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Antiviral Research (v.92, #1)
Antiviral Research, at its 30th anniversary, remembrances from its Editor-in-Chief
by Erik De Clercq (pp. 1-6).
Antiviral Research, at its 30th anniversary, remembrances from its Editor-in-Chief
by Erik De Clercq (pp. 1-6).
Antiviral Research, at its 30th anniversary, remembrances from its Editor-in-Chief
by Erik De Clercq (pp. 1-6).
Molecular strategies to design an escape-proof antiviral therapy by Ben Berkhout; Rogier W. Sanders (pp. 7-14).
Two antiviral approaches against the human immunodeficiency virus type 1 (HIV-1) were presented at the Antivirals Congress in Amsterdam. The common theme among these two separate therapeutic research lines is the wish to develop a durable therapy that prevents viral escape. We will present a brief overview of these two research lines and focus on our efforts to design an escape-proof anti-HIV therapy. The first topic concerns the class of HIV-1 fusion inhibitors, including the prototype T20 peptide and the improved versions T1249 and T2635, which were all developed by Trimeris–Roche. The selection of T20-resistant HIV-1 strains is a fairly easy evolutionary process that requires a single amino acid substitution in the peptide binding site of the viral envelope glycoprotein (Env) target. The selection of T1249-resistant HIV-1 strains was shown to require a more dramatic amino acid substitution in the viral Env protein, in particular the introduction of charged amino acid residues that cause resistance by charge-repulsion of the antiviral peptide. The third generation peptide T2635 remains active against all these HIV-1 escape variants because the charged residues within this peptide are “masked” by an introduced intra-helical salt bridge. This charge masking concept could facilitate the future design of escape-proof antiviral peptides. The second topic concerns the mechanism of RNA interference (RNAi) that we are currently employing to develop an antiviral gene therapy. One can make human T cells resistant to HIV-1 infection by a stable RNAi-inducing gene transfer, but the virus escapes under therapeutic pressure of a single inhibitor. Several options for a combinatorial RNAi attack to prevent viral escape will be discussed. The simultaneous use of multiple RNAi inhibitors turns out to be the most effective and durable strategy.
Keywords: HIV-1; Virus evolution; Drug resistance; Fusion inhibitor; RNA interference
Molecular strategies to design an escape-proof antiviral therapy by Ben Berkhout; Rogier W. Sanders (pp. 7-14).
Two antiviral approaches against the human immunodeficiency virus type 1 (HIV-1) were presented at the Antivirals Congress in Amsterdam. The common theme among these two separate therapeutic research lines is the wish to develop a durable therapy that prevents viral escape. We will present a brief overview of these two research lines and focus on our efforts to design an escape-proof anti-HIV therapy. The first topic concerns the class of HIV-1 fusion inhibitors, including the prototype T20 peptide and the improved versions T1249 and T2635, which were all developed by Trimeris–Roche. The selection of T20-resistant HIV-1 strains is a fairly easy evolutionary process that requires a single amino acid substitution in the peptide binding site of the viral envelope glycoprotein (Env) target. The selection of T1249-resistant HIV-1 strains was shown to require a more dramatic amino acid substitution in the viral Env protein, in particular the introduction of charged amino acid residues that cause resistance by charge-repulsion of the antiviral peptide. The third generation peptide T2635 remains active against all these HIV-1 escape variants because the charged residues within this peptide are “masked” by an introduced intra-helical salt bridge. This charge masking concept could facilitate the future design of escape-proof antiviral peptides. The second topic concerns the mechanism of RNA interference (RNAi) that we are currently employing to develop an antiviral gene therapy. One can make human T cells resistant to HIV-1 infection by a stable RNAi-inducing gene transfer, but the virus escapes under therapeutic pressure of a single inhibitor. Several options for a combinatorial RNAi attack to prevent viral escape will be discussed. The simultaneous use of multiple RNAi inhibitors turns out to be the most effective and durable strategy.
Keywords: HIV-1; Virus evolution; Drug resistance; Fusion inhibitor; RNA interference
Molecular strategies to design an escape-proof antiviral therapy by Ben Berkhout; Rogier W. Sanders (pp. 7-14).
Two antiviral approaches against the human immunodeficiency virus type 1 (HIV-1) were presented at the Antivirals Congress in Amsterdam. The common theme among these two separate therapeutic research lines is the wish to develop a durable therapy that prevents viral escape. We will present a brief overview of these two research lines and focus on our efforts to design an escape-proof anti-HIV therapy. The first topic concerns the class of HIV-1 fusion inhibitors, including the prototype T20 peptide and the improved versions T1249 and T2635, which were all developed by Trimeris–Roche. The selection of T20-resistant HIV-1 strains is a fairly easy evolutionary process that requires a single amino acid substitution in the peptide binding site of the viral envelope glycoprotein (Env) target. The selection of T1249-resistant HIV-1 strains was shown to require a more dramatic amino acid substitution in the viral Env protein, in particular the introduction of charged amino acid residues that cause resistance by charge-repulsion of the antiviral peptide. The third generation peptide T2635 remains active against all these HIV-1 escape variants because the charged residues within this peptide are “masked” by an introduced intra-helical salt bridge. This charge masking concept could facilitate the future design of escape-proof antiviral peptides. The second topic concerns the mechanism of RNA interference (RNAi) that we are currently employing to develop an antiviral gene therapy. One can make human T cells resistant to HIV-1 infection by a stable RNAi-inducing gene transfer, but the virus escapes under therapeutic pressure of a single inhibitor. Several options for a combinatorial RNAi attack to prevent viral escape will be discussed. The simultaneous use of multiple RNAi inhibitors turns out to be the most effective and durable strategy.
Keywords: HIV-1; Virus evolution; Drug resistance; Fusion inhibitor; RNA interference
A potential role for monoclonal antibodies in prophylactic and therapeutic treatment of influenza by Nicasio Mancini; Laura Solforosi; Nicola Clementi; Donata De Marco; Massimo Clementi; Roberto Burioni (pp. 15-26).
The role of humoral response in the effective control of infection by influenza viruses is well known, but the protection is usually limited to the infecting or vaccinating isolate and to few related strains. Recent studies have evidenced the existence of B-cell epitopes broadly conserved among different influenza subtypes recognized by monoclonal antibodies endowed with unprecedented broad activity. In this review, all major monoclonal antibodies directed against different influenza virus proteins are reported and their potential in the design of new anti-influenza prophylactic or therapeutic strategies is discussed.
Keywords: Monoclonal antibodies; Influenza viruses; Universal epitopes
A potential role for monoclonal antibodies in prophylactic and therapeutic treatment of influenza by Nicasio Mancini; Laura Solforosi; Nicola Clementi; Donata De Marco; Massimo Clementi; Roberto Burioni (pp. 15-26).
The role of humoral response in the effective control of infection by influenza viruses is well known, but the protection is usually limited to the infecting or vaccinating isolate and to few related strains. Recent studies have evidenced the existence of B-cell epitopes broadly conserved among different influenza subtypes recognized by monoclonal antibodies endowed with unprecedented broad activity. In this review, all major monoclonal antibodies directed against different influenza virus proteins are reported and their potential in the design of new anti-influenza prophylactic or therapeutic strategies is discussed.
Keywords: Monoclonal antibodies; Influenza viruses; Universal epitopes
A potential role for monoclonal antibodies in prophylactic and therapeutic treatment of influenza by Nicasio Mancini; Laura Solforosi; Nicola Clementi; Donata De Marco; Massimo Clementi; Roberto Burioni (pp. 15-26).
The role of humoral response in the effective control of infection by influenza viruses is well known, but the protection is usually limited to the infecting or vaccinating isolate and to few related strains. Recent studies have evidenced the existence of B-cell epitopes broadly conserved among different influenza subtypes recognized by monoclonal antibodies endowed with unprecedented broad activity. In this review, all major monoclonal antibodies directed against different influenza virus proteins are reported and their potential in the design of new anti-influenza prophylactic or therapeutic strategies is discussed.
Keywords: Monoclonal antibodies; Influenza viruses; Universal epitopes
Aprotinin and similar protease inhibitors as drugs against influenza by O.P. Zhirnov; H.D. Klenk; P.F. Wright (pp. 27-36).
Efforts to develop new antiviral chemotherapeutic approaches are focusing on compounds that target either influenza virus replication itself or host factor(s) that are critical to influenza replication. Host protease mediated influenza hemagglutinin (HA) cleavage is critical for activation of virus infectivity and as such is a chemotherapeutic target. Influenza pathogenesis involves a “vicious cycle” in which host proteases activate progeny virus which in turn amplifies replication and stimulates further protease activities which may be detrimental to the infected host. Aprotinin, a 58 amino acid polypeptide purified from bovine lung that is one of a family of host-targeted antivirals that inhibit serine proteases responsible for influenza virus activation. This drug and similar agents, such as leupeptin and camostat, suppress virus HA cleavage and limit reproduction of human and avian influenza viruses with a single arginine in the HA cleavage site. Site-directed structural modifications of aprotinin are possible to increase its intracellular targeting of cleavage of highly virulent H5 and H7 hemagglutinins possessing multi-arginine/lysine cleavage site. An additional mechanism of action for serine protease inhibitors is to target a number of host mediators of inflammation and down regulate their levels in virus-infected hosts. Aprotinin is a generic drug approved for intravenous use in humans to treat pancreatitis and limit post-operative bleeding. As an antiinfluenzal compound, aprotinin might be delivered by two routes: (i) a small-particle aerosol has been approved in Russia for local respiratory application in mild-to-moderate influenza and (ii) a proposed intravenous administration for severe influenza to provide both an antiviral effect and a decrease in systemic pathology and inflammation.
