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Characterization of an anthracycline-resistant human promyelocyte leukemia (HL-60) cell line with an elevated MDR-1
K Jönsson1, N Dahlberg, U Tidefelt
1Department of Medicine, Huddinge Hospital, Karolinska Institute, Sweden.
Abstract:
Multidrug resistance to a variety of cytotoxic drugs is due to decreased drug accumulation at the intracellular site of drug action. When due to increased energy-dependent drug efflux, this transport change is often associated with increased expression of an efflux pump for various lipophilic compounds, for example the P-glycoprotein which is the product of the MDR-1 gene. However, previously described HL-60 human promyelocytic leukemia cell lines resistant to the cytotoxic effect of anthracyclines have been reported not to express P-glycoprotein. We have isolated, by drug selection, an anthracycline-resistant HL-60 cell line that, in comparison to parental drug sensitive cells, exhibits a multidrug resistant phenotype including diminished intracellular drug retention, cross-resistance to multiple cytotoxic drugs, increased expression of a monoclonal antibody C219-reactive 180 kDa P-glycoprotein detected by Western blot analysis as well as increased expression of MDR-1 mRNA as determined by Northern blot and solution hybridization/RNAse protection analyses. Evidence is presented that the anthracycline-resistant HL-60 cells have amplified the MDR-1 gene.
Insights
This study identifies a multidrug-resistant HL-60 leukemia cell line that overexpresses P-glycoprotein and MDR-1 mRNA. This finding is linked to MDR-1 gene amplification, offering insights into drug resistance mechanisms.
Area of Science:
- Molecular Biology
- Cancer Research
- Pharmacology
Background:
- Multidrug resistance (MDR) in cancer often involves decreased intracellular drug accumulation.
- Increased energy-dependent drug efflux, mediated by efflux pumps like P-glycoprotein (MDR-1 gene product), is a common MDR mechanism.
- Previous anthracycline-resistant HL-60 cell lines reportedly did not express P-glycoprotein.
Purpose of the Study:
- To isolate and characterize an anthracycline-resistant HL-60 cell line.
- To investigate the molecular mechanisms underlying multidrug resistance in this cell line.
- To determine if P-glycoprotein expression and MDR-1 gene amplification are involved.
Main Methods:
- Drug selection to generate resistant cell lines.
- Phenotypic analysis for drug sensitivity and intracellular drug retention.
- Western blot analysis for P-glycoprotein expression (using C219 antibody).
- Northern blot and solution hybridization/RNAse protection assays for MDR-1 mRNA levels.
- Gene amplification analysis for the MDR-1 gene.
Main Results:
- An anthracycline-resistant HL-60 cell line was successfully isolated.
- The resistant cells exhibited cross-resistance to multiple cytotoxic drugs and diminished intracellular drug retention.
- Increased expression of 180 kDa P-glycoprotein and MDR-1 mRNA was observed in resistant cells.
- Evidence indicated amplification of the MDR-1 gene in the resistant HL-60 cells.
Conclusions:
- The isolated anthracycline-resistant HL-60 cell line demonstrates a multidrug-resistant phenotype.
- This resistance is associated with increased P-glycoprotein and MDR-1 mRNA expression.
- MDR-1 gene amplification appears to be the underlying mechanism for P-glycoprotein overexpression and multidrug resistance in this model.