内源性TLR9通过PARG-PARP1通路上调PD-L1导致肝癌免疫逃逸的机制研究

Toll样受体9激动剂单用在抗肿瘤临床试验中并没有取得预期效果，而与抗PD-1单抗联合却在黑色素瘤临床试验中产生协同抗肿瘤效果，其中机制还不明确。建立小鼠皮下瘤模型，我们发现TLR9激动剂联合抗PD-1单抗确实明显抑制小鼠肝癌生长，而TLR9单药却无此疗效，这与临床试验结果一致。体内外实验发现TLR9激活可明显上调肝癌细胞PD-L1转录水平，减少肿瘤组织内CD8+T细胞浸润。进一步，体外实验发现TLR9激活抑制PARP1表达但促进STAT3磷酸化修饰。据此提出“TLR9激活通过抑制PARP1表达，促进STAT3磷酸化从而上调PD-L1表达并影响抗肿瘤免疫效果”的假说。进一步我们将围绕TLR9/PARG/PARP1/p-STAT3/PD-L1轴展开具体机制研究，采用Co-IP、高通量质谱分析、芯片技术等方法明确TLR9激动剂单药临床效果不佳、联合免疫治疗效果显著的分子机制。

Clinical trials have shown that TLR9 agonist alone does not bring significant clinical benefits but produce synergistic antitumor effects when combining with anti-PD-1 monoclonal antibodies. However, the mechanism is unclear. By establishing the mouse subcutaneous tumor model, we found that TLR9 agonist combined with anti-PD-1 monoclonal antibody significantly inhibited the growth of mouse liver cancer, while TLR9 agonist alone could not significantly inhibit the growth of mouse liver cancer, which is consistent with the results of clinical trials. In vitro and in vivo experiments, we found that TLR9 activation could significantly increase the transcription level of PD-L1 in liver cancer cells and reduce CD8+ T cell infiltration. Further, we found that TLR9 activation inhibited PARP1 expression but promoted STAT3 phosphorylation modification. Based on these findings, we proposed that "TLR9 activation inhibits PARP1 expression and promotes STAT3 phosphorylation to up-regulate PD-L1 expression and induces immune escape". In future study, we will find out the mechanism around the “TLR9/PARG/PARP1/p-STAT3/ PD-L1 axis” by using Co-IP，high-throughput mass spectrometry analysis，chip technology to determine why the clinical outcome of the TLR9 agonist alone is not good, but producing boost anti-tumor effect when combined immunotherapy.