Discovery of potent and selective PKMYT1 inhibitors for the treatment of CCNE1-amplified cancer

PKMYT1 has recently emerged as a promising synthetic lethal target, having attracted considerable research interest over the past two years. To date, most reported inhibitors are structural analogs of RP-6306. In this study, we employed molecular dynamics (MD) simulation-guided strategies to design a novel series of selective and potent PKMYT1 inhibitors. Among these, compound A30 exhibited outstanding PKMYT1 kinase inhibitory activity (IC₅₀ = 0.003 μM) and strong antiproliferative effects in CCNE1-amplified tumor cells, demonstrating excellent selectivity at both the kinase and cellular levels. Further investigations revealed that compound A30 exerted a highly synergistic effect with gemcitabine in CCNE1-amplified Cancer models. Additionally, compound A30 inhibited colony formation in a concentration-dependent manner, induced Apoptosis, and induced S-phase cell cycle arrest. Pharmacokinetic profiling indicated that compound A30 exhibits liver microsomal stability, favorable plasma stability, minimal CYPs inhibition, and acceptable overall pharmacokinetic properties. Together, these findings establish that compound A30 represents a promising lead for the further development of selective PKMYT1 inhibitors.