Chemistry and Biology of Deoxyribose Oxidation in DNA

描述（由申请人提供）：为了开发氧化应激的生物标志物，我们建议通过定义DNA中脱氧核糖氧化的化学光谱和生物学后果来建立上一个资助期的结果。DNA氧化与人类疾病的病理生理密切相关，主要研究重点是核碱基损伤。然而，新出现的证据指出脱氧核糖氧化是氧化应激毒性的关键因素。我们将继续开发分析方法，定量分离DNA和暴露于氧化剂的细胞中的脱氧核糖氧化，并确定细胞对这种损伤的反应。目标1：发展定量脱氧核糖氧化产物的分析方法。我们建议继续开发分析方法来定量核糖内酯的1'-氧化基位，所有的2'-氧化基位，3'-氧化的3'-氧和5'-氧化的核苷-5'-醛。这将为我们提供一套几乎完整的候选生物标志物，用于各种脱氧核糖氧化的化学反应。目的2：脱氧核糖氧化的比较分析。在之前的资助期内，我们启动了由多种氧化剂引起的脱氧核糖氧化的比较研究，以确定损伤谱的决定因素。我们建议进一步表征由？-辐射，a粒子和FeEDTA，并包括低能电子的研究，这是电离辐射产生的最丰富的次级物质。目的3：定义细胞中脱氧核糖氧化的化学和生物学。使用Aim 1中开发的方法和Aim 2中确定的基准，我们将继续研究细胞中脱氧核糖氧化的生物学意义。这里有三个目标：(1)定义脱氧核糖引起的病变的生物学相关谱？-辐射、a-粒子和H2O2在细胞中的作用，并与Aim 4协同鉴定候选生物标志物；(2)继续与Bruce Demple教授合作，研究脱氧核糖氧化产物的修复特性；(3)根据对双链和单链断裂的独特反应的动力学和剂量依赖性，继续我们在上一个资助期内对酵母中脱氧核糖氧化的细胞反应的观察。目的4：确定脱氧核糖氧化产物的代谢命运。我们建议通过定义脱氧核糖氧化产物的代谢和生物转化命运来推进我们的研究。目的是确定与病变发生的化学转化和释放的物种作为生物标志物的潜在效用。公共卫生相关性：拟议研究的广泛目标是确定作为氧化应激生物标志物或指标的候选分子。DNA氧化与癌症和衰老的致病机制密切相关，这一领域的大部分研究都集中在DNA核碱基成分的损伤上。然而，越来越多的证据表明，脱氧核糖氧化是氧化应激和炎症遗传毒理学的关键因素。我们建议继续努力开发脱氧核糖损伤产物作为内源性和环境诱导的氧化应激和炎症的生物标志物，具有开发预测和临床有用工具的潜力。

DESCRIPTION (provided by applicant): With the goal of developing biomarkers of oxidative stress, we propose to build on results from the previous funding period by defining the chemical spectrum and biological consequences of deoxyribose oxidation in DNA. DNA oxidation is strongly associated with the pathophysiology of human disease, with a major research focus on nucleobase damage. However, emerging evidence points to deoxyribose oxidation as a critical factor in the toxicity of oxidative stress. We will continue to develop analytical methods to quantify deoxyribose oxidation in isolated DNA and cells exposed to oxidants, and to define the cellular responses to this damage. Aim 1: Develop analytical methods to quantify deoxyribose oxidation products. We propose to continue developing analytical methods to quantify the ribonolactone abasic site from 1'-oxidation, the erythose abasic site from 2'-oxidation, the 3'-oxo species from 3'-oxidation and the nucleoside-5'-aldehyde from 5'-oxidation. This will provide us with a nearly complete set of candidate biomarkers for the various chemistries of deoxyribose oxidation. Aim 2: Comparative analysis of deoxyribose oxidation. In the previous funding period, we initiated comparative studies of deoxyribose oxidation caused by a variety of oxidants to define the determinants of the damage spectrum. We propose to further characterize deoxyribose oxidation caused by ?-radiation, a-particles and FeEDTA, and to include studies of low energy electrons, the most abundant secondary species arising from ionizing radiation. Aim 3: Define the chemistry and biology of deoxyribose oxidation in cells. Using methods developed in Aim 1 and benchmarks determined in Aim 2, we will proceed to study the biological implications of deoxyribose oxidation in cells. There are three goals here: (1) define the biologically relevant spectrum of deoxyribose lesions caused by ?-radiation, a-particles and H2O2 in cells and to identify candidate biomarkers for development in coordination with Aim 4; (2) continue our collaboration with Prof. Bruce Demple to characterize the repair of deoxyribose oxidation products; and (3) pursue our observations of cellular responses to deoxyribose oxidation in yeast made in the previous funding period in terms of the kinetics and dose- dependence of the unique responses to double- and single-strand breaks. Aim 4: Define the metabolic fate of deoxyribose oxidation products. We propose to move our studies forward by defining the metabolic and biotransformational fate of deoxyribose oxidation products. The goal is to identify the chemical transformations that occur with the lesions and the potential utility of the released species as biomarkers. PUBLIC HEALTH RELEVANCE: The broad goal of the proposed studies is to identify candidate molecules for development as biomarkers or indicators of oxidative stress. DNA oxidation is strongly associated with the causative mechanisms of cancer and aging and the bulk of research in this area is focused on damage to the nucleobase components of DNA. However, emerging evidence points to deoxyribose oxidation as a critical factor in the genetic toxicology of oxidative stress and inflammation. We propose to continue our efforts to develop deoxyribose damage products as biomarkers of endogenous and environmentally-induced oxidative stress and inflammation, with the potential for developing predictive and clinically useful tools.