Gene gun dna vaccine pdf

Gene gun particlemediated dna vaccination was performed using a heliumdriven gene gun biorad, hercules, ca according to the protocol provided by the manufacturer. Ep dna immunization had a higher antihdv seroconversion rate of 80%, but the antihdv antibody responses were generally weak titer 400. Plasmid constructproduction of a fulllength hev orf2 based on the burmese hev strain sequence pcdna3. C57bl6 mice were immunized with empty plasmid, e7 dna or sige7lamp1 dna via gene gun. The dna vaccine administrated by particle bombardment induced higher. Serial dilutions were used to measure the end pointpositive titers. For dna vaccine concept to realize its promise and full potential, the delivery barrier has to be solved. Their safety in terms of adverse reactions after injection has been demonstrated in animal models 38, 39 as well as in human clinical trials. Pdf low dose and gene gun immunization with a hepatitis c.

Engineered nanodelivery systems to improve dna vaccine. Gene gun immunization uses gold particles as a carrier for dna molecules. The transfection efficiency is higher when using the gene gun, since the dna forces its entrance into the cells. Dna vaccine against west nile virus for use in horses was. 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. 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. By far the most efficient dna immunizations were achieved by using a gene gun to deliver dna coated gold beads to the epidermis. In human and animals, the gene gun method is used to deliver dna vaccines into the body. Gene gun using a gene gun directly shoots a piece of dna into the recipient plant tissue. In mice, 95% protection was achieved by two immunizations with beads loaded with as little as 0.

The recombinant plasmid administrated by gene gun achieved a medium. 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. 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. New physical and chemical approaches to facilitate dna entry into host cells deserve the continued attention of the scientific community. Because the gene gun delivers a small amount of dna efficiently, it is a promising approach for dna vaccination. Evaluation of the kinetic change of the immunogenicity of. Antigenic protein is presented by cell in which it is produced. An example of dna vaccine plasmid is pvac, which uses sv40 promoter. Protective efficacy of hepatitis e virus dna vaccine. Chickens inoculated via the im route displayed an increase in ndvspecific antibodies and cellmeditated immunity. Evaluation of vaccine efficacy in a syngeneic her2neu mouse tumor model. Enhancement of dna tumor vaccine efficacy by gene gun.

Elicited by gene gun immunization for humoral and cellmediated. The direct injection of genetic material into a living host causes a small amount of its cells to produce the introduced gene. This dna gene gun vaccine study is the first of its kind to be performed by the u. 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.

In the next study, 12 mice were immunized by intradermal gene gun inoculation 31. Vaxfectinformulated influenza dna vaccines encoding np. Plasmid dna was precipitated onto 1mmdiameter gold beads using a solution of 1. Modification of professional antigenpresenting cells with. Immune responses induced by gene gun or intramuscular. Pdf delivery of a dna vaccine for alzheimers disease by. Nevertheless, intramuscular injection is a variable technique. 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. 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. Gene vaccines, or the use of antigenencoding dnas to vaccinate, represent anewapproachto the developmentof subunit. In general, it is desirable to employ efficient routes for in vivo delivery of dna vaccines directly into dcs.

The study on dna vaccines first began in the 1990s, when the plasmid dna is injected into the skin. Gene gun inoculation requires lower dna doses than i. Recent advances in delivery of veterinary dna vaccines. Restifo national cancer institute, national institutes of health, building 10, bethesda, md. 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.

Many dna vaccines are injected into the muscle, however, a method using a gene gun is. It is also used to deliver plasmids into the neurons of rat. 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. In this study, we evaluated the immunogenicity and the protective efficacy of the sm23 dna vaccine using two different intradermal dna delivery methods. The protection elicited by the ha dna vaccine correlated with the titers of neutralizing antibody. 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. The dnacoated gold particles are driven into the epidermal layers of. 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. The dna vaccines elicit potent humoral and cellmediated responses and provide a promising method for treating rapidly mutating and evasive diseases such as cancer and human immunode.

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. 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. Tuberculosis dna vaccine encoding ag85a is immunogenic and. 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. 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. Accessory regions pertaining to the plasmid backbone may engage in a wide range of structural instability phenomena.

