review article new progress of epigenetic biomarkers in...

Review ArticleNew Progress of Epigenetic Biomarkers in Urological Cancer

Peng Wu1234 Ziyi Cao1245 and Song Wu1246

1Department of Urological Surgery The Affiliated Luohu Hospital of Shenzhen University Shenzhen UniversityShenzhen 518000 China2Shenzhen Following Precision Medical Institute Shenzhen Luohu Hospital Group Shenzhen 518000 China3First Clinical Medical College Anhui Medical University Hefei 230032 China4Shenzhen Second Peoplersquos Hospital 1st Affiliated Hospital of Shenzhen University Shenzhen 518037 China5Medical College Shenzhen University Shenzhen 518060 China6Shenzhen Gene Detection Public Service Platform of Clinical Application Shenzhen Luohu Hospital Group Shenzhen 518000 China

Correspondence should be addressed to Song Wu doctor wusong126com

Received 19 May 2016 Accepted 30 June 2016

Academic Editor Maria L Poeta

Copyright copy 2016 Peng Wu et alThis is an open access article distributed under the Creative Commons Attribution License whichpermits unrestricted use distribution and reproduction in any medium provided the original work is properly cited

Urological cancers consist of bladder kidney prostate and testis cancers and they are generally silenced at their early stage whichleads to the loss of the best opportunity for early diagnosis and treatment Desired biomarkers are scarce for urological cancersand current biomarkers are lack of specificity and sensitivity Epigenetic alterations are characteristic of nearly all kinds of humanmalignances including DNA methylation histone modification and miRNA regulation Besides the detection of these epigeneticconditions is easily accessible especially for urine best target for monitoring the diseases of urinary system Here we summarizesome new progress about epigenetic biomarkers in urological cancers hoping to provide new thoughts for the diagnosis treatmentand prognosis of urological cancers

1 Introduction

Urological cancers are comprised of bladder prostate renaland testis cancers which are among the 10 most frequentcancers in man except testis cancer So far the gold standarddiagnosis of urological cancer is pathological diagnosis andthe early screeningmethods are rare Some existing biomark-ers such as prostate-specific antigenmay be useful in prostatecancer screening but it is invasive and short of specificity andis causing overdiagnosis and overtreatment which limits itsapplication [1] Bladder cancer and renal cell carcinoma lackspecific predictive biomarkers and only some symptoms forinstance hematuria might have some effects in finding theexistence of cancer In the ear of personalized and precisemedicine searching noninvasive biomarkers to detect thepresence of urological cancers early has become an urgentneed In this review we focus on epigenetic-based biomarkersin urology system and summarize current state of researchon epigenetic alterations [2] holding the promise to providenew ideas for the diagnosis treatment and prediction ofurological cancer

2 Epigenetic Mechanisms and Cancer

DNA methylation mediated by DNA methyltransferases(DNMTs) using S-adenosylmethionine (SAM) as themethylsupplier and adding methyl group to the 5-carbon of thecytosine is the most well delved epigenetic modification inhuman diseases [3 4] This phenomenon mostly occurs atthe cytosine and guanine (CpG) dinucleotides [5] whichdispersed in the genome or in DNA repetitive region Butwithin the gene promoter regions there are clusters ofCpGs called CpG islands usually remaining unmethylated toenable gene expressionDNAmethylation and demethylationare known to be associated with tumorigenesis and result insilencing tumor suppressor genes and activating oncogenesrespectively [6] Chemically speaking DNA methylation isstable and can be accurately measured in almost all typesof specimens for instance plasma serum and specimens[7] Although DNA methylation is an ideal biomarker forclinical examination there are still some obstacles to beovercome includingDNA isolation efficiency leukocyteDNA

Hindawi Publishing CorporationDisease MarkersVolume 2016 Article ID 9864047 8 pageshttpdxdoiorg10115520169864047

2 Disease Markers

contamination and loss of DNA templates arising from theemployment of bisulfite treatment

Histones modification includes methylation carbonyla-tion phosphorylation ubiquitinylation sumoylation glyco-sylation and acetylation which are known as ldquothe histonecoderdquo together with DNAmethylation regulating the expres-sion of specific genes [8] The mechanism of regulation isthrough the addition of specific chemical groups to the N-terminal of basic amino acid residues at the tails of histonesaltering the affinity of the histone tails to the DNA andchanging the conformation of chromatin structure to regulatethe transcription of genes For instance histone acetylationenhances the transcriptional activity of gene for lowering theaffinity of histone tails to theDNA while histone deacetylasesresult in gene repression [9 10] Contrary to the DNAmethy-lation the histone methylation (addition of methyl groupsto H3 H4 lysine and arginine) can generate two effectseither activation or repression transcription depending onthe specific amino acid residues modified [11] In cancercell genome-wide histone modification and expression ofkey histone modulating enzymes have been reported but thedetailed mechanisms of their involvement in tumorigenesisare poorly understood [12 13]

Another epigenetic modification has been studied deeplyknown as microRNAs (miRNAs) miRNAs a class of non-coding RNAs which are about 18ndash25 nucleotides at lengthcould be synthesized and processed in the nucleus miRNAsalter the expression through binding to mRNAs then RNA-induced silencing complex takes part and regulates proteintranslation [14 15] It was estimated that at least 30 ofhuman genes are regulated by miRNA It is noteworthy thateach mRNA can be combined with multiple miRNAs andeach miRNA may regulate multiple mRNAs miRNAs playan important role in tumorigenesis activating oncogenes orrestraining tumor suppressor genes [14 15] The ways thatmiRNAs alter gene expression include deletion amplifica-tion mutation and chromosomal abnormalities [16] Somestudies found that miRNA can influence gene expression viatargeting a specific gene region for DNA methylation andhistone modifications [16 17] and the expression of miRNAcan be regulated by other epigenetic modifications such asDNA methylation [18 19] which indicated the interactionsbetween miRNAs and other epigenetic mechanisms

