Poly-?-Glutamic Acid Complexed With Alum Induces Cross-Protective Immunity of Pandemic H1N1 Vaccine.
ABSTRACT: The use of a good vaccine adjuvant may induce a higher immunogenicity profile of vaccine antigens. Here, we developed a new adjuvant by combining poly-?-glutamic acid (?-PGA) with alum (PGA/Alum) and investigated its ability to enhance the immunogenicity and the cross-reactive efficacy of pandemic H1N1 (pH1N1) influenza vaccine antigen. PGA/Alum enhanced antigen delivery to draining lymph nodes and antigen-specific immunogenicity in mice using OVA as a model antigen. It also greatly increased OVA-specific antibody production, cytotoxic T lymphocyte (CTL) activity, and antibody-dependent cellular cytotoxicity (ADCC). These abilities of PGA/Alum improved the protective efficacy of pH1N1 vaccine antigen by increasing hemagglutination-inhibition titers, enhancing ADCC and CTL activity, and speeding viral clearance following homologous viral challenge. Importantly, the cross-protective efficacy of pH1N1 vaccine against heterologous viruses [A/Puerto Rico/8/34 (H1N1) and A/Hong Kong/1/1968 (H3N2)] was significantly enhanced by PGA/Alum, and cross-reactive ADCC and CTL activities were observed. Together, our results strongly suggest that PGA/Alum may be a promising vaccine adjuvant for preventing influenza and other infectious diseases.
Project description:In 2009, the global outbreak of an influenza pandemic emphasized the need for an effective vaccine adjuvant. In this study, we examined the efficacy of poly-?-glutamic acid/chitosan (PC) nanogel as an adjuvant for the influenza vaccine. PC nanogel significantly enhanced antigen-specific cross-presentation and cytotoxic T lymphocyte (CTL) activity. Compared with alum, the protective efficacy of the pandemic H1N1 influenza (pH1N1) vaccine was substantially increased by PC nanogel, with increased hemagglutination-inhibition titers, CTL activity, and earlier virus clearance after homologous and heterosubtypic [A/Philippines/2/82 (H3N2)] virus challenges. However, CD8+ T cell-depleted mice displayed no protection against the heterosubtypic virus challenge after immunization with PC nanogel-adjuvanted pH1N1 vaccine. We also observed that using PC nanogel as a vaccine adjuvant had a dose-sparing effect and significantly enhanced the long-lasting protection of the pH1N1 vaccine. Together, these results suggest that PC nanogel is a promising vaccine adjuvant that could broadly prevent influenza virus infection.
Project description:Poly(gamma-glutamic acid) (gamma-PGA) nanoparticles (NPs) have previously been reported as an efficient antigen delivery system with adjuvant activity. In this study, the gene expression in murine bone marrow-derived dendritic cells (DCs) treated with gamma-PGA NPs was examined by oligonucleotide microarray analysis and compared with that in cells treated with other adjuvants. The gene expression of proinflammatory chemokines, cytokines, and costimulatory molecules was upregulated considerably in DCs treated with gamma-PGA NPs. The upregulation pattern was similar to that in DCs treated with lipopolysaccharide (LPS) but not to that in DCs treated with unparticulate gamma-PGA. The activation of DCs by gamma-PGA NPs was confirmed by real-time reverse transcriptase PCR (RT-PCR) analysis of genes related to Toll-like receptor (TLR) signaling. The effect of gamma-PGA NPs on DCs was not annihilated by treatment with polymyxin B, an inhibitor of LPS. Furthermore, the immunization of mice with gamma-PGA NPs carrying ovalbumin (OVA) as an antigen significantly induced antigen-specific CD8(+) T cells and antigen-specific production of interleukin-2, tumor necrosis factor alpha, and gamma interferon from the cells. Such activities of gamma-PGA NPs were more potent than those obtained with immunization with OVA plus aluminum hydroxide or OVA plus complete Freund's adjuvant. These results suggest that gamma-PGA NPs induce a CD8(+) T-cell response by activating innate immunity in a fashion different from that of LPS. Thus, gamma-PGA NPs may be an attractive candidate to be developed further as a vaccine adjuvant.
