Beta-platelet-derived growth factor receptor mediates motility and growth of Ewing's sarcoma cells.

A Üren,C J Sun,C Mackall,M Tsokos,P B Illei,A Passaniti,J A Toretsky,M S Merchant,Y Sun,M I Vitolo,M Ladanyi

doi:10.1038/sj.onc.1206330

Abstract

The Ewing's sarcoma family of tumors (ESFT) contain a translocation, t(11;22), which results in the novel oncogenic fusion protein EWS/FLI1. Platelet-derived growth factors (PDGF) and their receptors (PDGFR) are involved in the induction and proliferation of numerous solid tumors and are the potential candidates for novel targeted antitumor therapy. Since a relation was reported between PDGF-C and EWS/FLI1, we sought to characterize the PDGF signaling pathway in ESFT. Eight out of nine ESFT cell lines were found to express significant levels of beta-PDGFR. Interestingly, none of the tested cell lines expressed alpha-PDGFR, which is the receptor isotype required for PDGF-C binding. By immunohistochemical staining 47 of 52 (90.4%) archival tumor samples from patients with ESFT were positive for beta-PDGFR. ESFT cell lines were treated with PDGF-AA or PDGF-BB ligands to evaluate downstream signaling. Autophosphorylation of beta-PDGFR and tyrosine phosphorylation of PLC-gamma, PI3Kp85 and Shc were detected only in PDGF-BB-stimulated cells that express beta-PDGFR. Receptor function was further evaluated using chemotaxis assays that showed TC-32 cell migration towards PDGF-BB. A specific PDGFR kinase inhibitor AG1295 blocked beta-PDGFR activation, downstream signaling, growth in cell culture and chemotaxis of TC-32 cells. AG1295 also delayed tumor formation and prolonged survival in an ESFT animal model. We conclude that ESFT express beta-PDGFR and that this is a functional and potentially crucial signaling pathway. Therefore, beta-PDGFRs may provide a novel therapeutic target in ESFT that can be utilized to design better treatment modalities.

Highlights

The Ewing’s sarcoma family of tumors (ESFT) affect patients between ages of 3 and 40 years, with most cases occurring in the second decade of life (Grier, 1997; Denny, 1998)
Our demonstration of functional b-PDGF receptors (PDGFR) in ESFT cell lines as well as expression of this receptor in ESFT tumor specimens suggest a role for this signaling pathway in the ESFT phenotype. b-PDGFR in ESFT may contribute towards tumor proliferation and metastasis
Platelet-derived growth factors (PDGF)-B or PDGF-D, which are known to stimulate b-PDGFR do not appear to be secreted from the tumor cells, since conditioned media collected from ESFT cell lines did not activate b-PDGFR

Summary

Introduction

The Ewing’s sarcoma family of tumors (ESFT) affect patients between ages of 3 and 40 years, with most cases occurring in the second decade of life (Grier, 1997; Denny, 1998). ESFT contain a characteristic chromosomal translocation, t(11;22), which is present in 90– 95% of the tumors. The resulting fusion protein, EWS/ FLI1, contains the amino-terminal of EWS and the carboxy-terminal of FLI1. Patients presenting with localized disease have a significantly better survival rate (in the range 60–70%) than patients with metastatic disease (Miser et al, 1996). Treatment of metastatic tumors has been unsuccessful despite new high-dose chemotherapeutic regimens with stem cell rescue (Meyers et al, 2001). There is a need to identify and validate novel therapeutic targets that could interfere with Ewing’s sarcoma growth or metastasis

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Conclusion