基于当归多糖构建改善肿瘤微环境及协同抗肿瘤作用的纳米释药体系

Abnormal tumor microenvironment is a key factor for tumor development, less effectiveness of chemotherapy and radiotherapy. The good immunomodulatory activity of Angelica polysaccharide was proved by our previous research. This project is to construct a novel drug delivery system with dual functions of killing tumor cells and improving tumor microenvironment based on our previous study. After decoration, Angelica polysaccharide are graft copolymerized with Sulfadimethoxine. Then the intelligent polymer with acute response to tumor extracellular pH was obtained by adjusting the ratio of monomer and polysaccharide. Next, the nanoparticles are made by solvent selection dialysis method or emulsion dispersion method loaded with cell-penetrating peptides decorated Doxorubicin (Dox-TAT). Angelica polysaccharides act as not only carriers to targeted deliver drugs to tumor tissue, but also effector to improve tumor microenvironment and enhance immune function, resulting synergistic antitumor effect with antitumor drugs. The pH-sensitivity to tumor microenvironment pH, drug release profile, inhibitions of tumor cells and improvement of tumor microenvironment will be investigated in vitro and in vivo. And then the high efficiency of drugs into the cell mediated by TAT, overcoming drug resistance, the synergistic antitumor effects of Angelica polysaccharide and Dox will also be evaluated. This drug delivery system, with specific tissue targeting, high efficient cell penetrating and tumor microenvironment improvement, will provide a novel synergistic targeted strategy for tumor targeted nano drug release system.

异常的肿瘤微环境是降低放化疗疗效、促进肿瘤发展的关键因素。申请者研究证实当归多糖具有良好的免疫调节活性。本课题在前期研究基础上，以当归多糖为载体材料，修饰后与磺胺二甲氧嘧啶共聚，调节单体与多糖的比例，筛选对肿瘤组织pH有敏锐响应的智能高分子；溶剂选择透析法或乳液分散法制备载药纳米粒，包载穿膜肽修饰的化疗药物阿霉素（Dox-TAT），旨在构建兼有杀伤肿瘤细胞和改善肿瘤微环境双重功效的药物运载体系。当归多糖既作为载体材料将抗肿瘤药物靶向递送至肿瘤组织，又作为效应分子发挥免疫调节活性、与化疗药物产生协同抗肿瘤作用。体内外实验研究纳米微粒对肿瘤pH值的特异响应性及释药规律、抑制肿瘤生长与改善肿瘤微环境的作用及机制，评价穿膜肽介导药物高效入胞、克服耐药性及当归多糖与Dox协同抗肿瘤作用。本课题构建的既具肿瘤靶向和高效穿膜特性、又能改善肿瘤微环境的给药系统，为肿瘤靶向纳米释药体系提供新的协同设计策略。

化学药物治疗是肿瘤治疗的重要手段之一。但化疗药物缺乏肿瘤选择性，在杀伤肿瘤细胞的同时，亦杀伤正常组织细胞，特别是对免疫系统的破坏，不利于肿瘤的治疗。且化疗过程中忽视了肿瘤微环境在肿瘤演进过程中的重要作用。近来研究表明，异常的肿瘤微环境是降低肿瘤放化疗疗效、促进肿瘤发展和转移的关键因素。因此，将针对肿瘤细胞本身的传统治疗手段和改善肿瘤微环境联合应用，可为临床干预肿瘤提供更为有效、安全的新策略。. 本课题以当归多糖为载体材料，成功制备了当归多糖-基质金属蛋白酶敏感肽-阿霉素（AP-PP-DOX）纳米粒。对AP-PP-DOX纳米粒进行了结构表征：平均粒径和电位分 别是（139.00±3.32）nm和（-28.45±0.22）mV；通过TEM观察该纳米粒外观形态，发现该纳米粒结构圆整、粒径分布均一，平均粒径约100nm；利用紫外分光光度法测得该纳米粒平均载药量约为（17.00±1.72）%。体外模拟释药实验结果表明纳米粒在MMP-2（20nM）作用下4h释药率就高达60%左右，24h累积释药率高达74.5%，说明该纳米粒能在MMP-2作用下快速、高效释药。在MMP-2酶作用下AP-PP-DOX纳米粒对肿瘤细胞抑制率明显增加。激光共聚焦显微镜和流式细胞术的结果显示，AP-PP-DOX纳米粒在MMP-2酶作用下能成功释药并快速入胞。载体AP-PP能剂量依赖性的促进小鼠脾细胞增殖，具有潜在的增进机体免疫功能的的作用。载体AP-PP能剂量依赖性的促进小鼠脾细胞INF-γ、IL-2的分泌及其RNA转录水平，而抑制IL-10分泌及其RNA转录水平。说明载体AP-PP能够促进Th1型细胞分泌细胞因子INF-γ、IL-2，抑制Th2型细胞分泌细胞因子IL-10，具有潜在恢复肿瘤微环境Th1/Th2平衡的作用。此外，AP-PP具有促进CD4+T细胞分化、增强抗肿瘤免疫功能的作用。. 本课题将生物活性多糖既作为纳米释药体系的载体材料，又作为效应分子发挥其免疫调节活性，与化疗药物共同构建靶向肿瘤细胞及肿瘤微环境、协同抗肿瘤的纳米释药体系，可为肿瘤靶向纳米释药体系提供新的设计策略。