Immunogenicity and Protective Efficacy of Influenza A DNA Vaccines Encoding Artificial Antigens Based on Conservative Hemagglutinin Stem Region and M2 Protein in Mice

Background: Development of a universal vaccine capable to induce antibody responses against a broad range of influenza virus strains attracts growing attention. Hemagglutinin stem and the exposed fragment of influenza virus M2 protein are promising targets for induction of cross-protective humoral and cell-mediated response, since they contain conservative epitopes capable to induce antibodies and cytotoxic T lymphocytes (CTLs) to a wide range of influenza virus subtypes. Methods: In this study, we generated DNA vaccine constructs encoding artificial antigens AgH1, AgH3, and AgM2 designed on the basis of conservative hemagglutinin stem fragments of two influenza A virus subtypes, H1N1 and H3N2, and conservative M2 protein, and evaluate their immunogenicity and protective efficacy. To obtain DNA vaccine constructs, genes encoding the designed antigens were cloned into a pcDNA3.1 vector. Expression of the target genes in 293T cells transfected with DNA vaccine constructs has been confirmed by synthesis of specific mRNA. Results: Immunization of BALB/c mice with DNA vaccines encoding these antigens was shown to evoke humoral and T-cell immune responses as well as a moderated statistically significant cross-protective effect against two heterologous viruses A/California/4/2009 (H1N1pdm09) and A/Aichi/2/68 (H3N2). Conclusions: The results demonstrate a potential approach to creating a universal influenza vaccine based on artificial antigens.

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Immune effects against influenza A virus and a novel DNA vaccine with co-expression of haemagglutinin- and neuraminidase-encoding genes

Journal of Medical Microbiology ◽

10.1099/jmm.0.006825-0 ◽

2009 ◽

Vol 58 (7) ◽

pp. 845-854 ◽

Cited By ~ 8

Author(s):

Weidong Zhang ◽

Wanyi Li ◽

Yan Li ◽

Hong Li ◽

Baoning Wang ◽

...

Keyword(s):

Influenza Virus ◽

Dna Vaccine ◽

Influenza A ◽

Protective Efficacy ◽

Survival Rates ◽

Serum Antibody ◽

Lethal Challenge ◽

Vector Systems

The high variability of influenza virus causes difficulties in the control and prevention of influenza, thus seeking a promising approach for dealing with these problems is a hot topic. Haemagglutinin (HA) and neuraminidase (NA) are major surface antigens of the influenza virus, and provide effective protection against lethal challenges with this virus. We constructed a DNA vaccine (pHA-IRES2-NA) that co-expressed both HA and NA, and compared its protective efficacy and immunogenic ability with that of singly expressed HA or NA, or a mixture of the two singly expressed proteins. Our findings showed that both HA and NA proteins expressed by pHA-IRES2-NA could be detected in vivo and in vitro. The protection of DNA vaccines was evaluated by serum antibody titres, residual lung virus titres and survival rates of the mice. In the murine model, immunization of pHA-IRES2-NA generated significant anti-HA and anti-NA antibody, increased the percentage of CD8+ cells and gamma interferon-producing CD8+ cells and the ratio of Th1/Th2 (T helper) cells, which was comparable to the effects of immunization with HA or NA DNA alone or with a mixture of HA and NA DNA. All the mice inoculated by pHA-IRES2-NA resisted the lethal challenge by homologous influenza virus and survived with low lung virus titre. In addition, previous studies reported that co-expression allowed higher-frequency transduction compared to co-transduction of separated vector systems encoding different genes. The novel HA and NA co-expression DNA vaccine is a successful alternative to using a mixture of purified HA and NA proteins or HA and NA DNA.

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Efficacy of recombinant NP-M1 and NP-M1-CRT DNA vaccines against Influenza A viruses in mice C57/BL6

10.1101/2021.06.24.449692 ◽

2021 ◽

Author(s):

Hamidreza Attaran ◽

Wen He ◽

wei wang

Keyword(s):

Influenza Virus ◽

Murine Model ◽

Mammalian Cells ◽

Influenza A ◽

Matrix Protein ◽

Protective Efficacy ◽

Influenza Virus Infection ◽

Influenza Viruses ◽

Universal Vaccine

Effective vaccination against the influenza virus remains a challenge because of antigenic shift and drift in influenza viruses. Conservation is an important feature of the Nucleoprotein (NP)and Matrix protein 1(M1) qualifying them as potential candidates for developing a universal vaccine against the influenza A virus. Carliticulin (CRT), a member of heat shock protein (HSP) family, are conserved and widely distributed in many microorganisms and mammalian cells. In this study, a plasmid vector encoding the NP-M1-CRT sequence was constructed and compared with the NP-M1 sequence with respect to immunogenicity and protective efficacy in a murine model. The potency of the created construct for provoking humoral, cellular immune responses, and its protective immunity against the lethal influenza virus infection were then compared with commercial split vaccine and then evaluated in a murine model system. NP-M1-CRT as a DNA vaccine combined with in vivo electroporation could significantly improve the immunogenicity of constructed vectors. Serological evaluations demonstrated the potency of our approach to provoke strong anti-NP specific antibody responses. Furthermore, our strategy of immunization in prime-boost groups were able to provide protection against lethal viral challenge using H1N1 subtype. The ease of production of these types of vectors and the fact that they would not require annual updating and manufacturing may provide an alternative cost-effective approach to limit the spread of potential pandemic influenza viruses.

