Conventional Influenza Vaccines Research Articles

Impact of a mixed bacterial lysate (OM-85 BV) on the immunogenicity, safety and tolerability of inactivated influenza vaccine in children with recurrent respiratory tract infection

It is known that the immunogenicity and efficacy of conventional inactivated influenza vaccines (IIVs) are not completely satisfactory in children. The aim of this prospective, randomised, single-blind study was to compare the immune response to, and the effectiveness and safety of, an IIV (Fluarix, GlaxoSmithKline Biologicals, Rixensart, Belgium) administered to 68 children aged 36–59 months affected by recurrent respiratory tract infections (RRTIs) who were vaccinated with (n=33) or without (n=35) the mixed bacterial lysate OM-85 BV (Broncho-vaxom, Vifor Pharma, Geneva, Switzerland). OM-85 BV had no effect on seroconversion or seroprotection rates, geometric mean titres, or dendritic cells, which were not significantly different between the two groups. Moreover, OM-85 BV did not significantly increase the pool of the memory B cells that produce IgG and IgM antibodies against the influenza antigens. However, respiratory morbidity was significantly lower in the children treated with OM-85 BV (p<0.05), thus confirming its positive effect on the incidence of RRTIs. There was no difference in the incidence of adverse events between the two groups. These findings show that the immune response of children to influenza vaccine is not significantly influenced by the administration of OM-85 BV. However, the use of OM-85 before and at the same time as IIV seems to reduce respiratory morbidity, and seems to be safe and well tolerated.

Solid Bioneedle-Delivered Influenza Vaccines Are Highly Thermostable and Induce Both Humoral and Cellular Immune Responses

The potential of bioneedles to deliver influenza vaccines was investigated. Four influenza vaccine formulations were screened to determine the optimal formulation for use with bioneedles. The stability of the formulations after freeze-drying was checked to predict the stability of the influenza vaccines in the bioneedles. Subunit, split, virosomal and whole inactivated influenza (WIV) vaccine were formulated and lyophilized in bioneedles, and subsequently administered to C57BL/6 mice. Humoral and cellular immune responses were assessed after vaccination. The thermostability of lyophilized vaccines was determined after one-month storage at elevated temperatures. Bioneedle influenza vaccines induced HI titers that are comparable to those induced by intramuscular WIV vaccination. Delivery by bioneedles did not alter the type of immune response induced by the influenza vaccines. Stability studies showed that lyophilized influenza vaccines have superior thermostability compared to conventional liquid vaccines, and remained stable after one-month storage at 60°C. Influenza vaccines delivered by bioneedles are a viable alternative to conventional liquid influenza vaccines. WIV was determined to be the most potent vaccine formulation for administration by bioneedles. Lyophilized influenza vaccines in bioneedles are independent of a cold-chain, due to their increased thermostability, which makes distribution and stockpiling easier.

High Vaccination Rate Is Crucial

The article by Wutzler et al. (1) on targeted vaccine selection in influenza vaccination is of major importance in view of the imminent flu season. During the 2012/2013 flu season, the pediatric hospital at Hamburg-Eppendorf’s University Medical Center saw three deaths in children who had not been vaccinated in accordance with the recommendations from the Standing Committee on Vaccination Recommendations (STIKO), although they had severe underlying disorders. Wutzler et al say that the influenza rate can be lowered to 8.2% when using the conventional influenza vaccine and to 4.2% when using the attenuated live vaccine, which in theory corresponds to a 92% protective effect for the conventional vaccine compared with 96% for the newer, attenuated live vaccine. Regarding children up to age 4 living in Germany, the following theoretical calculation could be made with regard to mortality resulting from omitting the flu vaccination: number of children in the age group (about 2.6 million, according to Germany’s Federal Statistical Office) × excess hospital admission rate during the flu season 2012/2013 of 140/100 000 (2) × mortality in children with influenza admitted to hospital of 1.0–1.1% (3) × protective effect conferred by vaccination (conventional vaccine 92%, attenuated live vaccine 96%) (1)=2.6 million ×140/100 000 × 0.01 × 0.92 or 0.96 =about 33.5 or 35 children. Although this simplified calculation does not consider several epidemiologically relevant factors—such as contagiousness, virulence, the fact that STIKO recommends vaccination only for children with chronic disorders, and the fact that the attenuated live vaccine has license approval only for children older than 2 years—it still highlights that, in order to minimize the risk, the aim of a vaccination rate that is as high as possible is crucial for pediatric risk groups too, whereas the age-appropriate optimized vaccine yields additional benefit only by degrees. Parents should not be prompted to refuse the vaccination on the basis of insecurities caused by discussions of which might be the better vaccine.

