DNA vaccine

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The making of a DNA vaccine Making of a DNA vaccine.jpg
The making of a DNA vaccine

A DNA vaccine is a type of vaccine that transfects a specific antigen-coding DNA sequence into the cells of an organism as a mechanism to induce an immune response. [1] [2]

Contents

DNA vaccines work by injecting genetically engineered plasmid containing the DNA sequence encoding the antigen(s) against which an immune response is sought, so the cells directly produce the antigen, thus causing a protective immunological response. [3] DNA vaccines have theoretical advantages over conventional vaccines, including the "ability to induce a wider range of types of immune response". [4] Several DNA vaccines have been tested for veterinary use. [3] In some cases, protection from disease in animals has been obtained, in others not. [3] Research is ongoing over the approach for viral, bacterial and parasitic diseases in humans, as well as for cancers. [4] In August 2021, Indian authorities gave emergency approval to ZyCoV-D. Developed by Cadila Healthcare, it is the first DNA vaccine approved for humans. [5]

History

Conventional vaccines contain either specific antigens from a pathogen, or attenuated viruses which stimulate an immune response in the vaccinated organism. DNA vaccines are members of the genetic vaccines, because they contain a genetic information (DNA or RNA) that codes for the cellular production (protein biosynthesis) of an antigen. DNA vaccines contain DNA that codes for specific antigens from a pathogen. The DNA is injected into the body and taken up by cells, whose normal metabolic processes synthesize proteins based on the genetic code in the plasmid that they have taken up. Because these proteins contain regions of amino acid sequences that are characteristic of bacteria or viruses, they are recognized as foreign and when they are processed by the host cells and displayed on their surface, the immune system is alerted, which then triggers immune responses. [6] [7] Alternatively, the DNA may be encapsulated in protein to facilitate cell entry. If this capsid protein is included in the DNA, the resulting vaccine can combine the potency of a live vaccine without reversion risks.[ citation needed ]

In 1983, Enzo Paoletti and Dennis Panicali at the New York Department of Health devised a strategy to produce recombinant DNA vaccines by using genetic engineering to transform ordinary smallpox vaccine into vaccines that may be able to prevent other diseases. [8] They altered the DNA of cowpox virus by inserting a gene from other viruses (namely Herpes simplex virus, hepatitis B and influenza). [9] [10] In 1993, Jeffrey Ulmer and co-workers at Merck Research Laboratories demonstrated that direct injection of mice with plasmid DNA encoding a flu antigen protected the animals against subsequent experimental infection with influenza virus. [11] In 2016 a DNA vaccine for the Zika virus began testing in humans at the National Institutes of Health. The study was planned to involve up to 120 subjects aged between 18 and 35. Separately, Inovio Pharmaceuticals and GeneOne Life Science began tests of a different DNA vaccine against Zika in Miami. The NIH vaccine is injected into the upper arm under high pressure. Manufacturing the vaccines in volume remained unsolved as of August 2016. [12] Clinical trials for DNA vaccines to prevent HIV are underway. [13]

In August 2021, Indian authorities gave emergency approval to ZyCoV-D. Developed by Cadila Healthcare, it is the first DNA vaccine against COVID-19. [5]

Applications

As of 2021 no DNA vaccines have been approved for human use in the United States. Few experimental trials have evoked a response strong enough to protect against disease and the technique's usefulness remains to be proven in humans.

A veterinary DNA vaccine to protect horses from West Nile virus has been approved. [14] Another West Nile virus vaccine has been tested successfully on American robins. [15]

DNA immunization is also being investigated as a means of developing antivenom sera. [1] DNA immunization can be used as a technology platform for monoclonal antibody induction. [2]

Advantages

Disadvantages

Plasmid vectors

Vector design

DNA vaccines elicit the best immune response when high-expression vectors are used. These are plasmids that usually consist of a strong viral promoter to drive the in vivo transcription and translation of the gene (or complementary DNA) of interest. [18] Intron A may sometimes be included to improve mRNA stability and hence increase protein expression. [19] Plasmids also include a strong polyadenylation/transcriptional termination signal, such as bovine growth hormone or rabbit beta-globulin polyadenylation sequences. [6] [7] [20] Polycistronic vectors (with multiple genes of interest) are sometimes constructed to express more than one immunogen, or to express an immunogen and an immunostimulatory protein. [21]

Because the plasmid –carrying relatively small genetic code up to about 200 Kbp  –is the "vehicle" from which the immunogen is expressed, optimising vector design for maximal protein expression is essential. [21] One way of enhancing protein expression is by optimising the codon usage of pathogenic mRNAs for eukaryotic cells. Pathogens often have different AT-contents than the target species, so altering the gene sequence of the immunogen to reflect the codons more commonly used in the target species may improve its expression. [22]

Another consideration is the choice of promoter. The SV40 promoter was conventionally used until research showed that vectors driven by the Rous Sarcoma Virus (RSV) promoter had much higher expression rates. [6] More recently, expression and immunogenicity have been further increased in model systems by the use of the cytomegalovirus (CMV) immediate early promoter, and a retroviral cis-acting transcriptional element. [23] Additional modifications to improve expression rates include the insertion of enhancer sequences, synthetic introns, adenovirus tripartite leader (TPL) sequences and modifications to the polyadenylation and transcriptional termination sequences. [6] An example of DNA vaccine plasmid is pVAC, which uses SV40 promoter.

Structural instability phenomena are of particular concern for plasmid manufacture, DNA vaccination and gene therapy. [24] Accessory regions pertaining to the plasmid backbone may engage in a wide range of structural instability phenomena. Well-known catalysts of genetic instability include direct, inverted and tandem repeats, which are conspicuous in many commercially available cloning and expression vectors. Therefore, the reduction or complete elimination of extraneous noncoding backbone sequences would pointedly reduce the propensity for such events to take place and consequently the overall plasmid's recombinogenic potential. [25]

Mechanism of plasmids

Once the plasmid inserts itself into the transfected cell nucleus, it codes for a peptide string of a foreign antigen. On its surface the cell displays the foreign antigen with both histocompatibility complex (MHC) classes I and class II molecules. The antigen-presenting cell then travels to the lymph nodes and presents the antigen peptide and costimulatory molecule signalling to T-cell, initiating the immune response. [26]

Vaccine insert design

Immunogens can be targeted to various cellular compartments to improve antibody or cytotoxic T-cell responses. Secreted or plasma membrane-bound antigens are more effective at inducing antibody responses than cytosolic antigens, while cytotoxic T-cell responses can be improved by targeting antigens for cytoplasmic degradation and subsequent entry into the major histocompatibility complex (MHC) class I pathway. [7] This is usually accomplished by the addition of N-terminal ubiquitin signals. [27] [28] [29]

The conformation of the protein can also affect antibody responses. "Ordered" structures (such as viral particles) are more effective than unordered structures. [30] Strings of minigenes (or MHC class I epitopes) from different pathogens raise cytotoxic T-cell responses to some pathogens, especially if a TH epitope is also included. [7]

Delivery

DNA vaccine and Gene therapy techniques are similar. Gene therapy.jpg
DNA vaccine and Gene therapy techniques are similar.

DNA vaccines have been introduced into animal tissues by multiple methods. In 1999, the two most popular approaches were injection of DNA in saline: by using a standard hypodermic needle, or by using a gene gun delivery. [31] Several other techniques have been documented in the intervening years.

Saline injection

Injection in saline is normally conducted intramuscularly (IM) in skeletal muscle, or intradermally (ID), delivering DNA to extracellular spaces. This can be assisted either 1) by electroporation; [32] 2) by temporarily damaging muscle fibres with myotoxins such as bupivacaine; or 3) by using hypertonic solutions of saline or sucrose. [6] Immune responses to this method can be affected by factors including needle type, [16] needle alignment, speed of injection, volume of injection, muscle type, and age, sex and physiological condition of the recipient. [6]

Gene gun

Gene gun delivery ballistically accelerates plasmid DNA (pDNA) that has been absorbed onto gold or tungsten microparticles into the target cells, using compressed helium as an accelerant. [6] [21]

Mucosal surface delivery

Alternatives included aerosol instillation of naked DNA on mucosal surfaces, such as the nasal and lung mucosa, [21] and topical administration of pDNA to the eye [33] and vaginal mucosa. [21] Mucosal surface delivery has also been achieved using cationic liposome-DNA preparations, [7] biodegradable microspheres, [34] [21] attenuated Salmonalla , [35] Shigella or Listeria vectors for oral administration to the intestinal mucosa [36] and recombinant adenovirus vectors. [21]

