DNA adduct formation and molecular analysis of in vivo lacI mutations in the mammary tissue of Big Blue (R) rats treated with 7,12-dimethylbenz[a]anthracene

Recently we compared the lad and Hprt mutant frequencies (MFs) and types of mutations in lymphocytes of Big Blue(R) (BB) rats exposed to 7,12-dimethyl benz[a]anthracene (DMBA) under conditions that result in mammary gland tumo rs. In this study, we have examined the target mammary tissue for DMBA-indu ced DNA adducts, lad MF and types of lad mutations. seven-week-old female B E rats were given single doses of 0, 20 or 130 mg/kg DMBA by gavage and the DNA adducts and lad MFs in the mammary tissue were measured over a period of 14 days and 18 weeks, respectively, following treatment, The lacI MF in the mammary tissue increased for 10 weeks and then remained relatively cons tant; 130 mg/kg DMBA produced a 14-fold increase in the MF (255 +/- 50 x 10 (-6) p.f.u.) over control MF (18.3 +/- 4 x 10(-6) p.f.u.). P-32-post-labeli ng analysis of DNA from mammary tissue and splenic lymphocytes of treated r ats revealed two major adducts, Comparison of these adducts with DMBA stand ards indicated that the adducts formed by DMBA involved both G:C and A:T ba se pairs. DNA sequencing revealed that the majority of DMBA-induced lad mut ations were base pair sutbstitutions and that A:T-->T:A (44% of the indepen dent mutations) and G:C-->T:A (24% of the independent mutations:) transvers ions were the predominant types. Furthermore, the mutational results reveal ed a 'hotspot' for a G-->T mutation in codon 95 (GTG-->TTG) of the lad gene in mammary tissue. These results suggest that DMBA is highly mutagenic to lad in mammary tissue and that adducts with bath G:C and A:T base pairs par ticipate in forming mutations in DMBA-treated BB rats.