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Multiple mechanisms confer different drug-resistant phenotypes in pancreatic carcinoma cells
1Humboldt University Berlin, Charité Campus Mitte, Institute of Pathology, Schumannstr. 20/21, 10117 Berlin, Germany. hermann.lage@charite.de
Purpose:
Drug-resistant phenotypes of cancer cells may be caused by complex multimodal mechanisms of resistance. In order to gain further insighte into these mechanisms, a P-glycoprotein-mediated multidrug-resistant phenotype induced by daunorubicin-selection and an alternative drug resistance due to treatment with mitoxantrone were investigated in pancreatic carcinoma-derived cells.
Methods:
For assessing cross-resistance against various drugs, cell proliferation assays were performed. Drug accumulation was measured by flow cytometry. Messenger RNA expression was analyzed by Northern blot and RT-PCR, whereas protein expression was determined by Western blot. Catalytic activity of DNA-topoisomerases (Topo) II was determined by the decatenation assay.
Results:
In mitoxantrone-selected EPP85-181RNOV cells a decreased accumulation of mitoxantrone and daunorubicin was observed in the absence of P-glycoprotein, multidrug resistance protein or breast cancer resistance protein over-expression. An approximately twofold decrease of DNA topoisomerase II catalytic activity could be observed in both drug-resistance-exhibiting cell lines. The reduction of Topo II catalytic activity was reflected by decreased expression of Topo IIalpha and IIbeta mRNAs and proteins.
Conclusions:
The decreased drug accumulation in EPP85-181RNOV cells indicates that alternative transport events are occurring. The decreased catalytic activity and expression of Topo II indicate that modulation of Topo II catalytic activity contributes to both drug-resistant phenotypes in pancreatic carcinoma cells.
Insights
Pancreatic cancer cells develop drug resistance through alternative transport and reduced DNA topoisomerase II activity. This study investigates mechanisms of resistance to daunorubicin and mitoxantrone, revealing key molecular changes.
Area of Science:
- Oncology
- Molecular Biology
- Cancer Cell Biology
Background:
- Drug resistance is a major challenge in cancer therapy, often involving complex mechanisms.
- Multidrug resistance (MDR) phenotypes can arise from various cellular processes, including altered drug transport and target enzyme function.
Purpose of the Study:
- To investigate the mechanisms of P-glycoprotein-mediated multidrug resistance induced by daunorubicin and alternative drug resistance induced by mitoxantrone in pancreatic carcinoma cells.
- To elucidate the roles of drug accumulation, transporter proteins, and DNA topoisomerase II (Topo II) activity in acquired drug resistance.
Main Methods:
- Cell proliferation assays were used to assess cross-resistance to various drugs.
- Drug accumulation was quantified using flow cytometry.
- Messenger RNA (mRNA) and protein expression levels of Topo II were analyzed via Northern blot, RT-PCR, and Western blot.
- DNA topoisomerase II catalytic activity was measured using the decatenation assay.
Main Results:
- Mitoxantrone-selected cells showed decreased accumulation of daunorubicin and mitoxantrone without overexpression of known efflux pumps (P-glycoprotein, MRP, BCRP).
- Both drug-resistant cell lines exhibited an approximate twofold decrease in DNA topoisomerase II catalytic activity.
- Reduced Topo II activity correlated with decreased expression of Topo IIalpha and IIbeta mRNA and proteins.
Conclusions:
- Alternative transport mechanisms contribute to decreased drug accumulation in resistant pancreatic cancer cells.
- Modulation of DNA topoisomerase II catalytic activity and expression is a significant factor in the development of both daunorubicin- and mitoxantrone-induced drug resistance phenotypes.