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The unexpected emergence of oseltamivir-resistant A(H1N1) viruses in 2008 was facilitated in part by the establishment of permissive secondary neuraminidase (NA) substitutions that compensated for the fitness loss due to the NA-H275Y resistance substitution. These viruses were replaced in 2009 by oseltamivir-susceptible A(H1N1)pdm09 influenza viruses. Genetic analysis and screening of A(H1N1)pdm09 viruses circulating in Germany between 2009 and 2024 were conducted to identify any potentially synergistic or resistance-associated NA substitutions. Selected viruses were then subjected to further characterization in vitro. In the NA gene of circulating A(H1N1)pdm09 viruses, two secondary substitutions, NA-V241I and NA-N369K, were identified. These substitutions demonstrated a stable lineage in phylogenetic analysis since the 2010–2011 influenza season. The data indicate a slight increase in viral NA bearing two additional potentially synergistic substitutions, NA-I223V and NA-S247N, in the 2023–2024 season, which both result in a slight reduction in susceptibility to NA inhibitors. The accumulation of secondary synergistic substitutions in the NA of A(H1N1)pdm09 viruses increases the probability of the emergence of antiviral-resistant viruses. Therefore, it is crucial to closely monitor the evolution of circulating influenza viruses and to develop additional antiviral drugs against different target proteins. [ABSTRACT FROM AUTHOR]

Abstract Background Oseltamivir (OSM) was the first active oral therapeutic inhibitor approved by the Food and Drug Administration in 1999 for the clinical management of the influenza virus. It is an ester-type prodrug of OSM carboxylate in the market under the trade name Tamiflu™ capsules, i.e., oseltamivir phosphate. Because of the ubiquitous application to alleviate influenza virus (flu virus) symptoms, it is imperative to develop systematic analytical protocols for quality control laboratories, bioequivalence, and pharmacokinetic analysis. Main body of the abstract This review provides complete state-of-the-art analytical protocols for quantifying OSM, as published in scientific journals and official compendia. Several studies use LC–MS/MS and HPLC/UV. Additionally, there are reports on UPLC, HPTLC, capillary electrophoresis, FTIR, voltammetry, potentiometry, spectrophotometric, and spectrofluorometry protocols for the drug. Many analytical protocols have also been documented to analyze OSM from environmental water, surface water, sewage discharge, the Neya River and treated sewage effluent and surface water. Conclusion The present review concludes with significant remarks on the methodology used to analyze OSM. Despite the therapeutic applicability of the drug, there are a limited number of comprehensive documents on analytical protocols for determining its concentration in various matrices. This lack of information is elusive, as the applicability and effectiveness of these protocols are crucial for ensuring the quality, efficacy, and safety of OSM.

To assess the extent of susceptibility to the four neuraminidase inhibitors (NAIs) approved in Japan of the epidemic viruses in the 2022–23 influenza season in Japan, we measured the 50 % inhibitory concentration (IC 50) of oseltamivir, zanamivir, peramivir, and laninamivir in influenza virus isolates from patients. Viral isolation was done with specimens obtained prior to and after treatment, and the type/subtype was determined by RT-PCR using type- and subtype-specific primers. The IC 50 was determined by a neuraminidase inhibition assay using a fluorescent substrate. Virus isolates, one A(H1N1)pdm09 and 74 A(H3N2), were measured in the 2022–23 season. The geometric mean IC 50 s of the 74 A(H3N2) isolated prior to treatment were 0.78 nM, 0.66 nM, 2.08 nM, and 2.85 nM for oseltamivir, peramivir, zanamivir, and laninamivir, respectively, comparable to those of the previous ten studied seasons. No A(H3N2) with highly reduced sensitivity to any of the NAIs was found in the 2022-23 season prior to or after drug administration. These results indicate that the sensitivity to these four commonly used NAIs has been maintained, at least for A(H3N2), in the 2022–23 influenza season in Japan, after the 2020–21 and 2021–22 seasons when the prevalence of influenza was extremely low. [ABSTRACT FROM AUTHOR]

The unexpected emergence of oseltamivir-resistant A(H1N1) viruses in 2008 was facilitated in part by the establishment of permissive secondary neuraminidase (NA) substitutions that compensated for the fitness loss due to the NA-H275Y resistance substitution. These viruses were replaced in 2009 by oseltamivir-susceptible A(H1N1)pdm09 influenza viruses. Genetic analysis and screening of A(H1N1)pdm09 viruses circulating in Germany between 2009 and 2024 were conducted to identify any potentially synergistic or resistance-associated NA substitutions. Selected viruses were then subjected to further characterization in vitro. In the NA gene of circulating A(H1N1)pdm09 viruses, two secondary substitutions, NA-V241I and NA-N369K, were identified. These substitutions demonstrated a stable lineage in phylogenetic analysis since the 2010–2011 influenza season. The data indicate a slight increase in viral NA bearing two additional potentially synergistic substitutions, NA-I223V and NA-S247N, in the 2023–2024 season, which both result in a slight reduction in susceptibility to NA inhibitors. The accumulation of secondary synergistic substitutions in the NA of A(H1N1)pdm09 viruses increases the probability of the emergence of antiviral-resistant viruses. Therefore, it is crucial to closely monitor the evolution of circulating influenza viruses and to develop additional antiviral drugs against different target proteins.

Influenza can cause respiratory infections, leading to significant morbidity and mortality in humans. While current influenza vaccines offer varying levels of protection, there remains a pressing need for effective antiviral drugs to supplement vaccine efforts. Currently, the FDA-approved antiviral drugs for influenza include oseltamivir, zanamivir, peramivir, and baloxavir marboxil. These antivirals primarily target the virus, making them vulnerable to drug resistance. In this study, we evaluated the efficacy of the neuraminidase inhibitor, oseltamivir, against probenecid, which targets the host cells and is less likely to engender resistance. Our results show that probenecid has superior antiviral efficacy compared to oseltamivir in both in vitro replication assays and in vivo mouse models of influenza infection. [ABSTRACT FROM AUTHOR]

