Benzo(a)pyrene diolepoxide adducts to albumin in workers exposed to polycyclic aromatic hydrocarbons: association with specific CYP1A1, GSTM1, GSTP1 and EHPX genotypes

We investigated whether the presence of (+)-anti-benzo(a)pyrene diolepoxide adducts to serum albumin (BPDE-SA) among workers exposed to benzo(a)pyrene (BaP) and unexposed reference controls was influenced by genetic polymorphisms of cytochrome P4501A1 (CYP1A1), microsomal epoxide hydrolase (EHPX), glutathione S-transferases M1 (GSTM1) and P1 (GSTP1), all involved in BaP metabolism. Exposed workers had significantly higher levels of adducts (0.124 ± 0.02 fmol BPTmg^-1 SA, mean ± SE) and a higher proportion of detectable adducts (40.3%) than controls (0.051 ± 0.01 fmol BPT mg^-1 SA; 16.1%) (p = 0:014 and p = 0:012). Smoking increased adduct levels only in occupationally exposed workers with the GSTM1 deletion (GSTM1 null) (p = 0:034).

Smokers from the exposed group had higher adduct levels when they were CYP1A1 *1/*1 wild-type rather than heterozygous and homozygous for the variant alleles (CYP1A1 *1/*2 plus *2/*2) (p = 0:01). The dependence of BPDE-SA adduct levels and frequency on the CYP1A1 *1/*1 genotype was most pronounced in GSTM1-deficient smokers. Exposed workers with GSTM1 null/GSTP1 variant alleles had fewer detectable adducts than those with the GSTM1 null/GSTP1*A wild-type allele, supporting for the first time the recent in vitro finding that GSTP1 variants may be more effective in the detoxification of BPDE than the wild-type allele. Logistic regression analysis indicated that occupational exposure, wild-type CYP1A1*1/*1 allele and the combination of GSTM1 null genotype+EHPX genotypes associated with predicted low enzyme activity were significant predictors of BPDE-SA adducts. Though our findings should be viewed with caution because of the relatively limited size of the population analysed, the interaction between these polymorphic enzymes and BPDE-SA adducts seems to be specific for high exposure and might have an impact on the quantitative risk estimates for exposure to polycyclic aromatic hydrocarbons.     

Genetic polymorphisms in GSTM1, GSTT1, GSTP1, GSTM3 and the susceptibility to gallbladder cancer in North India

The glutathione S-transferase (GSTs) are polymorphic supergene family of detoxification enzymes that are involved in the metabolism of numerous potential carcinogens. Several allelic variants of polymorphic GSTs show impaired enzyme activity and are suspected to increase the susceptibility to various cancers. To find out the association of GST variants with risk of gallbladder cancer, the distribution of polymorphisms in the GST family of genes (GSTT1, GSTM1, GSTP1, and GSTM3) were studied in 106 cancer patients and 201 healthy controls. Genotypes were analysed by polymerase chain reaction (PCR) and PCR-restriction fragment length polymorphism (RFLP). The frequencies of GSTM1 null and GSTM3*BB genotypes did not differ between patients and controls. The overall frequency of GSTT1 null was lower in cases as compared with controls (p=0.003, Odds ratio (OR) = 0.2, 95% confidence interval (CI), 0.1-0.6). After sex stratification, the GSTT1 null frequency was reduced only in female patients (p=0.008, OR = 0.2, 95% CI = 0.1-0.6). However, the GSTP1, ile/val genotype and the val allele were significantly higher in cases than controls (p=0.013, OR = 1.9, 95% CI = 1.1-3.1; p=0.027, OR = 1.5, 95% CI = 1.0-2.1), respectively. To study gene-gene interactions, a combined risk of gallbladder cancer due to ile/val or val/val were calculated in combination with null alleles of GSTM1 and GSTT1 or the *B allele of GSTM3, but there was no enhancement of risk. Gallstones were present in 57.5% of patients with gallbladder cancer, but there were no significant differences between allelic/genotype frequencies of the studied GST genes polymorphisms between patients with or without gallstones. To best of our knowledge, this is the first paper showing ile/val genotypes and val allele of GSTP1 to be associated with higher risk of gallbladder cancer.     

