lipid-a and Influenza--Human

The discovery and wide spread use of vaccines have saved millions of lives in the past few decades. Vaccine adjuvants represent an integral part of the modern vaccines. Despite numerous efforts, however, only a handful of vaccine adjuvants is currently available for human use. A comprehensive understanding of the mechanisms of action of adjuvants is pivotal to harness the potential of existing and new adjuvants in mounting desirable immune responses to counter human pathogens. Decomposing the host response to vaccines and its components at systems level has recently been made possible owing to the recent advancements in Omics technology and cutting edge immunological assays powered by systems biology approaches. This approach has begun to shed light on the molecular signatures of several human vaccines and adjuvants. This review is an attempt to provide an overview of the recent efforts in systems analysis of vaccine adjuvants that are currently in clinic.

Topics: Adjuvants, Immunologic; alpha-Tocopherol; Animals; Drug Combinations; Glucosides; HIV Infections; Humans; Immunity, Innate; Immunogenicity, Vaccine; Influenza, Human; Lipid A; Liposomes; Malaria, Falciparum; Polysorbates; Squalene; Systems Analysis; Systems Biology; T-Lymphocytes, Helper-Inducer; Toll-Like Receptor 4; Vaccines

This is the second phase 1 study of a respiratory syncytial virus (RSV) vaccine containing RSV fusion protein (sF) adjuvanted with glucopyranosyl lipid A (GLA) in a squalene-based 2% stable emulsion (GLA-SE). In this randomized, double-blind study, 261 subjects aged ≥60 years received inactivated influenza vaccine (IIV), a vaccine containing 120 μg sF with escalating doses of GLA (1, 2.5, or 5 μg) in SE, or a vaccine containing 80 μg sF with 2.5 μg GLA in SE. Subjects receiving 120 μg sF with 2.5 or 5 μg GLA were also randomized to receive IIV or placebo. Immunity to RSV was assessed by detection of microneutralizing, anti-F immunoglobulin G, and palivizumab-competitive antibodies and F-specific gamma interferon enzyme-linked immunosorbent spot assay T-cell responses. Higher adjuvant doses increased injection site discomfort, but at the highest dose, the reactogenicity was similar to that of IIV. Significant humoral and cellular immune responses were observed. The 120 μg sF plus 5.0 μg GLA formulation resulted in the highest responses in all subjects and in older subjects. These results confirm previous observations of vaccine tolerability, safety, and immunogenicity and were used to select the 120 μg sF plus 5.0 μg GLA formulation for phase 2 evaluation. (This study has been registered at ClinicalTrials.gov under registration no. NCT02289820.).

Topics: Adjuvants, Immunologic; Adult; Aged; Aged, 80 and over; Antibodies, Neutralizing; Antibodies, Viral; Dose-Response Relationship, Immunologic; Double-Blind Method; Enzyme-Linked Immunospot Assay; Female; Glucosides; Humans; Immunity, Cellular; Immunity, Humoral; Immunogenicity, Vaccine; Immunoglobulin G; Influenza Vaccines; Influenza, Human; Interferon-gamma; Lipid A; Male; Middle Aged; Respiratory Syncytial Virus Infections; Respiratory Syncytial Virus Vaccines; Respiratory Syncytial Virus, Human; T-Lymphocytes; Viral Fusion Proteins

We conducted a phase I, multicenter, randomized, double-blind, placebo-controlled, multi-arm (10) parallel study involving healthy adults to evaluate the safety and immunogenicity of influenza A (H1N1) 2009 non-adjuvanted and adjuvanted candidate vaccines. Subjects received two intramuscular injections of one of the candidate vaccines administered 21 days apart. Antibody responses were measured by means of hemagglutination-inhibition assay before and 21 days after each vaccination. The three co-primary immunogenicity end points were the proportion of seroprotection >70%, seroconversion >40%, and the factor increase in the geometric mean titer >2.5.. A total of 266 participants were enrolled into the study. No deaths or serious adverse events were reported. The most commonly solicited local and systemic adverse events were injection-site pain and headache, respectively. Only three subjects (1.1%) reported severe injection-site pain. Four 2009 influenza A (H1N1) inactivated monovalent candidate vaccines that met the three requirements to evaluate influenza protection, after a single dose, were identified: 15 μg of hemagglutinin antigen without adjuvant; 7.5 μg of hemagglutinin antigen with aluminum hydroxide, MPL and squalene; 3.75 μg of hemagglutinin antigen with aluminum hydroxide and MPL; and 3.75 μg of hemagglutinin antigen with aluminum hydroxide and squalene.. Adjuvant systems can be safely used in influenza vaccines, including the adjuvant monophosphoryl lipid A (MPL) derived from Bordetella pertussis with squalene and aluminum hydroxide, MPL with aluminum hydroxide, and squalene and aluminum hydroxide.