3 Epigenetic Biomarkers in Bladder Cancer

Bladder cancer is the most common male urology malig-nancy in China and the total number of cases in malesfor 2015 is approximately 621 thousand [20] Currently thediagnosis of bladder is mainly invasive causing discomfortto patients and can only provide a generalized outcomefor patients so the demand for noninvasive screening anddiagnosis method is urgent Epigenetic biomarkers candecrease the use of invasive methods and provide the earlydiagnosis of bladder cancer allowing for more effectivetreatment For example RUNX3 gene that has been thoughtas tumor suppressor gene exhibited significant increase atmethylation levels in bladder cancer by analyzing 124 tumortissue samples This study shows that RUNX3 gene may

be a potential marker for detecting bladder cancer [21]Apart from diagnostic markers a prognostic indicator inpatients with non-muscle-invasive bladder cancer (NMIBC)was found by Yoon et al The clinical relevance of RSPH9was determined by quantitative pyrosequencing analysis of136 human bladder specimens (8 normal controls and 128NMIBCs) They concluded that RSPH9 methylation can beof value for the assessment of disease recurrence and anindependent prognostic indicator in NMIBC patients [22]Similarly Lin and Luo et al reported that the hypermethy-lation of PCDH10 (50 119899 = 117) and PCDH17 (52 119899 =151) was related to the development of bladder cancer and itwas an independent predictor of cancer-specific survival time[23 24]

With the deepening of research it was thought that therewas no single gene found to be methylated in vast majority ofbladder tumors More and more researches have focused onusing gene panels to early detect bladder cancer and predicttumor recurrence progression and metastasis A methyla-tion analysis of PCDH17 andPOU4F2was developed recentlyin 148 individuals by qMSP using urine sediment Thecombination of POU4F2PCDH17 detection had sensitivityand specificity of 9000 and 9396 in the validation of 312individuals [25] Renard et al found that 2 genes (TWIST1andNID2) were frequentlymethylated in urine samples frombladder cancer patients (93 specificity and 90 sensitivity)[26] Another study discovered that a methylation genepanel which consisted of 4 genes (CDH1 CDH13 RASSF1Aand APC) had significant correlations with poor prognosis(cancer with high grade advanced stages and aneuploidies)Likewise a study conducted by Yates et al demonstratedthat promoter methylation of RASSF1A CDH1 TNFSR25EDNRB and APC was associated with tumor progression[27]

Histone modifications were also found to relate to thepathogenesis of bladder cancer and regulation of cancer cellproliferation A study in 2011 identified global histoneH4K20trimethylation that could predict cancer-specific survival inpatients with muscle-invasive bladder cancer [28] A meta-analysis published by An et al showed that merged oddsratio of the effect between slow acetylation and bladdercancer was 131 (95 confidence odds ratio interval = 111ndash155) illustrating that slow acetylation modestly increasesthe risk of bladder cancer [29] Recently Jia et al foundthat the acetylation status of KLF4 can decide the genefunction as a tumor suppressor or oncogene in bladdercancer Their results indicated that deacetylated KLF4 canact as an oncogene accelerating bladder cancer proliferationon the contrary acetylation of KLF4 performs as a tumorsuppressor which may be a good target for bladder cancertherapy [30] In addition another study found that expressionlevels of H3K4me1 H4K20me1 H4K20me2 and H4K20me3were correlated with advanced pathological stage and thatH4K20me3 was an independent prognosis factor for thesurvival of patients with muscle-invasive bladder cancer [31]

miRNAs are synthesized and processed in the nucleusand released into cytoplasm binding to specific mRNA andinducing gene silence [32] Recently Jiang et al demonstrated

Disease Markers 3

Table 1 Overview of bladder cancer biomarkers

Biomarker Sample Type Diagnosis treatment or prognosisBladder cancer

RUNX3 Tissue DNA methylation DiagnosisRSPH9 Urine DNA methylation Diagnosis prognosisPCDH10 PCDH17 Urine DNA hypermethylation Treatment prognosisPCDH17 POU4F2 Urine DNA methylation AllTWIST1 NID2 Urine DNA methylation DiagnosisCDH1 CDH13 RASSF1A APC Urine DNA methylation PrognosisRASSF1A CDH1 TNFSR25 EDNRB APC Urine DNA methylation PrognosisH4K20 Tissue Histone modification PrognosisKLF4 Urine Histone modification TreatmentH4K20me3 Tissue Expression level PrognosismiR-422a-3pmiR-486-3pmiR-103a-3pmiR-27a-3p

Tissue (serum) Overexpression Prognosis

miRNA-146a-5p Urine Overexpression PrognosismiRNA-145 Urine Overexpression Prognosis

a four-miRNA panel (miR-422a-3p miR-486-3p miR-103a-3p and miR-27a-3p) that can be used for muscle-invasivebladder cancer (MIBC) prediction with an area under thereceiver operating characteristic curve (AUC) of 0894 (95CI 0864ndash0931) by MiSeq sequencing on serum from 207MIBC patients 285 non-muscle-invasive bladder cancer(NMIBC) patients and 193 controls [33] Sasaki et al foundthat miRNA-146a-5p is increased in bladder patients anddecreased in patients after transurethral resection What ismore levels of miRNA-146a-5p in urine are significantlyhigher in patients with high-grade tumors compared withthose with low-grade tumors Their results suggested thaturinary miRNA-146a-5p might be useful as a new noninva-sive diagnosticmarker therapeutic target or anticancer agent[34] A similar study conducted by Matsushita et al showsthat miRNA-145 (miR-145-5p guide-strand and miR-145-3p passenger-strand) plays pivotal roles in bladder cancer(BC) cells by regulating UHRF1 which is overexpressed inBC clinical specimens Further assay confirms that ectopicexpression of either miR-145-5p or miR-145-3p in BC cellsobviously suppressed cancer cell growth [35] Findings onepigenetic biomarkers in bladder cancer are summarized inTable 1

4 Epigenetic Biomarkers in Kidney Cancer

Kidney cancer is the thirdmost commonurologymalignancyin China and there were about 668 thousand people diag-nosed with renal carcinoma in 2015 Among these patients432 thousand are males and females only account for 35[20] Currently there are no widely accepted tumor markersfor clinical diagnosis of renal cell carcinoma The clinicaldiagnosis of renal cell carcinomamainly depends on imagingexamination and the definite diagnosis is confirmed bypathological examination Hauser et al demonstrated that

