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

基本信息

批准号：
8850443
负责人：
Kun Lu
金额：
$ 7.5万
依托单位：
UNIVERSITY OF GEORGIA
依托单位国家：
美国
项目类别：
财政年份：
2014
资助国家：
美国
起止时间：
2014-07-01 至 2017-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/8850443
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 Health 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 response

项目摘要

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量化学品，也是各种建筑产品燃烧和废气的副产品。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，甲醛还与白血病的诱发有关。甲醛与 DNA 和蛋白质高度反应，可诱发多种损伤，其中形成 DNA-蛋白质交联 (DPC) 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对鼻上皮远端器官的影响的能力，这仍然存在很大争议。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，并且使用该生物标志物生成的丰富数据集被广泛用于甲醛风险评估。然而，以高丰度形成的 DPC 在生物学上更为重要，因为它们作为极其庞大的 DNA 损伤会显着干扰 DNA 修复。然而，由于巨大的技术挑战，尚未建立量化甲醛诱导的 DPC 的生物标志物。以前测量 DPC 的方法要么使用灵敏度低的非化学特异性方法，要么依赖放射性甲醛进行暴露。该应用的目的是开发生物标志物和基于灵敏质谱的方法来测量甲醛诱导的 DPC。核心假设是，含半胱氨酸的交联可以作为生物标志物来量化甲醛诱导的 DPC，并且与 DNA 交联的蛋白质也可以作为生物标志物来评估 DPC 的形成。这一假设是根据申请人实验室产生的初步数据以及我们开发 DNA 损伤质谱分析的经验而制定的。我们将首先优化酶水解方法，将 DPC 消化成用于质谱检测的小交联，然后开发灵敏的质谱测定法来量化含半胱氨酸的交联，从而检验这一假设。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。该方法具有创新性，因为它新颖地应用了高灵敏度质谱法来开发新的甲醛特异性生物标志物来量化 DPC 的形成。这项研究意义重大，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，这是目前甲醛生物标志物研究的空白。这项研究的结果也将为未来的研究奠定基础，旨在利用 DPC 的这些新型生物标志物建立分子剂量测定法，以更好地了解甲醛致癌性及其癌症风险评估。