Keywords: Influenza; HA cleavage; Antivirals; Aprotinin; Protease inhibitors; Aerosol
Aprotinin and similar protease inhibitors as drugs against influenza by O.P. Zhirnov; H.D. Klenk; P.F. Wright (pp. 27-36).
Efforts to develop new antiviral chemotherapeutic approaches are focusing on compounds that target either influenza virus replication itself or host factor(s) that are critical to influenza replication. Host protease mediated influenza hemagglutinin (HA) cleavage is critical for activation of virus infectivity and as such is a chemotherapeutic target. Influenza pathogenesis involves a “vicious cycle” in which host proteases activate progeny virus which in turn amplifies replication and stimulates further protease activities which may be detrimental to the infected host. Aprotinin, a 58 amino acid polypeptide purified from bovine lung that is one of a family of host-targeted antivirals that inhibit serine proteases responsible for influenza virus activation. This drug and similar agents, such as leupeptin and camostat, suppress virus HA cleavage and limit reproduction of human and avian influenza viruses with a single arginine in the HA cleavage site. Site-directed structural modifications of aprotinin are possible to increase its intracellular targeting of cleavage of highly virulent H5 and H7 hemagglutinins possessing multi-arginine/lysine cleavage site. An additional mechanism of action for serine protease inhibitors is to target a number of host mediators of inflammation and down regulate their levels in virus-infected hosts. Aprotinin is a generic drug approved for intravenous use in humans to treat pancreatitis and limit post-operative bleeding. As an antiinfluenzal compound, aprotinin might be delivered by two routes: (i) a small-particle aerosol has been approved in Russia for local respiratory application in mild-to-moderate influenza and (ii) a proposed intravenous administration for severe influenza to provide both an antiviral effect and a decrease in systemic pathology and inflammation.
Keywords: Influenza; HA cleavage; Antivirals; Aprotinin; Protease inhibitors; Aerosol
Aprotinin and similar protease inhibitors as drugs against influenza by O.P. Zhirnov; H.D. Klenk; P.F. Wright (pp. 27-36).
Efforts to develop new antiviral chemotherapeutic approaches are focusing on compounds that target either influenza virus replication itself or host factor(s) that are critical to influenza replication. Host protease mediated influenza hemagglutinin (HA) cleavage is critical for activation of virus infectivity and as such is a chemotherapeutic target. Influenza pathogenesis involves a “vicious cycle” in which host proteases activate progeny virus which in turn amplifies replication and stimulates further protease activities which may be detrimental to the infected host. Aprotinin, a 58 amino acid polypeptide purified from bovine lung that is one of a family of host-targeted antivirals that inhibit serine proteases responsible for influenza virus activation. This drug and similar agents, such as leupeptin and camostat, suppress virus HA cleavage and limit reproduction of human and avian influenza viruses with a single arginine in the HA cleavage site. Site-directed structural modifications of aprotinin are possible to increase its intracellular targeting of cleavage of highly virulent H5 and H7 hemagglutinins possessing multi-arginine/lysine cleavage site. An additional mechanism of action for serine protease inhibitors is to target a number of host mediators of inflammation and down regulate their levels in virus-infected hosts. Aprotinin is a generic drug approved for intravenous use in humans to treat pancreatitis and limit post-operative bleeding. As an antiinfluenzal compound, aprotinin might be delivered by two routes: (i) a small-particle aerosol has been approved in Russia for local respiratory application in mild-to-moderate influenza and (ii) a proposed intravenous administration for severe influenza to provide both an antiviral effect and a decrease in systemic pathology and inflammation.
Keywords: Influenza; HA cleavage; Antivirals; Aprotinin; Protease inhibitors; Aerosol
Selective inhibition of Human Immunodeficiency Virus type 1 (HIV-1) by a novel family of tricyclic nucleosides by María-Cruz Bonache; Alessandra Cordeiro; Ernesto Quesada; Els Vanstreels; Dirk Daelemans; María-José Camarasa; Jan Balzarini; Ana San-Félix (pp. 37-44).
Nucleoside 1, with an unusual tricyclic carbohydrate moiety, specifically inhibits HIV-1 replication while being inactive against HIV-2 or other (retro) viruses. In an attempt to increase the inhibitory efficacy against HIV-1, and to further explore the structural features required for anti-HIV-1 activity, different types of modifications have been carried out on this prototype compound. These include substitution of the ethoxy group at the C-4″ position by alkoxy groups of different length, branching, conformational freedom or functionalization. In addition, the 4″-ethoxy group has been removed or substituted by other functional groups. The role of the tert-butyldimethylsilyl (TBDMS) group at the 2′ position has also been studied by preparing the corresponding 2′-deprotected derivative or by replacing it by other silyl ( tert-hexyldimethylsilyl) or acyl (acetyl) moieties. Finally, the thymine of the prototype compound has been replaced by N-3-methylthymine, uracil or thiophenyl. Some of these compounds were endowed with a 6- to 7-fold higher selectivity than the prototype 1. The tricyclic nucleosides here described represent a novel type of selective anti HIV-1 inhibitors, targeted at the HIV-1-encoded reverse transcriptase.
Keywords: Antiviral agents; AIDS; HIV-1 reverse transcriptase inhibitors; Nucleosides; Carbohydrates; Glycosylation
Selective inhibition of Human Immunodeficiency Virus type 1 (HIV-1) by a novel family of tricyclic nucleosides by María-Cruz Bonache; Alessandra Cordeiro; Ernesto Quesada; Els Vanstreels; Dirk Daelemans; María-José Camarasa; Jan Balzarini; Ana San-Félix (pp. 37-44).
Nucleoside 1, with an unusual tricyclic carbohydrate moiety, specifically inhibits HIV-1 replication while being inactive against HIV-2 or other (retro) viruses. In an attempt to increase the inhibitory efficacy against HIV-1, and to further explore the structural features required for anti-HIV-1 activity, different types of modifications have been carried out on this prototype compound. These include substitution of the ethoxy group at the C-4″ position by alkoxy groups of different length, branching, conformational freedom or functionalization. In addition, the 4″-ethoxy group has been removed or substituted by other functional groups. The role of the tert-butyldimethylsilyl (TBDMS) group at the 2′ position has also been studied by preparing the corresponding 2′-deprotected derivative or by replacing it by other silyl ( tert-hexyldimethylsilyl) or acyl (acetyl) moieties. Finally, the thymine of the prototype compound has been replaced by N-3-methylthymine, uracil or thiophenyl. Some of these compounds were endowed with a 6- to 7-fold higher selectivity than the prototype 1. The tricyclic nucleosides here described represent a novel type of selective anti HIV-1 inhibitors, targeted at the HIV-1-encoded reverse transcriptase.
Keywords: Antiviral agents; AIDS; HIV-1 reverse transcriptase inhibitors; Nucleosides; Carbohydrates; Glycosylation
Selective inhibition of Human Immunodeficiency Virus type 1 (HIV-1) by a novel family of tricyclic nucleosides by María-Cruz Bonache; Alessandra Cordeiro; Ernesto Quesada; Els Vanstreels; Dirk Daelemans; María-José Camarasa; Jan Balzarini; Ana San-Félix (pp. 37-44).
Nucleoside 1, with an unusual tricyclic carbohydrate moiety, specifically inhibits HIV-1 replication while being inactive against HIV-2 or other (retro) viruses. In an attempt to increase the inhibitory efficacy against HIV-1, and to further explore the structural features required for anti-HIV-1 activity, different types of modifications have been carried out on this prototype compound. These include substitution of the ethoxy group at the C-4″ position by alkoxy groups of different length, branching, conformational freedom or functionalization. In addition, the 4″-ethoxy group has been removed or substituted by other functional groups. The role of the tert-butyldimethylsilyl (TBDMS) group at the 2′ position has also been studied by preparing the corresponding 2′-deprotected derivative or by replacing it by other silyl ( tert-hexyldimethylsilyl) or acyl (acetyl) moieties. Finally, the thymine of the prototype compound has been replaced by N-3-methylthymine, uracil or thiophenyl. Some of these compounds were endowed with a 6- to 7-fold higher selectivity than the prototype 1. The tricyclic nucleosides here described represent a novel type of selective anti HIV-1 inhibitors, targeted at the HIV-1-encoded reverse transcriptase.
Keywords: Antiviral agents; AIDS; HIV-1 reverse transcriptase inhibitors; Nucleosides; Carbohydrates; Glycosylation
Inhibition of influenza virus-induced NF-kappaB and Raf/MEK/ERK activation can reduce both virus titers and cytokine expression simultaneously in vitro and in vivo by Ruth Pinto; Susanne Herold; Lidija Cakarova; Katrin Hoegner; Jürgen Lohmeyer; Oliver Planz; Stephan Pleschka (pp. 45-56).
Influenza virus (IV) infection can cause severe pneumonia and death. Therapeutic actions are limited to vaccines and a few anti-viral drugs. These target viral functions thereby selecting resistant variants. During replication IV activates the Raf/MEK/ERK-cascade and the transcription factor NF-kappaB. Both result in virus supportive and anti-viral effects by promoting viral genome transport for virus assembly and by inducing expression of pro-inflammatory host factors. Apart from tissue damage caused by the virus lytic replication, an imbalanced overproduction of anti-viral cytokines can cause severe lung damage as observed in human H5-type IV infections. Recently we showed that inhibition of NF-kappaB activity reduces the virus titer in vitro and in vivo. We have now analyzed whether inhibition of these pathways, allows simultaneous reduction of virus titers and virus-induced cytokines. The results show that inhibition of either pathway indeed leads to decreased virus titers and cytokine expression. This was not only true for infected permanent cells or primary mouse alveolar epithelial cells, but also in infected mice. Hereby we demonstrate for the first time in vitro and in vivo that virus titers and pro-inflammatory cytokine expression can be modulated simultaneously. This could provide a new rationale of future therapeutic strategies to treat IV pneumonia.
