Interferon Lambda Delays the Emergence of Influenza Virus Resistance to Oseltamivir

Influenza viruses are a leading cause of morbidity and mortality worldwide. These air-borne pathogens are able to cross the species barrier, leading to regular seasonal epidemics and sporadic pandemics. Influenza viruses also possess a high genetic variability, which allows for the acquisition of resistance mutations to antivirals. Combination therapies with two or more drugs targeting different mechanisms of viral replication have been considered an advantageous option to not only enhance the effectiveness of the individual treatments, but also reduce the likelihood of resistance emergence. Using an in vitro infection model, we assessed the barrier to viral resistance of a combination therapy with the neuraminidase inhibitor oseltamivir and human interferon lambda against the pandemic H1N1 A/Netherlands/602/2009 (H1N1pdm09) virus. We serially passaged the virus in a cell line derived from human bronchial epithelial cells in the presence or absence of increasing concentrations of oseltamivir alone or oseltamivir plus interferon lambda. While the treatment with oseltamivir alone quickly induced the emergence of antiviral resistance through a single mutation in the neuraminidase gene, the co-administration of interferon lambda delayed the emergence of drug-resistant influenza virus variants. Our results suggest a possible clinical application of interferon lambda in combination with oseltamivir to treat influenza.

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Zanamivir, at 600 Milligrams Twice Daily, Inhibits Oseltamivir-Resistant 2009 Pandemic H1N1 Influenza Virus in anIn VitroHollow-Fiber Infection Model System

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.01628-10 ◽

2011 ◽

Vol 55 (4) ◽

pp. 1740-1746 ◽

Cited By ~ 15

Author(s):

Ashley N. Brown ◽

James J. McSharry ◽

Qingmei Weng ◽

Jonathan R. Adams ◽

Robert Kulawy ◽

...

Keyword(s):

Influenza Virus ◽

Influenza A ◽

H1n1 Influenza ◽

Model System ◽

Hospitalized Patients ◽

Influenza Viruses ◽

Infection Model ◽

Pandemic H1n1 ◽

The Government

ABSTRACTIn 2009, a novel H1N1 influenza A virus emerged and spread worldwide, initiating a pandemic. Various isolates obtained from disparate parts of the world were shown to be uniformly resistant to the adamantanes but sensitive to the neuraminidase inhibitors oseltamivir and zanamivir. Over time, resistance to oseltamivir became more prevalent among pandemic H1N1 virus isolates, while most remained susceptible to zanamivir. The government has proposed the use of intravenous (i.v.) zanamivir to treat serious influenza virus infections among hospitalized patients. To use zanamivir effectively for patients with severe influenza, it is necessary to know the optimal dose and schedule of administration of zanamivir that will inhibit the replication of oseltamivir-sensitive and -resistant influenza viruses. Therefore, we performed studies using thein vitrohollow-fiber infection model system to predict optimal dosing regimens for zanamivir against an oseltamivir-sensitive and an oseltamivir-resistant virus. Our results demonstrated that zanamivir, at a dose of 600 mg given twice a day (Q12h), inhibited the replication of oseltamivir-sensitive and oseltamivir-resistant influenza viruses throughout the course of the experiment. Thus, our findings suggest that intravenous zanamivir, at a dose of 600 mg Q12h, could be used to treat hospitalized patients suffering from serious infections with oseltamivir-sensitive or -resistant influenza viruses.

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Macrocyclic peptides exhibit antiviral effects against influenza virus HA and prevent pneumonia in animal models

Nature Communications ◽

10.1038/s41467-021-22964-w ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Makoto Saito ◽

Yasushi Itoh ◽

Fumihiko Yasui ◽

Tsubasa Munakata ◽

Daisuke Yamane ◽

...

