Needle-free Vaccination Research Articles

Formulation, characterization and optimization of Hepatitis B surface antigen (HBsAg)-loaded chitosan microspheres for oral delivery

Context: Approximately 400 million persons worldwide have chronic hepatitis B. This is due to problems associated with vaccine delivery, stability and cost. Hence the present challenge in vaccinology is to develop safer, cheaper and easy-to-deliver forms of vaccines. A novel needle-free oral vaccine will be an ideal tool to fight this silent killer disease.Objective: The aim of this work was to prepare and evaluate chitosan-loaded HBsAg microspheres for oral delivery.Materials and methods: Chitosan microspheres were prepared by emulsion solvent evaporation technique. To overcome the enzymatic and permeation barrier, protease inhibitors and permeation enhancers were also added. Studies were conducted to find the effect of stabilizer concentration, stirring speed, cross-linking agent and polymer concentration on microsphere size and entrapment efficiency. Formulations were characterized for their particle size, entrapment efficiency. They were also evaluated for the in vitro drug release, in vivo performances and the effect of different storage conditions.Results: HBsAg-loaded chitosan microspheres with bacitracin as protease inhibitor showed better protective levels of immunity after oral administration comparing with aprotinin as protease inhibitor. Stability at room temperature up to a period of four months reduces incomplete vaccine coverage and logistic requirements.Conclusion: The study signifies the potential of the formulated chitosan microspheres for effective oral administration of HBsAg.

Progress Towards a Needle-Free Hepatitis B Vaccine

Hepatitis B virus (HBV) infection is a worldwide public health problem. Vaccination is the most efficient way to prevent hepatitis B. Despite the success of the currently available vaccine, there is a clear need for the development of new generation of HBV vaccines. Needle-free immunization is an attractive approach for mass immunization campaigns, since avoiding the use of needles reduces the risk of needle-borne diseases and prevents needle-stick injuries and pain, thus augmenting patient compliance and eliminating the need for trained medical personnel. Moreover, this kind of immunization was shown to induce good systemic as well as mucosal immunological responses, which is important for the creation of both a prophylactic and therapeutic vaccine. In order to produce a better, safer, more efficient and more suitable vaccine, adjuvants have been used. In this article, several adjuvants tested over the years for their potential to help create a needle-free vaccine against HBV are reviewed.

In Vivo Molecular Imaging Analysis of a Nasal Vaccine That Induces Protective Immunity against Botulism in Nonhuman Primates

Nasal administration is an effective route for a needle-free vaccine. However, nasally administered Ags have the potential to reach the CNS directly from the nasal cavity, thus raising safety concerns. In this study, we performed real-time quantitative tracking of a nasal vaccine candidate for botulism, which is a nontoxic subunit fragment of Clostridium botulinum type A neurotoxin (BoHc/A) effective in the induction of the toxin-neutralizing immune response, by using (18)F-labeled BoHc/A-positron-emission tomography, an in vivo molecular imaging method. This method provides results that are consistent with direct counting of [(18)F] radioactivity or the traditional [(111)In]-radiolabel method in dissected tissues of mice and nonhuman primates. We found no deposition of BoHc/A in the cerebrum or olfactory bulb after nasal administration of (18)F-labeled BoHc/A in both animals. We also established a real-time quantitative profile of elimination of this nasal vaccine candidate and demonstrated that it induces highly protective immunity against botulism in nonhuman primates. Our findings demonstrate the efficiency and safety of a nasal vaccine candidate against botulism in mice and nonhuman primates using in vivo molecular imaging.

Formulation and Evaluation of a Novel Needle-Free Hepatitis B Surface Antigen Delivery System

