RESUMEN
Enterovirus A71 can cause serious neurological disease in young children. Animal models for EV-A71 are needed to evaluate potential antiviral therapies. Existing models have limitations, including lack of lethality or crucial disease signs. Here we report the development of an EV-A71 model in 28-day-old mice. Virus was serially passaged until it produced consistent lethality and rear-limb paralysis. Onset of disease occurred between days 6-9 post-infection, with mortality following weight loss and neurological signs on days 9-14. In addition, a single administration of human intravenous immunoglobulin at doses of 200, 400 and 800 mg/kg at 4h post-infection was evaluated in the model. Protection from weight loss, neurological signs, and mortality (between 50 and 89%) were observed at doses of 400 mg/kg or greater. Based on these results, IVIG was selected for use as a positive control in this acute model, and suggest that IVIG is a potential therapeutic for EV-A71 infections.
Asunto(s)
Enterovirus Humano A , Infecciones por Enterovirus , Enterovirus , Enfermedades del Sistema Nervioso , Niño , Humanos , Ratones , Animales , Preescolar , Inmunoglobulinas Intravenosas/uso terapéutico , Modelos Animales de EnfermedadRESUMEN
Orthopoxviruses such as variola and monkeypox viruses continue to threaten the human population. Monkeypox virus is endemic in central and western Africa and outbreaks have reached as far as the U.S. Although variola virus, the etiologic agent of smallpox, has been eradicated by a successful vaccination program, official and likely clandestine stocks of the virus exist. Moreover, studies with ectromelia virus (the etiological agent of mousepox) have revealed that IL-4 recombinant viruses are significantly more virulent than wild-type viruses even in mice treated with vaccines and/or antivirals. For these reasons, it is critical that antiviral modalities are developed to treat these viruses should outbreaks, or deliberate dissemination, occur. Currently, 2 antivirals (brincidofovir and tecovirimat) are in the U.S. stockpile allowing for emergency use of the drugs to treat smallpox. Both antivirals have advantages and disadvantages in a clinical and emergency setting. Here we report on the efficacy of a recombinant immunoglobulin (rVIG) that demonstrated efficacy against several orthopoxviruses in vitro and in vivo in both a prophylactic and therapeutic fashion. A single intraperitoneal injection of rVIG significantly protected mice when given up to 14 days before or as late as 6 days post challenge. Moreover, rVIG reduced morbidity, as measured by weight-change, as well as several previously established biomarkers of disease. In rVIG treated mice, we found that vDNA levels in blood were significantly reduced, as was ALT (a marker of liver damage) and infectious virus levels in the liver. No apparent adverse events were observed in rVIG treated mice, suggesting the immunoglobulin is well tolerated. These findings suggest that recombinant immunoglobulins could be candidates for further evaluation and possible licensure under the FDA Animal Rule.
Asunto(s)
Antivirales/uso terapéutico , Inmunoglobulinas/uso terapéutico , Orthopoxvirus/efectos de los fármacos , Viruela/tratamiento farmacológico , Vaccinia/tratamiento farmacológico , Animales , Antivirales/administración & dosificación , Benzamidas , Línea Celular , Chlorocebus aethiops , Citosina/análogos & derivados , Femenino , Humanos , Isoindoles , Ratones , Ratones Endogámicos BALB C , Organofosfonatos , Viruela/prevención & control , Viruela/virología , Vacuna contra Viruela/administración & dosificación , Vacunas de ADN/administración & dosificación , Vaccinia/prevención & control , Vaccinia/virologíaRESUMEN
Antiviral countermeasures are needed to reduce the morbidity associated with Chikungunya virus (CHIKV) infection. This arbovirus reemerged in 2004 and causes periodic outbreaks in various areas throughout the world. While infection is rarely lethal, the majority of people infected with the virus develop a hallmark arthralgia as well as other disease manifestations. The virus is classified within three phylogenetic groups, namely, West African, East/Central/South African (ECSA), and Asian. Six strains of CHIKV covering the three phylogenetic groups were studied for their replication in cell culture, their ability to cause disease in susceptible mouse strains and susceptibility to antiviral treatment. Differential replication kinetics were observed for various CHIKV isolates in cell culture, which coincided with a decreased sensitivity to antiviral treatment as compared with ECSA and Asian clade viruses. This was confirmed in mouse infection studies with severe disease observed in mice infected with West African clade viruses, mild disease phenotype after infection with Asian clade viruses and an intermediate disease severity associated with ECSA virus infection. We also tested a broadly active antiviral, Favipiravir (T-705), which activity was inversely proportional to disease severity. These data suggest that some clades of CHIKV may cause more severe disease and may be more difficult to treat.
