Novel HIV 1 Envelope immunogens derived from broadly neutralizing plasmas

描述（由申请人提供）：我们研究的总体目标是设计能够引发广泛反应性中和抗体（NAb）的新型HIV包膜免疫原。尽管通过疫苗接种引发广泛中和抗体应答的尝试尚未成功，但在一些慢性感染的HIV+受试者中检测到了异常强效的广泛中和抗体应答。我们建议检测新的免疫原，这些免疫原来源于从充分表征的HIV+受试者中分离的包膜序列，该受试者的特异性广泛中和活性靶向CD 4受体结合位点（CD 4-BS）。我们假设，来自HIV-1感染受试者的HIV-1包膜免疫原（其血浆对CD 4-BS表现出强效、广泛的交叉中和活性）将引发能够广泛交叉中和的NAb。 这项建议分为两个不同的阶段。在第一阶段，我们提出以下建议：首先，构建和表征可溶性三聚体gp 140 Env蛋白，其来源于从充分表征的广泛中和血浆中分离的自体Env序列。其次，我们将使用这些三聚体gp 140 Env作为免疫原，并监测它们在动物中引发广泛中和抗体应答的能力。我们将仔细分析疫苗接种引起的NAb应答，以确定引起的NAb应答的质量和表位靶点。 在第二阶段，我们将从接种疫苗后产生广泛中和抗体的动物中分离出新的单克隆中和抗体。我们将利用噬菌体展示文库和抗原特异性单B细胞克隆从免疫动物的脾和骨髓组织中分离抗体。 将分离新型抗体和Fab并广泛表征其结合和中和活性。 有前途的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。本研究中的新型抗HIV中和抗体将为研究界提供有价值的新工具，并有助于确定HIV-1包膜刺突上的新中和表位。 公共卫生相关性：该提案将测试新的HIV-1候选疫苗，这些候选疫苗来自HIV-1+受试者的循环病毒，这些受试者在自然感染时产生了广泛中和抗体。我们将用HIV-1包膜蛋白免疫兔子，监测它们是否产生中和抗体，并在产生广泛中和抗体反应的动物中分离出新的中和抗体。该项目将评估自然产生广泛中和抗体的受试者中存在的HIV-1包膜序列可用于通过疫苗接种在其他人中引发相同类型的免疫应答的可能性。

DESCRIPTION (provided by applicant): 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. PUBLIC HEALTH RELEVANCE: This proposal will test new HIV-1 vaccine candidates derived from the circulating viruses of HIV-1+ subjects who developed broadly neutralizing antibodies in response to natural infection. We will immunize rabbits with HIV-1 Envelope proteins, monitor them for the development of neutralizing antibodies, and isolate new neutralizing antibodies in those animals that develop broadly neutralizing antibody responses. This project will evaluate the possibility that HIV-1 Envelope sequences that are present in subjects who naturally developed broadly neutralizing antibodies can be used to elicit the same type of immune response in others by vaccination.