VEGFR2抑制剂阿帕替尼在肺癌中对CD8^(+)T细胞功能影响的机制研究

Mechanistic Study of the Effect of VEGFR2 Inhibitor Apatinib on CD8^(+)T Cell Function in Lung Cancer

目的探索VEGFR2抑制剂阿帕替尼在肺癌中对CD8^(+)T细胞功能的影响及可能的机制。方法于2021年1—10月开展研究,采集1名健康志愿者外周血分离PBMC,再分离活化T细胞,分别使用IL-2、OKT3、阿帕替尼处理,使用ELISA检测CD8^(+)T细胞中IL-2,IFN-γ,PD-1,LAG-3,TIM-3,CLTA-4、NFAT1等的表达。使用CCK8检测阿帕替尼对A549细胞增殖的影响。结果IL-2、OKT3和阿帕替尼协同促进IL-2、INF-γ的表达,协同显著抑制PD-1、LAG-3、TIM-3的表达,协同显著抑制NFAT1表达,差异有统计学意义(P<0.001)。IL-2和OKT3有助于淋巴细胞杀伤肿瘤细胞,差异有统计学意义(t=-0.17,P=0.003)。IL-2、OKT3和阿帕替尼协同可以显著提高杀伤肿瘤细胞的水平,差异有统计学意义(t=-10.98,P<0.001)。结论阿帕替尼逆转T细胞失能与耗竭的关键在于诱导NFAT1的低量表达;NFAT1低量表达时,主要发挥激活靶基因IL-2表达、减少免疫耗竭失能相关的靶基因表达的作用,减少T细胞失能与耗竭,促进T细胞的活化。

Objective To explore the effect of VEGFR2 inhibitor apatinib on CD8^(+)T cell function in lung cancer and the possible mechanisms.Methods The study was conducted from January to October 2021,in which PBMC were collected from peripheral blood isolated from one healthy volunteer,and then activated T cells were isolated and treated with IL-2,OKT3,and apatinib,respectively,and the expression of IL-2,IFN-γ,PD-1,LAG-3,TIM-3,CLTA-4,and NFAT1 in CD8^(+)T cells were detected using ELISA.The effect of apatinib on the proliferation of A549 cells was detected using CCK8.Results IL-2,OKT3 and apatinib synergistically promoted IL-2,INF-γexpression and synergistically and significantly inhibited PD-1,LAG-3,expression and synergistically significantly inhibited NFAT1 expression,the difference was statistically significant(P<0.001).IL-2 and OKT3 contributed to tumor cell killing by lymphocytes,the difference was statistically significant(t=-0.17,P=0.003).Synergy of IL-2,OKT3 and apatinib significantly increased the level of tumor cell killing,the difference was statistically significant(t=-10.98,P<0.001).Conclusion The key to reversing T cell dysfunction and depletion by apatinib lies in the induction of low NFAT1 expression;when NFAT1 is expressed at low levels,it mainly plays the role of activating the expression of target gene IL-2 and reducing the expression of target genes related to immune depletion dysfunction,reducing T cell dysfunction and depletion and promoting T cell activation.