A Broad and Potent H1-Specific Human Monoclonal Antibody Produced in Plants Prevents Influenza Virus Infection and Transmission in Guinea Pigs

Although seasonal influenza vaccines block most predominant influenza types and subtypes, humans still remain vulnerable to waves of seasonal and new potential pandemic influenza viruses for which no immunity may exist because of viral antigenic drift and/or shift. Previously, we described a human monoclonal antibody (hMAb), KPF1, which was produced in human embryonic kidney 293T cells (KPF1-HEK) with broad and potent neutralizing activity against H1N1 influenza A viruses (IAV) in vitro, and prophylactic and therapeutic activities in vivo. In this study, we produced hMAb KPF1 in tobacco plants (KPF1-Antx) and demonstrated how the plant-produced KPF1-Antx hMAb possesses similar biological activity compared with the mammalian-produced KPF1-HEK hMAb. KPF1-Antx hMAb showed broad binding to recombinant HA proteins and H1N1 IAV, including A/California/04/2009 (pH1N1) in vitro, which was comparable to that observed with KPF1-HEK hMAb. Importantly, prophylactic administration of KPF1-Antx hMAb to guinea pigs prevented pH1N1 infection and transmission in both prophylactic and therapeutic experiments, substantiating its clinical potential to prevent and treat H1N1 infections. Collectively, this study demonstrated, for the first time, a plant-produced influenza hMAb with in vitro and in vivo activity against influenza virus. Because of the many advantages of plant-produced hMAbs, such as rapid batch production, low cost, and the absence of mammalian cell products, they represent an alternative strategy for the production of immunotherapeutics for the treatment of influenza viral infections, including emerging seasonal and/or pandemic strains.

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A broad and potent H1-specific human monoclonal antibody produced in plants prevents influenza virus infection and transmission in guinea pigs

10.1101/2020.01.13.902841 ◽

2020 ◽

Author(s):

Jun-Gyu Park ◽

Chengjin Ye ◽

Michael S. Piepenbrink ◽

Aitor Nogales ◽

Haifeng Wang ◽

...

Keyword(s):

Monoclonal Antibody ◽

Guinea Pigs ◽

Mammalian Cells ◽

Influenza A ◽

Human Monoclonal Antibody ◽

Influenza Virus Infection ◽

Influenza Viruses ◽

Antigenic Drift

AbstractAlthough seasonal influenza vaccines block most predominant influenza types and subtypes, humans still remain vulnerable to waves of seasonal and new potential pandemic influenza viruses for which no immunity may exist because of viral antigenic drift and/or shift, respectively. Previously, we have described a human monoclonal antibody (hMAb), KPF1, which was produced in human embryonic kidney 293T cells (KPF1-HEK) with broad and potent neutralizing activity against H1N1 influenza A viruses (IAV) in vitro, and prophylactic and therapeutic activities in vivo. In this study, we produced hMAb KPF1 in tobacco plants (KPF1-Antx) and demonstrate how the plant-produced KPF1-Antx hMAb possesses similar biological activity compared with the mammalian produced KPF1-HEK hMAb. KPF1-Antx hMAb shows broad binding to recombinant HA proteins and H1N1 IAV, including A/California/04/2009 (pH1N1) in vitro, that are comparable to those observed with KPF1-HEK hMAb. Importantly, prophylactic administration of KPF1-Antx hMAb to guinea pigs prevented pH1N1 infection and transmission in both prophylactic and therapeutic experiments, substantiating its clinical potential to prevent and treat H1N1 infections. Collectively, this study demonstrates, for the first time, that plant-produced influenza hMAbs have similar in vitro and in vivo biological properties to those produced in mammalian cells. Because of the many advantages of plant-produced hMAbs, such as rapid batch production, low cost, and the absence of mammalian cell products, they represent an alternative strategy for the production of immunotherapeutics for the treatment of influenza viral infections, including emerging seasonal and/or pandemic strains.

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Heterosubtypic Protection Conferred by the Human Monoclonal Antibody PN-SIA28 against Influenza A Virus Lethal Infections in Mice

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00118-15 ◽

2015 ◽

Vol 59 (5) ◽

pp. 2647-2653 ◽

Cited By ~ 1

Author(s):

Miguel Retamal ◽

Yacine Abed ◽

Chantal Rhéaume ◽

Francesca Cappelletti ◽

Nicola Clementi ◽

...

