8
CLINICAL STUDIES XRCC1genetic polymorphism Arg399Gln and hepatocellular carcinoma risk: a meta-analysis Fei Liu, Bo Li, Yonggang Wei, Lvnan Yan, Tianfu Wen, Jichun Zhao and Mingqing Xu Department of Liver and Vascular Surgery, West China Hospital, Sichuan University, Chengdu, Sichuan Province, China Keywords hepatocellular carcinoma – meta-analysis – polymorphism – XRCC1 Abbreviations CI, confidence interval; HBV, hepatitis B virus; HCC, hepatocellular carcinoma; HCV, hepatitis C virus; HWE, Hardy–Weinberg equilibrium; OR, odds ratio; SNP, single nucleotide polymorphisms; XRCC1, X-ray repair cross-complementing group 1. Correspondence Yonggang Wei, Department of Liver and Vascular Surgery, West China Hospital, Sichuan University, 37 Guo Xue Road, Chengdu, 610041, Sichuan Province, China. Tel: 186 28 85422476 Fax: 186 28 85423724 e-mail: [email protected] Received 17 October 2010 Accepted 21 February 2011 DOI:10.1111/j.1478-3231.2011.02508.x Abstract Background: Studies investigating the association between X-ray repair cross- complementing group 1 (XRCC1) genetic polymorphism Arg399Gln and hepatocellular carcinoma (HCC) risk report conflicting results. The aim of this study was to quantitatively summarize the evidence for such a relation- ship. Methods: Two investigators independently searched the Medline, Em- base, CNKI and Chinese Biomedicine Database. Summary odds ratios (ORs) and 95% confidence intervals (95% CIs) for XRCC1 polymorphism and HCC were calculated in a fixed-effects model (the Mantel–Haenszel method) and a random-effects model (the DerSimonian and Laird method) when appro- priate. The pooled ORs were performed for a codominant model (Gln/Gln vs. Arg/Arg, Arg/Gln vs. Arg/Arg), a dominant model (Gln/Gln1Arg/Gln vs. Arg/Arg) and a recessive model (Gln/Gln vs. Arg/Gln1Arg/Arg). Results: This meta-analysis included 11 case–control studies, which included 2208 HCC cases and 3265 controls. Overall, the variant genotypes (Gln/Gln and Arg/Gln) of Arg399Gln were not associated with HCC risk when compared with the wild-type Arg/Arg homozygote (Gln/Gln vs. Arg/Arg, OR = 1.01, 95% CI = 0.79–1.28; Arg/Gln vs. Arg/Arg, OR = 1.09, 95% CI = 0.81–1.45). Simi- larly, no associations were found in the dominant and recessive models (dominant model, OR = 1.12, 95% CI = 0.85–1.47; recessive model, OR = 0.99, 95% CI = 0.79–1.25). Limiting the analysis to the studies within Hardy–Weinberg equilibrium, the results were persistent and robust. When stratifying for ethnicity, country/region and source of controls, no evidence of a significant association was observed in any subgroup. No publication bias was found in the present study. Conclusion: No association is found between the XRCC1 polymorphism Arg399Gln and the risk of HCC. Hepatocellular carcinoma (HCC), which is the fifth most common cancer and the third leading cause of cancer- related death worldwide, is a global health problem (1). The estimated annual number of cases exceeds 500 000, with a mean annual incidence of around 3–4% (2). Patients with HCC have a poor prognosis, with a 5-year survival rate of 5% in developing countries in 2002 (1) because of the lack of effective therapy in most patients (3). Aetiologically, carcinogenesis of HCC is a complex, multistep and multifactor process, in which many factors are implicated. As we know, chronic infection with hepatitis B virus (HBV) or hepatitis C virus (HCV) is the most well-established environmental risk factor for HCC worldwide. However, only a fraction of HBsAg carriers eventually develop HCC and only 2.5% of HCV- infected individuals develop HCC later in life (4). The exact mechanism of hepatocarcinogenesis is still incom- pletely understood, and the risk factors for HCC still need to be further elucidated. Human cancer can be initiated by DNA damage caused by UV, ionizing radiation and environmental chemical agents. Importantly, to counteract the deleter- ious consequences of the DNA-damaging agents, evolution has moulded a number of DNA repair systems that as a whole take care of most of the insults infli- cted on a cell’s vital genetic information. The repairing of different types of DNA damages is important for safeguarding genomic integrity (5). Among these, the base excision repair (BER) pathway constitutes the primary defence against lesions generated by ionizing radiation and strong alkylating agents as well as lesions formed by endogenous DNA-damaging agents like viruses (6). The XRCC1 protein, which is encoded by the X-ray repair cross-complementing group 1 (XRCC1) gene, is an important component of the BER pathway. XRCC1 fixes base damage and DNA single-strand breaks caused by ionizing radiation and alkylating agents by directly Liver International (2011) 802 c 2011 John Wiley & Sons A/S Liver International ISSN 1478-3223

