Kimiyasu Shiraki

Favipiravir (T-705), a novel viral RNA polymerase inhibitor.

Favipiravir (T-705; 6-fluoro-3-hydroxy-2-pyrazinecarboxamide) is an antiviral drug that selectively inhibits the RNA-dependent RNA polymerase of influenza virus. It is phosphoribosylated by cellular enzymes to its active form, favipiravir-ribofuranosyl-5-triphosphate (RTP). Its antiviral effect is attenuated by the addition of purine nucleic acids, indicating the viral RNA polymerase mistakenly recognizes favipiravir-RTP as a purine nucleotide. Favipiravir is active against a broad range of influenza viruses, including A(H1N1)pdm09, A(H5N1) and the recently emerged A(H7N9) avian virus. It also inhibits influenza strains resistant to current antiviral drugs, and shows a synergistic effect in combination with oseltamivir, thereby expanding influenza treatment options. A Phase III clinical evaluation of favipiravir for influenza therapy has been completed in Japan and two Phase II studies have been completed in the United States. In addition to its anti-influenza activity, favipiravir blocks the replication of many other RNA viruses, including arenaviruses (Junin, Machupo and Pichinde); phleboviruses (Rift Valley fever, sandfly fever and Punta Toro); hantaviruses (Maporal, Dobrava, and Prospect Hill); flaviviruses (yellow fever and West Nile); enteroviruses (polio- and rhinoviruses); an alphavirus, Western equine encephalitis virus; a paramyxovirus, respiratory syncytial virus; and noroviruses. With its unique mechanism of action and broad range of antiviral activity, favipiravir is a promising drug candidate for influenza and many other RNA viral diseases for which there are no approved therapies.

Synergistic activity of amenamevir (ASP2151) with nucleoside analogs against herpes simplex virus types 1 and 2 and varicella-zoster virus.

ASP2151 (amenamevir) is a helicase-primase complex inhibitor with antiviral activity against herpes simplex virus HSV-1, HSV-2, and varicella-zoster virus (VZV). To assess combination therapy of ASP2151 with existing antiherpes agents against HSV-1, HSV-2, and VZV, we conducted in vitro and in vivo studies of two-drug combinations. The combination activity effect of ASP2151 with nucleoside analogs acyclovir (ACV), penciclovir (PCV), or vidarabine (VDB) was tested via plaque-reduction assay and MTS assay, and the data were analyzed using isobolograms and response surface modeling. In vivo combination therapy of ASP2151 with valaciclovir (VACV) was studied in an HSV-1-infected zosteriform spread mouse model. The antiviral activity of ASP2151 combined with ACV and PCV against ACV-susceptible HSV-1, HSV-2, and VZV showed a statistically significant synergistic effect (P<0.05). ASP2151 with VDB was observed to have additive effects against ACV-susceptible HSV-2 and synergistic effects against VZV. In the mouse model of zosteriform spread, the inhibition of disease progression via combination therapy was more potent than that of either drugs as monotherapy (P<0.05). These results indicate that the combination therapies of ASP2151 with ACV and PCV have synergistic antiherpes effects against HSV and VZV infections and may be feasible in case of severe disease, such as herpes encephalitis or in patients with immunosuppression.

Herpes Zoster and Recurrent Herpes Zoster

Abstract Background The incidence of recurrent herpes zoster (HZ) and the relationship between initial and recurrent HZ are not clear. Methods The Miyazaki Dermatologist Society has surveyed ~5000 patients with HZ annually since 1997. A questionnaire regarding HZ and its recurrence was completed by the dermatologists. Results A total of 34 877 patients with HZ were registered at 43 clinics between June 2009 and November 2015. Among 16 784 patients seen at 10 of the 43 clinics, 1076 patients (6.41%) experienced recurrence. Herpes zoster was more frequent in female than in male patients (5.27 vs 4.25 in 1000 person-years, P < .001), as was HZ recurrence (7.63% vs 4.73%, P < .001). Two and three recurrences were observed in 49 and 3 patients, respectively. Recurrence in the same dermatome was observed in 16.3% of patients, and more frequently this occurred in the left side (P = .027). The number of HZ-experienced persons increased with age, and one third of the population had experienced HZ by the age of 80. Conclusions Recurrent HZ was observed in 6.41% of patients, with a higher incidence in women. Moreover, HZ experience reduced the HZ incidence to 31.7% of the incidence in the HZ-naive population.

Subclinical generation of acyclovir-resistant herpes simplex virus with mutation of homopolymeric guanosine strings during acyclovir therapy.

