Study Of Low Dose Chemotherapy On Tumor Immune Micro-environment And Its Antitumor Effect With Anti-PD-1 Monoclonal Antibody In The Gastric Carcinoma Xenograft Model

Object: Exploring the effects of low dose chemotherapy on tumor immune microenvironment and its antitumor effect with anti-PD-1 monoclonal antibody in the gastric carcinoma xenograft model.Methods:In vitro: CCK-8 was used to analyze the inhibition effect of paclitaxel and 5-fluorouracil on MFC cell,and the PD-L1 expression on MFC cell’s membrane was analyzed by Flow cytometry.In vivo: Gastric carcinoma xenograft model was established.When the tumor reached 5-8 mm in diameter,mice were divided into 4 groups: NS group(mice received intraperitoneal of normal saline),PD-1 group(mice received intraperitoneal of anti-PD-1 monoclonal antibody),LDM groups(mice received intraperitoneal of low dose metronomic of chemotherapy);LDM+PD-1 groups(mice received intraperitoneal of low dose metronomic of chemotherapy and anti-PD-1 monoclonal antibody).Body weight,tumor volume of mice in each group were monitored every 2-3 days.Mice were killed on third day after the last anti-PD-1 monoclonal antibody.Flow cytometry was used to analyze the PD-L1 expression on MFC cell’s membrane and lymphocyte subsets in tumor tissues;The concentration of the cytokine IL-2 and IFN-γ in tumor tissue was measured using specific ELISA kit for mouse IL-2 and IFN-γ.Results:1 In vitro:1.1 Both paclitaxel and 5-fulorouracil inhibit the growth of MFC cell.The 50% inhibiting concentration(IC50)of paclitaxel and 5-Fluorouracil to MFC cell is 1.5940μg/ml、0.8810μg/ml respectively.1.2 Compare with 0 group(control group),both the low dose and the large dose of paclitaxel increase the PD-L1 expression on MFC cell’s membrane(40.15%±0.75%VS4.23%±1.81%、31.50%±3.50%VS4.23%±1.81%,P<0.05),and there is not statistical difference between the low dose and the large dose(40.15%±0.75%VS31.50%±3.50%,P<0.05);1.3 Compare with 0 group(control group),both the low dose and the large dose of 5-fluorouracil increase the PD-L1 expression on MFC cell’s membrane(26.50%±4.50%VS4.23%±1.81%、32.15%±0.15%VS4.23%±1.81%,P<0.05),and there is not statistical difference between the low dose and the large dose(26.50%±4.50% V32.15%±0.15%,P<0.05);2 In vivo:2.1 Low dose paclitaxel increases the PD-L1 expression and the effective immune cell CD8+T in tumor tissue of gastric carcinoma xenograft,but there is not significantly statistical differences in immune cell of CD4+T 、MDSC、PD-1+CD8+ when compares with the control group;Low dose 5-fluorouracil reduces the immune cell of MDSC、PD-1+CD8+T in tumor tissue of gastric carcinoma xenograft but there is not significantly statistical differences in the PD-L1 expression and the immune cell CD4+T、CD8+T in tumor tissue of gastric carcinoma xenograft when compares with the control group;2.2 In the analysis of the level of cytokine including IFN-γ and IL-2,there is not significantly statistical differences in low dose 5-fluorouracil group when compares with control group in tumor tissue of gastric carcinoma xenograft(P>0.05).2.3 Anti-PD-1 monoclonal antibody reduces the tumor growth on the model of gastric carcinoma xenograft when compares with the control group(P<0.05);But there are no significantly statistical differences between anti-PD-1 monoclonal antibody group and low dose chemotherapy combined with anti-PD-1 monoclonal antibody group(P>0.05).2.4 In the weight of mice,there is not significantly statistical differences between low dose paclitaxel and control group,But the weight of mice in the low dose 5-fluorouracil group and low dose 5-fluorouracil combined with anti-PD-1 monoclonal antibody group reduces after the chemotherapy.Conclusion:1 In vitro: Both paclitaxel and 5-fluorouracil inhibit the growth of the MFC cell;Both paclitaxel and 5-fluorouracil increase the PD-L1 expression on MFC cell’s membrane,and there is not difference between low dose and regular dose.2 In vivo:Low dose chemotherapy(low dose paclitaxel、low dose 5-fluorouracil)regulates the immune micro-environment on the gastric carcinoma xenograft model,but the antitumor effect dose not promote when combined with anti-PD-1 monoclonal antibody compares with anti-PD-1 monoclonal antibody.