Characterization of Epithelial Cell Adhesion Molecule as a Surface Marker on Undifferentiated Human Embryonic Stem Cells  

Abstract

Human embryonic stem cells (hESCs) have the capacity to remain pluripotent and self-renew indefinitely. To discover novel players in the maintenance of hESCs, we have previously reported the generation of monoclonal antibodies that bind to cell surface markers on hESCs, and not to mouse embryonic stem cells or differentiated embryoid bodies. In this study, we have identified the antigen target of one such monoclonal antibody as the epithelial cell adhesion molecule (EpCAM). In undifferentiated hESCs, EpCAM is localized to Octamer 4 (OCT4)-positive pluripotent cells, and its expression is down-regulated upon differentiation. To further understand its biological function in hESCs, endogenous EpCAM expression was silenced using small interfering RNA. EpCAM knockdown had marginal negative effects on OCT4 and TRA-1-60 expression, however cell proliferation was decreased by >40%.Examination of lineage marker expression showed marked upregulation of endoderm and mesoderm genes in EpCAM-silenced cells, under both pluripotent and differentiating conditions. These results were validated using a hESC line whose EpCAM expression has been stably knocked down. Data from the stable line confirmed that downregulation of EpCAM decreases cell growth and increases gene expression in the endoderm and mesoderm lineages. In vivo, hESCs lacking EpCAM were able to form teratomas containing tissues representing the three germ layers, and gene expression analysis yielded marked increase in the endoderm marker alpha fetoprotein compared with control. Together these data demonstrate that EpCAM is a surface marker on undifferentiated hESCs and plays functional roles in proliferation and differentiation. STEM CELLS 2010;28:29-35 Disclosure of potential conflicts of interest is found at the end of this article.

Cleavage and Cell Adhesion Properties of Human Epithelial Cell Adhesion Molecule (HEPCAM)

“…Based on the availability of the large extracellular domain for therapeutic antibodies, HEPCAM is a long-known target for cancer therapies (48 -50). More recently, Epcam was recognized as a marker for pluripotent stem cells in mice and humans (51,52) and for tissue stem cells (53,54). Frequent overexpression of HEPCAM was explained by its ability to foster proliferation and oncogenic behavior (14,23), which is based on regulated intramembrane proteolysis and formation of a signaling intracellular domain termed hEpICD (29).…”

“…It is conceivable that HEPCAM becomes actively retrieved from membranes of tumor cells via endocytosis and BACE1/proteasome action to warrant or even induce a mesenchymal shift in phenotype. Comparable erasure of Epcam was observed during differentiation processes of embryonic stem cells (51,52) and might be the result of transcriptional polycomb silencing (55) as well as post-translational degradation.…”

Mapping the first stages of mesoderm commitment during differentiation of human embryonic stem cells

“…Similar to many other epithelial systems, hESCs usually establish apical-basal polarity in relation to the feeder layers on which they are cocultured (10,11), and associate with each other through adhesion molecules such as E-cadherin and EpCAM (CD326) (11,12), which have also been shown to be required for maintenance of pluripotency (13,14). We hypothesized that the earliest stage of mesoderm commitment from pluripotent hESCs would be represented by a coordinated up-regulation of CD56 expression during the loss of epithelial cell adhesion.…”

EpCAM, a novel oncogenic receptor and its target therapy

“…EpCAM SiRNA treatment decreased cell migration by 91.8% and cell invasion by 96.4% in breast cancer cell line MDA-MB-231 in vitro, which associated with an increase in E-cadherin, beta-catenin, and especially alpha-catenin gene transcription [71].The results implicate that EpCAM expression can affect cell migration, invasion, and proliferation by enhancing E-cadherin-mediated cell-to-cell adhesion [71]. Recently, EpCAM has been identified as an additional marker for cancer-initiating stem cells [77][78][79][80].…”

“…Similarily, in vivo evaluation of tumorigenicity in hepatocellular carcinoma cell lines, using immunodeficient NOG mice, a smaller number of EpCAM+ cells (minimum 100) than EpCAM-cells are able to tumor formation. The introduction of exogenous EpCAM into EpCAM+ clones,but not into EpCAMclones, markedly enhanced their tumor-forming ability [79]. Also, EpCAM-positive hepatocellular carcinoma stem cells could efficiently initiate tumours in SCID mice [80].…”