GII.4型诺如病毒全人广谱中和单抗及其保守表位研究

The GII.4 genotype of norovirus (NoV) caused at least 5 pandemics of acute gastroenteritis worldwide in the last 30 years, which brought severe public health problem and heavy disease burden. Until now, vaccine and anti-viral drug are still unavailable. In our previous study, we discovered highly conserved central binding pocket (CBP) of HBGA binding interface among different GII.4 NoV cluster. In addition, the conserved neutralizing epitopes against broadly neutralizing mouse monoclonal antibodies were highly related to CBP region. Based on the findings from our previous studies, we hypotheses “CBP region is highly conserved neutralizing epitopes among different GII.4 clusters” and propose to screen human broadly neutralizing monoclonal antibodies (bnMAbs) against GII.4 NoV. In this project, we propose: 1) B cells will be isolated from individuals with highly neutralizing antibodies against different GII.4 clusters. GII.4 specific human monoclonal antibodies library will be generated using single B-cell RT-PCR technique. 2) Human bnMAbs panel will be constructed using NoV-HBGA blockade assay. 3) Corresponding conserved neutralizing epitopes against bnMAbs panle will be primarily identified using phase display. The bnMAbs against CBP will be further screened using CBP site-directed mutagenesis. This project will enable development of human broad-spectrum therapeutic antibodies and provide an important target to design universal vaccine against GII.4 NoV.

GII.4型诺如病毒（NoV）感染引发多次急性胃肠炎世界大流行，是严重的公共卫生问题，目前尚无特异性药物和疫苗。我们前期研究发现不同基因簇GII.4 NoV的受体结合界面形成高度保守中心口袋样结合域（CBP），广谱中和性鼠源单抗的中和表位亦与CBP紧密相关。本项目在前期工作基础上，提出“CBP是GII.4 NoV保守性中和抗原表位”假说并进行全人广谱中和单抗（bnMAbs）研究。拟开展：1）分离对不同GII.4基因簇具有高效中和作用的供体外周血，运用单个B细胞分选技术，获得单个特异性GII.4 NoV记忆B细胞；2）采用单细胞RT-PCR单抗技术建立全人单抗库，通过HBGA受体阻断实验获得bnMAbs盘；3）利用噬菌体展示技术初步确定其中和抗原表位，并通过CBP点突变进一步筛选CBP域全人单抗。项目将验证该假说为研发抗NoV治疗性单抗药物提供科学依据，并为“通用”的新型表位疫苗提供新靶点。

GII.4型诺如病毒（NoV）感染引发多次急性胃肠炎世界大流行，是严重的公共卫生问题。 .临床上尚无特异性药物和疫苗，本项目按研究计划完成了1、建立了不同GII.4基因簇用单细胞RT-PCR单抗技术建立全人单抗库，通过HBGA受体阻断实验获得bnMAbs盘；2、利用噬菌体展示技术初步确定其中和抗原表位，并通过CBP点突变进一步筛选CBP域全人单抗。此外，项目同时超额完成了：1、利用两次NoV暴发恢复期PBMC筛选了包含GII.4在内的广谱全人单抗库，鉴定2个相关抗原表位；2、并构建了表达九种不同 NoVs GⅡ/GI NoVs P 颗粒，通过对基因型的抗原相关性特征进行分析，首次证实了NoVs 免疫分型的假说。项目为研发抗NoV治疗性单抗药物提供科学依据，并为“通用”的新型表位疫苗提供新靶点，对 NoVs 的疫苗研制策略具有重大的指导意义。本项目发表SCI论文7篇，中文核心期刊5篇，申请专利3项，联合培养博士研究生1名，培养硕士研究生3名。