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

描述（由申请人提供）：IgG Fc受体(Fc？Rs)作用于自然杀伤细胞、单核细胞、巨噬细胞或树突状细胞可能在预防慢病毒感染中起关键作用。例如，一种广泛中和的单克隆抗体（IgG1 - b12）可以更全面地保护猕猴免受SHIV阴道攻击。R结合和抗体依赖细胞介导的病毒抑制（ADCVI）是允许发生的。此外，重组gp120 （rgp120）免疫人类引发ADCVI抗体反应，其强度与性传播HIV感染率的降低相关。因此，有效的HIV疫苗可能不仅取决于抗体和抗原之间的相互作用，还取决于Fc？r和它们的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，这两个特性会影响其与Fc？从而抑制HIV-1。由于IgG2与大多数Fc？Rs和唾液酸减少或焦点含量增加Fc-Fc？R亲和力，我们将检验以下假设：1)个体对rgp120疫苗的IgG2反应不同，rgp120疫苗引发的ADCVI活性与gp120特异性IgG2水平成反比；2) rgp120免疫诱导的抗gp120抗体在Fc糖基化模式方面存在差异，Fc糖基化是ADCVI活性的决定因素；3)唾液酸和IgG1 - b12的聚焦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，激发抗体，使其与Fc？我们将实现以下具体目标：1)测量接种rgp120诱导的gp120特异性IgG亚类，并确定亚类与ADCVI活性之间的关系。使用接种者的血清，用ELISA法定量gp120特异性IgG亚类。亚类分布将与先前测量的ADCVI反应相关；2)定量测定rgp120诱导的IgG Fc中唾液酸和病灶含量，确定Fc糖基化与抗hiv -1活性的关系。凝集素ELISA和其他方法将用于确定gp120亲和纯化的IgG的Fc糖基化模式-特别是集中和唾液化聚糖的频率；3)分离低唾液酸和低病灶的广泛反应性单抗IgG1 b12变体，并确定Fc糖基化变化对抗hiv -1活性的影响。将焦点液和唾液酸含量进行修饰，并测定中和作用和ADCVI功能。疫苗应答可能有意偏向于亚类或Fc糖型的特定分布。考虑到开发具有广泛反应性的疫苗诱导抗体的障碍，通过改变Fc来改善抗体功能的努力是开发艾滋病毒疫苗的一种有希望的、也许是必不可少的补充方法。我们建议研究可能对预防HIV感染至关重要的抗体的某些特性。这些研究可能导致提高抗体抑制艾滋病毒能力的方法，并最终开发出有效的艾滋病毒/艾滋病疫苗。

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.