High-Grade Glioma Formation Results from Postnatal Pten Loss or Mutant Epidermal Growth Factor Receptor Expression in a Transgenic Mouse Glioma Model Qingxia Wei, 1 Laura Clarke, 1 Danielle K. Scheidenhelm, 4 Baoping Qian, 1 Amanda Tong, 1 Nesrin Sabha, 1 Zia Karim, 1 Nicholas A. Bock, 2 Robert Reti, 1 Rolf Swoboda, 5 Enkhtsetseg Purev, 5 Jean-Francois Lavoie, 1 M. Livia Bajenaru, 4 Patrick Shannon, 3 Dorothee Herlyn, 5 David Kaplan, 1 R. Mark Henkelman, 2 David H. Gutmann, 4 and Abhijit Guha 1,4 1 Arthur & Sonia Labatt Brain Tumour Research Centre and 2 Mouse Imaging Centre, The Hospital for Sick Children; 3 Division of Neuropathology and Neurosurgery, University Health Network, Toronto, Ontario, Canada; 4 Department of Neurology, Washington University of Medicine, St. Louis, Missouri; and 5 The Wistar Institute, Philadelphia, Pennsylvania Abstract High-grade gliomas are devastating brain tumors associated with a mean survival of <50 weeks. Two of the most common genetic changes observed in these tumors are overexpression/ mutation of the epidermal growth factor receptor (EGFR) vIII and loss of PTEN/MMAC1 expression. To determine whether somatically acquired EGFRvIII expression or Pten loss accel- erates high-grade glioma development, we used a previously characterized RasB8 glioma-prone mouse strain, in which these specific genetic changes were focally introduced at 4 weeks of age. We show that both postnatal EGFRvIII expression and Pten inactivation in RasB8 mice potentiate high-grade glioma development. Moreover, we observe a con- cordant loss of Pten and EGFR overexpression in nearly all high-grade gliomas induced by either EGFRvIII introduction or Pten inactivation. This novel preclinical model of high- grade glioma will be useful in evaluating brain tumor thera- pies targeted to the pathways specifically dysregulated by EGFR expression or Pten loss. (Cancer Res 2006; 66(15): 7429-37) Introduction High-grade gliomas are the most common brain tumors in adults, accounting for f60% of all gliomas (1). The most malignant grade 4 astrocytoma (glioblastoma multiforme) is composed of poorly differentiated neoplastic astrocytes (a subtype of glial cell), developing from a preexisting low-grade astrocytoma (LGA) or arising de novo in an individual without a previously identified brain tumor. Those tumors that form de novo are termed primary glioblastoma multiforme, whereas those that progress from LGAs are termed secondary glioblastoma multiforme. Irrespective of their developmental origin, glioblastoma multiformes are clinically indistinguishable and associated with a dismal prognosis, with a mean survival of <1 year. Two of the most commonly observed single genetic changes in both primary and secondary glioblastoma multiformes are overexpression and amplification of the epidermal growth factor receptor (EGFR) with or without an activating mutation (2–5) and loss of PTEN expression (6–10). The most common EGFR mutation consists of an aberrantly spliced form that lacks exons 3 to 6 (EGFRvIII), resulting in a constitutively active receptor (11, 12). Because of the frequency of EGFR amplification/mutation in glioblastoma multiformes, small- molecule EGFR inhibitors and EGFRvIII antibodies are currently being studied as glioblastoma multiforme therapies (13, 14). Loss of PTEN expression, resulting in aberrant activation of the phospha- tidylinositol 3-kinase signaling pathway in glioblastoma multi- formes, has similarly led to therapeutic interest in targeting this signaling pathway. The importance of these two genetic alterations toward modulating the therapeutic response of glioblastoma multiformes is highlighted by recent data, suggesting that EGFR inhibitors are more effective in glioblastoma multiformes where EGFRvIII and PTEN are both expressed but are not effective when PTEN expression is lost (15). The frequency of these genetic events and the interest in developing biologically based therapies underscore the need to develop robust preclinical models of high-grade gliomas harboring these genetic changes. Several preclinical mouse glioma models have shown that aberrant expression of EGFRvIII or loss of Pten expression in embryonic and/or adult glial cells by themselves do not lead to glioma formation (16–19). However, mouse glial cells harboring additional glioma predisposing genetic alterations, in conjunction with increased EGFRvIII or decreased Pten expression, develop high-grade gliomas, supportive of their role in glioma progression. For example, glioma formation induced by a truncated version of the SV40 large T antigen (T 121 ) is markedly accelerated by breeding to Pten +/À mice (20). Similarly, two studies with introduction of EGFRvIII in glial cells, which are null for Ink4a inactivation, led to high-grade glioma formation (16, 17). Our previously described glioma-prone mice (RasB8 mice), in which activated oncogenic Ha- Ras (V 12 Ha-Ras) is expressed under regulation of the human glial fibrillary acidic protein (GFAP; hGFAP) promoter commencing at embryonic days E14 to E16, are also supportive of these findings (21, 22). RasB8 pups are viable but commence development of progressive diffuse astroglial hyperproliferation early after birth, with subsequent development of low-grade gliomas (30% of the mice at 2 months of age) and then high-grade gliomas between 4 to 6 months, with majority of the gliomas being of astrocytic lineage. In addition to pathologic progression to higher-grade gliomas, there is molecular progression with acquisition of additional genetic alterations, reminiscent of ‘‘secondary’’ human glioblasto- ma multiforme. These include Tp53 mutations in the low-grade gliomas and overexpression of EGFR and loss of Pten in the high- grade gliomas (21, 22). In contrast to V 12 Ha-Ras, hGFAP-regulated Requests for reprints: Abhijit Guha, Fellow of the Royal College of Surgeons of Canada, 4W-446 Toronto Western Hospital, 399 Bathurst Street, Toronto, Ontario, Canada M5T 2S8. Phone: 416-603-5740; Fax: 416-603-5298; E-mail: Abhijit.Guha@ uhn.on.ca. I2006 American Association for Cancer Research. doi:10.1158/0008-5472.CAN-06-0712 www.aacrjournals.org 7429 Cancer Res 2006; 66: (15). August 1, 2006 Research Article Research. on February 18, 2016. © 2006 American Association for Cancer cancerres.aacrjournals.org Downloaded from