Keyword(s):

Influenza Virus ◽

Cynomolgus Macaque ◽

Severe Pneumonia ◽

Neuraminidase Inhibitor ◽

Influenza Viruses ◽

Viral Envelope ◽

Viral Envelope Protein ◽

Wide Range ◽

Macrocyclic Peptides

AbstractMost anti-influenza drugs currently used, such as oseltamivir and zanamivir, inhibit the enzymatic activity of neuraminidase. However, neuraminidase inhibitor-resistant viruses have already been identified from various influenza virus isolates. Here, we report the development of a class of macrocyclic peptides that bind the influenza viral envelope protein hemagglutinin, named iHA. Of 28 iHAs examined, iHA-24 and iHA-100 have inhibitory effects on the in vitro replication of a wide range of Group 1 influenza viruses. In particular, iHA-100 bifunctionally inhibits hemagglutinin-mediated adsorption and membrane fusion through binding to the stalk domain of hemagglutinin. Moreover, iHA-100 shows powerful efficacy in inhibiting the growth of highly pathogenic influenza viruses and preventing severe pneumonia at later stages of infection in mouse and non-human primate cynomolgus macaque models. This study shows the potential for developing cyclic peptides that can be produced more efficiently than antibodies and have multiple functions as next-generation, mid-sized biomolecules.

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CS-8958, a Prodrug of the New Neuraminidase Inhibitor R-125489, Shows Long-Acting Anti-Influenza Virus Activity

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00333-08 ◽

2008 ◽

Vol 53 (1) ◽

pp. 186-192 ◽

Cited By ~ 152

Author(s):

Makoto Yamashita ◽

Takanori Tomozawa ◽

Masayo Kakuta ◽

Akane Tokumitsu ◽

Hatsumi Nasu ◽

...

Keyword(s):

Influenza Virus ◽

Acyl Chain ◽

Neuraminidase Inhibitor ◽

Influenza Viruses ◽

Infection Model ◽

Acyl Chain Length ◽

Long Acting ◽

Survival Effect ◽

First Time

ABSTRACT Two neuraminidase (NA) inhibitors, zanamivir (Relenza) and oseltamivir phosphate (Tamiflu), have been licensed for the treatment of and prophylaxis against influenza. In this paper, the new potent NA inhibitor R-125489 is reported for the first time. R-125489 inhibited the NA activities of various type A and B influenza viruses, including subtypes N1 to N9 and oseltamivir-resistant viruses. The survival effect of R-125489 was shown to be similar to that of zanamivir when administered intranasally in a mouse influenza virus A/Puerto Rico/8/34 infection model. Moreover, we found that the esterified form of R-125489 showed improved efficacy compared to R-125489 and zanamivir, depending on the acyl chain length, and that 3-(O)-octanoyl R-125489 (CS-8958) was the best compound in terms of its life-prolonging effect (P < 0.0001, compared to zanamivir) in the same infection model. A prolonged survival effect was observed after a single administration of CS-8958, even if it was given 7 days before infection. It is suggested that intranasally administered CS-8958 works as a long-acting NA inhibitor and shows in vivo efficacy as a result of a single intranasal administration.

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Biological Cost of Single and Multiple Norfloxacin Resistance Mutations in Escherichia coli Implicated in Urinary Tract Infections

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.49.6.2343-2351.2005 ◽

2005 ◽

Vol 49 (6) ◽

pp. 2343-2351 ◽

Cited By ~ 91

Author(s):

Patricia Komp Lindgren ◽

Linda L. Marcusson ◽

Dorthe Sandvang ◽

Niels Frimodt-Møller ◽

Diarmaid Hughes

Keyword(s):