Topics: Adjuvants, Immunologic; Adult; Aluminum Hydroxide; Antibodies, Viral; Antibody Formation; Double-Blind Method; Female; Hemagglutination Inhibition Tests; Hemagglutinin Glycoproteins, Influenza Virus; Humans; Influenza A Virus, H1N1 Subtype; Influenza Vaccines; Influenza, Human; Injections, Intramuscular; Lipid A; Male; Squalene; Vaccines, Inactivated; Young Adult

To identify an adjuvant that enhances antibody responses in respiratory secretions to inactivated influenza virus vaccine (IVV), a comparison was made of responses to intranasal vaccinations of mice with IVV containing monophosphoryl lipid A (MPL), type I interferon (IFN) or cholera toxin B (CTB). Antibody in nasal secretions and lung wash fluids from mice was increased after vaccination and lung virus was significantly reduced after challenge to a similar level in each adjuvant group. Interferon was selected for a trial in humans. Trivalent inactivated influenza vaccine was given intranasally to healthy adult volunteers alone or with 1 million units (Mu) or 10 Mu of alpha interferon. Vaccinations were well tolerated but neither serum hemagglutination-inhibiting nor neutralizing antibody responses among the vaccine groups were significantly different. Similarly, neither neutralizing nor IgA antibody responses in nasal secretions were significantly different. Thus, despite exhibiting a significant adjuvant effect in mice, interferon did not exhibit an adjuvant effect for induction of antibody in respiratory secretions of humans to inactivated influenza virus vaccine given intranasally.

Topics: Adjuvants, Immunologic; Administration, Intranasal; Adolescent; Adult; Animals; Antibodies, Viral; Cholera Toxin; Humans; Immunity, Mucosal; Influenza A virus; Influenza Vaccines; Influenza, Human; Interferon Type I; Lipid A; Mice; Mice, Inbred ICR; Neutralization Tests; Vaccines, Inactivated; Young Adult

Influenza A virus (IAV) poses a constant worldwide threat to human health. Although conventional vaccines are available, their protective efficacy is type or strain specific, and their production is time-consuming. For the control of an influenza pandemic in particular, agents that are immediately effective against a wide range of virus variants should be developed. Although pretreatment of various Toll-like receptor (TLR) ligands have already been reported to be effective in the defense against subsequent IAV infection, the efficacy was limited to specific subtypes, and safety concerns were also raised. In this study, we investigated the protective effect of an attenuated bacterial outer membrane vesicle -harboring modified lipid A moiety of lipopolysaccharide (fmOMV) against IAV infection and the underlying mechanisms. Administration of fmOMV conferred significant protection against a lethal dose of pandemic H1N1, PR8, H5N2, and highly pathogenic H5N1 viruses; this broad antiviral activity was dependent on macrophages but independent of neutrophils. fmOMV induced recruitment and activation of macrophages and elicited type I IFNs. Intriguingly, fmOMV showed a more significant protective effect than other TLR ligands tested in previous reports, without exhibiting any adverse effect. These results show the potential of fmOMV as a prophylactic agent for the defense against influenza virus infection.

Topics: Animals; Bacterial Outer Membrane; Escherichia coli; Female; Humans; Influenza A virus; Influenza Vaccines; Influenza, Human; Interferon Type I; Ligands; Lipid A; Macrophages; Mice; Mice, Inbred C57BL; Orthomyxoviridae Infections; Secretory Vesicles; Signal Transduction; Toll-Like Receptors

Elderly people are at high risk for influenza-related morbidity and mortality due to progressive immunosenescence. While toll-like receptor (TLR) agonist containing adjuvants, and other adjuvants, have been shown to enhance influenza vaccine-induced protective responses, the mechanisms underlying how these adjuvanted vaccines could benefit the elderly remain elusive. Here, we show that a split H1N1 influenza vaccine (sH1N1) combined with a TLR4 agonist, glucopyranosyl lipid adjuvant formulated in a stable oil-in-water emulsion (GLA-SE), boosts IgG2c:IgG1 ratios, enhances hemagglutination inhibition (HAI) titers, and increases protection in aged mice. We find that all adjuvanted sH1N1 vaccines tested were able to protect both young and aged mice from lethal A/H1N1/California/4/2009 virus challenge after two immunizations compared to vaccine alone. We show that GLA-SE combined with sH1N1, however, also provides enhanced protection from morbidity in aged mice given one immunization (based on change in weight percentage). While the GLA-SE-adjuvanted sH1N1 vaccine promotes the generation of cytokine-producing T helper 1 cells, germinal center B cells, and long-lived bone marrow plasma cells in young mice, these responses were muted in aged mice. Differential

