Oncolytic HSV–Infected Glioma Cells Activate NOTCH in Adjacent Tumor Cells Sensitizing Tumors to Gamma Secretase Inhibition

Purpose: To examine the effect of oncolytic herpes simplex virus (oHSV) on NOTCH signaling in central nervous system tumors. Experimental Design: Bioluminescence imaging, reverse phase protein array proteomics, fluorescence microscopy, reporter assays, and molecular biology approaches were used to evaluate NOTCH signaling. Orthotopic glioma-mouse models were utilized to evaluate effects in vivo. Results: We have identified that herpes simplex virus-1 (HSV-1; oncolytic and wild-type)-infected glioma cells induce NOTCH signaling, from inside of infected cells into adjacent tumor cells (inside out signaling). This was canonical NOTCH signaling, which resulted in activation of RBPJ-dependent transcriptional activity that could be rescued with dnMAML. High-throughput screening of HSV-1–encoded cDNA and miRNA libraries further uncovered that HSV-1 miR-H16 induced NOTCH signaling. We further identified that factor inhibiting HIF-1 (FIH-1) is a direct target of miR-H16, and that FIH-1 downregulation by virus encoded miR-H16 induces NOTCH activity. FIH-1 binding to Mib1 has been reported, but this is the first report that shows FIH-1 sequester Mib1 to suppress NOTCH activation. We observed that FIH-1 degradation induced NOTCH ligand ubiquitination and NOTCH activity. REMBRANDT and The Cancer Genome Atlas data analysis also uncovered a significant negative regulation between FIH-1 and NOTCH. Furthermore, combination of oHSV with NOTCH-blocking gamma secretase inhibitor (GSI) had a therapeutic advantage in two different intracranial glioma models treated with oncolytic HSV, without affecting safety profile of the virus in vivo. Conclusions: To our knowledge this is the first report to identify impact of HSV-1 on NOTCH signaling and highlights the significance of combining oHSV and GSI for glioblastoma therapy.