Biomarkers of formaldehyde based on DNA-protein cross-links

基于 DNA-蛋白质交联的甲醛生物标志物

• 批准号: 8747851
• 负责人: Kun Lu
• 金额: $ 7.45万
• 依托单位: UNIVERSITY OF GEORGIA
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-07-01 至 2016-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8747851
• 关键词: Animals; Archives; Biological Assay; Biological Markers; Biological Process; Cancer Etiology; Carcinogens; Cells; Chemicals; Cysteine; DNA; DNA Adducts; DNA Repair; DNA lesion; Data; Data Set; Deoxyguanosine; Detection; Distant; Dose; Electrophoresis; Environmental Pollutants; Enzymes; Exposure to; Formaldehyde; Foundations; Future; Glutathione; Goals; Heating; Human; Hydrolysis; International Agency for Research on Cancer; Laboratories; Lead; Lesion; Lysine; Malignant neoplasm of nasopharynx; Mass Spectrum Analysis; Measures; Methods; Molecular; Monitor; Nasal Epithelium; Organ; Pathway Analysis; Production; Proteins; Proteomics; Public Health; Radioactive; Reporting; Research; Risk Assessment; Testing; Time; Tissues; Toxic effect; base; cancer risk; carcinogenesis; carcinogenicity; crosslink; design; dosimetry; expectation; experience; exposed human population; innovation; leukemia; link protein; novel; programs; public health relevance; response

DESCRIPTION (provided by applicant): Formaldehyde is a widely used high production chemical that is also released as a byproduct of combustion and off-gassing of various building products. Formaldehyde has been classified as an animal and human carcinogen by the International Agency for Research on Cancer, causing nasopharyngeal cancer. Formaldehyde is also reported to be associated with the induction of leukemia. Formaldehyde is highly reactive with DNA and proteins to induce diverse lesions, with the formation of DNA-protein cross-links (DPC) as the primary genotoxic effect. Internal exposure monitoring and cancer risk assessment of formaldehyde have been challenging due to the lack of suitable biomarkers. This also limits our capability of examining formaldehyde distant effects in organs remote to nasal epithelium, which remains highly controversial. Recently, we have established N2-hydroxymethyl-deoxyguanosine as the first formaldehyde-specific DNA adduct biomarker, and the rich dataset generated using this biomarker is widely used in risk assessment of formaldehyde. However, DPC, formed at high abundance, are biologically more important because they significantly interfere with DNA repair as extremely bulky DNA lesions. However, no biomarkers have been established to quantify formaldehyde-induced DPC due to substantial technical challenges. Previous methods of measuring DPC either utilized non-chemical-specific methods with low sensitivity or relied on radioactive formaldehyde for exposure. The objective of this application is to develop biomarkers and sensitive mass spectrometry-based methods to measure formaldehyde-induced DPC. The central hypothesis is that cysteine-containing cross-links can be developed as biomarkers to quantify formaldehyde-induced DPC, and that proteins cross-linked with DNA can also serve as biomarkers to assess the formation of DPC. This hypothesis has been formulated on the basis of preliminary data produced in the applicant's laboratory and our experience on developing mass spectrometry assays for DNA lesions. We will test the hypothesis by first optimizing enzymatic hydrolysis approaches to digest DPC into small cross-links for mass spectrometry detection, and then developing sensitive mass spectrometry assays to quantify cysteine-containing cross-links. We will also isolate, identify and quantify proteins cross-linked with DNA following formaldehyde exposure as biomarkers of DPC. The approach is innovative because of the novel application of highly sensitive mass spectrometry to develop new formaldehyde-specific biomarkers to quantify DPC formation. The proposed research is significant, because it is expected to establish a set of novel biomarkers of formaldehyde exposure to measure formaldehyde- induced DPC, a current void in formaldehyde biomarker research. Results from this study will also lay a foundation for future studies aiming at establishing molecular dosimetry using these novel biomarkers of DPC to better understand formaldehyde carcinogenicity and its cancer risk assessment.

描述（由申请人提供）：甲醛是一种广泛使用的高产化学品，也是各种建筑产品燃烧和排气的副产品。甲醛已被国际癌症研究机构列为动物和人类致癌物，导致鼻咽癌。据报道，甲醛也与白血病的诱导有关。甲醛与DNA和蛋白质具有高度反应性，可诱导多种病变，DNA-蛋白质交联（DPC）的形成是主要的遗传毒性效应。由于缺乏合适的生物标志物，甲醛的内部暴露监测和癌症风险评估一直具有挑战性。这也限制了我们检查甲醛在远离鼻上皮的器官中的远距离影响的能力，这仍然是非常有争议的。最近，我们建立了N2-羟甲基脱氧鸟苷作为第一个甲醛特异性DNA加合物生物标志物，并使用该生物标志物产生的丰富数据集被广泛用于甲醛的风险评估。然而，在高丰度下形成的DPC在生物学上更重要，因为它们作为极其庞大的DNA损伤显著干扰DNA修复。然而，由于技术上的巨大挑战，尚未建立生物标志物来量化甲氨蝶呤诱导的DPC。以前测量DPC的方法要么利用灵敏度低的非化学特异性方法，要么依赖于放射性甲醛暴露。本申请的目的是开发生物标志物和灵敏的基于质谱的方法来测量甲醛诱导的DPC。中心假设是，含半胱氨酸的交联可以被开发为生物标志物，以量化甲酰胺诱导的DPC，并且与DNA交联的蛋白质也可以作为生物标志物，以评估DPC的形成。该假设是基于申请人实验室中产生的初步数据和我们在开发DNA损伤的质谱分析方面的经验而制定的。我们将首先通过优化酶水解方法将DPC消化成用于质谱检测的小交联来测试该假设，然后开发灵敏的质谱测定来量化含半胱氨酸的交联。我们还将分离、鉴定和定量甲醛暴露后与DNA交联的蛋白质作为DPC的生物标志物。该方法是创新的，因为高灵敏度质谱法的新应用，以开发新的甲酰胺特异性生物标志物，以量化DPC的形成。所提出的研究是有意义的，因为它有望建立一套新的甲醛暴露生物标志物来测量甲醛诱导的DPC，这是甲醛生物标志物研究中的一个空白。本研究的结果也将为未来的研究奠定基础，旨在建立分子剂量学使用这些新的生物标志物的DPC，以更好地了解甲醛致癌性及其癌症风险评估。