Potent Anti‐HIV Activity of Alkyl‐Modified DiPPro‐Nucleotides

Abstract

Two convergent approaches for synthesizing a new class of nucleoside diphosphate prodrugs bearing different nucleoside analogs are reported herein. The DiPPro‐nucleotides comprise an acyloxybenzyl group in combination with a lipophilic alkyl residue at the β‐phosphate or β‐phosphonate group, respectively. They are selectively cleaved to form their corresponding β‐alkylated nucleoside diphosphate derivatives in chemical and biological hydrolysis studies. In contrast, there is a selective but slow cleavage observed in the hydrolysis of the DiPPro‐compounds bearing two different, nonbioreversible alkyl moieties in human CD4+ T‐lymphocyte CEM/0 cell extracts. In these studies, the delivery of nucleoside monophosphates rather than nucleoside diphosphates is being observed, most likely due to a pure chemical phosphoranhydride cleavage of the β‐phosph(on)ate moiety. The antiviral evaluation of these two types of prodrugs reveals that these compounds exhibit marked anti‐HIV efficacy in HIV‐2‐infected thymidine kinase‐deficient CD4+ CEM T‐cells (CEM/TK−), with significantly better activities (up to 6700‐fold) against HIV‐2 replication than the parent nucleosides. Primer extension assays demonstrate that the β‐dialkylphosphate‐modified nucleoside derivatives, β‐monoalkylated‐diphosphates, and nucleoside diphosphates serve as substrates for HIV reverse transcriptase for the viral DNA elongation.

Antiviral Activity of Lipophilic Nucleoside Tetraphosphate Compounds

“…As expected, with d4TTP, the incorporation of d4TMP was detected, which led to the presence of the expected n+1 (26 nt) band (Figure A, lane 10). ,,,− Here, we could prove that δ-dialkylated compounds 4 and 5 , δ-monoalkylated nucleoside tetraphosphate derivatives 20 and 24 , and d4T4P were all substrates for HIV-RT (Figure A). In contrast, they all proved to be nonsubstrates for the cellular DNA polymerases α (Figure B) and γ (Figure D).…”

“…Encouraged by the previously obtained results that γ-phosphate-modified-d4TDPs 3 , γ-monoalkylated d4TTP derivatives as well as d4TDP were accepted as substrates by HIV-RT, ,,,− δ-dialkylated d4T tetraphosphate analogues 4 , 5 , δ-monoalkylated d4T tetraphosphate analogues 20 and 24 and d4T4P were investigated in primer extension assays in the presence of HIV-RT or the three different human DNA polymerases α, β and γ. D4TTP (for the compounds 4 – 9 ) and the natural thymidine triphosphate (TTP) were used as reference compounds. It was checked prior to the primer extension assay that compounds 4a , 4b , 5a , 20 , and 24 proved to be stable under the assay conditions.…”