RESUMEN
L: -FMAU [1-(2-fluoro-5-methyl-beta,L-arabinofuranosyl) uracil] has been shown to be an effective inhibitor of hepatitis B virus (HBV) and duck hepatitis B virus replication in cell culture and duck hepatitis B virus replication in acutely infected Peking ducks. The woodchuck hepatitis virus (WHV) and its natural host, the Eastern woodchuck (Marmota monax), have been established as a predictive model for the evaluation of antiviral therapies against chronic HBV infection. In this report, the antiviral activity of l-FMAU against WHV replication in chronically infected woodchucks is described. Four weeks of once-daily oral administration of L-FMAU significantly reduced viremia, antigenemia, intrahepatic WHV replication, and intrahepatic expression of woodchuck hepatitis virus core antigen (WHcAg) in a dose-dependent manner. At the highest dose administered (10 mg/kg/d), significant reductions of intrahepatic WHV RNA and covalently closed circular (ccc)WHV-DNA levels also were observed. The reduction in viremia was remarkably rapid at the higher doses of L-FMAU, with greater than 1,000-fold reductions in WHV-DNA serum levels observed after as little as 2 to 3 days of therapy. Following the withdrawal of therapy, a dose-related delay in viremia rebound was observed. At the highest doses used, viremia remained significantly suppressed in at least one half of the treated animals for 10 to 12 weeks' posttreatment. No evidence of drug-related toxicity was observed in the treated animals. L-FMAU is an exceptionally potent antihepadnaviral agent in vitro and in vivo, and is a suitable candidate for antiviral therapy of chronic HBV infection.
Asunto(s)
Antivirales/farmacología , Arabinofuranosil Uracilo/fisiología , Expresión Génica/efectos de los fármacos , Genes Virales/genética , Virus de la Hepatitis B de la Marmota/crecimiento & desarrollo , Virus de la Hepatitis B de la Marmota/genética , Hepatitis B Crónica/virología , Replicación Viral/efectos de los fármacos , Animales , Antígenos de Superficie/análisis , Arabinofuranosil Uracilo/análogos & derivados , Replicación del ADN/efectos de los fármacos , ADN Circular/antagonistas & inhibidores , ADN Viral/antagonistas & inhibidores , ADN Viral/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Antígenos de la Hepatitis/análisis , Virus de la Hepatitis B de la Marmota/efectos de los fármacos , Virus de la Hepatitis B de la Marmota/inmunología , Antígenos de la Hepatitis C/análisis , Marmota , ARN Viral/metabolismo , Factores de Tiempo , Viremia/prevención & controlRESUMEN
Cell culture studies in our laboratory previously demonstrated synergistic antiviral activity for the combinations of lamivudine and a novel recombinant hybrid human alpha B/D interferon (rHu alpha B/D IFN) against hepatitis B virus (HBV) replication. Based on these results, a study was designed to determine if an enhanced antiviral effect with this drug combination could be demonstrated in vivo using the woodchuck hepatitis virus (WHV)/woodchuck experimental model of chronic HBV infection. Both antiviral agents have been shown to be effective against WHV replication in WHV chronic carriers during previous studies by our laboratories. Two combination treatment regimens were compared to matched monotherapies in a placebo-controlled trial. The first used simultaneous administration of rHu alpha B/D IFN and lamivudine for 24 weeks. The other combination treatment regimen used a staggered dosing schedule of 12 weeks of administration of lamivudine alone, followed by 12 weeks of simultaneous dosing with both drugs, followed by 12 weeks of therapy with rHu alpha B/D IFN alone. Both treatment regimens with combinations of lamivudine and rHu alpha B/D IFN were more effective at reducing WHV replication in chronically infected wood-chucks than the corresponding monotherapies. Both combination treatments produced antiviral effects that were at least equal to that expected for additive activity based on estimations generated by Bliss Independence calculations. The staggered treatment regimen reduced viraemia and intrahepatic WHV replication significantly more than that expected for additive interactions, indicating synergistic antiviral effects. These studies demonstrate that combination therapy of chronic WHV infection has enhanced antiviral benefit over corresponding monotherapies and indicate that combination treatment of chronic HBV infection can be superior to therapies using a single antiviral agent.