Project description:This study describes the feasibility and adjuvant mechanism of a degradable emulsion for tuning adaptive immune responses to a vaccine antigen. We featured a mouse model with ovalbumin (OVA) as the antigen to deepen our understanding of the properties of a degradable emulsion-based adjuvant, dubbed PELC, interacting with immune cells and to elucidate their roles in vaccine immunogenicity in vivo. First, we demonstrated that the emulsion, which is stabilized by an amphiphilic bioresorbable polymer, shows degradation in mimic human body conditions and considerable tolerance in vivo. Then, we confirmed the model protein could be loaded into the emulsion and released from the matrix in a sustained manner, subsequently driving the production of antigen-specific antibodies. We also comprehended that PELC not only recruits antigen-presenting cells (APCs) to the injection site but also induces the activation of the recruited APCs and migration to the draining lymph nodes. As an adjuvant for cancer immunotherapy, PELC-formulated OVA could strongly enhance antigen-specific T-cell responses as well as anti-tumor ability with respected to non-formulated OVA, using OVA protein/EG7 cells as a tumor antigen/tumor cell model. Accordingly, our data paved the way for the clinical application of degradable emulsions based on amphiphilic bioresorbable polymers as vaccine adjuvants.
Project description:Single-component adjuvant is prone to eliciting a specific type of Th1 or Th2 response. So, the development of combinatorial adjuvants inducing a robust mixed Th1/Th2 response is a promising vaccination strategy against cancer. Here, we describe a novel combination of aluminum salts (alum), CpG oligodeoxynucleotide (CpG) and innate defense regulator peptide HH2 for improving anti-tumor immune responses. The CpG-HH2 complex significantly enhanced the production of IFN-?, TNF-? and IL-1?, promoted the uptake of antigen and strengthened the activation of p38, Erk1/2 and NF-?B in vitro, compared to CpG or HH2 alone. Immunization with NY-ESO-1 antigen plus alum-CpG-HH2 combinatorial adjuvant effectively inhibited tumor growth and reduced tumor burden in prophylactic and therapeutic tumor models and even in passive serum or cellular therapy. In addition, co-administration of NY-ESO-1 with alum-CpG-HH2 combinatorial adjuvant markedly activated NK cell cytotoxicity, induced antibody-dependent cellular cytotoxicity (ADCC), dramatically elicited cytotoxic T lymphocytes (CTLs) response, and increased infiltrating lymphocytes in tumors. Moreover, in vivo depletion of CD8+ T cells completely and depletion of NK cells partially blocked the anti-tumor activity of NY-ESO-1-alum-CpG-HH2 immunization. Overall, our results demonstrate a novel adjuvant combination for cancer vaccine with efficient immunomodulation by stimulating innate immunity and mediating adaptive immunity.
Project description:Vaccines are arguably one of the greatest advancements in modern medicine. Subunit vaccines comprise the majority of current preparations and consist of two main components-antigen and adjuvant. The antigen is a small molecule against which the vaccine induces an immune response to provide protection via the immunostimulatory ability of the adjuvant. Our laboratory has investigated the adjuvant properties of Toll-like receptor (TLR) ligand-based adjuvants, especially the outer membrane protein from Neisseria mengingitidis, PorB. In this current study we used PorB, along with CpG, an intracellular TLR9 agonist, and a non-TLR adjuvant, aluminum salts (Alum), to further investigate cellular mechanisms of adjuvanticity, focusing on the fate of intact antigen in the germinal center and association with follicular dendritic cells (FDCs). FDCs are located in the B cell light zone of the germinal center and are imperative for affinity maturation. They are stromal cells that retain whole intact antigen allowing recognition by the B cell receptor of the germinal center B cells. Our studies demonstrate that TLR ligands, but not Alum, increase the FDC network, while PorB and Alum increased colocalization of FDC and the model soluble antigen, ovalbumin (OVA). As PorB is the only adjuvant tested that induces both a higher number of FDCs and increased deposition of antigen on FDCs, it has the greatest ability to increase FDC-antigen interaction, essential for induction of B cell affinity maturation. These studies demonstrate a further mechanism and potential superiority of PorB as an adjuvant and its influence on antibody production.