Abstract Background This study assessed the differences in daily virus reduction and the residual infectivity after the recommended home stay period in Japan in patients infected with influenza and treated with baloxavir (BA), laninamivir (LA), oseltamivir (OS), and zanamivir (ZA). Methods We conducted an observational study on children and adults at 13 outpatient clinics in 11 prefectures in Japan during seven influenza seasons from 2013/2014 to 2019/2020. Virus samples were collected twice from influenza rapid test-positive patients at the first and second visit 4–5 days after the start of treatment. The viral RNA shedding was quantified using quantitative RT-PCR. Neuraminidase (NA) and polymerase acidic (PA) variant viruses that reduce susceptibility to NA inhibitors and BA, respectively, were screened using RT-PCR and genetic sequencing. Daily estimated viral reduction was evaluated using univariate and multivariate analyses for the factors such as age, treatment, vaccination status, or the emergence of PA or NA variants. The potential infectivity of the viral RNA shedding at the second visit samples was determined using the Receiver Operator Curve based on the positivity of virus isolation. Results Among 518 patients, 465 (80.0%) and 116 (20.0%) were infected with influenza A (189 with BA, 58 with LA, 181 with OS, 37 with ZA) and influenza B (39 with BA, 10 with LA, 52 with OS, 15 with ZA). The emergence of 21 PA variants in influenza A was detected after BA treatment, but NA variants were not detected after NAIs treatment. Multiple linear regression analysis showed that the daily viral RNA shedding reduction in patients was slower in the two NAIs (OS and LA) than in BA, influenza B infection, aged 0–5 years, or the emergence of PA variants. The residual viral RNA shedding potentially infectious was detected in approximately 10–30% of the patients aged 6–18 years after five days of onset. Conclusions Viral clearance differed by age, type of influenza, choice of treatment, and susceptibility to BA. Additionally, the recommended homestay period in Japan seemed insufficient, but reduced viral spread to some extent since most school-age patients became non-infectious after 5 days of onset.

Background: This study assessed the differences in daily virus reduction and the residual infectivity after the recommended home stay period in Japan in patients infected with influenza and treated with baloxavir (BA), laninamivir (LA), oseltamivir (OS), and zanamivir (ZA). Methods: We conducted an observational study on children and adults at 13 outpatient clinics in 11 prefectures in Japan during seven influenza seasons from 2013/2014 to 2019/2020. Virus samples were collected twice from influenza rapid test-positive patients at the first and second visit 4–5 days after the start of treatment. The viral RNA shedding was quantified using quantitative RT-PCR. Neuraminidase (NA) and polymerase acidic (PA) variant viruses that reduce susceptibility to NA inhibitors and BA, respectively, were screened using RT-PCR and genetic sequencing. Daily estimated viral reduction was evaluated using univariate and multivariate analyses for the factors such as age, treatment, vaccination status, or the emergence of PA or NA variants. The potential infectivity of the viral RNA shedding at the second visit samples was determined using the Receiver Operator Curve based on the positivity of virus isolation. Results: Among 518 patients, 465 (80.0%) and 116 (20.0%) were infected with influenza A (189 with BA, 58 with LA, 181 with OS, 37 with ZA) and influenza B (39 with BA, 10 with LA, 52 with OS, 15 with ZA). The emergence of 21 PA variants in influenza A was detected after BA treatment, but NA variants were not detected after NAIs treatment. Multiple linear regression analysis showed that the daily viral RNA shedding reduction in patients was slower in the two NAIs (OS and LA) than in BA, influenza B infection, aged 0–5 years, or the emergence of PA variants. The residual viral RNA shedding potentially infectious was detected in approximately 10–30% of the patients aged 6–18 years after five days of onset. Conclusions: Viral clearance differed by age, type of influenza, choice of treatment, and susceptibility to BA. Additionally, the recommended homestay period in Japan seemed insufficient, but reduced viral spread to some extent since most school-age patients became non-infectious after 5 days of onset. [ABSTRACT FROM AUTHOR]

Influenza can cause respiratory infections, leading to significant morbidity and mortality in humans. While current influenza vaccines offer varying levels of protection, there remains a pressing need for effective antiviral drugs to supplement vaccine efforts. Currently, the FDA-approved antiviral drugs for influenza include oseltamivir, zanamivir, peramivir, and baloxavir marboxil. These antivirals primarily target the virus, making them vulnerable to drug resistance. In this study, we evaluated the efficacy of the neuraminidase inhibitor, oseltamivir, against probenecid, which targets the host cells and is less likely to engender resistance. Our results show that probenecid has superior antiviral efficacy compared to oseltamivir in both in vitro replication assays and in vivo mouse models of influenza infection.

• Transplant patients experience prolonged respiratory virus infections. • Monotherapy with neuraminidase inhibitors failed to cure influenza virus infection. • Neuraminidase inhibitor monotherapy led to the emergence of drug-resistant strains. • The combination of zanamivir and baloxavir marboxil suppressed persistent Influenza infection. • The clearance of Influenza and HCoV-OC43 resulted in full respiratory recovery. Immunocompromised patients may experience prolonged shedding of influenza virus potentially leading to severe infections. Alternatives to monotherapy with neuraminidase inhibitors should be evaluated to entirely suppress viral replication and prevent drug-resistant mutations. We investigated the clinical and virological evolution in a case of persistent influenza A and human coronavirus OC43 (HCoV-OC43) coinfection in a hematopoietic stem cell transplant recipient after different therapeutic strategies. Successive oseltamivir and zanamivir monotherapies failed to control both infections, with positive results persisting for over 110 days each. This led to the emergence of highly resistant oseltamivir strains due to neuraminidase mutations (E119V and R292K) followed by a deletion (del245-248), while maintaining sensitivity to zanamivir. The intra-host viral diversity data showed that the treatments impacted viral diversity of influenza virus, but not of HCoV-OC43. Considering the patient's underlying condition and the impact of prolonged viral shedding on pulmonary function, eradicating the influenza virus was necessary. A 10-day regimen combining zanamivir and baloxavir-marboxil effectively controlled influenza virus replication and was associated with the clearance of HCoV-OC43, finally resulting in comprehensive respiratory recovery. These observations underscore the importance of further investigating combination treatments as the primary approach to achieve influenza eradication in immunocompromised patients. [ABSTRACT FROM AUTHOR]