XRCC1 genetic polymorphism Arg399Gln and hepatocellular carcinoma risk: a meta-analysis

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CL IN ICAL STUDIES

XRCC1genetic polymorphismArg399Gln and hepatocellular carcinomarisk: ameta-analysisFei Liu, Bo Li, Yonggang Wei, Lvnan Yan, Tianfu Wen, Jichun Zhao and Mingqing Xu

Department of Liver and Vascular Surgery, West China Hospital, Sichuan University, Chengdu, Sichuan Province, China

Keywords

hepatocellular carcinoma – meta-analysis –

polymorphism – XRCC1

Abbreviations

CI, confidence interval; HBV, hepatitis B virus;

HCC, hepatocellular carcinoma; HCV,

hepatitis C virus; HWE, Hardy–Weinberg

equilibrium; OR, odds ratio; SNP, single

nucleotide polymorphisms; XRCC1, X-ray

repair cross-complementing group 1.

Correspondence

Yonggang Wei, Department of Liver and

Vascular Surgery, West China Hospital, Sichuan

University, 37 Guo Xue Road, Chengdu,

610041, Sichuan Province, China.

Tel: 186 28 85422476

Fax: 186 28 85423724

e-mail: [email protected]

Received 17 October 2010

Accepted 21 February 2011

DOI:10.1111/j.1478-3231.2011.02508.x

AbstractBackground: Studies investigating the association between X-ray repair cross-complementing group 1 (XRCC1) genetic polymorphism Arg399Gln andhepatocellular carcinoma (HCC) risk report conflicting results. The aim ofthis study was to quantitatively summarize the evidence for such a relation-ship. Methods: Two investigators independently searched the Medline, Em-base, CNKI and Chinese Biomedicine Database. Summary odds ratios (ORs)and 95% confidence intervals (95% CIs) for XRCC1 polymorphism and HCCwere calculated in a fixed-effects model (the Mantel–Haenszel method) and arandom-effects model (the DerSimonian and Laird method) when appro-priate. The pooled ORs were performed for a codominant model (Gln/Gln vs.Arg/Arg, Arg/Gln vs. Arg/Arg), a dominant model (Gln/Gln1Arg/Gln vs.Arg/Arg) and a recessive model (Gln/Gln vs. Arg/Gln1Arg/Arg). Results: Thismeta-analysis included 11 case–control studies, which included 2208 HCCcases and 3265 controls. Overall, the variant genotypes (Gln/Gln and Arg/Gln)of Arg399Gln were not associated with HCC risk when compared withthe wild-type Arg/Arg homozygote (Gln/Gln vs. Arg/Arg, OR = 1.01, 95%CI = 0.79–1.28; Arg/Gln vs. Arg/Arg, OR = 1.09, 95% CI = 0.81–1.45). Simi-larly, no associations were found in the dominant and recessive models(dominant model, OR = 1.12, 95% CI = 0.85–1.47; recessive model,OR = 0.99, 95% CI = 0.79–1.25). Limiting the analysis to the studies withinHardy–Weinberg equilibrium, the results were persistent and robust. Whenstratifying for ethnicity, country/region and source of controls, no evidence ofa significant association was observed in any subgroup. No publication biaswas found in the present study. Conclusion: No association is found betweenthe XRCC1 polymorphism Arg399Gln and the risk of HCC.

