Anti-tumor effect of anti-programmed death 1 combined with OK432 in mouse hepatocellular carci-noma model
Objective To observe the anti-tumor effect of anti-programmed death 1(PD-1)com-bined with OK432 in mice with hepatocellular carcinoma,and to explore its anti-tumor mechanism.Methods Subcutaneous and ascites tumor models of H22 hepatocellular carcinoma were established in 8-10-week-old C57BL/6 male mice,respectively.Tumor volume and survival were monitored.IFN-γ,CD8+T,NK,and programmed death ligand 1(PD-L1)+cells in ascites were detected by flow cytometry,ELISPOT,and enzyme linked immunosorbent assay(ELISA)to evaluate immune activation and lymphocyte infiltration in the tumor microenvironment.NK1.1 and CD8a antibodies were used to deplete NK and CD8+T cells from tumor-bearing mice.Mouse survival was monitored to evaluate the role of CD8+T and NK cells in treatment.Survival analysis was performed using the Log-rank test,and measurement data be-tween groups were analyzed using one-way ANOVA.Results The tumor volume of the OK432+anti-PD-1 group was significantly inhibited as compared with the CON group,OK432 group,and anti-PD-1 group(F=19.000,P＜0.01).The median survival time of the OK432+anti-PD-1 group(35 days)was signifi-cantly longer than that of the CON group(15 days),OK432 group(21 days),and anti-PD-1 group(21 days)(x2=21.680,P＜0.01).The number of IFN-γ spots in OK432+anti-PD-1 group(319.7±48.4)was much more than that in CON group(8.8±8.9),OK432 group(173.2±38.6)and anti-PD-1 group(172.2±18.9,F=82.330,P＜0.01).Proportion of CD8+T and NK cells in ascites of OK432+anti-PD-1 group[(14.57±2.52)％]and[(6.52±1.43)％],respectively was significantly higher than that in CON group[(1.35±0.46)％and(0.85±0.54)％],OK432 group[(8.62±2.87)％and(4.04±0.70)％]and anti-PD-1 group[(5.57±2.23)％and(2.86±1.37)％](CD8*T:F=31.240,P＜0.01 and NK:F=23.870,P＜0.01).The concentration of IFN-γ in ascites of OK432+anti-PD-1 group[(1 434±407)pg/ml]was significantly higher than that of CON group[(306±151)pg/ml]and OK432 group[(1 180±565)pg/ml]and anti-PD-1 group[(649±130)pg/ml,F=9.837,P＜0.01].The proportion of PD-L1+cells in ascites of OK432+anti-PD-1 group[(22.68±5.43)％]was significant-ly higher than that of CON group[(2.09±0.64)％],OK432 group[(15.16±3.17)％]and anti-PD-1 group[(9.03±0.97)％,F=2.090,37.670,P＜0.05].After NK or CD8+T cell depletion,the survival time of the OK432+anti-PD-1+anti-NK1.1 group and the OK432+anti-PD-1+anti-CD8a group was 32 days and 21 days,that of the CON group 18 days,and that of the OK432+anti-PD-1 group 34 days,respectively.There was no significant difference between the OK432+anti-PD-1+anti-NK1.1 group and the OK432+anti-PD-1 group(x2=0.004,P＞0.05).The difference between the OK432+anti-PD-1+anti-CD8a group and the OK432+anti-PD-1 group was statistically significant(x2=14.520,P＜0.01).Conclusion OK432 combined with anti-PD-1 can significantly inhibit tumor growth,prolong the survival time of mice,and improve the efficacy of liver cancer treatment.The anti-tumor effects of this combination were primarily contributed to the activation of CD8*T cells.
Hepatocellular carcinomaOK432Programmed death 1 antibody
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