免疫细胞在肿瘤辐射远端效应中的作用及其分子机制研究

Radiotherapy is an important strategy for tumor therapy. Clinical phenomenon showed that radiotherapy on tumor could inhibit the growth of other metastatic tumor without irradiation and hence induces abscopal effects. However, the underlying mechanism of this abscopal effect is still obscured. This project will apply a variety of tumor-bearing mice models to study the tumor radiation induced abscopal effects (RIAE) not only on the attenuation of metastatic tumor but also on the damage to radiosensitive normal organs including lung, small intestine and bone marrow, to study the occurrence and development conditions of RIAE, and to study the different roles of immune system in RIAE on metastatic tumor ad normal tissues. Based on the findings of which kind tumors are much effective in inducing abscopal responses, this project will establish a cell co-culture system containing multi-cell lines of irradiated tumor cells, immune cells, and non-irradiated tumor cells or normal radiosensitive cells to investigate the functions of immune cells on RIAE and underlying molecular mechanisms. Especially, the roles of radiation damage related signal molecules (DAMPs, including exosomes, miRNA and functional proteins) released from irradiated tumor cells in RIAE will be paid great attentions. The circulation miRNAs, protein signals, and exosomes containing miRNA and functional proteins will be detected by a series of methods including chips, mass-spectrum, RT-PCR, Western blotting and so on to know the key signals and key genetic regulators in RIAE. Furthermore, the tumor cells effective in RIAE will be reconstructed by transfecting or knocking-out these key genes and then seeded in mice to confirm their functions on RIAE in feedback. The effect of targeted enhancement of immune function on anti-tumor treatment of radiotherapy will also be explored. The results of this study are expected to deepen the understanding of the role of immune function in RIAE and will provide a new experimental and theoretical basis for radiotherapy combined with immunotherapy.

放疗是一种重要的肿瘤治疗方法，临床发现，某些原发瘤放疗能够引发远端肿瘤的消减，产生辐射远端效应，但人们对其中的机理还了解得非常有限。本项目拟通过小鼠荷瘤鼠模型研究多种肿瘤照射后对转移瘤的远端效应，对肺肠骨髓等敏感组织的损伤作用，研究辐射远端效应的发生发展条件，研究其中免疫系统对肿瘤和正常组织作用的差异性；基于能够诱发显著辐射远端效应的肿瘤，利用由受辐射肿瘤细胞、免疫细胞、肿瘤或正常细胞组成的多细胞体系，研究免疫细胞在远端效应进程中的作用机制，重点研究受辐射肿瘤细胞释放的损伤相关信号分子DAMPs（包含外泌体、miRNA和功能蛋白等）在远端效应中的作用，检测分析筛选其中的关键信号因子与调控基因，并通过构建转基因或基因敲除肿瘤细胞，利用动物实验进行反馈验证，探讨靶向增强免疫细胞活性的辐射抗肿瘤效应。研究结果有望加深对辐射远端效应免疫机制的认识，为肿瘤放疗联合免疫治疗提供新的实验和理论基础。

放射联合免疫治疗是肿瘤治疗的研究热点之一，辐射远端效应在其中具有重要作用，但对相关分子机制的了解还非常有限。本项目研究了多种肿瘤照射后对转移瘤和脾肺等正常组织的远端效应，探索了其中的分子机理。研究表明，乳腺癌放疗引起的抗肿瘤远端效应依赖于巨噬细胞的存在，受照射乳腺癌细胞释放的HMGB1通过TLR-4途径促进巨噬细胞分泌TNF-α，通过PI3K-p110γ信号通路抑制癌细胞的增殖和迁移。利用PD-L1-/-小鼠模型，我们发现辐射联合PD-L1缺陷显著促进小鼠肺癌原发瘤和转移瘤中CD8+T细胞的激活，增强cGAS-STING介导的抗肿瘤远端效应。而自噬抑制剂氯喹可增强cGAS-STING调节的IFN-β释放，进一步激活CD8+T细胞释放IFN-γ，促进辐射诱导的ATAE。另一方面，本项目研究了胸腹部照射对远隔的肺、脾、睾丸、骨髓等组织的损伤效应。发现小鼠右胸射分割照射可引起远端生殖损伤效应（RIARE），TNF-α等炎症因子在其中发挥重要作用，同时，睾丸支持细胞中YWHAZ极性蛋白介导的Rac1蛋白核移位与TNF-α/p38 MAPK的激活密切相关，首次证明YWHAZ介导的Rac1核易位在RIARE中起中心作用。另外，胸部照射还增加了骨髓细胞损伤，受辐射肺细胞释放的TNF-α可导致肝脏释放SAA，它们可通过增加ROS诱导BMSCs的DNA损伤。对这些信号通路进行抑制或采用消炎药物西米替丁进行干预，可显著降低辐射远端效应。另外，本项目还以多种体外细胞共培养模型，研究证实巨噬细胞、受辐射细胞所释放外泌体等在辐射旁效应中具有重要介导作用，内质网、线粒体等细胞器的应激反应也是辐射旁效应发生发展的重要途径，并获知了其中的一些重要信号分子。本项目的这些发现为充分利用RIAE增强肿瘤免疫放疗联合治疗疗效提供了新的策略，同时，为辐射远后不良效应的防治提供了新的科学基础。