Wnt antagonist family genes can serve as biomarkers fordiagnosis staging and prognosis in kidney cancer usingtumor and serum DNA They utilized methylation-specificPCR detecting the level of genes panels comprised of sFRP-1 sFRP-2 sFRP-4 sFRP-5 Wif-1 and Dkk-3 in 62 RCCsamples and corresponding normal renal tissue The resultsindicated that Wnt antagonist family genes detection hadsensitivity of 790 and specificity of 758 and the serumDNAwas significantly correlated with tumor grade and stage[36] Besides other studies have reported that some genes arehighly specific for RCC patients in the level of DNA hyper-methylation including VHL (91) and RASSF1A (93) [37]Recently another similar outcome showed that two genesSMPD3 and FBXW10 are hypermethylated in ccRCC tissuesamples compared with paired normal tissues Interestinglyafter 5-aza-21015840-deoxycytidine treatment mRNA expression ofSMPD3 and FBXW10 was significantly upregulated indicat-ing that these two genes can be a target for treatment andprovide predictive information for clinical decisions [38]Furthermore a tumor suppressive gene DAB2IP reportedby Wang et al suggests that DAB2IP CpG1 methylationis a practical and repeatable biomarker for ccRCC whichcan provide prognostic value that complements the currentstaging system This research team validated the relationbetween CpG methylation biomarker (DAB2IP CpG1) andpoor overall survival in TCGA (a cohort of 318 ccRCCpatients) by pyrosequencing quantitative methylation assayin 224 ccRCCpatients frommultipleChinese centers (MCHCset) and 239 patients from University of Texas SouthwesternMedical Center [39]

Histone modifications in kidney cancer were found tohave significant relevance with the prognosis of renal cellcarcinoma (RCC) Patients with positive immunostainingof H3K4me2 and H3K18Ac but accounting for a lowerproportion generally had a shorter 1-year survival probability

4 Disease Markers

Table 2 Overview of kidney cancer biomarkers

Biomarker Sample Type Diagnosis treatment or prognosisKidney cancer

Wnt family genes Tissue (serum) DNA methylation Diagnosis prognosisVHL RASSF1A Tissue DNA methylation DiagnosisSMPD3 FBXW10 Tissue Hypermethylation DiagnosisDAB2IP Tissue Methylation PrognosisH3K4me2 H3K18Ac Tissue Histone modification PrognosishMOF Tissue Histone modification DiagnosisHDAC Tissue Histone modification TreatmentmiRNA-126 Tissue Downregulated TreatmentmiR-146a-5pmiR-128a-3pmiR-17-5p

Tissue Downregulated Prognosis

than those with more histone modifications [40] This studyreveals that histone modification is changed in the progressof kidney cancer Currently there is no curative treatmentfor advanced renal cancer Enhancing histone acetylation isa promising epigenetic-based therapy for cancer Seligson etal found that the combination treatment of ritonavir andpanobinostat can enhance histone acetylation and inhibitrenal cancer growth by suppressing the expression of histonedeacetylase (HDAC)They tested this combination inmurinesubcutaneous xenograft model using Caki-1 cells and foundthat after a 10-day treatment tumor growth was inhibitedsignificantly [41] Furthermore in a study with 21 clear cellrenal cell carcinoma (ccRCC) patients overexpression ofCA9 was detected in all patients Meanwhile the expressionlevels of hMOF gene (an acetyltransferase) frequently weredownregulated in 19 patients accompanied by the acetylationof histone H4K16 The study concluded that hMOF may beinvolved in the pathogenesis of kidney cancer and can be anew CA9-independent RCC diagnostic marker [42]

miRNAs are also involved in the development and pro-gression of renal cell carcinoma Zhang et al reported thatmiRNA-126 can inhibit tumor cell invasion and metastasisby downregulating ROCK1 in renal cell carcinoma Theyanalyzed 128 pairs of ccRCC and adjacent normal tissuesamples measured miRNA-126 expression levels and foundthe association between miRNA-126 and various clinico-pathological parameters [43] Using only one kind of miRNAmarker might be inaccurate for prognosis an analogousstudy concerning ccRCC presented that threemiRNAs (miR-146a-5p miR-128a-3p and miR-17-5p) were correlated withmetastasis of ccRCC patients Specifically they showed thatthe targeted genes were downregulated by miR-146a-5p andvalidated the interaction in cell culture experiments [44]

Findings on epigenetic biomarkers in kidney cancer aresummarized in Table 2

5 Epigenetic Biomarkers in Prostate Cancer

Screening of individuals without any cardinal symptom bythe PSA test has been the focus of increasing criticism

primarily due to potential overtreatment and less com-prehensive evaluation [45] Candidate biomarkers will beseparated with groups as molecular class soluble proteinsDNAmethylation mRNA microRNA and so forth [46ndash48]

Methylation levels of two genes PCDH17 and TCF21were quantified in a total of 12 cancer cell lines and 318clinical samples These two gene methylation levels provideda sensitivity rate of 96 for prostate cancerThehigh exposingof PCDH17 and TCF21 methylation in prostate cancer celllines was significantly different from primary tumor tissuesFurthermore methylation levels were meaningfully lower inbladder and prostate nontumorous tissues providing a bio-logical influence as cancer biomarkers [49 50] In additiondiagnostic coverage might be improved by using gene panelsincluding GSTP1ARFCDNK2AMGMT and GSTP1APCRARB2RASSF1A for urine and GSTP1PTGS2RPRMTIG1for serum samples [46]

It has been noticed that HOXB13 became overexpressedduring malignant progression of the prostatic tissue andplayed an important role in the pathogenesis of the prostategland as a novel biomarker for the prognosis of prostatecancer [51] ADAM19 (a disintegrin and metalloproteinase19) is a transmembrane and soluble protein concerned incell phenotype through cell adhesion and proteolysis Ithas been shown in special immunohistochemical studiesthat ADAM19 protein levels were more expressed comparedto normal prostate tissue during prostate cancer biopsies[52] As some reports said that the expression of SFRP1inversely correlates with the Gleason score survival rate andresponse for endocrine therapy expression are a favorablepredictive and prognostic biomarker [53] In other methodsPSF1 is expressed in high-grade prostate cancer and mayalso be a useful biomarker to identify patients for diagnosis[54] Engrailed-2 (EN2) protein a homeodomain-containingtranscription factor expressed in prostate cancer and secretedinto the urine showed a highly specific and sensitive effect asa kind of biomarker for prostate cancer [55] There are manyproteins which obtain the same function like downregulatedprotein SLC18A2 [56] and unregulated protein TRPM4 inprostate cancer [57] SOX2 was consistently downregulated

