NK cell dysregulation in metastatic melanoma.

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e21078 Background: The adaptive immune response is compromised in metastatic melanoma (MM), however little is known about the protective innate immune mechanisms including Natural Killer (NK) cells. NK cells can eliminate tumor cells through cytotoxicity and enhance adaptive immunity through the production of cytokines such as IFN-gamma. We propose that NK cell dysfunction may contribute towards melanoma progression. The aim of this study was to evaluate NK cell phenotype and correlate it with their function in MM subjects. Methods: We purified NK cells from the PBMCs of 10 MM donors (MD) and 10 healthy donors (HD) controls. They were analyzed for the percentage of NK cells (CD3-CD56+) and their subsets (CD56bright/CD16- - cytokine producers; CD56dim/CD16+ - cytotoxic; CD56bright/CD16+; CD56dim/CD16). These NK cells were characterized in terms of: cytotoxicity (perforin expression), cytokine production (IFN-g), activating receptors (NKp44 and NKp30) and immunoinhibitory receptors (PD-1) by flow cytometry, prior and after 24h stimulation with IL2, IL12 and IL15. Results: In unstimulated conditions, the cytotoxic subset (CD56dim/CD16+) was the most frequent (70%) in HD NK cells; in MD NK cells, the cytotoxic subset corresponded to only 40% while an as yet uncharacterized subset (CD56dim/CD16-) was the most frequent (60%). After stimulation, the cytotoxic subset decreased in both HD and MD NK cells; in HD there was an increase of the cytokine producers’ subset from 5% to 30%, while in MD the CD56dim/CD16- subset increased to almost 90%. When functionally characterized, MD NK cells decreased perforin levels after stimulation (from 75% to 50%), differently from HD NK cells. The production of IFN-g was higher in HD NK cells (25%) than in MD NK cells (15%) following stimulation. Finally, NKp44 was decreased in MD NK cells (10%), when compared to HD NK cells (25%) and PD1 was increased in MD NK cells (30%), when compared to HD NK cells (15%). Conclusions: NK cells from MM patients have significant changes in functionality (cytotoxicity and IFN-g) and in the expression of immunoinhibitory and activating receptors. Those changes suggest that these anti-tumor innate immune cells are dysregulated, contributing to an impaired immunosurveillance in MM.