Antiviral potencies of BV-araU and related nucleoside analogues against varicella-zoster virus in different cell lines

1-beta-D-Arabinofuranosyl-E-5-(2-bromovinyl)uracil (BV-araU) and nine other antiherpesviral nucleoside analogues were compared for their potencies against four strains of varicella-zoster virus (VZV) on three different cell lines: HEL cells, Vero cells, and MS cells established from a human malignant schwannoma. In contrast to the activity against herpes simplex virus type 1 previously reported, BV-araU showed extremely marked antiviral activity against VZV even on Vero cells. ED50, 50% plaque reduction dose, of BV-araU for VZV was 0.20-3.1 and 0.14-0.63 ng/ml on Vero cells and on HEL cells, respectively. Potency of BV-araU on MS cells was similar to that on these cell lines. There was not significant variation in anti-VZV activities of other nucleoside analogues on these three different cell lines except a few combinations of VZV strain and test compound.     

Antiviral Activity of 1-β-D-Arabinofuranosyl-E-5-(2-Bromovinyl)Uracil against Thymidine Kinase Negative Strains of Varicella-Zoster Virus

Mechanism of antiviral activity of 1-β-d -arabinofuranosyl-E-5-(2-bromovinyl)uracil (BV-araU) against the YSR strain of varicella-zoster virus (VZV), which is a mutant derived from the wild YS strain and is completely deficient in viral thymidine kinase (TK), was searched in comparison with antiviral activity of other thymidine analogues, guanosine analogue and thymidylate synthase (TS) inhibitor in human embryo lung fibroblast cells. Thymidine analogues, such as BV-araU, 5-iododeoxyuridine (IUDR), 1-β-d -arabinofuranosylthymine (araT), and guanosine analogue, such as 9-(2-hydroxyethoxymethyl)guanine (ACV), showed higher antiviral activity to the YS strain than to the YSR strain. Though, BV-araU also had the antiviral activity of a microgram level against the YSR strain. In contrast to these results, TS inhibitor, 5-fluorodeoxyuridine (FUDR), had higher antiviral activity to the YSR strain than to the YS strain. Highly synergistic antiviral activities of FUDR to the YS strain and the YSR strain were observed in combination with IUDR, araT, or ACV. However, weakly synergistic or additive inhibition to the YSR strain was shown in combination of BV-araU and FUDR, in spite of highly synergistic effect of this combination to the YS strain. The viral and cellular TS activity was partially inhibited by BV-araU monophosphate, but not by BV-araU. These results indicate that BV-araU is converted into BV-araU monophosphate by cellular TK, and the inhibition of TS activity by BV-araU monophosphate in the YSR strain-infected cells results in the suppression of viral replication.     

Enhancement of Antiviral Activity of Human Alpha-Defensin 5 against Herpes Simplex Virus 2 by Arginine Mutagenesis at Adaptive Evolution Sites

Herpes simplex virus 2 (HSV-2) infection is still one of the common causes of sexually transmitted diseases worldwide. The prevalence of HSV strains resistant to traditional nucleoside antiviral agents has led to the development of novel antiviral drugs. Human alpha-defensin 5 (HD5), a kind of endogenous antimicrobial peptide expressed in the epithelia of the small intestine and urogenital tract, displays natural antiviral activity. Based on arginine-rich features and adaptive evolution characteristics of vertebrate defensins, we conducted a screen for HD5 derivatives with enhanced anti-HSV-2 activity by a single arginine substitution at the adaptive evolution sites. Cell protection assay and temporal antiviral studies showed that HD5 and its mutants displayed affirmatory but differential anti-HSV-2 effects in vitro by inhibiting viral adhesion and entry. Inspiringly, the E21R-HD5 mutant had significantly higher antiviral activity than natural HD5, which is possibly attributed to the stronger binding affinity of the E21R-HD5 mutant with HSV-2 capsid protein gD, indicating that E21R mutation can increase the anti-HSV-2 potency of HD5. In a mouse model of lethal HSV-2 infection, prophylactic and/or therapeutic treatment with E21R-HD5 via intravaginal instillation remarkably alleviated the symptoms and delayed disease progress and resulted in about a 1.5-fold-higher survival rate than in the HD5 group. Furthermore, the E21R variant exhibited a 2-fold-higher antiviral potency against HIV-1 over parental HD5 in vitro. This study demonstrates that arginine mutagenesis at appropriate evolution sites may significantly enhance the antiviral activity of HD5, which also paves a facile way to search for potent antiviral drugs based on natural antimicrobial peptides.     

