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Accounts of Chemical Research May 2019Chemical damage to DNA is a key initiator of adverse biological consequences due to disruption of the faithful reading of the genetic code. For example, O-alkylguanine (...
Chemical damage to DNA is a key initiator of adverse biological consequences due to disruption of the faithful reading of the genetic code. For example, O-alkylguanine ( O-alkylG) DNA adducts are strongly miscoding during DNA replication when the damaged nucleobase is a template for polymerase-mediated translesion DNA synthesis. Thus, mutations derived from O-alkylG adducts can have severe adverse effects on protein translation and function and are an early event in the initiation of carcinogenesis. However, the low abundance of these adducts places significant limitations on our ability to relate their presence and biological influences with resultant mutations or disease risk. As a consequence, there is a critical need for novel tools to detect and study the biological role of alkylation adducts. Incorporating DNA bases with altered structures that are derived synthetically is a strategy that has been used widely to interrogate biological processes involving DNA. Such synthetic nucleosides have contributed to our understanding of DNA structure, DNA polymerase (Pol) and repair enzyme function, and to the expansion of the genetic alphabet. This Account describes our efforts toward creating and applying synthetic nucleosides directed at DNA adducts. We synthesized a variety of nucleosides with altered base structures that complement the altered hydrogen bonding capacity and hydrophilicity of O-alkylG adducts. The heterocyclic perimidinone-derived nucleoside Per was the first of such adduct-directed synthetic nucleosides; it specifically stabilized O-benzylguanine ( O-BnG) in a DNA duplex. Structural variants of Per were used to determine hydrogen bonding and base-stacking contributions to DNA duplex stability in templates containing O-BnG as well as O-methylguanine ( O-MeG) adducts. We created synthetic probes able to stabilize damaged over undamaged templates and established how altered hydrogen bonding or base-stacking properties impact DNA duplex stability as a function of adduct structures. This knowledge was then applied to devise a hybridization-based detection strategy involving gold nanoparticles that distinguish damaged from undamaged DNA by colorimetric changes. Furthermore, synthetic nucleosides were used as mechanistic tools to understand chemical determinants such as hydrogen bonding, π-stacking, and size and shape deviations that impact the efficiency and fidelity of DNA adduct bypass by DNA Pols. Finally, we reported the first example of amplifying alkylated DNA, accomplished by combining an engineered polymerase and synthetic triphosphate for which incorporation is templated by a DNA adduct. The presence of the synthetic nucleoside in amplicons could serve as a marker for the presence and location of DNA damage at low levels in DNA strands. Adduct-directed synthetic nucleosides have opened new concepts to interrogate the levels, locations, and biological influences of DNA alkylation.
Topics: Base Pairing; DNA Adducts; DNA Damage; DNA-Directed DNA Polymerase; Gold; Humans; Metal Nanoparticles; Nucleic Acid Hybridization; Nucleosides
PubMed: 30964643
DOI: 10.1021/acs.accounts.9b00054 -
International Journal of Molecular... Mar 2018A large number of chemicals and several physical agents, such as UV light and γ-radiation, have been associated with the etiology of human cancer. Generation of DNA... (Review)
Review
A large number of chemicals and several physical agents, such as UV light and γ-radiation, have been associated with the etiology of human cancer. Generation of DNA damage (also known as DNA adducts or lesions) induced by these agents is an important first step in the process of carcinogenesis. Evolutionary processes gave rise to DNA repair tools that are efficient in repairing damaged DNA; yet replication of damaged DNA may take place prior to repair, particularly when they are induced at a high frequency. Damaged DNA replication may lead to gene mutations, which in turn may give rise to altered proteins. Mutations in an oncogene, a tumor-suppressor gene, or a gene that controls the cell cycle can generate a clonal cell population with a distinct advantage in proliferation. Many such events, broadly divided into the stages of initiation, promotion, and progression, which may occur over a long period of time and transpire in the context of chronic exposure to carcinogens, can lead to the induction of human cancer. This is exemplified in the long-term use of tobacco being responsible for an increased risk of lung cancer. This mini-review attempts to summarize this wide area that centers on DNA damage as it relates to the development of human cancer.