Disease Markers 5

Table 3 Overview of prostate cancer biomarkers

Biomarker Sample Type Diagnosis treatment or prognosisProstate cancer

PCDH17 TCF21 Tissue DNA methylation DiagnosisGSTP1 ARF CDNK2A MGMT Urine DNA methylation DiagnosisGSTP1 APC RARB2 RASSF1A Urine DNA methylation DiagnosisGSTP1 PTGS2 RPRM TIG1 Tissue (serum) DNA methylation DiagnosisHOXB13 Tissue Overexpression PrognosisADAM19 Tissue Overexpression TreatmentSFRP1 Tissue Decreased expression Diagnosis prognosisPSF1 Tissue Overexpression Diagnosis prognosisEN2 Tissue urine Overexpression DiagnosisSLC18A2 Tissue Downregulated DiagnosisTRPM4 Tissue Overexpression PrognosisSUX2 Tissue Downregulated PrognosisXPO6 Tissue Overexpression Prognosis

except in cell clusters lyingwithin lymphnode- (LN-) positiveprostate cancer [58] Some other special genes could be foundto predict recurrence as XPO6 [59] and FMOD as biomarkerfor prostate cancer [45]

Other promising RNA markers are the transcripts offusion genes which between the androgen-regulated trans-membrane protease serine 2 gene (TMPSS2) and ERGtranscription factors through chromosomal rearrangementsbecome stable and produce a viable mRNA during transcrip-tion TMPSS2-ERG transcription factors have been identifiedas promising urinary novel biomarkers [60]

Several potential microRNAs for prostate cancer havebeen identified as important biomarkers CirculatingmiRNA-410-5p level was significantly higher in the prostatecancer patients than in normal patients [61] Recentlya present study compared miR-18a expression with theperipheral blood of patients benign prostatic hyperplasia(BPH) of patients and normal individuals to evaluate thepossibility of achieving noninvasive diagnosis for prostatecancer In summary the present results indicate that miR-18aexpression is significantly higher in peripheral blood ofpatients with prostate cancer compared with two othersPeripheral blood oncogenic miR-18a may serve as a potentialnoninvasive biomarker for prostate cancer tissues acting asan oncogenic miRNA [62] On the other hand miRNA-129 isa novel independent prognosic factor because of being down-regulated significantly in prostate cancer On the contraryoverexpression of miR-129 could develop tumor suppressivefunctions and prevent prostate cancer growth [63]

The curiousness of CTCs for specific tumor character-istics has drawn major attention over the past few yearsThe genomic profiles of CTCs have been found to be largelycomparable to primary tumors andor metastatic tissueindicating that CTCs are able to reflect tumor characteristicsincluding the extent of intratumoral and biological hetero-geneity CTCs have been shown to be tumorigenic and capa-ble of forming new metastases [64ndash67] Androgen receptorsplice variant-7 (AR-V7) in circulating tumor cells (CTCs)from metastatic castrate-resistant prostate cancer (mCRPC)

patients received enzalutamide or abiraterone The resultshocked us as none of the 18 patients with detectableAR-V7 inCTCs had prostate-specific antigen (PSA) responses Furtherthe median time to PSA progression after enzalutamide orabiraterone treatment was only 13-14 months in AR-V7-positive patients as compared to 53ndash61 months in AR-V7-negative patients AR-V7 in CTCs was also associated withshorter survival [68]

In 1960 prostate-specific antigen (PSA) was first discov-ered by Rubin Flocks Since the mid-80s PSA has been themost commonly used biomarker for prostate cancer to judgecurrent and future risk detect response to treatments anddetect recurrence in all stages of the disease Due to thiswork the Food and Drug Administration (FDA) in USAapproved the use of PSA for monitoring recurrence aftertreatment It was later known that it was human species-specific [69] Prostate-specific antigen (PSA) testing maybe used for PCA screening however significant problemsregarding specificity overdiagnosis and overtreatment limitthe acceptance of this marker [70] Recently macrophageinhibitory cytokine-1 (MIC-1) concentration along with thePSA assay could provide much improved specificity to theassay by a retrospective study It is not difficult to seethat MIC-1 concentration in serum was elevated in prostatecancer patients compared to normal and biopsy-negativeindividuals What is more the MIC-1 level was correlatedwith the progression of prostate cancer The analysis basedon MIC-1 and PSA concentrations in serum with the patientwith prostate cancer status improved the specificity of thediagnosis without compromising the high sensitivity of thePSA test alone and has potential for the prognosis for patienttherapy strategies [71] Findings on epigenetic biomarkers inprostate cancer are summarized in Table 3

6 Epigenetic Biomarkers in Testicular Cancer

Recent genomic studies have identified risk SNPs in testiculargerm cell tumors (TGCT) Increased PDE11A SPRY4 andBAK1 promoter methylation and decreased KITLG promoter

6 Disease Markers

methylation in familial TGCT cases versus healthy malefamily controls can be used to diagnose TGCT in the earlytime [72] It is reported that LINE-1 methylation may begender-specific with a strong correlation between LINE-1methylation levels associated with disease risk [73] Besidesthat if we knock down miR-199a-3p in a normal humantesticular cell line (HT) this leads to elevation of DNMT3A2(DNMT3A gene isoform 2) mRNA and protein levels Inclinical samples DNMT3A2 was significantly overexpressedin malignant testicular tumor which was inversely correlatedwith the expression of miR-199a-3p [74] The methylationprofile of cancer-associated genes in testicular cancer cor-relates with histological types and cancer-specific genesFurther methylation analysis in a larger cohort is needed toelucidate the role of genes role in testicular cancer develop-ment and potential for therapy early detection and diseasemonitoring [75]

7 Conclusions

In general these findings provide new directions for detec-tion diagnosis and prognosis of urological cancerwhichmayrevolutionize the clinical management of cancer patientsDNA methylation is the most explored modification inurological cancer and modifications in histone and miRNAare becoming a hot research topic Nevertheless the majorityof these markers are single target discovered in a smallamount of clinical cases and lack specificity therefore apanel consisting of multiple targets is needed and this kindof research will become a trend Besides there is plethoraof epigenetic biomarkers which has been identified butonly extremely rare biomarkers can be applied to clinicaldiagnosis and therapy targets The selected biomarker has tobe evaluated through rigid clinical trials in appropriate scaleand determine the relevance between the biomarkers andclinical practice We expect more novel publications aboutepigenetic biomarkers in urological malignances especiallyfrom large clinical cases

Competing Interests

The authors declare that there are no competing financialinterests

Authorsrsquo Contributions

Peng Wu and Ziyi Cao contributed equally to this work

References

[1] W J Catalona P C Southwick KM Slawin et al ldquoComparisonof percent free PSA PSA density and age-specific PSA cutoffsfor prostate cancer detection and stagingrdquo Urology vol 56 no2 pp 255ndash260 2000

