomics analysis, IP/MS of ternary complexes and the E‐STUB assay, to mechanistically characterize the effects of ligand residence time on targeted protein degradation and formulate hypotheses about the rate‐limiting step of degradation for each target. Our key finding is that target residence time is a major determinant of degrader activity, and this can be rapidly and rationally tuned through the synthesis of a minimal number of analogues to accelerate early degrader discovery and optimization.
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- Award ID(s):
- 2339705
- PAR ID:
- 10580425
- Publisher / Repository:
- Wiley
- Date Published:
- Journal Name:
- Angewandte Chemie International Edition
- Volume:
- 64
- Issue:
- 5
- ISSN:
- 1433-7851
- Format(s):
- Medium: X
- Sponsoring Org:
- National Science Foundation
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