Prospective Longitudinal Assessment of Culture-Independent Molecular Airway Markers of Nontuberculous Mycobacteria

大量证据表明，非结核分枝杆菌（NTM）是结核病的重要病原体。 囊性纤维化（CF）和其他阻塞性肺病。目前，气道培养是“黄金标准”， 所有的诊断和治疗决定都是基于这个。缺乏敏感和特异的NTM标记物， 气道阻碍了患者护理和临床试验设计。NTM感染的非培养依赖性标志物 克服标准NTM培养物的许多局限性的潜力，特别是非常缓慢的生长（向上）， 至8周），无法定量细菌负荷，以及由于需要去污而导致的低灵敏度 程序.挥发性代谢物分析和多重基因测序的初步数据 同时对相同的痰液样本进行检测，已经证明了快速、可定量地检测出 超过标准培养物敏感性的NTM。2018年，CF基金会赞助的预测 （囊性纤维化中NTM疾病的前瞻性评价，NCT 02073409）和患者（前瞻性 囊性纤维化中NTM治疗算法，NCT 02419989）试验将扩展至10个CF程序 在全国范围内标准化诊断和测试基于方案的NTM肺动脉高压治疗方法 疾病这些试验代表了一个理想的平台，以推进NTM的分子标记物的发展 疾病状态。作为对PAR-18-643 NHLBI临床辅助研究的回应，本提案将测试中心 假设：与痰培养相比，非培养依赖性标记物组在检测存在性方面具有优势 的细菌，预测临床意义和评估CF气道中NTM的治疗反应。作为 PREDICT和PATIENCE试验的时间敏感性辅助研究，目标1将测试痰液的效用 挥发性代谢物作为气道NTM的非培养依赖性标志物。目标2将测试痰液的效用 多重基因测序作为气道NTM的分子标志物，Aim 3将测试以下组合： 挥发性代谢物和多重基因测序数据结合标准培养结果， 受试者更可能发生NTM疾病和/或对治疗有反应。 如果成功的话，将对来自儿童和成人患者的样本进行横断面和纵向分析， 明确的NTM感染状态将揭示每种测定单独和组合的优点和缺点， 与标准培养物相比，在存在细菌共培养物的情况下， 感染这些标志物可用于NTM风险患者人群的定期筛查，以促进 早期检测此外，这些测定具有区分惰性感染和非惰性感染的潜力。 NTM疾病，以及对治疗反应的纵向监测和根除的确定 已经达到病原体的水平或已经达到亚临床阈值，这可能标志着 治疗长期目标是向CF患者提供经验证的NTM气道疾病标志物 人群，适用于具有NTM疾病风险的呼吸系统疾病的所有患者人群。

A preponderance of evidence has identified nontuberculous mycobacteria (NTM) as important pathogens in cystic fibrosis (CF) and other obstructive lung diseases. Currently, airway cultures are the “gold standard” by which all diagnosis and treatment decisions are based. The lack of sensitive and specific markers of NTM in the airway impedes patient care and clinical trial design. Culture-independent markers of NTM infection have the potential to overcome many of the limitations of standard NTM cultures, especially the very slow growth (up to 8 weeks), inability to quantitate bacterial burden, and low sensitivity due to required decontamination procedures. Preliminary data from analysis of volatile metabolites and multiplex gene sequencing performed simultaneously on the same sputum samples have demonstrated rapid, quantifiable detection of the presence of NTM that exceeds the sensitivity of standard cultures. In 2018, the CF Foundation-sponsored PREDICT (PRospective Evaluation of NTM Disease In CysTic Fibrosis, NCT02073409) and PATIENCE (Prospective Algorithm for Treatment of NTM in Cystic Fibrosis, NCT02419989) trials will be expanded to 10 CF Programs nationwide to standardize the diagnosis and test a protocol-based approach to treatment of NTM pulmonary disease. These trials represent an ideal platform to advance development of molecular markers of NTM disease status. In response to PAR-18-643 NHLBI Clinical Ancillary Studies, this proposal will test the Central Hypothesis: Culture-independent marker sets have advantages over sputum cultures for detecting presence of bacteria, predicting clinical significance and assessing treatment response of NTM in the CF airway. As a time-sensitive ancillary study to the PREDICT and PATIENCE trials, Aim 1 will test the utility of sputum volatile metabolites as culture-independent markers of airway NTM. Aim 2 will test the utility of sputum multiplex gene sequencing as molecular markers of airway NTM, and Aim 3 will test the combination of volatile metabolites and multiplex gene sequencing data combined with standard culture results to identify subjects more likely to develop NTM disease and/or respond to treatment. If successful, the cross-sectional and longitudinal analyses of samples from pediatric and adult patients with well-defined NTM infection status will reveal the strengths and weakness of each assay alone and combined, in comparison to standard culture for the detection and identification of NTM in the presence of bacterial co- infection. These markers can be applied to periodic screening of patient populations at risk for NTM to facilitate early detection. In addition, these assays have the potential to differentiate between indolent infection and NTM disease, as well as longitudinal monitoring of response to treatment and determination if eradication of the pathogen has been achieved or a subclinical threshold has been reached, which could signal end of treatment. The long-term goal is to deliver validated markers of NTM airway disease to the CF patient population, with applicability to all patient populations with respiratory conditions at risk for NTM disease.