Expression of Avian Influenza Virus (H5N1) Hemagglutinin and Matrix Protein 1 in Pichia pastoris and Evaluation of their Immunogenicity in Mice

The conventional avian influenza vaccines rely on development of neutralizing antibodies against the HA and NA antigens. However, these antigens are highly variable, and hence there is a need for better vaccine candidates which would offer broader protection in animals. The M1 of avian influenza is another major structural protein that has conserved epitopes that are reported to induce CD8+ T cells and can contribute to protection against morbidity and mortality from influenza. Hence in an effort to study the immune response of rM1 either alone or in combination with rHA, the hemagglutinin (HA) and matrix protein (M1) of A/Hatay/2004/H5N1 strain of avian influenza were expressed in Pichia pastoris as his-tagged proteins and purified through Ni-NTA chromatography. The His-tag was removed using TEV protease cleavage site and the immunogenicity of purified rHA and rM1 either alone or in combination was determined in mice. One group of mice was immunized with 5μg of purified rHA, the other group was immunized with rM1, and a third group of mice were immunized with 5μg of rHA and rM1. All the animals were boosted twice, once on 28days postimmunization (dpi) and the second on 42dpi. The immune response was evaluated by enzyme-linked immunosorbent assay (ELISA) and hemagglutination inhibition (HI) assay. The group of mice immunized with rHA and rM1 together showed significantly higher immune response against rHA and rM1 than mice immunized with either HA or M1 antigens. The addition of rM1 with rHA resulted in increased HI titer in animals immunized with both the antigens. These results suggest that the HA and M1 expressed in P. pastoris can be utilized in combination for the development of faster and cost-effective vaccines for circulating and newer strains of avian influenza and would aid in combating the disease in a pandemic situation, in which production time matters greatly.

Sublingual immunization with a subunit influenza vaccine elicits comparable systemic immune response as intramuscular immunization, but also induces local IgA and TH17 responses

Influenza is a vaccine-preventable disease that remains a major health problem world-wide. Needle and syringe are still the primary delivery devices, and injection of liquid vaccine into the muscle is still the primary route of immunization. Vaccines could be more convenient and effective if they were delivered by the mucosal route. Elicitation of systemic and mucosal innate and adaptive immune responses, such as pathogen neutralizing antibodies (including mucosal IgA at the site of pathogen entry) and CD4+ T-helper cells (especially the Th17 subset), have a critical role in vaccine-mediated protection. In the current study, a sublingual subunit influenza vaccine formulated with or without mucosal adjuvant was evaluated for systemic and mucosal immunogenicity and compared to intranasal and intramuscular vaccination. Sublingual administration of adjuvanted influenza vaccine elicited comparable antibody titers to those elicited by intramuscular immunization with conventional influenza vaccine. Furthermore, influenza-specific Th17 cells or neutralizing mucosal IgA were detected exclusively after mucosal immunization.

Survey of Human Antibody Responses to Influenza Virus Matrix Protein 2 by Use of a Sensitive Flow Cytometric Method

