Epithelial E- and P-cadherins: Role and clinical significance in cancer

E-cadherin and P-cadherin are major contributors to cell-cell adhesion in epithelial tissues, playing pivotal roles in important morphogenetic and differentiation processes during development, and in maintaining integrity and homeostasis in adult tissues. It is now generally accepted that alterations in these two molecules are observed during tumour progression of most carcinomas. Genetic or epigenetic alterations in E- and P-cadherin-encoding genes (CDH1 and CDH3, respectively), or alterations in their proteins expression, often result in tissue disorder, cellular de-differentiation, increased invasiveness of tumour cells and ultimately in metastasis. In this review, we will discuss the major properties of E- and P-cadherin molecules, its regulation in normal tissue, and their alterations and role in cancer, with a specific focus on gastric and breast cancer models.     

N-Cadherin Promotes Adhesion Between Invasive Breast Cancer Cells and the Stroma

Calcium-dependent cell adhesion molecules (cadherins) are involved in maintaining the epithelial structure of a number of tissues including the mammary gland. In breast and other tumor types, loss of E-cadherin expression has been seen in high grade tumors and correlates with increased invasiveness. Here we show high levels of expression of N-cadherin in the most invasive breast cancer cell lines which was inversely correlated with their expression of E-cadherin. A stromal cell line also expressed N-cadherin in accordance with its fibroblastic morphology. N-cadherin localized to areas of cell-cell contact in all cells that expressed it. Calcium-dependent intercellular adhesion of N-cadherin-expressing breast cancer and stromal cells was specifically inhibited by an anti N-cadherin monoclonal antibody. In addition, N-cadherin promoted the interaction of invasive breast cancer cells with mammary stromal cells: in contrast, E-cadherin expressing cell lines did not co-aggregate with stromal cells. The combined results suggest a functional role for N-cadherin in cohesion of breast tumor cells which, in addition promotes their interaction with the surrounding stromal cells, thereby facilitating invasion and metastasis.     

Molecular Plasticity of E-Cadherin and Sialyl Lewis X Expression,in Two Comparative Models of Mammary Tumorigenesis

Background

The process of metastasis involves a series of steps and interactions between the tumor embolus and the microenvironment. Key alterations in adhesion molecules are known to dictate progression from the invasive to malignant phenotype followed by colonization at a distant site. The invasive phenotype results from the loss of expression of the E-cadherin adhesion molecule, whereas the malignant phenotype is associated with an increased expression of the carbohydrate ligand-binding epitopes, (e.g. Sialyl Lewis ^x/a) that bind endothelial E-selectin of the lymphatics and vasculature.

Methodology

Our study analyzed the expression of two adhesion molecules, E-cadherin and Sialyl Lewis x (sLe^x), in both a canine mammary carcinoma and human inflammatory breast cancer (IBC) model, using double labelled immunofluorescence staining.

Results

Our results demonstrate that canine mammary carcinoma and human IBC exhibit an inversely correlated cellular expression of E-cadherin and sLe^x within the same tumor embolus.

Conclusions

Our results in these two comparative models (canine and human) suggest the existence of a biologically coordinated mechanism of E-cadherin and sLe^x expression (i.e. molecular plasticity) essential for tumor establishment and metastatic progression.     

N-Cadherin Promotes Adhesion Between Invasive Breast Cancer Cells and the Stroma

Calcium-dependent cell adhesion molecules (cadherins) are involved in maintaining the epithelial structure of a number of tissues including the mammary gland. In breast and other tumor types, loss of E-cadherin expression has been seen in high grade tumors and correlates with increased invasiveness. Here we show high levels of expression of N-cadherin in the most invasive breast cancer cell lines which was inversely correlated with their expression of E-cadherin. A stromal cell line also expressed N-cadherin in accordance with its fibroblastic morphology. N-cadherin localized to areas of cell-cell contact in all cells that expressed it. Calcium-dependent intercellular adhesion of N-cadherin-expressing breast cancer and stromal cells was specifically inhibited by an anti N-cadherin monoclonal antibody. In addition, N-cadherin promoted the interaction of invasive breast cancer cells with mammary stromal cells: in contrast, E-cadherin expressing cell lines did not co-aggregate with stromal cells. The combined results suggest a functional role for N-cadherin in cohesion of breast tumor cells which, in addition promotes their interaction with the surrounding stromal cells, thereby facilitating invasion and metastasis.     