Keywords: Abbreviations; PR8; A/PR/8/34 (H1N1); FPV; A/FPV/Bratislava/79 (H7N7); IV; influenza A virus; FFU; foci forming units; MOI; multiplicity of infection; A549; human alveolar epithelial cellsInfluenza virus; Signal transduction; NF-kappaB; MEK-inhibition; IKK-inhibition; Cytokines
Inhibition of influenza virus-induced NF-kappaB and Raf/MEK/ERK activation can reduce both virus titers and cytokine expression simultaneously in vitro and in vivo by Ruth Pinto; Susanne Herold; Lidija Cakarova; Katrin Hoegner; Jürgen Lohmeyer; Oliver Planz; Stephan Pleschka (pp. 45-56).
Influenza virus (IV) infection can cause severe pneumonia and death. Therapeutic actions are limited to vaccines and a few anti-viral drugs. These target viral functions thereby selecting resistant variants. During replication IV activates the Raf/MEK/ERK-cascade and the transcription factor NF-kappaB. Both result in virus supportive and anti-viral effects by promoting viral genome transport for virus assembly and by inducing expression of pro-inflammatory host factors. Apart from tissue damage caused by the virus lytic replication, an imbalanced overproduction of anti-viral cytokines can cause severe lung damage as observed in human H5-type IV infections. Recently we showed that inhibition of NF-kappaB activity reduces the virus titer in vitro and in vivo. We have now analyzed whether inhibition of these pathways, allows simultaneous reduction of virus titers and virus-induced cytokines. The results show that inhibition of either pathway indeed leads to decreased virus titers and cytokine expression. This was not only true for infected permanent cells or primary mouse alveolar epithelial cells, but also in infected mice. Hereby we demonstrate for the first time in vitro and in vivo that virus titers and pro-inflammatory cytokine expression can be modulated simultaneously. This could provide a new rationale of future therapeutic strategies to treat IV pneumonia.
Keywords: Abbreviations; PR8; A/PR/8/34 (H1N1); FPV; A/FPV/Bratislava/79 (H7N7); IV; influenza A virus; FFU; foci forming units; MOI; multiplicity of infection; A549; human alveolar epithelial cellsInfluenza virus; Signal transduction; NF-kappaB; MEK-inhibition; IKK-inhibition; Cytokines
Inhibition of influenza virus-induced NF-kappaB and Raf/MEK/ERK activation can reduce both virus titers and cytokine expression simultaneously in vitro and in vivo by Ruth Pinto; Susanne Herold; Lidija Cakarova; Katrin Hoegner; Jürgen Lohmeyer; Oliver Planz; Stephan Pleschka (pp. 45-56).
Influenza virus (IV) infection can cause severe pneumonia and death. Therapeutic actions are limited to vaccines and a few anti-viral drugs. These target viral functions thereby selecting resistant variants. During replication IV activates the Raf/MEK/ERK-cascade and the transcription factor NF-kappaB. Both result in virus supportive and anti-viral effects by promoting viral genome transport for virus assembly and by inducing expression of pro-inflammatory host factors. Apart from tissue damage caused by the virus lytic replication, an imbalanced overproduction of anti-viral cytokines can cause severe lung damage as observed in human H5-type IV infections. Recently we showed that inhibition of NF-kappaB activity reduces the virus titer in vitro and in vivo. We have now analyzed whether inhibition of these pathways, allows simultaneous reduction of virus titers and virus-induced cytokines. The results show that inhibition of either pathway indeed leads to decreased virus titers and cytokine expression. This was not only true for infected permanent cells or primary mouse alveolar epithelial cells, but also in infected mice. Hereby we demonstrate for the first time in vitro and in vivo that virus titers and pro-inflammatory cytokine expression can be modulated simultaneously. This could provide a new rationale of future therapeutic strategies to treat IV pneumonia.
Keywords: Abbreviations; PR8; A/PR/8/34 (H1N1); FPV; A/FPV/Bratislava/79 (H7N7); IV; influenza A virus; FFU; foci forming units; MOI; multiplicity of infection; A549; human alveolar epithelial cellsInfluenza virus; Signal transduction; NF-kappaB; MEK-inhibition; IKK-inhibition; Cytokines
Thiated derivatives of 2′,3′-dideoxy-3′-fluorothymidine: Synthesis, in vitro anti-HIV-1 activity and interaction with recombinant drug resistant HIV-1 reverse transcriptase forms by Agnieszka Miazga; François Hamy; Séverine Louvel; Thomas Klimkait; Zofia Pietrusiewicz; Anna Kurzyńska-Kokorniak; Marek Figlerowicz; Patrycja Wińska; Tadeusz Kulikowski (pp. 57-63).
Various thiated analogues of thymine 2′,3′-dideoxy-3′-fluoronucleoside (FLT) and their 5′-monophosphates and 5′-triphosphates were prepared with the use of modified multistep procedures. The thiated analogues of FLT and FLTMP were evaluated against the wild type and drug- and multidrug-resistant strains of HIV-1, using the replicative phenotyping format of the deCIPhR assay, and showed potent inhibition of drug-resistant HIV-1 strains at low cytotoxicity. Additionally, inhibition of recombinant drug resistant forms of reverse transcriptase from single and multiple HIV-1 mutants by the synthesized 5′-triphosphates was investigated. The strongest inhibition was observed for K103N and Δ67 mutants and the most potent anti-HIV-1 activity against drug resistant strains and the lowest cytotoxicity was exerted by S4FLTMP and FLTMP which may be regarded as potential anti-HIV/AIDS agents.
Keywords: Anti HIV-1 agents; NRTI; HIV-1 mutants
Thiated derivatives of 2′,3′-dideoxy-3′-fluorothymidine: Synthesis, in vitro anti-HIV-1 activity and interaction with recombinant drug resistant HIV-1 reverse transcriptase forms by Agnieszka Miazga; François Hamy; Séverine Louvel; Thomas Klimkait; Zofia Pietrusiewicz; Anna Kurzyńska-Kokorniak; Marek Figlerowicz; Patrycja Wińska; Tadeusz Kulikowski (pp. 57-63).
Various thiated analogues of thymine 2′,3′-dideoxy-3′-fluoronucleoside (FLT) and their 5′-monophosphates and 5′-triphosphates were prepared with the use of modified multistep procedures. The thiated analogues of FLT and FLTMP were evaluated against the wild type and drug- and multidrug-resistant strains of HIV-1, using the replicative phenotyping format of the deCIPhR assay, and showed potent inhibition of drug-resistant HIV-1 strains at low cytotoxicity. Additionally, inhibition of recombinant drug resistant forms of reverse transcriptase from single and multiple HIV-1 mutants by the synthesized 5′-triphosphates was investigated. The strongest inhibition was observed for K103N and Δ67 mutants and the most potent anti-HIV-1 activity against drug resistant strains and the lowest cytotoxicity was exerted by S4FLTMP and FLTMP which may be regarded as potential anti-HIV/AIDS agents.
Keywords: Anti HIV-1 agents; NRTI; HIV-1 mutants
Thiated derivatives of 2′,3′-dideoxy-3′-fluorothymidine: Synthesis, in vitro anti-HIV-1 activity and interaction with recombinant drug resistant HIV-1 reverse transcriptase forms by Agnieszka Miazga; François Hamy; Séverine Louvel; Thomas Klimkait; Zofia Pietrusiewicz; Anna Kurzyńska-Kokorniak; Marek Figlerowicz; Patrycja Wińska; Tadeusz Kulikowski (pp. 57-63).
Various thiated analogues of thymine 2′,3′-dideoxy-3′-fluoronucleoside (FLT) and their 5′-monophosphates and 5′-triphosphates were prepared with the use of modified multistep procedures. The thiated analogues of FLT and FLTMP were evaluated against the wild type and drug- and multidrug-resistant strains of HIV-1, using the replicative phenotyping format of the deCIPhR assay, and showed potent inhibition of drug-resistant HIV-1 strains at low cytotoxicity. Additionally, inhibition of recombinant drug resistant forms of reverse transcriptase from single and multiple HIV-1 mutants by the synthesized 5′-triphosphates was investigated. The strongest inhibition was observed for K103N and Δ67 mutants and the most potent anti-HIV-1 activity against drug resistant strains and the lowest cytotoxicity was exerted by S4FLTMP and FLTMP which may be regarded as potential anti-HIV/AIDS agents.
Keywords: Anti HIV-1 agents; NRTI; HIV-1 mutants
Packaging signals in the 5′-ends of influenza virus PA, PB1, and PB2 genes as potential targets to develop nucleic-acid based antiviral molecules by Simone Giannecchini; Helen M. Wise; Paul Digard; Valeria Clausi; Edoardo Del Poggetto; Liberio Vesco; Simona Puzelli; Isabella Donatelli; Alberta Azzi (pp. 64-72).