Keyword(s):

Monoclonal Antibody ◽

Body Weight ◽

Influenza Virus ◽

Influenza A ◽

Human Monoclonal Antibody ◽

Influenza A Viruses ◽

Virus Challenge ◽

Influenza A H1n1

ABSTRACTPN-SIA28 is a human monoclonal antibody (Hu-MAb) targeting highly conserved epitopes within the stem portion of the influenza virus hemagglutinin (HA) (N. Clementi, et al, PLoS One 6:e28001, 2011,http://dx.doi.org/10.1371/journal.pone.0028001). Previousin vitrostudies demonstrated PN-SIA28 neutralizing activities against phylogenetically divergent influenza A subtypes. In this study, the protective activity of PN-SIA28 was evaluated in mice inoculated with lethal influenza A/WSN/33 (H1N1), A/Quebec/144147/09 (H1N1)pdm09, and A/Victoria/3/75 (H3N2) viruses. At 24 h postinoculation (p.i.), animals received PN-SIA28 intraperitoneally (1 or 10 mg/kg of body weight) or 10 mg/kg of unrelated Hu-MAb (mock). Body weight loss and mortality rate (MR) were recorded for 14 days postinfection (p.i.). Lung viral titers (LVT) were determined at day 5 p.i. In A/WSN/33 (H1N1)-infected groups, all untreated and mock-receiving mice died, whereas MRs of 87.5% and 25% were observed in mice that received PN-SIA28 1 and 10 mg/kg, respectively. In influenza A(H1N1) pdm09-infected groups, an MR of 75% was recorded for untreated and mock-treated groups, whereas the PN-SIA28 1-mg/kg and 10-mg/kg groups had rates of 62.5% and 0%, respectively. In A/Victoria/3/75 (H3N2)-infected animals, untreated and mock-treated animals had MRs of 37.5% and 25%, respectively, and no mortalities were recorded after PN-SIA28 treatments. Accordingly, PN-SIA28 treatments significantly reduced weight losses and resulted in a ≥1-log reduction in LVT compared to the control in all infection groups. This study confirms that antibodies targeting highly conserved epitopes in the influenza HA stem region, like PN-SIA28, not only neutralize influenza A viruses of clinically relevant subtypesin vitrobut also, more importantly, protect from a lethal influenza virus challengein vivo.

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Hemagglutinin stalk-based monoclonal antibody elicits broadly reactivity against group 1 influenza A virus

Virology Journal ◽

10.1186/s12985-020-01458-z ◽

2020 ◽

Vol 17 (1) ◽

Author(s):

Jingjin Huang ◽

Nan Huang ◽

Menglu Fan ◽

Lingcai Zhao ◽

Yan Luo ◽

...

Keyword(s):

Monoclonal Antibody ◽

Broad Spectrum ◽

Influenza A ◽

Lethal Dose ◽

Influenza Viruses ◽

Antigenic Drift ◽

Linear Epitope ◽

Group 1

Abstract Background Influenza virus remains a continuous and severe threat to public health worldwide, and its prevention and treatment have always been a major international issue. Because of its ability to evade immune surveillance through rapid antigenic drift and antigenic shift, broad-spectrum vaccines seem increasingly important. Methods A mAb named 3C12 from an immortalized hybrid cell was generated via immunizing mice with HA2 protein from A/chicken/Anhui/BRI99/2016 (AH/BRI99/16, H9N2) generated by prokaryotic expression. Then, its broad-spectrum activity was analyzed by WB and IFA. Next, the minimal linear epitope was identified via analyzing the reaction of a series of HA truncations with 3C12. Finally, the protective effects of 3C12 were evaluated in vitro and in vivo infection experiments. Results The mAb could react with the viruses of subtypes H1, H2, H5, H8, H9, H12, H13, H16, and HA protein of H18 in group 1, but failed to react with viruses in group 2. The minimal linear epitope targeted by the mAb was 433NAELLVL439 in full length of HA and localized in the C-helix region of HA2 (residue 95-101, HA2 numbering). What’s more, the mAb 3C12 inhibited H1, H2, H5, H8, H9, H12, H13 and H16 virus-replication in vitro and also has shown effectiveness in preventing and treating disease in mice challenged with lethal dose of AH/BRI99/16 (H9N2) virus in vivo. These results suggested that the broadly reactive anti-HA stem mAb 3C12 exhibited prophylactic and therapeutic efficacy. Conclusions Here, we have demonstrated that the linear epitope identified in this study could be a novel target for developing broad-spectrum influenza diagnostics or vaccine design, and the HA2-based monoclonal antibody is indeed a promising strategy for broad-spectrum protection against seasonal and pandemic influenza viruses.

