Pulmonary Fibrosis and Telomerase Dysfunction

摘要 无偏见的基因组方法导致了新的疾病基因和变异的发现。致病性最强 在家族性肺纤维化（FPF）和特发性肺纤维化（IPF）患者中发现的罕见变异 导致端粒缩短。因此，这些疾病是已知的疾病谱的一部分， 端粒病或短端粒综合征。外周血细胞端粒长度预测临床 IPF患者的结局，包括生存期、疾病进展率和对某些药物的反应。 FPF和IPF患者中存在端粒缩短证据的比例高于 基因突变这项研究试图使用全基因组测序来识别遗传变异， 会导致肺纤维化的遗传易感性。本申请的基本假设是， 端粒酶功能异常是肺纤维化发生发展的重要机制。 该应用程序计划评估从发现队列中获得的全基因组序列（WGS）数据， 约950例不相关的FPF先证者和IPF患者。在目标1中，我们将从WGS数据估计端粒长度， 鉴定端粒基因中罕见的编码和非编码合格变体，评估基因型-表型 关系，并研究遗传结果对患者及其医生的回报。这一目标将使 在表型良好的患者中评估已知的端粒基因。由于大部分FPF和IPF 端粒长度<第10百分位数的患者没有可识别的端粒相关致病性或可能的 致病性变异，我们将利用WGS数据和五个独立的策略来识别新的候选基因： 通过基因组位置分析变异，通过基于基因的崩溃测试分析罕见变异， 通过GWAS分析常见变异，使用滑动窗口测试分析变异，并分析拷贝数 变体。将在重复队列中评价发现队列中鉴定的候选基因和变体。 这一目标有可能发现与肺纤维化和端粒生物学相关的新基因。在目标3中， 我们将探索三种功能性调查的途径：通过评估信息性调查中的共分离， 通过体外研究患者来源的淋巴细胞和其他细胞类型中的基因功能， 通过评估CRISPR/Cas9工程小鼠疾病模型。后一个目标将侧重于 阐明PARN的功能，PARN是一种在转录后水平上具有重要作用的去腺苷酸酶， 端粒酶RNA的成熟与肺纤维化的发展有关。总的来说，这个应用程序 计划使用WGS，大型FPF和IPF队列，以及尖端的统计分析和实验 方法来扩展我们对肺纤维化和人类端粒的遗传结构的知识， 相关疾病。

ABSTRACT Unbiased genomic approaches have led to discoveries of novel disease genes and variants. Most pathogenic rare variants found in patients with Familial Pulmonary Fibrosis (FPF) and Idiopathic Pulmonary Fibrosis (IPF) result in telomere shortening. Thus, these diseases are part of the spectrum of diseases known as telomeropathies or short telomere syndromes. Telomere lengths of peripheral blood cells predict clinical outcomes of IPF patients, including survival, rate of disease progression, and response to certain medications. A larger proportion of FPF and IPF patients have evidence of telomere shortening than are explained by genetic mutations. This study seeks to use whole genome sequencing to identify genetic variants that engender an inherited susceptibility to lung fibrosis. The underlying hypothesis of this application is that telomerase dysfunction is a key mechanism underlying development of pulmonary fibrosis. This application plans to evaluate whole genome sequence (WGS) data obtained for a discovery cohort of ~950 unrelated FPF probands and IPF patients. In Aim 1, we will estimate telomere length from the WGS data, identify rare coding and noncoding qualifying variants in the telomere genes, assess genotype-phenotype relationships, and study the return of genetic results to patients and their physicians. This aim will allow for assessment of known telomere genes in well-phenotyped patients. Since a large portion of FPF and IPF patients with telomere lengths <10th percentile have no identifiable telomere-related pathogenic or likely pathogenic variant, we will utilize WGS data and five independent strategies to identify novel candidate genes: analysis of variants by genomic location, analysis of rare variants by gene-based collapsing tests, analysis of common variants by GWAS, analysis of variants using a sliding window test, and analysis of copy number variants. Candidate genes and variants identified in the discovery cohort will be evaluated in replicate cohorts. This aim has the potential to discover new genes linked to pulmonary fibrosis and telomere biology. In Aim 3, we will explore three avenues of functional investigation: through assessment of co-segregation in informative kindreds, through in vitro studies of gene function in patient-derived lymphocytes and other cell types, and through evaluation of CRISPR/Cas9-engineered mouse models of disease. The latter aim will focus on elucidating the function of PARN, a de-adenylase that has an important role in the post-transcriptional maturation of telomerase RNA, with regard to the development of pulmonary fibrosis. Overall, this application plans to use WGS, large FPF and IPF cohorts, as well as cutting-edge statistical analyses and experimental approaches to extend our knowledge of the genetic architecture of pulmonary fibrosis and human telomere- related diseases.