Overexpression of EPH receptor B2 in malignant mesothelioma correlates with oncogenic behavior

Our plain-English summary of Overexpression of EPH receptor B2 in malignant mesothelioma correlates with oncogenic behavior, by C. Goparaju, J.S. Donington, T. Hsu, et al (Journal of Thoracic Oncology, 2013).

The study

• Authors: C. Goparaju, J.S. Donington, T. Hsu, R. Harrington, N. Hirsch, H.I. Pass
• Journal: Journal of Thoracic Oncology
• Published: 2013;8(9):1203-11
• Cited by: 19 articles, as counted by NIH iCite
• Read it: PubMed · Publisher

Abstract

Introduction: Malignant pleural mesothelioma (MM) is an aggressive asbestos-associated malignancy with limited therapeutic options. This study describes the overexpression of Ephrin B2 receptor (EPHB2) in MM cell lines and tumors, and the effect of its manipulation on proliferative and invasive qualities of the disease.

Methods: Using expression arrays, we investigated EPHB2 in MM tumors compared with normal mesothelium. EPHB2 and downstream target expression were evaluated using reverse-transcriptase polymerase chain reaction and immunoblotting methods. The biological significance of EPHB2 in MM was evaluated using in vitro functional assays with and without targeting by EPHB2-short hairpin RNA or blocking peptide in two mesothelioma cell lines, HP-1 and H2595.

Results: EPHB2 is overexpressed in all MM cell lines, but not in benign mesothelial cells, and is significantly elevated in MM tumor tissue compared with matched normal peritoneum. Targeted knockdown of EPHB2 in HP-1 and H2595 cell lines reduced its expression and that of EPHB2 downstream targets such as matrix metalloproteinase (MMP-2) and vascular endothelial growth factor, whereas caspase 2 and caspase 8 had increased expression. Inhibition of EPHB2 resulted in a significant decrease in scratch closure (1.25-fold-1.8-fold), proliferation (1.5-fold), and invasion (1.7-fold-1.8-fold) compared with the controls. Most notably, however, EPHB2 silencing resulted in a significant increase in apoptotic proteins and activity.

Conclusion: EPHB2 seems to play an important role in MM pathogenesis and these findings indicate that EPHB2 could serve as a potential novel therapeutic target for treatment of the disease.

In plain English

This laboratory study examined a protein, EPHB2 (ephrin B2 receptor), in malignant mesothelioma. It did not study asbestos exposure. Using gene-expression arrays on 34 matched pairs of mesothelioma tumor and normal peritoneum, taken from patients having extrapleural pneumonectomy, the authors found EPHB2 levels 3.75-fold higher in the tumors (p = 2.39E-05). EPHB2 was also higher in mesothelioma cell lines than in benign mesothelial cells. In two mesothelioma cell lines, H2595 and HP-1, the authors lowered EPHB2 with a genetic tool (shRNA), which cut its mRNA by 80% to 90%. In separate experiments they blocked it with a peptide. Lowering EPHB2 cut cell growth 1.4-fold to 2.0-fold and slowed closing of a scratch by 29% in H2595 and 25% in HP-1. It cut invasion through a gel barrier 2.3- to 2.7-fold in H2595 and 1.5- to 1.7-fold in HP-1, reduced colony formation, and raised dying (apoptotic) cells by 30% to 40%. Benign LP9 cells showed no changes. The peptide also reduced growth and invasion. The authors concluded that inhibiting EPHB2 had profound in vitro effects and called for more laboratory work.

We wrote this plain-English summary from the full article with the help of AI, and a second AI model checked each sentence against the article. It is not the authors' wording; the abstract and the article are their own account.

What the study did and found

• Design: Laboratory study: gene-expression analysis of human tumor tissue plus in vitro (cell culture) experiments on mesothelioma cell lines
• Who was studied: 34 matched malignant mesothelioma tumors and normal peritoneum from patients undergoing extrapleural pneumonectomy; 7 of these pairs were used for confirmation. Also eight mesothelioma cell lines and benign mesothelial cell lines. Experiments used the mesothelioma lines H2595 and HP-1 and the benign line LP9. Dates of collection are not given.
• Exposure: No asbestos exposure was studied. The experimental interventions were reducing EPHB2 with shRNA plasmids (compared with a scrambled control) and, separately, blocking EPHB2 with a blocking peptide at 0.2 and 2 µg/ml.
• Main results:
  • In the 34 matched pairs, EPHB2 gene expression was 3.75-fold higher in tumors than in matched normal peritoneum (p = 2.39E-05).
  • EPHB2 mRNA levels were higher in mesothelioma cell lines than in benign mesothelial cells.
  • In H2595 and HP-1 cells, shRNA reduced EPHB2 mRNA by 80% to 90%. Western blot showed a 90% reduction in EPHB2 protein.
  • Reducing EPHB2 in H2595 and HP-1 cells significantly lowered mRNA levels of MMP-2, VEGF, BCL2 and BIRC5 (survivin).
  • Reducing EPHB2 significantly raised expression of caspase 1 and caspase 8.
  • Apoptotic (dying) cells rose by 30% to 40% in HP-1 and H2595 cells compared with scrambled control.
  • Reducing EPHB2 produced no changes in benign LP9 mesothelial cells.
  • Cell density fell 1.4-fold to 2.0-fold (p < 0.0015) in both H2595 and HP-1 cells with EPHB2-shRNA compared with scrambled controls.
  • Scratch closure (cell migration) was reduced by 29% (p < 0.0005) in H2595 cells and by 25% (p < 0.0005) in HP-1 cells.
  • Invasion through Matrigel fell 2.3-fold to 2.7-fold in H2595 cells and 1.5-fold to 1.7-fold in HP-1 cells (p < 2.00E-05 for both), compared with scrambled controls.
  • In soft agar, reducing EPHB2 significantly reduced the size and number of colonies in both cell lines at 21 days.
  • With 0.2 µg/ml of blocking peptide, cell proliferation fell 2.1-fold (p = 0.031) and 2.56-fold (p = 0.003). The article does not say which figure belongs to which cell line.
  • With 2 µg/ml of blocking peptide, cell proliferation fell 2.5-fold in H2595 cells (p = 0.0002) and 2.8-fold in HP-1 cells (p = 0.003).
  • With 2 µg/ml of blocking peptide, invasion fell 2.0-fold in H2595 cells (p = 0.0002) and 1.6-fold in HP-1 cells (p = 3.41E-05).
• The authors' conclusion: The authors concluded that EPHB2 is overexpressed in malignant mesothelioma and that inhibiting it has profound in vitro effects on invasion and on apoptosis control. They suggested targeting EPHB2 might provide a novel therapy.
• Limitations the authors state:
  • The authors stated that further in vitro work with specific EPHB2 inhibitors is required to determine EPHB2's importance and its role as a marker of progression and metastasis.

Topics

The National Library of Medicine indexes this study under subjects our site covers: Lung cancer, Mesothelioma.

Medical terms in this study, explained in our Lexicon: Malignant Mesothelioma, Pleural Mesothelioma, Mesothelium, Mesothelioma, Mesothelial Cells, Peritoneum.

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