Abstract 6978: Discovery and characterization of BAY-805, a potent and selective DUB inhibitor of ubiquitin specific peptidase USP21

Abstract Deubiquitinases (DUBs) have recently become of high interest as some members of this protein class show potential for therapeutic interventions. The family of nearly 100 members is divided into 6 sub families. Their action is the cleavage of ubiquitin modifications from proteins, by which the function of the respective protein can change significantly. Due to this catalytic reaction all DUBs share, it is not clear so far how good specificity with potential small molecule inhibitors (SMOLs) can be achieved. Until today, only a handful of selective inhibitors were identified for a small number of protein deubiquitinases. Recent literature suggests that USP21 plays a prominent role in apoptosis, DNA repair, and signal transduction. It down-regulates TNFα-induced NF-κB activation through deubiquitinating of RIP115 and was reported to deubiquitinate RIG-I as well as STING to negatively regulate antiviral responses.16-18 Recent studies indicate the relevance of USP21 in promoting tumor development and growth, including in non-small cell lung cancer19, bladder carcinoma20, gastric cancer21, hepatocellular carcinoma22, basal-like breast cancer23, cervical cancer24, esophageal cancer25, colorectal cancer26 and pancreatic cancer27. We selected USP21 as an interesting target for oncological treatment with a focus on its immune-oncological potential. A uHTS hit finding campaign on the Bayer 4 million compound library was initiated to identify potential selective inhibitors for USP21. Selectivity was tested against 3 other USP family members and specificity by further interference assays. Binding was cross validated by thermal shift assay (TSA). Surprisingly only a very limited number of compounds passed all criteria, and one cluster was selected for optimization. The poster will show the identification of structural classes, the focus on one series and its optimization starting from the initial 12 µM HTS hit to a single digit nano-Molar probe. The probe BAY-805 shows high selectivity in biochemical assays against a larger number of USP family members and activity in a cell-based NF kB activation assay. This probe gives an opportunity to validate USP21 pharmacology under relevant conditions in cancer. Citation Format: Norbert Schmees, Fabian Göricke, Ulrike Scheib, Raphael Boehm, Namik Akkilic, Gerd Wohlfahrt, Kirstin Petersen, Victoria Vu, Leanna Smith, Niels Lindner, Joerg Weiske, Ulf Boemer, Krzysztof Brzezinka, Philip Lienau, Stefan Gradl, Hartmut Beck, Peter J. Brown, Vijayaratnam Santhakumar, Masoud Vedadi, Dalia Barsyte-Lovejoy, Cheryl Arrowsmith. Discovery and characterization of BAY-805, a potent and selective DUB inhibitor of ubiquitin specific peptidase USP21 [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 6978.