Antibody and viral determinants of protection against CMV infection following gB/MF59 vaccination

项目摘要/摘要 巨细胞病毒(CMV)垂直传播是先天性感染最常见的单一原因 在世界范围内，往往导致耳聋和神经发育迟缓的苦恼儿童。CMV相关 妊娠期原发母体感染后，神经系统并发症更加频繁和严重， 因此，预防孕妇在怀孕期间感染CMV的疫苗是一种潜在的策略 降低婴幼儿先天疾病的发生率。测试的最成功的孕产妇免疫方案 迄今为止，一种由CMV糖蛋白B(GB)与MF59佐剂结合组成的亚单位疫苗， 在多个II期临床试验中取得了50%的有希望的疗效。然而，改进这种疫苗的努力 由于对预防CMV感染的决定因素的不完全了解，已经减慢了速度。 以前的研究表明，母体巨细胞病毒中和的程度与 抗体反应和先天性巨细胞病毒感染的风险。与这些早期的研究一致，我们的实验室 利用临床队列和一种新的先天性非人类灵长类动物模型产生了令人信服的数据 CMV传播，表明具有中和CMV病毒能力的抗体可以预防 在子宫内传播。然而，目前尚不清楚抗体反应是否仅针对gB蛋白，以及 而不是其他CMV糖蛋白，足以构成预防先天性CMV的有效疫苗。 CMV gB晶体结构的最新发现及其靶向蛋白结构域的鉴定 中和抗体使研究GB/MF59诱导的保护性抗体反应成为可能。 重要的是，CMV表面糖蛋白如gB具有高度的结构多样性，这表明 这些蛋白质的突变可能是逃避疫苗引发的免疫反应的一种机制。我假设 母体免疫策略的疫苗效力取决于1)中和抗体 靶向gB结构域和2)从这些保护性免疫反应中逃避病毒免疫。在接下来的两年 作为我的博士论文项目的一部分，我建议使用新的方法和切割- 边缘疫苗学技术评估抗体和病毒决定因素的疫苗保护使用 独立的GB/MF59第二阶段试验队列。首先，我将评估中和抗体是否可以预测CMV 获取风险，并确定这些保护性抗体的表位特异性。第二，通过深入 病毒变异的测序和随后的病毒“筛子分析”，我将在 确定疫苗诱导的免疫反应的病毒免疫逃避是否与gB区基因座有关 疫苗失败。最后，我将设计并鉴定含有人的嵌合恒河猴CMV病毒 GB中和结构域，用于验证已识别的多态位点上的病毒免疫逃避。这些嵌合体 病毒将在未来用于我们实验室开发的非人类灵长类动物模型，为 为合理设计和测试以糖蛋白为基础的消除先天性巨细胞病毒感染的疫苗。

PROJECT SUMMARY/ABSTRACT Vertical transmission of cytomegalovirus (CMV) is the single most common cause of congenital infection worldwide, often resulting in deafness and neurodevelopmental delay for afflicted children. CMV related neurologic complications are more frequent and severe following primary maternal infection during pregnancy, and therefore a vaccine to prevent maternal acquisition of CMV during pregnancy is a potential strategy to reduce the incidence of infant congenital disease. The most successful maternal immunization regimen tested to date is a subunit vaccine consisting of CMV glycoprotein B (gB) combined with MF59 adjuvant, which achieved a promising 50% efficacy in multiple phase II clinical trials. However efforts to improve this vaccine have been slowed by an incomplete understanding of the determinants of protection against CMV infection. Previous studies have revealed a direct correlation between magnitude of the maternal CMV neutralizing antibody response and risk of congenital CMV infection. Concordant with these earlier studies, our laboratory has generated compelling data using both clinical cohorts and a novel nonhuman primate model of congenital CMV transmission, demonstrating that antibodies with the ability to neutralize CMV viruses may protect against in utero transmission. Yet it remains unclear whether antibody responses targeting the gB protein alone, and not other CMV glycoproteins, are sufficient for an effective vaccine to prevent congenital CMV. Recent discovery of the CMV gB crystal structure and identification of distinct protein domains targeted by neutralizing antibodies have made it possible to investigate gB/MF59-elicited protective antibody responses. Importantly, CMV surface glycoproteins such as gB have a high degree of structural diversity, suggesting that mutation of these proteins may be a mechanism to evade vaccine-elicited immune responses. I hypothesize that vaccine efficacy of a maternal gB immunization strategy is dependent upon 1) neutralizing antibodies targeting gB domains and 2) viral immune evasion from these protective immune responses. Over the next two years as a component of my Ph.D. dissertation project, I propose to employ novel approaches and cutting- edge vaccinology techniques to assess both antibody and viral determinants of vaccine protection using two independent gB/MF59 phase II trial cohorts. First, I will assess whether neutralizing antibodies predict CMV acquisition risk and define the epitope specificities of those protective antibodies. Secondly, through deep sequencing of viral variants and subsequent viral “sieve analysis,” I will investigate sequence diversity at the loci of gB domains to determine if viral immune evasion of vaccine-elicited immune responses is associated with vaccine failure. Finally, I will engineer and characterize chimeric rhesus CMV viruses containing human gB neutralizing domains for validation of viral immune evasion at identified polymorphic sites. These chimeric viruses will be used in the future in a nonhuman primate model developed by our laboratory, setting the stage for rational design and testing of a glycoprotein-based vaccine for the elimination of congenital CMV infection.