Hepatocellular carcinoma (HCC), which is the fifth mostcommon cancer and the third leading cause of cancer-related death worldwide, is a global health problem (1).The estimated annual number of cases exceeds 500 000,with a mean annual incidence of around 3–4% (2).Patients with HCC have a poor prognosis, with a 5-yearsurvival rate of 5% in developing countries in 2002 (1)because of the lack of effective therapy in most patients(3). Aetiologically, carcinogenesis of HCC is a complex,multistep and multifactor process, in which many factorsare implicated. As we know, chronic infection withhepatitis B virus (HBV) or hepatitis C virus (HCV) isthe most well-established environmental risk factor forHCC worldwide. However, only a fraction of HBsAgcarriers eventually develop HCC and only 2.5% of HCV-infected individuals develop HCC later in life (4). Theexact mechanism of hepatocarcinogenesis is still incom-pletely understood, and the risk factors for HCC stillneed to be further elucidated.

Human cancer can be initiated by DNA damagecaused by UV, ionizing radiation and environmentalchemical agents. Importantly, to counteract the deleter-ious consequences of the DNA-damaging agents,evolution has moulded a number of DNA repair systemsthat as a whole take care of most of the insults infli-cted on a cell’s vital genetic information. The repairingof different types of DNA damages is importantfor safeguarding genomic integrity (5). Among these,the base excision repair (BER) pathway constitutes theprimary defence against lesions generated by ionizingradiation and strong alkylating agents as well as lesionsformed by endogenous DNA-damaging agents likeviruses (6).

The XRCC1 protein, which is encoded by the X-rayrepair cross-complementing group 1 (XRCC1) gene, isan important component of the BER pathway. XRCC1fixes base damage and DNA single-strand breaks causedby ionizing radiation and alkylating agents by directly

Liver International (2011)802 c� 2011 John Wiley & Sons A/S

Liver International ISSN 1478-3223

interacting with polymerase-b, DNA ligase III and poly(ADP ribose) polymerase (PARP) (7).

The XRCC1 gene is located on chromosome 19q13.2(8), spans a genetic distance of 32 kb, comprises of 17 exonsand encodes a 70-kDa protein consisting of 633 aminoacids (9). Although there are more than 300 validatedsingle nucleotide polymorphisms (SNPs) in the XRCC1gene reported in the dbSNP database (http://www.ncbi.nlm.nih.gov/SNP), three of which are common (10) andlead to amino acid substitutions in XRCC1 at codon 194(exon 6, base C to T, amino acid Arg to Trp, dbSNP no.rs1799782), codon 280 (exon 9, base G to A, amino acidArg to His, dbSNP no. rs25489) and codon 399 (exon 10,base G to A, amino acid Arg to Gln, dbSNP no.rs25487),these non-conservative amino acid changes may alterXRCC1 function. This change in protein biochemistryleads to the supposition that variant alleles may diminishrepair kinetics, thereby influencing susceptibility to adversehealth effects, including cancer.

Over the last two decades, a number of case–controlstudies were conducted to investigate the association be-tween XRCC1 Arg399Gln polymorphism and HCC risk inhumans. But these studies reported conflicting results. Noquantitative summary of the evidence has ever been per-formed. The purpose of this meta-analysis was to quantita-tively summarize the evidence for such a relationship.