This study investigated whether quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR), using specific probes composed of locked nucleic acids (LNA/qRT-PCR), designed to evaluate H1N1 pdm09 H275Y, H3N2 E119V and R292K variant populations, could replace a neuraminidase (NA) inhibition assay to determine the 50% inhibitory concentration (IC50) of NA activity. For H1N1 pdm09, when the H275Y variant RNA load was 50% or 70% and the infective H275Y variant load was 40% or 70%, the IC50 were >10- and 100-fold higher, respectively, than that of the wild-type (WT) strain. For H3N2, when the E119V RNA load and infective E119V variant load were >90% and >60%, respectively, the IC50 of the mixed sample was >10-fold higher than that of the WT strain. The variant-mixed samples with a 70% or 80% R292K variant RNA load and a 60% or 70% infective R292K variant load exhibited >10- and 100-fold decreased susceptibility, respectively, compared with that of the WT. A positive correlation between the variant RNA load and infective variant load populations was observed. The LNA/qRT-PCR method can be used to improve the treatment and management of patients during antiviral therapy for influenza virus infection. [ABSTRACT FROM AUTHOR]

Large scale investigation of the clinical effectiveness of neuraminidase inhibitors for circulating influenza viruses are important along with the surveillance of virus susceptibility in vitro. The duration of fever and other influenza symptoms as markers of the clinical effectiveness of laninamivir octanoate hydrate (laninamivir) were investigated in the Japanese 2017/18 and 2018/19 influenza seasons and compared with the results of the previous six seasons. Influenza A(H1N1)pdm09, A(H3N2), and B were found in 14, 45, and 52 patients in the 2017/18 season and in 22, 62, and 0 in the 2018/19 season, respectively. The median duration of fever for B was significantly longer than for A(H1N1)pdm09 and A(H3N2) in the 2017/18 season (p = 0.0461) and for A(H3N2) than for A(H1N1)pdm09 in the 2018/19 season (p = 0.0290). However, the differences were subtle in both seasons for other symptoms, with no significant differences in their median duration in comparison of the circulating types/subtypes. Over the eight seasons with the previous six seasons added, the median durations of fever were consistently longer for B than for A, but the relation between the A subtypes was inconsistent. The median durations of fever were comparable over the eight seasons for the virus types/subtypes, as were the median durations of other symptoms. The percentage of febrile patients decreased in a similar pattern over the eight seasons for each type/subtype. The results confirmed that laninamivir has continued to be clinically effective against all types/subtypes of influenza viruses, with no safety issues. [ABSTRACT FROM AUTHOR]

Background: Influenza is difficult to distinguish clinically from other acute respiratory infections. Rapid laboratory diagnosis can help initiate early effective antiviral treatment and isolation. Implementing a novel point-of-care test (POCT) for influenza in the emergency department (ED) could improve treatment and isolation strategies and reduce the length of stay (LOS).Methods: In a prospective, controlled observational cohort study, we enrolled patients admitted due to acute respiratory illness to 2 public hospitals in Bergen, Norway, one using a rapid POCT for influenza (n = 400), the other (n = 167) using conventional rapid laboratory-based assay.Results: Prevalence of influenza was similar in the 2 hospitals (154/400, 38% vs 38%, 63/167; P = .863). Most patients in both hospitals received antiviral (83% vs 81%; P = .703) and antibiotic treatment (72% vs 62%; P = .149). Isolation was more often initiated in ED in the hospital using POCT (91% vs 80%; P = .025). Diagnosis by POCT was associated with shorter hospital stay; old age, diabetes, cancer, and use of antibiotics, particularly broad-spectrum antibiotics, were associated with prolonged stay.Conclusions: POCT implementation in ED resulted in improved targeted isolation and shorter LOS. Regardless of POCT use, most influenza patients received antivirals (>80%) and antibiotics (>69%). [ABSTRACT FROM AUTHOR]

Introduction and objective For centuries influenza has been one of the most prevalent viral infectious diseases and still remains an important cause of morbidity and mortality in many regions of the world, including Poland. The aim of the study was presentation of the scope of problems concerning influenza virus infections and the available options for controlling the disease. Review methods The publication was prepared based on a literature review in scientific information databases and on the websites of organizations operating in the field of public health. As part of the issue analysis, a systematic search was performed of current scientific data concerning the described problem. Brief description of the state of knowledge Four types of influenza virus are distinguished: A, B, C and D. Infections in humans are most often caused by influenza A and B. The best recognized virus is influenza A, which is characterized by high antigenic and genetic variability and zoonotic potential. Influenza A virus is responsible for local epidemics and pandemics and meets all the criteria of a pathogen that can cause a catastrophic biological threat on a global scale. Modern medicine has the ability to control influenza through protective vaccinations with multivalent vaccines and antiviral drugs, which include neuraminidase inhibitors and viral polymerase inhibitors. Summary Influenza is a major public health risk problem. The actual risk of a global epidemic in the future forces to undertake transdisciplinary and integrated actions striving at an effective immunoprophylaxis of influenza virus infections, as well as treatment of the disease and its complications. A global epidemic may go beyond the capabilities of individual countries. The development of a vaccine against pandemic influenza and its mass production in a short time has the utmost priority.

Four cases of oseltamivir-resistant influenza A(H1N1)pdm09 virus infection were detected among inhabitants of a border detention center in Texas, USA. Hemagglutinin of these viruses belongs to 6B.1A5A-156K subclade, which may enable viral escape from preexisting immunity. Our finding highlights the necessity to monitor both drug resistance and antigenic drift of circulating viruses.

Neuraminidase (NA) has been well-studied as a therapeutic target for Influenza. However, resistance to the influenza virus has been observed recently. Out of special interest in the utilization of dietary antivirals from citrus, in vitro inhibition activity against NA and in silico studies including molecular docking, molecular dynamic simulation, and a predictive ADMET study, were performed on five citrus-derived flavanones. Encouragingly, citrus-derived flavanones displayed comparable or even more potent in vitro inhibitory activity than oseltamivir carboxylate against NA. Orange peel extract exhibited higher activity than hesperidin. Among the tested compounds, neohesperidin, forming strong hydrogen-bonding interactions with key arginine residues, exhibited the most effective inhibitory activity against NAs from C. perfringens , consistent with the results of molecular dynamics simulations. Although the molecular docking results were inconsistent with the in vitro activity, the binding energy was identical against the wild-type and mutant, suggesting a lower likelihood of developing drug resistance. Moreover, predictive ADMET studies showed favorable pharmacokinetic properties for the tested compounds. Overall, citrus fruit peel emerges as a promising dietary supplement for prevention and treatment of influenza. These findings elucidate the impact of flavanones on NA activity, and the analysis of their binding modes provides valuable insights into the mechanism of NA inhibition. [Display omitted] • Citrus-derived flavanones exhibited similar or greater activity than OSC in NA inhibitory activity. • Five flavanones may exhibit a lower likelihood of developing drug resistance by molecular docking. • This study provides a partial theoretical basis for the application of citrus-derived extracts on dietary therapy in influenza patients. [ABSTRACT FROM AUTHOR]

