Abstract 4492: Covalent ligands that selectively disrupt CDK4-CDC37/HSP90 interactions in cancer cells

The Heat-Shock family of Proteins (HSPs)—which are conserved in prokaryotic and eukaryotic organisms—are molecular chaperones that regulate protein folding and stability, and in turn cellular homeostasis. In human cancers and hematological malignancies, HSPs are often upregulated. Additionally, many oncogenic proteins are HSP90 client proteins. Thus, decades of research have focused on inhibiting HSP90’s function as a potential anti-cancer therapeutic strategy. However, the clinical application of HSP90 inhibitors has been hampered by dose-limiting toxicity due to the systemic disruption and subsequent dysregulation of HSP90 client proteins. Hence, the selective targeting of individual cancer-relevant client:chaperone interactions could represent a more tractable therapeutic strategy. Using activity-based protein profiling (ABPP), we have identified a stereoselective and site-specific covalent liganding event at an isotype-restricted cysteine on CDK4 located in proximity to the interface for binding the HSP90 co-chaperone CDC37. Alterations to the CDK4/CDK6/Rb signaling pathway are common in almost all tumors, and typically arise via the overexpression of CCND1, or loss of the CDK4/CDK6-specific inhibitor p16INK4a—either event increases CDK4/CDK6 activity and leads to uncontrolled cell proliferation. Importantly, CDK4 stability is highly dependent on interactions with CDC37 and HSP90. We show that the covalent ligands selectively disrupt the CDK4:CDC37:HSP90 chaperone complex while sparing other kinase:CDC37:HSP90 interactions; the selective disruption of this chaperone complex leads to the partial degradation of CDK4 in breast cancer cells. The ligand-induced disruption of CDK4-chaperone interactions and CDK4 stability are both stereoselective (not observed with an inactive enantiomeric compound) and site-specific (not observed with a C135S mutant CDK4 protein). Given CDK4’s validated role in regulating the growth of hormone-sensitive cancer cells, compounds that can selectively disrupt chaperone regulation and stability of this kinase may have therapeutic potential as precision medicines. Citation Format: Zachary Eugene Potter, Bruno Melillo, Kristen E. DeMeester, Benjamin F. Cravatt. Covalent ligands that selectively disrupt CDK4-CDC37/HSP90 interactions in cancer cells [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 4492.