peramivir and Disease-Models--Animal

Influenza B viruses are important human pathogens that remain inadequately studied, largely because available animal models are poorly defined. Here, we developed an immunocompromised murine models for influenza B virus infection, which we subsequently used to study pathogenicity and to examine antiviral efficacy of the neuraminidase inhibitor peramivir. We studied three influenza B viruses that represent both the Yamagata (B/Massachusetts/2/2012 and B/Phuket/3073/2013) and Victoria (B/Brisbane/60/2008, BR/08) lineages. BR/08 was the most pathogenic in genetically modified immunocompromised mice [BALB scid and non-obese diabetic (NOD) scid strains] causing lethal infection without prior adaptation. The immunocompromised mice demonstrated prolonged virus shedding with modest induction of immune responses compared to BALB/c. Rather than severe virus burden, BR/08 virus-associated disease severity correlated with extensive virus spread and severe pulmonary pathology, stronger and persistent natural killer cell responses, and the extended induction of pro-inflammatory cytokines and chemokines. In contrast to a single-dose treatment (75 mg/kg/day), repeated doses of peramivir rescued BALB scid mice from lethal challenge with BR/08, but did not result in complete virus clearance. In summary, we have established immunocompromised murine models for influenza B virus infection that will facilitate evaluations of the efficacy of currently available and investigational anti-influenza drugs.

Topics: Acids, Carbocyclic; Animals; Antiviral Agents; Bronchoalveolar Lavage Fluid; Cyclopentanes; Cytokines; Disease Models, Animal; Female; Guanidines; Immunocompromised Host; Inflammation Mediators; Influenza B virus; Mice; Mice, Inbred NOD; Mice, SCID; Morbidity; Mortality; Orthomyxoviridae Infections; Viral Load; Virus Replication

High morbidity and mortality associated with human cases of highly pathogenic avian influenza (HPAI) viruses, including H5N1 influenza virus, have been reported. The purpose of the present study was to evaluate the antiviral effects of peramivir against HPAI viruses. In neuraminidase (NA) inhibition and virus replication inhibition assays, peramivir showed strong inhibitory activity against H5N1, H7N1 and H7N7 HPAI viruses with sub-nanomolar activity in enzyme assays. In H5N1 viruses containing the NA H275Y mutation, the antiviral activity of peramivir against the variant was lower than that against the wild-type. Evaluation of the in vivo antiviral activity showed that a single intravenous treatment of peramivir (10 mg/kg) prevented lethality in mice infected with wild-type H5N1 virus and also following infection with H5N1 virus with the H275Y mutation after a 5 day administration of peramivir (30 mg/kg). Furthermore, mice injected with peramivir showed low viral titers and low levels of proinflammatory cytokines in the lungs. These results suggest that peramivir has therapeutic activity against HPAI viruses even if the virus harbors the NA H275Y mutation.

Topics: Acids, Carbocyclic; Animals; Antiviral Agents; Cyclopentanes; Cytokines; Disease Models, Animal; Guanidines; Humans; Influenza A Virus, H5N1 Subtype; Influenza A Virus, H7N1 Subtype; Influenza A Virus, H7N7 Subtype; Influenza, Human; Lung; Mice; Mutation; Neuraminidase; Orthomyxoviridae Infections; Viral Load; Virus Replication

China-made Peramivir, an anti-influenza neuraminidase inhibitor drug, is manufactured and widely used in China. Although effective if initiated within 48 h of the onset of symptoms, yet we observed that this drug shows an inconclusive efficacy if treatment is delayed in clinical. Thus we evaluated the efficacy of delayed treatment of China-made Peramivir in a mouse model.. The mouse model of influenza infection was made and Peramivir was administered intravenously for 5 days following infection, and weight loss, lung index, viral shedding and survival rates were monitored.. Peramivir (60 mg/kg · d, repeated intravenous injections, quaque die (QD) × 5 days) enhanced survival rate and suppressed weight loss when treatment was initiated 24, 36, 48, or even 60 h post-infection (p.i.) (p < 0.01), compared with the virus-untreated group, and efficacy was abolished at 72 h p.i.. However the efficacy of delayed treatment was dose dependent, with the highest dose (90 mg/kg · d) even showing efficacy at 72 h p.i.. Furthermore, Peramivir (60 mg/kg · d, repeated intravenous injections, QD × 5 days) also reduced the lung virus titer 24 and 36 h p.i. on day 5, and even at 48 and 60 h p.i. on day 7 after infection, and the lung index was also improved. What is interesting that the concentration of the drug was maintained in blood after infected.. Delayed treatment with China-made Peramivir can reduce the severity of influenza disease, accelerate viral clearance and enhance the survival rate. This drug therefore shows good efficacy and is a promising candidate to control the influenza epidemic in China.

