Imatinib mesylate (STI-571), a c-Abl kinase inhibitor, indirectly blocks receptor tyrosine kinase activation and induces apoptosis in a human cholangiocarcinoma cell line

Abstract

Cholangiocarcinomas are highly malignant cancers arising from bile duct epithelial ceils. Therapy tor advanced cholangiocarcinoma is limited and the disease is often latal lmatinib mesylate (ST1-571), a selecnve c-Abl kmase inhibitor, has recently' been licensed for the treatment of gastrointestinal stromal tumors and chronic myelogenous leukemia However the ability of this drug to inhibit growth or induce apoptosis in human cholangiocarcinoma cells is unknown, Thus our AIM was to examine the ability of STI-571 to inhibit growth or induce apoptosis in KMCH cells, a human cholangiocarcinoma cell line. METHODS: Cell growth ,*'as examined using the MTS assay-. Apoptosis was assessed morphologically using the fluomscem dye DAPI and fluorescence microscupy. Caspase W was quantitated using CaspaTag and fluorescence microscopy. Immunoblot analysis was utillzed to examine cell lysates for tire presence of the epidermal growth factor receptor (EGFR), c-AN, and cMet (the receptor for bepatocyte growth factor), all tyrosine kinases, Phosphorylation of EGFR and c-Met was examined immunoprecipitating these receptors and performing ummmoblot analysis using phosphoprotein-speciflc antisera RESULTS: STI-571 reduced the growth of KMCH cells in a dose-dependent manner in concentrations ranging from 1020 mM The decrease in cell numbers could be accounted for an increase in cellular apoptosis, Indeed, at day 6, a concentration of 10 mM STI-571 induced a three-fold increase in apoptosis compared with control The induction of apoptosis was accompanied by a 4.5fold increase in caspase activity and abrogated the pancaspase inhibitor z-V.~(OMe)fmk ST1-571 treatment inhibited phosphorylation and therefore activation of the receptor tyrosine kinases, EGFR and c-Met. Because STI-571 does not inhibit these receptor tyrosine kniases directly, these observations implicate a mechanistic relationship between c-AN activ W and receptor tyrosine kinase activatinn. In CONCLUSION, STI-571 induces caspasedependent apoptosis in a human cholanglocarcinoma cell line. We have shown for the first time that c-AN inhibition is coupled to deactivation of the EGFR and c-Met receptor tyrosine kinases. STI-571 may be a potentially useful agent in the treatment of human cholangmcarcinoma