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Protective Efficacy and Immunogenicity of a Combinatory DNA Vaccine against Influenza A Virus and the Respiratory Syncytial Virus

PLoS ONE ◽

10.1371/journal.pone.0072217 ◽

2013 ◽

Vol 8 (8) ◽

pp. e72217 ◽

Cited By ~ 16

Author(s):

Viktoria Stab ◽

Sandra Nitsche ◽

Thomas Niezold ◽

Michael Storcksdieck genannt Bonsmann ◽

Andrea Wiechers ◽

...

Keyword(s):

Respiratory Syncytial Virus ◽

Influenza A Virus ◽

Dna Vaccine ◽

Influenza A ◽

Protective Efficacy ◽

Syncytial Virus

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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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Protective Efficacy of the Conserved NP, PB1, and M1 Proteins as Immunogens in DNA- and Vaccinia Virus-Based Universal Influenza A Virus Vaccines in Mice

Clinical and Vaccine Immunology ◽

10.1128/cvi.00091-15 ◽

2015 ◽

Vol 22 (6) ◽

pp. 618-630 ◽

Cited By ~ 29

Author(s):

Wenling Wang ◽

Renqing Li ◽

Yao Deng ◽

Ning Lu ◽

Hong Chen ◽

...

Keyword(s):

Influenza Virus ◽

Virus Strain ◽

Influenza A ◽

Protective Efficacy ◽

Lethal Dose ◽

Influenza Vaccines ◽

Strong Immune Response ◽

Influenza Virus Strain ◽

Vaccinia Viruses ◽

Viral Components

ABSTRACTThe conventional hemagglutinin (HA)- and neuraminidase (NA)-based influenza vaccines need to be updated most years and are ineffective if the glycoprotein HA of the vaccine strains is a mismatch with that of the epidemic strain. Universal vaccines targeting conserved viral components might provide cross-protection and thus complement and improve conventional vaccines. In this study, we generated DNA plasmids and recombinant vaccinia viruses expressing the conserved proteins nucleoprotein (NP), polymerase basic 1 (PB1), and matrix 1 (M1) from influenza virus strain A/Beijing/30/95 (H3N2). BALB/c mice were immunized intramuscularly with a single vaccine based on NP, PB1, or M1 alone or a combination vaccine based on all three antigens and were then challenged with lethal doses of the heterologous influenza virus strain A/PR/8/34 (H1N1). Vaccines based on NP, PB1, and M1 provided complete or partial protection against challenge with 1.7 50% lethal dose (LD50) of PR8 in mice. Of the three antigens, NP-based vaccines induced protection against 5 LD50and 10 LD50and thus exhibited the greatest protective effect. Universal influenza vaccines based on the combination of NP, PB1, and M1 induced a strong immune response and thus might be an alternative approach to addressing future influenza virus pandemics.

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Targeting Hemagglutinin: Approaches for Broad Protection against the Influenza A Virus

Viruses ◽

10.3390/v11050405 ◽

2019 ◽

Vol 11 (5) ◽

pp. 405 ◽

Cited By ~ 13

Author(s):

Zhang ◽

Xu ◽

Zhang ◽

Liu ◽

Xue ◽

...

Keyword(s):

Influenza A Virus ◽

Influenza A ◽

Protective Efficacy ◽

Structure And Function ◽

Antigenic Drift ◽

Influenza A Viruses ◽

Universal Vaccine ◽

Vaccine Candidates ◽

And Function ◽

Ha Protein

Influenza A viruses are dynamically epidemic and genetically diverse. Due to the antigenic drift and shift of the virus, seasonal vaccines are required to be reformulated annually to match with current circulating strains. However, the mismatch between vaccinal strains and circulating strains occurs frequently, resulting in the low efficacy of seasonal vaccines. Therefore, several “universal” vaccine candidates based on the structure and function of the hemagglutinin (HA) protein have been developed to meet the requirement of a broad protection against homo-/heterosubtypic challenges. Here, we review recent novel constructs and discuss several important findings regarding the broad protective efficacy of HA-based universal vaccines.