Asunto(s)
Antivirales/uso terapéutico , Virus de la Hepatitis B de la Marmota/efectos de los fármacos , Hepatitis B Crónica/virología , Interferón Tipo I/uso terapéutico , Lamivudine/uso terapéutico , Replicación Viral/efectos de los fármacos , Animales , Portador Sano , Quimioterapia Combinada , Femenino , Hepatitis B Crónica/tratamiento farmacológico , Humanos , Interferón-alfa , Marmota , ARN Viral/sangre , Proteínas Recombinantes , ViremiaRESUMEN
Acute hepatitis B virus (HBV) infections either resolve or progress to chronicity. Identification of early deviations in host-virus responses associated with these outcomes can further differentiate cause-effect mechanisms that initiate and maintain chronicity. Neonatal woodchucks were infected experimentally with the woodchuck hepatitis virus (WHV) at 3 days of age. At 8 or 14 weeks of age (i.e. , the early- or mid-acute stage of infection), whole blood and large surgical biopsies of the liver were obtained from infected animals and uninfected controls. These were stored for later correlating histopathologic responses and viral load with the subsequently determined outcome of infection. As of 1 year postinfection, half of the surgically treated infected woodchucks had developed self-limited infections, while the other half developed chronic infections. The self-limited outcome was characterized by decreased viral load in acute-phase liver and plasma and a generally robust acute hepatic inflammatory response. Comparisons at the same early time points revealed that the chronic outcome was characterized by increasing initial viral load in liver and plasma, and a detectable, but diminished, acute hepatic inflammation. These cotemporal comparisons indicate that there is an early host-response deviation during the acute phase of a developing chronic infection. Continued analysis of the tissues banked from this study will facilitate further temporal characterization of acute-phase mechanisms that determine resolution versus chronicity in WHV infection. Understanding such mechanisms may be useful in the rational design of therapy for established chronic HBV infection.
Asunto(s)
Virus de la Hepatitis B de la Marmota , Hepatitis B Crónica/etiología , Enfermedad Aguda , Animales , Animales Recién Nacidos , ADN Viral/análisis , Antígenos de la Hepatitis/análisis , Hepatitis B Crónica/inmunología , Hepatitis B Crónica/virología , Hígado/patología , Hígado/virología , Marmota , NecrosisRESUMEN
Acyclovir triphosphate is a potent inhibitor of hepatitis B virus DNA polymerase, but acyclovir treatment provides no benefit in patients with hepatitis B virus infection. This is due in part to the fact that hepatitis B virus, unlike herpes simplex virus, does not code for a viral thymidine kinase which catalyzes the initial phosphorylation of acyclovir. We synthesized 1-O-octadecyl-sn-glycero-3-phospho (3-P)-acyclovir and found that it was highly active in reducing hepatitis B virus replication in 2.2. 15 cells, while acyclovir was inactive. The greater antiviral activity of 1-O-octadecyl-sn-glycero-3-P-acyclovir appeared to be due to liver cell metabolism of the compound to acyclovir monophosphate (K. Y. Hostetler et al., Biochem. Pharmacol. 53:1815-1822, 1997). However, a closely related compound without a hydroxyl group at the sn-2 position of glycerol, 1-O-hexadecylpropanediol-3-P-acyclovir, was more active and selective in 2.2.15 cells in vitro. In this study, we treated woodchucks chronically infected with woodchuck hepatitis virus with increasing oral doses of 1-O-hexadecylpropanediol-3-P-acyclovir and assessed the response to therapy versus acyclovir or a placebo. At a dosage of 10 mg/kg of body weight twice a day, the test compound significantly inhibited viral replication in vivo, as indicated by a 95% reduction in serum woodchuck hepatitis virus DNA levels and by a 54% reduction in levels of woodchuck hepatitis virus replicative intermediates in the liver. Higher doses were somewhat less effective. In contrast, 20 mg of acyclovir/kg twice daily, a 5. 3-fold-higher molar dosage, had no demonstrable activity against woodchuck hepatitis virus. Oral 1-O-hexadecylpropanediol-3-P-acyclovir appeared to be safe and effective in chronic woodchuck hepatitis virus infection.
Asunto(s)
Aciclovir/análogos & derivados , Aciclovir/uso terapéutico , Antivirales/uso terapéutico , Hepatitis B/tratamiento farmacológico , Aciclovir/farmacología , Administración Oral , Animales , Antivirales/farmacología , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Femenino , Virus de la Hepatitis B de la Marmota/efectos de los fármacos , Masculino , Marmota/virología , Resultado del TratamientoRESUMEN
The woodchuck hepatitis virus (WHV) and its natural host, the Eastern woodchuck (Marmota monax), have been established as a model of hepatitis B virus (HBV)-induced disease. Several published studies have used this experimental animal model system to demonstrate potential antiviral therapies for chronic HBV infections. However, there has been little comparative information available on compounds used in clinical anti-HBV studies in WHV-infected woodchucks, thereby making interpretations of the potential relative effectiveness of new antiviral agents in humans more difficult. In this report, using a series of placebo-controlled studies, we compared the relative effectiveness of several nucleoside analogues that have been used in clinical trials for the treatment of chronic HBV infection against WHV replication in chronically infected woodchucks. Adenine-5'-arabinoside monophosphate (Ara-AMP [vidarabine]), ribavirin, (-)beta-L-2',3'-dideoxy-3'-thiacytidine (3TC [lamivudine]), and famciclovir (oral prodrug of penciclovir) induced depressions in viremia and intrahepatic WHV-DNA replication that were consistent with their relative effectiveness in anti-HBV human clinical trials. As observed in HBV-infected patients, 3' azido-3'-deoxythymidine (AZT [zidovudine]) had no effect on WHV replication in these studies. These experimental results more firmly establish chronic WHV infection in woodchucks as an accurate and predictive model for antiviral therapies against chronic HBV infection in humans and provide a baseline for comparative antiviral effects of other experimental antiviral agents in the WHV/woodchuck model system.