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Ethanol Extract of Caesalpinia decapetala Inhibits Influenza Virus Infection In Vitro and In Vivo

Viruses ◽

10.3390/v12050557 ◽

2020 ◽

Vol 12 (5) ◽

pp. 557 ◽

Cited By ~ 1

Author(s):

Li Zhang ◽

Jungang Chen ◽

Chang Ke ◽

Haiwei Zhang ◽

Shoujun Zhang ◽

...

Keyword(s):

Influenza Virus ◽

Influenza A ◽

Antiviral Agents ◽

Influenza Virus Infection ◽

Virus Titer ◽

Ethanol Extract ◽

Influenza Viruses ◽

Virus Infections

Influenza virus infections can lead to viral pneumonia and acute respiratory distress syndrome in severe cases, causing significant morbidity and mortality and posing a great threat to human health. Because of the diversity of influenza virus strains and drug resistance to the current direct antiviral agents, there have been no effective drugs as yet to cure all patients infected by influenza viruses. Natural products from plants contain compounds with diverse structures that have the potential to interact with multiple host and virus factors. In this study, we identified the ethanol extract of Caesalpinia decapetala (Roth) Alston (EEC) as an inhibitor against the replication of a panel of influenza A and B viruses both on human pulmonary epithelial A549 and human monocytic U937 cells. The animal study revealed that EEC administration reduces the weight loss and improves the survival rate of mice infected with lethal influenza virus. Also, EEC treatment attenuated lung injury and reduced virus titer significantly. In conclusion, we showed that EEC has antiviral activity both in vitro and in vivo, suggesting that the plant C. decapetala has the potential to be further developed as a resource of new anti-influenza drugs.

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Cross-Protection of Chicken Immunoglobulin Y Antibodies against H5N1 and H1N1 Viruses Passively Administered in Mice

Clinical and Vaccine Immunology ◽

10.1128/cvi.05075-11 ◽

2011 ◽

Vol 18 (7) ◽

pp. 1083-1090 ◽

Cited By ~ 19

Author(s):

Michael G. Wallach ◽

Richard J. Webby ◽

Fakhrul Islam ◽

Stephen Walkden-Brown ◽

Eva Emmoth ◽

...

Keyword(s):

Influenza Virus ◽

Influenza Pandemic ◽

Influenza Virus Infection ◽

Influenza Viruses ◽

Antigenic Drift ◽

Virus Infectivity ◽

Lethal Challenge ◽

H5n1 Influenza

ABSTRACTInfluenza viruses remain a major threat to global health due to their ability to undergo change through antigenic drift and antigenic shift. We postulated that avian IgY antibodies represent a low-cost, effective, and well-tolerated approach that can easily be scaled up to produce enormous quantities of protective antibodies. These IgY antibodies can be administered passively in humans (orally and intranasally) and can be used quickly and safely to help in the fight against an influenza pandemic. In this study, we raised IgY antibodies against H1N1, H3N2, and H5N1 influenza viruses. We demonstrated that, using whole inactivated viruses alone and in combination to immunize hens, we were able to induce a high level of anti-influenza virus IgY in the sera and eggs, which lasted for at least 2 months after two immunizations. Furthermore, we found that by use ofin vitroassays to test for the ability of IgY to inhibit hemagglutination (HI test) and virus infectivity (serum neutralization test), IgYs inhibited the homologous as well as in some cases heterologous clades and strains of viruses. Using anin vivomouse model system, we found that, when administered intranasally 1 h prior to infection, IgY to H5N1 protected 100% of the mice against lethal challenge with H5N1. Of particular interest was the finding that IgY to H5N1 cross-protected against A/Puerto Rico/8/34 (H1N1) bothin vitroandin vivo. Based on our results, we conclude that anti-influenza virus IgY can be used to help prevent influenza virus infection.

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Hemagglutinin Stalk-based Monoclonal Antibody Elicits Broadly Reactivity Against Group 1 Influenza A Virus

10.21203/rs.3.rs-97761/v1 ◽

2020 ◽

Author(s):

Jingjin Huang ◽

Nan Huang ◽

Menglu Fan ◽

Lingcai Zhao ◽

Yan Luo ◽

...

