Biomarkers of genetic damage and inflammation in blood and bronchoalveolar lavage fluid among former German uranium miners: a pilot study.

Former East German uranium miners who are known to have been exposed to radon are estimated to be at high risk for lung carcinogenesis. Among these miners over 200 occupationally caused lung cancer cases are expected to occur each year, resulting in a total of 7,000-24,000 excess lung cancer cases in the coming years. It is still unknown whether there is a correlation between biomarkers and the exposure of the uranium miners to ionizing radiation that might enable us to trace those miners with high lung cancer risk. The primary aim of this pilot study was to test the possibility of performing a biomarker study in this unique cohort of former uranium miners in spite of several limitations that had to be taken into consideration when comparing them with healthy controls, such as old age, age-dependent diseases and potential confounding artefacts from dissimilar smoking patterns. The second aim was to test a range of biomarkers for DNA damage and inflammation in leukocytes and bronchoalveolar fluid for their ability to detect biological effects. In this cohort of miners we found an increased frequency of chromosomal aberrations in blood lymphocytes and an increased prevalence of both fibronectin and tumour necrosis factor alpha in the bronchoalveolar fluid.

p53 mutations and codon 213 polymorphism of p53 in lung cancers of former uranium miners.

PURPOSE: There is a high prevalence of G-->T transversions of p53 in lung cancers of smokers. One study has reported a special "hotspot" mutation at codon 249 of p53 in lung cancers of former uranium miners. The aim of our study was to look for mutational spectra of p53 in former German uranium miners with lung cancers. METHODS: We investigated 16 patients with lung cancer who had worked as uranium miners in Germany and 13 lung cancer patients without a mining history of the same region. By means of the polymerase chain reaction and sequencing we looked for mutations in exons 5 7 of the p53 gene. RESULTS: We could not find any suggestion of hotspot mutations. The only G-->T mutation in former uranium miners was detected in the only nonsmoker. In 3 patients (19% of the total) we found a codon 213/3 polymorphism. CONCLUSIONS: The results indicate that G-->T transversions do not seem to be very common mutations in p53 in lung cancers probably caused by radiation. Therefore, p53 may be mutated early in lung cancer development if radiation exposure is a critical factor in carcinogenesis. In accordance with studies of thyroid cancer patients in the Chernobyl region, our results may indicate an overrepresentation of codon 213/3 polymorphism in p53 in radiation-caused cancers.

Changes in the classification of carcinogenic chemicals in the work area. Section III of the German List of MAK and BAT Values.

Carcinogenic chemicals in the work area are currently classified into three categories in section III of the German List of MAK and BAT Values (list of values on maximum workplace concentrations and biological tolerance for occupational exposures). This classification is based on qualitative criteria and reflects essentially the weight of evidence available for judging the carcinogenic potential of the chemicals. It is proposed that these categories - IIIA1, IIIA2, IIIB - be retained as Categories 1, 2, and 3, to correspond with European Union regulations. On the basis of our advancing knowledge of reaction mechanisms and the potency of carcinogens, these three categories are supplemented with two additional categories. The essential feature of substances classified in the new categories is that exposure to these chemicals does not contribute significantly to risk of cancer to man, provided that an appropriate exposure limit (MAK value) is observed. Chemicals known to act typically by nongenotoxic mechanisms and for which information is available that allows evaluation of the effects of low-dose exposures, are classified in Category 4. Genotoxic chemicals for which low carcinogenic potency can be expected on the basis of dose-response relationships and toxicokinetics, and for which risk at low doses can be assessed are classified in Category 5. The basis for a better differentiation of carcinogens is discussed, the new categories are defined, and possible criteria for classification are described. Examples for Category 4 (1,4-dioxane) and Category 5 (styrene) are presented.

European Consensus Statement on Lung Cancer: risk factors and prevention. Lung Cancer Panel.

This article is based on discussions of the lung cancer panel at the Hohenheim Consensus Meeting organized by the World Health Organization and the German Ministry of Health in November 1996. Panel members were international experts in the field of diet and cancer who discussed specific questions relating to lung cancer risk factors and prevention.

Elevated DNA single-strand breakage frequencies in lymphocytes of welders exposed to chromium and nickel.