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Enhanced Influenza Virus-Like Particle Vaccines Containing the Extracellular Domain of Matrix Protein 2 and a Toll-Like Receptor Ligand

Clinical and Vaccine Immunology ◽

10.1128/cvi.00153-12 ◽

2012 ◽

Vol 19 (8) ◽

pp. 1119-1125 ◽

Cited By ~ 37

Author(s):

Bao-Zhong Wang ◽

Harvinder S. Gill ◽

Sang-Moo Kang ◽

Li Wang ◽

Ying-Chun Wang ◽

...

Keyword(s):

Influenza Virus ◽

Fusion Protein ◽

Influenza A ◽

Matrix Protein ◽

Protective Efficacy ◽

Extracellular Domain ◽

Toll Like Receptor ◽

Influenza A Viruses ◽

Live Virus ◽

Content Type

ABSTRACTThe extracellular domain of matrix protein 2 (M2e) is conserved among influenza A viruses. The goal of this project is to develop enhanced influenza vaccines with broad protective efficacy using the M2e antigen. We designed a membrane-anchored fusion protein by replacing the hyperimmunogenic region ofSalmonella entericaserovar Typhimurium flagellin (FliC) with four repeats of M2e (4.M2e-tFliC) and fusing it to a membrane anchor from influenza virus hemagglutinin (HA). The fusion protein was incorporated into influenza virus M1-based virus-like particles (VLPs). These VLPs retained Toll-like receptor 5 (TLR5) agonist activity comparable to that of soluble FliC. Mice immunized with the VLPs by either intramuscular or intranasal immunization showed high levels of systemic M2-specific antibody responses compared to the responses to soluble 4.M2e protein. High mucosal antibody titers were also induced in intranasally immunized mice. All intranasally immunized mice survived lethal challenges with live virus, while intramuscularly immunized mice showed only partial protection, revealing better protection by the intranasal route. These results indicate that a combination of M2e antigens and TLR ligand adjuvants in VLPs has potential for development of a broadly protective influenza A virus vaccine.

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A DNA Vaccine That Targets Hemagglutinin to Antigen-Presenting Cells Protects Mice against H7 Influenza

Journal of Virology ◽

10.1128/jvi.01340-17 ◽

2017 ◽

Vol 91 (23) ◽

Cited By ~ 5

Author(s):

Tor Kristian Andersen ◽

Fan Zhou ◽

Rebecca Cox ◽

Bjarne Bogen ◽

Gunnveig Grødeland

Keyword(s):

Avian Influenza ◽

Dna Vaccine ◽

Influenza A ◽

Dna Vaccines ◽

Antigen Presenting Cells ◽

Dna Vaccination ◽

Lethal Challenge ◽

Highly Pathogenic ◽

Pandemic Threat ◽

Antigen Presenting

ABSTRACT Zoonotic influenza H7 viral infections have a case fatality rate of about 40%. Currently, no or limited human to human spread has occurred, but we may be facing a severe pandemic threat if the virus acquires the ability to transmit between humans. Novel vaccines that can be rapidly produced for global distribution are urgently needed, and DNA vaccines may be the only type of vaccine that allows for the speed necessary to quench an emerging pandemic. Here, we constructed DNA vaccines encoding the hemagglutinin (HA) from influenza A/chicken/Italy/13474/99 (H7N1). In order to increase the efficacy of DNA vaccination, HA was targeted to either major histocompatibility complex class II molecules or chemokine receptors 1, 3, and 5 (CCR1/3/5) that are expressed on antigen-presenting cells (APC). A single DNA vaccination with APC-targeted HA significantly increased antibody levels in sera compared to nontargeted control vaccines. The antibodies were confirmed neutralizing in an H7 pseudotype-based neutralization assay. Furthermore, the APC-targeted vaccines increased the levels of antigen-specific cytotoxic T cells, and a single DNA vaccination could confer protection against a lethal challenge with influenza A/turkey/Italy/3889/1999 (H7N1) in mice. In conclusion, we have developed a vaccine that rapidly could contribute protection against a pandemic threat from avian influenza. IMPORTANCE Highly pathogenic avian influenza H7 constitute a pandemic threat that can cause severe illness and death in infected individuals. Vaccination is the main method of prophylaxis against influenza, but current vaccine strategies fall short in a pandemic situation due to a prolonged production time and insufficient production capabilities. In contrast, a DNA vaccine can be rapidly produced and deployed to prevent the potential escalation of a highly pathogenic influenza pandemic. We here demonstrate that a single DNA delivery of hemagglutinin from an H7 influenza could mediate full protection against a lethal challenge with H7N1 influenza in mice. Vaccine efficacy was contingent on targeting of the secreted vaccine protein to antigen-presenting cells.

