In this study, we have examined whether polymorphisms in four DNA repair genes involved in the nucleotide excision (NER), base excision (BER), and double-strand break (DSBR) DNA repair pathways are implicated in the development of lung cancer in a Caucasian population from Asturias, Northern Spain. Our results support that polymorphisms in two different NER genes (XPC and XPD) increased the risk of developing lung cancer, so individuals homozygous for the XPC PAT+, XPD 312Asn or XPD 751Gln alleles have a higher risk of developing lung cancer (ORs 1.28, 1.52 and 1.38, respectively). This association was particularly important for ever smokers and patients with adenocarcinomas. On the other hand, no association was found between two genes that participate in the BER and DSBR repair processes (XRCC1 and XRCC3) and the risk of lung cancer. Additionally, interaction between XPC and XPD polymorphisms showed an increased risk of lung cancer (OR = 2.25). Similarly, interactions between XPC/XRCC3 and XPD/XRCC3 were observed, suggesting that coordination between both repair systems might contribute to the individual susceptibility to develop cancer.
Our study has several strengths, including high participation of eligible cases (rate 93.8%), quite large sample size from a homogeneous population of same ancestors (516 cases and 533 controls) and the fact that all our control subjects were under Hardy-Weinberg equilibrium. Nevertheless all our cases were pathology confirmed and finally we applied a severe quality control from genotyping. The main limitations of our study were hospital-based subjects, recall bias due to the fact that information on smoking exposure was obtained retrospectively, and especially possible false positive associations, due to multiple comparisons made, we cannot exclude the possibility that some of these associations may represent chance finding, because the power to detect interactions was limited. On the other hand, we have to bear in mind that 26% of controls were ETS exposed which could lead to underestimate our results. To limit selection bias, we carefully selected controls from patients admitted for various diagnoses that were thought to be unrelated to exposures of interest. Nevertheless, a recent paper from Campbell et al
] reported that European populations may display various levels of genetic substructure which may lead to false positive associations due to population stratification. In our study, we controlled for this possibility by matching individuals on the basis of European ancestry.
We have previously shown that the PAT+ allele is in complete linkage disequilibrium with the intron 11 A-allele [12
], reflecting the XPC
haplotype (PAT+/939Gln/intron 11 A
) with a reduced ability to repair DNA lesions and an increased risk of developing lung cancer. Previous functional analysis has shown that cells with the A/A
genotype at the splice acceptor site in intron 11 have a higher frequency of deletion of exon 12 [50
], suggesting that this mechanism might contribute to the reduced ability of individuals with this genotype to repair DNA lesions. Nevertheless, the effect of the Lys939Gln polymorphism on the biochemical activity of XPC is still under investigation.
Several reports have shown that polymorphisms in the XPC
gene increase the risk of different tumor types, including smoking-related cancers and cutaneous melanoma [13
]. For lung cancer, the number of studies is still very limited. A recent study carried out in an Asiatic population of 432 cases and 432 controls was unable to find any association between the XPC
PAT polymorphism and the risk of developing lung cancer [53
]. However, other reports studying the exon 15 polymorphism in Danish and Chinese populations have found an increased risk for developing lung cancer for the 939Gln
], similar to our results.
