Generation and Validation of Intracellular Ubiquitin Variant Inhibitors for USP7 and USP10

JOURNAL OF MOLECULAR BIOLOGY

卷:429

Article

Zhang, Wei^1,2  Sartori, Maria A.^1,2  Makhnevych, Taras^1,2  Federowicz, Kelly E.³  Dong, Xiaohui⁴  Liu, Li⁵  Nim, Satra^1,2  Dong, Aiping⁶  Yang, Jingsong⁷  Li, Yanjun⁶  Haddad, Dania^1,2  Ernst, Andreas^1,2  Heerding, Dirk³  Tong, Yufeng^6,8  Moffat, Jason^1,2  Sidhu, Sachdev S.^1,2 

[1] Univ Toronto, Banting & Best Dept Med Res, Donnelly Ctr Cellular & Biomol Res, 160 Coll St, Toronto, ON M5S 3E1, Canada

[2] Univ Toronto, Dept Mol Genet, 160 Coll St, Toronto, ON M5S 3E1, Canada

[3] GlaxoSmithKline Inc, Oncol Res & Dev, Canc Epigenet DPU, 1250 S Collegeville Rd, Collegeville, PA 19426 USA

[4] Guangdong Ocean Univ, Fisheries Coll, Dept Aquaculture, Zhanjiang 524088, Guangdong, Peoples R China

[5] Guangdong Ocean Univ, Fisheries Coll, Dept Marine Biol, Zhanjiang 524025, Guangdong, Peoples R China

[6] Univ Toronto, Struct Genom Consortium, Toronto, ON M5G 1L7, Canada

[7] GlaxoSmithKline Inc, Oncol Res & Dev, Immunooncol Combinat DPU, 1250 S Collegeville Rd, Collegeville, PA 19426 USA

[8] Univ Toronto, Dept Pharmacol & Toxicol, Toronto, ON M5G 1L7, Canada

关键词: USP7;    USP10;    p53;    inhibitor design;   

DOI  :  10.1016/j.jmb.2017.05.025

来源: Elsevier

【 摘 要 】

Post-translational modification of the p53 signaling pathway plays an important role in cell cycle progression and stress-induced apoptosis. Indeed, a large body of work has shown that dysregulation of p53 and its E3 ligase MDM2 by the ubiquitin-proteasome system (UPS) promotes carcinogenesis and malignant transformation. Thus, drug discovery efforts have focused on the restoration of wild-type p53 activity or inactivation of oncogenic mutant p53 by targeted inhibition of UPS components, particularly key deubiquitinases (DUBs) of the ubiquitin-specific protease (USP) class. However, development of selective small-molecule USP inhibitors has been challenging, partly due to the highly conserved structural features of the catalytic sites across the class. To tackle this problem, we devised a protein engineering strategy for rational design of inhibitors for DUBs and other UPS proteins. We employed a phage-displayed ubiquitin variant (UbV) library to develop inhibitors targeting the DUBs USP7 and USP10, which are involved in regulating levels of p53 and MDM2. We were able to identify UbVs that bound USP7 or USP10 with high affinity and inhibited deubiquitination activity. We solved the crystal structure of UbV.7.2 and rationalized the molecular basis for enhanced affinity and specificity for USP7. Finally, cell death was increased significantly by UbV.7.2 expression in a colon cancer cell line that was treated with the chemotherapy drug cisplatin, demonstrating the therapeutic potential of inhibiting USP7 by this approach. (c) 2017 Elsevier Ltd. All rights reserved.

【 授权许可】

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