Differential Mutagenic Response of Rat Liver and Lung to Nicotine-Derived Nitrosamine Ketone (NNK)

Abstract

Nitrosamines (NA) are chemical impurities that are present in tobacco, foods, more recently in some pharmaceuticals and are associated with genotoxicity and carcinogenicity. We evaluated the in vivo mutagenicity of nicotine-derived nitrosamine ketone (NNK) or 4-(methyl nitrosamino)-1-(3-pyridyl)-1-butanone, a model compound used as an anchor molecule to estimate carcinogenic potency of unknown nitrosamine impurities. Big Blue rats were treated with NNK at doses ranging from 0.001 to 30 mg/kg for 28 days, following which liver and lung tissue were harvested 3 days later for nuclear genomic DNA isolation. Mutations in liver and lung were assessed with the cII transgene assay and endogenous genomic loci using Duplex Sequencing (DupSeq), a highly validated error-corrected sequencing (ECS) technology. The no genotoxic effect level (NOGEL) was 1 mg/kg in liver and 0.1 mg/kg in lung while the benchmark dose (BMD) analysis for cII mutagenicity determined a BMDL50 of 1.3 mg/kg in liver and 0.12 mg/kg in lung, consistent with lung being the more sensitive target organ for carcinogenicity for NNK. ECS-derived mutagenicity was highly correlated with cII-derived mutagenicity. Interestingly, the types of mutations formed appeared to be tissue-specific with higher C > T transitions and lower T > G transversions in lung compared to liver, differences that may reflect tissue-specific DNA repair capacity and/or metabolic differences. Collectively, these data support the use of in vivo mutagenicity data─from both TGR cII and ECS methods─for human health and cancer risk characterization of nitrosamines and for estimating acceptable daily intakes for unknown nitrosamine drug substance related impurities.