Asunto(s)
Amidas/uso terapéutico , Antivirales/uso terapéutico , Fiebre Chikungunya/tratamiento farmacológico , Virus Chikungunya/efectos de los fármacos , Virus Chikungunya/patogenicidad , Pirazinas/uso terapéutico , Animales , Línea Celular , Fiebre Chikungunya/virología , Virus Chikungunya/clasificación , Femenino , Genotipo , Humanos , Ratones , Ratones Endogámicos DBA , Fenotipo , FilogeniaRESUMEN
There is a strong need for a new broad-spectrum antiinfluenza therapeutic, as vaccination and existing treatments are only moderately effective. We previously engineered a lectin, H84T banana lectin (H84T), to retain broad-spectrum activity against multiple influenza strains, including pandemic and avian, while largely eliminating the potentially harmful mitogenicity of the parent compound. The amino acid mutation at position 84 from histidine to threonine minimizes the mitogenicity of the wild-type lectin while maintaining antiinfluenza activity in vitro. We now report that in a lethal mouse model H84T is indeed nonmitogenic, and both early and delayed therapeutic administration of H84T intraperitoneally are highly protective, as is H84T administered subcutaneously. Mechanistically, attachment, which we anticipated to be inhibited by H84T, was only somewhat decreased by the lectin. Instead, H84T is internalized into the late endosomal/lysosomal compartment and inhibits virus-endosome fusion. These studies reveal that H84T is efficacious against influenza virus in vivo, and that the loss of mitogenicity seen previously in tissue culture is also seen in vivo, underscoring the potential utility of H84T as a broad-spectrum antiinfluenza agent.
Asunto(s)
Antivirales/administración & dosificación , Gripe Humana/tratamiento farmacológico , Lectinas/administración & dosificación , Lectinas/genética , Musa/genética , Proteínas de Plantas/administración & dosificación , Proteínas de Plantas/genética , Internalización del Virus/efectos de los fármacos , Animales , Humanos , Subtipo H1N1 del Virus de la Influenza A/efectos de los fármacos , Subtipo H1N1 del Virus de la Influenza A/genética , Subtipo H1N1 del Virus de la Influenza A/fisiología , Subtipo H3N2 del Virus de la Influenza A/efectos de los fármacos , Subtipo H3N2 del Virus de la Influenza A/genética , Subtipo H3N2 del Virus de la Influenza A/fisiología , Gripe Humana/virología , Masculino , Ratones , Musa/química , Musa/metabolismo , Mutación , Ingeniería de ProteínasRESUMEN
Enterovirus D68 (EV-D68) is a non-polio enterovirus that affects the respiratory system and can cause serious complications, especially in children and older people with weakened immune systems. As an emerging virus, there are no current antiviral therapies or vaccines available. Our goal was to develop a mouse model of human EV-D68 infection that mimicked the disease observed in humans and could be used for evaluation of experimental therapeutics. This is the first report of a respiratory disease model for EV-D68 infection in mice. We adapted the virus by 30 serial passages in AG129 mice, which are deficient in IFN- α/ß and -γ receptors. Despite a lack of weight loss or mortality in mice, lung function measured by plethysmography, showed an increase in enhanced pause (Penh) on days 6 and 7 post-infection. In addition, as virus adapted to mice, virus titer in the lungs increased 50-fold, and the pro-inflammatory cytokines MCP-1 and RANTES increased 15-fold and 2-fold in the lung, respectively. In addition, a time course of mouse-adapted EV-D68 infection was determined in lung, blood, liver, kidney, spleen, leg muscle, spinal cord and brain. Virus in the lung replicated rapidly after intranasal inoculation of adapted virus, 106 CCID50/mL by 4â¯h and 108.3 CCID50/mL by 24â¯h. Virus then spread to the blood and other tissues, including spinal cord and brain. This mouse model for EV-D68 infection includes enhanced pause (Penh) as an indicator of morbidity, and viremia, virus titers and proinflammatory cytokines in the lung, and lung histopathology as indicators of disease. Our mouse-adapted virus has a similar antiviral profile to the original isolate as well as another respiratory picornavirus, rhinovirus-14. This model will be valuable in evaluating experimental therapies in the future.
Asunto(s)
Modelos Animales de Enfermedad , Infecciones por Enterovirus/inmunología , Infecciones por Enterovirus/virología , Pulmón/virología , Infecciones del Sistema Respiratorio/virología , Animales , Antivirales/uso terapéutico , Quimiocinas/inmunología , Citocinas/inmunología , Enterovirus Humano D , Infecciones por Enterovirus/tratamiento farmacológico , Femenino , Masculino , Ratones , Infecciones del Sistema Respiratorio/tratamiento farmacológico , Infecciones del Sistema Respiratorio/inmunología , Carga Viral , ViremiaRESUMEN
Enterovirus D68 (EV-D68) is unique among enteroviruses because of the ability to cause severe respiratory disease as well as neurological disease. We developed separate models of respiratory and neurological disease following EV-D68 infection in AG129 mice that respond to antiviral treatment with guanidine. In four-week-old mice infected intranasally, EV-D68 replicates to high titers in lung tissue increasing the proinflammatory cytokines MCP-1 and IL-6. The respiratory infection also produces an acute viremia. In 10-day-old mice infected intraperitoneally, EV-D68 causes a neurological disease with weight-loss, paralysis, and mortality. In our respiratory model, treatment with guanidine provides a two-log reduction in lung virus titers, reduces MCP-1 and IL-6, and prevents histological lesions in the lungs. Importantly, viremia is prevented by early treatment with guanidine. In our neurological model, guanidine treatment protects mice from weight-loss, paralysis, and mortality. These results demonstrate the utility of these models for evaluation of antiviral therapies for EV-D68 infection.
