Assays for ROS-induced DNA damage

ROS 诱导的 DNA 损伤测定

• 批准号: 6550539
• 负责人: HAROLD C BOX
• 金额: $ 25万
• 依托单位: ZEPTOMETRIX CORPORATION
• 依托单位国家: 美国
• 财政年份: 1999
• 资助国家: 美国
• 起止时间: 1999-02-08 至 2004-07-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6550539
• 关键词: DNA damage; DNA repair; bioassay; chemical synthesis; formamide; free radical oxygen; guanine; high performance liquid chromatography; liquid chromatography mass spectrometry; nuclease; stable isotope; technology /technique development; thin layer chromatography; thymine

DESCRIPTION (provided by applicant):This project is a collaboration between ZeptoMetrix Corporation and Roswell Park Cancer Institute. The objective is to develop sensitive, quantitative and facile assays for measuring DNA damage caused by reactive oxygen species (ROS). ROS have been implicated in aging, cancer and several neurological diseases. Seven ROS-induced DNA lesions will be assayed, namely: the formamido remnant of pyrimidine bases, the 8-oxo-7, 9-dihydroguanine (8-oxo-guanine) lesion, the thymine glycol lesion, the 6-hydroxy-5,6-dihydrothymine lesion, the abasic lesion and two double base lesions. 32P-postlabeling or mass spectroscopic methods of detection are employed with the assays. ZeptoMetrix will make products commercially available that facilitate DNA measurements by either method. Products will include reference compounds, carriers and internal standards for assaying the aforementioned lesions. Carriers for 32P-postlabeling provide unequivocal identification of the DNA lesion of interest. Internal standards for MS measurements will be isotopically enriched to a higher mass than the DNA lesion to be quantitated. ZeptoMetrix anticipates that eventually the largest application of these products will be for evaluating the repair capabilities of cells and tissues. Products useful for repair studies will include reference compounds and duplex DNA substrates containing identified lesions. Liquid Chromatography -Mass Spectrometry (LC-MS) utilizing an electrospray interface will be developed with this application especially in mind. PROPOSED COMMERCIAL APPLICATION: Reference materials, internal standards, and carriers for measuring oxidative DNA damage will be synthesized. These products will be offered commercially for facilitating DNA damage and repair by 32P-postlabeling or by LC-MS.

描述(申请人提供)：该项目是ZeptoMetrix公司和罗斯威尔公园癌症研究所的合作项目。目的是建立一种灵敏、定量和简便的检测活性氧自由基(ROS)引起的DNA损伤的方法。ROS与衰老、癌症和几种神经疾病有关。将检测7种ROS诱导的DNA损伤，即：嘧啶碱基甲酰胺残留物、8-氧代-7，9-二氢鸟嘌呤(8-氧-鸟嘌呤)损伤、胸腺嘧啶二醇损伤、6-羟基-5，6-二氢胸腺嘧啶损伤、碱性损伤和两个双碱基损伤。分析采用了32P-后标记或质谱学检测方法。ZeptoMetrix将在商业上提供便于使用这两种方法进行DNA测量的产品。产品将包括用于检测上述病变的参考化合物、载体和内标。32P-后标记的载体提供了对感兴趣的DNA损伤的明确识别。用于MS测量的内部标准将被同位素浓缩到比要定量的DNA损伤更高的质量。ZeptoMetrix预计，这些产品最终最大的应用将是评估细胞和组织的修复能力。用于修复研究的产品将包括参考化合物和含有已识别损伤的双链DNA底物。利用电喷雾接口的液质联用(LC-MS)将特别考虑到这一应用。 建议的商业应用： 测量DNA氧化损伤的标准物质、内标和载体 将被合成。这些产品将以商业形式提供，以促进DNA损伤 并通过32P-后标记或LC-MS修复。