A meta-analysis of caspase-8-652 6N del polymorphisMand digestive tract cancer risk

Haina Du,Guoxin Song,Mingzhi Fang,Yongqian Shu,Xin Zhao,Lingjun Zhu,✉

1 Department of Oncology,the Third Af fi liated Hospital of Nanjing University of T.C.M,Nanjing,Jiangsu 210000,China;

2 Departments of Pathology,3Oncology,4Pneumology,the First Af fi liated Hospital of Nanjing Medical University,Nanjing,Jiangsu 210009,China.

AbstractCaspase-8(CASP8)isone key regulator of apoptosisof T lymphocytesand isencoded by the CASP8 gene.It has been reported that the six-nucleotide deletion polymorphism(-652 6N del)of the CASP8 gene had effect on some cancer risk.Few studiesexplored theassociation between CASP8 genepolymorphisMand digestive tractcancer risk.To evaluate theassociation between the CASP8-652 6N del polymorphisMand the risk of digestive tract cancer,we conducted this meta-analysis.We found that CASP8-652 6N del polymorphisMwas associated with a signi fi cantly reduced risk of digestive tract cancer in the co-dominant model(del/del vs.ins/ins:OR=0.82,95%CI=0.72-0.95;del/ins vs.ins/ins:OR=0.92,95%CI=0.87-0.97;doMinantmodel(del/ins+del/del vs.ins/ins:OR=0.91,95%CI=0.87-0.96,recessivemodel:del/del vs.del/ins+ins/ins:OR=0.85,95%CI=0.75-0.97).In thestrati fi ed analysisby cancer types,we found that all genetic models had protective effect on gastric cancer.Similar resultswereobserved for colorectal cancer under heterozygote comparison and dominant model,but not under homozygote comparison or recessivemodel.In addition,a signi fi cantly decreased risk was found on esophageal cancer for most genetic models,except heterozygote comparison.When strati fi ed by ethnicity and source of control,an evidently decreased risk was identi fi ed in the Asian populations and population-based studies.In conclusion,there exists an association between the CASP8-652 6N del polymorphisMand reduced digestive cancer risk,especially among Asians and populationbased studies.

Keywords:caspases-8,polymorphism,digestive tract cancer risk

Introduction

The activity of many genes in fluences a cell's likelihood of activating its apoptotic program[1],which is mainly regulated by caspases,a faMily of cysteine proteases,that play a crucial role in the development and progression of cancer[2].Several meta-analyses have con fi rmed that the gene variants of caspases-2,5,7,8,9 and 10 disturbed the apoptotic mechanisMand thus affected the risk of some cancers[3-10].

Caspase-8(CASP8)isakey regulator of apoptosisof T lymphocytes and is encoded by the CASP8 gene[8].

The CASP8(MACH,FLICE,Mch5)gene contains at least 11 exonsspanning～30 kb on human chromosome band 2q33-34[11]and encodes 479 aMino acids,including prodomain and caspase domain.There are 353 single-nucleotide polymorphisms(SNP)of the CASP8 reported in the dbSNP database[12].It has been reported that polymorphic variation in CASP8 in fluences cancer risk,such as the variant D302H(rs1045485),the 652 6N insertion/deletion(ins/del)promoter variant(rs3834129),and the IVS12-19G/A(rs3769818).Previous meta-analysis suggested CASP8-652 6N promoter polymorphisMis associated with reduced renal cell carcinoma risk[5]and breast cancer risk[10].However,there was no evidence on the association between the-652N ins/del and digestive tract cancer due to limited publications.Digestive tract cancers represent a homogeneous group of malignancies.Given the amount of data now available on the association between CASP8-652 6N promoter polymorphisMand digestive tract cancer susceptibility,we performed the meta-analysis based on published casecontrol studies.

