Chromosomal aberrations and SCEs as biomarkers of cancer risk.

Previous studies have suggested that the frequency of chromosomal aberrations (CAs), but not of sister chromatid exchanges (SCEs), predicts cancer risk. We have further examined this relationship in European cohorts comprising altogether almost 22,000 subjects, in the framework of a European collaborative project (CancerRiskBiomarkers). The present paper gives an overview of some of the results of the project, especially as regards CAs and SCEs. The results confirm that a high level of CAs is associated with an increased risk of cancer and indicate that this association does not depend on the time between CA analysis and cancer detection, i.e., is obviously not explained by undetected cancer. The present evidence indicates that both chromatid-type and chromosome-type CAs predict cancer, even though some data suggest that chromosome-type CAs may have a more pronounced predictive value than chromatid-type CAs. CA frequency appears to predict cancers at various sites, although there seems to be a particular association with gastrointestinal cancers. SCE frequency does not appear to have cancer predictive value, at least partly due to uncontrollable technical variation. A number of genetic polymorphisms of xenobiotic metabolism, DNA repair, and folate metabolism affect the level of CAs and might collectively contribute to the cancer predictivity of CAs. Other factors that may influence the association between CAs and cancer include, e.g., exposure to genotoxic carcinogens and internal generation of genotoxic species. Although the association between CA level and cancer is seen at the group level, an association probably also exists for the individual, although it is not known if an individual approach could be feasible. However, group level evidence should be enough to support the use of CA analysis as a tool in screening programs and prevention policies in occupational and environmental health.

Immortalization of normal human kidney epithelial cells by nickel(II).

The occupational and environmental hazards of nickel exposure are of great concern in environmental medicine. Nickel workers have increased risk of cancer of the nose, lung, larynx, and possibly the kidney. In the present investigation we have studied the effects of nickel ions on fetal human kidney cortex explants. The explants were continuously exposed to 5 micrograms/ml NiSO4. After 70-100 days in culture foci of phenotypically altered cells appeared. Immortalized cell lines were established and demonstrated to be of human epithelial origin. Tumorigenicity was not induced, but the cells demonstrated decreased requirement for serum, increased plating efficiency and saturation density, and formation of colonies in soft agar. Chromosome changes in the treated cells were observed. Worth mentioning are change in ploidy (3n) and abnormalities of chromosomes 1, 7, 9, 11, 13, 14 and 20; increased numbers of chromosome 17; and loss of normal chromosomes 20 and 22.

Chromosomal aberrations in lymphocytes predict human cancer independently of exposure to carcinogens. European Study Group on Cytogenetic Biomarkers and Health.

An increased risk of cancer in healthy individuals with high levels of chromosomal aberrations (CAs) in peripheral blood lymphocytes has been described in recent epidemiological studies. This association did not appear to be modified by sex, age, country, or time since CA test, whereas the role played by exposure to carcinogens is still uncertain because of the requisite information concerning occupation and lifestyle was lacking. We evaluated in the present study whether CAs predicted cancer because they were the result of past exposure to carcinogens or because they were an intermediate end point in the pathway leading to disease. A nested case-control study was performed on 93 incident cancer cases and 62 deceased cancer cases coming from two prospective cohort studies performed in Nordic countries (Denmark, Finland, Norway, and Sweden) and Italy. For each case, four controls matched by country, sex, year of birth, and year of CA test were randomly selected. Occupational exposure and smoking habit were assessed by a collaborative group of occupational hygienists. Logistic regression models indicated a statistically significant increase in risk for subjects with a high level of CAs compared to those with a low level in the Nordic cohort (odds ratio, 2.35; 95% confidence interval, 1.31-4.23) and in the Italian cohort (odds ratio, 2.66; 95% confidence interval, 1.26-5.62). These estimates were not affected by the inclusion of occupational exposure level and smoking habit in the regression model. The risk for high versus low levels of CAs was similar in subjects heavily exposed to carcinogens and in those who had never, to their knowledge, been exposed to any major carcinogenic agent during their lifetime, supporting the idea that chromosome damage itself is involved in the pathway to cancer. The results have important ramifications for the understanding of the role played by sporadic chromosome damage for the origin of neoplasia-associated CAs.

Cancer risk in humans predicted by increased levels of chromosomal aberrations in lymphocytes: Nordic study group on the health risk of chromosome damage.

