Abstract

1. Introduction

The introduction of vaccines to prevent infectious disease has had a transformational effect on human health. For example, the declaration of the global eradication of wild poliovirus type 2 was one of the highlights of the Global Commission for the Certification of Poliomyelitis Eradication (GCC) meeting in September 2015. Further, vaccination against measles, a highly contagious disease caused by morbillivirus in the paramyxovirus family has resulted in a 79% decrease in measles deaths globally from 2000 to 2014. Another example was the introduction of a meningitis A conjugate vaccine, which expedited the near elimination of the deadly disease in the African “meningitis belt”. These stunning accomplishments emphasize the importance and need for vaccines that profoundly contribute to well-being of society.

A vaccine can be generally defined as a biological preparation that contributes to active immunity against a particular disease. The pioneering work by Edward Jenner for the smallpox vaccination, followed by Louis Pasteur for an anthrax vaccine, lead to development of vaccines against many other infectious diseases. The World Health Organization (WHO) has reported vaccines for the prevention and control of 27 infections, with many other vaccines in the discovery pipeline. A candidate vaccine against malaria (RTS,S) recently completed phase III clinical trials involving approximately 15,000 infants and young children in seven sub-Saharan African countries. Two Ebola candidate vaccines are also in the final stages of clinical trials. The research community is also making sustained efforts toward the development of vaccines against diseases which are difficult to treat, such as cancer [1,2], hepatitis C, and tuberculosis [3,4,5].

2. Types of Vaccines

Vaccination involves exposure of an antigen, derived from a disease-causing agent, to the immune system with the aim to develop active immunity against the antigen. When the vaccinated individual comes in contact with the causative micro-organism, a strong protective immune response occurs. The optimal properties of any vaccine include long-lasting immunity, lack of autoimmunity or hypersensitivity, ease of administration, and storage. Additionally, vaccine components should be safe, and, specifically, the vaccine itself should not result in the disease state.

The different types of vaccines (Table 1) include live attenuated vaccines, which consist of live, weakened, or modified disease causing micro-organisms, which results in a limited infection that is sufficient to invoke immune response, but not long-lived enough to cause the actual disease state [6,7]. The attenuation is achieved by repeated culturing of disease causing agent in a foreign host. The less virulent mutant adapted to the foreign host can be used for vaccination. Killed/Inactivated vaccines work with help of different chemical methods, radiation, or heat. The pathogen is inactivated so it cannot replicate in the host and is used as the vaccinating agent [6,7]. Bacterial vaccines have generally used dead micro-organisms, while viral vaccines are composed of inactivated agents. Recombinant/DNA are experimental vaccines composed of genes encoding antigen inserted into a vector (recombinant vaccine) [6,7]. The DNA can be injected or attached to a carrier, such as metallic nanoparticles (DNA vaccine). In situ generation of antigenic protein can lead to strong immune response. Subunit vaccines contain a purified antigen instead of using whole microorganisms. Purified antigens could be toxoid, subcellular fragment, or surface molecules, which are transported by different carriers [6,7]. Immune response to subunit vaccine differs based on the antigen used. Protein antigens usually give rise to T-cell dependent adaptive immune response while polysaccharide antigens generate T-cell independent response. Conjugated vaccines can be defined as subclass of subunit vaccines as protein carriers are used to carry polysaccharide based antigen.

3. Immunology of Vaccines

Immunology is a science that studies the structure and function of the immune system, which itself is sub-classified into innate and adaptive immune systems. The innate immune system is non-specific and quickly forms the body’s first line of defense. Phagocytes, soluble peptides, and proteins are complement molecules, while natural killer (NK) cells are key elements of innate immune system. On the other hand, the adaptive immune system gives a pathogen or antigen a specific response along with immunological memory. The adaptive response takes days to weeks to develop. T lymphocytes and B lymphocytes are essential for the adaptive immune response.

Dendritic cells (DCs) play important roles in both innate as well as adaptive immunity. These antigen presenting cells (APCs) are the messengers that call for help from the adaptive immune response when an infection outruns innate immunity [8]. The immature dendritic cells function as phagocytes in innate immunity, participating in capture, uptake, and processing of antigen. While moving to secondary lymphoid tissue, these cells gain capacity to activate naïve T cells (CD4⁺ and CD8⁺). These APCs stop phagocytosis and present antigen on their surfaces with a high density during this migration. Such cells in lymph nodes are called mature or activated dendritic cells.

