云实属植物中具有免疫抑制作用的卡山烷型二萜的发现及其作用机制研究

Many autoimmune diseases, including rheumatoid arthritis, systemic lupus erythematosus, ankylosing spondylitis, are associated with high morbidity and mortality. The immunosuppressants targeting T cells are the main drugs for autoimmune diseases. However, due to its low selectivity, many immunosuppressive agents inhibit normal and abnormal cells, cause endocrine disorders and decline the function of liver and kidney and so on. In our previous study, four plants of Caesalpinia genus containing cassane diterpene have been shown to have immunosuppressive activity and non-toxic. Sixteen new cassane diterpene, including nine compounds possessing the selective inhibition of the proliferation of activated T cell, were isolated from Caesalpinia sinensis. These results suggested that the novel selective immunosuppressants will be found form the plants of Caesalpinia genus. So, on the base of our previous works, according to the mass spectrum information, cassane diterpenes from four species of Caesalpinia plants will be systematic separated and high-content compounds will be derived through chemical synthesis. The immunosuppressive activity of all compounds will be carried out with cell models and the better activity compounds will be re-assayed with the rat model. The selectivity of better activity compounds will be study with human resting naïve T cells, IL-4 treated-activated T cells and FLS. The mechanism of active compound will be expounded through detecting the cell cycle and the expression and transcription level of the markers in T cell. At the end, a new structure, high selective immunosuppressant will be found in this project.

包括类风湿性关节炎、系统性红斑狼疮等在内的多种自身免疫性疾病致残和致死率高。以淋巴细胞为靶标的免疫抑制剂是主要治疗药物。由于选择性低，药物对正常、异常淋巴细胞均有抑制，易引起肝、肾功能下降，内分泌紊乱等副作用。课题组前期发现4种富含卡山烷型二萜的云实属植物具有免疫抑制活性，且未显示毒性；从其中1种植物获得16个新卡山烷型二萜，9个对活化T淋巴细胞增殖有抑制而对正常淋巴细胞无作用，提示卡山烷型二萜具有选择性免疫抑制活性。基于此，本课题拟开展：①质谱指导分离4种植物中的卡山烷型二萜，并合成系列衍生物；②通过体内外模型筛选活性化合物，探讨构效关系；③利用人静息初始T细胞、IL-4处理的活化T细胞等模型研究活性化合物的选择性；④从IL-2产生、CD25与CD69表达、细胞周期变化等方面揭示活性化合物在T细胞活化、增殖过程中的作用，阐明其作用机制。以上研究有望发现选择性强、高效低毒新型免疫抑制剂。

T淋巴细胞是免疫抑制剂研究的主要靶标，本课题开展了4种植物的化学成分研究，评价了它们对ConA诱导的小鼠脾脏T淋巴细胞抑制活性，并探讨了2个化合物作用行为。取得如下进展：分离得到化合物103个，化合物结构主要为卡山烷二萜、卡山烷二萜生物碱、降卡山烷二萜，新卡山烷二萜60个；推导了内酯环型卡山烷二萜、卡山烷二萜生物碱和降卡山烷可能的生物合成途径；发现两个化合物具有免疫抑制活性，并计算了选择性值；研究发现两个卡山烷二萜对ConA诱导的小鼠脾脏T淋巴细胞的IFN-γ、IL-2、IL-4、IL-6因子和细胞凋亡均有不同程度的影响并呈剂量依赖关系；其中一个活性化合物的活性在DNFB诱导的小鼠迟发型超敏反应整体动物模型得到验证。上述结果为针对T淋巴细胞进行药物设计提供了重要的活性化合物及理论基础。