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Published May 2017 | Supplemental Material + Accepted Version
Journal Article Open

Capzimin is a potent and specific inhibitor of proteasome isopeptidase Rpn11


The proteasome is a vital cellular machine that maintains protein homeostasis, which is of particular importance in multiple myeloma and possibly other cancers. Targeting of proteasome 20S peptidase activity with bortezomib and carfilzomib has been widely used to treat myeloma. However, not all patients respond to these compounds, and those who do eventually suffer relapse. Therefore, there is an urgent and unmet need to develop new drugs that target proteostasis through different mechanisms. We identified quinoline-8-thiol (8TQ) as a first-in-class inhibitor of the proteasome 19S subunit Rpn11. A derivative of 8TQ, capzimin, shows >5-fold selectivity for Rpn11 over the related JAMM proteases and >2 logs selectivity over several other metalloenzymes. Capzimin stabilized proteasome substrates, induced an unfolded protein response, and blocked proliferation of cancer cells, including those resistant to bortezomib. Proteomic analysis revealed that capzimin stabilized a subset of polyubiquitinated substrates. Identification of capzimin offers an alternative path to develop proteasome inhibitors for cancer therapy.

Additional Information

© 2017 Macmillan Publishers Limited. received 7 November 2016; accepted 23 January 2017; published online 28 February 2017. We thank A. Martin (University of California, Berkeley) for the plasmid that expresses the Rpn11–Rpn8 dimer, E. Zeqiraj (University of Leeds) for providing purified BRISC complex, P. Coffino (The Rockefeller University) for providing the Rpn10-I27^(V13P)ext plasmid, T.M. Kapoor (The Rockefeller University) for providing the RPE1 WT and BTZ-resistant cell lines, National Cancer Institute (Developmental Therapeutics Program in the Division of Cancer Treatment and Diagnosis) for screening capzimin against 60 human cancer cell lines (http://dtp.cancer.gov), H. Park for testing the cell lines for mycoplasma contamination and Y. Zhang for advice on the competition studies. This work was supported by grants from the Caltech Gates Grubstake Fund and Amgen to R.J.D. and from the NIH (CA164803) to R.J.D. and S.M.C., T.Y. and S.H. were supported by the Gordon and Betty Moore Gordon Foundation, through Grant GBMF775, the Beckman Institute, and the NIH through Grant 1S10RR029594-01A1. R.J.D. is an Investigator of and was supported by the Howard Hughes Medical Institute. Author Contributions: J.L. designed, executed, and interpreted all experiments with capzimin, screened the MBP library, and drafted the manuscript. T.Y. executed and interpreted mass spectrometry proteomic analysis with capzimin. F.P. was responsible for the initial development of Rpn11 HTS substrate and fluorescence polarization assay and design and execution of Rpn11 HTS. A.L.M. triaged Rpn11 HTS hits. C.P. executed HDAC6, MMP, hCAII, and GLO1 assay and synthesis of 3021 and 3027 (capzimin). Y.M. synthesized 3027 (capzimin). S.C., G.M., B.B., E. Sergienko, and I.P. developed the HTS assay. K.N., S.V., E. Suyama, and L.H.S. performed the HTS assay. I.P., A.B.P., E. Sergienko, and T.D.Y.C. evaluated and selected hits. K.P.C. designed, executed, and interpreted experiments with zinc reporter. A.E.P. designed and interpreted experiments with zinc reporter. S.H. designed, interpreted and oversaw the mass spectrometric studies with capzimin and revised the manuscript. S.M.C. designed and oversaw capzimin syntheses as well as aided in interpretation of metalloenzyme inhibition studies. R.J.D. designed, interpreted, and oversaw the experiments for the entire study and drafted the manuscript. Data availability. All data generated or analyzed during this study are included in this published article (and its supplementary information files) or are available from the corresponding author upon reasonable request. Competing financial interests: R.J.D. is a founder, shareholder, consultant, and member of the Scientific Advisory Board of Cleave Biosciences, which is engaged in discovery and development of drugs that target enzymes involved in ubiquitin-dependent protein degradation. S.M.C. is a co-founder, has an equity interest, and receives income as member of the Scientific Advisory Board for Cleave Biosciences and is a co-founder, has an equity interest, and is a member of the Scientific Advisory Board for Forge Therapeutics. The terms of this arrangement have been reviewed and approved by the University of California, San Diego in accordance with its conflict of interest policies.

Attached Files

Accepted Version - nihms877167.pdf

Supplemental Material - nchembio.2326-S1.pdf

Supplemental Material - nchembio.2326-S2.pdf

Supplemental Material - nchembio.2326-S3.xlsx

Supplemental Material - nchembio.2326-S4.xlsx


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October 24, 2023