Design and evaluation of pan-CoV vaccines

摘要 作为泛冠状病毒疫苗(PanCoVac)联盟的一部分，研究项目1(RP1)的目标是 《泛冠状病毒疫苗的设计与评价》。B细胞和T细胞的综合特征 对接种疫苗的反应将由研究项目2(RP2)进行，即对泛冠状病毒的免疫反应 疫苗。在RP1的目标1(将免疫反应集中到尖峰蛋白的茎上)中，有几个 将测试提高针对保守的、免疫亚显性的抗体水平的策略 SARS-CoV-2刺突蛋白茎中的表位(针对主要病毒抗原)，同时避免强抗体 针对Spike蛋白头部的免疫优势表位的反应。这些战略包括 SARS-CoV-2免疫优势主干表位与免疫优势头部嵌合刺突 未见过的冠状病毒表位，通过以下方式“稀释”抗体对免疫优势表位的反应 使用关键氨基酸位置有多个突变的Spike蛋白的鸡尾酒，糖链屏蔽 免疫优势头部表位，即“反向抗原”，其中的尖峰蛋白将呈递给免疫 系统处于倒置方向(因此变得更容易访问)，并且缺乏无头部的尖峰蛋白 部分免疫优势头部表位。其中一些方法可能会进行组合测试，并且 可以用基于祖先重建(计算性的)的序列优化的干细胞表位进行测试 推断最可能的共同祖先序列的方法)。在目标2中(‘引起广泛的反应 对尖峰蛋白头部的免疫反应)，将测试指导免疫反应的策略 远离序列最多样化的表位，转向头部更保守的表位。此外, 将对保守的Hed表位进行测序-优化以供交叉保护抗体识别。这些 修饰将被引入不同的Spike蛋白，以及不同的、突变的Spike的疫苗鸡尾酒中 然后将测试蛋白质的免疫原性和保护效力。AIMS 1和AIMS 2中的所有抗原 与结构生物学家合作设计，并针对选定的抗原和/或抗原/抗体复合体， 将进行X射线结晶学和低温电子显微镜检查。在目标3(‘免疫原性和保护效力 ，新型疫苗将在小鼠身上进行免疫原性测试；样本来自 接种的小鼠将被提供给RP2进行B和T细胞分析。选定的候选疫苗(具有 更广泛的免疫反应)接下来将在小鼠和叙利亚仓鼠身上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性的影响。 此外，这些候选疫苗将在由提供的mrna脂质纳米粒疫苗平台上进行测试。 第一三共，一家制药公司。

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.