A Forward Genetics Screen Of Murine Brain Tumors Identifies Novel Candidate Genes Involved In Gliomagenesis

Proceedings: AACR 107th Annual Meeting 2016; April 16-20, 2016; New Orleans, LAGlioma is the most frequent malignant brain tumor in adults. Platelet-derived growth factor (PDGF) signaling is commonly activated in glioma. We have used a retrovirus-driven PDGFB-induced murine glioma model that causes tumors that closely resemble human gliomas of various grades. Knowing that retroviruses have a capacity to induce insertional mutagenesis, we have employed whole genome sequencing to identify potential genes that, together with PDGFB, drive glioma development.Gliomas were induced by RCAS virus injection into the brains of mice expressing the RCAS retroviral receptor from specific promoters. Genomic DNA from tumor cell lines was probed for retroviral tags and sequenced to identify genomic targets of the retrovirus. A streamlined analysis pipeline was developed for retrovirus integration detection and mapping to the reference mouse genome. Integration sites were analyzed and a common integration site (CIS) label was assigned to a gene, given that it was either tagged by a retrovirus more than once within a discovery set or found within the Retroviral Tagged Cancer Gene Database (RTCGD).In a small discovery subset of 15 murine gliomas, we have identified 40 CIS, of which 37 were validated by Sanger sequencing. When compared with previously identified CIS in RTCGD, 5.5% of them were shared with our older screen, where we overexpressed PDGFB from another retrovirus in order to induce glioma. Less CIS genes were shared with other published tumor models induced by viruses driven by other cancer genes/viruses.The majority of genes identified in our screen were tagged twice. However, Nfic, Cuecd1, Thra, Foxj1 and Nrxn1 were tagged three times, Ppfibp1 and Rhbg four times, and Mir29a/29b-1 seven times. As compared to control tumor lines, two top candidate genes, Mir29a and Ppfibp1, demonstrated significantly increased expression in tumor lines in were they were respectively tagged. Mir29a is often found downregulated in human tumors including gliomas, still high levels of Mir29a are sometimes found in certain aggressive cancers and in metastases.Interestingly, we found that specific PDGFR inhibition negatively regulates Mir29a, indicating a possible role for PDGF signaling in Mir29a regulation. Ppfibp1 has not been extensively studied in cancer. However, Ppfibp1 seems to have a subgroup-specific expression in human glioblastoma, making it an interesting candidate for further analysis.Here we present a new screening method that can be employed to identify genes involved in PDGFB-driven gliomagenesis. So far, we have identified 37 candidate genes by whole genome sequencing. Two of the most frequently tagged candidates, Mir29a and Ppfibp1 were upregulated as a consequence of retroviral mutagenesis. Their precise role in driving glioma formation in collaboration with PDGF is currently explored.Citation Format: Matko Cancer, Holger Weishaupt, Gabriela Rosen, Ignas Bunikis, Yiwen Jiang, Smitha Sreedharan, Sara Bolin, Ulf Gyllensten, Oren J. Becher, Lene Uhrbom, Adam Ameur, Fredrik J. Swartling. A forward genetics screen of murine brain tumors identifies novel candidate genes involved in gliomagenesis. [abstract]. In: Proceedings of the 107th Annual Meeting of the American Association for Cancer Research; 2016 Apr 16-20; New Orleans, LA. Philadelphia (PA): AACR; Cancer Res 2016;76(14 Suppl):Abstract nr 2688.