**1. DNA vaccine**

DNA vaccine is a third-generation vaccine, which encompasses a vector with eukaryotic cell promoter, and a gene, which encodes for an immunogenic protein. These vaccines have been shown to elicit a robust cytotoxic T cell in comparison with subunit vaccines. Also, DNA vaccine has the capacity to induce both cellular and humoral immune responses by utilizing MHC I and MHC II antigen presentation by DCs [1–3]. Although DNA vaccines are licensed for use in veterinary vaccines since 2005, they have their own limitation due to low transfection efficiency. As a result, they perform poorly in human clinical trials and require multiple booster doses to achieve desirable immune response [2, 4, 5]. With the advent of new adjuvant systems such as nanoparticles, the immunogenicity of DNA vaccines can be enhanced considerably [1]. DNA vaccines are being currently used against a wide variety of infectious diseases as well as cancer [3].

#### **1.1 Antigen presentation to T lymphocytes**

Surface receptors of T lymphocytes interact with antigens to mount an immune response [6]. For the antigenic features; alienation, molecular weight, the delivery route to the organism is very important. At the molecular level, these receptors interact with the phagocyte cell or infected target cell, which carries antigen on its surface bound to the major histocompatibility complex (MHC). While T cells do not recognize natural antigens, antigens must first be processed by antigen-presenting cells (APCs) and then presented to the T cells with the relevant MHC protein. T cells (cytotoxic T cells—Tc) with CD8 (cluster of differentiation 8) receptors recognize MHC I antigens, while T cells (helper T cells—Th) with CD4 receptor recognize antigens on MHC II. Macrophages are the first identified antigen-presenting cells. Then, dendritic cells and B cells were identified. T-cell receptors (TCRs) are proteins that have spread through the membrane extending from the cell surface to the outer periphery. Each cell carries thousands of the same receptor surface. TCRs recognize and bind only bound peptide antigens on MHC [7, 8].

#### *1.1.1 Major histocompatibility complex (MHC)*

MHC proteins are encoded by the respective gene in the genome of all the vertebrate animals. The human MHC proteins are called human leukocyte antigens (HLAs) (human MHC I antigens, HLA-A, B and C, human MHC II antigens, HLA-DR, DQ and DP). Because of the difference in MHC proteins between the tissue donor and recipient, they were first discovered with tissue transplant rejection. Even within a species, these proteins are not structurally the same because of differences in amino acid sequences, also known as polymorphisms. For this reason, it forms an important antigenic barrier in organ transplantation. MHC genes encode two classes of MHC proteins, class I and class II. While MHC class I protein is located on the surfaces of all the nucleated cells, MHC class II protein is located only on the surface of antigen-presenting cells (APCs), including B lymphocytes, macrophages and dendritic cells. The structure of the MHC I protein comprises of a relatively small size of the β-2 microglobulin protein with α1, α2 and α3 domains linked to each other by disulfide bonds. The α1 and α2 domains constitute the variable antigen-binding domain. MHC II protein is formed by α1, β1, α2 and β2 domains, each of which is attached to one of the non-covalent bonds, and α1 and β1 domains constitute the antigen-binding domain, which is a variable part [7, 9, 10].

MHC proteins carry proteins in the cell which they are in, on themself. Thus, if the cell is not infected, it will carry its own peptides on the MHC proteins. On the other hand, if the cell harbors a foreign pathogen or protein, it will contain foreign peptides on MHC proteins. The function of these MHC proteins is to allow T cells to recognize foreign antigen. The T cells constantly control the surface of other cells for foreign antigen presence and do not recognize foreign antigens unless they are presented through MHC proteins. No T cell interacts with MHC on a healthy cell surface, and self-attacking cells are eliminated during the development of tolerance. During antigen presentation, the MHC and peptide complex come out of the cell membrane and thus are recognized by the T cells. There are two different antigen presentations to T cells, MHC I and MHC II antigen presentation [7–9].

#### *1.1.2 Antigen presentation via MHC I protein*

Peptides that are processed endogenously in the cytoplasm of non-phagocytic cells are presented with MHC I proteins. These antigens are derived from viruses or intracellular pathogens that infect cells, also known as internal antigens. In this

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DNA vaccines [14].

targeting to desired cells [9, 16].