[2] S Sharma T K Kelly and P A Jones ldquoEpigenetics in cancerrdquoCarcinogenesis vol 31 no 1 pp 27ndash36 2009

[3] T Vaissiere C Sawan and Z Herceg ldquoEpigenetic interplaybetween histone modifications and DNA methylation in gene

silencingrdquo Mutation ResearchReviews in Mutation Researchvol 659 no 1-2 pp 40ndash48 2008

[4] L Lopez-Serra and M Esteller ldquoProteins that bind methylatedDNA and human cancer reading the wrong wordsrdquo BritishJournal of Cancer vol 98 no 12 pp 1881ndash1885 2008

[5] A S Perry R Foley K Woodson and M Lawler ldquoTheemerging roles of DNAmethylation in the clinical managementof prostate cancerrdquo Endocrine-Related Cancer vol 13 no 2 pp357ndash377 2006

[6] C H Waddington ldquoThe epigenotype 1942rdquo International Jour-nal of Epidemiology vol 41 no 1 pp 10ndash13 2012

[7] N B Y Tsui E K O Ng and Y M D Lo ldquoStability ofendogenous and added RNA in blood specimens serum andplasmardquo Clinical Chemistry vol 48 no 10 pp 1647ndash1653 2002

[8] H Easwaran H-C Tsai and S B Baylin ldquoCancer epigeneticstumor heterogeneity plasticity of stem-like states and drugresistancerdquoMolecular Cell vol 54 no 5 pp 716ndash727 2014

[9] C Sawan T Vaissiere R Murr and Z Herceg ldquoEpigeneticdrivers and genetic passengers on the road to cancerrdquoMutationResearchFundamental and Molecular Mechanisms of Mutagen-esis vol 642 no 1-2 pp 1ndash13 2008

[10] D J Weisenberger ldquoCharacterizing DNA methylation alter-ations from the cancer genome atlasrdquo Journal of ClinicalInvestigation vol 124 no 1 pp 17ndash23 2014

[11] E Dudziec S Miah H M Z Choudhry et al ldquoHypermethy-lation of CpG islands and shores around specific MicroRNAsand mirtrons is associated with the phenotype and presence ofbladder cancerrdquoClinical Cancer Research vol 17 no 6 pp 1287ndash1296 2011

[12] Z-G Luo Z-G Li S-L Gui B-J Chi and J-G MaldquoProtocadherin-17 promoter methylation in serum-derivedDNA is associated with poor prognosis of bladder cancerrdquoJournal of International Medical Research vol 42 no 1 pp 35ndash41 2014

[13] A Lennartsson and K Ekwall ldquoHistone modification patternsand epigenetic codesrdquo Biochimica et Biophysica Acta (BBA)mdashGeneral Subjects vol 1790 no 9 pp 863ndash868 2009

[14] T A Farazi J I Hoell P Morozov and T Tuschl ldquoMicroRNAsin human cancerrdquo Advances in Experimental Medicine andBiology vol 774 pp 1ndash20 2013

[15] C B Yoo and P A Jones ldquoEpigenetic therapy of cancer pastpresent and futurerdquo Nature Reviews Drug Discovery vol 5 no1 pp 37ndash50 2006

[16] M A Matzke and J A Birchler ldquoRNAi-mediated pathways inthe nucleusrdquo Nature Reviews Genetics vol 6 no 1 pp 24ndash352005

[17] F Sato S Tsuchiya S J Meltzer and K Shimizu ldquoMicroRNAsand epigeneticsrdquo The FEBS Journal vol 278 no 10 pp 1598ndash1609 2011

[18] B Weber C Stresemann B Brueckner and F Lyko ldquoMethyla-tion of humanmicroRNA genes in normal and neoplastic cellsrdquoCell Cycle vol 6 no 9 pp 1001ndash1005 2007

[19] A Lujambio and M Esteller ldquoHow epigenetics can explainhuman metastasis a new role for microRNAsrdquo Cell Cycle vol8 no 3 pp 377ndash382 2009

[20] WChen R Zheng PD Baade et al ldquoCancer statistics inChina2015rdquo CA Cancer Journal for Clinicians vol 66 no 2 pp 115ndash132 2016

[21] E-J Kim Y-J Kim P Jeong Y-S Ha S-C Bae andW-J KimldquoMethylation of the RUNX3 promoter as a potential prognosticmarker for bladder tumorrdquo Journal of Urology vol 180 no 3 pp1141ndash1145 2008

Disease Markers 7

[22] H-Y Yoon Y-J Kim J S Kim et al ldquoRSPH9 methylationpattern as a prognostic indicator in patients with non-muscleinvasive bladder cancerrdquo Oncology Reports vol 35 no 2 pp1195ndash1203 2016

[23] S Guil and M Esteller ldquoDNA methylomes histone codesand miRNAs tying it all togetherrdquo International Journal ofBiochemistry and Cell Biology vol 41 no 1 pp 87ndash95 2009

[24] Y L Lin Z G Li Z K He T Y Guan and J Q Ma ldquoClinicaland prognostic significance of protocadherin-10 (PCDH10)promoter methylation in bladder cancerrdquo Journal of Interna-tional Medical Research vol 40 no 6 pp 2117ndash2123 2012

[25] Y Wang Y Yu R Ye et al ldquoAn epigenetic biomarker com-bination of PCDH17 and POU4F2 detects bladder canceraccurately by methylation analyses of urine sediment DNA inHan Chineserdquo Oncotarget vol 7 no 3 pp 2754ndash2764 2016

[26] I Renard S Joniau B van Cleynenbreugel et al ldquoIdentifi-cation and validation of the methylated TWIST1 and NID2genes through real-timemethylation-specific polymerase chainreaction assays for the noninvasive detection of primary bladdercancer in urine samplesrdquo European Urology vol 58 no 1 pp96ndash104 2010

[27] D R Yates I Rehman M F Abbod et al ldquoPromoter hyperme-thylation identifies progression risk in bladder cancerrdquo ClinicalCancer Research vol 13 no 7 pp 2046ndash2053 2007

[28] A-C Schneider L C Heukamp S Rogenhofer et al ldquoGlobalhistone H4K20 trimethylation predicts cancer-specific survivalin patients with muscle-invasive bladder cancerrdquo BJU Interna-tional vol 108 no 8 B pp E290ndashE296 2011

