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Chemosensitisation by manganese superoxide dismutase inhibition is caspase-9 dependent and involves extracellular signal-regulated kinase 1/2.

Yeung BH, Wong KY, Lin MC, Wong CK, Mashima T, Tsuruo T, Wong AS - Br. J. Cancer (2008)

Bottom Line: Furthermore, our results revealed that this combination at the cellular level augmented activation of caspase-3 and caspase-9, but not caspase-8, suggesting involvement of an intrinsic apoptotic pathway.Akt activation was not affected.These results identify a novel chemoresistance mechanism in ovarian cancer, and show that combination of drugs capable of suppressing MnSOD with conventional chemotherapeutic agents may provide a novel strategy with a superior therapeutic index and advantage for the treatment of refractory ovarian cancer.

View Article: PubMed Central - PubMed

Affiliation: School of Biological Sciences, The University of Hong Kong, Hong Kong, PR China.

ABSTRACT
Chemoresistance and therapeutic selectivity are major obstacles to successful chemotherapy of ovarian cancer. Manganese superoxide disumutase (MnSOD) is an important antioxidant enzyme responsible for the elimination of superoxide radicals. We reported here that MnSOD was significantly elevated in ovarian cancer cells and its overexpression was one of the mechanisms that increased resistance to apoptosis in cancer cells. Knockdown of MnSOD by small-interfering RNA (siRNA) led to an increase in superoxide generation and sensitisation of ovarian cancer cells to the two front-line anti-cancer agents doxorubicin and paclitaxel whose action involved free-radical generation. This synergistic effect was not observed in non-transformed ovarian surface epithelial cells. Furthermore, our results revealed that this combination at the cellular level augmented activation of caspase-3 and caspase-9, but not caspase-8, suggesting involvement of an intrinsic apoptotic pathway. Evaluation of signalling pathways showed that MnSOD siRNA enhanced doxorubicin- and paclitaxel-induced phosphorylation of extracellular signal-regulated kinase 1/2. Akt activation was not affected. These results identify a novel chemoresistance mechanism in ovarian cancer, and show that combination of drugs capable of suppressing MnSOD with conventional chemotherapeutic agents may provide a novel strategy with a superior therapeutic index and advantage for the treatment of refractory ovarian cancer.

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Involvement of ROS production in anti-cancer drug-induced apoptosis in ovarian cancer cells. SKOV-3 cells transfected with manganese superoxide disumutase (MnSOD) small-interfering RNA (siRNA) were pre-treated with glutathione (GSH, 5 mM) or N-acetyl cysteine (NAC, 5 mM) for 30 min, and then the cells were treated with doxorubicin (DOX, 5 μM) or paclitaxel (PTX, 0.1 μM) for another 48 h. (A) Apoptosis was measured by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling (TUNEL) assay and 4′,6-diamidino-2-phenylindole (DAPI) staining. Experiments were repeated three times, and data are shown as mean±s.d. *P<0.05 vs untreated control cells. (B) O2•− level of DOX- and PTX-treated cells was measured by flow cytometry analysis using dihydroethidium. Solid histogram, untreated cells; open histogram, DOX- and PTX-treated cells.
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fig6: Involvement of ROS production in anti-cancer drug-induced apoptosis in ovarian cancer cells. SKOV-3 cells transfected with manganese superoxide disumutase (MnSOD) small-interfering RNA (siRNA) were pre-treated with glutathione (GSH, 5 mM) or N-acetyl cysteine (NAC, 5 mM) for 30 min, and then the cells were treated with doxorubicin (DOX, 5 μM) or paclitaxel (PTX, 0.1 μM) for another 48 h. (A) Apoptosis was measured by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling (TUNEL) assay and 4′,6-diamidino-2-phenylindole (DAPI) staining. Experiments were repeated three times, and data are shown as mean±s.d. *P<0.05 vs untreated control cells. (B) O2•− level of DOX- and PTX-treated cells was measured by flow cytometry analysis using dihydroethidium. Solid histogram, untreated cells; open histogram, DOX- and PTX-treated cells.

Mentions: Treatment of cells with antioxidants, such as GSH (5 mM) or NAC (5 mM), which can scavenge mitochondria-derived ROS, abolished the effects of MnSOD siRNA on DOX- and PTX-induced apoptosis (Figure 6A). Moreover, intracellular O2•− was actually generated by the treatment of OVCAR-3 (data not shown) and SKOV-3 (Figure 6B) cells with each anti-cancer drug, suggesting a role for ROS in the process.