Numb

Epithelial to mesenchymal transition (EMT) is a critical event in embryogenesis and plays a fundamental role in cancer progression and metastasis1. Numb has been shown to play an important role in the proper functions of Par protein complex and in cell-cell junctions, both of which are associated with EMT. However, the role of Numb in EMT has not been fully elucidated. Recently, we showed that Numb is capable of binding to both Par3 and E-cadherin. Intriguingly, the interaction of Numb with E-cadherin or the Par protein complex is dynamically regulated by tyrosine phosphorylation induced by HGF or Src. Knockdown of Numb by shRNA in MDCK cells led to an lateral to apical translocation of E-cadherin and β-catenin, active F-actin polymerization, mis-localization of Par3 and aPKC, a decrease in cell-cell adhesion, and an increase in cell migration and proliferation. These data suggest a diverse role for Numb in regulating cell-cell adhesion, polarity and migration during EMT.     

The role of Snail in prostate cancer

Prostate cancer is the second most frequently diagnosed cancer and the sixth leading cause of death from cancer in men. Epithelial-mesenchymal transition (EMT) is a process by which cancer cells invade and migrate, and is characterized by loss of cell-cell adhesion molecules such as E-cadherin and increased expression of mesenchymal proteins such as vimentin; EMT is also associated with resistance to therapy. Snail, a master regulator of EMT, has been extensively studied and reported in cancers such as breast and colon; however, its role in prostate cancer is not as widely reported. The purpose of this review is to put together recent facts that summarize Snail signaling in human prostate cancer. Snail is overexpressed in prostate cancer and its expression and activity is controlled via phosphorylation and growth factor signaling. Snail is involved in its canonical role of inducing EMT in prostate cancer cells; however, it plays a role in non-canonical pathways that do not involve EMT such regulation of bone turnover and neuroendocrine differentiation. Thus, studies indicate that Snail signaling contributes to prostate cancer progression and metastasis and therapeutic targeting of Snail in prostate cancer holds promise in �future.     

The role of Snail in prostate cancer

Prostate cancer is the second most frequently diagnosed cancer and the sixth leading cause of death from cancer in men. Epithelial-mesenchymal transition (EMT) is a process by which cancer cells invade and migrate, and is characterized by loss of cell-cell adhesion molecules such as E-cadherin and increased expression of mesenchymal proteins such as vimentin; EMT is also associated with resistance to therapy. Snail, a master regulator of EMT, has been extensively studied and reported in cancers such as breast and colon; however, its role in prostate cancer is not as widely reported. The purpose of this review is to put together recent facts that summarize Snail signaling in human prostate cancer. Snail is overexpressed in prostate cancer and its expression and activity is controlled via phosphorylation and growth factor signaling. Snail is involved in its canonical role of inducing EMT in prostate cancer cells; however, it plays a role in non-canonical pathways that do not involve EMT such regulation of bone turnover and neuroendocrine differentiation. Thus, studies indicate that Snail signaling contributes to prostate cancer progression and metastasis and therapeutic targeting of Snail in prostate cancer holds promise in ?future.     