Asunto(s)
Antivirales/uso terapéutico , Modelos Animales de Enfermedad , Enterovirus Humano D , Infecciones por Enterovirus/tratamiento farmacológico , Guanidina/uso terapéutico , Enfermedades del Sistema Nervioso/tratamiento farmacológico , Infecciones del Sistema Respiratorio/tratamiento farmacológico , Animales , Citocinas/metabolismo , Infecciones por Enterovirus/patología , Infecciones por Enterovirus/fisiopatología , Infecciones por Enterovirus/virología , Femenino , Humanos , Pulmón/metabolismo , Pulmón/patología , Pulmón/fisiopatología , Pulmón/virología , Masculino , Ratones , Enfermedades del Sistema Nervioso/patología , Enfermedades del Sistema Nervioso/fisiopatología , Enfermedades del Sistema Nervioso/virología , Infecciones del Sistema Respiratorio/patología , Infecciones del Sistema Respiratorio/fisiopatología , Infecciones del Sistema Respiratorio/virología , Resultado del Tratamiento , Carga Viral/efectos de los fármacos , Viremia/prevención & controlRESUMEN
2'-Fluoro-2'-deoxycytidine (2'-FdC) was reported to inhibit various viruses in vitro, including Borna disease, hepatitis C, Lassa fever, influenza and certain herpes viruses, and is inhibitory to influenza viruses in mice. We investigated the antiviral activity of 2'-FdC against several unrelated bunyaviruses in 50% cytopathic effect (CPE) inhibition assays and, with viruses that cause limited CPE, 90% virus yield reduction (VYR) assays. La Crosse (LACV), Maporal, Punta Toro, Rift Valley fever (RVFV), and San Angelo viruses were inhibited in CPE assays at 2.2-9.7⯵M concentrations. In VYR assays, Heartland and severe fever with thrombocytopenia syndrome (SFTSV) viruses were inhibited at 0.9 and 3.7⯵M, respectively. In contrast, ribavirin inhibited these viruses at an average of 47⯵M. Antiviral efficacy studies were also conducted in mice infected with RVFV, SFTSV, and LACV. Against RVFV, 2'-FdC (100 and 200â¯mg/kg/day) and ribavirin (100â¯mg/kg/day) treatments each delayed mortality by approximately 6 days compared to placebo. Liver, spleen, and serum viral titers were significantly reduced by antiviral treatments. 2'-FdC (100 and 200â¯mg/kg/day) prevented death in SFTSV-infected mice, but was not as effective as favipiravir (100â¯mg/kg/day) based on body weight loss during infection. The 100â¯mg/kg/day doses of 2'-FdC and favipiravir significantly reduced liver, spleen, and serum viral titers. 2'-FdC and ribavirin afforded no protection against LACV infection in mice, which is encephalitic and thus inherently more difficult to treat. Taken together, our data suggest that 2'-FdC may be a viable candidate for treating certain non-encephalitic bunyavirus infections such as those caused by phleboviruses.
Asunto(s)
Antivirales/administración & dosificación , Antivirales/farmacología , Infecciones por Bunyaviridae/tratamiento farmacológico , Virus ADN/efectos de los fármacos , Desoxicitidina/análogos & derivados , Virus ARN/efectos de los fármacos , Estructuras Animales/virología , Animales , Peso Corporal , Efecto Citopatogénico Viral , Virus ADN/crecimiento & desarrollo , Desoxicitidina/administración & dosificación , Desoxicitidina/farmacología , Modelos Animales de Enfermedad , Ratones , Pruebas de Sensibilidad Microbiana , Placebos/administración & dosificación , Virus ARN/crecimiento & desarrollo , Análisis de Supervivencia , Resultado del Tratamiento , Carga ViralRESUMEN
On September 22, 2008, a physician on Prince of Wales Island, Alaska, notified the Alaska Department of Health and Social Services (ADHSS) of an unusually high number of adult patients with recently diagnosed pneumonia (n = 10), including three persons who required hospitalization and one who died. ADHSS and CDC conducted an investigation to determine the cause and distribution of the outbreak, identify risk factors for hospitalization, and implement control measures. This report summarizes the results of that investigation, which found that the outbreak was caused by adenovirus 14 (Ad14), an emerging adenovirus serotype in the United States that is associated with a higher rate of severe illness compared with other adenoviruses. Among the 46 cases identified in the outbreak from September 1 through October 27, 2008, the most frequently observed characteristics included the following: male (70%), Alaska Native (61%), underlying pulmonary disease (44%), aged > or = 65 years (26%), and current smoker (48%). Patients aged > or = 65 years had a fivefold increased risk for hospitalization. The most commonly reported symptoms were cough (100%), shortness of breath (87%), and fever (74%). Of the 11 hospitalized patients, three required intensive care, and one required mechanical ventilation. One death was reported. Ad14 isolates obtained during the outbreak were identical genetically to those in recent community-acquired outbreaks in the United States which suggests the emergence of a new, and possibly more virulent Ad14 variant. Clinicians should consider Ad14 infection in the differential diagnosis for patients with community-acquired pneumonia, particularly when unexplained clusters of severe respiratory infections are detected.
