3-Nitrobenzanthrone (3NBA), an environmental pollutant and potent mutagen, causes DNA damage via the reaction of its metabolically activated form with the exocyclic amino groups of purines and the C-8 position of guanine. The present work describes a synthetic approach to the preparation of oligomeric 2′-deoxyribonucleotides containing a 2-(2′-deoxyguanosin-N2-yl)-3-aminobenzanthrone moiety, one of the major DNA adducts found in tissues of living organisms exposed to 3NBA. The NMR spectra indicate that the damaged oligodeoxyribonucleotide is capable of forming a regular double helical structure with the polyaromatic moiety assuming a single conformation at room temperature; the spectra suggest that the 3ABA moiety resides in the duplex minor groove pointing toward the 5′-end of the modified strand. Thermodynamic studies show that the dG(N2)-3ABA lesion has a stabilizing effect on the damaged duplex, a fact that correlates well with the long persistence of this damage in living organisms. 1. Introduction Nitroarenes are widespread environmental pollutants found in cigarette smoke, coal fly ash, exhausts from diesel and gasoline engines, and air particulate matter [1]. 3-Nitrobenzanthrone (3NBA, 3-nitro-7H-benz[de]anthracen-7-one), a member of this class of compounds, is one of the most potent bacterial mutagens known to date [2] and exhibits strong mutagenic activity in mammalian cells [3]. Human fibroblasts exposed to N-acetoxy-3-aminobenzanthrone, an active metabolite of 3NBA, show predominance of base substitution mutations mainly G → T transversions followed by G → A and A → G substitutions [4]. Transgenic mice treated with 3NBA exhibit increased mutation frequency in liver, colon, bladder, and bone marrow, with G → T transversions dominating this effect [5, 6]. Analysis of 3NBA-induced mutations in mouse fibroblasts harboring a functional copy of the human p53 gene also reveals a preponderance of G → T transversions, followed by adenine base substitutions [7]. In addition to mutations, 3NBA causes DNA double-strand breaks and induces the formation of micronuclei in human cell lines [8, 9]. Female F344 rats treated with 3NBA by intratracheal instillation show acute and subacute changes in their lungs just few weeks after treatment and develop squamous cell lung tumors within 9 to 12 months, depending on the amount of 3NBA administered [10]. Some epidemiological evidence links environmental 3NBA exposure to lung cancer in humans [11, 12]. Like other environmental nitroarenes, 3NBA undergoes metabolic transformation to reactive electrophilic
References
[1]
H. S. Rosenkranz, “Mutagenic nitroarenes, diesel emissions, particulate-induced mutations and cancer: an essay on cancer-causation by a moving target,” Mutation Research, vol. 367, no. 2, pp. 65–72, 1996.
[2]
T. Enya, H. Suzuki, T. Watanabe, T. Hirayama, and Y. Hisamatsu, “3-Nitrobenzanthrone, a powerful bacterial mutagen and suspected human carcinogen found in diesel exhaust and airborne particulates,” Environmental Science and Technology, vol. 31, no. 10, pp. 2772–2776, 1997.
[3]
P. T. Phousongphouang, A. J. Grosovsky, D. A. Eastmond, M. Covarrubias, and J. Arey, “The genotoxicity of 3-nitrobenzanthrone and the nitropyrene lactones in human lymphoblasts,” Mutation Research, vol. 472, no. 1-2, pp. 93–103, 2000.
[4]
H. Nishida, M. Kawanishi, T. Takamura-Enya, and T. Yagi, “Mutagenic specificity of N-acetoxy-3-aminobenzanthrone, a major metabolically activated form of 3-nitrobenzanthrone, in shuttle vector plasmids propagated in human cells,” Mutation Research, vol. 654, no. 1, pp. 82–87, 2008.
[5]
V. M. Arlt, L. Zhan, H. H. Schmeiser et al., “DNA adducts and mutagenic specificity of the ubiquitous environmental pollutant 3-nitrobenzanthrone in Muta Mouse,” Environmental and Molecular Mutagenesis, vol. 43, no. 3, pp. 186–195, 2004.
[6]
V. M. Arlt, J. Gingerich, H. H. Schmeiser, D. H. Phillips, G. R. Douglas, and P. A. White, “Genotoxicity of 3-nitrobenzanthrone and 3-aminobenzanthrone in Muta Mouse and lung epithelial cells derived from Muta？Mouse,” Mutagenesis, vol. 23, no. 6, pp. 483–490, 2008.
[7]
J. Vom Brocke, A. Krais, C. Whibley, M. C. Hollstein, and H. H. Schmeiser, “The carcinogenic air pollutant 3-nitrobenzanthrone induces GC to TA transversion mutations in human p53 sequences,” Mutagenesis, vol. 24, no. 1, pp. 17–23, 2009.
[8]
E. Lamy, F. Kassie, R. Gminski, H. H. Schmeiser, and V. Mersch-Sundermann, “3-Nitrobenzanthrone (3-NBA) induced micronucleus formation and DNA damage in human hepatoma (HepG2) cells,” Toxicology Letters, vol. 146, no. 2, pp. 103–109, 2004.