Materials and methods

Publication search

We searched the PubMed, Embase, CNKI (China Na-tional Knowledge Infrastructure) and Chinese Biomedi-cine databases for all articles on the association betweenXRCC1 polymorphisms and HCC risk (last search up-date 15 October 2010). The following key words wereused: ‘liver cancer’ or ‘hepatocellular carcinoma’, ‘X-rayrepair cross – complementing group 1’ or ‘XRCC1’ and‘polymorphism’ or ‘variant’. The search was withoutrestriction on language, conducted on human subjects.The reference lists of reviews and retrieved articles werehand searched at the same time. We did not considerabstracts or unpublished reports. If more than one articlewas published by the same author using the same caseseries, we selected the study where the most individualswere investigated.

Inclusion and exclusion criteria

We reviewed abstracts of all citations and retrieved studies.The following criteria were used to include publishedstudies: (i) case–control studies were conducted to evaluatethe association between XRCC1Arg399Gln polymorphismand HCC risk; (ii) sufficient genotypes data were presentedto calculate the odds ratios (ORs) and 95% confidenceintervals (CIs); and (iii) the paper should clearly describeHCC diagnoses and the sources of cases and controls. Themajor reasons for exclusion of studies were (i) no control;(ii) duplicate; and (iii) no sufficient data were reported.

Data extraction

Two investigators (F. L. and Y. W.) extracted informationfrom all eligible publications independently according tothe inclusion criteria listed above. Disagreements wereresolved by discussion between the two investigators. Thefollowing characteristics were collected from each study:first author, year of publication, country/region of thefirst or corresponding author, ethnicity, number of casesand controls, genotyping methods, matching variables,minor allele frequency (MAF) in controls and evidence ofHardy–Weinberg equilibrium (HWE) (Table 1). Differ-ent ethnicities were categorized as Asian and non-Asian.

Statistical analysis

We first assessed HWE in the controls for each studyusing a goodness-of-fit test (chi-square or Fisher’s exacttest), and a Po 0.05 was considered as significantdisequilibrium. The strength of the association betweenHCC and the XRCC1 Arg399Gln polymorphism wasestimated using ORs, with the corresponding 95% CIs.The pooled ORs were performed for a codominantmodel (Gln/Gln vs. Arg/Arg, Arg/Gln vs. Arg/Arg), adominant model (Gln/Gln1Arg/Gln vs. Arg/Arg) and arecessive model (Gln/Gln vs. Arg/Gln1Arg/Arg). We alsocarried out the stratified analyses by ethnicity, country/region, HWE in controls and source of controls.

Both the Cochran’s Q statistic (11) to test for hetero-geneity and the I2 statistic to quantify the proportion of thetotal variation because of heterogeneity (12) were calcu-lated. A P value of more than the nominal level of 0.10 forthe Q statistic indicated a lack of heterogeneity acrossstudies, allowing for the use of a fixed-effects model (theMantel–Haenszel method) (13); otherwise, the random-effects model (the DerSimonian and Laird method) wasused (14). To explore sources of heterogeneity acrossstudies, we performed logistic meta-regression analyses.We examined the following study characteristics: ethnicity,HWE in controls (yes/no), matched controls (yes/no),genotyping methods and study sample size (� 500 and4 500 subjects). Sensitivity analysis was performed toassess the stability of the results.

Several methods were used to assess the potentialpublication bias. A visual inspection of funnel plot asym-metry was conducted. Begg’s rank correlation method (15)and Egger’s weighted regression method (16) were used tostatistically assess publication bias (Po 0.05 was consid-ered statistically significant). All analyses were performedusing STATA software, version 11.0 (STATA Corp., CollegeStation, TX, USA). All the P values were two-sided.

Results

Characteristics of studies

A total of 15 published studies (17–31) with full-textarticles examined the association of polymorphismArg399Gln of the XRCC1 gene with HCC were found,four of which were the dissertations of postgraduate

Liver International (2011)c� 2011 John Wiley & Sons A/S 803

Liu et al. XRCC1 polymorphism and HCC risk

students (27–30). Four studies (17, 29–31) were excludedbecause they investigated the same population of re-ported articles. As summarized in Table 1, 11 eligiblearticles (18–28) [six in English (18–23) and five (24–28)in Chinese] were selected for this meta-analysis, includ-ing 2208 HCC cases and 3265 controls.

