Project 1: The impact of innate immune responses on the development of broadly neutralizing antibodies by vaccination

摘要-项目1 通过HIV疫苗接种诱导广泛中和抗体（bNAb）仍然是一个重大挑战。理性 具有天然HIV包膜三聚体的B细胞谱系疫苗设计已成为最有前途的疫苗 战略然而，尽管靶向相应生殖系B细胞受体的HIV Env糖蛋白三聚体已经被发现， 通过接种成功诱导自体2级中和抗体， 中和异源菌株的能力仍然有限。许多研究表明，疫苗诱导的 通过在疫苗方案中加入佐剂可以增强抗体应答。的潜力 然而，影响bNAb诱导的佐剂在很大程度上未被探索。项目1的长期目标是 鉴定佐剂可增强bNAb的诱导和成熟的机制。我们提出 初步数据表明，用靶向BG 505 GT 1.1的种系免疫恒河猴幼猴（RM） SOSIP与基于TLR7，8的佐剂3M-052在稳定乳液（SE）中混合导致bNAb的诱导 在1.5年时，5只动物中有3只动物的前体抗CD4结合位点（CD4bs），指示VRC01样bNAb。 成人RM研究报告称，在HIV疫苗方案中加入3M-052可促进诱导 以及在种系BCR基因敲入中具有基于TLR3或TLR4的佐剂的HIV疫苗 小鼠与增加的体细胞超突变（SHM）和VRC01样bNAb的诱导相关 前体然而，3M-052或其他基于TLR的佐剂支持Germination中心的机制是不确定的。 活性、SHM、HIV bNAb前体的诱导和/或长寿命浆细胞的发育仍然存在 未知项目1的目标是确定诱导bNAb的初始步骤的决定因素 前体我们假设，与BG 505 GT 1.1 SOSIP三聚体联合给予的佐剂改变了免疫原性。 Env特异性B细胞通过诱导特异性先天反应的发育途径， bNAb前体的亲和力成熟。了解B细胞向bNA B-发育的复杂过程 产生B细胞以响应HIV疫苗接种及其调节将需要广泛的基于组学的方法 该计划将适用。项目1将通过转录组学，单克隆抗体， 细胞（sc）RNA测序，通过CyTOF进行免疫组分析，以及可溶性免疫介质的蛋白质组分析 在血浆中。为了确定bNAb的形成是否会受到佐剂选择的影响，我们将询问 BG 505 GT1.1 SOSIP免疫后bNAb前体的产量是否取决于先天性 由3M-052-SE相对于基于皂苷的佐剂诱导的应答（目的1），并且依赖于佐剂- 淋巴结中的特异性免疫细胞活化和功能（目的2）。与项目2合作，我们将定义 这些先天性反应是如何被肠道细菌调节的（目标3）。结果预计将告知 开发疫苗初免的新靶向免疫调节方法，以优化诱导 的bNAb前体，并提供了一个框架的bNAb靶向HIV疫苗的评价和向下选择。

ABSTRACT - PROJECT 1 The induction of broadly neutralizing antibodies (bNAb) by HIV vaccination remains a major challenge. Rational B-cell-lineage vaccine design with native HIV envelope trimers has emerged as the most promising vaccine strategy. Yet, while HIV Env glycoprotein trimers targeting the respective germline B cell receptor have been successful in inducing autologous tier 2 neutralizing antibodies by vaccination, the induction of antibodies able to neutralize heterologous strains remains limited. Numerous studies have reported that vaccine-induced antibody responses can be enhanced by the inclusion of adjuvants in the vaccine regimen. The potential of adjuvants to impact the induction of bNAb, however, is largely unexplored. The long-term goal of Project 1 is to identify mechanisms by which adjuvants can enhance the induction and maturation of bNAbs. We present preliminary data that immunization of infant rhesus macaques (RM) with the germline targeting BG505 GT1.1 SOSIP mixed with the TLR7,8-based adjuvant 3M-052 in stable emulsion (SE) resulted in the induction of bNAb precursors against the CD4 binding site (CD4bs), indicative of VRC01-like bNAbs, in 3 of 5 animals by 1.5 years. Studies in adult RM have reported that the inclusion of 3M-052 in HIV vaccine regimens promotes the induction of long-lived plasma cells, and HIV vaccines with TLR3- or TLR4-based adjuvants in germline BCR knock-in mice have been associated with increased somatic hypermutation (SHM) and the induction of VRC01-like bNAb precursors. However, the mechanisms by which 3M-052 or other TLR-based adjuvants support germinal center activity, SHM, the induction of HIV bNAb precursors, and/or the development of long-lived plasma cells remain unknown. The objective of Project 1 is to identify the determinants of the initial steps in the induction of bNAb precursors. We hypothesize that adjuvants given in combination with BG505 GT1.1 SOSIP trimers alter the developmental pathway of Env-specific B cells through the induction of specific innate responses that will impact affinity maturation of bNAb precursors. Understanding the complex process of B cell development towards bNAb- producing B cells in response to HIV vaccination and its regulation will require broad omics-based approaches that this Program will apply. Project 1 will thoroughly assess innate immune responses via transcriptomics, single cell (sc) RNA-sequencing, immunome analysis by CyTOF, and proteome analysis of soluble immune mediators in plasma. To determine whether bNAb development can be impacted by the choice of adjuvant, we will ask whether the yield of bNAb precursors after BG505 GT1.1 SOSIP immunization differs dependent on innate responses induced by 3M-052-SE versus a saponin-based adjuvant (Aim 1), and dependent on adjuvant- specific immune cell activation and function in lymph nodes (Aim 2). In collaboration with Project 2, we will define how these innate responses are modulated by commensal bacteria (Aim 3). The results are expected to inform the development of new targeted immunomodulatory approaches of the vaccine prime to optimize the induction of bNAb precursors and provide a framework for bNAb-targeting HIV vaccine evaluation and down-selection.