Conventional influenza vaccines are designed to elicit antibodies to strainspecific antigens, leaving a public health gap when novel viruses break out unexpectedly. As examples, strain-matched vaccine became available too late in the pandemic of 2009 to protect against the fall wave, and drift viruses are sometimes divergent enough to cause vaccine failure (eg, A/Sydney in 1997). Available vaccines are also inadequate to protect against various zoonotic strains, including avian influenza A(H5N1), influenza A(H7N7), and most recently influenza A(H7N9). For this reason, there is much interest in developing vaccines based on conserved influenza virus features that can provide protection regardless of strain. These are usually designed for influenza A virus, but such vaccines can also be made for influenza B virus. Universal influenza vaccines can induce immune protection dependent upon antibody or T-cell responses or both, and the target antigens explored have included nucleoprotein, matrix proteins (M1 and M2), the hemagglutinin (HA) stem, polymerase PB1, and other antigens, as reviewed previously [1]. M2, the focus of Zhong et al in the current issue of the Journal, has long been known as a target of antibodies that reduce viral replication and spread [2, 3]. Vaccines based on many forms of M2 (fusion proteins, M2 multiple antigenic peptides, peptide conjugates, and M2 expressed from viral vectors) provide protective immunity in animals [4–10], and some have been tested in humans. A clinical trial of recombinant M2 shows that this antigen is immunogenic when administered with adjuvant [11]. Human anti-M2 antibodies induced by natural infection have been reported, but data are scanty. Black et al reported that 6 of 17 pairs of acute-phase and convalescent-phase serum specimens showed increased anti-M2 activity by enzyme-linked immunosorbent assay (ELISA), while 12 of 17 convalescent-phase serum specimens demonstrated some signal by Western blot [12]. In a study by Feng et al, an increase in anti-M2 activity was found for 11 of 24 such serum pairs [13]. In that study, of the antibodies detectable by assay on cell surface tetrameric M2, only a minority also recognized M2e peptide. Thus, the majority of antibodies appeared to be conformational [13]. These antibodies may be biologically very important, so assays on native M2 are needed. However, measuring antibodies to native M2 has presented technical problems, with high background encountered in cell surface ELISA. To overcome these problems, Zhong et al developed a flow cytometric assay (M2-FCA) using a panel of 293FT transfected cell lines (M2-293FT) stably expressing full-length tetrameric forms of M2 from various viral strains [14]. With this sensitive assay, some mouse antibodies recognize strain-specific epitopes and some see epitopes cross-reactive among viral strains [14]. The present article uses the assay to analyze collections of human sera. Transfection efficiency and expression levels are normalized through the use of a positive human serum pool as a control. Unlike most serological assays in which a titer reflects the dilution at which a defined end point is reached, the unit of M2 antibody is defined for a single (1:40) dilution of serum. This allows large numbers of sera to be analyzed within a single run, but does not measure titers or other antibody properties revealed by dilution series. Results for healthy donors of different ages showed that antibodies to M2 were found in a higher percentage of, and at higher levels, in adults aged ≥40 years, compared with younger donors. If anti-M2 antibodies were present, they usually recognized both seasonal and swine-origin M2. Received 5 December 2013; accepted 5 December 2013. Correspondence: Suzanne Epstein, PhD, OCTGT, CBER, FDA, 1401 Rockville Pk, HFM-730, Rockville, MD 20852-1448 (suzanne.epstein@fda.hhs.gov). The Journal of Infectious Diseases Published by Oxford University Press on behalf of the Infectious Diseases Society of America 2013. This work is written by (a) US Government employee(s) and is in the public domain in the US. DOI: 10.1093/infdis/jit813

Targeted vaccine selection in influenza vaccination.

The main target groups for influenza vaccination are the elderly, the chronically ill, infants, and toddlers. Influenza vaccines are needed that suit the immunological particularities of each of these age and risk groups. Recent years have seen the approval of influenza vaccines that are more immunogenic than before, but whose use in Germany is limited by the restriction of reimbursement to a small number of vaccines. The Medline database was selectively searched for pertinent literature. The suboptimal immunogenicity of conventional influenza vaccines that contain inactivated viral cleavage products and subunits can be markedly improved by the use of squalene-based adjuvant systems, by the integration of viral antigens in virosomal particles, or by intradermal administration. The vaccination of elderly persons with a vaccine containing the adjuvant MF59 was found to lower the risk of hospitalization for influenza or pneumonia by 25% compared to vaccination with a trivalent inactivated vaccine (TIV). On the other hand, the adjuvant ASO3 was found to be associated with an up to 17-fold increase in the frequency of narcolepsy among 4- to 18-year-olds. In a prospective study, a virosomal vaccine lowered the frequency of laboratory-confirmed influenza in vaccinated children by 88% compared to unvaccinated children (2 versus 18 cases per 1000 individuals). A live, attenuated influenza vaccine lowered the rate of disease in children up to age 7 by 48% compared to a TIV (4.2% versus 8.1%). The newer vaccines possess improved efficacy when used for primary and booster immunization in certain age and risk groups, and they are superior in this respect to conventional vaccines based on viral cleavage products and subunits. The risk/benefit profiles of all currently available vaccines vary depending on the age group or risk group in which they are used.

