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Sample records for linear dna vaccines

  1. DNA vaccines

    Science.gov (United States)

    Gregersen, Jens-Peter

    2001-12-01

    Immunization by genes encoding immunogens, rather than with the immunogen itself, has opened up new possibilities for vaccine research and development and offers chances for new applications and indications for future vaccines. The underlying mechanisms of antigen processing, immune presentation and regulation of immune responses raise high expectations for new and more effective prophylactic or therapeutic vaccines, particularly for vaccines against chronic or persistent infectious diseases and tumors. Our current knowledge and experience of DNA vaccination is summarized and critically reviewed with particular attention to basic immunological mechanisms, the construction of plasmids, screening for protective immunogens to be encoded by these plasmids, modes of application, pharmacokinetics, safety and immunotoxicological aspects. DNA vaccines have the potential to accelerate the research phase of new vaccines and to improve the chances of success, since finding new immunogens with the desired properties is at least technically less demanding than for conventional vaccines. However, on the way to innovative vaccine products, several hurdles have to be overcome. The efficacy of DNA vaccines in humans appears to be much less than indicated by early studies in mice. Open questions remain concerning the persistence and distribution of inoculated plasmid DNA in vivo, its potential to express antigens inappropriately, or the potentially deleterious ability to insert genes into the host cell's genome. Furthermore, the possibility of inducing immunotolerance or autoimmune diseases also needs to be investigated more thoroughly, in order to arrive at a well-founded consensus, which justifies the widespread application of DNA vaccines in a healthy population.

  2. DNA vaccines against influenza.

    Science.gov (United States)

    Stachyra, Anna; Góra-Sochacka, Anna; Sirko, Agnieszka

    2014-01-01

    Genetic vaccine technology has been considerably developed within the last two decades. This cost effective and promising strategy can be applied for therapy of cancers and for curing allergy, chronic and infectious diseases, such as a seasonal and pandemic influenza. Despite numerous advantages, several limitations of this technology reduce its performance and can retard its commercial exploitation in humans and its veterinary applications. Inefficient delivery of the DNA vaccine into cells of immunized individuals results in low intracellular supply of suitable expression cassettes encoding an antigen, in its low expression level and, in turn, in reduced immune responses against the antigen. Improvement of DNA delivery into the host cells might significantly increase effectiveness of the DNA vaccine. A vast array of innovative methods and various experimental strategies have been applied in order to enhance the effectiveness of DNA vaccines. They include various strategies improving DNA delivery as well as expression and immunogenic potential of the proteins encoded by the DNA vaccines. Researchers focusing on DNA vaccines against influenza have applied many of these strategies. Recent examples of the most successful modern approaches are discussed in this review.

  3. DNA vaccines and intradermal vaccination by DNA tattooing.

    Science.gov (United States)

    Oosterhuis, K; van den Berg, J H; Schumacher, T N; Haanen, J B A G

    2012-01-01

    Over the past two decades, DNA vaccination has been developed as a method for the induction of immune responses. However, in spite of high expectations based on their efficacy in preclinical models, immunogenicity of first generation DNA vaccines in clinical trials was shown to be poor, and no DNA vaccines have yet been licensed for human use. In recent years significant progress has been made in the development of second generation DNA vaccines and DNA vaccine delivery methods. Here we review the key characteristics of DNA vaccines as compared to other vaccine platforms, and recent insights into the prerequisites for induction of immune responses by DNA vaccines will be discussed. We illustrate the development of second generation DNA vaccines with the description of DNA tattooing as a novel DNA delivery method. This technique has shown great promise both in a small animal model and in non-human primates and is currently under clinical evaluation.

  4. DNA vaccine: the miniature miracle

    Directory of Open Access Journals (Sweden)

    Karthik Kaliaperumal

    2013-08-01

    Full Text Available DNA, the essential part of the life is making way in to new vaccine technology. Plasmid vectors from the bacteria have revolutionized the world of vaccine design by its new technology – DNA vaccines. Small portion of the nucleotides from the pathogen held under the control of promoter in a plasmid vector can be used as a vaccine. DNA vaccines alleviate the odds of the other vaccines by having good hold on both the faces of the immunity. The key to the success of DNA vaccine lies in the route of administration of the vaccine which can be done in many ways. Prime boost strategy is an approach used to boost the action of DNA vaccine. To date there are only four DNA vaccine available in the market. [Vet World 2013; 6(4.000: 228-232

  5. DNA fusion gene vaccines

    DEFF Research Database (Denmark)

    Holst, Peter Johannes; Bassi, Maria Rosaria; Thomsen, Allan Randrup

    2010-01-01

    DNA vaccines are versatile and safe, but limited immunogenicity has prevented their use in the clinical setting. Experimentally, immunogenicity may be enhanced by the use of new delivery technologies, by coadministration of cytokines and pathogen-associated molecular patterns, or by fusion...... of antigens into molecular domains that enhance antigen presentation. More specifically, the immunogenicity of DNA vaccines may benefit from increased protein synthesis, increased T-cell help and MHC class I presentation, and the addition of a range of specific cytokines and pathogen-associated molecular...... patterns that increase activation of the innate immune system. Importantly, viral-vectored vaccines that act through the induction of one or more of these factors also may benefit from cytokine coadministration and increased antigen presentation. In order to increase immunogenicity to the level achieved...

  6. Development of dengue DNA vaccines.

    Science.gov (United States)

    Danko, Janine R; Beckett, Charmagne G; Porter, Kevin R

    2011-09-23

    Vaccination with plasmid DNA against infectious pathogens including dengue is an active area of investigation. By design, DNA vaccines are able to elicit both antibody responses and cellular immune responses capable of mediating long-term protection. Great technical improvements have been made in dengue DNA vaccine constructs and trials are underway to study these in the clinic. The scope of this review is to highlight the rich history of this vaccine platform and the work in dengue DNA vaccines accomplished by scientists at the Naval Medical Research Center. This work resulted in the only dengue DNA vaccine tested in a clinical trial to date. Additional advancements paving the road ahead in dengue DNA vaccine development are also discussed.

  7. Ensuring safety of DNA vaccines

    Directory of Open Access Journals (Sweden)

    Wessels Stephen

    2005-09-01

    Full Text Available Abstract In 1990 a new approach for vaccination was invented involving injection of plasmid DNA in vivo, which elicits an immune response to the encoded protein. DNA vaccination can overcome most disadvantages of conventional vaccine strategies and has potential for vaccines of the future. However, today 15 years on, a commercial product still has not reached the market. One possible explanation could be the technique's failure to induce an efficient immune response in humans, but safety may also be a fundamental issue. This review focuses on the safety of the genetic elements of DNA vaccines and on the safety of the microbial host for the production of plasmid DNA. We also propose candidates for the vaccine's genetic elements and for its microbial production host that can heighten the vaccine's safety and facilitate its entry to the market.

  8. DNA vaccine for cancer immunotherapy.

    Science.gov (United States)

    Yang, Benjamin; Jeang, Jessica; Yang, Andrew; Wu, T C; Hung, Chien-Fu

    2014-01-01

    DNA vaccination has emerged as an attractive immunotherapeutic approach against cancer due to its simplicity, stability, and safety. Results from numerous clinical trials have demonstrated that DNA vaccines are well tolerated by patients and do not trigger major adverse effects. DNA vaccines are also very cost effective and can be administered repeatedly for long-term protection. Despite all the practical advantages, DNA vaccines face challenges in inducing potent antigen specific cellular immune responses as a result of immune tolerance against endogenous self-antigens in tumors. Strategies to enhance immunogenicity of DNA vaccines against self-antigens have been investigated including encoding of xenogeneic versions of antigens, fusion of antigens to molecules that activate T cells or trigger associative recognition, priming with DNA vectors followed by boosting with viral vector, and utilization of immunomodulatory molecules. This review will focus on discussing strategies that circumvent immune tolerance and provide updates on findings from recent clinical trials.

  9. DNA vaccines for aquacultured fish

    DEFF Research Database (Denmark)

    Lorenzen, Niels; LaPatra, S.E.

    2005-01-01

    Deoxyribonucleic acid (DNA) vaccination is based on the administration of the gene encoding the vaccine antigen, rather than the antigen itself. Subsequent expression of the antigen by cells in the vaccinated hosts triggers the host immune system. Among the many experimental DNA vaccines tested...... in various animal species as well as in humans, the vaccines against rhabdovirus diseases in fish have given some of the most promising results. A single intramuscular (IM) injection of microgram amounts of DNA induces rapid and long-lasting protection in farmed salmonids against economically important...... viruses such as infectious haematopoietic necrosis virus (IHNV) and viral haemorrhagic septicaemia virus (VHSV). DNA vaccines against other types of fish pathogens, however, have so far had limited success. The most efficient delivery route at present is IM injection, and suitable delivery strategies...

  10. DNA vaccines for aquacultured fish.

    Science.gov (United States)

    Lorenzen, N; LaPatra, S E

    2005-04-01

    Deoxyribonucleic acid (DNA) vaccination is based on the administration of the gene encoding the vaccine antigen, rather than the antigen itself. Subsequent expression of the antigen by cells in the vaccinated hosts triggers the host immune system. Among the many experimental DNA vaccines tested in various animal species as well as in humans, the vaccines against rhabdovirus diseases in fish have given some of the most promising results. A single intramuscular (IM) injection of microgram amounts of DNA induces rapid and long-lasting protection in farmed salmonids against economically important viruses such as infectious haematopoietic necrosis virus (IHNV) and viral haemorrhagic septicaemia virus (VHSV). DNA vaccines against other types of fish pathogens, however, have so far had limited success. The most efficient delivery route at present is IM injection, and suitable delivery strategies for mass vaccination of small fish have yet to be developed. In terms of safety, no adverse effects in the vaccinated fish have been observed to date. As DNA vaccination is a relatively new technology, various theoretical and long-term safety issues related to the environment and the consumer remain to be fully addressed, although inherently the risks should not be any greater than with the commercial fish vaccines that are currently used. Present classification systems lack clarity in distinguishing DNA-vaccinated animals from genetically modified organisms (GMOs), which could raise issues in terms of licensing and public acceptance of the technology. The potential benefits of DNA vaccines for farmed fish include improved animal welfare, reduced environmental impacts of aquaculture activities, increased food quality and quantity, and more sustainable production. Testing under commercial production conditions has recently been initiated in Canada and Denmark.

  11. DNA vaccines for aquacultured fish

    DEFF Research Database (Denmark)

    Lorenzen, Niels; LaPatra, S.E.

    2005-01-01

    in various animal species as well as in humans, the vaccines against rhabdovirus diseases in fish have given some of the most promising results. A single intramuscular (IM) injection of microgram amounts of DNA induces rapid and long-lasting protection in farmed salmonids against economically important...... viruses such as infectious haematopoietic necrosis virus (IHNV) and viral haemorrhagic septicaemia virus (VHSV). DNA vaccines against other types of fish pathogens, however, have so far had limited success. The most efficient delivery route at present is IM injection, and suitable delivery strategies...... for mass vaccination of small fish have yet to be developed. In terms of safety, no adverse effects in the vaccinated fish have been observed to date. As DNA vaccination is a relatively new technology, various theoretical and long-term safety issues related to the environment and the consumer remain...

  12. Biotechnology and DNA vaccines for aquatic animals

    Science.gov (United States)

    Kurath, G.

    2008-01-01

    Biotechnology has been used extensively in the development of vaccines for aquaculture. Modern molecular methods such as polymerase chain reaction (PCR), cloning and microarray analysis have facilitated antigen discovery, construction of novel candidate vaccines, and assessments of vaccine efficacy, mode of action, and host response. This review focuses on DNA vaccines for finfish to illustrate biotechnology applications in this field. Although DNA vaccines for fish rhabdoviruses continue to show the highest efficacy, DNA vaccines for several other viral and bacterial fish pathogens have now been proven to provide significant protection against pathogen challenge. Studies of the fish rhabdovirus DNA vaccines have elucidated factors that affect DNA vaccine efficacy as well as the nature of the fish innate and adaptive immune responses to DNA vaccines. As tools for managing aquatic animal disease emergencies, DNA vaccines have advantages in speed, flexibility, and safety, and one fish DNA vaccine has been licensed.

  13. DNA vaccines for aquacultured fish

    DEFF Research Database (Denmark)

    Lorenzen, Niels; LaPatra, S.E.

    2005-01-01

    of licensing and public acceptance of the technology. The potential benefits of DNA vaccines for farmed fish include improved animal welfare, reduced environmental impacts of aquaculture activities, increased food quality and quantity, and more sustainable production. Testing under commercial production...

  14. DNA vaccines for viral diseases

    Directory of Open Access Journals (Sweden)

    Donnelly J.J.

    1999-01-01

    Full Text Available DNA plasmids encoding foreign proteins may be used as immunogens by direct intramuscular injection alone, or with various adjuvants and excipients, or by delivery of DNA-coated gold particles to the epidermis through biolistic immunization. Antibody, helper T lymphocyte, and cytotoxic T lymphocyte (CTL responses have been induced in laboratory and domesticated animals by these methods. In a number of animal models, immune responses induced by DNA vaccination have been shown to be protective against challenge with various infectious agents. Immunization by injection of plasmids encoding foreign proteins has been used successfully as a research tool. This review summarizes the types of DNA vaccine vectors in common use, the immune responses and protective responses that have been obtained in animal models, the safety considerations pertinent to the evaluation of DNA vaccines in humans and the very limited information that is available from early clinical studies.

  15. The future of human DNA vaccines.

    Science.gov (United States)

    Li, Lei; Saade, Fadi; Petrovsky, Nikolai

    2012-12-31

    DNA vaccines have evolved greatly over the last 20 years since their invention, but have yet to become a competitive alternative to conventional protein or carbohydrate based human vaccines. Whilst safety concerns were an initial barrier, the Achilles heel of DNA vaccines remains their poor immunogenicity when compared to protein vaccines. A wide variety of strategies have been developed to optimize DNA vaccine immunogenicity, including codon optimization, genetic adjuvants, electroporation and sophisticated prime-boost regimens, with each of these methods having its advantages and limitations. Whilst each of these methods has contributed to incremental improvements in DNA vaccine efficacy, more is still needed if human DNA vaccines are to succeed commercially. This review foresees a final breakthrough in human DNA vaccines will come from application of the latest cutting-edge technologies, including "epigenetics" and "omics" approaches, alongside traditional techniques to improve immunogenicity such as adjuvants and electroporation, thereby overcoming the current limitations of DNA vaccines in humans.

  16. Oral DNA Vaccine in Chickens

    Directory of Open Access Journals (Sweden)

    Seyed Davoud Jazayeri

    2012-01-01

    Full Text Available Attenuated Salmonella has been used as a carrier for DNA vaccine. However, in vitro and in vivo studies on the bacteria following transfection of plasmid DNA were poorly studied. In this paper, eukaryotic expression plasmids encoding avian influenza virus (AIV subtype H5N1 genes, pcDNA3.1/HA, NA, and NP, were transfected into an attenuated Salmonella enteric typhimurium SV4089. In vitro stability of the transfected plasmids into Salmonella were over 90% after 100 generations. The attenuated Salmonella were able to invade MCF-7 (1.2% and MCF-10A (0.5% human breast cancer cells. Newly hatched specific-pathogen-free (SPF chicks were inoculated once by oral gavage with 109 colony-forming unit (CFU of the attenuated Salmonella. No abnormal clinical signs or deaths were recorded after inoculation. Viable bacteria were detected 3 days after inoculation by plating from spleen, liver, and cecum. Fluorescent in situ hybridization (FISH and polymerase chain reaction (PCR were carried out for confirmation. Salmonella was not detected in blood cultures although serum antibody immune responses to Salmonella O antiserum group D1 factor 1, 9, and 12 antigens were observed in all the inoculated chickens after 7 days up to 35 days. Our results showed that live attenuated S. typhimurium SV4089 harboring pcDNA3.1/HA, NA, and NP may provide a unique alternative as a carrier for DNA oral vaccine in chickens.

  17. Novel approaches to tuberculosis prevention: DNA vaccines.

    Science.gov (United States)

    Rivas-Santiago, Bruno; Cervantes-Villagrana, Alberto R

    2014-03-01

    It is estimated that there are approximately eight million new cases of active tuberculosis (TB) worldwide annually. There is only 1 vaccine available for prevention: bacillus Calmette-Guérin (BCG). This has variable efficacy and is only protective for certain extrapulmonary TB cases in children, therefore new strategies for the creation of novel vaccines have emerged. One of the promising approaches is the DNA vaccine, used as a direct vaccination or as a prime-boost vaccine. This review describes the experimental data obtained during the design of DNA vaccines for TB.

  18. Influenza DNA vaccine:an update

    Institute of Scientific and Technical Information of China (English)

    陈则

    2004-01-01

    @@ A series of global studies on the influenza DNA vaccine have revealed that it is capable of eliciting persistent humoral and cell mediated immune responses to influenza following delivery by various routes. DNA vaccines may not only serve as potentially safer alternatives to immunization with certain live virus vaccines, but may also provide a promising approach to the development of effective vaccines. The suggestions, based on our experiment, that both hemagglutinin (HA)- and neuraminidase (NA)- DNAs (or both HA and NA molecules) are highly protective against the influenza virus and are useful in the development of a more efficient vaccine against the influenza virus. In this article, we reviewed DNA vaccine against the influenza A and B viruses and the characteristics of the immune response induced by the DNA vaccine. Moreover, we discussed the importance of neutralizing antibodies to protect the host against a lethal influenza infection.

  19. Formulation and delivery of dermal DNA vaccines

    NARCIS (Netherlands)

    van den Berg, J.H.

    2009-01-01

    DNA vaccination is an appealing strategy of active vaccination, leading to the intracellular production of the encoding antigen which results in an efficient activation of an antigen specific immune response. Intradermal DNA tattooing was recently developed as a simple and robust method to induce

  20. Formulation and delivery of dermal DNA vaccines

    NARCIS (Netherlands)

    van den Berg, J.H.|info:eu-repo/dai/nl/304837016

    2009-01-01

    DNA vaccination is an appealing strategy of active vaccination, leading to the intracellular production of the encoding antigen which results in an efficient activation of an antigen specific immune response. Intradermal DNA tattooing was recently developed as a simple and robust method to induce an

  1. Micro- and nanoparticulates for DNA vaccine delivery.

    Science.gov (United States)

    Farris, Eric; Brown, Deborah M; Ramer-Tait, Amanda E; Pannier, Angela K

    2016-05-01

    DNA vaccination has emerged as a promising alternative to traditional protein-based vaccines for the induction of protective immune responses. DNA vaccines offer several advantages over traditional vaccines, including increased stability, rapid and inexpensive production, and flexibility to produce vaccines for a wide variety of infectious diseases. However, the immunogenicity of DNA vaccines delivered as naked plasmid DNA is often weak due to degradation of the DNA by nucleases and inefficient delivery to immune cells. Therefore, biomaterial-based delivery systems based on micro- and nanoparticles that encapsulate plasmid DNA represent the most promising strategy for DNA vaccine delivery. Microparticulate delivery systems allow for passive targeting to antigen presenting cells through size exclusion and can allow for sustained presentation of DNA to cells through degradation and release of encapsulated vaccines. In contrast, nanoparticle encapsulation leads to increased internalization, overall greater transfection efficiency, and the ability to increase uptake across mucosal surfaces. Moreover, selection of the appropriate biomaterial can lead to increased immune stimulation and activation through triggering innate immune response receptors and target DNA to professional antigen presenting cells. Finally, the selection of materials with the appropriate properties to achieve efficient delivery through administration routes conducive to high patient compliance and capable of generating systemic and local (i.e. mucosal) immunity can lead to more effective humoral and cellular protective immune responses. In this review, we discuss the development of novel biomaterial-based delivery systems to enhance the delivery of DNA vaccines through various routes of administration and their implications for generating immune responses.

  2. Vaccine development using recombinant DNA technology

    Science.gov (United States)

    Vaccines induce an immune response in the host that subsequently recognizes infectious agents and helps fight off the disease; vaccines must do this without causing the disease. This paper reviews the development of recombinant DNA technologies as a means of providing new ways for attenuating diseas...

  3. Improving DNA vaccine performance through vector design.

    Science.gov (United States)

    Williams, James A

    2014-01-01

    DNA vaccines are a rapidly deployed next generation vaccination platform for treatment of human and animal disease. DNA delivery devices, such as electroporation and needle free jet injectors, are used to increase gene transfer. This results in higher antigen expression which correlates with improved humoral and cellular immunity in humans and animals. This review highlights recent vector and transgene design innovations that improve DNA vaccine performance. These new vectors improve antigen expression, increase plasmid manufacturing yield and quality in bioreactors, and eliminate antibiotic selection and other potential safety issues. A flowchart for designing synthetic antigen transgenes, combining antigen targeting, codon-optimization and bioinformatics, is presented. Application of improved vectors, of antibiotic free plasmid production, and cost effective manufacturing technologies will be critical to ensure safety, efficacy, and economically viable manufacturing of DNA vaccines currently under development for infectious disease, cancer, autoimmunity, immunotolerance and allergy indications.

  4. DNA Vaccine Electroporation and Molecular Adjuvants

    Science.gov (United States)

    2016-03-16

    according to the manufacturer’s suggested values (see Note 6). 3. Draw DNA solution into syringe (see Note 7). 4. Anesthetize the animal with the...vaccination is an attractive method for inducing protective immunity to a variety of pathogens, but the low immunogenicity seen in larger animals and...for generating protective immunity against filovirus infection [11]. The demonstration of effective DNA vaccination in small animal models changed the

  5. DNA vaccination strategies against infectious diseases.

    Science.gov (United States)

    Watts, A M; Kennedy, R C

    1999-08-01

    DNA immunisation represents a novel approach to vaccine and immunotherapeutic development. Injection of plasmid DNA encoding a foreign gene of interest can result in the subsequent expression of the foreign gene products and the induction of an immune response within a host. This is relevant to prophylactic and therapeutic vaccination strategies when the foreign gene represents a protective epitope from a pathogen. The recent demonstration by a number of laboratories that these immune responses evoke protective immunity against some infectious diseases and cancers provides support for the use of this approach. In this article, we attempt to present an informative and unbiased representation of the field of DNA immunisation. The focus is on studies that impart information on the development of vaccination strategies against a number of human and animal pathogens. Investigations that describe the mechanism(s) of protective immunity induced by DNA immunisation highlight the advantages and disadvantages of this approach to developing vaccines within a given system. A variety of systems in which DNA vaccination has resulted in the induction of protective immunity, as well as the correlates associated with these protective immune responses, will be described. Particular attention will focus on systems involving parasitic diseases. Finally, the potential of DNA immunisation is discussed as it relates to veterinary medicine and its role as a possible vaccine strategy against animal coccidioses.

  6. Recent advances towards the clinical application of DNA vaccines.

    Science.gov (United States)

    Bins, A D; van den Berg, J H; Oosterhuis, K; Haanen, J B A G

    2013-04-01

    DNA vaccination is an attractive method for therapeutic vaccination against intracellular pathogens and cancer. This review provides an introduction into the DNA vaccination field and discusses the pre-clinical successes and most interesting clinical achievements thus far. Furthermore, general attributes, mechanism of action and safety of DNA vaccination will be discussed. Since clinical results with DNA vaccination so far show room for improvement, possibilities to improve the delivery and immunogenicity of DNA vaccines are reviewed. In the coming years, these new developments should show whether DNA vaccination is able to induce clinically relevant responses in patients.