Topics: Adjuvants, Immunologic; Aged; Animals; Antibodies, Viral; Cells, Cultured; Dendritic Cells; Female; Glucosides; Humans; Immunity; Immunization; Influenza A Virus, H1N1 Subtype; Influenza Vaccines; Influenza, Human; Lipid A; Mice; Mice, Inbred C57BL; Orthomyxoviridae Infections; Toll-Like Receptor 4

Annual vaccination is not effective in conferring cross-protection against antigenically different influenza viruses. Therefore, it is of high priority to improve the cross protective efficacy of influenza vaccines. We investigated the adjuvant effects of monophosphoryl lipid A (MPL) and oligodeoxynucleotide CpG (CpG) on promoting homologous protection and cross-protection after vaccination of C57BL/6 and BALB/c mice with inactivated split virus. Combination adjuvant effects of MPL and CpG on improving homologous and cross protective vaccine efficacy were evident as shown by higher levels of homologous and cross-reactive binding IgG and hemagglutination inhibiting antibodies. Combination adjuvant effects on enhancing the protective efficacy against homologous and heterosubtypic virus were demonstrated by less weight loss, lower airway inflammatory disease, and better control of viral loads as well as prevention of inflammatory cytokines and cellular infiltrates. Overall, the findings in this study suggest that a combination adjuvant of different toll-like receptor ligands exhibits a unique pattern of innate and adaptive immune responses, contributing to improved homologous and heterosubtypic cross-protection by inactivated split virion influenza vaccination.

Topics: Adjuvants, Immunologic; Animals; Antibodies, Viral; Body Weight; Cross Protection; Cytokines; Disease Models, Animal; Humans; Immunoglobulin G; Influenza Vaccines; Influenza, Human; Lipid A; Mice, Inbred BALB C; Mice, Inbred C57BL; Oligodeoxyribonucleotides; Vaccines, Inactivated; Vaccines, Subunit; Viral Load

Carbohydrate fatty acid sulphate esters (CFASEs) formulated in a squalane-in-water emulsion are effective adjuvants for humoral responses to a wide range of antigens in various animal species but rise in body temperature and local reactions albeit mild or minimal hampers application in humans. In rabbits, body temperature increased 1°C one day after intramuscular (IM) injection, which returned to normal during the next day. The effect increased with increasing dose of CFASE but not with the number of injections (up to 5). Antigen enhanced the rise in body temperature after booster immunization (P<0.01) but not after priming. Synthetic CFASEs are mixtures of derivatives containing no sulphate, one or multiple sulphate groups and the monosulphate derivatives (CMS) were isolated, incorporated in a squalane in-water emulsion and investigated. In contrast to CFASE, CMS adjuvant did not generate rise in body temperature or local reactions in rabbits immunized with a purified, recombinant malaria chimeric antigen R0.10C. In comparison to alum, CMS adjuvant revealed approximately 30-fold higher antibody titres after the first and >100-fold after the second immunization. In ferrets immunized with 7.5μg of inactivated influenza virus A/H7N9, CMS adjuvant gave 100-fold increase in HAI antibody titres after the first and 25-fold after the second immunisation, which were 10-20-fold higher than with the MF59-like AddaVax adjuvant. In both models, a single immunisation with CMS adjuvant revealed similar or higher titres than two immunisations with either benchmark, without detectable systemic and local adverse effects. Despite striking chemical similarities with monophospholipid A (MPL), CMS adjuvant did not activate human TLR4 expressed on HEK cells. We concluded that the synthetic CMS adjuvant is a promising candidate for poor immunogens and single-shot vaccines and that rise in body temperature, local reactions or activation of TLR4 is not a pre-requisite for high adjuvanticity.

