Associations of ERCC 4 rs 1800067 Polymorphism with Cancer Risk : an Updated Meta-analysis

DNA repair system play a pivotal role in maintaining normal functions of cells (Lindahl and Wood, 1999). In recent years, emerging numbers of studies investigated the relation of DNA damage and repair with the occurrence of cancer. Humans cells employ multiple and specific repair pathways such as nucleotide excision repair (NER) and base excision repair (BER) to repair DNA damage. NER is a versatile system that monitors and repairs a variety of DNA damage, including UV-induced cyclobutane pyrimidine dimers, bulky adducts and DNA crosslinks (de Laat et al., 1999). Excessive DNA damage may give rise to cancer, therefore NER exert important effect in the initiation and development of cancer. NER system is composed of multiple steps including damage recognition, damage demarcation and unwinding,


Introduction
DNA repair system play a pivotal role in maintaining normal functions of cells (Lindahl and Wood, 1999).In recent years, emerging numbers of studies investigated the relation of DNA damage and repair with the occurrence of cancer.Humans cells employ multiple and specific repair pathways such as nucleotide excision repair (NER) and base excision repair (BER) to repair DNA damage.NER is a versatile system that monitors and repairs a variety of DNA damage, including UV-induced cyclobutane pyrimidine dimers, bulky adducts and DNA crosslinks (de Laat et al., 1999).Excessive DNA damage may give rise to cancer, therefore NER exert important effect in the initiation and development of cancer.

Associations of ERCC4 rs1800067 Polymorphism with Cancer
Risk: an Updated Meta-analysis Quan Yuan, Jing-Wei Liu, Cheng-Zhong Xing, Yuan Yuan* damage incision and new strand ligation (Friedberg, 2001).Each step requires corresponding functional proteins, and various factors are involved in this complex and precise process, including ERCC1, ERCC2, ERCC3, ERCC4, ERCC5, XPA, XPC, DDB2 and so on.Excision repair cross-complementation group 4 (ERCC4), alternatively known as XPF, is an important member of NER system.The XPF-ERCC1 heterodimer is responsible for the 5' incision of the dual incision process in the NER pathway (Fagbemi et al., 2011).Besides, the XPF-ERCC1 heterodimer also exerts an important role in maintaining telomere stability and repairing interstrand cross-links (Niedernhofer et al., 2004).
Studies suggested that ERCC4 gene possessed hundreds of single nucleotide polymorphisms (SNPs), some of which have been linked to the susceptibilities to bladder cancer, lung cancer, breast cancer, colorectal cancer and so on.ERCC4 rs1800067 A/G polymorphism could cause the change from Arg to Gln, which may lead to the alternation of ERCC4 protein function and thus influencing the role of NER in carcinogenesis.A number of researches have been conducted to explore the association of ERCC4 rs1800067 A/G polymorphism with susceptibility to cancer.However, the results from individual studies were inconsistent (Huang et al., 2006;Yu et al., 2012;Wang et al., 2013;Steck et al., 2014).Previous meta-analysis did not find significant association of this polymorphism with cancer risk: Ding et al. found ERCC4 rs1800067 polymorphism was not significantly associated with risk of breast cancer in 2011 (Ding et al., 2011); Shi et al. (2012) did not linked the polymorphism to altered risk of cancer in 2012.
Recently, several studies continued to explore the association of ERCC4 rs1800067 polymorphism with risk of cancer but the results were controversial (Cheng et al., 2013;Santos et al., 2013;Wang et al., 2013;Wyss et al., 2013;Kohlhase et al., 2014;Steck et al., 2014).Aiming at elucidating the exact relation between ERCC4 rs1800067 A/G polymorphism with risk of cancer, we perform the current meta-analysis by collecting the data from published case-control studies concerning the role of rs1800067 polymorphism in carcinogenesis.

Identification and eligibility of relevant studies
Literatures of electronic databases including PubMed, Web of Science, EMBASE, Wanfang and Chinese National Knowledge Infrastructure (CNKI) were systematically searched using different combinations of the search terms including "XPF/ERCC4/Xeroderma pigmentosum group F", "polymorphism/mutation/variant" and "cancer/neoplasm/malignancy".References cited in each eligible literature were further searched manually for potentially available studies.When overlapping data exists, only the largest study was adopted.The author was contacted for specific raw data if the data provided in the publications were not sufficient.The last search date was March 5 th , 2014.