► S-ON reproducing the packaging signal in the 5′ end of PB1 and PA segment of influenza A virus proved inhibitory. ► PB2-derived S-ON selection of an H1N1 strain generated resistant viruses with mutations in the PB1, PB2, PA and M1 segments. ► Reverse genetics study suggested that alterations to RNA function in the packaging regions of PB1 and PA were crucial. ► Other mutations induced by S-ON treatment were markedly deleterious to virus fitness. ► Packaging signals in the influenza A virus polymerase segments provide feasible targets for nucleic acid-based antivirals.In a previous study a 15-mer phosphorothioate oligonucleotide (S-ON) derived from the packaging signal in the 5′ end of segment 1 (PB2) of influenza A virus (designated 5–15b) proved markedly inhibitory to virus replication. Here we investigated whether analogous inhibitory S-ONs targeting the 5′ end of segments 2 (PB1) and 3 (PA) could be identified and whether viral resistance to S-ONs can be developed. Similar to our earlier result, 20-mer S-ONs reproducing the 5′ ends of segments 2 or 3 (complementary to the 3′-coding regions of PB1 and PA, respectively) exerted a powerful antiviral activity against a variety of influenza A virus subtypes in MDCK cells. Serial passage of the A/Taiwan/1/86 H1N1 strain in the presence of S-ON 5–15b or its antisense as5–15b analogue showed that mutant viruses with reduced susceptibility to the S-ON could indeed be generated, although the resistant viruses displayed reduced replicative fitness. Sequencing the resistant viruses identified mutations in the PB1, PB2, PA and M1 genes. Introduction of these changes into the A/PR/8/34 H1N1 strain by reverse genetics, suggested that alterations to RNA function in the packaging regions of segments 2 and 3 were important in developing resistance to S-ON inhibition. However, many of the other sequence changes induced by S-ON treatment were markedly deleterious to virus fitness. We conclude that packaging signals in the influenza A virus polymerase segments provide feasible targets for nucleic acid-based antivirals that may be difficult for the virus to evade through resistance mutations.
Keywords: Influenza virus; Virus inhibition; Antiviral resistance; Polymerases-derived S-ON; Packaging
Packaging signals in the 5′-ends of influenza virus PA, PB1, and PB2 genes as potential targets to develop nucleic-acid based antiviral molecules by Simone Giannecchini; Helen M. Wise; Paul Digard; Valeria Clausi; Edoardo Del Poggetto; Liberio Vesco; Simona Puzelli; Isabella Donatelli; Alberta Azzi (pp. 64-72).
► S-ON reproducing the packaging signal in the 5′ end of PB1 and PA segment of influenza A virus proved inhibitory. ► PB2-derived S-ON selection of an H1N1 strain generated resistant viruses with mutations in the PB1, PB2, PA and M1 segments. ► Reverse genetics study suggested that alterations to RNA function in the packaging regions of PB1 and PA were crucial. ► Other mutations induced by S-ON treatment were markedly deleterious to virus fitness. ► Packaging signals in the influenza A virus polymerase segments provide feasible targets for nucleic acid-based antivirals.In a previous study a 15-mer phosphorothioate oligonucleotide (S-ON) derived from the packaging signal in the 5′ end of segment 1 (PB2) of influenza A virus (designated 5–15b) proved markedly inhibitory to virus replication. Here we investigated whether analogous inhibitory S-ONs targeting the 5′ end of segments 2 (PB1) and 3 (PA) could be identified and whether viral resistance to S-ONs can be developed. Similar to our earlier result, 20-mer S-ONs reproducing the 5′ ends of segments 2 or 3 (complementary to the 3′-coding regions of PB1 and PA, respectively) exerted a powerful antiviral activity against a variety of influenza A virus subtypes in MDCK cells. Serial passage of the A/Taiwan/1/86 H1N1 strain in the presence of S-ON 5–15b or its antisense as5–15b analogue showed that mutant viruses with reduced susceptibility to the S-ON could indeed be generated, although the resistant viruses displayed reduced replicative fitness. Sequencing the resistant viruses identified mutations in the PB1, PB2, PA and M1 genes. Introduction of these changes into the A/PR/8/34 H1N1 strain by reverse genetics, suggested that alterations to RNA function in the packaging regions of segments 2 and 3 were important in developing resistance to S-ON inhibition. However, many of the other sequence changes induced by S-ON treatment were markedly deleterious to virus fitness. We conclude that packaging signals in the influenza A virus polymerase segments provide feasible targets for nucleic acid-based antivirals that may be difficult for the virus to evade through resistance mutations.
Keywords: Influenza virus; Virus inhibition; Antiviral resistance; Polymerases-derived S-ON; Packaging
Packaging signals in the 5′-ends of influenza virus PA, PB1, and PB2 genes as potential targets to develop nucleic-acid based antiviral molecules by Simone Giannecchini; Helen M. Wise; Paul Digard; Valeria Clausi; Edoardo Del Poggetto; Liberio Vesco; Simona Puzelli; Isabella Donatelli; Alberta Azzi (pp. 64-72).
► S-ON reproducing the packaging signal in the 5′ end of PB1 and PA segment of influenza A virus proved inhibitory. ► PB2-derived S-ON selection of an H1N1 strain generated resistant viruses with mutations in the PB1, PB2, PA and M1 segments. ► Reverse genetics study suggested that alterations to RNA function in the packaging regions of PB1 and PA were crucial. ► Other mutations induced by S-ON treatment were markedly deleterious to virus fitness. ► Packaging signals in the influenza A virus polymerase segments provide feasible targets for nucleic acid-based antivirals.In a previous study a 15-mer phosphorothioate oligonucleotide (S-ON) derived from the packaging signal in the 5′ end of segment 1 (PB2) of influenza A virus (designated 5–15b) proved markedly inhibitory to virus replication. Here we investigated whether analogous inhibitory S-ONs targeting the 5′ end of segments 2 (PB1) and 3 (PA) could be identified and whether viral resistance to S-ONs can be developed. Similar to our earlier result, 20-mer S-ONs reproducing the 5′ ends of segments 2 or 3 (complementary to the 3′-coding regions of PB1 and PA, respectively) exerted a powerful antiviral activity against a variety of influenza A virus subtypes in MDCK cells. Serial passage of the A/Taiwan/1/86 H1N1 strain in the presence of S-ON 5–15b or its antisense as5–15b analogue showed that mutant viruses with reduced susceptibility to the S-ON could indeed be generated, although the resistant viruses displayed reduced replicative fitness. Sequencing the resistant viruses identified mutations in the PB1, PB2, PA and M1 genes. Introduction of these changes into the A/PR/8/34 H1N1 strain by reverse genetics, suggested that alterations to RNA function in the packaging regions of segments 2 and 3 were important in developing resistance to S-ON inhibition. However, many of the other sequence changes induced by S-ON treatment were markedly deleterious to virus fitness. We conclude that packaging signals in the influenza A virus polymerase segments provide feasible targets for nucleic acid-based antivirals that may be difficult for the virus to evade through resistance mutations.
Keywords: Influenza virus; Virus inhibition; Antiviral resistance; Polymerases-derived S-ON; Packaging
Antiherpes activity of glucoevatromonoside, a cardenolide isolated from a Brazilian cultivar of Digitalis lanata by Jéssica Wildgrube Bertol; Caroline Rigotto; Rodrigo Maia de Pádua; Wolfgang Kreis; Célia Regina Monte Barardi; Fernão Castro Braga; Cláudia Maria Oliveira Simões (pp. 73-80).
Cardiac glycosides, known ligands of the sodium pump, are widely used in the treatment of heart failure, such as digoxin and digitoxin. Besides this important activity, other biological activities, such as the antiviral activity, have been described for this group. HSV are responsible for many infections of oral, ocular and genital regions. Treatment with nucleoside analogs such as acyclovir is effective in most cases; however drug-resistance may arise due to prolonged treatment mainly in immunocompromised individuals. In this study, an antiherpes screening was performed with 65 cardenolide derivatives obtained from different sources, and one natural cardenolide, glucoevatromonoside, inhibited HSV-1 and HSV-2 replication at very low concentrations. This cardenolide showed viral inhibitory effects if added up to 12h p.i. and these effects appear to take place by the inhibition of viral proteins synthesis (ICP27, UL42, gB, gD), the blockage of virus release and the reduction of viral cell-to-cell spread. This compound also showed synergistic antiviral effects with acyclovir and anti-Na+K+ATPase activity, suggesting that cellular electrochemical gradient alterations might be involved in the mechanism of viral inhibition. These results suggest that cardenolides might be promising for future antiviral drug design.
Keywords: Glucoevatromonoside; Cardiac glycoside; Antiherpes; HSV-1
Antiherpes activity of glucoevatromonoside, a cardenolide isolated from a Brazilian cultivar of Digitalis lanata by Jéssica Wildgrube Bertol; Caroline Rigotto; Rodrigo Maia de Pádua; Wolfgang Kreis; Célia Regina Monte Barardi; Fernão Castro Braga; Cláudia Maria Oliveira Simões (pp. 73-80).
Cardiac glycosides, known ligands of the sodium pump, are widely used in the treatment of heart failure, such as digoxin and digitoxin. Besides this important activity, other biological activities, such as the antiviral activity, have been described for this group. HSV are responsible for many infections of oral, ocular and genital regions. Treatment with nucleoside analogs such as acyclovir is effective in most cases; however drug-resistance may arise due to prolonged treatment mainly in immunocompromised individuals. In this study, an antiherpes screening was performed with 65 cardenolide derivatives obtained from different sources, and one natural cardenolide, glucoevatromonoside, inhibited HSV-1 and HSV-2 replication at very low concentrations. This cardenolide showed viral inhibitory effects if added up to 12h p.i. and these effects appear to take place by the inhibition of viral proteins synthesis (ICP27, UL42, gB, gD), the blockage of virus release and the reduction of viral cell-to-cell spread. This compound also showed synergistic antiviral effects with acyclovir and anti-Na+K+ATPase activity, suggesting that cellular electrochemical gradient alterations might be involved in the mechanism of viral inhibition. These results suggest that cardenolides might be promising for future antiviral drug design.
Keywords: Glucoevatromonoside; Cardiac glycoside; Antiherpes; HSV-1
Antiherpes activity of glucoevatromonoside, a cardenolide isolated from a Brazilian cultivar of Digitalis lanata by Jéssica Wildgrube Bertol; Caroline Rigotto; Rodrigo Maia de Pádua; Wolfgang Kreis; Célia Regina Monte Barardi; Fernão Castro Braga; Cláudia Maria Oliveira Simões (pp. 73-80).
