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)
[48]
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)
[49]
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)
[50]
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)
[51]
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) ( )
[56]
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
[62]
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
[67]
Cathepsin D, AKR1B10, and AKR1C2 mRNA levelsWestern blotting, qRT-PCRdysregulation predicts progression to HGD/EACAKR1C2:
levels in BE ( ) but levels in EA ( )
[68]
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
PAPSS2
SIRT2
TRIM44
RT-PCR,
IHC
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%)
( )
[81]
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
[89]
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
[96]
Bcl-2IHCexpression = survival [97]
IHC, RT-PCR expression = survival, TN-stage, and recurrence ,
, and
[98]
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) ( )
[102]
expression = survival,
CD105angiolymphatic invasion
lymph node metastasis
T-stage
IHC distant metastasis [103]
expression = survival,
VEGFangiolymphatic invasion
lymph node metastasis
T-stage
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.