Activation of naïve T cells requires binding antigen to specific T cell receptors (TCR), along with a co-stimulatory signal. The receptor responsible for the co-stimulatory signal is called CD-28 and presents on T cells that bind to the B7 ligand on dendritic cells. Proliferation and differentiation of naïve T cells into effector cells is initiated by these intracellular signals. CD8⁺ T cells or CD4⁺ T cells form based on antigen presentation by either MHC-class I or MHC-class II molecules, respectively, on dendritic cells. In most cases, extracellular antigens are presented by MHC-class II on APCs, which in turns activate CD4⁺ T cells, also known as T helper cells. On the other hand, intracellular antigens presented by MHC-class I molecules on APCs lead to the activation of CD8⁺ T cells, also known as cytotoxic T cells.

Although the above-mentioned distinction is not absolute, the phenomenon of presentation of antigen located in the MHC-class II pathway to MHC-class I pathway is called “cross presentation”. Such presentation helps to develop both CD4⁺, as well as CD8⁺ response, against antigens. The understanding of cross-presentation is important to the success of producing an effective immune response to a vaccine [9]. There is a major focus on increasing cross-presentation of antigen in DCs targeting vaccines.

In the case of subunit vaccine preparation, polysaccharide antigens are poorly immunogenic. Therefore, immuno-adjuvants are added along with antigen in the vaccine carrier to augment the immune response. Another approach is to conjugate bacterial proteins to these antigens, as mentioned earlier. Immuno-adjuvants, especially Toll-like receptor (TLR) ligands are increasingly used in vaccine design [10,11]. Signals from TLRs on dendritic cells improve the processing and presentation by APCs. They also play a role in maturation of dendritic cells.

4. Minimal Subunit Vaccine Development

The classical approach of conjugating carbohydrate antigens to a protein, such as bovine serum albumin (BSA) and keyhole limpet hemocyanin (KLH), is reviewed elsewhere [12,13]. The problems associated with these conjugated vaccines include: Variation in antigen loading, immunogenic linkers, and antigenic carrier proteins lead to the development of minimal subunit vaccines. Research on these vaccines is happening in three areas: Minimal-peptide antigens, improved immune-adjuvant and the use of novel vaccine carriers.

Single molecule two-component vaccines with monomeric, trimeric tumor-associated carbohydrate antigens (TACAs), along with TLR ligands were developed [14,15,16,17,18]. To facilitate antibody class switching from IgM to IgG, a helper T-cell epitope was incorporated into a two-component vaccine to obtain a fully synthetic multi-component vaccine, including phospholipid-based liposomes [19,20], which serves as the carrier. Several polysaccharide antigens specific to cancer [21,22,23,24,25], HIV [26], and other infections [25,27], have been identified and incorporated into this type of vaccine. Investigating new immuno-adjuvants with good safety profiles is another way to potentially advance this platform. Adjuvants can enhancing antigen presentation through dendritic cell (DC) maturation, produce a depot effect, or induce APCs to release cytokines [28,29,30]. Different types of adjuvants, such as bacterial, gel-type, emulsifier, and synthetic, are known [31].

In recent years, advancement of vaccines has focused on one major area; i.e., vaccine carriers [32,33,34,35]. Liposomes, archeosomes, virosomes, virus-like particles (VLPs), polymeric, and inorganic micro-, nano-particles have all been developed. Some of these particulate carriers show adjuvant properties as well. Herein, we present recent case studies of advancements in these vaccine carriers with their physio-chemical attributes.

5. Liposomes

Liposomes are biocompatible, completely biodegradable, self-assembled vesicular structures composed of lipid bilayers. Liposomes have been extensively used as drug delivery vehicles for years, but, in last decade, their use as a vaccine carrier has increased (Figure 1). Considering their wide applications, liposomes have been extensively reviewed by many groups [36,37,38,39,40]. These reviews cover methods of preparation, physio-chemical properties, such as particle size, charge, lamellarity, and their effects on drug or vaccine delivery and different conjugation chemistry.

Open in a separate window

Figure 1

Variations with liposomal vaccine delivery system.

Liposomes are advantageous over other vaccine carriers due to their tolerability by the human body and lack of toxicity, along with their chemical and structural flexibility. Chemical flexibility refers to the ability of the liposome to encapsulate either a hydrophilic antigen or adjuvant or a lipophilic component, which can intercalate between the lipid molecules [41]. Surface conjugation of hydrophilic antigens is also possible, which facilitates a better uptake by phagocytes because of better accessibility of antigen [42,43,44,45,46]. Structural flexibility refers to the tuning of liposomal properties by modifying lipid composition. For example, the use of cationic lipids has increased, as cationic liposomes are known to improve cytosolic release of antigens by affecting endosomal membrane integrity [47,48,49,50].

