ISOLATION OF SLOW CYCLING STEM-LIKE MELANOMA SPHERES FROM mGluR1 POSITIVE MELANOMA CELL LINE

Human malignant melanoma is a highly aggressive and drug-resistant tumour of neuro-ectodermal origin. Tumor heterogeneity, undifferentiated molecular signatures, and increased tumorigenicity of melanoma subsets with embryonic-like differentiation plasticity, strongly suggests the presence and involvement of cancer a stem cells in the initiation and propagation of melanoma. Parallel to the role that normal stem cells play in organogenesis, cancer stem cells are thought to be crucial for tumorigenesis. Many investigators, indeed, propose that melanoma stem cells (MSCs) may be responsible for tumour chemoresistance, invasiveness, and neoplastic progression and that targeted abrogation of a MSCs population could therefore ultimately lead to stable remissions and perhaps cure of metastatic melanoma. However, the high percentage of cells expressing different markers of undifferentiated and partially differentiated stages that form the bulk of the melanoma tumour has made the isolation of melanoma stem cells a challenging goal. In fact, no peculiar marker of stemness has been identified so far that may consistently flag up a stem cell population different from the highly heterogenic melanoma cell population. Glutamate has been proposed to play an important role in the in biology of stem/progenitor cells (Melchiorri et al, 2007) and cancer-initiating stem cells (Nicoletti et al., 2007, Ciceroni et al, 2008). Results from previous studies suggest that the ectopic expression of the type 1 metabotropic glutamate receptor gene (GRM1) is sufficient to transform melanocytes in vitro and cause malignant melanoma in vivo (Pollock et al. 2003). In addition, it has been documented that the GRM1 gene is aberrantly expressed in about 60% of human melanomas cell lines and biopsies (Pollock et al. 2003, Ortiz P. 2007). By culturing the type 1 metabotropic glutamate receptor-positive metastatic melanoma cell line, C8161, under conditions that are known to allow the isolation of neural stem cells, we were able to isolate a highly tumorigenic, drug resistant, slow-cycling cell population that could represent a good cell model to study Melanoma resistance and recurrence both in vitro and in vivo