Two crucial enzymes involved in nucleic acid metabolism and cellular proliferation are targeted for inhibition by synthetic nucleoside analogs that are also aimed to serve as mechanistic probes. The cellular enzyme S-Adenosyl-L-homoeysteine (AdoHey) hydrolase affects cleavage of AdoHcy to adenosine and L-homocysteine. Since AdoHcy is a potent feedback inhibitor of crucial transmethylation enzymes, the design of inhibitors of AdoHcy hydrolase represents a rational approach for anticancer and antiviral chemotherapy. The major objective is to develop analogs of AdoHcy with 5',6'-olefin (or halovinyl) moieties incorporated in place of the sulfur atom. Such analogs should be substrates for the 'oxidative' and/or 'hydrolytic' activity of the enzyme but must not undergo elimination of the homocysteine surrogate. X-ray structures of such """"""""frozen"""""""" intermediates might provide important, but not yet available, information regarding key residues in the protein and their interactions with homocysteine moiety and/or the sequestered water molecule. Ribonueleofide reduetases (RNRs) are enzymes that execute 2'-deoxygenation of ribonucleotides in unique de novo biosynthetic pathways to DNA monomers. RNRs present attractive chemotherapeutic targets for intervention with replication of cancer cells and proliferation of viruses. The major objective is synthesis of geminal 2'-spirodifluorocyclopropyl nucleoside analogs as radical and ionic mechanistic probes. Ring opening of the spiro difluorocyclopropylcarbinyl radical will result in the generation of new radical species that might produce inactivation of RNR. The next target is 9-(2-thio-beta-D-arabinofuranosyl)adenine 5'-diphosphate which will be prepared in a stable mixed-disulfide form. This thionucleotides can serve as valuable probes for understanding the role of the cysteinyl 439 radical during enzymatic deoxygenation with natural substrates. It is essential to understand this mechanism to pursue the rational design of new mechanism-based inhibitor drugs. To accomplish our synthetic goals, we also plan to develop radical-mediated stannyl-, silyl- and germyldesulfonylations approaches for the synthesis of versatile, but scarcely available, (a-fluoro)vinyl stannanes, silanes and germanes. Chemical and biochemical basis for study of the proposed inhibitors and justification of the biological testing in collaboration with listed below group of distinguished scientists (at no cost to this grant application) are presented.
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