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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：
7615656
负责人：
ABRAHAM PINTER
金额：
$ 83.46万
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/7615656
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 Envelope Protein gp120 HIV-1 HIV-1 vaccine 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 Resistance Sampling Screening procedure Series Serum Silent Mutation Site Site-Directed Mutagenesis Sorting - Cell Movement Specificity Staging Structure Techniques Testing Treatment Protocols 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）：由于不能诱导保护性体液应答，HIV-1疫苗的进展受到阻碍。充分表征的中和表位或者免疫原性差（例如，b12，2F 5，2G 12）或对大多数原代分离株有效掩蔽（例如，V3、CD 4-bd和CD 4-i表位的抗体）。新的证据表明，即使是高度掩蔽的原代分离株也具有敏感的中和靶点，这些靶点可被自体患者血清识别，偶尔也可被异源血清识别。这表明，映射所涉及的表位可以鉴定能够诱导广泛中和活性的新靶标。我们已经确定了几个病人的血清，具有广泛的中和活性的主要分离株，我们已经开发了定位的目标表位的方法。我们现在提出鉴定介导自体和异源进化枝B和进化枝C Env中和的表位。具体目标是：1-筛选从北美和非洲队列获得的大量患者血清的交叉中和活性，并选择具有广泛中和特性的样品进行详细表征。2-通过检查中和敏感性和抗性Env之间形成的嵌合体的中和敏感性初步绘制靶表位，其中使用可用或工程化的限制性位点交换关键结构域。将通过交换较小区域和通过关键残基的诱变来进行所涉及的表位的更精细的作图。同时，在中和试验中，使用gp 120和gp 41抗原以及表达敏感Envs的各种结构域的融合蛋白作为阻断和免疫吸附试剂，对敏感表位进行免疫化学作图。3-从这些患者的EBV转化的B细胞培养物中以及用Env蛋白或表达潜在中和靶点的融合蛋白免疫的小鼠中分离具有广泛中和活性的单克隆抗体（mAb）。将采用结合和直接中和试验进行筛选。所得mAb将用于进一步确定所涉及表位的结构和分布。4-最后，用表达新靶点的融合蛋白免疫家兔。这些研究将评价表达序列的免疫原性，并检测针对这些免疫原诱导的抗体的中和特性。预计这些研究将确定暴露于典型的耐中和原代分离株并负责广泛中和的新靶标，并可能导致基于表达此类表位组合的免疫原的新疫苗方法。