IN-VIVO MUTAGENESIS INDUCED BY CC-1065 AND ADOZELESIN DNA ALKYLATION IN A TRANSGENIC MOUSE MODEL

Although considerable work has focused on characterizing the bonding c hemistry and sequence selective alkylation of DNA by cyclopropylpyrrol oindole compounds, little is known about the molecular consequence of their N-3-adenine adducts in whole animal systems. We have utilized a transgenic mouse system, harboring a lambda phage shuttle vector, to a ssess the mutagenic potential of the antitumor compounds CC-1065 and a dozelesin and, for the first time, to track the in vivo fate of their unique DNA modifications at the nucleotide level. Mice were inoculated with a single therapeutic dose of these agents and sacrificed at eith er 18 h, 3 days, or 15 days for extraction and analysis of liver DNA. Mutant frequencies obtained from drug treated and control animals were determined by in vitro packaging of the phage vector from genomic DNA followed by a colorimetric plaque assay to screen for phage in which the accompanying lacI repressor gene had mutated. Although undetectabl e at 18 h posttreatment, by 72 h a 3-fold increase in mutant frequency was observed in drug treated animals such that sequence analysis of d rug induced mutations could be performed and a direct comparison made between in vitro and in vivo DNA alkylation. Base substitution involvi ng guanine or cytosine accounted for 64% of the 41 mutations sequenced from drug treated animals. Only 7 of the mutations occurred at a cycl opropylpyrroloindole alkylation site while 23 occurred 1 to 4 nucleoti des from a potentially alkylated adenine.