Problem statement: Hepatitis B is a major public health problem, kill ing 1-2 million people annually, despite the introduction of an eff ective vaccine in 1982. Approximately 400 million persons worldwide have chronic hepatitis B. This is due to problems associated with vaccine delivery, stability and cost. Hence the present challenge in Vaccinology is to develop safer, cheaper and easy-t o- deliver forms of vaccines. A novel needle-free oral vaccine will be an ideal tool to fight with this s ilent killer disease. Approach: Hence a study was conducted with the aim of formulating and evaluating an effective oral HBsAg vaccine. HBsAg-loaded microspheres were prepared by microencapsulation using albumin as polymer and sodium taurocholate as permeation enhancer. Bacitracin and aprotinin were incorporated as protease inhibitors. After in vitro studies, optimized drug-loaded microspheres were encapsulated in hard gelatin capsules which we re further enteric coated with cellulose acetate phthalate. In vivo experiments were conducted in rabbits. Control gro up was administered with 20 µg of HBsAg intramuscularly and the test groups were given different formulations of oral vaccine as per the current vaccination schedule of 0, 1 and 6 mont hs. Immunogenicity was compared with IM injected vaccine control group. Results: HBsAg microspheres with albumin as polymer and bacitracin as protease inhibitor induced immune response with specific antibodies and cell mediated immunity better than the formulation with aprotinin as prote ase inhibitor (p = 0.047). Vaccine-related adverse events were absent in all immunized rabbits. Basic haematological, biochemical and physical parameters were within normal limits. Conclusion: The stability exhibited by this vaccine at room temperature can be extremely useful to overcome the incomplete vaccine coverage due to cold chain requirement. Impressive large logistical advantages and elimination of blood-borne infections are othe r major benefits, which may have a great impact on gl obal HBsAg immunization acceptance and compliance and on hepatitis B disease burden.

Targeted, Needle‐Free Vaccinations in Skin using Multilayered, Densely‐Packed Dissolving Microprojection Arrays

Targeting of vaccines to abundant immune cell populations within our outer thin skin layers using miniaturized devices-much thinner than a needle and syringe, could improve the efficacy of vaccines (and other immunotherapies). To meet this goal, a densely packed dissolving microprojection array (dissolving Nanopatch) is designed, achieving functional miniaturization by 1) formulating small microneedles (two orders of magnitude smaller than a standard needle and syringe) and 2) multiple layering of the payload within microprojections with tight tolerances (of the order of a micrometer). The formulation method is suitable to many vaccines because it is without harsh or complex chemical processes, and it is performed at low temperatures and at a neutral pH. When the formulated dNPs are applied to skin, consistent and robust penetration is achieved, rapidly targeting the skin strata of interest (<5 min; significantly faster than larger dissolving microneedles that have been previously reported). Resultant diffusion is significantly enhanced within the dermis compared with the epidermis. Using two different antigens (ovalbumin and a commercial trivalent influenza vaccine [Fluvax2008]), the administration of these dissolving patches generate robust systemic immune responses in a mouse model. To the authors' knowledge, this is the first report of successful vaccination with any form of dissolving microneedles. The patches made by this method therefore have the potential for pain-free, needle-free, and effective vaccination in humans.

Chaperoning vaccines

A cationic nanosized hydrogel (nanogel) shows controlled antigen delivery in vivo following intranasal administration and hence holds promise for a clinically effective adjuvant-free and needle-free vaccine system.

An Adjuvant for the Induction of Potent, Protective Humoral Responses to an H5N1 Influenza Virus Vaccine with Antigen-Sparing Effect in Mice

Intramuscular administration of inactivated influenza virus vaccine is the main vaccine platform used for the prevention of seasonal influenza virus infection. In clinical trials, inactivated H5N1 vaccines have been shown to be safe and capable of eliciting immune correlates of protection. However, the H5N1 vaccines are poorly immunogenic compared to seasonal influenza virus vaccines. Needle-free vaccination would be more efficient and economical in a pandemic, and the development of an effective and safe mucosal adjuvant will be an important milestone. A stabilized chemical analog of double-stranded RNA, PIKA, was previously reported to be a potent mucosal adjuvant in a murine model. While PIKA stimulates dendritic cells in vitro, little was known about its receptor and adjuvanting mechanism in vivo. In this study, we demonstrated that the immunostimulatory effect of PIKA resulted in an increased number of mature antigen-presenting cells, with the induction of proinflammatory cytokines at the inoculation site. In addition, coadministration of PIKA with a poorly immunogenic H5N1 subunit vaccine led to antigen sparing and quantitative and qualitative improvements of the immune responses over those achieved with an unadjuvanted vaccine in mice. The adjuvanted vaccine provided protection against lethal challenge with homologous and heterologous H5N1 wild-type viruses. Mice lacking functional TLR3 showed diminished cytokine production with PIKA stimulation, diminished antibody responses, and reduced protective efficacy against wild-type virus challenge following vaccination. These data suggest that TLR3 is important for the optimal performance of PIKA as an adjuvant. With its good safety profile and antigen-sparing effect, PIKA could be an attractive adjuvant for use in future pandemics.

