Cellular effects of imatinib on medullary thyroid cancer cells harboring multiple endocrine neoplasia Type 2A and 2B associated RET mutations.

J W B de Groot, I Plaza Menacho, H Schepers, L J Drenth-Diephuis, J Osinga, J Th M Plukker, Th P Links, B J L Eggen, R M W Hofstra

Department of Surgical Oncology, University Medical Center Groningen, Groningen, The Netherlands.

Surgery 2006 Jun

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Activating mutations in the RET gene, which encodes a tyrosine kinase receptor, often cause medullary thyroid carcinoma (MTC). Surgical resection is the only curative treatment; no effective systemic treatment is available. We evaluated imatinib, a tyrosine kinase inhibitor currently used to treat chronic myelogenous leukemia and gastrointestinal stromal tumors, as a potential drug for systemic treatment of MTC, in 2 MTC-derived cell lines expressing multiple endocrine neoplasia-associated mutant RET receptors. We determined RET expression and Y1062 phosphorylation using Western blot analysis and quantitative polymerase chain reaction. We determined the effects on cell proliferation by a 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide assay, and we used fluorescence-activated cell sorter analysis with annexin V/propidium iodide staining to study imatinib-induced cell-cycle arrest, apoptosis, and cell death. Imatinib inhibited RET Y1062 phosphorylation in a dose-dependent manner after 1.5 hours of exposure. After 16 hours both RET Y1062 phosphorylation and protein expression levels were affected. Dose-dependent decreases in cell proliferation of both cell lines after exposure to imatinib with inhibitory concentration of 50% levels of 23 +/- 2 micromol/L and 25 +/- 4 micromol/L were seen. These values are high, compared with those for chronic myelogenous leukemia and gastrointestinal stromal tumors. We further could show that imatinib induced cell-cycle arrest, and apoptotic and nonapoptotic cell death. Imatinib inhibits RET-mediated MTC cell growth affecting RET protein levels in vitro in a dose-dependent manner. The concentration of imatinib necessary to inhibit RET in vitro, however, makes it impossible to conclude that imatinib monotherapy will be a good option for systemic therapy of MTC.

Citation

J W B de Groot, I Plaza Menacho, H Schepers, L J Drenth-Diephuis, J Osinga, J Th M Plukker, Th P Links, B J L Eggen, R M W Hofstra. Cellular effects of imatinib on medullary thyroid cancer cells harboring multiple endocrine neoplasia Type 2A and 2B associated RET mutations. Surgery. 2006 Jun;139(6):806-14