能同时识别肿瘤相关抗原和痘苗病毒抗原的双TCR-T细胞亚群的鉴定及潜在应用

Accumulating evidence suggest that oncolytic virus-induced host T cell immunity plays an important role in the anti-tumour efficacy.Dissection of T cell responses against tumour cells and oncolytic virus will open new avenues to further potentiate the efficacy of oncolytic virus. Encouraging evidence in support of this application shows that intratumoral injection of Tumour-targeted replicating oncolytic vaccinia virus (TOVV) could result in a particular population of T cells: dual Cell receptor T cells specifically reacting with both tumour-associated antigens and viral proteins. Most strikingly, in vitro, the re-activated spleen cells with VV-specific peptides could effectively and specifically kill tumour cells. In this proposed project we will perform a series of in vitro, ex vivo and in vivo experiments and achieve the following five objectives: Evaluation of dual TCR T cell function on the efficacy of replicating VV for cancer treatment; Evaluation of the diversity of the dual TCR T cells specifically recognizing both VV B8R epitopes and TAAs; Screening of the epitopes of VV proteins that produce these dual TCR T cells; Evaluation of whether a particular antigen delivered by VV vector can induce dual TCR T cells against the antigen and VV and development of a new therapeutic cancer therapeutic regime by targeting dual TCR T cells. This project will provide a novel strategy to improve the efficacy of VV-based vaccination and cancer therapy.

越来越多的证据表明在肿瘤治疗过程中溶瘤病毒诱发的抗肿瘤细胞和病毒蛋白的T细胞免疫反应发挥着关键作用。解析机体对肿瘤细胞和溶瘤病毒的T细胞免疫反应将为提高溶肿瘤病毒疗效开辟新的方法。最近发现，瘤内注射肿瘤靶向复制型溶瘤痘苗病毒（VV）能产生一群特殊的T细胞亚群，这类T细胞有双T细胞受体，能同时与肿瘤相关抗原和病毒蛋白反应。更重要的是，用VV多肽激活用痘苗病毒治疗过的动物脾细胞能有效、特异地杀伤肿瘤细胞。本课题将通过一系列体外，间接体外和体内实验来评估溶肿瘤痘苗病毒在治疗肿瘤过程中所产生的双受体T细胞的功能作用；研究识别痘苗病毒B8R表位及肿瘤相关抗原的双受体T细胞的多样性；筛选能诱导产生双受体T细胞的痘苗病毒蛋白表位; 验证痘苗病毒载体表达的特殊抗原能否引发双受体T细胞；从而探讨一种新的靶向双受体T细胞的肿瘤治疗方法。本课题将为增强痘苗病毒载体介导的疫苗和溶肿瘤病毒的疗效提供新策略。

前人的研究发现肿瘤内部浸润了一些与肿瘤无关的靶向病毒的淋巴细胞，理论上如果肿瘤细胞表达这些病毒的表位肽将引起此群淋巴细胞对肿瘤的杀伤。近来有研究将病毒的抗原多肽经多次瘤内注射或者通过肿瘤特异结合抗体将病毒抗原带到肿瘤细胞表面来激起针对病毒的免疫记忆细胞对肿瘤细胞的杀伤，在动物实验中治疗效果比较明显。由于痘苗病毒(VV)是消灭天花运动中的主要天花疫苗，可以在人体内产生长期存在的免疫记忆淋巴细胞，我们推测如果肿瘤细胞可以表达痘苗病毒的表位肽将有利于肿瘤的治疗。本研究用李斯特株痘苗病毒免疫小鼠，发现在小鼠体内产生针对痘苗病毒的记忆T细胞，且病毒最强的免疫原性肽为B8R。表达B8R的DT6606肿瘤细胞在VV提前免疫的小鼠上生长明显减慢。为了进一步推动该结果的临床转化，本研究用基因治疗常用腺相关病毒（AAV）携带B8R基因感染肿瘤细胞，观察对肿瘤的治疗效果。为尽可能的减少副作用，还对AAV病毒的启动子进行改造，构建了肿瘤细胞特异表达的启动子。我们发现该方法对肿瘤治疗有一定效果，本研究第一次报道将AAV基因传递和肿瘤免疫治疗结合起来，拓展了肿瘤基因治疗的范畴。