Novel HIV 1 Envelope immunogens derived from broadly neutralizing plasmas

摘要 我们研究的总体目标是设计新型 HIV 包膜免疫原 能够引发广泛反应性中和抗体（NAb）。虽然尝试 通过疫苗接种引起广泛中和抗体反应 不成功的、异常有效的广泛中和抗体反应已被 在一些长期感染艾滋病毒的受试者中检测到。我们建议测试新的 源自从良好分离的包膜序列的免疫原 HIV 受试者的特征是其特殊的广泛中和活性针对 CD4 受体结合位点 (CD4-BS)。我们假设 HIV-1 包膜 来自 HIV-1 感染者的免疫原，其血浆表现出有效的、 对 CD4-BS 的广泛交叉中和活性将引发能够 广泛的交叉中和作用。 该提案分为两个不同的阶段。在第一阶段，我们 提出以下建议：首先，构建并表征可溶性三聚体 gp140 Env 源自自体 Env 序列的蛋白质，其分离自充分表征的 广泛中和等离子体。其次，我们将使用这些三聚 gp140 环境作为 免疫原并监测其引发广泛中和抗体反应的能力 动物。我们将仔细剖析疫苗接种引起的 NAb 反应 确定引发的 NAb 反应的质量和表位靶点。 在第二阶段，我们将从中分离出新的单克隆中和抗体 因接种疫苗而产生广泛中和抗体的动物。我们 将利用噬菌体展示文库和抗原特异性单 B 细胞克隆来 从免疫动物的脾脏和骨髓组织中分离抗体。 小说 抗体和 Fab 将被分离并广泛表征其结合和 中和活性。 有前景的 Fab 将被制成完整的免疫球蛋白 G 分子并广泛表征交叉中和潜力和结合 表位。这项研究中的新型抗 HIV 中和抗体将提供有价值的新成果 为研究界提供工具，并帮助定义新的中和表位 HIV-1 包膜尖峰。

ABSTRACT The overall aim of our studies is to design novel HIV Envelope immunogens capable of eliciting broadly reactive neutralizing antibodies (NAbs). Although attempts at eliciting broadly neutralizing antibody responses by vaccination have been unsuccessful, exceptionally potent broadly neutralizing antibody responses have been detected in some chronically infected HIV+ subjects. We propose to test new immunogens that are derived from Envelope sequences that were isolated from a well- characterized HIV+ subject whose exceptional broadly neutralizing activity targeted the CD4 receptor binding site (CD4-BS). We hypothesize that HIV-1 Envelope immunogens derived from HIV-1 infected subjects whose plasma exhibits potent, broad cross-neutralizing activity to the CD4-BS will elicit NAbs that are capable of broad cross-neutralization. This proposal is separated into two distinct phases. In the first phase, we propose the following: First, construct and characterize soluble trimeric gp140 Env proteins derived from autologous Env sequences isolated from well-characterized broadly neutralizing plasmas. Second, we will use these trimeric gp140 Envs as immunogens and monitor their ability to elicit broadly neutralizing antibody responses in animals. We will carefully dissect the NAb responses elicited by vaccination to determine both the quality and the epitope targets of the elicited NAb responses. In the second phase, we will isolate new monoclonal neutralizing antibodies from animals that develop broadly neutralizing antibodies in response to vaccinations. We will utilize both phage display libraries and antigen-specific single B cell cloning to isolate antibodies from spleen and bone marrow tissue of immunized animals. Novel antibodies and Fabs will be isolated and extensively characterized for binding and neutralizing activity. Promising Fabs will be made into full immunoglobulin G molecules and extensively characterized for cross-neutralizing potential and binding epitope. Novel anti-HIV neutralizing antibodies from this study will provide valuable new tools to the research community, and help to define new neutralization epitopes on the HIV-1 Envelope spike.