DETECTION OF THE MAJOR DNA-ADDUCTS OF BENZO[B]FLUORANTHENE IN MOUSE SKIN - ROLE OF PHENOLIC DIHYDRODIOLS

The primary mechanism of activation of benzo[b]fluoranthene (B[b]F) to a genotoxic agent does not involve the bay region dihydrodiol epoxide 12-epoxy-9,10,11,12-tetrahydrobenzo[b]fluoranthene (B[b]F-9,10-DE). P revious results have suggested that the major DNA adduct(s) formed in vivo with B[b]F are more polar than those formed from B[b]F-9,10-DE. E mploying newly-developed synthetic methods for the formation of B[b]F derivatives, 9,10-dihydro-5,9,10-trihydroxybenzo[b]fluoranthene (5-OH- B[b]F-9,10-diol) and 9,10-dihydro-6,9,10-trihydroxybenzo[b]fluoranthen e (6-OH-B[b]F-9,10-diol) were synthesized and evaluated for tumor-init iating activity at initiation doses of 0.04, 0.12, and 0.4 mumol per m ouse. In contrast to previous bioassay results, both of these phenolic dihydrodiols exhibited tumor-initiating activity. 5-OH-B[b]F-9,10-dio l was the more potent tumor initiator, inducing a 90 % incidence of tu mor-bearing mice with an average of 7.50 tumors per mouse at a total i nitiator dose of 0.4 mumol. 6-OH-B[b]F-9,10-diol was significantly les s active as a tumor initiator in this bioassay, inducing a 50% inciden ce of tumor-bearing mice with an average of 0.60 tumor per mouse at a total dose of 0.4 mumol. P-32-Postlabeling analysis was employed to ev aluate the DNA adducts formed in vivo in mouse skin from both 5- and 6 -OH-B[b]F-9,10-diol. HPLC analysis of B[b]F-DNA adducts formed in mous e skin demonstrated the presence of a single major adduct together wit h four additional minor adducts. The DNA adducts formed with 5-OH-B[b] F-9,10-diol had identical HPLC retention to the major and one of the m inor adducts observed for B[b]F. These two DNA adducts account for 58 % of the modified nucleotides detected in mouse skin following the top ical application of B[b]F. Methods for the preparation of -hydroxy-9,1 0,11,12-tetrahydrobenzo[b]fluoranthene (5-OH-B[b]F-9,10-DE) were devel oped. The DNA adducts formed with 5-OH-B[b]F-9,10-DE were similar to t he adducts formed with 5-OH-B[b]F-9,10-diol. The results of the DNA bi nding studies along with the tumor-initiating data indicate that 5-OH- B[b]F-9,10-diol is the major proximate tumorigenic metabolite of B[b]F on mouse skin. Studies performed with deoxyguanosine 3'-phosphate sug gest that the principal adducts formed with anti-5-OH-B[b]F-9,10-DE ar e the result of the interaction with deoxyguanosine.