Physical and immunogenic stability of spray freeze-dried influenza vaccine powder for pulmonary delivery: Comparison of inulin, dextran, or a mixture of dextran and trehalose as protectants

One of the advantages of dry influenza vaccines over conventional liquid influenza vaccines is that they can be used for alternative routes of administration. Previous studies showed that spray freeze-drying is an excellent technique to prepare vaccine containing powders for pulmonary delivery (J.P. Amorij, V. Saluja, A.H. Petersen, W.L.J. Hinrichs, A. Huckriede, H.W. Frijlink, Pulmonary delivery of an inulin-stabilized influenza subunit vaccine prepared by spray-freeze drying induces systemic, mucosal humoral as well as cell-mediated immune responses in BALB/c mice, Vaccine 25 (2007) 8707–8717; S.A. Audouy, G. van der Schaaf, W.L.J. Hinrichs, H.W. Frijlink, J. Wilschut, A. Huckriede. Development of a dried influenza whole inactivated virus vaccine for pulmonary immunization, Vaccine (2011)). The aim of this study was to investigate the physical and immunogenic stability of spray freeze-dried whole inactivated virus influenza vaccine prepared by using inulin, dextran, and a mixture of dextran and trehalose as protectants. Physical and biochemical characteristics of the vaccine powder were maintained at temperatures up to 30°C for 3months. In addition, in vivo data indicate that also, the immunogenic properties of the vaccine were maintained under these storage conditions. On the other hand, in vivo results also revealed that subtle changes in powder characteristics were induced during storage at 30°C. However, laser diffraction measurements showed that problems associated with these subtle changes can be overcome by using dry powder inhalers with an efficient powder dispersing capacity.

Influenza und Influenzaimpfstoffe für Kinder

Influenza is viral disease with very typical clinical symptoms of sudden onset, high fever, coughing, sore throat, muscle pain and headache. The morbidity is highest for children whereby infants substantially contribute to spreading the disease. Infants and babies are often severely affected and must be hospitalized. The World Health Organization (WHO) now recommends the implementation of routine influenza immunization of healthy children between 6 and 59 months old. As conventional trivalent influenza vaccines (TIV) are not optimal for protection young children more effective vaccines are needed. Since 2011 a live attenuated influenza vaccine has been approved for the age group 2-17 years old which provides children with better protection against influenza than conventional TIV. A further possibility for increased effectiveness is coupling TIV with an adjuvant. A virosomal vaccine is available for children over 6 months old. Antigen enhancement of TIV in infants and young children may lead to better protection rates.

Traditional and New Influenza Vaccines

The challenges in successful vaccination against influenza using conventional approaches lie in their variable efficacy in different age populations, the antigenic variability of the circulating virus, and the production and manufacturing limitations to ensure safe, timely, and adequate supply of vaccine. The conventional influenza vaccine platform is based on stimulating immunity against the major neutralizing antibody target, hemagglutinin (HA), by virus attenuation or inactivation. Improvements to this conventional system have focused primarily on improving production and immunogenicity. Cell culture, reverse genetics, and baculovirus expression technology allow for safe and scalable production, while adjuvants, dose variation, and alternate routes of delivery aim to improve vaccine immunogenicity. Fundamentally different approaches that are currently under development hope to signal new generations of influenza vaccines. Such approaches target nonvariable regions of antigenic proteins, with the idea of stimulating cross-protective antibodies and thus creating a "universal" influenza vaccine. While such approaches have obvious benefits, there are many hurdles yet to clear. Here, we discuss the process and challenges of the current influenza vaccine platform as well as new approaches that are being investigated based on the same antigenic target and newer technologies based on different antigenic targets.

