DNA & SOLUBLE MULTIMERIC IMMUNOGEN FOR HUMMORAL IMMUNITY

脱氧核糖核酸

• 批准号: 2751241
• 负责人: JON OSCHERWITZ
• 金额: $ 18.9万
• 依托单位: UNIVERSITY OF MICHIGAN AT ANN ARBOR
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-09-29 至 2000-09-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2751241
• 关键词: AIDS vaccines; G protein; chimeric proteins; gene targeting; human immunodeficiency virus; humoral immunity; laboratory mouse; neutralizing antibody; nucleic acid repetitive sequence; protein sequence; receptor coupling; vaccine development; vector vaccine; virus antigen; AIDS vaccines; G protein; chimeric proteins; gene targeting; human immunodeficiency virus; humoral immunity; laboratory mouse; neutralizing antibody; nucleic acid repetitive sequence; protein sequence; receptor coupling; vaccine development; vector vaccine; virus antigen

DESCRIPTION: (Adapted from applicant's abstract): Earlier work in a number of disease models has shown that immunogens containing tandemly repeated epitopes can be more immunogenic than a respective immunogen containing only a single copy of the sequence. The applicant's studies have revealed that antisera raised in mice to a recombinant immunogen in alum containing eight copies of an MN isolate V3PND sequence is able to neutralize ADA, a primary M-tropic isolate of HIV-I. However, despite hopes of generating immunogens able to induce antibodies capable of primary isolate neutralization, the hypervariable nature of the V3 loop of HIV would apparently necessitate incorporation of multiple strain specific V3 loop sequences in any prototype AIDS vaccine. Coincident with the increased understanding of the role of the V3 loop in determining cellular tropism, however, has come new insights into how the V3loop sequences interact with cellular coreceptors in the facilitation of HIV infection. CCR5 is a G-protein coupled receptor identified as integral for entry by M-tropic HIV isolates into macrophages. Since M-tropic viral isolates are especially implicated early in HIV infection and individuals who are homozygous for the 32 bp deletion which results in a non-functional-CCR5 receptor are protected from infection from HIV, yet appear to have a normal phenotype, the CCR5 extracellular receptor sequences represent attractive targets for humoral immunity to HIV-I. We have analyzed the CCR5 extracellular receptor sequence domains and have identified putative extracellular sequences which represent potential antibody targets. The major objectives of the current study are to: a) Develop tandem repeat soluble and DNA constructs representing sequences from the V3 loop of primary NSI M-tropic isolates; b) Analyze the ability of antisera to these constructs raised in BALB/c mice to neutralize homologous and heterologous primary NSI M-tropic HIV isolates; c) Develop tandem repeat soluble and DNA constructs representing sequences from the putative extracellular domains of CCR5; d) Analyze the ability of antisera to these constructs raised in BALB/c mice to neutralize homologous and heterologous primary NSI M-tropic HIV isolates and; e)Utilizing the results from targeting the V3 and CCR5 sequences, construct hybrid immunogens which contain sequences from both the V3 loop of primary NSI M-tropic viruses, and immunogenic sequences from the CCR5 coreceptor. The construction of these immunogens is enabled by our development of BMX7, a novel vector for cloning multi-determinant tandem repeat immunogens, and BMX7EX, a DNA expression vector which retains the essential cloning features of the BMX7 vector. These studies will ideally culminate in the development of immunogens capable of inducing antibodies able to neutralize M-tropic primary isolate infectivity and contribute significantly to our understanding of the immunology of peptide component DNA and soluble immunogens.

描述：（改编自申请人的摘要）： 疾病模型表明，含有串联的免疫原子重复 表位比仅包含的免疫原的表位更具免疫原性 序列的单个副本。 申请人的研究表明 抗血清在小鼠中升高到铝中的重组免疫原，其中八个 Mn分离株V3PND序列的副本能够中和ADA，主要 HIV-I的M-热带分离物。 但是，尽管希望产生免疫原 能够诱导能够进行原代分离株中和的抗体， HIV的V3环的高变量显然是必需的 在任何原型中掺入多种应变特异性V3环序列 艾滋病疫苗。 与对角色的理解不断增加 然而，V3循环确定细胞向潮流已经有了新的见解 进入V3loop序列如何与细胞共感受器相互作用 促进HIV感染。 CCR5是一种G蛋白偶联受体，被识别为以by M-热带HIV分离为巨噬细胞。 由于M-热带病毒分离株是 特别是在艾滋病毒感染的早期和 32 bp缺失的纯合子，导致非功能CCR5 受体免受HIV感染的保护，但似乎有正常 表型，CCR5细胞外受体序列代表吸引人 对HIV-I的体液免疫的靶标。 我们已经分析了CCR5 细胞外受体序列结构域，并已确定了推定 细胞外序列，代表潜在的抗体靶标。 这 当前研究的主要目标是：a）发展串联重复 可溶性和DNA构建体，代表来自V3环的序列 原发性NSI M-热分离株； b）分析抗血清对这些的能力 在BALB/C小鼠中提出的结构，以中和同源和异源 原发性NSI M-热带HIV分离株； c）发展串联重复可溶性和DNA 代表来自推定的细胞外域的序列的构造 CCR5; d）分析抗血清对这些构造的能力 BALB/C小鼠中和同源和异源原发性NSI M-热带 艾滋病毒分离株； e）利用靶向V3和CCR5的结果 序列，构造杂交免疫原，其中包含来自两个的序列 初级NSI M-冰球病毒的V3环，以及来自的免疫原性序列 CCR5共肽。 这些免疫原的结构由我们的 BMX7的开发，BMX7是一种克隆多确定串联的新型矢量 重复免疫原子和BMX7EX，它是保留的DNA表达载体 BMX7矢量的基本克隆特征。 这些研究理想情况下 最终导致能够诱导抗体的免疫原子的开发 能够中和M-热带原发性分离株感染力并有助于 我们对肽成分的免疫学的理解很大 DNA和可溶性免疫原。