Cytogenetic assays in peripheral blood lymphocytes (PBL) have been used extensively to survey the exposure of humans to genotoxic agents. The conceptual basis for this has been the hypothesis that the extent of genetic damage in PBL reflects critical events for carcinogenic processes in target tissues. Until now, no follow-up studies have been performed to assess the predictive value of these methods for subsequent cancer risk. In an ongoing Nordic cohort study of cancer incidence, 3182 subjects were examined between 1970 and 1988 for chromosomal aberrations (CA), sister chromatid exchange or micronuclei in PBL. In order to standardize for the interlaboratory variation, the results were trichotomized for each laboratory into three strata: low (1-33 percentile), medium (34-66 percentile), or high (67-100 percentile). In this second follow-up, a total of 85 cancers were diagnosed during the observation period (1970-1991). There was no significant trend in the standardized incidence ratio with the frequencies of sister chromatid exchange or micronuclei, but the data for these parameters are still too limited to allow firm conclusions. There was a statistically significant linear trend (P = 0.0009) in CA strata with regard to subsequent cancer risk. The point estimates of the standardized incidence ratio in the three CA strata were 0.9, 0.7, and 2.1, respectively. Thus, an increased level of chromosome breakage appears to be a relevant biomarker of future cancer risk.

Prader-Willi syndrome in a brother and sister without cytogenetic or detectable molecular genetic abnormality at chromosome 15q11q13.

We report on a 12-year-old boy and his 7-year-old sister with the Prader-Willi syndrome. They both had severe initial hypotonia with feeding problems and later developed an increasing appetite. Both sibs have almond-shaped eyes, triangular mouth, hypogonadism, retarded growth, and mental retardation. An older brother suffered from severe hypotonia and died at 7 days of age. The children have normal chromosomes by high-resolution technique and have inherited the same chromosomes 15 short arm polymorphisms from their parents. The family was informative for one of four DNA markers specific for the 15q11q13 region. No deletion was found using this marker. The parents were healthy and unrelated. Autosomal recessive inheritance or a paternally inherited submicroscopic deletion are possible explanations for the sib occurrence in this family.

Karyotypic changes in the preclinical and subsequent stages of malignant mesothelioma: a case report.

The karyotypic evolution was evaluated in cells from recurring pleural effusions in a patient previously exposed to asbestos. Pleural malignant mesothelioma (MM) was diagnosed 4 years after the first cytogenetic examination. The primary cytogenetic changes consisted of loss of chromosomes 1p,14,21, Y, both 22, and derivative chromosomes involving 1, 2, and 14. The modal chromosome number was 44. Sixty-seven percent of the cells had a normal karyotype. After 4 years of spontaneous remission, only 6% of the cells had a normal karyotype, 42% had the same karyotypic changes as found previously, whereas 52% had additional derivative chromosomes involving chromosomes 1, 3, 5, 7, 8, and 12, trisomy 7, 7p, and 11, and partial or whole monosomy 3, 8, and 9. The chromosomal changes are in agreement with the main findings in previous reports. The karyotype remained quite stable for 7 months in vitro. After 23 months in culture, all the cells were near-triploid. Cells established in culture were cytokeratin positive. All derivative and marker chromosomes identified in the cultured cells had previously been observed in direct preparations from the pleural effusions. We conclude that chromosomes 1, 14, 21, and 22 may be involved in the preclinical stage of development of asbestos-induced mesothelioma, whereas the later chromosomal changes may be related to progression of the tumor.

An inter-Nordic prospective study on cytogenetic endpoints and cancer risk. Nordic Study Group on the Health Risk of Chromosome Damage.

To investigate whether high rates of chromosomal aberrations (CAs), sister chromatid exchange (SCE), or micronuclei(MN) in peripheral lymphocytes indicate an increased risk for subsequent cancer, a prospective cohort study of 2,969 subjects cytogenetically examined between 1970 and 1988 in four Swedish, two Finnish, and two Norwegian laboratories was initiated. To standardize for the interlaboratory variation, the results of the three cytogenetic endpoints were trichotomized for each laboratory into "low" (1st to 33rd percentile), "medium" (34th to 66th percentile), and "high" (67th to 100th percentile]. Thirty-four cancers had been diagnosed in the cohort during the observation period (1970 to 1985). The point-estimates of the standardized morbidity ratio (SMR) in the three CA strata were 90, 92, and 180, respectively. This trend for a positive association was not statistically significant (p = 0.06). There was no significant trend between SMR and the trichotomized rates of SCE. In the subcohort examined for MN only two cases of cancer had been diagnosed until now. If subjects with "high" frequencies of CA or SCE have a two-fold (or greater) risk of developing cancer as compared with individuals who have "medium" or "low" frequencies, we estimate that there is a likelihood of 80% and 70%, respectively, that this will be detectable as significant (p less than or equal to 0.05) differences after a further follow-up period of 5 years. Weaker associations between cancer risk and the cytogenetic endpoints would not be possible to evaluate until even later follow-ups.