Inhibition of DNA synthesis in varicella-zoster virus-infected cells by BV-araU.

The inhibitory effect of BV-araU on DNA synthesis in human embryonic lung cells infected with varicella-zoster virus (VZV) or herpes simplex virus type 1 (HSV-1) was compared with that of acyclovir. Cellular uptake of [3H]thymidine and its incorporation into DNA was markedly stimulated by the infection with VZV or HSV-1, suggesting that the incorporation was mainly due to viral DNA synthesis. DNA synthesis in VZV-infected cells was dose-dependently suppressed by BV-araU and acyclovir, although cellular uptake of [3H]thymidine decreased in cells treated with a high concentration of drugs for an extended time. DNA synthesis in HSV-1-infected cells was also markedly inhibited by both drugs in a dose-dependent manner, without affecting cellular uptake of [3H]thymidine. The concentration of drugs inhibiting DNA synthesis was well correlated to their in vitro anti-VZV and anti-HSV-1 activities. The inhibitory concentration of BV-araU for DNA synthesis in VZV-infected cells was one-thousandth of that of acyclovir. Our results suggest that the antiviral action of BV-araU against VZV and HSV-1 is based on the inhibition of DNA synthesis in herpesvirus-infected cells.     

Anti-herpes simplex virus efficacies of 2-aminobenzamide derivatives as novel HSP90 inhibitors

After the widespread use of the acyclic purine nucleoside analogues for therapy of herpes simplex virus (HSV) infection since the 1980s, new antiviral strategies are urgently needed to counter the emergence of drug-resistant clinical isolates. In this report, we define the anti-HSV efficacies of three optimized 2-aminobenzamide derivatives in vitro and in vivo. The synthetic analogues SNX-25a, SNX-2112 and SNX-7081, which selectively bind to the N-terminal ATP pocket of heat shock protein 90 (HSP90), exhibited significant anti-HSV-1 and HSV-2 activities at non-cytotoxic concentrations in Vero cells, with EC(50) values close to that of acyclovir (ACV). The in vivo antiviral potentials were then confirmed using a herpes simplex keratitis (HSK) rabbit model, where eye gels containing 0.1% or 0.025% SNX-25a displayed the highest efficacies against HSV-1 infection, which were better than that obtained with 0.1% ACV. SNX-2112 and SNX-7081 gels were also effective against HSV-1 with different magnitude of activities. Our results for the first time confirmed the anti-HSV efficacies of these 2-aminobenzamide derivatives and suggest that with alternative mechanisms of action these novel HSP90 inhibitors, especially SNX-25a, could be potent as new anti-HSV clinical trial candidates.     

Potentiating effect of mizoribine on the anti-herpes virus activity of acyclovir

Pharmacological induction of low deoxyribonucleoside triphosphate (dNTP) levels in virus-infected cells could result in an increased antiviral effectiveness of some selective antiviral nucleoside analogues. That could be exploited as a new combined strategy in the treatment of herpes virus infections. From this point of view the alteration of antiherpes activity of acyclovir (ACV) in combination with mizoribine (N'-[beta-D-ribofuranosyl]-5-hydroxyimidazole-4-carboxamide) (MZR), an inhibitor which lowers the intracellular pool of dGTP, was studied. MZR applied alone at non-toxic concentrations had no effect on herpes simplex virus type 1 (HSV-1) replication in human embryonic skin-muscle fibroblasts (HESMF). The combination of MZR and ACV acts synergistically, as measured by the virus yield assay in the above mentioned system. The potentiating effect of MZR on the anti-HSV-1 activity of ACV was reversed by guanosine (Guo). In this case dGTP could be considered as the "key metabolite" responsible for the higher effectivity of the combination of drugs.     

