The origin and future protective activity of SARS-CoV-2 RBD specific neutralizing antibodies

项目总结 到目前为止，SARS-CoV-2已感染超过1.38亿人，导致超过280万人死亡，其中 预计疫情还将持续数月，病毒将在地方性疫情中持续数年， 新出现的关注变种(VOC)加剧了这一问题。尽管有几种疫苗正在广泛使用，但它 目前尚不清楚它们是否能够诱导对新出现的VOC产生有效的长期免疫。高效的抗病毒药物 SARS-CoV-2的病毒疗法仍然难以捉摸，尽管有几种针对SARS-CoV-2的单抗 S(Spike)蛋白的受体结合结构域(Rbd)已被授予轻至中度感染的EUA， 它们对严重疾病的有效性尚不明显。随着全球疫苗接种步伐的缓慢， 有限的抗病毒使用/效力，以及抗原漂移变异的出现，这一大流行的轨迹和 该病毒未来的死灰复燃令人极为担忧。对诱导机制的基本认识 而维持对SARS-CoV-2的保护性体液免疫将是其缓解的关键。病毒现在是 分为几个分支，出现了大量的VOC，以及包括我们的工作和其他一些迹象 这种抗原漂移是病毒逃离免疫压力和传播性增加的结果。漂移 在RBD内是最令人担忧的，因为它可以增强病毒的传染性，并否定 可能因以前接种疫苗或感染而发展成的nabs。已有大量报道称 患者反复感染SARS-CoV-2，以及完全接种疫苗的个人出现突破性感染， 突显了自然获得的SARS-CoV-2免疫系统的缺陷。利用我们合理设计的 RBD/RBD-ACE2融合蛋白变异体，我们鉴定了 人RBD特异性B细胞，并分离出几株有效的RBD特异性人中和单抗 (NmAbs)(IC50；50 ng/ml)抗SARS-CoV-2正在进入吸入性1/2期临床试验 在未来几个月内交货。我们假设在RBD内，高度保守的区域(RBD-CR)， 广泛有效的体液保护所需的表位被可变区(RBD- VR)是结构动态的，对抗原漂移高度敏感。此外，我们假设RBD-VR 缓解有效和广泛的RBD-CR特异性体液反应的发展 RBD-CR的免疫优势和直接封闭。这种RBD-CR/RBD-VR进化动态很可能 调节针对未来病毒变异的体液反应的持续保护(或失败)。我们将1)定义 人类RBD特异性中和抗体反应的本体论和表型多样性，2)定义 维持ACE2结合的动力学和抑制RBD进化的免疫压力，以及3) 测定RBD抗体对SARS-CoV-2漂移的耐受性和贡献。界定自然感染的限度和 疫苗接种诱导RBD中和抗体驱动抗原漂移并提供对分化的保护 SARS-CoV-2病毒将为下一代SARS-CoV-2疫苗和疗法的开发提供信息。

PROJECT SUMMARY SARS-CoV-2 has infected over 138 million people and resulted in over 2.8 million deaths so far, with the expectation the pandemic will continue for many more months, and the virus will persist endemically for years, exacerbated by emerging variants of concern (VoC). Although several vaccines are being used wide-spread, it is unclear if they will be able to induce effective long-term immunity against emerging VoC. Highly effective anti- viral therapeutics for SARS-CoV-2 remain elusive, although several monoclonal antibodies (mAbs) targeting the Receptor Binding Domain (RBD) of the Spike (S) protein have been granted EUA for mild to moderate infection, their effectiveness against severe disease has not yet been evident. With the slow pace of global vaccination, limited anti-viral use/efficacy, and the emergence of antigenic drift variants, the trajectory of this pandemic and future resurgences of the virus is of great concern. Fundamental understanding of the mechanisms of inducing and sustaining protective humoral immunity to SARS-CoV-2 will be critical to its mitigation. The virus is now classified into several clades, numerous VoC emerging, and indications including our work and others that some of this antigenic drift is the result of the virus escaping from immune pressure and increased transmissibility. Drift within the RBD is of the utmost concern as it can enhance the infectivity of the virus and negate the activity of NAbs that may have developed from previous vaccination or infection. Numerous reports have emerged of repeated SARS-CoV-2 infections in patients, and breakthrough infections in fully vaccinated individuals, highlighting the imperfection of naturally acquired SARS-CoV-2 immunity. Utilizing our rationally designed RBD/RBD-ACE2 fusion protein variants, we have identified epitopic and phenotypic heterogeneity amongst RBD-specific human B cells and have isolated several potent RBD-specific human neutralizing monoclonal Abs (NmAbs) (IC50<50 ng/ml) against SARS-CoV-2 which are entering into a Phase 1/2 clinical trial using inhaled delivery in the coming months. We hypothesize that within RBD, the highly conserved regions (RBD-CR), epitopes desirable for mediating broad and potent humoral protection, are surrounded by variable regions (RBD- VR) that are structurally dynamic and highly susceptible to antigenic drift. Further, we hypothesize that RBD-VR mitigate the development of potent and broad RBD-CR specific humoral responses through their immunodominance and direct occlusion of RBD-CR. This RBD-CR/RBD-VR evolutionary dynamic is likely to regulate the sustained protection (or failure) of humoral responses against future viral variants. We will 1) define the ontological and phenotypic diversity of the human RBD-specific neutralizing antibody response, 2) define the dynamics of maintenance of ACE2 binding and immunological pressure on constraining RBD evolution, and 3) determine RBD Ab tolerance for and contribution to SARS-CoV-2 drift. Defining the limits of natural infection and vaccination induced RBD neutralizing antibodies to drive antigenic drift and confer protection from divergent SARS-CoV-2 viruses will inform the development next generation SARS-CoV-2 vaccines and therapeutics.