The association between glutathione S-transferase M1 genotype and polycyclic aromatic hydrocarbon-DNA adducts in breast tissue.

A major goal in molecular epidemiology is to identify preventable environmental risk factors and susceptible subpopulations. In a hospital-based molecular epidemiological case-control study of breast cancer, we investigated the relationship between DNA damage from exposure to polycyclic aromatic hydrocarbons (PAHs) and susceptibility attributable to inherited deletion of the xenobiotic detoxifying gene, glutathione S-transferase M1 (GSTM1). Prior to breast surgery, women (n = 227) were enrolled and interviewed and donated a blood sample. PAH-DNA adduct levels were measured by immunohistochemistry in breast tissue samples retrieved from pathology blocks, and GSTM1 genotype was determined by PCR using WBC DNA. The GSTM1 analysis included 95 cases and 87 benign breast disease controls. GSTM1 genotype was not associated with breast cancer case-control status (odds ratio = 0.73; 95% confidence interval, 0.37-1.44). However, the GSTM1 null genotype predicted PAH-DNA adduct levels in malignant (beta = 0.407; P = 0.003) and nonmalignant (beta = 0.243; P = 0.05) breast tissue from cases. This relationship was not seen in tissue from controls (beta = 0.095; P = 0.341). When tissue from controls was compared with tumor tissue from cases, there was a significant case-control difference in PAH-DNA adduct levels among women who were GSTM1 null. There was no such case-control difference among women who were homozygous or heterozygous for GSTM1. There was an interaction between GSTM1 and case-control status on adduct levels in breast tissue (P = 0.002). The results suggest that genetic susceptibility to the formation of PAH-DNA adducts in breast tissue may play a role in breast cancer development.     

GSTM1, GSTP1, CYP1A1 and CYP2D6 polymorphisms in lung cancer patients from an environmentally polluted region of Poland: correlation with lung DNA adduct levels.

The CYP and GST genetic polymorphisms, controlling metabolism of xenobiotics, are considered to influence an individual's susceptibility to environmental and occupational carcinogens and predisposition to cancer. In the study, the effect of the GSTM1, GSTP1, CYP1A1 and CYP2D6 polymorphisms was investigated in relation to PAH-DNA adduct levels in non-tumourous lung tissue from non-small cell lung cancer (NSCLC) patients living in the industrialized region of Upper Silesia, Poland. The level of adducts among smokers was significantly elevated when compared to non-smokers (P = 0.0005). Adduct levels correlated inversely with age of patients (P = 0.00001). The GSTP1 and CYP2D6 polymorphisms had no influence on DNA adduct levels. There was a significant relationship between high adduct levels and the combined GSTM1 (null)/CYP1A1-Ile/Val genotype in the squamous cell carcinoma group (P = 0.028). An elevated number of adducts was found in patients with the GSTM1 (null)/CYP1Al-Ile/Val genotype compared to the GSTM1 (null)/CYP1A1-Ile/Ile carriers (P = 0.043). A higher frequency of the CYP1A1-Ile/Val and GSTM1 (null)/CYP1A1-Ile/Val genotypes was observed in patients with high adduct levels (P = 0.05 and P = 0.009, respectively). A significant prevalence of the GSTM1(null)/CYP1A1-Ile/Val carriers in the adenocarcinoma group was found (P = 0.003). Thus, our findings imply that the GSTMI and CYP1A1 exon 7 polymorphisms may influence PAH-DNA adduct levels in target tissue from NSCLC patients, especially in the squamous cell carcinoma group. Moreover, individuals carrying the GSTM1(null)/CYP1A1-Ile/Val genotype might exhibit a greater predisposition to a peripheral type of lung cancer.     