  7. Prediction and cloning linear Tcell epitopes of P14-3-3 antigen into pEGFP–N1 as a DNA vaccine model to induse immuno response against hydatidosis and it\\'s expression in CHO cell line

    Directory of Open Access Journals (Sweden)

    R mesri

    2015-11-01

    Full Text Available ABSTRACT Background & purpose: Hydatidosis is a zoonotic disease that caused by infection with the larvae of Echinococcus granulosus. Different antigens produced in larval stage of this parasite that recombinant vaccine base these antigens created significant immunity in infected animals. One of the important antigens is p14-3-3 that it's recombinant antigen created considerable immunity in mouse models. In this study according to the high immunity of antigen epitopes region the coding sequence of T-cell epitopes of P14-3-3 was cloned into pEGFP-N1vector in order to produce an effective DNA vaccine model to stimulate high level of Th1 immune response.   Material and method: In this study bioinformatics tools were used to prediction of linear T-Cell epitopes of Echinococcus granulosus P14-3-3 &zeta antigen. The nucleotide coding sequence of these epitopes was synthesized by PCR. the ampliqon was digested with XhoI restriction enzyme and cloned into pEGFP–N1 vector That has been purificated by modified sambrook method with CaCl2 and PEG6000..Positive colony was selected by direct colony PCR and confirmed by the sequencing.and evaluation of it's expression in Eukaryotic cells was done by transformed to CHO cell line with electroporation. Results: Linear T-cell epitopes of Echinococcus granulosus P14-3-3 after prediction,synthesis and amplification wae successfully cloned into pEGFP-N1 vector that purificated by new method with maximum vector and minimum RNA concentration.The expression of new constract in CHO cell line as a eukaryotic cells achivment by fluorescent microscope and will be used as a DNA vaccine model to evaluation immuno response in mouse models.   Discussion: Successfully cloning of The linear T-cell epitppes coding sequence of Echinococcus granulosus P14-3-3&zeta antigen into pEGFP-N1 verificated by sequencing and fluorscent microscope images demonstrated expression of recombinant protein in CHO cell line

  8. Polymer multilayer tattooing for enhanced DNA vaccination

    Science.gov (United States)

    Demuth, Peter C.; Min, Younjin; Huang, Bonnie; Kramer, Joshua A.; Miller, Andrew D.; Barouch, Dan H.; Hammond, Paula T.; Irvine, Darrell J.

    2013-04-01

    DNA vaccines have many potential benefits but have failed to generate robust immune responses in humans. Recently, methods such as in vivo electroporation have demonstrated improved performance, but an optimal strategy for safe, reproducible, and pain-free DNA vaccination remains elusive. Here we report an approach for rapid implantation of vaccine-loaded polymer films carrying DNA, immune-stimulatory RNA, and biodegradable polycations into the immune-cell-rich epidermis, using microneedles coated with releasable polyelectrolyte multilayers. Films transferred into the skin following brief microneedle application promoted local transfection and controlled the persistence of DNA and adjuvants in the skin from days to weeks, with kinetics determined by the film composition. These ‘multilayer tattoo’ DNA vaccines induced immune responses against a model HIV antigen comparable to electroporation in mice, enhanced memory T-cell generation, and elicited 140-fold higher gene expression in non-human primate skin than intradermal DNA injection, indicating the potential of this strategy for enhancing DNA vaccination.

  9. Development of DNA vaccines for fish

    DEFF Research Database (Denmark)

    Heppell, Joël; Lorenzen, Niels; Armstrong, Neil K.

    1998-01-01

    Disease control is one of the major concerns in the aquaculture industry. However, there are no vaccines available for the prevention of many piscine infectious diseases, especially those of viral and parasitic origin. DNA-based vaccination could circumvent several problems associated...... no permanent tissue damage. To further investigate the ability of DNA-based vaccines to induce protective immunity in fish, viral haemorrhagic septicaemia virus G and N genes were cloned individually into an expression plasmid. Both G and N proteins produced in transfected fish cells appeared identical...... protein, killing the transfected host cells and ablating further expression of G protein and luciferase. Finally, young rainbow trout injected with the G construct, alone or together with the N construct, were strongly protected against challenge with live virus. These results suggest that DNA vaccines...

  10. Antiparasitic DNA vaccines in 21st century.

    Science.gov (United States)

    Wedrychowicz, Halina

    2015-06-01

    Demands for effective vaccines to control parasitic diseases of humans and livestock have been recently exacerbated by the development of resistance of most pathogenic parasites to anti-parasitic drugs. Novel genomic and proteomic technologies have provided opportunities for the discovery and improvement of DNA vaccines which are relatively easy as well as cheap to fabricate and stable at room temperatures. However, their main limitation is rather poor immunogenicity, which makes it necessary to couple the antigens with adjuvant molecules. This paper review recent advances in the development of DNA vaccines to some pathogenic protozoa and helminths. Numerous studies were conducted over the past 14 years of 21st century, employing various administration techniques, adjuvants and new immunogenic antigens to increase efficacy of DNA vaccines. Unfortunately, the results have not been rewarding. Further research is necessary using more extensive combinations of antigens; alternate delivery systems and more efficient adjuvants based on knowledge of the immunomodulatory capacities of parasitic protozoa and helminths.

  11. HIV DNA Vaccine: Stepwise Improvements Make a Difference

    Directory of Open Access Journals (Sweden)

    Barbara K. Felber

    2014-05-01

    Full Text Available Inefficient DNA delivery methods and low expression of plasmid DNA have been major obstacles for the use of plasmid DNA as vaccine for HIV/AIDS. This review describes successful efforts to improve DNA vaccine methodology over the past ~30 years. DNA vaccination, either alone or in combination with other methods, has the potential to be a rapid, safe, and effective vaccine platform against AIDS. Recent clinical trials suggest the feasibility of its translation to the clinic.

  12. DNA vaccination in skin enhanced by electroporation.

    Science.gov (United States)

    Broderick, Kate E; Khan, Amir S; Sardesai, Niranjan Y

    2014-01-01

    DNA vaccines are a next generation branch of vaccines which offer major benefits over their conventional counterparts. However, to be effective in large mammals and humans, an enhancing delivery technology is required. Electroporation is a physical technique which results in improved delivery of large molecules through the cell membrane. In the case of plasmid DNA, electroporation enhances both the uptake and expression of the delivered DNA. The skin is an attractive tissue for DNA vaccination in a clinical setting due to the accessibility of the target, the ease of monitoring, and most importantly the immunocompetent nature of the dermis. Electroporation in the skin has the benefit of being minimally invasive and generally well tolerated. Previous studies have determined that optimized electroporation parameters (such as electrical field intensity, pulse length, pulse width, and plasmid formulation) majorly impact the efficiency of DNA delivery to the skin. We provide an overview of DNA vaccination in skin and muscle. In addition, we detail a protocol for the successful intradermal electroporation of plasmid DNA to guinea pig skin, an excellent dermatological animal model. The work detailed here suggests that the technique is safe and effective and could be highly applicable to a clinical setting.

  13. DNA-Based Vaccine Protects Against Zika in Animal Study

    Science.gov (United States)

    ... page: https://medlineplus.gov/news/fullstory_161959.html DNA-Based Vaccine Protects Against Zika in Animal Study ... In animals infected with Zika virus, the synthetic DNA-based vaccine was 100 percent effective in protecting ...

  14. DNA-Based Vaccine Guards Against Zika in Monkey Study

    Science.gov (United States)

    ... page: https://medlineplus.gov/news/fullstory_161106.html DNA-Based Vaccine Guards Against Zika in Monkey Study ... THURSDAY, Sept. 22, 2016 (HealthDay News) -- An experimental DNA-based vaccine protected monkeys from infection with the ...

  15. Lipopolysaccharide contamination in intradermal DNA vaccination : toxic impurity or adjuvant?

    NARCIS (Netherlands)

    Berg, J.H. van den; Quaak, S.G.L.; Beijnen, J.H.; Hennink, W.E.; Storm, G.; Schumacher, T.N.; Haanen, J.B.A.G.; Nuijen, B.

    2010-01-01

    Purpose: Lipopolysaccharides (LPS) are known both as potential adjuvants for vaccines and as toxic impurity in pharmaceutical preparations. The aim of this study was to assess the role of LPS in intradermal DNA vaccination administered by DNA tattooing. Method: Micewere vaccinated with a model DNA v

  16. Lipopolysaccharide contamination in intradermal DNA vaccination : toxic impurity or adjuvant?

    NARCIS (Netherlands)

    Berg, J.H. van den; Quaak, S.G.L.; Beijnen, J.H.; Hennink, W.E.; Storm, G.; Schumacher, T.N.; Haanen, J.B.A.G.; Nuijen, B.

    Purpose: Lipopolysaccharides (LPS) are known both as potential adjuvants for vaccines and as toxic impurity in pharmaceutical preparations. The aim of this study was to assess the role of LPS in intradermal DNA vaccination administered by DNA tattooing. Method: Micewere vaccinated with a model DNA

  17. Influenza Plasmid DNA Vaccines: Progress and Prospects.

    Science.gov (United States)

    Bicho, Diana; Queiroz, João António; Tomaz, Cândida Teixeira

    2015-01-01

    Current influenza vaccines have long been used to fight flu infectious; however, recent advances highlight the importance of produce new alternatives. Even though traditional influenza vaccines are safe and usually effective, they need to be uploaded every year to anticipate circulating flu viruses. This limitation together with the use of embryonated chicken eggs as the substrate for vaccine production, is time-consuming and could involve potential biohazards in growth of new virus strains. Plasmid DNA produced by prokaryote microorganisms and encoding foreign proteins had emerged as a promising therapeutic tool. This technology allows the expression of a gene of interest by eukaryotic cells in order to induce protective immune responses against the pathogen of interest. In this review, we discuss the strategies to choose the best DNA vaccine to be applied in the treatment and prevention of influenza. Specifically, we give an update of influenza DNA vaccines developments, all involved techniques, their main characteristics, applicability and technical features to obtain the best option against influenza infections.

  18. Vaccination strategies for SEIR models using feedback linearization. Preliminary results

    CERN Document Server

    De la Sen, M; Alonso-Quesada, S

    2011-01-01

    A linearization-based feedback-control strategy for a SEIR epidemic model is discussed. The vaccination objective is the asymptotically tracking of the removed-by-immunity population to the total population while achieving simultaneously the remaining population (i.e. susceptible plus infected plus infectious) to asymptotically tend to zero. The disease controlpolicy is designed based on a feedback linearization technique which provides a general method to generate families of vaccination policies with sound technical background.

  19. Innate Immune Signaling by, Genetic Adjuvants for, DNA Vaccination

    Directory of Open Access Journals (Sweden)

    Kouji Kobiyama

    2013-07-01

    Full Text Available DNA vaccines can induce both humoral and cellular immune responses. Although some DNA vaccines are already licensed for infectious diseases in animals, they are not licensed for human use because the risk and benefit of DNA vaccines is still controversial. Indeed, in humans, the immunogenicity of DNA vaccines is lower than that of other traditional vaccines. To develop the use of DNA vaccines in the clinic, various approaches are in progress to enhance or improve the immunogenicity of DNA vaccines. Recent studies have shown that immunogenicity of DNA vaccines are regulated by innate immune responses via plasmid DNA recognition through the STING-TBK1 signaling cascade. Similarly, molecules that act as dsDNA sensors that activate innate immune responses through STING-TBK1 have been identified and used as genetic adjuvants to enhance DNA vaccine immunogenicity in mouse models. However, the mechanisms that induce innate immune responses by DNA vaccines are still unclear. In this review, we will discuss innate immune signaling upon DNA vaccination and genetic adjuvants of innate immune signaling molecules.

  20. Innate Immune Signaling by, and Genetic Adjuvants for DNA Vaccination.

    Science.gov (United States)

    Kobiyama, Kouji; Jounai, Nao; Aoshi, Taiki; Tozuka, Miyuki; Takeshita, Fumihiko; Coban, Cevayir; Ishii, Ken J

    2013-01-01

    DNA vaccines can induce both humoral and cellular immune responses. Although some DNA vaccines are already licensed for infectious diseases in animals, they are not licensed for human use because the risk and benefit of DNA vaccines is still controversial. Indeed, in humans, the immunogenicity of DNA vaccines is lower than that of other traditional vaccines. To develop the use of DNA vaccines in the clinic, various approaches are in progress to enhance or improve the immunogenicity of DNA vaccines. Recent studies have shown that immunogenicity of DNA vaccines are regulated by innate immune responses via plasmid DNA recognition through the STING-TBK1 signaling cascade. Similarly, molecules that act as dsDNA sensors that activate innate immune responses through STING-TBK1 have been identified and used as genetic adjuvants to enhance DNA vaccine immunogenicity in mouse models. However, the mechanisms that induce innate immune responses by DNA vaccines are still unclear. In this review, we will discuss innate immune signaling upon DNA vaccination and genetic adjuvants of innate immune signaling molecules.

  1. Second Generation Therapeutic DNA Lymphoma Vaccines

    Science.gov (United States)

    2010-05-01

    ovalbumin (OVA) (Fig.1A: Protein Vaccine Constructs). Desired recombinant proteins were expressed in a SF9 insect cell/baculovirus expression...1500μg of purified protein per 108 virus infected SF9 cells) of mBD2-OVA protein (Fig. 1B) versus poor yielding (~250μg of purified protein per 108...virus infected SF9 DNA Vaccine Median Survival Time (Days*) Log-rank p-value vs PBS mBD2-gp100F 32.000 .017 MIP-3α-gp100F 32.000 .011 MCP-3-gp100F

  2. Linear IgA bullous dermatosis following influenza vaccination.

    Science.gov (United States)

    Alberta-Wszolek, Lauren; Mousette, Alyse M; Mahalingam, Meera; Levin, Nikki A

    2009-11-15

    Linear IgA Bullous Dermatosis (LABD) is an immune-mediated subepidermal vesiculobullous eruption characterized by linear deposits of IgA at the basement membrane zone. Most cases are idiopathic but medications, infections, and malignancies have also been reported to induce LABD. We report the case of a 54-year-old woman who developed LABD shortly after receiving an influenza vaccination.

  3. Preclinical and clinical safety studies on DNA vaccines.

    NARCIS (Netherlands)

    Schalk, Johanna A C; Mooi, Frits R; Berbers, Guy A M; Aerts, Leon A G J M van; Ovelgönne, Hans; Kimman, Tjeerd G

    2007-01-01

    DNA vaccines are based on the transfer of genetic material, encoding an antigen, to the cells of the vaccine recipient. Despite high expectations of DNA vaccines as a result of promising preclinical data their clinical utility remains unproven. However, much data is gathered in preclinical and

  4. Preclinical and clinical safety studies on DNA vaccines.

    NARCIS (Netherlands)

    Schalk, Johanna A C; Mooi, Frits R; Berbers, Guy A M; Aerts, Leon A G J M van; Ovelgönne, Hans; Kimman, Tjeerd G

    2007-01-01

    DNA vaccines are based on the transfer of genetic material, encoding an antigen, to the cells of the vaccine recipient. Despite high expectations of DNA vaccines as a result of promising preclinical data their clinical utility remains unproven. However, much data is gathered in preclinical and clini

  5. Interference of an ERM-vaccine with a VHS-DNA vaccine in rainbow trout

    DEFF Research Database (Denmark)

    Lorenzen, Ellen; Einer-Jensen, Katja; Rasmussen, Jesper Skou

    Simultaneous vaccination of fish against several diseases is often desirable in order to minimise cost and handling of the fish. Intramuscular DNA-vaccination of rainbow trout against viral haemorrhagic septicaemia virus (VHSV) has proved to provide very good protection. However, preliminary...... results showed that intraperitoneal injection of a commercial vaccine against Enteric Redmouth Disease (ERM) based on formalin-killed bacteria in oil adjuvant immediately followed by intramuscular injection of an experimental DNA-vaccine against VHSV, decreased the protective effect of the DNA-vaccine...... against challenge with VHSV 11 weeks post vaccination (pv). This experiment was performed with rainbow trout of 30 g injected with 0.5 g VHS-DNA vaccine. The experiment was later repeated with smaller fish (2.5g) and using two different doses of DNA-vaccine, 1 g and 0.05 g. Both doses provided good...

  6. Efficient vaccine against pandemic influenza: combining DNA vaccination and targeted delivery to MHC class II molecules.

    Science.gov (United States)

    Grødeland, Gunnveig; Bogen, Bjarne

    2015-06-01

    There are two major limitations to vaccine preparedness in the event of devastating influenza pandemics: the time needed to generate a vaccine and rapid generation of sufficient amounts. DNA vaccination could represent a solution to these problems, but efficacy needs to be enhanced. In a separate line of research, it has been established that targeting of vaccine molecules to antigen-presenting cells enhances immune responses. We have combined the two principles by constructing DNA vaccines that encode bivalent fusion proteins; these target hemagglutinin to MHC class II molecules on antigen-presenting cells. Such DNA vaccines rapidly induce hemagglutinin-specific antibodies and T cell responses in immunized mice. Responses are long-lasting and protect mice against challenge with influenza virus. In a pandemic situation, targeted DNA vaccines could be produced and tested within a month. The novel DNA vaccines could represent a solution to pandemic preparedness in the advent of novel influenza pandemics.

  7. Using Plasmids as DNA Vaccines for Infectious Diseases.

    Science.gov (United States)

    Tregoning, John S; Kinnear, Ekaterina

    2014-12-01

    DNA plasmids can be used to induce a protective (or therapeutic) immune response by delivering genes encoding vaccine antigens. That naked DNA (without the refinement of coat proteins or host evasion systems) can cross from outside the cell into the nucleus and be expressed is particularly remarkable given the sophistication of the immune system in preventing infection by pathogens. As a result of the ease, low cost, and speed of custom gene synthesis, DNA vaccines dangle a tantalizing prospect of the next wave of vaccine technology, promising individual designer vaccines for cancer or mass vaccines with a rapid response time to emerging pandemics. There is considerable enthusiasm for the use of DNA vaccination as an approach, but this enthusiasm should be tempered by the successive failures in clinical trials to induce a potent immune response. The technology is evolving with the development of improved delivery systems that increase expression levels, particularly electroporation and the incorporation of genetically encoded adjuvants. This review will introduce some key concepts in the use of DNA plasmids as vaccines, including how the DNA enters the cell and is expressed, how it induces an immune response, and a summary of clinical trials with DNA vaccines. The review also explores the advances being made in vector design, delivery, formulation, and adjuvants to try to realize the promise of this technology for new vaccines. If the immunogenicity and expression barriers can be cracked, then DNA vaccines may offer a step change in mass vaccination.

  8. Strategies and hurdles using DNA vaccines to fish.

    Science.gov (United States)

    Hølvold, Linn B; Myhr, Anne I; Dalmo, Roy A

    2014-01-01

    DNA vaccinations against fish viral diseases as IHNV at commercial level in Canada against VHSV at experimental level are both success stories. DNA vaccination strategies against many other viral diseases have, however, not yet yielded sufficient results in terms of protection. There is an obvious need to combat many other viral diseases within aquaculture where inactivated vaccines fail. There are many explanations to why DNA vaccine strategies against other viral diseases fail to induce protective immune responses in fish. These obstacles include: 1) too low immunogenicity of the transgene, 2) too low expression of the transgene that is supposed to induce protection, 3) suboptimal immune responses, and 4) too high degradation rate of the delivered plasmid DNA. There are also uncertainties with regard distribution and degradation of DNA vaccines that may have implications for safety and regulatory requirements that need to be clarified. By combining plasmid DNA with different kind of adjuvants one can increase the immunogenicity of the transgene antigen - and perhaps increase the vaccine efficacy. By using molecular adjuvants with or without in combination with targeting assemblies one may expect different responses compared with naked DNA. This includes targeting of DNA vaccines to antigen presenting cells as a central factor in improving their potencies and efficacies by means of encapsulating the DNA vaccine in certain carriers systems that may increase transgene and MHC expression. This review will focus on DNA vaccine delivery, by the use of biodegradable PLGA particles as vehicles for plasmid DNA mainly in fish.

  9. A comparative approach between heterologous prime-boost vaccination strategy and DNA vaccinations for rabies.

    Science.gov (United States)

    Borhani, Kiandokht; Ajorloo, Mehdi; Bamdad, Taravat; Mozhgani, Sayed Hamid Reza; Ghaderi, Mostafa; Gholami, Ali Reza

    2015-04-01

    Rabies is a widespread neurological zoonotic disease causing significant mortality rates, especially in developing countries. Although a vaccine for rabies is available, its production and scheduling are costly in such countries. Advances in recombinant DNA technology have made it a good candidate for an affordable vaccine. Among the proteins of rabies virus, the Glycoprotein (RVG) has been the major target for new vaccine development which plays the principal role in providing complete protection against RV challenge. The aim of this study is to produce recombinant RVG which could be a DNA vaccine candidate and to evaluate the efficiency of this construct in a prime-boost vaccination regimen, compared to commercial vaccine. Cloning to pcDNA3.1(+) and expression of rabies virus glycoprotein gene in BSR cell  line were performed followed by SDS-PAGE and Western blot analysis of the expressed glycoprotein. The resulting genetic construct was used as a DNA vaccine by injecting 80 µg of the plasmid to MNRI mice twice. Prime-Boost vaccination strategy was performed using 80 µg plasmid construct as prime dose and the second dose of an inactivated rabies virus vaccine. Production of rabies virus neutralizing antibody (RVNA) titers of the serum samples were determined by RFFIT. In comparisons between heterologous prime-boost vaccination strategy and DNA vaccinations, the potency of group D that received Prime-Boost vaccine with the second dose of pcDNA3.1(+)-Gp was enhanced significantly compared to the group C which had received pcDNA3.1(+)-Gp as first injection. In this study, RVGP expressing construct was used in a comparative approach between Prime-Boost vaccination strategy and DNA vaccination and compared with the standard method of rabies vaccination. It was concluded that this strategy could lead to induction of acceptable humoral immunity.

  10. A pilot study comparing the development of EIAV Env-specific antibodies induced by DNA/recombinant vaccinia-vectored vaccines and an attenuated Chinese EIAV vaccine.

    Science.gov (United States)

    Meng, Qinglai; Lin, Yuezhi; Ma, Jian; Ma, Yan; Zhao, Liping; Li, Shenwei; Yang, Kai; Zhou, Jianhua; Shen, Rongxian; Zhang, Xiaoyan; Shao, Yiming

    2012-12-01

    Data from successful attenuated lentiviral vaccine studies indicate that fully mature Env-specific antibodies characterized by high titer, high avidity, and the predominant recognition of conformational epitopes are associated with protective efficacy. Although vaccination with a DNA prime/recombinant vaccinia-vectored vaccine boost strategy has been found to be effective in some trials with non-human primate/simian/human immunodeficiency virus (SHIV) models, it remains unclear whether this vaccination strategy could elicit mature equine infectious anemia virus (EIAV) Env-specific antibodies, thus protecting vaccinated horses against EIAV infection. Therefore, in this pilot study we vaccinated horses using a strategy based on DNA prime/recombinant Tiantan vaccinia (rTTV)-vectored vaccines encoding EIAV env and gag genes, and observed the development of Env-specific antibodies, neutralizing antibodies, and p26-specific antibodies. Vaccination with DNA induced low titer, low avidity, and the predominant recognition of linear epitopes by Env-specific antibodies, which was enhanced by boosting vaccinations with rTTV vaccines. However, the maturation levels of Env-specific antibodies induced by the DNA/rTTV vaccines were significantly lower than those induced by the attenuated vaccine EIAV(FDDV). Additionally, DNA/rTTV vaccines did not elicit broadly neutralizing antibodies. After challenge with a virulent EIAV strain, all of the vaccinees and control horses died from EIAV disease. These data indicate that the regimen of DNA prime/rTTV vaccine boost did not induce mature Env-specific antibodies, which might have contributed to immune protection failure.

