This project involves the investigation of the metabolism of purine analogs in the procyclic and bloodstream forms of the African trypanosomes. Prior studies have shown that analogs of inosine, such as 9-deazainosine and formycin B, are very effective against these parasites as well as the related flagellated protozoans Trypanosoma cruzi and Leishmania donovani. Other analogs, such as allopurinol riboside and thiopurinol riboside, are much less effective against the former parasites when their effectiveness against the latter two pathogens is compared. In this proposal we will investigate the mechanisms responsible for the differences in toxicity of various purine analogs in the African trypanosome when compared to Leishmania and T. cruzi. These results will also be compared to what is known for these compounds in mammalian cells. These studies will encompass the following areas. Quantitative differences in the metabolism of these analogs in the African transport and specific enzymes responsible for converting these analogs to their active forms will be investigated. Secondly, the mechanism and sites of action of these drugs will be studied. In these studies the specific enzymes that appear to be affected by these drugs or their metabolites will be purified and characterized. Mutants resistant to these purine analogs will be isolated using somatic cell genetic techniques. These mutants will then be biochemically characterized and compared to the wild type with the goal of identifying specific sites involved either in the transport or activation of these drugs or the target sites that are affected by their metabolic products. Finally, we will continue our collaborative investigations with Dr. C. J. Bacchi of Pace University, New York, NY on synergism between purine analogs and polyamine inhibitors such as DFMO.
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