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Molecular targets for therapy

Development and preclinical validation of a novel covalent ubiquitin receptor Rpn13 degrader in multiple myeloma

Abstract

Proteasome inhibition is an effective treatment for multiple myeloma (MM); however, targeting different components of the ubiquitin–proteasome system (UPS) remains elusive. Our RNA-interference studies identified proteasome-associated ubiquitin-receptor Rpn13 as a mediator of MM cell growth and survival. Here, we developed the first degrader of Rpn13, WL40, using a small-molecule-induced targeted protein degradation strategy to selectively degrade this component of the UPS. WL40 was synthesized by linking the Rpn13 covalent inhibitor RA190 with the cereblon (CRBN) binding ligand thalidomide. We show that WL40 binds to both Rpn13 and CRBN and triggers degradation of cellular Rpn13, and is therefore first-in-class in exploiting a covalent inhibitor for the development of degraders. Biochemical and cellular studies show that WL40-induced Rpn13 degradation is both CRBN E3 ligase- and Rpn13-dependent. Importantly, WL40 decreases viability in MM cell lines and patient MM cells, even those resistant to bortezomib. Mechanistically, WL40 interrupts Rpn13 function and activates caspase apoptotic cascade, ER stress response and p53/p21 signaling. In animal model studies, WL40 inhibits xenografted human MM cell growth and prolongs survival. Overall, our data show the development of the first UbR Rpn13 degrader with potent anti-MM activity, and provide proof of principle for the development of degraders targeting components of the UPS for therapeutic application.

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Acknowledgments

The grant support for this investigation was provided by the National Institutes of Health Specialized Programs of Research Excellence (SPORE) grants P50100707, R01CA207237, and RO1CA050947. KCA is an American Cancer Society Clinical Research Professor.

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Authors

Contributions

DC conceptualized the project, designed and supervised all the research, analyzed the data, and wrote the manuscript; YS performed the majority of experiments, generated CRSPR-Cas9 Rpn13-knockout cells, and analyzed the data; LW performed molecule synthesis of WL40; PMCP designed and performed AlphaScreen assays for the CRBN and RPN13 and analyzed the data; AR performed flow cytometry and SCID mouse studies; TD carried out western blotting; SP and PF purified RPN13 protein. VKW and DL reviewed the manuscript. JQ designed biochemical experiments and molecule, and reviewed the manuscript; KCA provided clinical samples, reviewed the data, and wrote the manuscript.

Corresponding authors

Correspondence to Kenneth C. Anderson or Jun Qi or Dharminder Chauhan.

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Conflict of interest

KCA is on Advisory board of Celgene, Millenium-Takeda, Gilead, Janssen, and Bristol Myers Squibb, and is a Scientific Founder of Oncopep and C4 Therapeutics. DC is consultant to Stemline Therapeutic, Inc., and Equity owner in C4 Therapeutics. All the remaining authors declare no conflict of interest.

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Song, Y., Park, P.M.C., Wu, L. et al. Development and preclinical validation of a novel covalent ubiquitin receptor Rpn13 degrader in multiple myeloma. Leukemia 33, 2685–2694 (2019). https://doi.org/10.1038/s41375-019-0467-z

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