Potent Inhibition of NTPase/Helicase of the West Nile Virus by Ring-Expanded (“Fat”) Nucleoside Analogues

Abstract

A series of ring-expanded ("fat") nucleoside analogues (RENs) containing the 6-aminoimidazo[4,5-e][1,3]diazepine-4,8-dione ring system have been synthesized and screened for inhibition of NTPase/helicase of the West Nile Virus (WNV). To assess the selectivity of RENs against the viral enzymes, a truncated form of human enzyme Suv3((Delta)(1)(-)(159)) was also included in the study. Ring-expanded nucleosides 16, 17, and 19, which possess the long C(12), C(14), and C(18) side-chains, respectively, at position 6, as well as the ring-expanded heterocycle 39, which contains aralkyl substitution at position 1, were all found to have excellent profiles of activity and selectivity toward the viral versus human enzymes against the West Nile Virus (IC(50) ranging 1-10 muM). Compound 30, while being an equally potent inhibitor of WNV, was found to be somewhat less selective, whereas compound 36, which is an alpha-anomeric counterpart of 30, exhibited potent and selective inhibition of WNV (IC(50) 1-3 muM). The same compounds that showed potent inhibition of viral helicase activity completely failed to show any activity against the viral NTPase reaction even up to 500 muM. However, at concentrations >500 muM of RENs and the ATP concentrations >10 times the K(m) value of the enzyme, a significant activation of NTPase activity was observed. This activating effect underwent further dramatic enhancement (>1000%) by further increases in ATP concentration in the reaction mixture, suggesting that the viral helicase and NTPase reactions are not coupled. A tentative mechanistic model has been proposed to explain the observed results.

Understanding Helicases as a Means of Virus Control

“…Such compounds would not competitively inhibit ATP hydrolysis, but would modulate ATP hydrolysis and/or unwinding by binding an allosteric site. Several such compounds recently have been reported [186][187][188]. Representatives are shown along with HMC-HO4 in Fig.…”

“…[189] for a review), have also recently been examined as potential inhibitors of the WNV, JEV, and HCV helicases [187,188]. At least five different RENs inhibit WNV, one inhibits JEV, but none inhibit HCV helicase catalyzed DNA or RNA unwinding.…”

RNA helicases in infection and disease

“…220,221 Ring-expanded nucleoside and nucleotide analogs have been shown to selectively inhibit HCV, West Nile Virus, and Japanese encephalitis virus NS3 helicases in vitro without affecting the activity of a control host helicase. 222,223 Interestingly, different derivatives of these compounds showed differential effects on the three viral helicases tested, suggesting that selective inhibition is possible despite their structural similarity. The mechanism of inhibition by these and other nucleosides may involve an unidentified allosteric nucleoside binding site on the helicases.…”

Chemical and biological effects of substitution of the 2-position of ring-expanded (‘fat’) nucleosides containing the imidazo[4,5-e][1,3]diazepine-4,8-dione ring system: The role of electronic and steric factors on glycosidic bond stability and anti-HCV activity

“…Besides, since the target Flaviviridae are RNA viruses, synthesis of ribose nucleoside analogues as potential inhibitors would only be logical, notwithstanding the potent in vitro anti-HCV/WNV activities of a few 2′-deoxyribose analogues of RENs. 1,2 To this end, we synthesized the required diester precursors in the ribose series, namely ZP-115 and ZP-77, by standard Vorbrüggen ribosylation [17][18][19] of ZP-32 and ZP-72, respectively (Scheme 5). Whereas an attempted condensation reaction of ZP-115 with N-octylguanidine resulted in the formation of the deglycosylated, ring-closed heterocycle (ZP-16), the conversion of the 2-bromo group of ZP-77 into the corresponding p-methoxyphenyl derivative (ZP-108) via Suzuki coupling, 15 proceeded with no problem.…”

“…12,13 This unwinding activity is essential for the virus replication, and requires the hydrolysis of a molecule of nucleoside-5′-triphosphate (NTP), which is catalyzed by the viral NTPase. 1,2 While most of the active anti-Flaviviridae compounds were nucleoside analogues bearing a ribosyl or 2-deoxyribosyl sugar moiety at the 1-position of the heterocycle, there were also a few active compounds that simply possessed an aralkyl moiety in place of a sugar. This latter observation evoked our interest in exploring the structure-activity relationships (SAR) in various heterocyclic aglycons of the above nucleosides with an aralkyl moiety substituting for the sugar.…”