Synthetic HIV Vaccine Targeting Glycopeptide Neutralizing Epitopes

靶向糖肽中和表位的合成 HIV 疫苗

• 批准号: 8777748
• 负责人: LAI-XI WANG
• 金额: $ 48.52万
• 依托单位: UNIVERSITY OF MARYLAND BALTIMORE
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-07-11 至 2015-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8777748
• 关键词: AIDS/HIV problem; Alkynes; Antibodies; Antibody Affinity; Antibody Formation; Antigens; Azides; Bacteriophages; Binding; Biochemical; Biological Assay; Complex; Copper; Epidemic; Epitopes; Germ Lines; Glycopeptides; Goals; HIV; HIV Envelope Protein gp120; HIV vaccine; HIV-1; Health; Heterogeneity; Human; Immune Sera; Immune response; Immunization; Immunology; Individual; Knowledge; Lead; Light; Maps; Methods; Oryctolagus cuniculus; Pattern; Peptides; Polysaccharides; Reaction; Receptors, Antigen, B-Cell; Recombinants; Research; Role; Series; Serum; Site; Specificity; Speed; Structure; Synthetic Vaccines; Testing; Vaccines; Viral; Virus-like particle; base; combat; cycloaddition; density; design; glycosylation; immunogenicity; improved; insight; mannosyl(5)-N-acetyl(2)-glucose; mannosyl(9)-N-acetylglucosamine2; neutralizing antibody; novel; public health relevance; reconstitution; research study; social; vaccine development

DESCRIPTION (provided by applicant): An effective HIV vaccine capable of inducing broadly neutralizing antibodies remains a significant goal in combating the HIV/AIDS epidemic. Recent discovery of a new class of glycan-reactive broadly neutralizing antibodies (bNAbs), represented by PG9, PG16, PGT121, and PGT128, has shed light on design of novel epitope-based vaccine. These bNAbs neutralize primary HIV-1 strains with remarkable breadth and potency and appear to target glycopeptide epitopes located in the variable domains (V1/V2 or V3) of HIV-1 gp120. Mutational, biochemical and structural studies suggest that the epitopes of PG9 and PG16 are glycopeptides consist of two N-glycans (at the N160 and N156/N173 glycosylation sites) and a patch of peptide in the V1/V2 domain, while the epitopes of PGT128 appear to involve the N-glycans at the N332 and N301 sites. However, further characterization of the glycan specificity was complicated by the heterogeneity in glycosylation of gp120. We used a synthetic approach to constructing homogeneous V1V2 and V3 glycopeptides carrying defined glycans for further defining the glycan specificity, which led to the identification of a cyclic glycopeptide epitope carrying a Man5GlcNAc2 glycan at N160 and a sialylated N-glycan at N156/N173 site as the minimal epitopes for PG9/PG16. We also designed and synthesized cyclic V3 glycopeptides that confirm the essence of a Man9/Man8GlcNAc2 glycan at the N332 site for PGT128 recognition. With the ability to reconstitute these glycopeptide epitopes via synthesis, we propose to design novel immunogens and hypothesize that displaying the glycopeptide epitopes on bacteriophage Qb (a well-characterized virus-like particle) in a highly ordered repetitive pattern will lead to immunogens capable of eliciting glycopeptide-specific neutralizing antibodies. We will test this hypothesis through pursuing three specific aims. Aim 1 is to design and synthesize Qb-based immunogens carrying polyvalent V1V2 and V3 glycopeptide epitopes. Construction of the immunogens includes chemoenzymatic synthesis of alkyne-glycopeptides and their subsequent conjugation with azide-tagged Qb via the copper-catalyzed azide-alkyne cycloaddition (CuAAC) click reaction; Aim 2 is to evaluate the antigenicity of the synthetic immunogens by antibody binding studies; and Aim 3 is to evaluate the immunogenicity of the synthetic vaccines in rabbits. The anti-sera from immunized rabbits are analyzed by ELISAs and viral neutralization assays. These experiments will assess the magnitude, breadth and duration of neutralizing antibody responses from immunizations with the glycopeptide epitope-based immunogens. This study is likely to yield important new insights in the design of a truly effective HIV-1 vaccine.

描述（由申请人提供）：能够诱导广泛中和抗体的有效 HIV 疫苗仍然是对抗 HIV/AIDS 流行的重要目标。最近发现的一类新的聚糖反应性广泛中和抗体（bNAb），以 PG9、PG16、PGT121 和 PGT128 为代表，为新型基于表位的疫苗的设计提供了线索。这些 bNAb 具有显着的广度和效力中和初级 HIV-1 毒株，并且似乎靶向位于 HIV-1 gp120 可变结构域（V1/V2 或 V3）的糖肽表位。突变、生化和结构研究表明，PG9和PG16的表位是由两个N-聚糖（位于N160和N156/N173糖基化位点）和V1/V2结构域中的一段肽组成的糖肽，而PGT128的表位似乎涉及N332和N332处的N-聚糖。 N301 站点。然而，gp120 糖基化的异质性使聚糖特异性的进一步表征变得复杂。我们使用合成方法构建携带特定聚糖的均质 V1V2 和 V3 糖肽，以进一步定义聚糖特异性，从而鉴定出在 N160 处携带 Man5GlcNAc2 聚糖和在 N156/N173 位点处携带唾液酸化 N-聚糖的环状糖肽表位作为最小表位 PG9/PG16。我们还设计并合成了环状 V3 糖肽，证实了 N332 位点上用于 PGT128 识别的 Man9/Man8GlcNAc2 聚糖的本质。凭借通过合成重建这些糖肽表位的能力，我们建议设计新型免疫原，并假设以高度有序的重复模式在噬菌体 Qb（一种充分表征的病毒样颗粒）上展示糖肽表位将产生能够引发糖肽特异性中和抗体的免疫原。我们将通过追求三个具体目标来检验这一假设。目标 1 是设计和合成携带多价 V1V2 和 V3 糖肽表位的基于 Qb 的免疫原。免疫原的构建包括炔烃-糖肽的化学酶合成以及随后通过铜催化叠氮-炔烃环加成（CuAAC）点击反应与叠氮化物标记的Qb缀合；目标 2 是通过抗体结合研究评估合成免疫原的抗原性；目的3是评价合成疫苗在兔体内的免疫原性。通过 ELISA 和病毒中和测定对免疫兔的抗血清进行分析。这些实验将评估基于糖肽表位的免疫原免疫产生的中和抗体反应的程度、广度和持续时间。这项研究可能会为设计真正有效的 HIV-1 疫苗带来重要的新见解。