Polycyclic aromatic hydrocarbon-DNA adducts in prostate cancer

The formation of DNA adducts can lead to DNA replication errors and the potential for carcinogenesis. DNA adducts have been detected in prostate cells, but the distribution of adducts with respect to prostate cancer risk factors and histology is unknown. In a study of 130 Caucasian (n = 61) and African-American (n = 69) men with prostate cancer who underwent radical prostatectomy, we quantified polycyclic aromatic hydrocarbon (PAH)-DNA adducts in prostate tumor and adjacent nontumor cells by immunohistochemistry. A strong correlation between paired adduct levels in the two cell types was observed (r = 0.56; P < 0.0001); however, nontumor cells had a significantly higher level of adducts compared with tumor (0.30 absorbance units +/- 0.05 versus 0.17 absorbance units +/- 0.04; P < 0.0001). Variables significantly associated with PAH-DNA adduct levels in tumor cells included primary Gleason grade, tumor volume, and log-transformed prostate-specific antigen (PSA) at time of diagnosis. Tumors with a primary Gleason grade of 5 had significantly lower PAH-DNA adduct levels than tumor cells with a primary Gleason grade of 3 or 4 (P < 0.0001 for both). Tumors that involved 10% or less of the prostate gland had significantly higher PAH-DNA adduct levels than tumors that involved 15 to 20% of the prostate gland (P = 0.004). PSA levels were inversely associated with PAH-DNA adduct levels in tumor cells (P = 0.009). A similar, albeit less significant, inverse association was observed between PSA and PAH-DNA adduct levels in nontumor cells (P = 0.07). Interestingly, increasing primary Gleason grade was associated with increasing PAH-DNA adduct levels in adjacent nontumor cells (P = 0.008). Our results show that PAH-DNA adducts are present in the prostate but vary with regard to cellular histology. In prostate tumor cells, decreased cellular differentiation and increased tumor proliferation may reduce PAH-DNA adduct levels.     

Genetic polymorphisms of glutathione S-transferases M1 and T1 associated with susceptibility to aflatoxin-related hepatocarcinogenesis among chronic hepatitis B carriers: a nested case-control study in Taiwan

This study was conducted to investigate the modifying effect of glutathione S-transferase (GST) M1 and T1 polymorphisms on aflatoxin-induced hepatocarcinogenesis among chronic hepatitis B virus surface antigen (HBsAg) carriers. A total of 79 HBsAg-positive cases of hepatocellular carcinoma (HCC) diagnosed between 1991 and 1997 were identified and individually matched to one or two HBsAg-positive controls on age, gender, residence and date of recruitment from the same cancer screening cohort in Taiwan. Blood samples were tested for hepatitis B and C viral markers by enzyme immunoassay and for aflatoxin B(1) (AFB(1))-albumin adducts by competitive enzyme-linked immunosorbent assay. GSTM1 and GSTT1 genotypes were determined by PCR. There was a statistically significant relationship between detectable levels of AFB(1)-albumin adducts in serum and risk of HCC among chronic HBsAg carriers, with an adjusted odds ratio (OR) of 2.0 [95% confidence interval (CI) 1.1-3.7]. In addition, the effect of aflatoxin exposure on HCC risk was more pronounced among chronic HBsAg carriers with the GSTT1 null genotype (OR 3.7, 95% CI 1.5-9.3) than those who were non-null (OR 0.9, 95% CI 0.3-2.4). The interaction between serum AFB(1)-albumin adduct level and GSTT1 genotype was statistically significant (P = 0.03). For GSTM1 the effect of aflatoxin exposure on HCC risk in those with the null genotype was also greater (adjusted OR 2.8, 95% CI 1.0-7.8) than in those with the gene present (adjusted OR 1.8, 95% CI 0.8-4.5), but the difference was not significant (P = 0.91). Notably, when the interaction between aflatoxin exposure and GSTT1 genotype was considered, aflatoxin exposure by itself was not a significant determinant of HCC risk among chronic HBsAg carriers. These results demonstrate the importance of gene-environment interactions in the multifactorial development of HCC.     

Leukocyte polycyclic aromatic hydrocarbon-DNA adduct formation and colorectal adenoma

Consumption of charbroiled red meat and meat-derived polycyclic aromatic hydrocarbons (PAHs) has been associated with risk of colorectal adenoma, a precursor of colorectal cancer. Furthermore, leukocyte PAH-DNA adduct levels have been demonstrated to increase in response to charbroiled red meat intake but to date there have been no studies that have investigated the relationship between leukocyte PAH-DNA adduct levels and risk of colorectal adenoma. We investigated the relation of leukocyte PAH-DNA adduct formation and colorectal adenoma in a clinic-based case-control study of colorectal adenomas. The study comprised 82 cases of colorectal adenoma and 111 polyp-free controls, none of whom were current smokers. Leukocyte PAH-DNA adducts were measured by a sensitive chemiluminescence immunoassay using an antiserum elicited against DNA modified with (+/-)-7beta,8alpha-dihydroxy-9alpha,10alpha-epoxy-7,8,9,10-tetrahydro-benzo[a]pyrene that recognizes several PAHs bound to human DNA. Leukocyte PAH-DNA adduct levels were higher among colorectal adenoma cases (median, 1.4 adducts per 10(8) nucleotides) than polyp-free controls (median, 1.2 adducts per 10(8) nucleotides) (P = 0.02). There was a positive association between PAH-DNA adduct level and adenoma prevalence: each unit increase in PAH-DNA adduct level (per 10(8) nucleotides) was associated with an odds ratio (OR) of 1.5 [95% confidence interval (CI), 1.1-2.2]. In addition, a comparison of the lowest quartile for PAH-DNA adduct level with the highest quartile yielded an OR of 2.8 (95% CI, 1.2-6.5; P(trend) = 0.048) for risk of colorectal adenoma. These data support a link between PAH exposure and colorectal adenoma.     