Cardiac glycosides, known ligands of the sodium pump, are widely used in the treatment of heart failure, such as digoxin and digitoxin. Besides this important activity, other biological activities, such as the antiviral activity, have been described for this group. HSV are responsible for many infections of oral, ocular and genital regions. Treatment with nucleoside analogs such as acyclovir is effective in most cases; however drug-resistance may arise due to prolonged treatment mainly in immunocompromised individuals. In this study, an antiherpes screening was performed with 65 cardenolide derivatives obtained from different sources, and one natural cardenolide, glucoevatromonoside, inhibited HSV-1 and HSV-2 replication at very low concentrations. This cardenolide showed viral inhibitory effects if added up to 12h p.i. and these effects appear to take place by the inhibition of viral proteins synthesis (ICP27, UL42, gB, gD), the blockage of virus release and the reduction of viral cell-to-cell spread. This compound also showed synergistic antiviral effects with acyclovir and anti-Na+K+ATPase activity, suggesting that cellular electrochemical gradient alterations might be involved in the mechanism of viral inhibition. These results suggest that cardenolides might be promising for future antiviral drug design.
Keywords: Glucoevatromonoside; Cardiac glycoside; Antiherpes; HSV-1
Oseltamivir-resistant pandemic A(H1N1) 2009 influenza viruses detected through enhanced surveillance in the Netherlands, 2009–2010 by Adam Meijer; Marcel Jonges; Floor Abbink; Wim Ang; Janko van Beek; Matthias Beersma; Peter Bloembergen; Charles Boucher; Eric Claas; Gé Donker; Rianne van Gageldonk-Lafeber; Leslie Isken; Arjan de Jong; Aloys Kroes; Sander Leenders; Mariken van der Lubben; Ellen Mascini; Bert Niesters; Jan Jelrik Oosterheert; Albert Osterhaus; Rob Riesmeijer; Annelies Riezebos-Brilman; Martin Schutten; Fré Sebens; Foekje Stelma; Corien Swaan; Aura Timen; Annemarie van ’t Veen; Erhard van der Vries; Margreet te Wierik; Marion Koopmans (pp. 81-89).
Enhanced surveillance of infections due to the pandemic A(H1N1) influenza virus, which included monitoring for antiviral resistance, was carried out in the Netherlands from late April 2009 through late May 2010. More than 1100 instances of infection with the pandemic A(H1N1) influenza virus from 2009 and 2010 [A(H1N1) 2009] distributed across this period were analyzed. Of these, 19 cases of oseltamivir-resistant virus harboring the H275Y mutation in the neuraminidase (NA) were detected. The mean 50% inhibitory concentration (IC50) levels for oseltamivir- and zanamivir-susceptible A(H1N1) 2009 viruses were 1.4-fold and 2-fold, respectively, lower than for the seasonal A(H1N1) influenza viruses from 2007/2008; for oseltamivir-resistant A(H1N1) 2009 virus the IC50 was 2.9-fold lower. Eighteen of the 19 patients with oseltamivir-resistant virus showed prolonged shedding of the virus and developed resistance while on oseltamivir therapy. Sixteen of these 18 patients had an immunodeficiency, of whom 11 had a hematologic disorder. The two other patients had another underlying disease. Six of the patients who had an underlying disease died; of these, five had received cytostatic or immunosuppressive therapy. No indications for onward transmission of resistant viruses were found. This study showed that the main association for the emergence of cases of oseltamivir-resistant A(H1N1) 2009 virus was receiving antiviral therapy and having drug-induced immunosuppression or an hematologic disorder. Except for a single case of a resistant virus not linked to oseltamivir therapy, the absence of detection of resistant variants in community specimens and in specimens from contacts of cases with resistant virus suggested that the spread of resistant A(H1N1) 2009 virus was limited. Containment may have been the cumulative result of impaired NA function, successful isolation of the patients, and prophylactic measures to limit exposure.
Keywords: Influenza; Pandemic; A(H1N1); Antiviral; Resistance; Enhanced surveillance
Oseltamivir-resistant pandemic A(H1N1) 2009 influenza viruses detected through enhanced surveillance in the Netherlands, 2009–2010 by Adam Meijer; Marcel Jonges; Floor Abbink; Wim Ang; Janko van Beek; Matthias Beersma; Peter Bloembergen; Charles Boucher; Eric Claas; Gé Donker; Rianne van Gageldonk-Lafeber; Leslie Isken; Arjan de Jong; Aloys Kroes; Sander Leenders; Mariken van der Lubben; Ellen Mascini; Bert Niesters; Jan Jelrik Oosterheert; Albert Osterhaus; Rob Riesmeijer; Annelies Riezebos-Brilman; Martin Schutten; Fré Sebens; Foekje Stelma; Corien Swaan; Aura Timen; Annemarie van ’t Veen; Erhard van der Vries; Margreet te Wierik; Marion Koopmans (pp. 81-89).
Enhanced surveillance of infections due to the pandemic A(H1N1) influenza virus, which included monitoring for antiviral resistance, was carried out in the Netherlands from late April 2009 through late May 2010. More than 1100 instances of infection with the pandemic A(H1N1) influenza virus from 2009 and 2010 [A(H1N1) 2009] distributed across this period were analyzed. Of these, 19 cases of oseltamivir-resistant virus harboring the H275Y mutation in the neuraminidase (NA) were detected. The mean 50% inhibitory concentration (IC50) levels for oseltamivir- and zanamivir-susceptible A(H1N1) 2009 viruses were 1.4-fold and 2-fold, respectively, lower than for the seasonal A(H1N1) influenza viruses from 2007/2008; for oseltamivir-resistant A(H1N1) 2009 virus the IC50 was 2.9-fold lower. Eighteen of the 19 patients with oseltamivir-resistant virus showed prolonged shedding of the virus and developed resistance while on oseltamivir therapy. Sixteen of these 18 patients had an immunodeficiency, of whom 11 had a hematologic disorder. The two other patients had another underlying disease. Six of the patients who had an underlying disease died; of these, five had received cytostatic or immunosuppressive therapy. No indications for onward transmission of resistant viruses were found. This study showed that the main association for the emergence of cases of oseltamivir-resistant A(H1N1) 2009 virus was receiving antiviral therapy and having drug-induced immunosuppression or an hematologic disorder. Except for a single case of a resistant virus not linked to oseltamivir therapy, the absence of detection of resistant variants in community specimens and in specimens from contacts of cases with resistant virus suggested that the spread of resistant A(H1N1) 2009 virus was limited. Containment may have been the cumulative result of impaired NA function, successful isolation of the patients, and prophylactic measures to limit exposure.
Keywords: Influenza; Pandemic; A(H1N1); Antiviral; Resistance; Enhanced surveillance
Oseltamivir-resistant pandemic A(H1N1) 2009 influenza viruses detected through enhanced surveillance in the Netherlands, 2009–2010 by Adam Meijer; Marcel Jonges; Floor Abbink; Wim Ang; Janko van Beek; Matthias Beersma; Peter Bloembergen; Charles Boucher; Eric Claas; Gé Donker; Rianne van Gageldonk-Lafeber; Leslie Isken; Arjan de Jong; Aloys Kroes; Sander Leenders; Mariken van der Lubben; Ellen Mascini; Bert Niesters; Jan Jelrik Oosterheert; Albert Osterhaus; Rob Riesmeijer; Annelies Riezebos-Brilman; Martin Schutten; Fré Sebens; Foekje Stelma; Corien Swaan; Aura Timen; Annemarie van ’t Veen; Erhard van der Vries; Margreet te Wierik; Marion Koopmans (pp. 81-89).
Enhanced surveillance of infections due to the pandemic A(H1N1) influenza virus, which included monitoring for antiviral resistance, was carried out in the Netherlands from late April 2009 through late May 2010. More than 1100 instances of infection with the pandemic A(H1N1) influenza virus from 2009 and 2010 [A(H1N1) 2009] distributed across this period were analyzed. Of these, 19 cases of oseltamivir-resistant virus harboring the H275Y mutation in the neuraminidase (NA) were detected. The mean 50% inhibitory concentration (IC50) levels for oseltamivir- and zanamivir-susceptible A(H1N1) 2009 viruses were 1.4-fold and 2-fold, respectively, lower than for the seasonal A(H1N1) influenza viruses from 2007/2008; for oseltamivir-resistant A(H1N1) 2009 virus the IC50 was 2.9-fold lower. Eighteen of the 19 patients with oseltamivir-resistant virus showed prolonged shedding of the virus and developed resistance while on oseltamivir therapy. Sixteen of these 18 patients had an immunodeficiency, of whom 11 had a hematologic disorder. The two other patients had another underlying disease. Six of the patients who had an underlying disease died; of these, five had received cytostatic or immunosuppressive therapy. No indications for onward transmission of resistant viruses were found. This study showed that the main association for the emergence of cases of oseltamivir-resistant A(H1N1) 2009 virus was receiving antiviral therapy and having drug-induced immunosuppression or an hematologic disorder. Except for a single case of a resistant virus not linked to oseltamivir therapy, the absence of detection of resistant variants in community specimens and in specimens from contacts of cases with resistant virus suggested that the spread of resistant A(H1N1) 2009 virus was limited. Containment may have been the cumulative result of impaired NA function, successful isolation of the patients, and prophylactic measures to limit exposure.
Keywords: Influenza; Pandemic; A(H1N1); Antiviral; Resistance; Enhanced surveillance
Efficacy and tolerance of a combination of tenofovir disoproxil fumarate plus emtricitabine in patients with chronic hepatitis B: A European multicenter study by Si-Nafa Si-Ahmed; Pierre Pradat; Roeland Zoutendijk; Maria Buti; Vincent Mallet; Claire Cruiziat; Katja Deterding; Jérôme Dumortier; François Bailly; Rafael Esteban; Heiner Wedemeyer; Harry L. Janssen; Fabien Zoulim (pp. 90-95).
