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PDF file - 292K, Table S1 shows FACS analysis results of MET, CD133, and CD15 in the freshly isolated patient GBM cells, and Table S2 describes the list of patient-derived GBM cells and their derivatives used in this study. Patient information of 14 matched pre- and postradiated GBM specimens is shown in Table S3

PDF file - 6.4MB, Figure S1: High MET mRNA expression in GBM specimens portends poor patient survival. Figure S2: Dual immunofluorescence of MET and GSC enrichment markers (CD133 and CD15) on the frozen sections of GBM specimens. Figure S3: HGF/MET signaling in GBMs. Figure S4: METhigh cells generated xenograft tumors consisting of both METhigh and METlow/- cells. Figure S5: Immunohistochemical analysis of HGF protein in orthotopic xenograft tumors derived of from GBM 578 (A and C) and 464 (B and D). Figure S6: Effects of MET blockade on cell survival and clonogenicity of GSCs. Figure S7: In vitro migration and invasion assays using GSCs with MET targeting. Figure S8: Immunohistochemical analysis using human nuclei-specific antibody on the sections of GBM xenografts. Figure S9: Limiting dilution sphere forming assay to determine the clonogenic potentials of 822 GSCs after treatments of radiation and SU11274

Glioblastoma (GBM) patients have dismal median survival even with the most rigorous treatments currently available. Radiotherapy is the most effective non-surgical therapy for GBM patients; however, patients succumb due to tumor recurrence within a year. To develop a curative therapeutic approach, we need to better understand the underlying molecular mechanism of radiation resistance in GBM. Towards this goal, we developed an in vivo orthotopic GBM model system that mimics the radiation response of human GBM, using both established-GBM cell line and patient-derived freshly dissociated GBM specimen. In-vivo ionizing radiation (IR) treatment prolonged the survival of mice with intracranical tumor derived from U373MG, but failed to prevent tumor recurrence. U373MG and GBM578 cells isolated after in-vivo IR (U373-IR and 578-IR) were more clonogenic and enriched with stem cell-like characteristics, compared with mock-treated control tumor cells. Transcriptomic analyses and quantitative real-time reverse-transcription PCR analyses using these matched GBM cells before and after radiation treatment revealed that Wnt pathways were preferentially activated in post-IR GBM cells. U373-IR cells and 578-IR were enriched with cells positive for both active b-catenin (ABC) and Sox2 population, and this subpopulation was further increased after additional in-vitro radiation treatment, suggesting that radiation resistance of GBM is mediated due, in part, to the activation of stem cell-associated pathways including Wnt. Finally, pharmacological and siRNA inhibition of Wnt pathway significantly decreased the survival and clonogenicity of GBM cells and reduced their ABCþ/Sox2þ population. Together, these data suggest that Wnt activation is a molecular mechanism to confer GBM radioresistance and an important therapeutic target. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3458. doi:1538-7445.AM2012-3458

Ionizing radiation (IR) is a critical component in the treatments of breast cancer patients with all stages of the disease and a gold standard modality in patients with multiple brain metastasis, demonstrating improved overall survival (OS) in recent meta-analysis. However, the problem of recurrent or persistent disease exists due to tumor radio-resistance, and the development of distant metastasis after IR has presented a major obstacle to treatment. We investigated which molecule could be targeted for radio-sensitization. We found that c-Met over-expression and activation of downstream-signaling were induced in response to IR in a panel of human breast cancer cells by q-PCR, FACS, and Western blot analysis. The radio-sensitizing effect of c-Met silencing in vitro showed that increased c-Met activity is required for radio-resistance. In addition, c-Met inhibition combined with IR showed synergistic antitumor response including improved tumor regression and lengthened overall survival in breast cancer orthotopic and experimental brain metastasis xenograft models. Our preclinical findings establish a basis for a novel strategy to target the HGF/c-Met pathway with radiation when treating primary and brain metastatic breast cancer and possibly other c-Met pathway dependent cancers. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 3460. doi:1538-7445.AM2012-3460