Asunto(s)
Antivirales/uso terapéutico , Virus de la Hepatitis B de la Marmota , Hepatitis B Crónica/tratamiento farmacológico , 2-Aminopurina/análogos & derivados , 2-Aminopurina/uso terapéutico , Animales , Famciclovir , Hepatitis B Crónica/veterinaria , Humanos , Marmota , Ribavirina/uso terapéutico , Fosfato de Vidarabina/uso terapéutico , Replicación Viral/efectos de los fármacos , Zidovudina/uso terapéuticoRESUMEN
Emtricitabine [(-)FTC] [(-)-beta-2', 3'-dideoxy-5-fluoro-3'-thiacytidine] has been shown to be an effective inhibitor of hepatitis B virus (HBV) in cell culture, with a potency and selectivity that are essentially identical to those of lamivudine. The antiviral activity of oral administration of (-)FTC against WHV replication in chronically infected woodchucks, an established and predictive model for antiviral therapy against HBV, was examined in a placebo-controlled study. (-)FTC significantly reduced viremia and intrahepatic WHV replication in a dose-dependent manner that was comparable to the antiviral activity of lamivudine observed in previous studies conducted by our laboratories. No effect on the levels of hepatic WHV RNA or the levels of woodchuck hepatitis surface antigen or anti-woodchuck hepatitis surface and core antibodies in the serum of the treated animals was observed. No evidence of drug-related toxicity was observed in any of the animals treated.
Asunto(s)
Antivirales/administración & dosificación , Desoxicitidina/análogos & derivados , Virus de la Hepatitis B de la Marmota/efectos de los fármacos , Hepatitis B/tratamiento farmacológico , Administración Oral , Animales , Desoxicitidina/administración & dosificación , Emtricitabina , Virus de la Hepatitis B de la Marmota/fisiología , Marmota , Replicación Viral/efectos de los fármacosRESUMEN
Cell culture studies in our laboratory and others have previously demonstrated synergistic antiviral activity for combinations of 3TC (lamivudine) and penciclovir against Hepatitis B Virus (HBV) replication and the Duck Hepatitis B Virus (DHBV). Based on these results, a study was designed to determine if an enhanced antiviral effect with combinations of 3TC and famciclovir (FCV, oral prodrug of penciclovir) could be demonstrated in vivo using the Woodchuck Hepatitis Virus (WHV)/woodchuck experimental model of chronic HBV infection. Both antiviral agents have been shown to be effective against WHV replication in WHV chronic carriers in previous studies by our laboratories. The antiviral effects of four different combinations of lamivudine and FCV were found to be greater than those observed for the corresponding monotherapies. All four combination treatments produced antiviral effects that were at least equal to that expected for additive activity based on estimations generated by Bliss Independence calculations. Two of the combination treatments produced antiviral effects that were significantly greater than that expected for additive effects, indicative of synergistic antiviral interactions. These studies demonstrate that combination therapy of chronic WHV infection has enhanced antiviral benefit over corresponding monotherapies and indicate that combination treatment of chronic HBV infection can be superior to therapies using a single antiviral agent.