  11. DNA/MVA Vaccines for HIV/AIDS

    Directory of Open Access Journals (Sweden)

    Smita S. Iyer

    2014-02-01

    Full Text Available Since the initial proof-of-concept studies examining the ability of antigen-encoded plasmid DNA to serve as an immunogen, DNA vaccines have evolved as a clinically safe and effective platform for priming HIV-specific cellular and humoral responses in heterologous “prime-boost” vaccination regimens. Direct injection of plasmid DNA into the muscle induces T- and B-cell responses against foreign antigens. However, the insufficient magnitude of this response has led to the development of approaches for enhancing the immunogenicity of DNA vaccines. The last two decades have seen significant progress in the DNA-based vaccine platform with optimized plasmid constructs, improved delivery methods, such as electroporation, the use of molecular adjuvants and novel strategies combining DNA with viral vectors and subunit proteins. These innovations are paving the way for the clinical application of DNA-based HIV vaccines. Here, we review preclinical studies on the DNA-prime/modified vaccinia Ankara (MVA-boost vaccine modality for HIV. There is a great deal of interest in enhancing the immunogenicity of DNA by engineering DNA vaccines to co-express immune modulatory adjuvants. Some of these adjuvants have demonstrated encouraging results in preclinical and clinical studies, and these data will be examined, as well.

  12. DNA/MVA Vaccines for HIV/AIDS.

    Science.gov (United States)

    Iyer, Smita S; Amara, Rama R

    2014-01-01

    Since the initial proof-of-concept studies examining the ability of antigen-encoded plasmid DNA to serve as an immunogen, DNA vaccines have evolved as a clinically safe and effective platform for priming HIV-specific cellular and humoral responses in heterologous "prime-boost" vaccination regimens. Direct injection of plasmid DNA into the muscle induces T- and B-cell responses against foreign antigens. However, the insufficient magnitude of this response has led to the development of approaches for enhancing the immunogenicity of DNA vaccines. The last two decades have seen significant progress in the DNA-based vaccine platform with optimized plasmid constructs, improved delivery methods, such as electroporation, the use of molecular adjuvants and novel strategies combining DNA with viral vectors and subunit proteins. These innovations are paving the way for the clinical application of DNA-based HIV vaccines. Here, we review preclinical studies on the DNA-prime/modified vaccinia Ankara (MVA)-boost vaccine modality for HIV. There is a great deal of interest in enhancing the immunogenicity of DNA by engineering DNA vaccines to co-express immune modulatory adjuvants. Some of these adjuvants have demonstrated encouraging results in preclinical and clinical studies, and these data will be examined, as well.

  13. Therapeutic DNA vaccines against tuberculosis: a promising but arduous task

    Institute of Scientific and Technical Information of China (English)

    LI Jun-ming; ZHU Dao-yin

    2006-01-01

    Objective To review recent developments in therapeutic DNA vaccines against tuberculosis.Data sources The data used in this review were obtained mainly from the studies of therapeutic DNA vaccines against tuberculosis reported from 2000 to 2006.Study selection Relevant articles about studies of therapeutic DNA vaccines against tuberculosis were selected.Data extraction Data were mainly extracted from the 32 articles listed in the reference section of this review.Results Some DNA vaccines which previously showed to induce protective immunity against infection by Mycobacterium tuberculosis in a prophylactic manner are also surprisingly effective when used therapeutically,including persistent Mycobacterium tuberculosis and multidrug-resistant tuberculosis which are refractory to immune system and antibacterial chemotherapy alone. When used in combination with antibacterial drugs,therapeutic DNA vaccines could effectively eliminate residual bacteria in infected animals and shorten the therapy course of conventional chemotherapy. Detailed studies demonstrated that therapeutic effects of DNA vaccines may at least partly be due to the restoration of the Th1/Th2 balance. Some problems have also emerged along with these exciting results.Conclusions Therapeutic DNA vaccine is a promising strategy against tuberculosis, however developing an ideal DNA vaccine for therapy of tuberculosis will require further development.

  14. Licensed DNA Vaccines against Infectious Hematopoietic Necrosis Virus (IHNV).

    Science.gov (United States)

    Alonso, Marta; Leong, Jo-Ann C

    2013-04-01

    This article reviews some of the recent patents on DNA vaccines against fish viruses, in particular against the novirhabdovirus infectious hematopoitic necrosis virus (IHNV). Although very effective in protecting fish against IHNV, only one DNA vaccine has been approved to date for use in Canada. In Europe and in US, its commercialization is restricted due to safety concerns.

  15. Vector Design for Improved DNA Vaccine Efficacy, Safety and Production

    Directory of Open Access Journals (Sweden)

    James A. Williams

    2013-06-01

    Full Text Available DNA vaccination is a disruptive technology that offers the promise of a new rapidly deployed vaccination platform to treat human and animal disease with gene-based materials. Innovations such as electroporation, needle free jet delivery and lipid-based carriers increase transgene expression and immunogenicity through more effective gene delivery. This review summarizes complementary vector design innovations that, when combined with leading delivery platforms, further enhance DNA vaccine performance. These next generation vectors also address potential safety issues such as antibiotic selection, and increase plasmid manufacturing quality and yield in exemplary fermentation production processes. Application of optimized constructs in combination with improved delivery platforms tangibly improves the prospect of successful application of DNA vaccination as prophylactic vaccines for diverse human infectious disease targets or as therapeutic vaccines for cancer and allergy.

  16. Lipopolysaccharide contamination in intradermal DNA vaccination: toxic impurity or adjuvant?

    Science.gov (United States)

    van den Berg, Joost H; Quaak, Susanne G L; Beijnen, Jos H; Hennink, Wim E; Storm, Gert; Schumacher, Ton N; Haanen, John B A G; Nuijen, Bastiaan

    2010-05-05

    Lipopolysaccharides (LPS) are known both as potential adjuvants for vaccines and as toxic impurity in pharmaceutical preparations. The aim of this study was to assess the role of LPS in intradermal DNA vaccination administered by DNA tattooing. Mice were vaccinated with a model DNA vaccine (Luc-NP) with an increasing content of residual LPS. The effect of LPS on systemic toxicity, antigen expression and cellular immunity was studied. The presence of LPS in the DNA vaccine neither induced systemic toxicity (as reflected by IL-6 concentration in serum), nor influenced antigen expression (measured by intravital imaging). Higher LPS contents however, appeared to be associated with an elevated cytotoxic T-lymphocyte (CTL) response but without reaching statistical significance. Interestingly, the DNA tattoo procedure by itself was shown to induce a serum cytokine response that was at least as potent as that induced by parenteral LPS administration. LPS does not show toxicity in mice vaccinated by DNA tattooing at dose levels well above those encountered in GMP-grade DNA preparations. Thus, residual LPS levels in the pharmaceutical range are not expected to adversely affect clinical outcome of vaccination trials and may in fact have some beneficial adjuvant effect. The observed pro-inflammatory effects of DNA tattoo may help explain the high immunogenicity of this procedure. Copyright 2009 Elsevier B.V. All rights reserved.

  17. The past, current and future trends in DNA vaccine immunisations

    OpenAIRE

    Sidgi Syed Anwer Abdo Hasson; Juma Khalifa Zayid Al-Busaidi; Talal Abdulmalek Sallam

    2015-01-01

    This review focuses on DNA vaccines, denoting the last two decades since the early substantiation of preclinical protection was published in Science in 1993 by Ulmer et al. In spite of being safely administered and easily engineered and manufactured DNA vaccine, it holds the future prospects of immunization by inducing potent cellular immune responses against infectious and non-infectious diseases. It is well documented that injection of DNA plasmid encoding a desired gene of interest can res...

  18. ANTICANCER DNA VACCINATION: PRINCIPLE AND PERSPECTIVES OF THE METHOD

    Directory of Open Access Journals (Sweden)

    M. V. Stegantseva

    2017-01-01

    Full Text Available Conventional strategies for cancer treatment are close to their efficiency limits. Meanwhile, rapid development of experimental immunology and immunotherapy led to first successful experiences in antitumor vaccination. Over last decade, remarkable results were obtained by means of anticancer vaccination being implemented into clinical settings thus causing popularity and growth of interest to tumor-specific DNA vaccines. In this review, we discuss basic principles of a DNA vaccine construction, their structural characteristics and diversity, mechanisms of their biological action, pharmaceutical forms and delivery routes into the body. 

  19. Applications of nanoparticles for DNA based rabies vaccine.

    Science.gov (United States)

    Shah, Muhammad Ali A; Khan, Sajid Umar; Ali, Zeeshan; Yang, Haowen; Liu, Keke; Mao, Lanlan

    2014-01-01

    Rabies is a fatal encephalomyelitis. Most cases occur in developing countries and are transmitted by dogs. The cell culture vaccines as associated with high cost; therefore, have not replaced the unsafe brain-derived vaccines. In the developing countries these brain-derived rabies vaccines still can be seen in action. Moreover, there will be a need for vaccines against rabies-related viruses against which classical vaccines are not always effective. The worldwide incidence of rabies and the inability of currently used vaccination strategies to provide highly potent and cost-effective therapy indicate the need for alternate control strategies. DNA vaccines have emerged as the safest vaccines and best remedy for complicated diseases like hepatitis, HIV, and rabies. A number of recombinant DNA vaccines are now being developed against several diseases such as AIDS and malaria. Therefore, it can be a valuable alternative for the production of cheaper rabies vaccines against its larger spectrum of viruses. In this review we report published data on DNA-based immunization with sequences encoding rabies with special reference to nanotechnology.

  20. Preclinical and clinical development of DNA vaccines for prostate cancer.

    Science.gov (United States)

    Colluru, V T; Johnson, Laura E; Olson, Brian M; McNeel, Douglas G

    2016-04-01

    Prostate cancer is the most commonly diagnosed cancer in the United States. It is also the second leading cause of cancer-related death in men, making it one of the largest public health concerns today. Prostate cancer is an ideal disease for immunotherapies because of the generally slow progression, the dispensability of the target organ in the patient population, and the availability of several tissue-specific antigens. As such, several therapeutic vaccines have entered clinical trials, with one autologous cellular vaccine (sipuleucel-T) recently gaining Food and Drug Administration approval after demonstrating overall survival benefit in randomized phase III clinical trials. DNA-based vaccines are safe, economical, alternative "off-the-shelf" approaches that have undergone extensive evaluation in preclinical models. In fact, the first vaccine approved in the United States for the treatment of cancer was a DNA vaccine for canine melanoma. Several prostate cancer-specific DNA vaccines have been developed in the last decade and have shown promising results in early phase clinical trials. This review summarizes anticancer human DNA vaccine trials, with a focus on those conducted for prostate cancer. We conclude with an outline of special considerations important for the development and successful translation of DNA vaccines from the laboratory to the clinic.

  1. DNA-based influenza vaccines as immunoprophylactic agents toward universality.

    Science.gov (United States)

    Zhang, Han; El Zowalaty, Mohamed E

    2016-01-01

    Influenza is an illness of global public health concern. Influenza viruses have been responsible for several pandemics affecting humans. Current influenza vaccines have proved satisfactory safety; however, they have limitations and do not provide protection against unexpected emerging influenza virus strains. Therefore, there is an urgent need for alternative approaches to conventional influenza vaccines. The development of universal influenza vaccines will help alleviate the severity of influenza pandemics. Influenza DNA vaccines have been the subject of many studies over the past decades due to their ability to induce broad-based protective immune responses in various animal models. The present review highlights the recent advances in influenza DNA vaccine research and its potential as an affordable universal influenza vaccine.

  2. Comparative Study on the Immunogenicity between Hsp70 DNA Vaccine and Hsp65 DNA Vaccine in Human Mycobacterium Tuberculosis

    Institute of Scientific and Technical Information of China (English)

    DAI; Wuxing; HUANG; Hailang; YUAN; Ye; HU; Jiajie; HUANGFU; Yongmu

    2001-01-01

    The BALB/c mice were immunized with Hsp70 DNA and Hsp65 DNA vaccines in human Mycobacterium tuberculosis. Eight weeks after immunization, the eyeballs were removed, blood and spleen taken, and intraperitoneal macrophages were harvested. The lymphocytic stimulating index(SI) was used to measure the cellular proliferating ability and NO release to measure the phagocytic activity of the macrophages. With ELISA kit, the levels of interleukin-2 (IL-2) and interferon-γ(IFN-γ) in serum and the splenic lymphocytic cultured supernatant were detected. The results showed that after the mice were immunized with 100 μg/mouse of Hsp70 DNA vaccine intramuscularly, the splenic lymphocytic proliferating ability in the mice was significantly increased as compared with that in the control group, vector group and Hsp65 DNA vaccine group (P<0. 01); The contents of NO in the intraperitoneal macrophages of the mice were significantly lower than in the control group and Hsp65 DNA vaccine group (P<0. 01); The levels of serum IL-2 in the mice were significantly higher than in the control group, but there was no statistical difference between Hsp65 DNA group and vector group (P>0. 05); The contents of serum IFN-γ in the mice were significantly higher than in the control group, but significantly lower than in the Hsp65 DNA vaccine group (P<0. 05). It was indicated that immunization with Hsp70 DNA vaccine could obviously enhance the immune response, but its intensity seemed inferior to Hsp65 DNA vaccine. The anti-infection mechanisms and clinical use in the future of the vaccines of Hsp70 DNA and Hsp65 DNA are worth further studying.

  3. Production and pharmaceutical formulation of plasmid DNA vaccines

    NARCIS (Netherlands)

    van der Heijden, I.

    2013-01-01

    Research leading to the thesis ‘Production and pharmaceutical formulation of plasmid DNA vaccines‘ can be divided into two parts. The first part describes the development of a Good Manufacturing Practice (GMP) compliant plasmid DNA production process of pDNA vaccines for the treatment of Human papil

  4. Production and pharmaceutical formulation of plasmid DNA vaccines

    NARCIS (Netherlands)

    van der Heijden, I.

    2013-01-01

    Research leading to the thesis ‘Production and pharmaceutical formulation of plasmid DNA vaccines‘ can be divided into two parts. The first part describes the development of a Good Manufacturing Practice (GMP) compliant plasmid DNA production process of pDNA vaccines for the treatment of Human papil

  5. DNA Vaccines against Protozoan Parasites: Advances and Challenges

    Directory of Open Access Journals (Sweden)

    Eric Dumonteil

    2007-01-01

    Full Text Available Over the past 15 years, DNA vaccines have gone from a scientific curiosity to one of the most dynamic research field and may offer new alternatives for the control of parasitic diseases such as leishmaniasis and Chagas disease. We review here some of the advances and challenges for the development of DNA vaccines against these diseases. Many studies have validated the concept of using DNA vaccines for both protection and therapy against these protozoan parasites in a variety of mouse models. The challenge now is to translate what has been achieved in these models into veterinary or human vaccines of comparable efficacy. Also, genome-mining and new antigen discovery strategies may provide new tools for a more rational search of novel vaccine candidates.

  6. DNA vaccines: a rational design against parasitic diseases.

    Science.gov (United States)

    Carvalho, Joana A; Rodgers, Jean; Atouguia, Jorge; Prazeres, Duarte M F; Monteiro, Gabriel A

    2010-02-01

    Parasitic diseases are one of the most devastating causes of morbidity and mortality worldwide. Although immunization against these infections would be an ideal solution, the development of effective vaccines has been hampered by specific challenges posed by parasitic pathogens. Plasmid-based DNA vaccines may prove to be promising immunization tools in this area because vectors can be designed to integrate several antigens from different stages of the parasite life cycle or different subspecies; vaccines, formulations and immunization protocols can be tuned to match the immune response that offers protective immunity; and DNA vaccination is an affordable platform for developing countries. Partial and full protective immunity have been reported following DNA vaccination against the most significant parasitic diseases in the world.

  7. Current trends in separation of plasmid DNA vaccines: a review.

    Science.gov (United States)

    Ghanem, Ashraf; Healey, Robert; Adly, Frady G

    2013-01-14

    Plasmid DNA (pDNA)-based vaccines offer more rapid avenues for development and production if compared to those of conventional virus-based vaccines. They do not rely on time- or labour-intensive cell culture processes and allow greater flexibility in shipping and storage. Stimulating antibodies and cell-mediated components of the immune system are considered as some of the major advantages associated with the use of pDNA vaccines. This review summarizes the current trends in the purification of pDNA vaccines for practical and analytical applications. Special attention is paid to chromatographic techniques aimed at reducing the steps of final purification, post primary isolation and intermediate recovery, in order to reduce the number of steps necessary to reach a purified end product from the crude plasmid.

  8. Assessment of a DNA Vaccine Encoding Burkholderia pseudomallei Bacterioferritin

    Science.gov (United States)

    2007-08-01

    bacterioferritin gene from Brucella abortus, when delivered to mice as a DNA vaccine, evokes a potent Th1 immune response, including strong IFN-γ...blocking buffer containing goat anti-mouse IgG alkaline phosphatase conjugate (Sigma) at a dilution of 1:30000 for 1hr at room temperature. Following...Walravens, and J. J. Letesson. 2001. Induction of immune response in BALB/c mice with a DNA vaccine encoding bacterioferritin or P39 of Brucella

  9. DNA vaccination of small rainbow trout fry against VHSV

    DEFF Research Database (Denmark)

    Rasmussen, Jesper Skou; Lorenzen, Ellen; Kjær, Torben Egil;

    2012-01-01

    Small rainbow trout fry were DNA vaccinated by intramuscular injection at 0.25g and other fish later at 0.5g. Vaccine groups included pcDNA3-vhsG, heterologous vaccine (pcDNA3-ihnG), empty vector (pcDNA3) and unhandled fish. Fry vaccinated at 0.25g were challenged with VHSV by immersion at 3wpv, 11......wpv and 21wpv. The challenge at 3wpv was started 1wpv, however as no mortality was observed, the fish were re-challenged 3wpv using a modified setup. Fry vaccinated at 0.5g were challenged with VHSV by immersion at11wpv. By early challenge (3wpv) of fish vaccinated at 0.25g both homologous...... and heterologous challenge (5% mortality). At 11 wpv an unspecific protection with 30 % mortality was observed. At 21 wpv protection against VHSV had dropped further (50 % mortality). Protection against IHNV was better (10 % mortality) but equal for both homologous and heterologous vaccines confirming previous...

  10. The past, current and future trends in DNA vaccine immunisations

    Directory of Open Access Journals (Sweden)

    Sidgi Syed Anwer Abdo Hasson

    2015-05-01

    Full Text Available This review focuses on DNA vaccines, denoting the last two decades since the early substantiation of preclinical protection was published in Science in 1993 by Ulmer et al. In spite of being safely administered and easily engineered and manufactured DNA vaccine, it holds the future prospects of immunization by inducing potent cellular immune responses against infectious and non-infectious diseases. It is well documented that injection of DNA plasmid encoding a desired gene of interest can result in the subsequent expression of its products and lead to the induction of an immune response within a host. This is pertinent to prophylactic and therapeutic vaccination approach when the peculiar gene produces a protective epitope from a pathogen. The recent studies demonstrated by a number of research centers showed that these immune responses evoke protective immunity against several infectious diseases and cancers, which provides adequate support for the use of this approach. We attempt in this review to provide an informative and unbiased overview of the general principles and concept of DNA vaccines technology with a summary of a novel approach to the DNA vaccine, present investigations that describe the mechanism(s of protective immunity provoked by DNA immunization and to highlight the advantages and disadvantages of DNA immunisation.

  11. The past, current and future trends in DNA vaccine immunisations

    Institute of Scientific and Technical Information of China (English)

    Sidgi; Syed; Anwer; Abdo; Hasson; Juma; Khalifa; Zayid; Al-Busaidi; Talal; Abdulmalek; Sallam

    2015-01-01

    This review focuses on DNA vaccines, denoting the last two decades since the early substantiation of preclinical protection was published in Science in 1993 by Ulmer et al. In spite of being safely administered and easily engineered and manufactured DNA vaccine, it holds the future prospects of immunization by inducing potent cellular immune responses against infectious and non-infectious diseases. It is well documented that injection of DNA plasmid encoding a desired gene of interest can result in the subsequent expression of its products and lead to the induction of an immune response within a host. This is pertinent to prophylactic and therapeutic vaccination approach when the peculiar gene produces a protective epitope from a pathogen. The recent studies demonstrated by a number of research centers showed that these immune responses evoke protective immunity against several infectious diseases and cancers, which provides adequate support for the use of this approach. We attempt in this review to provide an informative and unbiased overview of the general principles and concept of DNA vaccines technology with a summary of a novel approach to the DNA vaccine, present investigations that describe the mechanism(s) of protective immunity provoked by DNA immunization and to highlight the advantages and disadvantages of DNA immunisation.

  12. Targeting DNA vaccines to myeloid cells using a small peptide.

    Science.gov (United States)

    Ye, Chunting; Choi, Jang Gi; Abraham, Sojan; Shankar, Premlata; Manjunath, N

    2015-01-01

    Targeting DNA vaccines to dendritic cells (DCs) greatly enhances immunity. Although several approaches have been used to target protein Ags to DCs, currently there is no method that targets DNA vaccines directly to DCs. Here, we show that a small peptide derived from the rabies virus glycoprotein fused to protamine residues (RVG-P) can target DNA to myeloid cells, including DCs, which results in enhanced humoral and T-cell responses. DCs targeted with a DNA vaccine encoding the immunodominant vaccinia B8R gene via RVG-P were able to restimulate vaccinia-specific memory T cells in vitro. Importantly, a single i.v. injection of B8R gene bound to RVG-P was able to prime a vaccinia-specific T-cell response that was able to rapidly clear a subsequent vaccinia challenge in mice. Moreover, delivery of DNA in DCs was enough to induce DC maturation and efficient Ag presentation without the need for adjuvants. Finally, immunization of mice with a DNA-vaccine encoding West Nile virus (WNV) prM and E proteins via RVG-P elicited high titers of WNV-neutralizing Abs that protected mice from lethal WNV challenge. Thus, RVG-P provides a reagent to target DNA vaccines to myeloid cells and elicit robust T-cell and humoral immune responses.