*Current State of the Art in DNA Vaccine Delivery and Molecular Adjuvants: Bcl-xL…*

pathway, for example, in a virus-infected cell, virus-associated proteins are primarily digested in the cytoplasmic proteasome. Peptide antigens of about 10 amino acids are delivered to the endoplasmic reticulum (ER), and a pore protein (TAP) produced by the two proteins acts in this stage. The peptides are bound to the MHC I protein, which results in the chaperone protein, which retains the MHC protein at that site. The resulting complex is released from the ER and goes to the cell surface and integrates into the membrane. This complex is recognized and bound by cytotoxic T cells. The CD8 receptor on the Tc cell surface strengthens it by adding the binding complex. This binding allows cytotoxic T cells to produce perforin and

The MHC II protein is produced only in cells that present phagocytic antigen. For example, if an extracellular pathogen such as a bacterium is engulfed and then peptide antigens are provided, then MHC II proteins are produced in the ER and accumulated therein blocked with Li proteins, which inhibit binding with other peptides. The MHC II-Li protein complex is transferred to lysozyme and then combined with the phagosome to form the phagolysosome. There are foreign pathogen antigens, pathogenic proteins including connective chaperones are digested to form peptides of 10–15 amino acids. The resulting pathogenic peptides are transferred to the cell surface by binding with MHC II. This complex is recognized by the TCR on the helper T cells; the CD4 coreceptor also binds to this complex and, through interaction with the Th cells, activates to produce cytokines. The produced cytokine activates antibody

The structure of plasmid DNA provides an advantage over other traditional protein-based or carbohydrate-based grafts in which it inherently possesses DNA vaccination. Immunogenesis from DNA vaccination takes a long time, and there is no pathogenicity caused by inactivated virus in DNA vaccines. The vaccine containing plasmid DNA (pDNA) can encode many immunogenic proteins of the same virus and can also encode similar proteins belonging to different infective agents [12]. Another important advantage is that the production of DNA vaccines is easier when compared to recombinant protein vaccines [13]. DNA vaccines prepared to make the cytokines more desirable to direct the immune system are more potent and suitable for stimulating the type of immunological response by cloning genes encoding the target cytokine and antigens into the same expression plasmids. DNA vaccines lead to sustained stimulation of antigen expression and the immune response leading to prolonged protective immunity [7]. Easy construction and manipulation of plasmid DNA are another important advantage of

DNA vaccines are stable at room temperature, are easy to obtain, are economical and are relatively more reliable than other vaccines. Plasmid DNA vaccines have been shown to be highly evaluated, safe and immunogenic in human clinical trials, even though they have no mental status [15]. However, it is necessary to increase transfection efficiencies of naked DNA vaccines, facilitate intracellular uptake, target into cells, and perform these operations with small amounts of DNA. Various gene delivery systems and adjuvant systems in the nanosphere are used to overcome these problems. During the use of nanotechnological adjuvant systems, the degradation of DNA is prevented, resulting in ultra-rapid delivery by

*DOI: http://dx.doi.org/10.5772/intechopen.82203*

cytotoxic proteins that kill infected cells [7, 8, 10, 11].

production by specific B-cell clones or causes inflammation [7, 8, 11].

**1.2 Advantages and disadvantages of DNA vaccination**

*1.1.3 Antigen presentation via MHC II protein*

*Current State of the Art in DNA Vaccine Delivery and Molecular Adjuvants: Bcl-xL… DOI: http://dx.doi.org/10.5772/intechopen.82203*

pathway, for example, in a virus-infected cell, virus-associated proteins are primarily digested in the cytoplasmic proteasome. Peptide antigens of about 10 amino acids are delivered to the endoplasmic reticulum (ER), and a pore protein (TAP) produced by the two proteins acts in this stage. The peptides are bound to the MHC I protein, which results in the chaperone protein, which retains the MHC protein at that site. The resulting complex is released from the ER and goes to the cell surface and integrates into the membrane. This complex is recognized and bound by cytotoxic T cells. The CD8 receptor on the Tc cell surface strengthens it by adding the binding complex. This binding allows cytotoxic T cells to produce perforin and cytotoxic proteins that kill infected cells [7, 8, 10, 11].

### *1.1.3 Antigen presentation via MHC II protein*

*Immune Response Activation and Immunomodulation*

**1.1 Antigen presentation to T lymphocytes**

*1.1.1 Major histocompatibility complex (MHC)*

recognize and bind only bound peptide antigens on MHC [7, 8].

