Adenovirus-mediated transfer of p53 and Fas ligand drastically enhances apoptosis in gliomas.
Various therapeutic approaches toward killing glioma cells by inducing apoptosis have been developed, but these approaches are often hampered by anti-apoptotic mechanisms. In this study, we attempted to develop a technique that overrides the resistance toward apoptosis in glioma cells. To date, p53- and Fas-mediated apoptotic pathways have been shown to be different. Therefore, we carried out a gene therapy that combines the pro-apoptotic effect of these two different pathways. The recombinant adenoviruses (Advs) for p53 and Fas ligand (FL) (Adv-p53 and Adv-FL, respectively) were constructed. Transfecting the p53 gene into glioma cell lines (A-172 and U251 glioma cells) led to overexpression of Bax, a protein that induces permeability transition; at the same time, this transfection brought about an overexpression of Fas. To intensify Fas-mediated apoptosis, we transferred the FL gene together with the p53 gene by Adv-mediated gene transduction into A-172 and U251 cells. Coinfecting Adv-p53 and Adv-FL into A-172 cells, which are relatively resistant to apoptosis by infection with Adv-p53 or Adv-FL alone (Adv-p53, multiplicity of infection (MOI) of 100: 8.5 +/- 0.7%; Adv-FL, MOI of 100: 3.0 +/- 0.1%) resulted in a drastic enhancement of the percentage of apoptotic cells (Adv-p53 and Adv-FL, each at an MOI of 30: 24.2 +/- 0.9%). Coinfection with Adv-p53 and Adv-FL in U251 cells resulted in a similar enhancement of the percentage of apoptotic cells (Adv-p53 and Adv-FL, each at an MOI of 30: 59.0 +/- 2.3%) compared with that seen in U251 cells transfected with Adv-p53 or Adv-FL alone (Adv-p53, MOI of 30: 3.1 +/- 0.3%; Adv-FL, MOI of 30: 18.1 +/- 0.3%). Regardless of whether a cell line is resistant or sensitive to FL- and p53-mediated apoptosis, coinfection with Adv-p53 and Adv-FL dramatically enhanced the degree of apoptosis of glioma cells. Our results indicate that the coinfection approach can be used as a modality for the gene therapy of gliomas, overriding the apoptosis-resistant mechanisms in glioma cells.
Shinoura
N
N
Department of Molecular Biotherapy Research, Cancer Chemotherapy Center, Japanese Foundation for Cancer Research, Tokyo.
Yoshida
Y
Y
Asai
A
A
Kirino
T
T
Hamada
H
H
D000256:Adenoviridae / Q000235:genetics*; D017209:Apoptosis; D001932:Brain Neoplasms / Q000378:metabolism / Q000628:therapy*; D002469:Cell Separation; D018076:DNA, Complementary / Q000378:metabolism; D004171:Diploidy; D004305:Dose-Response Relationship, Drug; D053222:Fas Ligand Protein; D005434:Flow Cytometry; D018014:Gene Transfer Techniques; D016158:Genes, p53 / Q000235:genetics*; D015316:Genetic Therapy / Q000379:methods*; D005910:Glioma / Q000378:metabolism / Q000628:therapy*; D006801:Humans; D015151:Immunoblotting; D007763:Lac Operon; D008562:Membrane Glycoproteins / Q000235:genetics*; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2; D014161:Transduction, Genetic; D014407:Tumor Cells, Cultured; D015854:Up-Regulation; D051028:bcl-2-Associated X Protein
C494826:BAX protein, human; D018076:DNA, Complementary; C505661:FASLG protein, human; D053222:Fas Ligand Protein; D008562:Membrane Glycoproteins; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D051028:bcl-2-Associated X Protein