A nanopore biosensor for leveling Mtb antigens in blood

用于平衡血液中 Mtb 抗原的纳米孔生物传感器

• 批准号: 10646134
• 负责人: Tony Y. Hu
• 金额: $ 75.26万
• 依托单位: TULANE UNIVERSITY OF LOUISIANA
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-06-15 至 2027-05-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10646134
• 关键词: Accounting; Adult; Algorithms; Antibodies; Antigens; Antitubercular Agents; Bacteriology; Biological Assay; Biological Markers; Biopsy; Biosensor; Blood; Blood specimen; Cause of Death; Cessation of life; Child; Childhood; Clinical; Clinical Data; Communicable Diseases; Containment; Data; Detection; Development; Devices; Diagnosis; Diagnostic; Diagnostic Sensitivity; Diagnostic Specificity; Diagnostic tests; Disease; Early Diagnosis; Early treatment; Epidemic; Event; Exhibits; Generations; Goals; Guidelines; HIV; Immunoprecipitation; Individual; Infection; Longterm Follow-up; Lung; Mass Spectrum Analysis; Measurement; Measures; Monitor; Mycobacterium Infections; Mycobacterium tuberculosis; Mycobacterium tuberculosis antigens; Nature; Nucleic Acids; Organizational Objectives; Patient Monitoring; Patient-Focused Outcomes; Patients; Peptides; Performance; Persons; Population; Population Heterogeneity; Prevalence; Procedures; Proteins; Protocols documentation; Pulmonary Tuberculosis; Radiology Specialty; Recommendation; Reporting; Research; Resource-limited setting; Resources; Sampling; Sensitivity and Specificity; Serum; Specificity; Specimen; Sputum; System; Technology; Test Result; Testing; Tissues; Translations; Tuberculosis; Tuberculosis diagnosis; Validation; Virulence Factors; Work; World Health Organization; biosignature; clinical application; clinical research site; cohort; cost; diagnostic assay; diagnostic platform; disease diagnosis; disorder control; fabrication; global health; improved; instrument; nanopore; non-tuberculosis mycobacteria; patient population; patient response; peripheral blood; portability; predictive modeling; programs; prototype; rapid diagnosis; research clinical testing; response; specific biomarkers; success; treatment response; trend; tuberculosis diagnostics; tuberculosis treatment; validation studies

Tuberculosis (TB) is a global health threat but can be difficult to diagnose and manage due to the sub-optimal performance and non-quantitative nature of frontline diagnostic assays, which require sputum or tissue biopsies and exhibit reduced performance when applied to diagnose paucibacillary or extrapulmonary TB cases. There is an unmet need for a rapid, non-sputum-based assay that can sensitively diagnose active TB and measure treatment responses in clinically diverse populations. We have previously reported that serum levels of two peptides derived from the Mycobacterium tuberculosis (Mtb) virulence factors CFP-10 and ESAT-6 can act as specific biomarkers of active TB and, using a mass spectrometry (MS) assay, validated their diagnostic performance in relevant cohorts of adults and children. This included HIV+/HIV-, pulmonary / extrapulmonary, and culture-positive / negative TB cases, as well as those with latent TB and nontuberculous mycobacteria infections. This MS assay had similar diagnostic sensitivity (88.6 vs 88.2%) and specificity (93.8 vs 97.2%) in adults and children and exceeded the reported performance of frontline tests in comparable populations. Serum Mtb antigen levels were also informative in monitoring anti-TB treatment responses. However, this MS assay is not suitable for use in resource limited settings. The proposed studies will therefore refine and evaluate the performance of a protein-based nanopore biosensor assay to diagnose active TB cases using the same serum biomarkers. This system is easy to operate, has low fabrication and instrument costs, and can perform high- throughput and ultra-sensitive measurements of specific Mtb-derived peptides. Its robust nature and portability also allow its use in resource-limited areas subject to high TB prevalence. Our results show that a nanopore assay can accurately detect CFP-10 and ESAT-6 peptides, and this data has significant diagnostic promise. Based on these findings, we propose that the portable protein nanopore biosensor assay that will be analyzed in these studies can improve TB diagnosis in adults and children, particularly in resource-limited areas with high TB prevalence. We will utilize this system to measure serum levels of Mtb antigen-derived peptide biomarkers in order to: (1) develop a sensitive and robust nanopore-based TB diagnostic assay; (2) validate this assay in well-organized cohorts, containing patients with pulmonary and extrapulmonary TB cases; and (3) evaluate how serum levels of Mtb antigens change in adult PTB cases in during anti-TB therapy. Given the success of these proof-of-concept studies, the long-term goal of the proposed research program is to build prototype devices for large-scale on-site clinical validation studies in high TB burden regions, and to modify and extend this system to detect other disease biomarkers. This research program should hasten the translation of a promising biosensor platform into a practical assay suitable for rapid translation to clinical applications for disease diagnosis.

结核病（TB）是全球健康的威胁，但由于前线诊断测定的次优性能和非定量性质，可能难以诊断和管理，这需要痰液或组织活检，并且在诊断可诊断paucibibaCillary或tulapullonary TB病例时表现出降低的性能。对快速，非输出的测定方法的需求未满足，可以敏感地诊断活跃的结核病并测量临床多样化种群中的治疗反应。我们先前已经报道说，从结核分枝杆菌（MTB）毒力因子CFP-10和ESAT-6衍生的两种肽的血清水平可以充当活性TB的特定生物标记物，并使用质谱法（MS）测定法（MS）测定法对成人和儿童的相关诊断进行了验证。其中包括HIV+ / HIV-，肺 /外肺外和培养阳性 /阴性结核病病例，以及潜在的结核病和无结核分枝杆菌感染。该MS分析在成人和儿童中具有相似的诊断敏感性（88.6 vs 88.2％）和特异性（93.8 vs 97.2％），并且超过了可比人群中报告的前线测试的报告。血清MTB抗原水平在监测抗TB治疗反应方面也有用。但是，此MS分析不适合在资源有限的设置中使用。因此，拟议的研究将完善并评估基于蛋白质的纳米生物传感器测定的性能，以使用相同的血清生物标志物诊断活跃的TB病例。该系统易于操作，制造和仪器成本较低，可以对特定MTB衍生的肽进行高吞吐量和超敏感的测量。它的强大性质和可移植性还允许其在较高的结核病患病率的资源有限区域中使用。我们的结果表明，纳米孔测定法可以准确检测CFP-10和ESAT-6肽，并且该数据具有显着的诊断前景。基于这些发现，我们建议在这些研究中进行分析的便携式蛋白纳米孔生物传感器测定可以改善成人和儿童的结核病诊断，尤其是在TB较高的资源有限区域中。我们将利用该系统来测量MTB抗原衍生的肽生物标志物的血清水平，以：（1）开发一种敏感且强大的基于纳米孔的TB诊断测定； （2）在组织良好的队列中验证该测定法，其中包含患有肺和肺外结核病病例的患者； （3）评估抗TB治疗期间成人PTB病例中MTB抗原的水平如何变化。鉴于这些概念验证研究的成功，拟议的研究计划的长期目标是在高结核病负担区域建立针对大规模现场临床验证研究的原型设备，并修改和扩展该系统以检测其他疾病生物标志物。该研究计划应加快将有希望的生物传感器平台转换为适合快速转化为疾病诊断临床应用的实用测定。