Formulation of bovine respiratory syncytial virus fusion protein with CpG oligodeoxynucleotide, cationic host defence peptide and polyphosphazene enhances humoral and cellular responses and induces a protective type 1 immune response in mice

Respiratory syncytial virus (RSV) is the leading cause of serious respiratory tract disease in children and calves; however, RSV vaccine development has been slow due to early observations that formalin-inactivated vaccines induced Th2-type immune responses and led to disease enhancement upon subsequent exposure. Hence, there is a need for novel adjuvants that will promote a protective Th1-type or balanced immune response against RSV. CpG oligodeoxynucleotides (ODNs), indolicidin, and polyphosphazene were examined for their ability to enhance antigen-specific immune responses and influence the Th-bias when co-formulated with a recombinant truncated bovine RSV (BRSV) fusion protein (ΔF). Mice immunized with ΔF co-formulated with CpG ODN, indolicidin, and polyphosphazene (ΔF/CpG/indol/PP) developed higher levels of ΔF-specific serum IgG, IgG1 and IgG2a antibodies when compared with ΔF alone, and displayed an increase in the frequency of gamma interferon-secreting cells and decreased interleukin (IL)-5 production by in vitro restimulated splenocytes, characteristic of a Th1 immune response. These results were observed in both C57BL/6 and BALB/c strains of mice. When evaluated in a BRSV challenge model, mice immunized with ΔF/CpG/indol/PP developed significantly higher levels of BRSV-neutralizing serum antibodies than mice immunized with the ΔF protein alone, and displayed significantly less pulmonary IL-4, IL-5, IL-13 and eotaxin and reduced eosinophilia after challenge. These results suggest that co-formulation of ΔF with CpG ODN, host defence peptide and polyphosphazene may result in a safe and effective vaccine for the prevention of BRSV and may have implications for the development of novel human RSV vaccines.

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Enhanced immune responses and protection by vaccination with respiratory syncytial virus fusion protein formulated with CpG oligodeoxynucleotide and innate defense regulator peptide in polyphosphazene microparticles

Vaccine ◽

10.1016/j.vaccine.2012.06.011 ◽

2012 ◽

Vol 30 (35) ◽

pp. 5206-5214 ◽

Cited By ~ 49

Author(s):

S. Garlapati ◽

R. Garg ◽

R. Brownlie ◽

L. Latimer ◽

E. Simko ◽

...

Keyword(s):

Respiratory Syncytial Virus ◽

Immune Responses ◽

Fusion Protein ◽

Innate Defense ◽

Virus Fusion ◽

Cpg Oligodeoxynucleotide ◽

Syncytial Virus

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Formulation with CpG Oligodeoxynucleotides Prevents Induction of Pulmonary Immunopathology following Priming with Formalin-Inactivated or Commercial Killed Bovine Respiratory Syncytial Virus Vaccine

Journal of Virology ◽

10.1128/jvi.79.4.2024-2032.2005 ◽

2005 ◽

Vol 79 (4) ◽

pp. 2024-2032 ◽

Cited By ~ 32

Author(s):

M. Oumouna ◽

J. W. Mapletoft ◽

B. C. Karvonen ◽

L. A. Babiuk ◽

S. van Drunen Littel-van den Hurk

Keyword(s):

Respiratory Syncytial Virus ◽

Immune Responses ◽

Virus Production ◽

Neutralizing Antibody ◽

Bovine Respiratory Syncytial Virus ◽

Interleukin 4 ◽

Cpg Odn ◽

Cpg Oligodeoxynucleotides ◽

Syncytial Virus

ABSTRACT Commercial killed bovine respiratory syncytial virus (K-BRSV) and formalin-inactivated BRSV (FI-BRSV) tend to induce Th2-type immune responses, which may not be protective and may even be detrimental during subsequent exposure to the virus. In this study we assessed the ability of CpG oligodeoxynucleotides (ODNs) to aid in the generation of effective and protective BRSV-specific immune responses. Mice were immunized subcutaneously with FI-BRSV formulated with CpG ODN, Emulsigen (Em), CpG ODN and Em, or non-CpG ODN and Em. Two additional groups were immunized with K-BRSV or K-BRSV and CpG ODN. After two vaccinations, the mice were challenged with BRSV. FI-BRSV induced Th2-biased immune responses characterized by production of serum immunoglobulin G1 (IgG1) and IgE, as well as interleukin-4 (IL-4), by in vitro-restimulated splenocytes. Formulation of FI-BRSV with CpG ODN, but not with non-CpG ODN, enhanced serum IgG2a and IFN-γ production by splenocytes, whereas serum IgE was reduced. Although the immune response induced by K-BRSV was not as strongly Th2 biased, the addition of CpG ODN to this commercial vaccine also resulted in a more Th1-type response. Furthermore, the addition of CpG ODN to the BRSV vaccine formulations resulted in enhanced neutralizing antibody responses. Significant production of IL-5, eotaxin, and eosinophilia was observed in the lungs of FI-BRSV- and K-BRSV-immunized mice. However, IL-5 and eotaxin levels, as well as the number of eosinophils, were decreased in the mice vaccinated with the CpG ODN-formulated vaccines. Finally, when formulated with CpG ODN, both FI-BRSV and K-BRSV significantly reduced virus production after challenge with BRSV.