Materials and methods

Search strategy and inclusion criteria

We carried out a search in PubMed and Embase databases with the follow ing keywords:('CASP8'or'caspase-8',or'rs3834129','cancer'or'neoplasm'or'carcinoma'or'tumor'and'polymorphism'or'single nucleotide polymorphisms'or'SNPs'(last search was updated on Dec 31,2015).The titlesand abstracts of all eligible studies were exaMined carefully,and the bibliographies of the selected papers were also checked manually.If thesamepatientpopulation wasincluded in multiple publications,only the most recent or complete study was used in this meta-analysis.

Selected studies in our meta-analysis had to meet all the follow ing criteria:(a)only case-control or cohort studies;(b)evaluation of the CASP8-652 6N promoter polymorphisMand digestivesysteMcancer risk;(c)data availableon genotype frequency or the valueof the OR.

Data extraction

For each study,the follow ing information was extracted by two evaluators:fi rst author's name,year of publication,country,ethnicity,source of control,samplesizeand genotyping resultsof casesand controls(Table 1).Data were independently extracted by two investigators.If consensus was not reached by discussion,the third investigator was rereading.

Statistical analysis

Our meta-analysis assessed the overall associationbetween CASP8-652 6N promoter polymorphisMand digestive systeMcancer risk.The pooled ORs were performed for homozygote comparison(del/del vs.ins/ins),heterozygote comparison(del/ins vs.ins/ins),doMinant model(del/ins+del/del vs.ins/ins),and recessive model(del/del vs.del/ins+ins/ins),respectively.P＜0.05 was considered as statistically signi ficant.Subgroup analyses were carried out by the cancer types and ethnicity,respectively.We used either the fi xed(Mantel-Haenszel)or random(DerSimonian-Laird)effects models to get pooled ORs.If the P was more than 0.05 for the Q-test,the heterogeneity was considered signi fi cant.When there was no heterogeneity among studies,thepooled ORestimateof each study was calculated by the fi xed-effects model[13].Otherw ise,the random-effects model was used[14].The Begg's test and the Egger's test were used to test possible publication bias in this meta-analysis.Hardy-Weinberg equilibrium(HWE)in the controls was evaluated using the chi-square test.Meta-analyses were performed using Stata 11.0 software.

Table 1 Characteristics of studies included in the meta-analysis

Results

Study characteristics

As shown in Table 1,18 separate studies including a total of 15,086 patientsand 16,374 controlswere fi nally retrieved.Among them,there were 3 esophageal cancer studies[12,15-16],3 gastric cancers[12,15,17]and 12 colorectal cancers(including studies)[15,18-23](Fig.1).In addition,there were 9 studies in Asians populations,9 in Caucasians populations and 13 were populationbased studies,and 5 hospital-based studies.The distribution of genotypes in the controls was consistent with HWE,except for only one study[16].

Meta-analysis results

Overall,a signi fi cant reduced risk of digestive tract cancer was associated with the CASP8-652 6N del polymorphisMfor the co-doMinant model(del/del vs.ins/ins:OR=0.82,95%CI=0.72-0.95,Fig.2A;del/ins vs.ins/ins:OR=0.92,95%CI=0.87-0.97,Fig.2B);doMinant model(del/ins+del/del vs.ins/ins:OR=0.91,95%CI=0.87-0.96,Fig.2C),recessive model(del/del vs.del/ins+ins/ins:OR=0.85,95%CI=0.75-0.97,Fig.2D).

In the strati fi ed analysisby cancer types,we found all genetic models had protective effect on gastric cancer:del/del vs.ins/ins:OR=0.43,95%CI=0.28-0.67;del/ins vs.ins/ins:0.79(0.63-0.99);del/ins+del/del vs.ins/ins:0.73(0.59-0.95);del/del vs.del/ins+ins/ins:0.50(0.33-0.77).Similar results were observed for colorectal cancer under heterozygotecomparison(OR=0.94,95%CI=0.88-0.99,Supplementary Fig.1A,availableonline)and dominant model(OR=0.93,95%CI=0.88-0.99,Supplementary Fig.1B,available online),but not under homozygote comparison or recessive model.In addition,a signi fi cant decreased risk was found on esophageal cancer for the mostly genetic models:del/del vs.ins/ins:0.56(0.40-0.78);del/ins+del/del vs.ins/ins:0.83(0.71-0.97);del/del vs.del/ins+ins/ins:0.57(0.41-0.79),except the heterozygote comparison.

