Sublethal concentrations of gemcitabine (2',2'-difluorodeoxycytidine) alter mitochondrial ultrastructure and function without reducing mitochondrial DNA content in BxPC-3 human pancreatic carcinoma cells.
2',2'-Difluorodeoxycytidine (gemcitabine), a pyrimidine nucleoside analog, is used therapeutically in the treatment of pancreatic, non-small cell lung, and breast cancer. The cytotoxic effect of gemcitabine is thought to be due to masked chain termination after the triphosphorylated anabolite of the drug is incorporated into nascent DNA strands. We tested the hypothesis that sublethal concentrations of gemcitabine inhibit DNA polymerase gamma and reduce mitochondrial DNA content in BxPC-3 and MOLT-4 cell lines, and we used 2',3'-dideoxycytidine, a known inhibitor of DNA polymerase gamma as a positive control. The 6-day BxPC-3 cell growth IC(50) for gemcitabine and 2',3'-dideoxycytidine was 0.003 microM (SD +/- 0.0005) and 14.5 microM (SD +/- 4.7), respectively, and in MOLT-4 cells was 0.002 microM (SD +/- 0.001) and 0.86 muM (SD +/- 0.23), respectively. These drug concentrations were anti-proliferative but non-cytotocidal. Electron photomicrographic studies showed deranged mitochondrial cristae patterns in BxPC-3 cells treated with either gemcitabine or 2',3'-dideoxycytidine for 6 days. Mitochondrial oxidative phosphorylation dysfunction was observed as reflected by increased lactate concentration in the media of cells exposed to gemcitabine, but to a much greater extent in cells exposed to 2',3'-dideoxycytidine. PCR analysis showed that gemcitabine did not reduce mitochondrial DNA content in either BxPC-3 or MOLT-4 cells, but 2',3'-dideoxycytidine did. The effect of gemcitabine on mitochondrial ultrastructure and function did not concomitantly yield a reduction in mitochondrial DNA content. Therefore, the molecular target(s) by which gemcitabine and 2',3'-dideoxycytidine produce mitochondrial abnormalities in these cells appear to be different.
Yeo
T K
TK
Section of Clinical Pharmacology, Department of Medicine, Dartmouth Medical School and Dartmouth Hitchcock Medical Center, Lebanon, NH 03756, USA.
Kintner
J
J
Armand
R
R
Perez
R
R
Lewis
L D
LD
D000964:Antimetabolites, Antineoplastic / Q000633:toxicity*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D000074002:DNA Polymerase gamma; D004272:DNA, Mitochondrial / Q000032:analysis / Q000187:drug effects*; D004259:DNA-Directed DNA Polymerase / Q000187:drug effects; D003841:Deoxycytidine / Q000031:analogs & derivatives* / Q000633:toxicity; D004305:Dose-Response Relationship, Drug; D004354:Drug Screening Assays, Antitumor; D006801:Humans; D020128:Inhibitory Concentration 50; D019344:Lactic Acid / Q000378:metabolism; D008928:Mitochondria / Q000187:drug effects* / Q000648:ultrastructure; D010190:Pancreatic Neoplasms / Q000188:drug therapy* / Q000473:pathology; D018894:Reverse Transcriptase Inhibitors / Q000494:pharmacology; D016047:Zalcitabine / Q000494:pharmacology; D000093542:Gemcitabine
D000964:Antimetabolites, Antineoplastic; D004272:DNA, Mitochondrial; D018894:Reverse Transcriptase Inhibitors; D003841:Deoxycytidine; D019344:Lactic Acid; D016047:Zalcitabine; D000074002:DNA Polymerase gamma; D004259:DNA-Directed DNA Polymerase; D000093542:Gemcitabine