Topics: DNA Adducts; DNA Damage; Humans; Mutagenesis; Neoplasms
PubMed: 29570697
DOI: 10.3390/ijms19040970 -
Journal of Separation Science Jan 2020The formation of DNA adducts by genotoxic agents is an early event in cancer development, and it may lead to gene mutations, thereby initiating tumor development. The... (Review)
Review
The formation of DNA adducts by genotoxic agents is an early event in cancer development, and it may lead to gene mutations, thereby initiating tumor development. The measurement of DNA adducts can provide critical information about the genotoxic potential of a chemical and its mechanism of carcinogenesis. In recent decades, liquid chromatography coupled with mass spectrometry has become the most important technique for analyzing DNA adducts. The improvements in resolution achievable with new chromatographic separation techniques coupled with the high specificity and sensitivity and wide dynamic range of new mass spectrometry systems have been used for both qualitative and quantitative analyses of DNA adducts. This review discusses the challenges in qualitative and quantitative analyses of DNA adducts by liquid chromatography coupled with mass spectrometry and highlights recent developments towards overcoming the limitations of liquid chromatography coupled with mass spectrometry methods. The key steps and new solutions, such as sample preparation, mass spectrometry fragmentation, and method validation, are summarized. In addition, the fundamental principles and latest advances in DNA adductomic approaches are reviewed.
Topics: Base Sequence; Chromatography, Liquid; DNA Adducts; Humans; Mass Spectrometry
PubMed: 31573133
DOI: 10.1002/jssc.201900737 -
Environmental and Molecular Mutagenesis Aug 2016Over two centuries ago, Sir Percival Pott, a London surgeon, published a pioneering treatise showing that soot exposure was the cause of high incidences of scrotal... (Review)
Review
Over two centuries ago, Sir Percival Pott, a London surgeon, published a pioneering treatise showing that soot exposure was the cause of high incidences of scrotal cancers occurring in young men who worked as chimney sweeps. Practicing at a time when cellular pathology was not yet recognized, Sir Percival nonetheless observed that the high incidence and short latency of the chimney sweep cancers, was fundamentally different from the rare scrotal cancers typically found in elderly men. Furthermore, his diagnosis that the etiology of these cancers was related to chimney soot exposure, was absolutely accurate, conceptually novel, and initiated the field of "occupational cancer epidemiology." After many intervening years of research focused on mechanisms of chemical carcinogenesis, briefly described here, it is clear that DNA damage, or DNA adduct formation, is "necessary but not sufficient" for tumor induction, and that many additional factors contribute to carcinogenesis. This review includes a synopsis of carcinogen-induced DNA adduct formation in experimental models and in the human population, with particular attention paid to molecular dosimetry and molecular cancer epidemiology. Environ. Mol. Mutagen. 57:499-507, 2016. © 2016 Wiley Periodicals, Inc.
Topics: Animals; Carcinogenesis; Carcinogens, Environmental; DNA Adducts; Environmental Exposure; Humans; Molecular Epidemiology; Neoplasms
PubMed: 27346877
DOI: 10.1002/em.22030 -
Oncogene Oct 2002DNA adducts associated with tobacco smoking could provide a marker of biologically effective dose of tobacco carcinogens and improve individual cancer risk prediction. A... (Review)
Review
DNA adducts associated with tobacco smoking could provide a marker of biologically effective dose of tobacco carcinogens and improve individual cancer risk prediction. A significant number of clinical and epidemiologic studies have reported associations of increased DNA adduct levels with the occurrence of the prevalent tobacco related cancers including cancer of the lung, head and neck, and bladder. The inducibility of DNA adducts following in vitro treatments using blood lymphocytes also appears to be a risk factor in the development of lung and head and neck cancer. Corroborative evidence pointing to the importance of DNA adducts in tobacco carcinogenesis include numerous studies showing associations of tobacco smoke exposure with the induction of DNA adducts in humans in vivo. Further effort is necessary, however, to more fully characterize the dose-response relationship between smoking and DNA adducts in exposed target and surrogate tissues. The relationship between gene polymorphisms thought to modify tobacco-related cancer risk and DNA adduct levels is complex. Results of some DNA adduct studies (both in vitro and in vivo) appear inconsistent with the epidemiologic findings. This is evident for polymorphisms involving both carcinogen metabolism (e.g. GSTP1) and DNA repair (e.g. XRCC1). Molecular studies of human tumors suggest associations of p53 mutation with DNA adducts and have revealed correlations of DNA adduct levels with somatic alterations (e.g. 3p21 LOH) that are thought to occur at the very earliest stages of tobacco carcinogenesis. More research is needed to assess the relationship between endogenous sources of DNA adducts and tobacco smoke exposure and the relative oncogenic effects of chemically stable versus unstable DNA adducts. Many potentially fruitful new avenues of cancer research are emerging that integrate DNA adduct analyses with assessments of smoking, genetics, diet and ambient air quality. These investigations aim to understand the multifactorial nature of interindividual variability in response to tobacco carcinogens. As these trends continue a variety of innovative study designs and approaches will become important in human populations.
