Gene gun dna vaccine pdf

The advantages of gene gun mediated delivery include signi. Immune responses induced by gene gun or intramuscular. The active constituent of a dna vaccine is a plasmid molecule that contains the gene for a component of a pathogenic organism under the control of a mammalian expression system, and possesses dna sequences necessary for replication and selection in bacteria. Use of gene gun for genetic immunotherapy springerlink. Using the gene gun delivery system, we tested several dna vaccines that employ intracellular targeting strategies for enhancing mhc class i and class ii presentation of encoded model antigen hpv. Conclusionthe low humoral immunogenicity of dna vaccine with lhdag cannot be ameliorated by different dosage, gene gun immunization, or in vivo ep intramuscular injection. A prototype gene gun accell was acquired from agracetus. Jan 01, 2009 a gene gun was used to cotransfect cells in vivo by cocoating the 2 plasmids onto the same pool of gold particles.

Dna vaccines genetic dna immunization is a novel technique used to efficiently stimulate humoral and cellular immune responses to protein antigens. Because the gene gun delivers a small amount of dna efficiently, it is a promising approach for dna vaccination. The dnacoated gold particles are driven into the epidermal layers of. Gene gun inoculation requires lower dna doses than i. For the np dna, boosted gene gun immunizations of 0. Gene guns have also been used to deliver dna vaccines. Cur gested that an epitope vaccine composed of a selfb cell rently, the two major delivery methods used for enhancing epitope of a. Annex 1 guidelines for assuring the quality and nonclinical. Dna vaccines were pioneered by several groups in the early 1990s. Gene gunmediated dna immunization primes development of. The direct injection of genetic material into a living host causes a small amount of its cells to produce the introduced gene. The dna vaccine in fullers lab was engineered by using genetic components of influenza virus the conserved areas which do not change.

Structural instability phenomena are of particular concern for plasmid manufacture, dna vaccination and gene therapy. Emphasis will be placed on epidermal powder immunization and particlemediated gene gun dna immunization, which use similar mechanical devices to deliver protein and dna vaccines, respectively, into the viable epidermis. By coating particles of a heavy metal with a gene of interest and firing these microprojectiles into cells using mechanical force, an integration of desired genetic information can be induced into cells. The method of dna immunization with a gene gun, also known as biolistic delivery, is superior to traditional techniques of immunization, in which preformed proteins or protein fragments dissociated from their membrane environment are delivered into the host in order to stimulate antibodies. Enhanced antitumor effect of a gene gun delivered dna vaccine encoding the human papillomavirus type 16 oncoproteins genetically fused to the herpes simplex virus glycoprotein d m. In this study, we evaluated the immunogenicity and the protective efficacy of the sm23 dna vaccine using two different intradermal dna delivery methods. The study on dna vaccines first began in the 1990s, when the plasmid dna is. Methods and protocols, third edition explores innovative approaches and technologies used to design, deliver, and enhance the efficacy of dna vaccines featuring applications which should be of great value in moving vaccines from research to clinic, this detailed volume includes sections on dna vaccine design and enhancement, delivery systems, production, purification, and. Dec 15, 1993 successful mucosal routes of vaccination included dna drops administered to the nares or trachea. We determined that the helper antigen plasmid was largely ineffective when delivered at the same dose as the tumor antigen plasmid but achieved dramatically increased efficacy at a very low. Gene gun delivered dna based immunizations mediate rapid production of murine monoclonal antibodies to the flt3 receptor katherine e. Chickens inoculated via the im route displayed an increase in ndvspecific antibodies and cellmeditated immunity. The gene gun is a unique means of introducing naked dna vaccines directly into cells, bypassing the cell membrane and its receptors.

The distinct th1 or th2biased immune responses elicited by intramuscular injection or gene gun delivery, respectively, are not fully. Several reports on rodents suggest that dna vaccines can be used to achieve mucosal immunity when applied to mucosal tissues. Antigenic protein is presented by cell in which it is produced. Dna vaccine, gene gun, hepatitis d virus, in vivo electroporation hdv. Enhancement of dna vaccine potency by linkage of antigen gene to an hsp70 gene1. In addition, the gene gun allows delivery of several genes simultaneously to the same target tissues, which enables studying the interaction among the gene products. Like recombinant vaccines, genes for the desired antigens are located and cloned. The past, current and future trends in dna vaccine. Dna vaccines consist of transient transfection of naked dna into target cells. Gene gunmediated dna vaccination enhances antigenspecific.