BACKGROUNDnSuppressive therapy in patients with genital herpes has been used in Japan since 2006. Susceptibility and resistance of herpes simplex virus (HSV)-2 to acyclovir were examined in genital isolates from patients receiving suppressive therapy and compared with those from those naïve to acyclovir and receiving episodic treatment with acyclovir.nnnOBJECTIVEnThe aim of this study was to analyze the effect of acyclovir use on the susceptibility to acyclovir and analysis of the thymidine kinase gene by acyclovir treatment.nnnMETHODSnGenital HSV isolates were obtained from three patients groups. Susceptibility to acyclovir, the frequency of acyclovir-resistant clones and mutations in the thymidine kinase gene of acyclovir-resistant clones were determined.nnnRESULTSnSusceptibility to ACV was significantly higher in isolates from patients receiving suppressive therapy than those naïve to acyclovir and receiving episodic treatment, but the frequencies of resistant clones were similar among the three groups. Mutation in guanosine homopolymeric strings (G-string mutation) was significantly more frequent in clones during episodic treatment and suppressive therapy than clones from patients naïve to ACV. The frequency of G-string mutation was significantly less frequent in isolates from patients naïve to ACV than those experienced ACV therapy.nnnCONCLUSIONnThe frequency of acyclovir-resistant mutants was not increased by episodic and suppressive therapy, but exposure to acyclovir significantly generated G-string mutations, possibly induced by acyclovir. Acyclovir therapy had no substantial effects on the susceptibility of HSV-2 or frequency of resistant virus but did generate subclinical G-string mutants in patients HSV-2.

Novel deletion in glycoprotein G forms a cluster and causes epidemiologic spread of herpes simplex virus type 2 infection

The herpes simplex virus type 2 (HSV‐2) glycoprotein G (gG‐2) gene of 106 clinical isolates was analyzed and six isolates were identified with 63 nucleotides comprising 21 amino acids (aa) deleted in the immunodominant region. Compared with strain HG52, variations in the gG‐2 gene were found at 276 and 27 sites in nucleotide and aa sequences, respectively, in the 106 strains. Significant variations in both nucleotides and aa were accumulated in the immunodominant region rather than in the other regions (Pu2009<u20090.001), indicating that the immunodominant region might be indispensable in vivo and a hot spot for variation. The frequency of 21 aa‐deleted strains (HSVΔ21/gG‐2) among clinical isolates was 5%, indicating the advantage of this deletion of gG‐2 for epidemiological expansion. Phylogenetic analysis of the 106 strains indicated that the HSVΔ21/gG‐2 strains formed a cluster among the various variations but that their genomes showed different endonuclease digestion patterns. The antibody titers to total HSV antigens of patients infected with wild HSV‐2 and HSVΔ21/gG‐2 were similar, but patients with HSVΔ21/gG‐2 had a lower antibody titer to gG‐2 than those with wild HSV‐2 (Pu2009<u20090.001). HSVΔ21/gG‐2 might be less immnunogenic and reduce antibody production to gG‐2, while its pathogenicity in humans was not distinguished in its clinical manifestations. Thus, infection with HSVΔ21/gG‐2 caused genital lesions similar to wild HSV‐2 infection, but evaded the immune response to gG‐2 to allow epidemiological spread, indicating the importance of this deletion in the immunodominant region of gG‐2 in the pathogenesis and transmission of genital herpes. J Med. Virol. 85:1818–1828, 2013.

Rapid detection of human cytomegalovirus UL97 and UL54 mutations for antiviral resistance in clinical specimens.

Drug‐resistant cytomegalovirus appears during prolonged anti‐cytomegalovirus therapy. Assays for human cytomegalovirus viral protein kinase (UL97) and viral DNA polymerase (UL54) gene mutations conferring drug resistance have been used rather than susceptibility assays to assess clinical specimens. In this study a sensitive system for genotype assay of UL97 and UL54 in clinical specimens with as few as six copies/µg of DNA was developed.

Profile of anti-herpetic action of ASP2151 (amenamevir) as a helicase-primase inhibitor