Inclusion and exclusion criteria
Studies included in this meta-analysis must meet the inclusion criteria as follows: case-control studies investigating the association between ERCC4 gene rs1800067 polymorphism and risk of cancer; studies with sufficient raw data for evaluating odds ratios (OR) and their 95% confidence interval (CI); studies published in English or Chinese.The main reasons for exclusion were no relevance; reviews or meta-analysis; duplicate publications; animal experiments; functional investigations; and not for specific polymorphisms.

Data extraction
Two authors (Quan Yuan and Jingwei Liu) independently extracted the data from the eligible studies.The following data was extracted from individual study: first author name, publication year, ethnicity of the population, type of studied cancer, the source of the control group, numbers of cases and controls and genotyping methods of ERCC4 polymorphism.The conflict was resolved after discussion and consensus was finally reached on all of the extracted information.

Statistical analysis
The statistical analysis was performed by Stata software (Version 11.0; StataCorp, College Station, TX).ORs and their corresponding 95%CI were applied to evaluate the strength of association between ERCC4 gene rs1800067 polymorphism and cancer risk.P value <0.05 was considered as statistically significant.Heterogeneity was assessed by using Q statistic (P <0.10 indicates significant heterogeneity between studies) and I-squared (I 2 ) value (Higgins and Thompson, 2002).A fixed-effects model using Mantel-Haenszel method (Mantel and Haenszel, 1959) was performed to calculate the pooled ORs when heterogeneity between studies was not significant.Otherwise, a random-effects model using DerSimonian and Laird (1986) method was used.Sensitivity analysis was carried out to explore heterogeneity when significant heterogeneity was indicated.Subgroup analyses were performed to explore the effects of ethnicities and cancer type.In addition, publication bias was evaluated by Begg andMazumdar (1994) test andEgger et al. (1997) test, respectively.P value < 0.1 for Begg's and Egger's tests suggests significant publication bias.
The main characteristics of the studies selected in this meta-analysis were summarized in

Quantitative data synthesis
Results of the association between ERCC4 rs1800067 A/G polymorphism and cancer risk was summarized in Table 2.There was no significant association between ERCC4 rs1800067 AA or AG genotypes and overall risk of cancer (AA vs. GG: OR=0.998, 95%CI=0.670-1.486,P=0.992; AG vs. GG: OR=0.970, 95%CI=0.888-1.061,P=0.508).Dominant genetic model also did not demonstrate significant association of (AA+AG) genotype carriers with altered risk of overall cancer compared with GG genotype carriers (OR=0.985,95%CI=0.909-1.068,P=0.719, Figure 2).In addition, no significant association was observed between A allele of ERCC4 rs1800067 A/G  Subgroup analysis was then performed to explore the effect of different ethnicities and cancer types.For different subgroups of ethnicities, no significant association was found between ERCC4 rs1800067 A/G polymorphism and altered cancer risk (Table 2).
For subgroup of glioma, AA genotype carriers were observed to be significantly associated with decreased risk of glioma compared with wild-type GG genotype individuals (OR=0.523,95%CI=0.275-0.993,P=0.048).For subgroup of lung cancer, A allele of ERCC4 rs1800067 A/G polymorphism was significantly associated with decreased risk of lung cancer compared with G allele (OR=0.806,95%CI=0.697-0.931,P=0.003).

Heterogeneity test, sensitivity analysis and publication bias
In some comparisons of ERCC4 rs1800067 A/G polymorphism and cancer risk, significant heterogeneities were observed where random effect model was used.Subgroup analysis reduced the heterogeneities in some genetic models (Table 2).We therefore performed sensitivity analysis to detect the influence of individual study on the pooled estimate by omitting one study from the pooled analysis each time.The outcomes suggested that no individual study significantly affected the pooled OR (figure not shown), indicating that the results of the meta-analysis were robust.The Begg's test and Egger's test were conducted to quantitatively assess the publication bias of the included studies.The detailed information for publication bias test was shown in Table 3.No significant publication bias  was observed in the present meta-analysis.Besides, the funnel plot which reflects the studies of the association of (AA+AG) genotype with cancer risk was presented in Figure 4.

