Evaluation of an Intranasal Virosomal Vaccine against Respiratory Syncytial Virus in Mice: Effect of TLR2 and NOD2 Ligands on Induction of Systemic and Mucosal Immune Responses

Abstract

IntroductionRSV infection remains a serious threat to newborns and the elderly. Currently, there is no vaccine available to prevent RSV infection. A mucosal RSV vaccine would be attractive as it could induce mucosal as well as systemic antibodies, capable of protecting both the upper and lower respiratory tract. Previously, we reported on a virosomal RSV vaccine for intramuscular injection with intrinsic adjuvant properties mediated by an incorporated lipophilic Toll-like receptor 2 (TLR2) ligand. However, it has not been investigated whether this virosomal RSV vaccine candidate would be suitable for use in mucosal immunization strategies and if additional incorporation of other innate receptor ligands, like NOD2-ligand, could further enhance the immunogenicity and protective efficacy of the vaccine.ObjectiveTo explore if intranasal (IN) immunization with a virosomal RSV vaccine, supplemented with TLR2 and/or NOD2-ligands, is an effective strategy to induce RSV-specific immunity.MethodsWe produced RSV-virosomes carrying TLR2 (Pam3CSK4) and/or NOD2 (L18-MDP) ligands. We tested the immunopotentiating properties of these virosomes in vitro, using TLR2- and/or NOD2-ligand-responsive murine and human cell lines, and in vivo by assessing induction of protective antibody and cellular responses upon IN immunization of BALB/c mice.ResultsIncorporation of Pam3CSK4 and/or L18-MDP potentiates the capacity of virosomes to activate (antigen-presenting) cells in vitro, as demonstrated by NF-κB induction. In vivo, incorporation of Pam3CSK4 in virosomes boosted serum IgG antibody responses and mucosal antibody responses after IN immunization. While L18-MDP alone was ineffective, incorporation of L18-MDP in Pam3CSK4-carrying virosomes further boosted mucosal antibody responses. Finally, IN immunization with adjuvanted virosomes, particularly Pam3CSK4/L18-MDP-adjuvanted-virosomes, protected mice against infection with RSV, without priming for enhanced disease.ConclusionMucosal immunization with RSV-virosomes, supplemented with incorporated TLR2- and/or NOD2-ligands, represents a promising approach to induce effective and safe RSV-specific immunity.

Nod2 is required for antigen-specific humoral responses against antigens orally delivered using a recombinant Lactobacillus vaccine platform

“…The critical role of NOD2 for lactobacilli-induced innate immune signaling was further linked to adaptive immune responses using an in vivo mucosal vaccination model. Consistent with other reports, we observed that robust antigen-specific humoral responses against our NCFM-based mucosal vaccines required NOD2 signaling in C57BL/6 and BALB/c mice [51,[53][54][55]. No specific vaccine correlates were identified here, but the significant increase of gut IL-1β in NCK2166 immunized C57BL/6 mice was also dependent on NOD2.…”

“…A growing body of evidence suggests a crucial role for NOD2 and the microbiota in mucosal vaccine responses. Recent studies have demonstrated synergy between NOD2 and TLR2 in the development of antigen-specific humoral responses [51,54]. While TLR2 has been shown to promote IgA class switching and plasma cell differentiation, Kim et al demonstrated the requirement of NOD2 for optimal humoral responses in a mucosal cholera toxin (CT) model [60,61].…”

Signaling via pattern recognition receptors NOD2 and TLR2 contributes to immunomodulatory control of lethal pneumovirus infection

“…A more recent study by Drake and colleagues (2013) extended the antiviral impact of TLR2/6/9 agonists to include parainfluenza virus. Furthermore, Shafique and colleagues (2013) have incorporated Pam 3 CSK 4 and L 18 -MDP into virosomes prepared from the human RSV pathogen, and reported increased systemic and mucosal antibody responses to live virus challenge.…”

Poly(U) and CpG ameliorate the unbalanced T cell immunity and pneumonia of mice with RSV vaccine-enhanced disease

“…Recently some researchers found that the ineffectivness of FIRSV was associated with its insufficient stimulation of host's innate immunity (11)(12)(13). Toll-like receptors (TLRs), which are considered to be doorkeepers in recognition of exogenous pathogens, play a critical role in induction of host's innate immunity (14).…”

“…As one of the most important Th2 cytokines, IL-4 could inhibit Th0 lymphocytes' differentiation into Th1 lymphocytes and promote reconstruction of the respiratory tract and development of pneumonia (26). Besides, IL-17 was reported to increase airway mucus by upregulating expression of the MUC5B gene and cause airway hyperesponsiveness (13,27). As the main source of IL-17, Th17 lymphocytes could also work synergistically with Th2 lymphocytes and inhibit the cytocidal effect of CD8 + T cells, which was consistent with the increased levels of Th2 and Th17 cytokines as well as the suppressed Th1 and CD8 + T cell cytokines of RVED mice in our data (28).…”