A series of novel benzoylhydrazone neuraminidase inhibitors were designed and synthesized on the base of pharmacophore-based virtual screening. [Display omitted] • An effective pharmacophore model for virtual screening was established and evaluated. • A series of NA inhibitors were designed and synthesized based on the characteristics of the pharmacophore model. • Some synthesized inhibitors exhibit excellent inhibitory activities against wild −type NA or mutant NA. • Molecular docking studies have shown that the 150-cavity and 430-cavity play an important role in discovery of novel novel benzoylhydrazone NA inhibitors. • This work provides unique insights in the discovery of potent NA inhibitors. Neuraminidase (NA) serves as a promising target for the exploration and development of anti-influenza drugs. In this work, lead compound 5 was discovered through pharmacophore-based virtual screening and molecular dynamics simulation, and 14 new compounds were obtained by modifying the lead compound 5 based on pharmacophore features. The biological activity test shows that 5n (IC 50 = 0.13 μM) has a better inhibitory effect on wild-type NA (H5N1), while 5i (IC 50 = 0.44 μM) has a prominent inhibitory effect on mutant NA (H5N1-H274Y), both of them are better than the positive control oseltamivir carboxylate (OSC). The analysis of docking results indicate that the good activities of compounds 5n and 5i may be attributed to the thiophene ring in 5n can stretch into the 150-cavity of NA, whereas the thiophene moiety in 5i can extend to the 430-cavity of NA. The findings of this study may be helpful for the discovery of new NA inhibitors. [ABSTRACT FROM AUTHOR]

Inspired by our earlier findings regarding neuraminidase (NA) inhibitors interacting with 150-cavity or 430-cavity of NA, sixteen novel polyheterocyclic NA inhibitors with 1,3,4-oxadiazole thioetheramide as core backbone were designed and synthesized based on the lead compound ZINC13401480. Of the synthesized compounds, compound N5 targeting 150-cavity exerts the best inhibitory activity against the wild-type H5N1 NA, with IC 50 value of 0.14 μM, which is superior to oseltamivir carboxylate (OSC) (IC 50 = 0.31 μM). Compound N10 targeting 430-cavity exhibits the best activity against the H5N1–H274Y mutant NA. Although the activity of N10 is comparable to that of OSC for wild-type H5N1 inhibition, it is approximately 60-fold more potent than OSC against the H274Y mutant, suggesting that it is not easy for the virus to develop drug resistance and is attractive for drug development. N10 (EC 50 = 0.11 μM) also exhibits excellent antiviral activity against H5N1, which is superior to the positive control OSC (EC 50 = 1.47 μM). Molecular docking study shows that the occupation of aromatic fused rings and oxadiazole moiety at the active site and the extension of the substituted phenyl to the 150-cavity or 430-cavity make great contributions to the good potency of this series of polyheterocyclic NA inhibitors. Some advancements in the discovery of effective target-specific NA inhibitors in this study may offer some assistance in the development of more potent anti-influenza drugs. Based on docking screening, a series of novel polyheterocyclic neuraminidase inhibitors with 1,3,4-oxadiazole thioetheramide as core backbone were designed and synthesized. [Display omitted] • Polyheterocyclic NA inhibitors were designed and synthesized with 1,3,4-oxadiazole thioetheramide as core backbone. • Some inhibitors exhibit excellent antiviral activity and inhibitory activities against H5N1 and H5N1-H274Y NA mutant. • Occupation of the active site and the 150- or 430-cavity makes great contributions to the good potency of the inhibitors. • This work provides a unique insight into the discovery of novel NA inhibitors against drug resistance. [ABSTRACT FROM AUTHOR]

There is an urgent need to develop and synthesize new anti-influenza drugs with activity against different strains, resistance to mutations, and suitability for various populations. Herein, we tested in vitro and in vivo the antiviral activity of new 1,2,3-triazole glycosides incorporating benzimidazole, benzooxazole, or benzotriazole cores synthesized by using a click approach. The Cu-catalyzation strategy consisted of 1,3-dipolar cycloaddition of the azidoalkyl derivative of the respective heterocyclic and different glycosyl acetylenes with five or six carbon sugar moieties. The antiviral activity of the synthesized glycosides against wild-type and neuraminidase inhibitor resistant strains of the avian influenza H5N1 and human influenza H1N1 viruses was high in vitro and in mice. Structure–activity relationship studies showed that varying the glycosyl moiety in the synthesized glycosides enhanced antiviral activity. The compound (2R,3R,4S,5R)-2-((1-(Benzo[d]thiazol-2-ylmethyl)-1H-1,2,3-triazol-4-yl)methoxy)tetrahydro-2H-pyran-3,4,5-triyl triacetate (Compound 9c) had a 50% inhibitory concentration (IC50) = 2.280 µM and a ligand lipophilic efficiency (LLE) of 6.84. The compound (2R,3R,4S,5R)-2-((1-((1H-Benzo[d]imidazol-2-yl)methyl)-1H-1,2,3-triazol-4-yl)methoxy)tetrahydro-2H-pyran-3,4,5-triyl triacetate had IC50 = 2.75 µM and LLE = 7.3 after docking analysis with the H5N1 virus neuraminidase. Compound 9c achieved full protection from H1N1 infection and 80% protection from H5N1 in addition to a high binding energy with neuraminidase and was safe in vitro and in vivo. This compound is suitable for further clinical studies as a new neuraminidase inhibitor. [ABSTRACT FROM AUTHOR]

Objectives: To investigate the susceptibility of epidemic influenza viruses to neuraminidase inhibitors (NAIs) and the emergence of resistant viruses after treatment, a prospective, observational study was done in the 2019-20 Japanese influenza season.Methods: Influenza viruses were isolated before and twice after treatment, the first at day 5 and the second at day 10. The 50% inhibitory concentrations (IC50s) to oseltamivir, zanamivir, peramivir, and laninamivir were measured and compared with those of 2010-11 to 2018-19 seasons. NA amino acid sequences were analyzed by next generation sequencing (NGS).Results: The IC50 geometric means of the NAIs for 128 A(H1N1)pdm09, 2 A(H3N2), and 33 B were comparable to those of the previous seasons. Only 2 (1.6%) A(H1N1)pdm09 with significantly high IC50 to oseltamivir were found pretreatment. No A(H3N2) or B had resistance. Treatment-emergent oseltamivir resistance was found in 2 among 33 oseltamivir-treated A(H1N1)pdm09, only at the second follow-up. The NGS indicated a rapid increase in the proportion of H275Y to wild type, from 0% to almost 100% between days 5 and 10.Conclusions: These results suggest the continued effectiveness of these NAIs for epidemic influenza in Japan. Treatment-emergent resistant H275Y A(H1N1)pdm09 viruses were detected after oseltamivir treatment, rapidly replacing the wild type. [ABSTRACT FROM AUTHOR]