The expression of core fucosylated E-cadherin in cancer cells and lung cancer patients： prognostic implications

It is well documented that the glycosylation of E-cadherin is correlated with cancer metastasis, but whether E-cadherin could be core fucosylated remains largely unknown. We found that E-cadherin was core fucosylated in highly metastatic lung cancer cells while absent in lowly metastatic lung cancer cells. Sinceα-1,6 Fucosyltransferase (α-1,6 FucT) is known to catalyze the reaction of core fucosylation, we investigated the biological function of core fucosylation on E-cadherin by α-1,6 FucT targeted RNAi and transfecting α-1,6 FucT expression vector. As a result, calcium dependent cell-cell adhesion mediated by E-cadherin was strengthened with the reduction of core fucosylation on E-cadherin after RNAi and was weakened with the elevated core fucosylation on E-cadherin after α-1,6 FucT over expression. Our data indicated that α-1,6 FucT could regulate E-cadherin mediated cell adhesion and thus play an important role in cancer development and progression. Computermodeling showed that core fucosylation on E-cadherin could significantly impair three-dimensional conformation of N-glycan on E-cadherin and produce conformational asymmetry so as to suppress the function of E-cadherin. Furthermore, the relationship between the expression of core fucosylated E-cadherin and clinicopathological background of lung cancer patients was explored in lung cancer tissue of patients. It turns out to demonstrate that core fucosylated E-cadherin could serve as a promising prognostic indicator for lung cancer patients.     

p120ctn and P-Cadherin but Not E-Cadherin Regulate Cell Motility and Invasion of DU145 Prostate Cancer Cells

Background

Adherens junctions consist of transmembrane cadherins, which interact intracellularly with p120ctn, ß-catenin and α-catenin. p120ctn is known to regulate cell-cell adhesion by increasing cadherin stability, but the effects of other adherens junction components on cell-cell adhesion have not been compared with that of p120ctn.

Methodology/Principal Findings

We show that depletion of p120ctn by small interfering RNA (siRNA) in DU145 prostate cancer and MCF10A breast epithelial cells reduces the expression levels of the adherens junction proteins, E-cadherin, P-cadherin, ß-catenin and α-catenin, and induces loss of cell-cell adhesion. p120ctn-depleted cells also have increased migration speed and invasion, which correlates with increased Rap1 but not Rac1 or RhoA activity. Downregulation of P-cadherin, β-catenin and α-catenin but not E-cadherin induces a loss of cell-cell adhesion, increased migration and enhanced invasion similar to p120ctn depletion. However, only p120ctn depletion leads to a decrease in the levels of other adherens junction proteins.

Conclusions/Significance

Our data indicate that P-cadherin but not E-cadherin is important for maintaining adherens junctions in DU145 and MCF10A cells, and that depletion of any of the cadherin-associated proteins, p120ctn, ß-catenin or α-catenin, is sufficient to disrupt adherens junctions in DU145 cells and increase migration and cancer cell invasion.     

Co-localization of Racl and E-Cadherin in Human Epidermal Keratinocytes

The Racl small GTP-binding protein is known to be involved in reorganization of the actin cytoskeleton and in regulation of intracellular signal transduction. The assembly and maintenance of cadherin-based cell-cell junctions in epidermal keratinocytes is thought to be dependent on activity of Racl. In this study we have generated green fluorescent protein (GFP)-tagged wild type, dominant negative and constitutively active Racl expression vectors and analyzed distribution of Racl following microinjection of human SCC12F epidermal keratinocytes. Wild type, dominant negative and constitutively active GFP-Racl proteins distribute to sites of cell-cell adhesion and co-localize with E-cadherin and the catenins. Disruption of cadherin-based junctions by reduction in extracellular calcium concentrations, or by use of antibodies to E-cadherin, results in redistribution of Racl away from sites of cell-cell interaction but the co-localization with E-cadherin is maintained. In addition, expression of constitutively active GFP-Racl results in formation of membrane ruffles on the apical surface of cells and intracellular vesicles. Interestingly, co-localization of Racl with E-cadherin is maintained in these structures. In contrast to previously published work we find that expression of dominant negative Racl neither disrupts cell-cell adhesion nor prevents assembly of new cadherin-based adhesion structures.     

