MOE International Joint Collaborative Research Laboratory for Animal Health and Food Safety & Jiangsu Engineering Laboratory of Animal Immunology, College of Veterinary Medicine, Nanjing Agricultural University, Nanjing, 210095, China.
Biotechnology Research laboratory, Jiangsu Lihua Animal Husbandry co. LTD, Changzhou, 213168, China.
Virol J. 2020 Dec 7;17(1):191. doi: 10.1186/s12985-020-01458-z.
Influenza virus remains a continuous and severe threat to public health worldwide, and its prevention and treatment have always been a major international issue. Because of its ability to evade immune surveillance through rapid antigenic drift and antigenic shift, broad-spectrum vaccines seem increasingly important.
A mAb named 3C12 from an immortalized hybrid cell was generated via immunizing mice with HA2 protein from A/chicken/Anhui/BRI99/2016 (AH/BRI99/16, H9N2) generated by prokaryotic expression. Then, its broad-spectrum activity was analyzed by WB and IFA. Next, the minimal linear epitope was identified via analyzing the reaction of a series of HA truncations with 3C12. Finally, the protective effects of 3C12 were evaluated in vitro and in vivo infection experiments.
The mAb could react with the viruses of subtypes H1, H2, H5, H8, H9, H12, H13, H16, and HA protein of H18 in group 1, but failed to react with viruses in group 2. The minimal linear epitope targeted by the mAb was NAELLVL in full length of HA and localized in the C-helix region of HA2 (residue 95-101, HA2 numbering). What's more, the mAb 3C12 inhibited H1, H2, H5, H8, H9, H12, H13 and H16 virus-replication in vitro and also has shown effectiveness in preventing and treating disease in mice challenged with lethal dose of AH/BRI99/16 (H9N2) virus in vivo. These results suggested that the broadly reactive anti-HA stem mAb 3C12 exhibited prophylactic and therapeutic efficacy.
Here, we have demonstrated that the linear epitope identified in this study could be a novel target for developing broad-spectrum influenza diagnostics or vaccine design, and the HA2-based monoclonal antibody is indeed a promising strategy for broad-spectrum protection against seasonal and pandemic influenza viruses.
流感病毒仍然是全球公共卫生的持续严重威胁，其预防和治疗一直是一个重大的国际问题。由于其通过快速抗原漂移和抗原转变逃避免疫监测的能力，广谱疫苗似乎越来越重要。
通过用原核表达的 A/chicken/Anhui/BRI99/2016（AH/BRI99/16，H9N2）的 HA2 蛋白免疫小鼠，产生了一种命名为 3C12 的源自永生化杂交细胞的 mAb。然后，通过 WB 和 IFA 分析其广谱活性。接下来，通过分析一系列 HA 截断与 3C12 的反应来确定最小线性表位。最后，在体外和体内感染实验中评估了 3C12 的保护作用。
该 mAb 可与亚型 H1、H2、H5、H8、H9、H12、H13、H16 和 H18 组 1 的 HA 蛋白的病毒发生反应，但与组 2 的病毒不发生反应。mAb 靶向的最小线性表位是全长 HA 中的 NAELLVL，位于 HA2 的 C-螺旋区（HA2 编号 95-101）。此外，mAb 3C12 抑制了 H1、H2、H5、H8、H9、H12、H13 和 H16 病毒在体外的复制，并且还在体内用致死剂量的 AH/BRI99/16（H9N2）病毒攻击的小鼠中显示出预防和治疗疾病的有效性。这些结果表明，广谱反应性抗 HA 茎 mAb 3C12 具有预防和治疗功效。
在这里，我们已经证明，本研究中鉴定的线性表位可能成为开发广谱流感诊断或疫苗设计的新靶标，并且基于 HA2 的单克隆抗体确实是针对季节性和大流行性流感病毒进行广谱保护的有前途的策略。
