Since the start of the coronavirus disease-2019 (COVID-19) pandemic, severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has caused more than 2 million deaths worldwide. Many vaccines have been deployed to date, but the continual evolution of the viral receptor-binding domain (RBD) has recently challenged their efficacy. In particular, SARS-CoV-2 variants originating in the U.K. (B.1.1.7), South Africa (B.1.351) and New York (B.1.526) have reduced neutralization activity from convalescent sera and compromised the efficacy of some antibody cocktails that received emergency use authorization. Whereas vaccines can be updated periodically to account for emerging variants, complementary strategies are urgently needed to avert viral escape. One potential alternative is the use of camelid VHHs (also known as nanobodies), which due to their small size can recognize protein domains that are often inaccessible to conventional antibodies. Here, we isolate anti-RBD nanobodies from llamas and nanomice we engineered to produce VHHs cloned from alpacas, dromedaries and camels. Through binding assays and cryo-electron microscopy, we identified two sets of highly neutralizing nanobodies. The first group expresses VHHs that circumvent RBD antigenic drift by recognizing a region outside the ACE2-binding site that is conserved in coronaviruses but is not typically targeted by monoclonal antibodies. The second group is almost exclusively focused to the RBD-ACE2 interface and fails to neutralize pseudoviruses carrying the E484K or N501Y substitutions. Notably however, they do neutralize the RBD variants when expressed as homotrimers, rivaling the most potent antibodies produced to date against SARS-CoV-2. These findings demonstrate that multivalent nanobodies overcome SARS-CoV-2 variant mutations through two separate mechanisms: enhanced avidity for the ACE2 binding domain, and recognition of conserved epitopes largely inaccessible to human antibodies. Therefore, while new SARS-CoV-2 mutants will continue to emerge, nanobodies represent promising tools to prevent COVID-19 mortality when vaccines are compromised.

中文翻译：

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来自骆驼科工程小鼠和美洲驼的多聚体纳米抗体有效地中和了SARS-CoV-2变体

自2019年冠状病毒病（COVID-19）大流行以来，严重的急性呼吸系统综合症冠状病毒2（SARS-CoV-2）已导致全球200万人死亡。迄今为止，已经部署了许多疫苗，但是病毒受体结合域（RBD）的不断发展最近对它们的功效提出了挑战。尤其是，源自英国（B.1.1.7），南非（B.1.351）和纽约（B.1.526）的SARS-CoV-2变体降低了来自恢复期血清的中和活性，并损害了某些抗体的功效已获得紧急使用授权的鸡尾酒。尽管可以定期更新疫苗以解决新出现的变异，但迫切需要补充策略来避免病毒逃逸。一种可能的替代方法是使用骆驼科VHH（也称为纳米抗体），由于其体积小，它可以识别常规抗体通常无法接近的蛋白质结构域。在这里，我们从羊驼和纳米小鼠中分离出了抗RBD纳米抗体，我们将其设计为生产从羊驼，单峰骆驼和骆驼克隆的VHH。通过结合测定和冷冻电子显微镜，我们鉴定出两组高度中和的纳米抗体。第一组表达通过识别ACE2结合位点之外在冠状病毒中保守但通常不被单克隆抗体靶向的区域来规避RBD抗原漂移的VHH。第二组几乎专门针对RBD-ACE2接口，无法中和带有E484K或N501Y替代的假病毒。然而，值得注意的是，当它们表达为同三聚体时，它们确实中和了RBD变体 可以与迄今为止针对SARS-CoV-2产生的最有效的抗体抗衡。这些发现表明，多价纳米抗体通过两种独立的机制克服了SARS-CoV-2变异：对ACE2结合域的亲和力增强，以及对人类抗体难以接近的保守表位的识别。因此，尽管新的SARS-CoV-2突变体将继续出现，但纳米抗体代表了在疫苗受损时防止COVID-19死亡的有前途的工具。