Characterization and Generalization of a Novel Immunogenic Target on HIV-1 gp120

HIV-1 gp120 新型免疫原性靶点的表征和推广

• 批准号: 8812773
• 负责人: Xueling Wu
• 金额: $ 23.7万
• 依托单位: AARON DIAMOND AIDS RESEARCH CENTER
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-03-01 至 2016-02-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8812773
• 关键词: Accounting; Amino Acids; Antibodies; Antibody Repertoire; Antibody Specificity; Antigens; B cell repertoire; Binding Sites; Clinical; Clinical Trials; Code; Complex; Core Protein; Data; Development; Enzyme-Linked Immunosorbent Assay; Epitopes; Excision; Exhibits; Face; Framework Regions; Genes; Genetic; Glycopeptides; Goals; HIV Envelope Protein gp120; HIV-1; Health; Human; Immune Targeting; Immune system; Immunoglobulin G; Immunoglobulin Genes; Immunoglobulin Somatic Hypermutation; Immunoglobulins; Individual; Infection; Length; Maps; Membrane; Memory B-Lymphocyte; Methods; Monoclonal Antibodies; Mutate; Neutralization Tests; Plasma; Point Mutation; Polysaccharides; Prevalence; Proteins; Recombinant Proteins; Recombinants; Sampling; Series; Serum; Site; Sorting - Cell Movement; Specificity; Staining method; Stains; Structure; Testing; Vaccination; Viral Physiology; Virus; antibody-dependent cell cytotoxicity; base; design; env Glycoproteins; glycosylation; immunogenic; immunogenicity; neutralizing antibody; neutralizing monoclonal antibodies; novel; success; vaccine development

DESCRIPTION: Successes in isolating human broadly neutralizing antibodies (bnAbs) and mapping the isolated monoclonal antibody (mAb) epitopes have helped to reveal several sites of vulnerability on the HIV-1 envelope glycoprotein (Env); however, the isolated bnAbs may only account for part of the donor sera neutralizing activity, indicating unknown targets on HIV-1 Env. Additionally, most of the isolated bnAbs have demonstrated unusual coding sequences implicating potential elicitation barriers. To progress on our quest to elicit protective antibodie through vaccination, we propose to identify and characterize new functional antibodies that may be more feasible for elicitation. Using a resurfaced gp120 stabilized core protein RSC3 as a probe, we previously identified the bnAb VRC01 and its class of mAbs that target the CD4-binding site (CD4bs) of gp120. By the same probing method, we recently identified a new mAb, VRC-PG05, from an infected individual who also developed the VRC01-class bnAb VRC-PG04. To date VRC-PG05 is the least somatically mutated neutralizing mAb against HIV-1 and its coding sequences do not seem uncommon in the human antibody repertoire. Atomic-level structure of VRC-PG05 in complex with gp120 revealed a novel glycopeptide epitope centering on two highly conserved glycans at N262 and N448 (based on HXB2 numbering) on gp120. Removal of either glycosylation site resulted in complete loss of VRC-PG05 activity, confirming the importance of the N262/N448 glycans for the mAb recognition. Based on the novelty of the glycopeptide epitope, the ability for co-elicitation with the CD4bs-directed functional mAbs, the relatively "normal" coding sequences, as well as the virus neutralization function, we consider VRC-PG05 and its epitope an attractive immune target and thus propose to generalize its findings to additional infected individuals. We hypothesize that 1) the newly discovered N262/N448 glycan site on gp120 is immunogenic, and antibodies targeting this site are frequently elicited in HIV-1-infected individuals; and 2) genetic sequences that encode functional antibodies targeting the N262/N448 glycan site do not require unusual features shared by most of the identified HIV-1 bnAbs. We have derived recombinant gp120 proteins with point mutations to detect the N262/N448 specificity by ELISA. We applied this monomeric gp120 ELISA to screen plasma samples from 85 HIV-1 seroconverters, and so far identified 17 (20%) who possessed antibodies with the N262/N448 specificity. These preliminary results support our first hypothesis regarding the immunogenicity of the newly discovered N262/N448 glycan site. From individuals with this antibody specificity, we propose to isolate the corresponding mAbs, determine the mAb anti-viral functions, and examine if the mAb coding sequences display any unusual features shared by most of the identified bnAbs. If successful, this project will yield new functional anti-HIV-1 mAbs that may be more feasible for elicitation through vaccination.

描述：成功分离人类广泛中和抗体 (bnAb) 并绘制分离的单克隆抗体 (mAb) 表位图谱有助于揭示 HIV-1 包膜糖蛋白 (Env) 上的几个脆弱位点；然而，分离的 bnAb 可能仅占供体血清中和活性的一部分，表明 HIV-1 Env 的目标未知。此外，大多数分离的 bnAb 已表现出不寻常的编码序列，暗示潜在的诱导障碍。为了在通过疫苗接种诱导保护性抗体的探索中取得进展，我们建议鉴定和表征可能更适合诱导的新功能性抗体。使用表面重构的 gp120 稳定核心蛋白 RSC3 作为探针，我们之前鉴定了 bnAb VRC01 及其靶向 gp120 的 CD4 结合位点 (CD4bs) 的 mAb 类。通过相同的探测方法，我们最近从一名感染者身上鉴定出了一种新的单克隆抗体 VRC-PG05，该感染者还开发了 VRC01 级 bnAb VRC-PG04。迄今为止，VRC-PG05 是体细胞突变最少的抗 HIV-1 中和单克隆抗体，其编码序列在人类抗体库中似乎并不罕见。 VRC-PG05 与 gp120 复合物的原子级结构揭示了一种新的糖肽表位，该表位以 gp120 上 N262 和 N448（基于 HXB2 编号）处的两个高度保守的聚糖为中心。任一糖基化位点的去除都会导致 VRC-PG05 活性完全丧失，证实了 N262/N448 聚糖对于 mAb 识别的重要性。基于糖肽表位的新颖性、与 CD4bs 导向的功能性单克隆抗体共诱导的能力、相对“正常”的编码序列以及病毒中和功能，我们认为 VRC-PG05 及其表位是一个有吸引力的免疫靶点，因此建议将其发现推广到其他感染个体。我们假设 1) gp120 上新发现的 N262/N448 聚糖位点具有免疫原性，并且在 HIV-1 感染者中经常引发针对该位点的抗体； 2) 编码靶向 N262/N448 聚糖位点的功能性抗体的基因序列不需要大多数已识别的 HIV-1 bnAb 所共有的不寻常特征。我们衍生了带有点突变的重组gp120蛋白，通过ELISA检测N262/N448特异性。我们应用这种单体 gp120 ELISA 筛选了 85 名 HIV-1 血清转换器的血浆样本，迄今为止已鉴定出 17 名 (20%) 人拥有具有 N262/N448 特异性的抗体。这些初步结果支持我们关于新发现的 N262/N448 聚糖位点的免疫原性的第一个假设。我们建议从具有这种抗体特异性的个体中分离出相应的 mAb，确定 mAb 的抗病毒功能，并检查 mAb 编码序列是否显示出大多数已识别的 bnAb 共有的任何异常特征。如果成功，该项目将产生新的 功能性抗 HIV-1 单克隆抗体通过疫苗接种可能更可行。