Broadly Reactive Human Monoclonal Antibodies Elicited following Pandemic H1N1 Influenza Virus Exposure Protect Mice against Highly Pathogenic H5N1 Challenge

ABSTRACT Broadly cross-reactive antibodies (Abs) that recognize conserved epitopes within the influenza virus hemagglutinin (HA) stalk domain are of particular interest for their potential use as therapeutic and prophylactic agents against multiple influenza virus subtypes, including zoonotic virus strains. Here, we characterized four human HA stalk-reactive monoclonal antibodies (MAbs) for their binding breadth and affinity, in vitro neutralization capacity, and in vivo protective potential against an highly pathogenic avian influenza virus. The monoclonal antibodies were isolated from individuals shortly following infection with (70-1F02 and 1009-3B05) or vaccination against (05-2G02 and 09-3A01) A(H1N1)pdm09. Three of the MAbs bound HAs from multiple strains of group 1 viruses, and one MAb, 05-2G02, bound to both group 1 and group 2 influenza A virus HAs. All four antibodies prophylactically protected mice against a lethal challenge with the highly pathogenic A/Vietnam/1203/04 (H5N1) strain. Two MAbs, 70-1F02 and 09-3A01, were further tested for their therapeutic efficacy against the same strain and showed good efficacy in this setting as well. One MAb, 70-1F02, cocrystallized with H5 HA and showed heavy-chain-only interactions similar to those seen with the previously described CR6261 anti-stalk antibody. Finally, we show that antibodies that compete with these MAbs are prevalent in serum from an individual recently infected with the A(H1N1)pdm09 virus. The antibodies described here can be developed into broad-spectrum antiviral therapeutics that could be used to combat infections by zoonotic or emerging pandemic influenza viruses. IMPORTANCE The rise in zoonotic infections of humans by emerging influenza viruses is a worldwide public health concern. The majority of recent zoonotic human influenza cases were caused by H7N9 and H5Nx viruses and were associated with high morbidity and mortality. In addition, seasonal influenza viruses are estimated to cause up to 650,000 deaths annually worldwide. Currently available antiviral treatment options include only neuraminidase inhibitors, but some influenza viruses are naturally resistant to these drugs, and others quickly develop resistance-conferring mutations. Alternative therapeutics are urgently needed. Broadly protective antibodies that target the conserved “stalk” domain of the hemagglutinin represent potential potent antiviral prophylactic and therapeutic agents that can assist pandemic preparedness. Here, we describe four human monoclonal antibodies that target conserved regions of influenza HA and characterize their binding spectrum as well as their protective capacity in prophylactic and therapeutic settings against a lethal challenge with a zoonotic influenza virus.

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Human monoclonal antibodies derived from a patient infected with 2009 pandemic influenza A virus broadly cross-neutralize group 1 influenza viruses

Biochemical and Biophysical Research Communications ◽

10.1016/j.bbrc.2014.05.060 ◽

2014 ◽

Vol 450 (1) ◽

pp. 42-48 ◽

Cited By ~ 6

Author(s):

Yang Pan ◽

Tadahiro Sasaki ◽

Ritsuko Kubota-Koketsu ◽

Yuji Inoue ◽

Mayo Yasugi ◽

...

Keyword(s):

Monoclonal Antibodies ◽

Influenza A Virus ◽

Pandemic Influenza ◽

Influenza A ◽

Influenza Viruses ◽

Human Monoclonal Antibodies ◽

Group 1

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Broadly inhibiting anti-neuraminidase monoclonal antibodies induced by trivalent influenza vaccine and H7N9 infection in humans

10.1101/682450 ◽

2019 ◽

Cited By ~ 1

Author(s):

Pramila Rijal ◽

Bei Bei Wang ◽

Tiong Kit Tan ◽

Lisa Schimanski ◽

Philipp Janesch ◽

...

Keyword(s):