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Features of immune response against influenza infection in animals vaccinated with recombinant cross-protective vaccine

Russian Journal of Infection and Immunity ◽

10.15789/2220-7619-2019-3-4-485-494 ◽

2019 ◽

Vol 9 (3-4) ◽

pp. 485-494

Author(s):

L. M. Tsybalova ◽

L. A. Stepanova ◽

A. V. Korotkov ◽

M. A. Shuklina ◽

M. V. Zaitseva ◽

...

Keyword(s):

Immune Response ◽

T Cells ◽

Influenza Virus ◽

Influenza A Virus ◽

Cd4 T Cells ◽

Influenza A ◽

Influenza Infection ◽

Sublethal Dose ◽

Universal Vaccine ◽

Toll Like Receptor 5

Generating cross-reactive vaccines aimed at targeting all human influenza A virus subtypes is among high priority tasks in contemporary vaccinology. Such vaccines will be primarily demanded during pre-pandemic period as well as used to prime some population cohorts prior to vaccination with standard vaccines containing area-relevant epidemic virus. Unlike routine approach universal vaccines do not induce a sterilizing immunity, but significantly ameliorate overt infection and probable complications. Our study was aimed at evaluating characteristics of immune response in experimental animals primed with a candidate universal vaccine challenged with sublethal influenza A virus infection. Mice were immunized intranasally with the recombinant protein FlgH2-2-4M2e containing conservative peptides derived from two influenza A virus proteins: M2 protein ectodomain and 76–130 amino acid sequence from the second hemagglutinin (HA2) subunit genetically linked to bacterial flagellin protein, which is a ligand for Toll-like receptor 5 (TLR5). Control mice received saline. Two weeks after immunization, mice from both groups were infected with a sublethal dose of A/Aichi/2/68 AN3N2 influenza virus strain. Level of immunoglobulins G and A in the blood sera and bronchoalveolar lavages (BAL) were determined two weeks after immunization and 1 month post infection. Percentage of lung CD4+ T and CD4+ Tem (CD44+CD62L–) cells secreting cytokines TNFα, IFNγ, IL-2 was determined. Immunized vs. control mice responded to sublethal infection with the influenza virus by insignificant weight loss and more pronounced production of vaccine peptide-specific (M2e and aa76–130 HA2) and pan-influenza A/Aichi/2/68 virus IgG and A in the blood sera and BAL. After challenge the number of CD4+ T cells secreting cytokines TNFα and/or IL-2 in immunized mice significantly exceeded counterpart T cells in unimmunized animals that was true for both CD4+T and CD4+ Tem cells. Memory CD4+ T cells were previously shown to play a key role in the prime-boost event and heterosubtypic immune response. Thus, we were able to demonstrate a priming effect for recombinant cross-protective vaccine used in our experiment.

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Immunization of Pigs with a Particle-Mediated DNA Vaccine to Influenza A Virus Protects against Challenge with Homologous Virus

Journal of Virology ◽

10.1128/jvi.72.2.1491-1496.1998 ◽

1998 ◽

Vol 72 (2) ◽

pp. 1491-1496 ◽

Cited By ~ 112

Author(s):

Michael D. Macklin ◽

Dennis McCabe ◽

Martha W. McGregor ◽

Veronica Neumann ◽

Todd Meyer ◽

...

Keyword(s):

Immune Response ◽

Influenza Virus ◽

Influenza A Virus ◽

Dna Vaccine ◽

Influenza A ◽

Swine Influenza Virus ◽

Swine Influenza ◽

Humoral Response ◽

Preclinical Model ◽

Homologous Virus

ABSTRACT Particle-mediated delivery of a DNA expression vector encoding the hemagglutinin (HA) of an H1N1 influenza virus (A/Swine/Indiana/1726/88) to porcine epidermis elicits a humoral immune response and accelerates the clearance of virus in pigs following a homotypic challenge. Mucosal administration of the HA expression plasmid elicits an immune response that is qualitatively different than that elicited by the epidermal vaccination in terms of inhibition of the initial virus infection. In contrast, delivery of a plasmid encoding an influenza virus nucleoprotein from A/PR/8/34 (H1N1) to the epidermis elicits a strong humoral response but no detectable protection in terms of nasal virus shed. The efficacy of the HA DNA vaccine was compared with that of a commercially available inactivated whole-virus vaccine as well as with the level of immunity afforded by previous infection. The HA DNA and inactivated viral vaccines elicited similar protection in that initial infection was not prevented, but subsequent amplification of the infection is limited, resulting in early clearance of the virus. Convalescent animals which recovered from exposure to virulent swine influenza virus were completely resistant to infection when challenged. The porcine influenza A virus system is a relevant preclinical model for humans in terms of both disease and gene transfer to the epidermis and thus provides a basis for advancing the development of DNA-based vaccines.

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