FUSION COMPETENT HIV VACCINE IMMUNOGENS

描述（改编自申请人的摘要）最近发现， HIV共受体为HIV包膜的研究提供了一个新的动态视角 蛋白质功能我们对这一概念进行了扩展，提出 瞬时包膜蛋白相互作用和构象变化， 在CD4和辅助受体介导的结合和融合过程中出现， 中和抗体的关键目标，尤其是抗体 能够中和HIV的原代分离株（PI）。在这方面，委员会注意到， 我们已经开发出新型的"融合能力"疫苗， 用于免疫转基因小鼠时的PI病毒中和抗体 对疫苗的人CD4和CCR5组分具有耐受性。 PI病毒中和的发现与我们的结果显著一致 使用呈现无功能包膜蛋白的可比疫苗。 在本建议中，我们将首先确定以下要求和限制： 通过"融合活性"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， 介导HIV结合和融合的构象， PI病毒中和的靶点。我们制定了一个具体的， 高亲和性"标签"，其允许分离化学交联的 CD4-相关复合物沿着结合和融合的途径。这些 将对复合物进行生物化学和免疫化学分析， CD4辅助受体时间和功能序列 交互.第三，这些综合大楼将在 转基因小鼠疫苗接种模型，以确定能够 引发PI病毒中和抗体。最后，"融合能力" 将使用疫苗和/或纯化的CD4-辅助受体复合物 以产生介导PI的单克隆抗体 病毒中和这些融合依赖性单克隆抗体将 用于更精细地剖析HIV包膜的分子途径 蛋白质结合和融合。 在这些研究结束时，我们将制定一个综合的 CD4辅助受体功能和免疫原性的描述， 对关键结构和表位的定义 在能够引发PI病毒中和的新型HIV疫苗中， 抗体的

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.