Protective immunity against HIV: the role of IgG subclass and glycosylation

描述（由申请方提供）：IgG Fc受体（Fc？Rs）在自然杀伤细胞、单核细胞、巨噬细胞或树突状细胞上的表达可能在预防慢病毒感染中起关键作用。例如，广泛中和的单克隆抗体（IgG 1 b12）提供了更完整的保护猕猴对阴道的挑战与SHIV时，Fc？允许发生R接合和抗体依赖性细胞介导的病毒抑制（ADCVI）。此外，人的重组gp 120（rgp 120）免疫激发ADCVI抗体应答，其幅度与性传播HIV感染率的降低相关。因此，有效的HIV疫苗很可能不仅取决于抗体和抗原之间的相互作用，而且还取决于Fc？Rs及其Fc配体。我们将研究抗体的两个生物学特性，IgG亚类和Fc糖基化，影响其与Fc结合的能力？Rs，从而抑制HIV-1。由于IgG 2与大多数Fc结合较差？RS和唾液酸或岩藻糖含量减少增加Fc-Fc？R亲和力，我们将检验以下假设：1）IgG 2对rgp 120疫苗的应答在个体之间不同，并且由rgp 120疫苗引起的ADCVI活性与gp 120特异性IgG 2水平成反比; 2）由rgp 120免疫引起的抗gp 120抗体在Fc糖基化模式方面不同，并且Fc糖基化是ADCVI活性的决定因素;和3）IgG 1b 12的唾液酸和岩藻糖含量的改变将增加中和和ADCVI活性的效力和广度。我们的长期目标是构建疫苗引发抗体，最佳地从事Fc？卢比我们将实现以下具体目标：1）测量由用rgp 120疫苗接种引起的gp 120特异性IgG亚类，并确定亚类与ADCVI活性之间的关联。使用来自接种受试者的血清，通过ELISA定量gp 120特异性IgG亚类。亚类分布将与先前测量的ADCVI应答相关; 2）定量由rgp 120引发的IgG Fc中的唾液酸和岩藻糖含量，并确定Fc糖基化与抗HIV-1活性之间的关联。将应用凝集素ELISA和其他方法来确定gp 120亲和纯化的IgG的Fc糖基化模式-特别是岩藻糖基化和唾液酸化聚糖的频率;和3）分离广泛反应性mAb IgG 1 b12的低唾液酸和低岩藻糖变体，并确定Fc糖基化变化对抗HIV-1活性的影响。岩藻糖和唾液酸含量将被改变，中和和ADCVI功能将被测量。疫苗应答可能有目的地偏向亚类或Fc糖型的特定分布。鉴于开发广泛反应性疫苗诱导抗体的障碍，通过改变Fc来改善抗体功能的努力代表了HIV疫苗开发的一种有前途的，也许是必不可少的补充方法。我们建议研究抗体的某些特性，这些特性可能对预防HIV感染至关重要。这些研究可能导致提高抗体抑制HIV的能力的方法，并最终导致有效的HIV/AIDS疫苗的开发。

DESCRIPTION (provided by applicant): Engagement of IgG Fc receptors (Fc?Rs) on natural killer cells, monocytes, macrophages, or dendritic cells may play a critical role in preventing lentivirus infection. For example, a broadly neutralizing monoclonal antibody (IgG1 b12) provides far more complete protection of macaques against vaginal challenge with SHIV when Fc?R engagement and antibody-dependent cell-mediated virus inhibition (ADCVI) are allowed to occur. Moreover, recombinant gp120 (rgp120) immunization of humans elicits an ADCVI antibody response whose magnitude correlates with reduced rates of sexually transmitted HIV infection. Thus, it is likely that an effective HIV vaccine will depend not only on interactions between antibody and antigen but also on interactions between Fc?Rs and their Fc ligands. We will investigate two biological properties of antibody, IgG subclass and Fc glycosylation, that impact its ability to bind to Fc?Rs and consequently to inhibit HIV-1. Since IgG2 binds poorly to most Fc?Rs and decreased sialic acid or fucose content increases Fc-Fc?R affinity, we will test the following hypotheses: 1) the IgG2 response to rgp120 vaccine varies between individuals, and the ADCVI activity elicited by rgp120 vaccine is inversely proportional to gp120-specific IgG2 levels; 2) anti-gp120 antibodies elicited by rgp120 immunization vary with respect to Fc glycosylation pattern, and Fc glycosylation is a determinant of ADCVI activity; and 3) modifications in the sialic acid and fucose content of IgG1 b12 will increase the potency and breadth of neutralizing and ADCVI activity. Our long-term goal is to construct vaccines eliciting antibodies that optimally engage Fc?Rs. We will accomplish the following specific aims: 1) Measure gp120-specific IgG subclasses elicited by vaccination with rgp120, and determine the association between subclass and ADCVI activity. Using sera from vaccinated subjects, gp120- specific IgG subclasses will be quantified by ELISA. Subclass distribution will be correlated with ADCVI responses measured previously; 2) Quantify the sialic acid and fucose content in IgG Fc elicited by rgp120, and determine the association between Fc glycosylation and anti-HIV-1 activity. Lectin ELISA and other methods will be applied to determine Fc glycosylation patterns-particularly the frequency of fucosylated and sialylated glycans - of gp120 affinity-purified IgG; and 3) Isolate low sialic acid and low fucose variants of the broadly reactive mAb IgG1 b12, and determine the affect of Fc glycosylation changes on anti-HIV-1 activity. Fucose and sialic acid content will be modified and neutralizing and ADCVI functions measured. It is likely that vaccine responses could be purposefully biased toward a specific distribution of subclasses or Fc glycoforms. Given the obstacles to developing broadly reactive, vaccine-induced antibodies, efforts at improving antibody function by altering Fc represents a promising, and perhaps essential, complementary approach to HIV vaccine development. We propose studying certain properties of antibody that might be crucial for preventing HIV infection. Such studies may lead to methods of improving the ability of antibody to inhibit HIV and eventually to the development of effective HIV/AIDS vaccines.