Cancer predictive value of cytogenetic markers used in occupational health surveillance programs.

It has not previously been clear whether cytogenetic biomarkers in healthy subjects will predict cancer. Earlier analyses of a Nordic and an Italian cohort indicated predictivity for chromosomal aberrations (CAS) but not for sister chromatid exchanges (SCES). A pooled analysis of the updated cohorts, forming a joint study base of 5271 subjects, will now be performed, allowing a more solid evaluation. The importance of potential effect modifiers, such as gender, age at testing, and time since testing, will be evaluated using Poisson regression models. Two other potential effect modifiers, occupational exposures and smoking, will be assessed in a case-referent study within the study base.

Heterogeneity and selection of neoplastic neurogenic rat cells in vivo and in vitro.

Analysis of chromosome numbers and flow cytometric measurements of relative DNA content have been performed on four neoplastic neurogenic rat cell lines obtained from fetal BD IX-rat brain cells exposed to N-ethyl-N-nitrosourea (EtNU) in vivo, which sequentially underwent neoplastic transformation in cell culture. In vitro, the cell lines showed varying degrees of ploidy (ranging from near-diploid to near-hexaploid values) and chromosome aberrations. When grown in vivo as solid tumours in BD IX-rats and in thymus deficient nude mice, or in intraperitoneal diffusion chambers in rats, the near diploid cell lines were stable. In the lines with higher ploidies the proportion of cells with near-diploid values increased. Upon retransfer to cell culture, the cell lines gradually changed to the original in vitro pattern. However, in some cases an elevated near-diploid fraction persisted during culture after retransfer. Despite the selection of malignant cells with different ploidy in vivo and in vitro, their ultrastructural morphology remained unchanged. Thus, in this system, near-diploid cells are favoured under conditions of in vivo growth.

Cytogenetic studies of stainless steel welders using the tungsten inert gas and metal inert gas methods for welding.

Cytogenetic damage was studied in lymphocytes from 23 welders using the Tungsten Inert Gas (TIG), and 21 welders using the Metal Inert Gas (MIG) and/or Metal Active Gas (MAG) methods on stainless steel (SS). A matched reference group I, and a larger reference group II of 94 subjects studied during the same time period, was established for comparison. Whole blood conventional cultures (CC), cultures in which DNA synthesis and repair were inhibited (IC), and the sister chromatid exchange (SCE) assay were applied in the study. For the CC a statistically significant decrease in chromosome breaks and cells with aberrations was found for both TIG/SS and MIG/MAG/SS welders when compared with reference group II. A non-significant decrease was found for the corresponding parameters for the two groups of welders when compared with their matched referents. A statistically significant negative association was found between measurements of total chromium (Cr) in inhaled air and SCE, and a weaker negative correlation with hexavalent Cr (Cr(VI)) in air. In conclusion, no cytogenetic damage was found in welders exposed to the TIG/SS and MIG/MAG/SS welding fumes with low content of Cr and Ni. On the contrary, a decline in the prevalence of chromosomal aberrations was indicated in the TIG/SS and MIG/MAG/SS welders, possibly related to the suggested enhancement of DNA repair capacity at slightly elevated exposures.

Chromosome damage in lymphocytes of stainless steel welders related to past and current exposure to manual metal arc welding fumes.

