Table 1: Summary of investigated and published biomarkers in the GERD-BE-EAC axis. The categorization is based on four groups according to their potential usage as A = Diagnostic Biomarker indicates the presence of disease, B = Progression Biomarker indicates the risk of developing cancer—progression in BE to EAC, C = Predictive Biomarker predicts response to therapy (CTX, RTX, photodynamic therapy), or D = Prognostic Biomarker indicates overall survival—prognostic in EAC (survival, recurrence).

BiomarkerMethodRemarks/findingsOR/RR/ valueRefs

A = Diagnostic BiomarkerTFF3novel nonendoscopic screening modality in a prospective cohort study (for maximal length of BE) 
(for circumferential length of BE)
TFF3 IHC, esophageal cytosponge samples for BE combined with IHC for TFF3biomarker to screen asymptomatic patients for BE;
TFF3 protein was expressed at the luminal surface of BE (not at normal esophageal or gastric mucosa)
Chromosomes 7 and 17 (copy number changes) ICDA & FISHchromosomal gains in early stages of BE;
valuable adjunct to conventional cytology to detect dysplasia or EAC
IND/LGD: 75% sensitivity, (76% specificity)
HGD/EAC: 85% sensitivity,
(84% specificity)
8q24 (C-MYC), 17q12 (HER2), and 20q13 (copy number changes)FISHchromosomal gains in early stages of BE;
represents a valuable adjunct to conventional cytology to detect dysplasia or EAC
LGD (50% sensitivity)
HGD (82% sensitivity)
EAC (100% sensitivity)
17q11.2 (ERBB2)Southern blotting, microarray analysisamplified copies of the ERBB2 gene in EAC10-fold amplification in 3 of 25 (12%) tumors [52]
Serum proteomic pattern analysismass spectrometryseveral limitations due to applied technology identified 10 of 11 normal’s; and 42 of 43 EAC’s correctly[53]

B = Progression BiomarkersP53 positivityIHClimited efficacy as a single progression biomarkerOR 11.7 (95% CI: 1.93–71.4)[54]
P53 positivityIHCpositive in 4/31 that regressed, 3/12 that persisted, and 3/5 that progressed to HGD or EAC RR not available[55]
DNA content abnormalities flow cytometryhigher relative risk for EAC in patients with tetraploidy (4N) or aneuploidy (>6%)tetraploidy: RR 7.5 (95% CI: 4–14) ( )
aneuploidy: RR 5.0 (95% CI: 2.7–9.4) ( )
4N fraction cut point of 6% for cancer risk RR 11.7 ( 95% CI: 6.2–22)
aneuploid DNA contents of 2.7N were predictive of higher cancer riskRR 9.5 (95% CI: 4.9–18)
DNA content abnormalities flow cytometrypresence of both 4N fraction of 6% and aneuploid DNA content of 2.7N is highly predictive for progression RR 23 (95% CI: 10–50) [57]
17p(p53) LOH associated with higher risk of progression to HGD + EACHGD: RR 3.6 ( )
flow cytometry, PCREAC: RR 16 ( ) [58]
combined LOH of 17p and 9p and DNA content abnormalities can best predict progression to EACRR 38.7 (95% CI: 10.8–138.5) not clinical applicable
LOH of 157p and 9p and DNA content abnormalitiesLOH of 17p aloneRR 10.6 (95% CI: 5.2–21.3)
flow cytometry, PCRLOH of 9p aloneRR 2.6 (95% CI: 1.1–6.0)
Aneuploidy alone RR 8.5 (95% CI: 4.3–17.0) [59]
Tetraploidy aloneRR 8.8 (95% CI: 4.3–17.7)
mutations of p16 and p53 loci (clonal diversity measurements)flow cytometry, PCRsignificant predictors for EAC progression, not clinical applicable [60]
EGFR IHCoverexpression in HGD/EAC35% of HGD/80% of EAC specimens[61]
MCM2IHCcorrelation between degree of dysplasia and level of ectopic luminal surface MCM2 expressionMCM2-positive
staining in 42% (19/45) of BE samples
Cyclin A IHCsurface expression of cyclin A in BE samples correlates with the degree of dysplasia OR 7.5 (95% CI: 1.8–30.7) ( )[63]
Cyclin D1IHCassociation with increased risk of EACOR 6.85 (95% CI: 1.57–29.91)[64]
hypermethylation of p16 (CDKI2A)association with increased risk of progression to HGD/EACOR 1.74 (95% CI: 1.33–2.2)
hypermethylation of RUNX3 association with increased risk of progression to HGD/EACOR 1.80 (95% CI: 1.08–2.81)
hypermethylation of HPP1 RT-PCRassociation with increased risk of progression to HGD/EACOR 1.77 (95% CI: 1.06–2.81) [41]
hypermethylation of p16 and APCPCRpredictor of progression to HGD/EACOR 14.97 (95% CI: 1.73–inf.)[65]
8 gene methylation panel RT-PCRage dependent; predicts 60.7% of progression to HGD/EAC within 2 yrsRR not available (90% specificity)[66]
Gene expression profilemicroarray analysis64 genes up regulated
110 genes down regulated in EAC
Cathepsin D, AKR1B10, and AKR1C2 mRNA levelsWestern blotting, qRT-PCRdysregulation predicts progression to HGD/EACAKR1C2:
levels in BE ( ) but levels in EA ( )
ICDAaneuploidy predicts progression to EAC60% with LGD; 73% with HGD, and 100% with EAC (total number of samples = 56)[69]
DNA abnormalitiesACISfrequency and severity of aneuploidy predicts progression to EACunstable aneuploidy in 95% with EAC[70]
DICMrelationship between DICM status and progression to HGD/EAC [71]
SNP-based genotyping in BE/EAC specimensflow cytometry, 33K SNP arraycopy gains, losses, and LOH increased in frequency and size between early and late stage of disease (BE)[72]