Transcutaneous vaccination using a hydrogel patch induces effective immune responses to tetanus and diphtheria toxoid in hairless rat

Transcutaneous immunization (TCI) targeting the Langerhans cells (LCs) of the epidermal layer is a promising needle-free, easy-to-use, and non-invasive vaccination method. We developed a hydrogel patch formulation to promote the penetration of antigenic proteins into the stratum corneum. Here, we investigated the characteristics of the immune responses induced by this vaccination method and the vaccine efficacy of TCI using a hydrogel patch containing tetanus and diphtheria toxoids. Our TCI system induced toxoid-specific IgG production in an antigen dose-, patch area-, and application period-dependent manner. Moreover, IgG subclass analysis indicated that our TCI predominantly elicited a Th2-type immune response rather than a Th1-type immune response. Importantly, our TCI system induced antigen-specific immune memory based on the booster effect and showed potent efficacy, comparable to that of subcutaneous immunization in toxin-challenge experiments. On the basis of these results, we are now performing translational research to apply TCI for tetanus and diphtheria.

Characterization of Stability and Nasal Delivery Systems for Immunization with Nanoemulsion-Based Vaccines

Many infectious diseases that cause significant morbidity and mortality, especially in the developing world, could be preventable through vaccination. The effort to produce safe, thermally stable, and needle-free mucosal vaccines has become increasingly important for global health considerations. We have previously demonstrated that a thermally stable nanoemulsion, a mucosal adjuvant for needle-free nasal immunization, is safe and induces protective immunity with a variety of antigens, including recombinant protein. The successful use of nanoemulsion-based vaccines, however, poses numerous challenges. Among the challenges is optimization of the formulation to maintain thermal stability and potency and another is accuracy and efficiency of dispensing the vaccines to the nasal mucosa in the anterior and turbinate region of the nasal cavity or potentially to the nasopharynx-associated lymphoid tissue. We have examined the effects of different diluents [phosphate-buffered saline (PBS) and 0.9% NaCl] on the stability and potency of nanoemulsion-based vaccines. In addition, we have determined the efficiency of delivering them using commercially available nasal spray devices (Pfeiffer SAP-62602 multidose pump and the BD Hypak SCF 0.5 ml unit dose Accuspray(TM)). We report the stability and potency of PBS-diluted ovalbumin-nanomeulsion mixtures for up to 8 months and NaCl-diluted mixtures up to 6 months when stored at room temperature. Significant differences in spray characteristics including droplet size, spray angle, plume width, and ovality ratios were observed between the two pumps. Further, we have demonstrated that the nanoemulsion-based vaccines are not physically or chemically altered and retain potency following actuation with nasal spray devices. Using either device, the measured spray characteristics suggest deposition of nanoemulsion-based vaccines in inductive tissues located in the anterior region of the nasal cavity. The results of this study suggest that nanoemulsion-based vaccines do not require specially engineered delivery devices and support their potential use as nasopharyngeal vaccine adjuvants.

Conjugation of a TLR5 agonist to the Plasmodium falciparum Circumsporozoite (CS) protein enhances murine and human Dendritic cells (DC) interaction in vitro and increases protective immunity following intranasal immunization in mice (37.4)

Abstract A needle-free malaria vaccine has been developed based on intranasal (I.N) immunization with a recombinant P. falciparum circumsporozoite (CS) protein conjugated to the TLR5 agonist flagellin. Mice immunized I.N, but not subcutaneously, with flagellin-modified CS developed sporozoite neutralizing antibodies that protect against challenge with a transgenic rodent parasite expressing P. falciparum CS repeats. To better understand the role of innate immune cells in the induction of protective immunity, we analyzed the interaction of flagellin-modified CS with murine bone marrow-derived dendritic cells (DC) and human monocyte-derived DC in vitro. The fusion protein was rapidly taken up by DC and remained detectable in the cytosol as long as 24 hours post antigen pulse. A murine DC line (D1), and a sub-population of human DC, matured and expressed increased levels of CD40 and CD86 following stimulation with flagellin-modified CS. In I.N. immunized mice, the Nasopharyngeal-associated Lymphoid Tissue (NALT) was enlarged 2-fold over naïve NALT, and increased levels of CD11c+ DC were observed by confocal microscopy and FACS analysis. NALT cell subpopulations exhibited a higher B:T cell ratio than those observed in lymph nodes. A better understanding of innate and adaptive immune responses elicited in NALT by TLR ligand-modified CS protein will help identify critical immune parameters required for the induction of sporozoite specific immunity that protects from malaria infection.

