Polymorphisms in CYP1A1, GSTM1, GSTT1 and lung cancer below the age of 45 years

E Taioli1, L Gaspari1, S Benhamou2, P Boffetta3, J Brockmoller4, D Butkiewicz5, I Cascorbi6, ML Clapper7, V Dolzan8, A Haugen9, A Hirvonen10, K Husgafvel-Pursiainen10, I Kalina11, P Kremers12, L Le Marchand13, S London14, A Rannug15, M Romkes16, B Schoket17, J Seidegard18, R C Strange19, I Stucker20, J To-Figueras21 and S Garte22

1 Molecular and Genetic Epidemiology Unit, Ospedale Maggiore IRCCS, Italy;
2 INSERM, Villejuif, France, and Geneva Cancer Registry;
3 IARC, Lyon, France;
4 Institute of Clinical Pharmacology, Georg August University, Goettingen, Germany;
5 Institute of Clinical Pharmacology, Gliwice, Poland;
6 Ernst Moritz Arndt University, Greifswald, Germany;
7 Fox Chase Cancer Center, Philadelphia, PA USA;
8 University of Ljubljana, Slovenia;
9 National Institute of Occupational Health, Oslo, Norway;
10 Finnish Institute of Occupational Health Helsinki, Finland;
11 PJ afárik University, Koice, Slovakia;
12 Institut de Pathologie, Liège-Belgium;
13 University of Hawaii-Honolulu, HA, USA;
14 National Institute for Environmental Health Sciences Research Triangle Park, NC USA;
15 Karolinska Institutet Stockholm, Sweden;
16 University of Pittsburgh, Pittsburgh, PA USA;
17 National Institute of Environmental Health, Budapest, Hungary;
18 Lund University, Lund, Sweden;
19 Keele University, Staffordshire UK;
20 INSERM, Villejuif, France;
21 Hospital Clinic Provincial, Barcelona, Spain;
22 EOHSI, UMDNJ, Picataway, NJ USA and Genetics Research Institute Milano, Italy.


    Abstract
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 Statistical analysis
 Results
 Discussion
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Background A genetic component of early-onset lung cancer has been suggested. The role of metabolic gene polymorphisms has never been studied in young lung cancer cases. Phase 1 and Phase 2 gene polymorphisms are involved in tobacco carcinogens’ metabolism and therefore in lung cancer risk.

Methods The effect of metabolic gene polymorphisms on lung cancer at young ages was studied by pooling data from the Genetic Susceptibility to Environmental Carcinogens (GSEC) database. All primary lung cancer cases of both sexes who were Caucasian and <=45 years of age at diagnosis, and the corresponding controls were selected. We obtained 261 cases and 1452 controls.

Results There was a marginally significant association between lung cancer and GSTT1 null genotype (OR=1.2; 95% CI:1.0–1.6), and a significant association between lung cancer and the homozygous CYP1A1 Msp1 variant allele (CYP1A1*2A and *2B) genotype (OR=4.7 95% CI:1.2–19.0). When data were stratified by smoking status, the association between CYP1A1 genotype and lung cancer was confined to never smokers.

Conclusions These results suggest that metabolic genetic factors play a role in lung cancer developing at young ages.


Accepted 26 April 2002

Lung cancer (LC) among smokers of middle and old ages is a major cause of death in western countries. However, LC at younger ages and among non-smokers is a rare disease: approximately 5% of LC are diagnosed under age 45 years, and 4–6% of total cases are never smokers. Although smoking is still a known risk factor at younger ages, it might be hypothesized that early-onset LC among both smokers and non-smokers may have a stronger genetic component in its pathology than that seen at older ages. Polymorphisms in genes involved in both Phase 1 and Phase 2 of xenobiotic metabolism have been associated with a modest increase in LC risk. These genes modulate the effect of tobacco by metabolism of carcinogens in tobacco smoke.1 The association of metabolic genes and LC, and their effect on the association between LC and smoking at younger ages, has never been studied because the rarity of these cases has not allowed gathering of sufficient subjects. Epidemiological data show that early-onset LC is associated with familial aggregation of other cancers,2 and suggest that a genetic component in early-onset LC may be present in non-smokers.3 We wanted to address the hypothesis that polymorphisms in Phase 1 and Phase 2 xenobiotic metabolizing genes are associated with lung cancer at young ages. We used data from the Genetic Susceptibility to Environmental Carcinogens (GSEC) database,4 a pooled analysis containing genotype data and epidemiological information on over 15 000 cancer cases and 15 000 controls.


