Eliciting neutralizing antibodies and B cell responses using novel HIV Env immunogens in non-human primates

有效的HIV-1疫苗可能需要中和抗体。然而，几乎没有候选疫苗 疫苗接种后有效地诱导广谱中和抗体(BNAbs)。最近，VRC工作组 已经在豚鼠和非人灵长类动物(NHP)中诱导了融合肽导向的bNAbs。同样，我们最近 异种NFL(未切割)三聚体脂质体PRIME诱导的兔bNAbs：在Scripps增强 在启动免疫中，我们用靶向N-糖链缺失来激活B细胞反应 (Dubrovskaya等人，豁免权2019年)。我们分离了两个概括了血清活性的兔bNAb。这些 BNAbs针对两个不同的环境漏洞位置。中和反应的激发是 通过靶向N-糖链缺失、高密度脂质体阵列和异源三聚体修复增强 助推环境。在对豚鼠的独立实验中，我们已经在多种动物中诱导出bNAb，也使用 N-糖链缺失，异源环境NFL PRIME：Boost方法。最近的这些结果令人鼓舞。 朝着一个长达30年的问题的成功解决迈进。近本土三聚体尖峰的新时代 模拟物，再加上颗粒阵列，结构信息设计和环境特异性B的高分辨率分析 细胞和淋巴结反应，为更有效地诱导bNAbs提供了新的机会。我们提出了一个 综合、多方面的方法融合了艾滋病毒环境三聚体设计、分析方面的世界领先者的专业知识 疫苗接种后的B细胞反应和抗体，NHP免疫组织分析和基于EM的正在进行的分析 免疫反应和高分辨率抗体：三聚体相互作用。我们将使用有序的三色素数：Booking 这在兔和豚鼠中引发了bNAbs，在NHP中引发了这样的反应，将Small的成功转化为 项目1(WYATT)中使用新型免疫原设计和演示的动物对NHP的免疫和NFL免疫 三聚体通过核心B(Silvestri)进入NHP。我们将使用EM评估的“实时”血清Fab-to-Trimer结合 补体核心C区多克隆免疫球蛋白表位定位(EMPEM) 项目2(卡尔松·海德斯坦)的克隆、测序和功能表达。与VRC合作 (Mascola)我们将定义非中和单抗来掩盖不需要的非中和表位，或者更好地 展示交叉中和单抗的表位。我们将比较NFL三聚体脂质体和细胞表面NFL三聚体 阵列表达来自于令人兴奋的mRNA脂质包埋技术。我们将评估免疫接种是否在 与成年猕猴相比，幼年NHP B细胞能更好地产生bNAbs 人类感染。继我们在三聚脂质体疫苗接种后发现第二层CD4bs导向的bNAb之后， 命名为E70，我们将通过在NHP中定向NFL三聚体去糖基化来靶向CD4bs。根据我们最近的调查 发现了非常广泛的中和疫苗诱导的单抗1C2，我们还将重点介绍gp41：120三聚体 界面。利用这些最初的线索，我们将采取多方面、跨组件的集成方法 用近乎天然的、未切割的NFL Env三聚体免疫后，指导NHP中bNAbs的诱导。

Neutralizing antibodies are likely to be required for an effective HIV-1 vaccine. However, few candidate vaccines efficiently elicit broadly neutralizing antibodies (bNAbs) following vaccination. Recently, the VRC working group has elicited fusion peptide-directed bNAbs in guinea pigs and non-human primates (NHPs). Similarly, we recently elicited bNAbs in rabbits following heterologous NFL (uncleaved) trimer-liposome prime:boosting at Scripps where we activated B cell responses with targeted N-glycan deletions in the priming immunizations (Dubrovskaya et al, Immunity 2019). We isolated two rabbit bNAbs that recapitulate the serum activity. These bNAbs are directed against two distinct sites of Env vulnerability. The elicitation of neutralizing responses was enhanced by targeted N-glycan deletion, high-density liposomal array and heterologous trimer restorative boosting Env. In independent experiments in guinea pigs, we have elicited bNAbs in multiple animals also using a N-glycan deletion, heterologous Env NFL prime:boost approach. These recent outcomes are encouraging inroads toward the successful solution of a 3-decade-long problem. The new era of near-native trimeric spike mimics, coupled with particulate array, structure-informed design and high-resolution analysis of Env-specific B cell and lymph node responses, afford new opportunities to more efficiently elicit bNAbs. We propose an integrated, multi-faceted approach blending the expertise of world leaders in HIV Env trimer design, analysis of B cell responses and Abs following vaccination, NHP immune tissue analysis and EM-based analysis of ongoing immune responses and high-resolution Ab:trimer interactions. We will use well-ordered trimer prime:boosting that elicited bNAbs in rabbits and guinea pigs to elicit such responses in NHPs, translating success in small animals to NHPs using novel immunogen design and presentation in Project 1 (Wyatt) and immunization of NFL trimers into NHPs via Core B (Silvestri). We will use “real-time” serum Fab-to-trimer binding evaluated by EM polyclonal IgG epitope mapping (EMPEM) in Core C (Ward) in complement with rapid mAb NGS-based mAb cloning, sequencing and functional expression in Project 2 (Karlsson Hedestam). In collaboration with the VRC (Mascola) we will define either non-neutralizing mAbs to mask unwanted non-neutralizing epitopes or to better display the epitopes of cross-neutralizing mAbs. We will compare NFL trimer-liposomes to cell surface NFL trimer array expressed from the exciting mRNA lipid encapsulation technology. We will assess if immunization in the juvenile NHP B cell repertoire compared to adult macaques will better generate bNAbs as is observed during human infection. To follow our discovery of a tier 2 CD4bs-directed bNAb following trimer-liposome vaccination, termed E70, we will target the CD4bs by directed NFL trimer deglycosylation in the NHPs. Based on our recent discovery of the very broadly neutralizing vaccine-induced mAb, 1C2, we will also focus on the gp41:120 trimer interface. Leveraging these initial leads, we will undertake a multifaceted, cross-component integrated approach to guide the elicitation of bNAbs in NHPs following vaccination with near-native, uncleaved NFL Env trimers.