Topics: Adjuvants, Immunologic; Animals; Antibodies, Viral; Body Temperature; Carbohydrates; Drug Compounding; Esters; Fatty Acids; Ferrets; HEK293 Cells; Hemagglutination Inhibition Tests; Humans; Immunity, Humoral; Influenza A Virus, H7N9 Subtype; Influenza Vaccines; Influenza, Human; Injections, Intramuscular; Lipid A; Orthomyxoviridae Infections; Polysorbates; Rabbits; Squalene; Toll-Like Receptor 4; Vaccination

The ongoing threat from Influenza necessitates the development of new vaccine and adjuvant technologies that can maximize vaccine immunogenicity, shorten production cycles, and increase global vaccine supply. Currently, the most successful adjuvants for Influenza vaccines are squalene-based oil-in-water emulsions. These adjuvants enhance seroprotective antibody titers to homologous and heterologous strains of virus, and augment a significant dose sparing activity that could improve vaccine manufacturing capacity. As an alternative to an emulsion, we tested a simple lipid-based aqueous formulation containing a synthetic TLR4 ligand (GLA-AF) for its ability to enhance protection against H5N1 infection. GLA-AF was very effective in adjuvanting recombinant H5 hemagglutinin antigen (rH5) in mice and was as potent as the stable emulsion, SE. Both adjuvants induced similar antibody titers using a sub-microgram dose of rH5, and both conferred complete protection against a highly pathogenic H5N1 challenge. However, GLA-AF was the superior adjuvant in ferrets. GLA-AF stimulated a broader antibody response than SE after both the prime and boost immunization with rH5, and ferrets were better protected against homologous and heterologous strains of H5N1 virus. Thus, GLA-AF is a potent emulsion-free adjuvant that warrants consideration for pandemic influenza vaccine development.

Topics: 1,2-Dipalmitoylphosphatidylcholine; Adjuvants, Immunologic; Animals; Antibody Formation; Dogs; Drug Combinations; Emulsions; Female; Ferrets; Humans; Immunity; Influenza A Virus, H5N1 Subtype; Influenza Vaccines; Influenza, Human; Lipid A; Madin Darby Canine Kidney Cells; Mice; Mice, Inbred C57BL; Orthomyxoviridae Infections; Pandemics; Th1 Cells

The antioxidant N-acetyl-l-cysteine (NAC) had been shown to inhibit replication of seasonal human influenza A viruses. Here, the effects of NAC on H9N2 swine influenza virus-induced acute lung injury (ALI) were investigated in mice. BALB/c mice were inoculated intranasally with 10(7) 50% tissue culture infective doses (TCID(50)) of A/swine/HeBei/012/2008/(H9N2) viruses with or without NAC treatments to induce ALI model. The result showed that pulmonary inflammation, pulmonary edema, MPO activity, total cells, neutrophils, macrophages, TNF-α, IL-6, IL-1β and CXCL-10 in BALF were attenuated by NAC. Moreover, our data showed that NAC significantly inhibited the levels of TLR4 protein and TLR4 mRNA in the lungs. Pharmacological inhibitors of TLR4 (E5564) exerted similar effects like those determined for NAC in H9N2 swine influenza virus-infected mice. These results suggest that antioxidants like NAC represent a potential additional treatment option that could be considered in the case of an influenza A virus pandemic.

Topics: Acetylcysteine; Acute Lung Injury; Animals; Antioxidants; Cytokines; Disease Models, Animal; Humans; Inflammation Mediators; Influenza A Virus, H9N2 Subtype; Influenza, Human; Lipid A; Lung; Male; Mice; Mice, Inbred BALB C; Orthomyxoviridae Infections; Peroxidase; Swine; Toll-Like Receptor 4; Virus Replication

A broad coverage influenza vaccine against multiple viral strains based on the viral nucleoprotein (NP) is a goal pursued by many laboratories. If the goal is to formulate the vaccine with recombinant NP it is essential to count on adjuvants capable of inducing cellular immunity. This work have studied the effect of the monophosphoryl lipid A and trehalose dimycolate, known as the Ribi Adjuvant System (RAS), in the immune response induced in mice immunized with recombinant NP. The NP was formulated with RAS and used to immunize BALB/c mice. Immunizations with NP-RAS increased the humoral and cellular immune responses compared to unadjuvanted NP. The predominant antibody isotype was IgG2a, suggesting the development of a Th1 response. Analysis of the cytokines from mice immunized with NP-RAS showed a significant increase in the production of IFN-g and a decreased production of IL-10 and IL-4 compared to controls without RAS. These results are similar to those usually obtained using Freund’s adjuvant, known to induce Th1 and CTL responses when co-administered with purified proteins, and suggest that a similar approach may be possible to enhance the performance of a T-cell vaccine containing NP.

