Novel epitopes that mediate broad neutralization of clade B and C HIV-1 isolates

介导 B 型和 C 型 HIV-1 分离株广泛中和的新表位

基本信息

批准号：
8069367
负责人：
ABRAHAM PINTER
金额：
$ 58.78万
依托单位：
UNIV OF MED/DENT OF NJ-NJ MEDICAL SCHOOL
依托单位国家：
美国
项目类别：
财政年份：
2008
资助国家：
美国
起止时间：
2008-05-01 至 2013-04-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8069367
关键词：
Adsorption African African American Antibodies Antigens Area Autologous B-Lymphocytes Binding Biological Assay C-terminal Carrier Proteins Cell Culture Techniques Cells Chimera organism Chimeric Proteins Doctor of Philosophy Engineering Enzyme-Linked Immunosorbent Assay Epitope Mapping Epitopes Fc domain Goals HIV HIV Envelope Protein gp120 HIV-1 Human Hybridomas Immune Sera Immunoglobulin Variable Region Individual Lead Maps Masks Mediating Memory B-Lymphocyte Methods Monoclonal Antibodies Mus Mutagenesis Oryctolagus cuniculus Patients Property Protocols documentation Reagent Regimen Resistance Sampling Screening procedure Series Serum Silent Mutation Site Site-Directed Mutagenesis Sorting - Cell Movement Specificity Staging Structure Techniques Testing Vaccines Virion Virus base cohort env Gene Products env Genes follow-up immunogenic immunogenicity improved interest neutralizing antibody novel novel vaccines research study response vaccine development

项目摘要

DESCRIPTION (provided by applicant): Progress towards an HIV-1 vaccine has been stymied by the inability to induce a protective humoral response. Well-characterized neutralization epitopes are either poorly immunogenic (e.g., b12, 2F5, 2G12) or effectively masked on the majority of primary isolates (e.g., antibodies to V3, CD4-bd and CD4-i epitopes). New evidence suggests that even highly masked primary isolates possess sensitive neutralization targets that are recognized by autologous patient sera and occasionally by heterologous sera. This suggests that mapping the epitopes involved may identify novel targets that are capable of inducing broadly neutralizing activities. We have identified several patient sera that possess broadly neutralizing activities for primary isolates, and we have developed methods for localizing the target epitopes. We now propose to identify epitopes that mediate neutralization of both autologous and heterologous clade B and clade C Envs. The Specific Aims are: 1- To screen a large number of patient sera obtained from both North-American and African cohorts for cross-neutralizing activities, and select samples with broad neutralizing properties for detailed characterization. 2- To map target epitopes initially by examining the neutralization sensitivities of chimeras formed between neutralization-sensitive and -resistant Envs, in which key domains were exchanged using available or engineered restriction sites. Finer mapping of the epitopes involved will be performed by exchanging smaller regions and by mutagenesis of key residues. In parallel, sensitive epitopes will be mapped immunochemically using gp120 and gp41 antigens and fusion proteins expressing various domains of sensitive Envs as blocking and immunoadsorption reagents in neutralization assays. 3- To isolate monoclonal antibodies (mAbs) with broad neutralizing activities from EBV-transformed B cell cultures obtained from these patients, and from mice immunized with Env proteins or fusion proteins expressing potential neutralization targets. Screening will be performed both with binding and direct neutralization assays. The resulting mAbs will be used to further define the structure and distribution of the epitopes involved. 4- Finally, to immunize rabbits with fusion proteins expressing novel targets. These studies will evaluate the immunogenicity of the expressed sequences and to test the neutralizing properties of antibodies induced against these immunogens. It is anticipated that these studies will define novel targets that are exposed on typical neutralization-resistant primary isolates and responsible for broad neutralization, and could lead to new vaccine approaches based on immunogens expressing combinations of such epitopes.

描述（由申请人提供）：无法诱导保护性的体液反应而阻碍了HIV-1疫苗的进展。特征良好的中和表位的免疫原性（例如B12、2F5、2G12）或有效地掩盖了大多数主要分离株（例如，抗V3，CD4-BD和CD4-I表位的抗体）。新的证据表明，即使是高度掩盖的原发性分离株也具有敏感的中和靶标，这些靶标被自体患者血清识别，偶尔会通过异源血清识别。这表明映射所涉及的表位可以确定能够诱导广泛中和活动的新目标。我们已经确定了几种具有广泛中和主要分离株活性的患者血清，并且我们开发了定位目标表位的方法。现在，我们建议确定介导自体和异源进化枝B和进化枝C envs中和的表位。具体目的是：1-筛选从北美和非洲人群中获得的大量患者血清进行跨中和活动，并选择具有广泛中和特性的样品以详细表征。 2-最初通过检查中和敏感和耐药性ENV中形成的嵌合体的中和敏感性来绘制目标表位，其中使用可用或工程限制位点交换了关键域。通过交换较小的区域和关键残基的诱变来进行涉及的表位的映射。同时，使用GP120和GP41抗原和融合蛋白在中和测定中表达各种敏感的Envs的融合蛋白，将敏感的表位映射为免疫化学映射。 3-分离从这些患者获得的EBV转换的B细胞培养物中的单克隆抗体（mAb），以及从这些患者获得的EBV转换的B细胞培养物，以及用ENV蛋白或融合蛋白免疫的小鼠表达潜在中和靶标。筛选将通过结合和直接中和测定进行。所得的mAB将用于进一步定义所涉及的表位的结构和分布。 4-最后，用表达新靶标的融合蛋白对兔子进行免疫。这些研究将评估表达序列的免疫原性，并测试针对这些免疫原子诱导的抗体的中和特性。可以预料，这些研究将定义新的靶标的新靶标，这些靶标在典型的中和抗性原发性分离株上暴露并负责广泛中和，并可能导致基于表达此类表位组合的免疫原的免疫原分的新疫苗方法。