Prodrugs of γ‐Alkyl‐Modified Nucleoside Triphosphates: Improved Inhibition of HIV Reverse Transcriptase

Abstract

The development of nucleoside triphosphate prodrugs is one option to apply nucleoside reverse transcriptase inhibitors.H erein, we report the synthesis and evaluation of d4TTP analogues,i nw hich the g-phosphate was modified covalently by lipophilic alkylr esidues,a nd acyloxybenzyl prodrugs of these g-alkyl-modified d4TTPs,w ith the aim of delivering of g-alkyl-d4TTP into cells.S elective formation of g-alkyl-d4TTP was proven with esterase and in CD4 +-cell extracts.I nc ontrast to d4TTP, g-alkyl-d4TTPs proved highly stable against dephosphorylation. Primer extension assays with HIV reverse transcriptase (RT) and DNA-polymerases a, b or g showed that g-alkyl-d4TTPs were substrates for HIV-RT only.I na ntiviral assays,c ompounds were highly potent inhibitors of HIV-1 and HIV-2 also in thymidine-kinasedeficient T-cell cultures (CEM/TK À). Thus,t he intracellular delivery of such g-alkyl-nucleoside triphosphates mayp otentially lead to nucleoside triphosphates with ahigher selectivity towards the viral polymerase that can act in virus-infected cells.

Membrane Permeable, Bioreversibly Modified Prodrugs of Nucleoside Diphosphate-γ-Phosphonates

“…It should be added that we proved that the γ-double bioreversibly masked Tri PPP ro-compounds 6 did not act as substrates for DNA polymerases, for example, HIV-RT or DNA pol β . In contrast, recently, we have shown that NTP analogues such as γ-alkylketobenzyl-d4TTP and γ-(alkyl-C18)-d4TTPs 9 that comprised one non-hydrolyzable lipophilic alkyl group were substrates for HIV-RT, and d4TMP 2 was incorporated in a primer extension assay into the primer strand while these compounds were nonsubstrates for cellular DNA polymerases α, β, and γ . More importantly, prodrug versions of compounds 9 such as γ-(AB;alkyl)-d4TTPs 8 showed even higher antiviral activity against HIV-2 in TK-deficient cell assays (CEM/TK – cells).…”

“…We disclosed recently a unique pronucleotide approach based on partially masking the negative charges and improving the lipophilicity of nucleoside triphosphates, namely, the Tri PPP ro-approach 5 . − At first, Tri PPP ro-compounds 6 comprising two lipophilic and bioreversible acyloxybenzyl (AB) masking groups at the γ-phosphate and d4T 1 as a nucleoside analogue were disclosed . The successful formation of a nucleoside triphosphate inside cells was proven in a cell uptake study involving fluorescent nucleoside analogues .…”

Photocaged and Mixed Photocaged Bioreversible‐Protected ATP Derivatives as Tools for the Controlled Release of ATP

“…As we have demonstrated earlier, γ‐modified nucleoside triphosphates can be synthesized in a three‐step protocol, starting from a nucleoside monophosphate and an appropriately masked H ‐phosphonate. [ 24 ]…”

“…As we have demonstrated earlier, γ-modified nucleoside triphosphates can be synthesized in a three-step protocol, starting from a nucleoside monophosphate and an appropriately masked H-phosphonate. [24] Following this protocol, the H-phosphonate was first activated by oxidative chlorination and subsequently reacted with inorganic phosphate to yield a non-symmetric pyrophosphate. Next, after stepwise activation this pyro-phosphate derivative was coupled with a nucleoside monophosphate, yielding the γ-masked nucleoside triphosphate.…”

Modified nucleoside triphosphates in bacterial research for in vitro and live-cell applications

“…181 The TriPPPro approach designed by Meier and co-workers relies on the addition of a lipophilic acyloxybenzyl moiety to two of the nucleophilic oxygen atoms of the NTP g-phosphate for successful internalization into cells. [182][183][184] The presence of remaining charged oxygen atoms on the aand b-phosphates is balanced by adjusting the lipophilicity of the acyloxybenzyl moiety. The advantage of this strategy is that only one hydrolysis step is required by the cells to release the active NTP, bypassing the phosphorylation metabolism steps.…”