Topics: Acids, Carbocyclic; Administration, Intravenous; Animals; Antiviral Agents; China; Cyclopentanes; Disease Models, Animal; Drug Administration Schedule; Female; Guanidines; Influenza A Virus, H1N1 Subtype; Injections, Intravenous; Lung; Male; Mice; Orthomyxoviridae Infections; Viral Load; Virus Shedding

Influenza virus infection increases susceptibility to bacterial infection and mortality in humans. Although the efficacy of approved intravenous peramivir, a neuraminidase (NA) inhibitor, against influenza virus infection has been reported, its efficacy against bacterial co-infection, which occurs during the period of viral shedding, was not fully investigated. To further understand the significance of treatment with peramivir, we assessed the efficacy of peramivir against a bacterial co-infection model in mice caused by clinically isolated influenza A(H1N1)pdm09 virus and Streptococcus pneumoniae.. Mice were infected with influenza A(H1N1)pdm09. Peramivir was intravenously administered after the viral infection. At 2days post viral infection, the mice were infected with S. pneumoniae. Peramivir efficacy was measured by the survival rates and viral titers, bacterial titers, or proinflammatory cytokine concentrations in lung homogenates.. Peramivir treatment reduced the mortality of mice infected with influenza virus and S. pneumoniae. The survival rate in the peramivir-treated group was significantly higher than that in the oseltamivir-treated group. Viral titers and proinflammatory cytokine responses in the peramivir-treated group were significantly lower than those in the oseltamivir-treated group until at 2days post viral infection. Bacterial titer was significantly lower in the peramivir-treated group than in the oseltamivir-treated group at 4days post viral infection.. These results demonstrated that peramivir inhibits viral replication, consequently leading to bacterial clearance and prevention of mortality during severe murine bacterial co-infection, which occurs during the period of viral shedding, with the efficacy of peramivir being superior to that of oseltamivir.

Topics: Acids, Carbocyclic; Administration, Intravenous; Animals; Antiviral Agents; Bacterial Load; Coinfection; Cyclopentanes; Cytokines; Disease Models, Animal; Dogs; Guanidines; Influenza A Virus, H1N1 Subtype; Madin Darby Canine Kidney Cells; Mice; Mice, Inbred BALB C; Orthomyxoviridae Infections; Oseltamivir; Pneumococcal Infections; Streptococcus pneumoniae; Viral Load; Virus Replication

The purpose of this study was to investigate the relationship between pharmacokinetic (PK) parameters of intravenous (IV) peramivir and in vivo antiviral activity pharmacodynamic (PD) outcomes in a mouse model of influenza virus infection. Peramivir was administrated to mice in three dosing schedules; once, twice and four times after infection of A/WS/33 (H1N1). The survival rate at day 14 after virus infection was employed as the antiviral activity outcome for analysis. The relationship between day 14 survival and PK parameters, including area under the concentration-time curve (AUC), maximum concentration (Cmax) and time that drug concentration exceeds IC95 (T(>IC95)), was estimated using a logistic regression model, and model fitness was evaluated by calculation of the Akaike information criterion (AIC) index. The AIC indices of AUC, Cmax and T(>IC95) were about 114, 151 and 124, respectively. The AIC of AUC and T(>IC95) were smaller than that of Cmax. Therefore, both AUC and T(>IC95) were the PK parameters that correlated best with the antiviral activity of peramivir IV against influenza virus infection in mice.