DNA damage (alkaline filter elution) and sister chromatid exchange (SCE) frequencies were measured in lymphocytes of 39 welders and 39 controls. The welders showed a significantly higher rate of DNA single-strand breakages and significantly elevated SCE values. These results are not in accordance with those of a former study in which only DNA-protein cross-links were measured. The different results may be explained on the basis of different exposure levels for chromium(VI) and nickel. Both methods are not specific but sensitive enough to measure genotoxic damage after occupational exposure to chromium(VI) and nickel in the range of threshold values for the workplace on a collective basis. Additionally, the results indicate that DNA single-strand breakage and DNA-protein cross-links show different increases depending on the exposure levels for chromium and nickel.

Changes in the classification of carcinogenic chemicals in the work area. (Section III of the German List of MAK and BAT values).

Carcinogenic chemicals in the work area were previously classified into three categories in section III of the German List of MAK and BAT values (the list of values on maximum workplace concentrations and biological tolerance for occupational exposures). This classification was based on qualitative criteria and reflected essentially the weight of evidence available for judging the carcinogenic potential of the chemicals. In the new classification scheme the former sections IIIA1, IIIA2, and IIIB are retained as categories 1, 2, and 3, to correspond with European Union regulations. On the basis of our advancing knowledge of reaction mechanisms and the potency of carcinogens, these three categories are supplemented with two additional categories. The essential feature of substances classified in the new categories is that exposure to these chemicals does not contribute significantly to the risk of cancer to man, provided that an appropriate exposure limit (MAK value) is observed. Chemicals known to act typically by non-genotoxic mechanisms, and for which information is available that allows evaluation of the effects of low-dose exposures, are classified in category 4. Genotoxic chemicals for which low carcinogenic potency can be expected on the basis of dose/response relationships and toxicokinetics and for which risk at low doses can be assessed are classified in category 5. The basis for a better differentiation of carcinogens is discussed, the new categories are defined, and possible criteria for classification are described. Examples for category 4 (1,4-dioxane) and category 5 (styrene) are presented.

DNA single strand breakage, DNA adducts, and sister chromatid exchange in lymphocytes and phenanthrene and pyrene metabolites in urine of coke oven workers.

OBJECTIVES: To investigate the specificity of biological monitoring variables (excretion of phenanthrene and pyrene metabolites in urine) and the usefulness of some biomarkers of effect (alkaline filter elution, 32P postlabelling assay, measurement of sister chromatid exchange) in workers exposed to polycyclic aromatic hydrocarbons (PAHs). METHODS: 29 coke oven workers and a standardised control group were investigated for frequencies of DNA single strand breakage, DNA protein cross links (alkaline filter elution assay), sister chromatid exchange, and DNA adducts (32P postlabelling assay) in lymphocytes. Phenanthrene and pyrene metabolites were measured in 24 hour urine samples. 19 different PAHs (including benzo(a)pyrene, pyrene, and phenanthrene) were measured at the workplace by personal air monitoring. The GSTT1 activity in erythrocytes and lymphocyte subpopulations in blood was also measured. RESULTS: Concentrations of phenanthrene, pyrene, and benzo(a)pyrene in air correlated well with the concentration of total PAHs in air; they could be used for comparisons of different workplaces if the emission compositions were known. The measurement of phenanthrene metabolites in urine proved to be a better biological monitoring variable than the measurement of 1-hydroxypyrene. Significantly more DNA strand breaks in lymphocytes of coke oven workers were found (alkaline filter elution assay); the DNA adduct rate was not significantly increased in workers, but correlated with exposure to PAHs in a semiquantitative manner. The number of sister chromatid exchanges was lower in coke oven workers but this was not significant; thus counting sister chromatid exchanges was not a good variable for biomonitoring of coke oven workers. Also, indications for immunotoxic influences (changes in lymphocyte subpopulations) were found. CONCLUSIONS: The measurement of phenanthrene metabolites in urine seems to be a better biological monitoring variable for exposure to PAHs than measurement of hydroxypyrene. The alkaline filter elution assay proved to be the most sensitive biomarker for genotoxic damage, whereas the postlabelling assay was the only one with some specificity for DNA alterations caused by known compounds.

Proposed changes in the classification of carcinogenic chemicals in the work area.