Polymer vehicle

A hybrid vehicle composed of bacteria cell and synthetic polymers has been employed for DNA vaccine delivery. An E. coli inner core and poly(beta-amino ester) outer coat function synergistically to increase efficiency by addressing barriers associated with antigen-presenting cell gene delivery which include cellular uptake and internalization, phagosomal escape and intracellular cargo concentration.[ jargon ] Tested in mice, the hybrid vector was found to induce immune response. [37] [38]

ELI immunization

Another approach to DNA vaccination is expression library immunization (ELI). Using this technique, potentially all the genes from a pathogen can be delivered at one time, which may be useful for pathogens that are difficult to attenuate or culture. [6] ELI can be used to identify which genes induce a protective response. This has been tested with Mycoplasma pulmonis, a murine lung pathogen with a relatively small genome. Even partial expression libraries can induce protection from subsequent challenge. [39]

Helpful tabular comparison

Table 2. Summary of plasmid DNA delivery methods
Method of deliveryFormulation of DNATarget tissueAmount of DNA
ParenteralInjection (hypodermic needle)Aqueous solution in saline IM (skeletal); ID; (IV, subcutaneous and intraperitoneal with variable success)Large amounts (approximately 100-200 μg)
Gene gunDNA-coated gold beadsED (abdominal skin); vaginal mucosa; surgically exposed muscle and other organsSmall amounts (as little as 16 ng)
Pneumatic (jet) injectionAqueous solutionEDVery high (as much as 300 μg)
Topical applicationAqueous solutionOcular; intravaginalSmall amounts (up to 100 μg)
Cytofectin-mediated[ jargon ] Liposomes (cationic); microspheres; recombinant adenovirus vectors; attenuated Shigella vector; aerosolised cationic lipid formulationsIM; IV (to transfect tissues systemically); intraperitoneal; oral immunization to the intestinal mucosa; nasal/lung mucosal membranesvariable
Table 3. Advantages and disadvantages of commonly used DNA vaccine delivery methods
Method of deliveryAdvantageDisadvantage
Intramuscular or Intradermal injection
  • No special delivery mechanism
  • Permanent or semi-permanent expression
  • pDNA spreads rapidly throughout the body
  • Inefficient site for uptake due to morphology of muscle tissue
  • Relatively large amounts of DNA used
  • Th1 response may not be the response required
Gene gun
  • DNA bombarded directly into cells
  • Small amounts DNA
  • Th2 response may not be the response required
  • Requires inert particles as carrier
Jet injection
  • No particles required
  • DNA can be delivered to cells mm to cm below skin surface
  • Significant shearing of DNA after high-pressure expulsion
  • 10-fold lower expression, and lower immune response
  • Requires large amounts of DNA (up to 300 μg)
Liposome-mediated delivery
  • High levels of immune response can be generated
  • Can increase transfection of intravenously delivered pDNA
  • Intravenously delivered liposome-DNA complexes can potentially transfect all tissues
  • Intranasally delivered liposome-DNA complexes can result in expression in distal mucosa as well as nasal muscosa and the generation of IgA antibodies

Dosage

The delivery method determines the dose required to raise an effective immune response. Saline injections require variable amounts of DNA, from 10 μg to 1 mg, whereas gene gun deliveries require 100 to 1000 times less. [40] Generally, 0.2 μg – 20 μg are required, although quantities as low as 16 ng have been reported. [6] These quantities vary by species. Mice for example, require approximately 10 times less DNA than primates. [7] Saline injections require more DNA because the DNA is delivered to the extracellular spaces of the target tissue (normally muscle), where it has to overcome physical barriers (such as the basal lamina and large amounts of connective tissue) before it is taken up by the cells, while gene gun deliveries drive/force DNA directly into the cells, resulting in less "wastage". [6] [7]

Immune response

Helper T cell responses

Antigen presentation stimulates T cells to become either "cytotoxic" CD8+ cells or "helper" CD4+ cells. Cytotoxic cells directly attack other cells carrying certain foreign or abnormal molecules on their surfaces. Helper T cells, or Th cells, coordinate immune responses by communicating with other cells. In most cases, T cells only recognize an antigen if it is carried on the surface of a cell by one of the body's own MHC, or major histocompatibility complex, molecules. Antigen presentation.svg
Antigen presentation stimulates T cells to become either "cytotoxic" CD8+ cells or "helper" CD4+ cells. Cytotoxic cells directly attack other cells carrying certain foreign or abnormal molecules on their surfaces. Helper T cells, or Th cells, coordinate immune responses by communicating with other cells. In most cases, T cells only recognize an antigen if it is carried on the surface of a cell by one of the body's own MHC, or major histocompatibility complex, molecules.

DNA immunization can raise multiple TH responses, including lymphoproliferation and the generation of a variety of cytokine profiles. A major advantage of DNA vaccines is the ease with which they can be manipulated to bias the type of T-cell help towards a TH1 or TH2 response. [41] Each type has distinctive patterns of lymphokine and chemokine expression, specific types of immunoglobulins, patterns of lymphocyte trafficking and types of innate immune responses.

Other types of T-cell help

The type of T-cell help raised is influenced by the delivery method and the type of immunogen expressed, as well as the targeting of different lymphoid compartments. [6] [42] Generally, saline needle injections (either IM or ID) tend to induce TH1 responses, while gene gun delivery raises TH2 responses. [41] [42] This is true for intracellular and plasma membrane-bound antigens, but not for secreted antigens, which seem to generate TH2 responses, regardless of the method of delivery. [43]

Generally the type of T-cell help raised is stable over time, and does not change when challenged or after subsequent immunizations that would normally have raised the opposite type of response in a naïve specimen. [41] [42] However, Mor et al.. (1995) [18] immunized and boosted mice with pDNA encoding the circumsporozoite protein of the mouse malarial parasite Plasmodium yoelii (PyCSP) and found that the initial TH2 response changed, after boosting, to a TH1 response.

Basis for different types of T-cell help

How these different methods operate, the forms of antigen expressed, and the different profiles of T-cell help is not understood. It was thought that the relatively large amounts of DNA used in IM injection were responsible for the induction of TH1 responses. However, evidence shows no dose-related differences in TH type. [41] The type of T-cell help raised is determined by the differentiated state of antigen presenting cells. Dendritic cells can differentiate to secrete IL-12 (which supports TH1 cell development) or IL-4 (which supports TH2 responses). [44] pDNA injected by needle is endocytosed into the dendritic cell, which is then stimulated to differentiate for TH1 cytokine (IL-12) production, [45] while the gene gun bombards the DNA directly into the cell, thus bypassing TH1 stimulation.

Practical uses of polarised T-cell help

Polarisation in T-cell help is useful in influencing allergic responses and autoimmune diseases. In autoimmune diseases, the goal is to shift the self-destructive TH1 response (with its associated cytotoxic T cell activity) to a non-destructive TH2 response. This has been successfully applied in predisease priming for the desired type of response in preclinical models [7] and is somewhat successful in shifting the response for an established disease. [46]

Cytotoxic T-cell responses

One of the advantages of DNA vaccines is that they are able to induce cytotoxic T lymphocytes (CTL) without the inherent risk associated with live vaccines. CTL responses can be raised against immunodominant and immunorecessive CTL epitopes, [47] as well as subdominant CTL epitopes, [34] [ jargon ] in a manner that appears to mimic natural infection. This may prove to be a useful tool in assessing CTL epitopes and their role in providing immunity.

Cytotoxic T-cells recognise small peptides (8-10 amino acids) complexed to MHC class I molecules. [48] These peptides are derived from cytosolic proteins that are degraded and delivered to the nascent MHC class I molecule within the endoplasmic reticulum (ER). [48] Targeting gene products directly to the ER (by the addition of an ER insertion signal sequence at the N-terminus) should thus enhance CTL responses. This was successfully demonstrated using recombinant vaccinia viruses expressing influenza proteins, [48] but the principle should also be applicable to DNA vaccines. Targeting antigens for intracellular degradation (and thus entry into the MHC class I pathway) by the addition of ubiquitin signal sequences, or mutation of other signal sequences, was shown to be effective at increasing CTL responses. [28]

CTL responses can be enhanced by co-inoculation with co-stimulatory molecules such as B7-1 or B7-2 for DNA vaccines against influenza nucleoprotein, [47] [49] or GM-CSF for DNA vaccines against the murine malaria model P. yoelii . [50] Co-inoculation with plasmids encoding co-stimulatory molecules IL-12 and TCA3 were shown to increase CTL activity against HIV-1 and influenza nucleoprotein antigens. [49] [51]

Humoral (antibody) response

Schematic diagram of an antibody and antigens Antibody.svg
Schematic diagram of an antibody and antigens

Antibody responses elicited by DNA vaccinations are influenced by multiple variables, including antigen type; antigen location (i.e. intracellular vs. secreted); number, frequency and immunization dose; site and method of antigen delivery.

