Globin Gene Expression in Sickle Cell Genotype-Specific iPS cells

镰状细胞基因型特异性 iPS 细胞中的球蛋白基因表达

• 批准号: 8294700
• 负责人: DAVID H K CHUI
• 金额: $ 121.89万
• 依托单位: BOSTON UNIVERSITY MEDICAL CAMPUS
• 依托单位国家: 美国
• 财政年份: 2011
• 资助国家: 美国
• 起止时间: 2011-07-05 至 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8294700
• 关键词: Accounting; Address; Adult; Affect; African American; Animals; Blood; Blood Cells; Cell Lineage; Cells; Clinical; Development; Disease; Embryo; Erythroid; Erythroid Cells; Erythroid Progenitor Cells; Erythropoiesis; FDA approved; Fetal Development; Fetal Hemoglobin; Gene Expression; Gene Expression Profile; Generations; Genes; Genetic; Genetic Variation; Genotype; Globin; Goals; Growth; Hemoglobin; Hereditary Disease; Histone Deacetylase Inhibitor; Human; In Vitro; Learning; Messenger RNA; Modeling; Molecular Profiling; Morbidity - disease rate; Orphan Disease; Other Genetics; Patients; Pharmaceutical Preparations; Phase; Preclinical Drug Evaluation; Production; Quantitative Trait Loci; Regulation; Residual state; Sickle Cell; Sickle Cell Anemia; Sickle Hemoglobin; Staging; Stem cells; System; Technology; Transgenes; Translating; Work; base; cytotoxicity; erythroid differentiation; fetal; genome-wide; high throughput screening; hydroxyurea; induced pluripotent stem cell; mortality; novel; novel therapeutics; polymerization; pre-clinical; response; sickle erythroid; sickling; small molecule; stem; vector

DESCRIPTION (provided by applicant): Sickle cell anemia, an orphan disease of African Americans, is noted for its extensive morbidity and high mortality. Only one FDA approved drug is available for its pathophysiologically-based treatment. This agent, hydroxyurea, works through its ability to induce fetal hemoglobin (HbF) expression, which thwarts sickle hemoglobin polymerization. Not all patients respond to this treatment, so additional HbF inducing drugs are needed. Our goal is to create the technology for developing high throughput sickle cell anemia-specific induced pluripotent stem cells (iPS) and characterize their directly differentiated progeny. Using a novel excisable reprogramming vector we will generate 'clinical grade' human iPS cells free of any residual reprogramming transgenes. These directly differentiated sickle iPS cells will be used to produce an unlimited supply of erythroid-lineage cells to better understand HbF genetic regulation and perform pre-clinical small molecule drug screens. Specifically, we hypothesize: 1) that 'clinical grade' disease-specific iPS cells can be efficiently and reproducibly derived from peripheral blood cells and induced to normal erythroid differentiation, which includes the expected the spectrum of p-like globin gene expression; 2) major HbF quantitative trait loci impact globin gene expression during embryonic, fetal, and adult erythropoiesis; 3) patients with markedly elevated HbF levels serve as natural models to identify and characterize genetic variations affecting HbF production; 4) high-potency inducers of HbF, either singly or in combination can be studied in iPS-derived erythroid precursor cells. Our aims are: 1) implement an efficient and 'scalable' system for the production of sickle cell anemia-specific iPS cells and use this to recapitulate erythroid-lineage ontogeny in vitro; 2) identify developmental gene expression profile differences between erythroid precursors that produce primarily HbF and those that produce primarily HbA or HbS; 3) determine the effects of the known major HbF quantitative trait loci on globin gene expression in iPS cells; 4) search for novel HbF genetic modifiers associated with HbF levels by examining gene expression in iPS derived erythroid cells; 5) determine which novel therapeutics discovered in high throughput screens can enhance and maintain high level HBG expression in iPS-derived sickle erythroid cells. Ultimately, we hope to translate these findings into clinically efficacious treatments.

描述（由申请人提供）： 镰状细胞性贫血是非裔美国人的一种罕见疾病，以其广泛的发病率和高死亡率而闻名。只有一种 FDA 批准的药物可用于基于病理生理学的治疗。这种名为羟基脲的药物能够诱导胎儿血红蛋白 (HbF) 表达，从而阻止镰状血红蛋白聚合。并非所有患者都对这种治疗有反应，因此需要额外的 HbF 诱导药物。我们的目标是创造开发高通量镰状细胞性贫血特异性诱导多能干细胞 (iPS) 的技术，并表征其直接分化的后代。使用新型可切除重编程载体，我们将产生不含任何残留重编程转基因的“临床级”人类 iPS 细胞。这些直接分化的镰状 iPS 细胞将用于生产无限量的红系细胞，以更好地了解 HbF 基因调控并进行临床前小分子药物筛选。具体来说，我们假设：1）“临床级”疾病特异性 iPS 细胞可以有效且可重复地从外周血细胞中衍生出来，并诱导正常红细胞分化，其中包括预期的 p 样珠蛋白基因表达谱； 2) 主要HbF数量性状位点影响胚胎、胎儿和成人红细胞生成过程中珠蛋白基因的表达； 3) HbF 水平显着升高的患者作为自然模型来识别和表征影响 HbF 产生的遗传变异； 4) 可以在 iPS 衍生的红系前体细胞中研究单独或组合的 HbF 高效诱导剂。我们的目标是：1）实施高效且“可扩展”的系统来生产镰状细胞性贫血特异性 iPS 细胞，并用其在体外重现红系谱系个体发育； 2) 确定主要产生 HbF 的红系前体与主要产生 HbA 或 HbS 的红系前体之间的发育基因表达谱差异； 3)确定已知主要HbF数量性状位点对iPS细胞中珠蛋白基因表达的影响； 4) 通过检查 iPS 衍生的红系细胞中的基因表达来寻找与 HbF 水平相关的新型 HbF 遗传修饰剂； 5) 确定在高通量筛选中发现的哪些新疗法可以增强和维持 iPS 衍生的镰状红细胞中高水平的 HBG 表达。最终，我们希望将这些发现转化为临床有效的治疗方法。