Type III hypersensitivity reactions to a B cell epitope antigen are abrogated using a depot forming vaccine platform.

Peptide antigens are combined with an adjuvant in order to increase immunogenicity in vivo. The immunogenicity and safety of a RSV vaccine formulated in a novel oil-based platform, DepoVax™ (DPX), was compared to an alum formulation. A peptide B cell epitope derived from RSV small hydrophobic ectodomain (SHe) served as the antigen. Both vaccines induced SHe-specific antibodies after immunization of mice. A single dose of the DPX-based formulation resulted in anti-SHe titres for up to 20 weeks. Boosting with Alum-SHe, but not with DPX-SHe, led to unexpected clinical signs such as decreased activity, cyanosis and drop in body temperature in mice but not in rabbits. The severity of adverse reactions correlated with magnitude of SHe-specific IgG immune responses and decreased complement component 3 plasma levels, indicating a type III hypersensitivity reaction. By RP-HPLC analysis, we found that only 8-20% of the antigen was found to be adsorbed to alum in vitro, indicating that this antigen is likely released systemically upon injection in vivo. Clinical signs were not observed in rabbits, indicating the response correlates with peptide dose relative to size of animal. These results suggest that peptide antigens targeted to produce B cell mediated response may result in increased incidence of type III hypersensitivity reactions when delivered in non-depot forming vaccines. The DPX formulation induced strong antibody titres to the antigen without causing adverse events, likely due to the strength of the depot in vivo, and demonstrates the potential safety and immunogenicity of this platform for B cell peptide antigens.

The GMZ2 malaria vaccine: from concept to efficacy in humans.

INTRODUCTION: GMZ2 is a recombinant protein consisting of conserved domains of GLURP and MSP3, two asexual blood-stage antigens of Plasmodium falciparum, and is designed with the aim of mimicking naturally acquired anti-malarial immunity. The rationale for combining these two antigens is based on a series of immune epidemiological studies from geographically diverse malaria endemic regions; functional in vitro studies; and pre-clinical studies in rodents and New World monkeys. GMZ2 adjuvanted with alhydrogel® (alum) was well tolerated and immunogenic in three phase 1 studies. The recently concluded phase 2 trial of GMZ2/alum, involving 1849 participants 12 to 60 month of age in four countries in West, Central and Eastern Africa, showed that GMZ2 is well tolerated and has some, albeit modest, efficacy in the target population. Areas covered: PubMed ( www.ncbi.nlm.nih.gov/pubmed ) was searched to review the progress and future prospects for clinical development of GMZ2 sub-unit vaccine. We will focus on discovery, naturally acquired immunity, functional activity of specific antibodies, sequence diversity, production, pre-clinical and clinical studies. Expert commentary: GMZ2 is well tolerated and has some, albeit modest, efficacy in the target population. More immunogenic formulations should be developed.

Vaccination strategies against Zika virus.

The epidemic emergence of Zika virus (ZIKV) in 2015-2016 has been associated with congenital malformations and neurological sequela. Current efforts to develop a ZIKV vaccine build on technologies that successfully reduced infection or disease burden against closely related flaviviruses or other RNA viruses. Subunit-based (DNA plasmid and modified mRNA), viral vectored (adeno- and measles viruses) and inactivated viral vaccines are already advancing to clinical trials in humans after successful mouse and non-human primate studies. Among the greatest challenges for the rapid implementation of immunogenic and protective ZIKV vaccines will be addressing the potential for exacerbating Dengue virus infection or causing Guillain-Barré syndrome through production of cross-reactive immunity targeting related viral or host proteins. Here, we review vaccine strategies under development for ZIKV and the issues surrounding their usage.

M2-based influenza vaccines: recent advances and clinical potential.

