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Functional Expression of Human Topoisomerase II in Yeast: Mutations at Amino Acids 450 or 803 of Topoisomerase II Result in Enzymes That Can Confer Resistance to Anti-Topoisomerase II Agents¹

Developmental Therapeutics Section, Division of Hematology/Oncology, Childrens Hospital, Los Angeles, California 90027 [Y. H., M. J., J. M., D. D., J. L. N.]; Department of Biochemistry and Molecular Biology, University of Southern California Medical School [Y. H., J. L. N.], Los Angeles, California 90033; and St. Jude Children's Research Hospital, Department of Molecular Pharmacology, Memphis, Tennessee 38101 [A. R., J. L. N.]

DNA topoisomerase II is the target of a variety of important antitumor agents, including etoposide, adriamycin, and amsacrine. We have constructed a system for analyzing the action of anti-topoisomerase II agents using the yeast Saccharomyces cerevisiae and have constructed vectors for expressing human topoisomerase II functionally in yeast. We have demonstrated that temperature-conditional yeast TOP2 mutants can be complemented by expression of wild-type human topoisomerase II. Furthermore, expression of human topoisomerase II in yeast results in a quantitatively unique pattern of sensitivity to amsacrine. We also have constructed mutations in human TOP2 based on previously identified mutations from a human cell line selected for resistance to teniposide. Our experiments demonstrate that mutation of either arginine 450 or proline 803 of human topoisomerase II can result in an enzyme that has altered sensitivity to anti-topoisomerase II agents, and that a human enzyme carrying both mutations confers a higher level of drug resistance than enzymes carrying either single mutation.

¹ This work was supported by NIH Grant CA52814, the Martell Foundation, and the American Lebanese Syrian Associated Charities.

² To whom requests for reprints should be addressed, at St. Jude Children's Research Hospital, Department of Molecular Pharmacology, 332 N. Lauderdale, Memphis TN 38101. Phone: (901) 495-2794; Fax: (901) 495-2176.

Received 7/21/94. Accepted 10/31/95.

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