Expression of the multidrug resistance associated protein and P-glycoprotein in doxorubicin-selected human myeloid leukemia cells.
Drug-resistant sublines of the human U-937 myeloid leukemia cell line were selected in doxorubicin concentrations of 10, 40, and 200 ng/mL (designated U-A10, U-A40, and U-A200, respectively). Northern blot analysis showed overexpression of the multidrug resistance-associated protein (MRP) gene, but not MDR1, in U-A10 cells as compared with parental U-937 cells. Prolonged passage of U-A10 cells in 10 ng/mL of doxorubicin had little effect on MRP RNA levels, but increased MDR1 expression. The U-A40 and U-A200 cells, derived by selection of U-A10 cells, showed high levels of both MRP and MDR1 expression. None of the drug-resistant cell lines showed MRP or MDR1 gene amplification as judged by Southern blot analysis. U-A10 cells exhibited minimal decreased net accumulation of anthracycline, whereas U-A40 and U-A200 cells showed more significantly decreased drug accumulation as compared with U-937 cells. Subcellular anthracycline accumulation in U-937 cells as determined by fluorescence microscopy showed daunorubicin fluorescence predominately in the nucleus. However, the drug-resistant cell lines showed minimal nuclear drug accumulation with marked redistribution of drug into a vesicular compartment. Treatment with sodium azide/2-deoxyglucose, 2,4-dinitrophenol, or monensin, but not verapamil, abolished the vesicular accumulation. These studies in doxorubicin-selected U-937 cells indicate that induction of MRP overexpression occurs before that for the MDR1 gene. In addition, the drug-resistant cells possess an energy-dependent redistribution of anthracyclines into a nonnuclear vesicular compartment.
Slapak
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Division of Cancer Pharmacology, Dana-Farber Cancer Institute, Harvard Medical School, Boston, MA 02115.
Mizunuma
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Kufe
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D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1 / Q000378:metabolism*; D002451:Cell Compartmentation; D004273:DNA, Neoplasm / Q000235:genetics; D003630:Daunorubicin / Q000378:metabolism; D004317:Doxorubicin / Q000494:pharmacology*; D018432:Drug Resistance, Multiple; D005784:Gene Amplification; D006801:Humans; D066298:In Vitro Techniques; D011506:Proteins / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D012334:RNA, Neoplasm / Q000378:metabolism; D014407:Tumor Cells, Cultured
D020168:ATP Binding Cassette Transporter, Subfamily B, Member 1; D004273:DNA, Neoplasm; D011506:Proteins; D012333:RNA, Messenger; D012334:RNA, Neoplasm; D004317:Doxorubicin; D003630:Daunorubicin