Fig.1 Flow diagraMof study identi fi cation.

When strati fi ed by ethnicity,two distinct results appeared in Asian populations and European populations.An evidently decreased risk for Asian populations in any genetic models(del/del vs.ins/ins:OR=0.62,95%CI=0.51-0.76,Supplementary Fig.2A,available online;del/ins vs.ins/ins:OR=0.87,95%CI=0.79-0.95,Supplementary Fig.2B,available online;del/ins+del/del vs.ins/ins:OR=0.83,95%CI=0.76-0.91,Supplementary Fig.2C,available online;del/del vs.del/ins+ins/ins:OR=0.61,95%CI=0.46-0,83,Supplementary Fig.2D,available online).In strati fi ed analysis by source of control,we found signi fi cant reduced CRC risk in population-based studies(del/del vs.ins/ins:OR=0.82,95%CI=0.70-0.96;del/ins vs.ins/ins:OR=0.92,95%CI=0.87-0.98;del/ins+del/del vs.ins/ins:OR=0.91,95%CI=0.86-0.96;del/del vs.del/ins+ins/ins:OR=0.83,95%CI=0.71-0.97),but not in hospital-based studies(fi gure not shown).

Fig.2 Odds ratios(ORs)for associations between CASP8 652 6N ins/del polymorphisMsand digestive systeMcancer risk based on all models.A:del/del vs.ins/ins,B:del/ins vs.ins/ins,C:del/del+del/ins vs.ins/ins;D:del/del vs.del/ins+ins/ins.

Publication bias

Begg's funnel plotand egger's testwereused to assess the publication bias.As shown in Fig.3,the shape of the funnel plot for theheterozygousmodel did not show any obvious asymmetry,which suggested no publication bias existing in all comparison models.And the P values were 0.914,95%CI were-1.461-1.317 in Egger's test,insinuating no publication bias(there were no publication bias in other genetic models,data not shown).

Sensitivity analysis

Sensitivity analysis was conducted repeatedly when the special studies had been left out at a time.For example,after excluding the study deviated froMHWE[16],the overall result did not been in fluenced signi fi cantly(heterozygousmodel:OR=0.84,95%CI:0.73-0.96).Thesensitivity analysis results revealed that no individual studies signi fi cantly affected the pooled ORs under heterozygous model of CASP8-652 6N polymorphism(Fig.4),suggesting that our results are stable.

Fig.3 Funnel plot analysis to detect publication bias for the CASP8-652 6N del polymorphisMs(del/ins vs.ins/ins).Each pointrepresents an individual study for the indicated association

Heterogeneity analysis

There was potential heterogeneity among studies in overall analysis and some subgroup analysis under homozygousand recessive model(Table 2).To explore the source of heterogeneity,we conducted a subgroup analysis by cancer types,ethnicity and the source of control.As a result,there presented little or no heterogeneity across studies.Furthermore,we found that colorectal cancer,Caucasian and population-based studies contributed to substantial heterogeneity.

Discussion

Fig.4 In fluenceanalysisof thesummary odds ratio coef fi cientson theassociation between CASP8-652 6N del polymorphisms(del/ins vs.ins/ins)with digestive tract cancer risk.Resultswere computed by omitting each study(leftcolumn)in turn.Bars,95%con fi dence interval.