Project description:There is an unmet medical need for new subunit vaccines that induce cytotoxic T-lymphocyte (CTL) responses to prevent infection with a number of pathogens. However, stimulation of CTL responses via clinically acceptable subcutaneous (s.c.) and intramuscular (i.m.) injection is challenging. Recently, we designed a liposomal adjuvant [cationic adjuvant formulation (CAF)09] composed of the cationic lipid dimethyldioctadecylammonium (DDA) bromide, a synthetic monomycoloyl glycerol analog and polyinosinic:polycytidylic acid, which induce strong CTL responses to peptide and protein antigens after intraperitoneal administration. By contrast, CAF09 does not stimulate CTL responses upon s.c. or i.m. injection because the vaccine forms a depot that remains at the injection site. Hence, we engineered a series of nanoemulsions (CAF24a-c) based on the active components of CAF09. The oil phase consisted of biodegradable squalane, and the surface charge was varied systematically by replacing DDA with zwitterionic distearoylphosphoethanolamine. We hypothesized that the nanoemulsions drain to the lymph nodes to a larger extent than CAF09, upon s.c. co-administration with the model antigen chicken egg ovalbumin (OVA). This results in an increased dose fraction that reaches the draining lymph nodes (dLNs) and subsequently activates cross-presenting dendritic cells (DCs), which can prime CTL responses. Indeed, the nanoemulsions induced antigen-specific CD8+ T-cell responses, which were significantly higher than those stimulated by OVA adjuvanted with CAF09. We explain this by the observed rapid localization of CAF24a in the dLNs and the subsequent association with conventional DCs, which promotes induction of CTL responses. Uptake of CAF24a was not specific for DCs, because CAF24a was also detected with B cells and macrophages. No measurable dose fraction of CAF09 was detected in the dLNs within the study period, and CAF09 formed a depot at the site of injection. Importantly, s.c. vaccination with OVA adjuvanted with CAF24a induced significant levels of specific lysis of antigen-pulsed splenocytes were induced after, which was not observed for OVA adjuvanted with CAF09. Thus, CAF24a is a promising adjuvant for induction of CTL responses upon s.c. and i.m. immunization, and it offers interesting perspectives for the design of vaccines against pathogens for which CTL responses are required to prevent infection.
Project description:Aluminum-containing adjuvants usually referred as Alum are considered as T helper type-2 (Th2) adjuvants, while agonists of toll-like receptors (TLRs) are viewed as adjuvants that favor Th1/Th17 immunity. Alum has been used in numerous vaccine formulations; however, its undesired pro-Th2 adjuvant activity constitutes a caveat for Alum-based vaccines. Combining Alum with TLR-dependent, pro-Th1/Th17 adjuvants might dampen the pro-Th2 activity and improve the effectiveness of vaccine formulations. Here, using the ovalbumin (OVA) model of allergic lung inflammation, we found that sensitization with the synthetic TLR9 agonist, which is composed of oligodeoxynucleotides containing CpG motifs adsorbed to Alum, inhibited the development of OVA-induced lung allergic Th2 responses without shifting toward a Th1 pattern. The conversion of T cell immunity from the polarized allergic Th2 response to a non-polarized form by sensitization with OVA/Alum/CpG was dependent on MyD88 signaling in myeloid cells. Notably, sensitization of IL-10-deficient mice with OVA/Alum/CpG resulted in the development of neutrophilic lung inflammation associated with IFN? production. However, in IL-10/IL-12-deficient mice, it resulted in neutrophilic inflammation dominated by IL-17 production. We conclude that OVA/Alum/CpG sensitization signaling via MyD88 and IL-10 molecules results in non-polarized immunity. Conversely, OVA/Alum/CpG sensitization in presence of MyD88 but absence of IL-10 or IL-10/IL-12 molecules results, respectively, in neutrophilic inflammation associated with IFN? or IL-17 production. Our work provides novel OVA models of lung inflammation and suggests that Alum/CpG-based formulations might be of potential use in anti-allergic or anti-infectious processes.
