Cell‐Active, Irreversible Covalent Inhibitors Targeting a Surface‐Exposed Non‐Catalytic Lysine on Aurora a Kinase by Using Squarate Chemistry

F) and demonstrated, for the first time, squarates' superior efficacy in engaging these challenging low-reactivity lysines. Amongst various AURKA CKIs, AL8 emerged as the first-in-class squarate-based, cell-active inhibitor, exhibiting excellent selectivity in both biochemical and cellular assays with long-residence time in covalently engaging endogenous AURKA. Detailed investigation of effects of leaving groups on squaric esters provided valuable insights for future development of lysine-reactive CKIs. Our finding has established squarate-containing ligands as a unique and readily tunable platform for covalent modification of surface-exposed, non-catalytic lysines in targeted kinase drug discovery.