In this proposal, we combine chemical and biochemical approaches to examine fundamental questions relating to the mechanism of action and toxicity of bleomycin. We recently isolated a series of nucleoside base-propenals produced by the action of bleomycin on DNA. These compounds proved highly cytotoxic and resemble bleomycin in their effects on tumor cells in culture. These observations lead to the development of a novel family of structurally-related site-directed inhibitors and suggest the possible involvement of base propenals in the toxic actions of bleomycin. We propose to explore the molecular basis by which bleomycin produces strand-scission in DNA and to define the mechanisms involved in the production of base propenals and free bases which are released during this process. Experiments have been designed to precisely identify products of reactions in which oligonucleotides of defined base sequence are cleaved by bleomycin. The action of """"""""activated"""""""" bleomycin on organic molecules will be utilized to establish chemical mechanisms involved. The reaction of ionizing radiation with DNA, a process which resembles the action of bleomycin, will also be examined. Finally, we propose a series of studies to test the hypothesis that base propenals produced by the action of bleomycin on nuclear DNA account for the cytostatic and cytotoxic properties of the drug. Results of these experiments should contribute to an understanding of radical mechanisms involved in the degradation of DNA by drugs, chemical and gamma-irradiation. They hold potential significance for the design and development of new analogs of bleomycin.
