Immunogen design to elicit polyclonal bNAb responses to the V3 glycan supersite

项目摘要 我们最近成功地设计了能够广泛中和HIV的免疫原 抗体（bNA B）前体，在免疫动物中扩增这些B细胞，并选择关键的 体细胞超突变对中和宽度的发展至关重要。虽然前景看好， 进展仅限于沿着漫长而复杂的进化途径的沿着第一步，因为bNAb通常 有大量的体细胞超突变虽然只有这些突变的一个子集是必要的， 获得广度，目前还不清楚哪些突变代表Env识别的共同特征， 可以用免疫原靶向的不同bNAb谱系。识别常见的成熟事件， 由靶向相同bNAb表位的多个谱系共享的bNAb发育对于 针对bNAb前体的不同库的疫苗设计策略。一旦血统从不同的开始 点与引发免疫原接合并扩大，增强免疫原将需要引导它们的 成熟，以获得广泛认可的组成部分。共享识别特征的识别是 只有通过比较靶向相同表位的大量bNAb谱系才有可能。通过我们目前的 HIVRAD，我们已经表明用于免疫原设计的灵长类bNA B B细胞谱系的一个来源可以是SHIV-1。 感染的恒河猴（RM）。我们已经证明，六分之一的感染了SHIV的RM已经继续 以产生bNAb应答。超过三分之一的这些应答已经靶向V3-聚糖表位， bNAb在人类感染中最常见的靶向部位。V3聚糖bNAb的结构测定 从SHIV感染的RM分离的人V3聚糖bNAb显示出与人V3聚糖bNAb的惊人相似性，表明显著的 V3聚糖表位识别的趋同进化。因此，我们现在有了一个可靠地诱导V3的系统 聚糖bNAb，其可以告知引发靶向V3聚糖表位的bNAb的免疫原的设计。的 项目3的中心假设是bNAb谱系如何识别V3聚糖存在共性 表位，并且免疫原可以被设计为选择这些共享的识别特征， 在顺序免疫方案中组合将引发bNAb。在目标1中，我们将确定 SHIV和人感染诱导的V3聚糖之间共享的V3聚糖表位识别机制 bNAb使用B细胞谱系追踪和快速结构分析来定义用于免疫原设计的模板。 保持战略定力在目标2中，我们将设计可以在恒河猴中引发多种V3聚糖谱系的免疫原， 使用首先选择具有长CDRH 3的B细胞的新设计目标对人进行筛选。在目标3中，我们将使用病毒 在哺乳动物细胞表面展示SHIV感染猴作为Env文库变体来源的测序， 设计增强免疫原，其选择V3聚糖bNAb识别中的共享成熟事件。项目 3将通过产生主要和增强免疫原来影响该领域，在RM中进行测试，这将导致疫苗 能够在人体中引发bNAb的候选物。

PROJECT SUMMARY We have demonstrated recent success in designing immunogens that can bind HIV broadly neutralizing antibodies (bNAb) precursors with high affinity, expand these B cells in immunized animals, and select for key somatic hypermutations essential for the development of neutralization breadth. Although promising, the progress has been limited to just the first steps along long and complex evolutionary pathways as bNAbs typically have extensive numbers of somatic hypermutation. While only a subset of these mutations is necessary for acquisition of breadth, it is unclear which mutations represent shared features of Env recognition between different bNAb lineages that can be targeted with immunogens. Identifying the common maturational events in bNAb development shared by multiple lineages that target the same bNAb epitope is of critical importance for vaccine design strategies that target a diverse pool of bNAb precursors. Once lineages from diverse starting points are engaged and expanded with priming immunogens, boosting immunogens will need to direct their maturation to acquire the components of broad recognition. The identification of shared recognition features is only possible by comparing a large set of bNAb lineages that target the same epitope. Through our current HIVRAD, we have shown one source of primate bNAb B cell lineages for immunogen design can be SHIV- infected rhesus macaques (RMs). We have demonstrated that one in six RMs infected by SHIVs have gone on to generate bNAb responses. Over one third of these responses have been targeted to the V3-glycan epitope, the most common site targeted by bNAbs in human infection. Structural determination of a V3 glycan bNAb isolated from a SHIV infected RM revealed striking similarities to a human V3 glycan bNAb indicating remarkable convergent evolution of V3 glycan epitope recognition. Thus, we now have a system for reliably inducing V3 glycan bNAbs that can inform the design of immunogens that elicit bNAbs targeting the V3 glycan epitope. The central hypothesis of Project 3 is that commonalities exist in how bNAb lineages recognize the V3 glycan epitope, and that immunogens can be designed to select for these shared recognition features which when combined in a sequential immunization regimen will elicit bNAbs. In Aim 1, we will identify the common mechanisms of V3 glycan epitope recognition shared between SHIV and human infection-induced V3 glycan bNAbs to define templates for immunogen design using B cell lineage tracing and a rapid structural determination. In Aim 2, we will design immunogens that can prime multiple V3 glycan lineages in rhesus and humans using a novel design goal of selecting first for B cells with long CDRH3s. In Aim 3, we will use viral sequencing of SHIV infected monkeys as a source for Env library variants in mammalian cell surface display to design boosting immunogens that select for shared maturational events in V3 glycan bNAb recognition. Project 3 will impact the field by generating prime and boosting immunogens, tested in RMs, that will lead to vaccine candidates capable of eliciting bNAbs in humans.