■严重急性呼吸道综合症冠状病毒2(SARS-CoV-2),2019年冠状病毒病的病原体(COVID-19),对全球健康和经济都产生了重大影响。针对SARS-CoV-2刺突(S)蛋白的S1亚基开发了许多病毒中和抗体,以阻断病毒与宿主细胞的结合,并被授权用于控制COVID-19大流行。然而,SARS-CoV-2S1亚基的频繁突变使得免疫规避变体的出现成为可能.为了应对这些挑战,已经研究了靶向相对保守的S2亚基及其表位的广泛中和抗体作为抗体治疗剂和通用疫苗。
■我们通过用β-丙内酯灭活的SARS-CoV-2(IAV)免疫BALB/c小鼠以产生B细胞杂交瘤来启动本研究。随后使用表达S2-ECD结构域的HEK293T细胞筛选这些杂交瘤。选择产生抗S2抗体的杂交瘤,我们对这些抗S2抗体作为抗病毒药物和用于研究和诊断的多功能工具的潜力进行了全面评估。
■在这项研究中,我们提出了一种新的S2特异性抗体,4A5,从用灭活的SARS-CoV-2免疫的BALB/c小鼠中分离。与其他β-CoV相比,4A5对SARS-CoV-2S2亚基表现出特异性亲和力。图4A5结合七肽-repeat1(HR1)和茎螺旋(SH)区之间的表位区段F1109-V1133。4A5表位在SARS-CoV-2变体中高度保守,在融合前和融合后S蛋白中均具有显着的构象特征。值得注意的是,4A5表现出针对变体的广泛中和活性并触发Fc增强的抗体依赖性细胞吞噬作用。
这些发现为新型抗体疗法和下一代疫苗设计提供了有希望的途径。4A5具有独特的结合特性和广泛的中和能力,为SARS-CoV-2变体带来的挑战提供了一个潜在的解决方案,并强调了靶向保守的S2亚基在对抗COVID-19中的重要性。
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative agent of coronavirus disease 2019 (COVID-19), had a major impact on both the global health and economy. Numerous virus-neutralizing antibodies were developed against the S1 subunit of SARS-CoV-2 spike (S) protein to block viral binding to host cells and were authorized for control of the COVID-19 pandemic. However, frequent mutations in the S1 subunit of SARS-CoV-2 enabled the emergence of immune evasive variants. To address these challenges, broadly neutralizing antibodies targeting the relatively conserved S2 subunit and its epitopes have been investigated as antibody therapeutics and universal vaccines.
We initiated this study by immunizing BALB/c mice with β-propiolactone-inactivated SARS-CoV-2 (IAV) to generate B-cell hybridomas. These hybridomas were subsequently screened using HEK293T cells expressing the S2-ECD domain. Hybridomas that produced anti-S2 antibodies were selected, and we conducted a comprehensive evaluation of the potential of these anti-S2 antibodies as antiviral agents and versatile tools for research and diagnostics.
In this study, we present a novel S2-specific antibody, 4A5, isolated from BALB/c mice immunized with inactivated SARS-CoV-2. 4A5 exhibited specific affinity to SARS-CoV-2 S2 subunits compared with those of other β-CoVs. 4A5 bound to epitope segment F1109-V1133 between the heptad-repeat1 (HR1) and the stem-helix (SH) region. The 4A5 epitope is highly conserved in SARS-CoV-2 variants, with a significant conformational feature in both pre- and postfusion S proteins. Notably, 4A5 exhibited broad neutralizing activity against variants and triggered Fc-enhanced antibody-dependent cellular phagocytosis.
These findings offer a promising avenue for novel antibody therapeutics and insights for next-generation vaccine design. The identification of 4A5, with its unique binding properties and broad neutralizing capacity, offers a potential solution to the challenge posed by SARS-CoV-2 variants and highlights the importance of targeting the conserved S2 subunit in combating the COVID-19.