Determinants of Neutralization Breadth in Early HIV-1 Infection

描述(由申请人提供)：诱导抗体中和(NAB)循环中的HIV-1毒株的能力将是疫苗对HIV-1表现出最佳保护效果的关键。为了确定能够刺激这些反应的表位，已经从少数血浆NAB水平较高的慢性感染患者中回收了具有广泛和强大中和能力的单抗，但在这种广泛和强大的中和表型之前的病毒和宿主B细胞事件尚未被定义。从这些广泛中和的人那里还没有得到早期感染的样本，所以关于NAB广度是如何发展的问题还不能回答。在这里，我们确定了5个感染HIV-1的A和C亚型血清转换者，其中17人在感染后3年内形成了相对有效的交叉NAB广度。这项建议将调查这些人的NAB广度是如何以及为什么发展的，但不是在其他广度水平较低或无法检测到的人中。我们的初步数据表明，这两个患者群体之间存在根本的病毒学和免疫学差异，这可以解释为什么早期的NAB是菌株特异性的，但后来NAB在一部分受试者中发展成不同的广度。这种对早期感染的关注与观察到的NAB广度通常要么存在要么不存在是一致的 感染后约3年。此外，我们的初步数据显示，在我们的小组中，来自广泛中和剂的自体NAB靶向V1V2和CD4结合位点，这两个靶点都是具有‘精英’异源中和活性的单抗。我们将利用储存的血浆和活的PBMC样本来确定5名有NAB广度的受试者和5名没有NAB广度的受试者的初始NAB靶点和病毒逃逸途径。我们将使用包膜(Env)gp140 B细胞探针和从这10名受试者感染后约3年的两个时间点收集的PBMC样本，从单个B细胞类型中回收单抗。单抗将在基因和功能上进行表征，一个具有代表性的子集，包括那些具有广度的单抗，也将被结晶。 据我们所知，对早期病毒和免疫决定因素中和广度的这种类型的比较调查还没有进行。拟议的目标还提供了恢复和鉴定广泛中和单抗的很大可能性，这些单抗可能代表那些从慢性感染中分离出来的单抗的早期版本。我们的假设是，初始靶向某些NAB表位，如CD4结合位点或V1V2，再加上随后逃逸途径的影响，导致HIV-1感染者亚群中NAB广度的发展。具体地说，目的是(I)确定初始中和抗体靶标，并确定5名新近感染的具有异源中和广度的受试者和5名缺乏中和广度的受试者的病毒逃逸途径，以及(Ii)确定具有菌株特异性的早期单抗和获得自体或异源中和广度的单抗的基因、功能和结构特征。

DESCRIPTION (provided by applicant): The ability to induce antibodies that neutralize (nAb) circulating HIV-1 strains will be critical for a vaccine to exhibit optimal protective efficacy aganst HIV-1. To define the epitopes that can stimulate these responses, monoclonal antibodies (mAb) with broad and potent neutralization capacity have been recovered from a few chronically infected individuals that exhibited superior plasma nAb, but the viral and host B cell events that preceded this broad and potent neutralization phenotype have not been defined. Samples from early infection have not been available from these broadly neutralizing individuals, so questions about how nAb breadth developed cannot yet be answered. Here we identified 5 subtype A and C HIV-1 infected seroconvertors, out of 17, who developed relatively potent cross-clade nAb breadth at ~3 years post-infection. This proposal will investigate how and why nAb breadth developed in these individuals, but not in others who had low or undetectable levels of breadth. Our Preliminary Data suggest that there are fundamental virologic and immune differences between these two patient populations that could explain why early nAb are strain-specific, but later nAb go on to develop heterologous breadth in a subset of subjects. This focus on early infection is consistent with the observation that nAb breadth is generally either present or absent by ~3 years post-infection. Furthermore, our Preliminary Data show that autologous nAb from broad neutralizers in our panel targeted V1V2 and the CD4 binding site, both targets of mAbs with 'elite' heterologous neutralizing activity. We will utilize stored plasma and viable PBMC samples to define the initial nAb targets and viral escape pathways in 5 subjects with nAb breadth and 5 subjects without. We will recover mAbs from single B cell sorts using envelope (Env) gp140 B cell probes and PBMC samples collected at two early time points, and ~3 years post-infection, from these 10 subjects. The mAbs will be characterized genetically and functionally, and a representative subset, including those with breadth, will also be crystallized. To our knowledge, this type of comparative investigation into the early viral and immune determinants of neutralization breadth has not been performed. The proposed aims also provide a strong likelihood of recovering and characterizing broadly neutralizing mAbs that could represent early versions of those isolated from chronic infection. Our hypothesis is that the initil targeting of certain nAb epitopes, such as the CD4 binding site or V1V2, combined with the subsequent influence of escape pathways, leads to the development of nAb breadth in a subset of HIV-1 infected individuals. Specifically, the aims are to (i) Identify the initial neutralizing antibody target and define the viral escape pathways in 5 recently infected subjects who developed heterologous neutralization breadth and 5 subjects that lack breadth and (ii) Determine the genotypic, functional, and structural characteristics of early mAbs that are strain-specific and those that have acquired autologous or heterologous neutralization breadth.