Maturation of Aluminium Adsorbed Antigens Contributes to the Creation of Homogeneous Vaccine Formulations.

Although aluminium-based vaccines have been used for almost over a century, their mechanism of action remains unclear. It is established that antigen adsorption to the adjuvant facilitates delivery of the antigen to immune cells at the injection site. To further increase our understanding of aluminium-based vaccines, it is important to gain additional insights on the interactions between the aluminium and antigens, including antigen distribution over the adjuvant particles.

The role of nanoparticle format and route of administration on self-amplifying mRNA vaccine potency.

The efficacy of RNA-based vaccines has been recently demonstrated, leading to the use of mRNA-based COVID-19 vaccines. The application of self-amplifying mRNA within these formulations may offer further enhancement to these vaccines, as self-amplifying mRNA replicons enable longer expression kinetics and more potent immune responses compared to non-amplifying mRNAs. To investigate the impact of administration route on RNA-vaccine potency, we investigated the immunogenicity of a self-amplifying mRNA encoding the rabies virus glycoprotein encapsulated in different nanoparticle platforms (solid lipid nanoparticles (SLNs), polymeric nanoparticles (PNPs) and lipid nanoparticles (LNPs)).

Conjugation of Mannans to Enhance the Potency of Liposome Nanoparticles for the Delivery of RNA Vaccines.

Recent approval of mRNA vaccines to combat COVID-19 have highlighted the potential of this platform. Lipid nanoparticles (LNP) is the delivery vehicle of choice for mRNA as they prevent its enzymatic degradation by encapsulation. We have recently shown that surface exposition of mannose, incorporated in LNPs as stable cholesterol-amine conjugate, enhances the potency of self-amplifying RNA (SAM) replicon vaccines through augmented uptake by antigen presenting cells (APCs).

Rational design of adjuvants for subunit vaccines: The format of cationic adjuvants affects the induction of antigen-specific antibody responses.

A range of cationic delivery systems have been investigated as vaccine adjuvants, though few direct comparisons exist. To investigate the impact of the delivery platform, we prepared four cationic systems (emulsions, liposomes, polymeric nanoparticles and solid lipid nanoparticles) all containing equal concentrations of the cationic lipid dimethyldioctadecylammonium bromide in combination with the Neisseria adhesin A variant 3 subunit antigen. The formulations were physicochemically characterized and their ability to associate with cells and promote antigen processing (based on degradation of DQ-OVA, a substrate for proteases which upon hydrolysis is fluorescent) was compared in vitro and their vaccine efficacy (antigen-specific antibody responses and IFN-γ production) and biodistribution (antigen and adjuvant) were evaluated in vivo.

Delivery of self-amplifying mRNA vaccines by cationic lipid nanoparticles: The impact of cationic lipid selection.

Self-amplifying RNA (SAM) represents a versatile tool that can be used to develop potent vaccines, potentially able to elicit strong antigen-specific humoral and cellular-mediated immune responses to virtually any infectious disease. To protect the SAM from degradation and achieve efficient delivery, lipid nanoparticles (LNPs), particularly those based on ionizable amino-lipids, are commonly adopted. Herein, we compared commonly available cationic lipids, which have been broadly used in clinical investigations, as an alternative to ionizable lipids.

Investigating the Impact of Delivery System Design on the Efficacy of Self-Amplifying RNA Vaccines.

messenger RNA (mRNA)-based vaccines combine the positive attributes of both live-attenuated and subunit vaccines. In order for these to be applied for clinical use, they require to be formulated with delivery systems. However, there are limited in vivo studies which compare different delivery platforms.

Mannosylation of LNP Results in Improved Potency for Self-Amplifying RNA (SAM) Vaccines.

Mannosylation of Lipid Nanoparticles (LNP) can potentially enhance uptake by Antigen Presenting Cells, which are highly abundant in dermal tissues, to improve the potency of Self Amplifying mRNA (SAM) vaccines in comparison to the established unmodified LNP delivery system. In the current studies, we evaluated mannosylated LNP (MLNP), which were obtained by incorporation of a stable Mannose-cholesterol amine conjugate, for the delivery of an influenza (hemagglutinin) encoded SAM vaccine in mice, by both intramuscular and intradermal routes of administration. SAM MLNP exhibited enhanced uptake in comparison to unglycosylated LNP from bone marrow-derived dendritic cells, and more rapid onset of the antibody response, independent of the route.

