SerpinB2基因在T细胞耗竭中的作用及机制研究

T cell exhaustion is a state of dysfunction that commonly occurs during chronic infections and cancer due to the persistence of antigen and inflammation，which is a main obstacle for the treatment of tumor immunotherapy. Exhausted T cell has an epigenetic map that is different from the effector and memory T cell, leading to the unsustainability of the immune checkpoint inhibitors (anti-PD1/PDL-1, anti-CTLA4) in reversing the T cells exhaustion and the the mechanism of T cell exhaustion is still unclear. Our previous studies have confirmed that leukemia-derived microvesicles (MV) can induce T cell exhaustion via miRNAs dilivery. During the induction process, Sequencing and bioinformatics analyses revealed that the NF-κB pathway related genes in the exhausted T cells were generally down regulated, and SerpinB2, serves as a negative feedback mechanism to avoid active-induced apoptosis and dysfunction, was also down-regulate and might contribute to the T cell exhaustion. Recent studies have implicated the important role of SerpinB2 in the regulation of Th1 and Th2 immune responses. Based on previous studies, we will further study the mechanism of SerpinB2 in the development of T cell exhaustion. It is emphasized that SerpinB2 can affect T cell exhaustion by regulating NF-κB pathway. The study will help to find out the underlying mechanism of T cell exhaustion in tumor microenvironment. SerpinB2 is expected to become a new therapeutic target for solving the key problem of T cell exhaustion.

T细胞耗竭是在慢性感染或肿瘤时出现的T细胞功能失调状态。近年来来大量旨在逆转T细胞耗竭的研究取得一定疗效，例如抗PD-1/PDL-1、抗CTLA4等免疫检查点阻断剂已用于治疗各类肿瘤，但是有效率低一直是这种新型免疫疗法的死穴。根本原因在于未能明确T细胞耗竭的关键机制。申请人课题组前期研究发现白血病微泡能够诱导T细胞耗竭，且对诱导过程中T细胞基因表达谱测序分析发现，耗竭T细胞NF-κB通路相关基因普遍下调，其中SrpinB2下调较为明显，对NF-κB通路具有负反馈调节作用。本课题拟在前期研究基础上，进一步研究SrpinB2在T细胞耗竭中的作用，重点证明SerpinB2通过表观遗传学修饰负反馈调控NF-κB信号通路，导致T细胞耗竭。本课题的完成，有助于发现肿瘤微环境中T细胞耗竭的深层机制，SerpinB2可作为监测T细胞功能状态的有效指标，有望成为解决T细胞耗竭这一难题的靶点。

免疫治疗是目前肿瘤领域的研究热点，尽管已在部分肿瘤展示了优异的疗效，但T细胞耗竭仍是限制免疫治疗疗效的关键卡脖子问题。申请人前期发现，微泡(MV)是一种在细胞间传递表观调控因子的新型交流方式，肿瘤来源MV能够诱导T细胞耗竭，本课题拟在前期工作基础上进一步探索SerpinB2等基因在T细胞耗竭中的作用。项目执行过程中，申请人对研究对象进行了小幅度调整，不仅聚焦单纯T细胞，而是将嵌合抗原受体T细胞(CAR-T)一并纳入研究，后者拥有更为直接的临床转化意义。通过本项目的资助，课题组首先基于长期关注的慢性髓性白血病干细胞表面的特有免疫表型分子进行了分析，选择了CD26与CD44v6两种分子分别构建了新型CAR-T产品，并在体内外验证了其对靶细胞的杀伤活性。在此基础上，证明肿瘤MV同样能够诱导经典的CD19 CAR-T以及上述新型CAR-T产品出现耗竭，导致体内外杀伤功能下降，细胞因子分泌水平下降。单细胞测序提示，肿瘤MV能够导致CAR-T细胞记忆亚型下降，效应亚型出现明显耗竭表型，其内在机制改变涉及核因子(NF-κB)水平下降，SerpinB2等上游分子水平同样下降。此外，申请人还基于代谢组学，初步探索了CAR-T细胞分泌的组胺对后续细胞因子释放综合征以及耗竭的影响。本课题的完成，有助于解析CAR-T细胞耗竭的全新机制，肿瘤MV的数目及内容物有望成为评判T细胞耗竭重要指标，为进一步促进和改善CAR-T的临床疗效提供优秀的前期基础。本项目共资助发表SCI论文6篇，包括Leukemia, Clinical and translational medicine等杂志，大于10分两篇，大于5分的1篇，明确标注受本项目资助。共毕业博士生2名，即将毕业博士1名；毕业硕士一名，形成博士论文2篇，硕士论文1篇，即将毕业博士论文1篇，团队成员基于本项目获得自然基金青年基金资助一项。