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Reduced topoisomerase II activity in multidrug-resistant human non-small cell lung cancer cell lines
E W Eijdems1, M de Haas, A J Timmerman
1Division of Molecular Biology, The Netherlands Cancer Institute, Amsterdam.
Abstract:
Multidrug-resistant (MDR) cell lines often have a compound phenotype, combining reduced drug accumulation with a decrease in topoisomerase II. We have analysed alterations in topoisomerase II in MDR derivatives of the human lung cancer cell line SW-1573. Selection with doxorubicin frequently resulted in reduced topo II alpha mRNA and protein levels, whereas clones selected with vincristine showed normal levels of topo II alpha. No alterations of topo II beta levels were detected. To determine the contribution of topo II alterations to drug resistance, topo II activity was analysed by the determination of DNA breaks induced by the topo II-inhibiting drug 4'-(9-acridinylamino)methane-sulphon-m-anisidide (m-AMSA) in living cells, as m-AMSA is not affected by the drug efflux mechanism in the SW-1573 cells. The number of m-AMSA-induced DNA breaks correlated well (r = 0.96) with in vitro m-AMSA sensitivity. Drug sensitivity, however, did not always correlate with reduced topo II mRNA or protein levels. In one of the five doxorubicin-selected clones m-AMSA resistance and a reduction in m-AMSA-induced DNA breaks were found in the absence of reduced topo II protein levels. Therefore, we assume that post-translational modifications of topo II also contribute to drug resistance in SW-1573 cells. These results suggest that methods that detect quantitative as well as qualitative alterations of topo II should be used to predict the responsiveness of tumours to cytotoxic agents. The assay we used, which measures DNA breaks as an end point of topo II activity, could be a good candidate.
Insights
Multidrug resistance in lung cancer cells involves changes in topoisomerase II (topo II). Altered topo II levels and activity contribute to resistance, suggesting DNA breaks assay can predict treatment response.
Area of Science:
- Molecular Biology
- Cancer Research
- Pharmacology
Background:
- Multidrug-resistant (MDR) cell lines often exhibit reduced drug accumulation and decreased topoisomerase II (topo II) activity.
- Topoisomerase II alpha and beta are key enzymes in DNA replication and repair, and their alterations are implicated in cancer drug resistance.
Purpose of the Study:
- To analyze alterations in topoisomerase II (topo II) in multidrug-resistant (MDR) derivatives of the human lung cancer cell line SW-1573.
- To determine the contribution of topo II alterations to drug resistance and evaluate a DNA breaks assay for predicting treatment responsiveness.
Main Methods:
- Analysis of topoisomerase II alpha mRNA and protein levels in doxorubicin- and vincristine-selected MDR SW-1573 cell lines.
- Measurement of DNA breaks induced by the topo II-inhibiting drug m-AMSA in living cells to assess topo II activity.
- Correlation of DNA breaks with in vitro drug sensitivity and topo II protein levels.
Main Results:
- Doxorubicin selection frequently reduced topo II alpha mRNA and protein, while vincristine selection did not affect topo II alpha levels. Topo II beta levels remained unchanged.
- The number of m-AMSA-induced DNA breaks strongly correlated with in vitro m-AMSA sensitivity (r = 0.96).
- Drug sensitivity did not consistently correlate with reduced topo II mRNA or protein levels, indicating potential post-translational modifications affecting topo II activity.
Conclusions:
- Alterations in topoisomerase II, including quantitative and qualitative changes, contribute to multidrug resistance in SW-1573 lung cancer cells.
- A DNA breaks assay measuring topoisomerase II activity serves as a reliable indicator of drug sensitivity and could predict tumor response to cytotoxic agents.