Altered phosphorylation, biosynthesis and degradation of the 170 kDa isoform of topoisomerase II in amsacrine-resistant human leukemia cells.

Resistance to amsacrine in HL-60/AMSA is 50-100 fold compared to the parental HL-60/S cells. Synthesis and phosphorylation of topoisomerase II (TOPO II) were 2-3 fold lower in HL-60/AMSA compared to HL-60/S cells metabolically labelled with [32P]-orthophosphoric acid or [35S]-L-methionine. Incubating cells in radiolabel-free media following metabolic labelling for 4 hr revealed: (a) dephosphorylation of topoisomerase II at 4 hr was 70% and 20% in HL-60/S and HL-60/AMSA cells, respectively; and (b) degradation of topoisomerase II at 4 hr was 40% and 10% in HL-60/S and HL-60/AMSA cells, respectively, while at 8 hr degradation was 80% and 50% in HL-60/S and HL-60/AMSA cells, respectively. The magnitude of topoisomerase II band depletion in immunoprecipitates of amsacrine-treated cells labelled with [35S]-L-methionine or [32P]-orthophosphoric acid, correlated with the differential amsacrine sensitivity of HL-60/S and HL-60/AMSA cells, suggesting that the amount of newly synthesized and phosphorylated topoisomerase II may be contributing to amsacrine resistance. Thus, the attenuated synthesis and phosphorylation of TOPO II in HL-60/AMSA may contribute to the resistance of these cells to amsacrine.