[9]
V. M. Arlt, K. J. Cole, and D. H. Phillips, “Activation of 3-nitrobenzanthrone and its metabolites to DNA-damaging species in human B lymphoblastoid MCL-5 cells,” Mutagenesis, vol. 19, no. 2, pp. 149–156, 2004.
[10]
E. Nagy, M. Zeisig, K. Kawamura et al., “DNA adduct and tumor formations in rats after intratracheal administration of the urban air pollutant 3-nitrobenzanthrone,” Carcinogenesis, vol. 26, no. 10, pp. 1821–1828, 2005.
[11]
P. Vineis, F. Forastiere, G. Hoek, and M. Lipsett, “Outdoor air pollution and lung cancer: recent epidemiologic evidence,” International Journal of Cancer, vol. 111, no. 5, pp. 647–652, 2004.
[12]
E. Garshick, F. Laden, J. E. Hart et al., “Lung cancer in railroad workers exposed to diesel exhaust,” Environmental Health Perspectives, vol. 112, no. 15, pp. 1539–1543, 2004.
[13]
V. M. Arlt, M. Stiborova, C. J. Henderson et al., “Environmental pollutant and potent mutagen 3-nitrobenzanthrone forms DNA adducts after reduction by NAD(P)H:quinone oxidoreductase and conjugation by acetyltranferases and sulfotransferases in human hepatic cytosols,” Cancer Research, vol. 65, no. 7, pp. 2644–2652, 2005.
[14]
M. Stiborová, H. Dra？ínská, J. Mizerovská et al., “The environmental pollutant and carcinogen 3-nitrobenzanthrone induces cytochrome P450 1A1 and NAD(P)H:quinone oxidoreductase in rat lung and kidney, thereby enhancing its own genotoxicity,” Toxicology, vol. 247, no. 1, pp. 11–22, 2008.
[15]
V. M. Arlt, M. Stiborova, A. Hewer, H. H. Schmeiser, and D. H. Phillips, “Human enzymes involved in the metabolic activation of the environmental contaminant 3-nitrobenzanthrone: evidence for reductive activation by human NADPH:Cytochrome P450 reductase,” Cancer Research, vol. 63, no. 11, pp. 2752–2761, 2003.
[16]
A. Seidel, D. Dahmann, H. Krekeler, and J. Jacob, “Biomonitoring of polycyclic aromatic compounds in the urine of mining workers occupationally exposed to diesel exhaust,” International Journal of Hygiene and Environmental Health, vol. 204, no. 5-6, pp. 333–338, 2002.
[17]
V. M. Arlt, A. Hewer, B. L. Sorg, H. H. Schmeiser, D. H. Phillips, and M. Stiborova, “3-Aminobenzanthrone, a human metabolite of the environmental pollutant 3-nitrobenzanthrone, forms DNA adducts after metabolic activation by human and rat liver microsomes: evidence for activation by cytochrome P450 1A1 and P450 1A2,” Chemical Research in Toxicology, vol. 17, no. 8, pp. 1092–1101, 2004.
[18]
V. M. Arlt, C. J. Henderson, C. R. Wolf, H. H. Schmeiser, D. H. Phillips, and M. Stiborova, “Bioactivation of 3-aminobenzanthrone, a human metabolite of the environmental pollutant 3-nitrobenzanthrone: evidence for DNA adduct formation mediated by cytochrome P450 enzymes and peroxidases,” Cancer Letters, vol. 234, no. 2, pp. 220–231, 2006.
[19]
V. M. Arlt, B. L. Sorg, M. Osborne et al., “DNA adduct formation by the ubiquitous environmental pollutant 3-nitrobenzanthrone and its metabolites in rats,” Biochemical and Biophysical Research Communications, vol. 300, no. 1, pp. 107–114, 2003.
[20]
C. A. Bieler, M. G. Cornelius, R. Klein et al., “DNA adduct formation by the environmental con taminant 3-Nitrobenzanthrone after intratracheal instillation in rats,” International Journal of Cancer, vol. 116, no. 6, pp. 833–838, 2005.
[21]
C. A. Bieler, M. G. Cornelius, M. Stiborova et al., “Formation and persistence of DNA adducts formed by the carcinogenic air pollutant 3-nitrobenzanthrone in target and non-target organs after intratracheal instillation in rats,” Carcinogenesis, vol. 28, no. 5, pp. 1117–1121, 2007.
[22]
V. M. Arlt, H. H. Schmeiser, M. R. Osborne et al., “Identification of three major DNA adducts formed by the carcinogenic air pollutant 3-nitrobenzanthrone in rat lung at the C8 and N2 position of guanine and at the N6 position of adenine,” International Journal of Cancer, vol. 118, no. 9, pp. 2139–2146, 2006.