INTRODUCTION: Current influenza vaccines can prevent disease caused by influenza viruses but require annual administration and almost yearly reformulation. An attractive alternative approach would be to use a vaccine that provides broad and, ideally, lifelong protection against all influenza A and B virus strains. The extracellular domain of matrix protein 2 (M2e) of influenza A viruses is conserved and thus fits well in such a broadly protective vaccine. Areas covered: Recent advances in M2e vaccine design, the mode of action of M2e-based immunity and clinical progress of M2-based influenza vaccines. Expert commentary: Many M2e vaccine have been successfully tested for efficacy against a panel of divergent influenza viruses in animal models. More recently, clinical studies have been conducted with M2e vaccine candidates, which demonstrated their safety and immunogenicity in humans. Efficacy studies in humans are still needed to provide evidence that an M2e-based vaccine can protect against human influenza.

Live virus vaccines based on a vesicular stomatitis virus (VSV) backbone: Standardized template with key considerations for a risk/benefit assessment.

The Brighton Collaboration Viral Vector Vaccines Safety Working Group (V3SWG) was formed to evaluate the safety of live, recombinant viral vaccines incorporating genes from heterologous viral and other microbial pathogens in their genome (so-called "chimeric virus vaccines"). Many such viral vector vaccines are now at various stages of clinical evaluation. Here, we introduce an attenuated form of recombinant vesicular stomatitis virus (rVSV) as a potential chimeric virus vaccine for HIV-1, with implications for use as a vaccine vector for other pathogens. The rVSV/HIV-1 vaccine vector was attenuated by combining two major genome modifications. These modifications acted synergistically to greatly enhance vector attenuation and the resulting rVSV vector demonstrated safety in sensitive mouse and non-human primate neurovirulence models. This vector expressing HIV-1 gag protein has completed evaluation in two Phase I clinical trials. In one trial the rVSV/HIV-1 vector was administered in a homologous two-dose regimen, and in a second trial with pDNA in a heterologous prime boost regimen. No serious adverse events were reported nor was vector detected in blood, urine or saliva post vaccination in either trial. Gag specific immune responses were induced in both trials with highest frequency T cell responses detected in the prime boost regimen. The rVSV/HIV-1 vector also demonstrated safety in an ongoing Phase I trial in HIV-1 positive participants. Additionally, clinical trial material has been produced with the rVSV vector expressing HIV-1 env, and Phase I clinical evaluation will initiate in the beginning of 2016. In this paper, we use a standardized template describing key characteristics of the novel rVSV vaccine vectors, in comparison to wild type VSV. The template facilitates scientific discourse among key stakeholders by increasing transparency and comparability of information. The Brighton Collaboration V3SWG template may also be useful as a guide to the evaluation of other recombinant viral vector vaccines.

Development of a recombinant, chimeric tetravalent dengue vaccine candidate.

Dengue is a significant threat to public health worldwide. Currently, there are no licensed vaccines available for dengue. Takeda Vaccines Inc. is developing a live, attenuated tetravalent dengue vaccine candidate (TDV) that consists of an attenuated DENV-2 strain (TDV-2) and three chimeric viruses containing the prM and E protein genes of DENV-1, -3 and -4 expressed in the context of the attenuated TDV-2 genome backbone (TDV-1, TDV-3, and TDV-4, respectively). TDV has been shown to be immunogenic and efficacious in nonclinical animal models. In interferon-receptor deficient mice, the vaccine induces humoral neutralizing antibody responses and cellular immune responses that are sufficient to protect from lethal challenge with DENV-1, DENV-2 or DENV-4. In non-human primates, administration of TDV induces innate immune responses as well as long lasting antibody and cellular immunity. In Phase 1 clinical trials, the safety and immunogenicity of two different formulations were assessed after intradermal or subcutaneous administration to healthy, flavivirus-naïve adults. TDV administration was generally well-tolerated independent of dose and route. The vaccine induced neutralizing antibody responses to all four DENV serotypes: after a single administration of the higher formulation, 24-67%% of the subjects seroconverted to all four DENV and >80% seroconverted to three or more viruses. In addition, TDV induced CD8(+) T cell responses to the non-structural NS1, NS3 and NS5 proteins of DENV. TDV has been also shown to be generally well tolerated and immunogenic in a Phase 2 clinical trial in dengue endemic countries in adults and children as young as 18 months. Additional clinical studies are ongoing in preparation for a Phase 3 safety and efficacy study.