The combination of tenofovir disoproxil fumarate (TDF) plus emtricitabine (FTC) is used extensively to treat HIV infection and also has potent activity against hepatitis B virus (HBV) infection. The aim of this study was to assess the efficacy and tolerance of TDF+FTC in patients with chronic hepatitis B (CHB).Seventy eight consecutive CHB patients from five European centers were included. All started a TDF+FTC combination between October 2005 and March 2010. Virological, biochemical, and clinical data were recorded during follow-up. Tolerance was also monitored. Patients were classified into either treatment simplification (TS), where efficacy of the previous treatment was obtained at TDF+FTC initiation, and treatment intensification (TI), where the previous line of therapy had failed.TDF+FTC was given as a TI to 54 patients (69%) and as a TS to 24 (31%). Among patients with TI, 83% were males. The median baseline HBV-DNA was 4.4log10IU/mL, and median alanine-transaminase (ALT) was 1.10×ULN. Sixty percent were HBeAg positive, 47% had significant fibrosis (⩾F3 Metavir equivalent), and 29% had confirmed cirrhosis. Median treatment duration was 76weeks (interquartile range 60–116). Kaplan–Meier analysis showed that, 48weeks after TI, the probability of being HBV-DNA becoming undetectable was 76%, and reached 94% at week 96. No viral breakthrough occurred. Patients with TS (87% males, median baseline HBV-DNA 1.1log10IU/mL, median ALT 0.79×ULN, 33% HBeAg positive, 61% with significant fibrosis) were treated for a median duration of 76weeks. In this subgroup, all patients but one remained HBV-DNA undetectable and no ALT flare-up occurred during follow-up. Creatinine levels did not show kidney-function deterioration in either group of patients.After a median follow-up of >76weeks, the TDF+FTC combination showed encouraging antiviral efficacy and a good safety profile in all patients with CHB. TDF+FTC may represent an interesting clinical option to simplify therapy and increase the barrier to resistance, which should be assessed in the long term.
Keywords: Abbreviations; ADV; adefovir; ALT; alanine aminotransferase; CHB; chronic hepatitis B; ETV; entecavir; FTC; emtricitabine; HBV; hepatitis B virus; HIV; human immunodeficiency virus; TDF; tenofovir; ULN; upper limit of normalChronic hepatitis B; Tenofovir; Emtricitabine; Combination therapy
Efficacy and tolerance of a combination of tenofovir disoproxil fumarate plus emtricitabine in patients with chronic hepatitis B: A European multicenter study by Si-Nafa Si-Ahmed; Pierre Pradat; Roeland Zoutendijk; Maria Buti; Vincent Mallet; Claire Cruiziat; Katja Deterding; Jérôme Dumortier; François Bailly; Rafael Esteban; Heiner Wedemeyer; Harry L. Janssen; Fabien Zoulim (pp. 90-95).
The combination of tenofovir disoproxil fumarate (TDF) plus emtricitabine (FTC) is used extensively to treat HIV infection and also has potent activity against hepatitis B virus (HBV) infection. The aim of this study was to assess the efficacy and tolerance of TDF+FTC in patients with chronic hepatitis B (CHB).Seventy eight consecutive CHB patients from five European centers were included. All started a TDF+FTC combination between October 2005 and March 2010. Virological, biochemical, and clinical data were recorded during follow-up. Tolerance was also monitored. Patients were classified into either treatment simplification (TS), where efficacy of the previous treatment was obtained at TDF+FTC initiation, and treatment intensification (TI), where the previous line of therapy had failed.TDF+FTC was given as a TI to 54 patients (69%) and as a TS to 24 (31%). Among patients with TI, 83% were males. The median baseline HBV-DNA was 4.4log10IU/mL, and median alanine-transaminase (ALT) was 1.10×ULN. Sixty percent were HBeAg positive, 47% had significant fibrosis (⩾F3 Metavir equivalent), and 29% had confirmed cirrhosis. Median treatment duration was 76weeks (interquartile range 60–116). Kaplan–Meier analysis showed that, 48weeks after TI, the probability of being HBV-DNA becoming undetectable was 76%, and reached 94% at week 96. No viral breakthrough occurred. Patients with TS (87% males, median baseline HBV-DNA 1.1log10IU/mL, median ALT 0.79×ULN, 33% HBeAg positive, 61% with significant fibrosis) were treated for a median duration of 76weeks. In this subgroup, all patients but one remained HBV-DNA undetectable and no ALT flare-up occurred during follow-up. Creatinine levels did not show kidney-function deterioration in either group of patients.After a median follow-up of >76weeks, the TDF+FTC combination showed encouraging antiviral efficacy and a good safety profile in all patients with CHB. TDF+FTC may represent an interesting clinical option to simplify therapy and increase the barrier to resistance, which should be assessed in the long term.
Keywords: Abbreviations; ADV; adefovir; ALT; alanine aminotransferase; CHB; chronic hepatitis B; ETV; entecavir; FTC; emtricitabine; HBV; hepatitis B virus; HIV; human immunodeficiency virus; TDF; tenofovir; ULN; upper limit of normalChronic hepatitis B; Tenofovir; Emtricitabine; Combination therapy
Efficacy and tolerance of a combination of tenofovir disoproxil fumarate plus emtricitabine in patients with chronic hepatitis B: A European multicenter study by Si-Nafa Si-Ahmed; Pierre Pradat; Roeland Zoutendijk; Maria Buti; Vincent Mallet; Claire Cruiziat; Katja Deterding; Jérôme Dumortier; François Bailly; Rafael Esteban; Heiner Wedemeyer; Harry L. Janssen; Fabien Zoulim (pp. 90-95).
The combination of tenofovir disoproxil fumarate (TDF) plus emtricitabine (FTC) is used extensively to treat HIV infection and also has potent activity against hepatitis B virus (HBV) infection. The aim of this study was to assess the efficacy and tolerance of TDF+FTC in patients with chronic hepatitis B (CHB).Seventy eight consecutive CHB patients from five European centers were included. All started a TDF+FTC combination between October 2005 and March 2010. Virological, biochemical, and clinical data were recorded during follow-up. Tolerance was also monitored. Patients were classified into either treatment simplification (TS), where efficacy of the previous treatment was obtained at TDF+FTC initiation, and treatment intensification (TI), where the previous line of therapy had failed.TDF+FTC was given as a TI to 54 patients (69%) and as a TS to 24 (31%). Among patients with TI, 83% were males. The median baseline HBV-DNA was 4.4log10IU/mL, and median alanine-transaminase (ALT) was 1.10×ULN. Sixty percent were HBeAg positive, 47% had significant fibrosis (⩾F3 Metavir equivalent), and 29% had confirmed cirrhosis. Median treatment duration was 76weeks (interquartile range 60–116). Kaplan–Meier analysis showed that, 48weeks after TI, the probability of being HBV-DNA becoming undetectable was 76%, and reached 94% at week 96. No viral breakthrough occurred. Patients with TS (87% males, median baseline HBV-DNA 1.1log10IU/mL, median ALT 0.79×ULN, 33% HBeAg positive, 61% with significant fibrosis) were treated for a median duration of 76weeks. In this subgroup, all patients but one remained HBV-DNA undetectable and no ALT flare-up occurred during follow-up. Creatinine levels did not show kidney-function deterioration in either group of patients.After a median follow-up of >76weeks, the TDF+FTC combination showed encouraging antiviral efficacy and a good safety profile in all patients with CHB. TDF+FTC may represent an interesting clinical option to simplify therapy and increase the barrier to resistance, which should be assessed in the long term.
Keywords: Abbreviations; ADV; adefovir; ALT; alanine aminotransferase; CHB; chronic hepatitis B; ETV; entecavir; FTC; emtricitabine; HBV; hepatitis B virus; HIV; human immunodeficiency virus; TDF; tenofovir; ULN; upper limit of normalChronic hepatitis B; Tenofovir; Emtricitabine; Combination therapy
Tripeptide inhibitors of dengue and West Nile virus NS2B–NS3 protease by Andreas Schüller; Zheng Yin; C.S. Brian Chia; Danny N.P. Doan; Hyeong-Kyu Kim; Luqing Shang; Teck Peng Loh; Jeffery Hill; Subhash G. Vasudevan (pp. 96-101).
► Tripeptide inhibitors with aldehyde warhead potently inhibit WNV protease but not DENV protease. ► DENV2 protease prefers P2-arginine over P2-lysine that is preferred by WNV NS2B-NS3. ► Phenylacetyl-KRR-H is the new low micromolar inhibitor of DENV2 protease.A series of tripeptide aldehyde inhibitors were synthesized and their inhibitory effect against dengue virus type 2 (DENV2) and West Nile virus (WNV) NS3 protease was evaluated side by side with the aim to discover potent flaviviral protease inhibitors and to examine differences in specificity of the two proteases. The synthesized inhibitors feature a varied N-terminal cap group and side chain modifications of a P2-lysine residue. In general a much stronger inhibitory effect of the tripeptide inhibitors was observed toward WNV protease. The inhibitory concentrations against DENV2 protease were in the micromolar range while they were submicromolar against WNV. The data suggest that a P2-arginine shifts the specificity toward DENV2 protease while WNV protease favors a lysine in the P2 position. Peptides with an extended P2-lysine failed to inhibit DENV2 protease suggesting a size-constrained S2 pocket. Our results generally encourage the investigation of di- and tripeptide aldehydes as inhibitors of DENV and WNV protease.
Keywords: Dengue virus; West Nile virus; NS2B–NS3 protease; Peptide inhibitor
Tripeptide inhibitors of dengue and West Nile virus NS2B–NS3 protease by Andreas Schüller; Zheng Yin; C.S. Brian Chia; Danny N.P. Doan; Hyeong-Kyu Kim; Luqing Shang; Teck Peng Loh; Jeffery Hill; Subhash G. Vasudevan (pp. 96-101).
