抗肿瘤多药耐药的胞内pH触发式多级靶向聚合物胶束的研究

Overcoming multidrug resistance of tumor cells to anticancer drugs is a major problem faced by the current chemotherapy. Single-targeting nanoformulations exhibited good targeting ability of tumor tissues and tumor cells. Therefore, the uncontrolled drug release in cells showed some limitations in multidrug resistance overcoming. Facing the key scientific issue of reversing multidrug resistance of PTX, a multistage targeting hyaluronic-deoxycholic acid-histidine polymeric micellar delivery system will be built in the project. The micellar system can achieve drug targeting delivery into tumor tissue and tumor cells, and pH triggered drug release in endosome for effectively multidrug resistance reversion in the combination of the EPR effect mediated passive targeting, CD44 receptor-active targeting and endosome pH-sensitive targeting. The pH triggered drug release behavior of polymeric micelles have been investigated for illustrating endosome triggered drug release characteristics. The pharmacokinetics and tissue distribution characteristics will be studied for evaluating the tumor targeting ability of micelles. Cellular uptake study will be investigated for illustrating cellular uptake mechanisms and intracellular drug distribution. In vitro antitumor activity and tumor inhibition pharmacodynamic study will be investigated in detailed for comprehensive evaluating the synergistic targeting effect of CD44 receptor targeting drug delivery and endosome pH-triggered drug released strategy of polymeric micellar system for multidrug resistance reversion, and the reversion mechanisms will be illustrated. We aim to achieve reversing the multidrug resistance of PTX, in order to provide guidance and reference for the multidrug resistance overcoming with multistage targeting drug delivery strategy.

克服肿瘤细胞对抗癌药产生的多药耐药性是目前化疗面临的一大难题。单一靶向纳米制剂虽表现出良好的肿瘤组织、肿瘤细胞靶向性，但胞内释药不可控，克服多药耐药存在局限性。本课题针对逆转紫杉醇耐药的关键问题，构建透明质酸-脱氧胆酸-组氨酸多级靶向聚合物胶束，联合EPR效应介导的被动靶向、CD44受体主动靶向和内涵体pH敏感靶向，多模式、多阶段实现肿瘤组织、肿瘤细胞靶向递药兼胞内触发释药，以逆转多药耐药。前期体外释药考察已初步证实胶束的内涵体触发释药特性。课题将通过药动学和组织分布研究，评价胶束的长循环作用和肿瘤组织靶向性；考察细胞摄取行为，阐明胶束靶向摄取机制和胞内药物转运过程；重点研究体外抗肿瘤活性和抗肿瘤药效学，综合评价CD44受体靶向递药和内涵体pH触发释药策略协同逆转紫杉醇多药耐药这一核心问题，并阐明其逆转机制。以期实现逆转紫杉醇耐药性，为多级靶向递药策略用于抗肿瘤多药耐药研究提供指导和借鉴。

P-糖蛋白介导的多药耐药（multidrug resistance，MDR）是紫杉醇（paclitaxel，PTX）临床化疗效果不佳的主要原因。本课题构建了肿瘤细胞内涵体pH触发式释药的多级靶向的载PTX的HA-DOCA-His胶束，应用多阶段、多种靶向“协同”机制，对胶束系统从血液长循环、肿瘤组织蓄积、细胞摄取和胞内释药这四个药物输送关键阶段进行“全程靶向”引导，实现多级靶向递药兼逆转PTX多药耐药的新策略。HA-DOCA-His胶束对PTX的载药量和包封率分别为18.9%和91.0%。HA-DOCA-His胶束在模拟生理环境及细胞外液具有良好的结构稳定性，释药缓慢；在内涵体（pH 5.0-6.0）环境中具有明显的pH敏感性，可实现触发释药。相比于PTX溶液和PTX/HA-DOCA胶束， PTX/HA-DOCA-His胶束在pH 5.8培养液环境中具有最佳的肿瘤细胞增殖抑制作用，表明受体介导的药物靶向输送和pH敏感靶向释药策略在可协同逆转MDR。HA-DOCA-His胶束在MCF-7和MCF-7/Adr细胞可借助CD44受体介导的靶向内吞机制入胞，启动内涵体环境的触发性释药行为，释放药物到达细胞质，提高其包载药物PTX的抗肿瘤活性，逆转PTX的MDR。与Taxol溶液相比，PTX-HA-DOCA-His胶束能够增加血药浓度曲线下面积（AUC）和半衰期（t1/2），表明PTX/HA-DOCA-His胶束能够提高PTX的生物利用度，并能显著延长PTX在血液中的循环时间。建立CD44受体表达丰富的MCF-7和MCF-7/Adr肿瘤小鼠模型，研究载药胶束对肿瘤小鼠的抑瘤作用。与生理盐水组相比，静脉注射Taxol和载药胶束后，肿瘤的体积及重量均明显下降。Taxol溶液，PTX-HA-DOCA胶束和PTX-HA-DOCA-His胶束对MCF-7肿瘤小鼠的抑瘤率分别为43.2%、60%、76.9%，对MCF-7/Adr肿瘤耐药小鼠的抑瘤率分别为28.1%, 48.8% 和 64.9%，表明HA-DOCA-His载药胶束的CD44受体主动靶向和His内涵体pH敏感靶向可协同提高PTX对多药耐药肿瘤的抗肿瘤功效。本课题对多级肿瘤靶向递药策略应用于克服MDR研究领域的研究思路和方法进行了积极的探索，为逆转抗肿瘤MDR提供一种新策略，也为新型多级靶向给药系统的构建提供指导和借鉴。