Vaccines that Replicate the Neutralization-Competent Structure of the gp41 MPER

复制 gp41 MPER 中和能力结构的疫苗

基本信息

批准号：
8263672
负责人：
WILLIAM DEGRADO
金额：
$ 70.12万
依托单位：
SIMON FRASER UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2012
资助国家：
美国
起止时间：
2012-03-01 至 2013-02-28
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8263672
关键词：
AIDS Vaccines Affinity Amino Acid Sequence Antibodies Antibody Formation Antigens B-Lymphocytes Behavior Binding Biological Assay Blocking Antibodies Cell membrane Cells Cultured Cells DNA DNA Vaccines Data Environment Epitopes Extravasation Failure Funding Future Gagging Genes Goals HIV HIV Envelope Protein gp41 HIV Infections HIV immunization HIV-1 Human Cell Line Immunization Immunodominant Epitopes In Vitro Infection prevention Laboratories Lipid Bilayers Lipids Liposomes Mediating Membrane Membrane Fusion Membrane Proteins Modeling Molecular Models Monkeys Mutation Oryctolagus cuniculus Peptides Protein Binding Proteins Relative (related person)Role Scaffolding Protein Serum Solutions Structure Testing Transmembrane Domain United States National Institutes of Health Vaccine Design Vaccines Vesicle Viral Virus Virus-like particle Work base design env Gene Products experience immunogenic immunogenicity improved molecular modeling multidisciplinary mutant nanoparticle neutralizing antibody neutralizing monoclonal antibodies novel particle response synthetic peptide

项目摘要

DESCRIPTION (provided by applicant): The membrane proximal external region (MPER) of HIV gp41 comprises a highly conserved region involved in HIV viral fusion. It is an important target of antibody (Ab)-mediated neutralization as it contains epitopes for two broadly (b) neutralizing (Nt) monoclonal (M) Abs (2F5 and 4E10) and two MAbs that neutralize a significant, but not truly broad, range of HIV isolates (Z13 and m66.6), making the MPER an obvious target for an AIDS vaccine. However, all attempts to produce an MPER-targeting vaccine have failed. Most of these vaccines have been of three types: (i) synthetic peptides; (ii) MPER sequence grafted onto protein scaffolds; or (iii) onto proteins displayed on virus-like particles. In explaiing these failures, we hypothesize that previous vaccines have not faithfully mimicked the neutralization competent structure (NCS) of the MPER. Our preliminary work has shown that the gp41 transmembrane domain (TMD) is required for the full exposure of the MPER, since its replacement with the TMD from another membrane protein, decreases binding by MAb 4E10. Drs. Scott and Lu previously collaborated on producing a DNA vaccine that expresses a gp41 fragment comprising the MPER and TMD, so as to present the MPER in the context of the cell membrane. However, repeated immunization of rabbits with this DNA vaccine elicited low titer Abs that cross-reacted weakly with MPER peptides and did not neutralize virus; boosting immunizations with a virus-like particle vaccine did not improve anti-MPER titers, probably because of low MPER copy number. We propose to design more effective protein-boost immunogens that will mimic the NCS. In specific aim 1 we plan to design liposome and nanoparticle immunogens that present the gp41 MPER+TMD in high copy number in lipid bilayers, so as to better mimic the NCS of the MPER without added proteins that might "distract" the Ab response. We will produce DNA, liposome and nanoparticle candidates that have optimized the: (i) MPER, (ii) TMD, and (iii) composition of the lipid environment, based on relative binding affinity by bNt MAbs, the absence of binding by non-Nt mutants of the bNt MAbs, their behavior in a novel membrane leakage assay, and structural stabilization. In specific aim 2 optimized DNA, liposome and nanoparticle candidates will be produced and tested for their ability to elicit MPER-binding activities and Nt Abs. The optimized liposome and nanoparticle vaccines will then be tested as protein boosts (i.e., following DNA priming) with the goal of maximizing MPER-specific Ab titers and Nt potency and breadth. In addition, antigenicity, immunogenicity and structural data will be used to develop a molecular model of the MPER NCS, which should support future vaccine design. PUBLIC HEALTH RELEVANCE: The membrane proximal external region of the envelope protein gp41 (MPER) is a highly conserved region on the trimeric spike of infectious HIV-1, and a target of antibodies that neutralize a broad range of HIV-1 isolates. When used as a passive vaccine in monkeys, these antibodies protect against infection by HIV-like viruses; however so far there is no vaccine that can elicit such antibodies. Our goal is to produce a vaccine that will mimic the structure of the MPER on the infectious spike, and in so doing, will elicit antibodies that prevent infection by a broad spectrum of HIV-1 isolates; providing the important first steps in achieving a successful AIDS vaccine.

