Project 2: PanbetaCoV protein vaccine design

摘要-项目2 动物贝塔冠状病毒(BetaCov)，如引起SARS-CoV-1、MERS和现在的SARS的那些病毒- CoV-2对人类健康构成重大、持续的威胁。疫苗可以提供预先存在的免疫力 未来的人畜共患病冠状病毒。目前有效的SARS-CoV-2疫苗的一个主要限制是它们 可能只会保护与中国使用的武汉分离株几乎相同的SARS-CoV-2分离株 疫苗。因此，世界仍然容易受到SARS-CoV-2变异逃逸变种的影响 以及未来可能发生大流行的动物贝塔冠状病毒溢出事件。疫苗研发 迫切需要解决这一脆弱性的方案。项目2将支持该计划开发的总体目标 预防未来贝塔冠状病毒大流行和SARS-CoV-2变异的疫苗 设计可诱导广谱中和抗体(BNab)的疫苗免疫原，以提供保护 对贝塔冠状病毒属的免疫力。存在结构保守的中和表位 在贝塔冠状病毒尖峰蛋白上，我们假设引起对这些蛋白的广泛保护性抗体的疫苗 网站可以开发。在初步研究中，我们分离出了交叉反应中和抗体 SARS-CoV-1或SARS-CoV-2-感染或康复的人类，并使用结构测定精确地 确定它们的BetaCoV bNab表位。我们的疫苗设计策略包括使用计算和结构- 基于设计技术来开发免疫原，优先呈现这些和另外 确定了bNab表位。每种免疫原设计都会展示在纳米颗粒的表面，以增强 抗原亲和力和抗原向生发中心和滤泡树突状细胞的转运。我们已经展示了一个 我们使用多聚化SARS-CoV-2受体结合域(RBD)的策略的概念验证- 铁蛋白纳米颗粒，可诱导高滴度抗体，中和不同的人类和 动物肉瘤病毒(2b组)在免疫猕猴体内。在目标1中，我们将分离新的抗体以确定 额外的bNab表位，并建立BetaCoV S蛋白表面的全谱区域 能向2b和2c组的动物BetaCov诱导bNAb。在目标2中，我们将利用尖端技术 计算机免疫原设计方法设计广泛中和表位的免疫原。这些 免疫原使尖峰蛋白固定在暴露中和表位或尖峰的构象中 亚基被截断或重新浮出水面，以去除不相关的、非中和的或贝塔冠状病毒株特异性的亚基 中和表位，同时保留广泛的中和表位。在目标3中，我们将免疫野生型小鼠 与这些新的免疫原结合，并评价它们诱导人BetaCoV bNAb的能力，以保护 动物来自病毒的挑战。项目2将通过产生交叉反应中和抗体来影响该领域 这可能被翻译成治疗学，阐明新的BetaCoV尖峰上的广泛中和表位 蛋白质，并确定了诱导保护性免疫的表位集中免疫原的设计原则 为了BetaCov。

Abstract – Project 2 Animal betacoronaviruses (betaCoVs), such as those that gave rise to SARS-CoV-1, MERS, and now SARS- CoV-2, represent a significant, continuous threat to human health. Vaccines can provide pre-existing immunity to future zoonotic coronaviruses. A major limitation of the current effective vaccines to SARS-CoV-2 is that they likely will only protect SARS-CoV-2 isolates that are nearly identical to the Wuhan isolate used in the vaccines. Thus, the world remains vulnerable to SARS-CoV-2 mutational escape variants and from future animal betacoronavirus spillover events that have pandemic potential. Vaccine development that addresses this vulnerability is urgently needed. Project 2 will support this Program’s overall goal to develop vaccines that will prevent future betaCoV pandemics and SARS-CoV-2 escape variants by designing vaccine immunogens that elicit broadly neutralizing antibodies (bnAb) to provide protective immunity across the betaCoV genus. Structurally conserved neutralizing epitopes exist on betaCoV spike proteins, and we hypothesize that vaccines that elicit broadly protective antibodies to these sites can be developed. In preliminary studies, we have isolated cross-reactive neutralizing antibodies from SARS-CoV-1 or SARS-CoV-2-infected or recovered humans and used structural determination to precisely define their betaCoV bnAb epitopes. Our vaccine design strategy involves using computational and structural- based design techniques to develop immunogens that preferentially present these and additionally identified bnAb epitopes. Each immunogen design will be displayed on the surface of nanoparticles to enhance antigen avidity and antigen trafficking to germinal centers and follicular dendritic cells. We have demonstrated a proof-of-concept for our strategy using a multimerized SARS-CoV-2 receptor binding domain (RBD)- ferritin nanoparticle that induces high titers of antibodies that neutralize diverse human and animal Sarbecoviruses (group 2b) in immunized macaques. In Aim 1, we will isolate new antibodies to define additional bnAb epitopes and establish the full spectrum of regions on the surface of betaCoV S proteins that are capable of inducing bnAbs to animal betaCoVs in groups 2b and 2c. In Aim 2, we will utilize cutting-edge computational immunogen design methods to design broadly neutralizing epitope-focused immunogens. These immunogens have the spike protein fixed in conformations that expose neutralizing epitopes or are spike subunits that are truncated or resurfaced to remove irrelevant, non-neutralizing, or betaCoV strain-specific neutralizing epitopes while retaining broadly neutralizing epitopes. In Aim 3, we will immunize wild-type mice with these novel immunogens and evaluate their ability to induce human betaCoV bnAbs that can protect animals from viral challenge. Project 2 will impact the field by generating cross-reactive neutralizing antibodies that could be translated into therapeutics, elucidating new broadly neutralizing epitopes on the betaCoV spike protein, and defining the design principles for epitope-focused immunogens that elicit protective immunity to betaCoVs.