The Experimental Study Of The Efficacy Of Chemotherapy Combined With Anti-PD-1 Antibody In The Treatment Of Lewis Lung Cancer Xenograft Mice And The Mechanism Of Regulating The Immune Function

Objective: 1.Establish the Lewis lung cancer model by C57BL/6 mice.2.To observe the anti-tumor effect of chemotherapy combineds with anti-PD-1antibody.3 Efficacy of chemotherapy combined with anti-PD-1 antibody therapy in the treatment of Lewis lung cancer transplanted tumor and its effect on immune function of tumor-bearing mice.Method: The Lewis lung cancer transplanted tumor mouse model was established,the tumor volume of the mouse back was about 62.5mm3-100mm3 and the C57BL/6xenograft mices were randomly divided into four groups: Control group,chemotherapy group(CDDP),immune group(PD-1inhibitor)and combined group(CDDP +PD-1inhibitor).The size of the transplanted tumor was measured every 2 days by vernier caliper,the tumor growth curve was drawn,and the survival time of the mice was dynamically supervised,and the survival curve was drawn.After treatment three days all mices executed for harvesting the mice peripheral blood,spleen,and whole tumor,spleen made for the single cell suspension by grinding method,part of the tumor tissue used the physical and enzyme digestion methods to obtain the single cell suspension which contains the lymphocytes,part of the tumor tissue samples immediately put into the-80 ℃ refrigerator,fluorescence-activated cell sorting(FACs)was used to detect the mice peripheral blood,spleen,CD3+ CD4+ T cells,CD3+ CD8+ T cells,and the tumor microenvironment CD3+ CD8+ T cells,detect the changes of CD4+ CD25+ Foxp3+Regulatory cells(Treg cells)in the spleen by FACs and ELISA was used to detect the expression of IL-10 and INF-γ in the tumor microenvironment.Results: 1.In the Lewis lung cancer mouse xenograft model,the volume of transplanted tumors in the three treatment groups was significantly smaller than that in the control group.The volume of transplanted tumors in the combined treatment group was significantly smaller than that in the chemotherapy group and the immunotherapy group.2.The median survival of the mice in the three treatment groups was significantly longer than that in the control group,and the combination treatment group was significantly longer than the chemotherapy group and the immunotherapy group.3.The effect of various treatments on the immune function of tumor-bearing mice:(1)Peripheral blood,spleen,tumor microenvironment CD3+CD8+ T cell ratio: compared with the control group,chemotherapy group,immunotherapy group,combined treatment group the proportion of CD3 +CD8+ T cells was significantly increased.Compared with the chemotherapy group and the immunotherapy group,the CD3+CD8+ T cells proportion in the combination treatment group were significantly increased.(2)Peripheral blood,spleen,tumor microenvironment proportion of CD3+CD4+ T cells: Compared with the control group,the proportion of CD3+CD4+ T cells in the immunotherapy group,chemotherapy group and combined treatment group was significantly lower;peripheral blood and spleen in the combined treatment group The CD3+CD4+ T cells in the tissues were significantly lower than those in the chemotherapy and immunotherapy group;however,the proportion of CD3+CD4+ T cells in the tumor microenvironment of the combined treatment group was higher than that in the immunotherapy group,but similar to the chemotherapy group.(3)The proportion of spleen CD4+CD25+FOXP3+ regulatory T cells(Treg cells): Compared with the control group,the proportion of CD4+CD25+FOXP3+Treg cells in the spleen of the chemotherapy group,immunotherapy group and combination group was significantly lower;Compared with the group chemotherapy group and the immunotherapy group,the proportion of Treg cells in the the combined treatment group was significantly lower(4)The expression levels of IL-10 and INF-γ in the tumor microenvironment: Compared with the control group,the IL-10 expression level in the tumor microenvironment of the chemotherapy group,the immunotherapy group and the combined treatment group was significantly decreased,and the expression level of INF-γ was increased.The level of IL-10 in the combined group was lower than that in the chemotherapy group and the immunotherapy group,and The level of INF-γ of the combined treatment group higher.Conclusion: Chemotherapy and anti-PD-1 inhibitor enhanced the anti-tumor effect of the body immune system by down-regulating the proportion of Treg cells,the level of IL-10 and up-regulating the proportion of CD8+ T cells,the level of INF-γ,etc.Chemotherapy combined with immunotherapy improved the anti-tumor immune function.In terms of inhibiting tumor growth and prolonging the survival of xenografts,were significantly better than chemotherapy and immunotherapy alone.