Evaluation of Heterogeneity
There was heterogeneity among studies in overall comparisons and also subgroup analyses in 3 XRCC6 polymorphisms: rs2267437, rs5751129, and rs132793. To explore sources of heterogeneity, we evaluated the following variables: ethnicities, cancer type, source of control, study quality, genotyping methods and sample size (≤500 and >500 subjects). Galbraith plot was also used to detect the possible sources of heterogeneity when none of the above variables could explain the heterogeneity.
For the rs2267437 polymorphism, there was significant heterogeneity in overall comparisons of allele model, heterozygote model, and dominant model. None of the possible variables could explain the heterogeneity. The studies potentially causing between-study heterogeneity were identified in the allele model8,9,27), heterozygote model8,14,25, and dominant model8,9,14,25 by the Galbraith plot (Figure 4B). However, the result was altered in allele model (OR = 1.13 95% CI 1.05–1.21) when the studies were excluded.
Meta-regression analysis indicated that the “source of control” could explain 100% of the τ2 in all comparison models in rs5751129 polymorphism, and the “cancer type” could explain 100% of the τ2 in allele model in rs132793 polymorphism. Furthermore, the combination of “ethnicity” and “cancer type” could explain 100% of the τ2 in heterozygote and dominant model in rs132793 polymorphism.
Sensitivity analyses were performed by sequential removal of each eligible study to assess the influence of each individual study on the pooled OR in each comparison in the polymorphisms of rs2267437, rs5751129, and rs132793. The omission of any study made no significant difference, indicating that the results of this meta-analysis were statistically reliable (Figure 5A).
Egger test reveals evidence of publication bias in rs5751129 polymorphism (Figure 5B). The trim and fill method showed that the funnel plot needs 4 more studies to be symmetrical (Figure 6) in allele model, recessive model and homozygous model, or 2 more in heterozygous and dominant model. But the results were altered in recessive model and homozygous model. The publication bias in rs132793 polymorphism was not checked because of the limited number of relevant studies.
In this study, we performed a systematic review of association between XRCC6 polymorphisms and cancer risks based on 20 case–control studies. This is also the first time to explore the individual association between the polymorphisms of rs5751129, rs132774, rs132793, and cancer risk. The results provided evidences that the SNPs in XRCC6 promoter region might associate with the cancer risks, while SNP in the XRCC6 intron might not. In addition, the XRCC6 promoter SNPs might play different roles in various cancers, indicating that XRCC6 gene may have different roles in different cancer development, and different mechanisms promote the development of various tumors.
XRCC6 gene was involved in multiple cellular pathways, including NHEJ pathway of DSB repair.40,41 During DNA repair procedure, Ku70 acted as a scaffold to recruit the other NHEJ factors to the damage site after its bound to DNA ends.4 However, inaccurate repair could lead to cellular aberrant function, apoptosis, and chromosomal rearrangements, which promote carcinogenesis finally.42 Besides the influence of XRCC6 gene on DSB repair and genomic stability, it has some other NHEJ-independent effects. Such as the Ku70 protein in cytoplasm can prevent Bax translocate to mitochondria, and suppresses cell apoptosis.43 The defects in Ku70 may also influence cell proliferation.44
It has been demonstrated that the rs2267437 polymorphism could influence the expression level and stability of the Ku70 protein in breast cancer cells and RCC tissues.14,15 The sequence variation in the rs2267437 may affect binding activity of the adjacent CACCC box with transcription factors, resulting in decreased Ku70 expression level, and the DSB repair activity thus was affected, finally leading to increased susceptibility to cancers.45 In our study, rs2267437 polymorphism was found to might increase the cancer risks in breast cancer, HCC and RCC, while may not function in lung cancer or some other cancers, and different ethnicities might influence the association.
The rs5751129 polymorphism locates between rs132770 and rs2267437 in the promoter region of XRCC6 gene. This polymorphism was proved to be functional in HCC and RCC, with the normal tissues with C allele having a lower expression level of XRCC6 mRNA or protein.11,16 Our results indicated that rs5751129 polymorphism might play the same role in various cancers. The variation may influence the expression level or stability of XRCC6 mRNA via alternative spicing or other mechanisms. Individuals carrying C allele may exhibit heritable decreased DNA repair capacity phenotypes compared with those carrying T allele, thus they might have less protective effects on normal tissues, and increased cancer risks. However, when stratified by HWE status of controls, we found that significantly elevated risks were observed in HWE inconsistent studies, but not in HWE consistent studies. Thus there may be possibility that the presence of the C allele is in linkage disequilibrium with another mutation located outside the coding region in the XRCC6 gene, which may be important for the Ku70 expression.