Human type 5 adenovirus-based tuberculosis vaccine: is the respiratory route of delivery the future?

Despite progress in managing TB, there were 8.6 million new cases in 2012. To control TB will require a more effective vaccine than BCG, new drugs and better diagnostic tests. Recombinant replication-defective adenoviruses expressing foreign DNA have been studied as vaccines. We developed and evaluated a recombinant replication-deficient human Ad5 vector expressing Ag85A (Ad5Ag85A) as a TB vaccine in animal models and a Phase I human study. Animal models of Ad5Ag85A show markedly improved protection over BCG alone and immunization via the respiratory route provides the best type of protection. In humans, intramuscular vaccination was safe; Ad5Ag85A was immunogenic and stimulated polyfunctional T cell responses, more potently in previously BCG-vaccinated volunteers. Pre-existing Ad5 antibodies did not dampen the response. Given its potency, Ad5-based TB vaccines are well-positioned to be delivered to the respiratory tract, induce local lung immunity to control TB, and inform innovative approaches to new TB vaccination strategies.

Safety, immunogenicity, and antibody persistence following an investigational Streptococcus pneumoniae and Haemophilus influenzae triple-protein vaccine in a phase 1 randomized controlled study in healthy adults.

We investigated a protein-based nontypeable Haemophilus influenzae (NTHi) and pneumococcal (HiP) vaccine containing pneumococcal histidine triad D (PhtD), detoxified pneumolysin (dPly), and NTHi protein D (PD) in adults. In a phase I study, 40 healthy 18- to 40-year-old subjects were randomized (2:2:1) to receive two HiP doses administered 60 days apart, with or without AS03 adjuvant (HiP-AS and HiP groups, respectively), or Engerix B (GlaxoSmithKline, Belgium) as a control. Safety, antibodies, and antigen-specific CD4(+) T-cell immune responses were assessed before and until 480 days after vaccination. No serious adverse events were reported, and no subject withdrew due to an adverse event. Local and systemic symptoms were reported more frequently in the HiP-AS group than in the other two groups. The frequency and intensity of local and systemic symptoms appeared to increase after the second dose of HiP-AS or HiP but not Engerix B. Antibody geometric mean concentrations (GMCs) for PhtD, dPly, and PD increased after each dose of HiP-AS or HiP, with higher GMCs being observed in the HiP-AS group (statistically significant for anti-PD after dose 1 and anti-Ply after dose 2). GMCs remained higher at day 420 than prior to vaccination in both the HiP-AS and HiP groups. Antigen-specific CD4(+) T cells increased after each dose but were unmeasurable by day 480. Two doses of an investigational PhtD-dPly-PD protein vaccine induced humoral immunity and antigen-specific CD4(+) T-cell responses after each dose, with generally higher responses when the vaccine was administered with AS03. HiP combined with AS03 appeared to be more reactogenic than the antigens alone. (This study has been registered at ClinicalTrials.gov under registration no. NCT00814489.).

Preclinical and clinical development of a YFV 17 D-based chimeric vaccine against West Nile virus.

Substantial success has been achieved in the development and implementation of West Nile (WN) vaccines for horses; however, no human WN vaccines are approved. This review focuses on the construction, pre-clinical and clinical characterization of ChimeriVax-WN02 for humans, a live chimeric vaccine composed of a yellow fever (YF) 17D virus in which the prM-E envelope protein genes are replaced with the corresponding genes of the WN NY99 virus. Pre-clinical studies demonstrated that ChimeriVax-WN02 was significantly less neurovirulent than YF 17D in mice and rhesus and cynomolgus monkeys. The vaccine elicited neutralizing antibody titers after inoculation in hamsters and monkeys and protected immunized animals from lethal challenge including intracerebral inoculation of high dose of WN NY99 virus. Safety, viremia and immunogenicity of ChimeriVax-WN02 were assessed in one phase I study and in two phase II clinical trials. No safety signals were detected in the three clinical trials with no remarkable differences in incidence of adverse events (AEs) between vaccine and placebo recipients. Viremia was transient and the mean viremia levels were low. The vaccine elicited strong and durable neutralizing antibody and cytotoxic T cell responses. WN epidemiology impedes a classical licensure pathway; therefore, innovative licensure strategies should be explored.

