Role of Melanoma Secreted Thrombospondin-1 on Induction of Immunosuppressive Regulatory T Cells through CD47

AIM: Regulatory T cells (Tregs) represent an important immune escape mechanism in advanced melanoma. Essential factors required for Treg induction by melanoma remain unidentified. The matricellular protein thrombospondin-1 (TSP-1) has recently been discovered to generate Tregs through its receptor, CD47. We speculated that melanoma might induce Tregs by secreting TSP-1. METHODS: We examined circulating levels of TSP-1 and Tregs in melanoma patients and normal subjects. We then examined TSP-1 secretion by metastatic melanoma cell lines in melanoma-conditioned media (MCM). The ability of MCM to induce functional Treg from normal human peripheral blood mononuclear cells (PBMCs), with or without TSP-1 or CD47 blockade was determined. Finally, we investigated the ability of recombinant TSP-1 or the CD47-binding peptide 4N1K alone to induce Tregs.RESULTS: Advanced melanoma patients had elevated levels of circulating TSP-1 which correlated with the proportion of peripheral Tregs. Furthermore, MCM contained high levels of TSP-1 and were able to induce Tregs from normal PBMCs. These Tregs were Foxp3+ and suppressed the proliferation of stimulated T responder cells. The amount of TSP-1 in the conditioned media was proportional to the degree of Treg induction and blockade of CD47 or TSP-1 abrogated this induction. However, recombinant TSP-1 or the CD47-binding peptide 4N1K were unable alone to induce Tregs. CONCLUSION: TSP-1 appears to be necessary but not sufficient alone for Treg induction by melanoma. This mechanism provides further understanding of the subversion of immune surveillance by melanoma and may provide a therapeutic target for immune therapies in this and in other malignancies.