Cytogenetic damage was studied in lymphocytes from 42 welders using the manual metal arc (MMA) method on stainless steel (SS). A detailed characterization of previous exposure by job interviews, and for current exposure with personal air sampling and biological monitoring of chromium (Cr) and nickel (Ni) in blood and urine, was done for 32 of these welders. A subgroup of 20 welders was studied before and after 1-4 months of MMA/SS welding. A matched reference group I, and a larger reference group II were established for comparison. A significant increase in chromatid breaks (1.4 vs. 0.9 and 0.8 for group I and II) and for cells with aberrations (2.2 vs. 1.6 in group II) was found in the welders. An even larger difference was found when comparing non-smoking welders with their non-smoking referents. No synergistic effect between smoking and MMA/SS welding fumes was observed for any type of aberrations. Current welding fume exposure during the week before sampling was not associated with increases in any type of cytogenetic damage. The results indicated that the increase in chromatid breaks was associated with cumulated welding fume exposure for more than a year, and with not using respirators. Exposure to MMA/SS welding fumes for up to 4 months gave a slight, but significant increase in chromatid breaks when using the welders as their own referents. However, when using matched referents in the study after exposure, no difference was found between these welders and their matched referents. No differences between the groups were observed in the DNA synthesis and repair-inhibited cultures or for SCE.

Enhanced cytogenetic detection of previous in vivo exposure to mutagens in human lymphocytes after treatment with inhibitors of DNA synthesis and DNA repair in vitro.

To increase the sensitivity of cytogenetic surveillance of exposure to mutagens in the peripheral lymphocyte assay, structural chromosome aberrations (CA) were studied after inhibition of DNA synthesis and DNA repair with hydroxyurea and caffeine in culture 3 h prior to harvesting. CA and sister-chromatid exchanges (SCE) from conventional cultures from the same subjects were used for comparison. Smoking was used as exposure parameter. Thirty-two smokers and 35 nonsmokers were studied. In the inhibited cultures a significantly higher number of aberrations was found in lymphocytes from smokers than nonsmokers: chromatid breaks (20.4 vs. 11.8, p = 0.0002), chromosome breaks (4.5 vs. 1.7, p = 0.0003), and the number of cells with aberrations (18.9 vs. 12.4, p = 0.0001), when 50 cells per subject were analyzed. In conventional cultures no increase in gaps, chromatid and chromosome breaks or number of cells with aberrations was found in smokers when 100 cells from each subject were studied. Smokers showed an increased number of SCE (6.8 vs. nonsmokers 5.9, p = 0.02). A significant positive linear correlation (r = 0.39, p = 0.01) was seen between SCE and the number of cells with chromatid breaks from inhibited cultures. The present results indicate that adding hydroxyurea and caffeine to lymphocyte cultures for the last 3 h prior to harvesting may enhance the detection of cytogenetic damage from previous in vivo exposure to mutagens.

Cancer predictive value of cytogenetic markers used in occupational health surveillance programs: a report from an ongoing study by the European Study Group on Cytogenetic Biomarkers and Health.

The cytogenetic endpoints in peripheral blood lymphocytes: chromosomal aberrations (CA), sister chromatid exchange (SCE) and micronuclei (MN) are established biomarkers of exposure for mutagens or carcinogens in the work environment. However, it is not clear whether these biomarkers also may serve as biomarkers for genotoxic effects which will result in an enhanced cancer risk. In order to assess this problem, Nordic and Italian cohorts were established, and preliminary results from these two studies indicated a predictive value of CA frequency for cancer risk, whereas no such associations were observed for SCE or MN. A collaborative study between the Nordic and Italian research groups, will enable a more thorough evaluation of the cancer predictivity of the cytogenetic endpoints. We here report on the establishment of a joint data base comprising 5271 subjects, examined 1965-1988 for at least one cytogenetic biomarker. Totally, 3540 subjects had been examined for CA, 2702 for SCE and 1496 for MN. These cohorts have been followed-up with respect to subsequent cancer mortality or cancer incidence, and the expected values have been calculated from rates derived from the general populations in each country. Stratified cohort analyses will be performed with respect to the levels of the cytogenetic biomarkers. The importance of potential effect modifiers such as gender, age at test, and time since test, will be evaluated using Poisson regression models. The remaining two potential effect modifiers, occupational exposures and smoking, will be assessed in a case-referent study within the study base.

Chromosome aberrations in lymphocytes of high-voltage laboratory cable splicers exposed to electromagnetic fields.

Thirteen high-voltage laboratory employees and 20 referents participated in a cross-sectional, matched-pairs study of cytogenetic damage. During cable testing the workers were exposed to static, alternating, or pulsed electric and magnetic fields. The alternating magnetic field levels of 50 Hz were 5-10 microT, occasionally much higher. Chromosome aberrations, sister chromatid exchanges, and aneuploidy were studied in peripheral blood lymphocytes. In addition, chromosome aberrations were investigated in lymphocyte cultures treated with hydroxyurea and caffeine, to inhibit deoxyribonucleic acid synthesis and repair. Among seven smoking laboratory employees the mean number of chromosome breaks/200 cells was 2.3, as compared with 0.7 for the job-matched referents. The comparable figures for inhibited cultures were 12.0 versus 6.0. No increase was detected in nonsmokers with either method. The other genetic parameters showed no differences between the exposed workers and the referents. The results support, to some extent, the hypothesis of an increased risk of genotoxic effects among high-voltage laboratory workers.

