DNA damage and mutagenesis induced by procarbazine in lambda lacZ transgenic mice: evidence that bone marrow mutations do not arise primarily through miscoding by O6-methylguanine.

The DNA damaging and mutagenic activities of procarbazine, a methylating drug employed in cancer chemotherapy and suspected of causing therapy-related leukaemia, were investigated in the liver and bone marrow of λlacZ transgenic mice (Muta TM Mouse). The drug was administered using two different protocols, a 'high-dose' one involving 5 daily doses of 200 mg/kg, expected to cause depletion of the repair enzyme O 6 -alkylguanine-DNA alkyltransferase (AGT) and thus favour the selective accumulation of the premutagenic lesion O 6 -methylguanine (O 6 -meG) relative to other adducts, and a 'low-dose' one involving 10 daily doses of 20 mg/kg procarbazine. Substantial accumulation of O 6 -meG was observed in both tissues examined 6h after the end of the 'high-dose' treatment, with the liver accumulating somewhat higher levels than the bone marrow (28.0 ± 1.8 fmol/μg DNA and 18.5 ± 1.1 fmol/μg DNA respectively). However, significant increases in mutant frequency 10 days after the end of treatment were observed only in the bone marrow, reaching a 16-fold increase over background following the 5 x 200 mg/kg treatment. Sequence analysis of the mutations induced after this treatment revealed a mixed spectrum, in which G:C→A:T transitions (characteristic of O 6 -meG miscoding) were only a secondary feature: Among 20 mutants analysed, only six such mutations were found, including three at CpG sites, which might have arisen from deamination of 5-methylcytosine. The other mutations observed included 1 A:T→G:C transition, five transversions (one G:C→T:A, one double G:C→C:G, two A:T→T:A, one A:T→C:G), five deletions and three insertions. The mechanistic and clinical significance of these findings is discussed.