TGR5通过NOD信号通路调控NSCLC细胞PD-L1表达及功能的作用机制研究

Anti-PD-1/PD-L1 immunotherapy has opened a new mode in the treatment of advanced NSCLC. However, the clinical response rate is only about 20% and the regulatory mechanisms of PD-L1 is still unclear.Our previous study found that the bile acid membrane receptor TGR5 was highly expressed in NSCLC cancer tissues, which was closely related to the poor prognosis of lung cancer patients. Moreover, TGR5 could promote the proliferation and migration of lung cancer cells in vitro and the tumor growth in vivo.Further study found that there was a significant negative correlation between TGR5 and PD-L1 expression in NSCLC tissue samples. TGR5 knockdown up-regulated the expression of PD-L1, while TGR5 over-expression down-regulated the expression of PD-L1. Next generation sequencing results showed that TGR5 affected NOD1 signaling pathway. So, the applicant speculated that TGR5 could regulate the expression and function of PD-L1 in NSCLC cells by acting on NOD1 pathway.The study will be intended to elucidate the regulatory effects of TGR5 on PD-L1 expression of NSCLC in the tumor immune microenvironment in vivo and in vitro and to explore the exact molecular mechanisms of TGR5 regulating NOD1 signaling pathway. The synergy of targeted TGR5 in anti-tumor immunotherapy will also be further discussed. Simutaneously, the relevant clinical studies will also be validated. This research will help to optimize the population with the best benefit of anti-PD-1/PD-L1immunotherapy, and provide a new marker for NSCLC immunotherapy.

抗PD-1／PD-L1免疫治疗已开启晚期NSCLC治疗的新模式，但临床反应率仅约20%，PD-L1调控机制仍不明确。申请人前期发现TGR5在NSCLC癌组织中高表达，与肺癌患者差预后相关；TGR5可促进肺癌细胞增殖、运动和体内肿瘤生长。进一步研究表明在肺癌组织中TGR5与PD-L1表达呈明显负相关；体外敲降TGR5上调PD-L1表达，而过表达TGR5则下调PD-L1表达；二代测序分析显示TGR5影响NOD1信号通路。申请人据此推测：TGR5可能通过NOD1通路调控NSCLC细胞PD-L1表达及功能。本课题拟从体内外全面阐明TGR5在肿瘤免疫微环境中对NSCLC细胞PD-L1表达的调控作用，剖析TGR5调控NOD1通路的分子机制，探讨靶向TGR5对抗肿瘤免疫治疗的增效作用，并进行临床相关验证。本研究有助于优化抗PD-1/PD-L1免疫治疗的最佳获益人群，为增效NSCLC免疫治疗提供新的靶点。

该项目从肿瘤微环境与肿瘤互动调控的动态视角出发，重点聚焦TGR5对肿瘤免疫抑制性微环境的功能作用，为拓展肺癌肿瘤免疫治疗思路，推进肺癌个体化用药策略提供重要的理论依据。该课题从TGR5与肿瘤相关巨噬细胞的相关性入手，明确了肺癌组织中浸润的TAM高表达TGR5，进一步明确了活化TGR5可下调促瘤M2型TAMs中PD-L1表达，体内实验明确TGR5-/-的BMDM解除了TAMs对CD8+T细胞的免疫抑制功能，提高了CD8+T细胞的抗肿瘤活性，提示TGR5可能通过诱导TAM促瘤表型，推动肺癌进程。因此，靶向促瘤TAM能够多环节加剧免疫抑制性微环境推动肿瘤，介导PD-1/PD-L1免疫检查点治疗耐药。我们又进一步拓展至微环境中另一重要的免疫细胞——树突状细胞，初步探讨TGR5对肿瘤免疫微环境中DCs分化及功能的调控作用，初步结果显示Triamterene作为TGR5抑制剂可促进DC细胞上MHCII的表达，提示其具有增强DC的抗原提呈能力；另外，干预小鼠骨髓来源的树突细胞（BMDC）中的TGR5后可降低其在肿瘤细胞条件培养基诱导下脂滴的生成，提示TGR5参与了BMDC中的脂质代谢调控作用，拟进一步深挖TGR5对肿瘤免疫微环境的重塑与编程作用。本课题重在探寻基于靶向TGR5增敏抗肿瘤免疫治疗PD-1/PD-L1单抗的联合用药策略，打破以PD-L1作为单一生物标记物的局限性，开发能更准确地预测临床效益和更有针对性地对肿瘤产生有效免疫应答的新的生物标记物。