Flotillins Directly Interact with γ-Catenin and Regulate Epithelial Cell-Cell Adhesion

Flotillin-1 and flotillin-2 are two homologous, membrane raft associated proteins. Although it has been reported that flotillins are involved in cell adhesion processes and play a role during breast cancer progression, thus making them interesting future therapeutic targets, their precise function has not been well elucidated. The present study investigates the function of these proteins in cell-cell adhesion in non-malignant cells. We have used the non-malignant epithelial MCF10A cells to study the interaction network of flotillins within cell-cell adhesion complexes. RNA interference was used to examine the effect of flotillins on the structure of adherens junctions and on the association of core proteins, such as E-cadherin, with membrane rafts. We here show that the cadherin proteins of the adherens junction associate with flotillin-2 in MCF10A cells and in various human cell lines. In vitro, flotillin-1 and flotillin-2 directly interact with γ-catenin which is so far the only protein known to be present both in the adherens junction and the desmosome. Mapping of the interaction domain within the γ-catenin sequence identified the Armadillo domains 6–8, especially ARM domain 7, to be important for the association with flotillins. Furthermore, depletion of flotillins significantly influenced the morphology of the adherens junction in human epithelial MCF10A cells and altered the association of E-cadherin and γ-catenin with membrane rafts. Taken together, these observations suggest a functional role for flotillins, especially flotillin-2, in cell-cell adhesion in non-malignant epithelial cells.     

Expression of N-cadherin by human squamous carcinoma cells induces a scattered fibroblastic phenotype with disrupted cell-cell adhesion

E-cadherin is a transmembrane glycoprotein that mediates calcium- dependent, homotypic cell-cell adhesion and plays an important role in maintaining the normal phenotype of epithelial cells. Disruption of E- cadherin activity in epithelial cells correlates with formation of metastatic tumors. Decreased adhesive function may be implemented in a number of ways including: (a) decreased expression of E-cadherin; (b) mutations in the gene encoding E-cadherin; or (c) mutations in the genes that encode the catenins, proteins that link the cadherins to the cytoskeleton and are essential for cadherin mediated cell-cell adhesion. In this study, we explored the possibility that inappropriate expression of a nonepithelial cadherin by an epithelial cell might also result in disruption of cell-cell adhesion. We showed that a squamous cell carcinoma-derived cell line expressed N-cadherin and displayed a scattered fibroblastic phenotype along with decreased expression of E- and P-cadherin. Transfection of this cell line with antisense N- cadherin resulted in reversion to a normal-appearing squamous epithelial cell with increased E- and P-cadherin expression. In addition, transfection of a normal-appearing squamous epithelial cell line with N-cadherin resulted in downregulation of both E- and P- cadherin and a scattered fibroblastic phenotype. In all cases, the levels of expression of N-cadherin and E-cadherin were inversely related to one another. In addition, we showed that some squamous cell carcinomas expressed N-cadherin in situ and those tumors expressing N- cadherin were invasive. These studies led us to propose a novel mechanism for tumorigenesis in squamous epithelial cells; i.e., inadvertent expression of a nonepithelial cadherin.     

Formation of E-cadherin-mediated cell-cell adhesion activates AKT and mitogen activated protein kinase via phosphatidylinositol 3 kinase and ligand-independent activation of epidermal growth factor receptor in ovarian cancer cells

E-cadherin is a well characterized adhesion molecule that plays a major role in epithelial cell adhesion. Based on findings that expression of E-cadherin is frequently lost in human epithelial cancers, it has been implicated as a tumor suppressor in carcinogenesis of most human epithelial cancers. However, in ovarian cancer development, our data from the current study showed that E-cadherin expression is uniquely elevated in 86.5% of benign, borderline, and malignant ovarian carcinomas irrespective of the degree of differentiation, whereas normal ovarian samples do not express E-cadherin. Thus, we hypothesize that E-cadherin may play a distinct role in the development of ovarian epithelial cancers. Using an E-cadherin-expressing ovarian cancer cell line OVCAR-3, we have demonstrated for the first time that the establishment of E-cadherin mediated cell-cell adhesions leads to the activation of Akt and MAPK. Akt activation is mediated through the activation of phosphatidylinositol 3 kinase, and both Akt and MAPK activation are mediated by an E-cadherin adhesion-induced ligand-independent activation of epidermal growth factor receptor. We have also demonstrated that suppression of E-cadherin function leads to retarded cell proliferation and reduced viability. We therefore suggest that the concurrent formation of E-cadherin adhesion and activation of downstream proliferation signals may enhance the proliferation and survival of ovarian cancer cells. Our data partly explain why E-cadherin is always expressed during ovarian tumor development and progression.     

