Abstract 3727: An orthotopic xenograft model for studying reirradiation and glioblastoma evolution
Authors: McAbee Joseph H. , Rath Barbara H. , Wu Xiaolin , Shankavaram Uma , Camphausen Kevin , Tofilon Philip J. 
Published: August 2019

Glioblastoma (GBM), the most common and malignant primary adult brain cancer, has a median survival of 15 months despite multimodal treatment involving surgical resection and chemo-radiotherapy. Ongoing clinical trials seek to examine the safety and benefit of retreatment with radiation therapy for recurrent glioblastoma. By adding a reirradiation protocol to a previously described glioma stem-like cell (GSC) initiated orthotopic xenograft model, this study seeks to better understand the impact of radiotherapy on both primary and recurrent GBM evolution and to establish an in vivo model for studying reirradiation. Therefore, we intracranially implanted CD133+ NSC11 cells, and other GSC lines, into nude mice. After 21 days, bioluminescence imaging was performed to confirm the presence of tumor prior to randomization into control and radiation therapy groups (3x5Gy). After treatment tumors were imaged weekly to track changes in BLI ratios. Once the average BLI ratio for the treated mice was found to be between 1 and 10, the mice were rerandomized into control (3x5Gy-Control) and radiation therapy groups (3x5Gy-3x5Gy). Following treatment, brain samples were collected at various time points out to morbidity to investigate changes in tumor morphology and histology. Further, tumors from morbid mice were collected for viral integration site analysis (VISA), whole-exome sequencing (WES), and NanoString gene expression analysis. Survival analysis demonstrated a significant survival advantage for mice undergoing radiation therapy (+34.2 days) compared to controls. A further survival advantage was found for mice undergoing reirradiation (+30.0 days) compared to mice receiving only one course of radiation. On gross examination of morphology and H&E/SOX2 staining, brains bearing irradiated tumors and reirradiated tumors contained tumor tissue that was more likely to efface olfactory bulb(s) and less infiltrative than control tumors. These histological changes were followed up with VISA which revealed that control tumors harbor fewer clones than in vitro lines and that irradiated tumors harbor the fewest clones of all. Gene expression and IPA analyses showed that pathways involved in cell movement, survival, and proliferation are differentially regulated between irradiated and control tumors. WES was performed to compare gene mutation patterns between reirradiated, irradiated, and control samples. Our results demonstrate that radiation, a central component of glioblastoma treatment, can have wide-ranging effects on the evolution of this dynamic tumor after initial presentation and recurrence. We have demonstrated for the first time the utility of a GSC-initiated orthotopic xenograft model for studying recurrent GBM biology and evolution. This reirradiation model may provide the opportunity to design and test more effective recurrent GBM treatment strategies that are centered around recurrent biology.

Citation Format: Joseph H. McAbee, Barbara H. Rath, Xiaolin Wu, Uma Shankavaram, Kevin Camphausen, Philip J. Tofilon. An orthotopic xenograft model for studying reirradiation and glioblastoma evolution [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 3727.