Development of iPS Cells for Treatment of Hemoglobinopathies

描述（由申请人提供）：拟议的POI是一项多机构资助，将开发一种基于干细胞的治疗方法，用于治疗镰状细胞病（SCD）和地中海贫血（¿-thal）以及其他血红蛋白病，使用患者来源的体细胞并将其重新编程为诱导多能干细胞（IPS）细胞，这些细胞将纠正其突变并最终分化为造血干细胞（hsc），以重建患者的造血系统。开发一种有效的细胞疗法来治疗血红蛋白病，这是世界上最常见的遗传性疾病，将大大改善非洲、地中海、中东人民中常见的SCD和b -地中海贫血患者的生活质量。这一提议将验证这样一个假设，即在这种PPG的背景下，通过对患者来源的iPS细胞的生成、修饰和造血分化，可以实现对这些疾病的有效细胞和基因治疗。这将通过以下项目来完成：项目1将涉及使用phiC31整合酶介导的，序列特异性整合携带2A肽连接Oct4， Sox2， Klf4和cMyc重编程cdna的质粒，或使用小激活双链RNA （saRNA）暂时增强这些重编程基因的表达，将患者的体细胞转化为IPS细胞。项目2将涉及在存在或不存在靶向锌指核酸酶（ZFNs）或其他巨核酶的情况下，通过序列特异性修饰，使用经典同源重组（HR）或基于寡核苷酸/多核苷酸的小片段同源置换（SFHR）来纠正体细胞和iPS细胞中引起突变的疾病。项目3将涉及将未校正和校正的iPS细胞暴露在指导造血分化的条件下，以产生具有移植和重建造血系统能力的造血干细胞。在PPG的过程中，所有项目都将开发无氙气系统，以优化安全性。项目中的科学将增加一个行政（核心A）和两个科学核心：核心B：细胞和分子生物学，核心C：细胞移植和分析。

DESCRIPTION (provided by applicant): The proposed POI is a multi-institutional grant that will develop a stem cell based therapy for the treatment of sickle cell disease (SCD) and ¿-thalassemia (¿-thal) as well as other hemoglobinopathies, using patient derived somatic cells and reprogramming them into induced pluripotent stem (IPS) cells that will have their mutations corrected and ultimately differentiated into hematopoietic stem cells (HSCs) to reconstitute the patient's hematopoietic system. Development of an effective cellular therapy for the treatment of hemoglobinopathies, the most common inherited diseases worldwide, would significantly improve the quality of life of individuals afflicted with SCD and B-thalassemia that are common among the peoples of Africa, the Mediterranean, the Middle East, and Asia as well as their descendents in the U.S. This proposal will test the hypothesis that an effective cellular and genetic therapy for these diseases can be achieved in the context of this PPG through the generation, modification, and the hematopoietic differentiation of patient derived iPS cells. This will be accomplished through the following Projects: Project 1 will involve the conversion of a patient's somatic cells into IPS cells using phiC31 Integrase-mediated, sequence-specific integration of a plasmid carrying 2A peptide linked Oct4, Sox2, Klf4, and cMyc reprogramming cDNAs or by using small activating double stranded RNA (saRNA) to transiently enhance the expression of these reprogramming genes. Project 2 will involve correction of the disease causing mutations in the somatic cells and the iPS cells by sequence specific modification using either classical homologous recombination (HR) or by oligo/polynucleotide-based small fragment homologous replacement (SFHR) in the presence or absence of targeted zinc finger nucleases (ZFNs) or other meganucleases. Project 3 will involve exposure of uncorrected and corrected iPS cells to conditions to direct hematopoietic differentiation to generate HSCs which have the capacity to engraft and reconstitute the hematopoietic system. In the course of this PPG, all Projects will develop xeno-free systems to optimize safety. The science in the Projects will be augmented by an administrative (Core A) and 2 scientific Cores: Core B: Cell and Molecular Biology, and Core C: Cell Transplantation and Analysis. PUBLIC HEALTH RELEVANCE: This Program Project will focus on the development of a therapy for sickle cell anemia and B-thalassemia the most common genetic diseases worldwide. It aims to devise new methods of treating these diseases by genetically correcting patient cells to generate stems cells for transplantation, thus avoiding rejection due to histo-incompatibility. Correcting these diseases would significantly improve the quality of life among afflicted individuals and decrease the social and economic burden that they impose on the healthcare system.