Abstract Background Guidelines emphasize prompt antiviral treatment in severe influenza patients. Although nearly a 50% of severe influenza present with pneumonia, the effect of early (≤ 2 days after illness onset) neuraminidase inhibitor (NAI) use on the clinical outcomes of influenza A-related pneumonia (FluA-p) has rarely been assessed. Furthermore, data about the administration of NAIs in the real-world management of Flu-p in China are limited. Methods Data of patients hospitalised with FluA-p from five teaching hospitals in China from 1 January 2013 to 31 December 2018 were reviewed retrospectively. The impact of early NAI therapy on the outcomes in FluA-p patients, and the indications of early NAI administration by clinicians were evaluated by logistic regression analysis. Results In total, 693 FluA-p patients were included. Of these patients, 33.5% (232/693) were treated early. After adjusting for weighted propensity scores for treatment, systemic corticosteroid and antibiotic use, a multivariate logistic regression model showed that early NAI therapy was associated with decreased risk for invasive ventilation [odds ratio (OR) 0.511, 95% confidence interval (CI) 0.312–0.835, p = 0.007) and 30-day mortality (OR 0.533, 95% CI 0.210–0.807, p

Abstract Background After the A/H1N1 influenza pandemic in 2009, two new drugs against influenza, namely laninamivir and peramivir, were released in 2010 in Japan. We investigated prescription trends of four neuraminidase inhibitors (NAIs), which are laninamivir, peramivir, oseltamivir, and zanamivir, and assessed clinical data related to influenza for 8 years. Methods Patients living in Osaka Prefecture and diagnosed with influenza responded to a postcard questionnaire that collected data regarding their demographic characteristics, symptoms including fever, prescribed drugs, and influenza type. Results Laninamivir was most prescribed to patients aged ≥ 10 years, and oseltamivir was most prescribed to patients aged

Background Baloxavir marboxil (baloxavir) is a single-dose, oral antiinfluenza drug with a novel mechanism of action. We compared the incidence of hospitalization in patients treated with baloxavir vs neuraminidase inhibitors. Methods In this retrospective, observational, cohort study, we used real-world patient data extracted from a Japanese health insurance claims database. The enrollment period was 1 October 2018 to 17 April 2019. On day 1, eligible patients (N = 339 007) received baloxavir, oseltamivir, zanamivir, or laninamivir. Baseline characteristics were standardized using the inverse probability of treatment weighting method. The primary end point was the incidence of hospitalization (days 2–14). Secondary end points included antibacterial use, secondary pneumonia, and additional antiinfluenza drug use. Results Compared with the baloxavir group, the incidence of hospitalization was greater in the oseltamivir group (risk ratio [RR] and 95% confidence interval [CI], 1.41 [1.00–2.00]; risk difference [RD] and 95% CI, 0.06 [.01–.12]) and zanamivir group (RR, 1.85 [1.23–2.78]; RD, 0.11 [.02–.20]). Oseltamivir-treated patients were less likely to require antibacterials than baloxavir-treated patients (RR, 0.87 [.82–.91]). However, oseltamivir-treated patients were more likely to be hospitalized with antibacterials (RR, 1.70 [1.21–2.38]) or antibacterial injection (RR, 1.67 [1.17–2.38]) than baloxavir-treated patients (post hoc analysis). Compared with baloxavir-treated patients, additional antiinfluenza drug use was greater in oseltamivir-, zanamivir-, and laninamivir-treated patients (RR, 1.51 [1.05–2.18], 2.84 [2.04–3.96], and 1.68 [1.35–2.10], respectively). Conclusions Baloxavir is an efficacious antiinfluenza treatment that may reduce hospitalization compared with oseltamivir and zanamivir. Clinical Trials Registration University hospital Medical Information Network Clinical Trials Registry (UMIN000038159). [ABSTRACT FROM AUTHOR]

Zanamivir is an antiviral agent useful in the treatment and prevention of type A and B kind of influenza viruses in pediatric and adult patients. A specific and accurate analytical method by liquid chromatography with tandem mass spectrometry was developed and validated for the estimation of zanamivir in biological matrices of humans. Ledipasvir was utilized as an internal standard. Chromatographic elution of zanamivir and ledipasvir were attained on C18 Thermo base deactivated silica Hypersil column (50 mm×4.6 mm, 5 µm) with mobile phase of same polarity, made up of 0.1 % formic acid and acetonitrile (35:65, v/v) processed at 0.7 ml/min flow rate. The method was linear over the range of 2.15-64.5 ng/ml with more than 0.999 r2 values, consisting of 2.15 ng/ml as lower limit of quantification. Analytes were subjected for pretreatment by solid-phase extraction procedure with average extraction recovery findings within 95.7±1.23 %. Method accuracy findings were present in between 96.49 % to 103.88 % and the assessed percentage relative standard deviation findings for within and between run precision were =6.81 %. Analytes were subjected for variable storage conditions for the assessment of stability and were stable at respective conditions. The developed liquid chromatography with tandem mass spectrometry technique for the quantitation of zanamivir in biological matrix was suitable for routine analysis of patient's blood samples for the pharmacokinetic studies and drug monitoring. [ABSTRACT FROM AUTHOR]

