To characterize proteins involved in melanoma dissemination, protein profiles from B16F10 and B16Bl6 cells were compared, as only B16Bl6 cells give pulmonary metastases after subcutaneous graft. As B16F10 and B16Bl6 cells had the same invasive capacities in vitro, we wondered whether their extracellular content could be different and correlate with their metastatic properties. We have shown that B16F10 and B16Bl6 culture cell supernatants have different modulatory effects on HT1080 fibrosarcoma cell invasion in Matrigel-coated chambers. B16Bl6 supernatants significantly enhanced HT1080 in vitro invasion as compared with B16F10 ones, suggesting differences in their protein profiles. Indeed, proteomic analysis allowed the identification of 18 differential proteins. Among the proteins with a higher concentration in B16Bl6 supernanants, lactate dehydrogenase B, M2 pyruvate kinase, cathepsin D, and galectin 1 were involved in the melanoma aggressiveness signature. Interestingly, several Gag retroviral proteins, as well as syntenin, were found mainly in the B16F10 secretome. Although its intracellular form is known as an aggressive melanoma marker, we show for the first time that syntenin was actively secreted and could reduce the invasion process, probably by protein interactions in the B16 model.