Human Cancer Biology EGFRvIII Deletion Mutations in Pediatric High-Grade Glioma and Response to Targeted Therapy in Pediatric Glioma Cell Lines

Purpose: The epidermal growth factor receptor (EGFR) is amplified and overexpressed in adult glioblastoma, with response to targeted inhibition dependent on the underlying biology of the disease. EGFR has thus far been considered to play a less important role in pediatric glioma, although extensive data are lacking. We have sought to clarify the role of EGFR in pediatric high-grade glioma (HGG). Experimental Design: We retrospectively studied a total of 90 archival pediatric HGG specimens for EGFR protein overexpression, gene amplification, and mutation and assessed the in vitro sensitivity of pediatric glioma cell line models to the small-molecule EGFR inhibitor erlotinib. Results: Amplification was detected in 11% of cases, with corresponding overexpression of the receptor. No kinase or extracellular domain mutations were observed; however, 6 of 35 (17%) cases harbored the EGFRvIII deletion, including two anaplastic oligodendrogliomas and a gliosarcoma overexpressing EGFRvIII in the absence of gene amplification and coexpressing platelet-derived growth factor receptor α. Pediatric glioblastoma cells transduced with wild-type or deletion mutant EGFRvIII were not rendered more sensitive to erlotinib despite expressing wild-type PTEN. Phosphorylated receptor tyrosine kinase profiling showed a specific activation of platelet-derived growth factor receptor α/β in EGFRvIII-transduced pediatric glioblastoma cells, and targeted coinhibition with erlotinib and imatinib leads to enhanced efficacy in this model. Conclusions: These data identify an elevated frequency of EGFR gene amplification and EGFRvIII mutation in pediatric HGG than previously recognized and show the likely necessity of targeting multiple genetic alterations in the tumors of these children. (Clin Cancer Res 2009;15(18):5753–61) Amplification, overexpression, and/or mutation of the epidermal growth factor receptor (EGFR) represent a compelling set of molecular genetic indicators for targeted therapy in adult glioblastoma. About 40% of glioblastomas show amplification of the EGFR gene locus, and about half of these tumors express a mutant receptor (EGFRvIII) that is constitutively active due to an in-frame truncation within the extracellular ligand-binding domain (1). In addition, novel missense mutations have been reported in the extracellular domain of tumors and cell lines (2), and recently, additional mutations have been described outside of these regions (e.g., in the transmembrane domains), although the significance of these is not yet clear (3). EGFRvIII is caused by deletion of exons 2 to 7, resulting in a protein that lacks a ligand-binding domain and is constitutively activated and is further resistant to down-regulation due to a low rate of receptor endocytosis (4). EGFRvIII has been shown Authors' Affiliations: Paediatric Oncology and Cancer Research UK Centre for Cancer Therapeutics, The Institute of Cancer Research; Paediatric Oncology, The Royal Marsden NHS Foundation Trust, Sutton, United Kingdom; Pharamacology and New Treatments of Cancer, Institut de Cancérologie, Gustave Roussy, Villejuif, France; Life and Health Science Research Institute, Universidade do Minho, Braga, Portugal; Breakthrough Breast Cancer, The Institute of Cancer Research; Neuropathology, Kings College Hospital, London, United Kingdom; Department of Pathology, Universidade Federal de Sao Paulo, Sao Paulo, Brazil; and Neuropathology, St Jude Children's Research Hospital, Memphis, Tennessee Received12/10/08; revised6/4/09; accepted6/17/09; publishedOnlineFirst 9/8/09. Grant support: Cancer Research UK grants C1178/A10294, C309/A2187, and C309/A8274;OakFoundation (L.Marshall); La FondationdeFrance (N.Gaspar); and Breakthrough Breast Cancer (J.S. Reis-Filho).We acknowledgeNHS funding to the National Institute for Health Research Biomedical Research Centre. P. Workman is a Cancer Research UK Life Fellow. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. Note: Supplementary data for this article are available at Clinical Cancer Research Online (http://clincancerres.aacrjournals.org/). D.A. Bax and N. Gaspar contributed equally to this work. Requests for reprints: Chris Jones, Paediatric Oncology, The Institute of Cancer Research, 15 Cotswold Road, Sutton, Surrey SM2 5NG, United Kingdom. Phone: 44-20-8722-4416; Fax: 44-20-8722-4321; E-mail. [email protected]. F 2009 American Association for Cancer Research. doi:10.1158/1078-0432.CCR-08-3210 5753 Clin Cancer Res 2009;15(18) September 15, 2009 www.aacrjournals.org Cancer Research. on January 13, 2018. © 2009 American Association for clincancerres.aacrjournals.org Downloaded from Published OnlineFirst September 8, 2009; DOI: 10.1158/1078-0432.CCR-08-3210