2'-Fluoro-arabinosylpyrimidines, with or without 5-substitutions, are among the most potent antiviral agents with a broad antiviral spectrum and exhibit a low toxicity toward the host cells. Some of these agents have also shown antineoplastic potentials. These analogs are resistant to glycosidic cleavage and are effective orally. They are highly soluble and are readily distributed throughout body water including that of the central nervous system. We propose to study two antiviral candidates, 2'-F-5-ethy-arabinosyluracil (FEAU) and 2'-F-3'-deoxy-5-methyl-arabinosyluracil (FdMAU), and two antitumor agents, 2'-F-arabinosylcytosine (FAC) and 2'-F-5-F-arabinosyluracil (FFAU). These in vitro studies will be conducted to determine the mechanism of action, the basis for selectivity, metabolic fates, membrane transport and the effects of combination with others agents. For mechanism and selectivity studies we plan to: 1) compare the relative rates of incorporation and isolate and purify DNA from leukemic cells or herpes simplex virus infected cells exposed to labeled drugs and identify incorporated moieties; 2) use CsCl isopycnic centrifugation or DNA hybridization assay for determining selective inhibition of viral DNA synthesis; 3) use sucrose gradient centrifugation and gel electrophoresis for determining the distribution and size of DNA molecules that are labeled; and 4) determine specificity of these analogs to serve as substrates for thymidine kinases and the potency of triphosphates of these analogs for inhibiting DNA polymerases of mammalian cells and/or virus specified polymerase. For metabolic studies we plan to use carbon-14 or tritium labeled analogs and to use HPLC and spectrometric techniques to study their anabolic and catabolic pathways in host and viral infected cells. The oil-layer rapid separation technique will be used for transport studies. The median-effect principle developed in this and other laboratories will be used for automated quantitation of multiple drug effects in terms of synergism, summation and antagonism. Effects of cytosine-analogs on cell differentiation will also be studied. The information obtained from the above studies along with the results from the toxicological and pharmacokinetic studies will be provided to clinical colleagues for the planning and designing of phase I trials.
NIH ExPorter
NIH ExPorter