The resistance of several leukaemic and myeloma cell lines (CCRF, L1210, HL-60, KG-1a and RPMI 8226) to VP-16 was found to increase with cell density and to be maximal (3.5- to 39-fold) in plateau phase cell cultures, as measured by clonogenic and MTT assays. Non-transformed confluent Flow 2000 human fibroblasts and Chinese hamster ovary (CHO) cells were also five- and 15-fold resistant to VP-16 respectively. The transition from log to plateau phase was accompanied by a drastic decrease in topoisomerase (topo) IIα content in CHO cells and human fibroblasts, while the leukaemic cells maintained constant cellular levels of topo IIα and topo IIβ. However, the nuclear topo IIα content was found to decrease as a result of translocation of the enzyme to the cytoplasmic compartment in the leukaemic cells. This was confirmed by subcellular fractionation experiments, Western blotting analyses and immunocytochemistry studies. The quantity of topo IIα in plateau phase cytoplasmic fractions ranged from 18% in L1210 cells to 50% in HL-60 and 8226 cells, as measured by both immunoblotting and quantification of the label in immunofluorescent images. The cytoplasmic fraction from plateau phase cells retained topo II catalytic activity, as measured by the decatenation of kinetoplast DNA. The nuclear–cytoplasmic ratio of topo IIα may be critical in determining the sensitivity of leukaemic cells to topo II inhibitors. Cytoplasmic trafficking of topo IIα was observed in plasma cells obtained from patients with multiple myeloma, and perhaps contributes to drug resistance in this disease.