Tungsten or gold beads are coated in the gene of interest and fired through a stopping screen, accelerated by helium, into the plant tissue. Dna immunization using a gene gun to deliver gold beads coated with dna plasmids. In general, it is desirable to employ efficient routes for in vivo delivery of dna vaccines directly into dcs. The dna vaccine conferred protection to 40% of chickens against ndv upon challenge. Genetic vaccination using naked plasmid dna is an immunization strategy both against infectious diseases and cancer. Nevertheless, intramuscular injection is a variable technique.

Gene gun has emerged as a novel tool for delivery of dna vaccines directly into dcs com. The dna vaccine administrated by particle bombardment induced higher. In this study, the potential of inducing mucosal immunity in cattle by immunization with a dna vaccine was demonstrated. Gene gun immunization uses gold particles as a carrier for dna molecules. In order to improve the efficacy of dna vaccines, particularly in large animals. Dna vaccine against west nile virus for use in horses was. Tuberculosis dna vaccine encoding ag85a is immunogenic and.

Ctl response in c57bl6 mice immunized with various recombinant dna vaccines. Intro duc tion immun ity is the d efens e mechan ism tha t work s in the. Two dna vaccine plasmids, pcdna3 sera 17382 and pcdna3 sera 17110 were coated onto gold particles. The direct injection of genetic material into a living host causes a small amount of its cells to produce the introduced gene products. Systemic administration of cpg oligodeoxynucleotide and. Gene gunmediated dna vaccination induces antitumor. Gene gunmediated dna vaccination induces antitumor immunity. Dna vaccine with a lhdag construct may not be a candidate hdv vaccine to generate antihdv humoral immunity. Analysis of humoral immunity of hepatitis d virus dna vaccine.

Through improving our understanding of basic immunology as it is related to dna vaccine technology, our. The concept of using dna immunization and other gene based vector tech. Serum samples were collected at weeks 4, 8, 12, and 23 from mice immunized at weeks 0 and 4. Enhanced antitumor effect of a gene gundelivered dna vaccine encoding the human papillomavirus type 16 oncoproteins genetically fused to the herpes simplex virus glycoprotein d m. The technique is based on bombardment of a targeted cellular surface.

While a gene gun might sound like a handy thing to have, the technology behind it is complex, delicate, and as of 2009, not safe for use in people. Both have required efforts to increase their potency either via manipulating the plasmid. The study on dna vaccines first began in the 1990s, when the plasmid dna is injected into the skin. Dna vaccines encoding iipadre generates potent padre. New physical and chemical approaches to facilitate dna entry into host cells deserve the continued attention of the scientific community. A major thrust in the application of gene transfer technology for cancer therapy has been the modulation of the immune response. Once in the nucleus, the antigen encoded by the dna vaccine needs to be expressed and presented on. Gene guns have also been used experimentally to deliver genetic vaccines, and to introduce dna into animal cells for the purpose of tagging them for microscopy or for genetic engineering of such cells.

Gene gun bombardment with dnacoated golden particles. It is also used to deliver plasmids into the neurons of rat. In the next study, 12 mice were immunized by intradermal gene gun inoculation 31. Since dna is highly stable, it can be stored at the dna enters myocytes 6 and antigen room temperature. The dna is injected into the muscle of the animal being vaccinated, usually with a gene gun that uses compressed gas to blow the dna into the muscle cells. One groups historical reflections on dna vaccine development. The protection elicited by the ha dna vaccine correlated with the titers of neutralizing antibody. The study on dna vaccines first began in the 1990s, when the plasmid dna is injected into the skin or muscle was reported to induce antibody responses to antigens. Apr 09, 2020 these are important issues because unlike other forms of gene therapy, which target very ill patients, dna vaccines are targeted at the young and the healthy. The plasmid dna vaccine above carries the genetic code for a piece of pathogen or tumor antigen. Dna immunization by intramuscular injection or gene gun. Dna vaccines have emerged as a potentially important form of antigenspecific immunotherapy because of their safety, ease of production, and stability.

