SURFACTANT PATHWAY GENE VARIANTS AMONG INFANTS WITH RESPIRATORY DISTRESS SYNDROME

描述(申请人提供)：新生儿呼吸窘迫综合征(RDS)是导致美国婴儿死亡和发病的最常见的呼吸系统原因。虽然RDS最常见的原因是肺表面活性物质产生的发育不成熟，但双胞胎、种族、性别和表面活性物质相关基因(SFTPB、SFTPC和ABCA3)的致命突变的研究表明了遗传机制。先前关于确定新生儿RDS的遗传机制的研究主要集中在单个候选基因中的常见遗传变异上，并且只解释了一小部分疾病的遗传性。对降低生殖适合性的复杂成人疾病的研究表明，多个基因中罕见的高渗透性变异的组合导致了疾病的遗传性。我们的实验室已经确定了表面活性物质相关基因的罕见变异与新生儿RDS的统计和功能相关性。这项建议的目标是找出过量的、罕见的功能变异，从统计上解释RDS缺失的遗传性，并开发方法来测试遗传变异扰乱表面活性物质代谢的生物机制。首先，我们将使用下一代测序来确定五个表面活性物质相关基因(SFTPC、ABCA3、LPCAT、CHPT和PCYT1B)之间基于群体的罕见遗传变异的频率，使用从密苏里州卫生部获得的1116个婴儿样本。其次，我们将确定在知情同意的情况下，与已有的DNA样本(N=940名非洲和欧洲血统的婴儿)相比，在患有RDS的婴儿中，这五个基因中多余的、罕见的、功能中断的变体是否在RDS婴儿中出现过多。第三，我们将使用基于细胞的功能分析来研究这些破坏性变体的潜在生物机制。这一综合研究策略是新颖的，因为我们将以更高的分辨率研究候选基因，使用基于软件的算法来评估变体功能，使用传统(回归)和新的(折叠方法，Bimbam，逻辑回归)统计方法来评估变体相互作用，并使用基于细胞的分析来研究已识别的变体和变体组合的功能。以细胞为基础的分析方法的发展将允许对通过统计关联确定的变异进行机械性研究，并为对受影响婴儿的诊断和治疗方法的长期发展提供基础。我的临床经验、先前的遗传流行病学教学和指导培训，以及使我能够发展使用替代细胞系统技能的综合培训计划，将使我能够发展成为一名独立的调查者，能够将统计观察与功能机制联系起来，并将这些发现转化为RDS婴儿的具体治疗方法。

DESCRIPTION (provided by applicant): Neonatal respiratory distress syndrome (RDS) is the most common respiratory cause of mortality and morbidity among infants in the United States. While RDS is most commonly attributed to a developmental immaturity of pulmonary surfactant production, a genetic mechanism is suggested by studies of twins, ethnicity, gender, and lethal mutations in surfactant-related genes (SFTPB, SFTPC, and ABCA3). Prior research to define the genetic mechanisms underlying neonatal RDS has focused on common genetic variants within single candidate genes and explains only a small proportion of disease heritability. Research in complex adult diseases that reduce reproductive fitness suggests that combinations of rare, highly penetrant variants in multiple genes account for disease heritability. Our laboratory has identified statistical and functional associations of rare variants in surfactant-related genes with neonatal RDS. The goals of this proposal are to identify gene loci with excess, rare, functional variants that statistically account for the missing heritability of RDS and to develop methods to test the biologic mechanisms by which genetic variants disrupt surfactant metabolism. First, we will use next generation sequencing to determine the population-based frequencies of rare genetic variants among five surfactant-related genes (SFTPC, ABCA3, LPCAT, CHPT, and PCYT1B) using 1,116 infant samples obtained from the Missouri Department of Health. Second, we will determine whether excess, rare, functionally disruptive variants in these five genes are overrepresented among infants with RDS compared to infants without RDS in already available DNA samples with informed consent (N=940 infants of African and European descent). Third, we will use cell-based functional assays to investigate the underlying biologic mechanisms of these disruptive variants. This comprehensive research strategy is novel because we will study candidate genes with higher resolution, use software based algorithms to assess variant functionality, use traditional (regression) and novel (Collapsing Methods, BimBam, logic regression) statistical approaches to evaluate variant interactions, and use cell-based assays to study the functionality of identified variants and combinations of variants. The development of cell-based assays will permit mechanistic study of variants identified through statistical associations and provide the basis for the long-term development of diagnostic and therapeutic approaches for affected infants. The combination of my clinical experience, prior didactic and mentored training in genetic epidemiology, and an integrated training plan that allows me to develop skills in the use of surrogate cell systems will enable me to develop into an independent investigator that can bridge statistical observation with functional mechanism and translate these findings into specific therapeutic approaches for infants with RDS.