Asunto(s)
2-Aminopurina/análogos & derivados , Antivirales/uso terapéutico , Virus de la Hepatitis B de la Marmota/efectos de los fármacos , Hepatitis B Crónica/tratamiento farmacológico , Lamivudine/uso terapéutico , Inhibidores de la Transcriptasa Inversa/uso terapéutico , 2-Aminopurina/uso terapéutico , Animales , Portador Sano , Modelos Animales de Enfermedad , Quimioterapia Combinada , Famciclovir , Marmota , Profármacos/uso terapéutico , Viremia/tratamiento farmacológico , Viremia/virología , Replicación Viral/efectos de los fármacosRESUMEN
Acute hepadnavirus infections either resolve or progress to chronicity. Factors that influence chronicity as an outcome of hepatitis B virus (HBV) infection in humans can be studied experimentally in the woodchuck model. Accordingly, several woodchuck hepatitis virus (WHV) inocula were characterized. Representative inocula had high titers of infectious virus (approximately 10(7.7)-10(9.5) woodchuck 50% infectious doses per milliliter [WID(50%)/mL] by subcutaneous inoculation), with 1 WID(50%) ranging between 21 and 357 physical virion particles. WHV7P1 (standard high dose, 5 x 10(6) WID(50%)) produced a 72% chronicity rate (i.e., percent chronic of total infected) in neonatal woodchucks (1-3 days old). Comparable doses of WHV8P1 resulted in a lower chronicity rate in neonates (34% chronic) indicating that it represented a strain different from WHV7P1. Neonatal woodchucks were more susceptible to chronic infection by high doses of WHV7P1 (range, 65%-75% chronic) compared with 8-week-old weanlings (33% chronic) and adult woodchucks (0% chronic; i.e., all resolved). High doses of cloned wild-type viruses also induced high rates of chronicity in neonates (70%-80% chronic). Chronicity rates in neonates were decreased for low doses of WHV7P1 (500 WID(50%), 9% chronic) and for high doses of a precore WHeAg-minus mutant WHV8 clone (17% chronic). Thus, both age and viral determinants can influence chronicity as an outcome of experimental WHV infection. Standardized inocula will enable the study of mechanisms that initiate and maintain chronic hepadnavirus infection and also provide a means for developing WHV carriers for therapeutic studies.
Asunto(s)
Envejecimiento , Virus de la Hepatitis B de la Marmota , Hepatitis B Crónica/virología , Marmota , Animales , Animales Recién Nacidos , Antígenos Virales/análisis , ADN Viral/análisis , Susceptibilidad a Enfermedades , Virus de la Hepatitis B de la Marmota/clasificación , Virus de la Hepatitis B de la Marmota/genética , Virus de la Hepatitis B de la Marmota/patogenicidad , Mutación , Especificidad de la Especie , DesteteRESUMEN
A recently developed transgenic mouse strain which expresses high levels of hepatitis B virus (HBV) was studied as a model for evaluation of potential chemotherapeutic agents. Lamivudine ([-]2'-deoxy-3'-thiacytidine), known to reduce hepatitis B viremia in human patients, and zidovudine (3'-azido-3'-deoxythymidine), previously shown to be ineffective for HBV infections in man, were used in parallel in this transgenic animal model. Orally administered lamivudine at dosages of 100, 50, and 25 mg/kg per day given once a day for 21 days significantly decreased serum and liver HBV DNA titers in a dose-responsive manner. Zidovudine (approximately 22 mg/kg per day) administered in the drinking water for 21 days was not effective in reducing these HBV parameters as compared to transgenic placebo-treated controls. The serum HBV DNA titers rebounded to high levels 1 week after cessation of lamivudine treatment. Male and female mice responded in a similar manner to these therapies. The results using this transgenic mouse model were similar to what would be predicted from treatment of HBV-infected human patients with lamivudine and zidovudine, and indicate these mice may be useful as a small animal chemotherapeutic model for study of potential HBV inhibitors.
Asunto(s)
Antivirales/uso terapéutico , Virus de la Hepatitis B/efectos de los fármacos , Hepatitis Viral Animal/tratamiento farmacológico , Lamivudine/uso terapéutico , Replicación Viral/efectos de los fármacos , Animales , ADN Viral/sangre , ADN Viral/efectos de los fármacos , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Evaluación de Medicamentos , Femenino , Antígenos de Superficie de la Hepatitis B/sangre , Antígenos de Superficie de la Hepatitis B/efectos de los fármacos , Antígenos e de la Hepatitis B/sangre , Antígenos e de la Hepatitis B/efectos de los fármacos , Virus de la Hepatitis B/genética , Humanos , Masculino , Ratones , Ratones Transgénicos , Factores de Tiempo , Resultado del Tratamiento , Zidovudina/uso terapéuticoRESUMEN
The synthesis and in vitro anti-hepatitis B virus (HBV) activity of two mononucleoside phosphotriester derivatives of acyclovir incorporating S-acyl-2-thioethyl (SATE) groups are reported. In contrast to the parent nucleoside, the described phosphotriesters emerged as potent and selective inhibitors of HBV replication in HepG2.2.15 cells. This result can be attributed to the unique cellular metabolism of the SATE pronucleotides giving rise to the delivery to acyclovir 5'-monophosphate inside the infected cells. Moreover, the in vitro anti-HBV activities of one of these bis(SATE)phosphotriesters and of (-)-beta-L-2',3'-dideoxy-3'-thiacytidine (lamivudine, 3TC) were compared alone and in combination. Analysis of the combination data indicates that 3TC and the studied SATE pronucleotide of acyclovir exhibited strong synergistic interactions. The present study provides an example where the use of a pronucleotide approach extends the antiviral spectrum of a nucleoside analogue. Given the potency of SATE pronucleotides of acyclovir against HBV in HepG2.2.15 cells, further studies including animal experiments seem warranted to evaluate the potential of these compounds as anti-HBV agents.
