In multiple endocrine neoplasia syndrome Type 2 (MEN2), medullary thyroid carcinoma (MTC) and pheochromocytoma (PC) are associated with hereditary activating germ-line mutations in the RET proto-oncogene. Also in a large percentage of sporadic MTCs and PCs, somatic RET mutations appear to be involved in tumor formation. In one single MEN2 family an extensive variety in disease expression may be observed, indicating that additional genetic events are responsible for progression of the disease towards a more aggressive phenotype. However, these additional mutations in both hereditary and sporadic MTC and PC development are largely unknown. Here, we show for the first time the presence of somatic mutations in the cell cycle regulator P18 in human RET-associated MTCs and PCs. Each of these mutations causes an amino acid substitution in the cyclin dependent kinaseinteracting region of P18 INK4C . Since these mutations partly inhibited P18 INK4C function and reduced its stability, our findings implicate P18 as a tumor suppressor gene involved in human MTC and PC development. ' 2008 Wiley-Liss, Inc.Key words: P18 INK4C; medullary thyroid carcinoma; RET; tumor suppressor gene; multistep tumorigenesis Multiple endocrine neoplasia Type 2 (MEN2) is an autosomal dominantly inherited cancer syndrome. The clinical expression of MEN2 varies, although all patients develop medullary thyroid carcinoma (MTC) or its precursor C-cell hyperplasia (CCH). MEN2 can be subdivided in MEN2A, MEN2B and familial MTC (FMTC). MEN2A is characterized by MTC, pheochromocytoma (PC), and hyperparathyroidism. MEN2B is characterized by MTC, PC and mucosal ganglioneuromas (in the colon, lips and tongue). FMTC patients solely develop MTC. MTC originates from the calcitonin producing neuroendocrine C-cells in the thyroid gland, and PC originates from the neuroendocrine chromaffin cells in the adrenal medulla. 1 MEN2 patients carry an activating germ-line mutation in the RET proto-oncogene, leading to constitutive activation of its encoded transmembrane receptor tyrosine kinase RET. The MEN2 phenotype correlates strongly with specific RET mutations. 2,3 In MEN2A the mutations affect cysteine residues in the extracellular domain of the protein. The most commonly affected Cysteine is C634. 4,5 In MEN2B, the most common mutation (in 95% of the cases) results in a Methionine to Threonine substitution at position 918 in the intracellular domain of the protein. [6][7][8] Somatic RET mutations have been detected in 23-69% of sporadic MTC, the vast majority of which has the M918T mutation, as well as in 10-15% of sporadic PC, with M918T in 50% of the cases. 9 Carriers of the same germ-line RET mutation, both related and unrelated, can develop MTC and PC at widely varying ages, suggesting that in addition to the mutated RET gene, other oncogenic events are required for the development of these tumors. The most frequently detected chromosomal alteration in MTC and PC is loss of a specific part or of the entire short arm of chromosome 1. [10][11][12] The most comm...
