南蛇藤素和扁蒴藤素显著下调HLA—B^＊2705启动子的活性

Caelastrol and pristimerin derived from Leigongteng inhibit promoter activity of HLA - B^ ＊ 2705 gene

目的:利用高通量药物筛选方式,为寻找潜在的脊柱关节炎(SpA)新的治疗药物提供理论依据。方法:选取12264小分子化合物分别使用293T-HLA-B27和HeLa-HLA-B27稳定细胞株,观察对HLA-B*2705启动子有调节作用的化合物,筛选能够下调HLA-B*2705启动子活性的阳性化合物:启动子活性>150%为激动剂,启动子活性<60%为抑制剂。并且对进一步筛选出的抑制剂进行细胞毒性试验及半数抑制浓度/半数效应浓度(IC50/EC50)检测,筛选出具有较好剂量-效应的阳性化合物。结果:(1)使用293T-HLA-B27细胞第1次筛选出624种阳性化合物,阳性率为5·1%;(2)使用HeLa-HLA-B27细胞株对上述624种化合物进行再次筛选,有70种化合物再次显示出对B*2705启动子活性的增强或者抑制作用;(3)进行EC50/IC50检测的40种化合物中,6种化合物为抑制剂,表现出较好的剂量-效应曲线,其中南蛇藤素和扁蒴藤素均为雷公藤类衍生物。结论:南蛇藤素和扁蒴藤素可下调HLA-B27表达,提示在今后针对HLA-B*2705相关的SpA患者治疗中,它们可能是值得研究的潜在有效化合物。

AIM ： To screen the effective chemicals, which can suppress the promoter activity of the HLA - B ^＊ 2705 gene as potential therapeutic agents. METHODS ： The HeLa - HLA - B27, 293T - HLA - B27 stable transfectants were used to monitor the effect of 12 264 chemicals through high throughput screening （HTS）. Chemicals which regulates HLA - B ^＊ 2705 promoter activity more than 150% or less than 60% were picked out for the further IC50/EC50 and cell viability detection. RESULTS： （ 1 ） The primary screening used by 293T - HLA - B27 stable transfectant yielded about 5.1% hits which either suppressed （556 chemicals） or enhanced （68 chemicals） the HLA - B ^＊ 2705 promoter activity. （ 2 ） A reconfirmation screening was carried out with these 624 of the candidates using transfected HeLa - HLA - B27 cells. Seventy hits were confirmed. （3） Based on the bioinformatics of those positive hits, 40 chemicals were selected for in - depth analysis by dose - response experiment and IC50/EC50 detection. Six suppressors showed potential pharmacological activities. Interestingly, two suppressors （celastrol and pristimerin） are derived from Leigongteng, a herbal medicine already used for several decades for treatment of immune regulatory and inflammatory diseases. Four active chemicals were computer designed with no relevance to the above structures. CONCLUSION： Chinese traditional herb Nansheteng and Leigongteng might be the potential drugs for HLA - B27 positive patients. These results provide new direction for research in both the therapeutics and the pathogenesis of spondyloarthritis.