Asunto(s)
Adenovirus Humanos/efectos de los fármacos , Ésteres/farmacología , Nucleósidos/farmacología , Serogrupo , Células A549 , Infecciones por Adenovirus Humanos/epidemiología , Infecciones por Adenovirus Humanos/virología , Adenovirus Humanos/patogenicidad , Anciano , Brotes de Enfermedades , Femenino , Fiebre , Humanos , Masculino , Nucleósidos/análogos & derivados , Análisis de Regresión , Infecciones del Sistema Respiratorio/virología , Estados UnidosRESUMEN
Cell culture antiviral experiments were conducted in order to understand the relationship between percentage data generated by plaque reduction (PR) and logarithmic data derived by virus yield reduction (VYR) assays, using three-dimensional MacSynergy II software. The relationship between percentage and logarithmic data has not been investigated previously. Interpretation of drug-drug interactions is based on a Volume of Synergy (VS) calculation, which can be positive (synergy), negative (antagonistic), or neutral (no or minimal interaction). Interactions of two known inhibitors of vaccinia virus replication, cidofovir and 6-azauridine, used in combination by PR assay yielded a VS value of 265, indicative of strong synergy. By VYR, the VS value was only 37, or weak synergy using the same criterion, even though profound log10 reductions in virus titer occurred at multiple drug combinations. These results confirm that the differences in VS values is dependent of the measurement scale, and not that the degree of synergy differed between the assays. We propose that for logarithmic data, the calculated VS values will be lower for significant synergy and antagonism and that volumes of >10 µM2log10 PFU/ml (or other units such as µM2log10 genomic equivalents/ml or µM2log10 copies/ml) and <-10 µM2log10 PFU/ml are likely to be indicative of strong synergy and strong antagonism, respectively. Data presented here show that the interaction of cidofovir and 6-azauridine was strongly synergistic in vitro.
Asunto(s)
Antivirales/farmacología , Sinergismo Farmacológico , Pruebas de Sensibilidad Microbiana , Virus Vaccinia/efectos de los fármacos , Animales , Azauridina/farmacología , Chlorocebus aethiops , Cidofovir , Citosina/análogos & derivados , Citosina/farmacología , Exactitud de los Datos , Interpretación Estadística de Datos , Interacciones Farmacológicas , Humanos , Organofosfonatos/farmacología , Programas Informáticos , Virus Vaccinia/crecimiento & desarrollo , Células Vero , Ensayo de Placa Viral , Replicación Viral/efectos de los fármacosRESUMEN
CONTEXT: Kalanchoe pinnata (Lam.) Pers. (Crassulaceae) is a succulent plant that is known for its traditional antivirus and antibacterial usage. OBJECTIVE: This work examines two compounds identified from the K. pinnata plant for their antivirus activity against human alphaherpesvirus (HHV) 1 and 2 and vaccinia virus (VACV). MATERIALS AND METHODS: Compounds KPB-100 and KPB-200 were isolated using HPLC and were identified using NMR and MS. Both compounds were tested in plaque reduction assay of HHV-2 wild type (WT) and VACV. Both compounds were then tested in virus spread inhibition and virus yield reduction (VYR) assays of VACV. KPB-100 was further tested in viral cytopathic effect (CPE) inhibition assay of HHV-2 TK-mutant and VYR assay of HHV-1 WT. RESULTS: KPB-100 and KPB-200 inhibited HHV-2 at IC50 values of 2.5 and 2.9 µg/mL, respectively, and VACV at IC50 values of 3.1 and 7.4 µg/mL, respectively, in plaque reduction assays. In virus spread inhibition assay of VACV KPB-100 and KPB-200 yielded IC50 values of 1.63 and 13.2 µg/mL, respectively, and KPB-100 showed a nearly 2-log reduction in virus in VYR assay of VACV at 20 µg/mL. Finally, KPB-100 inhibited HHV-2 TK- at an IC50 value of 4.5 µg/mL in CPE inhibition assay and HHV-1 at an IC90 of 3.0 µg/mL in VYR assay. DISCUSSION AND CONCLUSION: Both compounds are promising targets for synthetic optimization and in vivo study. KPB-100 in particular showed strong inhibition of all viruses tested.