GSTM1 null genotype as a risk factor for anti-BPDE-DNA adduct formation in mononuclear white blood cells of coke-oven workers

The influence of the genetic deletion polymorphism of glutathione S-transferase micro 1 (GSTM1 *0/*0) on levels of anti (+/-)-r-7,t-8-dihydroxy-t-9,10-oxy-7,8,9,10-tetrahydrobenzo[a]pyrene (anti-BPDE-DNA) adduct in the peripheral blood lymphocyte plus monocyte fraction (LMF) of coke-oven workers was investigated. A total of 95 male Polish coke-oven workers (60% current smokers) from two different plants comprised the sample population. Polycyclic aromatic hydrocarbons (PAH) exposure was assessed by means of the individual post-shift urinary excretion of 1-pyrenol (mean +/- S.D.: 6.93 +/- 7.20 micromol/mol creatinine; 70% of the subjects exceeded the proposed biological exposure index (BEI) 2.28 micromol/mol creatinine). Anti-BPDE-DNA adduct levels were detected by high performance liquid chromatography (HPLC)/fluorescence analysis of the anti-BPDE tetrol I-1 released after acid hydrolysis of DNA samples. Genotypes were determined by polymerase chain reaction (PCR) on the genomic DNA of each subject. Coke-oven workers without active GSTM1 (GSTM1 *0/*0, 33%) had significantly higher adduct levels than those with active GSTM1 (GSTM1*1/*1 and *1/*0) (5.90 +/- 5.59 versus 3.25 +/- 2.01 adducts/10(8) bases, Mann-Whitney U-test, z = 2.53, P = 0.011), PAH exposure in the two subgroups being similar (7.06 +/- 6.83 versus 6.67 +/- 8.00 1-pyrenol micromol/mol creatinine). The highest number of GSTM1 null subjects (12/23, 39%) belonged to the quartile with the highest adduct levels (i.e., >4.67 adducts/10(8) nucleotides). That is, coke-oven workers with GSTM1 *0/*0 genotype had a significantly higher risk of having high adduct levels than individuals with active GSTM1 genotype (Fisher exact test P = 0.0355; odds ratio (OR) = 4.145, 95% CI 1.0-18.8). Multiple linear regression analysis showed that the increase in anti-BPDE-DNA adduct levels in LMF was significantly related to the high occupational exposure to PAHs (benzo[a]pyrene (BaP)) of coke-oven workers (t = 3.087, P < 0.01) and to the lack of GSTM1 activity (t = 3.512, P < 0.001), rather than to the two other confounding factors of PAH intake, i.e. charcoal-broiled meat consumption and smoking habits. In conclusion, our results indicate the clear influence of the GSTM1 detoxifying genotype on anti-BPDE-DNA adduct formation in the LMF of coke-oven workers. This is invaluable for future environmental-occupational studies using this biomarker of PAH exposure.     

Interaction between smoking, GSTM1 deletion and colorectal cancer: results from the GSEC study

Cigarette smoking has inconsistently been associated with an increased risk of colorectal cancer. One of the enzymes responsible for the detoxification of the carcinogenic compounds present in tobacco smoke is glutathione S-transferase-μ (GST-μ). The gene that codes for this enzyme is GSTM1. In this study, we evaluated the associations and interaction between GSTM1 deletion, smoking behaviour and the development of colorectal cancer. We performed a pooled analysis within the International Collaborative Study on Genetic Susceptibility to Environmental Carcinogens (GSEC). We selected six studies on colorectal cancer, including 1130 cases and 2519 controls, and restricted our analyses to Caucasians because the number of patients from other races was too limited. In addition we performed a meta-analysis including the studies from the GSEC database and other studies identified on MEDLINE on the same subject. The prevalence of the GSTM1 null genotype was within the range reported in other studies: 51.8% of the cases had the GSTM1 null genotype versus 56.6% of the controls. No significant association between the GSTM1 null genotype and colorectal cancer was found (odds ratio 0.92, 95% confidence interval 0.73-1.14). Our results suggest a possible positive association between lack of the GST-μ enzyme and colorectal cancer for non-smoking women (odds ratio 1.47, 95% confidence interval 0.80-2.70). There was no interaction between the effects of smoking and GSTM1 genotype on colorectal cancer risk in men and women (χ²=0.007, p=0.97). Our findings do not support an association between the GSTM1 null genotype and colorectal cancer. In addition, we did not find any modification of the smoking-induced colorectal cancer risk by GSTM1 genotype     

