Active Notch1 confers a transformed phenotype to primary human melanocytes.

The importance of mitogen-activated protein kinase signaling in melanoma is underscored by the prevalence of activating mutations in N-Ras and B-Raf, yet clinical development of inhibitors of this pathway has been largely ineffective, suggesting that alternative oncogenes may also promote melanoma. Notch is an interesting candidate that has only been correlated with melanoma development and progression; a thorough assessment of tumor-initiating effects of activated Notch on human melanocytes would clarify the mounting correlative evidence and perhaps identify a novel target for an otherwise untreatable disease. Analysis of a substantial panel of cell lines and patient lesions showed that Notch activity is significantly higher in melanomas than their nontransformed counterparts. The use of a constitutively active, truncated Notch transgene construct (N(IC)) was exploited to determine if Notch activation is a "driving" event in melanocytic transformation or instead a "passenger" event associated with melanoma progression. N(IC)-infected melanocytes displayed increased proliferative capacity and biological features more reminiscent of melanoma, such as dysregulated cell adhesion and migration. Gene expression analyses supported these observations and aided in the identification of MCAM, an adhesion molecule associated with acquisition of the malignant phenotype, as a direct target of Notch transactivation. N(IC)-positive melanocytes grew at clonal density, proliferated in limiting media conditions, and also exhibited anchorage-independent growth, suggesting that Notch alone is a transforming oncogene in human melanocytes, a phenomenon not previously described for any melanoma oncogene. This new information yields valuable insight into the basic epidemiology of melanoma and launches a realm of possibilities for drug intervention in this deadly disease.

Loss-of-function fibroblast growth factor receptor-2 mutations in melanoma.

We report that 10% of melanoma tumors and cell lines harbor mutations in the fibroblast growth factor receptor 2 (FGFR2) gene. These novel mutations include three truncating mutations and 20 missense mutations occurring at evolutionary conserved residues in FGFR2 as well as among all four FGFRs. The mutation spectrum is characteristic of those induced by UV radiation. Mapping of these mutations onto the known crystal structures of FGFR2 followed by in vitro and in vivo studies show that these mutations result in receptor loss of function through several distinct mechanisms, including loss of ligand binding affinity, impaired receptor dimerization, destabilization of the extracellular domains, and reduced kinase activity. To our knowledge, this is the first demonstration of loss-of-function mutations in a class IV receptor tyrosine kinase in cancer. Taken into account with our recent discovery of activating FGFR2 mutations in endometrial cancer, we suggest that FGFR2 may join the list of genes that play context-dependent opposing roles in cancer.