Kinetics of antibody response

Humoral responses after a single DNA injection can be much longer-lived than after a single injection with a recombinant protein. Antibody responses against hepatitis B virus (HBV) envelope protein (HBsAg) have been sustained for up to 74 weeks without boost, while lifelong maintenance of protective response to influenza haemagglutinin was demonstrated in mice after gene gun delivery. [52] Antibody-secreting cells (ASC) migrate to the bone marrow and spleen for long-term antibody production, and generally localise there after one year. [52]

Comparisons of antibody responses generated by natural (viral) infection, immunization with recombinant protein and immunization with pDNA are summarised in Table 4. DNA-raised antibody responses rise much more slowly than when natural infection or recombinant protein immunization occurs. As many as 12 weeks may be required to reach peak titres in mice, although boosting can decrease the interval. This response is probably due to the low levels of antigen expressed over several weeks, which supports both primary and secondary phases of antibody response.[ clarification needed ] DNA vaccine expressing HBV small and middle envelope protein was injected into adults with chronic hepatitis. The vaccine resulted in specific interferon gamma cell production. Also specific T-cells for middle envelop proteins antigens were developed. The immune response of the patients was not robust enough to control HBV infection [53]

Table 4. Comparison of T-dependent antibody responses raise by DNA immunisations, protein inoculations and viral infections
 Method of Immunization
DNA vaccineRecombinant proteinNatural infection
Amount of inducing antigenngμg ? (ng-μg)
Duration of antigen presentationseveral weeks< 1 weekseveral weeks
Kinetics of antibody responseslow riserapid riserapid rise
Number of inoculations to obtain high avidity IgG and migration of ASC to bone marrowonetwoone
Ab isotype (murine models)C’-dependent or C’-independentC’-dependentC’-independent

Additionally, the titres of specific antibodies raised by DNA vaccination are lower than those obtained after vaccination with a recombinant protein. However, DNA immunization-induced antibodies show greater affinity to native epitopes than recombinant protein-induced antibodies. In other words, DNA immunization induces a qualitatively superior response. Antibodies can be induced after one vaccination with DNA, whereas recombinant protein vaccinations generally require a boost. DNA immunization can be used to bias the TH profile of the immune response and thus the antibody isotype, which is not possible with either natural infection or recombinant protein immunization. Antibody responses generated by DNA are useful as a preparative tool. For example, polyclonal and monoclonal antibodies can be generated for use as reagents.[ citation needed ]

Mechanistic basis for DNA-raised immune responses

DNA uptake mechanism

When DNA uptake and subsequent expression was first demonstrated in vivo in muscle cells, [54] these cells were thought to be unique because of their extensive network of T-tubules. Using electron microscopy, it was proposed that DNA uptake was facilitated by caveolae (or, non-clathrin coated pits). [55] However, subsequent research revealed that other cells (such as keratinocytes, fibroblasts and epithelial Langerhans cells) could also internalize DNA. [46] [56] The mechanism of DNA uptake is not known.

Two theories dominate – that in vivo uptake of DNA occurs non-specifically, in a method similar to phago- or pinocytosis, [21] or through specific receptors. [57] These might include a 30kDa surface receptor, or macrophage scavenger receptors.[ clarification needed ] The 30kDa surface receptor binds specifically to 4500-bp DNA fragments (which are then internalised) and is found on professional APCs and T-cells. Macrophage scavenger receptors bind to a variety of macromolecules, including polyribonucleotides and are thus candidates for DNA uptake. [57] [58] Receptor-mediated DNA uptake could be facilitated by the presence of polyguanylate sequences.[ clarification needed ][ citation needed ] Gene gun delivery systems, cationic liposome packaging, and other delivery methods bypass this entry method, but understanding it may be useful in reducing costs (e.g. by reducing the requirement for cytofectins), which could be important in animal husbandry.

Antigen presentation by bone marrow-derived cells

A dendritic cell Dendritic cell.JPG
A dendritic cell

Studies using chimeric mice have shown that antigen is presented by bone-marrow derived cells, which include dendritic cells, macrophages and specialised B-cells called professional antigen presenting cells (APC). [49] [59] After gene gun inoculation to the skin, transfected Langerhans cells migrate to the draining lymph node to present antigens. [7] After IM and ID injections, dendritic cells present antigen in the draining lymph node [56] and transfected macrophages have been found in the peripheral blood. [60]

Besides direct transfection of dendritic cells or macrophages, cross priming occurs following IM, ID and gene gun DNA deliveries. Cross-priming occurs when a bone marrow-derived cell presents peptides from proteins synthesised in another cell in the context of MHC class 1. This can prime cytotoxic T-cell responses and seems to be important for a full primary immune response. [7] [61]

Target site role

IM and ID DNA delivery initiate immune responses differently. In the skin, keratinocytes, fibroblasts and Langerhans cells take up and express antigens and are responsible for inducing a primary antibody response. Transfected Langerhans cells migrate out of the skin (within 12 hours) to the draining lymph node where they prime secondary B- and T-cell responses. In skeletal muscle, striated muscle cells are most frequently transfected, but seem to be unimportant in immune response. Instead, IM inoculated DNA "washes" into the draining lymph node within minutes, where distal dendritic cells are transfected and then initiate an immune response. Transfected myocytes seem to act as a "reservoir" of antigen for trafficking professional APCs. [21] [54] [61]

Maintenance of immune response

DNA vaccination generates an effective immune memory via the display of antigen-antibody complexes on follicular dendritic cells (FDC), which are potent B-cell stimulators. T-cells can be stimulated by similar, germinal centre dendritic cells. FDC are able to generate an immune memory because antibodies production "overlaps" long-term expression of antigen, allowing antigen-antibody immunocomplexes to form and be displayed by FDC. [7]

Interferons

Both helper and cytotoxic T-cells can control viral infections by secreting interferons. Cytotoxic T cells usually kill virally infected cells. However, they can also be stimulated to secrete antiviral cytokines such as IFN-γ and TNF-α, which do not kill the cell, but limit viral infection by down-regulating the expression of viral components. [62] DNA vaccinations can be used to curb viral infections by non-destructive IFN-mediated control. This was demonstrated for hepatitis B. [63] IFN-γ is critically important in controlling malaria infections [64] and is a consideration for anti-malarial DNA vaccines.

Immune response modulation

Cytokine modulation

An effective vaccine must induce an appropriate immune response for a given pathogen. DNA vaccines can polarise T-cell help towards TH1 or TH2 profiles and generate CTL and/or antibody when required. This can be accomplished by modifications to the form of antigen expressed (i.e. intracellular vs. secreted), the method and route of delivery or the dose. [41] [42] [65] [66] [67] It can also be accomplished by the co-administration of plasmid DNA encoding immune regulatory molecules, i.e. cytokines, lymphokines or co-stimulatory molecules. These "genetic adjuvants" can be administered as a:

In general, co-administration of pro-inflammatory agents (such as various interleukins, tumor necrosis factor, and GM-CSF) plus TH2-inducing cytokines increase antibody responses, whereas pro-inflammatory agents and TH1-inducing cytokines decrease humoral responses and increase cytotoxic responses (more important in viral protection). Co-stimulatory molecules such as B7-1, B7-2 and CD40L are sometimes used.

This concept was applied in topical administration of pDNA encoding IL-10. [33] Plasmid encoding B7-1 (a ligand on APCs) successfully enhanced the immune response in tumour models. Mixing plasmids encoding GM-CSF and the circumsporozoite protein of P. yoelii (PyCSP) enhanced protection against subsequent challenge (whereas plasmid-encoded PyCSP alone did not). It was proposed that GM-CSF caused dendritic cells to present antigen more efficiently and enhance IL-2 production and TH cell activation, thus driving the increased immune response. [50] This can be further enhanced by first priming with a pPyCSP and pGM-CSF mixture, followed by boosting with a recombinant poxvirus expressing PyCSP. [68] However, co-injection of plasmids encoding GM-CSF (or IFN-γ, or IL-2) and a fusion protein of P. chabaudi merozoite surface protein 1 (C-terminus)-hepatitis B virus surface protein (PcMSP1-HBs) abolished protection against challenge, compared to protection acquired by delivery of pPcMSP1-HBs alone. [30]

The advantages of genetic adjuvants are their low cost and simple administration, as well as avoidance of unstable recombinant cytokines and potentially toxic, "conventional" adjuvants (such as alum, calcium phosphate, monophosphoryl lipid A, cholera toxin, cationic and mannan-coated liposomes, QS21, carboxymethyl cellulose and ubenimex). [7] [21] However, the potential toxicity of prolonged cytokine expression is not established. In many commercially important animal species, cytokine genes have not been identified and isolated. In addition, various plasmid-encoded cytokines modulate the immune system differently according to the delivery time. For example, some cytokine plasmid DNAs are best delivered after immunogen pDNA, because pre- or co-delivery can decrease specific responses and increase non-specific responses. [69]

