State Key Laboratory of Cell Biology, Shanghai Institute of Biochemistry and Cell Biology, Center for Excellence in Molecular Cell Science, Chinese Academy of Sciences, Shanghai, 200031, China.
Shanghai Public Health Clinical Center, Shanghai Medical College, Fudan University, Shanghai, 201508, China.
Genome Med. 2021 Oct 14;13(1):164. doi: 10.1186/s13073-021-00985-w.
The receptor-binding domain (RBD) variants of SARS-CoV-2 could impair antibody-mediated neutralization of the virus by host immunity; thus, prospective surveillance of antibody escape mutants and understanding the evolution of RBD are urgently needed.
Using the single B cell cloning technology, we isolated and characterized 93 RBD-specific antibodies from the memory B cells of four COVID-19 convalescent individuals in the early stage of the pandemic. Then, global RBD alanine scanning with a panel of 19 selected neutralizing antibodies (NAbs), including several broadly reactive NAbs, was performed. Furthermore, we assessed the impact of single natural mutation or co-mutations of concern at key positions of RBD on the neutralization escape and ACE2 binding function by recombinant proteins and pseudoviruses.
Thirty-three amino acid positions within four independent antigenic sites (1 to 4) of RBD were identified as valuable indicators of antigenic changes in the RBD. The comprehensive escape mutation map not only confirms the widely circulating strains carrying important immune escape RBD mutations such as K417N, E484K, and L452R, but also facilitates the discovery of new immune escape-enabling mutations such as F486L, N450K, F490S, and R346S. Of note, these escape mutations could not affect the ACE2 binding affinity of RBD, among which L452R even enhanced binding. Furthermore, we showed that RBD co-mutations K417N, E484K, and N501Y present in B.1.351 appear more resistant to NAbs and human convalescent plasma from the early stage of the pandemic, possibly due to an additive effect. Conversely, double mutations E484Q and L452R present in B.1.617.1 variant show partial antibody evasion with no evidence for an additive effect.
Our study provides a global view of the determinants for neutralizing antibody recognition, antigenic conservation, and RBD conformation. The in-depth escape maps may have value for prospective surveillance of SARS-CoV-2 immune escape variants. Special attention should be paid to the accumulation of co-mutations at distinct major antigenic sites. Finally, the new broadly reactive NAbs described here represent new potential opportunities for the prevention and treatment of COVID-19.
SARS-CoV-2 的受体结合域（RBD）变体可能会损害宿主免疫产生的针对该病毒的抗体中和作用；因此，迫切需要对抗体逃逸突变体进行前瞻性监测，并了解 RBD 的进化情况。
我们使用单 B 细胞克隆技术，从 4 名 COVID-19 康复者的记忆 B 细胞中分离并鉴定了 93 种 RBD 特异性抗体。然后，我们使用一组 19 种具有代表性的中和抗体（NAb）对 RBD 进行了全局丙氨酸扫描，其中包括几种广泛反应性的 NAb。此外，我们通过重组蛋白和假病毒评估了 RBD 关键位置的单个自然突变或共同突变对中和逃逸和 ACE2 结合功能的影响。
鉴定出 RBD 四个独立抗原表位（1 至 4）的 33 个氨基酸位置，这些位置是 RBD 抗原变化的有价值的指标。全面的逃逸突变图谱不仅证实了广泛流行的携带重要免疫逃逸 RBD 突变（如 K417N、E484K 和 L452R）的株系，而且还发现了新的免疫逃逸赋予突变，如 F486L、N450K、F490S 和 R346S。值得注意的是，这些逃逸突变不会影响 RBD 与 ACE2 的结合亲和力，其中 L452R 甚至增强了结合。此外，我们表明，B.1.351 中存在的 RBD 共同突变 K417N、E484K 和 N501Y 对 NAb 和大流行早期的人类恢复期血浆的抵抗力更强，这可能是由于累加效应。相反，B.1.617.1 变体中存在的双重突变 E484Q 和 L452R 表现出部分抗体逃逸，没有证据表明存在累加效应。
本研究提供了针对中和抗体识别、抗原保守性和 RBD 构象的决定因素的全面了解。深入的逃逸图谱可能对 SARS-CoV-2 免疫逃逸变体的前瞻性监测具有价值。应特别注意在不同主要抗原表位处共同突变的积累。最后，这里描述的新型广泛反应性 NAb 代表了预防和治疗 COVID-19 的新潜在机会。
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