Asunto(s)
Aciclovir/análogos & derivados , Antivirales/farmacología , Virus de la Hepatitis B/efectos de los fármacos , Organofosfonatos/farmacología , Aciclovir/síntesis química , Aciclovir/química , Aciclovir/farmacología , Antivirales/síntesis química , Antivirales/química , Evaluación Preclínica de Medicamentos , Sinergismo Farmacológico , Virus de la Hepatitis B/crecimiento & desarrollo , Hepatoblastoma , Humanos , Lamivudine/farmacología , Neoplasias Hepáticas , Pruebas de Sensibilidad Microbiana , Organofosfonatos/síntesis química , Organofosfonatos/química , Profármacos/síntesis química , Profármacos/química , Profármacos/metabolismo , Inhibidores de la Transcriptasa Inversa/farmacología , Células Tumorales CultivadasRESUMEN
Robustaflavone, a naturally occurring biflavanoid isolated from Rhus succedanea, was found to be a potent inhibitor of hepatitis B virus (HBV) replication in 2.2.15 cells, with an effective concentration (EC50) of 0.25 microM, and a selectivity index (SI, IC50/EC90) of 153. Robustaflavone hexaacetate inhibited HBV replication with an EC50 of 0.73 microM, but exhibited no cytotoxicity at concentrations up to 1000 microM. Combinations of robustaflavone with penciclovir and lamivudine displayed synergistic anti-HBV activity, having the most pronounced effects when the combination ratios were similar to the ratio of EC50 potencies. Thus, a 1:1 combination of robustaflavone and penciclovir exhibited an EC50 of 0.11 microM and an SI of 684, while a 10:1 combination of robustaflavone and lamivudine exhibited an EC50 of 0.054 microM and an SI of 894. Statistical analyses of the combination data using the Combostat program confirmed that robustaflavone exhibited synergism with both penciclovir and lamivudine.
Asunto(s)
Antivirales/farmacología , Biflavonoides , Flavonoides/farmacología , Virus de la Hepatitis B/efectos de los fármacos , Aciclovir/análogos & derivados , Aciclovir/farmacología , Línea Celular , Sinergismo Farmacológico , Guanina , Virus de la Hepatitis B/fisiología , Lamivudine/farmacología , Pruebas de Sensibilidad Microbiana , Replicación Viral/efectos de los fármacosRESUMEN
The pharmacodynamics of (-)-beta-2',3'-dideoxy-3'-thiacytidine (3TC) was studied in chronically woodchuck hepatitis virus-infected woodchucks and compared to that in previous studies in hepatitis B virus (HBV)-infected humans. Net depletion rates of serum virus DNA in woodchucks receiving 3TC were modeled as a sum of an exponentially declining virus input and a first-order elimination. Preceding shoulders and pseudo-first-order virus half-lives in serum ranged from 1 to 7 days and were dose dependent. Higher plasma 3TC concentrations were needed in woodchucks for virus depletion similar to that attained in humans. Human HBV depletion curves from a previous clinical study with 3TC (>/=100 mg per day) were described by a biexponential relationship. The average half-life value in humans, normalized to fraction of area under the serum virus load-time curve, was similar to the average half-life value observed in woodchucks given the highest 3TC dose (2.4 and 2.0 days, respectively). On cessation of therapy, virus load rebounds in woodchucks were dose dependent and resembled posttherapy virus "flares" reported to occur in humans. The estimates of drug exposures that could lead to optimal antiviral effects presented indicate that 3TC should not be underdosed and compliance should be monitored. The study of chronically infected woodchucks may prove useful for optimizing drug regimens for hepadnavirus infections.