Safety and immunogenicity of a Sf9 insect cell-derived respiratory syncytial virus fusion protein nanoparticle vaccine.

OBJECTIVE: We performed a Phase 1 randomized, observer-blinded, placebo-controlled trial to evaluate the safety and immunogenicity of a recombinant respiratory syncytial virus (RSV) fusion (F) protein nanoparticle vaccine. METHODS: Six formulations with (5, 15, 30 and 60 µg) and without (30 and 60 µg) aluminum phosphate (AdjuPhos) were administered intramuscularly on day 0 and 30 in a dose escalating fashion to healthy adults 18-49 years of age. Solicited and unsolicited events were collected through day 210. Immunogenicity measures taken at day 0, 30 and 60 included RSV A and B microneutralization, anti-F IgG, antigenic site II peptide and palivizumab competitive antibodies. RESULTS: The vaccine was well-tolerated, with no evident dose-related toxicity or attributable SAEs. At day 60 both RSV A and B microneutralization was significantly increased in vaccinees versus placebo. Across all vaccinees there was a 7- to 19-fold increase in the anti-F IgG and a 7- to 24-fold increase in the antigenic site II binding and palivizumab competitive antibodies. CONCLUSIONS: The RSV F nanoparticle vaccine candidate was well tolerated without dose-related increases in adverse events. Measures of immunity indicate that neutralization, anti-RSV F IgG titers and palivizumab competing antibodies were induced at levels that have been associated with decreased risk of hospitalization. NCT01290419.

National surveillance for human and pet contact with oral rabies vaccine baits, 2001-2009.

OBJECTIVE: To determine the rate and absolute number of human and pet exposures to oral rabies vaccine (ORV) bait containing liquid vaccinia rabies glycoprotein recombinant vaccine and to evaluate factors that might affect human contact with bait to modify the program and reduce human exposure to the vaccine. DESIGN: Retrospective analysis of surveillance data (2001 to 2009). SAMPLE: Reports on human and pet contact with ORV baits in states with ORV surveillance programs. PROCEDURES: Data were collected from passive, multistate ORV surveillance systems in Alabama, Arizona, Florida, Georgia, Maine, Maryland, Massachusetts, New Hampshire, New Jersey, New York, North Carolina, Ohio, Pennsylvania, Tennessee, Texas, Vermont, Virginia, and West Virginia. Data collected included the nature of human or pet contact with bait and vaccine, the caller's knowledge of the ORV bait program, local human population density, and other relevant demographic data. RESULTS: All 18 states participated in the surveillance program for at least 1 year, for a combined 68 years of observation. One thousand four hundred thirty-six calls were reported, representing 3,076 found baits (6.89/100,000 baits dropped); 296 (20%) calls were related to human contact with ruptured bait, and 550 (38%) involved pet contact with the bait. Six adverse events in humans were reported, one of which required hospitalization. Fifty-nine adverse events in pets were noted, all of which were nonserious. CONCLUSIONS AND CLINICAL RELEVANCE: Findings from surveillance activities have been used to improve baiting strategies and minimize human and pet contact with ORV baits. Overall, human and pet contact with ORV baits was infrequent. Surveillance has led to early identification of persons exposed to ORV and rapid intervention.

Safety and immunogenicity of a recombinant nonglycosylated erythrocyte binding antigen 175 Region II malaria vaccine in healthy adults living in an area where malaria is not endemic.

