金刚烷胺二聚体佐剂对新型冠状病毒蛋白疫苗诱导小鼠体液免疫应答的效果  

Effect of amantadine dimer adjuvant on humoral immune response induced by SARS-CoV-2 protein vaccine in mice

作　　者：陈思思 金育 张玉芳 何春艳[1] 周正淼 刘野 CHEN Si-si;JIN Yu;ZHANG Yu-fang;HE Chun-yan;ZHOU Zheng-miao;LIU Ye(Institute of Medical Biology,Chinese Academy of Medical Sciences&Peking Union Medical College,Kunming 650118,Yunnan Province,China)

机构地区：[1]中国医学科学院北京协和医学院医学生物学研究所,云南昆明650118

出　　处：《中国生物制品学杂志》2023年第1期5-10,共6页Chinese Journal of Biologicals

基　　金：云南省创新疫苗技术与产业转化研发平台(202002AA100009);云南省医学领军人才(L-2018013)。

摘　　要：目的研究金刚烷胺二聚体佐剂对新型冠状病毒蛋白疫苗诱导小鼠体液免疫应答的效果。方法经取代反应连接、水解酸化反应连接、酰胺缩合反应连接合成金刚烷胺二聚体,作为佐剂与新型冠状病毒受体结合域(receptorbinding domain,RBD)蛋白免疫雌性BALB/c小鼠后,将小鼠随机分为5组,每组6只:R6A+RBD组[21μg(0.033μmol)金刚烷胺二聚体+10μg RBD]、Ada+RBD组[10μg(0.066μmol)金刚烷胺+10μg RBD]、Alu+RBD组(35μg铝佐剂+10μg RBD)、RBD组(10μg RBD)、Blank组(0.9%生理盐水)。分别在第0、14和28天肌肉注射免疫小鼠,于第2次免疫后7 d和末次免疫后14 d,经小鼠尾静脉采血分离血清,ELISA法检测小鼠血清中特异性IgG抗体水平。结果经薄层色谱法(thin layer chromatography,TLC)进行纯化,并利用电喷雾电离质谱(electrosprayionization-MS,ESIMS)正/负离子模式鉴定,获得的金刚烷胺二聚体为目标产物。2次免疫后,R6A+RBD组各稀释度小鼠血清抗体水平均高于Ada+RBD组,但均低于Alu+RBD组;3次免疫后,R6A+RBD组各稀释度小鼠血清抗体水平均显著高于Ada+RBD和Alu+RBD组,差异有统计学意义(F均>30,P均<0.0001和<0.01)。结论金刚烷胺二聚体佐剂能增强新冠蛋白疫苗诱导小鼠体液免疫应答,具有良好的佐剂效应,可作为备选佐剂,这种基于现有药物改造的策略为开发新型佐剂提供了新的思路。Objective To investigate the effect of amantadine dimer adjuvant on humoral immune response induced by SARS-CoV-2 crown protein vaccine in mice.Methods The amantadine dimer was synthesized by substitution reaction ligation,hydrolytic acidification reaction ligation and amide condensation reaction ligation,with which as adjuvant,female BALB/c mice were immunized with the receptor-binding domain(RBD).The mice were randomly divided into five groups,six for each as follows:R6A+RBD group[21μg(0.033μmol)amantadine dimer+10μg RBD],Ada+RBD group[10μg(0.066μmol)amantadine+10μg RBD],Alu+RBD group(35μg aluminum adjuvant+10μg RBD),RBD group(10μg RBD)and Blank group(0.9%normal saline),which were immunized i.m.on day 0,14 and 28 respectively.Serum samples were collected from tail vein of mice 7 d after the second dose and 14 d after the last dose and determined for specific IgG antibody levels by ELISA.Results The amantadine dimer was purified by thin layer chromatography(TLC)and identified by electrospray ionization-MS(ESI-MS)positive/negative ion mode.After two times of immunization,the antibody levels in sera at various dilutions of mice in R6A+RBD group were all higher than those of Ada+RBD group,while lower than those of Alu+RBD group.However,after three times of immunization,the antibody levels in sera at various dilutions of mice in R6A+RBD group were all significantly higher than those of Ada+RBD and Alu+RBD groups(each F>30,each P<0.0001 and each P<0.01).Conclusion Amantadine dimer adjuvant enhanced humoral immune response induced by SARS-CoV-2 protein vaccine in mice with good adjuvant effect,which may be used as an alternative adjuvant.This strategy based on existing drug transformation provided a new idea for the development of novel adjuvants.

关 键 词：金刚烷胺二聚体 佐剂 新冠疫苗 体液免疫 

分 类 号：R392[医药卫生—免疫学]