CYP1A1, GSTM1 and GSTT1 polymorphisms and lung cancer: a pooled analysis of gene-gene interactions

Gene-environment interactions have been extensively studied in lung cancer. It is likely that several genetic polymorphisms cooperate in increasing the individual risk. Therefore, the study of gene-gene interactions might be important to identify high-susceptibility subgroups. GSEC is an initiative aimed at collecting available data sets on metabolic polymorphisms and the risks of cancer at several sites and performing pooled analyses of the original data. Authors of published papers have provided original data sets. The present paper refers to gene-gene interactions in lung cancer and considers three polymorphisms in three metabolic genes: CYP1A1, GSTM1 and GSTT1. The present analyses compare the gene-gene interactions of the CYP1A1*2A, GSTM1 and GSTT1 polymorphisms from studies on lung cancer conducted in Europe and the USA between 1991 and 2000. Only Caucasians have been included. The data set includes 1466 cases and 1488 controls. The only clear-cut association was found with CYP1A1*2A. This association remained unchanged after stratification by polymorphisms in other genes (with an odds ratio [OR] of approximately 2.5), except when interaction with GSTM1 was considered. When the OR for CYP1A1*2A was stratified according to the GSTM1 genotype, the OR was increased only among the subjects who had the null (homozygous deletion) GSTM1 genotype (OR=2.8, 95% CI=0.9-8.4). The odds ratio for the interactive term (CYP1A1*2A by GSTM1) in logistic regression was 2.7 (95% CI=0.5-15.3). An association between lung cancer and the homozygous CYP1A1*2A genotype is confirmed. An apparent and biologically plausible interaction is suggested between this genotype and GSTM1.     

Polymorphisms in GSTT1 and p53 and urinary transitional cell carcinoma in south-western Taiwan: A preliminary study

Little is known about the relevance of genetic polymorphisms to arsenic-related bladder cancer. A preliminary case-control study was conducted to explore the association between genetic polymorphisms of GSTT1, p53 codon 72 and bladder cancer in southern Taiwan, a former high arsenic exposure area. Fifty-nine urinary transitional cell carcinoma (TCC) patients from a referral centre in south-western Taiwan and 81 community controls matched on residence were recruited from 1996 to 1999. A questionnaire was administered to obtain arsenic exposure and general health information. Genotypes of p53 codon 72 and GSTT1 were analysed by polymerase chain reaction-restriction fragment length polymerase. The combined variant genotypes (heterozygous or homozygous variant) of p53 codon 72 and GSTT1 null were observed in 29% of cases and in 44% of controls, respectively. In this preliminary study, bladder cancer risk was slightly elevated for subjects carrying the variant genotype of p53 codon 72 or in subjects carrying the GSTT1 null genotype. Variants in p53 codon 72 increased the risk of bladder cancer among smokers. However, the results were not statistically significant and larger confirmatory studies are needed to clarify the role of candidate gene polymorphisms and bladder cancer risk in arsenic exposed populations.     

Investigation of mutant frequency at the HPRT locus and changes in microsatellite sequences in healthy young adults

