Identification of new leads against ubiquitin specific protease-7 (USP7): a step towards the potential treatment of cancers

Sep 21, 2026

Ubiquitin-specific protease-7 (USP7) is highlighted as a key cancer drug target because it regulates multiple proteins and genes, including MDM2 and p53, thereby influencing cancer progression. A study was designed to express USP7 in a prokaryotic system and screen small molecules using biophysical techniques, especially STD-NMR. Twelve compounds were identified as interacting with USP7 and several destabilized the protein by lowering melting temperature in thermal shift assays. Molecular docking and simulation suggested binding to the substrate binding pocket, potentially blocking substrate entry. Four compounds showed anti-cancer activity against HCT116 cells, with IC50 values of 31–143 μM, down-regulating MDM2 and up-regulating p53 mRNA via qPCR. The work identifies negative USP7 modulators for further development as anti-cancer agents.

LanguageEnglish

CategoryResearch & Report

FormatPDF

Pages14

Reading Time35 min

AccessFree to download

Introduction

USP7 as a drug target in cancer

Materials and methods

USP7 expression and small-molecule screening

Biophysical, computational, and cellular assays

RSC Advances PAPER Cite this: RSC Adv., 2024, 14, 33080 Received 21st September 2024 Accepted 7th October 2024 DOI: 10.1039/d4ra06813k[rsc.li/rsc-advances](rsc.li/rsc-advances) Identiﬁcation of new leads against ubiquitin speciﬁc protease-7 (USP7): a step towards the potential treatment of cancers† Sumaira Javaid, *a Seema Zadi,a Muhammad Awais,a Atia-tul Wahab,a Humaira Zafar,*a Innokentiy Maslennikovd and M. Iqbal Choudhary*abc Ubiquitin-speciﬁc protease-7 (USP7) is an important drug target as it regulates multiple proteins and genes (such as MDM2 and p53) with roles in cancer progression. Its inhibition can hinder the function of oncogenes, increase tumor suppression, and enhance immune response. The current study was designed to express USP7 in a prokaryotic system, followed by screening of small molecules against it using biophysical methods, primarily STD-NMR technique. Among them, 12 compounds showed interaction with USP7 as inferred from NMR-based screening. These compounds further caused destabilization of USP7 by reducing its melting temperature (Tm) up to 6 °C in thermal shift assay. Molecular docking and simulation studies revealed that these compounds bind to the putative substrate binding pocket of USP7 and thus may block the entry of the substrate. Four compounds i.e., 4-hydroxydiphenyl amine (2), phenyl-(2,3,4-trihydroxyphenyl) methanone (3), 40-amino-20 ,50-diethoxy benzanilide (5), and hydroquinone (12), showed anti-cancer activity against colorectal cancerous cells (HCT116) with IC50 values in the range of 31–143 mM. These compounds also down-regulated the mRNA expression of the MDM2 gene and up-regulated the mRNA expression of the p53 gene in HCT116 cells, as studied using qPCR analysis. This study thereby identiﬁes several negative modulators of USP7 that can be studied further as potential anti-cancer agents. Introduction The ubiquitin system is central to control protein homeostasis by promoting proteasomal and lysosomal degradation and it also governs cell signaling networks by regulating protein interactions and activities. Deubiquitinases (DUBs) are enzymes that can remove ubiquitin by catalyzing the cleavage of the peptide bond between a protein and ubiquitin.1 The balance between the attachment or removal of ubiquitin from a protein is tightly regulated. However, dysregulation of any of these processes can contribute to diﬀerent pathological conditions, including cancer.2,3 The human genome expresses over 100 DUBs, which can be categorized as cysteine proteases and metalloproteases. These include six families of cysteine aDr. Panjwani Center for Molecular Medicine and Drug Research, International Center of Chemical and Biological Sciences, University of Karachi, Karachi 75270, Pakistan. E-mail: [sumairajavaid17@yahoo.com](sumairajavaid17@yahoo.com); [sumaira.javed@iccs.edu](sumaira.javed@iccs.edu); hamramalik@ [gmail.com](gmail.com); [iqbal.choudhary@iccs.edu](iqbal.choudhary@iccs.edu) bH. E. J. Research Institute of Chemistry, International Center for Chemical and Biological Sciences, University of Karachi, Karachi 75270, Pakistan cDepartment of Biochemistry, Faculty of Sciences, King Abdulaziz University, Jeddah 22252, Saudi Arabia dSchool of Pharmacy, Chapman University, Irvine, CA 92866, USA † Electronic supplementary information (ESI) available. See DOI: [https://doi.org/10.1039/d4ra06813k](https://doi.org/10.1039/d4ra06813k) proteases: ubiquitin-speci􀀁c proteases (USPs), ovarian tumor proteases (OTUs), ubiquitin C-terminal hydrolases (UCHs), the Josephin family, and the motif interacting with ubiquitin (MIU) -containing novel DUB family [MINDYs] . The family of metalloproteases includes JAB1/MPN/MOV134 (JAMMs) proteases.4–6 Among the ubiquitin-speci􀀁c proteases (USPs), USP7 has gained much interest as an anticancer drug target because of its involvement in multiple cellular pathways that regulate the expression of oncogenes and tumor suppressor genes. Moreove

FAQs

Why is USP7 considered an important target for cancer treatment?

USP7 regulates multiple proteins and genes involved in cancer progression, including MDM2 and p53. Inhibiting USP7 can suppress oncogenic functions, enhance tumor suppression, and support immune responses.

How were small molecules screened for interaction with USP7?

USP7 was expressed in a prokaryotic system, followed by screening using biophysical methods, primarily STD-NMR. Additional thermal shift assays assessed destabilization effects.

What cellular effects did the most active compounds have in HCT116 cells?

Four compounds showed anti-cancer activity against HCT116 cells and modulated gene expression: they down-regulated MDM2 mRNA and up-regulated p53 mRNA, measured by qPCR.

Sep 21, 2026

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