Immunostimulatory CpG motifs

Plasmid DNA itself appears to have an adjuvant effect on the immune system. [6] [7] Bacterially derived DNA can trigger innate immune defence mechanisms, the activation of dendritic cells and the production of TH1 cytokines. [45] [70] This is due to recognition of certain CpG dinucleotide sequences that are immunostimulatory. [66] [71] CpG stimulatory (CpG-S) sequences occur twenty times more frequently in bacterially-derived DNA than in eukaryotes. This is because eukaryotes exhibit "CpG suppression" – i.e. CpG dinucleotide pairs occur much less frequently than expected. Additionally, CpG-S sequences are hypomethylated. This occurs frequently in bacterial DNA, while CpG motifs occurring in eukaryotes are methylated at the cytosine nucleotide. In contrast, nucleotide sequences that inhibit the activation of an immune response (termed CpG neutralising, or CpG-N) are over represented in eukaryotic genomes. [72] The optimal immunostimulatory sequence is an unmethylated CpG dinucleotide flanked by two 5’ purines and two 3’ pyrimidines. [66] [70] Additionally, flanking regions outside this immunostimulatory hexamer must be guanine-rich to ensure binding and uptake into target cells.

The innate system works with the adaptive immune system to mount a response against the DNA encoded protein. CpG-S sequences induce polyclonal B-cell activation and the upregulation of cytokine expression and secretion. [73] Stimulated macrophages secrete IL-12, IL-18, TNF-α, IFN-α, IFN-β and IFN-γ, while stimulated B-cells secrete IL-6 and some IL-12. [21] [73] [74]

Manipulation of CpG-S and CpG-N sequences in the plasmid backbone of DNA vaccines can ensure the success of the immune response to the encoded antigen and drive the immune response toward a TH1 phenotype. This is useful if a pathogen requires a TH response for protection. CpG-S sequences have also been used as external adjuvants for both DNA and recombinant protein vaccination with variable success rates. Other organisms with hypomethylated CpG motifs have demonstrated the stimulation of polyclonal B-cell expansion. [75] The mechanism behind this may be more complicated than simple methylation – hypomethylated murine DNA has not been found to mount an immune response.

Most of the evidence for immunostimulatory CpG sequences comes from murine studies. Extrapolation of this data to other species requires caution – individual species may require different flanking sequences, as binding specificities of scavenger receptors vary across species. Additionally, species such as ruminants may be insensitive to immunostimulatory sequences due to their large gastrointestinal load.

Alternative boosts

DNA-primed immune responses can be boosted by the administration of recombinant protein or recombinant poxviruses. "Prime-boost" strategies with recombinant protein have successfully increased both neutralising antibody titre, and antibody avidity and persistence, for weak immunogens, such as HIV-1 envelope protein. [7] [76] Recombinant virus boosts have been shown to be very efficient at boosting DNA-primed CTL responses. Priming with DNA focuses the immune response on the required immunogen, while boosting with the recombinant virus provides a larger amount of expressed antigen, leading to a large increase in specific CTL responses.

Prime-boost strategies have been successful in inducing protection against malarial challenge in a number of studies. Primed mice with plasmid DNA encoding Plasmodium yoelii circumsporozoite surface protein (PyCSP), then boosted with a recombinant vaccinia virus expressing the same protein had significantly higher levels of antibody, CTL activity and IFN-γ, and hence higher levels of protection, than mice immunized and boosted with plasmid DNA alone. [77] This can be further enhanced by priming with a mixture of plasmids encoding PyCSP and murine GM-CSF, before boosting with recombinant vaccinia virus. [68] An effective prime-boost strategy for the simian malarial model P. knowlesi has also been demonstrated. [78] Rhesus monkeys were primed with a multicomponent, multistage DNA vaccine encoding two liver-stage antigens – the circumsporozoite surface protein (PkCSP) and sporozoite surface protein 2 (PkSSP2) – and two blood stage antigens – the apical merozoite surface protein 1 (PkAMA1) and merozoite surface protein 1 (PkMSP1p42). They were then boosted with a recombinant canarypox virus encoding all four antigens (ALVAC-4). Immunized monkeys developed antibodies against sporozoites and infected erythrocytes, and IFN-γ-secreting T-cell responses against peptides from PkCSP. Partial protection against sporozoite challenge was achieved, and mean parasitemia was significantly reduced, compared to control monkeys. These models, while not ideal for extrapolation to P. falciparum in humans, will be important in pre-clinical trials.

Enhancing immune responses

DNA

The efficiency of DNA immunization can be improved by stabilising DNA against degradation, and increasing the efficiency of delivery of DNA into antigen-presenting cells. [7] This has been demonstrated by coating biodegradable cationic microparticles (such as poly(lactide-co-glycolide) formulated with cetyltrimethylammonium bromide) with DNA. Such DNA-coated microparticles can be as effective at raising CTL as recombinant viruses, especially when mixed with alum. Particles 300 nm in diameter appear to be most efficient for uptake by antigen presenting cells. [7]

Alphavirus vectors

Recombinant alphavirus-based vectors have been used to improve DNA vaccination efficiency. [7] The gene encoding the antigen of interest is inserted into the alphavirus replicon, replacing structural genes but leaving non-structural replicase genes intact. The Sindbis virus and Semliki Forest virus have been used to build recombinant alphavirus replicons. Unlike conventional DNA vaccinations alphavirus vectors kill transfected cells and are only transiently expressed. Alphavirus replicase genes are expressed in addition to the vaccine insert. It is not clear how alphavirus replicons raise an immune response, but it may be due to the high levels of protein expressed by this vector, replicon-induced cytokine responses, or replicon-induced apoptosis leading to enhanced antigen uptake by dendritic cells.

See also

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The adaptive immune system, also known as the acquired immune system, or specific immune system is a subsystem of the immune system that is composed of specialized, systemic cells and processes that eliminate pathogens or prevent their growth. The acquired immune system is one of the two main immunity strategies found in vertebrates.

<span class="mw-page-title-main">Original antigenic sin</span> Immune phenomenon

Original antigenic sin, also known as antigenic imprinting, the Hoskins effect, immunological imprinting, or primary addiction is the propensity of the immune system to preferentially use immunological memory based on a previous infection when a second slightly different version of that foreign pathogen is encountered. This leaves the immune system "trapped" by the first response it has made to each antigen, and unable to mount potentially more effective responses during subsequent infections. Antibodies or T-cells induced during infections with the first variant of the pathogen are subject to repertoire freeze, a form of original antigenic sin.

<span class="mw-page-title-main">CD154</span> Protein-coding gene in humans

CD154, also called CD40 ligand or CD40L, is a protein that is primarily expressed on activated T cells and is a member of the TNF superfamily of molecules. It binds to CD40 on antigen-presenting cells (APC), which leads to many effects depending on the target cell type. In total CD40L has three binding partners: CD40, α5β1 integrin and integrin αIIbβ3. CD154 acts as a costimulatory molecule and is particularly important on a subset of T cells called T follicular helper cells. On TFH cells, CD154 promotes B cell maturation and function by engaging CD40 on the B cell surface and therefore facilitating cell-cell communication. A defect in this gene results in an inability to undergo immunoglobulin class switching and is associated with hyper IgM syndrome. Absence of CD154 also stops the formation of germinal centers and therefore prohibiting antibody affinity maturation, an important process in the adaptive immune system.

Antigenic variation or antigenic alteration refers to the mechanism by which an infectious agent such as a protozoan, bacterium or virus alters the proteins or carbohydrates on its surface and thus avoids a host immune response, making it one of the mechanisms of antigenic escape. It is related to phase variation. Antigenic variation not only enables the pathogen to avoid the immune response in its current host, but also allows re-infection of previously infected hosts. Immunity to re-infection is based on recognition of the antigens carried by the pathogen, which are "remembered" by the acquired immune response. If the pathogen's dominant antigen can be altered, the pathogen can then evade the host's acquired immune system. Antigenic variation can occur by altering a variety of surface molecules including proteins and carbohydrates. Antigenic variation can result from gene conversion, site-specific DNA inversions, hypermutation, or recombination of sequence cassettes. The result is that even a clonal population of pathogens expresses a heterogeneous phenotype. Many of the proteins known to show antigenic or phase variation are related to virulence.

<i>Murine respirovirus</i> Sendai virus, virus of rodents

Murine respirovirus, formerly Sendai virus (SeV) and previously also known as murine parainfluenza virus type 1 or hemagglutinating virus of Japan (HVJ), is an enveloped, 150-200 nm–diameter, negative sense, single-stranded RNA virus of the family Paramyxoviridae. It typically infects rodents and it is not pathogenic for humans or domestic animals.

