Killing of brain tumor cells by hypoxia-responsive element mediated expression of BAX.
The presence of radioresistant hypoxic cells in human brain tumors limits the overall effectiveness of conventional fractionated radiation therapy. Tumor-specific therapies that target hypoxic cells are clearly needed. We have investigated the expression of suicide genes under hypoxia by a hypoxia-responsive element (HRE), which can be activated through hypoxia-inducible factor-1 (HIF-1). We transfected plasmids containing multiple copies of HRE into U-87 MG and U-251 MG-NCI human brain tumor cells and tested their ability to induce LacZ gene expression under anoxia. Gene expression under anoxia versus oxia was increased about 12-fold for U-87 MG cells and about fourfold for U-251 MG-NCI cells. At intermediate hypoxic conditions, increased LacZ gene expression in U-87 MG cells was induced by the plasmid that contained three HREs, but not by the plasmid with two HREs. Lastly, when we placed a suicide gene BAX under the control of HREs, cells transfected with the BAX plasmids were preferentially killed through apoptosis under anoxia. Our studies demonstrate that HRE-regulated gene expression is active in brain tumor cells, and that the amount of increased gene expression obtained is dependent on the cell line, the HRE copy number, and the degree of hypoxia.
Brain Tumor Research Center of the Department of Neurological Surgery, University of California, San Francisco 94143-0520, USA.
H
H
Ruan
J
J
Wang
L
L
Hu
C S
CS
Lin
K R
KR
Lamborn
D F
DF
Deen
D001932:Brain Neoplasms / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000378:metabolism*; D015972:Gene Expression Regulation, Neoplastic; D005910:Glioma / Q000235:genetics*; D006801:Humans; D000860:Hypoxia; D051793:Hypoxia-Inducible Factor 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D020287:In Situ Nick-End Labeling; D007763:Lac Operon / Q000235:genetics; D008156:Luciferases / Q000378:metabolism; D009687:Nuclear Proteins / Q000378:metabolism*; D010100:Oxygen / Q000378:metabolism; D010957:Plasmids; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis*; D019253:Proto-Oncogene Proteins c-bcl-2; D014157:Transcription Factors; D014162:Transfection; D014407:Tumor Cells, Cultured; D051028:bcl-2-Associated X Protein; D001616:beta-Galactosidase / Q000378:metabolism
C494826:BAX protein, human; D004268:DNA-Binding Proteins; C497442:HIF1A protein, human; D051793:Hypoxia-Inducible Factor 1; D051795:Hypoxia-Inducible Factor 1, alpha Subunit; D009687:Nuclear Proteins; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D014157:Transcription Factors; D051028:bcl-2-Associated X Protein; D008156:Luciferases; D001616:beta-Galactosidase; D010100:Oxygen