&NA; The antiherpetic drugs acyclovir (ACV, valaciclovir) and penciclovir (famciclovir) are phosphorylated by viral thymidine kinase and terminate DNA synthesis. ASP2151 (amenamevir) and foscavir (PFA) directly inhibit viral helicase‐primase and DNA polymerase, respectively, and inhibit replication of herpes simplex virus (HSV) and varicella‐zoster virus. ACV, ASP2151, and PFA all inhibit HSV with a different mechanism of action and as a consequence, the kinetics of viral DNA accumulation and progeny virus production differ. This study focused on how viral DNA synthesis and its related events in the replication cycle would influence anti‐HSV action of ACV, ASP2151, and PFA. ASP2151 suppressed HSV replication more efficiently than ACV at 10 × 50% effective concentration of plaque formation (EC50), when treatments were started 0–24 h after infection. ASP2151 and PFA were more potent than ACV in suppressing viral DNA synthesis and infectious virus production when they were added up to 3 h following infection. The virus replicated in the presence of ACV was compared for the ratios of HSV DNA copy number to infectivity with that without ACV and infectivity of ACV‐treated virus was less efficient than that without ACV‐treatment. The EC50 of infected cells in the time course after infection was preserved in PFA, limited in ASP2151, and much increased for ACV, indicating that viral DNA synthesis had little effect on antiviral action of PFA and ASP2151 but reduced the susceptibility of ACV. ASP2151 showed a preferable profile as an anti‐herpetic agent with a better pharmacokinetic profile than ACV. HighlightsASP2151 and PFA were much potent than ACV in suppressing viral DNA.ASP2151 showed a stronger anti‐HSV activity than acyclovir in the early phase of infection.Viral DNA synthesis reduced anti‐HSV activity of ACV.Viral DNA synthesis had little effect on anti‐HSV action of ASP2151.

Functional differences between antiviral activities of sulfonated and intact intravenous immunoglobulin preparations toward varicella-zoster virus and cytomegalovirus

Intravenous immunoglobulin (IVIG) is used to treat severe viral infection, especially varicella-zoster virus (VZV) and cytomegalovirus (CMV) infections. The neutralization antibody titers of eleven IVIG preparations from four companies were examined using VZV and CMV with and without complement. The neutralizing antibody titers of intact IgG preparations were three to six times higher after addition of complement. The effectiveness of the sulfonated IgG preparation was not enhanced by complement, but desulfonated IgG regained enhanced neutralization activity with complement. Antibody-dependent cellular cytotoxicity (ADCC) toward VZV-infected cells was observed with both intact and sulfonated IVIG and guinea pig splenocytes, but ADCC toward CMV-infected cells was not, although NK cell activity toward cells infected with VZV or CMV was detected by splenocytes. Sulfonated IVIG had no complement-activated neutralization of VZV and CMV but retained ADCC toward VZV with less activity after dilution than with intact IVIG. Because sulfonated IVIG is converted to the intact form after intravenous administration, it would show complement-enhanced neutralization and ADCC activity similar to that of intact IVIG inxa0vivo. In this study we showed the effects of intact and sulfonated IgG on the functional activity of IgG against VZV and CMV.

Detection of cytomegalovirus in an immunocompetent adult presenting with acute retinal necrosis due to varicella-zoster virus: a case report

Background To report a case of simultaneous detection of cytomegalovirus (CMV) in acute retinal necrosis (ARN) due to varicella-zoster virus (VZV) in an immunocompetent adult. Methods A 65-year-old healthy woman presented with necrotizing retinitis. Vitreous, tears, saliva, and swabs of the auricular and forehead skin areas were collected and tested by polymerase chain reaction analysis at various time points during the treatment period. Results VZV and CMV DNA were detected in the vitreous, tears, saliva, and skin. CMV was present in fewer copies than VZV in the vitreous. The retinal lesion clinically improved rapidly only by acyclovir. According to the clinical findings and the clinical responses to acyclovir, CMV seems not to cause ARN in this case. Conclusion VZV and CMV DNA were detected in the vitreous of a patient with VZV ARN. CMV reactivation appeared to be asymptomatic.

Characterization of susceptibility variants of poliovirus grown in the presence of favipiravir

BACKGROUNDnT-705 (favipiravir) is a potent inhibitor of RNA-dependent RNA polymerases of influenza viruses and no favipiravir-resistant virus has been isolated. Poliovirus RNA polymerase has been well characterized and isolation of resistant virus was examined in poliovirus.nnnMETHODSnSusceptibility variants of poliovirus I (Sabin strain) were isolated during passages in the presence of favipiravir and characterized for their susceptibility and the sequence of RNA polymerase.nnnRESULTSnFive variants with 0.47-1.88 times the 50% inhibitory concentration for plaque formation of the parent poliovirus had amino acid variations in the 3D gene of the RNA polymerase. The distribution of amino acid variations was not related to ribavirin resistance, and two amino acid variation sites were found near the finger domain.nnnCONCLUSIONnFavipiravir as a chain terminator would not be incorporated and replicate to cause lethal mutagenesis as a mutagen like ribavirin, and resistant mutants were not isolated. A high replication level would generate mutations leading to favipiravir resistance as ribavirin resistance was generated, but generated mutations would be lethal to the RNA polymerase function.