NK cell anti-tumor and anti-viral function can be enhanced in vitro and exploited for successful cell mediated therapy

Abstract

Background & Aim Natural Killer (NK) cells respond to infection and tumor by releasing cytokines or by up-regulating the molecule TRAIL that induces target cells to apoptosis. The aim of the study was to discover if response to infection or tumor cells can be improved by differentiating NK cells in vitro with selected cytokines. Methods, Results & Conclusion Healthy donors NK cells were differentiated with two mixes of cytokines into cytotoxic TRAIL+ or cytokine-releasing IFNγ+TNFα+ cells and phenotype assessed by Flow Cytometry. Cytotoxicity was determined by CRA. NK immune function was studied in transwell co-cultures with tumor target cells +/- Hepatitis C virus (HCV); miRNome signature was investigated by Next Generation Sequencing and cytokine profiling by Multiplex. NK cell differentiation with any of the cytokine cocktails tested induced stable expression of activating receptors NKp30, NKp44 and NKp46. By cell-cell contact, cytotoxic TRAIL+ NK cells killed with the highest efficiency both tumor and infected target cells, while cytokine-releasing IFNγ+TNFα+ NK were less functional. Surprisingly, transwell co-culture assays demonstrated that TRAIL+ NK cells consistently eradicated HCV infection by releasing soluble factors and drastically diminished the levels of the tumor biomarker ∝FetoProtein released by tumor target cells; in parallel we observed that IFNγ+TNFα+ NK were not so efficient. By comparative miRNome analysis we underpinned a number of novel miRNA that will be herein described. Notably, hierarchical clustering showed systematic variations in the miRNA expression among the different groups. Protein analysis produced pattern of soluble factors, including soluble TRAIL protein implicated in the anti-tumor and anti- viral innate response. The pathways identified in activated TRAIL+ NK cells that specifically characterize their enhanced anti-viral and anti-tumor function compared to IFNγ+TNFα+ NK studied might represent a tool to license fully functional NK cells with translational potential for clinical applications. Directed pre-activation of NK cells prior infusion into patients might highly increase NK cells potential and thus contribute to successful immunotherapy.