Topics: Acids, Carbocyclic; Animals; Antiviral Agents; Cyclopentanes; Disease Models, Animal; Female; Guanidines; Humans; Influenza, Human; Mice; Mice, Inbred BALB C

Favipiravir, an influenza virus RNA polymerase inhibitor, and peramivir, an influenza virus neuraminidase inhibitor, were evaluated alone and in combination against pandemic influenza A/California/04/2009 (H1N1) virus infections in mice. Infected mice were treated twice daily for 5 d starting 4 h after virus challenge. Favipiravir was 40%, 70%, and 100% protective at 20, 40, and 100 mg/kg/d. Peramivir was 30% protective at 0.5 mg/kg/d, but ineffective at lower doses when used as monotherapy. Combinations of favipiravir and peramivir increased the numbers of survivors by 10-50% when the 0.025, 0.05, and 0.1 mg/kg/d doses of peramivir were combined with 20 mg/kg/d favipiravir and when all doses of peramivir were combined with 40 mg/kg/d favipiravir. Three-dimensional analysis of drug interactions using the MacSynergy method indicates strong synergy for these drug combinations. In addition, an increase in lifespan for groups of mice treated with drug combinations, compared to the most effective monotherapy group, was observed for the 0.025, 0.05, and 0.1 mg/kg/d doses of peramivir combined with favipiravir at the 20 mg dose level. Therefore, the 20 mg/kg/d dose of favipiravir was selected for further combination studies. Increased survival was exhibited when this dose was combined with peramivir doses of 0.1, 0.25 and 0.5 mg/kg/d (1 mg/kg/d of peramivir alone was 100% protective in this experiment). Improved body weight relative to either compound alone was evident using 0.25, 0.5, and 1 mg/kg/d of peramivir. Significant reductions in lung hemorrhage score and lung weight were evident on day 6 post-infection. In addition, virus titers were reduced significantly on day 4 post-infection by combination therapy containing favipiravir combined with peramivir at 0.25 and 0.5 mg/kg/d. These data demonstrate that combinations of favipiravir and peramivir perform better than suboptimal doses of each compound alone for the treatment of influenza virus infections in mice.

Topics: Acids, Carbocyclic; Amides; Animals; Antiviral Agents; California; Cyclopentanes; Disease Models, Animal; Drug Synergism; Drug Therapy, Combination; Female; Guanidines; Humans; Influenza A Virus, H1N1 Subtype; Influenza, Human; Mice; Mice, Inbred BALB C; Pandemics; Pyrazines

New and emerging influenza virus strains, such as the pandemic influenza A (H1N1) virus require constant vigilance for antiviral drug sensitivity and resistance. Efficacy of intramuscularly (IM) administered neuraminidase (NA) inhibitor, peramivir, was evaluated in mice infected with recently isolated pandemic A/California/04/2009 (H1N1, swine origin, mouse adapted) influenza virus. A single IM injection of peramivir (four dose groups), given 1h prior to inoculation, significantly reduced weight loss (p < 0.001) and mortality (p < 0.05) in mice infected with LD90 dose of pandemic A/California/04/2009 (H1N1) influenza virus compared to vehicle group. There was 20% survival in the vehicle-treated group, whereas in the peramivir-treated groups, survival increased in a dose-dependent manner with 60, 60, 90 and 100% survivors for the 1, 3, 10, and 30 mg/kg doses, respectively. Weight loss on day 4 in the vehicle-treated group was 3.4 gm, and in the peramivir-treated groups was 2.1, 1.5, 1.8 and 1.8 g for the 1, 3, 10 and 30 mg/kg dose groups, respectively. In the treatment model, peramivir given 24h after infection as a single IM injection at 50mg/kg dose, showed significant protection against lethality and weight loss. There was 13% survival in the vehicle-treated group while in the peramivir-treated group at 24, 48, and 72 h post infection, survival was 100, 40, and 50%, respectively. Survival in the oseltamivir groups (10 mg/kg/d twice a day, orally for 5 days) was 90, 30 and 20% at 24, 48 and 72 h, respectively. These data demonstrate efficacy of parenterally administered peramivir against the recently isolated pandemic influenza virus in murine infection models.

