Integrative Genomics of the Asthma-COPD Overlap

摘要 持续症状的哮喘患者发生固定性气道阻塞（AO）的风险很大。的 这项建议的首要假设是，这是哮喘患者的最大风险是发展 COPD，这种易感性是由特定的遗传和表观遗传变异引起的，这些变异影响了 支气管上皮（BE）中表达的关键哮喘和COPD易感基因的表达，包括 HHIP和FAM 13 A、TSLP和ORMDL 3。我们认为，这些目标的特征（和定义） 其他候选人）将有助于开发更有针对性的治疗方法，以对抗 气道重塑为了验证这些假设，我们提出了三个具体目标。在目标1中，我们将描述 哮喘易感者BE的mRNA和miRNA转录组学研究 表征的群组：（i）来自CAMP的表现出正常肺生长的哮喘患者与那些表现出异常肺生长的哮喘患者， 生长或早期下降;（ii）COPD中的哮喘患者肺功能正常（GOLD 0）与肺功能异常（GOLD 0） 功能（GOLD II-III）。将使用Illumina Hi-Seq 2000生成RNA-seq表达谱（n=175）。 我们假设，与肺功能正常的哮喘患者相比， 功能，证明BE基因表达的特定模式，包括增加ORMDL 3和TSLP 表达，并降低HHIP和FAM 13 A表达。我们将描述共表达网络 受这些基因的影响，以确定其他候选人，有助于肺功能下降。在目标2中， 将使用整合基因组学方法来绘制影响关键基因表达的遗传变异， 哮喘-COPD共表达网络的核心基因。这些研究将在BE进行 在目标1中采集样本，并补充哮喘桥（n=1548）和ECLIPSE（n=200）队列， 与可用的基因组数据（基因型，表达，甲基化）。将检测已识别的监管变体， 在COPD基因（n= 10，300）中作为COPD易感性位点，在CAMP中作为固定AO的决定因素 （n=968）。在目标3中，我们将描述候选基因失调的功能后果， 气液界面模型。我们将比较耐药和耐药人群对香烟烟雾的细胞反应， 敏感受试者（每组n=6，共24个样本），使用来自受试者的BE进行气液界面分析 CAMP和COPD基因组。我们将比较这些反应后，shRNA介导的敲低 候选基因（ORMDL 3，TSLP，HHIP和FAM 13 A），并正式测试这些反应是否 类似于哮喘和COPD。我们推测，在CAMP和COPDGene衍生的样品中，我们将 在易感受试者中观察到与正常哮喘患者不同的类似细胞反应， 肺功能，为这些基因影响肺功能的机制提供关键见解。

ABSTRACT Asthmatics with persistent symptoms are at great risk of developing fixed airways obstruction (AO). The overarching hypothesis of this proposal is that it is this subset of asthmatics that is at greatest risk of developing COPD and that this susceptibility is caused by specific genetic and epigenetic variants that influence the expression of key asthma- and COPD-susceptibility genes expressed in the bronchial epithelium (BE), including HHIP and FAM13A, TSLP and ORMDL3. We argue that characterization of these targets (and defining additional candidates) will facilitate the development of more directed therapies to counter the adverse effects of airway remodeling. To test these hypotheses, we propose three Specific Aims. In Aim 1, we will characterize the mRNA and miRNA transcriptomic profiles in BE of the susceptible asthmatic in subjects from two well- characterized cohorts: (i) asthmatics from CAMP who exhibit normal lung growth vs. those with abnormal growth or early decline; (ii) asthmatics in COPDGene with normal lung function (GOLD 0) vs. abnormal lung function (GOLD II-III). RNA-seq expression profiles (n=175) will be generated using the Illumina Hi-Seq 2000. We hypothesize that asthmatics with reduced lung function decline, compared to those with preserved lung function, demonstrate a specific pattern of BE gene expression, including increased ORMDL3 and TSLP expression, and reduced HHIP and FAM13A expression. We will characterize the co-expression network influenced by these genes to define additional candidates that contribute to lung function decline. In Aim 2, we will use an integrative genomics approach to map the genetic variants that influence the expression of key genes at the core of the asthma-COPD co-expression networks. These studies will be performed in the BE samples collected in Aim 1, and complemented by Asthma BRIDGE (n=1548) and ECLIPSE (n=200) cohorts, with available genomic data (genotype, expression, methylation). Identified regulatory variants will be tested for association as COPD-susceptibility loci in COPDGene (n=10,300), and as determinants of fixed AO in CAMP (n=968). In Aim 3, we will characterize the functional consequences of dysregulation of the candidate genes in an air-liquid interface model. We will compare cellular responses to cigarette smoke among resistant and susceptible subjects (n=6 per group, 24 samples total) in air liquid interface using BE derived from subjects from the CAMP and COPDGene cohorts. We will compare these responses following shRNA-mediated knockdown of candidate genes (ORMDL3, TSLP, HHIP and FAM13A) and formally test whether these responses are similar in asthma and COPD. We speculate that in both the CAMP and COPDGene derived samples, we will see similar cellular responses in the susceptible subjects that are distinct from those in asthmatics with normal lung function, providing key insights in to the mechanisms by which these genes impact lung function.