Identification of Differentially Expressed Genes in Esophageal Squamous Cell Carcinoma (ESCC) by cDNA Expression Array

Abstract

Purpose: This study aims to identify differentially expressed genes in esophageal squamous cell carcinoma (ESCC) through the use of a membrane-based cDNA array.

Experimental Design: Two newly established human ESCC cell lines (HKESC-1 and HKESC-2) and one corresponding to a morphologically normal, esophageal epithelium tissue specimen, prospectively collected from the HKESC-2-related patient, were screened in parallel using a cDNA expression array containing gene-specific fragments for 588 human genes spotted onto nylon membranes.

Results: The results of cDNA expression array showed that 53 genes were up-regulated 2-fold or higher and 8 genes were down-regulated 2-fold or higher in both ESCC cell lines at the mRNA level. Semiquantitative RT-PCR analysis of a subset of these differentially expressed genes gave results consistent with cDNA array findings. Four of the differentially expressed genes that belong to the categories of oncogenes/tumor suppressor genes (Fra-1 and Neogenin) and cell cycle-related genes (Id-1 and CDC25B) were studied more extensively for their protein expression by immunohistochemistry. The two ESCC cell lines and their corresponding primary tissues, 61 primary ESCC resected specimens and 16 matching, morphologically normal, esophageal epithelium tissues were analyzed. The immunostaining results showed that Fra-1, Neogenin, Id-1, and CDC25B were overexpressed in both ESCC cell lines and their corresponding primary tumors at the protein level, validating the microarray findings. The results of the clinical specimens showed that the Fra-1 gene was overexpressed in ESCC compared with normal esophageal epithelium in 53 of 61 cases (87%), Neogenin in 57 of 61 cases (93%), Id-1 in 57 of 61 cases (93%), and CDC25B in 48 of 61 cases (79%). Furthermore, the expression of Fra-1, Neogenin, and Id-1 in ESCC correlated with tumor differentiation.

Conclusions: Overall, this study demonstrates that multiple genes are differentially expressed in ESCC and provides the first evidence that oncogenes Fra-1 and Neogenin and cell cycle-related genes Id-1 and CDC25B are overexpressed in ESCC.