Escherichia Coli ◽

Urinary Tract ◽

Infection Model ◽

Resistance Mutations ◽

Topoisomerase Iv ◽

E Coli ◽

Tract Infections ◽

Subsequent Selection

ABSTRACT Resistance to fluoroquinolones in urinary tract infection (UTIs) caused by Escherichia coli is associated with multiple mutations, typically those that alter DNA gyrase and DNA topoisomerase IV and those that regulate AcrAB-TolC-mediated efflux. We asked whether a fitness cost is associated with the accumulation of these multiple mutations. Mutants of the susceptible E. coli UTI isolate Nu14 were selected through three to five successive steps with norfloxacin. Each selection was performed with the MIC of the selected strain. After each selection the MIC was measured; and the regions of gyrA, gyrB, parC, and parE, previously associated with resistance mutations, and all of marOR and acrR were sequenced. The first selection step yielded mutations in gyrA, gyrB, and marOR. Subsequent selection steps yielded mutations in gyrA, parE, and marOR but not in gyrB, parC, or acrR. Resistance-associated mutations were identified in almost all isolates after selection steps 1 and 2 but in less than 50% of isolates after subsequent selection steps. Selected strains were competed in vitro, in urine, and in a mouse UTI infection model against the starting strain, Nu14. First-step mutations were not associated with significant fitness costs. However, the accumulation of three or more resistance-associated mutations was usually associated with a large reduction in biological fitness, both in vitro and in vivo. Interestingly, in some lineages a partial restoration of fitness was associated with the accumulation of additional mutations in late selection steps. We suggest that the relative biological costs of multiple mutations may influence the evolution of E. coli strains that develop resistance to fluoroquinolones.

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Human Interferon Inducible Transmembrane Protein 3 (IFITM3) Inhibits Influenza Virus A Replication and Inflammation by Interacting with ABHD16A

BioMed Research International ◽

10.1155/2021/6652147 ◽

2021 ◽

Vol 2021 ◽

pp. 1-9

Author(s):

Liang Chen ◽

Limei Zhu ◽

Jun Chen

Keyword(s):

Influenza Virus ◽

Influenza A ◽

Transmembrane Protein ◽

Influenza Viruses ◽

Expression Level ◽

Hek293 Cells ◽

Human Interferon ◽

Control Group ◽

Mrna Levels ◽

Membrane Area

Studies have shown that human interferon inducible transmembrane protein (hIFITMs) family proteins have broad-spectrum antiviral capabilities. Preliminary studies in our laboratory have tentatively proved that hIFITMs have the effect of inhibiting influenza viruses. In order to further study its mechanism and role in the occurrence and development of influenza A, relevant studies have been carried out. Fluorescence quantitative polymerase chain reaction (PCR) detection technology was used to observe the effect of hIFITM3 on the replication of influenza A virus (IVA) and the interaction with hABHD16A. In HEK293 cells, overexpression of hIFITM3 protein significantly inhibited the replication of IVA at 24 h, 48 h, and 72 h; yeast two-hybrid experiment proved that hIFITM3 interacts with hABHD16A; laser confocal microscopy observations showed that hIFITM3 and hABHD16A colocalized in the cell membrane area; the expression level of inflammation-related factors in cells overexpressing hIFITM3 or hABHD16A was detected by fluorescence quantitative PCR, and the results showed that the mRNA levels of interleukin- (IL-) 1β, IL-6, IL-10, tumor necrosis factor- (TNF-) α, and cyclooxygenase 2 (COX2) were significantly increased. But when hIFITM3/hABHD16A was coexpressed, the mRNA expression levels of these cytokines were significantly reduced except COX2. When influenza virus infected cells coexpressing hIFITM3/hABHD16A, the expression level of inflammatory factors decreased compared with the control group, indicating that hIFITM3 can play an important role in regulating inflammation balance. This study confirmed that hIFITM3 has an effect of inhibiting IVA replication. Furthermore, it was found that hIFITM3 interacts with hABHD16A, following which it can better inhibit the replication of influenza virus and the inflammatory response caused by the disease process.

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A Host-Restricted Self-Attenuated Influenza Virus Provides Broad Pan-Influenza A Protection in a Mouse Model

Frontiers in Immunology ◽

10.3389/fimmu.2021.779223 ◽

2021 ◽

Vol 12 ◽

Author(s):

Minjin Kim ◽

Yucheol Cheong ◽

Jinhee Lee ◽

Jongkwan Lim ◽

Sanguine Byun ◽

...

