Investigating the neuropilin 2/semaphorin 3F pathway in melanocytes, melanoma, and associated therapies
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INTRODUCTION: Melanoma is the most deadly skin cancer with mortality dependent on the extent and location of metastases. Lymphatic metastasis occurs early in melanoma, and tumor-associated lymphatic vessel area correlates with melanoma progression. Recently, the discovery of checkpoint inhibitors has drastically changed the treatment strategy and survival rates in melanoma. Neuropilin-2 (NRP2) is a potential common target in melanoma cells, tumor-associated lymphatic endothelial cells (LECs) and tumor-infiltrating lymphocytes (TILs). NRP2 is a cell surface receptor with competing stimulatory ligands (VEGF-A/-C) and inhibitory ligands (SEMA3F/G). AIM: The goal of this study was to investigate the role of NRP2 in both melanoma cells and the melanoma microenvironment (LECs, TILs) and to examine the effect of semaphorin 3F (SEMA3F) on the tumor cells as well as an immune modulator. RESULTS: Mouse and human melanocytes expressed NRP2 but not other vascular endothelial growth factor (VEGF) receptor tyrosine kinases in vitro and in vivo. NRP2 protein expression, as analyzed by immunohistochemistry, was upregulated in human metastatic melanoma sections. Treatment of melanoma cells in vitro with SEMA3F inhibited migration and phosphorylation of protein kinase B (AKT) but did not inhibit cell viability. SEMA3F also increased programmed cell death-ligand 1 (PD-L1) expression in melanoma. Syngeneic B16F10 melanoma did not grow in global NRP2 knockout (KO) mice but did grow in wild-type mice. In addition, mice inoculated with B16F10 were treated with SEMA3F or phosphate-buffered saline (PBS) by mini-osmotic pumps. Resulting tumors were analyzed histologically for microvessel density and presence of TILs (number and subtype). CONCLUSIONS: Expression of NRP2 protein positively correlated with melanoma progression in human patient samples. NRP2 functions differently in melanoma tumor cells than in host stromal cells (endothelial cells [ECs], LECs). In melanoma, NRP2 is not a VEGF receptor but responds to the ligand, SEMA3F. Alternately, NRP2 appears to be an important VEGF-A/-C co-receptor in tumor-associated angiogenesis and lymphangiogenesis, as demonstrated in the NRP2 transgenic mice studies. SEMA3F inhibited tumor cell migration but increased PD-L1 expression. Systemic treatment with purified SEMA3F protein in melanoma preclinical trials inhibited melanoma growth and microvessel density. Taken together, these results suggest that exploiting the NRP2/SEMA3F signaling axis may be a novel treatment strategy to be used in combination with existing immunotherapy in melanoma.