Metabolomics-based discovery of small molecule biomarkers for noninvasive dengue

描述（由申请人提供）：流行性登革热（DF）和登革出血热（DHF/DSS）已经出现在整个热带世界，造成毁灭性的公共卫生后果。过去十年来，拉丁美洲严重登革热的急剧增加令人严重关切；30%的病例现在被诊断为严重登革热病毒感染。在许多发展中国家，DEN正在压倒公共卫生的临床护理能力。该提案的总体目标是为DENV感染的诊断和预后（D&P）提供重大改变。代谢组学方法将用于鉴定血清和非侵入性临床标本（尿液和唾液）中出现的候选代谢物小分子生物标志物（smb），以诊断DENV感染并预测严重疾病的进展。使用基于液相色谱-质谱（LC-MS）的代谢组学方法，对DEN患者急性期标本进行了初步研究，确定了血清、唾液和尿液中DF和DHF/SS的许多分子特征和候选smb。在该项目的R21阶段，我们将使用代谢指纹识别方法，在尼加拉瓜高质量儿科DEN医院和队列研究中回顾性收集的血清标本中确认现有的和识别新的候选smb，将开始表征smb及其代谢途径，将在医院研究中前瞻性收集的血清、唾液和尿液标本中调查这些smb对D&P的功效。并将确定区分DEN、严重DEN和非DEN疾病的一系列最重要的分子特征。我们将开发包括smb、临床体征和症状以及DENV感染D&P的临床实验室结果在内的算法。在项目的R33阶段，从医院研究和社区队列研究中前瞻性收集的血清、唾液和尿液样本将通过使用lc -串联质谱（LC-MS/MS）的代谢谱分析来确定候选smb。选定中小企业的第一代EIA检测将纳入尼加拉瓜的诊断方案。将确定候选中小企业和“第一代”中小企业检测的诊断和预后敏感性和特异性，以及基于中小企业诊断的首选临床标本。总体而言，这些研究将确定一组中小企业（例如5-10家），这些中小企业将用于制定用于诊所和医院用于DEN D&P的快速护理点（POC）测试的目标产品概况（TPP）。通过检测唾液和尿液中的smb来预测严重的DEN是一项创新，通过使用廉价、容易获得、无创的DEN临床标本进行D&P，为真正的诊断模式转变提供了机会。

DESCRIPTION (provided by applicant): Epidemic dengue fever (DF) and dengue hemorrhagic fever (DHF/DSS) have emerged throughout the tropical world with devastating public health consequences. A dramatic increase in severe dengue disease (DEN) in Latin America in the last decade is of grave concern; 30% of cases are now diagnosed as severe dengue virus (DENV) infections. DEN is overwhelming public health capacity for clinical care in much of the developing world. The overall goal of this proposal is to provide a major change in the diagnosis and prognosis (D&P) of DENV infections. A metabolomics approach will be used to identify candidate metabolite small molecule biomarkers (SMBs) that occur both in serum and in non-invasive clinical specimens (urine and saliva) that diagnose DENV infection and predict progression to severe disease. Preliminary studies using acute phase specimens from DEN patients have identified a number of molecular features and candidate SMBs of DF and DHF/SS in serum, saliva, and urine using liquid chromatography-mass spectrometry (LC-MS)-based metabolomics. In the R21 phase of this project, we will use a metabolic fingerprinting approach to confirm existing and identify new candidate SMBs in retrospectively collected serum specimens available from the high-quality pediatric DEN hospital-based and cohort studies in Nicaragua, will begin to characterize the SMBs and metabolic pathways involved, will investigate the efficacy for D&P of these SMBs in prospectively collected serum, saliva, and urine specimens in the hospital study, and will identify a portfolio of the most significant molecular features that differentiate DEN, severe DEN, and non-DEN disease. We will develop algorithms including SMBs, clinical signs and symptoms, and clinical laboratory results for the D&P of DENV infections. In the R33 phase of the project, prospectively collected serum, saliva, and urine samples from both the hospital study and a community-based cohort study will be analyzed by metabolic profiling using LC-tandem MS (LC-MS/MS) to identify candidate SMBs. First generation EIA tests for selected SMBs will be included in the diagnostic regimen in Nicaragua The diagnostic and prognostic sensitivity and specificity of the candidate SMBs and "first generation" SMB tests will be determined as will the preferred clinical specimen for SMB-based diagnoses. Overall these studies will identify a panel of SMBs (e.g. 5-10), which will be used to formulate the Target Product Profiles (TPP) for rapid point-of-care (POC) tests for use in clinics and hospitals for DEN D&P. Detection of SMBs in saliva and urine that are predictive of severe DEN is innovative and provides the opportunity for a true paradigm shift in diagnosis by using inexpensive, easily procured, non-invasive clinical specimens for D&P of DEN.