Involvement of the Aim2 inflammasome pathway in generating antibody responses elicited by DNA vaccination. (P4313)

Abstract Recent human study data suggests that DNA priming immunization can substantially enhance the protective antibody responses elicited by conventional influenza vaccines, but their mechanisms remain completely unknown. The current study explored the roles of innate immunity in regulating acquired immunity elicited by influenza vaccines. Innate pattern recognition receptors (PRRs), and their downstream signaling molecules, were investigated for their effect on DNA vaccination. Of particular interest is the absent in melanoma 2 (AIM2) protein pathway and its effects on the antigen-specific antibody response. AIM2 is a cytosolic DNA sensor important in both the maturation of the pro-inflammatory cytokines, IL-1b, IL-18, and an inflammatory form of cell death known as pyroptosis. Aim2 knockout mice and wild type controls were immunized with a DNA vaccine expressing the hemagglutinin antigen (HA) of H1 subtype influenza virus. Mouse serum anti-H1HA antibodies and B cell responses revealed that deletion of the Aim2 pathway led to a significant reduction of the anti-HA antibody response. Interestingly, non-antigen specific cytokine profile analysis did not demonstrate a dramatic change in cytokine signaling between wild type and knockout mice. Furthermore, deletion of IL-1 signaling did not significantly affect the anti-HA antibody responses. These results suggested that the Aim2 pathway plays an important role in shaping the antigen specific antibody response to DNA vaccines.

Fluad®-MF59®-Adjuvanted Influenza Vaccine in Older Adults.

Influenza directly or indirectly contributes to the four leading causes of global mortality, at rates that are highest in older adults. As the proportion of older adults in the Korean population is greater than in most other countries, influenza prevention is a greater public health priority in Korea than elsewhere. Conventional inactivated influenza vaccine (IIV) is less immunogenic and efficacious (-50%) in older than in young adults, but adjuvanting the vaccine with oil-in-water emulsion MF59® increases immunogenicity, resulting in comparatively higher levels of hemagglutination inhibition antibodies and greater protection against all influenza, as well as cases requiring hospitalization. A recent observational study demonstrated that the adjuvanted vaccine protected older adults against influenza in a year when nonadjuvanted IIV was ineffective. In another multiyear study, the adjuvanted vaccine was estimated to be 25% more effective in preventing pneumonia and influenza hospitalizations compared to nonadjuvanted vaccine. Although MF59-adjuvanted vaccine is transiently more reactogenic than nonadjuvanted vaccine, there is no evidence that it increases risks for serious adverse events, including those with an autoimmune etiology. Experience thus far indicates a favorable balance of benefit to risk for MF59. This may reflect the adjuvant's mechanism of action in which the squalene oil emulsion increases antibody responses to co-administered antigen without acting more generally as an immunopotentiator.

The age distribution of mortality due to influenza: pandemic and peri-pandemic

BackgroundPandemic influenza is said to 'shift mortality' to younger age groups; but also to spare a subpopulation of the elderly population. Does one of these effects dominate? Might this have important ramifications?MethodsWe estimated age-specific excess mortality rates for all-years for which data were available in the 20th century for Australia, Canada, France, Japan, the UK, and the USA for people older than 44 years of age. We modeled variation with age, and standardized estimates to allow direct comparison across age groups and countries. Attack rate data for four pandemics were assembled.ResultsFor nearly all seasons, an exponential model characterized mortality data extremely well. For seasons of emergence and a variable number of seasons following, however, a subpopulation above a threshold age invariably enjoyed reduced mortality. 'Immune escape', a stepwise increase in mortality among the oldest elderly, was observed a number of seasons after both the A(H2N2) and A(H3N2) pandemics. The number of seasons from emergence to escape varied by country. For the latter pandemic, mortality rates in four countries increased for younger age groups but only in the season following that of emergence. Adaptation to both emergent viruses was apparent as a progressive decrease in mortality rates, which, with two exceptions, was seen only in younger age groups. Pandemic attack rate variation with age was estimated to be similar across four pandemics with very different mortality impact.ConclusionsIn all influenza pandemics of the 20th century, emergent viruses resembled those that had circulated previously within the lifespan of then-living people. Such individuals were relatively immune to the emergent strain, but this immunity waned with mutation of the emergent virus. An immune subpopulation complicates and may invalidate vaccine trials. Pandemic influenza does not 'shift' mortality to younger age groups; rather, the mortality level is reset by the virulence of the emerging virus and is moderated by immunity of past experience. In this study, we found that after immune escape, older age groups showed no further mortality reduction, despite their being the principal target of conventional influenza vaccines. Vaccines incorporating variants of pandemic viruses seem to provide little benefit to those previously immune. If attack rates truly are similar across pandemics, it must be the case that immunity to the pandemic virus does not prevent infection, but only mitigates the consequences.