Apoptosis plays critical roles in a wide variety of physiologic processes during fetal development and in adult tissues[24].Defects in apoptotic cell death regulation contribute to many diseases,such as some cancers[24].Caspase8 isa central regulator of apoptosis or programmed cell death.Caspase-8 gene mutation can in fluence the rate of apoptosis,and thus affect cancer risk.The most widely reported mutation in CASP8 gene is the six-nucleotide deletion polymorphism(-652 6N del).The-652 6N del variant involves a six-nucleotide deletion in the promoter region of the CASP8 gene[15].The deletion abolishes an Sp1 binding siteand isassociated with decreased RNA expression in lymphocytes and lower CASP8 activity and activationinduced cell death of T lymphocytes[25].

Table 2 Associations between CASP8-652 6N ins/del genotype and digestive tract cancer risk

It is known that apoptosis plays an important role in cancer grow th,progression,and drug resistance[26].Previous studies found that genetic mutations of caspases 8 in tumor cells reduce the cell sensitivity to apoptosis,which may be related to the occurrence and development of tumors.Our meta-analysis based on 18 case-control studies suggested that the CASP8-652 6N del polymorphisMis associated with decreased risk of digestive tractcancer,which wassiMilar with the results of Yin etal.[27]and Zhang et al.[28].However,therewas potential heterogeneity under the four genetic models in digestive systeMcancer.Therefore,we also carried out the strati fi cation analysis by ethnicity,cancer type and source of control.In strati fi cation by cancer type,we also found the reduced cancer risk remained for subgroups of colorectal cancer,gastric cancer and esophageal cancer.What is more,siMilar results were found in Asians and population-based studies.Peng et al.[29]reached asimilar conclusion that CASP8-652 6N ins/del polymorphisMmay play a protective role in colorectal cancer development.But their sample size was relatively small,not having enough statistical power to explore the real association.Our study with a large sample size provides additional evidence of the association between CASP8-652 6N ins/del polymorphisMand colorectal cancer risk.

There was a signi fi cant reduced association between the-652 6N del polymorphisMand digestive tractcancer risk among Asian populations,but not in Caucasians.Oneexplanation may begenetic differencesand diverse living environments between Asian and Caucasian populations.On the other hand,the Asian population are relatively homogeneous,who mainly come froMHan Chinese.However,Caucasian population consisted of people froMvarious countries with diverse living environments.Difference between Asians and Caucasians may explain the distinct fi ndings.In addition,gene-gene and gene-environmental interactions may lead to the result bias.Among the included studies,13 were population-based samples and 5 were hospitalbased samples,respectively.Borderline signi fi cantly decreased risks were found for population-based studies.One explanation may be the hospital-based studies have some biases because such controls may be asampleofill-de fi ned referencepopulation,particularly when thegenotypesunder investigation wereassociated with the disease conditions[30].On the other hand,the insuf fi cient hospital-based samples tended to have underestimated cancer risks.

Some liMitations of this meta-analysis should be addressed.First,in the subgroup analyses of cancer type,the number of esophageal cancer and gastric cancer were relatively small,not having suf fi cient statistical power to explore the real association.Second,our resultswerebased on unadjusted estimates,because not all published studies presented adjusted ORs.More precise analysis should be carried out,which would allow for the adjustment by other variance,such assex,age,smoking status,drinking status,obesity and other lifestyles.Thirdly,the experiment method for each study was without restrictions,which affected the accuracy of the results to a certain extent.

In conclusion,our meta-analysis supports the association between the CASP8-652 6N del polymorphisms and reduced digestive tract cancer risks,especially in Asians.Future studies with large sample sizes are required to con fi rMcurrent fi ndings.

Acknow ledgments

This study is partially supported by the National Natural Science Foundation of China(NSFC:81472634),Health Department guidance project of Jiangsu Province(Z201201),the PrograMfor Developmentofinnovative Research TeaMin the First Af fi liated Hospital of NJMU and the Project Funded by the Priority Academic PrograMDevelopment of Jiangsu Higher Education Institutions(JX10231801),and the Summit of the Six Top Talents PrograMof Jiangsu Province(2013-WSN-034).