There were nine studies of Asian patients, one study ofAfrican patients and one study of Caucasian patients.Studies had been carried out in mainland China, Taiwan,India, Gambia and France. The cases definition used in theindividual studies were pathologically or clinically diag-nosed with HCC. Concerning controls’ selection, fourstudies were from healthy volunteer or blood donor, twostudies from HBV carriers, three studies from hospitalpatients without HCC (mixed by chronic hepatitis virus-positive and -negative individuals) and two studies wereseparately used a healthy population and chronic hepatitisvirus (HBV/HCV) carriers as controls. All studies extractedDNA from peripheral blood and a classic polymerase chainreaction–restriction fragment length polymorphism assaywas used in 10 out of 11 studies. Only one study (18)described the use of positive controls and a differentgenotyping assay to confirm the data. The genotypedistributions among the controls of all studies wereconsistent with HWE, except for three studies (18, 25, 27).

Quantitative synthesis

Table 2 lists the main results of this meta-analysis,and Figure 1 shows the association of HCC risk withXRCC1 polymorphism Arg399Gln. Overall, the variantgenotypes (Gln/Gln and Arg/Gln) of Arg399Gln werenot associated with HCC risk when compared with thewild-type Arg/Arg homozygote (Gln/Gln vs. Arg/Arg,OR = 1.01, 95% CI = 0.79–1.28; Arg/Gln vs. Arg/Arg, OR =1.09, 95% CI = 0.81–1.45). Similarly, no associationswere observed in the dominant and recessive models(dominant model, OR = 1.12, 95% CI = 0.85–1.47; reces-sive model, OR = 0.99, 95% CI = 0.79–1.25).

On the basis of the potential overestimation of the trueeffect of the polymorphism on the HCC risk, we strati-fied these studies according to ethnicity, country/region,source of controls and HWE in controls. Because of theinadequate sample populations available for Caucasianand African groups, different ethnicities were categorizedas Asian and non-Asian. Similarly, different countries/regions were categorized as mainland China and others.Different sources of controls were defined as chronichepatitis virus (HBV/HCV) positive and negative. Instratified analyses, the variant genotypes (Gln/Gln andArg/Gln) had no significant relationship with HCC in allof the subgroups, compared with the wild type. Similarresults were observed in the recessive model and thedominant model (Table 2).

Heterogeneity analysis

There was heterogeneity among studies in the overallcomparisons and also subgroup analyses in the heterozygoteTa

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Liver International (2011)804 c� 2011 John Wiley & Sons A/S

XRCC1 polymorphism and HCC risk Liu et al.

Table 2. Stratified analyses of the XRCC1 polymorphism Arg399Gln on hepatocellular carcinoma risk

Variables N�

Gln/Gln vs. Arg/Arg Arg/Gln vs. Arg/Arg Dominant model Recessive model

OR (95% CI) Pw OR (95% CI) Pw OR (95% CI) Pw OR (95% CI) Pw

Total 11 1.01 (0.79, 1.28) 0.19 1.09 (0.81, 1.45) o 0.001 1.12 (0.85, 1.47) o 0.001 0.99 (0.79, 1.25) 0.51

Ethnicity

Asian 9 1.05 (0.82, 1.35) 0.29 1.15 (0.84, 1.59) 0.001 1.19 (0.89, 1.59) o 0.001 1.01 (0.79, 1.29) 0.49

Non-Asian 2 0.64 (0.28, 1.45) 0.10 0.81 (0.32, 2.03) 0.045 0.80 (0.29, 2.19) 0.02 0.80 (0.37, 1.74) 0.21Country/region

Mainland China 6 0.87 (0.58, 1.31) 0.67 1.21 (0.65, 2.28) o 0.001 1.28 (0.81, 2.01) o 0.001 0.82 (0.55, 1.22) 0.90