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Intranasal Immunization of Mice with a Bovine Respiratory Syncytial Virus Vaccine Induces Superior Immunity and Protection Compared to Those by Subcutaneous Delivery or Combinations of Intranasal and Subcutaneous Prime-Boost Strategies

Clinical and Vaccine Immunology ◽

10.1128/cvi.00250-09 ◽

2009 ◽

Vol 17 (1) ◽

pp. 23-35 ◽

Cited By ~ 18

Author(s):

John W. Mapletoft ◽

Laura Latimer ◽

Lorne A. Babiuk ◽

Sylvia van Drunen Littel-van den Hurk

Keyword(s):

Respiratory Syncytial Virus ◽

Immune Responses ◽

Virus Replication ◽

Vaccination Strategy ◽

Bovine Respiratory Syncytial Virus ◽

Mucosal Vaccine ◽

Intranasal Delivery ◽

Cpg Odn ◽

Cpg Oligodeoxynucleotide ◽

Syncytial Virus

ABSTRACT Bovine respiratory syncytial virus (BRSV) infects cells of the respiratory mucosa, so it is desirable to develop a vaccination strategy that induces mucosal immunity. To achieve this, various delivery routes were compared for formalin-inactivated (FI) BRSV formulated with CpG oligodeoxynucleotide (ODN) and polyphosphazene (PP). Intranasal delivery of the FI-BRSV formulation was superior to subcutaneous delivery in terms of antibody, cell-mediated, and mucosal immune responses, as well as reduction in virus replication after BRSV challenge. Although intranasal delivery of FI-BRSV also induced higher serum and lung antibody titers and gamma interferon (IFN-γ) production in the lungs than intranasal-subcutaneous and/or subcutaneous-intranasal prime-boost strategies, no significant differences were observed in cell-mediated immune responses or virus replication in the lungs of challenged mice. Interleukin 5 (IL-5), eotaxin, and eosinophilia were enhanced after BRSV challenge in the lungs of subcutaneously immunized mice compared to unvaccinated mice, but not in the lungs of mice immunized intranasally or through combinations of the intranasal and subcutaneous routes. These results suggest that two intranasal immunizations with FI-BRSV formulated with CpG ODN and PP are effective and safe as an approach to induce systemic and mucosal responses, as well to reduce virus replication after BRSV challenge. Furthermore, intranasal-subcutaneous and subcutaneous-intranasal prime-boost strategies were also safe and almost as efficacious. In addition to the implications for the development of a protective BRSV vaccine for cattle, formulation with CpG ODN and PP could also prove important in the development of a mucosal vaccine that induces protective immunity against human RSV.

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Comparison of Immune Responses to Different Versions of VLP Associated Stabilized RSV Pre-Fusion F Protein

Vaccines ◽

10.3390/vaccines7010021 ◽

2019 ◽

Vol 7 (1) ◽

pp. 21 ◽

Cited By ~ 4

Author(s):

Lori Cullen ◽

Madelyn Schmidt ◽

Gretel Torres ◽

Adam Capoferri ◽

Trudy Morrison

Keyword(s):

Monoclonal Antibodies ◽

Respiratory Syncytial Virus ◽

Immune Responses ◽

Fusion Protein ◽

Neutralizing Antibodies ◽

Vaccine Development ◽

Vaccine Candidates ◽

F Protein ◽

Syncytial Virus ◽

Conformation Stability

Efforts to develop a vaccine for respiratory syncytial virus (RSV) have primarily focused on the RSV fusion protein. The pre-fusion conformation of this protein induces the most potent neutralizing antibodies and is the focus of recent efforts in vaccine development. Following the first identification of mutations in the RSV F protein (DS-Cav1 mutant protein) that stabilized the pre-fusion conformation, other mutant stabilized pre-fusion F proteins have been described. To determine if there are differences in alternate versions of stabilized pre-fusion F proteins, we explored the use, as vaccine candidates, of virus-like particles (VLPs) containing five different pre-fusion F proteins, including the DS-Cav1 protein. The expression of these five pre-F proteins, their assembly into VLPs, their pre-fusion conformation stability in VLPs, their reactivity with anti-F monoclonal antibodies, and their induction of immune responses after the immunization of mice, were characterized, comparing VLPs containing the DS-Cav1 pre-F protein with VLPs containing four alternative pre-fusion F proteins. The concentrations of anti-F IgG induced by each VLP that blocked the binding of prototype monoclonal antibodies using two different soluble pre-fusion F proteins as targets were measured. Our results indicate that both the conformation and immunogenicity of alternative VLP associated stabilized pre-fusion RSV F proteins are different from those of DS-Cav1 VLPs.

