High Resolution Sensitivity Charcterization Of Polycyclic Aromatic Hydrocarbon Dna Adducts Using Fluorescence Line Narrowing Spectrometry

Author : Joshua J. Klaene
Publisher :
ISBN 13 :
Total Pages : 217 pages
Book Rating : 4.:/5 (97 download)

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Download or read book DNA Adducts as Markers of Exposure and Disease written by Joshua J. Klaene and published by . This book was released on 2014 with total page 217 pages. Available in PDF, EPUB and Kindle. Book excerpt: DNA adducts serve as important biomarkers of exposure and disease. Because DNA adducts are present so early in the carcinogenic process, they are promising indicators of risk for early detection. Sensitive and specific analytical methods are necessary to detect DNA adducts that are present in very low abundance. Liquid chromatography-mass spectrometry has become a mainstay in DNA adduct analysis and rapid technological improvements continue to expand its utility. Chapter 1 reviews the process of carcinogenesis resulting from exposure to chemical carcinogens, in which DNA adduction is the initial event that triggers a cascade of effects to transform normal cells into cancer cells. This chapter illustrates the central role of DNA adducts in carcinogenesis and the metabolic processes that determine the fate of many chemical carcinogens. Chapter 2 shifts the focus to the analytical methods used to detect and quantitate DNA adducts. This chapter chronicles the transition from 32P-postlabeling to modern liquid chromatography - mass spectrometry that offers exceptional sensitivity and structural identification of DNA adducts. Chapter 3 describes the full validation of our nanoflow liquid chromatography electrospray ionization tandem mass spectrometric (nanoLC-ESI-MS/MS) method for the quantitation, by isotope dilution, of 4-aminobiphenyl DNA adducts in mouse bladder DNA. Using criteria from the FDA's current guidance for bioanalysis, the performance of our method was determined for accuracy, precision, selectivity, sensitivity, reproducibility, and stability. Then in Chapter 4, the validated nanoLC-ESI-MS/MS method was applied to series of dosing studies using a mouse model. The goal of this initiative was to identify the molecular mechanisms that explain why men have an increased risk (by 4-fold) for developing bladder cancer. The aromatic amine 4-aminobiphenyl (4-ABP) is a known bladder carcinogen but requires metabolic activation to be genotoxic. A phase 2 metabolic enzyme with gender-biased hepatic expression has been implicated as the determining factor in this gender disparity. Results provide insight into the regulation and activity of this metabolic enzyme and its role in 4-ABP-induced bladder cancer. Chapter 5 addresses the common issue of matrix effects in LC-MS analysis and details the strategies employed to characterize the sources of matrix effects and method optimization used to overcome this problem. The optimized method was then applied in two studies of DNA adducts resulting from PAH exposures. In the first study, the nano-LC-ESI-MS/MS method described above was used to determine dose-response dependence in a culture of human bronchial epithelial cells exposed to an activated PAH metabolite. In the second study, the mutagenic potential of atmospheric urban aerosols was investigated. Fine, respirable particulate material (PM 2.5) can be transported great distances by air masses and carry with them carcinogenic PAHs leading to unavoidable exposures of humans to environmental carcinogens. We modeled this exposure by directly administering PM to a metabolically competent human cell line and used nanoLC-MS/MS methodology including the "adductomics" approach of screening for PAH-DNA adducts with constant neutral loss (CNL) scanning. Then targeted analyses were done using reaction monitoring (MRM/SRM) and MSn for enhanced sensitivity and confirmation of PAH-DNA adducts. Identification of two PAH-DNA adducts confirms that PM functions as a vehicle to deliver harmful compounds to human cells resulting in DNA damage. These results have major implications for human health. Finally, a summary is provided in Chapter 6 along with future perspectives for the role of DNA adduct analysis in biomonitoring of human health.