Others 5 0.96 (0.57, 1.64) 0.06 1.06 (0.89, 1.25) 0.18 0.99 (0.75, 1.31) 0.05 1.09 (0.82, 1.46) 0.22

HWE in controls

Yes 7 1.16 (0.87, 1.53) 0.25 1.04 (0.77, 1.41) 0.005 1.14 (0.84, 1.54) o 0.001 1.15 (0.87, 1.51) 0.67

No 3 0.71 (0.45, 1.12) 0.41 1.25 (0.47, 3.28) 0.003 1.09 (0.48, 2.46) 0.002 0.68 (0.44, 1.06) 0.71

Source of controls

Healthy populationz 6 0.70 (0.45, 1.09) 0.23 1.04 (0.57, 1.88) o 0.001 0.98 (0.55, 1.72) o 0.001 0.69 (0.45, 1.05) 0.51

Hepatitis virus carrier‰ 4 1.17 (0.86, 1.61) 0.19 1.05 (0.88, 1.26) 0.22 0.98 (0.74, 1.31) 0.098 1.16 (0.85, 1.57) 0.43

�Number of comparisons.

wP value of Q-test for heterogeneity test. A random-effects model was used when P value for heterogeneity test o 0.1; otherwise, a fixed-effects model

was used.

zSubjects were chronic hepatitis virus (HBV/HCV) negative.

‰Subjects were chronic hepatitis virus (HBV/HCV) positive.

CI, confidence interval; HBV, hepatitis B virus; HCV, hepatitis C virus; HWE, Hardy–Weinberg equilibrium; OR, odds ratio.

Fig. 1. Forest plots of odds ratios (ORs) with 95% confidence interval (CI) for XRCC1 codon 399 polymorphism and risk for hepatocellular carcinoma.The centre of each square represents the OR, the area of the square is the number of sample and thus the weight used in the meta-analysis, and thehorizontal line indicates the 95% CI. (a) Gln/Gln vs. Arg/Arg. (b) Arg/Gln vs. Arg/Arg. (c) Gln/Gln+Arg/Gln vs. Arg/Arg. (d) Gln/Gln vs. Arg/Gln+ Arg/Arg.

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Liu et al. XRCC1 polymorphism and HCC risk

comparison (Arg/Gln vs. Arg/Arg) and the dominantmodel. To explore sources of heterogeneity across stu-dies, we assessed heterozygote comparison and dominantmodel comparison by ethnicity, HWE in controls,matched controls, genotyping methods and study samplesize. As a result, matched controls (Arg/Gln vs. Arg/Arg:P = 0.016; dominant model: P = 0.006), but not ethnicity,HWE in controls, genotyping methods or study samplesize, were found to contribute to substantial heterogene-ity. Moreover, meta-regression analyses indicated thatmatched controls could explain 67.30% (Arg/Gln vs.Arg/Arg) and 70.08% (dominant model) of the t2.

Sensitivity analysis

In the sensitivity analysis, the influence of each study onthe pooled OR was examined by repeating the meta-analysis while omitting each study, one at a time. Thisprocedure confirmed the stability of our overall results.In addition, when excluding the studies that were not inHWE, the results were persistent and robust (Table 2).

Publication bias

Funnel plot, Begg’s and Egger’s tests were performed toevaluate publication bias of the literature on HCC. Figure2 displays a funnel plot that examined the XRCC1polymorphism and overall HCC risk included in themeta-analysis in the homozygote comparison. The shapeof funnel plots did not reveal any evidence of funnel plotasymmetry. The statistical results still did not showpublication bias (Gln/Gln vs. Arg/Arg: Begg’s testP = 0.93, Egger’s test P = 0.26; Arg/Gln vs. Arg/Arg: Begg’stest P = 0.18, Egger’s test P = 0.39; dominant model:Begg’s test P = 0.33, Egger’s test P = 0.15; recessive model:Begg’s test P = 1.00, Egger’s test P = 0.36).

