A double-negative feedback loop between EpCAM and ERK contributes to the regulation of epithelial-mesenchymal transition in cancer.

Epithelial-mesenchymal transition (EMT) is an important biological process that has been implicated in cancer metastasis. Epithelial cell adhesion molecule (EpCAM) is expressed at the basolateral membrane of most normal epithelial cells but is overexpressed in many epithelial cancers. In our studies on the role of EpCAM in cancer biology, we observed that EpCAM expression is decreased in mesenchymal-like primary cancer specimens in vivo and following induction of EMT in cancer cell lines in vitro. Extracellular signal-related kinase (ERK) is a key regulator of EMT. We observed that EpCAM expression is decreased with activation of the ERK pathway in primary cancer specimens in vivo and in cancer cell lines in vitro. In experimental models, growth factor stimulation and/or oncogene-induced ERK2 activation suppressed EpCAM expression, whereas genetic or pharmacological inhibition of the ERK pathway restored EpCAM expression. In detailed studies of the EpCAM promoter region, we observed that ERK2 suppresses EpCAM transcription directly by binding to a consensus ERK2-binding site in the EpCAM promoter and indirectly through activation of EMT-associated transcription factors SNAI1, SNAI2, TWIST1 and ZEB1, which bind to E-box sites in the EpCAM promoter. Surprisingly, EpCAM appears to modulate ERK activity. Using multiple cell lines, we demonstrated that specific ablation of EpCAM resulted in increased ERK pathway activity and SNAI2 expression, migration and invasion, whereas forced expression of EpCAM resulted in decreased ERK pathway activity and SNAI2 expression, migration and invasion. These observations provide important insights into the regulation of EpCAM expression during EMT, demonstrate an unexpected role for EpCAM in the regulation of ERK and define a novel double-negative feedback loop between EpCAM and ERK that contributes to the regulation of EMT. These studies have important translational implications as both EpCAM and ERK are currently being targeted in human clinical trials.

Overexpression of CEBPbeta correlates with decreased TFF1 in gastric cancer.

CCAAT element binding protein beta (C/EBPbeta) is an important regulator of cell growth, differentiation and in promoting tumor invasiveness. C/EBPbeta is located on chromosome 20q, which is amplified in many solid tumors including gastric cancers (GC). We sought to characterize the status of C/EBPbeta expression in GCs, which was recently found to repres TFF1 gene. Microarray analysis revealed overexpression of C/EBPbeta in 25 of 27 (93%) GC when compared to 12 normal gastric tissue samples. RT-PCR analysis confirmed the overexpression of C/EBPbeta transcripts in 54 of 59 (91%) GC. In total, 15 of 18 gastric tumors exhibited at least fivefold higher C/EBPbeta transcript levels compared to their corresponding adjacent normal gastric tissue samples. Moreover, immunohistochemistry analysis demonstrated increased nuclear staining of C/EBPbeta in 10 of 13 GC and at least fourfold overexpression of C/EBPbeta in three primary GC compared to adjacent normal gastric tissue. Furthermore, a striking correlation of decreased TFF1 expression with increased C/EBPbeta was observed in the gastric tumors studied. Microarray analysis demonstrated a loss of TFF1 expression in all 27 GC cases examined, of which 25 exhibited high C/EBPbeta expression compared to normal gastric tissue. RT-PCR analysis revealed loss of TFF1 expression in 56 of 59 gastric tumors in which 54 of these tumors exhibited overexpression of C/EBPbeta. Immunohistochemical analysis revealed overexpression of C/EBPbeta in 10 of 13 gastric tumors that exhibited low expression of TFF1 at the protein level. Thus, overexpression of the transcription factor C/EBPbeta in the majority of GCs is a novel finding.