Associations between smoking, polymorphisms in polycyclic aromatic hydrocarbon (PAH) metabolism and conjugation genes and PAH-DNA adducts in prostate tumors differ by race.

Polycyclic aromatic hydrocarbon (PAH)-DNA adducts may induce mutations that contribute to carcinogenesis. We evaluated potential associations between smoking and polymorphisms in PAH metabolism [CYP1A1 Ile 462Val, CYP1B1 Ala 119Ser and Leu 432Val, microsomal epoxide hydrolase (mEH) Tyr 113His and His139Arg, CYP3A4 A(-392)G] and conjugation [glutathione S-transferase (GST) M1 null deletion, GSTP1 Ile 105Val] genes and PAH-DNA adduct levels (measured by immunohistochemistry) in tumor and nontumor prostate cells in 400 prostate cancer cases. Although no statistically significant associations were observed in the total sample, stratification by ethnicity revealed that Caucasian ever smokers compared with nonsmokers had higher adduct levels in tumor cells (mean staining intensity in absorbance units +/- SE, 0.1748 +/- 0.0052 versus 0.1507 +/- 0.0070; P = 0.006), and Caucasians carrying two mEH 139Arg compared with two 139His alleles had lower adducts in tumor (0.1320 +/- 0.0129 versus 0.1714 +/- 0.0059; P = 0.006) and nontumor (0.1856 +/- 0.0184 versus 0.2291 +/- 0.0085; P = 0.03) cells. African Americans with two CYP1B1 432Val compared with two 432Ile alleles had lower adducts in tumor cells (0.1600 +/- 0.0060 versus 0.1970 +/- 0.0153; P = 0.03). After adjusting for smoking status, carrying the putative "high-risk" genotype combination, the faster metabolism of PAH-epoxides to PAH-diol-epoxides (CYP1B1 432Val/Val and mEH 139Arg/Arg) with lower PAH-diol-epoxide conjugation (GSTP1 (105)Ile/Ile), was associated with increased adducts only in Caucasian nontumor cells (0.2363 +/- 0.0132 versus 0.1920 +/- 0.0157; P= 0.05). We present evidence, for the first time in human prostate that the association between smoking and PAH-DNA adducts differs by race and is modified by common genetic variants.     

Plasma antioxidant vitamins, chronic hepatitis B virus infection and urinary aflatoxin B1-DNA adducts in healthy males

Epidemiological evidence indicates that aflatoxin B1 (AFB1) intake is associated with an increased risk of hepatocellular carcinoma (HCC). The hepatocarcinogenesis is initiated by covalent binding of AFB1 to cellular DNA. To determine whether nutritional factors and hormonal status may influence the binding of AFB1 to hepatic DNA, a cross- sectional study was performed on a total of 42 male asymptomatic hepatitis B surface antigen (HBsAg) carriers and 43 male non-carriers in a cohort study on the multistage development of HCC in Taiwan. The major AFB1-DNA adduct in vivo, AFB1-N7-guanine, was measured by high- performance liquid chromatography in urine. Urinary AFB1-N7-guanine was detectable in 40% of the subjects. HBsAg carriers had a higher detection rate of urinary AFB1-DNA adducts than non-carriers and the difference was statistically significant after multivariate adjustment. After taking into account the total AFB1 urinary metabolite level, chronic HBsAg carrier status, and other potential confounders, plasma levels of cholesterol, alpha-tocopherol, and alpha- and beta-carotene were positively associated with the detection rate of the AFB1-DNA adducts in a dose-dependent manner, whereas plasma lycopene level was inversely related to the presence of the adducts in urine. The association of urinary AFB1-DNA adducts with the plasma levels of cholesterol, alpha-tocopherol, lycopene, and alpha- and beta-carotene was observed at both low and high exposure levels of AFB1. There was a synergistic interaction of plasma alpha-tocopherol with alpha- and beta- carotene on the adduct levels. No association with the adducts was found for plasma levels of retinol and testosterone. This study demonstrated different associations of antioxidant vitamins with AFB1- DNA adduct formation. The data consistent with our previous finding in cultured woodchuck hepatocytes that alpha-tocopherol and beta-carotene enhanced AFB1-DNA adduct formation suggest that prospective investigation of the relationship between plasma micronutrients and risk of AFB1-related HCC is warranted.      

