聚合物胶束纳米载体对肝转运体和CYP450酶功能的干预作用及其机制研究

Over the past two decades, there has been considerable interest in developing nanocarrierswith better efficacy and less side-effects through reducing the dose of the drug in bystander tissues and increasing the drug at the desired target site. Currently, little is known in the interaction between nanocarrier systems and liver transporters/metabolic enzymes. In this project, a series of polymeric micelle nanocarriers will be prepared with different chemical, phisical and morphological properties. After characterizing in vitro properties of these nanocarriers, their effects on drug cellular transport and metabolism will be investigated using in vitro models including HepaRG cell lines, sandwich-cultured rat hepatocytes and cocktail assays. In these studies, corresponding substrate will be selected to determine the effect of nanocarriers on OATPs, P-gp, BCRP, CYP3A4, CYP1A2, CYP2C9, CYP2C19 and CYP2D6. Furthermore, Hepatic metabolism, along with biliary excretion, will be examined by in situ liver perfusion model. The influence of Nanocarriers on drug disposition as well as potential drug interactions will be systematically studied. Subsequently, correlation among categories of polymeric micelle nanocarriers, physico-chemical characteristics, liver transporters and/or CYP450 function, and pharmacokinetic alteration will be illustrated and summarized. Finally, molecular biological techniques will be employed to clarify the in-depth mechanism of polymeric micelle nanocarriers on the regulation of function of liver transporters and metabolic enzymes as well as upstream molecular modulator such as FXR, AhR,CAR and etc. In summary, this project will provide theoretical and experimental basis for rational design and understanding in vivo pharmacokinetics, drug interaction and safety of nanocarriers.

纳米载体系统一直是近二十年来国内外药剂学的研究热点，其体内安全性问题正引起日益重视。目前关于纳米载体系统与肝脏转运体/代谢酶之间的相互作用缺乏深入了解。本项目以聚合物胶束为研究模型，用四类聚合物材料构建具不同性质的胶束载体，采用HepaRG、"三明治"大鼠肝细胞模型、Cocktail探针法及肝灌流等技术系统地考察不同聚合物胶束纳米载体对肝脏转运体（OATPs、P-gp、BCRP）和CYP450酶（CYP3A4、CYP1A2、CYP2C9、CYP2C19、CYP2D6）功能的干预作用，明确载体种类、各理化性质与肝脏药物转运和代谢能力及体内药动学特性之间的相关性；研究上述肝脏功能蛋白及上游调控因子FXR、AhR、CAR等的蛋白和基因表达水平，阐明其分子作用机制。为聚合物胶束纳米载体系统类制剂的研究、安全性评价和药物相互作用机制提供理论依据，也可为其他纳米载体系统的研究提供重要参考。

纳米载体系统是近年来药物治疗领域的研究热点，其体内安全性问题正引起日益重视。目前关于纳米载体系统与肝脏转运体/代谢酶之间的相互作用研究仍非常有限。本项目主要以mPEG-PCL、mPEG-PDLLA类等嵌段聚合物为材料制备两亲性聚合物纳米胶束，采用体外肝微粒体Cocktail探针法、转染细胞模型和整体动物实验评价了聚合物胶束纳米载体对肝脏重要CYP450酶（CYP1A1、CYP1A2、CYP2B1、CYP2C9、CYP2C19、CYP2D6、CYP3A4）功能和转运体（OATPs、NTCP、P-gp）功能的影响，并检测了胆汁酸内稳态、肝功能、相应功能蛋白及上游调控因子FXR、AhR、CAR的蛋白和基因表达水平，探索了其作用机制。研究结果发现，mPEGx-PCLy系列聚合物胶束和mPEG2k-PDLLA2k聚合物胶束对CYP450酶活性有一定的抑制作用和诱导作用，其抑制和诱导程度与聚合物浓度、亲/疏水嵌段分子量、疏水嵌段组成和酶亚型及种属有关；聚合物胶束在细胞水平和整体动物水平对转运体OATP/Oatp、NTCP/Ntcp和P-gp功能也产生一定的抑制或诱导作用，其作用程度与聚合物浓度、疏水段分子量及作用时间有关。本项目研究结果提示聚合物胶束纳米载体与体内代谢酶与转运体功能具有潜在影响，当与其底物药物合用时，可能存在胶束-药物相互作用。本项目可为临床上聚合物胶束载体药物的构建与合理安全应用提供实验依据。