MHC Class I-Presented T Cell Epitopes Identified by Immunoproteomics Analysis Are Targets for a Cross Reactive Influenza-Specific T Cell Response

Influenza virus infection and the resulting complications are a significant global public health problem. Improving humoral immunity to influenza is the target of current conventional influenza vaccines, however, these are generally not cross-protective. On the contrary, cell-mediated immunity generated by primary influenza infection provides substantial protection against serologically distinct viruses due to recognition of cross-reactive T cell epitopes, often from internal viral proteins conserved between viral subtypes. Efforts are underway to develop a universal flu vaccine that would stimulate both the humoral and cellular immune responses leading to long-lived memory. Such a universal vaccine should target conserved influenza virus antibody and T cell epitopes that do not vary from strain to strain. In the last decade, immunoproteomics, or the direct identification of HLA class I presented epitopes, has emerged as an alternative to the motif prediction method for the identification of T cell epitopes. In this study, we used this method to uncover several cross-specific MHC class I specific T cell epitopes naturally presented by influenza A-infected cells. These conserved T cell epitopes, when combined with a cross-reactive antibody epitope from the ectodomain of influenza M2, generate cross-strain specific cell mediated and humoral immunity. Overall, we have demonstrated that conserved epitope-specific CTLs could recognize multiple influenza strain infected target cells and, when combined with a universal antibody epitope, could generate virus specific humoral and T cell responses, a step toward a universal vaccine concept. These epitopes also have potential as new tools to characterize T cell immunity in influenza infection, and may serve as part of a universal vaccine candidate complementary to current vaccines.

Acceptability of Intanza® 15 μg intradermal influenza vaccine in Belgium during the 2010–2011 influenza season

Intradermal (ID) influenza vaccination induces an enhanced immune response in the elderly when compared with intramuscular (IM) vaccination. In 2009, an ID seasonal influenza vaccine (Intanza(®) [Sanofi Pasteur MSD, Lyon, France] 15 μg) was approved for use in individuals aged ≥ 60 years in Europe. This survey conducted in Belgium was the first in Europe to assess the acceptability of this vaccine in routine clinical practice by vaccinees and their general practitioners (GPs). GPs willing to use both the ID and IM influenza vaccines were selected during the 2010-2011 influenza season. Each GP recruited ≤ 10 patients aged ≥ 60 years who received the ID vaccine. Vaccinees and GPs completed questionnaires about their opinions on influenza vaccination and the acceptability of the ID influenza vaccine. In total, 105 GPs and 837 vaccinees completed questionnaires. A high proportion of vaccinees (40.3%) was aged ≥ 75 years, and 95.5% had been vaccinated the previous year. The majority of vaccinees was very satisfied (70.0%) or satisfied (27.9%) with the ID vaccine. The main reasons for the high satisfaction rate were that the injection was not very painful, administration was quick, and the vaccinee felt confident about the micro-needle injection system. Most vaccinees (91.1%) who had previously received IM influenza vaccination preferred the ID vaccine, and 98.5% of vaccinees reported they would consider receiving the ID vaccine the following year. The majority of GPs was very satisfied (78.6%) or satisfied (18.4%) with the ID vaccine, and most GPs (87.6%) expressed a preference for the ID vaccine over IM influenza vaccine. The ID influenza vaccine was well accepted by vaccinees and their GPs, who expressed a preference for the ID vaccine over conventional IM influenza vaccine. The availability of the ID influenza vaccine may help to improve uptake of seasonal influenza vaccination in the elderly.

