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The relationship between modulation of MDR and glutathione in MRP-overexpressing human leukemia cells
K V Grech1, R A Davey, M W Davey
1Department of Cell and Molecular Biology, University of Technology, Sydney, Gore Hill NSW, Australia.
Abstract:
Multidrug resistance-associated protein (MRP) causes multidrug resistance (MDR) involving the anthracyclines and epipodophyllotoxins. Many studies show modulation of anthracycline levels and cytotoxicity in MRP-overexpressing cells, but there is limited data on the modulation of etoposide levels and cytotoxicity in MRP-overexpressing or in P-glycoprotein-expressing cells. Etoposide accumulation was 50% reduced in both the CEM/E1000 MRP-overexpressing subline and the CEM/VLB100 P-glycoprotein-expressing subline compared to the parental CEM cells, correlating with similar resistance to etoposide (200-fold) of the two sublines. For the CEM/VLB100 subline, the P-glycoprotein inhibitor SDZ PSC 833, but not verapamil, was able to increase etoposide accumulation and cytotoxicity. For the CEM/E1000 subline, neither SDZ PSC 833 nor verapamil had any effect on etoposide accumulation. However, verapamil caused a 4-fold sensitization to etoposide in this subline, along with an 80% decrease in cellular glutathione (P < 0.05). Buthionine sulfoximine (BSO), which depletes glutathione, also caused a 2.5-fold sensitization to etoposide with no effect on accumulation in the CEM/E1000 subline. In contrast, SDZ PSC 833 was able to increase daunorubicin accumulation in the CEM/E1000 subline (P < 0.05), but had no effect on daunorubicin cytotoxicity, or cellular glutathione. These results show that modulation of etoposide cytotoxicity in MRP-overexpressing cells may be through changes in glutathione metabolism rather than changes in accumulation and confirm that changes in drug accumulation are not related to drug resistance in MRP-overexpressing cells.
Insights
Multidrug resistance-associated protein (MRP) affects etoposide resistance differently than P-glycoprotein. Etoposide resistance in MRP-overexpressing cells may involve glutathione metabolism, not just drug accumulation.
Area of Science:
- Biochemistry
- Cell Biology
- Pharmacology
Background:
- Multidrug resistance-associated protein (MRP) and P-glycoprotein (P-gp) are key mediators of multidrug resistance (MDR).
- While MRP's role in anthracycline resistance is established, its impact on etoposide levels and cytotoxicity is less understood.
- Limited data exists on etoposide modulation in P-gp-expressing cells.
Purpose of the Study:
- To investigate the mechanisms of etoposide resistance in MRP-overexpressing cells.
- To compare etoposide accumulation and cytotoxicity in MRP- and P-gp-expressing cells.
- To determine the role of drug accumulation versus glutathione metabolism in etoposide resistance.
Main Methods:
- Comparison of etoposide accumulation and cytotoxicity in parental, MRP-overexpressing (CEM/E1000), and P-gp-overexpressing (CEM/VLB100) cell lines.
- Treatment with P-gp inhibitor (SDZ PSC 833) and verapamil.
- Assessment of cellular glutathione levels following verapamil and buthionine sulfoximine (BSO) treatment.
- Measurement of daunorubicin accumulation and cytotoxicity.
Main Results:
- Etoposide accumulation and resistance were similar in both MRP- and P-gp-overexpressing cells.
- P-gp inhibition increased etoposide accumulation and cytotoxicity in P-gp-expressing cells.
- Neither P-gp inhibitor nor verapamil affected etoposide accumulation in MRP-overexpressing cells.
- Verapamil and BSO sensitized MRP-overexpressing cells to etoposide, linked to altered glutathione levels.
- SDZ PSC 833 increased daunorubicin accumulation in MRP-overexpressing cells but not cytotoxicity.
Conclusions:
- Etoposide resistance in MRP-overexpressing cells is primarily mediated by altered glutathione metabolism, not changes in drug accumulation.
- Drug accumulation is not the sole determinant of resistance in MRP-overexpressing cells.
- MRP and P-gp exhibit distinct mechanisms in modulating etoposide resistance.