GSTM1 and XRCC3 Polymorphisms： Effects on Levels of Aflatoxin B1-DNA Adducts

Objective  Aflatoxin B1 (AFB1), which can cause the formation of AFB1-DNA adducts, is a known human carcinogen. AFB1-exposure individuals with inherited susceptible carcinogen-metabolizing or repairing genotypes may experience an increased risk of genotoxicity. This study was designed to investigate whether the polymorphisms of two genes, the metabolic gene Glutathione S-transferase M1 (GSTM1) and DNA repair gene x-ray repair cross-complementing group 3 (XRCC3), can affect the levels of AFB1-DNA adducts in Guangxi Population (n= 966) from an AFB1-exposure area. Methods  AFB1-DNA adducts were measured by ELISA, and GSTM1 and XRCC3 codon 241 genotypes were identified by PCR-RFLP. Results  The GSTM1-null genotype [adjusted odds ratio (OR) = 2.09; 95% confidence interval (CI) = 1.61–2.71] and XRCC3 genotypes with 241 Met alleles [i.e., XRCC3-TM and -MM, adjusted ORs (95% CI) were 1.43 (1.08–1.89) and 2.42 (1.13–5.22), respectively] were significantly associated with higher levels of AFB1-DNA adducts. Compared with those individuals who did not express any putative risk genotypes as reference (OR = 1), individuals featuring all of the putative risk genotypes did experience a significantly higher DNA-adduct levels (adjusted ORs were 2.87 for GSTM1-null and XRCC3-TM; 5.83 for GSTM1-null and XRCC3-MM). Additionally, there was a positive joint effect between XRCC3 genotypes and long-term AFB1 exposure in the formation of AFB1-DNA adducts. Conclusion  These results suggest that individuals with susceptible genotypes GSTM1-null, XRCC3-TM, or XRCC3-MM may experience an increased risk of DNA damage elicited by AFB1 exposure. This work was supported by the National Natural Science Foundation of China (No.39860032), and the Youth Science Foundation of Guangxi (No.0833097).     

Aflatoxin B1-DNA adducts and hepatitis B virus antigens in hepatocellular carcinoma and non-tumorous liver tissue.

Studies were carried out to test the hypothesis that exposure to aflatoxin B1 (AFB1) is common among individuals with hepatocellular carcinoma (HCC) who are also chronically infected with hepatitis B virus (HBV). Experiments were also carried out to determine whether there is a close association between the presence of AFB1-DNA adducts and the expression of one or more HBV antigens in the tumor or non-tumor regions of the liver. Twenty-seven paired tumor and non-tumor liver tissues of HCC patients from Taiwan were analyzed. Monoclonal antibody 6A10, generated against the imidazole ring-opened persistent form of the major N-7 guanine adduct of AFB1, was used for adduct detection by both indirect immunofluorescence and competitive enzyme-linked immunosorbent assay. An avidin-biotin complex staining method was used for the detection of HBsAg and HBxAg in liver sections. A total of 8 (30%) HCC samples and 7 (26%) adjacent non-tumor liver tissue samples from Taiwan were positive for AFB1-DNA adducts. For HBsAg, 10 (37%) HCC samples and 22 (81%) adjacent non-tumorous liver samples were positive while 9 (33%) HCC samples and 11 (41%) adjacent non-tumor liver samples were HBxAg-positive. No association with AFB1-DNA adducts was observed for HBsAg and HBxAg. These results suggest that both AFB1 exposure and carrier status of HBsAg/HBxAg may be involved in the induction of HCC in Taiwan.     

Comparison of DNA adducts and sister chromatid exchange in lung cancer cases and controls

In a molecular epidemiological study of lung cancer cases (n = 81) and noncancer controls (n = 67), polycyclic aromatic hydrocarbon (PAH)-DNA adducts were evaluated in peripheral blood leukocytes from all subjects and in a smaller number of lung tissue specimens collected prior to or at surgery. Sister chromatid exchanges (SCE) in lymphocytes were also studied in a subset of cases and controls. Questionnaire, medical record, or tumor registry data provided a family history of cancer, as well as information on cigarette smoking, dietary and occupational exposure to PAHs, and other factors related to SCEs. In both cases and controls PAH-DNA adducts in leukocytes measured by an enzyme-linked immunosorbent assay were not significantly related to age, sex, ethnicity, amount of cigarette smoking, passive smoking, dietary charcoal, or caffeine consumption. Nor did family history of cancer or histological type of cancer significantly affect adduct levels. However, when subjects were stratified by smoking status (current, former, and nonsmoker), lung cancer cases who were current smokers had significantly higher levels of covalent adducts than current smoker controls. A seasonal variation was observed in PAH-DNA binding, with a peak in adduct levels during July-October. This peak corresponds to that seen in a prior study of aryl hydrocarbon hydroxylase inducibility by other investigators. The finding of significant levels of PAH-DNA adducts in former smokers and non-smokers supports an earlier observation that this marker is not smoking specific but reflects a pervasive and variable "background" exposure to PAH. These results are consistent with a genetically determined enhancement of PAH-DNA adduct formation in leukocytes of lung cancer cases which is evident in current smokers. The results in lung tissue are limited by the small number of samples. Adduct levels were not significantly increased in lung tissue of smokers compared with nonsmokers. An inverse linear correlation was seen between adduct values in lung tissue and age of the donors. SCEs were significantly related to pack years of smoking. However, there was no difference in the frequency of SCE between cases and controls; nor were SCE and DNA adducts significantly correlated in this small sample.     