Project description:<h4>Background</h4>Pre-pandemic development of an inactivated, split-virion avian influenza vaccine is challenged by the lack of pre-existing immunity and the reduced immunogenicity of some H5 hemagglutinins compared to that of seasonal influenza vaccines. Identification of an acceptable effective adjuvant is needed to improve immunogenicity of a split-virion avian influenza vaccine.<h4>Methods and findings</h4>Ferrets (N?=?118) were vaccinated twice with a split-virion vaccine preparation of A/Vietnam/1203/2004 or saline either 21 days apart (unadjuvanted: 1.9 µg, 7.5 µg, 30 µg, or saline), or 28 days apart (unadjuvanted: 22.5 µg, or alum-adjuvanted: 22.5 or 7.5 µg). Vaccinated animals were challenged intranasally 21 or 28 days later with 10(6) EID(50) of the homologous strain. Immunogenicity was measured by hemagglutination inhibition and neutralization assays. Morbidity was assessed by observed behavior, weight loss, temperature, cytopenias, histopathology, and viral load. No serum antibodies were detected after vaccination with unadjuvanted vaccine, whereas alum-adjuvanted vaccination induced a robust antibody response. Survival after unadjuvanted dose regimens of 30 µg, 7.5 µg and 1.9 µg (21-day intervals) was 64%, 43%, and 43%, respectively, yet survivors experienced weight loss, fever and thrombocytopenia. Survival after unadjuvanted dose regimen of 22.5 µg (28-day intervals) was 0%, suggesting important differences in intervals in this model. In contrast to unadjuvanted survivors, either dose of alum-adjuvanted vaccine resulted in 93% survival with minimal morbidity and without fever or weight loss. The rarity of brain inflammation in alum-adjuvanted survivors, compared to high levels in unadjuvanted vaccine survivors, suggested that improved protection associated with the alum adjuvant was due to markedly reduced early viral invasion of the ferret brain.<h4>Conclusion</h4>Alum adjuvant significantly improves efficacy of an H5N1 split-virion vaccine in the ferret model as measured by immunogenicity, mortality, morbidity, and brain invasion.
Project description:Vaccination remains the most effective public health tool to prevent infectious diseases. Many vaccines are marginally effective and need enhancement for immunocompromised, elderly, and very young populations. To enhance immunogenicity, we exploited the biphasic property of the (RADA)4 synthetic oligopeptide to create VacSIM (vaccine self-assembling immune matrix), a new delivery method. VacSIM solution can easily be mixed with antigens, organisms, and adjuvants for injection. Postinjection, the peptides self-assemble into hydrated nanofiber gel matrices, forming a depot with antigens and adjuvants in the aqueous phase. We believe the depot provides slow release of immunogens, leading to increased activation of antigen-presenting cells that then drive enhanced immunogenicity. Using recombinant hepatitis B virus surface antigen (rHBsAg) as a model immunogen, we compared VacSIM delivery to delivery in alum or complete Freund's adjuvant (CFA). Delivery of the rHBsAg antigen to mice via VacSIM without adjuvant elicited higher specific IgG responses than when rHBsAg was delivered in alum or CFA. Evaluating IgG subtypes showed a mixed Th1/Th2 type response following immunization with VacSIM, which was driven further toward Th1 with addition of CpG as the adjuvant. Increased specific IgG endpoint titers were observed in both C57BL/6 and BALB/c mice, representative of Th1 and Th2 environments, respectively. Restimulation of splenocytes suggests that VacSIM does not cause an immediate proinflammatory response in the host. Overall, these results suggest that VacSIM, as a new delivery method, has the potential to enhance immunogenicity and efficacy of numerous vaccines.
Project description:The effectiveness of current influenza vaccines is considered suboptimal, and 1 way to improve the vaccines is using adjuvants. However, the current pool of adjuvants used in influenza vaccination is limited due to safety concerns. Aloe vera, or aloe, has been shown to have immunomodulatory functions and to be safe for oral intake. In this study, we explored the potential of orally administered processed Aloe vera gel (PAG) as an adjuvant for influenza vaccines in C57BL/6 mice. We first evaluated its adjuvanticity with a split-type pandemic H1N1 (pH1N1) Ag by subjecting the mice to lethal homologous influenza challenge. Oral PAG administration with the pH1N1 Ag increased survival rates in mice to levels similar to those of alum and MF59, which are currently used as adjuvants in influenza vaccine formulations. Similarly, oral PAG administration improved the survival of mice immunized with a commercial trivalent influenza vaccine against lethal homologous and heterologous virus challenge. PAG also increased hemagglutination inhibition and virus neutralization Ab titers against homologous and heterologous influenza strains following immunization with the split-type pH1N1 Ag or the commercial trivalent vaccine. Therefore, this study demonstrates that PAG may potentially be used as an adjuvant for influenza vaccines.