以GLP_1受体为靶点的药物筛选细胞模型的建立和应用.pdf

药学学报 Acta Pharmaceutica Sinica 2009, 44 (3): 309−313 ·309 · 以 GLP-1 受体为靶点的药物筛选细胞模型的建立和应用 * 环 奕, 申竹芳 (中国医学科学院、北京协和医学院药物研究所, 北京 100050) 摘要: 建立以 GLP-1 信号通路为靶点的药物筛选细胞模型，用于筛选 GLP-1 受体激动剂类的新型抗糖尿病 药物。首先构建 GLP-1 受体信号通路调控的特异应答元件（RIP-CRE ）多拷贝序列及报告基因E-GFP 的重组载 体。将该载体转染胰岛 NIT-1 细胞株，以检测转染细胞对 GLP 类似物的反应性和特异性，并通过稳定转染和单 克隆培养，获得对 GLP-1 类似物特异应答的单克隆细胞株。该细胞模型在 GLP-1 类似物 Exendin 4 刺激下激活 表达报告基因，激活作用可以被 GLP-1 受体阻断剂 Exendin 9-39 完全阻断，且激活途径非 cAMP-PKA 依赖，具 有 GLP-1 受体特异性。构建该细胞模型，可以对肽类或非肽类 GLP-1 类似物进行高通量筛选。 关键词: GLP-1 受体激动剂; 高通量药物筛选; 细胞模型; 糖尿病 中图分类号: R965.1 文献标识码: A 文章编号: 0513-4870 (2009) 03-0309-05 A novel cell model targeted on GLP-1 receptor for application to anti-diabetic candidates screening * HUAN Yi, SHEN Zhu-fang (Institute of Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, China) Abstract: The aim of this project is to establish a GLP-1 signaling pathway targeted cell model, for screening the new class of GLP-1 receptor agonists as anti-diabetic candidates. Firstly construct a recombined plasmid with multi-copied specific response element (RIP-CRE) regulated by GLP-1 signaling pathway and E-GFP reporter gene. Transient transfect this recombined plasmid into islet cell NIT-1, then detect the responsibility of transfected cell to GLP-1 analogue, Exendin 4. For secondly, use stable transfection and monocloning cell culture to obtain a GLP-1 signaling-specific cell line. It in