Discussion

A wide variety of DNA damage may be induced bynormal endogenous metabolic processes or by environ-mental carcinogens. Most of these alterations, if notrepaired, may result in genetic instability, mutagenesisand cell death. DNA repair mechanisms are importantfor maintaining genome integrity and preventing carci-nogenesis. BER is the predominant DNA damage repairpathway for the processing of small base lesions, derivedfrom oxidation and alkylation’s damage. XRCC1 is one ofthe most important proteins in BER and is closelyassociated with BER pathway coordination by interactingwith most components of the BER short-patch pathway.Mutations of XRCC1 may increase the risk of cancers byimpairing the interaction of XRCC1 with other enzy-matic proteins and consequently altering DNA repairactivity (32, 33). In recent years, a large number ofmolecular epidemiological studies have been conductedto evaluate the role of polymorphisms Arg399Gln in theDNA repair gene XRCC1 on cancer risk; however, theresults remain conflicting rather than conclusive (34–42),including HCC (18–28).

To the best of our knowledge, this is the first systematicreview that has investigated the association of XRCC1codon 399 polymorphisms and HCC, and no evidencehas shown any association between Arg399Gln poly-morphism and HCC susceptibility. When stratifying forethnicity, country/region, HWE in controls and source ofcontrols, the results were similar. Our results were con-sistent with previous meta-analyses based on othercancer. For example, previous three meta-analyses hadconfirmed that Arg399Gln polymorphism was not asso-ciated with a risk of gastric cancer (43), colorectal cancer(44) and bladder cancer (45). However, Saadat andAnsari-Lari (46) concluded that Arg399Gln polymorph-ism had an association with breast cancer, Geng et al. (47)found that the Arg399Gln polymorphism was associatedwith a risk of prostate cancer among Asians and Kiyoharaet al. (48) found that the 399Gln/Gln genotype of theXRCC1 Arg399Gln polymorphism was associated withan increased risk of lung cancer among Asians but notamong Caucasians. Although the reasons for this appar-ent difference in risk with different tumours are as yetunknown, some possibilities should be considered.Firstly, those gene–variant associations vary in differentkinds of cancer and may result from the differentmechanisms of carcinogenesis among different kinds ofcancer. Secondly, different ethnic composition may con-tribute to the discrepancy. Different meta-analyses in-cluded different original studies that were performed indifferent races and the ethnic composition in differentmeta-analyses may be diverse. Thirdly, some methodolo-gical diversity, such as inclusion criteria, the quality oforiginal studies, selection bias, Type – error and smallsample size, can also contribute to the discrepancy.

Because the allele frequencies of polymorphismsand their effects on the cancer risk were diverse in the

Fig. 2. Funnel plot for publication bias test (Gln/Gln vs. Arg/Arg ).Each point represents a separate study for the indicated association.

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different ethnicities, we carried out subgroup analysis byethnicity. The results demonstrated that there was noassociation between XRCC1 Arg399Gln polymorphismand HCC risk in different ethnicities. The null result maybe because of the limited number of studies, with onlytwo studies from non-Asian (one Caucasian and oneAfrican) available in this meta-analysis. It is critical thatlarger and well-designed multicentric studies based onCaucasian and African-American patients should beperformed to re-evaluate the association.

It seemed that selection bias could have played a rolebecause the genotype distribution of Arg399Gln poly-morphism among control subjects disobeyed the law ofHWE in three studies (18, 25, 27). It is widely believedthat deviation from HWE may be as a result of geneticreasons including non-random mating, or the allelesreflect recent mutations that have not reached equili-brium, as well as methodological reasons includingbiased selection of subjects from the population orgenotyping errors (49, 50). Despite the reasons of dis-equilibrium, the results of genetic association studiesmight be spurious if the distribution of genotypes in thecontrol groups were not in HWE (51, 52). Hence, wecarried out subgroup analysis by HWE in controls. Whenexcluding the studies that were not in HWE, the resultswere persistent and robust, suggesting that this factorprobably had little effect on the overall estimates.

