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Characterization of actinomycin-D-resistant CHO cell lines exhibiting a multidrug-resistance phenotype and amplified
H Diddens1, V Gekeler, M Neumann
1Medizinisch-Naturwissenschaftliches Forschungzentrum, Universität Tübingen, FRG.
Abstract:
Actinomycin D (DACT)-resistant sublines of the Chinese hamster ovary cell line CHO-K1 were selected in vitro. Sublines were derived which expressed 5.2-fold (CHO 15DACT) and 35.8-fold (CHO 100DACT) resistance to DACT. The CHO 100DACT subline displayed marked cross-resistance to bleomycin, adriamycin, daunomycin, vinblastine, vincristine, VP 16 and VM 26. No cross-resistance was found to cisplatin or methotrexate. The resistant cells exhibited enhanced (collateral) sensitivity to prednisolone. Combination of prednisolone with vincristine resulted in a pronounced synergistic effect on sensitive cells, whereas in resistant cells the combined effect of both drugs was merely additive. Resistant cells, viably stained with the DNA-specific dye Hoechst 33342, exhibited decreased fluorescence intensities compared to parental cells. In contrast to sensitive cells the resistant sublines did not accumulate the mitochondria-specific dye rhodamine 123. Co-incubation with verapamil, however, effectively enhanced accumulation of the dye. The potential diagnostic value of these fluorescent compounds as marker dyes for the multidrug-resistance phenotype is discussed. Non-toxic doses of verapamil almost completely reversed the resistance to various drugs in CHO 100DACT cells. Specific DNA sequences were amplified in resistant cells, and the increase in resistance was paralleled by a concomitant increase in the copy number of these sequences, suggesting that the corresponding gene may be functionally linked to the multidrug-resistance phenotype.
Insights
Multidrug resistance in cancer cells was studied using Actinomycin D-resistant Chinese hamster ovary cells. Verapamil reversed resistance, and amplified DNA sequences correlated with multidrug resistance phenotype.
Area of Science:
- Cell Biology
- Molecular Biology
- Pharmacology
Background:
- Actinomycin D (DACT)-resistant Chinese hamster ovary (CHO-K1) cell sublines were developed in vitro.
- Selected sublines demonstrated significant DACT resistance (5.2-fold and 35.8-fold).
- The highly resistant CHO 100DACT subline showed cross-resistance to several chemotherapy drugs but not cisplatin or methotrexate.
Purpose of the Study:
- To investigate the characteristics of DACT-resistant CHO-K1 cell sublines.
- To explore cross-resistance patterns and collateral sensitivity.
- To evaluate the potential of fluorescent dyes and verapamil in multidrug resistance (MDR).
Main Methods:
- Selection of Actinomycin D-resistant CHO-K1 cell sublines.
- Assessment of drug cross-resistance and collateral sensitivity profiles.
- Analysis of fluorescent dye accumulation (Hoechst 33342, rhodamine 123) and verapamil's effect.
- Investigation of DNA sequence amplification in resistant cells.
Main Results:
- CHO 100DACT cells exhibited cross-resistance to multiple drugs (e.g., vincristine, adriamycin) and collateral sensitivity to prednisolone.
- Resistant cells showed decreased Hoechst 33342 fluorescence and failed to accumulate rhodamine 123, which was restored by verapamil.
- Verapamil effectively reversed drug resistance in CHO 100DACT cells.
- Specific DNA sequences were amplified in resistant cells, correlating with increased resistance.
Conclusions:
- The study characterized multidrug-resistant CHO-K1 cell lines with distinct resistance profiles.
- Fluorescent dyes show potential as diagnostic markers for MDR phenotype.
- Verapamil demonstrates significant potential in overcoming MDR.
- DNA sequence amplification is implicated in the MDR phenotype.