Particle Size Influences the Immune Response Produced by Hepatitis B Vaccine Formulated in Inhalable Particles

To test the hypothesis that particle size influences the magnitude of immune response produced by hepatitis B surface antigen (HBsAg) encapsulated in poly (lactic-co-glycolic acid) (PLGA) microspheres. Microspheres were prepared by a double-emulsion-solvent-evaporation method, and the particles were characterized for size, morphology, porosity and antigen content. Immunogenicity of encapsulated antigen and safety were studied in rats. Uptake of fluorescent-labeled particles by rat alveolar macrophages was studied by confocal microscopy. With increasing internal aqueous phase (IAP) volume of the microsphere, an increase in particle size and a decrease in particle density were observed. Particles with varying geometric diameters showed aerodynamic diameters between 1 and 6 mu. Addition of poly vinyl alcohol to the IAP resulted in particles with a porous surface. The integrity of HBsAg was maintained upon encapsulation in microspheres. Continuous release of the antigen was observed for formulations incubated in phosphate-buffered saline for 28 days. Immunogenicity increased as a function of particle size upon pulmonary administration. HBsAg encapsulated in approximately 5 mum particles elicited a significantly higher immune response compared to that encapsulated in approximately 12 mum particles. Similar to in vivo immune response data, fluorescent-labeled microspheres of smaller size were taken up more efficiently by rat alveolar macrophages compared to those of larger size. No significant increase in either tumor necrosis factor alpha level in bronchoalveolar lavage fluid or wet lung weight, indicators of inflammation, was observed in rats that received optimized formulations via the pulmonary route. HBsAg delivered in PLGA microspheres elicited an increase in immunogenicity, and the magnitude of immune response was more pronounced with smaller particles. Inhalable particles of HBsAg could be a viable approach to needle-free vaccination.

Formulation and Evaluation of a Novel Needle-Free Hepatitis B Surface Antigen Delivery System

Problem statement: Hepatitis B is a major public health problem, killing 1-2 million people annually, despite the introduction of an effective vaccine in 1982. Approximately 400 million persons worldwide have chronic hepatitis B. This is due to problems associated with vaccine delivery, stability and cost. Hence the present challenge in Vaccinology is to develop safer, cheaper and easy-to-deliver forms of vaccines. A novel needle-free oral vaccine will be an ideal tool to fight with this silent killer disease. Approach: Hence a study was conducted with the aim of formulating and evaluating an effective oral HBsAg vaccine. HBsAg-loaded microspheres were prepared by microencapsulation using albumin as polymer and sodium taurocholate as permeation enhancer. Bacitracin and aprotinin were incorporated as protease inhibitors. After in vitro studies, optimized drug-loaded microspheres were encapsulated in hard gelatin capsules which were further enteric coated with cellulose acetate phthalate. In vivo experiments were conducted in rabbits. Control group was administered with 20 µg of HBsAg intramuscularly and the test groups were given different formulations of oral vaccine as per the current vaccination schedule of 0, 1 and 6 months. Immunogenicity was compared with IM injected vaccine control group. Results: HBsAg microspheres with albumin as polymer and bacitracin as protease inhibitor induced immune response with specific antibodies and cell mediated immunity better than the formulation with aprotinin as protease inhibitor (p = 0.047). Vaccine-related adverse events were absent in all immunized rabbits. Basic haematological, biochemical and physical parameters were within normal limits. Conclusion: The stability exhibited by this vaccine at room temperature can be extremely useful to overcome the incomplete vaccine coverage due to cold chain requirement. Impressive large logistical advantages and elimination of blood-borne infections are other major benefits, which may have a great impact on global HBsAg immunization acceptance and compliance and on hepatitis B disease burden.