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. The study on dna vaccines first began in the 1990s, when the plasmid dna is. Although such dna vaccination by gene gun is effective in.

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. 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. Antigenic protein is formed in one cell but presented by different cell how dna vaccines work muscle cells. We used the gene gun method to vaccinate c57bl6 mice intradermally with dna vaccines containing the hpv16 e7 gene, the e7 gene. Enhancement of dna vaccine potency by linkage of antigen gene to an hsp70 gene1. Gene vaccines, or the use of antigenencoding dnas to vaccinate, represent anewapproachto the developmentof subunit vaccines 14, 29. 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.

Annex 1 guidelines for assuring the quality and nonclinical. Low dose and gene gun immunization with a hepatitis c virus nonstructural ns 3 dna based vaccine containing ns4a inhibit ns34aexpressing tumors in vivo. Both have required efforts to increase their potency either via manipulating the plasmid. Kilpatrick,1 thomas cutler,2 eric whrtehorn,2 robert j. Dna vaccines encoding iipadre generates potent padre.

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. Pdf low dose and gene gun immunization with a hepatitis. Dna vaccines genetic dna immunization is a novel technique used to efficiently stimulate humoral and cellular immune responses to protein antigens. In balbc mice, gg immunization could induce elevated antibody and cytotoxic t lymphocyte responses with plasmid doses 50fold lower than those. Dna vaccines were pioneered by several groups in the early 1990s. The gene gun is a unique means of introducing naked dna vaccines directly into cells, bypassing the cell membrane and its receptors.

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. The distinct th1 or th2biased immune responses elicited by intramuscular injection or gene gun delivery, respectively, are not fully. This protocol describes gene gun delivery of dna vaccine by a heliumdriven gene gun device. Genetic vaccination using naked plasmid dna is an immunization strategy both against infectious diseases and cancer. Dna vaccines have emerged as a potentially important form of antigenspecific immunotherapy because of their safety, ease of production, and stability. The plasmid dna vaccine above carries the genetic code for a piece of pathogen or tumor antigen. In the present study, we sought to improve the efficacy of our candidate dna vaccine by administering it with a gene gun. The plasmid vector is taken up into cells and transcribed in the nucleus 1. Gene gunmediated dna immunization primes development of. 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 advantages of gene gun mediated delivery include signi.

Once in the nucleus, the antigen encoded by the dna vaccine needs to be expressed and presented on. The latter entails coating small gold particles with vaccine dna, followed by airpressuremediated intradermal delivery. Tungsten or gold beads are coated in the gene of interest and fired through a stopping screen, accelerated by helium, into the plant tissue. 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. Conclusionthe low humoral immunogenicity of dna vaccine with lhdag cannot be ameliorated by different dosage, gene gun immunization, or in vivo ep intramuscular injection. Belyantseva abstract heliosr gene gun mediated transfection is a biolistic method for mechanical delivery of exogenous dna into cells in vitro or in vivo.

Gene gunmediated dna vaccination induces antitumor immunity. 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. Intro duc tion immun ity is the d efens e mechan ism tha t work s in the. Gene gun delivery of dna vaccines has proven to be effective for protective and therapeutic immunization in various animal models. Since dna is highly stable, it can be stored at the dna enters myocytes 6 and antigen room temperature. This method is also useful in studying the neurodegenerative diseases in humans like alzheimers disease. One groups historical reflections on dna vaccine development. Antigenic protein is presented by cell in which it is produced exogenous. It was originally developed using plant tissue as the target but can also be used with yeast, bacteria, mammalian cell lines, and insects. Systemic administration of cpg oligodeoxynucleotide and. Chapter 7 heliosr gene gun mediated transfection of the inner ear sensory epithelium inna a.