描述（由申请人提供）：HIV gp41 的近膜外部区域（MPER）包含参与 HIV 病毒融合的高度保守区域。它是抗体 (Ab) 介导的中和的重要靶标，因为它包含两种广泛 (b) 中和 (Nt) 单克隆 (M) Ab（2F5 和 4E10）的表位以及两种可中和显着但并非真正广泛的单克隆 (M) 单克隆抗体。一系列 HIV 分离株（Z13 和 m66.6），使 MPER 成为艾滋病疫苗的明显目标。然而，所有生产 MPER 靶向疫苗的尝试都失败了。大多数这些疫苗分为三种类型：(i) 合成肽； (ii) MPER序列移植到蛋白质支架上；或（iii）附着在病毒样颗粒上展示的蛋白质上。在解释这些失败时，我们假设以前的疫苗没有忠实地模仿 MPER 的中和能力结构 (NCS)。我们的初步工作表明，gp41 跨膜结构域 (TMD) 是 MPER 完全暴露所必需的，因为它被另一种膜蛋白的 TMD 取代，从而减少了 MAb 4E10 的结合。博士。 Scott和Lu之前合作生产了一种表达包含MPER和TMD的gp41片段的DNA疫苗，以便在细胞膜的背景下呈现MPER。然而，用这种 DNA 疫苗重复免疫兔子会产生低滴度抗体，与 MPER 肽交叉反应微弱，并且不能中和病毒；使用病毒样颗粒疫苗加强免疫并不能提高抗 MPER 滴度，可能是因为 MPER 拷贝数较低。我们建议设计更有效的蛋白质增强免疫原来模拟 NCS。在具体目标 1 中，我们计划设计脂质体和纳米颗粒免疫原，在脂质双层中以高拷贝数呈现 gp41 MPER+TMD，以便更好地模拟 MPER 的 NCS，而不添加可能“分散”Ab 反应的蛋白质。我们将生产 DNA、脂质体和纳米颗粒候选物，这些候选物已优化：(i) MPER、(ii) TMD 和 (iii) 脂质环境的组成，基于 bNt MAb 的相对结合亲和力，不存在非-MPER 结合。 bNt MAb 的 Nt 突变体、它们在新型膜渗漏测定中的行为以及结构稳定性。在具体目标 2 中，将生产优化的 DNA、脂质体和纳米颗粒候选物，并测试它们引发 MPER 结合活性和 Nt Ab 的能力。然后，优化的脂质体和纳米颗粒疫苗将作为蛋白质增强剂（即 DNA 引发后）进行测试，目标是最大化 MPER 特异性抗体滴度以及 Nt 效力和广度。此外，抗原性、免疫原性和结构数据将用于开发 MPER NCS 的分子模型，这应该支持未来的疫苗设计。公共卫生相关性：包膜蛋白 gp41 (MPER) 的膜近端外部区域是感染性 HIV-1 三聚体刺突上的高度保守区域，也是中和多种 HIV-1 分离株的抗体的靶标。当用作猴子的被动疫苗时，这些抗体可以防止艾滋病毒样病毒的感染；然而到目前为止，还没有疫苗可以引发这种抗体。我们的目标是生产一种疫苗模仿感染性刺突上 MPER 的结构，这样做将引发抗体，防止广谱 HIV-1 分离株感染；为成功研制艾滋病疫苗提供了重要的第一步。