Structure-Based Discovery of HS34: A Highly Selective and Orally Bioavailable CDK9 Inhibitor for Triple-Negative Breast Cancer.

Triple-negative breast cancer (TNBC) is a highly aggressive subtype that lacks effective targeted therapies and exhibits strong dependence on CDK9-driven transcription. Here, we report a structure-based drug design strategy exploiting CDK9-specific conformations to discover a novel series of potent inhibitors. This approach yielded HS34, a coumarin-based CDK9 inhibitor with low-nanomolar potency and exceptional selectivity. In cellular assays, HS34 displayed potent antiproliferative activity against TNBC cells, outperforming the reference inhibitor KB-0742. Mechanistically, HS34 suppresses RNAP II Ser2 phosphorylation, leading to the downregulation of short-lived survival and oncogenic proteins such as Mcl-1 and c-Myc, and consequently inducing apoptosis and blocking EMT-associated invasion. Furthermore, HS34 exhibits favorable DMPK properties, including high oral bioavailability and metabolic stability, which align with the significant antitumor efficacy observed in an orally treated xenograft model. Collectively, these findings establish HS34 as a selective CDK9 inhibitor and demonstrate that exploiting target-specific conformational features offers an effective strategy for kinase selectivity.