<span class="mw-page-title-main">Interleukin 15</span> Cytokine with structural similarity to Interleukin-2

Interleukin-15 (IL-15) is a protein that in humans is encoded by the IL15 gene. IL-15 is an inflammatory cytokine with structural similarity to Interleukin-2 (IL-2). Like IL-2, IL-15 binds to and signals through a complex composed of IL-2/IL-15 receptor beta chain (CD122) and the common gamma chain. IL-15 is secreted by mononuclear phagocytes following infection by virus(es). This cytokine induces the proliferation of natural killer cells, i.e. cells of the innate immune system whose principal role is to kill virally infected cells.

<span class="mw-page-title-main">Interleukin 21</span> Mammalian protein found in humans

Interleukin 21 (IL-21) is a protein that in humans is encoded by the IL21 gene.

<span class="mw-page-title-main">CD86</span> Mammalian protein found in Homo sapiens

Cluster of Differentiation 86 is a protein constitutively expressed on dendritic cells, Langerhans cells, macrophages, B-cells, and on other antigen-presenting cells. Along with CD80, CD86 provides costimulatory signals necessary for T cell activation and survival. Depending on the ligand bound, CD86 can signal for self-regulation and cell-cell association, or for attenuation of regulation and cell-cell disassociation.

<span class="mw-page-title-main">ICOSLG</span> Protein-coding gene in the species Homo sapiens

ICOS ligand is a protein that in humans is encoded by the ICOSLG gene located at chromosome 21. ICOSLG has also been designated as CD275.

<span class="mw-page-title-main">PRDM1</span> Protein-coding gene in the species Homo sapiens

PR domain zinc finger protein 1, or B lymphocyte-induced maturation protein-1 (BLIMP-1), is a protein in humans encoded by the gene PRDM1 located on chromosome 6q21. BLIMP-1 is considered a 'master regulator' of hematopoietic stem cells, and plays a critical role in the development of plasma B cells, T cells, dendritic cells (DCs), macrophages, and osteoclasts. Pattern Recognition Receptors (PRRs) can activate BLIMP-1, both as a direct target and through downstream activation. BLIMP-1 is a transcription factor that triggers expression of many downstream signaling cascades. As a fine-tuned and contextual rheostat of the immune system, BLIMP-1 up- or down-regulates immune responses depending on the precise scenarios. BLIMP-1 is highly expressed in exhausted T-cells – clones of dysfunctional T-cells with diminished functions due to chronic immune response against cancer, viral infections, or organ transplant.

<span class="mw-page-title-main">Lymphocyte-activation gene 3</span>

Lymphocyte-activation gene 3, also known as LAG-3, is a protein which in humans is encoded by the LAG3 gene. LAG3, which was discovered in 1990 and was designated CD223 after the Seventh Human Leucocyte Differentiation Antigen Workshop in 2000, is a cell surface molecule with diverse biological effects on T cell function but overall has an immune inhibitory effect. It is an immune checkpoint receptor and as such is the target of various drug development programs by pharmaceutical companies seeking to develop new treatments for cancer and autoimmune disorders. In soluble form it is also being developed as a cancer drug in its own right.

<span class="mw-page-title-main">Follicular B helper T cells</span>

Follicular helper T cells (also known as T follicular helper cells and abbreviated as TFH), are antigen-experienced CD4+ T cells found in the periphery within B cell follicles of secondary lymphoid organs such as lymph nodes, spleen and Peyer's patches, and are identified by their constitutive expression of the B cell follicle homing receptor CXCR5. Upon cellular interaction and cross-signaling with their cognate follicular (Fo B) B cells, TFH cells trigger the formation and maintenance of germinal centers through the expression of CD40 ligand (CD40L) and the secretion of IL-21 and IL-4. TFH cells also migrate from T cell zones into these seeded germinal centers, predominantly composed of rapidly dividing B cells mutating their Ig genes. Within germinal centers, TFH cells play a critical role in mediating the selection and survival of B cells that go on to differentiate either into long-lived plasma cells capable of producing high affinity antibodies against foreign antigen, or germinal center-dependent memory B cells capable of quick immune re-activation in the future if ever the same antigen is re-encountered. TFH cells are also thought to facilitate negative selection of potentially autoimmune-causing mutated B cells in the germinal center. However, the biomechanisms by which TFH cells mediate germinal center tolerance are yet to be fully understood.

A subunit vaccine is a vaccine that contains purified parts of the pathogen that are antigenic, or necessary to elicit a protective immune response. Subunit vaccine can be made from dissembled viral particles in cell culture or recombinant DNA expression, in which case it is a recombinant subunit vaccine.

Tolerogenic therapy aims to induce immune tolerance where there is pathological or undesirable activation of the normal immune response. This can occur, for example, when an allogeneic transplantation patient develops an immune reaction to donor antigens, or when the body responds inappropriately to self antigens implicated in autoimmune diseases. It must provide absence of specific antibodies for exactly that antigenes.

Intrastructural help (ISH) is where T and B cells cooperate to help or suppress an immune response gene. ISH has proven effective for the treatment of influenza, rabies related lyssavirus, hepatitis B, and the HIV virus. This process was used in 1979 to observe that T cells specific to the influenza virus could promote the stimulation of hemagglutinin specific B cells and elicit an effective humoral immune response. It was later applied to the lyssavirus and was shown to protect raccoons from lethal challenge. The ISH principle is especially beneficial because relatively invariable structural antigens can be used for the priming of T-cells to induce humoral immune response against variable surface antigens. Thus, the approach has also transferred well for the treatment of hepatitis B and HIV.

<span class="mw-page-title-main">Viral vector vaccine</span> Type of vaccine

A viral vector vaccine is a vaccine that uses a viral vector to deliver genetic material (DNA) that can be transcribed by the recipient's host cells as mRNA coding for a desired protein, or antigen, to elicit an immune response. As of April 2021, six viral vector vaccines, four COVID-19 vaccines and two Ebola vaccines, have been authorized for use in humans.

A genetic vaccine is a vaccine that contains nucleic acids such as DNA or RNA that lead to protein biosynthesis of antigens within a cell. Genetic vaccines thus include DNA vaccines, RNA vaccines and viral vector vaccines.

Bacteriophage AP205 is a plaque-forming bacteriophage that infects Acinetobacter bacteria. Bacteriophage AP205 is a protein-coated virus with a positive single-stranded RNA genome. It is a member of the family Fiersviridae, consisting of particles that infect Gram-negative bacteria such as E. coli.