Asunto(s)
Antivirales/farmacología , Kalanchoe/química , Extractos Vegetales/farmacología , Antivirales/administración & dosificación , Antivirales/aislamiento & purificación , Cromatografía Líquida de Alta Presión , Efecto Citopatogénico Viral/efectos de los fármacos , Células HeLa , Herpesvirus Humano 1/efectos de los fármacos , Herpesvirus Humano 2/efectos de los fármacos , Humanos , Concentración 50 Inhibidora , Espectroscopía de Resonancia Magnética , Espectrometría de Masas , Extractos Vegetales/administración & dosificación , Extractos Vegetales/aislamiento & purificación , Virus Vaccinia/efectos de los fármacosRESUMEN
Studies were conducted to determine the performance of four dyes in assessing antiviral activities of compounds against three RNA viruses with differing cytopathogenic properties. Dyes included alamarBlue® measured by absorbance (ALB-A) and fluorescence (ALB-F), neutral red (NR), Viral ToxGlo™ (VTG), and WST-1. Viruses were chikungunya, dengue type 2, and Junin, which generally cause 100, 80-90, and 50% maximal cytopathic effect (CPE), respectively, in Vero or Vero 76 cells Compounds evaluated were 6-azauridine, BCX-4430, 3-deazaguanine, EICAR, favipiravir, infergen, mycophenolic acid (MPA), ribavirin, and tiazofurin. The 50% virus-inhibitory (EC50) values for each inhibitor and virus combination did not vary significantly based on the dye used. However, dyes varied in distinguishing the vitality of virus-infected cultures when not all cells were killed by virus infection. For example, VTG uptake into dengue-infected cells was nearly 50% when visual examination showed only 10-20% cell survival. ALB-A measured infected cell viability differently than ALB-F as follows: 16% versus 32% (dengue-infected), respectively, and 51% versus 72% (Junin-infected), respectively. Cytotoxicity (CC50) assays with dyes in uninfected proliferating cells produced similar CC50 values for EICAR (1.5-8.9µM) and MPA (0.8-2.5µM). 6-Azauridine toxicity was 6.1-17.5µM with NR, VTG, and WST-1, compared to 48-92µM with ALB-A and ALB-F (P<0.001). Curiously, the CC50 values for 3-deazaguanine were 83-93µM with ALB-F versus 2.4-7.0µM with all other dyes including ALB-A (P<0.001). Overall, ALB minimized the toxicities detected with these two inhibitors. Because the choice of dyes affected CC50 values, this impacted on the resulting in vitro selectivity indexes (calculated as CC50/EC50 ratio).
Asunto(s)
Antivirales/farmacología , Supervivencia Celular/efectos de los fármacos , Colorantes , Efecto Citopatogénico Viral , Virus ARN/efectos de los fármacos , Virus/efectos de los fármacos , Animales , Virus Chikungunya/efectos de los fármacos , Virus Chikungunya/patogenicidad , Virus Chikungunya/fisiología , Chlorocebus aethiops , Colorantes/química , Virus del Dengue/efectos de los fármacos , Virus del Dengue/patogenicidad , Virus del Dengue/fisiología , Virus Junin/efectos de los fármacos , Virus Junin/patogenicidad , Virus Junin/fisiología , Oxazinas , Virus ARN/patogenicidad , Virus ARN/fisiología , Células Vero , Replicación Viral/efectos de los fármacos , XantenosRESUMEN
L-NG-monomethyl-arginine (L-NMMA) is an experimental compound that suppresses nitric oxide production in animals. The compound was combined with oseltamivir to treat lethal influenza A/California/04/2009 (H1N1) pandemic virus infections in mice. Treatments were given twice a day for five days starting 4 h (oseltamivir, by oral gavage) or three days (L-NMMA, by intraperitoneal route; corresponding to the time previously reported for nitric oxide induction in the animals) after infection. Low doses of oseltamivir were used in order to demonstrate synergy or antagonism. Oseltamivir monotherapy protected 70% of mice from death at 1 mg/kg/day. L-NMMA (40 and 80 mg/kg/day) was ineffective alone in preventing mortality. Compared to oseltamivir treatment alone, L-NMMA combined with oseltamivir was synergistically effective (as evaluated by three-dimensional MacSynergy analysis), resulting in survival increases from 20 to 70% when 40 or 80 mg/kg/day of L-NMMA was combined with 0.3 mg/kg/day of oseltamivir, and from 70 to 100% survival increases when these doses were combined with 1 mg/kg/day of oseltamivir. These data demonstrate that a nitric oxide inhibitor such as L-NMMA has the potential to be beneficial when combined with oseltamivir in treating influenza virus infections.
