We studied the behaviour of three novel human sporadic
melanoma cell lines (hmel1, hmel9, hmel11) extracted from
tumors with different degrees of
malignancy, concerning the cell signalling pathways controlled by MC1R, BRAF, NRAS and β-
catenins. The novel cell lines were compared to metastatic cell lines (HBL, LND1), wild type (wt) for MC1R and BRAF genes, that have been extensively characterised and were used as control. All the novel cell lines have silent or no MC1R mutations even though MC1R signalling is severely impaired. Conversely, they harbour BRAF mutations at the V600 residue. These mutations determine a constitutive ERK phosphorylation in all the three cell lines. Our new
melanoma cell lines were BRAF mutated in hetero- and homozygosis, even with a wild type MC1R, and unresponsive to
NDP-MSH treatment. Quantity and subcellular localization of β-
catenin were analyzed in both novel and control cell lines. In HBL and LND1 there were high levels of
beta-catenin distributed in the cytoplasm/nucleus, while in the novel
melanoma cell lines β-
catenins were less abundant and seemed to be located at the plasma membrane/cytoplasm and absent in the nucleus. We sequenced
beta-catenin cDNA for all the
melanoma cell lines, and found mutations in HBL, LND1 and hmel1, while hmel9 and hmel11 were wt. We found that
beta-catenin levels were not influenced by the RAS/RAF/MAPK pathway because inhibition with
PD98059 (a
MEK inhibitor) did not produce any effect on
beta-catenin stability and/or localization.