Detection of 1,N6-ethenodeoxyadenosine and 3,N4-ethenodeoxycytidine by immunoaffinity/32P-post-labelling in liver and lung DNA of mice treated with ethylcarbamate (urethane) or its metabolites

Fernando, R.Charles; Nair, J. Somarajan; Barbin, Alain; Miller, James A.; Bartsch, Helmut

doi:10.1093/carcin/17.8.1711

Cited by 72 publications

(43 citation statements)

References 28 publications

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“…This finding is consistent with data from previous in vivo studies that identified dAS and 3,N 4 -ethenodeoxycytidine in liver and lung DNA of mice treated with EC or VC (Fernando et al, 1996;Titis and Forkert, 2001). Both carbamate compounds also formed 7-(2Ј-oxoethyl)deoxyguanosine and dAS in liver DNA of rodents (Ribovich et al, 1982;Miller and Miller, 1983;Scherer et al, 1986).…”

Section: Downloaded Fromsupporting

confidence: 93%

“…In subsequent studies, VCO was synthesized and found to react with calf thymus DNA to form two major guanine adducts, N 2 ,3-ethenodeoxyguanosine and 7-(2Ј-oxoethyl)deoxyguanosine, and one minor adenine adduct, 1,N 6 -ethenodeoxyadenosine (dAS) (Park et al, 1993). More recent studies have identified the formation of dAS and 3,N 4 -ethenodeoxycytidine in liver and lung DNA of several strains of mice treated with EC or its metabolites (Fernando et al, 1996). These data supported the proposal that VCO is a reactive species responsible for mediating DNA alkylation, leading to the formation of DNA adducts.…”

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confidence: 99%

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Oxidation of Vinyl Carbamate and Formation of 1,N6-Ethenodeoxyadenosine in Murine Lung

Forkert

Kaufmann²,

Black³

et al. 2007

Drug Metabolism and Disposition

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ABSTRACT:Vinyl carbamate (VC) is derived from ethyl carbamate, a carcinogen formed in fermentation of food and alcoholic products. We have undertaken studies to test the hypothesis that an epoxide generated from VC oxidation leads to formation of 1,N 6 -ethenodeoxyadenosine (dAS). We have developed approaches using liquid chromatography-mass spectrometry and liquid chromatographytandem mass spectrometry for identification and quantitation of dAS. Peak rates were produced by lung microsomes and recombinant CYP2E1 at 3.0 and 2.5 mM VC, respectively. In inhibitory studies, incubations of VC were performed using lung microsomes from mice treated with the CYP2E1 inhibitor diallyl sulfone (100 mg/kg, p.o.). Results from these studies showed significantly decreased dAS formation in microsomes incubated with VC, with an inhibition of 70% at 3.0 mM. These findings suggested that CYP2E1 is a major enzyme mediating VC oxidation, leading to the formation of a metabolite that alkylates DNA to form the dAS adduct. Scanning and fragment ion analyses confirmed the identity of dAS based on the molecular ion [M ؉ H]

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confidence: 93%

mentioning

confidence: 99%

Oxidation of Vinyl Carbamate and Formation of 1,N6-Ethenodeoxyadenosine in Murine Lung

Forkert

Kaufmann²,

Black³

et al. 2007

Drug Metabolism and Disposition

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“…The liquid handling system consisted of two pumps. (2) (150 ϫ 4.6 mm, 5 m) protected with a C18 (ODS) guard column (4.0 ϫ 3.0 mm); column 2 (C2) was a Synergi Polar-RP column (75 ϫ 4.6 mm, 4 m); column 3 (C3) was a Synergi Polar-RP column (150 ϫ 4.6 mm, 4 m). All columns were from Phenomenex (Torrance, CA).…”

Section: Liquid Chromatographymentioning

confidence: 99%

“…To our knowledge, this is the first method described that allows simultaneous determination of Ade, dC, and dA in human urine samples. [2], are also generated endogenously by reactions of DNA with products derived from lipid peroxidation and oxidative stress [3]. These DNA adducts have miscoding potential and specific repair pathways supporting the hypothesis that -DNA adducts play a causal relationship in carcinogenesis [4].…”

