Redirecting hemoglobin expression during Human ES Cell differentiation

人胚胎干细胞分化过程中血红蛋白表达的重定向

• 批准号: 7814682
• 负责人: JAMES J BIEKER
• 金额: $ 65.76万
• 依托单位: ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-04-09 至 2012-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7814682
• 关键词: Address; Adult; Anemia; Area; Blood Transfusion; Categories; Cell Differentiation process; Cell Line; Cell physiology; Cells; Chemicals; Chromatin Structure; Clinical; Communities; Cosmids; Defect; Development; Developmental Gene; Disease; Economics; Embryo; Erythrocytes; Family; Gene Silencing; Genes; Globin; Goals; Hand; Health; Hematopoiesis; Hematopoietic; Hemoglobin; Hemoglobinopathies; Human; In Vitro; Lead; Leukemic Cell; Life; Life Expectancy; Modality; Molecular; Morbidity - disease rate; Mus; National Institute of Diabetes and Digestive and Kidney Diseases; Patients; Pattern; Process; Productivity; Quality of life; Regulator Genes; Reporter; Research Design; Sickle Cell Anemia; Small Molecule Chemical Library; Source; Stem cells; Switch Genes; Symptoms; System; Testing; Thalassemia; Therapeutic; Tissues; Transgenic Mice; cellular engineering; derepression; design; economic impact; embryonic stem cell; fetal; fetal globin; genetic analysis; human embryonic stem cell; human embryonic stem cell line; induced pluripotent stem cell; insight; interest; mutant; novel strategies; patient population; polymerization; promoter; prophylactic; public health relevance; sickling; small molecule; stem; stem cell differentiation; success; transcription factor

DESCRIPTION (provided by applicant): This application addresses broad Challenge Area (14) Stem Cells, and specific Challenge Topic 14-DK-104: In vitro differentiation of human Embryonic Stem Cells (ES)/Induced Pluripotent Stem Cells (iPS) to NIDDK relevant cells/tissues. Hemoglobinapathies, such as sickle cell disease and ¿-thalassemia, are anemias of varying intensity that continue to have a deleterious impact on the health of specific subpopulations in the US workforce, ultimately exerting a negative effect on our economic productivity. We have an interest in developing novel approaches to alter the normal hemoglobin switching mechanism so that patients with hemoglobinapathies might benefit from derepression of fetal or embryonic globin expression. Briefly, the idea described in the present proposal involves using human embryonic stem (ES) cells, engineered to express detectable reporters from their fetal and adult globin promoters, to screen for small molecules that lead to more efficient induction of adult globin, or reactivation of fetal globin, expression. The major advantages of this approach is that it avoids the historical use of the mouse to address what is a human gene regulatory pattern, and that it takes advantage of the normal hematopoietic process observed during ES cell differentiation, a process that cannot be recapitulated within human leukemic cell lines. As a result, this application proposes to define and utilize a novel approach to alter the normal hemoglobin switching mechanism via three interrelated aims: 1) establish efficient hES cell differentiation conditions that correctly recapitulate the normal ¿-like globin gene developmental sequence; 2) generate a correctly regulated reporter hES cell line that mimics this endogenous expression pattern; 3) use this line to screen a small molecule library for chemicals that alter/redirect ¿-like globin expression. A key aspect of this design is that the chemical screen in Aim 3, and thus the ultimate success of the proposal, is not solely dependent on establishment of adult ¿-producing cells in Aim 1. This fits well into the 14-DK-104 category, as it aims to identify small molecules that redirect hemoglobin expression in differentiating hES cells and is consistent with programmatic interests of the NIDDK. PUBLIC HEALTH RELEVANCE: The goals of this project are to establish a molecular and cellular baseline of marked ¿.like globin gene switching in human embryonic stem cells that will then provide a rigorous and directly relevant avenue for testing inducers of adult globin or reactivators of fetal globin expression.

描述(由申请人提供)：本申请涉及广泛的挑战领域(14)干细胞，以及特定的挑战主题14-DK-104：人类胚胎干细胞(ES)/诱导多能干细胞(IPS)向NIDDK相关细胞/组织的体外分化。血红蛋白疾病，如镰状细胞病和地中海贫血，是不同程度的贫血，继续对美国劳动力中特定亚群的健康产生有害影响，最终对我们的经济生产力产生负面影响。我们有兴趣开发新的方法来改变正常的血红蛋白转换机制，从而使患有血红蛋白疾病的患者可能受益于取消抑制胎儿或胚胎的珠蛋白表达。简而言之，本提案中描述的想法涉及使用人类胚胎干细胞(ES细胞)来筛选能够更有效地诱导成人珠蛋白表达或重新激活胎儿珠蛋白表达的小分子。人类胚胎干细胞经过改造，可以表达来自胎儿和成人的珠蛋白启动子。这种方法的主要优点是，它避免了历史上使用小鼠来解决什么是人类的基因调控模式，并且它利用了在ES细胞分化过程中观察到的正常造血过程，这一过程不能在人类白血病细胞系中重演。因此，本申请建议定义和利用一种新的方法来通过三个相互关联的目标来改变正常的血红蛋白交换机制：1)建立有效的HES细胞分化条件，正确地概括正常的类珠蛋白基因的发育序列；2)产生一个正确调控的报告HES细胞系，模拟这种内源性表达模式；3)使用该系来筛选小分子文库，以寻找改变/重定向类珠蛋白表达的化学物质。这一设计的一个关键方面是，目标3中的化学筛选，以及该提案的最终成功，并不完全依赖于在目标1中建立成体产生细胞。这非常适合14-DK-104类别，因为它旨在识别在分化的HES细胞中重新定向血红蛋白表达的小分子，并与NIDDK的计划目标一致。 公共卫生相关性：该项目的目标是建立人类胚胎干细胞中显著的类珠蛋白基因切换的分子和细胞基线，然后为测试成人珠蛋白的诱导剂或胎儿珠蛋白表达的再激活剂提供严格和直接相关的途径。