本课题将重点针对CAR-T免疫治疗中存在的关键问题，构建一种新型的基于双分子识别的Dox调控的增强型IL13 CAR-T。该新型CAR-T只有在靶细胞表面同时存在HER2及IL13Rα2两种分子时才能被激活，因此可以有效降低CAR-T 脱靶所导致的副作用；该CAR-T还同时表达PD-1 DNR （dominant negative receptor, DNR）分子，降低肿瘤细胞表面分子PD-L1对CAR-T的抑制作用，从而进一步增强CAR-T杀伤肿瘤的活性。此外，双分子识别所控制的CAR-T激活及PD-1 DNR 所致的CAR-T活性增强均受Dox的负调控，因此可以降低CAR-T打靶所致的副作用。本课题将在体外测试该新型CAR-T的基础上，采用恶性脑胶质瘤的动物模型在体内进一步评估该新型CAR-T的肿瘤免疫治疗效果，从而为CAR-T在恶性脑胶质瘤及其它肿瘤的临床免疫治疗中的应用奠定基础。

Although chimeric antigen receptor T (CAR-T) therapy plays an important role in tumor immunotherapy, it has some shortcomings. The project will establish a novel anti-HER2 scFv-tTA-synNotch receptor –based, Dox regulated and activity enhanced CAR-T by solving some key issues during CAR-T mediated tumor therapy: When the killing activity of CAR-T for tumors is enhanced, its side effects are also reduced. The novel CAR-T could be activated only the simultaneous existence of two molecules, Her2 and IL13Rα2, on the surface of target cell, and regulated by Dox, which hence could reduce off-target and on-target side effects caused by CAR-T activation. The novel CAR-T expresses Dox-controllable PD-1 dominant negative receptor that could block the inhibitory signal of CAR-T activation produced by the interaction between PD-1 and PD-L1, which therefore could further enhance CAR-T activity for killing tumor. The project will further investigate the therapeutic effects of the novel CAR-T in vivo using glioblastoma animal modelon the basis of validating the biological activity of CAR-T in vitro, which would lay a foundation for the clinical application of CAR-T in glioblastoma or other tumors. The project explores the role of CAR-T immunotherapy in glioblastoma on the basis of the previously finished project, which would open a new way for the therapy of glioblastoma.