MHC proteins are encoded by the respective gene in the genome of all the vertebrate animals. The human MHC proteins are called human leukocyte antigens (HLAs) (human MHC I antigens, HLA-A, B and C, human MHC II antigens, HLA-DR, DQ and DP). Because of the difference in MHC proteins between the tissue donor and recipient, they were first discovered with tissue transplant rejection. Even within a species, these proteins are not structurally the same because of differences in amino acid sequences, also known as polymorphisms. For this reason, it forms an important antigenic barrier in organ transplantation. MHC genes encode two classes of MHC proteins, class I and class II. While MHC class I protein is located on the surfaces of all the nucleated cells, MHC class II protein is located only on the surface of antigen-presenting cells (APCs), including B lymphocytes, macrophages and dendritic cells. The structure of the MHC I protein comprises of a relatively small size of the β-2 microglobulin protein with α1, α2 and α3 domains linked to each other by disulfide bonds. The α1 and α2 domains constitute the variable antigen-binding domain. MHC II protein is formed by α1, β1, α2 and β2 domains, each of which is attached to one of the non-covalent bonds, and α1 and β1 domains constitute the antigen-binding domain, which is a variable part [7, 9, 10]. MHC proteins carry proteins in the cell which they are in, on themself. Thus, if the cell is not infected, it will carry its own peptides on the MHC proteins. On the other hand, if the cell harbors a foreign pathogen or protein, it will contain foreign peptides on MHC proteins. The function of these MHC proteins is to allow T cells to recognize foreign antigen. The T cells constantly control the surface of other cells for foreign antigen presence and do not recognize foreign antigens unless they are presented through MHC proteins. No T cell interacts with MHC on a healthy cell surface, and self-attacking cells are eliminated during the development of tolerance. During antigen presentation, the MHC and peptide complex come out of the cell membrane and thus are recognized by the T cells. There are two different antigen

presentations to T cells, MHC I and MHC II antigen presentation [7–9].

Peptides that are processed endogenously in the cytoplasm of non-phagocytic cells are presented with MHC I proteins. These antigens are derived from viruses or intracellular pathogens that infect cells, also known as internal antigens. In this

*1.1.2 Antigen presentation via MHC I protein*

Surface receptors of T lymphocytes interact with antigens to mount an immune response [6]. For the antigenic features; alienation, molecular weight, the delivery route to the organism is very important. At the molecular level, these receptors interact with the phagocyte cell or infected target cell, which carries antigen on its surface bound to the major histocompatibility complex (MHC). While T cells do not recognize natural antigens, antigens must first be processed by antigen-presenting cells (APCs) and then presented to the T cells with the relevant MHC protein. T cells (cytotoxic T cells—Tc) with CD8 (cluster of differentiation 8) receptors recognize MHC I antigens, while T cells (helper T cells—Th) with CD4 receptor recognize antigens on MHC II. Macrophages are the first identified antigen-presenting cells. Then, dendritic cells and B cells were identified. T-cell receptors (TCRs) are proteins that have spread through the membrane extending from the cell surface to the outer periphery. Each cell carries thousands of the same receptor surface. TCRs

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The MHC II protein is produced only in cells that present phagocytic antigen. For example, if an extracellular pathogen such as a bacterium is engulfed and then peptide antigens are provided, then MHC II proteins are produced in the ER and accumulated therein blocked with Li proteins, which inhibit binding with other peptides. The MHC II-Li protein complex is transferred to lysozyme and then combined with the phagosome to form the phagolysosome. There are foreign pathogen antigens, pathogenic proteins including connective chaperones are digested to form peptides of 10–15 amino acids. The resulting pathogenic peptides are transferred to the cell surface by binding with MHC II. This complex is recognized by the TCR on the helper T cells; the CD4 coreceptor also binds to this complex and, through interaction with the Th cells, activates to produce cytokines. The produced cytokine activates antibody production by specific B-cell clones or causes inflammation [7, 8, 11].

#### **1.2 Advantages and disadvantages of DNA vaccination**

The structure of plasmid DNA provides an advantage over other traditional protein-based or carbohydrate-based grafts in which it inherently possesses DNA vaccination. Immunogenesis from DNA vaccination takes a long time, and there is no pathogenicity caused by inactivated virus in DNA vaccines. The vaccine containing plasmid DNA (pDNA) can encode many immunogenic proteins of the same virus and can also encode similar proteins belonging to different infective agents [12]. Another important advantage is that the production of DNA vaccines is easier when compared to recombinant protein vaccines [13]. DNA vaccines prepared to make the cytokines more desirable to direct the immune system are more potent and suitable for stimulating the type of immunological response by cloning genes encoding the target cytokine and antigens into the same expression plasmids. DNA vaccines lead to sustained stimulation of antigen expression and the immune response leading to prolonged protective immunity [7]. Easy construction and manipulation of plasmid DNA are another important advantage of DNA vaccines [14].

DNA vaccines are stable at room temperature, are easy to obtain, are economical and are relatively more reliable than other vaccines. Plasmid DNA vaccines have been shown to be highly evaluated, safe and immunogenic in human clinical trials, even though they have no mental status [15]. However, it is necessary to increase transfection efficiencies of naked DNA vaccines, facilitate intracellular uptake, target into cells, and perform these operations with small amounts of DNA. Various gene delivery systems and adjuvant systems in the nanosphere are used to overcome these problems. During the use of nanotechnological adjuvant systems, the degradation of DNA is prevented, resulting in ultra-rapid delivery by targeting to desired cells [9, 16].