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CpG Oligodeoxynucleotides Act as Adjuvants that Switch on T Helper 1 (Th1) Immunity

Journal of Experimental Medicine ◽

10.1084/jem.186.10.1623 ◽

1997 ◽

Vol 186 (10) ◽

pp. 1623-1631 ◽

Cited By ~ 735

Author(s):

Rose S. Chu ◽

Oleg S. Targoni ◽

Arthur M. Krieg ◽

Paul V. Lehmann ◽

Clifford V. Harding

Keyword(s):

Immune Response ◽

Immune Responses ◽

Cpg Odn ◽

T Helper ◽

T Helper 1 ◽

Cpg Oligodeoxynucleotides ◽

Cytokine Pattern ◽

Incomplete Freund’S Adjuvant ◽

Ifn Γ ◽

Hen Egg

Synthetic oligodeoxynucleotides (ODN) that contain unmethylated CpG motifs (CpG ODN) induce macrophages to secrete IL-12, which induces interferon (IFN)-γ secretion by natural killer (NK) cells. Since these cytokines can induce T helper 1 (Th1) differentiation, we examined the effects of coadministered CpG ODN on the differentiation of Th responses to hen egg lysozyme (HEL). In both BALB/c (Th2-biased) and B10.D2 (Th1-biased) mice, immunization with HEL in incomplete Freund's adjuvant (IFA) resulted in Th2-dominated immune responses characterized by HEL-specific secretion of IL-5 but not IFN-γ. In contrast, immunization with IFA-HEL plus CpG ODN switched the immune response to a Th1-dominated cytokine pattern, with high levels of HEL-specific IFN-γ secretion and decreased HEL-specific IL-5 production. IFA-HEL plus CpG ODN also induced anti-HEL IgG2a (a Th1-associated isotype), which was not induced by IFA-HEL alone. Control non–CpG ODN did not induce IFN-γ or IgG2a, excepting lesser increases in B10.D2 (Th1-biased) mice. Thus, CpG ODN provide a signal to switch on Th1-dominated responses to coadministered antigen and are potential adjuvants for human vaccines to elicit protective Th1 immunity.

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The respiratory syncytial virus fusion protein formulated with a novel combination adjuvant induces balanced immune responses in lambs with maternal antibodies

Vaccine ◽

10.1016/j.vaccine.2015.01.041 ◽

2015 ◽

Vol 33 (11) ◽

pp. 1338-1344 ◽

Cited By ~ 18

Author(s):

R. Garg ◽

L. Latimer ◽

V. Gerdts ◽

A. Potter ◽

S. van Drunen Littel-van den Hurk

Keyword(s):

Respiratory Syncytial Virus ◽

Immune Responses ◽

Fusion Protein ◽

Maternal Antibodies ◽

Virus Fusion ◽

Syncytial Virus

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Intranasal immunization of mice with a formalin-inactivated bovine respiratory syncytial virus vaccine co-formulated with CpG oligodeoxynucleotides and polyphosphazenes results in enhanced protection

Journal of General Virology ◽

10.1099/vir.0.83300-0 ◽

2008 ◽

Vol 89 (1) ◽

pp. 250-260 ◽

Cited By ~ 34

Author(s):

John W. Mapletoft ◽

Mustapha Oumouna ◽

Jennifer Kovacs-Nolan ◽

Laura Latimer ◽

George Mutwiri ◽

...

Keyword(s):

Respiratory Syncytial Virus ◽

Mucosal Immunity ◽

Neutralizing Antibodies ◽

Cell Mediated Immunity ◽

Cpg Odn ◽

Vaccine Platform ◽

Intranasal Immunization ◽

Cpg Oligodeoxynucleotides ◽

Enhanced Production ◽

Syncytial Virus

As respiratory syncytial virus (RSV) targets the mucosal surfaces of the respiratory tract, induction of both systemic and mucosal immunity will be critical for optimal protection. In this study, the ability of an intranasally delivered, formalin-inactivated bovine RSV (FI-BRSV) vaccine co-formulated with CpG oligodeoxynucleotides (ODN) and polyphosphazenes (PP) to induce systemic and mucosal immunity, as well as protection from BRSV challenge, was evaluated. Intranasal immunization of mice with FI-BRSV formulated with CpG ODN and PP resulted in both humoral and cell-mediated immunity, characterized by enhanced production of BRSV-specific serum IgG, as well as increased gamma interferon and decreased interleukin-5 production by in vitro-restimulated splenocytes. These mice also developed mucosal immune responses, as was evident from increased production of BRSV-specific IgG and IgA in lung-fragment cultures. Indeed, the increases in serum and mucosal IgG, and in particular mucosal IgA and virus-neutralizing antibodies, were the most critical differences observed between FI-BRSV formulated with both CpG ODN and PP in comparison to formulations with CpG ODN, non-CpG ODN or PP individually. Finally, FI-BRSV/CpG/PP was the only formulation that resulted in a significant reduction in viral replication upon BRSV challenge. Co-formulation of CpG ODN and PP is a promising new vaccine platform technology that may have applications in mucosal immunization in humans.

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