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Alfred K. Lam (ed.), Esophageal Adenocarcinoma: Methods and Protocols, Methods in Molecular Biology, vol. 1756,
https://doi.org/10.1007/978-1-4939-7734-5_23, © Springer Science+Business Media, LLC 2018
Chapter 23
RNA Interference-Mediated Gene Silencing in Esophageal
Adenocarcinoma
Farhadul Islam, Vinod Gopalan, and Alfred K. Lam
Abstract
RNA interference (RNAi) is a normal physiological mechanism in which a short effector antisense RNA
molecule regulates target gene expression. It is a powerful tool to silence a particular gene of interest in a
sequence-specific manner and can be used to target against various molecular pathways in esophageal
adenocarcinoma by designing RNAi targeting key pathogenic genes. RNAi-based therapeutics against
esophageal adenocarcinoma can be developed using different strategies including inhibition of overex-
pressed oncogenes, blocking cell division by interfering cyclins and related genes or enhancing apoptosis
by suppressing anti-apoptotic genes. In addition, RNAi against multidrug resistance genes or chemo-
resistance targets may provide promising cancer therapeutic options. Here, we describe RNAi technology
using MET, a proto-oncogene in esophageal adenocarcinoma cells, as a model target. Lentiviral particles
expressing MET shRNA was used to silence MET genes. Then, Western blot analysis was performed to
confirm MET knockdown.
Key words RNAi, Esophageal adenocarcinoma, Western blot, siRNA, shRNAs
1 Introduction
RNA interference (RNAi) is a group of naturally occurring gene
silencing mechanisms, in which a short antisense RNA (19–23
nucleotides) molecule inhibits the target gene expression by degrad-
ing after transcription of corresponding mRNA molecules [ 1 – 3 ].
This natural process is now being adopted to use as a powerful tool
to investigate gene functions by silencing the gene expression of
interest in a sequence-specific manner. Also, RNAi opens new
opportunities to develop therapeutics for clinically challenging dis-
eases (e.g., cancer) by specific inhibition of the key pathogenic
gene(s) [ 1 , 4 ]. The discovery of RNAi-mediated post-transcrip-
tional gene silencing was first reported in 1998 [ 5 ]. In plants, the
small RNA molecules detect and degrade target viral mRNA,
thereby exhibited defense against viral infection [ 6 ]. However, the
potential and efficacy of RNAi therapeutic intervention depend on