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（由申请方提供）：免疫应答的体液臂被认为是各种病毒疫苗引发的主要保护机制。诱导有效的中和抗体应答是HIV疫苗开发的主要目标之一。然而，很难确定中和抗体在遏制HIV原发感染、疾病自然史或疫苗接种中可能发挥的作用。我们认为，这些困难源于各种情况，有些是生物性的，有些是技术性的。为了克服技术上的局限性，我们开发了一种新的HIV-1进入检测方法，用于测量中和抗体（Nab）。该检测试剂盒基于对原代HIV-1分离株敏感的报告细胞系（JC 53-BL），可直接分析HIV进入步骤，并准确定量单个感染周期中的进入。此外，该测定法相对简单且稳健。最终，商业可行性将取决于该进入检测试剂盒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JC 53-BL细胞上的HIV-1感染的抑制是否模拟了对体内相关靶细胞的抑制。我们对使用该测定法获得的结果感到鼓舞，这些结果表明：（a）在接受融合抑制剂（T-20）单药治疗的患者中出现耐药HIV-1，和（B）Na B在体内发挥足够的病毒抑制压力，导致中和敏感性病毒被耐药病毒取代。我们的假设是-我们的进入检测是商业上可行的和临床相关的，因为它复制了大多数HIV-1毒株在天然靶细胞上的重要进入机制。 为了解决这一假设，我们提出：（1）分析进入试验的报告细胞系对不同HIV-1毒株感染的敏感性，和（2）确定基于JC 53-BL的进入试验检测异源Nab的灵敏度。