Strategic Design of Catalytic Lysine-Targeting Reversible Covalent BCR-ABL Inhibitors

Targeted covalent inhibitors have re-emerged as validated drugs to overcome acquired resistance in cancer treatment. Herein, by using a carbonyl boronic acid warhead, we report the structure-based design of BCR-ABL inhibitors via reversible covalent targeting of the catalytic lysine with improved single-digit nanomolar potency against both wild-type and mutant ABL kinases, especially ABL^T315I bearing the gatekeeper residue mutation. We show that, by using techniques including mass spectrometry, time-dependent biochemical assays and X-ray crystallography, the evolutionarily conserved lysine can be targeted selectively. Furthermore, we show that the selectivity depends largely on molecular recognition of the non-covalent pharmacophore in this class of inhibitors, probably due to the moderate reactivity of the warhead. We report the first co-crystal structures of covalent inhibitor-ABL kinase domain complexes, providing insights into the interaction of this warhead with the catalytic lysine. We also employed label-free mass spectrometry to evaluate potential off-targets of our compounds at proteome-wide level in different cancer cell lines.