AI-21HIF-1A TARGETING IN AN ORTHOTOPIC XENOGRAFT MODEL THAT RECAPITULATES THE HYPOXIC MICROENVIRONMENT OF GLIOBLASTOMA

INTRODUCTION: Tissue hypoxia and necrosis represent a pathological hallmark of glioblastoma. Hypoxia inducible factor 1a (HIF-1a) plays crucial roles in the malignant phenotypes of glioblastoma, and may offer a therapeutic target. However, developing HIF-1a targeted agents has been hampered by the lack of a preclinical model that recapitulates the disease relevant hypoxic microenvironment. MATERIALS AND METHODS: MGG123 neurosphere culture was established from a surgical specimen of a recurrent glioblastoma. Tumorigenicity of MGG123 cells was assessed by orthotopic xenografting in SCID-mice. The original tumor and xenografts were characterized by H&E staining and immunohistochemistry for HIF-1a, MIB-1, CD34, nestin and CD44. Hypoxic foci within xenografts were detected by pimonidazole immunofluorescence. In vitro, HIF-1a expression was evaluated by Western blot under normoxia and hypoxia after treatment with Digoxin and Ouabain, shown to inhibit HIF-1a synthesis. In vivo, mice bearing intracerebral MGG123 tumors were treated with intraperitoneal injections of digoxin, and its effects on HIF-1a expression, CD34+ vasculature, and overall survival were examined. RESULTS: MGG123 cells generated aggressive orthotopic xenografts that contain foci of pseudopalisading necrosis. The patient and xenografted tumors showed histopathological similarities as they both have high MIB-1 indices and are diffusely positive for CD44 and nestin. HIF-1a immunopositivity was specifically found in peudopalisading cells surrounding necrosis in both tumors. Within the xenografts, hypoxic areas were never found within 50mm proximity to CD34+ blood vessels. Hypoxia enhanced HIF-1a expression in cultured MGG123, and this was abrogated by treatment with Digoxin or Ouabain. In vivo, digoxin decreased HIF-1a expression and CD34+ vasculature within orthotopic MGG123 tumors, and extended overall survival of brain tumor-bearing mice. CONCLUSION: We established a preclinical glioblastoma model that uniquely mirrors the hypoxic microenvironment of human glioblastoma, and is suitable for developing HIF-1a targeted agents. Digoxin down-regulated HIF-1a and angiogenesis and increased overall survival in this clinically relevant glioblastoma model.