Structural and biophysical insights into the mode of covalent binding of rationally designed potent BMX inhibitors

Abstract

This journal is © The Royal Society of Chemistry 2020. ; The bone marrow tyrosine kinase in chromosome X (BMX) is pursued as a drug target because of its role in various pathophysiological processes. We designed BMX covalent inhibitors with single-digit nanomolar potency with unexploited topological pharmacophore patterns. Importantly, we reveal the first X-ray crystal structure of covalently inhibited BMX at Cys496, which displays key interactions with Lys445, responsible for hampering ATP catalysis and the DFG-out-like motif, typical of an inactive conformation. Molecular dynamic simulations also showed this interaction for two ligand/BMX complexes. Kinome selectivity profiling showed that the most potent compound is the strongest binder, displays intracellular target engagement in BMX-transfected cells with two-digit nanomolar inhibitory potency, and leads to BMX degradation PC3 in cells. The new inhibitors displayed anti-proliferative effects in androgen-receptor positive prostate cancer cells that where further increased when combined with known inhibitors of related signaling pathways, such as PI3K, AKT and Androgen Receptor. We expect these findings to guide development of new selective BMX therapeutic approaches. ; Funded under the Royal Society (URF to G. J. L. B., URF\R\180019), FCT Portugal (iFCT to G. J. L. B., IF/00624/2015, Postdoctoral Fellowship SFRH/BPD/95253/2013 to J. D. S., project PTDC/MED-QUI/28764/2017 to J. D. S., 02/SAICT/2017 grant 28333 to T. R., doctoral studentship SFRH/BD/143583/2019 to B. B. S., and DL 57/2016/CP1451/CT0025 to J. D. S.). This project has received funding from the European Union's Horizon 2020 research and innovation programme under grant agreements No. 807281 and 702428. The authors acknowledge the Wellcome Trust (104633/Z/14/Z, to D. W. and C. V. R.), Ministerio de Economía y Competitividad (project RTI2018-099592-B-C21 to F. C.), the Royal G. and Mae H. Westaway Family Memorial Fund (A. N. K.), National Science Foundation Graduate Research Fellowship (Grant No ...