Keyword(s):

Influenza Virus ◽

Mouse Model ◽

Neutralizing Antibodies ◽

Influenza A ◽

Protective Efficacy ◽

Influenza Viruses ◽

Cross Protection ◽

Infection Model ◽

Wild Type ◽

Group 1

Influenza virus infections can cause a broad range of symptoms, form mild respiratory problems to severe and fatal complications. While influenza virus poses a global health threat, the frequent antigenic change often significantly compromises the protective efficacy of seasonal vaccines, further increasing the vulnerability to viral infection. Therefore, it is in great need to employ strategies for the development of universal influenza vaccines (UIVs) which can elicit broad protection against diverse influenza viruses. Using a mouse infection model, we examined the breadth of protection of the caspase-triggered live attenuated influenza vaccine (ctLAIV), which was self-attenuated by the host caspase-dependent cleavage of internal viral proteins. A single vaccination in mice induced a broad reactive antibody response against four different influenza viruses, H1 and rH5 (HA group 1) and H3 and rH7 subtypes (HA group 2). Notably, despite the lack of detectable neutralizing antibodies, the vaccination provided heterosubtypic protection against the lethal challenge with the viruses. Sterile protection was confirmed by the complete absence of viral titers in the lungs and nasal turbinates after the challenge. Antibody-dependent cellular cytotoxicity (ADCC) activities of non-neutralizing antibodies contributed to cross-protection. The cross-protection remained robust even after in vivo depletion of T cells or NK cells, reflecting the strength and breadth of the antibody-dependent effector function. The robust mucosal secretion of sIgA reflects an additional level of cross-protection. Our data show that the host-restricted designer vaccine serves an option for developing a UIV, providing pan-influenza A protection against both group 1 and 2 influenza viruses. The present results of potency and breadth of protection from wild type and reassortant viruses addressed in the mouse model by single immunization merits further confirmation and validation, preferably in clinically relevant ferret models with wild type challenges.

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Characterization of Human Influenza Virus Variants Selected In Vitro in the Presence of the Neuraminidase Inhibitor GS 4071

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.42.12.3234 ◽

1998 ◽

Vol 42 (12) ◽

pp. 3234-3241 ◽

Cited By ~ 108

Author(s):

Chun Y. Tai ◽

Paul A. Escarpe ◽

Robert W. Sidwell ◽

Matthew A. Williams ◽

Willard Lew ◽

...

Keyword(s):

Influenza Virus ◽

Neuraminidase Inhibitor ◽

H3n2 Virus ◽

Wild Type Virus ◽

Type Virus ◽

Human Influenza ◽

Wild Type ◽

Viral Virulence ◽

High Level

ABSTRACT An oral prodrug of GS 4071, a potent and selective inhibitor of influenza neuraminidases, is currently under clinical development for the treatment and prophylaxis of influenza virus infections in humans. To investigate the potential development of resistance during the clinical use of this compound, variants of the human influenza A/Victoria/3/75 (H3N2) virus with reduced susceptibility to the neuraminidase inhibitor GS 4071 were selected in vitro by passaging the virus in MDCK cells in the presence of inhibitor. After eight passages, variants containing two amino acid substitutions in the hemagglutinin (A28T in HA1 and R124M in HA2) but no changes in the neuraminidase were isolated. These variants exhibited a 10-fold reduction in susceptibility to GS 4071 and zanamivir (GG167) in an in vitro plaque reduction assay. After 12 passages, a second variant containing these hemagglutinin mutations and a Lys substitution for the conserved Arg292 of the neuraminidase was isolated. The mutant neuraminidase enzyme exhibited high-level (30,000-fold) resistance to GS 4071, but only moderate (30-fold) resistance to zanamivir and 4-amino-Neu5Ac2en, the amino analog of zanamivir. The mutant enzyme had weaker affinity for the fluorogenic substrate 2′-(4-methylumbelliferyl)-α-d- N -acetylneuraminic acid and lower enzymatic activity compared to the wild-type enzyme. The viral variant containing the mutant neuraminidase did not replicate as well as the wild-type virus in culture and was 10,000-fold less infectious than the wild-type virus in a mouse model. These results suggest that although the R292K neuraminidase mutation confers high-level resistance to GS 4071 in vitro, its effect on viral virulence is likely to render this mutation of limited clinical significance.

