A new subunit vaccine based on nucleoprotein nanoparticles confers partial clinical and virological protection in calves against bovine respiratory syncytial virus

Human and bovine respiratory syncytial viruses (HRSV and BRSV) are two closely related, worldwide prevalent viruses that are the leading cause of severe airway disease in children and calves, respectively. Efficacy of commercial bovine vaccines needs improvement and no human vaccine is licensed yet. We reported that nasal vaccination with the HRSV nucleoprotein produced as recombinant ringshaped nanoparticles (NSRS) protects mice against a viral challenge with HRSV. The aim of this work was to evaluate this new vaccine that uses a conserved viral antigen, in calves, natural hosts for BRSV. Calves, free of colostral or natural anti-BRSV antibodies, were vaccinated with NSRS either intramuscularly, or both intramuscularly and intranasally using MontanideTM ISA71 and IMS4132 as adjuvants and challenged with BRSV. All vaccinated calves developed anti-N antibodies in blood and nasal secretions and N-specific cellular immunity in local lymph nodes. Clinical monitoring post-challenge demonstrated moderate respiratory pathology with local lung tissue consolidations for the non vaccinated calves that were significantly reduced in the vaccinated calves. Vaccinated calves had lower viral loads than the nonvaccinated control calves. Thus NSRS vaccination in calves provided cross-protective immunity against BRSV infection without adverse inflammatory reaction

Sub-Nucleocapsid Nanoparticles: A Nasal Vaccine against Respiratory Syncytial Virus

Background: Bronchiolitis caused by the respiratory syncytial virus (RSV) in infants less than two years old is a growing public health concern worldwide, and there is currently no safe and effective vaccine. A major component of RSV nucleocapsid, the nucleoprotein (N), has been so far poorly explored as a potential vaccine antigen, even though it is a target of protective anti-viral T cell responses and is remarkably conserved between human RSV A and B serotypes. We recently reported a method to produce recombinant N assembling in homogenous rings composed of 10–11 N subunits enclosing a bacterial RNA. These nanoparticles were named sub-nucleocapsid ring structure (N SRS). Methodology and Principal Findings: The vaccine potential of N SRS was evaluated in a well-characterized and widely acknowledged mouse model of RSV infection. BALB/c adult mice were immunized intranasally with N SRS adjuvanted with the detoxified E. coli enterotoxin LT(R192G). Upon RSV challenge, vaccinated mice were largely protected against virus replication in the lungs, with a mild inflammatory lymphocytic and neutrophilic reaction in their airways. Mucosal immunization with N SRS elicited strong local and systemic immunity characterized by high titers of IgG1, IgG2a and IgA anti-N antibodies, antigen-specific CD8+ T cells and IFN-c-producing CD4+ T cells. Conclusions/Significance: This is the first report of using nanoparticles formed by the recombinant nucleocapsid protein as an efficient and safe intra-nasal vaccine against RSV

Evaluation dans l'espèce cible d'une nouvelle stratégie vaccinale contre le virus respiratoire syncytial bovin, basée sur des assemblages nanoparticulaires de la protéine de nucléocapside

National audienc

Evaluation dans l'espèce cible d'une nouvelle stratégie vaccinale contre le virus respiratoire syncytial bovin, basée sur des assemblages nanoparticulaires de la protéine de nucléocapside

National audienc

Une nouvelle stratégie vaccinale contre le virus respiratoire syncytial, basée sur des assemblages nanoparticulaires de la protéine de nucléocapside

National audienc

Une nouvelle stratégie vaccinale contre le virus respiratoire syncytial, basée sur des assemblages nanoparticulaires de la protéine de nucléocapside

National audienc

T cell-mediated immune response to N SRS.

<p>Nasal vaccination with N SRS generated antigen-specific CD8⁺ T cells and IFN-γ producing CD4⁺ T cells. (A) Antigen-specific proliferation of CD4⁺ and CD8⁺ T splenocytes after 7 days restimulation with RSV-A2 (1 PFU/cell), N (10 µg/ml) or medium. Pooled splenocytes from non immunized (grey lines), LT(R192G) (black lines) or N SRS+LT(R192G) (red lines) immunized groups were stained with CFSE, cultured for 7 days and then labeled with anti-CD8-biot and anti-CD4-PE for flow-cytometry analysis. The data (100,000 events) were acquired with a FACScalibur and analyzed with Cell Quest-Pro. The CD8⁺ or CD4⁺ lymphocyte population was gated according to SSC/FCS and FL4 (CD8) or FL2 (CD4) fluorescence criteria and the fluorescence corresponding to CFSE was monitored in FL1. The percentage of proliferating cells (low CFSE staining) is indicated on the plot with the color corresponding to the immunization condition. (Data from one out of two experiments with similar results). (B) Two weeks after the booster immunization with LT(R192G) or N SRS+LT(R192G), spleen (white bars) and draining LN (cervical and sub-maxilliary LN, black bars) were dissected out and cell suspensions prepared. Splenocytes from individual mice and pooled draining LN cells from each group of mice were re-stimulated for 72 hr with N SRS (10 µg/ml) or medium (mock). IFN-γ secretion was measured in cell culture supernatant with a standardized specific sandwich ELISA assay (white bars represent the mean and SEM of 5 individual spleens, black bars represent the pool of LN, data from one out of three experiments). (C) The frequency of IFN-γ secreting splenocytes after 20 hr restimulation with N (10 µg/ml) was monitored by ELISPOT. Spleen cells from LT(R192G) or N SRS+LT(R192G) immunized mice were assayed for each mouse (each bar represents the mean and SEM of 5 mice). Depletion of CD4⁺ or CD8⁺ T cells was done by immuno-magnetic separation of pooled splenocytes from either LT(R192G) or N SRS+LT(R192G) groups. (Data from one out of two experiments with similar results).</p

Nasal vaccination with N SRS and LT(R192G) augments cellular infiltration in lung tissue.

<p>BALB/c mice were administered i.n. 10 µg N SRS and/or 5 µg LT(R192G), twice at two weeks interval. Two weeks after the second immunization, all animals were challenged with 10⁷ PFU hRSV strain A2, together with a control group of non-immunized mice. One group of mice was neither immunized nor infected. Lung were dissected out 5 days post challenge, embedded in paraffin, sectionned at 7 µm and stained with eosin-hematoxylin. One representative section per group is shown (original magnification 20×, bars 100 µm). (A) control group (no vaccine, no virus), (B) primary infection group (no vaccine, RSV); (C) adjuvant only group (LT(R192G), RSV), (D) N SRS vaccinated group (N SRS+LT(R192G), RSV). Areas with an infiltration of inflammatory cells are indicated with a white arrow. (E) Enlargement showing that the immune infiltrate in N SRS vaccinated group was composed predominantly of lymphocytes and some neutrophils (black arrows) (original magnification 63x, bars 20 µm).</p