苯并恶唑衍生物PO-291抑制活化T细胞增殖与功能的机制研究  

Research on principle of benzoxazole derivative PO-291 inhibiting T cell proliferation

作　　者：王思雨 熊静 周宏 罗兴燕 刘阳 赖翼[2] WANG Si-yu;XIONG Jing;ZHOU Hong;LUO Xing-yan;LIU Yang;LAI Yi(Center for Scientific Research,Chengdu Medical College,Chengdu 610500,China;School of Laboratory Medicine,Chengdu Medical College,Chengdu 610500,China)

机构地区：[1]成都医学院科研实验中心,四川成都610500 [2]成都医学院检验医学院,四川成都610500

出　　处：《中国药理学通报》2018年第7期925-933,共9页Chinese Pharmacological Bulletin

基　　金：国家自然科学基金资助项目(No 81302786);成都医学院自然基金项目(No CYTD15-01;CYZ15-13);大学生国家创新项目(No 201513705040)

摘　　要：目的探究PO-291抑制活化T细胞增殖与功能的机制。方法免疫磁珠法纯化人外周血T细胞,抗CD3/CD28抗体或同种异型抗原活化T细胞。流式细胞术检测细胞增殖、细胞凋亡、细胞活力、CD25和CD69表达以及细胞周期;ELISA检测细胞因子IL-2、IL-4、IL-6、IL-10、IL-17和IFN-γ的分泌水平;蛋白免疫印迹检测STAT5与p70S6K的表达和磷酸化。结果 PO-291抑制抗CD3/CD28抗体或同种异型抗原活化的T细胞增殖,IC_(50)分别为(8.09±1.04)μmol·L^(-1)、(8.01±0.95)μmol·L^(-1)。在完全抑制活化T细胞增殖的浓度下,PO-291不诱导T细胞凋亡,且在浓度达到160μmol·L^(-1)时,不影响静息T细胞和PBMC的细胞存活。PO-291不影响活化T细胞表达CD25、CD69和分泌IL-2,但阻滞T细胞周期于G_0/G_1期。PO-291不影响IL-2、IL-4、IL-10,但明显抑制IFN-γ、IL-6、IL-17的分泌。PO-291不影响STAT5和p70S6K表达,但抑制STAT5磷酸化,且可增强p70S6K磷酸化。结论 PO-291通过阻断JAK3/STAT5信号通路,抑制活化T细胞增殖,提示PO-291有望成为先导化合物,为开发用于器官移植以及自身免疫性疾病的新型药物提供新的方向。Objective To investigate the immunosuppressive activity of benzoxazole derivative PO-291 in inhibiting human activated T cell proliferation and function. Methods Human T cells were isolated and purified by the immunomagnetic microbeads and activated by anti-CD3/anti-CD28 mAbs or alloantigen.Cell proliferation,the expression of CD25 and CD69,cell cycle and apoptosis were measured by flow cytometry. Secretion levels,including IL-2,IL-4,IL-6,IL-10,IL-17 and IFN-γ were determined by ELISA. The expression and phosphorylation of STAT5 and p70S6K of activated T cells were detected by Western blot. Results PO-291 significantly inhibited human T cell proliferation with anti-CD3/anti-CD28 mAbs or alloantigen stimulation without obvious cytotoxicity. PO-291 did not affect CD25,CD69 and IL-2 expression,but induced T cell cycle arrest in G_0/G_1 phase. PO-291 significantly inhibited IL-17,IFN-γ and IL-6 expression,but not IL-2,IL-4 and IL-10. PO-291 did not affect STAT5 and p70S6K expression,but inhibited STAT5 phosphorylation and enhanced p70S6K phosphorylation. Conclusions PO-291 inhibits human activated T cell proliferation by affecting the JAK3/STAT5 pathway. PO-291 represents a potential lead compound for the design and development of new immunosuppressive drugs for the treatment of organ transplantation and autoimmune diseases.

关 键 词：PO-291 免疫抑制剂 T细胞增殖 细胞因子 流式细胞术 JAK3 

分 类 号：R329.24[医药卫生—人体解剖和组织胚胎学] R329.25[医药卫生—基础医学]