Systematic Review And Diagnosis Research Of Multiple Genes Methylation In Hepatocellular Carcinoma

Ojective:1. To evaluate the relationship of DAPK, RASSF1A andp16methylation with risk of hepatocellular carcinoma（HCC）by meta analysis.2. To investigate the detection values of promoter methylation status of cancersuppressor genes such as beclin1, RASSF1A, p16and DAPK in the screeningand early diagnosis of hepatocellular carcinoma（HCC）. Methods:1. Metaanalysis:⑴Literature related to the DAPK, RASSF1A and p16methylation inHCC retrieved from Pubmed, EMBASE, Ovid, Web of science, CBM, VIP,CNKI and Wangfang database.⑵Data was collected according to the inclusionand exclusion criteria, and analyzed by RevMan5.2software.2. Clinicalresearch case:⑴The tissues of37patients with HCC in the Affiliated Hospitalof Luzhou Medical College from April2012to June2013were colleted bysurgical resection, including the noncancerous tissues and5normal liver tissuesfrom surgical treatment of liver hemangioma adjacent tissues.⑵Themethylation specific PCR (MSP) method was used to detect the promotermethylation status of37HCC samples, corresponding tumor adjacentspecimens and5normal cases.⑶The area under the ROC curve (AUC) ofbeclin1, RASSF1A, p16, DAPK and stepwise multiple logistic regressionresults were compared and further analysed with the index of sensitivity,specificity, Youden’s index and positive likelihood ratio/negative likelihoodratio. Results:1. Meta analysis:⑴A total of31references were included in thestudy, involving DAPK gene3references and cumulative314cases,134HCC cases and180controls about DAPK gene methylation were brought into the study; a total of15references were included in the RASSF1A gene,involving cumulative1417cases,710HCC cases and707controls aboutRASSF1A gene methylation were brought into the study; a total of23referenceswere included in the p16gene, involving cumulative2245cases,1106HCCcases and1139controls about p16gene methylation were brought into the study.⑵The methylation rate of DAPK gene in HCC tissues were significantlyhigher than that of normal liver tissues[OR=6.69，95%CI（2.43~18.41），P<0.05], there was no significant difference between NCT[OR=0.87，95%CI（0.47~1.59），P>0.05] and cirrhosis tissues[OR=0.21，95%C（I0.01~8.56），P>0.05]; the methylation rate of RASSF1A gene in HCC tissues were allsignificantly higher than that of normal liver tissues[OR=50.23，95%CI（22.21~113.61），P<0.05], NCT[OR=7.63，95%CI（3.38~17.22）, P<0.05]and cirrhosis tissues[OR=17.18,95%CI （2.57~114.91）， P<0.05]; themethylation rate of p16gene in HCC tissues were all significantly higher thanthat of normal liver tissues[OR=21.18，95%CI（9.75~46.00），P<0.05],NCT[OR=7.97，95%CI （4.64~13.68）， P<0.05] and cirrhosistissues[OR=6.18，95%CI（4.16~9.18），P<0.05].2. Clinical research case:⑴The positive rates of promoter methylation of beclin1，RASSF1A，p16andDAPK genes in HCC were5.4%,94.6%,73.0%and35.1%and correspondingmethylation rate of NCT were0%,35.1％,24.3％�'�10.8％,5normal livertissues had no methylation detected. The methylation of RASSF1A, p16andDAPK genes was more frequent in HCC than that in NCT(P<0.05)．⑵Therewas no significant difference between the methylation rate of beclin1,RASSF1A, p16and DAPK genes and the gender, age, staging and presense ofHBV infection of HCC patients, but the methylation rate of RASSF1A was thehighest in the tissues with larger size（χ2=6.578，P＜0.05）, and DAPK gene inAFP＜400μg/L of positive rate HCC tissues have the highest methylation rate(χ2=5.106，P＜0.05).⑶Combined detection of promoter methylation ofRASSF1A and tumor marker AFP could get the higher sensitivity(81.8%) andspecificity (93.0%) in the diagnosis of HCC, the AUC is0.913, which washigher than that of single gene with specificity, Youden index, positivelikelihood/negative likelihood ration. Conclusion:1. The methylation ofRASSF1A, p16and DAPK genes are closely associated with HCC development.2. RASSF1A, p16gene have higher methylation rate in HCC tissues, moreovercombing detection of promoter methylation of RASSF1A and AFP may becomean effective index for early diagnosis of HCC.3. The methylation rate ofRASSF1A was associated with the tumor size, and the relationship with HCCclinical pathological features need further testing and research.4. Beclin1gene,with the lowest methylation rate (5.4%) may be not useful as a marker gene forthe diagnosis of HCC marker.