Design and evaluation of pan-CoV vaccines

总结 作为泛冠状病毒疫苗（PanCoVac）联盟的一部分，研究项目1（RP 1）的目标是 “泛冠状病毒疫苗的设计和评估”。B和T细胞的全面表征 疫苗接种反应将由研究项目2（RP 2，“对泛冠状病毒的免疫反应 疫苗。在RP 1的目标1（“将免疫反应集中在刺突蛋白的茎”）中， 将测试策略，以增加针对保守的免疫亚显性的抗体水平。 SARS-CoV-2刺突蛋白（主要病毒抗原）茎中的表位，同时避免强抗体 针对刺突蛋白头部中的免疫显性表位的应答。这些战略包括 由SARS-CoV-2免疫亚优势茎表位和免疫优势头组成的嵌合刺突 未遇到的冠状病毒的表位，“稀释”抗体反应的免疫显性表位， 使用在关键氨基酸位置具有多个突变的刺突蛋白的鸡尾酒， 免疫显性头部表位，“反向抗原”，其中刺突蛋白将呈递给免疫系统。 系统处于反向方向（因此变得更容易接近），而“无头”刺突蛋白缺乏 免疫显性头部表位的部分。这些方法中的一些可以组合测试， 可以用基于祖先重建的序列优化的干表位进行测试（计算 推断最可能的共同祖先序列的方法）。在目标2（“引起广泛反应 针对刺突蛋白头部的免疫应答“），将测试指导免疫应答的策略 远离最具序列多样性的表位而朝向头部中更保守的表位。此外，本发明还提供了一种方法， 保守的HED表位将被序列优化以被交叉保护性抗体识别。这些 修饰将被引入到不同的刺突蛋白中，以及不同的突变刺突蛋白的疫苗鸡尾酒。 然后测试蛋白质的免疫原性和保护效力。目标1和2中的所有抗原将被 与结构生物学家合作设计，并且对于选定的抗原和/或抗原/抗体复合物， 将进行X射线晶体学和Cryo-EM。在目标3（“免疫原性和保护效力， 广泛反应性抗原“），将在小鼠中测试新疫苗接种物的免疫原性;来自 将接种的小鼠提供给RP 2进行B和T细胞分析。选定的候选疫苗（ 更广泛的免疫反应）将在小鼠和叙利亚仓鼠中测试它们提供保护的能力 对抗不同的冠状病毒对于提供广泛保护的候选疫苗（与对照相比），我们 还将评估免疫反应的持久性，以及疫苗接种对病毒传播的影响。 此外，这些疫苗候选物将在mRNA脂质纳米颗粒疫苗平台中进行测试，该平台由 第一三共制药公司

SUMMARY As part of the Pan-Coronavirus Vaccine (PanCoVac) Consortium, the goal of Research Project 1 (RP1) is the ‘Design and evaluation of pan-CoV vaccines’. A comprehensive characterization of the B and T cell responses to vaccination will be carried out by Research Project 2 (RP2, ‘Immunological responses to pan-CoV vaccines. In Aim 1 of RP1 (‘Focusing immune responses towards the stem of the spike protein’), several strategies will be tested to increase the levels of antibodies directed at the conserved, immune-subdominant epitopes in the stem of the SARS-CoV-2 spike protein (to major viral antigen), while avoiding strong antibody responses directed at immunodominant epitopes in the head of the spike protein. These strategies include chimeric spikes composed of SARS-CoV-2 immune-subdominant stem epitopes and immunodominant head epitopes of unencountered coronaviruses, ‘outdiluting’ antibody responses to the immunodominant epitopes by using cocktails of spike proteins with multiple mutations in key amino acid positions, glycan-shielding of immunodominant head epitopes, ‘inverted antigens’ in which the spike protein will be presented to the immune system in an inverted orientation (and thus become more accessible), and ‘headless’ spike proteins lacking portions of the immunodominant head epitopes. Some of these approaches may be tested in combination, and may be tested with sequence-optimized stem epitopes based on ancestral reconstruction (a computational approach to deduce the most likely common progenitor sequence). In Aim 2 (‘Eliciting broadly reactive immune responses to the head of the spike protein’), strategies will be tested to direct the immune responses away from the most sequence-diverse epitopes and towards more conserved epitopes in the head. In addition, conserved heard epitopes will be sequenced-optimized to be recognized by cross-protective antibodies. These modifications will be introduced into diverse spike proteins, and vaccine cocktails of diverse, mutant spike proteins will then be tested for their immunogenicity and protective efficacy. All antigens in Aims 1 and 2 will be designed in collaboration with a structural biologist, and for selected antigens and/or antigen/antibody complexes, X-ray crystallography and Cryo-EM will be carried out. In Aim 3 (‘Immunogenicity and protective efficacy of broadly reactive antigens’), the novel vaccinates will be tested in mice for their immunogenicity; samples from vaccinated mice will be provided to RP2 for B and T cell analysis. Selected vaccine candidates (those with broader immune responses) will next be tested in mice and Syrian hamsters for their ability to provide protection against different coronaviruses. For candidate vaccines that provide broad protection (compared to controls), we will also assess the durability of immune responses, and the effect of vaccination on virus transmissibility. Moreover, these vaccinate candidates will be tested in an mRNA lipid nanoparticle vaccine platform provided by Daiichi Sankyo, a pharmaceutical company.