  13. Epitope DNA vaccines against tuberculosis: spacers and ubiquitin modulates cellular immune responses elicited by epitope DNA vaccine

    Institute of Scientific and Technical Information of China (English)

    Wang QM; Sun SH; Hu ZL; Zhou FJ; Yin M; Xiao CJ; Zhang JC

    2005-01-01

    Cell-mediated immune responses are crucial in the protection against tuberculosis. In this study, we constructed epitope DNA vaccines (p3-M-38) encoding cytotoxic T lymphocyte (CTL) epitopes of MPT64 and 38 kDa proteins of Mycobacterium tuberculosis. In order to observe the influence of spacer sequence (Ala-Ala-Tyr) or ubiquitin (UbGR) on the efficacy of the two CTL epitopes, we also constructed DNA vaccines, p3-M-S(spacer)-38, p3-Ub (UbGR)-M-S-38 and p3-Ub-M-38. The immune responses elicited by the four DNA vaccines were tested in C57BL/6 (H-2b) mice. The cytotoxicity of T cells was detected by LDH-release method and by enzyme-linked immunospot assay for epitope-specific cells secreting interferon-gamma. The results showed that DNA immunization with p3-M-38 vaccine could induce epitope-specific CD8+ CTL response and that the spacer sequence (AAY) only enhanced M epitope presentation. The protein-targeting sequence (UbGR) enhanced the immunogenicity of the two epitopes. The finding that defined spacer sequences at C-terminus and protein-targeting degradation modulated the immune response of epitope string DNA vaccines will be of importance for the further development of multi-epitope DNA vaccines against tuberculosis.

  14. DNA vaccines, electroporation and their applications in cancer treatment.

    Science.gov (United States)

    Lee, Si-Hyeong; Danishmalik, Sayyed Nilofar; Sin, Jeong-Im

    2015-01-01

    Numerous animal studies and recent clinical studies have shown that electroporation-delivered DNA vaccines can elicit robust Ag-specific CTL responses and reduce disease severity. However, cancer antigens are generally poorly immunogenic, requiring special conditions for immune response induction. To date, many different approaches have been used to elicit Ag-specific CTL and anti-neoplastic responses to DNA vaccines against cancer. In vivo electroporation is one example, whereas others include DNA manipulation, xenogeneic antigen use, immune stimulatory molecule and immune response regulator application, DNA prime-boost immunization strategy use and different DNA delivery methods. These strategies likely increase the immunogenicity of cancer DNA vaccines, thereby contributing to cancer eradication. However, cancer cells are heterogeneous and might become CTL-resistant. Thus, understanding the CTL resistance mechanism(s) employed by cancer cells is critical to develop counter-measures for this immune escape. In this review, the use of electroporation as a DNA delivery method, the strategies used to enhance the immune responses, the cancer antigens that have been tested, and the escape mechanism(s) used by tumor cells are discussed, with a focus on the progress of clinical trials using cancer DNA vaccines.

  15. Potent T cell Responses Induced by Single DNA Vaccine Boosted with Recombinant Vaccinia Vaccine

    Institute of Scientific and Technical Information of China (English)

    Lianxing Liu; Chao Qiu; Yang Huang; Jianqing Xu; Yiming Shao

    2013-01-01

    Plasmid DNA,an effective vaccine vector,can induce both cellular and humoral immune responses.However,plasmid DNA raises issues concerning potential genomic integration after injection.This issue should be considered in preclinical studies.Tiantan vaccinia virus (TV) has been most widely utilized in eradicating smallpox in China.This virus has also been considered as a successful vaccine vector against a few infectious diseases.Potent T cell responses through T-cell receptor (TCR) could be induced by three injections of the DNA prime vaccine followed by a single injection of recombinant vaccinia vaccine.To develop a safer immunization strategy,a single DNA prime followed by a single recombinant Tiantan vaccinia (rTV) AIDS vaccine was used to immunize mice.Our data demonstrated that one DNA prime/rTV boost regimen induced mature TCR activation with high functional avidity,preferential T cell Vβ receptor usage and high sensitivity to anti-CD3 antibody stimulation.No differences in T cell responses were observed among one,two or three DNA prime/rTV boost regimens.This study shows that one DNA prime/rTV boost regimen is sufficient to induce potent T cell responses against HIV.

  16. Nanoconfined circular and linear DNA - equilibrium conformations and unfolding kinetics

    CERN Document Server

    Alizadehheidari, M; Noble, C; Reiter-Schad, M; Nyberg, L K; Fritzsche, J; Mehlig, B; Tegenfeldt, J O; Ambjörnsson, T; Persson, F; Westerlund, F

    2016-01-01

    Studies of circular DNA confined to nanofluidic channels are relevant both from a fundamental polymer-physics perspective and due to the importance of circular DNA molecules in vivo. We here observe the unfolding of DNA from the circular to linear configuration as a light-induced double strand break occurs, characterize the dynamics, and compare the equilibrium conformational statistics of linear and circular configurations. This is important because it allows us to determine to which extent existing statistical theories describe the extension of confined circular DNA. We find that the ratio of the extensions of confined linear and circular DNA configurations increases as the buffer concentration decreases. The experimental results fall between theoretical predictions for the extended de Gennes regime at weaker confinement and the Odijk regime at stronger confinement. We show that it is possible to directly distinguish between circular and linear DNA molecules by measuring the emission intensity from the DNA....

  17. Successive site translocating inoculation potentiates DNA/recombinant vaccinia vaccination.

    Science.gov (United States)

    Ren, Yanqin; Wang, Na; Hu, Weiguo; Zhang, Xiaoyan; Xu, Jianqing; Wan, Yanmin

    2015-12-15

    DNA vaccines have advantages over traditional vaccine modalities; however the relatively low immunogenicity restrains its translation into clinical use. Further optimizations are needed to get the immunogenicity of DNA vaccine closer to the level required for human use. Here we show that intramuscularly inoculating into a different limb each time significantly improves the immunogenicities of both DNA and recombinant vaccinia vaccines during multiple vaccinations, compared to repeated vaccination on the same limb. We term this strategy successive site translocating inoculation (SSTI). SSTI could work in synergy with genetic adjuvant and DNA prime-recombinant vaccinia boost regimen. By comparing in vivo antigen expression, we found that SSTI avoided the specific inhibition of in vivo antigen expression, which was observed in the limbs being repeatedly inoculated. Employing in vivo T cell depletion and passive IgG transfer, we delineated that the inhibition was not mediated by CD8(+) T cells but by specific antibodies. Finally, by using C3(-/-) mouse model and in vivo NK cells depletion, we identified that specific antibodies negatively regulated the in vivo antigen expression primarily in a complement depended way.

  18. Next generation sequencing of DNA-launched Chikungunya vaccine virus

    Energy Technology Data Exchange (ETDEWEB)

    Hidajat, Rachmat; Nickols, Brian [Medigen, Inc., 8420 Gas House Pike, Suite S, Frederick, MD 21701 (United States); Forrester, Naomi [Institute for Human Infections and Immunity, Sealy Center for Vaccine Development and Department of Pathology, University of Texas Medical Branch, GNL, 301 University Blvd., Galveston, TX 77555 (United States); Tretyakova, Irina [Medigen, Inc., 8420 Gas House Pike, Suite S, Frederick, MD 21701 (United States); Weaver, Scott [Institute for Human Infections and Immunity, Sealy Center for Vaccine Development and Department of Pathology, University of Texas Medical Branch, GNL, 301 University Blvd., Galveston, TX 77555 (United States); Pushko, Peter, E-mail: ppushko@medigen-usa.com [Medigen, Inc., 8420 Gas House Pike, Suite S, Frederick, MD 21701 (United States)

    2016-03-15

    Chikungunya virus (CHIKV) represents a pandemic threat with no approved vaccine available. Recently, we described a novel vaccination strategy based on iDNA® infectious clone designed to launch a live-attenuated CHIKV vaccine from plasmid DNA in vitro or in vivo. As a proof of concept, we prepared iDNA plasmid pCHIKV-7 encoding the full-length cDNA of the 181/25 vaccine. The DNA-launched CHIKV-7 virus was prepared and compared to the 181/25 virus. Illumina HiSeq2000 sequencing revealed that with the exception of the 3′ untranslated region, CHIKV-7 viral RNA consistently showed a lower frequency of single-nucleotide polymorphisms than the 181/25 RNA including at the E2-12 and E2-82 residues previously identified as attenuating mutations. In the CHIKV-7, frequencies of reversions at E2-12 and E2-82 were 0.064% and 0.086%, while in the 181/25, frequencies were 0.179% and 0.133%, respectively. We conclude that the DNA-launched virus has a reduced probability of reversion mutations, thereby enhancing vaccine safety. - Highlights: • Chikungunya virus (CHIKV) is an emerging pandemic threat. • In vivo DNA-launched attenuated CHIKV is a novel vaccine technology. • DNA-launched virus was sequenced using HiSeq2000 and compared to the 181/25 virus. • DNA-launched virus has lower frequency of SNPs at E2-12 and E2-82 attenuation loci.

  19. Ebola Vaccination Using a DNA Vaccine Coated on PLGA-PLL/γPGA Nanoparticles Administered Using a Microneedle Patch.

    Science.gov (United States)

    Yang, Hung-Wei; Ye, Ling; Guo, Xin Dong; Yang, Chinglai; Compans, Richard W; Prausnitz, Mark R

    2017-01-01

    Ebola DNA vaccine is incorporated into PLGA-PLL/γPGA nanoparticles and administered to skin using a microneedle (MN) patch. The nanoparticle delivery system increases vaccine thermostability and immunogenicity compared to free vaccine. Vaccination by MN patch produces stronger immune responses than intramuscular administration. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  20. DNA addition using linear self-assembly

    Institute of Scientific and Technical Information of China (English)

    ZHAO Jian; QIAN LuLu; LIU Qiang; ZHANG ZhiZhou; HE Lin

    2007-01-01

    This paper presents a DNA algorithm which adds two nonnegative binary integers using self-assembly in constant steps. The approach has the benefit of greater experimental simplicity when compared with previous DNA addition algorithms. For the addition of two binary n-bit integers, O(n) is different from DNA strands and only O(1) biochemical experimental procedures are required.

  1. Cationic influenza virosomes as an adjuvanted delivery system for CTL induction by DNA vaccination

    NARCIS (Netherlands)

    Jamali, Abbas; Holtrop, Marijke; de Haan, Aalzen; Hashemi, Hamidreza; Shenagari, Mohammad; Memarnejadian, Arash; Roohvand, Farzin; Sabahi, Farzaneh; Kheiri, Masumeh Tavassoti; Huckriede, Anke

    2012-01-01

    DNA vaccines have emerged as an attractive approach to induce CTL responses against cancer and infectious agents in recent years. Although CTL induction by DNA vaccination would be a valuable strategy for controlling viral infections, increasing the potency of DNA vaccines is mandatory before DNA

  2. Overview of recent DNA vaccine development for fish

    Science.gov (United States)

    Kurath, G.; ,

    2005-01-01

    Since the first description of DNA vaccines for fish in 1996, numerous studies of genetic immunisation against the rhabdovirus pathogens infectious haematopoietic necrosis virus (IHNV) and viral haemorrhagic septicaemia virus (VHSV) have established their potential as both highly efficacious biologicals and useful basic research tools. Single small doses of rhabdovirus DNA constructs provide extremely strong protection against severe viral challenge under a variety of conditions. DNA vaccines for several other important fish viruses, bacteria, and parasites are under investigation, but they have not yet shown high efficacy. Therefore, current research is focussed on mechanistic studies to understand the basis of protection, and on improvement of the nucleic acid vaccine applications against a wider range of fish pathogens.

  3. DNA vaccine prime and recombinant FPV vaccine boost: an important candidate immunization strategy to control bluetongue virus type 1.

    Science.gov (United States)

    Li, Junping; Yang, Tao; Xu, Qingyuan; Sun, Encheng; Feng, Yufei; Lv, Shuang; Zhang, Qin; Wang, Haixiu; Wu, Donglai

    2015-10-01

    Bluetongue virus (BTV) is the causative agent of bluetongue (BT), an important sheep disease that caused great economic loss to the sheep industry. There are 26 BTV serotypes based on the outer protein VP2. However, the serotypes BTV-1 and BTV-16 are the two most prevalent serotypes in China. Vaccination is the most effective method of preventing viral infections. Therefore, the need for an effective vaccine against BTV is urgent. In this study, DNA vaccines and recombinant fowlpox virus (rFPV) vaccines expressing VP2 alone or VP2 in combination with VP5 or co-expressing the VP2 and VP5 proteins of BTV-1 were evaluated in both mice and sheep. Several strategies were tested in mice, including DNA vaccine prime and boost, rFPV vaccine prime and boost, and DNA vaccine prime and rFPV vaccine boost. We then determined the best vaccine strategy in sheep. Our results indicated that a strategy combining a DNA vaccine prime (co-expressing VP2 and VP5) followed by an rFPV vaccine boost (co-expressing VP2 and VP5) induced a high titer of neutralizing antibodies in sheep. Therefore, our data suggest that a DNA vaccine consisting of a pCAG-(VP2+VP5) prime and an rFPV-(VP2+VP5) boost is an important candidate for the design of a novel vaccine against BTV-1.

  4. A DNA vaccine against yellow fever virus: development and evaluation.

    Science.gov (United States)

    Maciel, Milton; Cruz, Fábia da Silva Pereira; Cordeiro, Marli Tenório; da Motta, Márcia Archer; Cassemiro, Klécia Marília Soares de Melo; Maia, Rita de Cássia Carvalho; de Figueiredo, Regina Célia Bressan Queiroz; Galler, Ricardo; Freire, Marcos da Silva; August, Joseph Thomas; Marques, Ernesto T A; Dhalia, Rafael

    2015-04-01

    Attenuated yellow fever (YF) virus 17D/17DD vaccines are the only available protection from YF infection, which remains a significant source of morbidity and mortality in the tropical areas of the world. The attenuated YF virus vaccine, which is used worldwide, generates both long-lasting neutralizing antibodies and strong T-cell responses. However, on rare occasions, this vaccine has toxic side effects that can be fatal. This study presents the design of two non-viral DNA-based antigen formulations and the characterization of their expression and immunological properties. The two antigen formulations consist of DNA encoding the full-length envelope protein (p/YFE) or the full-length envelope protein fused to the lysosomal-associated membrane protein signal, LAMP-1 (pL/YFE), aimed at diverting antigen processing/presentation through the major histocompatibility complex II precursor compartments. The immune responses triggered by these formulations were evaluated in H2b and H2d backgrounds, corresponding to the C57Bl/6 and BALB/c mice strains, respectively. Both DNA constructs were able to induce very strong T-cell responses of similar magnitude against almost all epitopes that are also generated by the YF 17DD vaccine. The pL/YFE formulation performed best overall. In addition to the T-cell response, it was also able to stimulate high titers of anti-YF neutralizing antibodies comparable to the levels elicited by the 17DD vaccine. More importantly, the pL/YFE vaccine conferred 100% protection against the YF virus in intracerebrally challenged mice. These results indicate that pL/YFE DNA is an excellent vaccine candidate and should be considered for further developmental studies.

  5. A DNA vaccine against yellow fever virus: development and evaluation.

    Directory of Open Access Journals (Sweden)

    Milton Maciel

    2015-04-01

    Full Text Available Attenuated yellow fever (YF virus 17D/17DD vaccines are the only available protection from YF infection, which remains a significant source of morbidity and mortality in the tropical areas of the world. The attenuated YF virus vaccine, which is used worldwide, generates both long-lasting neutralizing antibodies and strong T-cell responses. However, on rare occasions, this vaccine has toxic side effects that can be fatal. This study presents the design of two non-viral DNA-based antigen formulations and the characterization of their expression and immunological properties. The two antigen formulations consist of DNA encoding the full-length envelope protein (p/YFE or the full-length envelope protein fused to the lysosomal-associated membrane protein signal, LAMP-1 (pL/YFE, aimed at diverting antigen processing/presentation through the major histocompatibility complex II precursor compartments. The immune responses triggered by these formulations were evaluated in H2b and H2d backgrounds, corresponding to the C57Bl/6 and BALB/c mice strains, respectively. Both DNA constructs were able to induce very strong T-cell responses of similar magnitude against almost all epitopes that are also generated by the YF 17DD vaccine. The pL/YFE formulation performed best overall. In addition to the T-cell response, it was also able to stimulate high titers of anti-YF neutralizing antibodies comparable to the levels elicited by the 17DD vaccine. More importantly, the pL/YFE vaccine conferred 100% protection against the YF virus in intracerebrally challenged mice. These results indicate that pL/YFE DNA is an excellent vaccine candidate and should be considered for further developmental studies.

  6. Immunotherapy of Trypanosoma cruzi infection with DNA vaccines in mice.

    Science.gov (United States)

    Dumonteil, Eric; Escobedo-Ortegon, Javier; Reyes-Rodriguez, Norma; Arjona-Torres, Arletty; Ramirez-Sierra, Maria Jesus

    2004-01-01

    The mechanisms involved in the pathology of chronic chagasic cardiomyopathy are still debated, and the controversy has interfered with the development of new treatments and vaccines. Because of the potential of DNA vaccines for immunotherapy of chronic and infectious diseases, we tested if DNA vaccines could control an ongoing Trypanosoma cruzi infection. BALB/c mice were infected with a lethal dose (5 x 10(4) parasites) as a model of acute infection, and then they were treated with two injections of 100 microg of plasmid DNA 1 week apart, beginning on day 5 postinfection. Control mice had high levels of parasitemia and mortality and severe cardiac inflammation, while mice treated with plasmid DNA encoding trypomastigote surface antigen 1 or Tc24 had reduced parasitemia and mild cardiac inflammation and >70% survived the infection. The efficacy of the immunotherapy also was significant when it was delayed until days 10 and 15 after infection. Parasitological analysis of cardiac tissue of surviving mice indicated that most mice still contained detectable parasite kinetoplast DNA but fewer mice contained live parasites, suggesting that there was efficient but not complete parasite elimination. DNA vaccine immunotherapy was also evaluated in CD1 mice infected with a low dose (5 x 10(2) parasites) as a model of chronic infection. Immunotherapy was initiated on day 70 postinfection and resulted in improved survival and reduced cardiac tissue inflammation. These results suggest that DNA vaccines have strong potential for the immunotherapy of T. cruzi infection and may provide new alternatives for the control of Chagas' disease.

  7. Structural changes of linear DNA molecules induced by cisplatin

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Zhiguo, E-mail: cn.zguoliu@yahoo.com [State Engineering Laboratory of Bio-Resource Eco-Utilization, Harbin 150040 (China); Engineering Research Center of Forest Bio-preparation, Ministry of Education, Northeast Forestry University, Harbin 150040 (China); Key Laboratory of Forest Plant Ecology of Ministry of Education, Northeast Forestry University, Harbin 150040 (China); Liu, Ruisi; Zhou, Zhen; Zu, Yuangang; Xu, Fengjie [State Engineering Laboratory of Bio-Resource Eco-Utilization, Harbin 150040 (China); Engineering Research Center of Forest Bio-preparation, Ministry of Education, Northeast Forestry University, Harbin 150040 (China); Key Laboratory of Forest Plant Ecology of Ministry of Education, Northeast Forestry University, Harbin 150040 (China)

    2015-02-20

    Interaction between long DNA molecules and activated cisplatin is believed to be crucial to anticancer activity. However, the exact structural changes of long DNA molecules induced by cisplatin are still not very clear. In this study, structural changes of long linear double-stranded DNA (dsDNA) and short single-stranded DNA (ssDNA) induced by activated cisplatin have been investigated by atomic force microscopy (AFM). The results indicated that long DNA molecules gradually formed network structures, beads-on-string structures and their large aggregates. Electrostatic and coordination interactions were considered as the main driving forces producing these novel structures. An interesting finding in this study is the beads-on-string structures. Moreover, it is worth noting that the beads-on-string structures were linked into the networks, which can be ascribed to the strong DNA–DNA interactions. This study expands our knowledge of the interactions between DNA molecules and cisplatin. - Highlights: • We investigate structural changes of dsDNA and ssDNA induced by cisplatin. • AFM results indicated long dsDNA formed network, beads-on-string and aggregates. • ssDNA can form very similar structures as those of long linear dsDNA. • A possible formation process of theses novel structure is proposed.

  8. Effect of Cisplatin on the Flexibility of Linear DNA

    Institute of Scientific and Technical Information of China (English)

    JI Chao; ZHANG Ling-Yun; HOU Xi-Miao; DOU Shuo-Xing; WANG Peng-Ye

    2011-01-01

    With the aid of an atomic force microscope (AFM), we study the interaction between linear DNA fragment and cisplatin. For different cisplatin concentrations, the AFM used to observe the conformation of DNA has a gradual change. The contour length, the end-to-end distance and the local bend angles of the linear DNA fragment can be accurately measured. The persistence length of DNA interacting with cisplatin is decreased with the increasing cisplatin concentration. Furthermore, it is demonstrated that the local bend angles of DNA chains are increased by the binding interaction of cisplatin.%@@ With the aid of an atomic force microscope (AFM), we study the interaction between linear DNA fragment and cisplatin.For different cisplatin concentrations, the AFM used to observe the conformation of DNA has a gradual change.The contour length, the end-to-end distance and the local bend angles of the linear DNA fragment can be accurately measured.The persistence length of DNA interacting with cisplatin is decreased with the increasing cisplatin concentration.Furthermore, it is demonstrated that the local bend angles of DNA chains are increased by the binding interaction of cisplatin.

  9. Functional demonstration of adaptive immunity in zebrafish using DNA vaccination

    DEFF Research Database (Denmark)

    Lorenzen, Niels; Lorenzen, Ellen; Einer-Jensen, Katja

    studies have documented existence of a classical innate immune response, there is mainly indirect evidence of functional adaptive immunity. To address this aspect, groups of zebrafish were vaccinated with DNA-vaccines against the rhabdoviruses VHSV, IHNV and SVCV. Seven weeks later, the fish were...... challenged with SVCV by immersion. Despite some variability between replicate aquaria, there was a protective effect of the homologous vaccine and no effect of the heterologous vaccines. The results therefore confirm the existence of not only a well developed but also a fully functional adaptive immune......Due to the well characterized genome, overall highly synteny with the human genome and its suitability for functional genomics studies, the zebrafish is considered to be an ideal animal model for basic studies of mechanisms of diseases and immunity in vertebrates including humans. While several...

  10. Prophylactic and therapeutic DNA vaccines against Chagas disease.

    Science.gov (United States)

    Arce-Fonseca, Minerva; Rios-Castro, Martha; Carrillo-Sánchez, Silvia del Carmen; Martínez-Cruz, Mariana; Rodríguez-Morales, Olivia

    2015-01-01

    Chagas disease is a zoonosis caused by Trypanosoma cruzi in which the most affected organ is the heart. Conventional chemotherapy has a very low effectiveness; despite recent efforts, there is currently no better or more effective treatment available. DNA vaccines provide a new alternative for both prevention and treatment of a variety of infectious disorders, including Chagas disease. Recombinant DNA technology has allowed some vaccines to be developed using recombinant proteins or virus-like particles capable of inducing both a humoral and cellular specific immune response. This type of immunization has been successfully used in preclinical studies and there are diverse models for viral, bacterial and/or parasitic diseases, allergies, tumors and other diseases. Therefore, several research groups have been given the task of designing a DNA vaccine against experimental infection with T. cruzi. In this review we explain what DNA vaccines are and the most recent studies that have been done to develop them with prophylactic or therapeutic purposes against Chagas disease.