Topics: Cell Transformation, Neoplastic; DNA Adducts; DNA Repair; Humans; Lung Neoplasms; Risk Factors; Nicotiana
PubMed: 12379880
DOI: 10.1038/sj.onc.1205799 -
Advances in Experimental Medicine and... 2014The formation of DNA adducts is considered essential for tumor initiation. Quantification of DNA adducts may be achieved by various techniques of which LC-MS/MS-based... (Review)
Review
The formation of DNA adducts is considered essential for tumor initiation. Quantification of DNA adducts may be achieved by various techniques of which LC-MS/MS-based multiple reaction monitoring has become the most prominent in the past decade. Adducts of single nucleosides are analyzed following enzymatic break-down of a DNA sample following adduct enrichment usually by solid-phase extraction. LC-MS/MS quantification is carried out using stable isotope-labeled internal reference substances. An upcoming challenge is the use of DNA adducts as biomarkers either for internal exposure to electrophilic genotoxins or for the approximation of cancer risk. Here we review recent studies in which DNA adducts were quantified by LC-MS/MS in DNA samples from human matrices. We outline a possible way for future research to aim at the development of an "adductome" approach for the characterization of DNA adduct spectra in human tissues. The DNA adduct spectrum reflects the inner exposure of an individual's tissue to electrophilic metabolites and, therefore, should replace the conventional and inaccurate external exposure in epidemiological studies in the future.
Topics: Animals; Biomarkers, Tumor; DNA Adducts; DNA, Neoplasm; Humans; Mass Spectrometry; Molecular Epidemiology; Neoplasms
PubMed: 24952193
DOI: 10.1007/978-3-319-06068-2_18 -
Mutation Research Mar 1999The quantitative relationship between DNA adducts and tumor incidence is evaluated in this review. All available data on DNA adduct levels determined after repeated... (Review)
Review
The quantitative relationship between DNA adducts and tumor incidence is evaluated in this review. All available data on DNA adduct levels determined after repeated administration of a carcinogen to rats or mice have been compiled. The list comprised 27 chemicals, of all major structural classes of carcinogens. For the correlation with tumor incidence, the DNA adduct levels measured at the given dose were normalized to the dose which resulted in a 50% tumor incidence under the conditions of a 2-year bioassay (TD50 dose). In rat liver, the calculated adduct concentration 'responsible' for a 50% hepatocellular tumor incidence spanned from 53 to 2083 adducts per 108 nucleotides, for aflatoxin B1, tamoxifen, IQ, MeIQx, 2,4-diaminotoluene, and dimethylnitrosamine (in this order). In mouse liver, the respective figures were 812 to 5543 adducts per 108 nucleotides, for ethylene oxide, dimethylnitrosamine, 4-aminobiphenyl, and 2-acetylaminofluorene. The observed span (40-fold in rats, 7-fold in mice) reflects differences between the various DNA adducts to lead to critical mutations. If additional carcinogens fit in with this astonishingly narrow range, the measurement of DNA adduct levels in target tissue has the potential to be not only an exposure marker but an individual cancer risk marker. For toremifen and styrene, low levels of DNA adducts were detected in rat liver at the end of a negative long-term bioassay. This shows that the limit of detection of DNA adducts can be well below the limit of detection of an increased tumor incidence. For a cancer risk assessment at low levels of DNA damage, treatment-related adducts must be discussed in relation to the background DNA damage and its inter- and intraindividual variability.
Topics: Animals; Carcinogens; DNA Adducts; DNA, Neoplasm; Humans; Mice; Neoplasms; Neoplasms, Experimental; Rats; Risk Factors
PubMed: 10064864
DOI: 10.1016/s0027-5107(99)00022-6 -
Mutation Research Mar 1999Reports on intraindividual changes of DNA adduct levels in humans are rare. Most of the data available in the literature are from polycyclic aromatic hydrocarbons (PAHs)... (Review)
Review
Reports on intraindividual changes of DNA adduct levels in humans are rare. Most of the data available in the literature are from polycyclic aromatic hydrocarbons (PAHs) and are measured in white blood cells with 32P-postlabeling or immunochemical assays. Surprisingly, environmental exposure can have a larger effect on PAH adduct levels than occupational exposure, food or smoking. Highest (13-fold) summer/winter increments, due to indoor heating were observed in Gliwice, Poland. Further studies of environmental PAH exposure confirm the environmental influence on intraindividual changes in PAHs-DNA adduct levels: studies of the Teplice program, (Czech Republic) and studies with US soldiers, stationed in Germany who went for a 8-week period of duty to Kuwait. Variations in occupational exposure, e.g., changing of anode material in aluminium plants (elevation factor 3.94), layoffs, reduced working hours in iron foundries or vacation also led to intraindividual changes in PAH adduct levels. Increase in PAH adduct levels after consumption of charcoal broiled meat evidently depends on individual susceptibility, e.g., polymorphism. In one person a 7.4-fold increment was observed. PAH adduct levels were not significantly influenced by smoking cessation whereas sister chromatid exchanges significantly decreased. Changes in occupational exposure to styrene in lamination plants, e.g., due to vacation, did not significantly influence styrene-O6-dG adduct levels in lymphocytes and granulocytes as determined by 32P-postlabeling. Increase of N7-methylguanine and O6-methylguanine levels were followed in white blood cells during treatment of cancer patients with dacarbazine and allowed insights into pharmacokinetic properties. According to a rough estimation the high increment in the PAHs-DNA adduct level of about 13 observed in Gliwice (see above) would result in a tentative increase in cancer risk from about 1 death/107 inhabitants to approximately 10 deaths/107 inhabitants which, in general, is considered as acceptable.
