Application of high-throughput approaches in the study of complex disorders

高通量方法在复杂疾病研究中的应用

• 批准号: 7594270
• 负责人: Owen M Rennert
• 金额: $ 24.26万
• 依托单位: EUNICE KENNEDY SHRIVER NATIONAL INSTITUTE OF CHILD HEALTH & HUMAN DEVELOPMENT
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7594270
• 关键词: African American; Age related macular degeneration; Age-Years; Angiotensins; Antithrombin III; Apolipoprotein E; Asians; Biological Assay; Biomedical Research; Blindness; CX3CL1 gene; Caucasians; Caucasoid Race; Complex; Developed Countries; Developing Countries; Development; Diagnosis; Disease; Enzymes; Factor V; Fibrinogen; Genes; Genetic Polymorphism; Genetic screening method; Goals; Hispanics; Hospitalization; Incidence; Individual; Inherited; Institutes; Legal patent; Methods; Methylenetetrahydrofolate reductase (NADPH); Morbidity - disease rate; Mutation; Numbers; Oligonucleotide Microarrays; Oligonucleotides; Pacific Island Americans; Pathogenesis; Plasminogen Activator Inhibitor 1; Polymerase Chain Reaction; Population; Predictive Value; Predictive Value of Tests; Predisposition; Process; Prophylactic treatment; Protein C; Prothrombin; Protocols documentation; Reaction; Recurrence; Reporting; Risk; Risk Factors; Screening procedure; Spottings; Stratification; TLR4 gene; Technology; Testing; Thrombophilia; United States; Variant; Venous Thrombosis; aryldialkylphosphatase; base; citrate carrier; density; design; fibulin; human CX3CR1 protein; mortality

Venous thrombosis (VT) is one of the leading causes of mortality and morbidity resulting in approximately 300,000 hospitalizations and 50,000 fatalities per year in the United States with an incidence of 141 per 100 000 African-Americans (1.4 per 1000), 104 per 100 000 Caucasians (1 per 1000), 55/100 000 in Hispanics (0.6per 1000) and 21 per 100 000 Asian/Pacific Islanders (0.2 per 1000). It is, however, an avoidable disease if currently available prophylactic treatment is instituted. Our calculations demonstrated that concurrent use of a panel of 11 genetic tests increases the positive predictive value of testing for venous thrombosis at least 30-fold. We have devised an approach (Method Evolved for Recognition of Thrombophilia MERT, patent pending) that will allow prediction and accurate assessment of hereditary thrombophilia in several ethnic populations by rapid, concurrent screening of an array of all known 145 venous thrombosis-associated recurrent mutations and polymorphisms in nine molecules antithrombin III (AT III), protein C, protein S, fibrinogen, factor V (FV), prothrombin (factor II), methylenetetrahydrofolate reductase (MTHFR), angiotensin 1-converting enzyme (ACE) and plasminogen activator inhibitor-1 (PAI-1) genes . MERT will help us develop stratification protocols for risk-adapted prophylaxis. We have designed 291 oligonucleotide 25mer probes to be spotted onto the microarray and achieved to amplify 40 amplicons covering the variation sequences from nine different genes in a single amplification reaction by Multiplex PCR assay. We are now in the process of verifying the analytic validity of our method. We applied a similar approach to the design of an oligo-array for screening for susceptibility to development of age-related macular degeneration (Method Evolved for Recognition and Testing of Age-Related Macular Degeneration-MERT-ARMD, patent pending). Age-related macular degeneration (ARMD) is the most common cause of severe vision loss in the United States and developed countries among people 65 years of age and older. It has been suggested that ARMD is a multifactorial disorder. Our previous reports described screening for one or more polymorphisms associated with ARMD. In general, the use of these assays is limited because they have a low predictive value and detect mutations prevalent only in Caucasian populations. We have designed a MERT-ARMD that will concurrently screen 105 known age-related macular degeneration-associated mutations and polymorphisms in 16 molecules (CFH, LOC387715, BF, C2, ABCR, Fibulin 5, VMD2, TLR4, CX3CR1, CST3, MnSOD, MEHE, paraoxonase, APOE, ELOVL4 and hemicentin-1 genes), using hybridization-based, high-density oligonucleotide array technology.

Venous thrombosis (VT) is one of the leading causes of mortality and morbidity resulting in approximately 300,000 hospitalizations and 50,000 fatalities per year in the United States with an incidence of 141 per 100 000 African-Americans (1.4 per 1000), 104 per 100 000 Caucasians (1 per 1000), 55/100 000 in Hispanics (0.6per 1000) and 21 per 100 000 Asian/Pacific岛民（每1000份0.2）。但是，如果目前进行了预防性治疗，则可以避免疾病。我们的计算表明，同时使用一组11个基因检测会增加静脉血栓形成的阳性预测值，至少30倍。我们已经设计了一种方法（进化用于识别血栓形成梅尔特，专利申请的方法），该方法将通过快速，同时筛选所有已知的145个静脉血栓形成相关的复发性突变和多态性（nine Molecyles III），以预测和准确评估几个族裔人群中的遗传性血栓形成，并同时筛选一系列已知的145个静脉血栓形成相关的突变和多态性。 S，纤维蛋白原，因子V（FV），凝结凝结蛋白（因子II），甲基苯甲酸叶酸酸盐还原酶（MTHFR），血管紧张素1-转化酶（ACE）和纤溶酶原激活剂抑制剂-1（PAI-1）基因。 MERT将帮助我们制定针对风险适应性预防的分层方案。我们已经设计了291个寡核苷酸25mer探针，可以在微阵列上发现并实现了40个扩增子，涵盖了通过多重PCR分析的单个扩增反应中9个不同基因的变异序列。现在，我们正在验证方法的分析有效性。 我们采用了类似的方法来设计寡阵阵列，以筛选与年龄相关的黄斑变性的敏感性（方法是为了识别和测试与年龄相关的黄斑变性臂识别和测试，申请专利）。与年龄相关的黄斑变性（ARMD）是美国和发达国家在65岁及以上的人群中造成严重视力丧失的最常见原因。有人建议ARMD是一种多因素疾病。我们以前的报告描述了与ARMD相关的一种或多种多态性的筛查。通常，这些测定法的使用受到限制，因为它们的预测价值低，并且仅在高加索人群中检测突变。我们设计了一种MERT-ARMD，该ARMD将同时筛选16个分子中与年龄相关的黄斑变性相关的突变和多态性（CFH，LOC387715，BF，C2，C2，ABCR，Fibulin，Fibulin 5，vmd2，vmd2，tlr4，tlr4，tlr4，cx3cr1，cx3cr1，cst3，mnssoson，mnsson，para，para and para and para and para and para，半开蛋白-1基因），使用基于杂交的高密度寡核苷酸阵列技术。