The impact of glutathione s-transferase M1 and cytochrome P450 1A1 genotypes on white-blood-cell polycyclic aromatic hydrocarbon-dna adduct levels in humans

Carcinogenic polycyclic aromatic hydrocarbons(a) form DNA adducts via a complex metabolic activation pathway that includes cytochrome P450(a) 1A1, whereas intermediate metabolites can be detoxified by conjugation through pathways including glutathione s-transferase M1 (GSTM1). PAH-DNA adducts can be measured in peripheral white blood cells (a) and should reflect the net effect of competing activation and detoxification pathways and ONA repair as well as exposure. We have previously shown that WBC PAH-DNA adducts measured by an enzyme-linked immunosorbent assay (a) were associated with recent, frequent consumption of charbroiled food among 47 nonsmoking wildland fire-fighters who provided two blood samples 8 wk apart. In the investigation reported here, which was performed in the same population, we measured the association between the GSTM1 null genotype, which results in loss of enzyme activity, and PAH-DNA adduct levels, hypothesizing that subjects with this genotype would have higher levels of DNA adducts because of their decreased ability to detoxify PAH metabolites. However, PAH-DNA adduct levels were nonsignificantly lower in subjects with the GSTM1 null genotype (n = 28) compared with other subjects (n=19) (median 0.04 fmol/μg DNA vs 0.07 fmol/μg DNA, respectively, P = 0.45, Wilcoxon rank-sum test). Adduct levels were also lower in the nine subjects heterozygous or homozygous for the CYP1A1 exon 7 polymorphism (which codes for a valine rather than isoleucine and is thought to be associated with greater CYP1A1 activity) compared with the 38 wild-type subjects (P = 0.12). In the entire group, there was a positive association between consuming charbroiled food and PAH-DNA adduct formation (r = 0.24, P = 0.02, Spearman rank-order correlation). This association was weaker in the subgroup of subjects with the GSTM1 null genotype (r = 0.03, P = 0.84) and stronger among the remaining subjects (r = 0.57, P = 0.0002). These results suggest that the GSTM1 null genotype and CYPT1A1 exon 7 polymorphism are not associated with increased susceptibility for PAH-DNA adduct formation in peripheral WBCs measured by ELISA in nonsmoking populations.© 1995 Wiley-Liss, Inc     

DNA adducts in tumour, normal peripheral lung and bronchus, and peripheral blood lymphocytes from smoking and non-smoking lung cancer patients: correlations between tissues and detection by 32P-postlabelling and immunoassay

Smoking is a major risk factor for lung cancer. This comparative study of smoking-related carcinogen-DNA adducts in pulmonary tissues and peripheral blood lymphocytes aims to further explore the primary DNA damaging processes by cigarette smoke in target and surrogate tissues. Samples of tumour and normal peripheral lung tissue, normal bronchial tissue and peripheral blood lymphocytes were obtained from a total of 85 lung cancer patients who underwent lung resection. Bulky DNA adducts were determined by 32P-postlabelling, and polycyclic aromatic hydrocarbon (PAH)-DNA adducts were detected by (+/-)-7beta, 8alpha-dihydroxy-9alpha,10alpha-epoxy-7,8,9,10-tetrahydrobenzo[a]pyrene-DNA chemiluminescence immunoassay (BPDE-DNA CIA) in smaller subsets of tissue samples subject to availability of DNA. Bulky DNA adduct levels ranged between 0.3 and 27.8 adducts/10(8) nucleotides (nt) with mean adduct levels between 2.8 and 11.5 adducts/10(8) nt. Mean PAH-DNA adduct levels were 2.6-6.2 adducts/10(8) nt. Significantly higher bulky DNA adduct levels were detected in smokers' lungs as compared with non-smokers' (P < 0.02). PAH-DNA adduct levels appeared higher in the lungs of smokers compared with non-smokers but the difference was not significant. Lung tumour contained on average a 50% lower DNA adduct level compared with normal lung tissue. A statistically significant positive correlation was found between the DNA adduct levels of the corresponding tumour and normal lung tissue samples in both smokers and non-smokers using both methodologies. Bulky DNA adduct levels in normal lung and blood lymphocytes correlated significantly in non-smokers only (r = 0.55, P = 0.023). In lung tumour DNA samples there was a weak correlation between values obtained by 32P-postlabelling and by the BPDE-DNA immunoassay (r = 0.27, P = 0.054). However, with normal lung DNA samples, values obtained by the two assays did not correlate.     

