RESUMO
Over-expression and aberrant activation of tyrosine kinases occur frequently in human cancers. Various tyrosine kinase inhibitors (TKIs) are under clinical use, but acquisition of resistance to these drugs is a major problem. Here, we studied the interaction between two drug-resistant mutants of fibroblast growth factor receptor 1 (FGFR1), N546K and V561M, and four ATP-competitive inhibitors, ponatinib, dovitinib, PD173074 and BGJ-398. Among these protein-drug systems, the only marked reduction in affinity was that of PD173074 for the V561M mutant. We also examined the interaction of these FGFR1 variants to AMP-PNP, a nonhydrolyzable analogue of ATP, and showed that N546K showed increased affinity for the ATP analogue as compared with the wild type. These findings will help to clarify the mechanism of drug resistance in mutant tyrosine kinases.
Assuntos
Inibidores de Proteínas Quinases/farmacologia , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/genética , Adenilil Imidodifosfato/metabolismo , Benzimidazóis/metabolismo , Benzimidazóis/farmacologia , Resistência a Medicamentos/genética , Fluorometria , Humanos , Imidazóis/metabolismo , Imidazóis/farmacologia , Espectroscopia de Ressonância Magnética , Modelos Moleculares , Mutação , Conformação Proteica , Piridazinas/metabolismo , Piridazinas/farmacologia , Pirimidinas/metabolismo , Pirimidinas/farmacologia , Quinolonas/metabolismo , Quinolonas/farmacologia , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/antagonistas & inibidores , Receptor Tipo 1 de Fator de Crescimento de Fibroblastos/química , Espectrometria de FluorescênciaRESUMO
Tyrosine kinases are key enzymes that play critical roles in growth signaling, the abnormal activation of which is associated with various human cancers. Activation of tyrosine kinases is mediated by tyrosine phosphorylation in the activation-loop, which transforms the catalytic domain to the active state conformation. Cancer mutations are supposed to transform the conformation of the catalytic domain into the active-form independent of the phosphorylation state of the activation-loop. Here, we report structural and biophysical analyses of cancer mutations of the tyrosine kinase domain of fibroblast growth factor receptor 1 (FGFR1). Based on the nuclear magnetic resonance analyses, phosphorylation of the activation-loop exhibited cooperative structural transition in the activation-loop, C-helix and P-loop regions, whereas cancer mutations induced structural transformation at either one or two of these regions.