Apoptosis by Retrovirus- and Adenovirus-Mediated Gene Transfer of Fas Ligand to Glioma Cells: Implications for Gene Therapy

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Abstract
Astrocytic tumors frequently express Fas/APO-1 (Fas), in sharp contrast to surrounding normal brain cells, providing a potential window through which selective killing of tumor cells could be pursued. To assess this possibility, we transduced Fas into U251, a glioma cell line resistant to anti-Fas antibody-mediated apoptosis, and obtained transfectants with high levels of Fas expression. Anti-Fas antibody showed significantly enhanced cytotoxicity for the transfectants, suggesting that U251 cells maintained an intercellular cascade of Fas-mediated apoptosis. When U251 transfectants with high-level Fas expression were transduced with Fas ligand-encoding gene via retrovirus, they were unaffected by exposure to anti-Fas antibody or Fas ligand adenovirus (Adeno-FL). Thus, retroviral induction of Fas ligand into the glioma cells with high levels of Fas led to the selection of cells that were resistant to Fas-dependent apoptosis. These resistant U251 transfectants were susceptible to FADD adenovirus (Adeno-FADD)-induced apoptosis, indicating that a cascade of death signals was blocked at the steps between Fas ligand and FADD. As for adenoviral transduction of Fas ligand into gliomas, gliomas with a relatively high level of expression of Fas were remarkably sensitive to Adeno-FL-induced apoptosis. Besides, Adeno-FADD induced pronounced apoptosis in all glioma cells. Our data suggest the possibility of using adenovirus-mediated transduction of Fas ligand and FADD genes as a therapeutic approach to target gliomas. Malignant astrocytomas express higher levels of Fas/APO-1 (Fas) than do normal brain tissues known to be resistant to Fas ligand-mediated apoptosis. To evaluate whether gliomas are susceptible to Fas ligand-mediated apoptosis, U251 glioma cells expressing high levels of Fas were exposed to Fas ligand by retrovirus and adenovirus infection. Fas ligand adenovirus (Adeno-FL) was lethal to U251 cells with high Fas expression, but Fas ligand retrovirus led to the selection of cells that were resistant to Fas-mediated apoptosis. This resistance might be explained by a blockage of apoptotic pathways at the steps between Fas ligand and FADD. As for gene therapy for malignant astrocytomas by Adeno-FL in vitro, gliomas with a relatively high expression of Fas were highly sensitive to Adeno-FL-mediated apoptosis. Moreover, adenovirus-mediated gene transduction of FADD (Adeno-FADD) was lethal to all glioma cells. These data support the potential usefulness of Adeno-FL and Adeno-FADD as a novel approach to the treatment of gliomas.