Erythrocyte binding antigen region II (EBA-175) is a conserved antigen of Plasmodium falciparum that is involved in binding of the parasite to the host's erythrocytes. We evaluated the safety and immunogenicity of a recombinant EBA-175 vaccine with aluminum phosphate adjuvant in healthy young adults living in the United States. Eighteen subjects/group received ascending doses (5, 20, 80, or 160 µg) of the vaccine at 0, 1, and 6 months; 8 subjects received placebo. Most of the injection site and systemic reactions were mild to moderate in intensity. After 2 or 3 doses of the vaccine at any concentration, antibody levels measured by enzyme-linked immunosorbent assay were significantly higher than those for the placebo group. Sera from subjects who received 3 doses of the vaccine at any concentration inhibited the growth of erythrocyte-stage P. falciparum at low levels compared to sera from placebo recipients or preimmune sera. In conclusion, the EBA-175 vaccine with adjuvant was safe and immunogenic in malaria-naïve subjects.

GI-5005, a yeast vector vaccine expressing an NS3-core fusion protein for chronic HCV infection.

The HCV nonstructural protein 3 (NS3) and core protein are highly conserved among various HCV genotypes, and several B- and T-cell epitopes have been characterized with these antigens. The immunotherapeutic vaccine GI-5005, being developed by GlobeImmune Inc, is a Tarmogen (targeted molecular immunogen) consisting of recombinant Saccharomyces cerevisiae yeast expressing an HCV NS3-core fusion protein designed to elicit antigen-specific host CD4+ and CD8+ T-cell responses for the treatment of chronic HCV infection. GI-5005 has demonstrated robust immunogenicity in preclinical in vitro and in vivo models. In a phase Ib clinical trial, GI-5005 monotherapy was well tolerated and displayed efficacy in patients with chronic HCV infection. At the time of publication, interim data were available from a completed phase II trial that evaluated a triple therapy of GI-5005 in combination with the standard-of-care (SOC; PEGylated-IFNalpha and ribavirin) regimen, compared with the SOC regimen alone. Triple therapy resulted in improved early virological responses in all treatment-naïve patients. End-of-trial results, including data of sustained virological responses, are required to better evaluate the efficacy of GI-5005 for the improvement of viral clearance and to compare the efficacy of the agent with other approaches such as NS3 protease inhibitors.

[Development and preclinical study of new generation virosomal split influenza vaccine "Grifor"].

New Russian virosomal split vaccine against influenza "Grifor" was developed. The vaccine is represented by mix of highly purified protective external and internal antigens of influenza A (H1N1 and H3N2) and B viruses. Developed technology of manufacture allowed to provide presentation of external antigens of influenza virus in the form of virosomes, and presentation of internal antigens in the form of micelles with maximal preservation of their antigenic activity. Using electron microscopy, electrophoresis in 10% polyacrilamide gel with sodium dodecyl sulfate, and polymerase chain reaction, morphologic and biochemical properties of the vaccine were studied. Preclinical study, including assessment of antigenic characteristics of "Grifor" vaccine compared to vaccine "Vaxigrip" (France), was performed. It was established that administration of the vaccine did not result in death of experimental animals, decrease of body mass, development of pathologic (including inflammatory, dystrophic and necrobiotic) changes in viscera or render adverse effects on blood hematologic and biochemical parameters and on the immune system. The vaccine was not pyrogenic and allergenic, did not have local irritating effects. Obtained results supported the appropriateness of conducting the clinical trials of "Grifor" vaccine on limited number of volunteers.

A therapeutic vaccine for nicotine dependence: preclinical efficacy, and Phase I safety and immunogenicity.

Nicotine is the principal addictive component in tobacco, and following uptake acts in the central nervous system. The smoking-cessation efforts of most smokers fail because a single slip often delivers sufficient nicotine to the brain to reinstate the drug-seeking behaviour. Blocking nicotine from entering the brain by induction of specific antibodies may be an effective means to prevent such relapses. The hapten nicotine was coupled to virus-like particles (VLP) formed by the coat protein of the bacteriophage Qb. In preclinical experiments, this Nicotine-Qb VLP (NicQb) vaccine induced strong antibody responses. After intravenous nicotine challenge, vaccinated mice exhibited strongly reduced nicotine levels in the brain compared with control mice. In a phase I study, 32 healthy non-smokers were immunized with NicQb. The vaccine was safe and well-tolerated. All volunteers who received NicQb showed nicotine-specific IgM antibodies at day 7 and nicotine-specific IgG antibodies at day 14. Antibody levels could be boosted by a second injection or the addition of Alum as an adjuvant and the antibodies had a high affinity for nicotine. These data suggest that antibodies induced by NicQb may prevent relapses by sequestering nicotine in the blood of immunized smokers.