  11. Optimised electroporation mediated DNA vaccination for treatment of prostate cancer.

    LENUS (Irish Health Repository)

    Ahmad, Sarfraz

    2010-01-01

    ABSTRACT: BACKGROUND: Immunological therapies enhance the ability of the immune system to recognise and destroy cancer cells via selective killing mechanisms. DNA vaccines have potential to activate the immune system against specific antigens, with accompanying potent immunological adjuvant effects from unmethylated CpG motifs as on prokaryotic DNA. We investigated an electroporation driven plasmid DNA vaccination strategy in animal models for treatment of prostate cancer. METHODS: Plasmid expressing human PSA gene (phPSA) was delivered in vivo by intra-muscular electroporation, to induce effective anti-tumour immune responses against prostate antigen expressing tumours. Groups of male C57 BL\\/6 mice received intra-muscular injections of phPSA plasmid. For phPSA delivery, quadriceps muscle was injected with 50 mug plasmid. After 80 seconds, square-wave pulses were administered in sequence using a custom designed pulse generator and acustom-designed applicator with 2 needles placed through the skin central to the muscle. To determine an optimum treatment regimen, three different vaccination schedules were investigated. In a separate experiment, the immune potential of the phPSA vaccine was further enhanced with co- administration of synthetic CpG rich oligonucleotides. One week after last vaccination, the mice were challenged subcutaneously with TRAMPC1\\/hPSA (prostate cancer cell line stably expressing human PSA) and tumour growth was monitored. Serum from animals was examined by ELISA for anti-hPSA antibodies and for IFNgamma. Histological assessment of the tumours was also carried out. In vivo and in vitro cytotoxicity assays were performed with splenocytes from treated mice. RESULTS: The phPSA vaccine therapy significantly delayed the appearance of tumours and resulted in prolonged survival of the animals. Four-dose vaccination regimen provided optimal immunological effects. Co - administration of the synthetic CpG with phPSA increased anti-tumour responses

  12. Programmable multimetallic linear nanoassemblies of ruthenium-DNA conjugates

    OpenAIRE

    Irvoas, Joris; Noirot, Arielle; Chouini-Lalanne, Nadia; Reynes, Olivier; Garrigues, Jean-Christophe; Sartor, Valérie

    2012-01-01

    International audience; A new ruthenium-DNA conjugates family was synthesized, made up of a ruthenium complex bound to one or two identical DNA strands of 14-58 nucleotides. The formation of controlled linear nanoassemblies containing one to seven ruthenium complexes is described.

  13. Survivin minigene DNA vaccination is effective against neuroblastoma.

    Science.gov (United States)

    Fest, Stefan; Huebener, Nicole; Bleeke, Matthias; Durmus, Tahir; Stermann, Alexander; Woehler, Anja; Baykan, Bianca; Zenclussen, Ana C; Michalsky, Elke; Jaeger, Ines S; Preissner, Robert; Hohn, Oliver; Weixler, Silke; Gaedicke, Gerhard; Lode, Holger N

    2009-07-01

    The inhibitor of apoptosis protein survivin is highly expressed in neuroblastoma (NB) and survivin-specific T cells were identified in Stage 4 patients. Therefore, we generated a novel survivin minigene DNA vaccine (pUS-high) encoding exclusively for survivin-derived peptides with superior MHC class I (H2-K(k)) binding affinities and tested its efficacy to suppress tumor growth and metastases in a syngeneic NB mouse model. Vaccination was performed by oral gavage of attenuated Salmonella typhimurium SL7207 carrying pUS-high. Mice receiving the pUS-high in the prophylactic setting presented a 48-52% reduction in s.c. tumor volume, weight and liver metastasis level in contrast to empty vector controls. This response was as effective as a survivin full-length vaccine and was associated with an increased target cell lysis, increased presence of CD8(+) T-cells at the primary tumor site and enhanced production of proinflammatory cytokines by systemic CD8(+) T cells. Furthermore, depletion of CD8(+) but not CD4(+) T-cells completely abrogated the pUS-high mediated primary tumor growth suppression, demonstrating a CD8(+) T-cell mediated effect. Therapeutic vaccination with pUS-high led to complete NB eradication in over 50% of immunized mice and surviving mice showed an over 80% reduction in primary tumor growth upon rechallenge in contrast to controls. In summary, survivin-based DNA vaccination is effective against NB and the rational minigene design provides a promising approach to circumvent potentially hazardous effects of using full length antiapoptotic genes as DNA vaccines.

  14. Tailoring DNA vaccines: designing strategies against HER2 positive cancers

    Directory of Open Access Journals (Sweden)

    Cristina eMarchini

    2013-05-01

    Full Text Available The crucial role of HER2 in epithelial transformation and its selective overexpression on cancer tissues makes it an ideal target for cancer immunotherapies such as passive immunotherapy with Trastuzumab. There are, however, a number of concerns regarding the use of monoclonal antibodies which include resistance, repeated treatments, considerable costs and side effects that make active immunotherapies against HER2 desirable alternative approaches. The efficacy of anti-HER2 DNA vaccination has been widely demonstrated in transgenic cancer-prone mice, which recapitulate several features of human breast cancers. Nonetheless, the rational design of a cancer vaccine able to trigger a long lasting immunity, and thus prevent tumor recurrence in patients, would require the understanding of how tolerance and immunosuppression regulate antitumor immune responses and, at the same time, the identification of the most immunogenic portions of the target protein. We herein retrace the findings that led to our most promising DNA vaccines that, by encoding human/rat chimeric forms of HER2, are able to circumvent peripheral tolerance. Preclinical data obtained with these chimeric DNA vaccines have provided the rationale for their use in an ongoing phase I clinical trial (EudraCT 2011-001104-34.

  15. Immunogenicity of DNA and Recombinant Sendai Virus Vaccines Expressing the HIV-1 gag Gene

    Institute of Scientific and Technical Information of China (English)

    Xia FENG; Shuang-qing YU; Tsugumine Shu; Tetsuro Matano; Mamoru Hasegawa; Xiao-li WANG; Hong-tao MA; Hong-xia LI; Yi ZENG

    2008-01-01

    Combinations of DNA and recombinant-viral-vector based vaccines are promising AIDS vaccine methods because of their potential for inducing cellular immune responses. It was found that Gag-specific cytotoxic lymphocyte (CTL) responses were associated with lowering viremia in an untreated HIV-1 infected cohort. The main objectives of our studies were the construction of DNA and recombinant Sendal virus vector (rSeV) vaccines containing a gag gene from the prevalent Thailand subtype B strain in China and trying to use these vaccines for therapeutic and prophylactic vaccines. The candidate plasmid DNA vaccine pcDNA3.1(+)-gag and recombinant Sendai virus vaccine (rSeV-gag) were constructed separately. It was verified by Western blotting analysis that both DNA and rSeV-gag vaccines expressed the HIV-1 Gag protein correctly and efficiently. Balb/c mice were immunized with these two vaccines in different administration schemes. HIV-1 Gag-specific CTL responses and antibody levels were detected by intracellular cytokine staining assay and enzyme-linked immunosorbant assay (ELISA) respectively. Combined vaccines in a DNA prime/rSeV-gag boost vaccination regimen induced the strongest and most long-lasting Gag-specific CTL and antibody responses. It maintained relatively high levels even 9 weeks post immunization. This data indicated that the prime-boost regimen with DNA and rSeV-gag vaccines may offer promising HIV vaccine regimens.

  16. Nanobiotechnological approaches to delivery of DNA vaccine against fungal infection.

    Science.gov (United States)

    Ribeiro, A M; Souza, A C O; Amaral, A C; Vasconcelos, N M; Jeronimo, M S; Carneiro, F P; Faccioli, L H; Felipe, M S S; Silva, C L; Bocca, A L

    2013-02-01

    Vaccines play an essential role in keeping humans healthy. Innovative approaches to their use include the utilization of plasmid DNA encoding sequences to express foreign antigens. DNAhsp65 from Mycobacterium leprae is suitable for this purpose due to its ability to elicit a powerful immune response. Controlled release systems represent a promising approach to delivering vaccines. In this work, we used liposomes or PLGA systems to deliver DNAhsp65 to treat the pulmonary fungal infection Paracoccidioidomycosis. Both formulations modulated a protective immune response and reduced the pulmonary fungal burden even in the groups receiving less than four times the amount of the DNAhps65 entrapped within the nanoparticles. Although both systems had the same effective therapeutic results, the advantage of the liposome formulation was that it was administered intranasally, which may be more easily accepted by patients. These systems are a great alternative to be considered as adjuvant vaccine therapy for systemic mycosis.

  17. Immunization against Egyptian Schistosoma mansoni infection by multivalent DNA vaccine

    Institute of Scientific and Technical Information of China (English)

    Mahmoud H Romeih; Hanem M Hassan; Tarek S Abou Shousha; Mohamed A Saber

    2008-01-01

    The development of multivalent vaccines consisting of several antigens is a novel approach to creating broad-range protection against different parasite strains and parasite life cycle stages. We have previously confirmed that the schistosome Sm21.7 and SmFimbrin (SmFim) proteins could induce protection in mice. Therefore, this study aimed to construct the multivalent DNA vaccine Sm21.7-SmFim/pBudCE4.1 and evaluate its immune efficacy. The open reading frames of two Schistosoma mansoni genes, Sm21.7 and SmFim, were inserted into the eukaryotic expression plasmid pBudCE4.1 designed for the independent expression of two genes in mammalian cells. To evaluate the in vitro expression of the multivalent Sm21.7-SmFim/pBudCE4.1 DNA vaccine and its immunological effect in mice, the recombinant plasmid Sm21.7-SmFim/pBudCE4.1 was used to transfect 293T cells, and the expression of mRNA and proteins was examined using reverse transcription-polymerase chain reaction and Western blot analysis. Then the ability of Sm21.7.SmFim/pBudCE4.1 to protect against S. mansoni challenge infections was analyzed according to worm burden and egg reduction rates after vaccination of mice. Vaccinated mice showed a significant level of protection (56%), and a decrease in the number and size, and change in the cellular profile, of granulomas. Egg reduction in liver and intestine was 41.53% and 55.63%,respectively, as determined relative to mice that received the empty vector only. In addition to reductions in worm viability,worm fecundity and egg hatching ability were observed following challenge infection in the immunized group.Results showed that Sm21.7-SmFim/pBudCE4.1 could express Sm 21.7 and SmFim mRN A and proteins. Enzyme-linked immunosorbent assay and Western blot analysis indicated that immunized mice generated specific immunoglobulin G against Sm21.7-SmFim/pBudCE4.1. These results suggest that vaccination with multivalent S. mansoni DNA vaccine (SmFim-Sm21.7/pBudCE4.1) not only induces a

  18. Transposon leads to contamination of clinical pDNA vaccine.

    Science.gov (United States)

    van der Heijden, I; Gomez-Eerland, R; van den Berg, J H; Oosterhuis, K; Schumacher, T N; Haanen, J B A G; Beijnen, J H; Nuijen, B

    2013-07-11

    We report an unexpected contamination during clinical manufacture of a Human Papilomavirus (HPV) 16 E6 encoding plasmid DNA (pDNA) vaccine, with a transposon originating from the Escherichia coli DH5 host cell genome. During processing, presence of this transposable element, insertion sequence 2 (IS2) in the plasmid vector was not noticed until quality control of the bulk pDNA vaccine when results of restriction digestion, sequencing, and CGE analysis were clearly indicative for the presence of a contaminant. Due to the very low level of contamination, only an insert-specific PCR method was capable of tracing back the presence of the transposon in the source pDNA and master cell bank (MCB). Based on the presence of an uncontrolled contamination with unknown clinical relevance, the product was rejected for clinical use. In order to prevent costly rejection of clinical material, both in-process controls and quality control methods must be sensitive enough to detect such a contamination as early as possible, i.e. preferably during plasmid DNA source generation, MCB production and ultimately during upstream processing. However, as we have shown that contamination early in the process development pipeline (source pDNA, MCB) can be present below limits of detection of generally applied analytical methods, the introduction of "engineered" or transposon-free host cells seems the only 100% effective solution to avoid contamination with movable elements and should be considered when searching for a suitable host cell-vector combination.

  19. Development of DNA vaccines for fish

    DEFF Research Database (Denmark)

    Heppell, Joël; Lorenzen, Niels; Armstrong, Neil K.;

    1998-01-01

    with traditional methods of immunization, but little is known on its efficacy in fish. The luciferase and lacZ reporter genes were used to characterize expression of plasmid-encoded genes in rainbow trout and zebra fish injected intramuscularly. For a given dose of DNA, the luciferase activity was higher in fish...

  20. Immunogenicity of a DNA-launched replicon-based canine parvovirus DNA vaccine expressing VP2 antigen in dogs.

    Science.gov (United States)

    Dahiya, Shyam S; Saini, Mohini; Kumar, Pankaj; Gupta, Praveen K

    2012-10-01

    A replicon-based DNA vaccine encoding VP2 gene of canine parvovirus (CPV) was developed by cloning CPV-VP2 gene into a replicon-based DNA vaccine vector (pAlpha). The characteristics of a replicon-based DNA vaccine like, self-amplification of transcripts and induction of apoptosis were analyzed in transfected mammalian cells. When the pAlpha-CPV-VP2 was injected intradermal as DNA-launched replicon-based DNA vaccine in dogs, it induced CPV-specific humoral and cell mediated immune responses. The virus neutralization antibody and lymphocyte proliferative responses were higher than conventional CPV DNA vaccine and commercial CPV vaccine. These results indicated that DNA-launched replicon-based CPV DNA vaccine was effective in inducing both CPV-specific humoral and cellular immune responses and can be considered as effective alternative to conventional CPV DNA vaccine and commercial CPV vaccine. Crown Copyright © 2012. Published by Elsevier India Pvt Ltd. All rights reserved.

  1. Synthetic DNA approach to cytomegalovirus vaccine/immune therapy.

    Science.gov (United States)

    Wu, Stephan J; Villarreal, Daniel O; Shedlock, Devon J; Weiner, David B

    2015-01-01

    There is no licensed vaccine or cure for human cytomegalovirus (CMV), a ubiquitous β-herpes virus that infects 60-95 % of adults worldwide. Infection is a major cause of congenital abnormalities in newborns, contributes to development of childhood cerebral palsy and medulloblastoma, can result in severe disease in immunocompromised patients, and is a major impediment during successful organ transplantation. While CMV has been increasingly associated with numerous inflammatory diseases and cancers, only recently has it been correlated with increased risk of heart disease in adults, the number-one killer in the USA. These data, among others, suggest that subclinical CMV infection, or microinfection, in healthy individuals may play more of a causative role than an epiphenomenon in development of CMV-associated pathologies. Due to the myriad of diseases and complications associated with CMV, an efficacious vaccine would be highly valuable in reducing human morbidity and mortality as well as saving billions of dollars in annual health-care costs and disability adjusted life years (DALY) in the developing world. Therefore, the development of a safe efficacious CMV vaccine or immune therapy is paramount to the public health. This review aims to provide a brief overview on aspects of CMV infection and disease and focuses on current vaccine strategies. The use of new synthetic DNA vaccines might offer one such approach to this difficult problem.

  2. Preventive and therapeutic DNA vaccination partially protect dogs against an infectious challenge with Trypanosoma cruzi.

    Science.gov (United States)

    Quijano-Hernández, Israel A; Castro-Barcena, Alejandro; Vázquez-Chagoyán, Juan C; Bolio-González, Manuel E; Ortega-López, Jaime; Dumonteil, Eric

    2013-04-26

    American trypanosomiasis, or Chagas disease, is caused by Trypanosoma cruzi, and a vaccine would greatly improve disease control. While some studies in mice suggest that a vaccine is feasible, limited efficacy has been observed in dogs. We evaluated here the safety and efficacy of a DNA vaccine encoding TSA-1 and Tc24 antigens in a dog model of acute T. cruzi infection. Mongrel dogs were immunized with two doses of 500 μg of DNA vaccine, two weeks apart, and infected with T. cruzi (SylvioX10/4 strain) two weeks after the second vaccine dose. Another group of dogs was infected first and treated with the vaccine. Disease progression was monitored for up to 70 days post-infection. The vaccine did not induce any critical change in blood parameters, nor exacerbation of disease in vaccinated animals. On the contrary, it prevented anemia and a decrease in lymphocyte counts following T. cruzi infection in vaccinated dogs. Both preventive and therapeutic vaccination significantly reduced parasitemia, cardiac inflammation and cardiac parasite burden, and tended to reduce the development of cardiac arrhythmias. These results indicate that a preventive or therapeutic DNA vaccine encoding TSA-1 and Tc24 antigens is safe and may reduce both parasite transmission and the clinical progression of Chagas disease in vaccinated dogs. This DNA vaccine may thus be an excellent veterinary vaccine candidate. These data also further strengthen the feasibility of a Chagas disease vaccine for humans.

  3. Enhancement Effect of CpG DNA on the Somatostatin DNA Vaccine in Mice

    Institute of Scientific and Technical Information of China (English)

    XUE Chun-lin; MAO Da-gan; YANG Li-guo; CHENG Bao

    2007-01-01

    To study the immune effect of CpGDNA on somatostatin (SS) DNA vaccine, the 20-day-old experimental mice were immunized with 20 μg SS eukaryotic expression plasmid pES/2SS with different adjuvants in equal dose, such as the synthetic CpG-ODN, the pE-CpG plasmid, E. coli DNA and the crude liposome. A booster was given two weeks later. The results showed that the body weight gain of female mice in the SS immunized group was higher than that of the control (P<0.05). The levels of antibodies against SS, IgG2a/IgG1, spleen lymphocyte proliferation activity and the concentrations of GH and IGF- Ⅰ in the DNA vaccine groups combined with CpGDNA were significantly increased compared to that of the group immunized with DNA vaccine alone. All these suggested the recombinant SS expression plasmid can stimulate animals to produce antibodies against SS, and CpGDNA adjuvant can enhance the immune effect of DNA vaccine against SS and influence the concentration of GH and IGF- Ⅰ.

  4. Superparamagnetic nanoparticles for effective delivery of malaria DNA vaccine.

    Science.gov (United States)

    Al-Deen, Fatin Nawwab; Ho, Jenny; Selomulya, Cordelia; Ma, Charles; Coppel, Ross

    2011-04-05

    Low efficiency is often observed in the delivery of DNA vaccines. The use of superparamagnetic nanoparticles (SPIONs) to deliver genes via magnetofection could improve transfection efficiency and target the vector to its desired locality. Here, magnetofection was used to enhance the delivery of a malaria DNA vaccine encoding Plasmodium yoelii merozoite surface protein MSP1(19) (VR1020-PyMSP1(19)) that plays a critical role in Plasmodium immunity. The plasmid DNA (pDNA) containing membrane associated 19-kDa carboxyl-terminal fragment of merozoite surface protein 1 (PyMSP1(19)) was conjugated with superparamagnetic nanoparticles coated with polyethyleneimine (PEI) polymer, with different molar ratio of PEI nitrogen to DNA phosphate. We reported the effects of SPIONs-PEI complexation pH values on the properties of the resulting particles, including their ability to condense DNA and the gene expression in vitro. By initially lowering the pH value of SPIONs-PEI complexes to 2.0, the size of the complexes decreased since PEI contained a large number of amino groups that became increasingly protonated under acidic condition, with the electrostatic repulsion inducing less aggregation. Further reaggregation was prevented when the pHs of the complexes were increased to 4.0 and 7.0, respectively, before DNA addition. SPIONs/PEI complexes at pH 4.0 showed better binding capability with PyMSP1(19) gene-containing pDNA than those at neutral pH, despite the negligible differences in the size and surface charge of the complexes. This study indicated that the ability to protect DNA molecules due to the structure of the polymer at acidic pH could help improve the transfection efficiency. The transfection efficiency of magnetic nanoparticle as carrier for malaria DNA vaccine in vitro into eukaryotic cells, as indicated via PyMSP1(19) expression, was significantly enhanced under the application of external magnetic field, while the cytotoxicity was comparable to the benchmark nonviral

  5. Immunogenicity of candidate chimeric DNA vaccine against tuberculosis and leishmaniasis.

    Science.gov (United States)

    Dey, Ayan; Kumar, Umesh; Sharma, Pawan; Singh, Sarman

    2009-08-13

    Mycobacterium tuberculosis and Leishmania donovani are important intracellular pathogens, especially in Indian context. In India and other South East Asian countries, both these infections are highly endemic and in about 20% cases co-infection of these pathogens is reported. For both these pathogens cell mediated immunity plays most important role. The available treatment of these infections is either prolonged or cumbersome or it is ineffective in controlling the outbreaks and spread. Therefore, potentiation of a common host defense mechanism can be used to prevent both the infections simultaneously. In this study we have developed a novel chimeric DNA vaccine candidate comprising the esat-6 gene of M. tuberculosis and kinesin motor domain gene of L. donovani. After developing this novel chimera, its immunogenicity was studied in mouse model. The immune response was compared with individual constructs of esat-6 and kinesin motor domain. The results showed that immunization with chimeric DNA vaccine construct resulted in stronger IFN-gamma and IL-2 response against kinesin (3012+/-102 and 367.5+/-8.92pg/ml) and ESAT-6 (1334+/-46.5 and 245.1+/-7.72pg/ml) in comparison to the individual vaccine constructs. The reciprocal immune response (IFN-gamma and IL-2) against individual construct was lower (kinesin motor domain: 1788+/-36.48 and 341.8+/-9.801pg/ml and ESAT-6: 867.0+/-47.23 and 170.8+/-4.578pg/ml, respectively). The results also suggest that using the chimeric construct both proteins yielded a reciprocal adjuvant affect over each other as the IFN-gamma production against chimera vaccination is statistically significant (pleishmaniasis and tuberculosis and have important implication in future vaccine design.

  6. Potent tetravalent replicon vaccines against botulinum neurotoxins using DNA-based Semliki Forest virus replicon vectors.

    Science.gov (United States)

    Yu, Yun-Zhou; Guo, Jin-Peng; An, Huai-Jie; Zhang, Shu-Ming; Wang, Shuang; Yu, Wei-Yuan; Sun, Zhi-Wei

    2013-05-07

    Human botulism is commonly associated with botulinum neurotoxin (BoNT) serotypes A, B, E and F. This suggests that the greatest need is for a tetravalent vaccine that provides protection against all four of these serotypes. In current study, we investigated the feasibility of generating several tetravalent vaccines that protected mice against the four serotypes. Firstly, monovalent replicon vaccine against BoNT induced better antibody response and protection than that of corresponding conventional DNA vaccine. Secondly, dual-expression DNA replicon pSCARSE/FHc or replicon particle VRP-E/FHc vaccine was well resistant to the challenge of BoNT/E and BoNT/F mixture as a combination vaccine composed of two monovalent replicon vaccines. Finally, the dual-expression DNA replicon or replicon particle tetravalent vaccine could simultaneously and effectively neutralize and protect the four BoNT serotypes. Protection correlated directly with serum ELISA titers and neutralization antibody levels to BoNTs. Therefore, replicon-based DNA or particle might be effective vector to develop BoNT vaccines, which might be more desirable for use in clinical application than the conventional DNA vaccines. Our studies demonstrate the utility of combining dual-expression DNA replicon or replicon particle vaccines into multi-agent formulations as potent tetravalent vaccines for eliciting protective responses to four serotypes of BoNTs.

