SPECIFICITY OF INTERCALATION REACTIONS

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A wide variety of compounds bind to DNA by intercalation. While the average affinity of these intercalators for DNA is similar, not all intercalators are effective in the treatment of cancer. The molecular basis for this behavior is sought. What makes an intercalating compound an effective cancer drug? My hypothesis is that the intercalators effective in cancer chemotherapy recognize some feature of the DNA in proliferating cells that the less effective intercalators do not, either a particular DNA conformation or sequence. Previous work on the cancer drug daunomycin indicated that one possible distinguishing trait is the ability to discriminate "free" DNA from DNA structured into nucleosomes. The proposed research will explore the nucleosome as a drug receptor in three ways. First, the binding of a wide variety of intercalators to "free" DNA and to nucleosomes will be studied, to see if the clinically effective compounds show discriminating behavior analogous to daunomycin. Second, stopped-flow and temperature-jump studies will be used to develop plausible reaction mechanisms for the binding of ethidium and daunomycin to nucleosomes. Finally, photochemical crosslinking experiments will be performed to localize drug binding sites within the nucleosomal DNA, in order to establish if all possible intercalation sites are freely accessible. In a separate aspect of the proposed research, the sequence specificity of anthracycline antibiotics widely used in cancer chemotherapy, will be explored using photochemical crosslinking, modern "footprinting" techniques, and affinity chromatography. Finally, the influence of intercalators on the B to Z transition will be studied, to define the role of intercalators as allosteric effectors of DNA conformational changes. The information obtained from these studies will enhances our fundamental understanding of an important class of ligand-DNA interactions, and should provide a foundation for the rational design of new chemotherapeutic compounds of greater specificity and potency.