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A Temperature Sensitive Topoisomerase II Allele Confers Temperature Dependent Drug Resistance on Amsacrine and Etoposide: A Genetic System for Determining the Targets of Topoisomerase II Inhibitors¹

Cancer Pharmacology Section, Division of Hematology/Oncology, Children's Hospital of Los Angeles, Los Angeles, California 90027 [J. L. N., Y-X. L.], and Department of Biochemistry, University of Southern California Medical School, Los Angeles, California 90033 [J. L. N., Y. H.]

We have developed a simple system for determining the specific contribution of topoisomerase II targeting to the cytotoxic activity of a drug. We have constructed yeast strains that are permeable to anti-topoisomerase II drugs, carry a DNA repair mutation, rad52, and also have a temperature sensitive topoisomerase II mutation, top2-1. Strains carrying the top2-1 mutation have nearly normal topoisomerase II activity at 25°C but less than 10% of wild type activity at 36°C. We find that at a semipermissive temperature (30°C), there is sufficient topoisomerase II activity for viability; but since the topoisomerase II activity is greatly reduced, the cells are very resistant to anti-topoisomerase II drugs. Conversely, such cells are hypersensitive to the topoisomerase I inhibitor camptothecin. These results provide strong support for the model that drug stabilized DNA cleavage, rather than a lack of enzyme activity, is responsible for cell killing by eukaryotic anti-topoisomerase II agents. They also show that there is a minimum level of topoisomerase II activity in yeast that is consistent with viability but also allows a high degree of resistance to anti-topoisomerase II agents.

¹ This work was supported by Grant CA52814 from the National Cancer Institute and by the Martell Foundation for Leukemia, Cancer and AIDS.

² Special fellow of the Leukemia Society of America. To whom requests for reprints should be addressed, at Childrens Hospital, Los Angeles, Division of Hematology/Oncology, Box 54, 4650 Sunset Blvd., Los Angeles, CA 90027.

Received 7/ 3/92. Accepted 10/19/92.

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