Single cycle reporter assay for quantifying HIV-1 Nab

用于量化 HIV-1 Nab 的单循环报告基因检测

• 批准号: 6694007
• 负责人: JOHN Christopher KAPPES
• 金额: $ 18.73万
• 依托单位: TRANZYME, INC.
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-09-02 至 2005-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6694007
• 关键词: antiviral antibody; clinical research; human immunodeficiency virus 1; human tissue; immunologic assay /test; neutralizing antibody; reporter genes; technology /technique development; virus infection mechanism; virus integration; virus protein

DESCRIPTION (provided by applicant): The humoral arm of the immune response is considered to be the major protective mechanism elicited by a variety of viral vaccines. The induction of a potent neutralizing antibody response is one of the principal goals in HIV vaccine development. However, it has been remarkably difficult to ascertain what affect neutralizing antibodies may play in the containment of HIV primary infection, the natural history of disease or vaccination. We suggest that these difficulties stem from a combination of circumstances, some biologic and some technical. To overcome the technical limitations, we have developed a novel HIV-1 entry assay for measuring neutralizing antibodies (Nab). This assay is based on a reporter cell line (JC53-BL) that is sensitive to primary HIV-1 isolates, directly analyzes HIV entry steps and accurately quantifies entry in a single cycle of infection. Moreover, the assay is relatively simple and robust. Ultimately, the commercial viability will depend on whether the results produced from this entry assay have biologic, immunologic and clinical relevance. That is - whether the inhibition of HIV-1 infection by Nab in vitro, on JC53-BL cells, mimics inhibition on relevant target cells in vivo. We are encouraged by results obtained using this assay that have show (a) the emergence of resistant HIV-1 in patients receiving fusion inhibitor (T-20) monotherapy, and (b) Nab exert sufficient virus inhibitory pressure, in vivo, to result in the replacement of neutralization sensitive virus by resistant virus. Our hypothesis is - our entry assay is commercially viable and clinically relevant because it replicates important entry mechanisms of most HIV-1 strains on natural target cells. To address this hypothesis we propose: (1) To analyze the susceptibility of the entry assay's reporter cell line to infection by diverse strains of HIV-1 and (2) To determine the sensitivity of the JC53-BL-based entry assay for detecting heterologous Nab.

描述（由申请人提供）：免疫反应的体液臂被认为是多种病毒疫苗引发的主要保护机制。诱导有效的中和抗体反应是艾滋病毒疫苗开发的主要目标之一。然而，要确定中和抗体在遏制 HIV 原发感染、疾病自然史或疫苗接种方面可能发挥的作用非常困难。我们认为这些困难源于多种情况的结合，一些是生物学的，一些是技术的。为了克服技术限制，我们开发了一种新型 HIV-1 进入测定法，用于测量中和抗体 (Nab)。该检测基于对原代 HIV-1 分离株敏感的报告细胞系 (JC53-BL)，可直接分析 HIV 进入步骤并准确量化单个感染周期中的进入情况。此外，该测定相对简单且稳健。最终，商业可行性将取决于该入门测定产生的结果是否具有生物学、免疫学和临床相关性。也就是说，Nab 在体外对 JC53-BL 细胞上的 HIV-1 感染的抑制是否模仿了体内对相关靶细胞的抑制。我们对使用该测定获得的结果感到鼓舞，这些结果表明（a）接受融合抑制剂（T-20）单一疗法的患者中出现了耐药性 HIV-1，以及（b）Nab 在体内发挥了足够的病毒抑制压力，从而导致用抗性病毒替代中和敏感病毒。我们的假设是——我们的进入检测具有商业可行性和临床相关性，因为它复制了大多数 HIV-1 毒株对天然靶细胞的重要进入机制。 为了解决这一假设，我们建议：(1) 分析进入测定的报告细胞系对不同 HIV-1 毒株感染的敏感性；(2) 确定基于 JC53-BL 的进入测定检测异源病毒的敏感性纳布。