[29] Y An H Li K J Wang et al ldquoMeta-analysis of the relationshipbetween slow acetylation of N-acetyl transferase 2 and the riskof bladder cancerrdquo Genetics and Molecular Research vol 14 no4 pp 16896ndash16904 2015

[30] Z-M Jia X Ai J-F Teng Y-PWang B-JWang andX Zhangldquop21 and CK2 interaction-mediated HDAC2 phosphorylationmodulates KLF4 acetylation to regulate bladder cancer cellproliferationrdquoTumor Biology vol 37 no 6 pp 8293ndash8304 2016

[31] H Wang R Albadine A Magheli et al ldquoIncreased EZH2protein expression is associated with invasive urothelial carci-noma of the bladderrdquoUrologic Oncology Seminars and OriginalInvestigations vol 30 no 4 pp 428ndash433 2012

[32] M Fabbri and G A Calin ldquoEpigenetics and miRNAs in humancancerrdquo Advances in Genetics vol 70 pp 87ndash99 2010

[33] X Jiang L Du W Duan et al ldquoSerum microRNA expressionsignatures as novel noninvasive biomarkers for prediction andprognosis of muscle-invasive bladder cancerrdquoOncotarget vol 7no 24 pp 36733ndash36742 2016

[34] H Sasaki M Yoshiike S Nozawa et al ldquoExpression levelof urinary MicroRNA-146a-5p is increased in patients withbladder cancer and decreased in those after transurethralresectionrdquo Clinical Genitourinary Cancer 2016

[35] R Matsushita H Yoshino H Enokida et al ldquoRegulationof UHRF1 by dual-strand tumor-suppressor microRNA-145(miR-145-5p and miR-145-3p) inhibition of bladder cancer cellaggressivenessrdquo Oncotarget vol 7 no 19 2016

[36] S Hauser T Zahalka G Fechner S C Muller and J EllingerldquoSerumDNA hypermethylation in patients with kidney cancerresults of a prospective studyrdquo Anticancer Research vol 33 no10 pp 4651ndash4656 2013

[37] M de Martino T Klatte A Haitel and M Marberger ldquoSerumcell-free DNA in renal cell carcinoma a diagnostic and prog-nostic markerrdquo Cancer vol 118 no 1 pp 82ndash90 2012

[38] J Wang J Li J Gu et al ldquoAbnormal methylation status ofFBXW10 and SMPD3 and associations with clinical character-istics in clear cell renal cell carcinomardquoOncology Letters vol 10no 5 pp 3073ndash3080 2015

[39] Z R Wang J H Wei J C Zhou et al ldquoValidation of DAB2IPmethylation and its relative significance in predicting outcomein renal cell carcinomardquo Oncotarget vol 7 no 21 2016

[40] A D Goldberg C D Allis and E Bernstein ldquoEpigenetics alandscape takes shaperdquo Cell vol 128 no 4 pp 635ndash638 2007

[41] D B Seligson S Horvath M A McBrian et al ldquoGlobal levelsof histone modifications predict prognosis in different cancersrdquoTheAmerican Journal of Pathology vol 174 no 5 pp 1619ndash16282009

[42] Y Wang R Zhang D Wu et al ldquoEpigenetic change in kidneytumor downregulation of histone acetyltransferase MYST1 inhuman renal cell carcinomardquo Journal of Experimental ampClinicalCancer Research vol 32 no 1 article 8 2013

[43] G M Zhang L Luo X M Ding et al ldquoMicroRNA-126 inhibitstumor cell invasion and metastasis by downregulating ROCK1in renal cell carcinomardquoMolecular Medicine Reports vol 13 pp5029ndash2036 2016

[44] Z Wotschofsky L Gummlich J Liep et al ldquoIntegratedmicroRNA and mRNA signature associated with the transitionfrom the locally confined to themetastasized clear cell renal cellcarcinoma exemplified by miR-146-5prdquo PLOS ONE vol 11 no2 article e0148746 2016

[45] A Bettin I Reyes and N Reyes ldquoGene expression profilingof prostate cancer-associated genes identifies fibromodulin aspotential novel biomarker for prostate cancerrdquoThe InternationalJournal of Biological Markers vol 31 no 2 pp e153ndashe162 2016

[46] C Jeronimo and R Henrique ldquoEpigenetic biomarkers inurological tumors a systematic reviewrdquo Cancer Letters vol 342no 2 pp 264ndash274 2014

[47] S Dijkstra I L Birker F P Smit et al ldquoProstate cancerbiomarker profiles in urinary sediments and exosomesrdquo TheJournal of Urology vol 191 no 4 pp 1132ndash1138 2014

[48] V Urquidi C J Rosser and S Goodison ldquoMolecular diagnostictrends in urological cancer biomarkers for non-invasive diag-nosisrdquo Current Medicinal Chemistry vol 19 no 22 pp 3653ndash3663 2012

[49] V L Costa R Henrique S A Danielsen et al ldquoTCF21 andPCDH17 methylation an innovative panel of biomarkers for asimultaneous detection of urological cancersrdquo Epigenetics vol6 no 9 pp 1120ndash1130 2011

[50] L Van Neste R J Hendriks S Dijkstra et al ldquoDetectionof high-grade prostate cancer using a urinary molecularbiomarker-based risk scorerdquo European Urology 2016

[51] A Ouhtit M N Al-Kindi P R Kumar I Gupta S Shan-muganathan and Y Tamimi ldquoHoxb13 a potential prognosticbiomarker for prostate cancerrdquo Frontiers in Bioscience vol 8 no1 pp 40ndash45 2016

[52] G Hoyne C Rudnicka Q-X Sang et al ldquoGenetic and cellularstudies highlight that A Disintegrin andMetalloproteinase 19 isa protective biomarker in human prostate cancerrdquo BMCCancervol 16 no 1 article 151 2016

[53] L Zheng D Sun W Fan Z Zhang Q Li and T JiangldquoDiagnostic value of SFRP1 as a favorable predictive andprognostic biomarker in patients with prostate cancerrdquo PLoSONE vol 10 no 2 article e0118276 2015

[54] H Tahara H Naito K Kise et al ldquoEvaluation of PSF1 as aprognostic biomarker for prostate cancerrdquo Prostate Cancer andProstatic Diseases vol 18 no 1 pp 56ndash62 2015

8 Disease Markers

[55] R Morgan A Boxall A Bhatt et al ldquoEngrailed-2 (EN2) atumor specific urinary biomarker for the early diagnosis ofprostate cancerrdquo Clinical Cancer Research vol 17 no 5 pp1090ndash1098 2011