Asunto(s)
Virus de la Hepatitis B de la Marmota/efectos de los fármacos , Virus de la Hepatitis B/efectos de los fármacos , Lamivudine/farmacología , Animales , Relación Dosis-Respuesta a Droga , Femenino , Humanos , Masculino , Marmota , Viremia/tratamiento farmacológico , Viremia/virologíaRESUMEN
Woodchucks were used to study the antiviral activity and toxicity of fialuridine (FIAU; 1,-2'deoxy-2'fluoro-1-beta-D-arabinofuranosyl-5-iodo-uracil). In an initial experiment, groups of six chronic woodchuck hepatitis virus (WHV) carrier woodchucks received daily doses of FIAU by intraperitoneal injection for 4 weeks. At 0.3 mg/kg/d, the antiviral effect was equivocal, but at 1.5 mg/kg/d, FIAU had significant antiviral activity. No evidence of drug toxicity was observed during the 4-week period of treatment or during posttreatment follow-up. In a second experiment, groups of nine WHV carriers or uninfected woodchucks were given 1.5 mg/kg/d of FIAU orally for 12 weeks, and the results compared with placebo-treated controls. After 4 weeks, the serum WHV-DNA concentration in the FIAU-treated carrier group was two to three logs lower than that in the placebo-treated group. After 12 weeks of FIAU treatment, serum WHV DNA was not detectable by conventional dot-blot analysis, hepatic WHV-DNA replicative intermediates (RI) had decreased 100-fold, and hepatic expression of WHV core antigen was remarkably decreased. No evidence of toxicity was observed after 4 weeks, but, after 6 to 7 weeks, food intake decreased and, after 8 weeks, the mean body weights of woodchucks treated with FIAU were significantly lower than controls. Anorexia, weight loss, muscle wasting, and lethargy became progressively severe, and all FIAU-treated woodchucks died or were euthanized 78 to 111 days after treatment began. Hepatic insufficiency (hyperbilirubinemia, decreased serum fibrinogen, elevated prothrombin time), lactic acidosis, and hepatic steatosis were characteristic findings in the final stages of FIAU toxicity in woodchucks. The syndrome of delayed toxicity in woodchucks was similar to that observed previously in humans treated with FIAU, suggesting that the woodchuck should be valuable in future investigations of the molecular mechanisms of FIAU toxicity in vivo and for preclinical toxicological evaluation of other nucleoside analogs before use in patients.
Asunto(s)
Antivirales/uso terapéutico , Arabinofuranosil Uracilo/análogos & derivados , Hepatitis B/tratamiento farmacológico , Animales , Anorexia/inducido químicamente , Antivirales/efectos adversos , Arabinofuranosil Uracilo/efectos adversos , Arabinofuranosil Uracilo/farmacocinética , Arabinofuranosil Uracilo/uso terapéutico , Portador Sano/virología , ADN Viral/análisis , Hepatitis B/sangre , Hepatitis B/patología , Antígenos del Núcleo de la Hepatitis B/análisis , Virus de la Hepatitis B/genética , Virus de la Hepatitis B/fisiología , Hígado/metabolismo , Hígado/patología , Marmota , Músculos/efectos de los fármacos , Fases del Sueño , Factores de Tiempo , Replicación Viral/efectos de los fármacosRESUMEN
Many transgenic mice carrying either portions of the hepatitis B virus (HBV) genome, or the complete genome, have been developed as models because HBV does not infect any other organisms besides humans and chimpanzees to cause a productive infection and disease. Some of these models have been useful in evaluating chemotherapeutic agents such as interferon-alpha, interleukins-2 and -12, other cytokines and nucleoside analogues for efficacy against HBV. A recently developed transgenic mouse (Guidotti et al., Journal of Virology 69:6158-6169.) which supports the replication of high levels of infectious HBV, provides the opportunity to evaluate the effect of antiviral drugs on various portions of the HBV life cycle in the whole animal. Evaluation of lamivudine, zidovudine and interferon-alpha B/D (IFN-alpha) in this HBV transgenic mouse model are described. Lamivudine and IFN-alpha were highly efficacious in reducing serum HBV DNA. As might be predicted, zidovudine was not efficacious. IFN-alpha was more effective in reducing virus titres in male mice as compared to female mice. This gender difference was not due to lower ability of female mice to express the virus. One anticipates that as this high level HBV transgenic-expressing mouse becomes more fully developed as a chemotherapeutic model, questions about the efficacy of different agents, routes of administration, synergy of antiviral combinations and novel drug therapies will be answered.
Asunto(s)
Modelos Animales de Enfermedad , Hepatitis B/tratamiento farmacológico , Ratones Transgénicos , Administración Oral , Animales , ADN Viral/sangre , Relación Dosis-Respuesta a Droga , Genoma Viral , Hepatitis B/virología , Antígenos de Superficie de la Hepatitis B/sangre , Virus de la Hepatitis B de la Marmota/genética , Antígenos e de la Hepatitis B/sangre , Virus de la Hepatitis B/genética , Virus de la Hepatitis B/inmunología , Humanos , Interferón-alfa/uso terapéutico , Lamivudine/uso terapéutico , Ratones , Reacción en Cadena de la Polimerasa , Factores Sexuales , Factores de Tiempo , Carga Viral , Replicación Viral , Zidovudina/administración & dosificación , Zidovudina/uso terapéuticoRESUMEN
Preclinical aspects of a potent anti-hepatitis B virus (HBV) L-nucleoside, 1-(2-fluoro-5-methyl-beta-L-arabino-furanosyl)uracil (L-FMAU) are described. L-FMAU was prepared from L-ribose derivatives via either L-xylose or L-arabinose. L-FMAU shows potent antiviral activity against hepatitis B virus (EC50 5.0 microM in H1 cells) with high selectivity in vitro. L-FMAU is not incorporated into mitochondrial DNA and no significant lactic acid production was observed in vitro. L-FMAU is phosphorylated by thymidine kinase as well as deoxycytidine kinase, ultimately to the triphosphate, which inhibits HBV DNA polymerase as the mechanism of antiviral action. Preliminary in vivo toxological studies suggest no apparent toxicity for 30 days at 50 mg/kg/day in mice and for 3 months in woodchucks (10 mg/kg/day). L-FMAU also has respectable bioavailability in rats. L-FMAU shows potent anti-HBV activity in vivo against woodchuck hepatitis virus in chronically infected woodchucks and there is no significant virus rebound after cessation of the drug treatment.
