肝癌胚胎抗原Glypican-3纳米疫苗递送系统的设计及其在肝癌免疫治疗中的应用

本项目拟制备一种具有树突状细胞（DC）靶向性、内涵体逃逸特性，能够将包载的肝癌胚胎抗原缓-控释放的可示踪纳米粒子，应用于肝癌免疫治疗。以甘露糖、脱镁叶绿酸盐-A和肝癌胚胎抗原（磷脂酰肌醇蛋白聚糖-3，GPC-3）修饰三嵌段共聚物PEG-PPG-PEG，分别制备具有DC靶向性、光敏性和可示踪功能，以及pH敏感的乳化剂，以聚乙烯亚胺修饰PLGA为载体材料，采用双乳化-溶剂挥发法制备包载GPC-3纳米粒子，皮下给药后，此纳米粒子可通过甘露糖受体介导的内吞方式被DC包吞，通过活体成像可示踪纳米粒子随DC迁移过程，纳米粒子进入DC内涵体/溶酶体，在较低pH环境下，纳米粒子表面以pH敏感键C=N偶联的抗原突释，内部包载的抗原缓释，在体外激光刺激下，光敏剂产生ROS，协同PEI“质子海绵效应”抗原可实现内涵体逃逸，进行抗原交叉呈递，同时激活CD4+辅助性T细胞和CD8+细胞毒性T细胞，进行肝癌免疫治疗。

The aim of the project is to prepare hepatocellular oncofetal antigen loaded nanoparticles with DC targeting effect, photosensitivity and pH sensitivity for liver cancer immunotherapy. Mannose, pheophorbide-A and glypican -3 (GPC-3) modified triblock copolymer PEG-PPG-PEG were used as emulsifier with DC targeting effect, photosensitivity and pH sensitivity. polyethyleneimine (PEI) modified poly(D,L-lactide-co-glycolide) (PLGA) was choosed as carrier material, GPC-3 loaded nanoparticles were preparation using double emulsion solvent evaporation method. After subcutaneous administration, GPC-3 loaded nanoparticles could be uptake by DC via mannose receptor-mediated endocytosis. The track of DC migration with GPC-3 loaded nanoparticles is studied using near infrared imaging. GPC-3 antigen coupling on the surface of GPC-3 loaded nanoparticles with pH sensitive C=N bond could initial burst release in endosome/lysosomes low pH environment, while GPC-3 loaded in the core of GPC-3 loaded nanoparticles was sustained release. The endo-lysosomal membranes could be destruction by the reactive oxygen species (ROS) generated from activated photosensitizers collaborative PEI proton sponge effect. And then, GPC-3 antigen could be released into the cytoplasm for antigen cross presentation. CD4+ helper T cells and CD8+ cytotoxic T lymphocyte could be simultaneous activated for liver cancer immunotherapy.