Inhibition of transforming activity of the ret/ptc1 oncoprotein by a 2-indolinone derivative

ret-derived oncogenes are frequently and specifically expressed in thyroid tumors. in contrast to the ret receptor, ret oncoproteins are characterized by ligand-independent tyrosine-kinase activity and tyrosine phosphorylatio n, In this study, novel synthetic arylidene 2-indolinone compounds were eva luated as inhibitors of the ret/ptc I tyrosine kinase, Four compounds inhib ited ret/ptc I activity in immunokinase assay (IC50 27 - 42 mu M) including one (1,3-dihydro-5,6-dimethoxy-3-[(4-hydroxyphenyl) methylene)-2H-indol-2- one) (Cpd I)that selectively inhibited the anchorage-independent growth of NIH3T3 transformants expressing the ret/ptc1 gene (NIH3T3(ptc1) cells). Fol lowing exposure to Cpd I, the transformed phenotype of NIH3T3(ptc1) cells w as reverted, within 24 hr, to a normal fibroblast-like morphology in adhere nt-cell culture. In these cells, the constitutive tyrosine phosphorylation of ret/ptc I, of the transducing adaptor protein shc and of a series of co- immunoprecipitated peptides became much reduced, as demonstrated by immunop recipitation/Western-blot analyses. Data presented provide additional evide nce that ret/ptc1 is directly implicated in malignant transformation, and d emonstrate the ability of Cpd I to interfere in the signal transduction pat hway constitutively activated by the ret/ptc I oncoprotein. These results c onfirm the interest of the arylidene 2-indolinone class of tyrosine-kinase inhibitors as tools for the study of ret signaling and the control of cell proliferation in ret- and ret/ptcs-associated diseases. (C) 2000 Wiley-Liss , Inc.