Lipidomic Screening For Functional Surfactant Gene Mutations

功能性表面活性剂基因突变的脂质组学筛选

• 批准号: 9021312
• 负责人: Francis Sessions Cole
• 金额: $ 45.57万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-03-01 至 2019-02-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9021312
• 关键词: 1 year old; ABCA3 gene; Adenovirus Vector; Alveolar; Animals; Award; Binding; Biological Models; Cause of Death; Cell Culture System; Cell Culture Techniques; Cells; Complementary DNA; Computer Simulation; Defect; Development; Diagnostic; Electron Microscopy; Electrospray Ionization; Epithelial; Epitopes; Family; Fetal Lung; Frequencies; Gene Mutation; Gene Silencing; Genes; Genetic; Genomics; Goals; Health; Heritability; Human; Individual; Infant; Kinetics; Label; Link; Lipids; Mass Fragmentography; Mass Spectrum Analysis; Measurement; Mediating; Methods; Modeling; Morbidity - disease rate; Mutation; Phase; Phenotype; Phospholipids; Premature Infant; Property; RNA; Regulation; Resistance; Risk; Sensitivity and Specificity; Site-Directed Mutagenesis; Surface; Surface Tension; System; Testing; Treatment Failure; United States; Variant; abstracting; alveolar lamellar body; base; burden of illness; disorder risk; endoplasmic reticulum stress; fetal; gender disparity; high throughput screening; improved; innovation; lipid transport; member; mutation screening; neonatal respiratory distress; precursor cell; racial disparity; reconstitution; respiratory; respiratory distress syndrome; screening; small hairpin RNA; small molecule; species difference; surfactant; surfactant deficiency; surfactant function; surfactant production; surfactant replacement; tandem mass spectrometry; tissue culture; trafficking; transduction efficiency; treatment strategy

DESCRIPTION (provided by applicant): Genetic regulation of surfactant deficiency has been suggested by heritability of neonatal respiratory distress syndrome, persistence of gender and racial disparities in disease risk, and the frequency of surfactant replacement treatment failures. Individually rare, collectively common (2-4/100 infants) mutations in the ATP- binding cassette sub-family A member 3 gene (ABCA3) disrupt surfactant function through diverse mechanisms including reduced lipid transport, ABCA3 misfolding or altered trafficking, or induction of increased endoplasmic reticulum stress and increase risk for neonatal respiratory distress syndrome in term and late preterm infants. Our goal is to develop and implement a human model system that uses static and dynamic lipidomic signatures for functional screening of ABCA3 mutations and that could be used for testing small molecules to correct mutation-encoded, functional defects in any gene expressed in the human alveolar type 2 cell. In the R21 Phase of this Award, using adenoviral vectors with high transduction efficiency and cargo capacity and highly sensitive and specific mass spectrometry based lipidomic profiling, we will test the hypothesis that ABCA3 gene silencing and rescue with wild-type ABCA3 cDNA reconstitute surfactant phospholipid signatures in human, primary alveolar type 2 cells. In the R33 Phase of this Award, using mass spectrometry-based lipidomic profiling, electron microscopy, and surface activity measurements, we will examine disruption of surfactant lipidomic signatures, lipid turnover rates, and lipid secretion kinetics, lamellar body phenotype, and surfactant function by previously characterized and uncharacterized ABCA3 mutations associated with increased risk for neonatal respiratory distress syndrome. The overall impact of this Award will provide a human model system for functional, lipidomics-based screening of genomic hits associated with surfactant deficiency and for development of small- molecule based strategies to correct mutation-encoded, functional surfactant defects.

描述(由申请人提供)：新生儿呼吸窘迫综合征的遗传性、疾病风险的性别和种族差异以及表面活性物质替代治疗失败的频率表明表面活性物质缺乏的遗传调节。 个别罕见、集体常见(2-4/100名婴儿)的ATP结合盒A亚家族成员3基因(ABCA3)突变通过多种机制扰乱表面活性物质的功能，包括脂质转运减少，ABCA3错误折叠或改变转运，或诱导内质网应激增加，增加足月儿和早产儿新生儿呼吸窘迫综合征的风险。我们的目标是开发和实现一种人体模型系统，该系统使用静态和动态脂组特征进行ABCA3突变的功能筛选，并可用于测试小分子以纠正在人类肺泡2型细胞中表达的任何基因中的突变编码的功能缺陷。在该奖项的R21阶段，我们将使用具有高转导效率和运载能力的腺病毒载体以及基于高灵敏和特定质谱学的脂质体图谱，验证ABCA3基因沉默和野生型ABCA3基因拯救在人原代肺泡2型细胞中重建表面活性磷脂特征的假设。在该奖项的R33阶段，我们将使用基于质谱学的脂体图谱、电子显微镜和表面活性测量，检查表面活性物质脂体特征、脂类周转率和脂类分泌动力学、板层体表型和表面活性物质功能的破坏，这些突变与新生儿呼吸窘迫综合征的风险增加有关。该奖项的总体影响将提供一个人体模型系统，用于基于脂类组学的功能性筛选与表面活性物质缺乏相关的基因组点击，并用于开发基于小分子的策略来纠正突变编码的功能性表面活性物质缺陷。