PEPT1和P-gp介导白藜芦醇与乌苯美司发生药物相互作用的分子药代动力学机制

The anticancer effects of bestatin and resveratrol have attracted considerable attention. Previous reports indicate that gastrointestinal oligopeptide transporter (PEPT1), P-glycoprotein (P-gp) and organic anion transporters (OAT1/3) were involved in the transport of bestatin. Recently, we found that resveratrol induced PEPT1 expression through activating of the Nrf2 pathway. Moreover, resveratrol has been found to reverse multidrug resistance (MDR) due to P-gp inhibition and downregulation of MDR1 gene. Thereby, the present research was designed (1) to investigate the drug-drug interactions (DDI) between bestatin and resveratrol; (2) to further clarify molecular pharmacokinetic bases of the DDI mediated by intestinal, hepatic and renal transporters (such as P-gp, PEPT1, OAT1, OAT3); and (3) to assess the effect of NF-κB and/or Nrf2 signaling pathway-mediated transporter regulation on MDR reverse. The involved transporter-based molecular pharmacokinetic mechanisms were then studied through in vivo rat pharmacokinetics study, in situ rat jejunal and liver perfusion technique, in vitro everted intestinal sac preparation, in vitro uptake study using rat/human kidney slices, uptake studies and bi-directional transport assays in Caco-2 cells and transporter gene transfected cells, MTT assay, qRT-PCR, Western blotting and docking methods. The present study will suggest that health care professionals and consumers should be aware of the potential for transporter based DDI and MDR.

白藜芦醇和乌苯美司均为抗癌药物。前期工作首次发现乌苯美司不仅是小肠PEPT1和P-gp的底物，还为肾脏OATs的底物。最近发现，白藜芦醇不仅可激活Nrf-2，促进PEPT1表达，还可抑制P-gp。本研究建立LC-MS-MS内标法同时检测白藜芦醇和乌苯美司，采用in vivo、小肠和肝灌流、翻转小肠袢、新鲜大鼠和人肾切片摄取及Caco-2细胞、PEPT1、OAT1/OAT3、OATP 1B1/1B3及MRP2基因单、双转染细胞等和RT-PCR、Western blot、分子对接等分子生物学方法，阐明多转运体靶点介导白藜芦醇和乌苯美司在吸收、肝肾排泄过程的药物相互作用（DDI）及在分布环节、NF-κB和Nrf2对转运体表达调控环节逆转抗肿瘤药物多药耐药（MDR）的分子药代动力学机制。本课题从DDI和MDR二方面呼吁临床重视转运体介导的DDI和MDR，对指导临床安全合理用药有着重要的应用价值。

白藜芦醇和乌苯美司均为抗癌药物。前期工作首次发现乌苯美司不仅是小肠PEPT1和P-gp的底物，还为肾脏OATs的底物。最近发现，白藜芦醇不仅可激活Nrf-2，促进PEPT1表达，还可抑制P-gp。本研究建立了LC-MS-MS内标法同时检测白藜芦醇和乌苯美司，采用in vivo、小肠和肝灌流、翻转小肠袢、新鲜大鼠和人肾切片摄取及Caco-2细胞、PEPT1、OAT1/OAT3和RT-PCR、Western blot等分子生物学方法，阐明多转运体靶点介导白藜芦醇和乌苯美司在吸收、肝肾排泄过程的药物相互作用（DDI）及在分布环节、NF-κB和Nrf2对转运体表达调控环节逆转抗肿瘤药物多药耐药（MDR）的分子药代动力学机制。主要研究内容、重要结果、关键数据及其科学意义如下：.一、肠道转运体介导的白藜芦醇的肠道吸收过程及白藜芦醇与乌苯美司肠道DDI.1.成功建立了LC-MS/MS测定方法定量分析生物样本中乌苯美司和白藜芦醇.2.确定乌苯美司和白藜芦醇在大鼠肠道的DDI靶点为小肠寡肽转运体Pept1。（1）白藜芦醇使乌苯美司在大鼠肠道的吸收增加；（2）白藜芦醇促进大鼠肠道Pept1和蛋白水平的表达；（3）白藜芦醇促进乌苯美司在Caco-2细胞的摄取，从而说明乌苯美司和白藜芦醇相互作用的靶点在人小肠上皮细胞。.二、白藜芦醇通过抑制PI3K/Akt通路降低P-gp表达，增强乌苯美司对K562/ADR细胞的抗增殖作用.三、肾转运体OAT1/3介导的白藜芦醇的肾处置过程及白藜芦醇与乌苯美司在肾DDI的靶点研究表明白藜芦醇可能是OAT1/3的底物或抑制剂。.四、白藜芦醇通过上调Pept1增强JBP485的抑制炎症和抗凋亡作用，减轻吲哚美辛诱导的大鼠小肠损伤.本课题的结果为临床安全合理联合用药提供了理论和实验证据。