Modeling targeted inhibition of wild type and mutated c-MET receptor

Abstract

Listen

Translate article

14010 Background: c-MET belongs to the semaphorin superfamily of signaling proteins, containing three protein families (semaphorins, plexins, c-MET and RON) that have central roles in cell signaling. The c-MET receptor tyrosine kinase is involved in regulating cell growth/proliferation, survival, angiogenesis, cell scattering, cell motility and migration. Mutations in c-MET have been identified in various human cancers including lung cancer and papillary renal cell carcinomas. c-MET mutations occur within the extracellular seven-blade β-propeller fold sema domain (E168D, L229F, S325G, N375S), juxtamembrane domain (R988C, T1010I), and kinase domain (M1268T). We hypothesized that these mutations would have differential effects on the kinase inhibition. Methods: We modeled the various c-MET mutations from different functional domains of the receptor using a G418-resistant stable Cos-7 transfection cell system to determine their effect on sensitivity to a selective c-MET kinase inhibitor SU11274. Sensitivity to SU11274 inhibition was assayed by phospho-immunoblotting using phospho- specific antibody against the major tyrosine kinase phosphorylation epitopes pY1234/1235 of the c-MET kinase in vitro. Results: First, we identified that mutations in the sema and juxtamembrane domain were activating as defined by ligand-independent constitutive receptor activation. SU11274 was capable of inhibiting ligand induced signaling through the wild-type c-MET as well as mutant c-MET receptors harboring mutations in the sema, juxtamembrane and tyrosine kinase domain. However, SU11274 inhibition of mutant c-MET was mutation-dependent, with the juxtamembrane domain mutations R988C and T1010I resulting in a receptor form that was less sensitive to SU11274. Mutations in the sema and kinase domain also resulted in varying sensitivity to inhibition by SU11274 inhibition. Conclusions: Mutations in the sema and juxtamembrane domain of c-MET result in receptor activation. The small molecule inhibitor SU11274 is active against wild type and mutated c- MET receptor. Further studies to characterize the signaling effects and the mechanism of sensitivity and resistance of c-MET mutations to specific inhibitors are crucial in the successful development of therapeutic c-MET and mutant c-MET inhibitors. No significant financial relationships to disclose.