Topics: Acids, Carbocyclic; Animals; Antiviral Agents; Body Weight; Cyclopentanes; Disease Models, Animal; Female; Guanidines; Influenza A Virus, H1N1 Subtype; Injections, Intramuscular; Mice; Mice, Inbred BALB C; Orthomyxoviridae Infections; Rodent Diseases; Survival Analysis

Oseltamivir and peramivir are being considered for combination treatment of serious influenza virus infections in humans. Both compounds are influenza virus neuraminidase inhibitors, and since peramivir binds tighter to the enzyme than oseltamivir carboxylate (the active form of oseltamivir), the possibility exists that antagonistic interactions might result when using the two compounds together. To study this possibility, combination chemotherapy experiments were conducted in vitro and in mice infected with influenza A/NWS/33 (H1N1) virus. Treatment of infected MDCK cells was performed with combinations of oseltamivir carboxylate and peramivir at 0.32-100μM for 3 days, followed by virus yield determinations. Additive drug interactions with a narrow region of synergy were found using the MacSynergy method. In a viral neuraminidase assay with combinations of inhibitors at 0.01-10nM, no significant antagonistic or synergistic interactions were observed across the range of concentrations. Infected mice were treated twice daily for 5 days starting 2h prior to virus challenge using drug doses of 0.05-0.4mg/kg/day. Consistent and statistically significant increases in the numbers of survivors were seen when twice daily oral oseltamivir (0.4mg/kg/day) was combined with twice daily intramuscular peramivir (0.1 and 0.2mg/kg/day) compared to single drug treatments. The data demonstrate that combinations of oseltamivir and peramivir perform better than suboptimal doses of each compound alone to treat influenza infections in mice. Treatment with these two compounds should be considered as an option.

Topics: Acids, Carbocyclic; Animals; Antiviral Agents; Cell Line; Cyclopentanes; Disease Models, Animal; Drug Evaluation, Preclinical; Drug Interactions; Drug Therapy, Combination; Guanidines; Humans; Influenza A Virus, H1N1 Subtype; Mice; Mice, Inbred BALB C; Neuraminidase; Orthomyxoviridae Infections; Oseltamivir

Efficacy of combination of the intramuscularly administered neuraminidase (NA) inhibitor, peramivir, and the orally administered M2 ion channel blocker, rimantadine was evaluated in mouse influenza A/Victoria/3/75 (H3N2) model. Mice were challenged with a sub-lethal virus dose (0-40% mortality in placebo group) and changes in body weights were analyzed by three-dimensional effect analysis to assess mode of drug interactions. Compounds were administered in a 5-day treatment course starting 1h before viral inoculation. The peramivir and rimantadine doses ranged from 0.3-3 mg/kg/d and 5-30 mg/kg/d, respectively. The maximum mean weight loss of 5.19 g was observed in the vehicle-infected group on day 10. In the 1 and 3 mg/kg/d peramivir monotherapy groups, the weight losses were 4.3 and 3.55 g, respectively. In the rimantadine monotherapy group, the weight losses were 3.43, 2.1, and 1.64 g for the 5, 10, and 30 mg/kg/d groups, respectively. Combination of 1mg/kg/d peramivir with 5 and 10 mg/kg/d rimantadine produced weight losses of 1.69 and 0.69 (p<0.05 vs. vehicle and individual agent), respectively, whereas the combination of 3.0 mg/kg/d peramivir with 10 and 30 mg/kg/d rimantadine did not show any weight loss (p<0.05 vs. vehicle and individual agent). The three-dimensional analysis of the weight loss for the majority of the drug combinations of peramivir and rimantadine tested demonstrated synergistic antiviral effects.

Topics: Acids, Carbocyclic; Animals; Antiviral Agents; Cyclopentanes; Disease Models, Animal; Dose-Response Relationship, Drug; Drug Administration Routes; Drug Administration Schedule; Drug Synergism; Female; Guanidines; Influenza A Virus, H3N2 Subtype; Mice; Mice, Inbred BALB C; Orthomyxoviridae Infections; Rimantadine; Survival Rate; Weight Loss