References

  1. 1 2 Henrique Roman Ramos and Paulo Lee Ho. "Developing Snake Antivenom Sera by Genetic Immunization: A Review". Clinical Toxinology in Asia Pacific and Africa. 2: 401–414.
  2. 1 2 Liu S, Wang S, Lu S (April 2016). "DNA immunization as a technology platform for monoclonal antibody induction". Emerging Microbes & Infections. 5 (4): e33. doi:10.1038/emi.2016.27. PMC   4855071 . PMID   27048742.
  3. 1 2 3 "DNA vaccines". World Health Organization.
  4. 1 2 Khan KH (March 2013). "DNA vaccines: roles against diseases". Germs. 3 (1): 26–35. doi:10.11599/germs.2013.1034. PMC   3882840 . PMID   24432284.
  5. 1 2 "India gives emergency approval for world's first COVID-19 DNA vaccine". Reuters. 2021-08-20. Retrieved 2021-08-22.
  6. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 Alarcon JB, Waine GW, McManus DP (1999). "DNA Vaccines: Technology and Application as Anti-parasite and Anti-microbial Agents". Advances in Parasitology Volume 42. Vol. 42. pp. 343–410. doi:10.1016/S0065-308X(08)60152-9. ISBN   9780120317424. PMID   10050276.
  7. 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 Robinson HL, Pertmer TM (2000). DNA vaccines for viral infections: basic studies and applications. Advances in Virus Research. Vol. 55. pp. 1–74. doi:10.1016/S0065-3527(00)55001-5. ISBN   9780120398553. PMID   11050940.
  8. ↑ White LO, Gibb E, Newham HC, Richardson MD, Warren RC (July 1979). "Comparison of the growth of virulent and attenuated strains of Candida albicans in the kidneys of normal and cortison-treated mice by chitin assay". Mycopathologia. 67 (3): 173–177. doi:10.1007/bf00470753. PMID   384256. S2CID   31914107.
  9. ↑ Paoletti E, Lipinskas BR, Samsonoff C, Mercer S, Panicali D (January 1984). "Construction of live vaccines using genetically engineered poxviruses: biological activity of vaccinia virus recombinants expressing the hepatitis B virus surface antigen and the herpes simplex virus glycoprotein D". Proceedings of the National Academy of Sciences of the United States of America. 81 (1): 193–197. Bibcode:1984PNAS...81..193P. doi: 10.1073/pnas.81.1.193 . PMC   344637 . PMID   6320164.
  10. ↑ US Patent 4722848 - Method for immunizing animals with synthetically modified vaccinia virus
  11. ↑ Ulmer JB, Donnelly JJ, Parker SE, Rhodes GH, Felgner PL, Dwarki VJ, et al. (March 1993). "Heterologous protection against influenza by injection of DNA encoding a viral protein". Science. 259 (5102): 1745–1749. Bibcode:1993Sci...259.1745U. doi:10.1126/science.8456302. PMID   8456302.
  12. ↑ Regalado A (2 August 2016). "The U.S. government has begun testing its first Zika vaccine in humans". MIT Technology Review Magazine. Retrieved 2016-08-06.
  13. ↑ Chen Y, Wang S, Lu S (February 2014). "DNA Immunization for HIV Vaccine Development". Vaccines. 2 (1): 138–159. doi: 10.3390/vaccines2010138 . PMC   4494200 . PMID   26344472.
  14. ↑ Ledgerwood JE, Pierson TC, Hubka SA, Desai N, Rucker S, Gordon IJ, et al. (May 2011). "A West Nile virus DNA vaccine utilizing a modified promoter induces neutralizing antibody in younger and older healthy adults in a phase I clinical trial". The Journal of Infectious Diseases. 203 (10): 1396–1404. doi:10.1093/infdis/jir054. PMC   3080891 . PMID   21398392.
  15. ↑ Kilpatrick AM, Dupuis AP, Chang GJ, Kramer LD (May 2010). "DNA vaccination of American robins (Turdus migratorius) against West Nile virus". Vector Borne and Zoonotic Diseases. 10 (4): 377–380. doi:10.1089/vbz.2009.0029. PMC   2883478 . PMID   19874192.
  16. 1 2 Sedegah M, Hedstrom R, Hobart P, Hoffman SL (October 1994). "Protection against malaria by immunization with plasmid DNA encoding circumsporozoite protein". Proceedings of the National Academy of Sciences of the United States of America. 91 (21): 9866–9870. Bibcode:1994PNAS...91.9866S. doi: 10.1073/pnas.91.21.9866 . JSTOR   2365723. PMC   44918 . PMID   7937907.
  17. ↑ Harmon BT, Aly AE, Padegimas L, Sesenoglu-Laird O, Cooper MJ, Waszczak BL (May 2014). "Intranasal administration of plasmid DNA nanoparticles yields successful transfection and expression of a reporter protein in rat brain". Gene Therapy. 21 (5): 514–521. doi: 10.1038/gt.2014.28 . PMID   24670994. S2CID   5560134.
  18. 1 2 Mor G, Klinman DM, Shapiro S, Hagiwara E, Sedegah M, Norman JA, et al. (August 1995). "Complexity of the cytokine and antibody response elicited by immunizing mice with Plasmodium yoelii circumsporozoite protein plasmid DNA". Journal of Immunology. 155 (4): 2039–2046. doi: 10.4049/jimmunol.155.4.2039 . PMID   7636255. S2CID   37290980.
  19. ↑ Leitner WW, Seguin MC, Ballou WR, Seitz JP, Schultz AM, Sheehy MJ, Lyon JA (December 1997). "Immune responses induced by intramuscular or gene gun injection of protective deoxyribonucleic acid vaccines that express the circumsporozoite protein from Plasmodium berghei malaria parasites". Journal of Immunology. 159 (12): 6112–6119. doi: 10.4049/jimmunol.159.12.6112 . PMID   9550412. S2CID   37685499.
  20. ↑ Böhm W, Kuhröber A, Paier T, Mertens T, Reimann J, Schirmbeck R (June 1996). "DNA vector constructs that prime hepatitis B surface antigen-specific cytotoxic T lymphocyte and antibody responses in mice after intramuscular injection". Journal of Immunological Methods. 193 (1): 29–40. doi:10.1016/0022-1759(96)00035-X. PMID   8690928.
  21. 1 2 3 4 5 6 7 8 9 10 11 Lewis PJ, Babiuk LA (1999). DNA vaccines: a review. Vol. 54. Academic Press. pp. 129–88. doi:10.1016/S0065-3527(08)60367-X. ISBN   978-0-12-039854-6. PMID   10547676.{{cite book}}: |journal= ignored (help)
  22. ↑ André S, Seed B, Eberle J, Schraut W, Bültmann A, Haas J (February 1998). "Increased immune response elicited by DNA vaccination with a synthetic gp120 sequence with optimized codon usage". Journal of Virology. 72 (2): 1497–1503. doi:10.1128/JVI.72.2.1497-1503.1998. PMC   124631 . PMID   9445053.
  23. ↑ Muthumani K, Zhang D, Dayes NS, Hwang DS, Calarota SA, Choo AY, et al. (September 2003). "Novel engineered HIV-1 East African Clade-A gp160 plasmid construct induces strong humoral and cell-mediated immune responses in vivo". Virology. 314 (1): 134–146. doi: 10.1016/S0042-6822(03)00459-8 . PMID   14517067.
  24. ↑ Oliveira PH, Prather KJ, Prazeres DM, Monteiro GA (September 2009). "Structural instability of plasmid biopharmaceuticals: challenges and implications". Trends in Biotechnology. 27 (9): 503–511. doi:10.1016/j.tibtech.2009.06.004. PMID   19656584.
  25. ↑ Oliveira PH, Mairhofer J (September 2013). "Marker-free plasmids for biotechnological applications - implications and perspectives". Trends in Biotechnology. 31 (9): 539–547. doi:10.1016/j.tibtech.2013.06.001. PMID   23830144.
  26. ↑ Kutzler MA, Weiner DB (October 2008). "DNA vaccines: ready for prime time?". Nature Reviews. Genetics. 9 (10): 776–788. doi:10.1038/nrg2432. PMC   4317294 . PMID   18781156.
  27. ↑ Rodriguez F, Zhang J, Whitton JL (November 1997). "DNA immunization: ubiquitination of a viral protein enhances cytotoxic T-lymphocyte induction and antiviral protection but abrogates antibody induction". Journal of Virology. 71 (11): 8497–8503. doi:10.1128/JVI.71.11.8497-8503.1997. PMC   192313 . PMID   9343207.
  28. 1 2 Tobery TW, Siliciano RF (March 1997). "Targeting of HIV-1 antigens for rapid intracellular degradation enhances cytotoxic T lymphocyte (CTL) recognition and the induction of de novo CTL responses in vivo after immunization". The Journal of Experimental Medicine. 185 (5): 909–920. doi:10.1084/jem.185.5.909. PMC   2196169 . PMID   9120397.
  29. ↑ Huebener N, Fest S, Strandsby A, Michalsky E, Preissner R, Zeng Y, et al. (July 2008). "A rationally designed tyrosine hydroxylase DNA vaccine induces specific antineuroblastoma immunity". Molecular Cancer Therapeutics. 7 (7): 2241–2251. doi:10.1158/1535-7163.MCT-08-0109. PMID   18645033. S2CID   35652424.
