Runx3抑制GATA3活性改善CAR-T细胞在实体瘤微环境耗竭的研究

Exhaustion plays a vital role restricting CAR-T cells effect targeting solid tumors and is one of the key points in CAR-T area. Our team has already conducted a clinical trial of CAR-T cells targeting CEA positive carcinoma and confirmed CAR-T cell infiltration in tumor tissues. These resident T cells were dysfunction and limited of effect. In prior experiments, we overexpressed Runx3 and observed revised exhaustion with enhanced cytokine expression including IL-2 and IFN-γ. Series of researches have convinced the positive regulation of transcriptional factor Runx3 in T cells especially in differentiation and tumor residency while the mechanism of regulating exhaustion remains unclear. It is evidenced that Runx3 combined with GATA3 and attenuated the transcriptional activity during T cell differentiation. GATA3 has been revealed promoting T cell exhaustion by inhibiting cytokines expression including IL-2 and IFN-γ. Thus, we suppose that Runx3 may improve T cell exhaustion through attenuating GATA3 activity. In order to validate this hypothesis, we plan to control expression of Runx3 in CAR-T cells and administrated series of experiments focusing on GATA3, exhaustion and cell effect in vitro and vivo. The result is a potential method elevating CAR-T cell effect targeting solid tumors.

细胞耗竭是制约CAR-T细胞在实体瘤疗效的重要原因之一，是目前该领域研究的重点。本研究团队前期开展CAR-T细胞治疗CEA阳性肿瘤的临床试验，但治疗效果有限。我们探究发现CAR-T细胞在肿瘤组织存在浸润，但其表现耗竭细胞特征。课题组预实验发现CAR-T细胞过表达Runx3可提高细胞因子表达而改善其耗竭，研究已证明Runx3在调控T细胞分化、肿瘤定植的相关机制，但其调控耗竭的机制未见报道。文献报道Runx3可通过直接结合GATA3并减弱其转录活性而调控T细胞分化，而GATA3是T细胞重要转录因子，被证实通过抑制IL-2，IFN-γ等细胞因子表达而促进T细胞耗竭。因此我们提出科学猜想：Runx3通过抑制GATA3转录活性改善CAR-T细胞耗竭。本项目拟在体内体外模型中调控Runx3表达并研究GATA3、耗竭相关表型及细胞功能变化从而论证猜想。研究结果可为提升CAR-T细胞在实体瘤疗效提供依据。

功能不足是CAR-T细胞在实体肿瘤应用受限的重要原因，如何提高CAR-T细胞的功能是亟需解决的科学问题。本研究依据申报计划，从Runx3改善CAR-T细胞的功能入手，通过在CAR-T上过表达RUNX3，发现RUNX3在成熟的T细胞上不会影响CD4CD8表型的分化，同时还可以维持CAR-T细胞的低分化状态，从而改善CAR-T细胞在抗原刺激下引起的耗竭。同时RUNX3还可以保留CAR-T细胞的效能，另外降低CAR-T发挥功能过程中细胞因子的释放，改善了CAR-T在体内的安全性。此外，本研究通过延伸，还从T细胞正常发育、分化、发挥对病毒和肿瘤细胞的杀伤功能出发，寻找对应的关键内源性分子CD27，证实添加CD27信号的CAR-T细胞在体外试验种具有较强的增殖、细胞因子释放及长期存续能力。本课题的研究对于CAR-T细胞功能提升提供了思路。