Susceptibility of Human Lymphocytes to 2-Nitrofluorene-Induced Gene Mutation is Strongly Dependent on Metabolic Genotypes

Abstract

Genetic polymorphisms in drug metabolism contribute largely to adverse drug reactions in susceptible individuals due to excess production of reactive metabolites and active oxygen species. We conducted an in vitro study that clearly demonstrates that functional impact of common polymorphisms in the metabolic genes, such as N-acetyltransferase 2 (NAT2), can be easily reflected in mutant induction in the gene coding for hypoxanthineguanine phosphoribosyl transferase (HPRT) in T-lymphocytes isolated from human peripheral blood. NAT2 is involved in the metabolic activation of 2-nitrofluorene (2-NF) to the known aromatic amide carcinogen N-acetyl- 2-aminofluorene. Subsequent deactivation through glutathione conjugation involves glutathione S-transferase M1 (GSTM1). We obtained a clear dose-related increase in the HPRT mutant frequency after treating mitogen-stimulated lymphocytes isolated from a normal blood donor with 2-NF (up to 5 fold at 400 μg/mL, 24h exposure), while no HPRT mutant induction was observed using cells from another blood donor. The susceptible cells turned out to have the NAT2 rapid and GSTM1 null genotype combination (capable of activation, with insufficient deactivation), while the resistant cells had the NAT2 slow and GSTM1 positive combination. Although the contribution of GSTM1 genotype is less clear, our finding suggests that functional polymorphisms in key metabolic genes do affect induction of gene mutations at the HPRT locus and at least the NAT2 genotype plays a critical role in determining the susceptibility of human cells to genotoxicity of 2-NF. Further, human peripheral T-lymphocytes and the in vitro HPRT gene mutation assay can be utilized to study the functional impact of common genetic polymorphisms in drug metabolism and to identify risk genotypes susceptible for drug toxicity and somatic mutations.