Asunto(s)
Antivirales/farmacología , Arabinofuranosil Uracilo/análogos & derivados , Virus de la Hepatitis B/efectos de los fármacos , Animales , Antivirales/toxicidad , Arabinofuranosil Uracilo/química , Arabinofuranosil Uracilo/farmacología , Arabinofuranosil Uracilo/toxicidad , Disponibilidad Biológica , Línea Celular , Desoxicitidina Quinasa/metabolismo , Evaluación Preclínica de Medicamentos , Patos , Infecciones por Hepadnaviridae/tratamiento farmacológico , Infecciones por Hepadnaviridae/virología , Virus de la Hepatitis B del Pato/efectos de los fármacos , Virus de la Hepatitis B de la Marmota/efectos de los fármacos , Hepatitis B Crónica/tratamiento farmacológico , Hepatitis B Crónica/virología , Ácido Láctico/sangre , Ácido Láctico/metabolismo , Dosificación Letal Mediana , Marmota , Ratones , Fosforilación , Ratas , Timidina Quinasa/metabolismoRESUMEN
An arabinogalactan conjugate containing a 9 kDa fragment of arabinogalactan and adenine-9-beta-D-arabinofuranoside 5'-monophosphate (araAMP), denoted AG(9 kDa)-araAMP, has been synthesized and characterized. In 2.2.15 (human hepatoblastoma) cells, the attachment of araAMP to AG(9 kDa), a ligand of the asialoglycoprotein receptor, decreased the effective concentration for inhibiting extracellular hepatitis B virus (HBV) production by 90% (EC90) from 17 to 0.9 microM adenine arabinoside (araA) equivalents, and increased the cytotoxic concentration (CC50) from 188 to > 17 300 microM araA equivalents. Hence, the selectivity index (CC50/EC90) of araA was improved from 11 (188/17) to > 19200 (17 300/0.9) by conjugation with the 9 kDa fragment of arabinogalactan. AG(9 kDa)-araAMP did not affect the production of viral RNA or viral proteins. In the woodchuck hepatitis model, AG(9 kDa)-araAMP inhibited woodchuck hepatitis virus (WHV) DNA replication at a dose of 0.3 mg of araA equivalents per kg; in this case, AG(9 kDa)-araAMP was 20-30 times more potent than was unconjugated araA. AG(9 kDa)-araAMP was effective by intramuscular or subcutaneous administration. The reduction in HBV DNA levels obtained in 2.2.15 cells and of WHV DNA levels in woodchucks was sustained after treatment with AG(9 kDa)-araAMP ceased. In both cases, viral DNA gradually returned to pre-treatment levels.
Asunto(s)
Antivirales/farmacología , Galactanos/farmacología , Fosfato de Vidarabina/farmacología , Animales , Antivirales/síntesis química , ADN Viral/análisis , Virus de la Hepatitis B de la Marmota/efectos de los fármacos , Virus de la Hepatitis B/efectos de los fármacos , Marmota , Peso MolecularRESUMEN
Penciclovir [9-(4-hydroxy-3-hydroxymethylbut-1-yI)guanine], an effective antiherpesvirus agent, was found to be a potent and selective antiviral agent against intracellular hepatitis B virus (HBV) replication (drug concentration at which a 10-fold decrease in HBV DNA from the average level in an untreated culture was observed [EC90], 1.6 microM) and extracellular virion release (EC90, 0.7 microM) by cultured human hepatoblastoma (2.2.15) cells. Acyclovir and three other related 9-alkoxypurines with activity against either herpesviruses or human immunodeficiency virus were uniformly inactive against HBV. The activity of penciclovir is discussed in relation to recent findings related to its mode of action against HBV.
