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The aim of this proposal is to create a new class of antiviral nucleotide prodrugs, which combine the potency and selectivity of L-nucleoside inhibitors for reverse transcriptase (RT) with advantages of boranophosphate substitution (in which an oxygen of the a-phosphate is replaced with a borano (BH3) group). We hypothesize that these prodrugs will have good cellular membrane permeability and, after hydrolysis of the lipophilic carriers inside the cell, would generate the (alpha-P-borano) mono-, di- and triphosphates of the corresponding nucleoside analogs. The boronated analogs are proposed to be stable to phosphodiesterases and phosphatases, yet good substrates for nucleotide kinases and selective for viral RTs. Moreover, the (alpha-P- borano) triphosphates are proposed to be effective inhibitors of the HIV-RT ATP-dependent and pyrophosphorolytic removal of nucleotide chain terminators; and a boranophosphate linkage incorporated into viral DNA is proposed to exhibit increased stability toward repair mechanisms that contribute to drug resistance.
Our Specific Aims i nclude 1) To synthesize and characterize (alpha-P-borano) analogs of potent viral replication inhibitors, including D- and L-enantiomers of 2',3'-dideoxynucleosides (2',3'-ddN), 2',3'-dideoxy- 2',3'-didehydrothymidine (d4T), 2'-deoxy-3'-thiacytidine triphosphate (3TC), and 2',3'-dideoxy-2'- fluoroarabinonucleosides (2'-FddAraN). 2) To study these boronated analogs as substrates and inhibitors of nucleoside mono- and diphosphate kinases, viral reverse transcriptases, and mammalian polymerases; to determine structure-function relationships; to select the most potent and selective polymerase chain terminators for future pharmacological studies; and to study how well boranophosphates block the repair of viral DNA and their mechanism of blockage. 3) To design new nucleotide prodrugs by conjugating selected alpha-P-boronated analogues with appropriate functional groups to facilitate their cell penetration and chemical inactivation of target molecules; and to investigate the chemical, biochemical, and biophysical properties of the analogs that are most essential for biochemical and therapeutic applications; 4) To investigate cell uptake and stability to phosphodiesterases and phosphatases; and to collaborate with other laboratories to study the activity of the new prodrugs against human viral diseases in cell culture. We anticipate that these studies will lead to better understanding of the mechanisms of nucleoside activation, viral replication, and drug resistance, and provide a basis for the rational design of more efficient and less toxic antiviral agents.
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