Chemotherapy of varicella-zoster virus by a novel class of highly specific anti-VZV bicyclic pyrimidine nucleosides

(E)-5-(2-bromovinyl)-2'-deoxyuridine (BVDU) is a potent inhibitor of herpes simplex virus type 1 (HSV-1) and varicella-zoster virus (VZV). Its mechanism of action is based on a specific conversion to its 5'-mono- and 5'-diphosphate derivative by HSV-1- and VZV-encoded thymidine kinase, and after further conversion to its 5'-triphosphate derivative, inhibition of the viral DNA polymerase and eventual incorporation into the viral DNA. Recently, a new structural class of bicyclic pyrimidine nucleoside analogues (designated BCNAs) with highly specific and selective anti-VZV activity in cell culture has been discovered. The compounds need a long alkyl or alkylaryl side-chain at the base moiety for pronounced biological activity. This property makes these compounds highly lipophilic. They are also endowed with fluorescent properties when exposed to light with short UV wavelength. In striking contrast to BVDU, the members of this class of compounds are active only against VZV, but not against any other virus, including the closely related HSV-1, HSV-2 and cytomegalovirus. The most active compounds inhibit VZV replication at subnanomolar concentrations and are not toxic at high micromolar concentrations. The compounds lose their antiviral activity against thymidine kinase (TK)-deficient VZV strains, pointing to a pivotal role of the viral TK in their activation (phosphorylation). Kinetic studies with purified enzymes revealed that the compounds were recognized by VZV TK as a substrate, but not by HSV-1 TK, nor by cytosolic or mitochondrial TK. VZV TK is able to phosphorylate the test compounds not only to their corresponding 5'-mono- but also to their 5'-diphosphate derivatives. These data may readily explain and rationalize the anti-VZV selectivity of the BCNAs. There is no clear-cut correlation between the antiviral potency of the compounds and their affinity for VZV TK, pointing to a different structure/activity relationship of the eventual antiviral target of these compounds. The compounds are stable in solution and, in contrast to BVDU, not susceptible to degradation by thymidine phosphorylase. The bicyclic pyrimidine nucleoside analogues represent an entirely new class of highly specific anti-VZV compounds that should be further pursued for clinical development.     

Current status of natural products from plants as anti-herpes simplex virus 1 agents

Nucleoside analogues have been the mainstay of clinical treatment of herpes simplex virus 1 (HSV-1) infections since their development. However, the emergence of drug resistant strains has underlined the urgency of the discovery of novel anti-HSV-1 drugs. Natural products, which provided many novel drug leads, are known to be an important source of anti-HSV-1 agents. Herein, we present an overview of natural products with anti-HSV-1 activities isolated from a variety of plants reported in recent years. Several different compounds, mainly belonging to the three groups of polysaccharides, polyphenols and terpenes, showed antiviral effects against HSV-1, indicating their potential to be promising anti-HSV-1 agents.     

ACYCLIC/CARBOCYCLIC GUANOSINE ANALOGUES AS ANTI-HERPESVIRUS AGENTS

Several guanosine analogues, i.e. acyclovir (and its oral prodrug valaciclovir), penciclovir (in its oral prodrug form, famciclovir) and ganciclovir, are widely used for the treatment of herpesvirus [i.e. herpes simplex virus type 1 (HSV-1), and type 2 (HSV-2),varicella-zoster virus (VZV) and/or human cytomegalovirus (HCMV)] infections. In recent years, several new guanosine analogues have been developed, including the 3-membered cyclopropylmethyl and-methenyl derivatives (A-5021 and synguanol) and the 6-membered D-and L-cyclohexenyl derivatives. The activity of the acyclic/carbocyclic guanosine analogues has been determined against a wide spectrum of viruses, including the HSV-1, HSV-2, VZV, HCMV, and also human herpesviruses type 6 (HHV-6), type 7 (HHV-7) and type 8 (HHV-8), and hepatitis B virus (HBV). The new guanosine analogues (i.e. A-5021 and D- and L-cyclohexenyl G) were found to be particularly active against those viruses (HSV-1, HSV-2, VZV) that encode for a specific thymidine kinase (TK), suggesting that their antiviral activity (at least partially) depends on phosphorylation by the virus-induced TK. Marked antiviral activity was also noted with A-5021 against HHV-6 and with D- and L-cyclohexenyl G against HCMV and HBV. The antiviral activity of the acyclic/carbocyclic nucleoside analogues could be markedly potentiated by mycophenolic acid, a potent inhibitor of inosine 5′-monophosphate (IMP) dehydrogenase. The new carbocyclic guanosine analogues (i.e. A-5021 and D- andL-cyclohexenyl G) hold great promise, not only as antiviral agents for the treatment of herpesvirus infections, but also an antitumor agents for the combined gene therapy/chemotherapy of cancer, provided that (part of) the tumor cells have been transduced by the viral (HSV-1, VZV) TK gene.     

