Vesicular anthracycline accumulation in Doxorubicin-selected U-937 cells: Participation of lysosomes

The U-A10 cell line, a doxorubicin-selected variant of human U-937 myeloid leukemia cells, exhibits a redistribution of anthracyclines into a expanded vesicular compartment. The acidic nature of this compartment was confirmed by vital staining with a pH sensitive dye, LysoSensor yellow/blue DND-160. Identification of the vesicular compartment was performed by immunofluorescence analysis. Staining for the LAMP-1 and LAMP-2 antigens showed that the vesicles are enlarged lysosomes that are eccentrically placed near the nucleus of U-A10 cells. By contrast, the expression of the multidrug resistance-associated protein and the P-glycoprotein wits observed predominately on the plasma membrane of the drug-resistant cells. The accumulation of daunorubicin into cellular compartments was quantified using radiolabeled drug. Exposing cells to (3)[H]-daunorubicin and then isolating intact nuclei showed that nuclei from U-A10 cells accumulated twofold to threefold less anthracycline than nuclei from U-937 cells, However, when nuclei were isolated first and then exposed to (3)[H]-daunorubicin, little difference in net nuclear drug accumulation was detected. Cytoplasts prepared from U-A10 and U-937 cells were exposed to (3)[H]daunorubicin to measure cytoplasmic drug accumulation.