Identification of a Point Mutation in the Topoisomerase II Gene from a Human Leukemia Cell Line Containing an Amsacrine-resistant Form of Topoisomerase II

HL-60/AMSA is a human leukemia cell line that is SO- to 100-fold more resistant to the cytotoxic actions of the topoisomerase II-reactive intercalator amsacrine than is its drug-sensitive 111-hi) parent line. Previously, we have shown that the topoisomerase II from HL-60/AMSA is also resistant to inhibition by amsacrine and other intercalating agents. We therefore sought the molecular basis for the resistance of the topoi somerase II of HL-60/AMSA and, by inference, of the HL-60/AMSA line itself. We report the cloning and sequencing of the topoisomerase II genes from both the sensitive and resistant leukemia cell lines using polymerase chain reaction technology. We have identified a single base change associated with the drug-resistant form of topoisomerase II. This mutation is present in both cloned HL-60/AMSA complementary DNA and extracted HL-60/AMSA genomic DNA. A rapid assay for this mutation in clinical samples has been developed and applied to the DNA of cells from both normal volunteers and leukemia patients. Thus far, the HL-60/AMSA genotype has not been identified in the cells from any individual, suggesting that this genotype is indeed a mutation and not an alleile form of topoisomerase II. The novel assay developed will allow a rapid search for the prevalence of this mutation in clinical samples from patients with leukemia who have relapsed following intercalator therapy.