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的HIV包膜免疫原 能够引起广泛反应的中和抗体(NAB)。尽管尝试 在通过接种疫苗引起广泛的中和抗体反应方面， 不成功的，异常有效的广谱中和抗体反应 在一些慢性感染艾滋病毒的受试者中检测到。我们建议测试新的 从井中分离的包膜序列中提取的免疫原- 具有艾滋病毒主题的特征，其特殊的广泛中和活动针对 CD_4受体结合位点(CD_4-BS)。我们假设HIV-1信封 来自HIV-1感染者的免疫原，其血浆表现出效力， 广泛的对CD4-BS的交叉中和活性将诱导能够 广泛的交叉中和。 这项提议分为两个不同的阶段。在第一阶段，我们 提出如下建议：第一，构建和鉴定可溶性三聚体gp140环境蛋白 来自自体膜蛋白序列的蛋白质从鉴定良好的 宽泛的中和等离子体。其次，我们将使用这些三聚体gp140 env作为 免疫原并监测其诱导广谱中和抗体反应的能力 动物。我们将仔细分析疫苗接种引起的NAB反应，以 确定激发的NAB反应的质量和表位靶点。 在第二阶段，我们将分离出新的单克隆中和抗体 在接种疫苗后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个B细胞克隆来 从免疫动物的脾和骨髓组织中分离抗体。小说 抗体和抗体将被分离和广泛鉴定以结合和 中和活性。有希望的纤维将被制成完全的免疫球蛋白G 分子，并广泛表征交叉中和潜力和结合 表位。本研究中的新型抗HIV中和抗体将提供有价值的新的 工具，并帮助定义新的中和表位 HIV-1信封尖峰。

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.