Asunto(s)
Antivirales/farmacología , Inhibidores Enzimáticos/farmacología , Subtipo H1N1 del Virus de la Influenza A/efectos de los fármacos , Infecciones por Orthomyxoviridae/tratamiento farmacológico , Oseltamivir/farmacología , omega-N-Metilarginina/farmacología , Animales , Modelos Animales de Enfermedad , Sinergismo Farmacológico , Femenino , Subtipo H1N1 del Virus de la Influenza A/aislamiento & purificación , Ratones , Ratones Endogámicos BALB C , Infecciones por Orthomyxoviridae/virologíaRESUMEN
Adamantanes such as amantadine (1) and rimantadine (2) are FDA-approved anti-influenza drugs that act by inhibiting the wild-type M2 proton channel from influenza A viruses, thereby inhibiting the uncoating of the virus. Although adamantanes have been successfully used for more than four decades, their efficacy was curtailed by emerging drug resistance. Among the limited number of M2 mutants that confer amantadine resistance, the M2-V27A mutant was found to be the predominant mutant under drug selection pressure, thereby representing a high profile antiviral drug target. Guided by molecular dynamics simulations, we previously designed first-in-class M2-V27A inhibitors. One of the potent lead compounds, spiroadamantane amine (3), inhibits both the M2-WT and M2-V27A mutant with IC50 values of 18.7 and 0.3 µM, respectively, in in vitro electrophysiological assays. Encouraged by these findings, in this study we further examine the in vitro and in vivo antiviral activity of compound 3 in inhibiting both amantadine-sensitive and -resistant influenza A viruses. Compound 3 not only had single to sub-micromolar EC50 values against M2-WT- and M2-V27A-containing influenza A viruses in antiviral assays, but also rescued mice from lethal viral infection by either M2-WT- or M2-V27A-containing influenza A viruses. In addition, we report the design of two analogs of compound 3, and one was found to have improved in vitro antiviral activity over compound 3. Collectively, this study represents the first report demonstrating the in vivo antiviral efficacy of inhibitors targeting M2 mutants. The results suggest that inhibitors targeting drug-resistant M2 mutants are promising antiviral drug candidates worthy of further development.
Asunto(s)
Adamantano/análogos & derivados , Amantadina/farmacología , Antivirales/farmacología , Farmacorresistencia Viral , Virus de la Influenza A/efectos de los fármacos , Compuestos de Espiro/farmacología , Proteínas de la Matriz Viral/antagonistas & inhibidores , Proteínas de la Matriz Viral/genética , Adamantano/química , Adamantano/farmacología , Animales , Antivirales/química , Perros , Humanos , Gripe Humana/tratamiento farmacológico , Concentración 50 Inhibidora , Células de Riñón Canino Madin Darby , Simulación de Dinámica Molecular , Mutación , Infecciones por Orthomyxoviridae/tratamiento farmacológico , Compuestos de Espiro/química , Relación Estructura-ActividadRESUMEN
JNJ63623872 (formerly known as VX-787) is an inhibitor of influenza A virus polymerases through interaction with the viral PB2 subunit. This interaction blocks the cap-snatching activity of the virus that is essential for virus replication. Previously published work has documented antiviral activity of JNJ63623872 in cell culture and mouse infection studies. In this report, we extend the in vivo observations by comparing the efficacies of JNJ63623872 and oseltamivir in mice infected with influenza A/California/04/2009 (H1N1pdm) and A/Victoria/3/75 (H3N2) viruses. Animals received JNJ63623872 or oseltamivir orally twice daily for 10 days starting 2 h pre-infection. JNJ63623872 (2, 6, and 20 mg/kg/day) and oseltamivir (20 mg/kg/day) completely prevented death in the H1N1pdm virus infection. Weight loss at nadir was only 12% in mice receiving 2 mg/kg/day of JNJ63623872 compared to 23% and 32%, respectively, in oseltamivir-treated (20 mg/kg/day) and placebo groups. Lung hemorrhage scores, lung weights, and lung virus titers on day 6 were reduced in a dose-responsive manner by JNJ63623872 treatments, whereas oseltamivir treatments were not as effective. JNJ63623872 was less active against H3N2 virus infection, with more body weight loss occurring and only 30% survival at the 2-mg/kg/day dose. Lung scores, lung weights, and H3N2 viral titers in lungs of mice were reduced less by JNJ63623872 treatments compared to the H1N1pdm infection. Nevertheless, the 20-mg/kg/day dose of JNJ63623872 was more effective than oseltamivir (20 mg/kg/day) in improving body weight and reducing the severity of lung infection. JNJ63623872 appears to be an important new drug candidate to treat influenza A H1N1pdm and H3N2 virus infections.
Asunto(s)
Antivirales/uso terapéutico , Indoles/uso terapéutico , Subtipo H1N1 del Virus de la Influenza A/efectos de los fármacos , Subtipo H3N2 del Virus de la Influenza A/efectos de los fármacos , Infecciones por Orthomyxoviridae/tratamiento farmacológico , Oseltamivir/uso terapéutico , Animales , Descubrimiento de Drogas , Quimioterapia Combinada , Indoles/administración & dosificación , Pulmón/virología , Ratones , Infecciones por Orthomyxoviridae/virología , Oseltamivir/administración & dosificación , Piridinas , Pirimidinas , Pirroles , Carga Viral/efectos de los fármacos , Replicación Viral/efectos de los fármacosRESUMEN
Compounds were evaluated for antiviral activity in rhabdomyosarcoma (RD) cells against a recent 2014 clinical isolate of enterovirus D68 (EV-D68), a 1962 strain of EV-68D, rhinovirus 87 (RV-87, serologically the same as EV-D68), and enterovirus 71 (EV-71). Test substances included known-active antipicornavirus agents (enviroxime, guanidine HCl, pirodavir, pleconaril, and rupintrivir), nucleobase/nucleoside analogs (3-deazaguanine and ribavirin), and three novel epidithiodiketopiperazines (KCN-2,2'-epi-19, KCN-19, and KCN-21). Of these, rupintrivir was the most potent, with 50% inhibition of viral cytopathic effect (EC50) and 90% inhibition (EC90) of virus yield at 0.0022-0.0053 µM against EV-D68. Enviroxime, pleconaril and the KCN compounds showed efficacy at 0.01-0.3 µM; 3-deazaguanine and pirodavir inhibited EV-D68 at 7-13 µM, and guanidine HCl and ribavirin were inhibitory at 80-135 µM. Pirodavir was active against EV-71 (EC50 of 0.78 µM) but not against RV-87 or EV-D68, and all other compounds were less effective against EV-71 than against RV-87 and EV-D68. The most promising compound inhibiting both virus infections at low concentrations was rupintrivir. Antiviral activity was confirmed for the ten compounds in virus yield reduction (VYR) assays in RD cells, and for enviroxime, guanidine HCl, and pirodavir by cytopathic effect (CPE) assays in A549, HeLa-Ohio-1, and RD cells. These studies may serve as a basis for further pre-clinical discovery of anti-enterovirus inhibitors. Furthermore, the antiviral profiles and growth characteristics observed herein support the assertion that EV-D68 should be classified together with RV-87.
