Robust Immunity and Heterologous Protection against Influenza in Mice Elicited by a Novel Recombinant NP-M2e Fusion Protein Expressed in E. coli

The Ohio State University, United States of America
PLoS ONE (Impact Factor: 3.23). 12/2012; 7(12):e52488. DOI: 10.1371/journal.pone.0052488
Source: PubMed


The 23-amino acid extracellular domain of matrix 2 protein (M2e) and the internal nucleoprotein (NP) of influenza are highly conserved among viruses and thus are promising candidate antigens for the development of a universal influenza vaccine. Various M2e- or NP-based DNA or viral vector vaccines have been shown to have high immunogenicity; however, high cost, complicated immunization procedures, and vector-specific antibody responses have restricted their applications. Immunization with an NP-M2e fusion protein expressed in Escherichia coli may represent an alternative strategy for the development of a universal influenza vaccine.
cDNA encoding M2e was fused to the 3' end of NP cDNA from influenza virus A/Beijing/30/95 (H3N2). The fusion protein (NM2e) was expressed in E. coli and isolated with 90% purity. Mice were immunized with recombinant NM2e protein along with aluminum hydroxide gel and/or CpG as adjuvant. NM2e plus aluminum hydroxide gel almost completely protected the mice against a lethal (20 LD(50)) challenge of heterologous influenza virus A/PR/8/34.
The NM2e fusion protein expressed in E. coli was highly immunogenic in mice. Immunization with NM2e formulated with aluminum hydroxide gel protected mice against a lethal dose of a heterologous influenza virus. Vaccination with recombinant NM2e fusion protein is a promising strategy for the development of a universal influenza vaccine.

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Available from: Baoying Huang, Apr 15, 2014
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    • "We used CpG and Al(OH) 3 to improve the immunogenicity of the NM2e fusion protein of the influenza virus in a previous report; the adjuvant showed potential efficacy, strengthened the immune response and elicited cross protection after NM2e immunization in mice. A combination of Al(OH) 3 and CpG in the formulation of NM2e induced the highest protection in mice (Wang et al., 2012). In this study, CpG and Al(OH) 3 were employed separately or in combination with 10 mg NP expressed in E. coli to improve its Fig. 3. IgG1 and IgG2a isotypes of serum from NP-immunized mice. "
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    ABSTRACT: The highly conserved internal nucleoprotein (NP) is a promising antigen to develop a universal influenza A virus vaccine. In this study, mice were injected intramuscularly with Escherichia coli-derived NP protein alone or in combination with adjuvant alum (Al(OH)3), CpG or both. The results showed that the NP protein formulated with adjuvant was effective in inducing a protective immune response. Additionally, the adjuvant efficacy of Al(OH)3 was stronger than that of CpG. Optimal immune responses were observed in BALB/c mice immunized with a combination of NP protein plus Al(OH)3 and CpG. These mice also showed maximal resistance following challenge with influenza A virus PR8 strain. Most importantly, 10µg NP formulated with Al(OH)3 and CpG induced higher protection than did 90µg NP. These findings indicated that a combination of Al(OH)3 and CpG may be an efficient adjuvant in the NP formulation.
    Virology 09/2014; 468-470C:265-273. DOI:10.1016/j.virol.2014.08.008 · 3.32 Impact Factor
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    • "Thus, the development of vaccines able to trigger strong CD4+ responses could be central for the induction of memory responses capable of combating divergent influenza viruses through multiple pathways. In this context, there is experimental evidence on NP-based vaccines which promote CD4+ T cell responses contributing towards protective immunity [22], [66]. This phenomenon is not restricted only to influenza, since numerous infectious models have demonstrated the importance of CD4+ T cells in cellular mediated protection [67], [68]. "
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    ABSTRACT: There is a critical need for new influenza vaccines able to protect against constantly emerging divergent virus strains. This will be sustained by the induction of vigorous cellular responses and humoral immunity capable of acting at the portal of entry of this pathogen. In this study we evaluate the protective efficacy of intranasal vaccination with recombinant influenza nucleoprotein (rNP) co-administrated with bis-(3',5')-cyclic dimeric adenosine monophosphate (c-di-AMP) as adjuvant. Immunization of BALB/c mice with two doses of the formulation stimulates high titers of NP-specific IgG in serum and secretory IgA at mucosal sites. This formulation also promotes a strong Th1 response characterized by high secretion of INF-γ and IL-2. The immune response elicited promotes efficient protection against virus challenge. These results suggest that c-di-AMP is a potent mucosal adjuvant which may significantly contribute towards the development of innovative mucosal vaccines against influenza.
    PLoS ONE 08/2014; 9(8):e104824. DOI:10.1371/journal.pone.0104824 · 3.23 Impact Factor
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    • "route (Figure 2C) showing clearly the effectiveness of PapMV nanoparticles to improve the mucosal antibody response in the lung. The ELISA also revealed that the adjuvanted vaccine broaden the immune response to the TIV antigens through a significant increased in the lungs of the IgG2a titers directed to the influenza NP (Figure 2D), a highly conserved protein through all the strains of influenza often used in experimental influenza vaccines [7,14,15]. NP has also been identified as a key antigen to trigger cross-protection to influenza viruses [8,16-18]. It is therefore a good strategy to increase the immune response directed to this antigen using PapMV nanoparticles to broaden and improve protection to this virus. "
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    ABSTRACT: Background Trivalent inactivated flu vaccines (TIV) are currently the best means to prevent influenza infections. However, the protection provided by TIV is partial (about 50%) and it is needed to improve the efficacy of protection. Since the respiratory tract is the main site of influenza replications, a vaccine that triggers mucosal immunity in this region can potentially improve protection against this disease. Recently, PapMV nanoparticles used as an adjuvant in a formulation with TIV administered by the subcutaneous route have shown improving the immune response directed to the TIV and protection against an influenza challenge. Findings In the present study, we showed that intranasal instillation with a formulation containing TIV and PapMV nanoparticles significantly increase the amount of IgG, IgG2a and IgA in lungs of vaccinated mice as compared to mice that received TIV only. Instillation with the adjuvanted formulation leads to a more robust protection against an influenza infection with a strain that is lethal to mice vaccinated with the TIV. Conclusions We demonstrate for the first time that PapMV nanoparticles are an effective and potent mucosal adjuvant for vaccination.
    Journal of Nanobiotechnology 05/2014; 12(19). DOI:10.1186/1477-3155-12-19 · 4.12 Impact Factor
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