Point-of-care immunoassay for early diagnosis of pertussis

用于早期诊断百日咳的即时免疫分析

• 批准号: 8780071
• 负责人: Amanda Burnham-Marusich
• 金额: $ 29.21万
• 依托单位: DXDISCOVERY, INC.
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-06-01 至 2016-05-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8780071
• 关键词: Antibodies; Antigen Targeting; Antigens; Bacteria; Benchmarking; Binding Sites; Bioinformatics; Biological; Biological Assay; Bordetella pertussis; Case Study; Cells; Cessation of life; Clinic; Clinical Trials; Communities; Containment; Coughing; DNA; Detection; Development; Diagnosis; Diagnostic; Disease; Disease Outbreaks; Early Diagnosis; Epitopes; Equipment; Goals; Immunoassay; Infant; International; Lateral; Life; Lung diseases; Metric; Monoclonal Antibodies; Nasopharynx; Patient Care; Patients; Pertussis; Pharyngitis; Phase; Proteins; Public Health; Resources; Respiratory Tract Infections; Sampling; Sensitivity and Specificity; Small Business Innovation Research Grant; Specificity; Staging; Streptococcal Infections; Swab; Symptoms; Target Populations; Technology; Test Result; Testing; Time; United States; Vaccines; Validation; antigen binding; base; clinically relevant; cost; design; effective therapy; improved; meetings; novel; pediatrician; phase 1 study; phase 2 study; point of care; polyclonal antibody; preclinical study; prototype; public health relevance; rapid diagnosis; research clinical testing; research study; response; success; transmission process

DESCRIPTION (provided by applicant): Pertussis is a serious and potentially life-threatening respiratory disease caused by the bacterium Bordetella pertussis. Worldwide, 18.4 million people become ill with pertussis each year, and 254,000 of these patients die. Despite high vaccine coverage in the United States, there were still over 48,000 pertussis cases reported in the U.S. in 2012. Rapid diagnosis of pertussis is critical because treatment and outbreak containment are only effective if initiated early in the disease. Currently, there are no assays that can rapidly and accurately detect early-stage pertussis. The goal is to develop a rapid, point-of-care (POC) immunoassay to detect B. pertussis during early disease. The target population will be infants who present with the non-specific, respiratory tract infection symptoms of early pertussis (prior to onset of paroxysmal cough). The approach will be detection of B. pertussis antigens from nasopharyngeal swabs by lateral flow immunoassay (LFI). The product will be an assay relevant to i) U.S. pediatrician offices, and ii) international clinics in low-resource settings: a sensitive and specific test that delivers results in 5-10min at very low cost without specialized equipment or user expertise. In preliminary studies, we used a bioinformatics analysis to identify five high priority B. pertussis protein antigens for further stdy. Specific Aim 1 will develop epitope-specific polyclonal antibodies (pAbs) for detection of B. pertussis antigens. pAbs will be screened to select those that i) react with B. pertussis proteins, ii) react with B. pertussis but not other bacteria found in the nasopharynx, iii) can function in pairs to bind the antigen in the sandwich format necessary for LFI construction, and iv) detect antigen in patient nasopharyngeal samples. Only pAbs that meet all the criteria will be advanced to LFI prototype development in Specific Aim 2. Specific Aim 2 will determine the limit of detection of B. pertussis by LFI by optimizing the physical and biological components of the LFI prototypes. Based on previously determined B. pertussis loads in infant NP swabs of 106 bacteria per swab, the benchmark for sensitivity is detection of 105 CFU per swab. Together, the experiments of Phase I will determine which B. pertussis antigen epitopes can produce pAb pairs that yield LFI prototypes with the required sensitivity. A Phase II study would develop monoclonal antibodies (mAbs) against the validated epitopes, incorporate them into an LFI, and perform detailed pre-clinical studies. Successful navigation of a full clinical trial in Phase III would ultimately yield a POC immunoassay that could dramatically increase early pertussis diagnosis, limit pertussis transmission during community outbreaks, and save patient lives.

描述（由申请人提供）：百日咳是一种严重且潜在的威胁生命的呼吸道疾病，是由Bordetella bordtussis细菌引起的。在全球范围内，每年有1,840万人患有百日咳，其中254,000人死亡。尽管在美国，疫苗的覆盖范围很高，但2012年美国仍有超过48,000例百日咳病例。迅速诊断百日咳是至关重要的，因为治疗和疫情遏制只有在疾病早期开始时才有效。当前，没有任何测定能够快速，准确地检测到早期百日咳。目的是开发快速的，即保健（POC）的免疫测定方法，以检测早期疾病期间百日咳。目标人群将是婴儿早期百日咳的非特异性呼吸道感染症状（阵发性咳嗽之前）。该方法将是通过横向流量免疫测定法（LFI）从鼻咽拭子中检测到百日咳抗原。该产品将是与I）美国儿科医生办公室有关的测定法，以及II）在低资源环境中的国际诊所：一种敏感和特定的测试，在没有专业设备或用户专业知识的情况下以非常低的成本提供了5-10分钟的结果。在初步研究中，我们使用了生物信息学分析来识别五个高优先级B.百日咳蛋白抗原以进行进一步的Stdy。特定的目标1将开发表位特异性多克隆抗体（PAB），以检测百日咳抗原抗原。将对PAB进行筛选以选择i）与百日咳蛋白质反应的那些 ii）与百日咳芽孢杆菌反应，但没有在鼻咽中发现的其他细菌，iii）可以成对起作用，以结合LFI构造所需的夹层格式的抗原，而iv）在患者鼻咽样样品中检测抗原。在特定的目标2中，只有符合所有标准的PAB才能将其推进到LFI原型的开发中。特定的目标2将通过优化LFI原型的物理和生物学成分来确定LFI对百日咳芽孢杆菌的检测极限。基于先前确定的B.百日咳载荷，每拭子106个细菌的木棉中的百日咳载荷，敏感性的基准是每拭子105 cfu。同时，第一阶段的实验将确定哪些百日咳抗原表位可以产生PAB对，从而产生具有所需灵敏度的LFI原型。 II期研究将针对经过验证的表位开发单克隆抗体（mAb），将其纳入LFI，并进行详细的临床前研究。在第三阶段进行全面临床试验的成功导航最终将产生POC免疫测定，该试验可能会大大增加百日咳诊断，限制社区疫情期间的百日咳传播并挽救患者的生命。