Growth and apoptosis pathways in human cutaneous melanoma: in vitro and in vivo studies by using biological and proteomics approaches

Purpose. Melanoma is the most aggressive cutaneous cancer without effective treatment. Diagnosis is achieved very often too late and prognosis is poor. Aim of this work is to identify proteomic pathways potentially involved in melanoma aggressiveness. Materials and Methods. We analyzed 5 human metastatic melanoma cell lines and human keratinocyte and melanocyte cell lines as a control. Their proliferation and apoptotic behaviors under serum stimulation and starvation were analyzed in order to identify the most aggressive one. Melanoma cell proteome from the most aggressive cell line (A375), compared to the less aggressive one (SK mel 28), was analyzed by means of two complementary approaches: 1) multiplexed assay to measure the levels of 27 cytokines both in cell extracts and in conditioned media; 2) proteomic study through LC-MS/MS analysis of cell extracts. Data obtained were analyzed using bioinformatic analysis. Results. A375 cells were found to possess the highest growth rate both under serum stimulation and under serum starvation, while SK mel 28 cells under similar conditions were significantly less aggressive, confirmed also by invasion assays. The effect was markedly cell-density dependent, suggesting that cell-cell interaction and/or secretory signals dependent phenomena are important. Proteome analyses indicated that several proteins are differentially expressed and possibly related the aggressiveness. Some of these proteins have been identified as transport and proteasome components. The Bio-Plex analysis of the melanoma cell lines under study indicated that melanoma cells contain significantly (p<0.001) different levels of some inflammatory cytokines and angiogenic growth factors: the most significantly modified factors were IL-6, Il-7, RANTES and VEGF, suggesting a novel interesting viewpoint to explain melanoma cell aggressiveness. Conclusions. The reported results show that transport, proteasome components and an altered cytokine balance may be responsible for human melanoma aggressiveness.