The Study of NKG2D Ligand Expression and NK Cell Cytotoxicity after HDAC Inhibitors in Lung Cancer Cell Lines

4646 Background: Natural killer (NK) cell can directly kill tumor cells without the aid of other immune cells. The killing activity is determined by the balance of stimulatory signals and inhibitory signals. A signal through NKG2D receptor can overcome inhibitory signals such as major histocompatibility complex (MHC) class I, and activate NK cells. Induction of NKG2D ligand in tumor cells by some known drugs would lead to easier clinical application than genetic engineering in increasing the tumor cell’s susceptibility to NK cell killing. Histone acetylation is one of epigenetic mechanism of gene expression control. Histone deacetylase (HDAC) inhibitor showed anti-tumor effect and has been approved in some cancers. The anti-tumor mechanism of HDAC inhibitor is not fully understood. There would be other mechanisms of anti-tumor effect besides epigenetic control of gene transcription. Immunomodulatory effect of HDAC inhibitor is a possible mechanism of action. Methods: We evaluated the ability of valproate (VPA) and vorinostat (SAHA) to induce NKG2D ligand in eight NSCLC cell lines (A549, NCI-H23, NCI-H157, NCI-H358, NCI-H522, NCI-H1975, PC9, SNU2292), and NK-cell cytotoxicity assay. We also investigated the ability of NKG2D ligand induction in vivo mouse model (C57BL/6 mouse with RMA cell lines). Results: 1) We showed that NSCLC cell lines expressed NKG2D ligands with varying degree between cell lines. ULBP-2 and MICA expression was more prevalent than other NKG2D ligands at basal status. 2) VPA and SAHA increased NKG2D ligand surface expression in NSCLC cell lines. Among five NKG2D ligands, ULBP-2 and MICA were predominantly and universally induced in surface of NSCLC cell lines. 3) The surface expression of NKG2D ligands lead to increased NK cell cytotoxicity against tumor cell lines, and this increased cytotoxicity was abrogated by blocking NKG2D receptor, suggesting the increased cytotoxicity by NK cells is mediated by NKG2D signaling pathways. 4) NKG2D ligand induction was independent of apoptosis process. 5) We found that induction of NKG2D ligand occurs not only in vitro but also in vivo. Conclusion: We showed that HDAC inhibitor can kill tumor cells by inducing surface NKG2D ligand induction in NSCLC cell lines. The future research of anti-tumor effect by HDAC inhibitor should include this immune-facilitated mechanism, especially in clinical research.