In contrast to conventional inactivated influenza vaccines, 4xM2e.HSP70c fusion protein fully protected mice against lethal dose of H1, H3 and H9 influenza A isolates circulating in Iran

Ideal vaccines against influenza viruses should elicit not only a humoral response, but also a cellular response. Mycobacterium tuberculosis HSP70 (mHSP70) have been found to promote immunogenic APCs function, elicit a strong cytotoxic T lymphocyte (CTL) response, and prevent the induction of tolerance. Moreover, it showed linkage of antigens to the C-terminus of mHSP70 (mHSP70c) can represent them as vaccines resulted in more potent, protective antigen specific responses in the absence of adjuvants or complex formulations. Hence, recombinant fusion protein comprising C-terminus of mHSP70 genetically fused to four tandem repeats of the ectodomain of the conserved influenza matrix protein M2 (M2e) was expressed in Escherichia coli, purified under denaturing condition, refolding, and then confirmed by SDS–PAGE, respectively. The recombinant fusion protein, 4xM2e.HSP70c, retained its immunogenicity and displayed the protective epitope of M2e by ELISA and FITC assays. A prime-boost administration of 4xM2e.HSP70c formulated in F105 buffer by intramuscular route in mice (Balb/C) provided full protection against lethal dose of mouse-adapted H1N1, H3N2, or H9N2 influenza A isolates from Iran compared to 0–33.34% survival rate of challenged unimmunized and immunized mice with the currently in use conventional vaccines designated as control groups. However, protection induced by immunization with 4xM2e.HSP70c failed to prevent weight loss in challenged mice; they experienced significantly lower weight loss, clinical symptoms and higher lung viral clearance in comparison with protective effects of conventional influenza vaccines in challenged mice. These data demonstrate that C-terminal domain of mHSP70 can be a superior candidate to deliver the adjuvant function in M2e-based influenza A vaccine in order to provide significant protection against multiple influenza A virus strains.

Involvement of Type I Interferon pathway in generating antibody responses elicited by DNA vaccination. (52.7)

Abstract Recent human study data suggests that DNA immunization can substantially enhance the protective antibody responses elicited by conventional influenza vaccines, but the mechanism of such actions is completely unknown. The current study was designed to explore the roles of innate immunity in regulating acquired immunity elicited by influenza vaccines. Innate pattern recognition receptors (PRRs), and their downstream signaling molecules, were investigated for their effect on DNA vaccination. Of particular interest was the type I interferon (IFN) pathway and its effects on the antigen-specific antibody response. Wild type C57BL/6 mice and selected Interferon Regulatory Factors (IRFs) knockout mice were immunized with DNA vaccines expressing the hemagglutinin antigen (HA) of H1 subtype influenza virus. Mouse serum anti-H1HA antibodies and non-antigen specific cytokine profile analysis showed that deletion of the IRF3 pathway alone did not result in a significantly decreased antibody response but deletion of both the IRF3 and IRF7 pathways led to a dramatic reduction of the anti-HA antibody response. Interestingly, deletion of IL-1 signaling did not significantly affect the anti-HA antibody responses. These results suggested that the IFN pathway does play an important role in shaping the antigen specific antibody response to DNA vaccines and may require the involvement of more than one key regulatory molecule.