Like recombinant vaccines, genes for the desired antigens are located and cloned. Dna vaccine with a lhdag construct may not be a candidate hdv vaccine to generate antihdv humoral immunity. Ctl response in c57bl6 mice immunized with various recombinant dna vaccines. Enhanced antitumor effect of a gene gundelivered dna. The gene gun is a device used to transfect cells with foreign dna by bombarding the target cells with dna coated microparticles. Serum samples were collected at weeks 4, 8, 12, and 23 from mice immunized at weeks 0 and 4. 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. Immunological properties of gene vaccines delivered by different. Dna vaccinations were performed predominantly using two different methods.

Cur gested that an epitope vaccine composed of a selfb cell rently, the two major delivery methods used for enhancing epitope of a. The past, current and future trends in dna vaccine. The concept of using dna immunization and other gene based vector tech. Analysis of humoral immunity of hepatitis d virus dna. In order to improve the efficacy of dna vaccines, particularly in large animals. In summary, our results demonstrate the benefit of combined therapy with gene gun delivered antitumor dna vaccines and systemic administration of cpgodn or lms.

Several reports on rodents suggest that dna vaccines can be used to achieve mucosal immunity when applied to mucosal tissues. Currently, dna vaccines have received increased attention and are. 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. Dna vaccines delivered by microneedle and tattoo gun induce. Dec 15, 1993 successful mucosal routes of vaccination included dna drops administered to the nares or trachea. Two dna vaccine plasmids, pcdna3 sera 17382 and pcdna3 sera 17110 were coated onto gold particles. We and others have successfully used plasmid dna pdnabased vaccines to induce. There are some unique properties of dna vaccines which represent. By far the most efficient dna immunizations were achieved by using a gene gun to deliver dnacoated gold beads to the epidermis.

Structural instability phenomena are of particular concern for plasmid manufacture, dna vaccination and gene therapy. The particles pass through the plant cells, leaving the dna inside. Cattle were immunized with a plasmid encoding bovine herpesvirus 1 bhv1 glycoprotein b, which was delivered with a gene gun either intradermally or intravulvomucosally. Gene gun immunization uses the particle bombardment or particlemediated dna delivery technology that was developed as a physical gene transfer method for various in vivo, ex vivo, or in vitro applications.

Gene gun has emerged as a novel tool for delivery of dna vaccines directly into dcs com. A prototype gene gun accell was acquired from agracetus. 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. 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. Gene guns have also been used to deliver dna vaccines. In this study, the potential of inducing mucosal immunity in cattle by immunization with a dna vaccine was demonstrated. The dna vaccine in fullers lab was engineered by using genetic components of influenza virus the conserved areas which do not change.

Through improving our understanding of basic immunology as it is related to dna vaccine technology, our. Dna immunization using a gene gun to deliver gold beads coated with dna plasmids. Gene gun delivered dna based immunizations mediate rapid production of murine monoclonal antibodies to the flt3 receptor katherine e. A dna vaccine uses a gene from a virus or bacteria to stimulate the immune system. 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. Gene gunmediated dna vaccination induces antitumor. Analysis of humoral immunity of hepatitis d virus dna vaccine. A major thrust in the application of gene transfer technology for cancer therapy has been the modulation of the immune response. Enhancement of dna vaccine potency by linkage of antigen.

Use of gene gun for genetic immunotherapy springerlink. Sm23 has already been shown to elicit protective immune responses following immunization with peptides or dna constructs. Better vaccines through innovative dna immunization asu. 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. Immune responses induced by gene gun or intramuscular injection. 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 technique is based on bombardment of a targeted cellular surface. 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. 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. Recent work has suggested that direct transfection of apcs plays a predominant role after gene gun administration of dna. For the np dna, boosted gene gun immunizations of 0.

Gene gun bombardment with dnacoated golden particles. Dna vaccine, gene gun, hepatitis d virus, in vivo electroporation hdv. 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. Genetic dna immunization is a novel technique used to efficiently stimulate humoral and cellular immune responses to protein antigens. 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. Dna immunization by intramuscular injection or gene gun. 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. Gene gunmediated dna vaccination enhances antigenspecific. Dna vaccines consist of transient transfection of naked dna into target cells. The direct injection of genetic material into a living host causes a small amount of its cells to produce the introduced gene products. The dna vaccine conferred protection to 40% of chickens against ndv upon challenge.