  7. Immunisation against PCV2 structural protein by DNA vaccination of mice

    DEFF Research Database (Denmark)

    Kamstrup, Søren; Barfoed, Annette Malene; Frimann, Tine;

    2004-01-01

    -protective levels around weaning at 3-5-weeks of age. If immunoprophylaxis is to be effective, an immunisation method capable of breaking through maternal immunity must be employed. In this study, we have developed and investigated the potential of a DNA vaccination approach to be one such method. The gene encoding...... the capsid protein of PCV2 was cloned in a DNA vaccination plasmid and expression of capsid protein was demonstrated in vitro. Mice were gene gun vaccinated three timesand all mice responded serologically by raising antibodies against PCV2. The results suggest, that DNA based vaccination might offer...... opportunities for vaccination of piglets against PCV2....

  8. Clinical development of intramuscular electroporation: providing a "boost" for DNA vaccines.

    Science.gov (United States)

    Khan, Amir S; Broderick, Kate E; Sardesai, Niranjan Y

    2014-01-01

    The development of effective vaccines has helped to eradicate or control the spread of numerous infectious diseases. However, there are many more diseases that have proved more difficult to eliminate using conventional vaccines. The recent innovation of DNA vaccines may provide a "boost" to the development efforts. While the early efforts of DNA vaccines in the clinic were disappointing, the use of in vivo electroporation has helped to provide some basis for optimism. Now, there are several ongoing clinical studies of vaccines against such diseases as malaria, HIV, hepatitis C, and even various types of cancer. This review will highlight three recently published clinical studies using intramuscular DNA administration with electroporation.

  9. [Recent advances in DNA vaccines against allergic airway disease: a review].

    Science.gov (United States)

    Ou, Jin; Xu, Yu; Shi, Wendan

    2013-12-01

    DNA vaccine is used in infectious diseases initially, and later is applied in neoplastic diseases, allergic diseases and other fields with the further understanding of DNA vaccine and the development of genetic engineering. DNA vaccine transfers the genes encoding exogenous antigens to plasmid vector and then is introduced into organism. It controls the antigen proteins synthesis, thus induces specific humoral and cellular immune responses. So it has a broad application prospect in allergic diseases. Compared with the traditional protein vaccines used in specific immunotherapy, DNA vaccine has many advantages, including high purity and specificity, and improvement of patients' compliance etc. However, there are still two unsolved problems. First, the transfection rate of unmodified naked DNA plasmid is not high, Second, it's difficult to induce ideal immune response. In this study, we will review the progress of DNA vaccine applications in respiratory allergic diseases and its various optimization strategies.

  10. DNA vaccine protects ornamental koi (Cyprinus carpio koi) against North American spring viremia of carp virus

    Science.gov (United States)

    Emmenegger, E.J.; Kurath, G.

    2008-01-01

    The emergence of spring viremia of carp virus (SVCV) in the United States constitutes a potentially serious alien pathogen threat to susceptible fish stocks in North America. A DNA vaccine with an SVCV glycoprotein (G) gene from a North American isolate was constructed. In order to test the vaccine a challenge model utilizing a specific pathogen-free domestic koi stock and a cold water stress treatment was also developed. We have conducted four trial studies demonstrating that the pSGnc DNA vaccine provided protection in vaccinated fish against challenge at low, moderate, and high virus doses of the homologous virus. The protection was significant (p vaccine construct containing a luciferase reporter gene and to non-vaccinated controls in fish ranging in age from 3 to 14 months. In all trials, the SVCV-G DNA immunized fish were challenged 28-days post-vaccination (546 degree-days) and experienced low mortalities varying from 10 to 50% with relative percent survivals ranging from 50 to 88%. The non-vaccinated controls and mock construct vaccinated fish encountered high cumulative percent mortalities ranging from 70 to 100%. This is the first report of a SVCV DNA vaccine being tested successfully in koi. These experiments prove that the SVCV DNA (pSGnc) vaccine can elicit specific reproducible protection and validates its potential use as a prophylactic vaccine in koi and other vulnerable North American fish stocks.

  11. Targeted Collection of Plasmid DNA in Large and Growing Animal Muscles 6 Weeks after DNA Vaccination with and without Electroporation

    Directory of Open Access Journals (Sweden)

    Daniel Dory

    2015-01-01

    Full Text Available DNA vaccination has been developed in the last two decades in human and animal species as a promising alternative to conventional vaccination. It consists in the injection, in the muscle, for example, of plasmid DNA encoding the vaccinating polypeptide. Electroporation which forces the entrance of the plasmid DNA in cells at the injection point has been described as a powerful and promising strategy to enhance DNA vaccine efficacy. Due to the fact that the vaccine is composed of DNA, close attention on the fate of the plasmid DNA upon vaccination has to be taken into account, especially at the injection point. To perform such studies, the muscle injection point has to be precisely recovered and collected several weeks after injection. This is even more difficult for large and growing animals. A technique has been developed to localize precisely and collect efficiently the muscle injection points in growing piglets 6 weeks after DNA vaccination accompanied or not by electroporation. Electroporation did not significantly increase the level of remaining plasmids compared to nonelectroporated piglets, and, in all the cases, the levels were below the limit recommended by the FDA to research integration events of plasmid DNA into the host DNA.

  12. Development of a human live attenuated West Nile infectious DNA vaccine: conceptual design of the vaccine candidate.

    Science.gov (United States)

    Yamshchikov, Vladimir

    2015-10-01

    West Nile virus has become an important epidemiological problem attracting significant attention of health authorities, mass media, and the public. Although there are promising advancements toward addressing the vaccine need, the perspectives of the commercial availability of the vaccine remain uncertain. To a large extent this is due to lack of a sustained interest for further commercial development of the vaccines already undergoing the preclinical and clinical development, and a predicted insignificant cost effectiveness of mass vaccination. There is a need for a safe, efficacious and cost effective vaccine, which can improve the feasibility of a targeted vaccination program. In the present report, we summarize the background, the rationale, and the choice of the development pathway that we selected for the design of a live attenuated human West Nile vaccine in a novel infectious DNA format.

  13. Intranasal DNA Vaccine for Protection against Respiratory Infectious Diseases: The Delivery Perspectives

    Directory of Open Access Journals (Sweden)

    Yingying Xu

    2014-07-01

    Full Text Available Intranasal delivery of DNA vaccines has become a popular research area recently. It offers some distinguished advantages over parenteral and other routes of vaccine administration. Nasal mucosa as site of vaccine administration can stimulate respiratory mucosal immunity by interacting with the nasopharyngeal-associated lymphoid tissues (NALT. Different kinds of DNA vaccines are investigated to provide protection against respiratory infectious diseases including tuberculosis, coronavirus, influenza and respiratory syncytial virus (RSV etc. DNA vaccines have several attractive development potential, such as producing cross-protection towards different virus subtypes, enabling the possibility of mass manufacture in a relatively short time and a better safety profile. The biggest obstacle to DNA vaccines is low immunogenicity. One of the approaches to enhance the efficacy of DNA vaccine is to improve DNA delivery efficiency. This review provides insight on the development of intranasal DNA vaccine for respiratory infections, with special attention paid to the strategies to improve the delivery of DNA vaccines using non-viral delivery agents.

  14. Enhancement of HIV-1 DNA vaccine immunogenicity by BCG-PSN, a novel adjuvant.

    Science.gov (United States)

    Sun, Jing; Hou, Jue; Li, Dingfeng; Liu, Yong; Hu, Ningzhu; Hao, Yanling; Fu, Jingjing; Hu, Yunzhang; Shao, Yiming

    2013-01-07

    Although the importance of DNA vaccines, especially as a priming immunization has been well established in numerous HIV vaccine studies, the immunogenictiy of DNA vaccines is generally moderate. Novel adjuvant is in urgent need for improving the immunogenicity of DNA vaccine. Polysaccharide and nucleic acid fraction extracted by hot phenol method from Mycobacterium bovis bacillus Calmette-Guérin, known as BCG-PSN, is a widely used immunomodulatory product in China clinical practice. In this study, we evaluated whether the BCG-PSN could serve as a novel adjuvant of DNA vaccine to trigger better cellular and humoral immune responses against the HIV-1 Env antigen in Balb/C mouse model. The BCG-PSN was mixed with 10 μg or 100 μg of pDRVI1.0gp145 (HIV-1 CN54 gp145 gene) DNA vaccine and intramuscularly immunized two or three times. We found that BCG-PSN could significantly improve the immunogenicity of DNA vaccine when co-administered with DNA vaccine. Further, at the same vaccination schedule, BCG-PSN co-immunization with 10 μg DNA vaccine could elicit cellular and humoral immune responses which were comparable to that induced by 100 μg DNA vaccine alone. Moreover, our results demonstrate that BCG-PSN can activate TLR signaling pathways and induce Th1-type cytokines secretion. These findings suggest that BCG-PSN can serve as a novel and effective adjuvant for DNA vaccination. Copyright © 2012 Elsevier Ltd. All rights reserved.

  15. Epitope-driven DNA vaccine design employing immunoinformatics against B-cell lymphoma: a biotech's challenge.

    Science.gov (United States)

    Iurescia, Sandra; Fioretti, Daniela; Fazio, Vito Michele; Rinaldi, Monica

    2012-01-01

    DNA vaccination has been widely explored to develop new, alternative and efficient vaccines for cancer immunotherapy. DNA vaccines offer several benefits such as specific targeting, use of multiple genes to enhance immunity and reduced risk compared to conventional vaccines. Rapid developments in molecular biology and immunoinformatics enable rational design approaches. These technologies allow construction of DNA vaccines encoding selected tumor antigens together with molecules to direct and amplify the desired effector pathways, as well as highly targeted vaccines aimed at specific epitopes. Reliable predictions of immunogenic T cell epitope peptides are crucial for rational vaccine design and represent a key problem in immunoinformatics. Computational approaches have been developed to facilitate the process of epitope detection and show potential applications to the immunotherapeutic treatment of cancer. In this review a number of different epitope prediction methods are briefly illustrated and effective use of these resources to support experimental studies is described. Epitope-driven vaccine design employs these bioinformatics algorithms to identify potential targets of vaccines against cancer. In this paper the selection of T cell epitopes to develop epitope-based vaccines, the need for CD4(+) T cell help for improved vaccines and the assessment of vaccine performance against tumor are reviewed. We focused on two applications, namely prediction of novel T cell epitopes and epitope enhancement by sequence modification, and combined rationale design with bioinformatics for creation of new synthetic mini-genes. This review describes the development of epitope-based DNA vaccines and their antitumor effects in preclinical research against B-cell lymphoma, corroborating the usefulness of this platform as a potential tool for cancer therapy. Achievements in the field of DNA vaccines allow to overcome hurdles to clinical translation. In a scenario where the vaccine

  16. DNA vaccination of pigs with open reading frame 1-7 of PRRS virus

    DEFF Research Database (Denmark)

    Barfoed, Annette Malene; Blixenkrone-Møller, Merete; Jensen, Merethe Holm

    2004-01-01

    We cloned all open reading frames of a Danish isolate of porcine reproductive and respiratory syndrome (PRRS) virus in DNA vaccination vectors. Pigs were vaccinated using a gene gun with each single construct (ORF1, ORF2, ORF3, ORF4, ORF5, ORF6, or ORF7) or combinations thereof. Vaccination...

  17. Phase 1 study of pandemic H1 DNA vaccine in healthy adults.

    Directory of Open Access Journals (Sweden)

    Michelle C Crank

    Full Text Available A novel, swine-origin influenza A (H1N1 virus was detected worldwide in April 2009, and the World Health Organization (WHO declared a global pandemic that June. DNA vaccine priming improves responses to inactivated influenza vaccines. We describe the rapid production and clinical evaluation of a DNA vaccine encoding the hemagglutinin protein of the 2009 pandemic A/California/04/2009(H1N1 influenza virus, accomplished nearly two months faster than production of A/California/07/2009(H1N1 licensed monovalent inactivated vaccine (MIV.20 subjects received three H1 DNA vaccinations (4 mg intramuscularly with Biojector at 4-week intervals. Eighteen subjects received an optional boost when the licensed H1N1 MIV became available. The interval between the third H1 DNA injection and MIV boost was 3-17 weeks. Vaccine safety was assessed by clinical observation, laboratory parameters, and 7-day solicited reactogenicity. Antibody responses were assessed by ELISA, HAI and neutralization assays, and T cell responses by ELISpot and flow cytometry.Vaccinations were safe and well-tolerated. As evaluated by HAI, 6/20 developed positive responses at 4 weeks after third DNA injection and 13/18 at 4 weeks after MIV boost. Similar results were detected in neutralization assays. T cell responses were detected after DNA and MIV. The antibody responses were significantly amplified by the MIV boost, however, the boost did not increased T cell responses induced by DNA vaccine.H1 DNA vaccine was produced quickly, was well-tolerated, and had modest immunogenicity as a single agent. Other HA DNA prime-MIV boost regimens utilizing one DNA prime vaccination and longer boost intervals have shown significant immunogenicity. Rapid and large-scale production of HA DNA vaccines has the potential to contribute to an efficient response against future influenza pandemics.Clinicaltrials.gov NCT00973895.

  18. DNA vaccine against visceral leishmaniasis: a promising approach for prevention and control.

    Science.gov (United States)

    Kumar, A; Samant, M

    2016-05-01

    The visceral leishmaniasis (VL) caused by Leishmania donovani parasite severely affects large populations in tropical and subtropical regions of the world. The arsenal of drugs available is limited, and resistance is common in clinical field isolates. Therefore, vaccines could be an important alternative for prevention against VL. Recently, some investigators advocated the protective efficacy of DNA vaccines, which induces the T cell-based immunity against VL. The vaccine antigens are selected as conserved in various Leishmania species and provide a viable strategy for DNA vaccine development. Our understanding for DNA vaccine development against VL is not enough and much technological advancement is required. Improved formulations and methods of delivery are required, which increase the uptake of DNA vaccine by cells; optimization of vaccine vectors/encoded antigens to augment and direct the host immune response in VL. Despite the many genes identified as vaccine candidates, the disappointing potency of the DNA vaccines in VL underscores the challenges encountered in the efforts to translate efficacy in preclinical models into clinical realities. This review will provide a brief background of DNA vaccines including the insights gained about the design, strategy, safety issues, varied candidates, progress and challenges that play a role in their ability against VL.

  19. Therapeutic Effects of DNA Vaccine on Allergen-Induced Allergic Airway Inflammation in Mouse Model

    Institute of Scientific and Technical Information of China (English)

    Guoping Li; Zhigang Liu; Nanshan Zhong; Bin Liao1; Ying Xiong

    2006-01-01

    Vaccination with DNA encoding Dermatophagoides pteronyssinus group 2 (Der p 2) allergen previously showed its effects of immunologic protection on Der p 2 allergen-induced allergic airway inflammation in mice. In present study, we investigated whether DNA vaccine encoding Der p 2 could exert therapeutic role on allergen-induced allergic airway inflammation in mouse model and explored the mechanism of DNA vaccination in asthma specific-allergen immunotherapy. After sensitized and challenged by Der p 2, the BALB/c mice were immunized with DNA vaccine. The degrees of cellular infiltration were scored. IgE levels in serum and IL-4/lL-13 levels in BALF were determined by ELISA. The lung tissues were assessed by histological examinations. Expressions of STAT6 and NF-κB in lung were determined by immunohistochemistry staining. Vaccination of mice with DNA vaccine inhibited the development of airway inflammation and the production of mucin induced by allergen, and reduced the level of Der p 2-specific IgE level. Significant reductions of eosinophii infiltration and levels of IL-4and IL-13 in BALF were observed after vaccination. Further more, DNA vaccination inhibited STAT6 and NF-κBexpression in lung tissue in Der p 2-immunized mice. These results indicated that DNA vaccine encoding Der p 2allergen could be used for therapy of allergen-induced allergic airway inflammation in our mouse model.

  20. Preparation, characterization, and in ovo vaccination of dextran-spermine nanoparticle DNA vaccine coexpressing the fusion and hemagglutinin genes against Newcastle disease.

    Science.gov (United States)

    Firouzamandi, Masoumeh; Moeini, Hassan; Hosseini, Seyed Davood; Bejo, Mohd Hair; Omar, Abdul Rahman; Mehrbod, Parvaneh; El Zowalaty, Mohamed E; Webster, Thomas J; Ideris, Aini

    2016-01-01

    Plasmid DNA (pDNA)-based vaccines have emerged as effective subunit vaccines against viral and bacterial pathogens. In this study, a DNA vaccine, namely plasmid internal ribosome entry site-HN/F, was applied in ovo against Newcastle disease (ND). Vaccination was carried out using the DNA vaccine alone or as a mixture of the pDNA and dextran-spermine (D-SPM), a nanoparticle used for pDNA delivery. The results showed that in ovo vaccination with 40 μg pDNA/egg alone induced high levels of antibody titer (P0.05). Higher antibody titer was observed in the group immunized with 40 μg pDNA/egg at 4 weeks postvaccination. The findings also showed that vaccination with 40 μg pDNA/egg alone was able to confer protection against Newcastle disease virus strain NDIBS002 in two out of seven SPF chickens. Although the chickens produced antibody titers 3 weeks after in ovo vaccination, it was not sufficient to provide complete protection to the chickens from lethal viral challenge. In addition, vaccination with pDNA/D-SPM complex did not induce high antibody titer when compared with naked pDNA. Therefore, it was concluded that DNA vaccination with plasmid internal ribosome entry site-HN/F can be suitable for in ovo application against ND, whereas D-SPM is not recommended for in ovo gene delivery.

  1. Transient global T cell activation after vaccination of rhesus macaques with a DNA-poxvirus vaccine regimen for HIV.

    Science.gov (United States)

    Soares, Andreia; Müller, Tracey L; Chege, Gerald K; Williamson, Anna-Lise; Burgers, Wendy A

    2015-07-09

    Persistent T cell activation following immunization with HIV vaccines may increase HIV acquisition risk. We investigated the magnitude and kinetics of T cell activation following vaccination of rhesus macaques with a candidate HIV vaccine consisting of a recombinant DNA and MVA vaccination regimen. We show that global CD4+ and CD8+ T cell activation, as measured by the expression of Ki67 and Bcl-2, peaked one week after boosting with MVA, but then waned rapidly to pre-vaccination levels. Furthermore, increased frequencies of CD4+ CCR5+ T cells, which represent potential HIV target cells, were short-lived and decreased to baseline levels within two months. Activated CD4+ T cells were predominantly of a central memory phenotype, and activated CD8+ T cells were distributed between central and effector memory phenotypes. Thus, only transient changes in T cell activation occurred following poxvirus vaccination, indicating a lack of persistent immune activation. Copyright © 2015 Elsevier Ltd. All rights reserved.

  2. Vaccination with pcDNA3-15/60 Naked DNA Encoding the Surface Proteinof Sporozoites in Cryptosporidium parvum

    Institute of Scientific and Technical Information of China (English)

    HEHong-xuan; ZHANGXi-chen; YINJi-gang; LIJian-hua; YANGJu

    2004-01-01

    The CP15/60 gene encoding the CP15/60 surface protein of sporozoites in Cryptosporidium parvum was obtained by PCR so as to research the nucleic vaccine against C.parvum. The eukaryotic expressing vector pcDNA3-15/60 was constructed by inserting CP15/60 gene into pcDNA3 (+) in Xho Ⅰ and EcoR Ⅰ. A vaccination protocol was the adult pregnant goats inoculated intranasally with the pcDNA3-15/60 plasmid and their offspring were infected with C.parvum oocysts. The results showed that the pcDNA3-15/60 plasmid can induce the immune response of goats and the vaccinated goats can transfer the immunity to offspring conferring protection against C.parvum infection. These suggested that the recombinant plasmid could be a DNA vaccine candidate.

  3. Vaccination with pcDNA3-15/60 Naked DNA Encoding the Surface Protein of Sporozoites in Cryptosporidium parvum

    Institute of Scientific and Technical Information of China (English)

    HE Hong-xuan; ZHANG Xi-chen; YIN Ji-gang; LI Jian-hua; YANG Ju

    2004-01-01

    The CP15/60 gene encoding the CP15/60 surface protein of sporozoites in Cryptosporidium parvum was obtained by PCR so as to research the nucleic vaccine against C.parvum. The eukaryotic expressing vector pcDNA3-15/60 was constructed by inserting CP15/60 gene into pcDNA3 (+) in Xho Ⅰ and EcoR Ⅰ. A vaccination protocol was the adult pregnant goats inoculated intranasally with the pcDNA3-15/60 plasmid and their offspring were infected with C.parvum oocysts. The results showed that the pcDNA3-15/60 plasmid can induce the immune response of goats and the vaccinated goats can transfer the immunity to offspring conferring protection against C.parvum infection. These suggested that the recombinant plasmid could be a DNA vaccine candidate.

  4. Advances in research of DNA vaccine%DNA 疫苗研究进展

    Institute of Scientific and Technical Information of China (English)

    郭杨(综述); 方刚(审校)

    2013-01-01

    DNA疫苗是20世纪90年代初出现的一种新型疫苗,近年来发展迅速,在预防和治疗病毒性疾病及肿瘤等方面效果显著。同传统的疫苗相比,DNA疫苗具有免疫效果好、生产成本低、临床应用方便等优点,但同样存在安全性的担忧。对DNA疫苗的发展及其作用机制、优势进行了综述,并对DNA疫苗的安全性提出了自己的观点与看法,可供DNA疫苗的研究者参考。%As a novel vaccine set up in early 1990 s, DNA vaccine has been developed rapidly in recent years and played an important role in the prevention and treatment of viral diseases and tumor .Compared with the traditional vaccines , it had some advantages , such as good immune effect , low production cost , and convenient usage for the clinical application , but it could also be found safety concerns .To provide the references for DNA vaccine researchers , the development , mecha-nism, and advantage are reviewed in this paper , and my personal viewpoint about its safety is presented as well .

  5. Protective antibody titres and antigenic competition in multivalent Dichelobacter nodosus fimbrial vaccines using characterised rDNA antigens.

    Science.gov (United States)

    Raadsma, H W; O'Meara, T J; Egerton, J R; Lehrbach, P R; Schwartzkoff, C L

    1994-03-01

    The relationship between K-agglutination antibody titres and protection against experimental challenge with Dichelobacter nodosus, the effect of increasing the number of D. nodosus fimbrial antigens, and the importance of the nature of additional antigens in multivalent vaccines on antibody response and protection against experimental challenge with D. nodosus were examined in Merino sheep. A total of 204 Merino sheep were allocated to one of 12 groups, and vaccinated with preparations containing a variable number of rDNA D. nodosus fimbrial antigens. The most complex vaccine contained ten fimbrial antigens from all major D. nodosus serogroups, while the least complex contained a single fimbrial antigen. In addition to D. nodosus fimbrial antigens, other bacterial rDNA fimbrial antigens (Moraxella bovis Da12d and Escherichia coli K99), and bovine serum albumin (BSA) were used in some vaccines. Antibody titres to fimbrial antigens and BSA were measured by agglutination and ELISA tests, respectively. Antibody titres were determined on five occasions (Weeks 0, 3, 6, 8, and 11 after primary vaccination). All sheep were exposed to an experimental challenge with virulent isolates of D. nodosus from either serogroup A or B, 8 weeks after primary vaccination. For D. nodosus K-agglutinating antibody titres, a strong negative correlation between antibody titre and footrot lesion score was observed. This relationship was influenced by the virulence of the challenge strain. Increasing the number of fimbrial antigens in experimental rDNA D. nodosus fimbrial vaccines resulted in a linear decrease in K-agglutinating antibody titres to individual D. nodosus serogroups. Similarly, a linear decrease in protection to challenge with homologous serogroups was observed as the number of D. nodosus fimbrial antigens represented in the vaccine increased. The reduction in antibody titres in multicomponent vaccines is thought to be due to antigenic competition. The level of competition

  6. Subtype C gp140 Vaccine Boosts Immune Responses Primed by the South African AIDS Vaccine Initiative DNA-C2 and MVA-C HIV Vaccines after More than a 2-Year Gap.