[56] C Haldrup A-S Lynnerup T M Storebjerg et al ldquoLarge-scaleevaluation of SLC18A2 in prostate cancer reveals diagnosticand prognostic biomarker potential at three molecular levelsrdquoMolecular Oncology vol 10 no 6 pp 825ndash837 2016

[57] K D Berg D Soldini M Jung et al ldquoTRPM4 protein expres-sion in prostate cancer a novel tissue biomarker associated withrisk of biochemical recurrence following radical prostatectomyrdquoVirchows Archiv vol 468 pp 345ndash355 2016

[58] M V Russo S Esposito M G Tupone et al ldquoSOX2 boostsmajor tumor progression genes in prostate cancer and is afunctional biomarker of lymph node metastasisrdquo Oncotargetvol 7 no 11 pp 12372ndash12385 2016

[59] J Hao Y T Chiang P W Gout and Y Wang ldquoElevated XPO6expression as a potential prognostic biomarker for prostatecancer recurrencerdquo Frontiers in Bioscience vol 8 no 1 pp 44ndash55 2016

[60] F Sanguedolce A CormioM Brunelli et al ldquoUrine TMPRSS2ERG fusion transcript as a biomarker for prostate cancerliterature reviewrdquo Clinical Genitourinary Cancer vol 14 no 2pp 117ndash121 2016

[61] JWangH YeD Zhang et al ldquoMicroRNA-410-5p as a potentialserum biomarker for the diagnosis of prostate cancerrdquo CancerCell International vol 16 no 1 article 12 2016

[62] G Al-Kafaji Z T Al-Naieb and M Bakhiet ldquoIncreasedoncogenic microRNA-18a expression in the peripheral bloodof patients with prostate cancer a potential novel non-invasivebiomarkerrdquo Oncology Letters vol 11 no 2 pp 1201ndash1206 2016

[63] S Xu X M Yi W Q Zhou et al ldquoDownregulation of miR-129 in peripheral blood mononuclear cells is a diagnostic andprognostic biomarker in prostate cancerrdquo International Journalof Clinical and Experimental Pathology vol 8 no 11 pp 14335ndash14344 2015

[64] A E Dago A Stepansky A Carlsson et al ldquoRapid phenotypicand genomic change in response to therapeutic pressure inprostate cancer inferred by high content analysis of singlecirculating tumor cellsrdquo PLoS ONE vol 9 no 8 article e1017772014

[65] J G Lohr V A Adalsteinsson K Cibulskis et al ldquoWhole-exome sequencing of circulating tumor cells provides a windowinto metastatic prostate cancerrdquo Nature Biotechnology vol 32no 5 pp 479ndash484 2014

[66] M J M Magbanua E V Sosa J H Scott et al ldquoIsolationand genomic analysis of circulating tumor cells from castrationresistantmetastatic prostate cancerrdquoBMCCancer vol 12 article78 2012

[67] W Onstenk W de Klaver R de Wit M Lolkema J Foekensand S Sleijfer ldquoThe use of circulating tumor cells in guidingtreatment decisions for patients with metastatic castration-resistant prostate cancerrdquoCancer Treatment Reviews vol 46 pp42ndash50 2016

[68] O Sartor and Y Dong ldquoAndrogen receptor variant-7 animportant predictive biomarker in castrate resistant prostatecancerrdquo Asian Journal of Andrology vol 17 no 3 pp 439ndash4402015

[69] T Dellavedova ldquoProstatic specific antigen From its early daysuntil becoming a prostate cancer biomarkerrdquoArchivos Espanolesde Urologia vol 69 no 1 pp 19ndash23 2016

[70] J Ellinger S CMuller andD Dietrich ldquoEpigenetic biomarkersin the blood of patients with urological malignanciesrdquo ExpertReview ofMolecular Diagnostics vol 15 no 4 pp 505ndash516 2015

[71] J Li R W Veltri Z Yuan C S Christudass and W Man-decki ldquoMacrophage inhibitory cytokine 1 biomarker serumimmunoassay in combination with PSA is a more specificdiagnostic tool for detection of prostate cancerrdquo PLoS ONE vol10 no 4 Article ID e0122249 2015

[72] L Mirabello C P Kratz S A Savage and M H GreeneldquoPromoter methylation of candidate genes associated withfamilial testicular cancerrdquo International Journal of MolecularEpidemiology and Genetics vol 3 no 3 pp 213ndash227 2012

[73] L Mirabello S A Savage L Korde S M Gadalla and M HGreene ldquoLINE-1 methylation is inherited in familial testicularcancer kindredsrdquo BMCMedical Genetics vol 11 article 77 2010

[74] B-F Chen S Gu Y-K Suen L Li and W-Y ChanldquomicroRNA-199a-3p DNMT3A and aberrant DNA methyla-tion in testicular cancerrdquo Epigenetics vol 9 no 1 pp 119ndash1282014

[75] M Brait L Maldonado S Begum et al ldquoDNA methylationprofiles delineate epigenetic heterogeneity in seminoma andnon-seminomardquo British Journal of Cancer vol 106 no 2 pp414ndash423 2012

Submit your manuscripts athttpwwwhindawicom

Stem CellsInternational

Hindawi Publishing Corporationhttpwwwhindawicom Volume 2014


MEDIATORSINFLAMMATION

of


Behavioural Neurology

EndocrinologyInternational Journal of



Disease Markers


BioMed Research International

OncologyJournal of



Oxidative Medicine and Cellular Longevity


PPAR Research

The Scientific World JournalHindawi Publishing Corporation httpwwwhindawicom Volume 2014

Immunology ResearchHindawi Publishing Corporationhttpwwwhindawicom Volume 2014

Journal of

ObesityJournal of



Computational and Mathematical Methods in Medicine

OphthalmologyJournal of


Diabetes ResearchJournal of



Research and TreatmentAIDS


Gastroenterology Research and Practice


Parkinsonrsquos Disease

Evidence-Based Complementary and Alternative Medicine

Volume 2014Hindawi Publishing Corporationhttpwwwhindawicom

2 Disease Markers

contamination and loss of DNA templates arising from theemployment of bisulfite treatment