Monoclonal Antibodies ◽

Influenza Vaccine ◽

Influenza Viruses ◽

List Type ◽

Human Monoclonal Antibodies ◽

Human Antibodies ◽

Trivalent Influenza Vaccine ◽

H7n9 Infection ◽

Group 2 ◽

Group 1

AbstractThe majority of antibodies induced by influenza neuraminidase (NA), like those against hemagglutinin (HA), are relatively specific to viruses isolated within a limited time-window as seen in serological studies and the analysis of many murine monoclonal antibodies. We report three broadly reactive human monoclonal antibodies (mAbs) targeting N1 NA. Two were isolated from a young adult vaccinated with trivalent influenza vaccine (TIV), which inhibited N1 NA from viruses isolated from human over a period of a hundred years. The third antibody isolated from a child with acute mild H7N9 infection inhibited both group 1 N1 and group 2 N9 NAs. In addition, the antibodies cross-inhibited the N1 NAs of highly pathogenic avian H5N1 influenza viruses. These antibodies are protective in prophylaxis against seasonal H1N1 viruses in mice. This study demonstrates that human antibodies to N1 NA with exceptional cross-reactivity can be recalled by vaccination and highlights the importance of standardizing the NA antigen in seasonal vaccines to offer optimal protection.ImportanceAntibodies to the influenza NA can provide protection against influenza disease. Analysis of human antibodies to NA lags behind that for HA. We show that human monoclonal antibodies against NA induced by vaccination and infection can be very broadly reactive and able to inhibit a wide spectrum of N1 NAs between 1918 and 2018. This suggests that antibodies to NA may be a useful therapy, and that efficacy of influenza vaccines could be enhanced by ensuring appropriate content of NA antigen.Highlights of the paperAntibodies that inhibit influenza viruses with N1 neuraminidase (NA), with broad reactivity for viruses isolated between 1918-2018, can be isolated from human recipients of seasonal influenza vaccineAntibodies targeting N1 NA of human seasonal H1N1 viruses can cross-react with a variety of avian N1 neuraminidasesAcute H7N9 infection can recall memory B cells to N1 NA and elicit cross-reactive antibodies to the group 1 N1 and group 2 N9 NAsAntibodies to N1 NA with this broad reactivity protect against lethal virus challenge

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Subclade 2.2.1-Specific Human Monoclonal Antibodies That Recognize an Epitope in Antigenic Site A of Influenza A(H5) Virus HA Detected between 2015 and 2018

Viruses ◽

10.3390/v11040321 ◽

2019 ◽

Vol 11 (4) ◽

pp. 321

Author(s):

Moe Okuda ◽

Seiya Yamayoshi ◽

Ryuta Uraki ◽

Mutsumi Ito ◽

Taiki Hamabata ◽

...

Keyword(s):

Amino Acids ◽

Monoclonal Antibodies ◽

Influenza A ◽

Antigenic Site ◽

Influenza Viruses ◽

Interspecies Transmission ◽

Single Amino Acid ◽

Human Monoclonal Antibodies ◽

Highly Pathogenic ◽

H5n1 Vaccine

Highly pathogenic avian H5 influenza viruses persist among poultry and wild birds throughout the world. They sometimes cause interspecies transmission between avian and mammalian hosts. H5 viruses possessing the HA of subclade 2.3.4.4, 2.3.2.1, 2.2.1, or 7.2 were detected between 2015 and 2018. To understand the neutralizing epitopes of H5-HA, we characterized 15 human monoclonal antibodies (mAbs) against the HA of H5 viruses, which were obtained from volunteers who received the H5N1 vaccine that contains a subclade 2.2.1 or 2.1.3.2 virus as an antigen. Twelve mAbs were specific for the HA of subclade 2.2.1, two mAbs were specific for the HA of subclade 2.1.3.2, and one mAb was specific for the HA of both. Of the 15 mAbs analyzed, nine, which were specific for the HA of subclade 2.2.1, and shared the VH and VL genes, possessed hemagglutination inhibition and neutralizing activities, whereas the others did not. A single amino acid substitution or insertion at positions 144–147 in antigenic site A conferred resistance against these nine mAbs to the subclade 2.2.1 viruses. The amino acids at positions 144–147 are highly conserved among subclade 2.2.1, but differ from those of other subclades. These results show that the neutralizing epitope including amino acids at positions 144–147 is targeted by human antibodies, and plays a role in the antigenic difference between subclade 2.2.1 and other subclades.

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Hemagglutinin stalk domain from H5N1 strain as a potentially universal antigen.

Acta Biochimica Polonica ◽

10.18388/abp.2014_1876 ◽

2014 ◽

Vol 61 (3) ◽

Cited By ~ 5

Author(s):

Karolina Uranowska ◽

Jolanta Tyborowska ◽

Anna Jurek ◽

Bogusław Szewczyk ◽

Beata Gromadzka

Keyword(s):

Influenza Virus ◽

Influenza A Virus ◽

Neutralizing Antibodies ◽

Influenza A ◽

Virus Antigen ◽

Cross Reactivity ◽

Influenza Viruses ◽

Health Concern ◽

Stalk Domain ◽

Major Surface

Influenza A virus infections are the major public health concern and cause significant morbidity and mortality each year worldwide. Vaccination is the main strategy of influenza epidemic prevention. However, seasonal vaccines induce strain-specific immunity and must be reformulated annually based on prediction of the strains that will circulate in the next season. Thus, it is essential to develop vaccines that would induce broad and persistent immunity to influenza viruses. Hemagglutinin is the major surface antigen of the influenza virus. Recent studies revealed the importance of HA stalk-specific antibodies in neutralization of different influenza virus strains. Therefore, it is important to design an immunogen that would focus the immune response on the HA stalk domain in order to elicit neutralizing antibodies. In the present study, we report characterization of a conserved truncated protein, potentially a universal influenza virus antigen from the H5N1 Highly Pathogenic Avian Influenza A virus strain. Our results indicate that exposure of the HA stalk domain containing conserved epitopes results in cross reactivity with different antibodies (against group 1 and 2 HAs). Additionally, we conclude that HA stalk domain contains not only conformational epitopes recognized by universal FI6 antibody, but also linear epitopes recognized by other antibodies.