Asunto(s)
Aciclovir/análogos & derivados , Antivirales/farmacología , Virus de la Hepatitis B/efectos de los fármacos , Aciclovir/farmacología , Línea Celular , ADN Polimerasa Dirigida por ADN/metabolismo , Didesoxinucleósidos/farmacología , Guanina , Virus de la Hepatitis B/enzimología , Hepatoblastoma , Humanos , Lamivudine/farmacología , Pruebas de Sensibilidad Microbiana , Replicación ViralRESUMEN
The HBV-producing human hepatoblastoma cell line, 2.2.15, has been shown to be an accurate model of chronic cellular viral infection and a predictive model of antiviral response for in vivo hepadnaviral infection. Our laboratory has utilized the 2.2.15 cell line in a standardized assay to examine treatment schemes which use combinations of clinically relevant nucleoside analogues, novel methods to deliver potentially useful nucleoside combinations, and treatments which simultaneously target different parts of the HBV replication pathway. For example, the combination of 3TC (lamivudine) with either alpha interferon or penciclovir significantly enhances the antiviral effectiveness of these agents against HBV replication in 2.2.15 cell culture.
Asunto(s)
Antivirales/administración & dosificación , Virus de la Hepatitis B/efectos de los fármacos , Aciclovir/análogos & derivados , Aciclovir/uso terapéutico , Sinergismo Farmacológico , Quimioterapia Combinada , Guanina , Hepatitis B/tratamiento farmacológico , Hepatoblastoma , Humanos , Interferón-alfa/uso terapéutico , Lamivudine , Células Tumorales Cultivadas , Replicación Viral/efectos de los fármacos , Zalcitabina/análogos & derivados , Zalcitabina/uso terapéuticoRESUMEN
Antisense oligonucleotides are currently being used in numerous laboratories as potential anticancer and antiviral agents. The unique replication cycle of hepatitis B virus (HBV) contains several different steps which are potentially amenable to modulation by these molecules. We have examined the ability of 56 different single-stranded, oligodeoxyribonucleotides (14-23 nucleotides in length), which target several HBV-specific functions, to inhibit HBV replication in the human hepatoblastoma cell line, 2.2.15. None of the oligonucleotides examined were toxic at concentrations up to 500 microM. Oligonucleotides directed against the HBV surface antigen (HBsAg) gene (S gene), the preS1 open reading frame, and the HBV core antigen (HBcAg) gene (C gene) were effective at depressing virus production, while molecules targeting the HBV e antigen (HBeAg) open reading frame and the HBV polymerase (POL) gene were ineffective. Oligonucleotides directed against the HBV encapsidation signal/structure (epsilon) comprised some of the most effective antiviral molecules against HBV. None of 5 oligonucleotides complementary (i.e., 'sense' orientation) to the antisense oligonucleotides targeting HBsAg, HBcAg, HBeAg, preS1 and POL had any measurable effect on HBV production. The relative effectiveness of oligonucleotides targeting the S and C genes on HBV replication was highly correlated with an effect on HBsAg or HBcAg levels, respectively. None of the antisense oligonucleotides examined affected either the levels or the sizes of HBV-specific RNA transcripts. Since antisense oligonucleotides can exert biologic effects on HBV in 2.2.15 cell cultures in a sequence-specific manner which are consistent with predicted modes of action, such molecules may have practical applications in the therapy of chronic HBV infection.
Asunto(s)
Antivirales/farmacología , Virus de la Hepatitis B/efectos de los fármacos , Oligonucleótidos Antisentido/farmacología , Secuencia de Bases , ADN Viral/efectos de los fármacos , Genes Virales , Antígenos de la Hepatitis B/biosíntesis , Antígenos del Núcleo de la Hepatitis B/biosíntesis , Antígenos de Superficie de la Hepatitis B/biosíntesis , Antígenos e de la Hepatitis B/biosíntesis , Virus de la Hepatitis B/crecimiento & desarrollo , Proteínas Virales/efectos de los fármacos , Replicación Viral/efectos de los fármacosRESUMEN
The nucleoside analog 2,4-diamino-7-(2-deoxy-2-fluoro-beta-D- arabinofuranosyl)pyrrolo[2,3-d]pyrimidine (T70080) and several related compounds were evaluated for anti-hepatitis B virus (HBV) activity by using cultured 2.2.15 cells. T70080 reduced episomal viral replication in these cells by 50% at a concentration of 0.7 microgram/ml. At the same time, T70080 reduced cellular proliferation by 50% at a concentration in excess of 100 micrograms/ml, yielding a therapeutic index of > 143. In cells cultured for 12 days in the presence of 10 or 50 micrograms of T70080 per ml and then with drug-free medium, for an additional 12 days, viral DNA replication was completely inhibited initially but resumed between 6 and 12 days post-drug removal. In view of the potent anti-HBV activity shown, T70080 is a good candidate for further evaluation as a treatment of human HBV infection.