Influenza virus, rhinovirus, and adenovirus frequently cause viral pneumonia, an important cause of morbidity and mortality especially in the extreme ages of life. During the last two decades, three outbreaks of coronavirus-associated pneumonia, namely Severe Acute Respiratory Syndrome, Middle-East Respiratory Syndrome, and the ongoing Coronavirus Infectious Disease—2019 (COVID-19) were reported. The rate of diagnosis of viral pneumonia is increasingly approaching 60% among children identified as having community-acquired pneumonia (CAP). Clinical presentation ranges from mild to severe pneumonitis complicated by respiratory failure in severe cases. The most vulnerable patients, the elderly and those living with cancer, report a relevant mortality rate. No clinical characteristics can be useful to conclusively distinguish the different etiology of viral pneumonia. However, accessory symptoms, such as anosmia or ageusia together with respiratory symptoms suggest COVID-19. An etiologic-based treatment of viral pneumonia is possible in a small percentage of cases only. Neuraminidase inhibitors have been proven to reduce the need for ventilatory support and mortality rate while only a few data support the large-scale use of other antivirals. A low-middle dose of dexamethasone and heparin seems to be effective in COVID-19 patients, but data regarding their possible efficacy in viral pneumonia caused by other viruses are conflicting. In conclusion, viral pneumonia is a relevant cause of CAP, whose interest is increasing due to the current COVID-19 outbreak. To set up a therapeutic approach is difficult because of the low number of active molecules and the conflicting data bearing supportive treatments such as steroids. [ABSTRACT FROM AUTHOR]

Four cases of oseltamivir-resistant influenza A(H1N1)pdm09 virus infection were detected among inhabitants of a border detention center in Texas, USA. Hemagglutinin of these viruses belongs to 6B.1A5A-156K subclade, which may enable viral escape from preexisting immunity. Our finding highlights the necessity to monitor both drug resistance and antigenic drift of circulating viruses. [ABSTRACT FROM AUTHOR]

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Background: UK Public Health England (PHE) guidelines recommend the liberal use of neuraminidase inhibitors (NAIs) in hospitalised adults with suspected influenza and are aligned with international guidelines. NAI use is recommended to start as early as possible and empirical use is recommended whilst awaiting laboratory results. Current UK hospital physician knowledge, attitudes, and practises regarding the use of NAIs, and levels of adherence to guideline recommendations are not known. Methods: This online, cross-sectional questionnaire-based survey of self-reported prescribing practice using clinical scenarios, was distributed to secondary care physicians involved in the assessment of adults presenting to hospital with suspected influenza. The primary outcome measure was adherence to PHE guidelines. Results: There were 237 respondents to the survey. 157 (67%) of 233 respondents reported awareness of PHE guidelines. Adherence to guidelines in the clinical scenarios ranged from 56% (95% CI 49–63%) to 72% (95% CI 66–79%) with considerable variability between specialities (p = 0.0008). Not treating suspected cases was common as was withholding of NAIs whilst awaiting laboratory results, despite the acknowledgment of prolonged turnaround times. 73 of 220 (33%) respondents reported that concerns about NAI efficacy influenced their prescribing. Conclusions: Concordance with national guidelines for the treatment of influenza is sub-optimal. Lack of guideline awareness and concerns over the effectiveness of NAIs are contributing factors. This study highlights a disparity between public health policy and clinical practice and suggests that strategies that promote rapid diagnostic testing and adherence to treatment guidelines are required. Study registration: This study is registered with the ISRCTN:18249297.

Seasonal influenza epidemics result in substantial health care burden annually. Early initiation of antiviral treatment of influenza has been shown to reduce the risk of complications and duration of illness. Pregnant and postpartum women may be at increased risk for influenza-associated complications; however, pregnant women have been generally excluded from clinical trials of antiviral treatment of influenza. In this review, we summarize the available evidence on the clinical effectiveness and safety of antiviral treatment of pregnant women with influenza. Observational data show a reduction of severe outcomes when pregnant and postpartum women are treated with oseltamivir and other neuraminidase inhibitors without increased risk of adverse maternal, fetal, or neonatal outcomes. Due to lack of safety and efficacy data for baloxavir treatment of pregnant and postpartum women, baloxavir is currently not recommended for use in these populations. [ABSTRACT FROM AUTHOR]

Few treatment options are available for oseltamivir‐resistant influenza. It has been proposed that baloxavir can be effective in this setting due to its distinct mechanism of action but clinical experience is lacking for immunocompromised patients. We report two such cases treated with baloxavir after failure of oseltamivir and detection of oseltamivir resistance mutations. Baloxavir/zanamivir combination therapy was effective in one patient, but persistent viral shedding was noted with baloxavir monotherapy in the other patient. [ABSTRACT FROM AUTHOR]

Purpose Baloxavir marboxil is a novel antiviral agent for influenza, introduced into clinical practice in 2018. A concern remains about the variant virus with reduced susceptibility after baloxavir exposure and its clinical consequences such as healthcare‐seeking behavior. Methods: Using a healthcare database in Japan, we compared the medical resource use following baloxavir and neuraminidase inhibitors (NAIs) treatment among children aged 7–15 years. The study period was from December 2018 to March 2019. The primary endpoint was the composite of hospitalization, laboratory and radiological tests, and antibiotic use over 1–9 days of antiviral treatment. As exploratory analyses, secondary outcomes being each single component of the primary composite were assessed and subgroup analyses comparing baloxavir with each NAI were done. Results: Data from 115 867 prescriptions in 115 238 children were analyzed (median age: 10 years; severe influenza risk in 26%; baloxavir accounting for 43%). Overall, baloxavir use did not increase subsequent medical resource utilization in the composite endpoint (adjusted odds ratio [aOR]: 1.04; 95% confidence interval [CI]: 0.99–1.09; P = 0.14), as were likelihoods of other secondary outcomes. In the subgroup analysis, baloxavir use was associated with higher medical resource use than oseltamivir (aOR: 1.21; 95% CI: 1.13–1.31; P < 0.001) and lower resource use than zanamivir (aOR: 0.93; 95% CI 0.86–1.00; P = 0.040). Conclusions: Based on a single‐year experience in Japan, prescribing baloxavir rather than NAIs did not increase medical resource utilization within 9 days of treatment, except in one exploratory comparison with oseltamivir. [ABSTRACT FROM AUTHOR]