Carcinogenic chemicals in the work area are currently classified into three categories in Section III of the German List of MAK and BAT Values. This classification is based on qualitative criteria and reflects essentially the weight of evidence available for judging the carcinogenic potential of the chemicals. It is proposed that these Categories--IIIA1, IIIA2, and IIIB--be retained as Categories 1, 2, and 3, to conform with EU regulations. On the basis of our advancing knowledge of reaction mechanisms and the potency of carcinogens, it is now proposed that these three categories be supplemented with two additional categories. The essential feature of substances classified in the new categories is that exposure to these chemicals does not convey a significant risk of cancer to man, provided that an appropriate exposure limit (MAK value) is observed. It is proposed that chemicals known to act typically by nongenotoxic mechanisms and for which information is available that allows evaluation of the effects of low-dose exposures be classified in Category 4. Genotoxic chemicals for which low carcinogenic potency can be expected on the basis of dose-response relationships and toxicokinetics and for which risk at low doses can be assessed will be classified in Category 5. The basis for a better differentiation of carcinogens is discussed, the new categories are defined, and possible criteria for classification are described. Examples for Category 4 (1,4-dioxane) and Category 5 (styrene) are presented. The proposed changes in classifying carcinogenic chemicals in the work area are presented for further discussion.

Phenylguanine found in urine after benzene exposure.

Comparative investigations with synthetic N7-phenylguanine were carried out to clarify whether this compound is eliminated via the urine of rats as a benzene-derived nucleic acid adduct. As sensitive methods for detecting trace amounts of the compound, gas chromatography-mass spectroscopy, high performance liquid chromatography, and two immunoassays (enzyme-linked immunosorbent assay and fluoroimmunoassay) with appropriate monoclonal antibodies were used. The results indicate the excretion of several benzene-related guanine adducts slightly different from N7-phenylguanine that may possibly be hydroxylated. These adducts differ also from (O6-, N2- and C8-phenylguanine, respectively.

32P-postlabeling analysis of DNA adducts in different populations.

Blood samples were obtained from different populations exposed occupationally or by lifestyle habits to polycyclic aromatic hydrocarbons (PAH). DNA adducts were determined by 32P-postlabeling assay either in white blood cells (WBC) or lymphocytes. The level of DNA adducts ranged from 1.5 per 10(9) nucleotides in one of the control groups up to 7.1 per 10(9) nucleotides in one group of PAH-exposed workers. Comparison of the adduct thin layer chromatography (TLC) profiles revealed individual variation in both pattern and level of DNA adducts. Significant differences of adduct levels were detected between one group of PAH-exposed coke-oven workers and the corresponding control group. Only a weak influence of the smoking habits on the amount of adducts was detectable in occupationally exposed or unexposed individuals.

Biomonitoring of urinary aromatic amines and arylamine hemoglobin adducts in exposed workers and nonexposed control persons.

The renal excretion of arylamines in occupationally exposed and nonexposed subjects was measured by a gas chromatography-electron capture detector method. Additionally, in the occupationally exposed persons hemoglobin adduct levels of arylamines were determined by a liquid chromatography-electrochemical detector method, together with the individual acetylator status. The aromatic amines aniline, p-toluidine, 2-naphthylamine, and 4-chloro-o-toluidine were detected in the urine of nonsmoking subjects who were not occupationally exposed to arylamines. Significantly higher concentrations of aniline, o-toluidine, m-toluidine, 2-naphthylamine, and 4-methyl-1,3-phenylenediamine could be observed in the urine of smoking control persons in comparison to nonsmokers. Comparison of smokers and nonsmokers in a group of workers primarily exposed to aniline and 4-chloroaniline revealed significant differences (P < 0.05) in the formation of 4-aminodiphenyl hemoglobin adducts and in the renal excretion of 2-naphthylamine. The slow acetylators in this group produced significantly more hemoglobin adducts of aniline and 4-chloroaniline than did the fast acetylators. In slow acetylators among the smoking workers there was a significant increase in the formation of 4-aminodiphenyl hemoglobin adducts and in the renal excretion of 4-chloroaniline and m-toluidine. The results indicate that there are influences of smoking habits and acetylator status on the levels of arylamine hemoglobin adducts as well as urinary arylamine concentrations. Hemoglobin adducts seem to be good parameters for monitoring aniline and 4-chloroaniline exposure at the workplace, especially if the acetylator polymorphism can be taken into account. 4-Aminodiphenyl hemoglobin adducts might be good parameters for monitoring individual smoking habits. The determination of urinary arylamine concentrations provides additional information concerning acute exposures to aromatic amines.