Mixed mucosal-parenteral immunizations with the broadly conserved pathogenic Escherichia coli antigen SslE induce a robust mucosal and systemic immunity without affecting the murine intestinal microbiota.

Emergence and dissemination of multidrug resistance among pathogenic Escherichia coli have posed a serious threat to public health across developing and developed countries. In combination with a flexible repertoire of virulence mechanisms, E. coli can cause a vast range of intestinal (InPEC) and extraintestinal (ExPEC) diseases but only a very limited number of antibiotics still remains effective against this pathogen.

Alum/Toll-Like Receptor 7 Adjuvant Enhances the Expansion of Memory B Cell Compartment Within the Draining Lymph Node.

Vaccination is one of the most cost-effective health interventions and, with the exception of water sanitization, no other action has had such a major effect in mortality reduction. Combined with other approaches, such as clean water, better hygiene, and health education, vaccination contributed to prevent millions of cases of deaths among children under 5 years of age. New or improved vaccines are needed to fight some vaccine-preventable diseases that are still a threat for the public health globally, as reported also in the Global Vaccine Action Plan (GVAP) endorsed by the World Health Assembly in 2012.

Addition of a TLR7 agonist to an acellular pertussis vaccine enhances Th1 and Th17 responses and protective immunity in a mouse model.

A resurgence of whooping cough (pertussis) has been observed in recent years in a number of developed countries, despite widespread vaccine coverage. Although the exact reasons of the recurrence of pertussis are not clear, there are a number of potential causes, like antigenic variation in the circulating strains of Bordetella pertussis, changes in surveillance and diagnostic tools, and potential differences in protection afforded by current acellular pertussis (aP) vaccines compared to more reactogenic whole cell (wP) vaccines, which they replaced. Studies in animal models have shown that induction of cellular as well as humoral immune responses are key to conferring effective and long lasting protection against B.

Optimizing adjuvants for intradermal delivery of MenC glycoconjugate vaccine.

Intradermal vaccine delivery is a promising alternative to the conventional intramuscular route. The skin layer is immunologically supported by a densely network of antigen presenting cells, while the skeletal muscle is loaded with a relatively sparse population of APCs. Nevertheless, the vaccine to be suitable for intradermal delivery needs a new formulation to facilitate either smaller injection volumes or the introduction into new delivery devises as micro-needles.

Novel adjuvant Alum-TLR7 significantly potentiates immune response to glycoconjugate vaccines.

Although glycoconjugate vaccines are generally very efficacious, there is still a need to improve their efficacy, especially in eliciting a strong primary antibody response. We have recently described a new type of vaccine adjuvant based on a TLR7 agonist adsorbed to alum (Alum-TLR7), which is highly efficacious at enhancing immunogenicity of protein based vaccines. Since no adjuvant has been shown to potentiate the immune response to glycoconjugate vaccines in humans, we investigated if Alum-TLR7 is able to improve immunogenicity of this class of vaccines.

The adjuvant effect of TLR7 agonist conjugated to a meningococcal serogroup C glycoconjugate vaccine.

Conjugation of a small molecule immunopotentiator to antigens has been proposed to deliver the ligand to the receptor, localize its action and minimize systemic inflammation. However, the effect of conjugation of Toll like receptor 7 agonists (TLR7a) on the immunogenicity of carbohydrate-based vaccines is unknown. In this study we synthesized an anti-Neisseria meningitidis serogroup C (MenC) glycoconjugate vaccine composed of MenC oligosaccharide antigens covalently linked to the carrier protein CRM197, to which a TLR7a was in turn conjugated.

The preparation and characterization of PLG nanoparticles with an entrapped synthetic TLR7 agonist and their preclinical evaluation as adjuvant for an adsorbed DTaP vaccine.

The design of safe and potent adjuvants able to enhance and modulate antigen-specific immunity is of great interest for vaccine research and development. In the present study, negatively charged poly(lactide-co-glycolide) (PLG) nanoparticles have been combined with a synthetic immunepotentiator molecule targeting the Toll-like receptor 7. The selection of appropriate preparation and freeze-drying conditions resulted in a PLG-based adjuvant with well-defined and stable physico-chemical properties.

Positive Contribution of Adjuvanted Influenza Vaccines to the Resolution of Bacterial Superinfections.

Background: Most preclinical studies assess vaccine effectiveness in single-pathogen infection models. This is unrealistic given that humans are continuously exposed to different commensals and pathogens in sequential and mixed infections. Accordingly, complications from secondary bacterial infection are a leading cause of influenza-associated morbidity and mortality.