    Science.gov (United States)

    Gray, Glenda E; Mayer, Kenneth H; Elizaga, Marnie L; Bekker, Linda-Gail; Allen, Mary; Morris, Lynn; Montefiori, David; De Rosa, Stephen C; Sato, Alicia; Gu, Niya; Tomaras, Georgia D; Tucker, Timothy; Barnett, Susan W; Mkhize, Nonhlanhla N; Shen, Xiaoying; Downing, Katrina; Williamson, Carolyn; Pensiero, Michael; Corey, Lawrence; Williamson, Anna-Lise

    2016-06-01

    A phase I safety and immunogenicity study investigated South African AIDS Vaccine Initiative (SAAVI) HIV-1 subtype C (HIV-1C) DNA vaccine encoding Gag-RT-Tat-Nef and gp150, boosted with modified vaccinia Ankara (MVA) expressing matched antigens. Following the finding of partial protective efficacy in the RV144 HIV vaccine efficacy trial, a protein boost with HIV-1 subtype C V2-deleted gp140 with MF59 was added to the regimen. A total of 48 participants (12 U.S. participants and 36 Republic of South Africa [RSA] participants) were randomized to receive 3 intramuscular (i.m.) doses of SAAVI DNA-C2 of 4 mg (months 0, 1, and 2) and 2 i.m. doses of SAAVI MVA-C of 1.45 × 10(9) PFU (months 4 and 5) (n = 40) or of a placebo (n = 8). Approximately 2 years after vaccination, 27 participants were rerandomized to receive gp140/MF59 at 100 μg or placebo, as 2 i.m. injections, 3 months apart. The vaccine regimen was safe and well tolerated. After the DNA-MVA regimen, CD4(+) T-cell and CD8(+) T-cell responses occurred in 74% and 32% of the participants, respectively. The protein boost increased CD4(+) T-cell responses to 87% of the subjects. All participants developed tier 1 HIV-1C neutralizing antibody responses as well as durable Env binding antibodies that recognized linear V3 and C5 peptides. The HIV-1 subtype C DNA-MVA vaccine regimen showed promising cellular immunogenicity. Boosting with gp140/MF59 enhanced levels of binding and neutralizing antibodies as well as CD4(+) T-cell responses to HIV-1 envelope. (This study has been registered at ClinicalTrials.gov under registration no. NCT00574600 and NCT01423825.).

  7. Transcriptome profiles associated to VHSV infection or DNA vaccination in turbot (Scophthalmus maximus).

    Science.gov (United States)

    Pereiro, Patricia; Dios, Sonia; Boltaña, Sebastián; Coll, Julio; Estepa, Amparo; Mackenzie, Simon; Novoa, Beatriz; Figueras, Antonio

    2014-01-01

    DNA vaccines encoding the viral G glycoprotein show the most successful protection capability against fish rhabdoviruses. Nowadays, the molecular mechanisms underlying the protective response remain still poorly understood. With the aim of shedding light on the protection conferred by the DNA vaccines based in the G glycoprotein of viral haemorrhagic septicaemia virus (VHSV) in turbot (Scophthalmus maximus) we have used a specific microarray highly enriched in antiviral sequences to carry out the transcriptomic study associated to VHSV DNA vaccination/infection. The differential gene expression pattern in response to empty plasmid (pMCV1.4) and DNA vaccine (pMCV1.4-G860) intramuscular administration with regard to non-stimulated turbot was analyzed in head kidney at 8, 24 and 72 hours post-vaccination. Moreover, the effect of VHSV infection one month after immunization was also analyzed in vaccinated and non-vaccinated fish at the same time points. Genes implicated in the Toll-like receptor signalling pathway, IFN inducible/regulatory proteins, numerous sequences implicated in apoptosis and cytotoxic pathways, MHC class I antigens, as well as complement and coagulation cascades among others were analyzed in the different experimental groups. Fish receiving the pMCV1.4-G860 vaccine showed transcriptomic patterns very different to the ones observed in pMCV1.4-injected turbot after 72 h. On the other hand, VHSV challenge in vaccinated and non-vaccinated turbot induced a highly different response at the transcriptome level, indicating a very relevant role of the acquired immunity in vaccinated fish able to alter the typical innate immune response profile observed in non-vaccinated individuals. This exhaustive transcriptome study will serve as a complete overview for a better understanding of the crosstalk between the innate and adaptive immune response in fish after viral infection/vaccination. Moreover, it provides interesting clues about molecules with a potential

  8. Immunogenicity and protective efficacy of a vaxfectin-adjuvanted tetravalent dengue DNA vaccine.

    Science.gov (United States)

    Porter, Kevin R; Ewing, Daniel; Chen, Lan; Wu, Shuenn-Jue; Hayes, Curtis G; Ferrari, Marilyn; Teneza-Mora, Nimfa; Raviprakash, Kanakatte

    2012-01-05

    A prototype dengue-1 DNA vaccine was shown to be safe and immunogenic in a previous Phase 1 clinical trial. Anti-dengue-1 neutralizing antibody responses were detectable only in the group of volunteers receiving the high dose of nonadjuvanted vaccine and the antibody titers were low. Vaxfectin(®), a lipid-based adjuvant, enhances the immunogenicity of DNA vaccines. We conducted a nonhuman primate study to evaluate the effect of Vaxfectin(®) on the immunogenicity of a tetravalent dengue DNA vaccine. Animals were immunized on days 0, 28 and 84, with each immunization consisting of 3mg of Vaxfectin(®)-adjuvanted tetravalent dengue DNA vaccine. The use of Vaxfectin(®) resulted in a significant increase in anti-dengue neutralizing antibody responses against dengue-1, -3 and -4. There was little to no effect on T cell responses as measured by interferon gamma ELISPOT assay. Animals immunized with the Vaxfectin(®)-formulated tetravalent DNA vaccine showed significant protection against live dengue-2 virus challenge compared to control animals (0.75 mean days of viremia vs 3.3 days). Animals vaccinated with nonadjuvanted DNA had a mean 2.0 days of viremia. These results support further evaluation of the Vaxfectin(®)-adjuvanted tetravalent dengue DNA vaccine in a Phase 1 clinical trial.

  9. Use of DNA vaccination for determination of onset of adaptive immunity in rainbow trout fry

    DEFF Research Database (Denmark)

    Rasmussen, Jesper Skou; Lorenzen, Ellen; Kjær, Torben Egil

    2013-01-01

    that intramuscular injection of the DNA vaccine encoding the viral glycoprotein G induced protective immunity to VHS in rainbow trout fry of 0.5g.However, the vaccine is known to induce both innate and adaptive protection. The present work therefore aimed at determination of which type of protection the DNA vaccine...... to innate cross-reactive antiviral mechanisms of shorter duration. The critical size for induction of an adaptive immune response in rainbow trout to this type of vaccination thus appears to be between 0.25 and 0.5g. This work was supported by the “DAFINET” grant from the Danish Council for Strategic...... the duration and nature of the protective immunity induced by the vaccines in the fish. The present work aimed at determination of the smallest size at which specific immunity could be induced in rainbow trout fry by DNA vaccination against viral haemorrhagic septicaemia (VHS). Earlier experiments revealed...

  10. Evaluation of a DNA Vaccine for Immunocontraceptive Potential Against Zona Pellucida Glycoproteins in Cattle

    Directory of Open Access Journals (Sweden)

    C. A. Foley

    2007-01-01

    Full Text Available Holstein cows were administered zona pellucida (ZP DNA vaccine and used to determine the potential of recombinant rabbit ZP glycoproteins (rZP as immunocontraceptive antigens. Zona pellucida proteins were purified and quantified. Cows were assigned to one of four treatment groups in which plasmids encoding rabbit ZP proteins were administered, i.d., using a gene gun (ZP55, n=2; ZP75, n=2; Hep55, n=2; and Control, n=3. Blood samples were taken before initial vaccination, once weekly for 5 wk and at 148 wk post-immunization. An ELISA was developed to assess anti-ZP titer levels in cow serum and ovarian function in cows was monitored using trans-rectal ultrasonography. Four of the six cows in ZP treatment groups developed antibody titer levels with similar linear responses over time. These cows also experienced reduced ovarian function as indicated by decreases in follicular and luteal activity. Estrous activity was observed in all cows and decreased in ZP treatment cows in comparison to Controls. Further research is needed to determine the relationship between ZP immunocontraception and ovarian function. Still, this study provides a basis for future researchers to use in developing a contraceptive vaccine for cattle.

  11. DNA vaccine initiates replication of live attenuated chikungunya virus in vitro and elicits protective immune response in mice.

    Science.gov (United States)

    Tretyakova, Irina; Hearn, Jason; Wang, Eryu; Weaver, Scott; Pushko, Peter

    2014-06-15

    Chikungunya virus (CHIKV) causes outbreaks of chikungunya fever worldwide and represents an emerging pandemic threat. Vaccine development against CHIKV has proved challenging. Currently there is no approved vaccine or specific therapy for the disease. To develop novel experimental CHIKV vaccine, we used novel immunization DNA (iDNA) infectious clone technology, which combines the advantages of DNA and live attenuated vaccines. Here we describe an iDNA vaccine composed of plasmid DNA that encode the full-length infectious genome of live attenuated CHIKV clone 181/25 downstream from a eukaryotic promoter. The iDNA approach was designed to initiate replication of live vaccine virus from the plasmid in vitro and in vivo. Experimental CHIKV iDNA vaccines were prepared and evaluated in cultured cells and in mice. Transfection with 10 ng of iDNA was sufficient to initiate replication of vaccine virus in vitro. Vaccination of BALB/c mice with a single 10 μg of CHIKV iDNA plasmid resulted in seroconversion, elicitation of neutralizing antibodies, and protection from experimental challenge with a neurovirulent CHIKV. Live attenuated CHIKV 181/25 vaccine can be delivered in vitro and in vivo by using DNA vaccination. The iDNA approach appears to represent a promising vaccination strategy for CHIK and other alphaviral diseases. © The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  12. Immunogenicity of Virus Like Particle Forming Baculoviral DNA Vaccine against Pandemic Influenza H1N1.

    Directory of Open Access Journals (Sweden)

    Yong-Dae Gwon

    Full Text Available An outbreak of influenza H1N1 in 2009, representing the first influenza pandemic of the 21st century, was transmitted to over a million individuals and claimed 18,449 lives. The current status in many countries is to prepare influenza vaccine using cell-based or egg-based killed vaccine. However, traditional influenza vaccine platforms have several limitations. To overcome these limitations, many researchers have tried various approaches to develop alternative production platforms. One of the alternative approach, we reported the efficacy of influenza HA vaccination using a baculoviral DNA vaccine (AcHERV-HA. However, the immune response elicited by the AcHERV-HA vaccine, which only targets the HA antigen, was lower than that of the commercial killed vaccine. To overcome the limitations of this previous vaccine, we constructed a human endogenous retrovirus (HERV envelope-coated, baculovirus-based, virus-like-particle (VLP-forming DNA vaccine (termed AcHERV-VLP against pandemic influenza A/California/04/2009 (pH1N1. BALB/c mice immunized with AcHERV-VLP (1×107 FFU AcHERV-VLP, i.m. and compared with mice immunized with the killed vaccine or mice immunized with AcHERV-HA. As a result, AcHERV-VLP immunization produced a greater humoral immune response and exhibited neutralizing activity with an intrasubgroup H1 strain (PR8, elicited neutralizing antibody production, a high level of interferon-γ secretion in splenocytes, and diminished virus shedding in the lung after challenge with a lethal dose of influenza virus. In conclusion, VLP-forming baculovirus DNA vaccine could be a potential vaccine candidate capable of efficiently delivering DNA to the vaccinee and VLP forming DNA eliciting stronger immunogenicity than egg-based killed vaccines.

  13. Immune response to dna vaccine expressing transferrin binding protein a gene of Pasteurella multocida

    Directory of Open Access Journals (Sweden)

    Satparkash Singh

    2011-06-01

    Full Text Available Haemorrhagic Septicaemia (HS, an acute and fatal disease of cattle and buffalo is primarily caused by serotype B:2 or E:2 of Pasteurella multocida. The transferrin binding protein A (TbpA has been found to act as immunogen and potent vaccine candidate in various Gram negative bacteria including P. multocida. The present study was carried out to evaluate the potential of this antigen as a DNA vaccine against HS in mice model. The tbpA gene of P. multocida serotype B:2 was cloned in a mammalian expression vector alone and along with murine IL2 gene as immunological adjuvant to produce monocistronic and bicistronic DNA vaccine constructs, respectively. The immune response to DNA vaccines was evaluated based on serum antibody titres and lymphocyte proliferation assay. A significant increase in humoral and cell mediated immune responses was observed in mice vaccinated with DNA vaccines as compared to non immunized group. Additionally, the bicistronic DNA vaccine provided superior immune response and protection level following challenge as compared to monocistronic construct. The study revealed that DNA vaccine presents a promising approach for the prevention of HS.

  14. Immune response to dna vaccine expressing transferrin binding protein a gene of Pasteurella multocida.

    Science.gov (United States)

    Singh, Satparkash; Singh, Vijendra Pal; Cheema, Pawanjit Singh; Sandey, Maninder; Ranjan, Rajeev; Gupta, Santosh Kumar; Sharma, Bhaskar

    2011-04-01

    Haemorrhagic Septicaemia (HS), an acute and fatal disease of cattle and buffalo is primarily caused by serotype B:2 or E:2 of Pasteurella multocida. The transferrin binding protein A (TbpA) has been found to act as immunogen and potent vaccine candidate in various Gram negative bacteria including P. multocida. The present study was carried out to evaluate the potential of this antigen as a DNA vaccine against HS in mice model. The tbpA gene of P. multocida serotype B:2 was cloned in a mammalian expression vector alone and along with murine IL2 gene as immunological adjuvant to produce monocistronic and bicistronic DNA vaccine constructs, respectively. The immune response to DNA vaccines was evaluated based on serum antibody titres and lymphocyte proliferation assay. A significant increase in humoral and cell mediated immune responses was observed in mice vaccinated with DNA vaccines as compared to non immunized group. Additionally, the bicistronic DNA vaccine provided superior immune response and protection level following challenge as compared to monocistronic construct. The study revealed that DNA vaccine presents a promising approach for the prevention of HS.

  15. What you always needed to know about electroporation based DNA vaccines

    DEFF Research Database (Denmark)

    Gothelf, Anita Birgitte; Gehl, Julie

    2012-01-01

    Vaccinations are increasingly used to fight infectious disease, and DNA vaccines offer considerable advantages, including broader possibilities for vaccination and lack of need for cold storage. It has been amply demonstrated, that electroporation augments uptake of DNA in both skin and muscle......, and it is foreseen that future DNA vaccination may to a large extent be coupled with and dependent upon electroporation based delivery. Understanding the basic science of electroporation and exploiting knowledge obtained on optimization of DNA electrotransfer to muscle and skin, may greatly augment efforts...... on vaccine development. The purpose of this review is to give a succinct but comprehensive overview of electroporation as a delivery modality including electrotransfer to skin and muscle. As well, this review will speculate and discuss future uses for this powerful electrotransfer technology....

  16. Chicken HSP70 DNA vaccine inhibits tumor growth in a canine cancer model.

    Science.gov (United States)

    Yu, Wen-Ying; Chuang, Tien-Fu; Guichard, Cécile; El-Garch, Hanane; Tierny, Dominique; Laio, Albert Taiching; Lin, Ching-Si; Chiou, Kuo-Hao; Tsai, Cheng-Long; Liu, Chen-Hsuan; Li, Wen-Chiuan; Fischer, Laurent; Chu, Rea-Min

    2011-04-18

    Immunization with xenogeneic DNA is a promising cancer treatment to overcome tolerance to self-antigens. Heat shock protein 70 (HSP70) is over-expressed in various kinds of tumors and is believed to be involved in tumor progression. This study tested a xenogeneic chicken HSP70 (chHSP70) DNA vaccine in an experimental canine transmissible venereal tumor (CTVT) model. Three vaccination strategies were compared: the first (PE) was designed to evaluate the prophylactic efficacy of chHSP70 DNA vaccination by delivering the vaccine before tumor inoculation in a prime boost setting, the second (T) was designed to evaluate the therapeutic efficacy of the same prime boost vaccine by vaccinating the dogs after tumor inoculation; the third (PT) was similar to the first strategy (PE), with the exception that the electroporation booster injection was replaced with a transdermal needle-free injection. Tumor growth was notably inhibited only in the PE dogs, in which the vaccination program triggered tumor regression significantly sooner than in control dogs (NT). The CD4(+) subpopulation of tumor-infiltrating lymphocytes and canine HSP70 (caHSP70)-specific IFN-γ-secreting lymphocytes were significantly increased during tumor regression in the PE dogs as compared to control dogs, demonstrating that specific tolerance to caHSP70 has been overcome. In contrast, no benefit of the therapeutic strategy (T) could be noticed and the (PT) strategy only led to partial control of tumor growth. In summary, antitumor prophylactic activity was demonstrated using the chHSP70 DNA vaccine including a boost via electroporation. Our data stressed the importance of DNA electroporation as a booster to get the full benefit of DNA vaccination but also of cancer immunotherapy initiation as early as possible. Xenogeneic chHSP70 DNA vaccination including an electroporation boost is a potential vaccine to HSP70-expressing tumors, although further research is still required to better understand true

  17. Comprehensive gene expression profiling following DNA vaccination of rainbow trout against infectious hematopoietic necrosis virus

    Science.gov (United States)

    Purcell, Maureen K.; Nichols, Krista M.; Winton, James R.; Kurath, Gael; Thorgaard, Gary H.; Wheeler, Paul; Hansen, John D.; Herwig, Russell P.; Park, Linda K.

    2006-01-01

    The DNA vaccine based on the glycoprotein gene of Infectious hematopoietic necrosis virus induces a non-specific anti-viral immune response and long-term specific immunity against IHNV. This study characterized gene expression responses associated with the early anti-viral response. Homozygous rainbow trout were injected intra-muscularly (I.M.) with vector DNA or the IHNV DNA vaccine. Gene expression in muscle tissue (I.M. site) was evaluated using a 16,008 feature salmon cDNA microarray. Eighty different genes were significantly modulated in the vector DNA group while 910 genes were modulated in the IHNV DNA vaccinate group relative to control group. Quantitative reverse-transcriptase PCR was used to examine expression of selected immune genes at the I.M. site and in other secondary tissues. In the localized response (I.M. site), the magnitudes of gene expression changes were much greater in the vaccinate group relative to the vector DNA group for the majority of genes analyzed. At secondary systemic sites (e.g. gill, kidney and spleen), type I IFN-related genes were up-regulated in only the IHNV DNA vaccinated group. The results presented here suggest that the IHNV DNA vaccine induces up-regulation of the type I IFN system across multiple tissues, which is the functional basis of early anti-viral immunity.

  18. A novel non-integrative single-cycle chimeric HIV lentivector DNA vaccine.

    Science.gov (United States)

    Moussa, Maha; Arrode-Brusés, Géraldine; Manoylov, Iliyan; Malogolovkin, Alexander; Mompelat, Dimitri; Ishimwe, Honorine; Smaoune, Amel; Ouzrout, Bilel; Gagnon, Jean; Chebloune, Yahia

    2015-05-05

    Novel HIV vaccine vectors and strategies are needed to control HIV/AIDS epidemic in humans and eradicate the infection. DNA vaccines alone failed to induce immune responses robust enough to control HIV-1. Development of lentivirus-based DNA vaccines deficient for integration and with a limited replication capacity is an innovative and promising approach. This type of vaccine mimics the early stages of virus infection/replication like the live-attenuated viruses but lacks the inconvenient integration and persistence associated with disease. We developed a novel lentivector DNA vaccine "CAL-SHIV-IN(-)" that undergoes a single round of replication in the absence of integration resulting in augmented expression of vaccine antigens in vivo. Vaccine gene expression is under control of the LTRs of a naturally attenuated lentivirus, Caprine arthritis encephalitis virus (CAEV) the natural goat lentivirus. The safety of this vaccine prototype was increased by the removal of the integrase coding sequences from the pol gene. We examined the functional properties of this lentivector DNA in cell culture and the immunogenicity in mouse models. Viral proteins were expressed in transfected cells, assembled into viral particles that were able to transduce once target permissive cells. Unlike the parental replication-competent SHIV-KU2 that was detected in DNA samples from any of the serial passage infected cells, CAL-SHIV-IN(-) DNA was detected only in target cells of the first round of infection, hence demonstrating the single cycle replication of the vaccine. A single dose DNA immunization of humanized NOD/SCID/β2 mice showed a substantial increase of IFN-γ-ELISPOT in splenocytes compared to the former replication and integration defective Δ4SHIV-KU2 DNA vaccine. Copyright © 2015 Elsevier Ltd. All rights reserved.

  19. An active DNA vaccine against infectious pancreatic necrosis virus (IPNV) with a different mode of action than fish rhabdovirus DNA vaccines.

    Science.gov (United States)

    Cuesta, A; Chaves-Pozo, E; de Las Heras, A I; Saint-Jean, S Rodríguez; Pérez-Prieto, S; Tafalla, C

    2010-04-26

    Although there are some commercial vaccines available against infectious pancreatic necrosis virus (IPNV), the disease still continues to be a major problem for aquaculture development worldwide. In the current work, we constructed a DNA vaccine against IPNV (pIPNV-PP) by cloning the long open reading frame of the polyprotein encoded by the viral RNA segment A. In vitro, the vaccine is properly translated giving the functional IPNV polyprotein since preVP2, VP2 and VP3 proteins were detected because of the VP4-protease cleavage. EPC cells transfected with the vaccine plasmid expressed the viral proteins and induced the expression of type I interferon (IFN)-induced Mx genes. Furthermore, IPNV synthesized proteins seemed to assemble in virus-like particles as evidenced by electron microscopy. In vivo, rainbow trout specimens were intramuscularly injected with the vaccine and expression of immune-relevant genes, the presence of neutralizing antibodies and effect on viral load was determined. The pIPNV-PP vaccine was expressed at the injection site and up-regulated MHC Ialpha, MHC IIalpha, type-I interferon (IFN), Mx, CD4 and CD8alpha gene expression in the muscle, head kidney or spleen, although to a much lower extent than the up-regulations observed in response to an effective DNA vaccine against viral hemorrhagic septicaemia virus (VHSV). However, the IPNV vaccine was also very effective in terms of acquired immunity since it elicited neutralizing antibodies (in 6 out of 8 trout fingerlings) and decreased 665-fold the viral load after IPNV infection. The effectiveness of this new IPNV DNA vaccine and its possible mechanism of action are discussed and compared to other viral vaccines.