alleles in the XPD
gene have been associated with a reduced capacity to repair BPDE and UV-induced damage in host cell reactivation assays [48
] and with a higher DNA adduct, chromosomal aberrations, and single-strand breaks level which is interpreted as lower repair efficiency [27
]. Our results confirm an association between these polymorphisms and the risk of developing lung cancer, and extend previous findings [17
Our results for the stratified analysis are supported by biological evidence. Tobacco smoke increases the risk of lung cancer and increases the risk for all histological types of this cancer, including adenocarcinoma [61
]. Our results showed higher risk for adenocarcinoma, although the reason for the observed histology-dependent difference in the genetic effect conferred by these polymorphisms is unknown, being perhaps a bit too hypothetical, it may be attributable to differences in the carcinogenesis pathways among the histological types of lung cancer. Various lines of evidence have suggested that the histological type of lung cancer may be determined by the particular initiating agent to which an individual is exposed [62
], which need to be verified in further studies. Therefore, genetic factors involved in susceptibility could be different between the histological subtypes of lung cancer [21
Contrary to the results observed with polymorphisms in genes that participate in the NER mechanism, the polymorphisms studied in XRCC1
, implicated in other DNA repair processes such as BER and DSBR, were not associated to the global individual susceptibility to develop lung cancer. Previous studies of XRCC1 Arg399Gln polymorphism have shown contradictory results, several reports have found association with different types of cancer, including colorectal, breast, lung or melanoma [64
], while other reports have failed to find association with some of these pathologies, or even found a protective effect [71
]. Our data showed no association between XRCC1 Arg399Gln and lung cancer risk, but 399Gln/Gln
genotype showed a not significant increased risk for light smokers, suggesting any kind of effect modification as Hung et al. concluded for all smoking related cancers [74
]. These results fit in studies showing 399Gln
allele may be associated with higher mutagen sensitivity and higher levels of DNA adducts [75
] who reported that never smokers carriers of 399Gln
had higher DNA adduct levels than current smokers.
allele has previously been associated with less efficient DNA repair [75
], as well as an increased number of centrosomes and binucleated cells [76
]. However, it has also been shown that the common and the variant XRCC3
alleles are functionally equivalent in the double-strand break repair pathway [77
], which may explain the lack of association between XRCC3 Thr241Met polymorphism and lung cancer risk shown in several studies [41
]. In the Caucasian population, there are inconclusive and conflicting results: several studies have found an increased risk for non small cell carcinoma and lung cancer [19
], while other studies have shown a protective effect, once more for non small cell carcinoma and ever smokers [20
]. Our study showed a statistically significant protective effect for squamous cell carcinoma, but it is difficult to assess the effect of this single common sequence variant because it might not be detectable in population association studies being necessary larger samples.
We have found that polymorphisms in NER genes increase the risk of developing lung cancer, while no association was found between polymorphisms in BER and DSBR genes and lung cancer risk. These results might reflect differences in the etiology of different carcinomas, or a more important role of the NER repair pathways in the development of lung cancer. In this regard, numerous studies have shown that most DNA lesions caused by tobacco-smoke carcinogens are repaired by the NER mechanism [8
], suggesting that this particular cancer could be more susceptible to polymorphisms affecting genes implicated in the NER pathway.
Although the relative risks for individuals carrying the polymorphisms in XPC and XPD genes are modest (ORs < 1.52), these polymorphisms could account for a large proportion of lung cancers, as they are very common in the population. In fact, between 9% and 16% of individuals are homozygous for the high-risk genotypes (XPC PAT+/+ or XPD 751Gln/Gln). In this regard, we observed a borderline combined effect between these polymorphisms and the risk of lung cancer, as individuals homozygous for both risk genotypes showed a further increase in the risk of developing lung cancer than that observed for the individual polymorphisms (adjusted OR = 2.25; 95% CI 0.83–6.13, P = 0.202). This combined effect of XPC and XPD polymorphisms could support the hypothesis for this population that changes in genes implicated in the NER repair pathway contribute to the susceptibility of developing lung cancer, and the combination of genotypes with a reduced ability to repair DNA lesions could result in a higher risk of developing this disease.
Similarly, when we combined XRCC3 241Met/Met genotype with the XPC PAT+/+ or the XPD 751Gln/Gln genotypes, an increased risk was observed (Table ). These results could suggest that the DSBR mechanism might also play a role in the development of lung cancer when combined with certain NER genes genotypes. Indeed, smoking induces a great variety of DNA damage, which must be repaired by more than one repair pathway, being NER the main pathway and DSBR the second, thus the combined occurrence of genetic variants in these two repair pathways might contribute to a greater risk of lung cancer. The approach of using combined analysis of polymorphisms may represent an alternative way of analyzing the overall effect of the different genetic variants as well as the potential joint effect of these genes.