#Zanamivir - #Amantadine Conjugate: A Dual-Action Agent with Broad-Spectrum Synergistic #Antiviral Efficacy

 

Abstract

Influenza A virus is a highly contagious respiratory pathogen, and its rapid and continuous adaptive mutations for immune escape have limited the efficacy of existing vaccines and antiviral drugs. Here, we report a multimechanism anti-influenza platform based on the conjugation of zanamivir (ZMV) with amantadine (Aman). Aman acts as a hydrophobic tag that promotes the degradation of neuraminidase and concurrently enhances the physicochemical properties of ZMV, leading to improved membrane permeability and a significantly prolonged half-life. Meanwhile, the ZMV moiety counteracts Aman-induced cytotoxic autophagy. The resulting conjugate, compound 7j, exhibits potent activity against a wide range of neuraminidase and M2 ion channel mutations. Notably, a single intravenous dose of 7j fully protected mice from a lethal H1N1 challenge. Our work demonstrates that the rational fusion of ZMV and Aman achieves synergistic multimechanistic antiviral activity with enhanced efficacy and safety, offering a new strategy for the development of next-generation anti-influenza drugs.

Source: 

Link: https://pubs.acs.org/doi/10.1021/acs.jmedchem.5c03547

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#Baloxavir outperforms #oseltamivir, #favipiravir, and #amantadine in treating lethal #influenza #H5N1 HA clade 2.3.4.4b #infection in #mice

 

Abstract

Intercontinental spread of highly pathogenic avian influenza A(H5N1) viruses poses significant pandemic risks and necessitates strong protective countermeasures. We evaluated the therapeutic efficacy of the neuraminidase inhibitor oseltamivir, the polymerase inhibitors baloxavir and favipiravir, and an ion-channel blocker amantadine, against severe influenza A(H5N1) virus infection in female BALB/c mice. Baloxavir (≥10 mg/kg, 1 dose) fully protected mice from death, significantly reduced virus respiratory replication, and prevented neuroinvasion. Oseltamivir (≥100 mg/kg/day for 5 days) provided limited survival benefits, reduced lung titers but failed to prevent viral neuroinvasion. Favipiravir (≥100 mg/kg/day for 5 days) provided partial protection, although did not reduce viral titers in lungs and brain. Amantadine provided no benefits. Although all drugs inhibited A(H5N1) viruses in vitro, in vivo correlations did not extend beyond baloxavir. Our results indicate that baloxavir is the most reliable treatment to address both respiratory replication and systemic spread of contemporary A(H5N1) viruses in mice and should be considered in pandemic planning.

Source: 

Link: https://www.nature.com/articles/s41467-026-69721-5

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Genetic diversity of #H5N1 avian #influenza viruses isolated from #birds and #seals in #Russia in 2023

Abstract

Thousands of outbreaks of the highly pathogenic avian influenza A(H5N1) virus in birds and an increasing number of mammal infections are registered annually. In 2023, multiple avian influenza outbreaks were registered among wild birds, poultry and seals in Russia. The genetic characterization of seventy-seven avian viruses and three viruses from seals showed that they belonged to the 2.3.4.4b clade and represented four distinct reassortant genotypes. The majority of viruses represented genotype BB, which was widespread in Europe in 2023. Viruses from seals and four viruses from birds, isolated from outbreaks in the Far East region, belonged to the G1 (A3) genotype and had the amino acid substitution N319K in the NP protein, previously associated with an increased virulence for mammals. In addition, one virus of the G10 genotype and two viruses, representing a previously undescribed genotype (designated as Ru-23-G4) were identified. The viruses analyzed showed normal inhibition by neuraminidase inhibitors. Seven viruses had genetic markers of amantadine resistance. All the influenza A(H5N1) viruses studied showed a binding preference for α2-3-linked sialic acids, suggesting a low risk of transmission among humans. Nevertheless, monitoring of reassortment and mammalian adaptation mutations is essential for the timely identification of viruses with increased pandemic potential.