In an attempt to understand the inter-individual variation that occurs in in vivo mutant frequency at the HPRT locus, we have examined the effect of polymorphisms in genes for metabolic enzymes on the mutation rate. In the same population of human volunteers, the background variant frequency in a number of microsatellite sequences was studied to determine individual variation in the capacity to repair mismatches in these sequences. The HPRT mutant frequency of T-cells isolated from a group of 49 healthy, non-smoking adults varied from 0.25 to 9.64×10⁻⁶. The frequency of polymorphisms in CYP1A1, GSTM1 and NAT2 among these individuals was similar to those published, and when subjected to univariate analysis these polymorphisms showed no influence on the HPRT mutant frequency. However, there was a significant interaction between the GSTM1 null genotype and the slow acetylator status in NAT2 (P<0.05) which was associated with higher mutant frequency. Analysis of 30 microsatellite sequences in 20 HPRT proficient clones per individual showed only six alterations in total, giving an overall mutation rate per allele of 0.01%, whilst three alterations were found in five HPRT deficient clones per individual examined for changes in 10 microsatellites, giving an overall mutation rate per allele of 0.3%. Thus, the alterations detected are probably due to background mutations and not to differences in mismatch repair capacity.     

Simultaneous determination of urinary 1- and 2-naphthols, 3- and 9-phenanthrols, and 1-pyrenol in coke oven workers

A method was developed for simultaneous quantification of urinary 1- and 2-naphthols, 3- and 9-phenanthrols and 1-pyrenol using gas chromatography with mass spectrometry (GC-MS). This method was applied to urine samples from coke oven workers (n =28) and controls (n =22) from Northern China. Geometric mean levels of urinary 1-naphthol (58.8 μg l^-1), 2-naphthol (34.1 μg l^-1), 3-phenanthrol (7.35 μg l^-1), 9-phenanthrol (1.28 μg l^-1) and 1-pyrenol (25.4 μg l^-1) were significantly higher among coke oven workers than controls. All the substances tested were highest among top-of-oven workers, who had 15-fold higher 1-naphthol, eight-fold higher 2-naphthol and 20-fold higher 1-pyrenol levels compared with controls. Using multiple linear regression models, 72.5% of the variation in 1- and 2-naphthol and 82.8% of the variation in 1-pyrenol were explained by the concentration of naphthalene or pyrene in the urine, the work category and the smoking intensity. Cigarette consumption significantly contributed to levels of urinary 1-pyrenol and naphthols, particularly 2-naphthol. A negative relationship between work category and the ratio of naphthols/1-pyrenol was observed among smokers. Our results suggest that urinary naphthols and phenanthrols reflect polycyclic aromatic hydrocarbon (PAH) exposure as well as the widely used 1-pyrenol, and that interactions between cigarette smoking and PAH exposure result in different patterns of metabolism for individual PAHs.     

Association between metabolic gene polymorphisms and susceptibility to peripheral nerve damage in workers exposed to n-hexane: A preliminary study

Chronic exposure to n-hexane may result in peripheral neuropathy. 2,5-Hexanedione (2,5-HD) has been identified as a toxic metabolite of n-hexane. The CYP2E1, CYP1A1 and GST genes are involved in the formation of 2,5-hexanedione from n-hexane as well as the elimination of 2,5-HD-formed electrophile, and these genes are highly polymorphic in the general population. A nested case-control study in an industrial cohort was conducted to evaluate the associations between polymorphisms in these metabolic genes and n-hexane-induced peripheral nerve damage. The study subjects included 22 cases, who worked in a printing factory with symptoms of peripheral nerve damage, and 163 controls, who came from the same factory of cases. DNA was extracted from blood samples and genotyping was conducted for CYP2E1 Pst, CYP2E1 Dra, CYP2E1 Ins96, CYP1A1 Msp, GSTT1 null, GSTM1 null and GSTP1 105V. Unconditional logistic regression was applied to estimate the odds ratio and 95% confidence intervals. There were no significant differences between the two groups regarding age, sex, smoking and alcohol status. A significant association between Dra polymorphism and peripheral nerve damage was found. The frequency of CYP2E1 Dra homozygous mutation in the case group (18.2%) was higher than that in the control group (3.7%, p=0.015). Individuals with homozygote genotype (CC) of CYP2E1 Dra had a significantly higher risk of peripheral nerve damage compared with those with DD genotype (adjusted OR = 5.58, 95% CI = 1.32-23.65) after n-hexane exposure duration, sex, age, smoking and alcohol status were adjusted. No significant association was found that CYP2E1 Pst, CYP2E1 Ins96, CYP1A1 Msp, GSTT1, GSTM1, GSTP gene polymorphisms associated with the susceptibility of peripheral nerve damage. These findings suggested that CYP2E1 gene might increase the susceptibility to n-hexane-induced peripheral damage.     