Asunto(s)
Antivirales/farmacología , Enterovirus Humano A/efectos de los fármacos , Enterovirus Humano D/efectos de los fármacos , Rhinovirus/efectos de los fármacos , Células A549 , Antimetabolitos/farmacología , Bencimidazoles/farmacología , Enterovirus Humano A/crecimiento & desarrollo , Enterovirus Humano D/crecimiento & desarrollo , Guanina/análogos & derivados , Guanina/farmacología , Células HeLa , Humanos , Oxadiazoles/farmacología , Oxazoles , Oximas , Picornaviridae/efectos de los fármacos , Piperazinas/farmacología , Piperidinas/farmacología , Piridazinas/farmacología , Rabdomiosarcoma , Rhinovirus/crecimiento & desarrollo , Ribavirina/farmacología , SulfonamidasRESUMEN
Iminosugars that are competitive inhibitors of endoplasmic reticulum (ER) α-glucosidases have been demonstrated to have antiviral activity against a diverse set of viruses. A novel iminosugar, UV-4B, has recently been shown to provide protection against lethal infections with dengue and influenza A (H1N1) viruses in mice. In the current study, the breadth of activity of UV-4B against influenza was examined ex vivo and in vivo. Efficacy of UV-4B against influenza A and B viruses was shown in primary human bronchial epithelial cells, a principal target tissue for influenza. Efficacy of UV-4B against influenza A (H1N1 and H3N2 subtypes) and influenza B was demonstrated using multiple lethal mouse models with readouts including mortality and weight loss. Clinical trials are ongoing to demonstrate safety of UV-4B and future studies to evaluate antiviral activity against influenza in humans are planned.
Asunto(s)
1-Desoxinojirimicina/análogos & derivados , Antivirales/administración & dosificación , Virus de la Influenza A/efectos de los fármacos , Virus de la Influenza B/efectos de los fármacos , Infecciones por Orthomyxoviridae/tratamiento farmacológico , 1-Desoxinojirimicina/administración & dosificación , 1-Desoxinojirimicina/farmacología , Animales , Antivirales/farmacología , Peso Corporal , Células Cultivadas , Modelos Animales de Enfermedad , Células Epiteliales/virología , Humanos , Ratones , Infecciones por Orthomyxoviridae/patología , Infecciones por Orthomyxoviridae/virología , Análisis de Supervivencia , Resultado del TratamientoRESUMEN
Broadly neutralizing antibodies targeting a highly conserved region in the hemagglutinin (HA) stem protect against influenza infection. Here, we investigate the protective efficacy of a protein (HB36.6) computationally designed to bind with high affinity to the same region in the HA stem. We show that intranasal delivery of HB36.6 affords protection in mice lethally challenged with diverse strains of influenza independent of Fc-mediated effector functions or a host antiviral immune response. This designed protein prevents infection when given as a single dose of 6.0 mg/kg up to 48 hours before viral challenge and significantly reduces disease when administered as a daily therapeutic after challenge. A single dose of 10.0 mg/kg HB36.6 administered 1-day post-challenge resulted in substantially better protection than 10 doses of oseltamivir administered twice daily for 5 days. Thus, binding of HB36.6 to the influenza HA stem region alone, independent of a host response, is sufficient to reduce viral infection and replication in vivo. These studies demonstrate the potential of computationally designed binding proteins as a new class of antivirals for influenza.
