Glycan dependent epitopes of HIV broadly neutralizing antibodies

描述(由申请人提供)：尽管世界各地的许多团体共同努力生产艾滋病毒疫苗，但迄今仍没有有效的疫苗。疫苗的靶标是包膜尖峰蛋白(gp120/gp41)，它介导与CD4T细胞表面受体的结合。从疫苗的角度来看，问题是Spike蛋白高度糖基化，有多达25个或更多的N-连接的糖链，形成了一个防止对潜在蛋白质的免疫反应的“糖链屏障”。然而，许多艾滋病毒感染者会产生广泛的中和抗体(BNAbs)，以密集的糖链为靶标。分析这些依赖于糖的bNAb表明，它们与Spike蛋白上的不同区域结合，并识别不同类型的多糖。特别是，一些抗体对未加工的“高甘露糖”型多聚糖表现出高度的特异性，而另一些抗体对更多加工的“复合型”多聚糖表现出专一性。目前知识中的一个主要差距是在完整病毒的gp120/gp41上的每个糖基化位点上存在什么类型的糖链。这是了解依赖于糖的bNAbs的最佳表位的关键信息。在这个项目中，我们试图更好地定义依赖于糖的bNAbs的表位，并利用这些信息来开发免疫原，以诱导针对糖保护层的中和免疫反应。我们将使用一种新的糖蛋白组学方法来分析和确定实验室细胞系和外周血单核细胞中产生的HIV病毒gp120/gp41上每个糖基化位点的糖类型(高甘露糖或复合体)。我们将利用这些信息来生产携带定义的多糖的合成支架，用于表征bNAbs的最佳表位，并产生接近完整病毒上gp120糖基化的重组gp120免疫原。合成支架和糖基化优化的重组gp120三聚体都将被测试以诱导依赖于糖的gp120抗体介导的中和免疫反应。

DESCRIPTION (provided by applicant): There is as yet no effective vaccine to HIV despite concerted efforts by numerous groups worldwide to produce one. The target for vaccines is the envelope spike protein (gp120/gp41), which mediates binding to receptors on the surface of CD4 T cells. The problem from a vaccine perspective is that the spike protein is heavily glycosylated, with up to 25 or more N-linked glycans, creating a 'glycan shield' that prevent an immune response to the underlying protein. Yet, many HIV infected individuals develop broadly neutralizing antibodies (bnAbs) that target the dense array of glycans. Analysis these glycan- dependent bnAbs show that they bind to different regions on the spike protein, and that they recognize different types of glycans. In particular, some antibodies exhibit high specificity for unprocessed 'high mannose' type glycans, while others exhibit specificity for more highly processed 'complex type' glycans. A major gap in current knowledge is what types of glycans are present at each glycosylation site on gp120/gp41 on the intact virus. This is critical information for understanding the optimal epitopes of glycan-dependent bnAbs. In this project we seek to better define the epitopes of glycan-dependent bnAbs, and use the information to develop immunogens that elicit a neutralizing immune response that targets the glycan shield. We will use a novel glyco-proteomics approach to analyze and determine type of glycan (high mannose or complex) for each glycosylation site on gp120/gp41 from HIV viruses produced in laboratory cell lines and peripheral blood mononuclear cells. We will use this information to produce synthetic scaffolds that carry defined glycans for characterization of the optimal epitopes of bnAbs, and to generate recombinant gp120 immunogens that approximate the glycosylation of gp120 on intact virus. Both the synthetic scaffolds and glycosylation optimized recombinant gp120 trimers will be tested for induction of neutralizing immune responses mediated by glycan-dependent gp120 antibodies.