Microencapsulation of Vaccine Antigens and Adjuvants for Mucosal Targeting

Delivery of vaccine antigens that can trigger potent mucosal immune response is one of the effective strategies to overcome a wide array of infectious diseases. Microencapsulation of vaccine antigens with Poly(lactide-co-glycolic acids) (PLGA), an FDA approved biodegradable polymer, has been investigated for targeted M-cell uptake. While PLGA possesses many attractive properties, a successful PLGA based mucosal-targeted vaccine has yet to be introduced. This review focuses on the physiochemical properties important in the preparation of antigen-loaded PLGA microparticles, properties that influence M-cell specific uptake, and the induction of effective immune responses. In addition, a possible role of microparticle properties in immune adjuvant activity is discussed. A careful consideration of these factors may yet lead to the development of an effective needle-free mucosal vaccine using polymer microparticles. Keywords: M-cell targeting, PLGA, microparticles

Enhanced humoral and cellular immune responses after sublingual immunization against human papillomavirus 16 L1 protein with adjuvants

Needle-free nonparenteral vaccines offer a number of practical advantages, especially in developing countries. To address the effects of vaccine administration route, we tested mucosal and systemic immune responses against human papillomavirus 16 L1(HPV16L1) protein using intranasal, intravaginal, transdermal, sublingual (SL) and intramuscular routes. The SL route provided the most effective mucosal secretory IgA (sIgA) and serum IgG responses. After a 150 μg antigen dose via the SL route, saliva sIgA levels were 7.2- and 5.8-fold higher than those achieved via intravaginal and transdermal routes, respectively. Notably, SL administration even produced 4.6-fold higher levels of vaginal sIgA levels than did intravaginal delivery of 150 μg antigen. To enhance the immunogenicity of SL vaccines, we tested the adjuvanticity of nine molecules: three toll-like receptor agonists, three nucleotide-binding oligomerization-domain agonists, vitamin D3, poly-γ-glutamic acid and cholera toxin subunit B (CTB). Among the molecules tested, CTB provided the most enhanced mucosal sIgA and systemic IgG induction. SL-applied CTB enhanced the production of interleukin-4 and interferon-γ from stimulated CD4+ T cells. Moreover, interferon-γ-producing CD8+ T cell responses were increased 1.7-fold after co-treatment with SL CTB and HPV16L1. These results suggest the potential of the SL route for delivery of HPV16L1 vaccines using CTB as an adjuvant.

From single-dose vaccine delivery systems to nanovaccines

The application of micro- and nanotechnologies to the development of vaccine delivery systems started in the 70s. From that time onwards, significant advances in this field have opened up opportunities in the research of new adjuvants and the design of safer and more effective vaccines. The versatility of vaccine delivery vehicles represents an important feature in vaccine development. Indeed, the modulation of the composition, particle size and/or surface properties, offers a wide range of possibilities for overcoming natural barriers and controlling antigen release. Thus, it is currently accepted that besides their ability to enhance the low immunogenicity of new subunit and DNA vaccines, these delivery carriers represent a promising strategy for needle-free and single-dose vaccination. In this review article we aim to present the historical advances made in the field of vaccine delivery, particularly the progress made in the last two decades, describing our specific contribution, ranging from the idea of single-dose vaccination to the current concept of nanovaccines. Overall, this review illustrates the potential of nanosciences and nanotechnologies to solve the current problems of vaccination and, thus, to expand the immunization coverage worldwide.

Intercell and GSK in Alliance for Novel Patch Vaccines

GlaxoSmithKline (GSK) Biologicals SA and Austrian vaccine maker Intercell announced an agreement for a strategic alliance to develop and commercialise innovative needle-free patch-based vaccines.

