Partial selective inhibition of HIV-1 reverse transcriptase and human DNA polymerases γ and β by thiated 3′-fluorothymidine analogue 5′-triphosphates

Abstract

a b s t r a c t 3 -Deoxy-3 -fluorothymidine (FLT, alovudine ® ) belongs to the most potent agents inhibiting HIV-1 replication. Its 5 -triphosphate (FLTTP) is a potent inhibitor of HIV-1 reverse transcriptase (HIV RT). Unfortunately, FLT exerts substantial hematologic toxicity both in vitro and in vivo. It was suggested that this toxicity may be related to inhibition of human DNA polymerases, especially mitochondrial DNA polymerase ␥, by nucleoside analogue 5 -triphosphates leading to termination of DNA synthesis and mitochondrial dysfunction. To decrease the toxicity of FLT, its thiated analogues, 4-SFLT and 2-SFLT, were previously synthesized and shown to be potent inhibitors of HIV-1 with low in vitro cytotoxicity. To explain this phenomenon in the present study the synthesis of 5 -triphosphates of thiated FLT analogues was undertaken and their interaction with recombinant HIV-1 RT and human DNA polymerases ␥ (pol ␥) and ␤ (pol ␤) was investigated. It was shown that 3 -deoxy-3 -fluoro-4-thiothymidine 5 -triphosphate (4-SFLTTP) and 3 -deoxy-3 -fluoro-2-thiothymidine 5 -triphosphate (2-SFLTTP) were, similarly to FLTTP, potent competitive inhibitors of HIV-1 RT, with K Of special interest is that 2-SFLTTP, showing syn conformation, is a less potent inhibitor of human mitochondrial pol ␥ than 4-SFLTTP and FLTTP, both in the anti conformation, and has a higher inhibitory activity against HIV-1 RT than 4-SFLTTP. Moreover, the parent nucleoside 2-SFLT possessing the syn conformation shows a more potent anti-HIV-1 activity and a better selectivity index than its 4-thio isomer in the anti conformation (Matthes et al., 1989;Poopeiko et al., 1995), 2-SFLT is a potent and selective anti-HIV-1 agent with the selectivity index 4-fold higher than that of FLT.Findings regarding the mechanisms of antiviral and cytotoxic activities of FLT and its thioanalogues are discussed.

Advances in Nucleoside and Nucleotide Analogues in Tackling Human Immunodeficiency Virus and Hepatitis Virus Infections

“…In stark comparison to its predecessor, 2‐SFLT showed a near 1000‐fold decrease in cytotoxicity with no signs of mitochondrial toxicity and still showed an EC 50 value of 0.269 μM [55] . This retention of anti‐HIV‐1 activity without the basic drawback of mitochondrial damage is an exceptional improvement on its parent molecule [55] …”

“…[55] In stark comparison to its predecessor, 2-SFLT showed a near 1000-fold decrease in cytotoxicity with no signs of mitochondrial toxicity and still showed an EC 50 value of 0.269 μM. [55] This retention of anti-HIV-1 activity without the basic drawback of mitochondrial damage is an exceptional improvement on its parent molecule. [55] Censavudine/festinavir/BMS-986001/OBP-601 or 2',3'-didehydro-3'-deoxy-4'-ethynylthymidine (or 4'-ethynyl stavudine/4'ethynyl-d4T) is currently under Phase IIb of clinical trials.…”

“…It traces its lineage from the flawed alovudine (FLT) which was withdrawn from clinical trials due to its hematologic toxicity [54] . To counter these drawbacks of FLT and its analogues, in the most recent study, carbonyl oxygen on the thymine ring was replaced with a sulfur atom, thus forming FLT's successor 3‐deoxy‐3‐fluoro‐2‐thiothymidine (2‐SFLT) [55] . In stark comparison to its predecessor, 2‐SFLT showed a near 1000‐fold decrease in cytotoxicity with no signs of mitochondrial toxicity and still showed an EC 50 value of 0.269 μM [55] .…”

“…To counter these drawbacks of FLT and its analogues, in the most recent study, carbonyl oxygen on the thymine ring was replaced with a sulfur atom, thus forming FLT's successor 3‐deoxy‐3‐fluoro‐2‐thiothymidine (2‐SFLT) [55] . In stark comparison to its predecessor, 2‐SFLT showed a near 1000‐fold decrease in cytotoxicity with no signs of mitochondrial toxicity and still showed an EC 50 value of 0.269 μM [55] . This retention of anti‐HIV‐1 activity without the basic drawback of mitochondrial damage is an exceptional improvement on its parent molecule [55] …”

Balancing Antiviral Potency and Host Toxicity: Identifying a Nucleotide Inhibitor with an Optimal Kinetic Phenotype for HIV-1 Reverse Transcriptase

“…Previous data on the interactions of these inhibitors with WT pol ␥ were limited to IC 50 of 2.7 M (Cheng et al, 1987) and K i of 50 nM (Wiń ska et al, 2010) for FLT-TP and IC 50 of 100 M for Ed4T-TP [100-fold higher than that for d4T-TP (Yang et al, 2007)]. Steady-state kinetic data reflect only the ratelimiting step of catalysis, which is product release for many polymerases, including WT pol ␥.…”