Keyword(s):

Monoclonal Antibody ◽

Broad Spectrum ◽

Influenza A ◽

Lethal Dose ◽

Influenza Viruses ◽

Antigenic Drift ◽

Linear Epitope ◽

Group 1

Abstract Background: Influenza virus remains a continuous and serious threat to public health worldwide, and its prevention and treatment have always been a major international issue. Because of its ability to evade immune surveillance through rapid antigenic drift and antigenic shift, broad-spectrum vaccines seem increasingly important. Methods: A mAb named 3C12 from an immortalized hybrid cell was generated via immunizing mice with HA2 protein from A/chicken/Anhui/BRI99/2016 (AH/BRI99/16, H9N2) generated by prokaryotic expression. Then, its broad-spectrum activity was analyzed by WB and IFA. Next, the minimal linear epitope was identified via analyzing the reaction of a series of HA truncations with 3C12. Finally, the protective effects of 3C12 were evaluated in vitro and in vivo infection experiments.Results: The mAb could react with the viruses of subtypes H1, H2, H5, H8, H9, H12, H13, H16, and HA protein of H18 in group 1, but failed to react with viruses in group 2. The minimal linear epitope targeted by the mAb was 433NAELLVL439 in full length of HA and localized in the C-helix region of HA2 (residue 95-101, HA2 numbering). What’s more, the mAb 3C12 inhibited H1, H2, H5, H8, H9, H12, H13 and H16 virus-replication in vitro and also has shown effectiveness in preventing and treating disease in mice challenged with lethal dose of AH/BRI99/16 (H9N2) virus in vivo. These results suggested that the broadly reactive anti-HA stem mAb 3C12 exhibited prophylactic and therapeutic efficacy.Conclusions: Here, we have demonstrated that the linear epitope identified in this study could be a novel target for developing broad-spectrum influenza diagnostics or vaccine design, and the HA2-based monoclonal antibody is indeed a promising strategy for broad-spectrum protection against seasonal and pandemic influenza viruses.

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Human Monoclonal Antibody Derived from Transchromosomic Cattle Neutralizes Multiple H1 Clades of Influenza A Virus by Recognizing a Novel Conformational Epitope in the Hemagglutinin Head Domain

Journal of Virology ◽

10.1128/jvi.00945-20 ◽

2020 ◽

Vol 94 (22) ◽

Author(s):

Rongyuan Gao ◽

Chithra C. Sreenivasan ◽

Zizhang Sheng ◽

Ben M. Hause ◽

Bin Zhou ◽

...

Keyword(s):

Monoclonal Antibody ◽

Influenza Virus ◽

Neutralizing Antibodies ◽

Influenza A ◽

Human Monoclonal Antibody ◽

Influenza Virus Infection ◽

Influenza Viruses ◽

Head Domain ◽

Potent Neutralization

ABSTRACT Influenza remains a global health risk and challenge. Currently, neuraminidase (NA) inhibitors are extensively used to treat influenza, but their efficacy is compromised by the emergence of drug-resistant variants. Neutralizing antibodies targeting influenza A virus surface glycoproteins are critical components of influenza therapeutic agents and may provide alternative strategies to the existing countermeasures. However, the major hurdle for the extensive application of antibody therapies lies in the difficulty of generating nonimmunogenic antibodies in large quantities rapidly. Here, we report that one human monoclonal antibody (MAb), 53C10, isolated from transchromosomic (Tc) cattle exhibits potent neutralization and hemagglutination inhibition titers against different clades of H1N1 subtype influenza A viruses. In vitro selection of antibody escape mutants revealed that 53C10 recognizes a novel noncontinuous epitope in the hemagglutinin (HA) head domain involving three amino acid residues, glycine (G), serine (S), and glutamic acid (E) at positions 172, 207, and 212, respectively. The results of our experiments supported a critical role for substitution of arginine at position 207 (S207R) in mediating resistance to 53C10, while substitutions at either G172E or E212A did not alter antibody recognition and neutralization. The E212A mutation may provide structural stability for the epitope, while the substitution G172E probably compensates for loss of fitness introduced by S207R. Our results offer novel insights into the mechanism of action of MAb 53C10 and indicate its potential role in therapeutic treatment of H1 influenza virus infection in humans. IMPORTANCE Respiratory diseases caused by influenza viruses still pose a serious concern to global health, and neutralizing antibodies constitute a promising area of antiviral therapeutics. However, the potential application of antibodies is often hampered by the challenge in generating nonimmunogenic antibodies in large scale. In the present study, transchromosomic (Tc) cattle were used for the generation of nonimmunogenic monoclonal antibodies (MAbs), and characterization of such MAbs revealed one monoclonal antibody, 53C10, exhibiting a potent neutralization activity against H1N1 influenza viruses. Further characterization of the neutralization escape mutant generated using this MAb showed that three amino acid substitutions in the HA head domain contributed to the resistance. These findings emphasize the importance of Tc cattle in the production of nonimmunogenic MAbs and highlight the potential of MAb 53C10 in the therapeutic application against H1 influenza virus infection in humans.