Chemosensitisation by manganese superoxide dismutase inhibition is caspase-9 dependent and involves extracellular signal-regulated kinase 1/2.

Yeung BH, Wong KY, Lin MC, Wong CK, Mashima T, Tsuruo T, Wong AS - Br. J. Cancer (2008)

Involvement of ROS production in anti-cancer drug-induced apoptosis in ovarian cancer cells. SKOV-3 cells transfected with manganese superoxide disumutase (MnSOD) small-interfering RNA (siRNA) were pre-treated with glutathione (GSH, 5 mM) or N-acetyl cysteine (NAC, 5 mM) for 30 min, and then the cells were treated with doxorubicin (DOX, 5 μM) or paclitaxel (PTX, 0.1 μM) for another 48 h. (A) Apoptosis was measured by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling (TUNEL) assay and 4′,6-diamidino-2-phenylindole (DAPI) staining. Experiments were repeated three times, and data are shown as mean±s.d. *P<0.05 vs untreated control cells. (B) O2•− level of DOX- and PTX-treated cells was measured by flow cytometry analysis using dihydroethidium. Solid histogram, untreated cells; open histogram, DOX- and PTX-treated cells.
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Related In: Results  -  Collection

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fig6: Involvement of ROS production in anti-cancer drug-induced apoptosis in ovarian cancer cells. SKOV-3 cells transfected with manganese superoxide disumutase (MnSOD) small-interfering RNA (siRNA) were pre-treated with glutathione (GSH, 5 mM) or N-acetyl cysteine (NAC, 5 mM) for 30 min, and then the cells were treated with doxorubicin (DOX, 5 μM) or paclitaxel (PTX, 0.1 μM) for another 48 h. (A) Apoptosis was measured by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling (TUNEL) assay and 4′,6-diamidino-2-phenylindole (DAPI) staining. Experiments were repeated three times, and data are shown as mean±s.d. *P<0.05 vs untreated control cells. (B) O2•− level of DOX- and PTX-treated cells was measured by flow cytometry analysis using dihydroethidium. Solid histogram, untreated cells; open histogram, DOX- and PTX-treated cells.
Mentions: Treatment of cells with antioxidants, such as GSH (5 mM) or NAC (5 mM), which can scavenge mitochondria-derived ROS, abolished the effects of MnSOD siRNA on DOX- and PTX-induced apoptosis (Figure 6A). Moreover, intracellular O2•− was actually generated by the treatment of OVCAR-3 (data not shown) and SKOV-3 (Figure 6B) cells with each anti-cancer drug, suggesting a role for ROS in the process.

Bottom Line: Furthermore, our results revealed that this combination at the cellular level augmented activation of caspase-3 and caspase-9, but not caspase-8, suggesting involvement of an intrinsic apoptotic pathway.Akt activation was not affected.These results identify a novel chemoresistance mechanism in ovarian cancer, and show that combination of drugs capable of suppressing MnSOD with conventional chemotherapeutic agents may provide a novel strategy with a superior therapeutic index and advantage for the treatment of refractory ovarian cancer.

View Article: PubMed Central - PubMed

Affiliation: School of Biological Sciences, The University of Hong Kong, Hong Kong, PR China.

ABSTRACT
Chemoresistance and therapeutic selectivity are major obstacles to successful chemotherapy of ovarian cancer. Manganese superoxide disumutase (MnSOD) is an important antioxidant enzyme responsible for the elimination of superoxide radicals. We reported here that MnSOD was significantly elevated in ovarian cancer cells and its overexpression was one of the mechanisms that increased resistance to apoptosis in cancer cells. Knockdown of MnSOD by small-interfering RNA (siRNA) led to an increase in superoxide generation and sensitisation of ovarian cancer cells to the two front-line anti-cancer agents doxorubicin and paclitaxel whose action involved free-radical generation. This synergistic effect was not observed in non-transformed ovarian surface epithelial cells. Furthermore, our results revealed that this combination at the cellular level augmented activation of caspase-3 and caspase-9, but not caspase-8, suggesting involvement of an intrinsic apoptotic pathway. Evaluation of signalling pathways showed that MnSOD siRNA enhanced doxorubicin- and paclitaxel-induced phosphorylation of extracellular signal-regulated kinase 1/2. Akt activation was not affected. These results identify a novel chemoresistance mechanism in ovarian cancer, and show that combination of drugs capable of suppressing MnSOD with conventional chemotherapeutic agents may provide a novel strategy with a superior therapeutic index and advantage for the treatment of refractory ovarian cancer.

Show MeSH
Related in: MedlinePlus