  30. 1 2 Wunderlich G, Moura IC, del Portillo HA (October 2000). "Genetic immunization of BALB/c mice with a plasmid bearing the gene coding for a hybrid merozoite surface protein 1-hepatitis B virus surface protein fusion protects mice against lethal Plasmodium chabaudi chabaudi PC1 infection". Infection and Immunity. 68 (10): 5839–5845. doi:10.1128/IAI.68.10.5839-5845.2000. PMC   101545 . PMID   10992493.
  31. ↑ Weiner DB, Kennedy RC (July 1999). "Genetic vaccines". Scientific American. 281 (1): 50–57. Bibcode:1999SciAm.281a..50W. doi:10.1038/scientificamerican0799-50. PMID   10396782. Archived from the original on 2009-03-25. Retrieved 2007-11-21.
  32. ↑ Widera G, Austin M, Rabussay D, Goldbeck C, Barnett SW, Chen M, et al. (May 2000). "Increased DNA vaccine delivery and immunogenicity by electroporation in vivo". Journal of Immunology. 164 (9): 4635–4640. doi: 10.4049/jimmunol.164.9.4635 . PMID   10779767.
  33. 1 2 Daheshia M, Kuklin N, Kanangat S, Manickan E, Rouse BT (August 1997). "Suppression of ongoing ocular inflammatory disease by topical administration of plasmid DNA encoding IL-10". Journal of Immunology. 159 (4): 1945–1952. doi: 10.4049/jimmunol.159.4.1945 . PMID   9257860. S2CID   43203331.
  34. 1 2 Chen Y, Webster RG, Woodland DL (March 1998). "Induction of CD8+ T cell responses to dominant and subdominant epitopes and protective immunity to Sendai virus infection by DNA vaccination". Journal of Immunology. 160 (5): 2425–2432. doi: 10.4049/jimmunol.160.5.2425 . PMID   9498786. S2CID   2250871.
  35. ↑ Lode HN, Huebener N, Zeng Y, Fest S, Weixler S, Gaedicke G (December 2004). "DNA minigene vaccination for adjuvant neuroblastoma therapy". Annals of the New York Academy of Sciences. 1028 (1): 113–121. Bibcode:2004NYASA1028..113L. doi:10.1196/annals.1322.012. PMID   15650237. S2CID   27240738.
  36. ↑ Sizemore DR, Branstrom AA, Sadoff JC (October 1995). "Attenuated Shigella as a DNA delivery vehicle for DNA-mediated immunization". Science. 270 (5234): 299–302. Bibcode:1995Sci...270..299S. doi:10.1126/science.270.5234.299. PMID   7569980. S2CID   12532901.
  37. ↑ Nealon, Cory (25 November 2014). "A hybrid vehicle that delivers DNA". The State University of New York at Buffalo. Retrieved 16 December 2014.
  38. ↑ Jones CH, Ravikrishnan A, Chen M, Reddinger R, Kamal Ahmadi M, Rane S, et al. (August 2014). "Hybrid biosynthetic gene therapy vector development and dual engineering capacity". Proceedings of the National Academy of Sciences of the United States of America. 111 (34): 12360–12365. Bibcode:2014PNAS..11112360J. doi: 10.1073/pnas.1411355111 . PMC   4151754 . PMID   25114239.
  39. ↑ Barry MA, Lai WC, Johnston SA (October 1995). "Protection against mycoplasma infection using expression-library immunization". Nature. 377 (6550): 632–635. Bibcode:1995Natur.377..632B. doi: 10.1038/377632a0 . PMID   7566175. S2CID   4306972.
  40. ↑ Fynan EF, Webster RG, Fuller DH, Haynes JR, Santoro JC, Robinson HL (December 1993). "DNA vaccines: protective immunizations by parenteral, mucosal, and gene-gun inoculations". Proceedings of the National Academy of Sciences of the United States of America. 90 (24): 11478–11482. Bibcode:1993PNAS...9011478F. doi: 10.1073/pnas.90.24.11478 . PMC   48007 . PMID   8265577.
  41. 1 2 3 4 5 Feltquate DM, Heaney S, Webster RG, Robinson HL (March 1997). "Different T helper cell types and antibody isotypes generated by saline and gene gun DNA immunization". Journal of Immunology. 158 (5): 2278–2284. doi: 10.4049/jimmunol.158.5.2278 . PMID   9036975. S2CID   41368723.
  42. 1 2 3 4 Boyle CM, Morin M, Webster RG, Robinson HL (December 1996). "Role of different lymphoid tissues in the initiation and maintenance of DNA-raised antibody responses to the influenza virus H1 glycoprotein". Journal of Virology. 70 (12): 9074–9078. doi:10.1128/JVI.70.12.9074-9078.1996. PMC   191015 . PMID   8971047.
  43. ↑ Sällberg M, Townsend K, Chen M, O'Dea J, Banks T, Jolly DJ, et al. (July 1997). "Characterization of humoral and CD4+ cellular responses after genetic immunization with retroviral vectors expressing different forms of the hepatitis B virus core and e antigens". Journal of Virology. 71 (7): 5295–5303. doi:10.1128/JVI.71.7.5295-5303.1997. PMC   191766 . PMID   9188598.
  44. ↑ Banchereau J, Steinman RM (March 1998). "Dendritic cells and the control of immunity". Nature. 392 (6673): 245–252. Bibcode:1998Natur.392..245B. doi:10.1038/32588. PMID   9521319. S2CID   4388748.
  45. 1 2 Jakob T, Walker PS, Krieg AM, Udey MC, Vogel JC (September 1998). "Activation of cutaneous dendritic cells by CpG-containing oligodeoxynucleotides: a role for dendritic cells in the augmentation of Th1 responses by immunostimulatory DNA". Journal of Immunology. 161 (6): 3042–3049. doi: 10.4049/jimmunol.161.6.3042 . PMID   9743369. S2CID   35107733.
  46. 1 2 Raz E, Tighe H, Sato Y, Corr M, Dudler JA, Roman M, et al. (May 1996). "Preferential induction of a Th1 immune response and inhibition of specific IgE antibody formation by plasmid DNA immunization". Proceedings of the National Academy of Sciences of the United States of America. 93 (10): 5141–5145. Bibcode:1996PNAS...93.5141R. doi: 10.1073/pnas.93.10.5141 . PMC   39421 . PMID   8643542.
  47. 1 2 Fu TM, Friedman A, Ulmer JB, Liu MA, Donnelly JJ (April 1997). "Protective cellular immunity: cytotoxic T-lymphocyte responses against dominant and recessive epitopes of influenza virus nucleoprotein induced by DNA immunization". Journal of Virology. 71 (4): 2715–2721. doi:10.1128/JVI.71.4.2715-2721.1997. PMC   191393 . PMID   9060624.
  48. 1 2 3 Restifo NP, Bacík I, Irvine KR, Yewdell JW, McCabe BJ, Anderson RW, et al. (May 1995). "Antigen processing in vivo and the elicitation of primary CTL responses". Journal of Immunology. 154 (9): 4414–4422. doi:10.4049/jimmunol.154.9.4414. PMC   1952186 . PMID   7722298.
  49. 1 2 3 Iwasaki A, Stiernholm BJ, Chan AK, Berinstein NL, Barber BH (May 1997). "Enhanced CTL responses mediated by plasmid DNA immunogens encoding costimulatory molecules and cytokines". Journal of Immunology. 158 (10): 4591–4601. doi:10.4049/jimmunol.158.10.4591. PMID   9144471. S2CID   41779568.
  50. 1 2 Weiss WR, Ishii KJ, Hedstrom RC, Sedegah M, Ichino M, Barnhart K, et al. (September 1998). "A plasmid encoding murine granulocyte-macrophage colony-stimulating factor increases protection conferred by a malaria DNA vaccine". Journal of Immunology. 161 (5): 2325–2332. doi: 10.4049/jimmunol.161.5.2325 . PMID   9725227. S2CID   21038927.
  51. ↑ Tsuji T, Hamajima K, Fukushima J, Xin KQ, Ishii N, Aoki I, et al. (April 1997). "Enhancement of cell-mediated immunity against HIV-1 induced by coinnoculation of plasmid-encoded HIV-1 antigen with plasmid expressing IL-12". Journal of Immunology. 158 (8): 4008–4013. doi: 10.4049/jimmunol.158.8.4008 . PMID   9103472. S2CID   38099483.
  52. 1 2 Justewicz DM, Webster RG (October 1996). "Long-term maintenance of B cell immunity to influenza virus hemagglutinin in mice following DNA-based immunization". Virology. 224 (1): 10–17. doi: 10.1006/viro.1996.0501 . PMID   8862394.
  53. ↑ Mancini-Bourgine M, Fontaine H, Bréchot C, Pol S, Michel ML (May 2006). "Immunogenicity of a hepatitis B DNA vaccine administered to chronic HBV carriers". Vaccine. 24 (21): 4482–4489. doi:10.1016/j.vaccine.2005.08.013. PMID   16310901.
  54. 1 2 Wolff JA, Dowty ME, Jiao S, Repetto G, Berg RK, Ludtke JJ, et al. (December 1992). "Expression of naked plasmids by cultured myotubes and entry of plasmids into T tubules and caveolae of mammalian skeletal muscle". Journal of Cell Science. 103. 103 (4): 1249–1259. doi:10.1242/jcs.103.4.1249. PMID   1487500.