The most commonly used delivery strategies are physical methods, such as electroporation, sonoporation 25, 26, dna tattooing, or gene gun, which are able to overcome the extra and intracellular barriers to transport dna into the nucleus. Vaxfectinformulated influenza dna vaccines encoding np. Gene gun particlemediated dna vaccination was performed using a heliumdriven gene gun biorad, hercules, ca according to the protocol provided by the manufacturer. Restifo national cancer institute, national institutes of health, building 10, bethesda, md. In mice, 95% protection was achieved by two immunizations with beads loaded with as little as 0. Elicited by gene gun immunization for humoral and cellmediated. Successful mucosal routes of vaccination included dna drops administered to the nares or trachea. In genetic engineering, a gene gun or biolistic particle delivery system is a device used to deliver exogenous dna, rna, or protein to cells. Many dna vaccines are injected into the muscle, however, a method using a gene gun is. Genetic dna immunization is a novel technique used to efficiently stimulate humoral and cellular immune responses to protein antigens. Gene vaccines, or the use of antigenencoding dnas to vaccinate, represent anewapproachto the developmentof subunit.

Ep dna immunization had a higher antihdv seroconversion rate of 80%, but the antihdv antibody responses were generally weak titer 400. Better vaccines through innovative dna immunization asu. Immunological properties of gene vaccines delivered by different. C57bl6 mice were immunized with empty plasmid, e7 dna or sige7lamp1 dna via gene gun. In summary, our results demonstrate the benefit of combined therapy with gene gun delivered antitumor dna vaccines and systemic administration of cpgodn or lms. Belyantseva abstract heliosr gene gun mediated transfection is a biolistic method for mechanical delivery of exogenous dna into cells in vitro or in vivo. Dna vaccines into dendritic cells via gene gun provides an opportunity to modify the quality and quantity of dna transfected dendritic cells and influence vaccine potency. Pdf low dose and gene gun immunization with a hepatitis. This method is also useful in studying the neurodegenerative diseases in humans like alzheimers disease. Gene vaccines, or the use of antigenencoding dnas to vaccinate, represent anewapproachto the developmentof subunit vaccines 14, 29. For dna vaccine concept to realize its promise and full potential, the delivery barrier has to be solved. An example of dna vaccine plasmid is pvac, which uses sv40 promoter. Enhanced antitumor effect of a gene gundelivered dna.

Immune responses induced by gene gun or intramuscular injection. Recent work has suggested that direct transfection of apcs plays a predominant role after gene gun administration of dna. Pdf delivery of a dna vaccine for alzheimers disease by. By far the most efficient dna immunizations were achieved by using a gene gun to deliver dna coated gold beads to the epidermis. The recombinant plasmid administrated by gene gun achieved a medium. Antigenic protein is presented by cell in which it is produced exogenous. The particles pass through the plant cells, leaving the dna inside.

Chapter 7 heliosr gene gun mediated transfection of the inner ear sensory epithelium inna a. The gene gun is a device used to transfect cells with foreign dna by bombarding the target cells with dna coated microparticles. The reason for this is due to the gene gun s ability to inject the dna loaded particles directly into the cytosol of the target cells. Enhancement of dna tumor vaccine efficacy by gene gun. There are some unique properties of dna vaccines which represent. When the dna vaccine is administered to a patient, the machinery in their cells makes a viral or bacterial protein which their immune system recognises as being foreign to the body. Antibody titers from mice immunized with pcdna3 sera 17382 or pcdna3 sera 17110 dna vaccines by intradermal gene gun injection. Adsorbed plasmid dna into gold particles ballastically accelerated into body with gene gun. This is one way fullers dna vaccine gets around the genetic drift, or changes, that occur in influenza strains from year to year, and challenge for.