Acyclic/carbocyclic guanosine analogues as anti-herpesvirus agents

Several guanosine analogues, i.e. acyclovir (and its oral prodrug valaciclovir), penciclovir (in its oral prodrug form, famciclovir) and ganciclovir, are widely used for the treatment of herpesvirus [i.e. herpes simplex virus type 1 (HSV-1), and type 2 (HSV-2), varicella-zoster virus (VZV) and/or human cytomegalovirus (HCMV)] infections. In recent years, several new guanosine analogues have been developed, including the 3-membered cyclopropylmethyl and -methenyl derivatives (A-5021 and synguanol) and the 6-membered D- and L-cyclohexenyl derivatives. The activity of the acyclic/carbocyclic guanosine analogues has been determined against a wide spectrum of viruses, including the HSV-1, HSV-2, VZV, HCMV, and also human herpesviruses type 6 (HHV-6), type 7 (HHV-7) and type 8 (HHV-8), and hepatitis B virus (HBV). The new guanosine analogues (i.e. A-5021 and D- and L-cyclohexenyl G) were found to be particularly active against those viruses (HSV-1, HSV-2, VZV) that encode for a specific thymidine kinase (TK), suggesting that their antiviral activity (at least partially) depends on phosphorylation by the virus-induced TK. Marked antiviral activity was also noted with A-5021 against HHV-6 and with D- and L-cyclohexenyl G against HCMV and HBV. The antiviral activity of the acyclic/carbocyclic nucleoside analogues could be markedly potentiated by mycophenolic acid, a potent inhibitor of inosine 5'-monophosphate (IMP) dehydrogenase. The new carbocyclic guanosine analogues (i.e. A-5021 and D- and L-cyclohexenyl G) hold great promise, not only as antiviral agents for the treatment of herpesvirus infections, but also an antitumor agents for the combined gene therapy/chemotherapy of cancer, provided that (part of) the tumor cells have been transduced by the viral (HSV-1, VZV) TK gene.     

Design and Synthesis of Novel 1′,3′-Dioxolane 5′-Deoxyphosphonic Acid Purine Analogues as Potent Antiviral Agents

Electronic parameters of 1′,3 ′-oxygen play significant roles in steering the conformation of nucleoside phosphonic acid analogues. To investigate the relationship of two oxygen atoms with antiviral enhancement, novel 1′,3 ′-dioxolane 5 ′-deoxyphosphonic acid purine analogues were synthesized via de novo acyclic stereoselective route from acrolein and glycolic acid. The synthesized nucleoside phosphonic acid analogues 14 and 19 were subjected to antiviral screening against several viruses, such as HIV-1, HSV-1, HSV-2, and HCMV. The guanine analogue 19 exhibits in vitro anti-HIV-1 activity similar to that of 9-[2-(phosphonomethoxy)ethyl]adenine (PMEA) in MT-4 cells.     

Synthesis and Antiviral Activity Evaluation of 2′,5′,5′-Trifluoro-Apiosyl Nucleoside Phosphonic Acid Analogs

Racemic synthesis of novel 2′,5′,5′-trifluoro-apiose nucleoside phosphonic acid analogs were performed as potent antiviral agents. Phosphonation was performed by direct displacement of triflate intermediate with diethyl (lithiodifluoromethyl) phosphonate to give the corresponding (α,α-difluoroalkyl) phosphonate. Condensation successfully proceeded from a glycosyl donor with persilylated bases to yield the nucleoside phosphonate analogs. Deprotection of diethyl phosphonates provided the target nucleoside analogs. An antiviral evaluation of the synthesized compounds against various viruses such as HIV, HSV-1, HSV-2, and HCMV revealed that the pyrimidine analogues have significant anti-HCMV activity.     

New antiherpetic nucleoside from a Basidiomycete

Antiviral activity was characterized from the culture broth of the Basidiomycete Macrocystidia cucumis (Pers. ex Fr.) Heim. When the stationary phase was reached (21 d), the culture broth was shown through an ELISA assay to contain antiviral activity against herpes simplex virus type 1 (HSV-1), as assessed in baby hamster kidney cells (BHK-21). Once the presence of the anti-HSV-1 activity in the culture broth was demonstrated, we proceeded with the purification and isolation of the active principle using a semi-preparative HPLC technique. The activity was associated with a purine nucleoside designated McA. This compound displayed no cytotoxicity at antivirally effective concentrations and proved to be a novel nucleoside analogue.     

