Antitumor Mechanism Of Oncolytic Viruses Regulating NK Cell Immune Checkpoint Molecules

In addition to the direct lysis of tumor cells by oncolytic viruses,the efficacy of the host's immune response to the virus is also crucial.Studies have shown that the infiltration of natural killer cells(NK)and T cells is related to the improvement of prognosis.The oncolytic virus administered in the tumor can attract immune cells,make the tumor microenvironment in the lesion develop in a favorable direction and increase the infiltration of CD8 + T cells.At present,most immunotherapy programs focus on enhancing the T cell immune response,but Recently,there has been a strong interest in the use of NK cells for treatment and intervention in tumor progression.As a kind of immune cell,NK cells express various activating and inhibiting germline-encoded receptors,so they can directly target and kill exogenous cells without MHC specificity.It can enhance the function of NK cells in tumor progression.An expected approach in immunotherapy,which aims to initiate a multi-layered immune response to achieve high-level therapeutic effects.Checkpoint inhibitors are currently one of the effective treatments for cancer.The use of therapeutic monoclonal antibodies(m Abs)can block the immunosuppressive "checkpoint" receptors,which has made unprecedented progress in controlling tumor progression.By blocking the inhibitory pathway of effector lymphocytes,the anti-tumor activity of these immune cells is enhanced.At the same time,further combining the blocking of inhibitory signals with the transmission of activation signals can improve the efficacy of immunotherapy.In many human cancers,the NKG2 A receptor is expressed on NK and T cells in the tumor bed,and studies have also found that its ligand HLA-E is often overexpressed in tumors.A study showed that in several mouse models of tumors,the expression of NKG2 A is associated with poor clinical outcomes.Therefore,targeting NKG2 A with blocking antibodies has the unique advantage of enhancing the immune response of NK cells.Using UV-inactivated oncolytic herpes simplex virus type 2(UVoHSV2)to stimulate NK cells to study the anti-tumor effect and mechanism induced by NK cells.In addition,the effect of antibody treatment against checkpoint(NKG2A)combined with UV-o HSV2 stimulation on the anti-tumor activity of NK cells was studied in vitro and in vivo.Early research by the team showed that the lymphocytes activated by UVoHSV2 had better therapeutic effects on the murine colon cancer model(CT26 model)and murine breast cancer model(4T1 model),and the tumors in the CT26 model disappeared.This article further studies the in vitro killing effect of human NK cells on BGC823 tumor cells.UVoHSV2 stimulates NK cells to release IFN-? to help kill tumor cells.At the same time,in order to further study the immune and antitumor process of NK cells,NK92 cells were selected as the model.As an activated NK cell model,NK92 has a killing effect on BGC823(E: T = 5: 1,44.76 ± 1.84)and the killing effect of UVoHSV2 stimulated NK92 cells against BGC823 and A549 is significantly enhanced(E:T=5:1,69.64±1.29),the release of IFN-?(223.86±4.15pg/ml)of NK92 cells after UVoHSV2 stimulation similar to NK cells increased.The full-length gene of TLR2 protein was obtained in vitro.A plasmid vector expressing TLR2 was constructed.TLR2 protein was over-expressed by electrotransfection of NK92 cells and the TLR2 protein was blocked with Anti-TLR2 antibody.It was verified that UVoHSV2 activated NK92 cells through TLR2 protein and enhanced IFN-? release and further enhance the killing of NK92 cells to BGC823 tumor cells.Western blot method was used to verify that TLR2 protein can induce an increase in the level of p65 phosphorylation in NK92 cells.When the NF-?B inhibitor BYA 11-7082 was used to block the NF-?B signaling pathway of NK92 cells,the phosphorylation level decreased At the same time,BYA 11-7082 also reduced the final IFN-? release from(573.52 ± 4.38 pg / ml)to(263.87 ± 6.48 pg / ml),indicating that UVoHSV2 activates the NF-?B signaling pathway through the TLR2 protein.At the same time,we found that UVoHSV2 can increase the expression of NKG2 A on NK cells.After Anti-NKG2 A antibody blocked the function of NKG2 A protein,the ability of NK cells and NK92 cells to kill BGC823 cells in vitro was enhanced.Among them,UVoHSV2 stimulated the killing rate in NK92 group From(54.36 ± 1.93)to(68.07 ± 1.32),BGC823 cell tumors were used to establish nude mouse tumor models.Anti-NKG2 A antibody incubation enhanced the antitumor activity of NK92 cells stimulated by UVoHSV2.The results show that UVoHSV2 can effectively activate NK cells to enhance their antitumor activity in vitro,and release IFN-? through the Toll-like receptor 2(TLR2)/ NF-?B signaling pathway.Further UVoHSV2 stimulation upregulated the checkpoint molecule NKG2 A.Anti-NKG2 A antibody treatment can further enhance the in vitro and in vivo antitumor effects of UVoHSV2-stimulated NK92 cells.