Revisiting the Roles of Pro-Metastatic EpCAM in Cancer

Abstract

Epithelial cell adhesion molecule (EpCAM) is a cell surface protein that was discovered as a tumour marker of epithelial origins nearly four decades ago. EpCAM is expressed at basal levels in the basolateral membrane of normal epithelial cells. However, EpCAM expression is upregulated in solid epithelial cancers and stem cells. EpCAM can also be found in disseminated tumour cells and circulating tumour cells. Various OMICs studies have demonstrated that EpCAM plays roles in several key biological processes such as cell adhesion, migration, proliferation and differentiation. Additionally, EpCAM can be detected in the bodily fluid of cancer patients suggesting that EpCAM is a pathophysiologically relevant anti-tumour target as well as being utilized as a diagnostic/prognostic agent for a variety of cancers. This review will focus on the structure-features of EpCAM protein and discuss recent evidence on the pathological and physiological roles of EpCAM in modulating cell adhesion and signalling pathways in cancers as well as deliberating the clinical implication of EpCAM as a therapeutic target.

Keywords: adhesion; biomarker; cancer; circulating tumour cell; metastasis.

Figure 1

Epithelial cell adhesion molecule (EpCAM) protein structure and splice variant expression in cancer. (A) The secondary structure of EpCAM which consists of signal peptide (SP, blue), N-domain (ND, pink), Thyroglobulin type-1 domain (TY, lime green), C-domain (CD, grey), transmembrane domain (TM, grey) and intracellular part (EpIC, white). Three-dimensional illustration and surface representation of the EpCAM cleaved extra-cellular domain (EpEX) (PDB code: 4MZV) color-coded as in the secondary structure. (B) Schematic of EpCAM gene structure and the splice variants extracted from Ensembl database (http://www.ensembl.org). The predominant isoform, EpCAM-201, consists of 9 exons. Isoforms color-coded in green are those encode for EpCAM protein. (C) Bar-plot shows the commonly expressed EPCAM isoforms (from 0% to 100%) across the TCGA-Pan-cancer analysis.

Figure 2

Post-translational modifications of EpCAM. (A) EpCAM harbours six disulphide bridges that cluster around the N-domain (ND) and Thyroglobulin type-1 domain (TY). Proteolytic cleavages can take place at two locations. i.e., after Ala23 or Ala21 by a signal peptidase; and in the TY-repeat, between Arg80 and Arg81. Additionally, EpCAM undergoes a two-step regulated intramembrane proteolysis (RIP), whereby EpCAM is first cleaved close to the plasma membrane at the EpEX1 site, leading to the shedding of the EpEX domain; followed by a second cleavage at the EpICD site. In another proposed RIP pathway, the first cuts are made to the ectodomain via consecutive cleavages at two sites (EpEX2), followed by the EpICD cleavage. (B) The presence of three N-glycosylation sites located on Asn74, Asn111 and Asn198 of EpCAM have been confirmed by numerous reports. Other post-translational modifications that have been discovered include: O-linked glycosylations at Thr171 and Thr172; phosphorylations at Tyr214, Tyr215 and Tyr297; ubiquitylations at Lys168, Lys 218, Lys 299 and Lys303; and acetylation at Lys179.

Figure 3

EpCAM mRNA expression in normal and cancer tissues and survival analysis. (A) Dot plots representing EpCAM expression, determined by RNA-Sequencing platform, in tumour and normal tissue samples from the TCGA and GTEx databases, respectively. The data were analysed and visualized using GEPIA2 web tool [65]. Cancer types that are labelled red have upregulated EpCAM expression compared to the normal tissues whereas cancer types labelled green are those that have lower EpCAM expression than their normal counterparts. (B) Heatmap showing the correlation between EpCAM gene isoforms expression and patients’ survival across cancer types from TCGA databases. The data were analysed and visualized using GEPIA2 web tool as in Figure 2A. The hazard ratios are in the logarithmic scale (log10); the red and blue blocks denote higher and lower hazard ratios, respectively. Blocks with highlighted border indicate statistically significant correlations.

Figure 4

EpCAM localization and modulation of its downstream targets in cancer. The membrane bound EpCAM undergoes regulated intermembrane proteolysis that leads to the secretion of solubilised extracellular domain, EpEX and the liberation of cytoplasmic intracellular domain, EpICD. The cytoplasmic EpICD forms transcriptional complex with WNT signalling downstream effector to activate genes that are associated with cells proliferation and stemness maintenance. The solubilized EpEX acts as ligand to stimulate the oncogenic signalling pathways like the PI3K/AKT/mTOR via its binding to receptor tyrosine kinase and activate the expression of genes involve in supporting cancer cells growth and fitness. The membrane bound EpCAM can also function in promoting cell adhesion as well as being expressed on the exosome surface to mediate cell-cell communication.