    Material and Methods
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 Material and Methods
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The GSEC study is a pooled analysis of case-control studies involving metabolic susceptibility genes such as those belonging to the CYP, GST, and NAT families. The study was initiated in 1996, as described elsewhere,4 and now includes individual data on over 30000 subjects, approximately half controls and half cases, of which 5526 are LC cases. We selected from the main data set all the primary Caucasian LC cases of both sexes, aged <=45 years at diagnosis, and their corresponding controls. We obtained 261 cases (5% of available LC, 174 males, 87 females), and 1452 controls (998 males, 454 females), from 21 different studies. Variables selected from the data set are: age, sex, histology, smoking status (never and ever smoker), packs of cigarettes x years of smoking, polymorphisms in CYP1A1, GSTM1, GSTT1 genes. Other variables, such as occupational exposure and family history of cancer, as well as data on other metabolic genes were available, but only for a small fraction of the cases, and therefore were not considered in this analysis. Information on smoking was available for 251 cases and 1309 controls, and on histology for 253 cases. Genotype information was available for GSTM1 (236 cases), GSTT1 (144 cases), CYP1A1 Msp1 (144 cases) and Exon 7 (110 cases) polymorphisms. Genetic analysis for all polymorphisms was done using the same polymerase chain reaction methodology. Standard methodology includes blinding of case/ control status in all studies that are part of the GSEC database.


    Statistical analysis
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Data were analysed using the SAS statistical package, version 7.0. Odds ratios (OR) and 95% CI were calculated, and adjusted for possible confounding factors, such as study, age and gender, by multiple logistic regression. Genotype distribution were compared by {chi}2 according to the Monte Carlo simulation.5 The Breslow-Day test for homogeneity of OR across studies was calculated6 in order to identify significant heterogeneities. The A2455G (Ile/Val) base change in the Exon 7 of the CYP1A1 gene (in CYP1A1*2B and *2C alleles) was analysed in all the available data but one data set, because of methodological differences in detection of this polymorphism. The genetic polymorphisms analysed in the present paper are not in linkage disequilibrium, as previously shown.7


    Results
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The description of cases and controls is reported in Table 1Go. Cases were ever smokers more often than controls (86% versus 50%), and among smokers, the mean value of pack-years was higher among cases than controls. The age- and study-adjusted OR of LC with smoking was 6.2 (95% CI: 4.2–8.9). Of the cases, 32% had a diagnosis of adenocarcinoma, 25% of squamous cell carcinoma, 16% of small cell carcinoma, 7% of large cell carcinoma, 10% of mixed tumours and 10% had other histological diagnoses.


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Table 1 Description of subjects included in the pooled analysis
 
Among the genotypes considered in this analysis (Table 2Go), there was a marginally significant association between LC and GSTT1 null genotype (OR=1.2; 95% CI: 1.0–1.6), and an association between LC and the homozygous variant of the CYP1A1 Msp1 variant polymorphism (OR=4.7, 95% CI:1.2–19.0). Although the A2455G base change in Exon 7 was more frequent among cases than controls, the association was not statistically significant. There was no evidence of heterogeneity among studies for the reported associations: the P-value was 0.7 for CYP1A1 Msp1 polymorphism, 0.6 for CYP1A1 Exon 7 A2455G polymorphism, 0.4 for GSTM1 deletion, 0.9 for GSTT1 deletion.


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Table 2 Association between gene polymorphisms and lung cancer
 
Since these genes are involved in metabolism of tobacco constituents, and their role seems to be more relevant to exposure to low dose of carcinogens,8 the data were stratified by smoking status: the association between the two CYP1A1 polymorphisms and LC seems to be higher among never smokers (Table 3Go). The OR was 3.2 (95% CI:0.8–11.8) for the Msp1 polymorphism, and 4.2 (95% CI:1.1–15.7) for the heterozygous Exon 7 A2455G polymorphism among never smokers, while the respective OR were 0.9 (95% CI:0.5–1.8) and 1.6 (95% CI:0.8–3.4) among ever smokers. When the analysis was repeated by histological type, no difference in genotype distribution with histology was observed. The lack of difference with histological type was still present when the data were stratified by gender, although the small number of females prevented any formal statistical analysis. We did not observe any significant difference in amount of smoking according to genotype among either cases or controls.


