Esophageal adenocarcinoma (EAC) arises after normal squamous mucosa undergo
es metaplasia to specialized columnar epithelium (intestinal metaplasia or
Barrett's esophagus), which can then ultimately progress to dysplasia and s
ubsequent malignancy. Epigenetic studies of this model have thus far been l
imited to the DNA methylation analysis of a few genes. In this study, we an
alyzed a panel of 20 genes using a quantitative, high-throughput methylatio
n assay, MethyLight. Ne used this broader approach to gain insight into con
cordant methylation behavior between genes and to generate epigenomic finge
rprints for the different histological stages of EAC. Our study included a
total of 104 tissue specimens from 51 patients with different stages of Bar
rett's esophagus and/or associated adenocarcinoma. We screened 84 of these
samples with the full panel of 20 genes and found distinct classes of methy
lation patterns in the different types of tissue. The most informative gene
s,were those with an intermediate frequency of significant hypermethylation
[ranging from 15% (CDKN2A) to 60% (MGMT) of the samples]. This group could
be further subdivided into three classes, according to the absence (CDKN2A
, ESRI, and MYODI) or presence (CALCA, MGMT, and TIMP3) of methylation in n
ormal esophageal mucosa and stomach, or the infrequent methylation of norma
l esophageal mucosa accompanied by methylation in all normal stomach sample
s (APC). The other genes were less informative, because the frequency of hy
permethylation was below 5% (ARF, CDH1, CDKN2B, GSTP1, MLH1, PTGS2, and THB
S1), completely absent (CTNNB1, RBI, TGFBR2, and TYMS1), or ubiquitous (H1C
1 and MTHFR), regardless of tissue type. Each class undergoes unique epigen
etic changes at different steps of disease progression of EAC, suggesting a
step-wise loss of multiple protective barriers against CpG island hypermet
hylation. The aberrant hypermethylation occurs at many different loci in th
e same tissues, suggestive of an overall deregulation of methylation contro
l in EAC tumorigenesis. However, we did not find evidence for a distinct gr
oup of tumors with a CpG island methylator phenotype. Finally,we found that
normal and metaplastic tissues from patients with evidence of associated d
ysplasia or cancer had a significantly higher incidence of hypermethylation
than similar tissues from patients with no further progression of their di
sease. The fact that the samples from these two groups of patients were his
tologically indistinguishable, yet molecularly distinct, suggests that the
occurrence of such hypermethylation may provide a clinical tool to identify
patients with premalignant Barrett's who are at risk for further progressi
on.