Modification of professional antigenpresenting cells with. Although the quality and safety considerations for vaccines for veterinary use differ from those for human use, experience with veterinary dna vaccines can provide valuable information for the control and use of human dna vaccines. It was originally developed using plant tissue as the target but can also be used with yeast, bacteria, mammalian cell lines, and insects. Vaccination with dna may provide a viable alternative or may be used in conjunction with proteinbased subunit vaccines to maximize the efficacy of a human. By far the most efficient dna immunizations were achieved by using a gene gun to deliver dnacoated gold beads to the epidermis. Serial dilutions were used to measure the end pointpositive titers. Antigenic protein is formed in one cell but presented by different cell how dna vaccines work muscle cells. Since dna vaccines are able to induce both humoral and cellular immune responses, an important consideration in the application of dna vaccines is the immunization routes employed.

Gene gun delivery of dna vaccines has proven to be effective for protective and therapeutic immunization in various animal models. This dna gene gun vaccine study is the first of its kind to be performed by the u. Protective efficacy of hepatitis e virus dna vaccine. Gene gun using a gene gun directly shoots a piece of dna into the recipient plant tissue.

Immunogenicity and protective efficacy of a dna vaccine encoding ag85a from mycobacterium tuberculosis were compared in balbc and c57bl b6 and b10 mice immunized by intramuscular i. While both have been under development since the 1990s, in recent years, significant excitement has turned to mrna despite the licensure of several veterinary dna vaccines. The first clinical trail, initiated to monitor the safety and efficacy of a dna vaccine against hiv1 infection, demonstrated that dna plasmid vaccines were safe and were capable of inducing detectable immune cellular and antibody responses 4042. In balbc mice, gg immunization could induce elevated antibody and cytotoxic t lymphocyte responses with plasmid doses 50fold lower than those. The plasmid vector is taken up into cells and transcribed in the nucleus 1. The delivery of plasmids into rat neurons through the use of a gene gun, specifically drg neurons, is also used as a pharmacological precursor in studying the effects of neurodegenerative diseases such as alzheimers disease. Engineered nanodelivery systems to improve dna vaccine. The potency of dna vaccines encoding influenza virus antigens in many of these animal studies was demonstrated by the low amounts of dna required to induce immunity, using either im injection of the vaccine or by use of the gene gun fynan et al. Evaluation of the kinetic change of the immunogenicity of. Accessory regions pertaining to the plasmid backbone may engage in a wide range of structural instability phenomena.

Analysis of humoral immunity of hepatitis d virus dna. The transfection efficiency is higher when using the gene gun, since the dna forces its entrance into the cells. Kilpatrick,1 thomas cutler,2 eric whrtehorn,2 robert j. Pdf low dose and gene gun immunization with a hepatitis c. A dna vaccine uses a gene from a virus or bacteria to stimulate the immune system. The relative importance of these two distinct pathways in tcell priming by dna vaccines is the subject of current debate, and it may be dependent upon the route of dna administration. Evaluation of vaccine efficacy in a syngeneic her2neu mouse tumor model.

Dna vaccines delivered by microneedle and tattoo gun induce. Plasmid constructproduction of a fulllength hev orf2 based on the burmese hev strain sequence pcdna3. We used the gene gun method to vaccinate c57bl6 mice intradermally with dna vaccines containing the hpv16 e7 gene, the e7 gene. Although such dna vaccination by gene gun is effective in. Their safety in terms of adverse reactions after injection has been demonstrated in animal models 38, 39 as well as in human clinical trials. Enhancement of dna vaccine potency by linkage of antigen. Dna vaccines showed great promise in preclinical models of infectious and malignant diseases, but their potency was insufficient in clinical trials and is needed to be improved. The latter entails coating small gold particles with vaccine dna, followed by airpressuremediated intradermal delivery. In the present study, we sought to improve the efficacy of our candidate dna vaccine by administering it with a gene gun.

This protocol describes gene gun delivery of dna vaccine by a heliumdriven gene gun device. There has been a veritable explosion of information regarding the components of the immune response that are required to generate a meaningful cellular response to tumorassociated antigens taas capable of eliciting rejection of established tumor. No dna vaccine has been licenced for use in humans yet although some dna vaccines are now in clinical trials, none are licenced for use so they are an unproven method. Gene gun delivery of dna which propels the dna coated gold particles into the epidermis83, 84 resulted in a more th2 biased antibody isotype response and efficient humoral and cellular responses76, 77. Dna vaccinations were performed predominantly using two different methods.