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Transgenic Chicks Expressing Interferon-Inducible Transmembrane Protein 1 (IFITM1) Restrict Highly Pathogenic H5N1 Influenza Viruses

International Journal of Molecular Sciences ◽

10.3390/ijms22168456 ◽

2021 ◽

Vol 22 (16) ◽

pp. 8456

Author(s):

Mohammed A. Rohaim ◽

Mohammad Q. Al-Natour ◽

Mohammed A. Abdelsabour ◽

Rania F. El Naggar ◽

Yahia M. Madbouly ◽

...

Keyword(s):

Influenza Virus ◽

Mammalian Cells ◽

Transmembrane Protein ◽

Wide Spectrum ◽

Lethal Dose ◽

Influenza Viruses ◽

Highly Pathogenic ◽

Zoonotic Infections ◽

H5n1 Influenza ◽

Transgenic Chickens

Mammalian cells utilize a wide spectrum of pathways to antagonize the viral replication. These pathways are typically regulated by antiviral proteins and can be constitutively expressed but also exacerbated by interferon induction. A myriad of interferon-stimulated genes (ISGs) have been identified in mounting broad-spectrum antiviral responses. Members of the interferon-induced transmembrane (IFITM) family of proteins are unique among these ISGs due to their ability to prevent virus entry through the lipid bilayer into the cell. In the current study, we generated transgenic chickens that constitutively and stably expressed chicken IFITM1 (chIFITM1) using the avian sarcoma-leukosis virus (RCAS)-based gene transfer system. The challenged transgenic chicks with clinical dose 104 egg infective dose 50 (EID50) of highly pathogenic avian influenza virus (HPAIV) subtype H5N1 (clade 2.2.1.2) showed 100% protection and significant infection tolerance. Although challenged transgenic chicks displayed 60% protection against challenge with the sub-lethal dose (EID50 105), the transgenic chicks showed delayed clinical symptoms, reduced virus shedding, and reduced histopathologic alterations compared to non-transgenic challenged control chickens. These finding indicate that the sterile defense against H5N1 HPAIV offered by the stable expression of chIFITM1 is inadequate; however, the clinical outcome can be substantially ameliorated. In conclusion, chIFITM proteins can inhibit influenza virus replication that can infect various host species and could be a crucial barrier against zoonotic infections.

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Vaccine Efficacy of Inactivated, Chimeric Hemagglutinin H9/H5N2 Avian Influenza Virus and Its Suitability for the Marker Vaccine Strategy

Journal of Virology ◽

10.1128/jvi.01693-16 ◽

2017 ◽

Vol 91 (6) ◽

Cited By ~ 4

Author(s):

Se Mi Kim ◽

Young-Il Kim ◽

Su-Jin Park ◽

Eun-Ha Kim ◽

Hyeok-il Kwon ◽

...

Keyword(s):