Smoking-associated bulky DNA adducts in bronchial tissue related to CYP1A1 MspI and GSTM1 genotypes in lung patients

Relationships between smoking status and levels of bulky DNA adducts were investigated in bronchial tissue of lung patients in relation to their GSTM1 and CYP1A1 MspI genotypes. A total of 150 Hungarian patients undergoing pulmonary surgery were included in the study, 124 with lung malignancies and 26 with non-malignant lung conditions. There were significant relationships between smoking status and bulky DNA adduct levels, as determined by 32P-post-labelling analysis, in macroscopically normal bronchial tissues. There was a highly significant difference in the adduct levels of a combined group consisting of current smokers and short-term ex-smokers (< or = 1 year abstinence) compared with life-time non-smokers and long-term ex- smokers (> 1 year abstinence) (P = 0.0001). The apparent half-life was estimated to be 1.7 years for bulky DNA adducts in the bronchial tissue from ex-smokers. There were no statistically significant correlations between (i) daily cigarette dose and DNA adduct levels in current smokers, (ii) DNA adduct level and histological type of lung cancer, or (iii) GSTM1 and CYP1A1 MspI genotypes and DNA adduct levels after adjustment for either smoking status or malignancy. By multiple logistic regression analysis, smoking and GSTM1 null genotype were found to be risk factors for squamous cell carcinoma. However, bulky DNA adduct levels in bronchial tissue did not appear to be a statistically-significant risk factor for the major histological types of lung cancer.      

淋巴瘤患者多环芳烃-DNA加合物及代谢酶基因细胞色素氧化酶P4501A1与谷胱苷肽硫转移酶M1基因多态性的研究

目的 探讨多环芳烃（PAH）-DNA加合物和代谢酶基因细胞色素氧化酶P4501A1（CYP1A1）与谷胱苷肽硫转移酶M1（GSTM1）基因多态性与淋巴瘤发病的关系.方法 通过竞争性酶联免疫吸附法测定54例淋巴瘤患者及34例对照组骨髓液中PAH-DNA加合物含量,采用限制性片段长度多态性PCR法（PCR-RFLP）测定上述标本CYP1A1、GSTM1基因多态性.结果 淋巴瘤患者骨髓液中PAH-DNA加合物含量为（2498±1 250） pg/ml,较对照组的（1 882±797） pg/ml增加,差异有统计学意义（t=0.006,P＜0.05）;淋巴瘤患者GSTM1基因型缺失占85.2％,对照组占58.8％,差异有统计学意义（x2=7.73,P＜0.05）,GSTM1基因型缺失者患淋巴瘤风险是GSTM1表达者的4.03倍（95％ CI1.51～10.76,P＜ 0.05）;CYP1A1变异型是野生型患淋巴瘤风险的1.36倍（95％CI 0.56～ 3.31,P＞ 0.05）,GSTM1缺失者PAH-DNA加合物水平≥2 200 pg/ml时患淋巴瘤的危险性增加（OR=9.53,95％CI 2.397～ 37.990,P＜ 0.05）.结论 PAH-DNA加合物可能参与淋巴瘤的发病,GSTM1缺失与淋巴瘤发生有关,并且增加患淋巴瘤的风险.     

Biomarkers of polycyclic aromatic hydrocarbon-DNA damage and cigarette smoke exposures in paired maternal and newborn blood samples as a measure of differential susceptibility.

