A pathway for intrinsic drug resistance in leukemia cells.

Abstract

Drug resistance is a major cause of failure in the treatment of cancer using chemotherapy. The efflux of drug away from its target site within the cell is one resistance mechanism that has not been fully characterized. The subcellular transport of the anticancer agent doxorubicin was evaluated in three human leukemia cell lines displaying different levels of chemosensitivity. Of the cells tested, the K562 cell line exhibited the greatest intrinsic resistance to doxorubicin. Compared to the other cell lines, K562 cells accumulated the greatest mass of doxorubicin within the nucleus, the drug's site of action, but drug also egressed from the nucleus faster. The efflux of drug from the nucleus occurred more rapidly in living K562 cells than isolated nuclei, indicating the presence of a facilitated nuclear efflux mechanism in these cells. Following drug efflux from the nucleus, sequestration of drug in cytosolic vesicles was observed. The vesicles were identified as multivesicular bodies (MVBs), components of the endocytic pathway. Upon drug binding, the MVBs changed in morphology and were seen fusing with the plasma membrane, providing an exit route for drug from the cell. Expression of the drug transporter P-glycoprotein was not found to be required for rapid nuclear efflux, nor was the sequestration of drug within cytoplasmic vesicles. In studies evaluating drug efflux in permeabilized cells, the presence of cytoplasmic membranes was shown to promote egress of drug from the nucleus. The addition of cellular membranes to isolated nuclei was found to restore rapid nuclear efflux in an energy-independent manner, confirming that perinuclear membranes participate in drug efflux from the target site. This thesis argues for the importance of understanding general biophysical phenomena governing subcellular drug transport in the development of chemotherapeutic strategies to overcome drug resistance.