FoxP3介导的胃癌细胞-Treg的交互作用对胃癌术前新辅助放化疗后抗肿瘤免疫应答的影响及机制

Neoadjuvant radiochemotherapy of gastric cancer is an emerging adjuvant treatment, and it increases the rate of radical surgical resection. During clinical treatments, several off-target foci or lymph nodes might be shrinking when performing radiochemotherapy to targeted cancer, which indicating that radiochemotherapy is able to change the immune status of human body, however, the underlying mechanism is unknown. Our previous research found that FoxP3 expressed in gastric cancer cells and regulatoryT cells (Treg) , and they are associated with different prognosis of gastric cancer respectively. Moreover, the cross-talk between gastric cancer cells and lymphocytes will influence the immune status and microenvironment of the cancer. Thus, this project will identify the differences of in situ antineoplastic immune response in sample of gastric cancer and peripheral circulating blood after radiochemotherapy. It will also investigate the influence about the number and function of Treg within the interaction between gastric cancer cells and Treg cells under the radiochemotherapy through co-culture in vitro, and further explore the function and mechanism of the antineoplastic immune response mediated by FoxP3. The result of this project will not only explain how the cross-talk between gastric cancer cells and Treg cells affecting antineoplastic immune response after radiochemotherapy and the mechanism behind them, but also it will provide new ideas to improve the effects of radiochemotherapy, which has important clinical significance.

胃癌新辅助放化疗是一门渐兴起的辅助治疗手段，可提高根治性手术切除率。临床上观察到对胃癌进行放化疗的同时，一些非靶区的小病灶或淋巴结也会随之缩小，提示放化疗可以改变整个机体的免疫状态，但具体机制尚未阐明。我们前期研究发现FoxP3可分别表达于胃癌细胞和调节性T细胞（Treg），具有不同的预后提示作用；且胃癌细胞和淋巴细胞的相互作用可对肿瘤免疫微环境产生影响。本项目拟检测放化疗前后胃癌组织和癌旁组织标本中以及外周循环血中免疫细胞及细胞因子数量和类型的变化；并通过体外胃癌细胞和Treg共培养实验，检测放化疗后Treg数量和功能的变化，并阐述此过程中FoxP3所参与的调节抗肿瘤免疫应答的机制。该项目研究成果不仅有利于阐释FoxP3介导的胃癌细胞-Treg交互作用对放化疗后抗肿瘤免疫应答的影响机制，而且能为提高放化疗疗效提供新思路，具有重要的临床意义。

我国胃癌发病率居第4位，死亡率居第3位。多数胃癌确诊时已是III期，不具有根治性手术治疗的指征。FoxP3 以往被认为是Treg的特异性标志，但近来发现它在肿瘤细胞中表达，具有重要的功能。放疗后可使循环中的Treg明显下降，无瘤生存时间延长。但是，另有证据发现Treg比细胞毒性T细胞更耐受离子照射，放疗后局部原位组织中Treg数目不降低是致放疗耐受的主要原因；直肠癌间质中FoxP3+Treg越低，化疗敏感性越好。我们前期研究发现胃癌组织中胃癌细胞和淋巴细胞中均表达FoxP3，而且胃癌组织标本中胃癌细胞FoxP3的表达与预后良好有关，而高密度浸润的Treg与预后差有关。胃癌细胞和外周单个核淋巴细胞（PBMC）共培养后能促进FoxP3增多，说明Treg的形成增多，反过来，Treg也通过分泌抑制性细胞因子抑制了胃癌细胞的生长。那么FoxP3过表达的胃癌细胞在放化疗中有何作用还值得探索。本研究中，我们通过回顾性比较D2胃切除术后辅助化疗或辅助化疗的局晚期胃癌患者的总生存率的差异，发现辅助放化疗能改善pIIIA和pIIIB期患者总生存（OS），且对淋巴结阳性比值<50%患者优势明显。 然而，在pIIIC期或有癌结节的患者中获益不明显。体内实验，我们首先用腺病毒转染AGS胃癌细胞，发现过表达FoxP3的胃癌能抑制生长，促进凋亡和DNA损伤，抑制迁移，进一步发现其与上皮间质转化（EMT）及CTNNB1分子相关。过表达FoxP3基因的胃癌细胞在放化疗的条件下，肿瘤抑制更明显，并能促进DNA损伤的相关分子的表达。最后，我们检测了免疫相关分子mRNA水平。发现癌组织中FoxP3的表达高于癌旁组织，而PD1、PD-L1、PD-L2的表达低于癌旁组织。说明肿瘤原位癌组织的免疫可能被抑制。免疫磁珠分选胃癌患者中CD4+的细胞，与胃癌细胞共培养。发现FoxP3过表达组淋巴细胞中Treg比率增加，而加了TGF-β1后Treg的比率降低。研究结果有助于更好地指导D2胃切除术后III期病人的个体化治疗。FoxP3在放化疗促凋亡机制中发挥重要作用。放疗作为一种局部治疗，能促进机体释放更多的抗原，也许能打破这一状态。