Identification of EGFR expression status association with metastatic lymph node density (ND) by expression microarray analysis of advanced gastric cancer

Metastatic lymph node density (ND) has been reproducibly proven to be a prognostic factor in gastric cancer. The molecular mechanisms that underlie this aggressiveness are underexplored. Here, we aimed to identify molecules associated with this unique phenotype. Tumor specimens from patients with stage III gastric cancer with high or low ND (n = 4 for both) were compared at the mRNA level using Affymetrix microarray (harboring 54,675 genes). The expression data were prioritized, and genes that correlated with ND were selected. Ultimately, the EGFR was validated as such a candidate molecule in patients with primary advanced gastric cancer who underwent standard treatment (n = 167). Expression data of the microarray were prioritized based on gene expression ratio and frequency of gene expression. The first priority genes to be selected were genes that are known to be amplified in cancer, which included NKX2.1, CHST9, CTNND2, SLC25A27, FGFR2, EGFR, and PTGER1. Of these genes, the EGFR gene was of particular interest. EGFR expression in primary gastric cancer was examined using immunohistochemistry (IHC). The Student's t-test elucidated a significant difference in EGFR expression between IHC 2+/3+ and IHC 1+ according to ND (P = 0.0035). The Chi-square test also indicated a significant difference between high and low levels of EGFR immunohistochemical staining (IHC2+/3+ and IHC1+, respectively) and ND status (P = 0.0023). According to the least squares method, as ND increased, the risk that EGFR staining levels changed from IHC 1+ to IHC 2+ also increased. In this study, we determined that high EGFR expression may underlie the aggressive mechanism of advanced gastric cancer with high ND.