► Tripeptide inhibitors with aldehyde warhead potently inhibit WNV protease but not DENV protease. ► DENV2 protease prefers P2-arginine over P2-lysine that is preferred by WNV NS2B-NS3. ► Phenylacetyl-KRR-H is the new low micromolar inhibitor of DENV2 protease.A series of tripeptide aldehyde inhibitors were synthesized and their inhibitory effect against dengue virus type 2 (DENV2) and West Nile virus (WNV) NS3 protease was evaluated side by side with the aim to discover potent flaviviral protease inhibitors and to examine differences in specificity of the two proteases. The synthesized inhibitors feature a varied N-terminal cap group and side chain modifications of a P2-lysine residue. In general a much stronger inhibitory effect of the tripeptide inhibitors was observed toward WNV protease. The inhibitory concentrations against DENV2 protease were in the micromolar range while they were submicromolar against WNV. The data suggest that a P2-arginine shifts the specificity toward DENV2 protease while WNV protease favors a lysine in the P2 position. Peptides with an extended P2-lysine failed to inhibit DENV2 protease suggesting a size-constrained S2 pocket. Our results generally encourage the investigation of di- and tripeptide aldehydes as inhibitors of DENV and WNV protease.
Keywords: Dengue virus; West Nile virus; NS2B–NS3 protease; Peptide inhibitor
Tripeptide inhibitors of dengue and West Nile virus NS2B–NS3 protease by Andreas Schüller; Zheng Yin; C.S. Brian Chia; Danny N.P. Doan; Hyeong-Kyu Kim; Luqing Shang; Teck Peng Loh; Jeffery Hill; Subhash G. Vasudevan (pp. 96-101).
► Tripeptide inhibitors with aldehyde warhead potently inhibit WNV protease but not DENV protease. ► DENV2 protease prefers P2-arginine over P2-lysine that is preferred by WNV NS2B-NS3. ► Phenylacetyl-KRR-H is the new low micromolar inhibitor of DENV2 protease.A series of tripeptide aldehyde inhibitors were synthesized and their inhibitory effect against dengue virus type 2 (DENV2) and West Nile virus (WNV) NS3 protease was evaluated side by side with the aim to discover potent flaviviral protease inhibitors and to examine differences in specificity of the two proteases. The synthesized inhibitors feature a varied N-terminal cap group and side chain modifications of a P2-lysine residue. In general a much stronger inhibitory effect of the tripeptide inhibitors was observed toward WNV protease. The inhibitory concentrations against DENV2 protease were in the micromolar range while they were submicromolar against WNV. The data suggest that a P2-arginine shifts the specificity toward DENV2 protease while WNV protease favors a lysine in the P2 position. Peptides with an extended P2-lysine failed to inhibit DENV2 protease suggesting a size-constrained S2 pocket. Our results generally encourage the investigation of di- and tripeptide aldehydes as inhibitors of DENV and WNV protease.
Keywords: Dengue virus; West Nile virus; NS2B–NS3 protease; Peptide inhibitor
4-[1-(4-Fluorobenzyl)-4-hydroxy-1 H-indol-3-yl]-2-hydroxy-4-oxobut-2-enoic acid as a prototype to develop dual inhibitors of HIV-1 integration process by Laura De Luca; Rosaria Gitto; Frauke Christ; Stefania Ferro; Sara De Grazia; Francesca Morreale; Zeger Debyser; Alba Chimirri (pp. 102-107).
In recent years several potent HIV-1 integrase (IN) inhibitors have been identified and after the successful clinical use of raltegravir, they have gained a definitive place in the treatment of HIV-1 infection. Yet, there is a continuous effort to design newer inhibitors that target different steps in the integration process. Furthermore, the increased understanding of IN structural biology has opened novel approaches to inhibit IN, such as targeting its multimerization or interaction with cellular cofactors. On these bases, we have concentrated our research on the identification of small molecules able to inhibit two different stages of the integration process: the IN strand-transfer phase and the IN–LEDGF/p75 interaction. We found that the 4-[1-(4-fluorobenzyl)-4-hydroxy-1 H-indol-3-yl]-2-hydroxy-4-oxobut-2-enoic acid (CHI-1043) is an interesting anti-HIV agent exhibiting dual inhibitory effects. This work has suggested the possibility of also constructing an integration dual inhibitor using a design-in strategy.
Keywords: HIV-1 integrase; INSTI; IN–LEDGF/p75; Dual inhibitor
4-[1-(4-Fluorobenzyl)-4-hydroxy-1 H-indol-3-yl]-2-hydroxy-4-oxobut-2-enoic acid as a prototype to develop dual inhibitors of HIV-1 integration process by Laura De Luca; Rosaria Gitto; Frauke Christ; Stefania Ferro; Sara De Grazia; Francesca Morreale; Zeger Debyser; Alba Chimirri (pp. 102-107).
In recent years several potent HIV-1 integrase (IN) inhibitors have been identified and after the successful clinical use of raltegravir, they have gained a definitive place in the treatment of HIV-1 infection. Yet, there is a continuous effort to design newer inhibitors that target different steps in the integration process. Furthermore, the increased understanding of IN structural biology has opened novel approaches to inhibit IN, such as targeting its multimerization or interaction with cellular cofactors. On these bases, we have concentrated our research on the identification of small molecules able to inhibit two different stages of the integration process: the IN strand-transfer phase and the IN–LEDGF/p75 interaction. We found that the 4-[1-(4-fluorobenzyl)-4-hydroxy-1 H-indol-3-yl]-2-hydroxy-4-oxobut-2-enoic acid (CHI-1043) is an interesting anti-HIV agent exhibiting dual inhibitory effects. This work has suggested the possibility of also constructing an integration dual inhibitor using a design-in strategy.
Keywords: HIV-1 integrase; INSTI; IN–LEDGF/p75; Dual inhibitor
4-[1-(4-Fluorobenzyl)-4-hydroxy-1 H-indol-3-yl]-2-hydroxy-4-oxobut-2-enoic acid as a prototype to develop dual inhibitors of HIV-1 integration process by Laura De Luca; Rosaria Gitto; Frauke Christ; Stefania Ferro; Sara De Grazia; Francesca Morreale; Zeger Debyser; Alba Chimirri (pp. 102-107).
In recent years several potent HIV-1 integrase (IN) inhibitors have been identified and after the successful clinical use of raltegravir, they have gained a definitive place in the treatment of HIV-1 infection. Yet, there is a continuous effort to design newer inhibitors that target different steps in the integration process. Furthermore, the increased understanding of IN structural biology has opened novel approaches to inhibit IN, such as targeting its multimerization or interaction with cellular cofactors. On these bases, we have concentrated our research on the identification of small molecules able to inhibit two different stages of the integration process: the IN strand-transfer phase and the IN–LEDGF/p75 interaction. We found that the 4-[1-(4-fluorobenzyl)-4-hydroxy-1 H-indol-3-yl]-2-hydroxy-4-oxobut-2-enoic acid (CHI-1043) is an interesting anti-HIV agent exhibiting dual inhibitory effects. This work has suggested the possibility of also constructing an integration dual inhibitor using a design-in strategy.
Keywords: HIV-1 integrase; INSTI; IN–LEDGF/p75; Dual inhibitor
Antiherpetic activity of a sulfated polysaccharide from Agaricus brasiliensis mycelia by Francielle Tramontini Gomes de Sousa Cardozo; Carla Maísa Camelini; Alessandra Mascarello; Márcio José Rossi; Ricardo José Nunes; Célia Regina Monte Barardi; Margarida Matos de Mendonça; Cláudia Maria Oliveira Simões (pp. 108-114).
► Sulfation of Agaricus brasiliensis mycelial polysaccharide to obtain its sulfated derivative (MI-S). ► MI-S showed selectivity indices higher than 439, 208, and 562 for HSV-1 KOS, HSV-1 29R, and HSV-2 strain 333, respectively. ► MI-S presented multiple modes of anti-HSV action. ► MI-S inhibited HSV attachment, penetration, protein expression, and cell-to-cell spread. ► MI-S displayed synergistic antiviral effect with acyclovir.Sulfated polysaccharides are good candidates for drug discovery in the treatment of herpetic infections. Agaricus brasiliensis (syn A. subrufescens, A. blazei) is a Basidiomycete fungus native to the Atlantic forest region of Southeastern Brazil. Herein we report the chemical modification of a polysaccharide extracted from A. brasiliensis mycelia to obtain its sulfated derivative (MI-S), which presented a promising inhibitory activity against HSV-1 [KOS and 29R (acyclovir-resistant) strains] and HSV-2 strain 333, with selectivity indices (SI=CC50/IC50) higher than 439, 208, and 562, respectively. The mechanisms underlying this inhibitory activity were scrutinized by plaque assay with different methodological strategies. MI-S had no virucidal effects, but inhibited HSV-1 and HSV-2 attachment, penetration, and cell-to-cell spread, as well as reducing the expression of HSV-1 ICP27, UL42, gB, and gD proteins. MI-S also presented synergistic antiviral effect with acyclovir. These results suggest that MI-S presents multiple modes of anti-HSV action.
Keywords: Agaricus brasiliensis; Sulfated polysaccharide; HSV-1; HSV-2; Antiherpes activity; Mechanism of action
Antiherpetic activity of a sulfated polysaccharide from Agaricus brasiliensis mycelia by Francielle Tramontini Gomes de Sousa Cardozo; Carla Maísa Camelini; Alessandra Mascarello; Márcio José Rossi; Ricardo José Nunes; Célia Regina Monte Barardi; Margarida Matos de Mendonça; Cláudia Maria Oliveira Simões (pp. 108-114).