Total gene deletions and mutant frequency of the HPRT gene as indicators of radiation exposure in Chernobyl liquidators

This study was conducted to determine the utility of deletion spectrum and mutant frequency (MF) of the hypoxanthine phosphoribosyl transferase gene (HPRT) as indicators of radiation exposure in Russian Liquidators who served in 1986 or 1987 in the clean up effort following the nuclear power plant accident at Chernobyl. HPRT MF was determined using the cloning assay for 117 Russian Controls and 122 Liquidators whose blood samples were obtained between 1991 and 1998. Only subjects from whom mutants were obtained for deletion analysis are included. Multiplex PCR analysis was performed on cell extracts of 1080 thioguanine resistant clones from Controls and 944 clones from Liquidators. Although the deletion spectra of Liquidators and Controls were similar overall, the Liquidator deletion spectrum was heterogeneous over time. Most notable, the proportion of total gene deletions was higher in 1991–1992 Liquidators than in Russian Controls (χ²=10.5, p=0.001) and in 1993–1994 Liquidators (χ²=8.3, p=0.004), and was marginally elevated relative to 1995–1996 Liquidators (χ²=3.3, p=0.07). This type of mutation has been highly associated with radiation exposure. Total gene deletions were not increased after 1992. Band shift mutations were also increased in the 1991–1992 Liquidators but were associated with increased MF of both Liquidators and Controls (p=0.009), not with increased MF in 1991–1992 Liquidators (p=0.7), and hence are not believed to be associated with radiation exposure. Regression analysis demonstrated that relative to Russian Controls HPRT MF was elevated in Liquidators overall when adjusted for age and smoking status (37%, p=0.0001), and also was elevated in Liquidators sampled in 1991–1992 (72%, p=0.0076), 1993–1994 (22%, p=0.037), and 1995–1996 (62%, p=0.0001). In summary, HPRT MF was found to be the more sensitive and persistent indicator of radiation exposure, but the specificity of total gene deletions led to detection of probable heterogeneity of radiation exposure within the exposed population.     

Association between nucleotide excision repair gene polymorphisms and chromosomal damage in coke-oven workers

The associations between several genetic polymorphisms of nucleotide excision repair genes (NER) and chromosome damage level were studied among 140 coke-oven workers exposed to a high level of polyaromatic hydrocarbons (PAHs) and 66 non-exposed workers. Seven polymorphisms with functional potential in five NER genes (ERCC1, ERCC2, ERCC4, ERCC5 and ERCC6) were genotyped in the 206 study subjects. Multivariate analysis of covariance revealed that coke-oven workers with the ERCC1 19007 CC genotype had significantly higher cytokinesis-block micronucleus frequency (CBMN) (10.5±6.8‰) than those with CT (8.1±6.6‰, p=0.01) or TT (6.6±3.7‰, p=0.05) or CT+TT genotypes (7.5±6.3‰, p=0.004). The ERCC6 A3368G polymorphism was also associated with CBMN frequency among coke-oven workers. Subjects with the AA genotype have a significantly higher CBMN frequency (10.0±6.9‰) than those with AG (6.7±4.2‰, p=0.05) or AG+GG genotypes (6.6±4.1‰, p=0.02). Stratification analysis revealed the significant associations between ERCC1 C19007T and ERCC6 A3368G, and the CBMN frequencies were only found among older workers. In addition, a significant association between ERCC2 G23591A polymorphism and CBMN frequencies was also found among older coke-oven workers. The results suggest that polymorphisms of ERCC1 C19007T, ERCC6 A3368G and ERCC2 G23591A are associated with the CBMN frequencies among coke-oven workers     

GST, CYP and PON1 polymorphisms in farmers attributing ill health to organophosphate-containing sheep dip