Technology evaluation: ChimeriVax-DEN, Acambis/Aventis.

Acambis, in collaboration with Aventis Pasteur, is developing a chimeric vaccine based on a recombinant yellow fever vaccine for the potential prevention of dengue virus infection. The vaccine is undergoing phase I clinical trials.

Plant-based vaccines.

Plant systems are reviewed with regard to their ability to express and produce subunit vaccines. Examples of different types of expression systems producing a variety of vaccine candidates are illustrated. Many of these subunit vaccines have been purified and shown to elicit an immune response when injected into animal models. This review also includes vaccines that have been administered orally in a non-purified form as a food or feed product. Cases are highlighted which demonstrate that orally delivered plant-based vaccines can elicit immune responses and in some case studies, confer protection. Examples are used to illustrate some of the inherent advantages of a plant-based system, such as cost, ease of scale-up and convenience of delivery. Also, some of the key steps are identified that will be necessary to bring these new vaccines to the market.

Effect of monophosphoryl lipid A (MPL) on T-helper cells when administered as an adjuvant with pneumocococcal-CRM197 conjugate vaccine in healthy toddlers.

As new vaccines are developed, novel adjuvants may play an important role in eliciting an effective immune response. We evaluated the safety and adjuvant properties of monophosphoryl lipid A (MPL in 129 healthy toddlers immunized with two doses of nine-valent pneumococcal-CRM(197) protein conjugate vaccine (PCV9) combined with 10, 25, or 50 micro g of MPL with or without alum (AlPO(4)). Vaccine-specific humoral and cell-mediated responses were examined following the second dose of study vaccine. All doses of MPL were well-tolerated and a dose-dependent effect of MPL on specific cellular responses was observed. The 10 micro g MPL dose significantly enhanced CRM(197)-specific T-cell proliferation (P=0.02) and interferon-gamma (INF-gamma) production (P=0.009) compared to responses of controls who received PCV9 with AlPO(4). In contrast, CRM(197)-specific T-cell proliferation and interferon-gamma production of the 50 micro g MPL/AlPO(4) group were decreased when compared to controls although these differences did not reach statistical significance. IL-5 and IL-13 responses after immunization showed a similar pattern with increased production in the 10 micro g MPL group and decreased production in the 50 micro g MPL/AlPO(4) group compared to controls. There were no differences in serum IgG antibody concentrations to the nine vaccine pneumococcal capsular polysaccharides and carrier protein between the MPL-containing and control vaccine groups. These findings demonstrate a dose-dependent effect of MPL on T-helper cell type 1 (TH-1) responses to the carrier protein and also suggest an effect on T-helper cell type 2 (TH-2) responses.

Human immune responses to a novel norwalk virus vaccine delivered in transgenic potatoes.

A new approach for delivering vaccine antigens is the use of inexpensive, plentiful, plant-based oral vaccines. Norwalk virus capsid protein (NVCP), assembled into virus-like particles, was used as a test antigen, to determine whether immune responses could be generated in volunteers who ingested transgenic potatoes. Twenty-four healthy adult volunteers received 2 or 3 doses of transgenic potato (n=20) or 3 doses of wild-type potato (n=4). Each dose consisted of 150 g of raw, peeled, diced potato that contained 215-751 microgram of NVCP. Nineteen (95%) of 20 volunteers who ingested transgenic potatoes developed significant increases in the numbers of specific IgA antibody-secreting cells. Four (20%) of 20 volunteers developed specific serum IgG, and 6 (30%) of 20 volunteers developed specific stool IgA. Overall, 19 of 20 volunteers developed an immune response of some kind, although the level of serum antibody increases was modest.