Stabilized gp41 Structural Motif for HIV-1 Vaccine

HIV-1 疫苗的稳定 gp41 结构基序

• 批准号: 7284745
• 负责人: Min Lu
• 金额: $ 25.2万
• 依托单位: WEILL MEDICAL COLL OF CORNELL UNIV
• 依托单位国家: 美国
• 财政年份: 2007
• 资助国家: 美国
• 起止时间: 2007-05-01 至 2009-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7284745
• 关键词: Animals; Antibodies; Antigens; Biomedical Research; Cavia; Coiled-Coil Domain; Conserved Sequence; Cysteine; Development; Epitopes; Genetic; Goals; HIV-1; HIV-1 vaccine; Helix (Snails); Immunization; Individual; Knowledge; Membrane; Molecular Conformation; Numbers; Oryctolagus cuniculus; Particulate; Peptides; Property; Proteins; Research; Side; Structure; Surface; Thermodynamics; Today; Vaccination; Variant; design; dimer; disulfide bond; ear helix; human monoclonal antibodies; immunogenicity; nanoscale; neutralizing antibody; novel; response

DESCRIPTION (provided by applicant): The development of immunogens capable of eliciting broadly neutralizing antibodies to HIV-1 remains one of the most difficult challenges confronting biomedical research today. Nonetheless, a small number of human monoclonal antibodies (mAbs) have been isolated from HIV-1-infected individuals that can neutralize primary isolates of HIV-1 from different genetic subtypes. Of these rare mAbs, two (2F5 and 4E10) recognize epitopes on the membrane-proximal region of the gp41 ectodomain. However, all attempts to elicit comparable anti-gp41 antibodies by vaccination with peptides or simple linear epitopes have thus far failed. Our approach to immunogen design is guided by our recent identification and structural determination of a novel two-stranded coiled-coil domain (C55) that includes the membrane-proximal epitope region of gp41. However, we have determined that the C55 dimer structure is too unstable to be suitable for immunogenicity studies. The overall goal of this research plan is to gain a detailed understanding of the structural and thermodynamic properties of the C55 coiled-coil domain, and to use this knowledge to design and produce stable immunogens for the elicitation of broadly neutralizing HIV-1 antibodies. Our central hypothesis is that presentation of the highly conserved sequences recognized by 2F5 and 4E10 in this new structural state opens a promising avenue for the discovery and development of novel immunogens to induce broadly reactive neutralizing antibodies. Specific aims of this research are: (1) To identify and develop stabilized versions of the C55 coiled-coil domain encompassing the 2F5 and 4E10 epitopes. We will identify and incorporate apolar side chain substitutions into the buried hydrophobic core of the dimer to stabilize the coiled-coil structure. We will also screen cysteine substitutions to introduce a disulfide bond between the two interacting helices. Our emphasis is to generate stable molecules that preserve both the dimer conformation and the critical surface-exposed residues of the membrane-proximal region. (2) To evaluate the immunological responses elicited by stabilized C55 coiled-coil variants in small animals. We will conduct immunogenicity studies in rabbits and guinea-pigs to determine whether the stabilized dimer structure can elicit neutralizing antibodies. These studies will involve immunization of the animals using soluble proteins. We will also evaluate the immunogenicity of the stable coiled-coil molecules captured onto nanometer-sized beads as particulate immunogens.

描述（由申请人提供）：能够引发广泛中和 HIV-1 抗体的免疫原的开发仍然是当今生物医学研究面临的最困难的挑战之一。尽管如此，还是从 HIV-1 感染者中分离出了少量人单克隆抗体 (mAb)，这些抗体可以中和来自不同遗传亚型的 HIV-1 初级分离株。在这些罕见的 mAb 中，有两种（2F5 和 4E10）可识别 gp41 胞外域近膜区域上的表位。然而，迄今为止，所有通过肽或简单线性表位疫苗接种来引发类似抗 gp41 抗体的尝试都失败了。我们的免疫原设计方法以我们最近对一种新型双链卷曲螺旋结构域 (C55) 的鉴定和结构测定为指导，该结构域包括 gp41 的近膜表位区域。然而，我们确定C55二聚体结构太不稳定，不适合免疫原性研究。该研究计划的总体目标是详细了解 C55 卷曲螺旋结构域的结构和热力学特性，并利用这些知识设计和生产稳定的免疫原，以引发广泛中和 HIV-1 抗体。我们的中心假设是，2F5 和 4E10 在这种新结构状态下识别的高度保守序列的呈现，为发现和开发新型免疫原以诱导广泛反应性中和抗体开辟了一条有希望的途径。这项研究的具体目标是： (1) 识别和开发包含 2F5 和 4E10 表位的 C55 卷曲螺旋结构域的稳定版本。我们将识别非极性侧链取代并将其合并到二聚体的埋藏疏水核心中，以稳定卷曲螺旋结构。我们还将筛选半胱氨酸取代，以在两个相互作用的螺旋之间引入二硫键。我们的重点是产生稳定的分子，既保留二聚体构象，又保留膜近端区域的关键表面暴露残基。 (2) 评估稳定的 C55 卷曲螺旋变体在小动物中引起的免疫反应。我们将在兔子和豚鼠中进行免疫原性研究，以确定稳定的二聚体结构是否可以引发中和抗体。这些研究将涉及使用可溶性蛋白质对动物进行免疫。我们还将评估捕获到纳米大小的珠子上作为颗粒免疫原的稳定卷曲螺旋分子的免疫原性。