Discovery of potent, selective, and orally bioavailable monocyclic pyridine-based PKMYT1 inhibitors that synergize with CHK1 inhibition in replication-stressed cancer cells.

PKMYT1 mediates the inhibitory phosphorylation of CDK1 and represents a promising synthetic lethal target in p53-deficient or CCNE1-amplified malignancies. A series of PKMYT1 inhibitors featuring a novel monocyclic pyridine core structure was discovered, exhibiting improved selectivity and enhanced drug-like properties. Among these, compound 18 demonstrated potent PKMYT1 inhibitory activity (IC50 = 2.3 nM) and high selectivity over the homologous kinase WEE1 and other related kinases, as well as good permeability and metabolic stability. In vivo pharmacokinetic studies in mice revealed that compound 18 possessed a favorable pharmacokinetic profile, including high exposure (AUC0-t = 4877 h·ng/mL) and excellent oral bioavailability (F = 75.9%). Furthermore, the combination of compound 18 with the CHK1 inhibitor prexasertib demonstrated robust synergistic antitumor activity against triple-negative breast cancer (TNBC) and ovarian cancer cells. Collectively, compound 18 emerges as a promising preclinical lead candidate for targeted cancer therapy.