Derivation and Correction of Thalassemic Pluripotent Stem Cells

地中海贫血多能干细胞的衍生和校正

• 批准号: 7799411
• 负责人: David W Russell
• 金额: $ 45.09万
• 依托单位: UNIVERSITY OF WASHINGTON
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-09-01 至 2014-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7799411
• 关键词: Adult; Autologous Transplantation; Biological Assay; CD34 gene; Cell Line; Cells; Clinical; Coculture Techniques; DNA Methylation; Deoxyribonucleases; Dependovirus; Derivation procedure; Development; Disease; Engraftment; Epigenetic Process; Erythroid; Erythroid Cells; Excision; Fibroblasts; Funding; Gene Delivery; Gene Targeting; Genes; Genetic Recombination; Globin; Grant; Hematopoietic; Hematopoietic stem cells; Hemoglobin F Disease; Hereditary Disease; Human; Hypersensitivity; Immunodeficient Mouse; In Vitro; Integrase; Kanamycin Kinase; Lentivirus Vector; Long Terminal Repeats; Mediating; Mesenchymal; Methods; Monitor; Mus; Mutation; Patients; Phenotype; Pluripotent Stem Cells; Point Mutation; Production; Proteins; Recovery; Research; Retroviral Vector; Site; Source; Spumavirus; Stem cells; Stromal Cells; Subfamily lentivirinae; Surface; System; Testing; Thalassemia; Therapeutic; Thymidine Kinase; Transgenes; Transplantation; adeno-associated viral vector; arm; base; beta Globin; blastomere structure; cell type; design; disease-causing mutation; erythroid differentiation; fetal; fusion gene; gene correction; gene therapy; genome-wide; genotoxicity; histone modification; homologous recombination; human embryonic stem cell; improved; induced pluripotent stem cell; mutant; pluripotency; recombinase; research study; small hairpin RNA; vector

Human induced pluripotent stem cells (iPSCs) have the potential to treat many diseases by autologous transplantation. However, before they can be used clinically, efficient and reproducible methods are required for their derivation and differentiation into therapeutic cell types. In the case of genetic diseases, methods for correcting disease-causing mutations also need to be developed. Here we will use this approach to design a treatment for thalassemia, which is caused by mutations in globin genes. This approach avoids the potential genotoxic complications of conventional gene therapy with retroviral vectors and should achieve consistent, regulated globin expression from the endogenous locus. We will derive IPSCs from the adult cells of patients with thalassemia using lentivirus and foamy virus vectors that express reprogramming transgenes. Some of the vectors will be designed for transient gene delivery to create transgene-free iPSCs. These IPSCs will be differentiated into hematopoietic cells to determine the best adult cell type and reprogramming vectors for creating iPSCs with hematopoietic potential. Adeno-associated virus gene targeting vectors will be used to correct the globin mutations in these thalassemic IPSCs, and globin expression will be studied after their differentiation into erythroid progeny. This research plan capitalizes on recent advances in the derivation of patient-specific stem cells, which in combination with gene correction constitutes a new paradigm for the treatment of genetic diseases.

人诱导多能干细胞（iPSC）具有通过自体移植治疗多种疾病的潜力。 移植然而，在它们可以用于临床之前，需要高效和可重复的方法 用于它们的衍生和分化成治疗性细胞类型。在遗传性疾病的情况下， 还需要开发纠正致病突变的方法。在这里，我们将使用这种方法来设计一个 地中海贫血的治疗，这是由珠蛋白基因突变引起的。这种方法避免了潜在的 基因毒性并发症的常规基因治疗与逆转录病毒载体， 从内源基因座调节珠蛋白表达。我们将从患者的成体细胞中获得IPSC 用慢病毒和泡沫病毒载体表达重编程转基因治疗地中海贫血。一些 这些载体将被设计用于瞬时基因递送以产生无转基因的iPSC。这些IPSC将 分化成造血细胞，以确定最佳的成人细胞类型和重编程载体， 产生具有造血潜能的iPSC。腺相关病毒基因靶向载体将用于 纠正这些地中海贫血IPSC中的珠蛋白突变，并在其发生后研究珠蛋白表达。 分化成红系后代。这项研究计划利用了最近的进展，在推导 患者特异性干细胞，与基因校正相结合，构成了一个新的范例， 治疗遗传性疾病。