Polymerase chain reaction analysis of cisplatin-induced mitochondrial DNA damage in human ovarian carcinoma cells.
The purpose of this study was to determine whether the observed synergistic interaction between cisplatin and valinomycin (VM) in human ovarian carcinoma is the result of mitochondrial DNA (mtDNA) damage. A polymerase chain reaction (PCR)-based method was used to quantitate the lesion frequencies produced by cisplatin, VM and/or drug combination in a 1.1 kbp segment of mtDNA and a 0.536 kbp segment of the nuclear-located beta-globin gene in human ovarian CaOV-3 carcinoma cells. Our data indicates that the nuclear DNA (nDNA) received more cisplatin-induced damage at doses of 25 microM or less than did mtDNA. At higher cisplatin doses (50 microM or more), however, the damage was relatively equal in both segments. VM alone produced little or no damage on mtDNA, yet a significant amount of damage was detected within nDNA. However, when 1 microM VM was used in combination with low doses of cisplatin (0-40 microM), extensive mtDNA damage was detected as compared with the absence of detectable damage on nDNA. In mtDNA, the lesion frequency was 5.45 lesions/10 kb/10 microM cisplatin in the presence of 1 microM VM, whereas no detectable lesions were induced by cisplatin alone. This drug combination produced no detectable damage on DNA, indicating that cisplatin-induced mtDNA damage could be the basis for the observed synergistic interaction with VM. These results also correlate well with our recent in vivo study with the nude mice model of human ovarian cancer treated with a cisplatin/liposomal VM drug combination. Furthermore, this report shows evidence for the role of mitochondria and mtDNA as alternative targets for drug action in cancer therapy.
Daoud
S S
SS
Department of Pharmaceutical Sciences, Washington State University, Pullman 99164, USA.
Clements
M K
MK
Small
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D001483:Base Sequence; D002467:Cell Nucleus / Q000378:metabolism; D002945:Cisplatin / Q000494:pharmacology*; D004249:DNA Damage; D004272:DNA, Mitochondrial / Q000187:drug effects*; D004273:DNA, Neoplasm / Q000187:drug effects*; D004357:Drug Synergism; D005260:Female; D006801:Humans; D008969:Molecular Sequence Data; D015345:Oligonucleotide Probes; D010051:Ovarian Neoplasms / Q000378:metabolism*; D016012:Poisson Distribution; D016133:Polymerase Chain Reaction; D014407:Tumor Cells, Cultured; D014634:Valinomycin / Q000494:pharmacology
D004272:DNA, Mitochondrial; D004273:DNA, Neoplasm; D015345:Oligonucleotide Probes; D014634:Valinomycin; D002945:Cisplatin