Transfection with mdm2-antisense or wtp53 results in radiosensitization and an increased apoptosis of a soft tissue sarcoma cell line.
Soft tissue sarcomas (STS) are mostly resistant after radiation treatment and are characterized by a rather low rate of apoptosis. The aim of this study was to test, in the p53 mutant STS cell line US8-93, the effect of a combined treatment with DNA transfection--either with mdm2 antisense oligodesoxynucleotides (mdm2-AS) or with a wild-type p53-plasmid (wtp53)--and the effects of irradiation on radiosensitivity. Mdm2-sense oligodesoxynucleotides (mdm2-SE) and a GFP-plasmid (GFP) were applied as controls. In order to evaluate the treatment radiation sensitization (clonogenic survival), apoptotic cell death and P53/MDM2-protein expression were determined. A moderately increased radiation sensitization was observed when comparing clonogenic survival after 2 Gy irradiation between cells transfected either with the control mdm2-SE (48%) or with mdm-2 AS (30%). At the same irradiation dose, clonogenic survival of wtp53-plasmid transfected cells (32%) was about 2-fold less than in the cells transfected with the control GFP-plasmid (61%). This enhancement factor of radiation sensitization was increased by about 3-fold at 4 Gy irradiation. Furthermore, an increase in apoptotic cells was already detectable by up to 7.7% (mdm2-AS) in comparison to 3.1% (mdm2-SE control) 72 hours after transfection. In parallel, the percentage of apoptotic cells could be further elevated after subsequent irradiation with 12 Gy by up to 15% (mdm2-AS) compared to 5.7% (mdm2-SE control). A striking result was obtained with the combined treatment of a wtp53 and 12 Gy irradiation which produced in 25% and 38.9% of apoptotic cells 48 hours and 72 hours after transfection, respectively. We can therefore conclude that the sensitivity of radiation therapy is enhanced by DNA transfection with wtp53 or mdm-2 AS ODNs for the correction of the p53-mdm2 balance in STS in vitro.
Grünbaum
U
U
Institute of Pathology, Martin Luther University Halle-Wittenberg, Halle/Saale, Germany.
Meye
A
A
Bache
M
M
Bartel
F
F
Würl
P
P
Schmidt
H
H
Dunst
J
J
Taubert
H
H
D017209:Apoptosis; D015153:Blotting, Western; D002460:Cell Line; D002470:Cell Survival; D004307:Dose-Response Relationship, Radiation; D006579:Heterozygote; D006801:Humans; D009687:Nuclear Proteins; D016376:Oligonucleotides, Antisense; D010957:Plasmids / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000235:genetics*; D051736:Proto-Oncogene Proteins c-mdm2; D011836:Radiation Tolerance; D012509:Sarcoma / Q000378:metabolism*; D013997:Time Factors; D014162:Transfection; D014407:Tumor Cells, Cultured; D016159:Tumor Suppressor Protein p53 / Q000235:genetics*
D009687:Nuclear Proteins; D016376:Oligonucleotides, Antisense; D011518:Proto-Oncogene Proteins; D016159:Tumor Suppressor Protein p53; C497206:MDM2 protein, human; D051736:Proto-Oncogene Proteins c-mdm2