Nasal cancer in leather workers: an occupational disease.

Nasal cancer has a number of causative agents; exposures to most of the established nasal carcinogens occur in the workplace. An increased risk of nasal cancer has been ascertained in shoe-making and shoe-repairing, but the results for leather goods manufacture and leather tanning don't provide adequate evidence of carcinogenicity. Findings from two epidemiological studies carried out in Italy (a case/control study and a case/series report) add further information on this issue. The case/control study was performed in the provinces of Siena (Tuscany), Verona and Vicenza (Venetia) including 96 cases and 378 controls. A significant increased risk (Odds Ratio: 6.8; 90% C.I. = 1.9-25) of sinonasal cancer was found for the employ in the whole leather industry; Odds Ratio of 8.3 (C.I. = 1.9-36) and 5.0 (C.I. = 0.92-28) were associated with shoe-making and leather tanning. The case/series report is based on 110 patients accepted in some Italian hospitals during last three years (1990-1993): 26 cases had worked in the leather industry; adenocarcinoma is the most frequent type observed. Chromium salts and natural tannins are indicated as possible aetiological agents.

Genotoxicity of wood dust in a human embryonic lung cell line.

Wood dust exposure has been found to be an occupational hazard, being linked to an enhanced incidence of various neoplasias. Here we performed an experiment to evaluate the ability of solvent extracts of natural woods to induce chromosome aberrations in respiratory cells in culture. Human embryonic lung cells, MRC-5, grown in Dulbecco's medium were exposed to various concentrations of the dust extracts of pesticide-free (untreated) beech, oak and pine woods. Three concentrations per extract with and without metabolic activation (S9) and 100 metaphase cells per dose were examined for possible structural aberrations. Although no dose-dependent activity could be found with any extract in the presence of S9, most aberrations observed were of the chromatid type caused by oak wood. Dose-dependent chromosomal breaks caused by oak and chromatid breaks caused by both beech and oak were observed in the absence of S9. These data might support the early hypothesis that hard wood dust per se contains some in vivo genotoxic and thus possibly carcinogenic components.

Sister chromatid exchange frequencies in lymphocytes of oral cancer patients seem to be influenced by drinking habits.

Sister chromatid exchange (SCE) values were determined in the lymphocytes of 24 oral cancer patients before therapy and in the lymphocytes of 24 control persons standardized with respect to sex, age and smoking habits. Oral cancer patients showed significantly elevated SCE values (mean 7.82 versus 6.42). In both groups the highest SCE values were found in the subgroups with the highest alcohol consumption. A significant correlation between SCE and gamma-glutamyltranspeptidase (GGT) values by Spearman correlation analysis was detected in the combined group (cancer patients and control persons) (n = 32, r = 0.40, P = 0.023). The SCE values in the oral cancer patients were weakly correlated (Pearson) to DNA adduct levels (n = 22, r = 0.39, P = 0.068) and DNA single-strand breakage frequencies (n = 12, r = 0.56, P = 0.054) in lymphocytes. The correlation (Pearson) between SCE values and DNA strand breakage values in lymphocytes was significant (n = 10, r = 0.67, P = 0.036) in smoking cancer patients. The increase of SCE values with respect to alcohol drinking habits underlines epidemiologic findings that alcohol is an important co-carcinogen in many cancers, especially in oral cancers. Because of the influences on SCE and adduct levels in lymphocytes, alcohol drinking habits should be controlled as broadly as possible in biomarker studies.

Genotoxic and cytotoxic effects of 4-aroyl-1-nitrosohydrazine-carboxamides on O6-alkylguanine-DNA alkyltransferase-positive and -negative human cell lines.

Five different representatives (I-V) of a new class of bifunctional alkylating agents, the 4-aroyl-1-nitrosohydrazinecarboxamides ("nitrososemicarbazides"), were evaluated for their potential interaction with DNA and for their cytotoxic activity in vitro to O6-alkylguanine-DNA alkyltransferase-positive (Mer+) and -negative (Mer-) human cell lines. The HeLa MR cell line (Mer-) showed up to 20-fold higher sensitivity at IC50 (dose that inhibits colony formation by 50%) to agents I-V than did the HeLa S3 cell line (Mer+) in a colony-formation assay. These data were compared to those obtained by treatment of the two cell lines with carmustine, a currently used antitumor drug. In Mer+ cells comparable results to those with carmustine were obtained with compounds III, IV and V; in Mer- cells compounds I and II showed nearly the same effects as carmustine. Whether compounds I-V produce DNA strand breaks and/or DNA-protein cross-links was investigated using an alkaline filter elution technique. In this assay all compounds produced DNA single-strand breaks; no correlation could be detected between the strand breakage frequency and cytostatic, mutagenic and antitumor activity.

