GENETICS VARIATION IN 5LO PRODUCT PRODUCTION

5LO 产品生产中的遗传变异

• 批准号: 2897325
• 负责人: Jeffrey Mark Drazen
• 金额: $ 21.68万
• 依托单位: BRIGHAM AND WOMEN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-07-01 至 2004-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/2897325
• 关键词: asthma; blood chemistry; clinical research; diagnostic respiratory lavage; enzyme inhibitors; fatty acid biosynthesis; gene deletion mutation; gene expression; gene mutation; genetic mapping; genetic promoter element; genetic susceptibility; genetic transcription; human subject; leukotrienes; lipoxygenase; nucleic acid sequence; respiratory airflow disorder

Asthma is a heterogenous syndrome with multiple potential pathways leading to the airway obstruction which is its hallmark. Among the substances which have an established role as mediators of the airway obstruction in asthma are the leukotrienes. The leukotrienes are the products of the catalytic action of a number of enzymes among which is 5-Lipoxygenase (5-LO). Given the perceived heterogeneity of the effector mechanisms in asthma and the established importance of the cysteinyl leukotrienes as one of the families of effector molecules in asthma, an understanding of the factors responsible for variations among individuals in the regulation of leukotriene synthesis has applications to both basic and clinical science. This proposal addresses one of the potential sites of regulation of 5-LO product production, i.e., regulation due to the presence of genetic mutations in the 5-LO core promoter. We have discovered a series of mutations in which there is the addition of or the deletion of Sp1/Egr-1 binding motifs (i.e.,- GGGCGG-) in the 5-LO core promoter; these mutations modify gene transcription in reporter gene constructs. Our preliminary data, reviewed herein, indicate that patients with asthma harboring the mutant forms of the core promoter have a diminished salutary therapeutic response to 5-LO inhibition. From this observation we have indirectly asserted that the lesser therapeutic response to 5- LO inhibition reflects decreased synthesis of 5-LO products by patients who are homozygous for mutant forms of the 5-LO core promoter. The goal of the proposed research is to determine if this assertion, based on analysis of the results of a large clinical trial, is true at the level of 5-LO product formation in individual patients. To test this hypothesis we will pursue two specific aims. In the first aim we will identify cohorts of patients, with atopic asthma of equivalent severity, who harbor either the wild-type or the mutant 5-LO core promoter. These subjects will be used to ascertain the quantitative production of LTE4, cysteinyl leukotrienes and lipoxins from basal and A23 187 challenged neutrophils and eosinophils isolated from the peripheral blood, or from alveolar macrophages isolated by bronchoalveolar lavage. 5-LO product production will also be studied in each of the genotypically defined cohorts of asthma patients by measurement of urinary LTE4 before and after inhaled whole lung allergen stimulation. In addition to whole lung allergen challenge, patients will undergo pulmonary segmental allergen challenge and the recovery of 5-LO products from the challenged versus the control segment compared. If the hypothesis is correct, then patients harboring the mutant forms of the core promoter should have diminished 5-LO product production when compared to patients with the wild type form of the core promoter. Furthermore, after allergen challenge (both whole lung and segmental), the yield of 5-LO products should be less in patients with the mutant forms of the core promoter than in patients with the wild type form of the core promoter.

哮喘是一种异质综合征，具有多种潜在途径，导致气道阻塞是其标志。在哮喘中，作为气道阻塞的介体具有确定作用的物质是白细胞。白细胞是许多酶的催化作用的产物，其中5-脂氧酶（5-lo）。 鉴于哮喘中效应机制的异质性以及白细胞三烯作为哮喘中效应分子家族的确定重要性，因此对负责白细胞合成调节的个体变异因素的理解适用于基础和临床科学。 该建议介绍了5-LO产品生产的调节的潜在位点之一，即由于5-LO核心启动子中存在基因突变而导致的调节。我们发现了一系列突变，其中添加或缺失了5-lo核心启动子中SP1/EGR-1结合基序（即 - GGGCGG-）；这些突变改变了报告基因构建体中的基因转录。我们的初步数据在本文中进行了综述，表明携带核心启动子突变形式的哮喘患者对5-LO抑制作用的有益治疗反应减少。从这一观察结果中，我们间接地断言，对5-LO抑制的治疗反应较小，反映出对5-lo核心启动子突变形式纯合的患者对5-LO产物的合成降低。拟议的研究的目的是确定基于对大型临床试验结果的分析，在个别患者的5-LO产品形成水平上，是否基于对大型临床试验结果的分析。为了检验这一假设，我们将追求两个具体的目标。在第一个目标中，我们将确定患者的同类群体，以及具有同等严重程度的特应性哮喘，他们携带野生型或突变体5-LO核心启动子。这些受试者将用于确定LTE4的定量产生，基底和A23 187的LTE4，白细胞藻和脂氧蛋白挑战了从外周血中分离出的中性粒细胞和嗜酸性粒细胞，或者是从通过Bronchoalchoalchoalvealvealvealvealvealvealvealvealveaveage分离出的肺泡巨噬细胞。还将通过测量吸入整个肺部过敏原刺激前后的尿液LTE4来研究5-LO产品的产生。 除了整个肺过敏原挑战外，患者还将受到肺部节段过敏原挑战，并从受到挑战的对照段与对照段相比，从挑战性段中恢复5-LO产物。如果该假设正确，则与具有核心启动子的野生型形式的患者相比，具有核心启动子突变形式的突变形式的患者应减少5-LO产品的产生。此外，在过敏原挑战（整个肺和分段）之后，与核心启动子的野生型形式的患者相比，具有核心启动子突变形式的5-LO产物的产率应较少。