Anti-BPDE

Recent advances in PAH carcinogenesis research over the past decade have led to identification of diol epoxide metabolites as the principal active forms of the PAH investigated to date Q,2). Benzo-(a)pyrene (BP) has been most intensively investigated, and it has been demonstrated that a diol epoxide metabolite anti-BPDE is the active intermediate which binds covalently to DNA in human and other mammalian tissues 0,4). Anti-BPDE was also demonstrated to be a powerful mutagen in both bacterial and mammalian cells (15) These findings stimulated an outpouring of research directed towards elucidation of the molecular mechanism of PAH carcinogenesis. 

Resolution of the enantiomers of anti-BPDE was achieved by reaction of the racemic dihydrodiol with (-)-menthoxyacetyl chloride followed by preparative HPLC separation and basic methanolysis to give the optically pure (+) and (-) dihydrodiols (18-20). Epo-... 

A molecular model is proposed for stereoselectivity during the process of covalent binding of (+)-trans-7,8- dihydroxy- anti-9,10- epoxy-7,8,9,10- tetrahydro-benzo[a]pyrene, denoted by (+)-anti-BPDE or BPDE l(+) ... 

Benzo[n]pyrene is an extensively studied PAH compound. The mutagenic metabolites of benzo[a]pyrene are the (+)-7R,85,95,101 -flm/-benzo[fl]pyrene-7,8-dihydrodiol-9,10-epoxide, (+)-a //-BPDE, and the (-)-7R,8S,9S,10R enantiomer, ( )-anti-BPDE (Figure 22.16A,B). DNA damage occurs mainly by adduct formation between the CIO position of anti-BPDE to the N2 position of guanine. Four stereoisomeric bulky adducts are produced 105 (+)-trans-anti-BPDE-N2-dG, 1 OR (+)-cis-anti-BPE>E-N2-dG, 1 OR (-)-trans-anti-BPDE-N2-dG, and 105 (-)-cis-anti-BPDE-A2-dG. In vitro, the reaction of (+)-anti-BPDE with DNA forms predominantly the (+)-trans-anti-BPE)E-N2-dG adduct, while the reaction of ( )-anti-BPDE produces mainly the ( )-trans-anti-BPDE-N2-dG adduct. In cells, the major benzo[a]pyrene DNA adduct is (+)-trans-anti-BPDE-N2-dG. 

Purified yeast Pol is able to perform limited nucleotide insertions opposite several DNA lesions such as TT (6-4) photoproduct, AAF-dG adduct, and (+) or ( )-trans-anti-BPDE-N2-dG adduct. Furthermore, Pol also catalyzes extension synthesis from opposite many types of lesions with varying efficiencies, including an AP site, cis-syn TT dimer, (64) photoproduct, AAF-dG adduct, (+) or (-)-trans-an//-BPDE-/V2-dG adduct, and an acrolein-derived dG adduct. Therefore, it has been proposed that Pol functions both as an insertion polymerase and an extension polymerase. It appears that the extension activity of Pol is versatile. Thus, it is believed that Pol is a major extension polymerase during translesion synthesis in eukaryotes. 

Polr performs error-prone translesion synthesis opposite (+)- and (-)-trans-anti-BPDE-A -dG DNA adducts by predominantly inserting A opposite the lesion in vitro. This polymerase is more active in response to the former isomeric lesion. In yeast cells, Polr, Pol , and Revl are all required for G -> T transvertion mutations. The likely mechanism is A insertion opposite the lesion by Polr followed by extension synthesis by Pol . Revl probably plays a noncatalytic role in such a mutagenic bypass of the BPDE lesions. 

B. E. Flingerty and S. Broyde, Biopolymers, 24,2279 (1985). Carcinogen-Base Stacking and Base-Base Stacking in dCpdG Modified by (+) and (-) Anti BPDE. 

