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

描述：艾滋病毒-1/艾滋病仍然是一个严重的全球公共卫生问题，估计每天有7000名新感染者。一种有效的艾滋病毒-1疫苗对于长期控制艾滋病毒-1大流行是必要的。这项研究的目标是开发一种安全有效的疫苗，以防止艾滋病毒-1的感染和传播。来自HIV-1感染者的几种有效的广中和单抗(BNAbs)与HIV-1包膜蛋白gp120亚基上的高甘露糖碳水化合物结构结合，例如2G12和一系列新发现的抗体PGT125-131和PGT135，它们是迄今发现的最有效和最具交叉反应的bNAbs之一。对这些bNAbs的鉴定表明，gp120的糖链保护膜是一个可行的疫苗靶点。我们通过对一株酵母菌株进行遗传修饰来靶向HIV-1Env多糖屏蔽，并证明了在酿酒酵母(TM代表三重突变体)中消除三种碳水化合物处理酶会导致Man8GlcNAc2碳水化合物结构的统一产生，这是PGT bNAbs和2G12识别的表位中糖链的主要形式。我们已经证明，用整个TM酵母细胞以及纯化的、高度糖基化的2G12/PGT128反应性TM酵母蛋白免疫兔子，而不使用任何gp120或HIV-1相关蛋白，可以诱导出识别合成的高甘露糖多糖以及来自广泛的HIV-1分离株和所有测试病毒粒子的单体gp120蛋白的抗体。基因支架诱导的抗体也能有效地中和携带高甘露糖N-糖链的不同亚型的HIV-1，尽管到目前为止还没有观察到对主要病毒株的有效中和活性。我们假设，在异源蛋白上适当地呈现高甘露糖簇，结合优化的免疫策略，可以诱导出针对HIV-1环境蛋白糖壳上的关键中和表位的抗体，并有效地中和初级分离株，因此提出了以下具体目标： 1.利用生物信息学方法，结合转基因TM酵母，从全基因组筛选的高度糖化蛋白中选择能够有效支持PGT bNAb结合的先导免疫原进行免疫试验。 2.优化免疫策略，利用先导免疫原和可溶性HIV-1 gp140三聚体的分步支架Prime-Env Boost策略诱导中和抗体。

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.