Identification of suitable target/E3 ligase pairs for PROTAC development using a rapamycin-induced proximity assay (RiPA)
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Abstract
The development of proteolysis targeting chimeras (PROTACs), which induce the degradation of target proteins by bringing them into proximity with cellular E3 ubiquitin ligases, has revolutionized drug development. While the human genome encodes more than 600 different E3 ligases, current PROTACs use only a handful of them, drastically limiting their full potential. Furthermore, many PROTAC development campaigns fail because the selected E3 ligase candidates are unable to induce degradation of the particular target of interest. As more and more ligands for novel E3 ligases are discovered, the chemical effort to identify the best E3 ligase for a given target is exploding. Therefore, a genetic system to identify degradation-causing E3 ligases and suitable target/E3 ligase pairs is urgently needed. Here, we used the well-established dimerization of the FKBP12 protein and FRB domain by rapamycin to bring the target protein WDR5 into proximity with candidate E3 ligases. Strikingly, this rapamycin-induced proximity assay (RiPA) revealed that VHL, but not Cereblon, is able to induce WDR5 degradation - a finding previously made by PROTACs, demonstrating its predictive power. By optimizing the steric arrangement of all components and fusing the target protein with a minimal luciferase, RiPA can identify the ideal E3 for any target protein of interest in living cells, significantly reducing and focusing the chemical effort in the early stages of PROTAC development.
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- Authors
- Adhikari, B., Schneider, K., Diebold, M., Sotriffer, C., & Wolf, E.
- Affiliations
- Christian-Albrechts-Universität zu Kiel (DE)
- University of Würzburg (DE)
- Corresponding author
- Wolf, Elmar
- Venue
- eLife
- Publisher
- eLife Sciences Publications, Ltd
- Published
- 2024-12-06
- Volume
- 13
- ISSN
- 2050-084X (ISSN-L)
- OpenAlex
- W4405113972
- Cited by
- 6
- References
- 54
- Citations by year
- 2026: 2 · 2025: 4
- FWCI
- 0.82
- Type
- journal article
- Language
- en
- Open access
- gold
- doi.org
- Licence
- Locations
- eLife (publishedVersion, cc-by)
- bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion, cc-by)
- Multimedialen Archiv und Publikationsserver der Christian-Albrechts-Universität zu Kiel (Christian-Albrechts-Universität zu Kiel) (submittedVersion, cc-by)
- Funders
- Deutsche Forschungsgemeinschaft (WO 2108/2-1, CRC 387)
- European Research Council (PROTAC-PDAC)
- Deutsche Krebshilfe (TACTIC)
- Bundesministerium für Bildung und Forschung (DEGRON)
- German Research Foundation (GRK 2243)
- Topics
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Protein Degradation and Inhibitors
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Ubiquitin and proteasome pathways
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Chemical Synthesis and Analysis
- Keywords
- PROTACs, E3 ubiquitin ligase, VHL, cereblon, protein degradation
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