[23]
T. Takamura-Enya, M. Kawanishi, T. Yagi, and Y. Hisamatsu, “Structural identification of DNA adducts derived from 3-nitrobenzanthrone, a potent carcinogen present in the atmosphere,” Chemistry, vol. 2, no. 9, pp. 1174–1185, 2007.
[24]
T. Zaliznyak, R. Bonala, F. Johnson, and C. De Los Santos, “Structure and stability of duplex DNA containing the 3-(deoxyguanosin- N2-yl)-2-acetylaminofluorene (dG(N2)-AAF) lesion: a bulky adduct that persists in cellular DNA,” Chemical Research in Toxicology, vol. 19, no. 6, pp. 745–752, 2006.
[25]
X. S. Cui, L. C. Eriksson, and L. M？ller, “Formation and persistence of DNA adducts during and after a long-term administration of 2-nitrofluorene,” Mutation Research, vol. 442, no. 1, pp. 9–18, 1999.
[26]
K. J. Breslauer, “Extracting thermodynamic data from equilibrium melting curves for oligonucleotide order-disorder transitions,” Methods in Enzymology, vol. 259, pp. 221–242, 1995.
[27]
F. Delaglio, S. Grzesiek, G. W. Vuister, G. Zhu, J. Pfeifer, and A. Bax, “NMRPipe: a multidimensional spectral processing system based on UNIX pipes,” Journal of Biomolecular NMR, vol. 6, no. 3, pp. 277–293, 1995.
[28]
T. D. Goddard and D. G. Kneller, SPARKY 3, University of California, San Francisco, Calif, USA.
[29]
R. R. Bonala, M. C. Torres, S. Attaluri, C. R. Iden, and F. Johnson, “Incorporation of N2-deoxyguanosine metabolic adducts of 2-aminonaphthalene and 2-aminofluorene into oligomeric DNA,” Chemical Research in Toxicology, vol. 18, no. 3, pp. 457–465, 2005.
[30]
F. De Riccardis and F. Johnson, “Chemical synthesis of cross-linked purine nucleosides,” Organic Letters, vol. 2, no. 3, pp. 293–295, 2002.
[31]
M. Cosman, C. De los Santos, R. Fiala et al., “Solution conformation of the major adduct between the carcinogen (+)-anti- benzo[a]pyrene diol epoxide and DNA,” Proceedings of the National Academy of Sciences of the United States of America, vol. 89, no. 5, pp. 1914–1918, 1992.
[32]
C. De los Santos, M. Cosman, B. E. Hingerty et al., “Influence of benzo[a]pyrene diol epoxide chirality on solution conformations of DNA covalent adducts: the (-)-trans-anti-[BP]G · C adduct structure and comparison with the (+)-trans-anti-[BP]G · C enantiomer,” Biochemistry, vol. 31, no. 23, pp. 5245–5252, 1992.
[33]
L. A. Marky, D. Rentzeperis, N. P. Luneva, M. Cosman, N. E. Geacintov, and D. W. Kupke, “Differential hydration thermodynamics of stereoisomeric DNA-benzo[a]pyrene adducts derived from diol epoxide enantiomers with different tumorigenic potentials,” Journal of the American Chemical Society, vol. 118, no. 16, pp. 3804–3810, 1996.
[34]
S. Gopalakrishnan, T. M. Harris, and M. P. Stone, “Intercalation of aflatoxin B1 in two oligodeoxynucleotide adducts: comparative 1H NMR analysis of d(ATCAFBGAT)·d(ATCGAT) and d(ATAFBGCAT)2,” Biochemistry, vol. 29, no. 46, pp. 10438–10448, 1990.
[35]
H. Mao, Z. Deng, F. Wang, T. M. Harris, and M. P. Stone, “An intercalated and thermally stable FAPY adduct of aflatoxin B1 in a DNA duplex: structural refinement from 1H NMR,” Biochemistry, vol. 37, no. 13, pp. 4374–4387, 1998.
[36]
D. Gunz, M. T. Hess, and H. Naegeli, “Recognition of DNA adducts by human nucleotide excision repair. Evidence for a thermodynamic probing mechanism,” Journal of Biological Chemistry, vol. 271, no. 41, pp. 25089–25098, 1996.
[37]
N. E. Geacintov, S. Broyde, T. Buterin et al., “Thermodynamic and structural factors in the removal of bulky DNA adducts by the nucleotide excision repair machinery,” Biopolymers, vol. 65, no. 3, pp. 202–210, 2002.
[38]
Y. Cai, S. Ding, N. E. Geacintov, and S. Broyde, “Intercalative conformations of the 14R(+)- and 14S(-)- trans-anti-DB[a,l]P- N6-dA adducts: molecular modeling and MD simulations,” Chemical Research in Toxicology, vol. 24, no. 4, pp. 522–531, 2011.
[39]
R. J. Isaacs and H. P. Spielmann, “A model for initial DNA lesion recognition by NER and MMR based on local conformational flexibility,” DNA Repair, vol. 3, no. 5, pp. 455–464, 2004.