Source: Scientific Reports, https://www.nature.com/articles/s41598-025-00417-4

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#Antiviral Susceptibility of #Influenza A(#H5N1) Clade 2.3.2.1c and 2.3.4.4b Viruses from #Humans, 2023-2024

Abstract

During 2023-2024, highly pathogenic avian influenza A(H5N1) viruses from clade 2.3.2.1c caused human infections in Cambodia and from clade 2.3.4.4b caused human infections in the Americas. We assessed the susceptibility of those viruses to approved and investigational antiviral drugs. Except for 2 viruses isolated from Cambodia, all viruses were susceptible to M2 ion channel-blockers in cell culture-based assays. In the neuraminidase inhibition assay, all viruses displayed susceptibility to neuraminidase inhibitor antiviral drugs oseltamivir, zanamivir, peramivir, laninamivir, and AV5080. Oseltamivir was ≈4-fold less potent at inhibiting the neuraminidase activity of clade 2.3.4.4b than clade 2.3.2.1c viruses. All viruses were susceptible to polymerase inhibitors baloxavir and tivoxavir and to polymerase basic 2 inhibitor pimodivir with 50% effective concentrations in low nanomolar ranges. Because drug-resistant viruses can emerge spontaneously or by reassortment, close monitoring of antiviral susceptibility of H5N1 viruses collected from animals and humans by using sequence-based analysis supplemented with phenotypic testing is essential.

Source: US National Library of Medicine, https://pubmed.ncbi.nlm.nih.gov/40064473/

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#Antiviral Mechanisms and #Preclinical Evaluation of #Amantadine #Analogs that Continue to Inhibit #Influenza A Viruses with M2S31N-Based Drug Resistance

Abstract

To better manage seasonal and pandemic influenza infections, new drugs are needed with enhanced activity against amantadine- and rimantadine-resistant influenza A virus (IAV) strains containing the S31N variant of the viral M2 ion channel (M2S31N). Here we tested 36 amantadine analogs against a panel of viruses containing either M2S31N or the parental, M2 S31 wild-type variant (M2WT). We found that several analogs, primarily those with sizeable lipophilic adducts, inhibited up to three M2S31N-containing viruses with activities at least 5-fold lower than rimantadine, without inhibiting M2S31N proton currents or modulating endosomal pH. While M2WT viruses in passaging studies rapidly gained resistance to these analogs through the established M2 mutations V27A and/or A30T, resistance development was markedly slower for M2S31N viruses and did not associate with additional M2 mutations. Instead, a subset of analogs, exemplified by 2-propyl-2-adamantanamine (38), but not 2-(1-adamantyl)piperidine (26), spiro[adamantane-2,2’-pyrrolidine] (49), or spiro[adamantane-2,2’-piperidine] (60), inhibited cellular entry of infectious IAV following pre-treatment and/or H1N1 pseudovirus entry. Conversely, an overlapping subset of the most lipophilic analogs including compounds 26, 49, 60, and others, disrupted viral M2-M1 protein colocalization required for intracellular viral assembly and budding. Finally, a pilot toxicity study in mice demonstrated that 38 and 49 were tolerated at 30 mg/kg. Together, these results indicate that amantadine analogs act on multiple, complementary mechanisms to inhibit replication of M2S31N viruses.

Source: Antiviral Research, https://www.sciencedirect.com/science/article/abs/pii/S0166354225000300?via%3Dihub

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#Combination #chemotherapy, a potential #strategy for reducing the emergence of #drug-resistant #influenza A variants

Natalia A. Ilyushina {a b}, Nicolai V. Bovin {c}, Robert G. Webster {a d}, Elena A. Govorkova  {a b}

a} Department of Infectious Diseases, St. Jude Children's Research Hospital, 332 N. Lauderdale, Memphis, TN 38105-2794, USA; b} The D.I. Ivanovsky Institute of Virology, Gamaleya 16, Moscow 123098, Russia; c} Shemyakin Institute of Bioorganic Chemistry, 16/10 Miklukho-Maklaya, Moscow 117997, Russia; d} Department of Pathology, University of Tennessee, Memphis, TN 38105, USA

Received 21 November 2005, Accepted 26 January 2006, Available online 21 February 2006.

