Efficacy of Favipiravir (T-705) in Rabies Postexposure Prophylaxis

Abstract

Rabies is a fatal encephalitis caused by rabies virus (RABV), and no antiviral drugs for RABV are currently available. We report for the first time the efficacy of favipiravir (T-705) against RABV in vitro and in vivo. T-705 produced a significant, 3-4 log10 reduction in the multiplication of street and fixed RABV strains in mouse neuroblastoma Neuro-2a cells, with half-maximal inhibitory concentrations of 32.4 µM and 44.3 µM, respectively. T-705 significantly improved morbidity and mortality among RABV-infected mice when orally administered at a dose of 300 mg/kg/day for 7 days, beginning 1 hour after inoculation. T-705 significantly reduced the rate of virus positivity in the brain. Furthermore, the effectiveness of T-705 was comparable to that of equine rabies virus immunoglobulin for postexposure prophylaxis. Collectively, our results suggest that T-705 is active against RABV and may serve as a potential alternative to rabies immunoglobulin in rabies postexposure prophylaxis.

Keywords: T-705; antiviral drug; postexposure prophylaxis; rabies.

Figure 1.

Antiviral activity of T-705 against rabies virus (RABV) in mouse neuroblastoma cell lines. A, Expression of HPRT in NA and N2a cells. HPRT and β-actin proteins were detected by Western blots. The relative molecular mass is indicated on the left. B, The 1088 or challenge virus standard (CVS) RABV strain was inoculated into NA or N2a cells at a multiplicity of infection (MOI) of 0.01. The cells were incubated for 96 hours in the presence (1000 µM) or absence (0 µM) of T-705. Each virus titer in the supernatant was determined using the focus assay. C, Expression of HPRT in NA cells transfected with the empty or hprt vector. The indicated proteins were detected by Western blots. D, Efficacy of T-705 against NA cells transfected with the hprt vector. The 1088 strain was inoculated into NA cells transfected with the empty or hprt vector or into N2a cells. The cells were incubated for 96 hours in the presence (1000 µM) or absence (0 µM) of T-705. Each virus titer in the supernatant was determined using the focus assay. Data represent the mean ± SD (n = 3). *P < .05, by the Student t test; **P < .05, by 2-way analysis of variance for interaction. Abbreviations: FFU, focus-forming units; NS, not significant.

Figure 2.

Determination of the inhibitory concentration of T-705 against rabies virus in N2a cells. N2a cells were inoculated with 1088 (A) or CVS (B) at a multiplicity of infection of 0.01 and incubated for 96 hours with the indicated concentration of T-705. Each virus titer in the supernatant was determined using the focus assay. Data represent the mean ±SD (n = 3). A sigmoidal dose response was fitted using GraphPad Prism. Each 50% inhibitory concentration (IC₅₀) with its 95% confidential interval (CI) is indicated. Abbreviation: CVS, challenge virus standard.

Figure 3.

Efficacy of postexposure T-705 administration for 7 days in mice infected with 1088. A, Mice were intramuscularly inoculated with 1088 and orally administered T-705 (30, 100, or 300 mg/kg/day) or 0.5% methylcellulose (as a control) daily for 7 days (days 0–6) beginning 1 hour after inoculation. Mice were monitored for 28 days. “Sick” indicates that mice showed significant body weight loss or neurological signs. Surviving sick mice had begun to gain body weight but with sequelae, such as limb paralysis. For the survival curves, a significant difference was observed between the control and T-705 (300 mg/kg/day) groups (P < .01, by the log-rank test) but not between the control and T-705 (30 or 100 mg/kg/day) groups (P ≥ .05, by the log-rank test). B, The VNA titers in sera of surviving mice. Sera were collected at 28 days after inoculation, and the titers were determined by a rapid fluorescent focus inhibition test.

Figure 4.

Virus titers in the brains of infected mice administered T-705. Mice were intramuscularly inoculated with 1088 and orally administered T-705 (300 mg/kg/day) or 0.5% methylcellulose daily for 7 days (days 0–6) beginning 1 hour after inoculation. Brain samples were collected at 8 and 11 days after inoculation, and the titers were determined. P values were determined using the Fisher exact test (treatment and virus-positive rate in the brain). *P < .05. Abbreviations: FFU, focus-forming units; NS not significant.

Figure 5.

Efficacy of postexposure T-705 administration for 14 days in mice infected with 1088. Mice were intramuscularly inoculated with 1088 and orally administered T-705 (300 mg/kg/day) or 0.5% methylcellulose (as a control) daily for 14 days, beginning 1 hour (days 0–13) or 4 days (days 4–17) after inoculation. Mice were monitored for 28 days. “Sick” indicates that mice showed significant body weight loss or neurological signs. Surviving sick mice had begun to gain body weight but with sequelae, such as limb paralysis. For the survival curves, a significant difference was observed between the T-705 (days 0–13) and control (days 0–13) or T-705 (days 4–17) groups (P < .01, by the log-rank test) but not between the control (days 0–13) and T-705 (days 4–17) groups (P = .96, by the log-rank test). B, The VNA titers in sera of surviving mice. Sera were collected at 28 days after inoculation, and the titers were determined by the rapid fluorescent focus inhibition test.

Figure 6.

Comparison of the efficacy of T-705 and equine rabies immunoglobulin (ERIG) as postexposure treatments. Mice were intramuscularly inoculated with 1088 and orally administered T-705 (300 mg/kg/day) daily for 7 days (days 0–6) beginning 1 hour (0 days), 1 day, or 2 days after inoculation, or were intramuscularly injected with 40 IU/kg of ERIG at the virus inoculation site at 1 hour (on 0 day), 1 day, or 2 days after inoculation. Inoculated mice without any treatments were used as controls. Mice were monitored for 28 days. “Sick” indicates that mice showed significant body weight loss or neurological signs. Surviving sick mice had begun to gain body weight but with sequelae, such as limb paralysis. For the survival curves, log-rank tests were performed, and the P values are indicated in the figure. A P value of <.0056 (after Bonferroni correction) was considered significant.