RETROVIRUS PARTICLE DISPLAY FOR IMMUNOGEN OPTIMIZATION

用于免疫原优化的逆转录病毒颗粒显示

• 批准号: 2760177
• 负责人: SAMUEL C KAYMAN
• 金额: $ 24.15万
• 依托单位: PUBLIC HEALTH RESEARCH INSTITUTE
• 依托单位国家: 美国
• 财政年份: 1998
• 资助国家: 美国
• 起止时间: 1998-09-30 至 2000-09-29
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2760177
• 关键词: AIDS vaccines; HIV envelope protein gp120; antigen antibody reaction; chimeric proteins; gene expression; genetic library; human immunodeficiency virus 1; humoral immunity; laboratory rat; murine leukemia virus; protein folding; site directed mutagenesis; vaccine development; virion; virus antigen

We propose to use a novel murine leukemia virus (MuLV)-based retroviral display system that we have recently developed (mss in prep) to improve our HIV-1 humoral vaccine candidate, which is based on the V1V2 domain of gpl20. We have recently obtained evidence indicating that the V1V2 domain of the HIV-1 envelope protein contains highly conserved epitopes that mediate potent neutralization of primary HIV-1. Our subunit vaccine candidate is a fusion protein that expresses the isolated V1V2 domain of gpl20 from a V2 consensus North American clade B isolate in the absence of other HIV sequences, using the N-terminal domain of the MuLV envelope protein to promote proper folding and secretion. Immunization of rats or macaques with this V1V2 fusion protein induces antibodies that have potent neutralizing activity for all primary virus isolates tested, including representatives of clades B, C, D and E. However, these antibodies are present in the vaccine sera at low concentration compared to antibodies against less conserved, non- neutralizing epitopes. Not only are these non-neutralizing epitopes immunodominant, but our recent evidence indicates that antibodies with enhancing activity are also induced. We will isolate variant V1V2 sequences that retain the neutralization epitopes but no longer present the non-neutralization epitopes by enrichment from random libraries using our novel MuLV retroviral particle display system. Since the desired epitopes are glycan-dependent conformational structures, standard phage display systems cannot be used for this purpose. Available antibodies against either conserved neutralization epitopes or non-neutralization epitopes will be used to separate MuLV virions bearing variants of V1V2 that express the desired epitopes from those that do not. This will involve a negative enrichment step to remove V1V2 sequence variants that express non-neutralizing epitopes followed by a positive enrichment step to require that the V1V2 variants isolated do express the potent neutralization targets in V1V2. Once sequences with improved immunoreactivity are identified, they will be expressed in our standard soluble V1V2 fusion protein system for further evaluation as immunogens. This novel methodology promises to advance our development of a V1V2 domain-targeted subunit vaccine that can induce a protective humoral response against HIV-1.

我们建议使用一种新型鼠白血病病毒（MULV）的逆转录病毒 我们最近开发的显示系统（准备中的MSS） 我们的HIV-1体液疫苗候选者，该疫苗基于V1V2结构域 GPL20。 我们最近获得了证据，表明V1V2 HIV-1包膜蛋白的结构域含有高度保守的表位 介导原代HIV-1的有效中和。 我们的亚基 疫苗候选者是一种表达孤立V1V2的融合蛋白 从V2共识北美进化枝B分离的GPL20的域 使用其他HIV序列，使用的N末端结构域 mulv包膜蛋白促进适当的折叠和分泌。 用这种V1V2融合蛋白诱导大鼠或猕猴的免疫接种 对所有原发病毒具有有效中和活性的抗体 测试的分离物，包括B，C，D和E的代表。 但是，这些抗体存在于低疫苗血清中 浓度与抗体抗体相比较少保守，非 - 中和表位。 这些非中和表位不仅是 免疫主导，但我们最近的证据表明 增强活动也被诱导。 我们将隔离变体V1V2 保留中和表位但不再存在的序列 从随机库中富集的非中和表位 使用我们新颖的MULV逆转录病毒颗粒显示系统。 自从 所需的表位是聚糖依赖性构象结构， 标准噬菌体显示系统不能用于此目的。 可用的抗体针对任何一种保守的中和表位 或非中和表位将用于分离Mulv病毒体 V1V2的轴承变体，表达来自这些的所需表位 那不是。 这将涉及一个负面的步骤以删除 表达非中和表位的V1V2序列变体跟随 通过一个积极的富集步骤，要求隔离V1V2变体 在V1V2中表达有效的中和目标。 一旦序列 通过确定了改善的免疫反应性，将表达它们 在我们的标准可溶性V1V2融合蛋白系统中 评估为免疫原。这种新颖的方法学有望促进我们的 可以诱导的V1V2域靶向亚基疫苗的开发 对HIV-1的保护性体液反应。