Topics: Adult; Circadian Rhythm; DNA Adducts; Genetic Variation; Humans
PubMed: 10064865
DOI: 10.1016/s0027-5107(99)00023-8 -
Environmental Health Perspectives Oct 1996Human DNA adduct formation (covalent modification of DNA with chemical carcinogens) is a promising biomarker for elucidating the molecular epidemiology of cancer.... (Review)
Review
Human DNA adduct formation (covalent modification of DNA with chemical carcinogens) is a promising biomarker for elucidating the molecular epidemiology of cancer. Classes of compounds for which human DNA adducts have been observed include polycyclic aromatic hydrocarbons (PAHs), nitrosamines, mycotoxins, aromatic amines, heterocyclic amines, ultraviolet light, and alkylating cancer chemotherapeutic agents. Most human DNA adduct exposure monitoring has been performed with either 32P-postlabeling or immunoassays, neither of which is able to chemically characterize specific DNA adducts. Recently developed combinations of methods with chemical and physical end points have allowed identification of specific adducts in human tissues. Studies are presented that demonstrate that high ambient levels of benzo[a]pyrene are associated with high levels of DNA adducts in human blood cell DNA and that the same DNA adduct levels drop when the ambient PAH levels decrease significantly. DNA adduct dosimetry, which has been achieved with some dietary carcinogens and cancer chemotherapeutic agents, is described, as well as studies correlating DNA adducts with other biomarkers. It is likely that some toxic, noncarcinogenic compounds may have genotoxic effects, including oxidative damage, and that adverse health outcomes other than cancer may be correlated with DNA adduct formation. The studies presented here may serve as useful prototypes for exploration of other toxicological end points.
Topics: Biomarkers; DNA Adducts; Environmental Monitoring; Gas Chromatography-Mass Spectrometry; Humans; Immunoassay; Luminescent Measurements
PubMed: 8933030
DOI: 10.1289/ehp.96104s5883 -
Regulatory Toxicology and Pharmacology... Dec 2000This article, based on a presentation on DNA adduct detection given at a Genetic Toxicology Association workshop, is an overview of methods used for testing compounds... (Review)
Review
This article, based on a presentation on DNA adduct detection given at a Genetic Toxicology Association workshop, is an overview of methods used for testing compounds for DNA adduct formation. A DNA adduct study may be initiated on a case by case basis when there are conflicting results within the standard battery of genetic toxicology tests or when tumors are detected in the animal bioassay for nongenotoxic compounds. Methods for adduct detection include the 32P-postlabeling assay, the use of radioactive test chemicals, physicochemical methods, and immunoassays. Of these, the 32P-postlabeling assay and the use of radiochemicals are discussed in greater detail, since only these two methods are readily applicable to test a compound for the formation of uncharacterized DNA adducts. The other methods are applicable to those adducts that have been chemically characterized or that contain a fluorophore or electrochemically active groups. Evaluation of mutagenic and carcinogenic risk from DNA adducts would require the understanding of various parameters, including the chemical nature, quantity and stability of adducts, proliferation rates for target cells to fix adducts into mutations, mutagenic and repair efficiencies of adducts, and the extent of modifications in critical genes. Since such data cannot be readily obtainable, the toxicological risk from uncharacterized adducts is difficult to assess.
Topics: Animals; Carcinogens; Chromatography, Thin Layer; DNA Adducts; Humans; Immunoassay; Mutagens; Phosphorus Radioisotopes
PubMed: 11162719
DOI: 10.1006/rtph.2000.1430