The results of meta-analyses often depend on controlselection procedures (53). Different control sources maybe a confounding factor that may impact the conclusionof our study because of case–control studies. For in-stance, some studies used a healthy population as thereference group; whereas, others selected chronic hepati-tis virus carriers without HCC as the reference group. Inorder to eliminate interference from the confoundingfactor, we performed subgroup analysis by the source ofcontrols. Our results showed that there was no significantassociation between XRCC1 Arg399Gln polymorphismand HCC risk in different controls, which confirmed thereliability of our overall results.

One of the major concerns in a sound meta-analysis isthe degree of heterogeneity that exists between the compo-nent studies because non-homogeneous data are liable toresult in misleading results. In the present study, the Q-testand I2 statistics were carried out to test the significanceof heterogeneity. Obvious heterogeneity between studieswas observed in overall comparisons and also some sub-group analyses for some models, and then meta-regressionanalysis was used to explore the sources of heterogeneity.We found that matched controls did contribute to poten-tial heterogeneity. Another important issue for any meta-analysis is publication bias because of selective publicationof reports. In the current study, Funnel plot, Begg’s andEgger’s tests were performed to evaluate this problem.Both the shape of funnel plots and the statistical results didnot show publication bias. It is worth mentioning that theresults held when the sensitivity analysis was performed,which implied that the results were reliable.

However, there are still some limitations in this meta-analysis. Firstly, this paper was limited by analysing a singleSNPs site and lack of haplotype analyses. As mentionedabove, there are three common SNPs sites (Arg399Gln,Arg280His and Arg194Trp) in the XRCC1 gene. Data frommultiple polymorphisms within a gene can be combined tocreate haplotypes, the set of multiple alleles on a singlechromosome. The analysis of haplotype can increase thepower to detect disease associations because of higherheterozygosity and stronger linkage disequilibrium withdisease-causing mutations (54–56). However, only onestudy (18) in this meta-analysis reported haplotype ana-lyses, although several studies analysed the three commonSNPs sites in the XRCC1 gene. Secondly, we did not test forgene-to-environment interactions because of the issue ofmultiple testing and the lack of sufficient studies. It ispossible for specific environmental and lifestyle factors toalter those associations between gene polymorphisms andcancer risk. For example, Lao et al. (57) concluded that theGln/Gln genotype of Arg399Gln was associated with adecreased risk of bladder cancer among ever smokers whilethe Arg399Gln polymorphism was not associated withbladder cancer risk in the total population. Thirdly, therewas significant between-study heterogeneity from studiesin overall comparisons and also subgroup analyses in theheterozygote comparison and the dominant model, andthe genotype distribution in the control group also showeddeviation from HWE in some studies.

In conclusion, this meta-analysis evaluates the relation-ship between genetic polymorphisms and HCC risk andreveals that the XRCC1 polymorphism Arg399Gln is notassociated with altered susceptibility to HCC. Becauselimited studies were from non-Asian populations, it iscritical that larger and well-designed multicentric studiesbased on Caucasian and African-American patients shouldbe performed to re-evaluate the association. Moreover,further studies estimating the effect of haplotypes andgene–environment interactions may eventually provide abetter, comprehensive understanding of the associationbetween the XRCC1 polymorphisms and HCC risk.

Acknowledgements

Fei Liu designed research/study, performed research/study, collected data, analysed data, wrote the paper,reviewed/edited the manuscript. Bo Li performed re-search/study, collected data, analyzed data, reviewed/edited the manuscript. Yonggang Wei collected data,analysed data, reviewed/edited the manuscript. LvnanYan, Tianfu Wen, Jichun Zhao designed research/study,reviewed/edited the manuscript. Mingqing Xu reviewed/edited the manuscript.

Conflict of interest statement: none declared.

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