基于P-gp途径研究蟾毒灵逆转结肠癌多药耐药的分子机制

项目名称： 基于P-gp途径研究蟾毒灵逆转结肠癌多药耐药的分子机制

项目编号： No.81473482

项目类型： 面上项目

立项/批准年度： 2015

项目学科： 医药、卫生

项目作者： 殷佩浩

作者单位： 上海中医药大学

项目金额： 75万元

中文摘要： 多药耐药是导致肿瘤化疗失败的关键因素，P-糖蛋白（P-gp）是产生耐药的主要靶分子之一。我们前期发现中药蟾酥中活性成分蟾毒灵能够通过抑制P-gp的表达，从而促进结肠癌细胞凋亡。提示蟾毒灵可能作为一种P-gp抑制剂，通过协同其它化疗药，增强抗肿瘤疗效。鉴此，本项目应用膜转运功能测定、荧光标记、HPLC等观察蟾毒灵对肿瘤细胞P-gp-ATPase、P-gp在细胞膜与核上的定位与表达、P-gp介导药物转运等影响，阐明蟾毒灵抑制P-gp介导的药物外排机制。应用Western blot、RT-PCR、免疫组化等观察蟾毒灵对小鼠移植瘤模型P-gp介导的细胞凋亡和肿瘤生长的作用；并研究蟾毒灵对相关信号分子如JNK、ERK、p38-MAPK、P13K/AKT及相应转录因子如多药耐药基因1（MDR1）的影响，揭示蟾毒灵作为P-gp抑制剂逆转结肠癌耐药的机制，为临床治疗耐药性结肠癌提供理论依据和新的治疗策略。

中文关键词： 结肠癌；蟾毒灵；P-糖蛋白；分子机制；多药耐药

英文摘要： Multidrug resistance is one of the most formidable challenges in the field of cancer chemotherapy. P-glycoprotein belongs to the large ATP-binding cassette transporter superfamily of membrane tranporters and mediates resistance to a variety of anticancer drugs. In our previous study, we found that bufalin, a major bioactive component isolated from the skin and parotid glands of venenum bufois, could inhibit P-gp expression and induce apoptosis in colon cancer cells. Thus we hypothesize that bufalin, a potential P-gp inhibitor, act as a sensitzer in anti-cancer chemotherapy. On this hypothesis, the present project designs to investigate the effect of bufalin on cellular P-gp-ATPase activity, P-gp localization and expression. P-gp-mediated drug active transport will be investigated by using fluorescent probe, transport assay, HPLC technique, RT-PCR and Western blot in Lovo and Caco-2 cells. Moreover, the influences of bufalin on cell apoptosis and proliferation mediated by multidrug resistance molecule P-gp/MDR1, and the related signaling molecules such as MAPK (p38-MAPK, ERK, JNK), and P13K/AKT were investigated by mean of immunohistochemistry, RT-PCR, Western blot in murine xenograft model. This project will provide the experimental and theoretical evidences and new treatment strategies for the clinical application of bufalin in therapy of colon cancer.

英文关键词： colon cancer;bufalin;P-glycoprotein;molecular mechanisms;multidrug resistance