ASSAY FOR MUTATIONS IN STEM CELLS AND THEIR DERIVATIVES

描述（由申请人提供）：基于干细胞的疗法在治疗许多衰弱性疾病和病症方面具有很大的前景。然而，这种方法的成功依赖于维持这些细胞的遗传完整性，在诱导分化之前和之后，在体外产生特异性的、功能性的细胞类型，和/或移植分化的衍生物来减轻患者体内的缺陷。目前，评估干细胞或其衍生物遗传完整性的主要方法是核型分析。该方法检测染色体数目的异常或染色体结构的严重缺陷，但不能检测点突变，包括碱基替换、小缺失或小插入，这些突变是导致细胞功能异常（包括肿瘤发生）的很大一部分遗传畸变的原因。该提案将支持基于转基因的测定方法的开发和验证，以确定人类和非人类灵长类干细胞及其衍生物中点突变的频率和频谱。该分析将促进高度相关的遗传完整性评估，在各种不同干细胞治疗方法的开发过程中提供急需的质量控制和安全保证水平。随着这项工作进入第二阶段，多种潜在的产品和工具将从独特的非人类灵长类动物恒河猴和狒狒细胞以及新的和现有的人类细胞系中衍生出来。

DESCRIPTION (provided by applicant): Stem cell-based therapies hold great promise for the treatment of numerous debilitating diseases and conditions. However, the success of this approach is dependent upon maintenance of genetic integrity in these cells, both before and after induction of differentiation to yield specific, functional cell types in vitro, and/or transplantation of differentiated derivatives to mitigate defects in patients in vivo. Currently, the primary method available to assess the genetic integrity of stem cells or their derivatives is karyotyping. This method detects abnormalities in chromosome number or gross defects in chromosomal structure, but does not detect point mutations, including base substitutions, small deletions or small insertions, that are responsible for a significant proportion of genetic aberrations leading to abnormal cellular function, including tumorigenesis. This proposal will support the development and validation of a transgene-based assay to determine the frequency and spectrum of point mutations in human and nonhuman primate stem cells and their derivatives. This assay will facilitate a highly relevant assessment of genetic integrity that will provide a critically needed level of quality control and safety assurance during the development of a variety of different stem cell-based therapeutic approaches. As the work evolves into Phase II, multiple potential products and tools will be derived from unique non-human primate rhesus and baboon cells and from new and existing human cell lines.