The lambda-Int family recombinases, Cre and Flp are inhibited by topoisomerase I targeting anticancer drugs

RF Frøhlich, BSc,a SG Hansen, BSc,a FF Andersen, MSc,a JR Olesen, MSc,a M Jayaram, PhD,b O Westergaard, PhD,a BR Knudsen, Ph.D.,a

aDepartment of Molecular and Structural Biology, University of Aarhus, Aarhus, Denmark, bSection of Molecular Genetics and Microbiology, University of Texas, Austin, TX

The topoisomerase I targeting drug class, camptothecins, are among the most promising antineoplastic compounds available today. A severe problem in the application of camptothecins, however, is the development of drug resistant cancer cells, posing the need for design of new drugs that can be used in different combinations. For this purpose it is necessary to expand the knowledge on drug action. One approach for dissecting the molecular action of the camptothecins relies on the close structural and mechanistic relationship between type IB topoisomerases and the lambda-Int recombinases. Although catalyzing different reactions, i.e. relaxation and recombination, these enzymes act by introducing transient single strand breaks in DNA using chemically identical reaction schemes. In our research we have probed the functional relationship between the lambda-Int recombinase and type IB topoisomerase active site and the mode of drug action by investigating the sensitivity of recombinases Cre and Flp towards CPT and a new structurally unrelated compound, NSC-314622. We show that catalysis mediated by the Flp recombinase is inhibited in both cleavage and religation, whereas only the religation step for the Cre recombinase is affected. For both enzymes the recombination efficiency is largely decreased upon addition of drug. These results suggest that simple members of the lambda-Int family may be exploited as a model to dissect the molecular action of the topoisomerase I directed cancer drugs. Furthermore, the recombinases are widely used for gene transfer in genetic studies in mammalian cells, and regulation of their recombinatory activity by camptothecins or related compounds would be valuable for such studies.

KEY WORDS: camptothecin, recombination, type IB topoisomerase.

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Paper presented at the International Symposium on Predictive Oncology and Intervention Strategies; Paris, France; February 9 - 12, 2002; in the section on Multidrug Resistance - MDR.