Influenza Virus ◽

Avian Influenza ◽

Recombinant Vaccine ◽

Influenza Viruses ◽

Avian Influenza Viruses ◽

Lethal Challenge ◽

Virus Shedding ◽

Immune Mediated ◽

Hpai H5n1 ◽

Stalk Domain

ABSTRACT In order to produce a dually effective vaccine against H9 and H5 avian influenza viruses that aligns with the DIVA (differentiating infected from vaccinated animals) strategy, we generated a chimeric H9/H5N2 recombinant vaccine that expressed the whole HA1 region of A/CK/Korea/04163/04 (H9N2) and the HA2 region of recent highly pathogenic avian influenza (HPAI) A/MD/Korea/W452/14 (H5N8) viruses. The chimeric H9/H5N2 virus showed in vitro and in vivo growth properties and virulence that were similar to those of the low-pathogenic avian influenza (LPAI) H9 virus. An inactivated vaccine based on this chimeric virus induced serum neutralizing (SN) antibodies against both H9 and H5 viruses but induced cross-reactive hemagglutination inhibition (HI) antibody only against H9 viruses. Thus, this suggests its compatibility for use in the DIVA strategy against H5 strains. Furthermore, the chimeric H9/H5N2 recombinant vaccine protected immunized chickens against lethal challenge by HPAI H5N8 viruses and significantly attenuated virus shedding after infection by both H9N2 and HPAI H5N8 viruses. In mice, serological analyses confirmed that HA1- and HA2 stalk-specific antibody responses were induced by vaccination and that the DIVA principle could be employed through the use of an HI assay against H5 viruses. Furthermore, each HA1- and HA2 stalk-specific antibody response was sufficient to inhibit viral replication and protect the chimeric virus-immunized mice from lethal challenge with both mouse-adapted H9N2 and wild-type HPAI H5N1 viruses, although differences in vaccine efficacy against a homologous H9 virus (HA1 head domain immune-mediated protection) and a heterosubtypic H5 virus (HA2 stalk domain immune-mediated protection) were observed. Taken together, these results demonstrate that the novel chimeric H9/H5N2 recombinant virus is a low-pathogenic virus, and this chimeric vaccine is suitable for a DIVA vaccine with broad-spectrum neutralizing antibody against H5 avian influenza viruses. IMPORTANCE Current influenza virus killed vaccines predominantly induce antihemagglutinin (anti-HA) antibodies that are commonly strain specific in that the antibodies have potent neutralizing activity against homologous strains but do not cross-react with HAs of other influenza virus subtypes. In contrast, the HA2 stalk domain is relatively well conserved among subtypes, and recently, broadly neutralizing antibodies against this domain have been isolated. Therefore, in light of the need for a vaccine strain that applies the DIVA strategy utilizing an HI assay and induces broad cross-protection against H5N1 and H9N2 viruses, we generated a novel chimeric H9/H5N1 virus that expresses the entire HA1 portion from the H9N2 virus and the HA2 region of the heterosubtypic H5N8 virus. The chimeric H9/H5N2 recombinant vaccine protected immunized hosts against lethal challenge with H9N2 and HPAI H5N1 viruses with significantly attenuated virus shedding in immunized hosts. Therefore, this chimeric vaccine is suitable as a DIVA vaccine against H5 avian influenza viruses.

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Human monoclonal antibodies with broad-spectrum neutralizing activity against influenza viruses

Current Issues with Influenza Management ◽

10.2217/fmeb2013.13.284 ◽

2015 ◽

pp. 40-53

Author(s):

Yang Pan ◽

Ririn Ramadhany ◽

Tadahiro Sasaki ◽

Kazuyoshi Ikuta

Keyword(s):

Monoclonal Antibodies ◽

Broad Spectrum ◽

Influenza Viruses ◽

Human Monoclonal Antibodies ◽

Neutralizing Activity

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Antibodies to Highly Pathogenic A/H5Nx (Clade 2.3.4.4) Influenza Viruses in the Sera of Vietnamese Residents

Pathogens ◽

10.3390/pathogens10040394 ◽

2021 ◽

Vol 10 (4) ◽

pp. 394

Author(s):

Tatyana Ilyicheva ◽

Vasily Marchenko ◽

Olga Pyankova ◽

Anastasia Moiseeva ◽

Tran Thi Nhai ◽

...

Keyword(s):

Influenza Virus ◽

Avian Influenza ◽

Wide Spectrum ◽

Influenza Viruses ◽

Serum Samples ◽

Socialist Republic ◽

Highly Pathogenic ◽

Virus Variant ◽

Human Sera ◽

Hemagglutinin Inhibition

To cause a pandemic, an influenza virus has to overcome two main barriers. First, the virus has to be antigenically new to humans. Second, the virus has to be directly transmitted from humans to humans. Thus, if the avian influenza virus is able to pass the second barrier, it could cause a pandemic, since there is no immunity to avian influenza in the human population. To determine whether the adaptation process is ongoing, analyses of human sera could be conducted in populations inhabiting regions where pandemic virus variant emergence is highly possible. This study aimed to analyze the sera of Vietnamese residents using hemagglutinin inhibition reaction (HI) and microneutralization (MN) with A/H5Nx (clade 2.3.4.4) influenza viruses isolated in Vietnam and the Russian Federation in 2017–2018. In this study, we used sera from 295 residents of the Socialist Republic of Vietnam collected from three groups: 52 samples were collected from households in Nam Dinh province, where poultry deaths have been reported (2017); 96 (2017) and 147 (2018) samples were collected from patients with somatic but not infectious diseases in Hanoi. In all, 65 serum samples were positive for HI, at least to one H5 virus used in the study. In MN, 47 serum samples neutralizing one or two viruses at dilutions of 1/40 or higher were identified. We postulate that the rapidly evolving A/H5Nx (clade 2.3.4.4) influenza virus is possibly gradually adapting to the human host, insofar as healthy individuals have antibodies to a wide spectrum of variants of that subtype.

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