基于活性氧响应性细胞膜仿生纳米载体的蛋白质药物传输系统及其用于肺纤维化治疗

Protein drugs have attractive prospects in the treatment of lung fibrosis and other major human diseases, developing safe and efficient carrier materials to maintain their activities as well as achieve the targeted and controlled release is the key to fully exert their therapeutic efficacy. The project is proposed to build a biodegradable and reactive oxygen species (ROS)-responsive biomimetic nanocarrier to achieve the efficient delivery of protein drugs for improving fibrosis treatment. A well-known biodegradable natural polysaccharide, chitosan is modified by introducing cell membrane mimicked phosphorylcholine groups with zwitterionic charge and arylboronic ester modified side groups with ROS-induced hydrophilic-shift property, which can assemble with protein drugs to form nanocomplexes, but release them induced by oxidative stress in the pathological environment of fibrosis. And cell membrane-mimicking nanostructures can be benificial to maintaining protein activity, inhibiting the body's defense mechanism to activate and improving transmembrane transport. We will investigate the synthesis of ROS-responsive biomimetic chitosan derivatives, and the effect of their structure on the loading, activity and ROS-responsive release of protein drugs, as well as the biocompatibility, interaction with cells, in vivo behavior and the inhibition effect on lung fibrosis of biomimetic nanocomplexes, to reveal the interrelation of ROS-responsive biomimetic nanocarriers, protein, cell and tissue. The study will provide the basis for the design and application of the safe and efficient protein drug delivery systems and have important scientific significance.

蛋白质药物在治疗肺纤维化等人类重大疾病方面具有诱人前景，发展安全高效的载体材料保持其活性并实现靶向控释是充分发挥其疗效的关键。本项目提出构建一种生物降解型活性氧（ROS）响应性仿生纳米载体，用于实现抗纤维化蛋白质药物的高效化。在壳聚糖上分别引入亲水性仿细胞膜结构的磷酸胆碱和响应ROS发生疏水-亲水转变的芳基硼酸酯修饰侧基，蛋白质药物可与其组装形成纳米复合物，并在纤维化的氧化应激病理环境下定位释放，而细胞膜仿生纳米结构有利于维持蛋白质活性，抑制机体防御机制激活及促进跨细胞膜转运。系统研究ROS响应性仿生壳聚糖衍生物的合成，其结构对蛋白质药物的负载及响应性释放行为的影响，以及纳米复合物的生物相容性、与细胞相互作用、在体行为及肺纤维化抑制效果，阐明ROS响应性仿生载体与蛋白质及机体组织细胞间的相互作用规律，为安全高效的蛋白质药物等诊断治疗物质传输系统的设计及应用提供依据，具有重要科学意义。

蛋白质药物在治疗肺纤维化等人类重大疾病方面具有诱人前景，但如何保障其高效发挥治疗作用仍然是亟待解决的难题。本项目从仿生出发，提出构建可生物降解的，能有效负载并维持蛋白质活性，并在氧化应激病理环境下定位释放的活性氧（ROS）响应性仿生纳米系统，用于实现抗纤维化蛋白质药物的高效化。系统研究了仿细胞膜结构的两性磷酸胆碱（PC）和带双正电荷的磷酸二胆碱（PdC）修饰壳聚糖衍生物（PCCs和PdCCs），及具有可响应ROS发生疏水-亲水转变的芳基硼酸酯修饰壳聚糖衍生物（PCCs-BSer和乙二醇壳聚糖GC-NBC）和聚天冬氨酸衍生物（PASP-BSer）等智能高分子的合成及ROS响应性纳米系统的构建、理化和生物学性能规律；分别以小分子药物和牛血清白蛋白为模型研究其载药和响应性释放；以Calu-3上皮细胞层为细胞模型和博来霉素诱导的肺纤维化大鼠为动物模型，研究了负载重组成纤维细胞生长因子受体胞外段（rhFGFR）（治疗蛋白ED-1）的纳米系统经肺给药的跨上皮细胞层行为机制及肺纤维化抑制效果。结果表明：不同电荷特性的PCCs和PdCCs均具有特异的结合水结构，可有效维持蛋白质构象，并表现出良好的生物相容性；ROS响应性PCCs-BSer、GC-NBC和PASP-BSer均可自组装形成生物相容性良好的ROS响应性纳米系统，且GC-NBC可进一步与含儿茶酚结构的疏水性多酚（如槲皮素）偶联构成抗炎性的ROS/pH双响应性纳米载体系统；细胞和动物实验结果证实，肺给药方式施加rhFGFR纳米药物可以通过穿胞及胞间方式经肺上皮细胞层发挥作用，有效抑制肺纤维化。综上所述，我们成功制备了多种可实现蛋白质药物高效化的ROS响应性纳米仿生载体，较系统地研究了其与蛋白和机体组织细胞间相互作用，可为安全高效的蛋白质药物传输系统的设计及应用提供依据，促进纳米医学等学科的发展。