In the event of an influenza outbreak, antivirals including the neuraminidase (NA) inhibitors, peramivir, oseltamivir, and zanamivir may provide valuable benefit when vaccine production is delayed, limited, or cannot be used. Here we demonstrate the efficacy of a single intramuscular injection of peramivir in the mouse influenza model. Peramivir potently inhibits the neuraminidase enzyme N9 from H1N9 virus in vitro with a 50% inhibitory concentration (IC(50)) of 1.3+/-0.4 nM. On-site dissociation studies indicate that peramivir remains tightly bound to N9 NA (t(1/2)>24h), whereas, zanamivir and oseltamivir carboxylate dissociated rapidly from the enzyme (t(1/2)=1.25 h). A single intramuscular injection of peramivir (10mg/kg) significantly reduces weight loss and mortality in mice infected with influenza A/H1N1, while oseltamivir demonstrates no efficacy by the same treatment regimen. This may be due to tight binding of peramivir to the N1 NA enzymes similar to that observed for N9 enzyme. Additional efficacy studies indicate that a single injection of peramivir (2-20mg/kg) was comparable to a q.d.x 5 day course of orally administered oseltamivir (2-20mg/kg/day) in preventing lethality in H3N2 and H1N1 influenza models. A single intramuscular injection of peramivir may successfully treat influenza infections and provide an alternate option to oseltamivir during an influenza outbreak.

Topics: Acids, Carbocyclic; Animals; Antiviral Agents; Cyclopentanes; Disease Models, Animal; Drug Evaluation, Preclinical; Female; Guanidines; Influenza A Virus, H1N1 Subtype; Injections, Intramuscular; Mice; Neuraminidase; Orthomyxoviridae Infections

An inbred murine model (BALB/c) was utilized to assess the protective effect of the immunomodulator methyl inosine 5'-monophosphate (MIMP) against infection with influenza A/PR/8/34 (H1N1) virus. Contrary to the data reported for outbred mice (NMRI) infected with the aerosolized virus (Masihi, Hadden, 2003. J. Int. Immunopharmacol. 3, 1205-1215), there were no improvements in the outcomes of infection in the inbred animals treated with MIMP intranasally 1 day before the challenge and/or orally after the challenge for 5 days (up to 10 mg/kg/day). Nevertheless, complete protection against lethality was afforded by the treatment with the neuraminidase inhibitor peramivir given once daily for 5 days after the challenge (10 mg/kg/day). We speculate that the rapid progression of the disease in inbred mice caused by the intranasal challenge may render the MIMP-treatment ineffective. Our results emphasize the need for careful consideration of murine strains and routes of virus challenge in the design of experiments utilizing lethal influenza virus infection.

Topics: Acids, Carbocyclic; Age Factors; Animals; Antiviral Agents; Body Weight; Cyclopentanes; Disease Models, Animal; Female; Guanidines; Influenza A Virus, H1N1 Subtype; Inosine Monophosphate; Mice; Mice, Inbred BALB C; Orthomyxoviridae Infections

The cyclopentane influenza virus neuraminidase inhibitor RWJ-270201 was evaluated against influenza A/NWS/33 (H1N1), A/Shangdong/09/93 (H3N2), A/Victoria/3/75 (H3N2), and B/Hong Kong/05/72 virus infections in mice. Treatment was by oral gavage twice daily for 5 days beginning 4 h pre-virus exposure. The influenza virus inhibitor oseltamivir was run in parallel, and ribavirin was included in studies with the A/Shangdong and B/Hong Kong viruses. RWJ-270201 was inhibitory to all infections using doses as low as 1 mg/kg/day. Oseltamivir was generally up to 10-fold less effective than RWJ-270201. Ribavirin was also inhibitory but was less tolerated by the mice at the 75-mg/kg/day dose used. Disease-inhibitory effects included prevention of death, lessening of decline of arterial oxygen saturation, inhibition of lung consolidation, and reduction in lung virus titers. RWJ-270201 and oseltamivir, at doses of 10 and 1 mg/kg/day each, were compared with regard to their effects on daily lung parameters in influenza A/Shangdong/09/93 virus-infected mice. Maximum virus titer inhibition was seen on day 1, with RWJ-270201 exhibiting the greater inhibitory effect, a titer reduction of >10(4) cell culture 50% infective doses (CCID(50))/g. By day 8, the lung virus titers in mice treated with RWJ-270201 had declined to 10(1.2) CCID(50)/g, whereas titers from oseltamivir-treated animals were >10(3) CCID(50)/g. Mean lung consolidation was also higher in the oseltamivir-treated animals on day 8. Both neuraminidase inhibitors were well tolerated by the mice. RWJ-270201 was nontoxic at doses as high as 1,000 mg/kg/day. These data indicate potential for the oral use of RWJ-270201 in the treatment of influenza virus infections in humans.