Previously we reported that in sheep dippers exposed to organophosphates the frequency of paraoxonase (PON1) polymorphisms differed between those with or without self-reported ill health. We have now examined whether polymorphisms in other genes involved in xenobiotic metabolism alter disease risk in this population. There were elevated but non-significant risks associated with the CYP2D6 WT genotype (odds ratio (OR) 1.47, 95% CI 0.83-2.60), or a GSTP1*B or *C allele (OR 1.37, 95% CI 0.88-2.01) or being GSTM1*2/GSTT1*2 homozygous (OR 1.61, 95% CI 0.74-3.48). Similar results were generally obtained after the exclusion of subjects to obtain a more homogenous case-referent population: for double null GSTM1 and GSTT1 homozygotes the OR was 2.06 (95% CI 0.85-2.04). In those also likely to have been exposed to diazinon, risks associated with a GSTP1*B or *C allele (OR 1.82, 95% CI 0.92-3.63) or a GSTM1*2/GSTT1*2 homozygous (OR 2.60, 95% CI 0.72-10.42) were elevated but not to a significant extent. Risk associated with PON1 genotype and phenotype varied with CYP2D6 and GSTP1 genotype but not consistently with a priori hypotheses. Further work is necessary to delineate more clearly pathways of organophosphate activation and non-PON1 pathways of detoxification and to confirm whether CYP and GST polymorphisms alter disease risk in populations exposed to organophosphates.     

Mismatch repair deficient human cells: spontaneous and MNNG-induced mutational spectra in the HPRT gene

We have determined both the spontaneous and N-methyl-N′-nitro-N-nitrosoguanidine (MNNG)-induced mutational spectra in the HPRT gene of human cells (MT1) defective in the mismatch repair gene hMSH6 (GTBP). Eight of nine exons and nine of sixteen intronic flanking sequences were scanned, encompassing >900 bp of the HPRT gene. Mutant hotspots were detected and separated by differences in their melting temperatures using constant denaturant capillary electrophoresis (CDCE) or denaturing gradient gel electrophoresis (DGGE).

A key finding of this work is that a high proportion of all HPRT inactivating mutations is represented by a small number of hotspots distributed over the exons and mRNA splice sites. Thirteen spontaneous hotspots and sixteen MNNG-induced hotspots accounted for 55% and 48% of all 6TG^R point mutations, respectively. MNNG-induced hotspots were predominantly G:C→A:T transitions. The spontaneous spectrum of cells deficient in hMSH6 contained transversions (A:T→T:A, G:C→T:A, A:T→C:G), transitions (A:T→G:C), a plus-one insertion, and a minus-one deletion. Curiously, G:C→A:T transitions, which dominate human germinal and somatic point mutations were absent from the spontaneous hMSH6 spectra.     

Comparison of DNA adduct levels associated with oxidative stress in human pancreas

DNA adducts associated with oxidative stress are believed to involve the formation of endogenous reactive species generated by oxidative damage and lipid peroxidation. Although these adducts have been reported in several human tissues by different laboratories, a comparison of the levels of these adducts in the same tissue samples has not been carried out. In this study, we isolated DNA from the pancreas of 15 smokers and 15 non-smokers, and measured the levels of 1,N⁶-etheno(2′-deoxy)guanosine (edA), 3,N⁴-etheno(2′-deoxy)cytidine (edC), 8-oxo-2′-deoxyguanosine (8-oxo-dG), and pyrimido[1,2-]purin-10(3H)-one (m₁G). Using the same DNA, the glutathione S-transferase (GST) M1, GSTT1, and NAD(P)H quinone reductase-1 (NQO₁) genotypes were determined in order to assess the role of their gene products in modulating adduct levels through their involvement in detoxification of lipid peroxidation products and redox cycling, respectively. The highest adduct levels observed were for m₁G, followed by 8-oxo-dG, edA, and edC, but there were no differences in adduct levels between smokers and non-smokers and no correlation with the age, sex or body mass index of the subject. Moreover, there was no correlation in adduct levels between edA and eC, or between edA or edC and m₁G or 8-oxo-dG. However, there was a significant correlation (r=0.76; p<0.01) between the levels of 8-oxo-dG and m₁G in human pancreas DNA. Neither GSTM1 nor NQO₁ genotypes were associated with differences in any of the adduct levels. Although the sample set was limited, the data suggest that endogenous DNA adduct formation in human pancreas is not clearly derived from cigarette smoking or from (NQO₁)-mediated redox cycling. Further, it appears that neither GSTM1 nor GSTT1 appreciably protects against endogenous adduct formation. Together with the lack of correlation between m₁G and edA or edC, these data indicate that the malondialdehyde derived from lipid peroxidation may not contribute significantly to m₁G adduct formation. On the other hand, the apparent correlation between m₁G and 8-oxo-dG and their comparable high levels are consistent with the hypothesis that m₁G is formed primarily by reaction of DNA with a base propenal, which, like 8-oxo-dG, is thought to be derived from hydroxyl radical attack on the DNA.     