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Table 3 Association between gene polymorphisms and lung cancer according to smoking status
 

    Discussion
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This pooled analysis of LC in subjects under 45 years suggests that Phase 1 and Phase 2 metabolic gene polymorphisms may have a role in cancer susceptibility in this age group. In particular, we observed a significant association between the homozygous variant of the CYP1A1 Msp1 polymorphism and LC in young patients, although the results should be viewed with caution due to the small number of subjects in this genotype group. A meta-analysis of published studies in Caucasians of all ages9 shows a non-significant association between the CYP1A1 Msp1 variant allele containing genotypes (*2A and *2B) and LC risk. However, since the analysis was performed by pooling the information on the heterozygous and homozygous variant allele containing genotypes together, the assessment of the effect of the homozygous variant genotype on cancer risk could not be assessed separately. Furthermore, no stratification by age was possible in the meta-analysis, whereas one of the advantages of the pooled data analysis (where original data on each subject are available) is the possibility of analysis of relevant sub-groups. From these data, it seems that at least the CYP1A1 Msp1 polymorphism plays a more relevant role at younger ages than later in life.

Additional information derived from the present analysis is that the association between LC and CYP1A1 genotype appears to be potentially relevant for never smokers. This interpretation is possible because some metabolic gene variants (such as CYP1A1 variants) seem to exert their effect at low doses of exposure to carcinogens.8,10 Similar data have been reported in the meta-analysis, when the data are stratified by dose of exposure to environmental carcinogens.9 Never smokers included in our study could be exposed to low doses of environmental carcinogens from other sources, such as passive smoking, dietary polyaromatic hydrocarbons or occupation. Another explanation for our results could be that LC at young age in never smokers is a disease with a strong genetic component, and therefore the effect of metabolic gene polymorphisms becomes more evident in this subpopulation. There is evidence that familiarity for LC is associated with increased risk of early-onset, but not late-onset, LC.11 A recent publication on risk factors for LC in young women12 shows a smaller association of LC with smoking, especially at very young ages (below age 35). No other known risk factors for LC at young ages are reported in the literature. Our data should be viewed with caution due to the relatively small number of subjects in the never smoking group, and the very few subjects homozygous variant for CYP1A1 among never smokers.

Our analysis does not suggest a major role for Phase 2 genetic polymorphisms GSTM1 and GSTT1 in this age group. Moreover, no combined effect was observed for Phase 1 and Phase 2 genes, in contrast with some published data suggesting an effect of both GSTM1 deletion and CYP1A1 polymorphism on LC risk in women.14

One of the limitations of the present analysis is the difference in laboratory methods for detection of the Exon7 CYP1A1 polymorphisms. Most of the data sets included in the GSEC database contain information on this polymorphism obtained with a method15 that may not accurately distinguish between the Exon 7 variant alleles having a C2455 base change (CYP1A1*2A and *2B) and the more recently described CYP1A1*4 allele having a C2453 base change. Newer studies using more precise methods16 may give different results compared with the previous studies that used the old method. In the current analysis, only one study used the alternative, more precise method, and this study was excluded when analysing the CYP1A1 polymorphisms, for purposes of consistency and homogeneity among studies. Once a sufficient number of subjects have been typed using the new method, a new pooled analysis using these data will be necessary in order to give a more precise estimate of the role of the CYP1A1 Exon 7 polymorphism in the individual LC risk.

There are other limitations in this study, such as uncertainty about standardization of some epidemiological information such as smoking history, and the impossibility of conducting more detailed analysis on environmental exposure. For example, we were unable to study in detail the effect of metabolic polymorphisms at different smoking doses, or other sophisticated analysis of interaction. Population stratification among Caucasians should not be an issue, as we and others have previously shown7,17 that inter-ethnic differences in allele frequencies for these genes in Caucasians are not large. Heterogeneities among studies, such as patient recruitment, distribution of histological types, proportion of subjects successfully genotyped out of the total population, cannot be addressed in this pooled analysis.

However, this is the largest study conducted so far on young LC cases including the measure of metabolic gene polymorphisms, and was possible because of the ongoing collaborative project on genetic susceptibility to environmental carcinogens. Pooled analyses allow investigation of relatively large numbers of cases of rare cancers, and also of the role of genetic factors in particular subgroups of the population at risk. Our study suggests that, in addition to smoking, the main risk factor for LC, metabolic gene polymorphisms may play a role in LC at young ages.


KEY MESSAGES

  • Smoking is a risk factor for lung cancer at younger ages.
  • Metabolic gene polymorphisms appear to modulate smoking-related lung cancer risk.
  • CYP1A1 Msp1 polymorphisms are associated with lung cancer at younger age.

 


    Acknowledgments
 
This work was partly supported by EC grant No. 96/CAN/33919.


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 Abstract
 Material and Methods
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