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Evaluation of the Biological Properties and Cross-Reactive Antibody Response to H10 Influenza Viruses in Ferrets

Journal of Virology ◽

10.1128/jvi.00895-17 ◽

2017 ◽

Vol 91 (19) ◽

Cited By ~ 6

Author(s):

Troy C. Sutton ◽

Elaine W. Lamirande ◽

Rita Czako ◽

Kanta Subbarao

Keyword(s):

Weight Loss ◽

Antibody Response ◽

Viral Replication ◽

Vaccine Strain ◽

Biological Properties ◽

Sialic Acids ◽

Influenza Viruses ◽

Species Barrier ◽

Reactive Antibody

ABSTRACT The recent outbreak of avian origin H10N7 influenza among seals in northern Europe and two fatal human infections with an avian H10N8 virus in China have demonstrated that H10 viruses can spread between mammals and cause severe disease in humans. To gain insight into the potential for H10 viruses to cross the species barrier and to identify a candidate vaccine strain, we evaluated the in vitro and in vivo properties and antibody response in ferrets to 20 diverse H10 viruses. H10 virus infection of ferrets caused variable weight loss, and all 20 viruses replicated throughout the respiratory tract; however, replication in the lungs was highly variable. In glycan-binding assays, the H10 viruses preferentially bound “avian-like” α2,3-linked sialic acids. Importantly, several isolates also displayed strong binding to long-chain “human-like” α2,6-linked sialic acids and exhibited comparable or elevated neuraminidase activity relative to human H1N1, H2N2, and H3N2 viruses. In hemagglutination inhibition assays, 12 antisera cross-reacted with ≥14 of 20 H10 viruses, and 7 viruses induced neutralizing activity against ≥15 of the 20 viruses. By combining data on weight loss, viral replication, and the cross-reactive antibody response, we identified A/mallard/Portugal/79906/2009 (H10N7) as a suitable virus for vaccine development. Collectively, our findings suggest that H10 viruses may continue to sporadically infect humans and other mammals, underscoring the importance of developing an H10 vaccine for pandemic preparedness. IMPORTANCE Avian origin H10 influenza viruses sporadically infect humans and other mammals; however, little is known about viruses of this subtype. Thus, we characterized the biological properties of 20 H10 viruses in vitro and in ferrets. Infection caused mild to moderate weight loss (5 to 15%), with robust viral replication in the nasal tissues and variable replication in the lung. H10 viruses preferentially bind “avian-like” sialic acids, although several isolates also displayed binding to “human-like” sialic acid receptors. This is consistent with the ability of H10 viruses to cross the species barrier and warrants selection of an H10 vaccine strain. By evaluating the cross-reactive antibody response to the H10 viruses and combining this analysis with viral replication and weight loss findings, we identified A/mallard/Portugal/79906/2009 (H10N7) as a suitable H10 vaccine strain.

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Mutations in the NS1 Protein of Swine Influenza Virus Impair Anti-Interferon Activity and Confer Attenuation in Pigs

Journal of Virology ◽

10.1128/jvi.79.12.7535-7543.2005 ◽

2005 ◽

Vol 79 (12) ◽

pp. 7535-7543 ◽

Cited By ~ 176

Author(s):

Alicia Solórzano ◽

Richard J. Webby ◽

Kelly M. Lager ◽

Bruce H. Janke ◽

Adolfo García-Sastre ◽

...