Human and experimental evidence indicates that the developing fetus may be more susceptible than the adult to the effects of certain carcinogens, including some polycyclic aromatic hydrocarbons (PAHs). Factors that can modulate susceptibility include proliferation rates, detoxification capabilities, and DNA repair capacity. Biomarkers can facilitate quantification of age-related susceptibility among human populations. In this study, we report on three biomarkers measured in paired blood samples collected at birth from 160 Polish mothers and newborns: 70 pairs from Krakow (a city with high air pollution including PAHs) and 90 pairs from Limanowa (an area with lower ambient pollution but greater indoor coal use). Biomarkers were: WBC aromatic-DNA adducts by (32)P-postlabeling and PAH-DNA adducts by ELISA (as indicators of DNA damage from PAHs and other aromatics) and plasma cotinine (as an internal dosimeter of cigarette smoke). Correlations were assessed by Spearman's rank test, and differences in biomarker levels were assessed by the Wilcoxon signed-ranks test. A significant correlation between paired newborn/maternal samples was seen for aromatic-DNA adduct levels (r = 0.3; P < 0.001) and plasma cotinine (r = 0.8; P < 0.001) but not PAH-DNA adduct levels (r = 0.14; P = 0.13). Among the total cohort, levels of the three biomarkers were higher in newborn samples compared with paired maternal samples. The difference was significant for aromatic-DNA adduct levels (16.6 +/- 12.5 versus 14.21 +/- 15.4/10(8) nucleotides; P = 0.002) and plasma cotinine (14.2 +/- 35.5 versus 8.3 +/- 24.5 ng/ml; P < 0.001) but not for PAH-DNA adduct levels (7.9 +/- 9.9 versus 5.9 +/- 8.2/10(8) nucleotides; P = 0.13). When analyses were restricted to the 80 mother/newborn pairs from whom the blood sample was drawn concurrently (within 1 h of each other), levels of all of the three biomarkers were significantly higher in the newborn compared with paired maternal blood samples (P < 0.05). Results suggest reduced detoxification capabilities and increased susceptibility of the fetus to DNA damage, especially in light of experimental evidence that transplacental exposures to PAHs are 10-fold lower than paired maternal exposures. The results have implications for risk assessment, which currently does not adequately account for sensitive subsets of the population.     

Polycyclic aromatic hydrocarbon--DNA adducts in white blood cells from lung cancer patients: no correlation with adduct levels in lung.

Smokers of cigarettes are exposed to a number of carcinogens, including polycyclic aromatic hydrocarbons (PAHs), and are at a high risk for lung cancer. PAHs exert their carcinogenic activity after metabolic activation to reactive intermediates that can damage DNA through adduct formation. Measuring DNA adducts in peripheral white blood cells (WBC) could serve as a means of monitoring human exposure to genotoxic agents and subsequently risk assessment. In this study, DNA from WBC obtained from 39 lung cancer patients was examined for PAH-DNA adducts both in an ELISA using a polyclonal antibody against benzo[a]pyrene 7,8-diol-9,10-epoxide (BPDE)-DNA and the 32P-post-labeling technique. The ELISA results showed BPDE-DNA antigenicity in WBC DNA from 12/38 (32%) patients and adduct levels ranged from 1.5 to greater than 150 adducts in 10(8) nucleotides. The autoradiographs of chromatograms of 32P-post-labeled digests of WBC DNA from the 38 patients showed a variety of adduct spots; relative adduct labeling (RAL) values ranged from 0.3 to 407 adducts in 10(8) nucleotides. In 18 of the 38 (47%) persons an adduct spot was detected that co-chromatographed with the major BPDE-DNA adduct (BPDE-dG); RAL values ranged from 0.03 to 382 adducts in 10(8) nucleotides. Correlations were not significant between the adduct levels in WBC and smoking habits, age or sex. From 20 patients of the same group lung tissue was collected at surgery and examined for PAH-DNA adducts by ELISA and 32P-post-labeling assay. No significant correlation was found between DNA adduct levels in blood and lung. This finding stresses the limitations of the use of WBC as a surrogate for adduct levels in the target organ.     

The relationship between genetic damage from polycyclic aromatic hydrocarbons in breast tissue and breast cancer

A number of polycyclic aromatic hydrocarbons (PAH) are widespread environmental contaminants that cause mammary cancer experimentally. We investigated whether exposure and susceptibility to PAH, as measured by PAH-DNA adducts in breast tissue, are associated with human breast cancer. We carried out a hospital-based case-control study using immunohistochemical methods to analyze PAH-DNA adducts in tumor and nontumor breast tissue from cases and benign breast tissue from controls. The subjects were white, African-American and Latina women without prior cancer or treatment, including 119 women with breast cancer and 108 with benign breast disease without atypia. PAH-DNA adducts measured in breast tumor tissue of 100 cases and in normal tissue from 105 controls were significantly associated with breast cancer (OR=4.43, 96% CI 1.09-18.01) after controlling for known breast cancer risk factors and current active and passive smoking, and dietary PAH. There was substantial interindividual (17-fold) variability in adducts overall, with 27% of cases and 13% of controls having elevated adducts. The odds ratio for elevated adducts in tumor tissue compared with control tissue was 2.56 (1. 05-6.24), after controlling for potential confounders. Adduct levels in tumor tissue did not vary by stage or tumor size. Among 86 cases with paired tumor and nontumor tissue, adducts levels in these two tissues were highly correlated (r=0.56, P<0.001). However, the corresponding associations between case-control status and adducts measured in nontumor tissue from 90 cases and in normal tissue from 105 controls were positive but not statistically significant. Overall, neither active nor passive smoking, or dietary PAH were significantly associated with PAH-DNA adducts or breast cancer case-control status. These results suggest that genetic damage reflecting individual exposure and susceptibility to PAH may play a role in breast cancer; but more research is needed to determine whether the findings are relevant to causation or progression of breast cancer.     