Synthesis and antiviral evaluation of novel open-chain analogues of neplanocin A

Novel acyclic nucleoside analogues were designed and synthesized as open-chain analogues of neplanocin A. The coupling of the allylic bromide with purine bases using cesium carbonate afforded a series of novel acyclic nucleosides. The synthesized compounds Ia-II were evaluated for their antiviral activity against various viruses such as HIV HSV-1, HSV-2, and ECMV.     

Synthesis and in vitro anti-HSV-1 activity of a novel Hsp90 inhibitor BJ-B11

In this study, a novel Hsp90 inhibitor BJ-B11, was synthesized and evaluated for in vitro antiviral activity against several viruses. Possible anti-HSV-1 mechanisms were also investigated. BJ-B11 displayed no antiviral activity against coxsackievirus B₃ (CVB₃), human respiratory syncytial virus (RSV) and influenza virus (H1N1), but exhibited potent anti-HSV-1 and HSV-2 activity with EC₅₀ values of 0.42 ± 0.18 μM and 0.60 ± 0.21 μM, respectively. Additionally, the inhibitory effects of BJ-B11 against HSV-1 were likely to be introduced at early stage of infection. Our results indicate that BJ-B11 with alternative mechanisms of action is potent as an anti-HSV clinical trial candidate.     

Concise synthesis and antiviral activity of novel unsaturated acyclic pyrimidine nucleosides

Novel acyclic nucleoside analogues were designed and synthesized as open-chain analogues of neplanocin A. The coupling of the allylic bromide with pyrimidine bases using cesium carbonate afforded a series of novel acyclic nucleosides. The synthesized compounds 15-22 were evaluated for their antiviral activity against various viruses such as HIV, HSV-1, HSV-2, and HCMV.     

Synthesis and evaluation of anti-HSV activity of new 5-alkynyl-2'-deoxyuridines

Eight 5-alkynyl-2'deoxyuridines containing different bulky substituents have been prepared and tested against HSV-1 in Vero cells. The compounds show positive antiviral activity. There is no obvious correlation between activity and substituent size. The nature of the linker between uracil and a substituent appears to be more important for antiviral properties: nucleosides containing arylethynyl groups show higher activity.     

Fluorosubstitution and 7-alkylation as prospective modifications of biologically active 6-aryl derivatives of tricyclic acyclovir and ganciclovir analogues

A series of fluorine containing tricyclic analogues of acyclovir (ACV, 1) and ganciclovir (GCV, 2) were synthesized and evaluated for their activity against herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2) and cytostatic activity against HSV-1 thymidine kinase (TK) gene-transduced human osteosarcoma tumour cells. It was found that fluorine substitution reduced the antiviral activity, but most of the new compounds were pronounced cytostatic agents with potency and selectivity similar to those of parental ACV and GCV. Compounds 12, 13 and 16 seem to be promising as labeled substrates for (19)F NMR studies of the HSV TK-ligand interaction and/or monitoring of their metabolites in cells expressing HSV TK.     

SYNTHESIS AND EVALUATION OF ANTI-HSV ACTIVITY OF NEW 5-ALKYNYL-2′-DEOXYURIDINES

Eight 5-alkynyl-2′deoxyuridines containing different bulky substituents have been prepared and tested against HSV-1 in Vero cells. The compounds show positive antiviral activity. There is no obvious correlation between activity and substituent size. The nature of the linker between uracil and a substituent appears to be more important for antiviral properties: nucleosides containing arylethynyl groups show higher activity.     

Modifications on the heterocyclic base of ganciclovir,penciclovir, acyclovir - syntheses and antiviral properties

АBSTRACT

Esters of the antiherpetic drugs ganciclovir, penciclovir with the bile acids (cholic, chenodeoxycholic and deoxycholic) and amino acid esters of acyclovir were generated and evaluated for their in vitro antiviral activity against herpes simplex viruses type 1 and type 2 (HSV-1, HSV-2). The antiviral assays demonstrated that modified analogs of ACV and PCV are less active compared to the initial substances against HSV-1and HSV-2. CC50 for ganciclovir-deoxycholate corresponded to the CC50 of the other analogs and its activity is lower than ganciclovir. Obtained results show that tested modification do not improve bioavailability of nucleoside analogs in cells.