The Expression And Immune Modulating Function Of PD-Ls And PD-1in Extranodal NK/T-cell Lymphoma

BackgroundNon-Hodgkin’s lymphoma(NHL) is a malignant proliferative disease thatoriginates from immune cells and their precursor cells of human immune system.Extranodal NK/T-cell lymphoma（ENKTL）is a subtype of NHL. SO far, there are nostandard therapeutic outline. It has poor treatment effect and poor prognosis.Therefore, an optimal therapeutic strategy is needed.ObjectiveThe expression of PD-L1and PD-L2were detected in ENKTL cell lines (YTSand SNK-6) and pathologic tissues. Furthermore, the immune modulating function ofPD-Ls/PD-1were explored in ENKTL.MethodsNormal NK-cell and allogeneic CD8+T-cell were isolated by magnetic activatedcell sorting （MACS) from healthy volunteers (HVs). The expression of PD-L1,PD-L2mRNA were analyzed in ENKL cell lines (SNK-6, YTS) by using Real-timefluorescence quantitative PCR (RQ-PCR) and normal NK cells were used as control. Immunohistochemistry (IHC) was used to examine the expression of PD-L1, PD-L2and PD-1in the specimens of30ENKL patients and25rhinitis patients. Thecorrelation between these proteins and Clinical histopathological parameters wereanalyzed subsequently. By applying flow cytometry (FCM) to investigate theexpression of PD-1on CD4+T and CD8+T subsets in the peripheral blood of20ENKL patients and10HVs. Functional studies of SNK-6and activated CD8+T cellby PHA were performed to analyze the functional activity of PD-L1/PD-1in thetumor immunity of ENKTL with or without anti-PD-L1antibody.Results1．The expression levels of PD-L1and PD-L2mRNA in ENKTL cell linesSNK-6and YTS both were significantly higher than that in normal NK cells. Thedifferences have statistics significance (all P<0.05). PD-1expression on CD4+T andCD8+T subsets of20ENKL patients prior to therapy were both significantly higherthan10healthy volunteers (HVs)(P<0.05). Notably, PD-1expression wasdown-regulated along with chemotherapy. The proteins of PD-L1and PD-L2constitutively expressed in30ENKL specimens were significantly higher than in20rhinitis specimens (60.0%(18/30)、63.3%(19/30) VS22.2%(4/18)、16.7%(3/18), allP<0.05).2．Furthermore, the count of PD-1+Tumor-infiltrating T lymphocytes (TIL) wasnegatively correlated with both PD-L1and PD-L2expression. PD-L1and PD-L2expression were closely correlated with clinical stage(P <0.05). PD-L1expressionwas positively correlated with lactate dehydrogenase (LDH) and Ki-67level (P <0.05). Furthermore, PD-L1expression was inversely correlated with IPI(P <0.05).However, there were no significant correlation between PD-L2expression and IPI,LDH and Ki-67level(P＞0.05). However, there were no significant correlationbetween PD-L1, PD-L2expression and other Clinical histopathological parameterssuch as gender, age and the level of β2-MG.3．In functional studies, there were no significant inhibitory effect of PD-L1/PD-1on CD8+T cells cytotoxicity assessed by SNK-6cells proliferation and CD8+T cells apoptosis. But the signal pathway of PD-L1/PD-1significantly inhibit theactivity of CD8+T cells secretion Th1cytokines such as IL-2and IFN-r. And thiscould be restored at the presence of PD-L1blocking antibody.Conclusion1．PD-Ls and their receptor PD-1were aberrantly overexpressed in humanENKTL. Furthermore, there were greatly clinical significance of PD-Ls/PD-1.2．PD-L1/PD-1signal pathway could down-regulate the activity of CD8+Tlymphocyte and it’s function would be restored by blocking the signal pathway.PD-L1/PD-1is expected to be a novel target for immunotherapy of human ENKTL.