具有程序化靶向性和多重生物响应性的仿病毒脂质聚合复合物基因输送载体研究

In order to solve the two major problems in gene delivery system, including low in vivo transfection efficiency and poor targeting capacity, the project designs and constructs a lipid polymeric compound with biomimic viral "envelop” and “capsid". The "envelope” is composed of hyaluronic acid, which is modified by two targeting peptides. The peptides are conjugated to main chain directly or indirectly with PEG as spacer. Using the differences in steric effect and affinity, the delivery system could procedurally target to specific cells and induce endocytosis. In this process, the first bioresponse occurs that the disulfide bonds between hyaluronic acid and PEG is cleaved, and hyaluronic acid is degraded by hyaluronidase in tumor site. It exposes the “capsid" layer for easy lysosomal escape. "Envelope" layer is composed of peptide dendrimer-lipidic carriers, with high density of positive charge for effectively compressing gene. The second bioresponse happens after cellular internalization that the disulfide bond in lipids were broken, resulting in gene release. At the same time, the tumor specific promoter is used in plasmid gene transcription. Ingenious system with "dual response" and "dual targeting" are designed in the “capsid” and "envelope" layer of gene delivery system, in order to simulate the infection process of virus to host cells. Through the systematic study on lipid-polymer carriers, the structure-activity relationship, endocytosis process, targeting mechanism, in vitro and in vivo transfection effect will be studied.

本项目针对基因载体系统体内转染效率低、靶向性差两大问题，设计构建一种模仿病毒“包膜”、“衣壳”层的脂质聚合复合物。“包膜”层由两种靶向配体修饰的透明质酸构成，配体直接或间隔PEG键和在透明质酸主链上，利用空间位阻和亲和力的差异，程序化的与靶细胞相互作用并引导内吞；在此过程中，发生第一次生物响应，即以还原敏感二硫键键和在透明质上的PEG断裂，透明质酸被肿瘤部位富集的透明质酸酶降解，暴露“衣壳”层以利于溶酶体逃逸。“衣壳”层由肽类树状大分子为亲水头部的阳离子脂质载体构成，具有高密度正电荷，能有效压缩基因；载体进入细胞后发生第二次生物响应，脂质载体中的二硫键断裂，释放基因。同时利用肿瘤特异性的启动子调控质粒基因转录。系统巧妙的将“双响应”、“双靶向”设计入“衣壳”和“包膜”层，模拟了病毒感染宿主细胞的过程，通过系统考察可对脂质聚合物载体的构效关系、入胞过程和机理、体内外转染效果的相关性进行研究。

本项目针对基因载体系统体内转染效率低、靶向性差两大问题，设计构建一种模仿病毒“包膜”、“衣壳”层的脂质聚合复合物。“包膜”层由两种靶向配体修饰的透明质酸构成，配体直接或间隔PEG键和在透明质酸主链上，利用空间位阻和亲和力的差异，程序化的与靶细胞相互作用并引导内吞；在此过程中，发生第一次生物响应，即以还原敏感二硫键键和在透明质上的PEG断裂，透明质酸被肿瘤部位富集的透明质酸酶降解，暴露“衣壳”层以利于溶酶体逃逸。“衣壳”层由肽类树状大分子为亲水头部的阳离子脂质载体构成，具有高密度正电荷，能有效压缩基因；载体进入细胞后发生第二次生物响应，脂质载体中的二硫键断裂，释放基因。同时利用肿瘤特异性的启动子调控质粒基因转录。系统巧妙的将“双响应”、“双靶向”设计入“衣壳”和“包膜”层，模拟了病毒感染宿主细胞的过程，通过系统考察可对脂质聚合物载体的构效关系、入胞过程和机理、体内外转染效果的相关性进行研究。