Discussion
As a key member of NER system, ERCC4 polymorphism might be related to cancer susceptibilities.The relation of DNA repair gene polymorphism and cancer risk has been widely studies (Li et al., 2013;Yang et al., 2013).Previous studies concerning the association between ERCC4 rs1800067 A/G polymorphism and risk of cancer were controversial.By conducting the present meta-analysis, we suggested that ERCC4 rs1800067 A/G polymorphism might not be associated with risk of overall cancer.However, individuals with AA genotype were associated with significantly reduced risk of glioma compared with wild-type GG genotype; A allele was associated with significantly reduced risk of lung cancer.
Human ERCC4 gene, also named as XPF, is located at chromosome 6p13.12,consisting of eleven exons and ten introns.ERCC4 exert its functions in the irreversible dual-incision process during NER by forming an obligate heterodimer complex with ERCC1 and then operating 5' incision to the DNA lesion (Nouspikel, 2009).More importantly, the catalyzing domain that determines NER activity is in ERCC4 (Enzlin and Scharer, 2002).Additionally, low expression of ERCC4 has been reported to be related with an elevated risk of head and neck cancer (Wei et al., 2005).Key polymorphism of ERCC4 might affect the expression and function of ERCC4 protein and its role in NER system, thus altering individual's susceptibility to cancer.Driven by this hypothesis, an increasing number of studies investigated the association between ERCC4 rs1800067 A/G polymorphism and risk of cancer in recent years.However, the results from individual studies were inconclusive.
By collecting and analyzing the data from published twenty-six articles including thirty studies conducted on the association of ERCC4 rs1800067 polymorphism with cancer risk, we found that ERCC4 rs1800067 AA or AG genotypes was not significantly associated with overall risk of cancer (AA vs. GG: OR=0.998,P=0.992; AG vs. GG: OR=0.970,P=0.508).Dominant genetic model also did not demonstrate significant association of (AA+AG) genotype carriers with altered risk of overall cancer compared with GG genotype carriers (OR=0.985,P=0.719).In addition, no significant association was observed between A allele of ERCC4 rs1800067 A/G polymorphism and altered cancer risk compared with G allele (OR=0.952,P=0.381), indicating that this polymorphism might not be associated with overall risk of cancer.Subgroup analysis found that individuals with AA genotype were associated with significantly reduced risk of glioma compared with wild-type GG genotype; A allele was associated with significantly reduced risk of lung cancer, suggesting that ERCC4 rs1800067 A/G polymorphism might be involved in the carcinogenesis of glioma and lung cancer.Considering the limited number of studies for certain subgroups, future large-scale investigations are still needed to draw a more reliable conclusion.
Our meta-analysis had advantages than the previously published meta-analysis conducted on the association of ERCC4 rs1800067 polymorphism and cancer risk.This meta-analysis included all types of cancer into pooled analysis, while the Ding et al. (2011) meta-analysis only investigated breast cancer susceptibility.In addition, Shi et al. (2012) meta-analysis did not find significant association of ERCC4 rs1800067 polymorphism with risk of overall cancer or in any subgroup.In the present meta-analysis, we included twenty-six articles including thirty studies (19514 cases and 20777 controls) and revealed significant association between ERCC4 rs1800067 polymorphism and certain cancer subtypes.
Several limitations should also be acknowledged in the present meta-analysis.First, the studied sample was relatively not large for certain subgroup analysis.Second, obvious heterogeneity was observed in the comparisons of ERCC4 rs1800067 A/G polymorphism and risk of cancer, which could not be fully explained by subgroup analysis.Third, the ethnicities of all the included studies were mainly Caucasians, which may limit the generalizability of our conclusion.Fourth, other important raw data such as age, sex and family history were not available for each individual study so that we could not obtain results with adjustments by other co-variables.
In conclusion, to be concluded, this meta-analysis indicated that ERCC4 rs1800067 A/G polymorphism might not be associated with risk of overall cancer.However, individuals with AA genotype were associated with significantly reduced risk of glioma compared with wild-type GG genotype; A allele was associated with significantly reduced risk of lung cancer compared with G allele.Future large-scale studies performed in multiple populations are warranted to confirm our results.

Figure 1 .
Figure 2. Forest Plot for the Association between ERCC4 rs1800067 A/G Polymorphism and Cancer Risk ((AA+AG) vs. GG) Figure 4. Funnel Plot for Studies of Association between ERCC4 rs1800067 A/G Polymorphism and Cancer Risk ((AA+AG) vs. GG)

Table 1 .
All the included studies were case-control designed published in English.Twenty-six articles including thirty studies (19514 cases and 20777 controls) were eventually included for meta-analysis.The types of cancer studied in relation to ERCC4 rs1800067 polymorphism covered breast cancer, colorectal cancer, thyroid cancer, head and neck cancer, glioma, laryngeal cancer, lung cancer and pancreatic cancer.The ethnicities of the included studies DOI:http://dx.doi.org/10.7314/APJCP.2014.15.18.7639ERCC4Polymorphism and Cancer Risk -an Updated Meta-analysishad Caucasian, Asian, African American and Latino.