► Sulfation of Agaricus brasiliensis mycelial polysaccharide to obtain its sulfated derivative (MI-S). ► MI-S showed selectivity indices higher than 439, 208, and 562 for HSV-1 KOS, HSV-1 29R, and HSV-2 strain 333, respectively. ► MI-S presented multiple modes of anti-HSV action. ► MI-S inhibited HSV attachment, penetration, protein expression, and cell-to-cell spread. ► MI-S displayed synergistic antiviral effect with acyclovir.Sulfated polysaccharides are good candidates for drug discovery in the treatment of herpetic infections. Agaricus brasiliensis (syn A. subrufescens, A. blazei) is a Basidiomycete fungus native to the Atlantic forest region of Southeastern Brazil. Herein we report the chemical modification of a polysaccharide extracted from A. brasiliensis mycelia to obtain its sulfated derivative (MI-S), which presented a promising inhibitory activity against HSV-1 [KOS and 29R (acyclovir-resistant) strains] and HSV-2 strain 333, with selectivity indices (SI=CC50/IC50) higher than 439, 208, and 562, respectively. The mechanisms underlying this inhibitory activity were scrutinized by plaque assay with different methodological strategies. MI-S had no virucidal effects, but inhibited HSV-1 and HSV-2 attachment, penetration, and cell-to-cell spread, as well as reducing the expression of HSV-1 ICP27, UL42, gB, and gD proteins. MI-S also presented synergistic antiviral effect with acyclovir. These results suggest that MI-S presents multiple modes of anti-HSV action.
Keywords: Agaricus brasiliensis; Sulfated polysaccharide; HSV-1; HSV-2; Antiherpes activity; Mechanism of action
Antiherpetic activity of a sulfated polysaccharide from Agaricus brasiliensis mycelia by Francielle Tramontini Gomes de Sousa Cardozo; Carla Maísa Camelini; Alessandra Mascarello; Márcio José Rossi; Ricardo José Nunes; Célia Regina Monte Barardi; Margarida Matos de Mendonça; Cláudia Maria Oliveira Simões (pp. 108-114).
► Sulfation of Agaricus brasiliensis mycelial polysaccharide to obtain its sulfated derivative (MI-S). ► MI-S showed selectivity indices higher than 439, 208, and 562 for HSV-1 KOS, HSV-1 29R, and HSV-2 strain 333, respectively. ► MI-S presented multiple modes of anti-HSV action. ► MI-S inhibited HSV attachment, penetration, protein expression, and cell-to-cell spread. ► MI-S displayed synergistic antiviral effect with acyclovir.Sulfated polysaccharides are good candidates for drug discovery in the treatment of herpetic infections. Agaricus brasiliensis (syn A. subrufescens, A. blazei) is a Basidiomycete fungus native to the Atlantic forest region of Southeastern Brazil. Herein we report the chemical modification of a polysaccharide extracted from A. brasiliensis mycelia to obtain its sulfated derivative (MI-S), which presented a promising inhibitory activity against HSV-1 [KOS and 29R (acyclovir-resistant) strains] and HSV-2 strain 333, with selectivity indices (SI=CC50/IC50) higher than 439, 208, and 562, respectively. The mechanisms underlying this inhibitory activity were scrutinized by plaque assay with different methodological strategies. MI-S had no virucidal effects, but inhibited HSV-1 and HSV-2 attachment, penetration, and cell-to-cell spread, as well as reducing the expression of HSV-1 ICP27, UL42, gB, and gD proteins. MI-S also presented synergistic antiviral effect with acyclovir. These results suggest that MI-S presents multiple modes of anti-HSV action.
Keywords: Agaricus brasiliensis; Sulfated polysaccharide; HSV-1; HSV-2; Antiherpes activity; Mechanism of action
Fluoroquinolones inhibit human polyomavirus BK (BKV) replication in primary human kidney cells by Biswa Nath Sharma; Ruomei Li; Eva Bernhoff; Tore Jarl Gutteberg; Christine Hanssen Rinaldo (pp. 115-123).
Reactivation of human polyomavirus BK (BKV) may cause polyomavirus-associated nephropathy or polyomavirus-associated hemorrhagic cystitis in renal- or bone marrow-transplant patients, respectively. Lack of treatment options has led to exploration of fluoroquinolones that inhibit topoisomerase II and IV in prokaryotes and possibly large T-antigen (LT-ag) helicase activity in polyomavirus. We characterized the effects of ofloxacin and levofloxacin on BKV replication in the natural host cells – primary human renal proximal tubular epithelial cells (RPTECs). Ofloxacin and levofloxacin inhibited BKV load in a dose-dependent manner yielding a ∼90% inhibition at 150μg/ml. Ofloxacin at 150μg/ml inhibited LT-ag mRNA and protein expression from 24h post infection (hpi). BKV genome replication was 77% reduced at 48hpi and a similar reduction was found in VP1 and agnoprotein expression. At 72hpi, the reduction in genome replication and protein expression was less pronounced. A dose-dependent cytostatic effect was noted. In infected cells, 150μg/ml ofloxacin led to a 26% and 6% inhibition of cellular DNA replication and total metabolic activity, respectively while 150μg/ml levofloxacin affected this slightly more, particularly in uninfected cells. Cell counting and xCELLigence results revealed that cell numbers were not reduced. In conclusion, ofloxacin and levofloxacin inhibit but do not eradicate BKV replication in RPTECs. At a concentration of ofloxacin giving ∼90% inhibition in BKV load, no significant cytotoxicity was observed. This concentration can be achieved in urine and possibly in the kidneys. Our results support a mechanism involving inhibition of LT-ag expression or functions but also suggest inhibition of cellular enzymes.
Keywords: Polyomavirus BK; Ofloxacin; Levofloxacin; Large T-antigen; Replication; RPTECs
Fluoroquinolones inhibit human polyomavirus BK (BKV) replication in primary human kidney cells by Biswa Nath Sharma; Ruomei Li; Eva Bernhoff; Tore Jarl Gutteberg; Christine Hanssen Rinaldo (pp. 115-123).
Reactivation of human polyomavirus BK (BKV) may cause polyomavirus-associated nephropathy or polyomavirus-associated hemorrhagic cystitis in renal- or bone marrow-transplant patients, respectively. Lack of treatment options has led to exploration of fluoroquinolones that inhibit topoisomerase II and IV in prokaryotes and possibly large T-antigen (LT-ag) helicase activity in polyomavirus. We characterized the effects of ofloxacin and levofloxacin on BKV replication in the natural host cells – primary human renal proximal tubular epithelial cells (RPTECs). Ofloxacin and levofloxacin inhibited BKV load in a dose-dependent manner yielding a ∼90% inhibition at 150μg/ml. Ofloxacin at 150μg/ml inhibited LT-ag mRNA and protein expression from 24h post infection (hpi). BKV genome replication was 77% reduced at 48hpi and a similar reduction was found in VP1 and agnoprotein expression. At 72hpi, the reduction in genome replication and protein expression was less pronounced. A dose-dependent cytostatic effect was noted. In infected cells, 150μg/ml ofloxacin led to a 26% and 6% inhibition of cellular DNA replication and total metabolic activity, respectively while 150μg/ml levofloxacin affected this slightly more, particularly in uninfected cells. Cell counting and xCELLigence results revealed that cell numbers were not reduced. In conclusion, ofloxacin and levofloxacin inhibit but do not eradicate BKV replication in RPTECs. At a concentration of ofloxacin giving ∼90% inhibition in BKV load, no significant cytotoxicity was observed. This concentration can be achieved in urine and possibly in the kidneys. Our results support a mechanism involving inhibition of LT-ag expression or functions but also suggest inhibition of cellular enzymes.
Keywords: Polyomavirus BK; Ofloxacin; Levofloxacin; Large T-antigen; Replication; RPTECs
Fluoroquinolones inhibit human polyomavirus BK (BKV) replication in primary human kidney cells by Biswa Nath Sharma; Ruomei Li; Eva Bernhoff; Tore Jarl Gutteberg; Christine Hanssen Rinaldo (pp. 115-123).
Reactivation of human polyomavirus BK (BKV) may cause polyomavirus-associated nephropathy or polyomavirus-associated hemorrhagic cystitis in renal- or bone marrow-transplant patients, respectively. Lack of treatment options has led to exploration of fluoroquinolones that inhibit topoisomerase II and IV in prokaryotes and possibly large T-antigen (LT-ag) helicase activity in polyomavirus. We characterized the effects of ofloxacin and levofloxacin on BKV replication in the natural host cells – primary human renal proximal tubular epithelial cells (RPTECs). Ofloxacin and levofloxacin inhibited BKV load in a dose-dependent manner yielding a ∼90% inhibition at 150μg/ml. Ofloxacin at 150μg/ml inhibited LT-ag mRNA and protein expression from 24h post infection (hpi). BKV genome replication was 77% reduced at 48hpi and a similar reduction was found in VP1 and agnoprotein expression. At 72hpi, the reduction in genome replication and protein expression was less pronounced. A dose-dependent cytostatic effect was noted. In infected cells, 150μg/ml ofloxacin led to a 26% and 6% inhibition of cellular DNA replication and total metabolic activity, respectively while 150μg/ml levofloxacin affected this slightly more, particularly in uninfected cells. Cell counting and xCELLigence results revealed that cell numbers were not reduced. In conclusion, ofloxacin and levofloxacin inhibit but do not eradicate BKV replication in RPTECs. At a concentration of ofloxacin giving ∼90% inhibition in BKV load, no significant cytotoxicity was observed. This concentration can be achieved in urine and possibly in the kidneys. Our results support a mechanism involving inhibition of LT-ag expression or functions but also suggest inhibition of cellular enzymes.
Keywords: Polyomavirus BK; Ofloxacin; Levofloxacin; Large T-antigen; Replication; RPTECs