HGF/c-Met通路激活诱导PD-L1表达在非小细胞肺癌免疫逃逸中的作用及机制研究

It is urgent to identify the subgroup of lung cancer that most likely to benefit from anti-PD-1 antibody immunotherapy. Our previous study demonstrated that the expression of PD-L1 is associated with the efficacy of anti-PD-1 antibody. And EGFR mutation could induce the expression of PD-L1, while EGFR-TKI could down-regulate the expression of PD-L1 in mutant sensitive cells. We also found lung cancer cells harboring c-Met over-expression were resistant to EGFR-TKI and expressed the higher expression of PD-L1. So, whether or not c-Met regulate PD-L1? Its mechanism and function remain poorly understood. The preliminary experiment found activation of HGF/c-Met pathway which is one of the resistant mechanisms of EGFR-TKI，could positively regulate the expression of PD-L1. Therefore, we assume that activation of HGF/c-Met could induce up-regulation of PD-L1, which mediates immune escape in lung cancer. In this study, we will investigate the correlation and the detailed regulatory mechanism between c-Met and PD-L1 expression in the tissue and cells of lung cancer. Moreover, we will investigate the effect of c-Met-induced PD-L1 on immunologic function and the efficacy of anti-PD-1/PD-L1 treatment to lung cancer with c-Met amplification in co-culture system in vitro and in mouse models of lung cancer with c-Met amplification. This project will provide the new therapeutic strategy for EGFR-TKI resistant lung cancer with c-Met amplification.

寻找抗PD-1抗体免疫治疗在肺癌中的优势人群亟待解决。我们前期研究显示肺癌中PD-L1表达与抗PD-1抗体疗效相关。EGFR突变可诱导PD-L1表达，而EGFR-TKI可下调突变敏感细胞的PD-L1。我们近期发现c-Met过表达肺癌细胞对EGFR-TKI耐药且PD-L1表达升高，那么肺癌中c-Met是否调节PD-L1，其机制和功能仍不清楚？预实验发现EGFR-TKI耐药机制中HGF/c-Met通路激活可正性调控PD-L1表达。我们假设：HGF/c-Met通路激活诱导PD-L1上调介导肺癌免疫逃逸作用。本课题将在肺癌组织和细胞中研究c-Met与PD-L1表达的相关性及调控机制。通过体外共培养体系及c-Met扩增肺癌小鼠模型研究c-Met过表达诱导PD-L1对免疫功能的影响及阻断PD-1/PD-L1通路治疗c-Met扩增肺癌的疗效，为c-Met扩增EGFR-TKI耐药肺癌患者提供新的治疗策略。

如何筛选免疫检查点抑制剂(PD-1/PD-L1抗体)治疗在非小细胞肺癌（NSCLC）中的优势人群亟待解决。我们前期研究显示PD-L1表达受癌基因调控，那么NSCLC中癌基因如c-Met或KRAS是否调节PD-L1，其调节机制和对T细胞功能的影响仍不清楚？.本项目主要研究NSCLC中HGF/c-Met通路和肺腺癌中KRAS突变对PD-L1的表达调控和对下游信号通路的影响。并且研究了c-Met或KRAS过表达的肺癌细胞与DC-CIK细胞共培养下，c-Met或ERK的抑制剂和PD-1抗体分别对T细胞凋亡的影响，及对c-Met和KRAS过表达的NSCLC细胞增殖的影响。 .研究结果:216例NSCLC患者基线时肿瘤组织的PD-L1和c-Met蛋白表达阳性率分别为50.9%和21.2%。c-Met阳性比c-Met阴性患者的PD-L1的HSCORE高。21例患者EGFR-TKI耐药后肿瘤组织中c-Met与PD-L1的HSCORE均高于EGFR-TKI耐药前。NSCLC细胞中c-Met可在转录水平调控PD-L1。在c-Met表达的H1993细胞和DC-CIK共培养体系中，PD-1抗体和c-Met抑制剂可减少CD3+T细胞凋亡。实时监测在H1993细胞和DC-CIK体外共培养体系中用PD-1抗体或c-Met抑制剂可降低H1993细胞增殖指数。我们研究还证明了在肺腺癌组织和细胞系中PD-L1表达与KRAS突变呈正相关。KRAS调控PD-L1表达是通过p-ERK信号而不是p-AKT通路。在DC-CIK与KRAS过表达细胞共培养中，KRAS过表达或突变可使DC-CIK细胞系中CD3+T细胞的凋亡率增加，从而使肺癌细胞发生免疫逃逸。PD-1抗体或ERK抑制剂通过阻断PD-1/PD-L1通路可逆转CD3+T细胞的凋亡，而使用AKT抑制剂却不能逆转上述过程。在体外DC-CIK细胞与KRAS突变肺癌细胞共培养体系中，抑制PD-1/PD-L1通过恢复T细胞的抗肿瘤免疫力，可减少KRAS突变肺癌细胞的增殖率。.本项目为EGFR-TKI耐药中c-Met扩增的NSCLC患者或KRAS突变的肺腺癌患者使用PD-1/PD-L1抗体提供理论指导。ERK抑制剂通过PD-1/PD-L1通路恢复T细胞的免疫力，可能是KRAS突变肺癌患者新的治疗药物。但PD-1抗体和ERK抑制剂的联合使用目前在体外未显示出优势。