247Alfred K. Lam (ed.), Esophageal Adenocarcinoma: Methods and Protocols, Methods in Molecular Biology, vol. 1756,
https://doi.org/10.1007/978-1-4939-7734-5_21, © Springer Science+Business Media, LLC 2018
Chapter 21
Epigenetics: DNA Methylation Analysis in Esophageal
Adenocarcinoma
Farhadul Islam, Johnny C. Tang, Vinod Gopalan, and Alfred K. Lam
Abstract
The aberrant DNA methylation has been noted to occur at promoter of tumor suppressor, cell adhesion,
DNA repair, and other growth regulating genes during the progression of nonneoplastic esophageal
mucosa to Barrett esophagus to esophageal adenocarcinoma. Methylation-mediated silencing of individual
gene or concurrent loss of a number of genes plays crucial roles in dysplasia-metaplasia-neoplasia sequence
of esophageal adenocarcinoma. In addition, promoter methylation of genes had shown significant prog-
nostic potential in patients with esophageal adenocarcinoma. Thus, determination of methylation status of
genes of interest can be used as a molecular marker for risk stratification and/or better prognosis of
patients with esophageal adenocarcinoma. There are a number of methods including bead array, PCR and
sequencing, pyrosequencing, methylation-specific PCR, and PCR with high-resolution melt curve avail-
able to determine the methylation status of particular gene of interest. Herein, we describe the polymerase
chain reaction followed by sequencing-based protocol for identifying DNA methylation status in esopha-
geal adenocarcinoma.
Key words DNA methylation, Bisulfite conversion, PCR, Esophageal adenocarcinoma, Barrett
esophagus1 Introduction
The epigenetic regulation of gene expression plays important roles
in embryonic development, imprinting, X-chromosome inactivation,
and tissue differentiation in mammals [ 1 , 2 ]. Alteration of epigen-
etic landscape, consisting of DNA methylation, histone modification,
nucleosome positioning, and microRNAs, leads to aberrant expres-
sion of genes in normal and cancerous cells [ 1 – 3 ]. These global
changes in epigenetics are the hallmark of malignant transformation
and lead to cancer [ 2 ]. DNA methylation is one of the most com-
mon epigenetic alterations involved in the pathogenesis of different
cancers including esophageal adenocarcinoma [ 4 , 5 ].
Numerous genetic and epigenetic changes occur during the dys-
plasia-metaplasia-neoplasia sequence in the pathogenesis of esopha-
geal adenocarcinoma [ 4 , 5 ]. Among the epigenetic alterations,