trans-3,4-dihydroxy-anti-1,2-epoxy-1,2,3,4-tetrahydrodibenz[a,j]acridine involvement in dibenz[a,j]acridine DNA adduct formation in mouse skin consistent with Ha-ras mutation patterns in tumors

Dibenz[a,j]acridine (DBA), is a N-heteropolycyclic aromatic environmental c arcinogen found in complex combustion mixtures. The major route of DBA meta bolic activation is reportedly through the trans-3,4-dihydroxy-3,4-dihydroD BA (DBA-3,4-DHD). The present studies were undertaken to determine the role of trans-3,4-dihydroxy-anti-1,2-epoxy-1,2,3,4-tetrahydroDBA (DBADE) in DBA activation pathway(s), the DNA bases involved in the binding of DBA to DNA , and whether the adducts produced are consistent with the mutation pattern in the Ha-ras gene. DBA (300 mug) or 50 mug synthesized (+/-)-DBADE was ap plied to the back of female Hsd:ICR(Br) mice. The mice were sacrificed 48 h later, and skin DNA was isolated, hydrolyzed, and analyzed with P-32-postl abeling. Of the four adducts produced in vivo, adduct 1 was the major adduc t for DBA (> 50%) and adduct 2 was the major adduct for DBADE (89%). After the reaction of (+/-)-DBADE with purine nucleotides or calf thymus (CT) DNA in vitro, 100% of the DBADE-2 ' -dAMP adducts and 94% of DBADE-CT DNA addu cts were chromatographically identical on TLC with adduct 2 and 86% of the DBADE-2 ' -dGMP adducts were chromatographically consistent with adduct 1 b y 32P-postlabeling. Papillomas were induced on the backs of mice by a singl e application of 0.2 mu mol of DBA followed by twice-weekly application of 12-o-tetra-decanoylphorbol-13-acetate (TPA, 2 mug) for 24-26 weeks. Skin ca rcinomas were induced by twice weekly applications of DBA (0.1 mu mol) on t he backs of mice. A to T and G to T transversions were found in codons 12, 13, and 61 of the Ha-ras gene in the treated mouse skin carcinoma and papil loma DNA, The mutational spectra in the Ha-ras gene are consistent with the DNA binding of DBA to dG or dA in vivo. Thus, this research has indicated that DBADE plays an important role in DBA metabolic activation and DNA bind ing in mouse skin, and an alternative pathway through a bis-dihydrodiol-epo xide of DBA may also be involved.