Specificity of alpha-cyano-beta-hydroxy-beta-methyl-N-[4-(trifluoromethoxy)phenyl]-propenamide as an inhibitor of the epidermal growth factor receptor tyrosine kinase

The epidermal growth factor receptor (EGFR) tyrosine kinase has an essentia l function for the survival of human breast canter cells, In a systematic e ffort to design potent and specific inhibitors of this receptor family prot ein tyrosine kinase (PTK) as antibreast cancer agents, we recently reported the construction of a three-dimensional homology model of the EGFR kinase domain. In this model, the catalytic site is defined by two beta-sheets tha t form an interface at the cleft between the NH2-terminal and COOH-terminal lobes of the kinase domain. Our modeling studies revealed a distinct, rema rkably planar triangular binding pocket within the kinase domain with appro ximate dimensions of 15 Angstrom x 12 Angstrom x 12 Angstrom, and the thick ness of the binding pocket is -7 Angstrom with an estimated volume of simil ar to 600 Angstrom(3) available for inhibitor binding. Molecular docking st udies had identified alpha-cyano-beta-hydroxy-beta-methyl-N-[4-(trifluorome thoxy)phenyl]-propenamide (LFM-A12) as our lead inhibitor, with an estimate d binding constant of 13 mu M, which subsequently inhibited EGFR kinase in vitro with an IC50 value of 1.7 mu M. LFM-A12 was also discovered to be a h ighly specific inhibitor of the EGFR. Even at very high concentrations rang ing from 175-350 mu M, this inhibitor did not affect the enzymatic activity of other PTKs, including the Janus kinases JAK1 and JAK3, the Src family k inase HCK, the Tec family member Bruton's tyrosine kinase, SYK kinase, and the receptor family PTK insulin receptor kinase. This observation is in con trast to the activity of a quinazoline inhibitor tested as a control, 4-(3- bromo, 4-hydroxyanilino)-6,7-dimethoxyquinazoline, which was shown to inhib it EGFR and other tyrosine kinases such as HCK, JAK3, and SYK.