Topics: Animals; Antibodies, Viral; Cell Wall Skeleton; Cord Factors; Female; Humans; Immunity, Cellular; Immunization; Influenza Vaccines; Influenza, Human; Interferon-gamma; Interleukin-10; Interleukin-4; Lipid A; Mice; Mice, Inbred BALB C; Nucleocapsid Proteins; RNA-Binding Proteins; T-Lymphocytes, Cytotoxic; Th1 Cells; Viral Core Proteins

Recent studies have demonstrated that therapies targeting the innate immune system have the potential to provide transient, non-specific protection from a variety of infectious organisms; however, the potential of enhancing the efficacy of such treatments using nano-scale delivery platforms requires more in depth evaluation. As such, we employed a nanolipoprotein (NLP) platform to enhance the efficacy of innate immune agonists. Here, we demonstrate that the synthetic Toll-like receptor (TLR) agonists monophosphoryl lipid A (MPLA) and CpG oligodeoxynucleotides (CpG) can be readily incorporated into NLPs. Conjugation of MPLA and CpG to NLPs (MPLA:NLP and CpG:NLP, respectively) significantly enhanced their immunostimulatory profiles both in vitro and in vivo compared to administration of agonists alone, as evidenced by significant increases in cytokine production, cell surface expression of activation markers, and upregulation of immunoregulatory genes. Importantly, enhancement of cytokine production by agonist conjugation to NLPs was also observed in primary human dendritic cells. Furthermore, BALB/c mice pretreated with CpG:NLP constructs survived a lethal influenza challenge whereas pretreatment with CpG alone had no effect on survival.

Topics: Animals; Cell Line; Cytokines; Dendritic Cells; Humans; Immunologic Factors; Influenza, Human; Lipid A; Male; Mice; Mice, Inbred BALB C; Nanoparticles; Oligodeoxyribonucleotides

Earlier we showed that the structural requirements for adjuvanticity among the aminoalkyl glucosaminide 4-phosphate (AGP) class of synthetic immunostimulants may be less strict than those for other endotoxic activities, including the induction of nitric oxide synthase in murine macrophages and cytokine production in human whole blood. The known role of nitric oxide and pro-inflammatory cytokines in the activation of host defenses against infection prompted us to examine the ability of certain AGPs to enhance non-specific resistance in mice to Listeria monocytogenes and influenza infections as well as to stimulate the production of pro-inflammatory cytokines in mouse splenocytes, human PBMCs, and human U937 histiocytic lymphoma cells. Intranasal administration of RC-524 or RC-529 to mice 2 days prior to a lethal influenza challenge provided significant protection in each case. Similarly, the intravenous administration of these AGPs induced resistance to L. monocytogenes infection as measured by survival or reduction of bacteria in the spleen. Activation of the innate immune response by AGPs appears to involve activation of Toll-like receptor 4 (TLR4) because RC-524 failed to elicit a protective effect in C3H/HeJ mice which have a defect in TLR4 signaling or induce significant cytokine levels in C3H/HeJ splenocytes. Both AGPs also stimulated pro-inflammatory cytokine release in human cell cultures in a dose-dependent manner.

Topics: Administration, Intranasal; Animals; Glycolipids; Humans; Immunity, Innate; Inflammation; Influenza, Human; Leukocytes, Mononuclear; Lipid A; Listeria monocytogenes; Listeriosis; Lymphoma, Large B-Cell, Diffuse; Membrane Glycoproteins; Mice; Nitric Oxide; Receptors, Cell Surface; Spleen; Toll-Like Receptor 4; Toll-Like Receptors; Tumor Cells, Cultured

Old and young Balb/c mice, 24--26 and 2--4 months old, respectively, were infected with a 0.1 LD50 of influenza A/Taiwan/1/86 (A/H1N1) virus by small particle aerosol. Lung virus titers were determined 4, 6, 8, 12, and 17 days later. Old mice had significantly higher virus titers than young mice (P < 0.05-0.0001) and shed virus up to Day 17, while young mice were free of virus by Day 12. Splenic MHC class I CD8+ CTL activity (P < 0.08--0.001) and IFN-gamma production (0.1-0.008) measured on Days 8, 12, and 17 were significantly lower among old mice than among young mice. Coadministration of liposomal influenza vaccine with monophosphoryl lipid A (MPL) resulted in enhanced CD8+ CTL response and IFN-gamma production among old mice (35 and 12,000 times, respectively). These results demonstrate that MPL stimulates CTL and Th1 cytokines (IFN-gamma) in aged mice and may serve to reverse age-related CD8+ CTL deficiency and reduce severe influenza disease in elderly human populations.

Topics: Adjuvants, Immunologic; Aging; Animals; Disease Models, Animal; Histocompatibility Antigens Class I; Humans; Influenza A virus; Influenza Vaccines; Influenza, Human; Interferon-gamma; Lipid A; Liposomes; Lung; Mice; Mice, Inbred BALB C; T-Lymphocytes, Cytotoxic; Tumor Cells, Cultured