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confidence: 99%

Quantification of urinary etheno-DNA adducts by column-switching LC/APCI-MS/MS

Hillestrøm

Weimann

Poulsen

2006

J. Am. Soc. Mass Spectrom.

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Lipid peroxidation induced etheno-DNA adducts are promutagenic and have been suggested to play a causal role in the development of human cancers. Therefore, human biomonitoring of etheno-DNA adducts in urine has been suggested as a potential marker for oxidative stress-related DNA damage. For quantitative determination, a column-switching LC/APCI-MS/MS method was developed for simultaneous analysis of Ade, dC, and dA in human urine. Quantitative validation parameters (precision, within-day repeatability, and betweenday reproducibility) yielded satisfactory results below 10%. Limit of quantification for Ade, dC, and dA was 5.3 fmol, 7.5 fmol, and 1.3 fmol on column, respectively. Mean urinary excretion rates of a six healthy volunteers were 45.8 pmol Ade/24 h, 96.8 pmol dC/24 h, and 18.1 pmol dA/24 h. The demonstrated levels of performance suggest a future applicability of this method to studies of cancer and other diseases related to oxidative stress in humans. To our knowledge, this is the first method described that allows simultaneous determination of Ade, dC, and dA in human urine samples. [2], are also generated endogenously by reactions of DNA with products derived from lipid peroxidation and oxidative stress [3]. These DNA adducts have miscoding potential and specific repair pathways supporting the hypothesis that -DNA adducts play a causal relationship in carcinogenesis [4]. Furthermore, several studies have found elevated -DNA adduct levels in disorders known as risk factors for cancers [5,6]. Therefore -DNA adducts have been proposed as biomarkers for human cancers associated with certain lifestyles or chronic inflammations [4]. Upon DNA repair, -DNA adducts are excreted in the urine and could, in that respect, be used to investigate the body burden of DNA damage.The analytical procedures for urinary DNA adducts are challenging indeed. The presence of many interfering substances in concentrations far greater than those of the DNA adducts demands high selectivity and low limit of quantitation for unequivocal identification and quantification. Methods using high-performance liquid chromatography (HPLC) with fluorescence detection have been used for dA determination [4,7] after immunoaffinity clean-up and a very tedious sample preparation. Furthermore, the internal standard was measured separately. Several etheno-DNA adducts have been determined in urine from healthy humans using gas chromatography-electron capture negative chemical ionization mass spectrometry (GC/EC-NCI/ MS) [8 -11]. Gas chromatography offers the advantage of greater peak resolution compared to conventional LC. However, the requirement for derivatization before analysis is a major limitation. Recently, LC-MS have been employed in determination of etheno adducts from human urine using electrospray ionization (ESI) [12][13][14]. Atmospheric pressure chemical ionization (APCI) has been used for analysis of dA and 1,N 2 -dG in vitro [15] and of dA in human urine [16].In this paper, we report that solid-phase extraction (SPE) for preconcent...

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“…It produced significant increases in the lacZ mutant frequency in the liver and lung in Muta TM Mouse transgenic model (Williams et al, 1998) and in the lambda/cII mutant frequency in BigBlue® lacI/cII transgenic mice (Mirsalis et al, 2005). It induced DNA adducts in mouse liver and lung (Fernando et al, 1996). Forkert and Lee (1997) demonstrated that urethane metabolism in lung microsomes is mediated by CYP2E1 and the carboxylesterase isozyme hydrolase A.…”

Section: Occurence Of Unique In Vivo Positivesmentioning

confidence: 99%

Scientific opinion on genotoxicity testing strategies applicable to food and feed safety assessment

2011

EFS2

298

136

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The Scientific Committee reviewed the current state-of-the-science on genotoxicity testing and provided a commentary and recommendations on genotoxicity testing strategies. A step-wise approach is recommended for the generation and evaluation of data on genotoxic potential, beginning with a basic battery of in vitro tests, comprising a bacterial reverse mutation assay and an in vitro micronucleus assay. Consideration should be given to whether specific features of the test substance might require substitution of one or more of the recommended in vitro tests by other in vitro or in vivo tests in the basic battery. In the event of negative in vitro results, it can be concluded that the substance has no genotoxic potential. In case of inconclusive, contradictory or equivocal results, it may be appropriate to conduct further testing in vitro. In case of positive in vitro results, review of the available relevant data on the test substance and, where necessary, an appropriate in vivo study to assess whether the genotoxic potential observed in vitro is expressed in vivo is recommended. Suitable in vivo tests are the mammalian erythrocyte micronucleus test, transgenic rodent assay, and Comet assay. The approach to in vivo testing should be step-wise. If the first in vivo test is positive, no further testing is necessary and the substance should be considered as an in vivo genotoxin. If the test is negative, it may be possible to conclude that the substance is not an in vivo genotoxin. However, in some cases, a second in vivo test may be necessary (e.g. if the first test is negative but more than one endpoint in the in vitro tests are positive, an in vivo test on a second endpoint may be necessary). The combination of assessing different endpoints in different tissues in the same animal in vivo should also be considered.

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Detection of 1,N⁶-ethenodeoxyadenosine and 3,N⁴-ethenodeoxycytidine by immunoaffinity/³²P-post-labelling in liver and lung DNA of mice treated with ethylcarbamate (urethane) or its metabolites

Cited by 72 publications

References 28 publications

Oxidation of Vinyl Carbamate and Formation of 1,N6-Ethenodeoxyadenosine in Murine Lung

Oxidation of Vinyl Carbamate and Formation of 1,N6-Ethenodeoxyadenosine in Murine Lung

Quantification of urinary etheno-DNA adducts by column-switching LC/APCI-MS/MS

Scientific opinion on genotoxicity testing strategies applicable to food and feed safety assessment

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