Project-002

项目-002

• 批准号: 10794919
• 负责人: Richard Thomas Wyatt
• 金额: $ 50.33万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-02-04 至 2026-01-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10794919
• 关键词: Project; 002

Summary Circulating (tier 2) HIV-1 variants are highly resistant to antibody-mediated neutralization, making broadly effective vaccine design a major challenge. Encouragingly, work in our previous HIVRAD program demonstrated that Ab responses capable of cross-neutralizing multiple heterologous tier 2 viruses were elicited by targeted N-glycan deletion priming and heterologous glycan restorative Env trimer-liposome boosting. The isolation of two monoclonal antibodies with broadly neutralizing activity from these studies and high-resolution structures in complex with native-like trimers revealed that one mAb targeted the conserved CD4 binding site (CD4bs) and the other targeted the gp41:gp120 interface region, substantiated this result. In the current application, we will build on these efforts by evaluating responses elicited by well-ordered, novel, trimer-based immunogens inoculated into Indian origin rhesus macaques. In addition to the use pf protein-based trimers, we will test trimer platforms based on administration of mRNA lipid nanoparticles as described in Project 1. In Project 2, we will characterize B cell responses elicited by vaccine regimens evaluated in Indian origin rhesus macaques in Core B by rapid, high-throughput monoclonal antibody (mAb) isolation to define the targeted epitopes and guide both the choice of boosting immunogens and, if needed, trimer redesign by eliminating or masking unwanted non-neutralizing immunodominant responses for subsequent immunization studies. We will interact with the investigators in Core C, who also will generate information about immunodominant Ab responses through their negative stain electron microscopy (nsEM)-based method for evaluation of vaccine-induced serum responses. Using the Env-specific mAbs, we will further determine how Env-specific antibody lineages evolve over time using Next Generation Sequencing (NGS) and IgDiscover, a computational tool optimized for use in rhesus macaques. We will generate individualized databases of macaque germline VDJ alleles for precise gene assignments, which is necessary for correct conclusions about Ab affinity maturation and SHM levels. We will further use the IgDiscover results and the new haplotype module to investigate if certain alleles, or combinations of alleles, predispose to elicitation of neutralizing Ab responses in either adult of juvenile macaques. We will investigate the level of expansion of neutralizing and non-neutralizing Ab lineages over time and ask if vaccine-induced neutralizing Ab lineages persist in long-lived immune compartments such as memory B cells and plasma cells, which are critical for the long-term protective effects of vaccines.

总结 循环（2级）HIV-1变异体对抗体介导的中和具有高度抗性，使得广泛的 有效疫苗设计是一项重大挑战。令人鼓舞的是，在我们以前的HIVRAD计划中工作 证明了能够交叉中和多种异源2级病毒的Ab应答， 通过靶向N-聚糖缺失引发和异源聚糖恢复性Env三聚体-脂质体引发 增强从这些研究中分离出两种具有广泛中和活性的单克隆抗体 与天然样三聚体复合的高分辨率结构显示，一种mAb靶向 保守的CD 4结合位点（CD 4 bs）和另一个靶向gp 41：gp 120界面区， 证实了这一结果。在当前的应用程序中，我们将通过评估响应来构建这些工作 由接种到印度起源的恒河猴中的有序的、新颖的、基于三聚体的免疫原引起。 除了使用基于蛋白质的三聚体之外，我们还将测试基于以下施用的三聚体平台： mRNA脂质纳米颗粒，如项目1中所述。 在项目2中，我们将描述印度来源的疫苗方案引起的B细胞应答 通过快速、高通量单克隆抗体（mAb）分离， 定义靶向表位，并指导加强免疫原的选择，如果需要的话，指导三聚体的选择 通过消除或掩蔽不需要的非中和免疫显性应答进行重新设计， 随后的免疫研究。我们将与核心C的研究人员互动，他们也将产生 免疫显性抗体反应负染电镜观察 用于评价疫苗诱导的血清应答的基于nsEM的方法。使用Env特异性mAb， 我们将进一步确定Env特异性抗体谱系如何随着时间的推移而演变， 测序（NGS）和IgDiscover，一种优化用于恒河猴的计算工具。我们将 生成猕猴生殖系VDJ等位基因的个性化数据库，用于精确的基因分配， 对于正确的Ab亲和力成熟和SHM水平的结论是必要的。我们将进一步利用 IgDiscover结果和新的单倍型模块，以调查某些等位基因或 等位基因，倾向于在成年或幼年猕猴中引发中和抗体应答。我们 将研究中和和非中和抗体谱系随时间的扩增水平， 问疫苗诱导的中和抗体谱系是否持续存在于长寿命的免疫区室中， 如记忆B细胞和浆细胞，它们对疫苗的长期保护作用至关重要。