MAGE-3多肽纳米疫苗对小鼠胃癌种植瘤抑瘤效应研究

Preparation and antitumor effects of nanovaccines with MAGE-3 peptides in transplanted gastric cancer in mice

背景与目的:纳米颗粒作为疫苗载体可保护抗原免受酶解,增强免疫原性,是一类极具开发潜力的新型疫苗载体。本实验制备负载CD4+CD8+T细胞表位MAGE-3多肽抗原的纳米疫苗,探讨其相关特性及抗肿瘤免疫。方法:利用自组装技术制备多肽/Chit-DC(壳聚糖-脱氧胆酸)载药纳米胶束,透射电镜观察纳米微观形态,荧光分光光度法计算负载率、载药量,并测定药物释放规律。流式细胞仪检测DC(树突状细胞)对药物的吞噬率,酶联免疫斑点实验(ELISPOT)和细胞毒性实验检测MAGE-3多肽纳米疫苗激活机体细胞免疫反应的状况。动物实验观察其体内抑瘤效应。结果:成功制备多肽/Chit-DC纳米胶束,药物包封率约为37%,载药量为17%。载药纳米颗粒中的多肽在pH7.4的PBS中释放缓慢,于48h达释放平台;在2mg/mL溶菌酶溶液中,药物有一定的突释现象,于24h达释放平台。ELISPOT和细胞毒性实验显示MAGE-3多肽纳米疫苗可以激活体内免疫反应而产生针对MAGE-3的CTL,特异性杀伤表达MAGE-3的肿瘤细胞。体内抑瘤实验显示,多肽纳米疫苗组相对肿瘤抑制率为37.81%。结论:MAGE-3多肽/Chit-DC纳米疫苗能有效激活体内的抗肿瘤免疫效应,抑制MFC小鼠前胃癌细胞的生长。

Background and Objective：As a prospective vaccine carrier,nanoparticles can protect antigens from degradation and enhance immune response. This study prepared nanovaccines with MAGE-3-derived CD4^＋-CD8^＋T cell epitope peptides,and investigated its character and antitumor effects on transplanted gastric cancer in mice.Methods：We adopted the self-assembly method to prepare peptide/chitosan conjugated with deoxycholic acid （chitosan-deoxycholic acid） nanoparticles. We observed the appearance of the chitosan-deoxycholic acid nanoparticles through a transmission electron microscope （TEM） and analyzed the peptide content and its release pattern by fluorescence spectrophotometry. We observed tumor-suppression efficacy in vivo through animal experiments.Results：We successfully prepared nanoparticles with MAGE-3 peptide antigen,and its encapsulation efficiency and loading level were about 37% and 17.0%,respectively. These nanoparticles presented a delayed release pattern in phosphate buffered saline （PBS） at pH 7.4,and the full release time was about 48 h. In 2 mg/mL lysozyme,the nanoparticles showed a sudden release,and the full release time was about 24 h. ELISPOT and cytotoxic experiments showed that the MAGE-3 peptide loaded nanoparticles could stimulate immune response in vivo and could generate MAGE-3-targeted cytotoxic T lymphocytes （CTLs）,and kill MAGE-3-specific tumor cells. Tumor suppression experiments showed that the regression ratio of the peptide-loaded nanoparticles group was 37.81%. Conclusion：MAGE-3 peptide/chitosan-deoxycholic acid vaccine-loaded nanoparticles can stimulate antitumor immune response in vivo and can regress the growth of mouse forestomach carcinoma cell line MFC.