可溶性PD-L1在肺癌免疫逃逸中的作用及其机制

The costimulatory molecule PD-1/PD-L1 convey negative regulatory signals by inhibiting T and B cell proliferation, activation and cytokine secretion, which have important biological significance in maintaining immune tolerance, terminating immune response properly and timely. Membrane-bound form and soluble PD-1/PD-L1 is closely related to the development, progression and prognosis of the malignant tumor. The study of soluble PD-L1 (sPD-L1) worldwide remains scarce. sPD-L1 and lung cancer are selected as the entry point and object of study in this project. Specific sPD-L1 ELISA detection kit developed by our group is used to reveal the characteristics of sPD-L1 expression in human peripheral blood and the mechanism of sPD-L1 formation is explored. Using confocal microscope, flow cytometry and methods for studying cell function and signal transduction, the role of sPD-L1 in regulating T and B cell function and its molecular mechanism are to be elucidated. In addition, clinical samples of lung cancer such as tissue and serum are collected for analyzing the expression of sPD-L1 in the peripheral blood of patients with lung cancer, foci of the tumor and changes in expression level before and after treatment. In addition, tumor-bearing mice model is to be establised to explore the role of the factor in lung cancer immune escape by blockage of the pathway.

协同刺激信号PD-1/PD-L1通过抑制细胞增殖、活化和细胞因子的分泌，在T、B等细胞的免疫应答中发挥负性调控作用，对有效维持免疫耐受和保证免疫应答的适时终止具有重要的生物学意义。细胞膜型和可溶性PD-1/PD-L1抑制途径与肿瘤的发生、发展及预后密切相关。国内外对可溶性PD-L1（sPD-L1）的研究尚为空白。本项目以负性sPD-L1和肺癌为切入点和研究对象，利用自主研发的特异性检测sPD-L1的ELISA试剂盒，揭示sPD-L1在人外周血中的表达特性及产生机制，并利用Confocal、FCM、细胞功能和信号传导等实验方法阐明该因子对T、B等细胞的调节作用及其分子机制。收集临床肺癌标本，分析sPD-L1在肺癌患者外周血、肿瘤局灶及治疗前后血清中的表达特性，建立小鼠肺癌模型，通过体内阻断干预研究探讨该因子在肺癌免疫逃逸中的作用。

协同刺激分子 PD-1/PD-L 在介导T、B 等细胞的免疫应答中发挥了重要的负性调控作用。膜型和可溶性PD-1/PD-L1介导的负性共刺激信号与肿瘤的发生、发展及预后密切相关。本项目以sPD-L1和肺癌为切入点和研究对象，利用自主研发的特异性检测sPD-L1的ELISA试剂盒，并利用Confocal、FCM、细胞功能和信号传导等实验方法阐明该因子对T、B等细胞的调节作用及其分子机制。通过收集临床肺癌标本，分析sPD-L1在肺癌患者外周血中的表达特性，分析该因子与肺癌发生、发展的相关性；通过T淋巴细胞与CD14+单核细胞共培养体系探讨PD-1/PD-L1抑制途径对胸腔局部T淋巴细胞的生物学功能的影响，揭示sPD-L1与PD-1/PD-L1协同刺激信号和结核性胸腔积液疾病发生、发展的关系。研究发现：1. mPD-L1+肺癌细胞中可分泌sPD-L1，sPD-L1抑制信号通过磷酸化SHP-1、p-SHP-2发挥作用，使T淋巴细胞周期抑制于G1期；2.肺癌患者外周血sPD-L1表达增高，与肺癌患者TNM临床分期及是否存在远处转移存在一定相关性，并与临床疗效呈负相关；3.低浓度顺铂联合抗PD-Ll抗体可有效促进肿瘤微环境中T细胞的增殖，增加Thl型细胞因子的分泌，提高机体抗肿瘤的能力；4.肺癌患者外周血中CD4+CD25+CD127low Treg 表面 PD-1的表达较健康人增高，且与临床分期有关；5.结核性胸腔积液中sPD-L1含量显著高于恶性组和非结核、非恶性组，反转录PCR结果显示结核性胸腔积液中PD-L1 mRNA表达增高，且与MMP-3表达水平相关，高表达PD-L1的胸腔积液单核细胞能通过PD-1/PD-L1信号抑制T淋巴细胞的增殖和活化；6.厄洛替尼可下调EGFR敏感突变NSCLC细胞株膜型及可溶性PD-L1的表达；厄洛替尼、抗PD-L1单抗联合对EGFR突变NSCLC细胞株共培养中T细胞的细胞增殖及凋亡影响大于单药；在晚期肺腺癌患者经EGFR-TKI治疗2月后，EGFR敏感突变组sPD-L1水平降低；7.通过荟萃分析发现Axin2 -148 C/T 多态性可降低肿瘤发生风险，培美曲塞为基础的双药治疗较单药在非小细胞肺癌的二线治疗中可提高PFS、ORR，对OS无影响，PD-L1还不能成为非小细胞肺癌预后的标志物。