. doi: 10.1016/j.virs.2022.11.005
Citation: Haoneng Tang, Yong Ke, Yunji Liao, Yanlin Bian, Yunsheng Yuan, Ziqi Wang, Li Yang, Hang Ma, Tao Sun, Baohong Zhang, Xiaoju Zhang, Mingyuan Wu, Jianwei Zhu. Mutational escape prevention by combination of four neutralizing antibodies that target RBD conserved regions and stem helix .VIROLOGICA SINICA, 2022, 37(6) : 860-873.  http://dx.doi.org/10.1016/j.virs.2022.11.005

分别靶向RBD三个保守表位和S2茎螺旋表位的四种中和抗体联合作用防止病毒突变逃逸

  • 新的新冠病毒突变株每隔数月就快速出现。这些突变株已经表现出强大的适应能力来逃避免疫系统。为了进一步认识新冠病毒的适应能力以便寻求策略来减缓新突变株的出现,在此我们研究了病毒在广谱中和抗体和它们联用条件下的适应性。首先,我们选取了四种广谱中和抗体,其中包括泛沙贝病毒和泛贝塔冠状病毒的中和抗体,它们识别RBD上的不同保守区域或S2亚基上的保守茎螺旋区域。通过竞争性结合分析,我们证明了它们能够同时结合上刺突蛋白。然后,一种具有复制能力的新冠病毒刺突蛋白假型化的水疱性口炎病毒被用于研究病毒的适应性。在单个抗体或抗体组合的选择压力下,进行了二十次连续病毒传代。我们发现病毒并不难适应广谱中和抗体,即便是对于泛沙贝病毒和泛贝塔冠状病毒中和抗体亦是如此。然而病毒越来越难逃逸两个/三个/四个抗体的组合。而且,高通量测序所揭示的病毒群体中的突变显示,在三个/四个组合抗体的选择压力下,病毒突变不倾向于出现在贝塔冠状病毒中高度保守的区域(茎螺旋区域),但在单个/两个抗体的选择压力下则并非如此。重要的是,靶向RBD保守区域和茎螺旋的中和抗体的联合使用协同地防止了逃逸突变的出现。这些研究将指导未来疫苗和治疗药物开发的攻关,并提供了设计RBD-茎螺旋串联疫苗的逻辑依据,这种疫苗设计可能有助于阻碍新突变株的产生。

Mutational escape prevention by combination of four neutralizing antibodies that target RBD conserved regions and stem helix

  • New variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) appear rapidly every few months. They have showed powerful adaptive ability to circumvent the immune system. To further understand SARS-CoV-2's adaptability so as to seek for strategies to mitigate the emergence of new variants, herein we investigated the viral adaptation in the presence of broadly neutralizing antibodies and their combinations. First, we selected four broadly neutralizing antibodies, including pan-sarbecovirus and pan-betacoronavirus neutralizing antibodies that recognize distinct conserved regions on receptor-binding domain (RBD) or conserved stem-helix region on S2 subunit. Through binding competition analysis, we demonstrated that they were capable of simultaneously binding. Thereafter, a replication-competent vesicular stomatitis virus pseudotyped with SARS-CoV-2 spike protein was employed to study the viral adaptation. Twenty consecutive passages of the virus under the selective pressure of individual antibodies or their combinations were performed. It was found that it was not hard for the virus to adapt to broadly neutralizing antibodies, even for pan-sarbecovirus and pan-betacoronavirus antibodies. The virus was more and more difficult to escape the combinations of two/three/four antibodies. In addition, mutations in the viral population revealed by high-throughput sequencing showed that under the selective pressure of three/four combinational antibodies, viral mutations were not prone to present in the highly conserved region across betacoronaviruses (stem-helix region), while this was not true under the selective pressure of single/two antibodies. Importantly, combining neutralizing antibodies targeting RBD conserved regions and stem helix synergistically prevented the emergence of escape mutations. These studies will guide future vaccine and therapeutic development efforts and provide a rationale for the design of RBD-stem helix tandem vaccine, which may help to impede the generation of novel variants.

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    Mutational escape prevention by combination of four neutralizing antibodies that target RBD conserved regions and stem helix

      Corresponding author: Baohong Zhang, bhzhang@sjtu.edu.cn
      Corresponding author: Xiaoju Zhang, zhangxiaoju@zzu.edu.cn
      Corresponding author: Mingyuan Wu, wumingyuan@sjtu.edu.cn
      Corresponding author: Jianwei Zhu, jianweiz@sjtu.edu.cn
    • a Engineering Research Center of Cell and Therapeutic Antibody, Ministry of Education, Shanghai, 200240, China;
    • b School of Pharmacy, Shanghai Jiao Tong University, Shanghai, 200240, China;
    • c School of Agriculture and Biology, Shanghai Jiao Tong University, Shanghai, 200240, China;
    • d Shanghai Municipal Veterinary Key Laboratory, Shanghai, 200240, China;
    • e Department of Respiratory and Critical Care Medicine, Zhengzhou University People's Hospital/Henan Provincial People's Hospital, Zhengzhou, 450003, China;
    • f Jecho Institute, Shanghai, 200240, China

    Abstract: New variants of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) appear rapidly every few months. They have showed powerful adaptive ability to circumvent the immune system. To further understand SARS-CoV-2's adaptability so as to seek for strategies to mitigate the emergence of new variants, herein we investigated the viral adaptation in the presence of broadly neutralizing antibodies and their combinations. First, we selected four broadly neutralizing antibodies, including pan-sarbecovirus and pan-betacoronavirus neutralizing antibodies that recognize distinct conserved regions on receptor-binding domain (RBD) or conserved stem-helix region on S2 subunit. Through binding competition analysis, we demonstrated that they were capable of simultaneously binding. Thereafter, a replication-competent vesicular stomatitis virus pseudotyped with SARS-CoV-2 spike protein was employed to study the viral adaptation. Twenty consecutive passages of the virus under the selective pressure of individual antibodies or their combinations were performed. It was found that it was not hard for the virus to adapt to broadly neutralizing antibodies, even for pan-sarbecovirus and pan-betacoronavirus antibodies. The virus was more and more difficult to escape the combinations of two/three/four antibodies. In addition, mutations in the viral population revealed by high-throughput sequencing showed that under the selective pressure of three/four combinational antibodies, viral mutations were not prone to present in the highly conserved region across betacoronaviruses (stem-helix region), while this was not true under the selective pressure of single/two antibodies. Importantly, combining neutralizing antibodies targeting RBD conserved regions and stem helix synergistically prevented the emergence of escape mutations. These studies will guide future vaccine and therapeutic development efforts and provide a rationale for the design of RBD-stem helix tandem vaccine, which may help to impede the generation of novel variants.

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