Abstract 3458: Activated BRAF induces pilocytic astrocytomas in mice
Authors: Gronych Jan , Korshunov Andrey , Bageritz Josephine , Milde Till , Jugold Manfred , Jones David T. , Hambardzumyan Dolores , Remke Marc , Hartmann Christian , Witt Hendrik , Witt Olaf , Heiland Sabine , Bendszus Martin , Holland Eric C. , Pfister Stefan , Lichter Peter 
Published: June 2012

Abstract
Pilocytic astrocytoma (PA) comprises the most common primary brain tumor in children. Although this WHO grade I neoplasm shows a relatively benign biologic behavior many patients suffer from therapy-related long term neurologic impairment and up to 20% of patients experience tumor recurrence. Due to the slow growing nature of PA, adjuvant radiation and chemotherapy are only moderately effective. In order to pre-clinically evaluate novel therapy approaches, we aimed to establish a murine model of these tumors. Based on our previous work which showed that the MAPK intermediate BRAF is altered by duplication, activating mutation (V600E) or translocation of its kinase domain in a vast majority of cases, we applied somatic gene transfer using the Replication-Competent ASLV long terminal repeat (LTR) with a Splice acceptor (RCAS)/Tv-a technique to ectopically express different BRAF variants in Nestin-positive neural progenitors in the brains of newborn mice. While expression of either the BRAF i) full length wildtype, ii) full length V600E or iii) isolated kinase domain wildtype did not induce neoplastic lesions in mice, expression of the iv) isolated V600E mutated kinase domain induced tumors either in the cerebral hemispheres or in the brainstem depending on the site of injection. Although showing no clinical symptoms after an observation period of 4 months, post mortem analysis revealed presence of tumors in &gt;90% of cases. By histopathologic analysis, presence of strongly GFAP-positive lesions with a proliferation index (Ki67) below 1% could be identified. The slow tumor growth compared to other tumors induced with this technique together with presence of piloid-like tumor cells and eosinophilic structures (Rosenthal fibers) closely recapitulate clinical and biological features of human PA. Further analysis using primary murine Ntv-a astrocytes transduced with either of the four constructs in vitro revealed that expression of the isolated BRAF V600E kinase domain led to a twofold increase in proliferation compared to full length BRAF V600E or wild type BRAF kinase domain, while cells transduced with either full length wild type BRAF or GFP (control) showed almost no proliferation. Furthermore, this increase in proliferation could be markedly decreased by in vitro treatment with the kinase inhibitor Sorafenib at a concentration of 5 µM. This could be correlated with the amount of phosphorylation of the MAP kinases MEK and ERK, the downstream targets of BRAF. Treatment of mice with Sorafenib upon tumor induction is currently being conducted using ultrahigh field magnetic resonance imaging to monitor tumor growth and therapy response.
Taken together, our data provide first in vivo evidence for the oncogenic role of BRAF in pediatric pilocytic astrocytomas and it will be of paramount clinical importance to use this first animal model for pre-clinical testing of novel treatment approaches.
Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 3458. doi:10.1158/1538-7445.AM2011-3458