Genetic Scaffolds Targeting the HIV Glycan Shield for an HIV Vaccine.

HIV-1/艾滋病仍然是全球严重的公共卫生问题，估计每天有7，000例新感染。有效的HIV-1疫苗对于长期控制HIV-1流行病是必要的。本研究的目的是开发一种安全有效的疫苗来预防HIV-1的获得和传播。来自HIV-1感染患者的几种有效和广泛中和的单克隆抗体（bNAb）结合HIV-1包膜（Env）蛋白的gp 120亚基上的高甘露糖碳水化合物结构，例如2G 12和一系列新鉴定的抗体PGT 125 -131和PGT 135，它们是迄今为止鉴定的最有效和广泛交叉反应性的bNAb。这些bNAb的鉴定表明gp 120的聚糖屏蔽是可行的疫苗靶标。我们已经通过基因修饰酵母菌株来靶向HIV-1 Env聚糖屏蔽，并且已经表明消除S.酿酒酵母（TM为三重突变体）导致Man 8 GlcNAc 2碳水化合物结构的均匀产生，Man 8 GlcNAc 2碳水化合物结构是PGT bNAb和2G 12识别的表位中聚糖的主要形式。我们已经证明，用完整的TM酵母细胞以及纯化的、高度糖基化的、2G 12/PGT 128反应性TM酵母蛋白免疫兔子，而不使用任何gp 120或HIV-1相关蛋白、识别合成的高甘露糖聚糖以及来自广泛的HIV-1分离物和所有测试的病毒体的单体gp 120蛋白的抗体以碳水化合物依赖性方式。遗传支架引发的抗体也有效地中和来自携带高甘露糖N-聚糖的不同亚型的HIV-1，尽管迄今为止尚未观察到对主要病毒株的有效中和活性。我们假设，在异源蛋白上适当呈现高甘露糖簇与优化的免疫策略相结合，可以引发特异性靶向HIV-1 Env聚糖盾上关键中和表位的抗体，并有效中和原代分离株，因此提出以下特定目的： 1.使用生物信息学方法结合遗传修饰的TM酵母从高度糖基化蛋白的全基因组筛选中选择可有效支持PGT bNAb结合的先导免疫原用于免疫试验。 2.优化免疫策略，并使用逐步支架初免-Env加强策略与领先的免疫原和可溶性HIV-1 gp 140三聚体引发中和抗体。

DESCRIPTION: HIV-1/AIDS remains a serious public health problem globally with an estimated 7,000 new infections each day. An effective HIV-1 vaccine will be necessary for the long-term control of the HIV-1 pandemic. The goal of this study is to develop a safe and effective vaccine to prevent HIV-1 acquisition and transmission. Several potent and broadly neutralizing monoclonal antibodies (bNAbs) derived from HIV-1 infected patients bind to high mannose carbohydrate structures on the gp120 subunit of the HIV-1 envelope (Env) protein, such as 2G12 and a series of newly identified antibodies PGT125-131 and PGT135 that are among the most potent and broadly cross-reactive bNAbs identified to date. The identification of these bNAbs suggests that the glycan shield of gp120 is a viable vaccine target. We have targeted the HIV-1 Env glycan shield by genetically modifying a yeast strain and have shown that the elimination of three carbohydrate processing enzymes in S. cerevisiae (TM for triple mutant) results in the uniform production of the Man8GlcNAc2 carbohydrate structures that are the major form of glycans in the epitopes recognized by the PGT bNAbs and 2G12. We have shown that immunization of rabbits with whole TM yeast cells as well as purified, highly glycosylated, 2G12/PGT128 reactive TM yeast proteins, without the use of any gp120 or HIV-1 related proteins, elicits antibodies that recognize synthetic high-mannose glycans as well as monomeric gp120 proteins from a wide array of HIV-1 isolates and all tested virions in a carbohydrate dependent manner. The genetic scaffold elicited antibodies also potently neutralize HIV-1 from different subtypes that carry high-mannose N-glycans although potent neutralizing activity to primary virus strains has not been observed so far. We hypothesize that appropriate presentation of high mannose clusters on heterologous proteins in a combination with optimized immunization strategies can elicit antibodies that specifically target the key neutralizing epitope on the glycan shield of HIV-1 Env and potently neutralize primary isolates, and so propose the following Specific Aims: 1. Select lead immunogens that can efficiently support PGT bNAb binding from genome-wide screens of highly glycosylated proteins using a bioinformatics approach in combination with the genetically modified TM yeast for immunization trials. 2. Optimize immunization strategies and elicit neutralizing antibodies using a stepwise scaffold prime-Env boost strategy with the lead immunogens and a soluble HIV-1 gp140 trimer.