PLASMA MEMBRANE SIALIDASE NEU3 SILENCING EFFECTS ON THE MOLECULAR PHENOTYPE OF MELANOMA CELLS

Human melanoma has been shown to be marked by an atypical cellular ganglioside profile and by the upregulation of plasma membrane sialidase NEU3. In this PhD thesis, I analyzed NEU3 silencing effects on the two human primary melanoma cell lines, named L3 and L6. These cell lines were stably transfected with a lentiviral vector. Previous results already demonstrated that NEU3 silenced melanoma cells reduced their migration potential and their growth in soft agar medium. Based on these data, my PhD work was focused towards the molecular features and signaling pathways alterations induced by NEU3 silencing in primary melanoma cell lines. Whole genome microarray analysis revealed the presence of differentially expressed genes above all associated with migration, motility, and control of cell death (G0 biological processes enrichment analysis). Some of these genes were validated by Real Time PCR: MAL, SEMA 3B, SEMA 3C and SEMA 5A. NEU3 silenced clones of both cell lines, 3C, 6A and 6B underwent to the upregulation of MAL, SEMA 3B and 3C. In contrast, SEMA 5A was downregulated. Different expression of markers related to epithelial mesenchymal transitions (N and E cadherin, MITF, vimentin, claudin 1, zo 1) was also revealed in NEU3 silenced clones. This proved the acquisition of a different molecular phenotype after NEU3 silencing that could explain the less motile properties. Moreover, we analyzed the activation of signaling pathways involved in these processes and we found a less activation of PI3K/AKT/PRAS40 axis and p38 kinases. Significantly, both these signaling pathways are involved in melanoma migration and differentiation control. All these results suggest that NEU3 upregulation could enhance melanoma malignancy by altering specific signaling pathways that are involved in cell motility and cell differentiation and thus NEU3 could be a novel target for treating melanoma.