Concentrations of benzene in blood and S-phenylmercapturic and t,t-muconic acid in urine in car mechanics.

Different parameters of biological monitoring were applied to 26 benzene-exposed car mechanics. Twenty car mechanics worked in a work environment with probably high benzene exposures (exposed workers); six car mechanics primarily involved in work organization were classified as non-exposed. The maximum air benzene concentration at the work places of exposed mechanics was 13 mg/m3 (mean 2.6 mg/m3). Elevated benzene exposure was associated with job tasks involving work on fuel injections, petrol tanks, cylinder blocks, gasoline pipes, fuel filters, fuel pumps and valves. The mean blood benzene level in the exposed workers was 3.3 micrograms/l (range 0.7-13.6 micrograms/l). Phenol proved to be an inadequate monitoring parameter within the exposure ranges investigated. The muconic and S-phenylmercapturic acid concentrations in urine showed a marked increase during the work shift. Both also showed significant correlations with benzene concentrations in air or in blood. The best correlations between the benzene air level and the mercapturic and muconic acid concentrations in urine were found at the end of the work shift (phenylmercapturic acid concentration: r = 0.81, P < 0.0001; muconic acid concentration: r = 0.54, P < 0.05). In conclusion, the concentrations of benzene in blood and mercapturic and muconic acid in urine proved to be good parameters for monitoring benzene exposure at the workplace even at benzene air levels below the current exposure limits. Today working as a car mechanic seems to be one of the occupations typically associated with benzene exposure.

DNA-protein cross-links and sister chromatid exchange frequencies in lymphocytes and hydroxyethyl mercapturic acid in urine of ethylene oxide-exposed hospital workers.

The lymphocytes of 25 hospital workers exposed to ethylene oxide and of a standardized control group were investigated for DNA damage (measured by alkaline filter elution) and sister chromatid exchange (SCE) frequencies. Additionally, the excretion of hydroxyethyl mercapturic acid (HEMA) in the 24-h urine of ten workers and ten control persons was determined. The peak levels of ethylene oxide in air during the first 8 min after opening of the sterilization unit were measured. Peak levels of ethylene oxide in the air of up to 417 ppm after opening of the sterilization unit were detected. In the alkaline filter elution assay we found significantly reduced elution rates in the exposed workers, indicating DNA-protein cross-links. The reduction of the elution rates through HVLP filters correlated significantly with the exposure classification (low, medium, high) (r = -0.45, P < 0.05) and the ethylene oxide peak level after opening of the sterilization unit (r = -0.42, P < 0.05). The SCE frequencies in the standardized control group were significantly elevated. With respect to (n = 78) historic control SCE values of our institute, the SCE values of the disinfectors were not significantly elevated (6.54 vs 6.27). The ethylene oxide-exposed workers did not have a greater percentage of high-frequency SCE cells. The mean HEMA concentration in the urine of the exposed workers was significantly elevated, but there were wide variations in HEMA concentrations and no correlation to ethylene oxide exposure. We conclude that the alkaline filter elution assay may be a sensitive parameter for ethylene oxide-exposed workers.(ABSTRACT TRUNCATED AT 250 WORDS)

DNA strand breakage and DNA adducts in lymphocytes of oral cancer patients.

In lymphocytes of 12 oral cancer patients (and two control groups) the frequencies of DNA single-strand breaks and DNA-protein cross-linking were determined by alkaline filter elution. We found elevated DNA elution rates, which must be interpreted as an increased strand breakage frequency. There were significant correlations between the DNA strand breakage frequency and smoking habits. Using the 32P-postlabelling assay we determined the DNA adduct level in lymphocytes of 23 oral cancer patients (and two control groups). No significant influence of smoking habit on the DNA adduct level could be detected. There was a significant correlation between the DNA adduct level and the gamma-glutamyltranspeptidase (GGT) value, suggesting systemic influences of alcohol drinking habits on the adduct level.