Urban benzene exposure and oxidative DNA damage: influence of genetic polymorphisms in metabolism genes

Abstract

Benzene has been implicated as an environmental risk factor in leukaemia and other haematological diseases. Relationships between urban benzene exposure, oxidative DNA damage and polymorphisms in metabolism enzymes were examined in 40 volunteers living and working in Copenhagen. Personal exposures to benzene, toluene and methyl tert-butyl ether (MTBE) were monitored during a 5-day period. DNA damage was measured by 7-hydro-8-oxo-2′-deoxyguanosine (8-oxodG) in lymphocyte DNA and urine and by comet assay with use of fapyguanine glycosylase (FPG) and endonuclease III (ENDO). Excretion of the benzene metabolites trans,trans-muconic acid (ttMA) and S-phenylmercapturic acid (S-PMA) were measured in urine. Polymorphisms in glutathione-S-transferases T1 (GSTT1), M1 (GSTM1) and P1 (GSTP1) and NAD(P)H:quinone oxidoreductase (NQO) were determined. Median exposures to benzene, toluene and MTBE were 2.5, 18.7 and 0.86 μg/m³. No significant correlations between external benzene exposure and any of the biomarkers were found. However, a significant correlation between S-PMA excretion and 8-oxodG in lymphocytes was found (R_s=0.39). Men were found to excrete significantly more ttMA than the women did and ttMA excretion in men was found to be significantly associated with external benzene exposure (R=0.53, P=0.025). In addition, ttMA and S-PMA excretion was significantly higher in subjects with the NQO+/−genotype compared with subjects with the wild type (P=0.004 and P=0.011, respectively). Even though there are some limitations in this study due to the low range of benzene exposure and biomarker concentrations as well as a small number of subjects, these results could suggest that even at ambient concentrations exposure to benzene could have genotoxic effects in susceptible individuals.

Introduction

In several European cities the annual average concentrations of outdoor benzene have been determined to be in the range of 1 and 50 μg/m³ depending on traffic density (Skov et al., 2001). In addition, environmental tobacco smoke is known to be an important indoor source of benzene (Wallace, 1996). Benzene has been found to be mutagenic and carcinogenic in various animal experiments and epidemiological studies, especially associated to bone marrow toxicity and leukaemia (Yin et al., 1996, Savitz and Andrews, 1997). The risk of developing leukaemia has been estimated to approximately six cases per million among people who experience lifelong exposure to benzene concentrations of 1 μg/m³ in air (WHO Working Group, 1996). This estimated risk is determined based on ambient benzene concentration. However, it has been shown that ambient benzene concentrations cannot simply be used to estimate personal exposure (Cocheo et al., 2000) and, therefore, the risk estimates are associated with a great deal of uncertainty. Biomarkers of internal and biologically effective exposure as well as susceptibility may thus be of help.

Metabolism plays an important role in benzene toxicity (Snyder and Hedli, 1996). Benzene is primarily metabolised in the liver to a variety of ring-hydroxylated and ring-opened metabolites (Snyder and Hedli, 1996). These metabolites can be transported to the bone marrow where secondary metabolism occurs which can contribute to the toxicity (Subrahmanyam et al., 1991). Several of the metabolites have been found to bind to DNA and benzene-derived quinone metabolites are known to generate reactive oxygen species (ROS) through redox cycling (Fig. 1), which can lead to oxidative damage to, e.g. DNA (Subrahmanyam et al., 1991). Experimental studies have found that exposure to benzene induce oxidative DNA damage measured by 8-oxodG in lymphocytes and bone marrow and DNA damage measured by the comet assay (Tuo et al., 1996, Tuo et al., 1999). The detoxification enzyme NQO that converts quinones into less toxic compounds through a two-electron reduction could theoretically protect against a benzene-induced oxidative stress (Moran et al., 1999) and epidemiological studies have found humans genetically deficient in NQO to be more susceptible to benzene exposure (Smith, 1999). Several intermediate benzene metabolites can be conjugated by GSTs (Fig. 1) (Snyder et al., 1993). The glutathionyl conjugate of benzene oxide is further metabolised to S-PMA and excreted in urine. GSTM1, GSTP1 and GSTT1 are polymorphic enzymes and associations with several cancer types have been found, usually with increased risk in the null genotype (Rossi et al., 1999, Autrup, 2000). With respect to benzene the GSTM1 wild type was reported to confer increased signs of bone marrow toxicity in occupationally exposed subjects, suggesting that glutathione conjugation could increase toxicity (Hsieh et al., 1999). In addition, it has been reported that excretion of the metabolite ttMA is increased in subjects with GSTT1 null genotype (Rossi et al., 1999).

The aim of this study was to investigate relationships between personal benzene exposure and biomarkers of internal as well as biologically effective dose and susceptibility in 40 volunteers living and working in Copenhagen. The internal dose was measured in terms of ttMA and S-PMA, and DNA damage was measured by 8-oxodG and strand breaks in lymphocytes as well as urinary 8-oxodG excretion. Genotypes of GSTT1, GSTM1, GSTP1 and NQO1 were also determined.