Asunto(s)
Anticuerpos Antivirales/inmunología , Proteínas Portadoras/inmunología , Glicoproteínas Hemaglutininas del Virus de la Influenza/inmunología , Virus de la Influenza A/inmunología , Gripe Humana/inmunología , Animales , Anticuerpos Neutralizantes/inmunología , Proteínas Portadoras/química , Proteínas Portadoras/metabolismo , Citocinas/metabolismo , Humanos , Virus de la Influenza A/genética , Gripe Humana/virología , Ratones , Modelos Moleculares , Mutación , Replicación ViralRESUMEN
UNLABELLED: Arbidol (ARB) is a synthetic antiviral originally developed to combat influenza viruses. ARB is currently used clinically in several countries but not in North America. We have previously shown that ARB inhibits in vitro hepatitis C virus (HCV) by blocking HCV entry and replication. In this report, we expand the list of viruses that are inhibited by ARB and demonstrate that ARB suppresses in vitro infection of mammalian cells with Ebola virus (EBOV), Tacaribe arenavirus, and human herpesvirus 8 (HHV-8). We also confirm suppression of hepatitis B virus and poliovirus by ARB. ARB inhibited EBOV Zaire Kikwit infection when added before or at the same time as virus infection and was less effective when added 24 h after EBOV infection. Experiments with recombinant vesicular stomatitis virus (VSV) expressing the EBOV Zaire glycoprotein showed that infection was inhibited by ARB at early stages, most likely at the level of viral entry into host cells. ARB inhibited HHV-8 replication to a similar degree as cidofovir. Our data broaden the spectrum of antiviral efficacy of ARB to include globally prevalent viruses that cause significant morbidity and mortality. IMPORTANCE: There are many globally prevalent viruses for which there are no licensed vaccines or antiviral medicines. Some of these viruses, such as Ebola virus or members of the arenavirus family, rapidly cause severe hemorrhagic diseases that can be fatal. Other viruses, such as hepatitis B virus or human herpesvirus 8 (HHV-8), establish persistent infections that cause chronic illnesses, including cancer. Thus, finding an affordable, effective, and safe drug that blocks many viruses remains an unmet medical need. The antiviral drug arbidol (ARB), already in clinical use in several countries as an anti-influenza treatment, has been previously shown to suppress the growth of many viruses. In this report, we expand the list of viruses that are blocked by ARB in a laboratory setting to include Ebola virus, Tacaribe arenavirus, and HHV-8, and we propose ARB as a broad-spectrum antiviral drug that may be useful against hemorrhagic viruses.
Asunto(s)
Antivirales/farmacología , Indoles/farmacología , Virus/efectos de los fármacos , Animales , Línea Celular , Humanos , Pruebas de Sensibilidad Microbiana , Modelos Moleculares , Estructura Molecular , Internalización del Virus/efectos de los fármacos , Replicación Viral/efectos de los fármacosRESUMEN
Favipiravir is approved in Japan to treat novel or re-emerging influenza viruses, and is active against a broad spectrum of RNA viruses, including Ebola. Ribavirin is the only other licensed drug with activity against multiple RNA viruses. Recent studies show that ribavirin and favipiravir act synergistically to inhibit bunyavirus infections in cultured cells and laboratory mice, likely due to their different mechanisms of action. Convalescent immune globulin is the only approved treatment for Argentine hemorrhagic fever caused by the rodent-borne Junin arenavirus. We previously reported that favipiravir is highly effective in a number of small animal models of Argentine hemorrhagic fever. We now report that addition of low dose of ribavirin synergistically potentiates the activity of favipiravir against Junin virus infection of guinea pigs and another arenavirus, Pichinde virus infection of hamsters. This suggests that the efficacy of favipiravir against hemorrhagic fever viruses can be further enhanced through the addition of low-dose ribavirin.
Asunto(s)
Amidas/farmacología , Antivirales/farmacología , Fiebres Hemorrágicas Virales/tratamiento farmacológico , Pirazinas/farmacología , Virus ARN/efectos de los fármacos , Ribavirina/farmacología , Animales , Arenavirus/efectos de los fármacos , Chlorocebus aethiops , Cricetinae , Virus del Dengue/efectos de los fármacos , Modelos Animales de Enfermedad , Sinergismo Farmacológico , Femenino , Cobayas , Orthohantavirus/efectos de los fármacos , Virus de la Fiebre Hemorrágica de Crimea-Congo/efectos de los fármacos , Fiebre Hemorrágica Americana/tratamiento farmacológico , Fiebre Hemorrágica Ebola/tratamiento farmacológico , Fiebres Hemorrágicas Virales/sangre , Fiebres Hemorrágicas Virales/veterinaria , Fiebres Hemorrágicas Virales/virología , Virus Junin/efectos de los fármacos , Masculino , Mesocricetus , Ratones , Células VeroRESUMEN
A key effector route of the Sugar Code involves lectins that exert crucial regulatory controls by targeting distinct cellular glycans. We demonstrate that a single amino-acid substitution in a banana lectin, replacing histidine 84 with a threonine, significantly reduces its mitogenicity, while preserving its broad-spectrum antiviral potency. X-ray crystallography, NMR spectroscopy, and glycocluster assays reveal that loss of mitogenicity is strongly correlated with loss of pi-pi stacking between aromatic amino acids H84 and Y83, which removes a wall separating two carbohydrate binding sites, thus diminishing multivalent interactions. On the other hand, monovalent interactions and antiviral activity are preserved by retaining other wild-type conformational features and possibly through unique contacts involving the T84 side chain. Through such fine-tuning, target selection and downstream effects of a lectin can be modulated so as to knock down one activity, while preserving another, thus providing tools for therapeutics and for understanding the Sugar Code.