  55. ↑ Anderson RG, Kamen BA, Rothberg KG, Lacey SW (January 1992). "Potocytosis: sequestration and transport of small molecules by caveolae". Science. 255 (5043): 410–411. Bibcode:1992Sci...255..410A. doi:10.1126/science.1310359. PMID   1310359.
  56. 1 2 Casares S, Inaba K, Brumeanu TD, Steinman RM, Bona CA (November 1997). "Antigen presentation by dendritic cells after immunization with DNA encoding a major histocompatibility complex class II-restricted viral epitope". The Journal of Experimental Medicine. 186 (9): 1481–1486. doi:10.1084/jem.186.9.1481. PMC   2199124 . PMID   9348305.
  57. 1 2 Bennett RM, Gabor GT, Merritt MM (December 1985). "DNA binding to human leukocytes. Evidence for a receptor-mediated association, internalization, and degradation of DNA". The Journal of Clinical Investigation. 76 (6): 2182–2190. doi:10.1172/JCI112226. PMC   424340 . PMID   3001145.
  58. ↑ Bennet RM, Hefeneider SH, Bakke A, Merritt M, Smith CA, Mourich D, Heinrich MC (May 1988). "The production and characterization of murine monoclonal antibodies to a DNA receptor on human leukocytes". Journal of Immunology. 140 (9): 2937–2942. doi: 10.4049/jimmunol.140.9.2937 . PMID   2452195. S2CID   22923379.
  59. ↑ Corr M, Lee DJ, Carson DA, Tighe H (October 1996). "Gene vaccination with naked plasmid DNA: mechanism of CTL priming". The Journal of Experimental Medicine. 184 (4): 1555–1560. doi:10.1084/jem.184.4.1555. PMC   2192808 . PMID   8879229.
  60. ↑ Chattergoon MA, Robinson TM, Boyer JD, Weiner DB (June 1998). "Specific immune induction following DNA-based immunization through in vivo transfection and activation of macrophages/antigen-presenting cells". Journal of Immunology. 160 (12): 5707–5718. doi: 10.4049/jimmunol.160.12.5707 . PMID   9637479. S2CID   33499198.
  61. 1 2 Torres CA, Iwasaki A, Barber BH, Robinson HL (May 1997). "Differential dependence on target site tissue for gene gun and intramuscular DNA immunizations". Journal of Immunology. 158 (10): 4529–4532. doi: 10.4049/jimmunol.158.10.4529 . PMID   9144463. S2CID   45069087.
  62. ↑ Franco A, Guidotti LG, Hobbs MV, Pasquetto V, Chisari FV (August 1997). "Pathogenetic effector function of CD4-positive T helper 1 cells in hepatitis B virus transgenic mice". Journal of Immunology. 159 (4): 2001–2008. doi: 10.4049/jimmunol.159.4.2001 . PMID   9257867. S2CID   20528634.
  63. ↑ Mancini M, Hadchouel M, Davis HL, Whalen RG, Tiollais P, Michel ML (October 1996). "DNA-mediated immunization in a transgenic mouse model of the hepatitis B surface antigen chronic carrier state". Proceedings of the National Academy of Sciences of the United States of America. 93 (22): 12496–12501. Bibcode:1996PNAS...9312496M. doi: 10.1073/pnas.93.22.12496 . PMC   38020 . PMID   8901610.
  64. ↑ Doolan DL, Hoffman SL (July 1999). "IL-12 and NK cells are required for antigen-specific adaptive immunity against malaria initiated by CD8+ T cells in the Plasmodium yoelii model". Journal of Immunology. 163 (2): 884–892. doi: 10.4049/jimmunol.163.2.884 . PMID   10395683. S2CID   41651105.
  65. ↑ Cardoso AI, Blixenkrone-Moller M, Fayolle J, Liu M, Buckland R, Wild TF (November 1996). "Immunization with plasmid DNA encoding for the measles virus hemagglutinin and nucleoprotein leads to humoral and cell-mediated immunity". Virology. 225 (2): 293–299. doi: 10.1006/viro.1996.0603 . PMID   8918915.
  66. 1 2 3 Sato Y, Roman M, Tighe H, Lee D, Corr M, Nguyen MD, et al. (July 1996). "Immunostimulatory DNA sequences necessary for effective intradermal gene immunization". Science. 273 (5273): 352–354. Bibcode:1996Sci...273..352S. doi:10.1126/science.273.5273.352. PMID   8662521. S2CID   9333197.
  67. ↑ Weiss R, Leitner WW, Scheiblhofer S, Chen D, Bernhaupt A, Mostböck S, et al. (October 2000). "Genetic vaccination against malaria infection by intradermal and epidermal injections of a plasmid containing the gene encoding the Plasmodium berghei circumsporozoite protein". Infection and Immunity. 68 (10): 5914–5919. doi:10.1128/IAI.68.10.5914-5919.2000. PMC   101554 . PMID   10992502.
  68. 1 2 Sedegah M, Weiss W, Sacci JB, Charoenvit Y, Hedstrom R, Gowda K, et al. (June 2000). "Improving protective immunity induced by DNA-based immunization: priming with antigen and GM-CSF-encoding plasmid DNA and boosting with antigen-expressing recombinant poxvirus". Journal of Immunology. 164 (11): 5905–5912. doi: 10.4049/jimmunol.164.11.5905 . PMID   10820272.
  69. ↑ Barouch DH, Santra S, Steenbeke TD, Zheng XX, Perry HC, Davies ME, et al. (August 1998). "Augmentation and suppression of immune responses to an HIV-1 DNA vaccine by plasmid cytokine/Ig administration". Journal of Immunology. 161 (4): 1875–1882. doi: 10.4049/jimmunol.161.4.1875 . PMID   9712056. S2CID   36488254.
  70. 1 2 Krieg AM, Yi AK, Matson S, Waldschmidt TJ, Bishop GA, Teasdale R, et al. (April 1995). "CpG motifs in bacterial DNA trigger direct B-cell activation". Nature. 374 (6522): 546–549. Bibcode:1995Natur.374..546K. doi:10.1038/374546a0. PMID   7700380. S2CID   4261304.
  71. ↑ Klinman DM, Yamshchikov G, Ishigatsubo Y (April 1997). "Contribution of CpG motifs to the immunogenicity of DNA vaccines". Journal of Immunology. 158 (8): 3635–3639. doi: 10.4049/jimmunol.158.8.3635 . PMID   9103425. S2CID   41861994.
  72. ↑ Krieg AM, Wu T, Weeratna R, Efler SM, Love-Homan L, Yang L, et al. (October 1998). "Sequence motifs in adenoviral DNA block immune activation by stimulatory CpG motifs". Proceedings of the National Academy of Sciences of the United States of America. 95 (21): 12631–12636. Bibcode:1998PNAS...9512631K. doi: 10.1073/pnas.95.21.12631 . PMC   22882 . PMID   9770537.
  73. 1 2 Klinman DM, Yi AK, Beaucage SL, Conover J, Krieg AM (April 1996). "CpG motifs present in bacteria DNA rapidly induce lymphocytes to secrete interleukin 6, interleukin 12, and interferon gamma". Proceedings of the National Academy of Sciences of the United States of America. 93 (7): 2879–2883. Bibcode:1996PNAS...93.2879K. doi: 10.1073/pnas.93.7.2879 . PMC   39727 . PMID   8610135.
  74. ↑ Yi AK, Chace JH, Cowdery JS, Krieg AM (January 1996). "IFN-gamma promotes IL-6 and IgM secretion in response to CpG motifs in bacterial DNA and oligodeoxynucleotides". Journal of Immunology. 156 (2): 558–564. doi: 10.4049/jimmunol.156.2.558 . PMID   8543806. S2CID   42145608.
  75. ↑ Barwick BG, Scharer CD, Martinez RJ, Price MJ, Wein AN, Haines RR, et al. (May 2018). "B cell activation and plasma cell differentiation are inhibited by de novo DNA methylation". Nature Communications. 9 (1): 1900. Bibcode:2018NatCo...9.1900B. doi:10.1038/s41467-018-04234-4. PMC   5953949 . PMID   29765016.
  76. ↑ Letvin NL, Montefiori DC, Yasutomi Y, Perry HC, Davies ME, Lekutis C, et al. (August 1997). "Potent, protective anti-HIV immune responses generated by bimodal HIV envelope DNA plus protein vaccination". Proceedings of the National Academy of Sciences of the United States of America. 94 (17): 9378–9383. Bibcode:1997PNAS...94.9378L. doi: 10.1073/pnas.94.17.9378 . PMC   23198 . PMID   9256490.
  77. ↑ Sedegah M, Jones TR, Kaur M, Hedstrom R, Hobart P, Tine JA, Hoffman SL (June 1998). "Boosting with recombinant vaccinia increases immunogenicity and protective efficacy of malaria DNA vaccine". Proceedings of the National Academy of Sciences of the United States of America. 95 (13): 7648–7653. Bibcode:1998PNAS...95.7648S. doi: 10.1073/pnas.95.13.7648 . PMC   22711 . PMID   9636204.
  78. ↑ Rogers WO, Baird JK, Kumar A, Tine JA, Weiss W, Aguiar JC, et al. (September 2001). "Multistage multiantigen heterologous prime boost vaccine for Plasmodium knowlesi malaria provides partial protection in rhesus macaques". Infection and Immunity. 69 (9): 5565–5572. doi:10.1128/IAI.69.9.5565-5572.2001. PMC   98670 . PMID   11500430.

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