CAF01 liposomes as a mucosal vaccine adjuvant: In vitro and in vivo investigations

Mucosal administration of vaccines has many advantages compared to parenteral vaccination. Needle-free mucosal vaccination would be easily applicable, target the vaccine to the entry point of many pathogens, and reduce the risk of infection with other pathogens during vaccination as compared to invasive methods. CAF01 is a novel liposome-based vaccine adjuvant with remarkable immunostimulatory activity. The potential of CAF01 liposomes as adjuvant for mucosal vaccines was investigated using the Calu-3 epithelial cell culture in vitro model. Thus, the mucosal permeability of the antigen as well as the epithelial integrity and the metabolic activity of the well-differentiated cells were investigated after exposure to CAF01. Finally, the adjuvant was tested for nasal administration in mice, combined with an influenza vaccine. The results suggest that CAF01 enhanced transport of antigen through the mucus layer on Calu-3 cells, increasing the concentration of antigen in the cell layer, as well as the transport across the epithelial cells. Furthermore CAF01 was well tolerated by the Calu-3 cells and the in vivo studies demonstrated increased cell-mediated immunity (CMI) as well as humoral immune responses in mice after nasal application of the influenza vaccine when combined with CAF01. CAF01 is thus a promising adjuvant for mucosal delivery.

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AUSTRALIA – Australia's First Full Genome Project to be Conducted on Corals. AUSTRALIA – Scientists May have Discovered New Potential Cure for Cancer. AUSTRALIA – First Genetically-engineered Malaria Vaccine To Enter Human Trials. CHINA – Lead Poisoning Sickens 600 Kids in China. CHINA – Groundbreaking Treatment for Oxygen-deprived Newborns. CHINA – China Builds First Heavy Ion Therapy Center for Cancer Patients. CHINA – Creating Live Mice from Skin Cells. INDIA – Human Clinical Trial in 2010 for Needle-free Measles Vaccination. INDIA – Indian Wonder Herb can Treat Male Infertility. JAPAN – Flood Resistant High-yield Rice Developed. SINGAPORE – Minimally Invasive Option for Knee Cartilage Repair. SINGAPORE – Novel Immunization Method for Malaria Offers Insights into Human Anti-Malaria Immune Response. TAIWAN – Taiwan Researchers Identify Sites of Breast Cancer Genes. TAIWAN – Taiwanese Researchers Develop Cell Therapy For Immunodeficiency. TAIWAN – Remote Healthcare Services for High-risk Patients. TAIWAN – Marine-derived Compounds Holds New Treatment Premise for Neuropathic Pain. OTHER REGIONS — UNITED STATES – New No-needle Approach to Prevent Blood Clots.

Oral Vaccine Delivery – New Strategies and Technologies

Although most commercial vaccines are delivered by injection, there is an increasing interest in needle-free vaccine delivery for reasons including the ability to elicit immune responses at mucosal surfaces, ease of administration, and the ability to administer vaccines without the need for trained medical professionals. This review summarizes strategies and technologies that are being used to improve oral vaccine absorption. Peptides and proteins, which comprise important vaccine components, exhibit unfavorable physicochemical properties including degradation in the gastrointestinal tract, and poor transport across the intestinal wall, which hinder oral vaccine development. Approaches to overcome these obstacles aim to provide new vaccines and delivery systems that are capable of eliciting protective immune responses, and are making an impact on current vaccine development.

A genital tract peptide epitope vaccine targeting TLR-2 efficiently induces local and systemic CD8+ T cells and protects against herpes simplex virus type 2 challenge

The next generation of needle-free mucosal vaccines is being rationally designed according to rules that govern the way in which the epitopes are recognized by and stimulate the genital mucosal immune system. We hypothesized that synthetic peptide epitopes extended with an agonist of Toll-like receptor 2 (TLR-2), that are abundantly expressed by dendritic and epithelial cells of the vaginal mucosa, would lead to induction of protective immunity against genital herpes. To test this hypothesis, we intravaginally (IVAG) immunized wild-type B6, TLR-2 (TLR2−/−) or myeloid differentiation factor 88 deficient (MyD88−/−) mice with a herpes simplex virus type 2 (HSV-2) CD8+ T-cell peptide epitope extended by a palmitic acid moiety (a TLR-2 agonist). IVAG delivery of the lipopeptide generated HSV-2-specific memory CD8+ cytotoxic T cells both locally in the genital tract draining lymph nodes and systemically in the spleen. Moreover, lipopeptide-immunized TLR2−/− and MyD88−/− mice developed significantly less HSV-specific CD8+ T-cell response, earlier death, faster disease progression, and higher vaginal HSV-2 titers compared to lipopeptide-immunized wild-type B6 mice. IVAG immunization with self-adjuvanting lipid-tailed peptides appears to be a novel mucosal vaccine approach, which has attractive practical and immunological features.