GSTM1 and GSTT1 polymorphisms as modifiers of age at diagnosis of hereditary nonpolyposis colorectal cancer (HNPCC) in a homogeneous cohort of individuals carrying a single predisposing mutation

The variability in phenotype that occurs for so-called ‘single-gene disorders’ may be because of germline alterations in numerous primary and “modifier” genes. Within HNPCC families harbouring the same primary predisposing mutation, differences exist in the site of cancer, age of onset of disease symptoms and, consequently, survival until diagnosis of disease. The current study investigated a cohort of 129 individuals, from 13 different families, who harbour the identical nonsense mutation (C1528T) in the hMLH1 gene, predisposing them primarily to Lynch I syndrome. This cohort was screened for previously described polymorphisms in the glutathione-S-transferase genes, viz. GSTT1 and GSTM1. Male null carriers for both GSTT1 and GSTM1 were approximately three times more at risk of developing cancer at an earlier age when compared to non-null males. This work, particularly because of the relatively large “homogeneous” primary mutation cohort, provides evidence that genotypic changes distinct from the primary ‘HNPCC-causing’ mutation, influence the survival period until diagnosis of disease. It provides an impetus for expanding the study to include a wider range of candidate modifier genes. Such work may potentially lead to the development of individualised interval screening regimens for individuals with varying modifier genotypes—an attractive option in a resource-poor country.     

Simultaneous determination of urinary 1- and 2-naphthols, 3- and 9-phenanthrols,and 1-pyrenol in coke oven workers

A method was developed for simultaneous quantification of urinary 1- and 2-naphthols, 3- and 9-phenanthrols and 1-pyrenol using gas chromatography with mass spectrometry (GC-MS). This method was applied to urine samples from coke oven workers (n =28) and controls (n =22) from Northern China. Geometric mean levels of urinary 1-naphthol (58.8 μg l^?1), 2-naphthol (34.1 μg l^?1), 3-phenanthrol (7.35 μg l^?1), 9-phenanthrol (1.28 μg l^?1) and 1-pyrenol (25.4 μg l^?1) were significantly higher among coke oven workers than controls. All the substances tested were highest among top-of-oven workers, who had 15-fold higher 1-naphthol, eight-fold higher 2-naphthol and 20-fold higher 1-pyrenol levels compared with controls. Using multiple linear regression models, 72.5% of the variation in 1- and 2-naphthol and 82.8% of the variation in 1-pyrenol were explained by the concentration of naphthalene or pyrene in the urine, the work category and the smoking intensity. Cigarette consumption significantly contributed to levels of urinary 1-pyrenol and naphthols, particularly 2-naphthol. A negative relationship between work category and the ratio of naphthols/1-pyrenol was observed among smokers. Our results suggest that urinary naphthols and phenanthrols reflect polycyclic aromatic hydrocarbon (PAH) exposure as well as the widely used 1-pyrenol, and that interactions between cigarette smoking and PAH exposure result in different patterns of metabolism for individual PAHs.     

