Given the importance of ubiquitin-specific protease 7 (USP7) in oncogenic pathways, identification of USP7 inhibitors has attracted considerable interest. Despite substantial efforts, however, the development of validated deubiquitinase (DUB) inhibitors that exhibit drug-like properties and a well-defined mechanism of action has proven particularly challenging. In this article, we describe the identification, optimization and detailed characterization of highly potent (IC₅₀ < 10 nM), selective USP7 inhibitors together with their less active, enantiomeric counterparts. We also disclose, for the first time, co-crystal structures of a human DUB enzyme complexed with small-molecule inhibitors, which reveal a previously undisclosed allosteric binding site. Finally, we report the identification of cancer cell lines hypersensitive to USP7 inhibition (EC₅₀ < 30 nM) and demonstrate equal or superior activity in these cell models compared to clinically relevant MDM2 antagonists. Overall, these findings demonstrate the tractability and druggability of DUBs, and provide important tools for additional target validation studies.

考虑到泛素特异性蛋白酶7（USP7）在致癌途径中的重要性，USP7抑制剂的鉴定引起了相当大的兴趣。尽管付出了巨大的努力，但是，开发出具有药物样特性和明确作用机制的经过验证的去泛素酶（DUB）抑制剂已被证明特别具有挑战性。在本文中，我们描述了高效（IC _50）的鉴定，优化和详细表征小于10 nM），选择性USP7抑制剂及其活性较低的对映体对应物。我们还首次公开了与小分子抑制剂复合的人DUB酶的共晶体结构，该结构揭示了以前未公开的变构结合位点。最后，我们报告了对USP7抑制高度敏感（EC ₅₀ <30 nM）的癌细胞系的鉴定，并证明与临床相关的MDM2拮抗剂相比，在这些细胞模型中具有同等或更高的活性。总体而言，这些发现证明了DUB的易处理性和可药物处理性，并为进行其他靶标验证研究提供了重要工具。