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Primate iPS lines using Retrovirus and TAT Protein Transduction

使用逆转录病毒和 TAT 蛋白转导的灵长类 iPS 系

基本信息

批准号：
7799770
负责人：
COLIN Edward BISHOP
金额：
$ 18.32万
依托单位：
WAKE FOREST UNIVERSITY HEALTH SCIENCES
依托单位国家：
美国
项目类别：
财政年份：
2009
资助国家：
美国
起止时间：
2009-05-01 至 2012-04-30
项目状态：
已结题

项目摘要

DESCRIPTION (provided by applicant): The overriding hypothesis of this application is that the generation and characterization of non Human primate (NHP) induced pluripotent stem cell lines (iPS) will hasten the development of replacement tissues and organs in Humans. Macaque monkeys share an extremely close phylogenetic relationship with Man, and develop natural or experimentally induced disease states which accurately mimic the Human condition. NHPs can provide a unique preclinical model to test the usefulness of iPS cells and regenerative medicine approaches to the treatment and cure of disease, where other models may fail. The ability to generate autologous, pluripotent stem cells from skin fibroblasts, by relatively simple reprogramming steps, is a quantum scientific advance which will have major ramifications in many areas of basic research and applied clinical medicine. It is highly unlikely, however, that the use of viral vectors in the reprogramming protocol (as it exists now) will be acceptable for use in patients. It is vital, therefore, that alternative ways of reprogramming, without virus, be investigated. The short 11 amino acid protein transduction domain (PTD) derived from the HIV transactivator, TAT has been used to deliver large (~110kD), biologically active proteins directly into cells in vitro and in vivo. TAT fusion proteins and peptides have also been used to treat mouse models of cancer, inflammation and other diseases. It is our hypothesis that recombinant TAT and be able to reprogram cells into a pluripotent state. Once well characterized, lines derived by viral or protein transduction methods can be used in NHP preclinical models for a number of different diseases. We therefore propose to: (1) generate induced pluripotent stem cell lines (iPS) from the non-human primate, Macaca fascicularis (cynomolgus macaque) (2) To determine if M. fascicularis iPS cells can be produced non-virally using recombinant proteins modified with an HIV derived, TAT cell penetrating motif to aid in protein translocation across the plasma membrane and (3) characterize and compare iPS lines generated by viral and protein transduction.

描述（由申请人提供）：本申请的首要假设是非人灵长类动物（NHP）诱导的多能干细胞系（iPS）的产生和表征将加速人类替代组织和器官的发育。猕猴与人类有着极其密切的系统发育关系，并发展出自然或实验诱导的疾病状态，这些疾病状态准确地模拟了人类的状况。NHP可以提供一个独特的临床前模型来测试iPS细胞和再生医学方法在治疗和治愈疾病方面的有用性，而其他模型可能会失败。通过相对简单的重编程步骤，从皮肤成纤维细胞产生自体多能干细胞的能力是一个巨大的科学进步，将在基础研究和应用临床医学的许多领域产生重大影响。然而，在重编程方案中使用病毒载体（如现在所存在的）对于患者来说是非常不可能的。因此，至关重要的是，研究不使用病毒的重编程替代方法。来自HIV反式激活因子达特的短的11个氨基酸的蛋白转导结构域（PTD）已被用于在体外和体内将大的（~ 110 kD）生物活性蛋白直接递送到细胞中。达特融合蛋白和肽也已用于治疗癌症、炎症和其他疾病的小鼠模型。我们假设重组达特能够将细胞重编程为多能状态。一旦得到很好的表征，通过病毒或蛋白转导方法获得的细胞系可用于许多不同疾病的NHP临床前模型。因此，我们建议：（1）从非人灵长类动物食蟹猴（Macaca fascicularis）（cynomolgus macaque）产生诱导的多能干细胞系（iPS）。可以使用用HIV衍生的达特细胞穿透基序修饰的重组蛋白质非病毒地产生fascicularis iPS细胞，以帮助蛋白质跨质膜易位，和（3）表征和比较通过病毒和蛋白质转导产生的iPS系。