Assessment of interactions between PAH exposure and genetic polymorphisms on PAH-DNA adducts in African American, Dominican, and Caucasian mothers and newborns.

Polycyclic aromatic hydrocarbons (PAH) are widespread pollutants commonly found in air, food, and drinking water. Benzo[a]pyrene is a well-studied representative PAH found in air from fossil fuel combustion and a transplacental carcinogen experimentally. PAHs bind covalently to DNA to form DNA adducts, an indicator of DNA damage, and an informative biomarker of potential cancer risk. Associations between PAH-DNA adduct levels and both cancer risk and developmental deficits have been seen in previous experimental and epidemiologic studies. Several genes have been shown to play an important role in the metabolic activation or detoxification of PAHs, including the cytochrome P450 genes CYP1A1 and CYP1B1 and the glutathione S-transferase (GST) genes GSTM1, and GSTT2. Genetic variation in these genes could influence susceptibility to adverse effects of PAHs in polluted air. Here, we have explored interactions between prenatal PAH exposure and 17 polymorphisms in these genes (rs2198843, rs1456432, rs4646903, rs4646421, rs2606345, rs7495708, rs2472299, rs162549, rs1056837, rs1056836, rs162560, rs10012, rs2617266, rs2719, rs1622002, rs140194, and gene deletion GSTM1-02) and haplotypes on PAH-DNA adducts in cord blood of 547 newborns and in maternal blood of 806 mothers from three different self-described ethnic groups: African Americans, Dominicans, and Caucasians. PAHs were measured by personal air monitoring of mothers during pregnancy. Significant interactions (p < 0.05) were observed between certain genetic polymorphisms and CYP1A1 haplotype and PAHs in mothers and their newborns in the three ethnic groups. However, with our limited sample size, the current findings are suggestive only, warranting further study.     

Semiquantitation of polycyclic aromatic hydrocarbon-DNA adducts in human esophagus by immunohistochemistry and the automated cellular imaging system

It has been suggested that ingestion of polycyclic aromatic hydrocarbons (PAHs) may contribute to the high incidence and mortality of esophageal cancer in Linxian, China. To explore this relationship a semiquantitative immunohistochemical staining method was developed for localization of PAH-DNA adducts. Nuclear color intensity (bright field average pink intensity per nucleus for >1000 cells) was measured using the ChromaVision Automated Cellular Imaging System (ACIS). Paraffin-embedded sections of cultured human keratinocytes exposed to increasing concentrations of 7beta,8alpha-dihydroxy-9alpha,10alpha-epoxy-7,8,9,10-tetrahydro-benzo[a]pyrene (BPDE) were incubated with BPDE-DNA antiserum and served as an internal positive control (standard curve). Values for nuclear staining intensity correlated directly with BPDE exposure concentration (r(2) = 0.99) and were reproducible. DNA adduct levels determined by BPDE-DNA chemiluminescence immunoassay in DNA from BPDE-exposed keratinocytes, correlated with BPDE exposure concentrations (r(2) = 0.99), showing that nuclear staining intensity determined by ACIS correlated directly with BPDE-DNA adduct levels determined by chemiluminescence immunoassay. The ACIS methodology was applied to 5 human samples from Linxian, and significantly positive nuclear PAH-DNA adduct staining was observed in this group when compared with esophageal tissue from 4 laboratory-housed monkey controls and 6 samples obtained at autopsy from smokers and nonsmokers in the United States. Nuclear PAH-DNA staining was absent from Linxian samples when serial sections were incubated with normal rabbit serum (negative control) and was significantly reduced on incubation with BPDE-DNA antiserum absorbed previously with the immunogen BPDE-DNA. These results appear to support the hypothesis that high PAH exposure levels may be etiologically associated with the development of esophageal cancer in Linxian.