The rs132770 polymorphism locates closer than rs5751129 polymorphism to the translation starting point in the XRCC6 promoter. Our meta-analysis indicating that rs132770 polymorphism might affect RCC risk in a different way from in other cancers, the RCC pathogenesis mechanism might be distinguished from other cancers when rs132770 polymorphism involved. The results were consistent with the finding that the A allele of the rs132770 polymorphism could increase the expression levels of the Ku70 mRNA in normal tissue of RCC patients.21 Xu17 also collected 3 studies but failed to find any association. The negative results in Xu study might be due to the limited studies.
It is interesting that the rs132793 polymorphism was found to play opposite roles in different cancers: it might decrease breast cancer risk, whereas increase cancer risk in “other cancers”, indicating that the rs132793 may play opposite roles in breast cancer contrast to other cancers. The rs132793 locates in the sequences downstream of stop code of XRCC6 gene, and until recently, the role of rs132793 in tumorigenesis is unclear. We hypothesized that the rs132793 polymorphism might influence breast cancer risk via some mechanisms different from other cancers. Functional studies should be carried out to explore the mechanisms involving rs132793 polymorphism in tumorigenesis in the future. The different ethnicities of the study population might influence the genetic effect of the rs132793 polymorphism on cancer susceptibility; anyhow, it seems not substantial in our study.
Our results implied that rs132774 polymorphism have no association with cancer risk in Chinese population. To validate the results, more population-based studies in other populations should be carried out in the future.
There was heterogeneity among studies in rs2267437, rs5751129, and rs132793 polymorphisms. “Source of control” and “ethnicity”, “cancer type” might explain 100% of the τ2 in the rs5751129 and rs132793 polymorphism, respectively. However, the combination of “ethnicities”, “source of control”, and “sample size” could explain only 17.6% of the τ2 in heterozygote model in rs2267437 polymorphism, implying that there may be other reasons accounting for the heterogeneity in the rs2267437 polymorphism. Publication bias was found in the rs5751129 polymorphism and trim and fill method could reduce the influence of publication bias in all models except recessive and homozygous model, implying that more cautions should be paid when elucidate the role of the rs5751129 polymorphism in cancer susceptibility. Anyhow, sensitivity analysis proved that the results of this meta-analysis were statistically reliable. Therefore, a methodologically preferable design, such as using population-based controls, is crucial to avoid selection bias and heterogeneity.
The limitations of our meta-analysis should also be discussed. First, the non-English articles were excluded in our study, which thus may bias the results of our results. Second, some low-quality studies with deviation from HWE in the control group were included in our meta-analysis. Subgroup analysis were carried out by HWE status of controls in rs5751129 polymorphism and rs132770 polymorphism, and the opposite results were found in rs5751129 polymorphism between HWE consistent/inconsistent groups, implying that low-quality studies might influence the results in rs5751129 polymorphism. Third, the number of included studies was relatively small in some subgroups, thus the specific results should be interpreted with caution.
In conclusion, this meta-analysis showed some evidence of the XRCC6 SNP polymorphisms and cancer risk, supporting the existence of association between XRCC6 polymorphisms and cancer risks in different ethnicities and cancer types. The rs2267437 polymorphism was found to be associated with a significant increase in risks of overall cancers, breast cancer, RCC and HCC, and it might increase the cancer risk in Asian population. The rs5751129 polymorphism might increase the cancer risk in overall cancers, and the rs132770 polymorphism might be associated with the increased cancer risk in RCC. Furthermore, the rs132793 polymorphism could decrease breast cancer risk and increase risks in “other cancers”. However, there was no obvious association between XRCC6 polymorphisms and risks of some “other cancer”. This may be due to the limited studies included or limited sample size in “other cancers”. Therefore, more studies with large sample are required to further confirm the results in the future. Functional studies are also needed to elucidate the roles of XRCC6 promoter polymorphisms in cancer pathogenesis. Moreover, gene–environment and gene–gene interaction analyses, as well as haplotype analysis should be carried out to clarify the role of the XRCC6 genes in cancer. Our studies may perhaps supplement for the disease monitoring of cancers in the future, and additional studies to determine the exact molecular mechanism might provide us with interventions to protect the susceptible subgroups.
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