Translesion synthesis with DNA Pol of the A-acetyl-2-aminofluorene adduct of guanosine (88) is inefficient with templates containing (88). In the presence of the Revl protein, translesion synthesis occurs and dCTP is the major nucleotide incorporated opposite it, and studies with a mutant DNA Pol I gave similar results. Benzo[a]pyrene is a potent environmental carcinogen, which when metabolised leads to u t -benzo[a]pyrene-7,8-dihydrodiol-9,10-epoxide anti-BPDE). With dG, the major lesion is (+)-tra w-a h-B[a]P-A -dG, (89), and is usually repaired by the nucleotide excision repair (NER) pathway. The translesion synthesis past (89) has been examined with a number of polymerases. With human RNA Pol II, (89) is a block to synthesis, whilst DNA Pol k preferentially incorporated the correct nucleotide. In yeast cells, Pol induced a large number of mutations involving Pol p, whilst Pol p alone contributed to 1-3 deletions or insertions. The NER of (89) with UvrB proteins was also studied. ... 

Treatment with 2% horse USERIA serum incubation with rabbit anti-BPDE-DNA antiserum incubation with alkaline phosphatase-conjugated goat antirabbit IgG. PNPP, radiolabeled PNPP, and MgClg separation of hydrolyzed radiolabeled PNPP measurement of radioactivity... 

Pavanello, S., Favretto, D., Bmgnone, R, Mastrangelo, G., Dal Pra, G., and Clonfero, E. (1999). HPLC/ fluorescence determination of anti-BPDE—DNA adducts in mononuclear white blood cells from PAH-exposed humans. Carcinogenesis 20, 431 35. 

Hingerty, B. E. Broyde, S. (1985). Carcinogen-base stacking and base-base stacking in depdg modified by (+) and (—) anti-bpde. Biopolymers 24, 2279-2299. 

Shibutani, S., Margulis, L.A., Geacintov, N.E., and Grollman, A.P. (1993) Translesional synthesis on a DNA template containing a single stereoisomer of dG-(+)- or dG-(-)-anti-BPDE (7,8-dihydroxy-anti-9,10-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene). Biochemistry, 32, 7531-7541. 

Zhao, B., Wang, J., Geacintov, N.E., and Wang, Z. (2006) Polq, Pol and Revl together are required for G to T transversion mutations induced by the (+)- and (-)-trans-anti-BPDE-N2-dG DNA adducts in yeast cells. Nucleic Acids Res., 34, 417-425. 

DATS which are potent inhibitors of BP-induced fore-stomach cancer in mice, resulted in a significant increase, as compared with control, in bodi hepatic (3.0-, 3.2-and 4.4-fold, respectively) and fore-stomach (1.5-, 2.7-and 2.7-fold, respectively) glutathione transferase (GST) activity toward anti-7P,8a-dihy oxy-9a, 1 Oa-oxy-7,8,9,10-tetrahydrobenzo(a)pyrene (anti-BPDE), which is the ultimate carcinogen of BP [102, 107]. On the contrary, this activity was not increased in either organ by dipropyl sulfide (DPS), which is ineffective against BP-induced fore-stomach cancer. The pulmonary GST activity was not increased by any of the tested OSCs. Even though epoxide hydrolase (EH) activity was differentially altered by these OSCs, a correlation between chemopreventive efficacy of OSCs and their effects on EH activity was not apparent [102]. The chemopreventive efficacy of these OSCs correlated with their ability to increase the expression of GST n. For example, DAS treatment resulted in approximate increases of 1.7- and 2.2-fold in hepatic and fore-stomach GST n expression, respectively, over the control. Treatment of mice with DATS, which is a relatively more potent inhibitor of BP-induced fore-stomach cancer than DAS, resulted in about 3.8- and 3,2-fold increases, respectively, in hepatic and fore-stomach GST n expression over the control. On the contrary, the expression of hepatic and fore-stomach GST n was increased only marginally (10-20%) upon DPS administration [107],... 

Pavanello et al. (1999) used the HPLC/fluorescence method to detect the anti -BPDE-DNA adduct in mononuclear leukocytes exposed to PAHs. The authors concluded that this methodology can be used for the evaluation of the chronic exposure to PAHs and that the inhalatory via contributed, substantially, for the adducts formation in the lymphoi de plus monocyte fraetion in eoal factories workers and chimney cleaners. 

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