Mutagenesis Advance Access originally published online on September 2, 2008
Mutagenesis 2009 24(1):17-23; doi:10.1093/mutage/gen049
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The carcinogenic air pollutant 3-nitrobenzanthrone induces GC to TA transversion mutations in human p53 sequences
Division of Molecular Toxicology 1Division of Genetic Alterations in Carcinogenesis, Deutsches Krebsforschungszentrum (German Cancer Research Center), Im Neuenheimer Feld 280, D-69120 Heidelberg, Germany 2Molecular Epidemiology Unit, Leeds Institute of Genetics, Health and Therapeutics, University of Leeds, Leeds LS2 9JT, UK
3-Nitrobenzanthrone (3-NBA) is a potent mutagen and a suspected human carcinogen present in particulate matter of diesel exhaust and ambient air pollution. Employing an assay with human p53 knock-in (Hupki) murine embryonic fibroblasts (HUFs), we examined p53 mutations induced by 3-NBA and its active metabolite, N-hydroxy-3-aminobenzanthrone (N-OH-3-ABA). Twenty-nine immortalized cultures (cell lines) from 89 HUF primary cultures exposed at passage 1 for 5 days to 2 µM 3-NBA harboured 22 different mutations in the human DNA-binding domain sequence of the Hupki p53 tumour suppressor gene. The most frequently observed mutation was GC to TA transversion (46%), corroborating previous mutation studies with 3-NBA, and consistent with the presence of persistent 3-NBA–guanosine adducts found in DNA of exposed rodents. Six of the transversions found solely in 3-NBA-treated HUFs have not been detected thus far in untreated HUFs, but have been found repeatedly in human lung tumours. 32P-post-labelling adduct analysis of DNA from HUF cells treated with 2 µM 3-NBA for 5 days showed a pattern similar to that found in vivo, indicating the metabolic competence of HUF cells to metabolize 3-NBA to electrophilic intermediates. Total DNA binding was 160 ± 56 per 107 normal nucleotides with N2-guanosine being the major adduct. In contrast, identical treatment with N-OH-3-ABA resulted in a 100-fold lower level of specific DNA adducts and no carcinogen-specific mutation pattern in the Hupki assay. This indicates that the level of DNA adduct formation by the mutagen is critical to obtain specific mutation spectra in the assay. Our results are consistent with previous experiments in Muta Mouse and are compatible with the possibility that diesel exhaust exposure contributes to mutation load in humans and to lung cancer risk.
* To whom correspondence should be addressed. Tel: 49 (0)6221 42 33 48; Fax: 49 (0)6221 42 33 75; Email: h.schmeiser{at}dkfz.de
Received on May 27, 2008; revised on July 22, 2008; accepted on July 31, 2008.