Abstract

Rapid development of resistant influenza variants after amantadine treatment is one of the main drawbacks of M2 blockers. On the other hand, the emergence of variants with low susceptibility to the neuraminidase (NA) inhibitors is limited. In the present study we examined whether combination therapy with two classes of anti-influenza drugs can affect the emergence of resistant variants in vitro. We observed that virus yields of human A/Nanchang/1/99 (H1N1), A/Panama/2007/99 (H3N2), and A/Hong Kong/156/97 (H5N1) viruses in MDCK cells were significantly reduced (P < 0.005) when the cells were treated with the combination of amantadine and low doses of oseltamivir carboxylate (≤1 μM). After five sequential passages in MDCK cells, the M2 protein of viruses cultivated with amantadine alone mutated at positions V27A and S31N/I. Viruses cultivated with oseltamivir carboxylate (≥0.001 μM) possessed mutations in the hemagglutinin (HA) protein. These variants showed reduced efficiency of binding to sialic acid receptors and decreased sensitivity to NA inhibitor in plaque reduction assay. Importantly, no mutations in the HA, NA, and M2 proteins were detected when the drugs were used in combination. Our results suggest that combination chemotherapy with M2 blocker and NA inhibitor reduced the emergence of drug-resistant influenza variants in vitro. This strategy could be an option for the control of influenza virus infection, and combinations with other novel drugs should be explored.

Source: Antiviral Research, https://www.sciencedirect.com/science/article/abs/pii/S0166354206000349

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#Critical #Illness in an #Adolescent with #Influenza A(#H5N1) Virus #Infection

To the Editor:

Highly pathogenic avian influenza A(H5N1) viruses are circulating among wild birds and poultry in British Columbia, Canada.1 These viruses are also recognized to cause illness in humans. Here, we report a case of critical illness caused by influenza A(H5N1) virus infection in British Columbia.

On November 4, 2024, a 13-year-old girl with a history of mild asthma and an elevated body-mass index (the weight in kilograms divided by the square of the height in meters) of greater than 35 presented to an emergency department in British Columbia with a 2-day history of conjunctivitis in both eyes and a 1-day history of fever. She was discharged home without treatment, but cough, vomiting, and diarrhea then developed, and she returned to the emergency department on November 7 in respiratory distress with hemodynamic instability. On November 8, she was transferred, while receiving bilevel positive airway pressure, to the pediatric intensive care unit at British Columbia Children’s Hospital with respiratory failure, pneumonia in the left lower lobe, acute kidney injury, thrombocytopenia, and leukopenia (...). A nasopharyngeal swab obtained at admission was positive for influenza A but negative for A(H1) and A(H3) by the BioFire Respiratory Panel 2.1 assay (BioFire Diagnostics). Reflex testing of the specimen with the Xpert Xpress CoV-2/Flu/RSV plus assay (Cepheid) revealed an influenza A cycle threshold (Ct) value of 27.1. This finding indicates a relatively high viral load for which subtyping would be expected; the lack of subtype identification suggested infection with a novel influenza A virus. Oseltamivir treatment was started on November 8 (Table S2), and the use of eye protection, N95 respirators, and other precautions against droplet, contact, and airborne transmission were implemented.

A reverse-transcriptase–polymerase-chain-reaction (RT-PCR) test specific for influenza A(H5)2 was positive on the day of admission. The patient had signs of respiratory deterioration — chest radiographs were consistent with progression to acute respiratory distress syndrome (...) — which prompted tracheal intubation and initiation of venovenous extracorporeal membrane oxygenation (ECMO) on November 9. Continuous renal replacement therapy was initiated on November 10. Combination antiviral treatment with amantadine (initiated on November 9) and baloxavir (initiated on November 11) was added to ongoing treatment with oseltamivir. Bacterial cultures of blood (samples obtained at admission) and endotracheal aspirate (obtained after intubation) yielded no growth.

