PROTEIN-ASSOCIATED DNA BREAKS AS INDICATOR OF TOPOISOMERASE INHIBITION

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DNA topoisomerases have been identified as major targets for cancer chemotherapy. Two types of topoisomerases have been isolated from eukaryotic cell nuclei. Both topoisomerases 1 & II relax DNA supercoiling and play a crucial role in DNA replication and transcription. However, topoisomerase II is the only enzyme able to anchor DNA loops to the nuclear matrix and to decatenate newly replicated DNA circles. Two families of anticancer drugs inhibit topoisomerase II: DNA intercalators, such as amsacrine (m-AMSA), adriamycin and ellipticine derivatives, and demethylepipodophyllotoxins, such as etoposide (VP-16) and teniposide (VM-26). Drug-induced topoisomerase II inhibition results in the formation of protein-associated DNA breaks, which can be detected by alkaline elution. We have shown now, that the specific topoisomerase I inhibitor, camptothecin produces also protein-associated DNA breaks. Our recent finding that the DNA breaks induced by topoisomerase inhibitors are associated with strong and poorly reversible DNA synthesis inhibition, and that aphidicolin prevents the cytotoxicity of topoisomerase inhibitors suggest that drug-induced topoisomerase complexes could exert their cytotoxic action by producing stable topoisomerase-DNA complexes which would destabilize DNA replication complexes. Finally, we have found that a human breast cancer cell line (MCF-7) resistant to adriamycin has topoisomerase 11 modifications, which are associated with the drug uptake reduction usually encountered in pleiotropic resistant cell lines.