Objective: To review the pharmacology, clinical trial data, and clinical implications for the intravenous formulation of zanamivir.Data Sources: MEDLINE, PubMed, EMBASE, and Google Scholar were searched during November 2019 to July 2020. Search terms zanamivir and neuraminidase inhibitor were used.Study Selection and Data Extraction: All human trials and major reports from compassionate use programs with the intravenous zanamivir (IVZ) formulation were assessed and reviewed here.Data Synthesis: IVZ was found to be similar but not superior to oral oseltamivir in hospitalized patients when studied in populations with very low baseline oseltamivir resistance. IVZ provides an effective alternative for critically ill patients when oral antiviral therapy is not preferred or when oseltamivir resistance is increased.Relevance To Patient Care and Clinical Practice: IVZ was recently authorized for use by the European Medicines Agency, and it is eligible for consideration in emergency use protocols and US stockpile inclusion. It will be of particular interest in critically ill patients especially during influenza seasons with appreciable oseltamivir and peramivir resistance.Conclusions: The available information suggests that the intravenous formulation of zanamivir offers a viable alternative treatment for critically ill patients with influenza, especially when resistance to other agents is present. [ABSTRACT FROM AUTHOR]

Neuraminic acid (Neu5Ac, also known as sialic acid) is an important monosaccharide found in glycoproteins and glycolipids which plays a vital role in regulation of physiological functions and pathological conditions. The study of sialoglycans has benefitted from the development of glycomimetic probes and inhibitors for proteins and enzymes that interact with and modify neuraminic acid in glycan chains. Methods to access sialoside intermediates with high yield are needed to facilitate the design of new targets. Here, we report the synthesis of C5-azido thiosialosides using a mild method to deprotect the C5-acetamido functional group followed by the use of a diazo-transfer reagent. We examined two diazo-transfer strategies and compared their yields and tolerance of acetate protecting groups. The same methods and comparisons were also performed for the 2,3-dehydro-5- N -acetylneuraminic acid (DANA) scaffold which is commonly used to generate inhibitors of neuraminidase (sialidase) enzymes. We found that C5-azido derivatives of both thiosialosides and DANA could be produced in five or six steps with yields up to 76 % and 83 %, respectively. Diazo-transfer reagents compared in this study were trifluoromethanesulfonyl azide (TfN 3) and imidazole-1-sulfonyl azide (ImzSO 2 N 3). We found that both reagents were compatible with this method and showed comparable yields. Finally, we show that C5-azido derivatives can help to avoid O , N -acyl protecting group migration which was observed in C5-NHAc analogs. [Display omitted] • Improved synthetic routes to access sialosides or DANA analogs with a C5-azide. • C5-azido DANA analogs avoid protecting group migrations found for N -Acyl intermediates. • Preparation of the C5-azido DANA in 6 steps and up to 80% yield. [ABSTRACT FROM AUTHOR]

In this research, we employed a deep reinforcement learning (RL)-based molecule design platform to generate a diverse set of compounds targeting the neuraminidase (NA) of influenza A and B viruses. A total of 60,291 compounds were generated, of which 86.5 % displayed superior physicochemical properties compared to oseltamivir. After narrowing down the selection through computational filters, nine compounds with non-sialic acid-like structures were selected for in vitro experiments. We identified two compounds, DS-22-inf-009 and DS-22-inf-021 that effectively inhibited the NAs of both influenza A and B viruses (IAV and IBV), including H275Y mutant strains at low micromolar concentrations. Molecular dynamics simulations revealed a similar pattern of interaction with amino acid residues as oseltamivir. In cell-based assays, DS-22-inf-009 and DS-22-inf-021 inhibited IAV and IBV in a dose-dependent manner with EC 50 values ranging from 0.29 μM to 2.31 μM. Furthermore, animal experiments showed that both DS-22-inf-009 and DS-22-inf-021 exerted antiviral activity in mice, conferring 65 % and 85 % protection from IAV (H1N1 pdm09), and 65 % and 100 % protection from IBV (Yamagata lineage), respectively. Thus, these findings demonstrate the potential of RL to generate compounds with promising antiviral properties. • Deep reinforcement learning was used to design anti-influenza agents. • All designed compounds have non-sialic acid-like structures. • DS-22-inf-009 and -021 showed antiviral activity against influenza viruses in vitro. • Both compounds protected mice from influenza A and B viruses. [ABSTRACT FROM AUTHOR]

Neuraminidase (NA) cleaves the glycosidic bond linkages of sialic acids to release the mature virions from infected cells and has been an attractive therapeutic target for anti-influenza agents. In our ongoing investigation of NA inhibitors in mushroom extracts, we found that the extract the fruiting body of Glaziella splendens potently inhibited neuraminidase. The fruiting bodies of G. splendens were extracted and partitioned successively with hexane, ethyl acetate, and butanol. The ethyl acetate soluble-layer was subjected to silica gel and Sephadex LH-20 column chromatographies, and MPLC to obtain five compounds (1–5). Their structures were determined by spectroscopic methods. NA inhibitory activity of these compounds was evaluated using NAs from recombinant rvH1N1, H3N2, and H5N1 influenza A viruses. One compound (1) was elucidated as a new azaphilone derivative, and four compounds (2–5) were identified as entonaemin A, comazaphilone D, rubiginosin A, and entonaemin B, respectively. Compounds 3 and 4 showed considerable inhibitory activity against three types of neuraminidases with the IC50 values of 30.9, 41.8, and 35.7 µM for 3 and 46.5, 50.4, and 29.9 µM for 4, respectively. This study reveals that the fruiting bodies of G. splendens possess azaphilone derivatives with the NA inhibitory activity. This is the first report on the isolation of neuraminidase inhibitors from the fruiting bodies of G. splendens.

Victor Yu Glanz,1 Veronika A Myasoedova,2 Andrey V Grechko,3 Alexander N Orekhov2,4 1Department of Genetics, Cytology and Bioengineering, Faculty of Biology and Medicine, Voronezh State University, Voronezh, Russia; 2Laboratory of Angiopathology, Institute of General Pathology and Pathophysiology, Moscow, Russia; 3Federal Research and Clinical Center of Intensive Care Medicine and Rehabilitology, Moscow, Russia; 4Institute for Atherosclerosis Research, Skolkovo Innovative Center, Moscow, Russia Abstract: The demand for novel anti-influenza drugs persists, which is highlighted by the recent pandemics of influenza affecting thousands of people across the globe. One of the approaches to block the virus spreading is inhibiting viral sialidase (neuraminidase). This enzyme cleaves the sialic acid link between the newly formed virions and the host cell surface liberating the virions from the cell and maintaining the cycle of infection. Viral neuraminidases appear therefore as attractive therapeutic targets for preventing further spread of influenza infection. Compared to ion channel blockers that were the first approved anti-influenza drugs, neuraminidase inhibitors are well tolerated and target both influenza A and B viruses. Moreover, neuraminidase/sialidase inhibitors may be useful for managing some other human pathologies, such as cancer. In this review, we discuss the available knowledge on neuraminidase or sialidase inhibitors, their design, clinical application, and the current challenges. Keywords: sialidase, neuraminidase, neuraminidase inhibitor, influenza, drug design