Because of concern for cytokine-mediated hemodynamic instability, plasma exchange was performed daily from November 14 through November 16. Serial influenza A–specific RT-PCR tests showed increasing Ct values, which suggested a decline in the viral RNA load in serum and a decline in viral RNA in upper- and lower-respiratory specimens shortly after the initiation of antiviral treatment, with the first negative RT-PCR result for serum obtained on November 16 (...). It is notable that lower-respiratory specimens consistently yielded lower Ct values than upper-respiratory specimens, a finding that suggested higher viral levels in the lower-respiratory tract (...).

Influenza A(H5N1) virus was cultured from respiratory specimens obtained between November 8 and November 12 but not from subsequent respiratory specimens or from any serum specimens (...). No evidence of reduced susceptibility to any of the three antiviral agents used in treatment was observed in serial respiratory specimens by either genomic analysis or phenotypic testing with the NA-Star influenza neuraminidase inhibitor resistance detection kit (ThermoFisher Scientific) (...). The patient’s respiratory status improved, ECMO was discontinued on November 22, and the patient’s trachea was extubated on November 28.

The viral genome sequence obtained from a tracheal-aspirate specimen collected on November 9 (8 days after the onset of symptoms) was reconstructed as described previously.3 The virus was typed as clade 2.3.4.4b, genotype D1.1,4 most closely related to viruses detected in wild birds in British Columbia around the same time (...). Markers of adaptation to humans were detected in the tracheal-aspirate specimen collected on November 9: the E627K mutation was detected (52% allele frequency) in the polymerase basic 2 (PB2) gene product, and analysis of the H5 hemagglutinin (HA) gene yielded ambiguous calls in the codons for amino acid residues E186 (E190 according to H3 mature HA numbering) — 28% allele frequency for E186D — and Q222 (Q226 according to H3 mature HA numbering) — 35% allele frequency for Q222H. The mutations in the H5 HA gene have previously been shown to increase binding to α2-6–linked sialic acids, which act as receptors that facilitate viral entry into cells in the human respiratory tract and enable viral replication.5

Highly pathogenic avian influenza A(H5N1) virus infection acquired in North America can cause severe human illness. Evidence for changes to HA that may increase binding to human airway receptors is worrisome.

Agatha N. Jassem, Ph.D., British Columbia Centre for Disease Control, Vancouver, BC, Canada; Ashley Roberts, M.D., British Columbia Children’s Hospital, Vancouver, BC, Canada; John Tyson, Ph.D., James E.A. Zlosnik, Ph.D., Shannon L. Russell, Ph.D., British Columbia Centre for Disease Control, Vancouver, BC, Canada; Jessica M. Caleta, M.Sc., Public Health Agency of Canada, Winnipeg, MB, Canada; Eric J. Eckbo, M.D., British Columbia Centre for Disease Control, Vancouver, BC, Canada; Ruimin Gao, Ph.D., Taeyo Chestley, Ph.D., Public Health Agency of Canada, Winnipeg, MB, Canada; Jennifer Grant, M.D., British Columbia Centre for Disease Control, Vancouver, BC, Canada; Timothy M. Uyeki, M.D., M.P.H., Centers for Disease Control and Prevention, Atlanta, GA; Natalie A. Prystajecky, Ph.D., British Columbia Centre for Disease Control, Vancouver, BC, Canada; Chelsea G. Himsworth, D.V.M., Ph.D., British Columbia Ministry of Agriculture and Food, Abbotsford, BC, Canada; Elspeth MacBain, M.D., British Columbia Children’s Hospital, Vancouver, BC, Canada; Charlene Ranadheera, Ph.D., Public Health Agency of Canada, Winnipeg, MB, Canada; Lynne Li, M.D., British Columbia Children’s Hospital, Vancouver, BC, Canada; Linda M.N. Hoang, M.D., British Columbia Centre for Disease Control, Vancouver, BC, Canada; Nathalie Bastien, Ph.D., Public Health Agency of Canada, Winnipeg, MB, Canada; David M. Goldfarb, M.D., British Columbia Children’s Hospital, Vancouver, BC, Canada.

Source: New England Journal of Medicine, https://www.nejm.org/doi/full/10.1056/NEJMc2415890

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