Histones modification includes methylation carbonyla-tion phosphorylation ubiquitinylation sumoylation glyco-sylation and acetylation which are known as ldquothe histonecoderdquo together with DNAmethylation regulating the expres-sion of specific genes [8] The mechanism of regulation isthrough the addition of specific chemical groups to the N-terminal of basic amino acid residues at the tails of histonesaltering the affinity of the histone tails to the DNA andchanging the conformation of chromatin structure to regulatethe transcription of genes For instance histone acetylationenhances the transcriptional activity of gene for lowering theaffinity of histone tails to theDNA while histone deacetylasesresult in gene repression [9 10] Contrary to the DNAmethy-lation the histone methylation (addition of methyl groupsto H3 H4 lysine and arginine) can generate two effectseither activation or repression transcription depending onthe specific amino acid residues modified [11] In cancercell genome-wide histone modification and expression ofkey histone modulating enzymes have been reported but thedetailed mechanisms of their involvement in tumorigenesisare poorly understood [12 13]

Another epigenetic modification has been studied deeplyknown as microRNAs (miRNAs) miRNAs a class of non-coding RNAs which are about 18ndash25 nucleotides at lengthcould be synthesized and processed in the nucleus miRNAsalter the expression through binding to mRNAs then RNA-induced silencing complex takes part and regulates proteintranslation [14 15] It was estimated that at least 30 ofhuman genes are regulated by miRNA It is noteworthy thateach mRNA can be combined with multiple miRNAs andeach miRNA may regulate multiple mRNAs miRNAs playan important role in tumorigenesis activating oncogenes orrestraining tumor suppressor genes [14 15] The ways thatmiRNAs alter gene expression include deletion amplifica-tion mutation and chromosomal abnormalities [16] Somestudies found that miRNA can influence gene expression viatargeting a specific gene region for DNA methylation andhistone modifications [16 17] and the expression of miRNAcan be regulated by other epigenetic modifications such asDNA methylation [18 19] which indicated the interactionsbetween miRNAs and other epigenetic mechanisms

3 Epigenetic Biomarkers in Bladder Cancer

Bladder cancer is the most common male urology malig-nancy in China and the total number of cases in malesfor 2015 is approximately 621 thousand [20] Currently thediagnosis of bladder is mainly invasive causing discomfortto patients and can only provide a generalized outcomefor patients so the demand for noninvasive screening anddiagnosis method is urgent Epigenetic biomarkers candecrease the use of invasive methods and provide the earlydiagnosis of bladder cancer allowing for more effectivetreatment For example RUNX3 gene that has been thoughtas tumor suppressor gene exhibited significant increase atmethylation levels in bladder cancer by analyzing 124 tumortissue samples This study shows that RUNX3 gene may

be a potential marker for detecting bladder cancer [21]Apart from diagnostic markers a prognostic indicator inpatients with non-muscle-invasive bladder cancer (NMIBC)was found by Yoon et al The clinical relevance of RSPH9was determined by quantitative pyrosequencing analysis of136 human bladder specimens (8 normal controls and 128NMIBCs) They concluded that RSPH9 methylation can beof value for the assessment of disease recurrence and anindependent prognostic indicator in NMIBC patients [22]Similarly Lin and Luo et al reported that the hypermethy-lation of PCDH10 (50 119899 = 117) and PCDH17 (52 119899 =151) was related to the development of bladder cancer and itwas an independent predictor of cancer-specific survival time[23 24]

With the deepening of research it was thought that therewas no single gene found to be methylated in vast majority ofbladder tumors More and more researches have focused onusing gene panels to early detect bladder cancer and predicttumor recurrence progression and metastasis A methyla-tion analysis of PCDH17 andPOU4F2was developed recentlyin 148 individuals by qMSP using urine sediment Thecombination of POU4F2PCDH17 detection had sensitivityand specificity of 9000 and 9396 in the validation of 312individuals [25] Renard et al found that 2 genes (TWIST1andNID2) were frequentlymethylated in urine samples frombladder cancer patients (93 specificity and 90 sensitivity)[26] Another study discovered that a methylation genepanel which consisted of 4 genes (CDH1 CDH13 RASSF1Aand APC) had significant correlations with poor prognosis(cancer with high grade advanced stages and aneuploidies)Likewise a study conducted by Yates et al demonstratedthat promoter methylation of RASSF1A CDH1 TNFSR25EDNRB and APC was associated with tumor progression[27]

Histone modifications were also found to relate to thepathogenesis of bladder cancer and regulation of cancer cellproliferation A study in 2011 identified global histoneH4K20trimethylation that could predict cancer-specific survival inpatients with muscle-invasive bladder cancer [28] A meta-analysis published by An et al showed that merged oddsratio of the effect between slow acetylation and bladdercancer was 131 (95 confidence odds ratio interval = 111ndash155) illustrating that slow acetylation modestly increasesthe risk of bladder cancer [29] Recently Jia et al foundthat the acetylation status of KLF4 can decide the genefunction as a tumor suppressor or oncogene in bladdercancer Their results indicated that deacetylated KLF4 canact as an oncogene accelerating bladder cancer proliferationon the contrary acetylation of KLF4 performs as a tumorsuppressor which may be a good target for bladder cancertherapy [30] In addition another study found that expressionlevels of H3K4me1 H4K20me1 H4K20me2 and H4K20me3were correlated with advanced pathological stage and thatH4K20me3 was an independent prognosis factor for thesurvival of patients with muscle-invasive bladder cancer [31]

miRNAs are synthesized and processed in the nucleusand released into cytoplasm binding to specific mRNA andinducing gene silence [32] Recently Jiang et al demonstrated

Disease Markers 3











4 Disease Markers













Disease Markers 5












6 Disease Markers


7 Conclusions


Competing Interests




References























Disease Markers 7


































8 Disease Markers



























of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of
















Disease Markers 3











4 Disease Markers













Disease Markers 5












6 Disease Markers


7 Conclusions


Competing Interests




References























Disease Markers 7


































8 Disease Markers



























of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of
















4 Disease Markers













Disease Markers 5












6 Disease Markers


7 Conclusions


Competing Interests




References























Disease Markers 7


































8 Disease Markers



























of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of
















Disease Markers 5












6 Disease Markers


7 Conclusions


Competing Interests




References























Disease Markers 7


































8 Disease Markers



























of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of
















6 Disease Markers


7 Conclusions


Competing Interests




References























Disease Markers 7


































8 Disease Markers



























of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of
















Disease Markers 7


































8 Disease Markers



























of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of
















8 Disease Markers



























of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of





















of






Disease Markers



OncologyJournal of





PPAR Research



Journal of

ObesityJournal of
















review article new progress of epigenetic biomarkers in...

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