There are different methods of preparation of liposomes, such as physical dispersion, solvent dispersion, and detergent solubilization, which are reviewed elsewhere [51,52,53,54]. The general procedure for liposomal formulation involves dissolution of lipid content in an organic solvent. The organic solvent is then evaporated to obtain a thin film of lipid. After drying, the film is hydrated with an aqueous system containing hydrophilic antigen and adjuvants. The resulting vesicular structures are then subjected to freeze-thaw cycles, sonication, or membrane extrusion to ensure entrapment efficiency, size, and lamellarity according to application. In the case of hydrophobic components, they are dissolved in organic solvents along with lipids.

6. Polymeric Nanoparticles

Polymeric nanoparticles are also under investigation for their potential use as vaccine carrier platforms [83,84,85,86,87,88]. Apart from similar advantages, such as biocompatibility, safety, and flexibility of liposomal carrier systems, a wide variety of natural, as well as synthetic, bio-degradable polymers and block co-polymers create opportunities to tailor and improve these materials (Figure 2). By varying nanoparticle size, shape, surface charge, type of polymer, and polymer concentration, researchers can optimize polymeric nano-particulate carriers. Recent advances in these areas are discussed further.

Open in a separate window

Figure 2

Polymers used in vaccine delivery.

7. Inorganic Nanoparticles

Recently, Zhang and co-workers presented promising CD8⁺ T cell results with polyelectrolyte multilayers (PEM) assembled on gold nanoparticles (AuNPs) [107]. PEMs are self-assembled structures constructed via layer-by-layer (LbL) deposition using electrostatic interactions between positively and negatively charged electrolytes. Positively charged antigen and negatively charged immuno-adjuvant on gold nanoparticles resulted in a new vaccine platform. Greater control over antigen loading is a key advantage in vaccine design. The concept of PEMs was further exploited by Chiu, et al. to develop vaccine capsules made up of PEMs without any vehicle [108]. Calcium carbonate was used as solid support for PEM deposition, which was later removed.

The ease of synthesis of these inorganic nanoparticles, with precise control over mono-dispersity, size and shape, higher cargo loading, and colloidal stability, outweigh some limitations, such as their non-biodegradability. Recently, AuNPs have been used in immunotherapy as they are inert and can be easily functionalized with desired molecules. Chiodo, et al. synthesized AuNPs with tetra- and pentamannosides in order to mimic clusters of HIV gp120 [109]. Similar efforts have been made to develop AuNP-based vaccines for cancer [110,111,112,113], influenza [114], malaria [115], FMD [116], and HIV [117], by conjugating respective antigens on the surface. Sungsuwan, et al. developed lipid-coated iron oxide nanaoparticles with mucin-1 (MUC-1) antigens for cancer therapy [118]. The antigen-modified lipid was used to coat iron oxide nanoparticles to bind antigens on the surface without covalent modification. Meningitis A capsular polysaccharide fragments carrying iron oxide particle have been recently developed by Ramella, et al. [119]. Carbon nanoparticles, carbon nanotubes, silica, as well as calcium phosphate nanoparticles, are also under investigation for vaccine development.

The optical and photothermal properties of inorganic particles have been exploited in drug delivery as a tumor imaging and targeting tool. Based on recent studies, these treatments can have applications in immune therapy [120,121,122]. Heat shock proteins and tumor antigens released from dying tumor cells can activate the immune system. Koboyashi, et al. observed anti-tumor immunity via magnetic-nanoparticle-induced hyperthermia [123]. Application of an external magnetic field to the targeted magnetic nanoparticles increases tumor cells temperature without causing any harm to normal cells. Along with expected tumor cell death, the treatment resulted in an unexpected tumor specific response. These observations could lead to treatment addressing disease states, as well as immunotherapy.

8. Conclusions and Future Directions

This review summarizes recent developments in particulate carries for subunit vaccines. Development of a wide variety of nanoparticles with target specific modifications has a profound impact on the efficacy of vaccines. Understanding the fluid dynamics of these carriers, based on their physio-chemical variations, is the key to a better bio-distribution and antigen presentation. With advances in technology, investigators can achieve better control over the synthetic parameters of these vehicles to ensure reduced toxicity, antigen stability, and enhanced immunogenicity.

With the availability of new multifunctional carriers, the possibilities for novel vaccines have been greatly expanded. Comparison of different adjuvant loadings may be more accessible with these newer material. The synergistic effects of multiple immuno adjuvants may be addressed with DoE and simple conjugation chemistry to these nano-carrier. Parallel immune response studies of different nano-carriers, having the same antigen loading and adjuvant, could be interesting and may help in further development of hybrid nano-particles. In conclusion, recent studies highlighted in this review represent a step forward in dealing with current challenges in vaccinology and present new directions for future vaccine design.

Acknowledgments

Abbreviations

Author Contributions

Abhishek Vartak and Prof.Steven Sucheck both wrote, revised and proofread this review article.

Conflicts of Interest

The authors declare no conflict of interest.

References