Gene Expression Biomarkers for Diagnosis of Lyme Disease

描述(由申请人提供)：莱姆病是美国最常见的扁虱传播传染病，其发病率和范围都在增加。早期诊断和治疗莱姆病是防止疾病进展和后遗症的关键。目前的实验室诊断主要是血清学分析，它们在一些关键领域失败了。它们的特异性不够，对早期莱姆病不敏感，也无法确定治疗反应。更重要的是，由于血清学检测是基于抗体的存在，它们不能区分活动性感染和免疫记忆，因为先前已解决的伯氏疏螺旋体感染。需要新的范例和分析方法来弥补我们诊断能力中的这些差距。这里我们提出了一种新的方法。我们将利用差异基因表达分析来确定莱姆病患者血液中对伯氏杆菌的特征反应，长期目标是开发一种可靠的基于基因的活动性感染诊断测试。具体地说，我们建议：1)使用微阵列分析来鉴定从活动期、培养确认的莱姆病患者中分离的外周血单个核细胞(PBMC)中差异表达的基因，并将结果与来自健康捐赠者的PBMC进行比较；2)使用预测因子选择的监督学习方法，识别一组预测伯氏杆菌感染(而不是其他常见血液传播细菌病原体)的判别基因；3)通过实时RT-PCR在症状缓解的患者经过标准抗生素治疗后获得的成对急性期和恢复期患者样本中评估被鉴定为伯氏杆菌特异指示基因的基因的表达水平。这将提供有关成功治疗的患者随着时间的推移基因表达的信息，并应该定义具有活跃的、与已解决的伯氏杆菌感染相反的特征的鉴别基因的子集。这些研究有望确定一套明确的指示基因，这些基因在活动性莱姆病患者中受到显著和独特的调控，而不是那些成功使用抗生素治疗的患者或患有其他常见血液传播细菌感染的患者。一旦确定，这个分类基因的子集可以作为开发一种敏感和特异的伯氏杆菌感染诊断试验的基础。 公共卫生相关性：莱姆病是美国最常见的扁虱传播疾病，是由伯氏疏螺旋体感染引起的，目前的诊断基于血清学测试，缺乏敏感性和特异性，特别是在早期疾病中，并且不区分活动性感染和先前解决的伯氏疏螺旋体感染。拟议的研究有望确定一套明确的指示基因，这些基因在活动期莱姆病患者中受到显著和独特的调控，而不是其他常见的血液传播细菌感染患者或有或没有持续症状的经抗生素治疗、已解决的莱姆病患者。一旦确定，这个分类基因的子集可以用作开发一种敏感和特异的基于基因的活动性伯氏杆菌感染诊断试验的基础。

DESCRIPTION (provided by the applicant): Lyme disease is the most frequently reported tick-borne infectious disease in the United States and both its incidence and range are increasing. Early diagnosis and treatment of Lyme disease is critical to prevent disease progression and sequellae. Current laboratory diagnostics are primarily serologic assays and they fail in a number of key areas. They are insufficiently specific, not sensitive in early Lyme disease and cannot determine treatment response. More importantly, because serological assays are based on the presence of antibodies, they are incapable of discriminating between active infection and immunological memory due to prior, resolved infection with Borrelia burgdorferi. New paradigms and assays are needed to address these gaps in our diagnostic capabilities. Here we propose a new approach. We will utilize differential gene expression analysis to identify a signature response to B. burgdorferi in the blood of Lyme disease patients, with the long-term objective of developing a reliable gene-based diagnostic test for active infection. Specifically we propose to: 1) use microarray analysis to identify genes differentially expressed in peripheral blood mononuclear cells (PBMCs) isolated from patients with active, culture-confirmed Lyme disease and compare the results to PBMCs from healthy donors; 2) identify a set of discriminator genes predictive of B. burgdorferi infection, as opposed to infection by other common blood-borne bacterial pathogens, using a supervised learning approach of predictor selection; 3) assess expression levels of genes identified as B. burgdorferi-specific indicator genes by real-time RT-PCR in paired acute and convalescent patient samples obtained following standard antibiotic treatment in patients with resolution of symptoms. This will provide information on the gene expression over time in successfully-treated patients and should define a subset of discriminator genes characteristic of active, as opposed to resolved, B. burgdorferi infection. These studies are expected to result in the identification of a defined set of indicator genes which are significantly and uniquely regulated in patients with active Lyme disease as opposed to patients that have been successfully treated with antibiotics or individuals with other common blood-borne bacterial infections. Once identified, this subset of classifier genes can serve as the basis for development of a sensitive and specific diagnostic test for active B. burgdorferi infection. PUBLIC HEALTH RELEVANCE: Lyme disease, the most frequently reported tick-borne disease in the United States, is caused by infection with the spirochete Borrelia burgdorferi and diagnosis is currently based on serological tests which lack sensitivity and specificity, especially in early disease, and do not discriminate between active infection and prior, resolved infection with B. burgdorferi. The proposed studies are expected to result in the identification of a defined set of indicator genes which are significantly and uniquely regulated in patients with active Lyme disease as opposed to patients with other common blood-borne bacterial infections or individuals with antibiotic-treated, resolved Lyme disease with or without ongoing symptoms. Once identified, this subset of classifier genes could be employed as the basis for development of a sensitive and specific gene-based diagnostic test for active B. burgdorferi infection.