Topics: Acetamides; Acids, Carbocyclic; Animals; Antiviral Agents; Cyclopentanes; Disease Models, Animal; Female; Guanidines; Influenza A virus; Influenza B virus; Lung; Mice; Mice, Inbred C57BL; Neuraminidase; Orthomyxoviridae Infections; Oseltamivir; Respiratory Function Tests; Ribavirin; Virus Replication

We examined RWJ-270201 in a lethal model of influenza in BALB/c mice. The aim was to delineate the pharmacodynamically linked variable for the drug. Challenge was performed with influenza virus A/Shongdong/09/93 (H3N2). Treatment was administered by gavage. Five doses (1 to 10 mg/kg of body weight) and three schedules (every 24, 12, and 8 h) were evaluated with 10 mice per group. There were 39 placebo-treated mice. Drug exposure was evaluated for infected mice. Exposures were calculated after population modeling of all the plasma concentration-time data simultaneously using the NPEM3 program. Evaluation of dose and schedule with Kaplan-Meier analysis and Cox proportional hazards modeling demonstrated that schedule offered no explanatory power relative to dose alone. Evaluation of peak concentration, trough concentration, and area under the concentration-time curve (AUC) by the same methods revealed that AUC was the dynamically linked variable. Again, schedule offered no further explanatory power when included in the model with AUC. This indicates that AUC is the linked variable and that the anti-influenza effect of RWJ-270201 is independent of schedule. We conclude that once-daily dosing of RWJ-270201 should be evaluated in clinical trials of influenza therapy.

Topics: Acids, Carbocyclic; Animals; Antiviral Agents; Cyclopentanes; Disease Models, Animal; Female; Guanidines; Mice; Mice, Inbred BALB C; Neuraminidase; Orthomyxoviridae Infections; Treatment Outcome

The orally administered neuraminidase (NA) inhibitor RWJ-270201 was tested in parallel with zanamivir and oseltamivir against a panel of avian influenza viruses for inhibition of NA activity and replication in tissue culture. The agents were then tested for protection of mice against lethal H5N1 and H9N2 virus infection. In vitro, RWJ-270201 was highly effective against all nine NA subtypes. NA inhibition by RWJ-270201 (50% inhibitory concentration, 0.9 to 4.3 nM) was superior to that by zanamivir and oseltamivir carboxylate. RWJ-270201 inhibited the replication of avian influenza viruses of both Eurasian and American lineages in MDCK cells (50% effective concentration, 0.5 to 11.8 microM). Mice given 10 mg of RWJ-270201 per kg of body weight per day were completely protected against lethal challenge with influenza A/Hong Kong/156/97 (H5N1) and A/quail/Hong Kong/G1/97 (H9N2) viruses. Both RWJ-270201 and oseltamivir significantly reduced virus titers in mouse lungs at daily dosages of 1.0 and 10 mg/kg and prevented the spread of virus to the brain. When treatment began 48 h after exposure to H5N1 virus, 10 mg of RWJ-270201/kg/day protected 50% of mice from death. These results suggest that RWJ-270201 is at least as effective as either zanamivir or oseltamivir against avian influenza viruses and may be of potential clinical use for treatment of emerging influenza viruses that may be transmitted from birds to humans.

Topics: Acetamides; Acids, Carbocyclic; Animals; Antiviral Agents; Body Weight; Brain; Cyclopentanes; Disease Models, Animal; Dogs; Female; Guanidines; Influenza A virus; Influenza A Virus, H5N1 Subtype; Influenza A Virus, H9N2 Subtype; Lung; Mice; Mice, Inbred BALB C; Neuraminidase; Orthomyxoviridae Infections; Oseltamivir; Pyrans; Sialic Acids; Treatment Outcome; Virus Replication; Zanamivir