BackgroundHepatitis B virus infection remains an important global health concern despite the availability of safe and effective prophylactic vaccines. Limitations to these vaccines include requirement for refrigeration and three immunizations thereby restricting use in the developing world. A new nasal hepatitis B vaccine composed of recombinant hepatitis B surface antigen (HBsAg) in a novel nanoemulsion (NE) adjuvant (HBsAg-NE) could be effective with fewer administrations.Methodology and Principal FindingsPhysical characterization indicated that HBsAg-NE consists of uniform lipid droplets (349+/−17 nm) associated with HBsAg through electrostatic and hydrophobic interactions. Immunogenicity of HBsAg-NE vaccine was evaluated in mice, rats and guinea pigs. Animals immunized intranasally developed robust and sustained systemic IgG, mucosal IgA and strong antigen-specific cellular immune responses. Serum IgG reached ≥106 titers and was comparable to intramuscular vaccination with alum-adjuvanted vaccine (HBsAg-Alu). Normalization showed that HBsAg-NE vaccination correlates with a protective immunity equivalent or greater than 1000 IU/ml. Th1 polarized immune response was indicated by IFN-γ and TNF-α cytokine production and elevated levels of IgG2 subclass of HBsAg-specific antibodies. The vaccine retains full immunogenicity for a year at 4°C, 6 months at 25°C and 6 weeks at 40°C. Comprehensive pre-clinical toxicology evaluation demonstrated that HBsAg-NE vaccine is safe and well tolerated in multiple animal models.ConclusionsOur results suggest that needle-free nasal immunization with HBsAg-NE could be a safe and effective hepatitis B vaccine, or provide an alternative booster administration for the parenteral hepatitis B vaccines. This vaccine induces a Th1 associated cellular immunity and also may provide therapeutic benefit to patients with chronic hepatitis B infection who lack cellular immune responses to adequately control viral replication. Long-term stability of this vaccine formulation at elevated temperatures suggests a direct advantage in the field, since potential excursions from cold chain maintenance could be tolerated without a loss in therapeutic efficacy.

The currently available commercial human anthrax vaccine requires multiple injections for efficacy and has side effects due to its alum adjuvant. These factors limit its utility when immunizing exposed populations in emergent situations. We evaluated a novel mucosal adjuvant that consists of a nontoxic, water-in-oil nanoemulsion (NE). This material does not contain a proinflammatory component but penetrates mucosal surfaces to load antigens into dendritic cells. Mice and guinea pigs were intranasally immunized with recombinant Bacillus anthracis protective antigen (rPA) mixed in NE as an adjuvant. rPA-NE immunization was effective in inducing both serum anti-PA immunoglobulin G (IgG) and bronchial anti-PA IgA and IgG antibodies after either one or two mucosal administrations. Serum anti-PA IgG2a and IgG2b antibodies and PA-specific cytokine induction after immunization indicate a Th1-polarized immune response. rPA-NE immunization also produced high titers of lethal-toxin-neutralizing serum antibodies in both mice and guinea pigs. Guinea pigs nasally immunized with rPA-NE vaccine were protected against an intradermal challenge with ϳ1,000 times the 50% lethal dose (ϳ1,000؋ LD 50 ) of B. anthracis Ames strain spores (1.38 ؋ 10 3 spores), which killed control animals within 96 h. Nasal immunization also resulted in 70% and 40% survival rates against intranasal challenge with 10؋ LD 50 and 100؋ LD 50 (1.2 ؋ 10 6 and 1.2 ؋ 10 7 ) Ames strain spores. Our results indicate that NE can effectively adjuvant rPA for intranasal immunization. This potentially could lead to a needle-free anthrax vaccine requiring fewer doses and having fewer side effects than the currently available human vaccine.

Influenza A viral infection begins by hemagglutinin glycoproteins on the viral envelope binding to cell membrane sialic acid (SA). Free SA monomers cannot block hemagglutinin adhesion in vivo because of toxicity. Polyvalent, generation 4 (G4) SA-conjugated polyamidoamine (PAMAM) dendrimer (G4-SA) was evaluated as a means of preventing adhesion of 3 influenza A subtypes (H1N1, H2N2, and H3N2). In hemagglutination-inhibition assays, G4-SA was found to inhibit all H3N2 and 3 of 5 H1N1 influenza subtype strains at concentrations 32-170 times lower than those of SA monomers. In contrast, G4-SA had no ability to inhibit hemagglutination with H2N2 subtypes or 2 of 5 H1N1 subtype strains. In vivo experiments showed that G4-SA completely prevented infection by a H3N2 subtype in a murine influenza pneumonitis model but was not effective in preventing pneumonitis caused by an H2N2 subtype. Polyvalent binding inhibitors have potential as antiviral therapeutics, but issues related to strain specificity must be resolved.

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