Rapid doxorubicin efflux from the nucleus of drug-resistant cancer cells following extracellular drug clearance

Following extracellular drug clearance, we analyzed the rate of doxorubicin efflux from the nucleus of three human leukemic cells (K562, Molt4 and CCRF-CEM) and related it to their differential sensitivity to this drug, after a short drug pulse. For many pulse-chase regimes, K562 cell viability was least affected by doxorubicin. In K562 cells, nuclear drug accumulation was greatest, but nuclear drug egress was also greatest. P-glycoprotein over-expression in a doxorubicin-resistant, K562/DOX sub-line did not facilitate doxorubicin efflux from the nucleus. In K562 cells, doxorubicin accumulated in multivesicular bodies (MVBs) through a pH-dependent mechanism. Inhibiting drug sequestration in MVBs did not affect nuclear efflux. The rates of doxorubicin efflux from the nuclei of live and digitonin-permeabilized K562 cells were similar. However, extracting cytoplasmic membranes with Triton X-100 significantly inhibited nuclear drug efflux following extracellular drug clearance. Our results are consistent with drug efflux from the nucleus being primarily mediated by an ATP-independent, passive diffusion mechanism. The effect of membrane extraction suggests that nonspecific drug absorption to cytoplasmic membranes plays a role in facilitating nuclear efflux in K562 cells, perhaps by lowering the concentration of free doxorubicin from a perinuclear diffusion boundary layer.