Multiplex immunoassay for in vitro characterization of acellular pertussis antigens in combination vaccines.

Vaccines characterization is required to ensure physical, chemical, and biological integrity of antigens and adjuvants. Current analytical methods mostly require complete antigen desorption from aluminum-based adjuvants and are not always suitable to distinguish individual antigens in multivalent formulations. Here, Luminex technology is proposed to improve the analytics of vaccine characterization.

CD8 T-cell priming upon mRNA vaccination is restricted to bone-marrow-derived antigen-presenting cells and may involve antigen transfer from myocytes.

Self-amplifying mRNAs (SAM(®) ) are a novel class of nucleic acid vaccines, delivered by a non-viral delivery system. They are effective at eliciting potent and protective immune responses and are being developed as a platform technology with potential to be used for a broad range of targets. However, their mechanism of action has not been fully elucidated.

The potential of adjuvants to improve immune responses against TdaP vaccines: A preclinical evaluation of MF59 and monophosphoryl lipid A.

The successful approach of combining diphtheria, tetanus and pertussis antigens into a single vaccine has become a cornerstone of immunization programs. Yet, even if vaccination coverage is high, a resurgence of pertussis has been reported in many countries suggesting current vaccines may not provide adequate protection. To induce better tailored and more durable immune responses against pertussis vaccines different approaches have been proposed, including the use of novel adjuvants.

Fabrication of cell culture-derived influenza vaccine dissolvable microstructures and evaluation of immunogenicity in guinea pigs.

Microstructure patches provide an opportunity for simple, effective, and safe vaccine administration, while achieving the desired immune response. We have evaluated the MicroCor transdermal system for cell culture-derived trivalent influenza vaccine administration. Influenza monovalent purified bulk vaccines (monobulks) (H1N1, H3N2, B) were concentrated by tangential flow filtration, lyophilized, and formulated with biocompatible excipients to form the microstructure array dissolvable tips.

Rational Design of Adjuvant for Skin Delivery: Conjugation of Synthetic β-Glucan Dectin-1 Agonist to Protein Antigen.

The potential benefits of skin delivery of vaccines derive from the presence of a densely connected network of antigen presenting cells in the skin layer, most significantly represented by Langerhans cells and dermal dendritic cells. Targeting these cells by adjuvant conjugated to an antigen should result in enhanced immunogenicity of a vaccine. Since one of the most widely used adjuvants is an insoluble salt of aluminum (aluminum hydroxide) that cannot be used for skin delivery due to reactogenicity, we focused our attention on agonists of receptors present on skin dendritic cells, including the Dectin-1 receptor.

Vaccine composition formulated with a novel TLR7-dependent adjuvant induces high and broad protection against Staphylococcus aureus.

Both active and passive immunization strategies against Staphylococcus aureus have thus far failed to show efficacy in humans. With the attempt to develop an effective S. aureus vaccine, we selected five conserved antigens known to have different roles in S.

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.

Protective efficacy induced by recombinant Clostridium difficile toxin fragments.

Clostridium difficile is a spore-forming bacterium that can reside in animals and humans. C. difficile infection causes a variety of clinical symptoms, ranging from diarrhea to fulminant colitis.

The adjuvant effect of MF59 is due to the oil-in-water emulsion formulation, none of the individual components induce a comparable adjuvant effect.

MF59 is a safe and effective vaccine adjuvant that has been used in a licensed seasonal influenza vaccine for 15 years. The purpose of the present studies was to directly address a question that has been asked of us on many occasions: "which is the adjuvant active component of MF59?". Since we have recently gained a number of insights on how MF59 works as an adjuvant, we were able to use these approaches to evaluate if the individual components of MF59 (squalene oil, the surfactants Span 85 and Tween 80 or the citrate buffer) showed any direct immunostimulatory activity.

Preparation of highly concentrated influenza vaccine for use in novel delivery approaches.

Vaccine antigens are usually available only as dilute solutions, which are difficult to formulate into various novel delivery systems, which often require highly concentrated antigens. To address this problem, we have utilized tangential flow filtration (TFF), a simple and scalable process to prepare highly concentrated vaccine antigens. Here, we describe the optimization of TFF to concentrate hemagglutinin (HA) of egg-derived influenza antigens, from 2008 to 2009 seasonal vaccine, to concentrations up to 28 mg/mL.