Abstract C176: Combining oncolytic herpes simplex virus and chemotherapy: G47 and temozolomide synergize in killing glioma stem cells

Abstract

Abstract Malignant gliomas, such as glioblastoma, are amongst the most common primary brain tumors in adults. Along with debulking surgery and post-operative radiation, the alkylating agent temozolomide (TMZ) is the standard of care for glioblastoma patients. However, the benefits of TMZ are correlated with tumors having transcriptional silencing of the O6-methylguanine methyltransferase (MGMT) gene, which occurs in approximately half of tumors. Unfortunately, regardless of MGMT status, recurrence or progression are inevitable, necessitating novel therapeutic approaches. Glioma stem cells (GSCs) have been implicated as a potential perpetrator of glioma recurrence. We have established human GSCs from surgically resected glioblastomas that vary in MGMT expression and TMZ sensitivity. Five human GSC lines were examined, two of which were MGMT-positive (confirmed by western blotting). MGMT-negative GSCs were more sensitive to TMZ (IC50<100 µM) than their counterparts grown in serum-containing media (representative of the bulk tumor) or established glioma cell lines such as U87 or U373. MGMT-positive GSCs were extremely resistant to TMZ (IC50>1000 µM). When MGMT was inactivated by O6-Benzylguanine (O6-BG), GSC sensitivity to TMZ approximately doubled. Oncolytic viruses destroy tumor cells by different mechanisms from conventional therapeutics, including TMZ, and therefore should interact positively. We have previously reported that clinically tested oncolytic herpes simplex virus (oHSV) G207 combined with TMZ synergistically kills established glioma cell lines. However, G207 does not replicate in GSCs. Therefore, we examined a newer generation oHSV, G47, which was effective at killing GSCs regardless of MGMT status. The interaction between TMZ and G47 on GSC killing was assessed by Chou-Talalay analysis. For MGMT-negative GSCs, the combination was synergistic, while it was antagonistic for MGMT-positive GSCs. When MGMT was inactivated with O6-BG, the interaction became synergistic for MGMT-positive GSCs. Interestingly, viral replication, entry, or spread was not affected by combination treatment. For those combinations that were synergistic, there was a large increase in H2AX, indicative of DNA double-strand breaks. Finally, the effect of combination therapy was assessed in two in vivo GSC models: GBM8 (MGMT-negative) and BT74 (MGMT-positive). Intracerebral tumors were established in athymic mice and treated with chemotherapy and/or intracerebral injection of G47 on the second day of chemotherapy. TMZ + G47 was more effective than either agent alone in extending survival in mice bearing GBM8 tumors. In the BT74 model, TMZ + O6-BG + G47 was more effective than TMZ+O6-BG or G47 alone. These studies demonstrate that GSCs can be both sensitive and resistant to TMZ, correlating with their MGMT status. All GSCs were sensitive to killing by G47 and this was synergistically enhanced by TMZ or TMZ+O6-BG, depending upon the MGMT status. Importantly, the combination was very efficacious in vivo in both models, suggesting that this combination strategy be tested clinically. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):C176.