Design, Synthesis, and Pharmacological Profiling of Serine/Threonine Kinase 33 (STK33) Inhibitors for Nonhormonal Male Contraception.

Serine/threonine kinase 33 (STK33) is a male germ-specific nonhormonal contraceptive target that is both genetically and chemically validated. Building on our previously reported reversible male contraceptive inhibitor, CDD-2807, we conducted systematic structure-activity relationship (SAR) and structure-property relationship (SPR) studies on the 3-(biphenylethynyl)-1H-indazole scaffold to address key drug development liabilities, most notably poor kinetic solubility. These efforts identified CDD-3094, which exhibits a balanced profile characterized by retained nanomolar cellular potency, >100 μM kinetic solubility, comparable metabolic stability, reduced human ether-à-go-go-related gene (hERG) channel inhibition, and a favorable cellular kinase profile, with 278 of 300 kinases (92.7%) displaying <30% occupancy at 1000 nM. Leveraging X-ray crystallography of the STK33/CDD-3094 complex, we performed structure-based modifications to specifically target the primary off-target liability, CDC-like kinase 4 (CLK4). This work culminated in the discovery of compound 40, which achieved 446-fold biochemical selectivity for STK33 over CLK4. While its cellular potency is attenuated to >1 μM, this analog provides a valuable biochemical tool for investigating STK33 selectivity and highlights the challenges of balancing kinase discrimination with intracellular activity. Collectively, these studies establish a foundation for developing soluble STK33 inhibitors, facilitating further preclinical refinement of nonhormonal male contraceptives.