Selecting HIV recombination libraries for stable Env trimer immunogens

选择稳定的 Env 三聚体免疫原的 HIV 重组文库

• 批准号: 9040773
• 负责人: MICHAEL B ZWICK
• 金额: $ 53.41万
• 依托单位: SCRIPPS RESEARCH INSTITUTE, THE
• 依托单位国家: 美国
• 财政年份: 2012
• 资助国家: 美国
• 起止时间: 2012-05-24 至 2018-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9040773
• 关键词: AIDS/HIV problem; Adjuvant; Animals; Antibodies; Antibody Response; Antigen Mimicry; Antigens; Binding; Biological Assay; CD4 Positive T Lymphocytes; Cavia; Cell Line; Cells; Chimera organism; Collaborations; Cytoplasmic Tail; Detergents; Development; Engineering; Enzyme-Linked Immunosorbent Assay; Epitopes; Flow Cytometry; Genetic; Genetic Recombination; Genome; Genotype; Goals; HIV; HIV Envelope Protein gp120; HIV-1; HIV-1 vaccine; Heating; Helper-Inducer T-Lymphocyte; Heterogeneity; Immune Sera; Immunization; In Vitro; Individual; Lead; Libraries; Ligands; Membrane; Methods; Molecular; Molecular Cloning; Molecular Conformation; Oryctolagus cuniculus; Peptide Hydrolases; Peripheral Blood Mononuclear Cell; Play; Property; RNA-Directed DNA Polymerase; Regimen; Resistance; Reverse Transcriptase Polymerase Chain Reaction; Scheme; Serum; Sorting - Cell Movement; Specificity; Surface; Time; Vaccination; Vaccines; Variant; Vertebral column; Virion; Virus; Virus-like particle; Work; base; cell growth; design; env Glycoproteins; genetic information; immunogenic; improved; mimetics; mutant; neutralizing antibody; non-Native; pandemic disease; particle; prospective; receptor; response; screening; sound; vaccine development; vaccine trial

DESCRIPTION (provided by applicant): A preferable solution to the HIV/AIDS global pandemic is an effective vaccine. A major priority for HIV-1 vaccine development is to identify an immunogen that can elicit high titers of neutralizing antibody (nAb) against the envelope glycoprotein (Env) spikes on circulating primary viruses. The functional (native) form of Env on the surface of virions and infected cells is a gp120-gp41 heterotrimer. A critical barrier in eliciing HIV-1 nAb is not one of eliciting anti-Env Ab in general, but rather that anti-Env responses are disproportionately elicited to irrelevant forms of Env and its subunits, rather than to the native Env trimer. Factors that likely contribute to the elicitation of weak and/or isolate specific nAb responses to HIV immunogens are sequence variability and heterogeneity in Env, as well as lability and low copy number of the trimeric, native form that is found on membrane surfaces. The goal of this project is to identify, characterize and develop virus-like particles (VPs or VLPs immunogens that display Env trimers that are more stable, more homogeneously native and in higher copy number than that of wild-type HIV-1. From diverse Env libraries created using DNA recombination methods Env VPs will be subjected to a variety of destabilizing conditions and then selected for binding to trimer-specific nAbs and nAb precursors in native and receptor-activated conformations while counter-screening against non-neutralizing Abs. To identify VPs with improved yield and native Env trimer content, we will engineer and select from diverse gp41 cytoplasmic tail and Gag libraries, again using nAbs. We will immunize rabbits using selected VLPs with stable, high copy number Env trimers and determine serum nAb titers against related strains and heterologous isolates in HIV-1 neutralization assays. We will probe lead Env immunogens using serum Ab and a panel of mAbs in a variety of binding and neutralization assays to ascertain the extent of antigenic mimicry of wildtype Env trimers. Detailed molecular characterization of the stable Env trimers and antisera should allow further optimization of lead immunogens and immunization regimens to elicit crossreactive nAb responses against different primary isolates of HIV-1. We expect the proposed studies to positively contribute to the development of an Env trimer-based vaccine.

描述（由申请人提供）：艾滋病毒/艾滋病全球流行的最佳解决方案是有效的疫苗。 HIV-1 疫苗开发的一个主要优先事项是确定一种免疫原，该免疫原可以引发针对循环原发病毒上的包膜糖蛋白 (Env) 尖峰的高滴度中和抗体 (nAb)。病毒颗粒和感染细胞表面的 Env 功能（天然）形式是 gp120-gp41 异源三聚体。引发 HIV-1 nAb 的关键障碍不是引发一般的抗 Env Ab，而是抗 Env 反应不成比例地引发不相关形式的 Env 及其亚基，而不是针对天然 Env 三聚体。可能有助于引发针对 HIV 免疫原的微弱和/或分离的特异性 nAb 反应的因素是 Env 中的序列变异性和异质性，以及膜表面上发现的三聚体天然形式的不稳定性和低拷贝数。 该项目的目标是鉴定、表征和开发病毒样颗粒（VP 或 VLP 免疫原，其显示的 Env 三聚体比野生型 HIV-1 更稳定、更均匀、拷贝数更高。从使用 DNA 重组方法创建的不同 Env 文库中，Env VP 将经历各种不稳定条件，然后被选择用于与三聚体特异性 nAb 和 nAb 结合 天然构象和受体激活构象的前体，同时针对非中和抗体进行反筛选。为了鉴定具有提高的产量和天然 Env 三聚体含量的 VP，我们将再次使用 nAb 从不同的 gp41 细胞质尾和 Gag 文库中进行设计和选择。我们将使用选定的具有稳定、高拷贝数 Env 三聚体的 VLP 对兔子进行免疫，并测定针对相关菌株的血清 nAb 滴度和 HIV-1 中和试验中的异源分离株。我们将在各种结合和中和测定中使用血清 Ab 和一组 mAb 来探测先导 Env 免疫原，以确定野生型 Env 三聚体的抗原模拟程度。稳定的 Env 三聚体和抗血清的详细分子表征应允许进一步优化先导免疫原和免疫方案，以引发 针对 HIV-1 不同主要分离株的交叉反应 nAb 反应。我们预计拟议的研究将对基于 Env 三聚体的疫苗的开发做出积极贡献。