Differential expression of E-cadherin gene in human neuroepithelial tumors

Cadherins are cell-to-cell adhesion molecules that play an important role in the establishment of adherent-type junctions by mediating calcium-dependent cellular interactions. The CDH1 gene encodes the transmembrane glycoprotein E-cadherin which is important in maintaining homophilic cell-cell adhesion in epithelial tissues. E-cadherin interacts with catenin proteins to maintain tissue architecture. Structural defects or loss of expression of E-cadherin have been reported as a common feature in several human cancer types. This study aimed to evaluate the expression of E-cadherin and their correlation with clinical features in microdissected brain tumor samples from 81 patients, divided into 62 astrocytic tumors grades I to IV and 19 medulloblastomas, and from 5 white matter non-neoplasic brain tissue samples. E-cadherin (CDH1) gene expression was analyzed by quantitative real-time polymerase chain reaction. Mann-Whitney, Kruskal-Wallis, Kaplan-Meir, and log-rank tests were performed for statistical analyses. We observed a decrease in expression among pathological grades of neuroepithelial tumors. Non-neoplasic brain tissue showed a higher expression level of CDH1 gene than did neuroepithelial tumors. Expression of E-cadherin gene was higher in astrocytic than embryonal tumors (P = 0.0168). Low-grade malignancy astrocytomas (grades I-II) showed higher CDH1 expression than did high-grade malignancy astrocytomas (grades III-IV) and medulloblastomas (P < 0.0001). Non-neoplasic brain tissue showed a higher expression level of CDH1 gene than grade I malignancy astrocytomas, considered as benign tumors (P = 0.0473). These results suggest that a decrease in E-cadherin gene expression level in high-grade neuroepithelial tumors may be a hallmark of malignancy in dedifferentiated tumors and that it may be possibly correlated with their progression and dissemination.     

Uncoupling of the ERα regulated morphological phenotype from the cancer stem cell phenotype in human breast cancer cell lines

The CD24^low/−CD44⁺EpCAM⁺ phenotype is associated with breast cancer initiating cells. To investigate if these putative breast cancer stem cell markers are regulated by estrogen receptor alpha (ERα) we have determined the expression levels of EpCAM, CD44 and CD24 in several well characterized breast cancer cell lines. The expression levels of the three adhesion proteins were quantitatively different in the cell lines but the composite CD24^low/−CD44⁺EpCAM⁺ breast cancer stem cell phenotype was shown to exist as a small fraction, between 0.1% and 1.2%, in all breast cancer cell lines tested. Experimental silencing of ERα resulted in a reduced epithelial appearance and partial reduction of CD24 mRNA, while levels of CD44 and EpCAM were unaltered. Moreover, knockdown of ERα led to a change in the morphology of the cells similar to the epithelial to mesenchymal transition phenotype and was associated with decreased E-cadherin expression. Our findings offer new insights into the regulation of the breast cancer stem cell phenotype by ERα and suggest that treatments targeting the breast cancer stem cell adhesion molecules and the ERα pathway may be complementary.     

E-cadherin downregulation in cancer: fuel on the fire?

The development, maintenance and repair of tissue requires an exquisite balance between cell proliferation, cell adhesion and cell motility. Equally, tumour initiation and progression are characterized by not only the abnormal expression of genes involved in cell proliferation and survival but also by genes responsible for the control of cell adhesion and cell motility. Central to the process of cell-cell adhesion in epithelial tissues is E-cadherin. Loss of E-cadherin function in tumours results in the rapid progression of relatively benign adenomas to invasive, metastatic carcinomas. Germline mutation of the E-cadherin gene predisposes to diffuse, poorly differentiated gastric cancer, and its downregulation in sporadic tumours is associated with poor clinical prognosis.