The relationship between biomarkers of oxidative DNA damage, polycyclic aromatic hydrocarbon DNA adducts, antioxidant status and genetic susceptibility following exposure to environmental air pollution in humans

Polycyclic aromatic hydrocarbons (PAHs) appear to be significant contributors to the genotoxicity and carcinogenicity of air pollution present in the urban environment for humans. Populations exposed to environmental air pollution show increased levels of PAH DNA adducts and it has been postulated that another contributing cause of carcinogenicity by environmental air pollution may be the production of reactive oxygen species following oxidative stress leading to oxidative DNA damage. The antioxidant status as well as the genetic profile of an individual should in theory govern the amount of protection afforded against the deleterious effects associated with exposure to environmental air pollution. In this study we investigated the formation of total PAH (bulky) and B[a]P DNA adducts following exposure of individuals to environmental air pollution in three metropolitan cities and the effect on endogenously derived oxidative DNA damage. Furthermore, the influence of antioxidant status (vitamin levels) and genetic susceptibility of individuals with regard to DNA damage was also investigated. There was no significant correlation for individuals between the levels of vitamin A, vitamin E, vitamin C and folate with M₁dG and 8-oxodG adducts as well as M₁dG adducts with total PAH (bulky) or B[a]P DNA adducts. The interesting finding from this study was the significant negative correlation between the level of 8-oxodG adducts and the level of total PAH (bulky) and B[a]P DNA adducts implying that the repair of oxidative DNA damage may be enhanced. This correlation was most significant for those individuals that were non smokers or those unexposed to environmental air pollution. Furthermore the significant inverse correlation between 8-oxodG and B[a]P DNA adducts was confined to individuals carrying the wild type genotype for both the GSTM1 and the GSTT1 gene (separately and interacting). This effect was not observed for individuals carrying the null variant.     

Analysis of genetic variants of the poly(ADP-ribose) polymerase-1 gene in breast cancer in French patients

Previous studies have provided evidence for an association between exposure to high levels of air pollution and increased DNA damage in human sperm. In these studies DNA damage was measured using the sperm chromatin structure assay (SCSA) wherein the percentage of sperm with abnormal chromatin/fragmented DNA is determined and expressed as % DNA fragmentation index (%DFI). Here we extend these observations to address the following hypothesis: men who are homozygous null for glutathione-S-transferase M1 (GSTM1−) are less able to detoxify reactive metabolites of carcinogenic polycyclic aromatic hydrocarbons (c-PAHs) found in air pollution. Consequently they are more susceptible to the effects of air pollution on sperm chromatin. Using a longitudinal study design in which men provided semen samples during periods of both low (baseline) and episodically high air pollution, this study revealed a statistically significant association between GSTM1 null genotype and increased SCSA-defined %DFI (beta = 0.309; 95% CI: 0.129, 0.489). Furthermore, GSTM1 null men also showed higher %DFI in response to exposure to intermittent air pollution (beta = 0.487; 95% CI: 0.243, 0.731). This study thus provides novel evidence for a gene–environment interaction between GSTM1 and air pollution (presumably c-PAHs). The significance of the findings in this study with respect to fertility status is unknown. However, it is biologically plausible that increases in %DFI induced by such exposures could impact the risk of male sub/infertility, especially in men who naturally exhibit high levels of %DFI.     

Metabolic gene polymorphisms and p53 mutations in healthy centenarians and younger controls

To obtain insights into the genetic mechanisms of ageing, we studied the frequency of the simultaneous presence of polymorphisms in phase I and phase II genes and of several p53 germline mutations in a group of 66 nonagenarians and centenarians in good health, selected from a larger sample of a multicentre Italian study in Northern Italy, and in a sample of 150 young healthy volunteers of the same ethnic group. We found a statistically significant difference in the frequency of the GSTT1 deletion and the p53 genotypes: the absence of any p53 polymorphisms and of GSTT1 deletion, and the simultaneous presence of the three p53 polymorphisms and of GSTT1 deletion, were much more frequent in young subjects than in centenarians (41.5% versus 26.9% and 8.8% versus 3.8%, respectively). One hypothesis to explain this difference is that subjects with both GSTT1 deletion and p53 polymorphisms may accumulate carcinogens and may have reduced DNA repair ability, and thus are more at risk for cancer. Another possible explanation is that both metabolic genes and p53 act on pathways related to cell ageing and death, and therefore certain composite genetic patterns could represent a generic mechanism of protection against ageing, not just against the development of chronic diseases. It is likely that longevity is related to a complex genetic trait as well as to certain environmental exposures.