Silencing the receptor EphA2 suppresses the growth and haptotaxis of malignant mesothelioma cells

Abstract

BACKGROUND.The over‐expression of the ephrin‐A1 ligand receptor EphA2 is associated with the growth and metastatic potential of tumors. Although EphA2 is expressed in a variety of tumors, its expression and function in malignant mesothelioma (MM) remain unknown. The authors hypothesized that expression of the receptor EphA2 in MM cells (MMCs) plays a key role in the growth and haptotactic migration of MM. They also hypothesized that silencing EphA2 expression by using small‐interfering RNA (siRNA) inhibits the proliferation and haptotaxis of MMCs and induces apoptosis in MMCs.METHODS.The expression of EphA2 in MMCs and in normal pleural mesothelial cells (PMCs) was studied by using real‐time quantitative polymerase chain reaction analysis and Western blot analysis. The growth of MMCs was determined with the WST‐1 cell‐proliferation assay. The haptotactic migration of MMCs and PMCs was determined with a Boyden chamber assay. Expression of caspases was determined with calorimetric assays.RESULTS.The results demonstrated that silencing the receptor EphA2 by siRNA significantly reduced the proliferation and haptotactic migration of MMCs compared with controls. Over‐expression of EphA2 with plasmid pcDNA/EphA2 enhanced the proliferation and haptotaxis of MMCs significantly. Knocking down EphA2 expression initiated caspase‐9–mediated apoptosis in MMCs.CONCLUSIONS.The current results suggested that constitutive expression of EphA2 may contribute to the aggressive behavior and cellular survival of MMCs. EphA2 may be an effective therapeutic target in patients with mesothelioma. Silencing the receptor EphA2 gene is a novel approach for the containment of growth and migration of tumor in patients with malignant mesothelioma. Cancer 2006. © 2006 American Cancer Society.

EphA2 Induces Metastatic Growth Regulating Amoeboid Motility and Clonogenic Potential in Prostate Carcinoma Cells

“…The ability of EphA2 to induce tumorigenesis and metastatic dissemination has been correlated with the increased skill of EphA2-overexpressing cells to grow in soft agar, to invade into Matrigel (7) and to increase the resistance to anoikis (9,12). Among the previously mentioned characteristics concurring to determine a more aggressive phenotype, we would like to introduce 2 new features: the acquisition of an amoeboid motility style due to EphA2 overexpression and the increase in clonogenic potential.…”

“…In some cellular models, EphA2 expression represents a way for cells to escape from anoikis as shown in pancreatic adenocarcinoma cells (9), or to avoid caspase-9-mediated apoptosis as evidentiated in malignant mesothelioma cells (12). To test whether in PC3 cells the expression of EphA2 could confer resistance to anoikis, we analyzed cell survival to 24 hours and 72 hours suspension by flow cytometry analysis.…”

“…Furthermore, overexpression of dominant-negative EphA2 mutant or a kinase inactive form results in decreased tumor volume and increased tumor apoptosis (10,11). Recently, Nasreen and colleagues (12) showed that silencing EphA2 expression by using short interfering RNA (siRNA) inhibits the proliferation and haptotaxis of malignant mesothelioma cells. In agreement, Landen and colleagues (13) showed that the therapeutic delivery of EphA2 siRNA into an orthotopic mouse model of ovarian cancer reduced tumor growth when compared with a nonsilencing siRNA.…”

EphrinA1 inhibits malignant mesothelioma tumor growth via let-7 microRNA-mediated repression of the RAS oncogene

“…MMCs were counted and subcultured in 60-mm cell culture dishes (5 Â 10 5 cells per dish) as reported earlier. 23 …”

“…MMC proliferation MMC proliferation was measured by a colorimetric assay kit (WST-1; Roche Diagnostics, Indianapolis, IN), as reported earlier. 23 In brief, MMCs were plated in 96-well microplate, at a density of 1 Â 10 4 cells per well.…”