Chromosome aberrations in chloralkali workers previously exposed to mercury vapor.

Chromosome aberrations and micronuclei in peripheral lymphocytes were studied in 29 male chloralkali workers previously exposed to mercury vapor and in two matched reference groups comprising 29 nitrate fertilizer workers and 29 customs and police officers. The study was performed using whole-blood cultures with and without hydroxyurea and caffeine to inhibit deoxyribonucleic acid synthesis and repair, respectively. No significant differences in the frequencies of chromosome aberrations and micronuclei were observed. However, a nonsignificant increase in chromosome breaks and dicentrics was found in the subgroups with high urinary mercury peak levels or high cumulative mercury exposure. An increased prevalence of "high" scores of chromatid breaks in the inhibited cultures, exceeding the 75th percentile of all of the subjects studied, was observed for the chloralkali workers when compared with both reference groups. No evident cytogenetic effects were observed among the chloralkali workers with the methods used in the present study.

Birth weight and environmental pollution in the county of Telemark, Norway.

The possible effect of environmental pollution on fetal growth was examined in 3,231 consecutively liveborn single infants (>/=37 weeks' gestation) of Caucasian parents born between 1986 and 1988. The parents lived in an industrial area or in less polluted urban and rural residential areas. Information about lifestyle, health, and work exposures was collected from the parents. A significantly lower arithmetic mean birth weight was observed for newborns in the industrial residential area (3,517 g, SD, 482), compared with the urban (3,592 g, SD 495) and rural (3,618 g, SD 517) areas (P < 0.05). Even controlling for gestational age, sex, parity, maternal smoking habits, and social class, residential area still had a significant effect on birth weight. Among other factors examined, only maternal psychological stress at work had a significant effect on birth weight. If the observed association reflects a causal relationship, birth weight may represent a potential outcome parameter for surveillance of effects on humans of environmental exposures.

Transformation of human kidney epithelial cells to tumorigenicity by nickel(II) and V-HA-RAS oncogene.

Nickel is a toxic metal of environmental concern that has been found to be carcinogenic in man and animals. Primary human kidney (NHKE) cells were immortalized or rescued from senescence after exposure to NiSO4. The cell lines (IHKE) displayed abnormal karyotype and anchorage independent growth was observed. However, none of the IHKE cells produced tumor upon injection into athymic nude mice. Transfer of the v-Ha-ras oncogene into IHKE cells induced conversion of the immortalized cells into cell lines (THKE) that were tumorigenic when transplanted into athymic nude mice. Ha-ras DNA was present in the transformed cell lines and expressed at high level.

Two chromosomal syndromes in the same family: monosomy and trisomy for part of the short arm of chromosome 10.

A grandfather with balanced translocation t(1:10) gave rise to three possible combinations involving chromosome 10: balanced translocation and trisomy for part of the short arm 10p13 leads to pter in the second generation, and mono- and trisomy 10p13 leads to pter in the third generation. The clinical signs and symptoms of the present case with monosomy 10p13 leads to pter are compared with those of 9 earlier reported cases with a deleted 10p. Together they represent a clinically recognizable syndrome with antimongoloid eye slant, ptosis, epicanthus, high arched or cleft palate, flat nasal bridge, micrognathia, small round and low-set ears, wide spaced nipples, cardiac and urinary tract abnormalities, hand and foot anomalies, hypoplasia/absence of the olfactory bulbs/tracts, psychomotor and growth retardation. More than 20 cases of the trisomy 10p syndrome have been described earlier. The most constant clinical findings are mental retardation, dolichocephaly, frontal bossing, broad nasal bridge, cheilo-palatoschisis, retrognathia and variable internal malformations. We found, however, the clinical characteristics in this syndrome more variable than for the monosomy 10p13 leads to pter syndrome. Our two cases, representing the eldest and the youngest described, have rather few of the typical characteristics, and few in common with each other. This indicates the difficulty in making this diagnosis on clinical features only without a cytogenetic verification.