Tools to Study Mammalian Mutagenesis

描述（由申请人提供）： 尽管正在进行广泛的工作，以了解癌症和化学品和其他试剂的致癌特性，我们仍然有能力有效地识别致癌物，并阐明其作用模式的差距。设计用于检查突变和致癌机制的测定的当前状态尤其受到其高成本和低通量能力的限制。该实验室开发了一种基于内源性Pig-a基因的基因突变检测。Pig-a基因产物对于糖基磷脂酰肌醇（GPI）锚的生物合成是必需的。产生无功能GPI锚的突变阻止某些蛋白质在细胞表面上表达，这代表了可以通过流式细胞术测量的表型。本文提出的工作将通过创建一套代表研究致癌作用的关键模式突变的平台的测定来扩展我们的开发工作。通过设计基于Pig-a的方法来检查培养物、实验室啮齿动物和人类受试者中的细胞，将实现真正全面的桥接生物标志物。所提出的突变评估工具的桥接能力和高通量性质将对包括学术、监管和工业环境在内的重要研究领域做出重大贡献。

DESCRIPTION (provided by applicant): Despite the extensive work being performed to understand cancer and carcinogenic properties of chemicals and other agents, there are still gaps in our ability to efficiently identify carcinogens, and elucidate their mode(s) of action. The current state of assays designed to examine mutation and carcinogenic mechanisms are especially limited by their high costs and low throughput capacities. This laboratory has developed a gene mutation assay that is based on the endogenous Pig-a gene. The Pig-a gene product is essential for the biosynthesis of glycosyl phosphatidylinositol (GPI) anchors. Mutations giving rise to nonfunctional GPI anchors prevent certain proteins from being expressed on the cell surface, and this represents a phenotype that can be measured by flow cytometry. The work proposed herein will extend our development efforts by creating a suite of assays that represent a platform for studying a key mode of carcinogenic action, mutation. By devising Pig-a based methods that examine cells in culture, laboratory rodents, and human subjects, a truly comprehensive bridging biomarker will be realized. The bridging capabilities and high throughput nature of the proposed mutation assessment tools will contribute significantly to important areas of research that include academic, regulatory, and industrial settings.