The coronaviruses plaguing humanity in the 21st century share much in common: a spontaneous route of origin from wild animals, a propensity to take human life, and, importantly, a highly conserved set of biological machinery necessary for viral replication. Most recently, the SARS-CoV-2 is decimating economies around the world and has claimed over two million human lives, reminding the world of a need for an effective drug against present and future coronaviruses. To date, attempts to repurpose clinically approved antiviral medications show minimal promise, highlighting the need for development of new antiviral drugs. Nucleotide analog inhibitors are a promising therapeutic candidate, but early data from clinical studies suggests these compounds have limited efficacy. However, novel compounds targeting the main protease responsible for critical steps in viral assembly are gaining considerable interest because they offer the potential for broad-spectrum coronavirus therapy. Here, we review the literature regarding potential inhibitors for the main protease of coronaviruses, especially SARS-CoV-2, analyze receptor-drug interactions, and draw conclusions about candidate inhibitors for future outbreaks. Promising candidates for development of a broad-spectrum coronavirus protease inhibitor include the neuraminidase inhibitor 3K, the peptidomimetic inhibitor 11a and 11b, the α-ketoamide inhibitor 13b, the aldehyde prodrug, and the phosphate prodrug developed by Pfizer. In silico and in vitro analyses have shown that these inhibitors strongly interact with the active site of the main protease, and to varying degrees, prevent viral replication via interactions with the largely conserved active site pockets. [ABSTRACT FROM AUTHOR]

Laninamivir (LAN) is a long-acting neuraminidase (NA) inhibitor (NAI) with a similar binding profile in the influenza NA enzyme active site as those of other NAIs, oseltamivir (OS), zanamivir (ZAN), and peramivir, and may share common resistance markers with these NAIs. We screened viruses with NA substitutions previously found during OS and ZAN selection in avian influenza viruses (AIVs) of the N3 to N9 subtypes for LAN susceptibility. Of the 72 NA substitutions, 19 conferred resistance to LAN, which ranged from 11.2- to 549.8-fold-decreased inhibitory activity over that of their parental viruses. Ten NA substitutions reduced the susceptibility to all four NAIs, whereas the remaining 26 substitutions yielded susceptibility to one or more NAIs. To determine whether the in vitro susceptibility of multi-NAI-resistant AIVs is associated with in vivo susceptibility, we infected BALB/c mice with recombinant AIVs with R292K (ma81K-N3R292K) or Q136K (ma81K-N8Q136K) NA substitutions, which impart in vitro susceptibility only to LAN or OS, respectively. Both ma81KN3R292K and ma81K-N8Q136K virus-infected mice exhibited reduced weight loss, mortality, and lung viral titers when treated with their susceptible NAIs, confirming the in vitro susceptibility of these substitutions. Together, LAN resistance profiling of AIVs of a range of NA subtypes improves the understanding of NAI resistance mechanisms. Furthermore, the association of in vitro and in vivo NAI susceptibility indicates that our models are useful tools for monitoring NAI susceptibility of AIVs. [ABSTRACT FROM AUTHOR]

Puerarin is a major isofiavone compound isolated from the root of Pueraria lobata. It was reported that puerarin had antioxidant, antiinflammatory, antitumor, cholesterol lowering, liver protective, and neuroprotective properties. However, few studies have explored the antiviral effect of puerarin and its target mechanism related to influenza virus. Here, the antiinfluenza activity of puerarin in vitro and in vivo and its mode of action on the potential inhibition of neuraminidase (NA) were investigated. Puerarin displayed an inhibitory effect on A/FM/1/1947(H1N1) (EC50 = 52.06 μM). An indirect immunofluorescence assay indicated that puerarin blocked the nuclear export of viral NP. The inhibition of NA activity confirmed that puerarin can block the release of newly formed virus particles from infected cells. Puerarin (100 and 200 mg/kg/d) exhibited effective antiviral activity in mice, conferring 50% and 70% protection from death against H1N1, reducing virus titers, and effectively alleviating inflammation in the lungs. The molecular docking results showed that puerarin had a strong binding affinity with NA from H1N1. The results of the molecular dynamics simulation revealed that puerarin had higher stable binding at the 150‐loop region of the NA protein. These results demonstrated that puerarin acts as a NA blocker to inhibit influenza A virus both in cellular and animal models. Thus, puerarin has potential utility for the treatment of the influenza virus infection. [ABSTRACT FROM AUTHOR]

There is an urgent need to develop and synthesize new anti-influenza drugs with activity against different strains, resistance to mutations, and suitability for various populations. Herein, we tested in vitro and in vivo the antiviral activity of new 1,2,3-triazole glycosides incorporating benzimidazole, benzooxazole, or benzotriazole cores synthesized by using a click approach. The Cu-catalyzation strategy consisted of 1,3-dipolar cycloaddition of the azidoalkyl derivative of the respective heterocyclic and different glycosyl acetylenes with five or six carbon sugar moieties. The antiviral activity of the synthesized glycosides against wild-type and neuraminidase inhibitor resistant strains of the avian influenza H5N1 and human influenza H1N1 viruses was high in vitro and in mice. Structure–activity relationship studies showed that varying the glycosyl moiety in the synthesized glycosides enhanced antiviral activity. The compound (2R,3R,4S,5R)-2-((1-(Benzo[d]thiazol-2-ylmethyl)-1H-1,2,3-triazol-4-yl)methoxy)tetrahydro-2H-pyran-3,4,5-triyl triacetate (Compound 9c) had a 50% inhibitory concentration (IC50) = 2.280 µM and a ligand lipophilic efficiency (LLE) of 6.84. The compound (2R,3R,4S,5R)-2-((1-((1H-Benzo[d]imidazol-2-yl)methyl)-1H-1,2,3-triazol-4-yl)methoxy)tetrahydro-2H-pyran-3,4,5-triyl triacetate had IC50 = 2.75 µM and LLE = 7.3 after docking analysis with the H5N1 virus neuraminidase. Compound 9c achieved full protection from H1N1 infection and 80% protection from H5N1 in addition to a high binding energy with neuraminidase and was safe in vitro and in vivo. This compound is suitable for further clinical studies as a new neuraminidase inhibitor.