Immunogenicity, Safety and Reactogenicity of a Mammalian Cell-Culture–Derived Influenza Vaccine in Healthy Children and Adolescents Three to Seventeen Years of Age

The safety and immunogenicity of the cell-culture-derived seasonal trivalent influenza vaccine ([CCIV]; Optaflu) has been reported previously in adults and the elderly. In this study, we compared the safety, reactogenicity and immunogenicity of CCIV with a conventional egg-derived trivalent influenza vaccine (TIV) in a healthy pediatric population. A total of 3604 subjects were randomized to receive 2 doses of CCIV or TIV (3-8 years, n = 2630) at a 28-day interval or a single vaccination (9-17 years, n = 974). Antibody levels on days 1, 29 and 50 were measured by hemaglutination inhibition assay using egg-derived and cell-derived test antigens. Adverse reactions were solicited via memory aids for 7 days after each injection, and unsolicited adverse events/serious adverse events were collected for 6 months postvaccination. Noninferiority of CCIV versus TIV was demonstrated for most immunogenicity measures, particularly by using cell-derived antigen in the hemaglutination inhibition assay. In 3- to 8-year-olds (the primary objective), both CCIV and TIV met all 3 Committee for Medicinal Products for Human Use immunogenicity criteria for A/H1N1 and A/H3N2 strains. Lower immune responses were observed against the B strain, fulfilling Committee for Medicinal Products for Human Use criteria only for geometric mean ratio (TIV, CCIV) and seroconversion rate (TIV, CCIV [cell-derived antigen]). Both CCIV and TIV were safe and well tolerated, with no differences in local and systemic solicited reactions or in unsolicited adverse events/serious adverse events. CCIV produced in mammalian cell culture is a safe, well-tolerated and immunogenic alternative to conventional egg-derived influenza vaccines for children and adolescents.

Seguridad de la vacuna antigripal con el adyuvante MF59. Revisión sistemática de la literatura

ObjectiveTo assess the safety of the MF59-adjuvanted influenza vaccine in adults versus conventional influenza vaccines. DesignSystematic literature review. Data sourcesMEDLINE, EMBASE, CRD (Center for Review Dissemination), ECRI, Hayes, Health Technology Assessment Agencies databases, the USA clinical trial register ClinicalTrials.gov, and manual search of specialized journals. MeSH terms were “influenza”, “vaccine”, “adults”, “safety”, and free terms were “adjuvant vaccine” “Chiromas”, “Fluad”, and “MF59”. MethodsInclusion criteria were studies including adults vaccinated with MF59-adjuvanted influenza vaccine, compared with conventional vaccines or placebo group, and including results on vaccine safety. The quality of studies included studies was assessed using the CASPe checklist. ResultsWe selected 12 clinical trials with moderate-high quality. Local and systemic adverse effects were scarce and mild. There were no significant differences (P<.01) between the vaccines tested for most reactions. The most frequent and significant adverse effects of the adjuvant vaccine were pain (25.5%), heat (19.9%), local induration (11.2%), chills (8.9%), fever (7.8%), and myalgia (7.5%). ConclusionsThe MF59-adjuvanted vaccine has a good safety profile. The local and systemic adverse effects that may result are typical of any injected vaccine and their frequency is similar to that of conventional influenza vaccines.

Pandemic H1N1 Influenza Infection and Vaccination in Humans Induces Cross-Protective Antibodies that Target the Hemagglutinin Stem

Most monoclonal antibodies (mAbs) generated from humans infected or vaccinated with the 2009 pandemic H1N1 (pdmH1N1) influenza virus targeted the hemagglutinin (HA) stem. These anti-HA stem mAbs mostly used IGHV1-69 and bound readily to epitopes on the conventional seasonal influenza and pdmH1N1 vaccines. The anti-HA stem mAbs neutralized pdmH1N1, seasonal influenza H1N1 and avian H5N1 influenza viruses by inhibiting HA-mediated fusion of membranes and protected against and treated heterologous lethal infections in mice with H5N1 influenza virus. This demonstrated that therapeutic mAbs could be generated a few months after the new virus emerged. Human immunization with the pdmH1N1 vaccine induced circulating antibodies that when passively transferred, protected mice from lethal, heterologous H5N1 influenza infections. We observed that the dominant heterosubtypic antibody response against the HA stem correlated with the relative absence of memory B cells against the HA head of pdmH1N1, thus enabling the rare heterosubtypic memory B cells induced by seasonal influenza and specific for conserved sites on the HA stem to compete for T-cell help. These results support the notion that broadly protective antibodies against influenza would be induced by successive vaccination with conventional influenza vaccines based on subtypes of HA in viruses not circulating in humans.