泽漆汤减少肿瘤微环境中S100A9调控G-MDSC细胞亚群抑制肺癌的分子机制研究

Myeloid-derived suppressor cells (MDSC) are heterogeneous cell populations, which play immune suppressive properties for T cells,mediating tumor immune escape. Classic traditional Chinese medicine formula Ze-Qi-Tang has definite curative effect on lung cancer . However, the molecular mechanism of inhibitive effect of Ze-Qi-Tang on lung cancer has remained unknown. Our previous research showed the survival carve of orthotopic lung cancer model mice could be significantly prolonged by Ze-Qi-Tang. Moreover,the subpopulation of G-MDSC and S100A9 gene (associating with STAT3 signaling pathway) were both down-regulated by Ze-Qi-Tang. Besides, the proliferation of CD8+T cells were also promoted at meanwhile. Nevertheless, in vitro, lung cancer cells and MDSC were not directly inhibited by Ze-Qi-Tang and the proliferation of T cells were not directly promoted by it either. Based on these work, we propose a hypothesis that in tumor microenvironment the suppressive effect of Ze-Qi-Tang on lung cancer cells via down-regulating S100A9 to inhibit G-MDSC to enhance the killing ability of CD8+T cells to reshape the tumor microenvironment. To verify the hypothesis, this topic will be around: 1 The overall analysis of tumor suppressive effect of Ze-Qi-Tang ; 2 Whether G-MDSC is the target for Ze-Qi-Tang to inhibit tumor growth; 3 The molecular mechanism of Ze-Qi-Tang via down-regulating S100A9 to inhibit G-MDSC in tumor microenvironment, taking advantage of the technology of molecular biology and molecular immunology to reveal the mechanism of tumor suppressive effect of Ze-Qi-Tang, and providing modern scientific support for the clinical treatment of lung cancer.

髓源抑制细胞（MDSC）是肿瘤微环境中对T细胞具有免疫抑制特性的异质细胞群，介导肿瘤免疫逃逸。中医经方泽漆汤在临床上对肺癌有明确疗效，但其作用机制不明。本课题组前期研究发现泽漆汤可延长肺癌原位模型小鼠生存期，下调小鼠体内G-MDSC细胞亚群数量及与STAT3通路相关的S100A9基因，增加CD8+T细胞浸润，但体外泽漆汤对肺癌细胞、MDSC及T细胞并没有直接抑制或促进增殖作用。据此提出“泽漆汤下调肿瘤微环境中S100A9抑制G-MDSC细胞亚群，从而增强CD8+T细胞杀伤活性重塑肿瘤微环境抑制肺癌细胞生长”的科学假说（图1.1）。为验证假说，本课题将围绕：1泽漆汤整体抑瘤效果分析；2泽漆汤是否以G-MDSC为作用靶点发挥抑瘤功效；3泽漆汤通过下调肿瘤微环境中S100A9以抑制G-MDSC的分子机制，拟采用分子生物、分子免疫学等技术来揭示泽漆汤抑瘤功效机理，为其临床治疗肺癌提供现代科学支持。

肺癌作为最常见的恶性肿瘤之一，在全球范围内发病率与死亡率居高不下。中医药素来擅长调控机体内环境，对肺癌的治疗具有其独特优势。在传统医学中，认为肺癌的病机以正虚、痰邪为主。基于此，选用益气化痰代表经方泽漆汤治疗肺癌，探究其通过调控肿瘤免疫而实现抑瘤功效的潜在作用机制。本课题组首先从整体层面出发，通过肺内注射法成功建立原位肺癌小鼠模型，检测泽漆汤对全免疫鼠与免疫缺陷鼠的抑瘤效果以及用药安全性，发现泽漆汤显著延长全免疫肺癌原位模型小鼠生存期、抑制其肺内荧光信号增长，但对免疫缺陷鼠无明显作用，提示泽漆汤是通过调节肿瘤微环境中的肿瘤免疫来间接发挥抑瘤效应。随后，从细胞层面进一步探究了泽漆汤对模型小鼠免疫微环境中MDSCs与T细胞的影响，发现泽漆汤能减少肿瘤微环境中G-MDSCs的数量、抑制G-MDSCs对T细胞的免疫抑制能力并显著促进G-MDSCs的凋亡，应用G-MDSCs耗竭抗体时，泽漆汤对肺癌荷瘤小鼠生存期的延长作用，对其肺内荧光信号增长的抑制作用等均被剥夺。最后，在分子层面，泽漆汤通过抑制肿瘤微环境中的STAT3/S100A9信号通路进而促进了G-MDSCs凋亡相关蛋白Bcl2与Caspase3的表达，以重塑肿瘤免疫微环境。本研究揭示了经方泽漆汤通过抑制STAT3/S100A9/Bcl2/Caspase3信号级联促进肿瘤微环境中G-MDSCs的凋亡及下调其免疫抑制活性，从而延长肺癌小鼠生存期。本项目以益气化痰治癌为理论指导，一方面为泽漆汤临床治疗肺癌提供有力的现代科学依据，另一方面也为以G-MDSCs细胞亚群为靶点的肿瘤免疫治疗提供中医药新思路。