Keyword(s):

Influenza Virus ◽

Nonstructural Protein ◽

Swine Influenza Virus ◽

Swine Influenza ◽

Influenza Viruses ◽

Model Systems ◽

Ns1 Protein ◽

Wild Type ◽

Virulence Attenuation

ABSTRACT It has been shown previously that the nonstructural protein NS1 of influenza virus is an alpha/beta interferon (IFN-α/β) antagonist, both in vitro and in experimental animal model systems. However, evidence of this function in a natural host has not yet been obtained. Here we investigated the role of the NS1 protein in the virulence of a swine influenza virus (SIV) isolate in pigs by using reverse genetics. The virulent wild-type A/Swine/Texas/4199-2/98 (TX/98) virus and various mutants encoding carboxy-truncated NS1 proteins were rescued. Growth properties of TX/98 viruses with mutated NS1, induction of IFN in tissue culture, and virulence-attenuation in pigs were analyzed and compared to those of the recombinant wild-type TX/98 virus. Our results indicate that deletions in the NS1 protein decrease the ability of the TX/98 virus to prevent IFN-α/β synthesis in pig cells. Moreover, all NS1 mutant viruses were attenuated in pigs, and this correlated with the amount of IFN-α/β induced in vitro. These data suggest that the NS1 protein of SIV is a virulence factor. Due to their attenuation, NS1-mutated swine influenza viruses might have a great potential as live attenuated vaccine candidates against SIV infections of pigs.

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Laninamivir-Interferon Lambda 1 Combination Treatment Promotes Resistance by Influenza A Virus More Rapidly than Laninamivir Alone

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00301-20 ◽

2020 ◽

Vol 64 (7) ◽

Author(s):

Simone E. Adams ◽

Vladimir Y. Lugovtsev ◽

Anastasia Kan ◽

Nicolai V. Bovin ◽

Raymond P. Donnelly ◽

...

Keyword(s):

Influenza Virus ◽

Influenza A Virus ◽

Combination Treatment ◽

Influenza A ◽

Influenza Virus Infection ◽

The United States ◽

Epithelial Cell Line ◽

Drug Resistant ◽

Interferon Lambda

ABSTRACT Each year, 5% to 20% of the population of the United States becomes infected with influenza A virus. Combination therapy with two or more antiviral agents has been considered a potential treatment option for influenza virus infection. However, the clinical results derived from combination treatment with two or more antiviral drugs have been variable. We examined the effectiveness of cotreatment with two distinct classes of anti-influenza drugs, i.e., neuraminidase (NA) inhibitor, laninamivir, and interferon lambda 1 (IFN-λ1), against the emergence of drug-resistant virus variants in vitro. We serially passaged pandemic A/California/04/09 [A(H1N1)pdm09] influenza virus in a human lung epithelial cell line (Calu-3) in the presence or absence of increasing concentrations of laninamivir or laninamivir plus IFN-λ1. Surprisingly, laninamivir used in combination with IFN-λ1 promoted the emergence of the E119G NA mutation five passages earlier than laninamivir alone (passage 2 versus passage 7, respectively). Acquisition of this mutation resulted in significantly reduced sensitivity to the NA inhibitors laninamivir (∼284-fold) and zanamivir (∼1,024-fold) and decreased NA enzyme catalytic activity (∼5-fold) compared to the parental virus. Moreover, the E119G NA mutation emerged together with concomitant hemagglutinin (HA) mutations (T197A and D222G), which were selected more rapidly by combination treatment with laninamivir plus IFN-λ1 (passages 2 and 3, respectively) than by laninamivir alone (passage 10). Our results show that treatment with laninamivir alone or in combination with IFN-λ1 can lead to the emergence of drug-resistant influenza virus variants. The addition of IFN-λ1 in combination with laninamivir may promote acquisition of drug resistance more rapidly than treatment with laninamivir alone.

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