Detection of initiating potential of non-genotoxic carcinogens in a two-stage hepatocarcinogenesis study in rats

We previously reported a toxicogenomics-based prediction model for hepatocarcinogens in which the expression patterns of signature genes following repeated doses of either genotoxic or non-genotoxic compounds were similar. Based on the results of our prediction model, we hypothesized that repeated doses of non-genotoxic carcinogens might have initiating potential. Here, we conducted a two-stage hepatocarcinogenesis study in rats exposed to the initiating agent nitrosodiethylamine (DEN), and hepatotoxic compounds thioacetamide (TAA), methapyrilene (MP) and acetaminophen (APAP) for 1-2 weeks followed by the liver tumor promoter phenobarbital (PB). The duration of initial treatment was determined based on positive results from our prediction model. Combined treatment of 3 or 30 mg/kg of genotoxic DEN and PB induced marked increases in altered hepatocellular foci and a DEN dose-dependent increase in the number and area of glutathione S-transferase-placental form (GST-P)-positive foci. A low number of altered hepatocellular foci were also observed in rats treated with TAA at a dose of 45 mg/kg. MP at a dose of 100 mg/kg induced a very low number of foci, but APAP did not. Hierarchical clustering analysis using gene expression data revealed that 2-week treatment with TAA at a dose of 30 mg/kg and MP at 45 mg/kg induced specific expression of DNA damage-related genes, similar to 1-week treatment with DEN at a dose of 30 mg/kg. These results suggest that TAA and MP induce DNA damage, which partially supports our hypothesis. Although this study does not indicate whether tumor growth in response to these compounds can be assessed in this model, our results suggest that cumulative treatment with non-genotoxic TAA might have initiating potential in the liver.