C = Predictive Biomarkersp16 allelic loss FISHdecreased response to photodynamic therapyOR 0.32 (95% CI: 0.10–0.96) [73]
DNA ploidy abnormalities ICDADNA ploidy as a covariate value for recurrenceHR 6.3 (1.7–23.4) ( )[74]
HSP27IHCassociation between low HSP27 expression and no response to neoadjuvante chemotherapy and [75]
Ephrin B3 receptormicroarrayresponse prediction in EAC in patients with Ephrin B3 receptor positive versus Ephrin B3 receptor negativeResponse rate <50%: 3 (15.8) versus 16 (84.2) ( )[76]
Genetic polymorphismsqRT-PCRassociation between individual single nucleotide polymorphisms
and clinical outcomes
comprehensive panel of genetic polymorphisms on clinical outcomes in 210 esophageal cancer patients[77]
P21IHCalteration in expression correlated with better CTX-response [78]
P53IHCalteration in expression correlated with better CTX-response [79]
ERCC1IHCERCC1-positivity predicts CTX-resistance and poor outcome [80]

D = Prognostic BiomarkersDCK
prognostic 4-gene signature in EAC predicts 5-year survival0/4 genes dysregulated: 58%
(95% CI: 36%–80%)
1-2/4 genes dysregulated: 26%
(95% CI: 20%–32%)
3-4/4 genes dysregulated: 14%
(95% CI: 4%–24%)
( )
p16 loss
C-MYC gain
FISHassociation between therapy response status and FISH positivity [82]
ASS expressionmicroarrayslow expression correlates with lymph node metastasis [83]
microRNA expression profilesmiRNA microarray, qRT-PCRassociation with prognosis (e.g. low levels of mir-375 in EAC → worse prognosis)HR = 0.31 (95% CI: 0.15–0.67) ( )[84]
Genomic alterations MLPAreverse association between survival and DNA copy number alterations (>12 aberrations low mean survival) [85]
Cyclin D1FISH, IHC2 of 3 genotypes confers to survival [86]
IHCexpression = survival [87]
EGFRIHC expression = survival [88]
Ki-67IHClow levels of staining (<10%)
= survival
Her2/neuFISHamplification = survival [90, 91]
IHClow levels = survival [92]
TGF-αIHC, ISHhigh levels = tumor progression and lymph node metastasis and [93]
qRT-PCRoverexpression = survival [94]
TGF-β1ELISAhigh plasma levels = survival [95]
APCRT-PCRhigh plasma levels of methylation
= survival
Bcl-2IHCexpression = survival [97]
IHC, RT-PCR expression = survival, TN-stage, and recurrence ,
, and
IHCstrong staining = survival [99]
COX-2IHCstrong staining = survival, distant metastasis, and recurrence , , and [100]
NF-κBIHCactivated NF-κB = survival, and
disease free survival
and [101]
TelomeraseSouthern blot analysis, RT-PCRhigher telomere-length ratio
= survival
RR of death: 3.4
(CI: 1.3–8.9) ( )
expression = survival,
CD105angiolymphatic invasion
lymph node metastasis
IHC distant metastasis [103]
expression = survival,
VEGFangiolymphatic invasion
lymph node metastasis
distant metastasis
CadherinIHC level = survival [89]
uPAELISA uPA = survival [104]
TIMPIHC, RT-PCR expression = survival, and disease stage and [105]
Promoter hypermethylation of multiple genesIHC, methylation specific PCRif >50% of gene profile methylated
= survival, and earlier recurrence
and [106]
MGMT hypermethylationIHC, methylation specific PCRcorrelation with higher tumor differentiation [107]

ACIS: automated cellular imaging system; ASS: argininosuccinate synthase; APC: adenomatous polyposis coli; BE: barrett’s esophagus; COX: cyclooxygenase; DCK: deoxycytidine kinase; DICM: digital image cytometry; EAC: esophageal adenocarcinoma; EGFR: epidermal growth factor receptor; ELISA: enzyme-linked immunosorbent assay; FISH: fluorescence in-situ-hybridization; ICDA: image cytometric DNA analysis; HSP27: Heat-shock protein 27; IHC: immunohistochemistry; LOH: loss of heterozygosity; PAPSS2: 3′-phosphoadenosine 5′-phosphosulfate synthase 2; PCR: polymerase chain reaction; qRT: quantitative reverse transcriptase; MLPA: multiplex ligation dependent probe amplification; NF-κB: nuclear factor kappa B; SIRT2: Sirtuin 2; SNP: single nucleotide polymorphism; TFF3: Trefoil factor 3; TGF: transforming growth factor; TIMP: tissue inhibitors of metalloproteinases; TRIM44: Tripartite motif-containing 44; uPA: urokinase-type plasminogen activator; VEGF: vascular endothelial growth factor.