FUSION COMPETENT HIV VACCINE IMMUNOGENS

描述(改编自申请人的摘要)最近发现的 HIV辅助受体为HIV包膜提供了一种新的、动态的视觉 蛋白质的功能。我们对这一概念进行了扩展，以建议 瞬时包膜蛋白相互作用和构象变化 在CD4和辅受体介导的结合和融合过程中出现可能暴露 中和抗体的关键靶点，特别是抗体 能够中和艾滋病毒的初级分离株(PI)。在这方面， 我们已经开发出新型的融合能力强的疫苗，这种疫苗可以诱导出 猪瘟病毒中和抗体免疫转基因小鼠的研究 它们对疫苗中的人类CD4和CCR5成分具有耐受性。 PI病毒中和的发现与我们的结果有显著差异 使用提供不起作用的包膜蛋白的可比疫苗。 在这份提案中，我们将首先定义以下要求和限制 用“融合能力”疫苗中和PI病毒。其次，我们 将扩展这些研究以确定分子结构和 介导HIV结合和融合的构象，并作为 PI病毒中和的目标。我们已经开发了一种特定的和 高亲和力的‘标签’，允许分离化学交联物 CD4相关的复合体沿着结合和融合的途径进行。这些 将对复合体进行生化和免疫化学分析，以确定 包膜-CD4-辅受体的时间和功能序列 互动。第三，这些建筑群将在 转基因小鼠疫苗模型以确定能够 诱发PI病毒中和抗体。最后，“核聚变能力” 将使用疫苗和/或纯化的包膜-CD4-辅受体复合体 在转基因小鼠中产生介导猪流感的单抗 病毒中和。这些依赖于融合的单抗将 被用来更精细地解剖HIV包膜的分子途径 蛋白质结合和融合。 在这些研究的结论中，我们将开发出一个完整的 包膜-CD4-辅受体功能和免疫原性的描述， 关于包含的关键结构和表位的定义 在能够诱导PI病毒中和的新型HIV疫苗中 抗体。

DESCRIPTION (Adapted from applicant's abstract) The recent discovery of HIV coreceptors has provided a new and dynamic vision of HIV envelope protein function. We have expanded on this concept to propose that the transient envelope protein interactions and conformational changes that arise during CD4- and coreceptor-mediated binding and fusion may expose critical targets for neutralizing antibodies, especially antibodies capable of neutralizing primary isolates (PI) of HIV. In this regard, we have developed novel 'fusion-competent' vaccines that elicit potent PI virus neutralizing antibodies when used to immunize transgenic mice that are tolerant to the human CD4 and CCR5 components of the vaccine. The finding of PI virus neutralization is in marked contract our results using comparable vaccines that present non-functioning envelope protein. In this proposal, we will firstly define the requirements and limits of PI virus neutralization by 'fusion-competent' vaccines. Secondly, we will extend these studies to identify the molecular structures and conformations that mediate HIV binding and fusion, and that serve as targets for PI virus neutralization. We have developed a specific and high-affinity 'tag' that permits the isolation of chemically crosslinked CD4-associated complexes along the pathway of binding and fusion. These complexes will be analyzed biochemically and immunochemically to define the temporal and functional sequence of envelope-CD4-coreceptor interactions. Thirdly, these complexes will be assessed in the transgenic mouse vaccination model to define structures capable of eliciting PI virus neutralizing antibodies. Finally, 'fusion-competent' vaccines and/or purified envelope-CD4-coreceptor complexes will be used in transgenic mice to generate monoclonal antibodies that mediate PI virus neutralization. These fusion-dependent monoclonal antibodies will be used to dissect more finely the molecular pathway of HIV envelope protein binding and fusion. At the conclusion of these studies, we will have developed an integrated description of envelope-CD4-coreceptor function and immunogenicity, towards the definition of critical structures and epitopes for inclusion in novel HIV vaccines capable of eliciting PI virus neutralizing antibodies.