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Δ12-Prostaglandin J2 Is a Potent Inhibitor of Influenza A Virus Replication

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.44.1.200-204.2000 ◽

2000 ◽

Vol 44 (1) ◽

pp. 200-204 ◽

Cited By ~ 21

Author(s):

Francesca Pica ◽

Anna Teresa Palamara ◽

Antonio Rossi ◽

Alessandra De Marco ◽

Carla Amici ◽

...

Keyword(s):

Influenza Virus ◽

Heat Shock ◽

Influenza A Virus ◽

Influenza A ◽

Potent Inhibitor ◽

Influenza Virus Infection ◽

H1n1 Infection ◽

Shock Proteins

ABSTRACT 9-Deoxy-Δ9,Δ12-13,14-dihydro-prostaglandin D2 (Δ12-PGJ2), a natural cyclopentenone metabolite of prostaglandin D2, is shown to possess therapeutic efficacy against influenza A virus A/PR8/34 (H1N1) infection in vitro and in vivo. The results indicate that the antiviral activity is associated with induction of cytoprotective heat shock proteins and suggest novel strategies for treatment of influenza virus infection.

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Influenza A Virus Vaccination: Immunity, Protection, and Recent Advances Toward A Universal Vaccine

Vaccines ◽

10.3390/vaccines8030434 ◽

2020 ◽

Vol 8 (3) ◽

pp. 434 ◽

Cited By ~ 2

Author(s):

Christopher E. Lopez ◽

Kevin L. Legge

Keyword(s):

Immune Response ◽

Influenza Virus ◽

Virus Infection ◽

Influenza A ◽

Influenza Virus Infection ◽

Influenza Viruses ◽

Antigenic Drift ◽

Influenza Vaccines ◽

Virus Infections ◽

Universal Vaccine

Influenza virus infections represent a serious public health threat and account for significant morbidity and mortality worldwide due to seasonal epidemics and periodic pandemics. Despite being an important countermeasure to combat influenza virus and being highly efficacious when matched to circulating influenza viruses, current preventative strategies of vaccination against influenza virus often provide incomplete protection due the continuous antigenic drift/shift of circulating strains of influenza virus. Prevention and control of influenza virus infection with vaccines is dependent on the host immune response induced by vaccination and the various vaccine platforms induce different components of the local and systemic immune response. This review focuses on the immune basis of current (inactivated influenza vaccines (IIV) and live attenuated influenza vaccines (LAIV)) as well as novel vaccine platforms against influenza virus. Particular emphasis will be placed on how each platform induces cross-protection against heterologous influenza viruses, as well as how this immunity compares to and contrasts from the “gold standard” of immunity generated by natural influenza virus infection.

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Novel Ranking System for Identifying Efficacious Anti-Influenza Virus PB2 Inhibitors

Antimicrobial Agents and Chemotherapy ◽

10.1128/aac.00781-15 ◽

2015 ◽

Vol 59 (10) ◽

pp. 6007-6016 ◽

Cited By ~ 8

Author(s):

Alice W. Tsai ◽

Colleen F. McNeil ◽

Joshua R. Leeman ◽

Hamilton B. Bennett ◽

Kwame Nti-Addae ◽

...

Keyword(s):

Influenza Virus ◽

Influenza A Virus ◽

Influenza A ◽

Therapeutic Agents ◽

Influenza Viruses ◽

Antigenic Drift ◽

Whole Body ◽

Infection Model ◽

Virus Infections

ABSTRACTThrough antigenic drift and shifts, influenza virus infections continue to be an annual cause of morbidity in healthy populations and of death among elderly and at-risk patients. The emergence of highly pathogenic avian influenza viruses such as H5N1 and H7N9 and the rapid spread of the swine-origin H1N1 influenza virus in 2009 demonstrate the continued need for effective therapeutic agents for influenza. While several neuraminidase inhibitors have been developed for the treatment of influenza virus infections, these have shown a limited window for treatment initiation, and resistant variants have been noted in the population. In addition, an older class of antiviral drugs for influenza, the adamantanes, are no longer recommended for treatment due to widespread resistance. There remains a need for new influenza therapeutic agents with improved efficacy as well as an expanded window for the initiation of treatment. Azaindole compounds targeting the influenza A virus PB2 protein and demonstrating excellentin vitroandin vivoproperties have been identified. To evaluate thein vivoefficacy of these PB2 inhibitors, we utilized a mouse influenza A virus infection model. In addition to traditional endpoints, i.e., death, morbidity, and body weight loss, we measured lung function using whole-body plethysmography, and we used these data to develop a composite efficacy score that takes compound exposure into account. This model allowed the rapid identification and ranking of molecules relative to each other and to oseltamivir. The ability to identify compounds with enhanced preclinical properties provides an opportunity to develop more-effective treatments for influenza in patients.

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