  20. Improvement in efficacy of DNA vaccine encoding HIV-1 Vif by LIGHT gene adjuvant.

    Science.gov (United States)

    Du, Jiani; Wu, Xiaoyu; Long, Fengying; Wen, Jiejun; Hao, Wenli; Chen, Ran; Kong, Xiaobo; Qian, Min; Jiang, Wenzheng

    2013-02-01

    DNA vaccine can induce the prolonged immune responses against the encoded antigen with the appropriate adjuvant. To study the immunogenicity of the HIV-1 vif DNA vaccine in inducing the humoral and cellular immune responses and the immunoadjuvant effect of LIGHT, which is a member of TNF superfamily and can stimulate the proliferation of naïve T cells as a co-stimulatory molecule, DNA vaccine plasmid pcDNA-Vif was constructed by inserting HIV-1 vif gene into the downstream of CMV promoter in eukaryotic expression vector pcDNA3.1(+). In vitro expression of HIV-1 Vif in pcDNA-Vif-transfected HeLa cells was confirmed in transcriptional and protein level by RT-PCR and Western blot, respectively. After BALB/c mice were injected muscularly with DNA vaccines for three times, the specific immune responses were analyzed. The data showed that anti-Vif antibody response, Vif-specific T cell proliferation, and CTL activities were induced in the mice that were inoculated with HIV-1 vif DNA vaccine plasmid. Interestingly, stronger humoral and cellular immune responses were detected in mice that were immunized with plasmid pcDNA-Vif and pcDNA-LIGHT together compared to the single immunization with plasmid pcDNA-Vif alone. Together, the results of the study suggest that candidate HIV-1 DNA vaccine can elicit HIV-1 Vif-specific immune responses in mice and that LIGHT plays the role of immunoadjuvant in co-immunization with DNA vaccine.

  1. Interleukin-13 receptor α2 DNA prime boost vaccine induces tumor immunity in murine tumor models

    Directory of Open Access Journals (Sweden)

    Puri Raj K

    2010-11-01

    Full Text Available Abstract Background DNA vaccines represent an attractive approach for cancer treatment by inducing active T cell and B cell immune responses to tumor antigens. Previous studies have shown that interleukin-13 receptor α2 chain (IL-13Rα2, a tumor-associated antigen is a promising target for cancer immunotherapy as high levels of IL-13Rα2 are expressed on a variety of human tumors. To enhance the effectiveness of DNA vaccine, we used extracellular domain of IL-13Rα2 (ECDα2 as a protein-boost against murine tumor models. Methods We have developed murine models of tumors naturally expressing IL-13Rα2 (MCA304 sarcoma, 4T1 breast carcinoma and D5 melanoma tumors transfected with human IL-13Rα2 in syngeneic mice and examined the antitumor activity of DNA vaccine expressing IL-13Rα2 gene with or without ECDα2 protein mixed with CpG and IFA adjuvants as a boost vaccine. Results Mice receiving IL-13Rα2 DNA vaccine boosted with ECDα2 protein were superior in exhibiting inhibition of tumor growth, compared to mice receiving DNA vaccine alone, in both prophylactic and therapeutic vaccine settings. In addition, prime-boost vaccination significantly prolonged the survival of mice compared to DNA vaccine alone. Furthermore, ECDα2 booster vaccination increased IFN-γ production and CTL activity against tumor expressing IL-13Rα2. The immunohistochemical analysis showed the infiltration of CD4 and CD8 positive T cells and IFN-γ-induced chemokines (CXCL9 and CXCL10 in regressing tumors of immunized mice. Finally, the prime boost strategy was able to reduce immunosuppressive CD4+CD25+Foxp3+ regulatory T cells (Tregs in the spleen and tumor of vaccinated mice. Conclusion These results suggest that immunization with IL-13Rα2 DNA vaccine followed by ECDα2 boost mixed with CpG and IFA adjuvants inhibits tumor growth in T cell dependent manner. Thus our results show an enhancement of efficacy of IL-13Rα2 DNA vaccine with ECDα2 protein boost and offers an

  2. The safety of DNA vaccines%DNA 疫苗接种的安全性

    Institute of Scientific and Technical Information of China (English)

    靳彦文; 马清钧

    2001-01-01

    Three major issues have been raised with respect to the safety of DNA vaccines .These are the potential for inducing a transformational event, possible unexpected adverse consequences of the persistent expression of a foreign antigen and the formation of anti DNA antibodies. The review based on the relevent research articles give a summary of knowledge of DNA vaccines' safety published during past years.%DNA疫苗的安全性问题主要有三个方面:转入体内的外源DNA有可能整合到宿主细胞基因组上,使宿主细胞抑癌基因失活或癌基因活化,使宿主细胞转化成癌细胞;外源抗原持续表达产生的不良后果;质粒DNA诱导的自身免疫反应。本文综合近年来有关文献对DNA疫苗安全性的研究作一概括性介绍。

  3. Antiviral immunity in fish – functional analysis using DNA vaccination as a tool

    DEFF Research Database (Denmark)

    Lorenzen, Niels; Lorenzen, Ellen; Einer-Jensen, Katja

    2013-01-01

    CpG motifs in the plasmid backbone sequence might play a role, the viral G protein appears to have an inherent ability to stimulate innate immune mechanisms by receptors and pathways that still remain to be characterized in detail. Immunity to VHS in rainbow trout can be induced by DNA vaccination......In fish, DNA vaccines encoding the glycoproteins (G proteins) of the salmonid rhabdoviruses VHSV and IHNV have proved very efficient under experimental conditions. Nano-gram amounts of plasmid DNA can induce long-lasting protective immunity when delivered by intramuscular injection in rainbow trout...... fingerlings. Vaccination of fish at an early stage appears advantageous, since larger fish require higher doses of vaccine to be protected. Even in fish with an average size of 0.5 g at the time of vaccination, good protection can be obtained. Interestingly, immunity is established already a few days after...

  4. Use of DNA and recombinant canarypox viral (ALVAC) vectors for equine herpes virus vaccination.

    Science.gov (United States)

    Minke, J M; Fischer, L; Baudu, Ph; Guigal, P M; Sindle, T; Mumford, J A; Audonnet, J C

    2006-05-15

    In this study, experimental canarypox virus (ALVAC) and plasmid DNA recombinant vaccines expressing the gB, gC and gD glycoproteins of EHV-1 were assessed for their ability to protect conventional ponies against a respiratory challenge with EHV-1. In addition, potential means of enhancing serological responses in horses to ALVAC and DNA vaccination were explored. These included co-administration of the antigen with conventional adjuvants, complexation with DMRIE-DOPE and co-expression of the antigen along with equine GM-CSF. Groups of EHV primed ponies were vaccinated twice intra-muscularly with one dose of the appropriate test vaccine at an interval of 5 weeks. Two to 3 weeks after the second vaccination, ponies were infected intra-nasally with the virulent Ab4 strain of EHV-1 after which they were observed clinically and sampled for virological investigations. The results demonstrated that DNA and ALVAC vaccination markedly reduced virus excretion after challenge in terms of duration and magnitude, but failed to protect against cell-associated viremia. Noteworthy was the almost complete absence of virus excretion in the group of ponies vaccinated with ALVAC-EHV in the presence of Carbopol adjuvant or DNA plasmid formulated with aluminium phosphate. The administration of the DNA vaccine in the presence of GM-CSF and formulated in DMRIE-DOPE and of the ALVAC vaccine in the presence of Carbopol adjuvant significantly improved virus neutralising antibody responses to EHV-1. These findings indicate that DNA and ALVAC vaccination is a promising approach for the immunological control of EHV-1 infection, but that more research is needed to identify the immunodominant protective antigens of EHV-1 and their interaction with the equine immune system.

  5. Production and purification of plasmid DNA vaccines: is there scope for further innovation?

    Science.gov (United States)

    Xenopoulos, Alex; Pattnaik, Priyabrata

    2014-12-01

    The demand for plasmid DNA (pDNA) has vastly increased over the past decade in response to significant advances that have been made in its application for gene therapy and vaccine development. Plasmid DNA-based vaccines are experiencing a resurgence due to success with prime-boost immunization strategies. The challenge has always been poor productivity and delivery of pDNA. Plasmid DNA-based vaccines have traditionally required milligram scale of GMP-grade product for vaccination due to the relatively low efficacy and duration of gene expression. However, efforts to increase pDNA vaccine effectiveness are evolving in genetic manipulations of bacterial host, improvements in product recovery and innovative delivery methods. This review summarizes recent advances in large-scale pDNA vaccine manufacturing, ranging from upstream processing, downstream processing and formulation, as such information is usually not available to the scientific community. The article will highlight technology gaps and offer insight on further scope of innovation.

  6. Strategies to enhance immunogenicity of cDNA vaccine encoded antigens by modulation of antigen processing

    NARCIS (Netherlands)

    Platteel, Anouk C M; Marit de Groot, A; Andersen, Peter; Ovaa, Huib; Kloetzel, Peter M; Mishto, Michele; Sijts, Alice J A M

    2016-01-01

    Most vaccines are based on protective humoral responses while for intracellular pathogens CD8(+) T cells are regularly needed to provide protection. However, poor processing efficiency of antigens is often a limiting factor in CD8(+) T cell priming, hampering vaccine efficacy. The multistage cDNA va

  7. Induction of strain-transcending immunity against Plasmodium chabaudi adami malaria with a multiepitope DNA vaccine.

    Science.gov (United States)

    Scorza, T; Grubb, K; Smooker, P; Rainczuk, A; Proll, D; Spithill, T W

    2005-05-01

    A major goal of current malaria vaccine programs is to develop multivalent vaccines that will protect humans against the many heterologous malaria strains that circulate in endemic areas. We describe a multiepitope DNA vaccine, derived from a genomic Plasmodium chabaudi adami DS DNA expression library of 30,000 plasmids, which induces strain-transcending immunity in mice against challenge with P. c. adami DK. Segregation of this library and DNA sequence analysis identified vaccine subpools encoding open reading frames (ORFs)/peptides of >9 amino acids [aa] (the V9+ pool, 303 plasmids) and >50 aa (V50+ pool, 56 plasmids), respectively. The V9+ and V50+ plasmid vaccine subpools significantly cross-protected mice against heterologous P. c. adami DK challenge, and protection correlated with the induction of both specific gamma interferon production by splenic cells and opsonizing antibodies. Bioinformatic analysis showed that 22 of the V50+ ORFs were polypeptides conserved among three or more Plasmodium spp., 13 of which are predicted hypothetical proteins. Twenty-nine of these ORFs are orthologues of predicted Plasmodium falciparum sequences known to be expressed in the blood stage, suggesting that this vaccine pool encodes multiple blood-stage antigens. The results have implications for malaria vaccine design by providing proof-of-principle that significant strain-transcending immunity can be induced using multiepitope blood-stage DNA vaccines and suggest that both cellular responses and opsonizing antibodies are necessary for optimal protection against P. c. adami.

  8. Protection against Vibrio alginolyticus in crimson snapper Lutjanus erythropterus immunized with a DNA vaccine containing the ompW gene.

    Science.gov (United States)

    Cai, Shuang-Hu; Lu, Yi-Shan; Jian, Ji-Chang; Wang, Bei; Huang, Yu-Cong; Tang, Ju-Fen; Ding, Yu; Wu, Zao-He

    2013-09-24

    The outer membrane proteins of Vibrio alginolyticus play an important role in the virulence of the bacterium and are potential candidates for vaccine development. In the present study, the ompW gene was cloned, expressed and purified. A DNA vaccine was constructed by inserting the ompW gene into a pcDNA plasmid. Crimson snapper Lutjanus erythropterus (Bloch) were injected intramuscularly with the recombinant plasmid pcDNA-ompW. The expression of the DNA vaccine was detected in gill, head kidney, heart, liver, spleen and injection site muscle of crimson snapper by RT-PCR 7 and 28 d post-vaccination. The ELISA results demonstrated that the DNA vaccine produced an observable antibody response in all sera of the vaccinated fish. In addition, crimson snapper immunized with the DNA vaccine showed a relative percentage survival (RPS) of 92.53%, indicating effective protection against V. alginolyticus infection.

  9. Cysteine proteases as potential antigens in antiparasitic DNA vaccines

    DEFF Research Database (Denmark)

    Jørgensen, Louise von Gersdorff; Buchmann, Kurt

    2011-01-01

    En litteraturgennemgang af muligheder for at bruge cystein proteaser som antigener i antiparasitære vacciner.......En litteraturgennemgang af muligheder for at bruge cystein proteaser som antigener i antiparasitære vacciner....

  10. Multiple factors affect immunogenicity of DNA plasmid HIV vaccines in human clinical trials.

    Science.gov (United States)

    Jin, Xia; Morgan, Cecilia; Yu, Xuesong; DeRosa, Stephen; Tomaras, Georgia D; Montefiori, David C; Kublin, James; Corey, Larry; Keefer, Michael C

    2015-05-11

    Plasmid DNA vaccines have been licensed for use in domesticated animals because of their excellent immunogenicity, but none have yet been licensed for use in humans. Here we report a retrospective analysis of 1218 healthy human volunteers enrolled in 10 phase I clinical trials in which DNA plasmids encoding HIV antigens were administered. Elicited T-cell immune responses were quantified by validated intracellular cytokine staining (ICS) stimulated with HIV peptide pools. HIV-specific binding and neutralizing antibody activities were also analyzed using validated assays. Results showed that, in the absence of adjuvants and boosting with alternative vaccines, DNA vaccines elicited CD8+ and CD4+ T-cell responses in an average of 13.3% (95% CI: 9.8-17.8%) and 37.7% (95% CI: 31.9-43.8%) of vaccine recipients, respectively. Three vaccinations (vs. 2) improved the proportion of subjects with antigen-specific CD8+ responses (p=0.02), as did increased DNA dosage (p=0.007). Furthermore, female gender and participants having a lower body mass index were independently associated with higher CD4+ T-cell response rate (p=0.001 and p=0.008, respectively). These vaccines elicited minimal neutralizing and binding antibody responses. These findings of the immunogenicity of HIV DNA vaccines in humans can provide guidance for future clinical trials. Copyright © 2015 Elsevier Ltd. All rights reserved.

  11. Advances and challenges in the development of therapeutic DNA vaccines against hepatitis B virus infection.

    Science.gov (United States)

    Cova, Lucyna

    2014-01-01

    Despite the existence of an effective prophylactic vaccine, chronic hepatitis B virus (HBV) infection remains a major public health problem. Because very weak and functionally impaired virus-specific immune responses play a key role in the persistence of HBV infection, the stimulation of these responses appears to be of particular importance for virus clearance. In this regard DNA-based vaccination has emerged as novel, promising therapeutic approach for chronic hepatitis B. This review provides an update of preclinical studies in animal models (mouse, chimpanzee, duck, woodchuck), which evaluated the ability of DNA vaccines targeting hepadnaviral proteins to induce potent and sustained immune responses in naïve animals and to enhance virus clearance and break immune tolerance in chronic virus-carriers. Different strategies have been developed and evaluated in these models to optimize DNA vaccine including genetic adjuvants, combination with antiviral drugs, prime-boost regimens and plasmid delivery. The delivery of DNA by in vivo electroporation appears to be of particular interest for increase of vaccine potency in both small and large animal models. Based on the promising results generated in preclinical studies, first clinical trials of DNA vaccines have been initiated, although effective therapy of chronic hepatitis B awaits further improvements in vaccine efficacy.

  12. Assessing Linearity of the Parasite Varroa destructor DNA Amplification

    Directory of Open Access Journals (Sweden)

    ODAGIU Antonia

    2009-12-01

    Full Text Available The importance of honeybee products make of disease prevention and control in honeybees one of the mainconcerns of beekeepers in the world. The PCR – RT reaction represents an alternative for amplification performed inorder to realize the Varroa destructor O. genotypization, very important stage in haoneybee resistance to parasitedescription and also in management of the treatments. The linearity data is a very important parameter and very usefulin determination of the amplification of the parasite DNA and success of the genotypization process. The amplificationefficiency was very satisfactory, fact revealed by the value of the regression line y = - 2.3103 * 26.552 together withcoefficient of determination equal (r2 = 0.9691, meaning that more than 96% of the reaction efficiency may beexplained by the process liniarity. The implementation of the RT-PCR method was successful and it represents apremise for validation process evolution.

  13. Transcriptome Profiles Associated to VHSV Infection or DNA Vaccination in Turbot (Scophthalmus maximus)

    OpenAIRE

    Patricia Pereiro; Sonia Dios; Sebastián Boltaña; Julio Coll; Amparo Estepa; Simon Mackenzie; Beatriz Novoa; Antonio Figueras

    2014-01-01

    DNA vaccines encoding the viral G glycoprotein show the most successful protection capability against fish rhabdoviruses. Nowadays, the molecular mechanisms underlying the protective response remain still poorly understood. With the aim of shedding light on the protection conferred by the DNA vaccines based in the G glycoprotein of viral haemorrhagic septicaemia virus (VHSV) in turbot (Scophthalmus maximus) we have used a specific microarray highly enriched in antiviral sequences to carry out...

  14. Construction and Nonclinical Testing of a Puumala Virus Synthetic M Gene-Based DNA Vaccine

    Science.gov (United States)

    2012-12-12

    DNA vaccine constructsf Amino acid positiona PUUV M segment amino acid DNA vaccine amino acid DTK/ Ufa -97b K27c P360d Hallnas B1e PUU- M(x22) PUU- M...G 1097 S S S S L L S a M gene segment open reading frame amino acid position. b DTK/ Ufa -97, PUUV strain DTK/ Ufa (GenBank accession no. BAF49040). c

  15. Evaluation of cellular responses for a chimeric HBsAg-HCV core DNA vaccine in BALB/c mice

    Directory of Open Access Journals (Sweden)

    Maryam Yazdanian

    2015-01-01

    Conclusion: Fusion of HBsAg to HCVcp in the context of a DNA vaccine modality could augment Th1-oriented cellular and CTL responses toward a protective epitope, comparable to that of HCVcp (subunit HCV vaccine immunization.

  16. Distribution and expression in vitro and in vivo of DNA vaccine against lymphocystis disease virus in Japanese flounder (Paralichthys olivaceus)

    Institute of Scientific and Technical Information of China (English)

    郑风荣; 孙修勤; 刘洪展; 吴兴安; 钟楠; 王波; 周国栋

    2010-01-01

    Lymphocystis disease,caused by the lymphocystis disease virus (LCDV),is a significant worldwide problem in fish industry causing substantial economic losses.In this study,we aimed to develop the DNA vaccine against LCDV,using DNA vaccination technology.We evaluated plasmid pEGFP-N2-LCDV1.3 kb as a DNA vaccine candidate.The plasmid DNA was transiently expressed after liposome transfection into the eukaryotic COS 7 cell line.The distribution and expression of the DNA vaccine (pEGFP-N2-LCDV1.3kb) were also ana...

  17. Immunogenicity and protective efficacy study using combination of four tuberculosis DNA vaccines

    Institute of Scientific and Technical Information of China (English)

    蔡宏; 田霞; 呼西旦; 潘怡; 李国利; 庄玉辉; 朱玉贤

    2003-01-01

    Immune response and protective efficacy for the combination of four tuberculosis DNA vaccines were evaluated in this study. We obtained 1:200 antibody titers against Ag85B 21d after mice were vaccinated for the first time by four recombinant eukaryotic expression vectors containing coding sequences for Ag85B, MPT-64, MPT-63 and ESAT-6. The titers of Ag85B were elevated to 1:102400 after the second injection and decreased to 1:12800 after the third injection. Antibody titers for MPT-64 and MPT-63 reached 1:25600 21 d after the first vaccination, and were then decreased following the second and third injections. No antigen-specific antibody titer against ESAT-6 was detected in sera harvested from immunized mice at any time. These DNA vaccines evoked specific IFN-λ responses in the spleens of vaccinated mice as well. When challenged with M. tuberculosis H37Rv, we found that the lungs of the vaccinated mice produced 99.8% less bacterial counts than that of the empty-vector control group and the bacterial counts were also significantly less than that of the BCG group. Histopathological analyses showed that the lungs of vaccinated mice produced no obvious caseation while over 50%-70% of the pulmonary parenchyma tissue produced central caseation in the vector control group. Our results indicated that the combination of four tuberculosis DNA vaccines may generate high levels of immune responses and result in better animal protection.

  18. Alphavirus vectors: applications for DNA vaccine production and gene expression.

    Science.gov (United States)

    Lundstrom, K

    2000-01-01

    Replication-deficient alphavirus vectors have been developed for efficient high-level transgene expression. The broad host range of alphaviruses has allowed infection of a wide variety of mammalian cell lines and primary cultures. Particularly, G protein-coupled receptors have been expressed at high levels and subjected to binding and functional studies. Expression in suspension cultures has greatly facilitated production of large quantities of recombinant proteins for structural studies. Injection of recombinant alphavirus vectors into rodent brain resulted in local reporter gene expression. Highly neuron-specific expression was obtained in hippocampal slice cultures in vivo. Additionally, preliminary studies in animal models suggest that alphavirus vectors can be attractive candidates for gene therapy applications. Traditionally alphavirus vectors, either attenuated strains or replication-deficient particles, have been used to elicit efficient immune responses in animals. Recently, the application of alphaviruses has been extended to naked nucleic acids. Injection of DNA as well as RNA vectors has demonstrated efficient antigen production. In many cases, protection against lethal challenges has been obtained after immunization with alphavirus particles or nucleic acid vectors. Alphavirus vectors can therefore be considered as potentially promising vectors for vaccine production.

  19. Design and Construction of Shrimp Antiviral DNA Vaccines Expressing Long and Short Hairpins for Protection by RNA Interference.

    Science.gov (United States)

    Chaudhari, Aparna; Pathakota, Gireesh-Babu; Annam, Pavan-Kumar

    2016-01-01

    DNA vaccines present the aquaculture industry with an effective and economically viable method of controlling viral pathogens that drastically affect productivity. Since specific immune response is rudimentary in invertebrates, the presence of RNA interference (RNAi) pathway in shrimps provides a promising new approach to vaccination. Plasmid DNA vaccines that express short or long double stranded RNA in vivo have shown protection against viral diseases. The design, construction and considerations for preparing such vaccines are discussed.

  20. Early life DNA vaccination with the H gene of Canine distemper virus induces robust protection against distemper

    DEFF Research Database (Denmark)

    Jensen, Trine Hammer; Nielsen, Line; Aasted, Bent;

    2009-01-01

    Young mink kits (n = 8)were vaccinated withDNA plasmids encoding the viral haemagglutinin protein (H) of a vaccine strain of Canine distemper virus (CDV). Virus neutralising (VN) antibodieswere induced after 2 immunisations and after the third immunisation all kits had high VN antibody titres...... demonstrate that early life DNA vaccination with the H gene of a CDV vaccine strain induced robust protective immunity against a recent wild type CDV....

  1. Effective Protection Induced by a Monovalent DNA Vaccine against Dengue Virus (DV Serotype 1 and a Bivalent DNA Vaccine against DV1 and DV2 in Mice

    Directory of Open Access Journals (Sweden)

    Xiaoyan Zheng

    2017-05-01

    Full Text Available Dengue virus (DV is the causal pathogen of dengue fever, which is one of the most rapidly spread mosquito-borne disease worldwide and has become a severe public health problem. Currently, there is no specific treatment for dengue; thus, a vaccine would be an effective countermeasure to reduce the morbidity and mortality. Although, the chimeric Yellow fever dengue tetravalent vaccine has been approved in some countries, it is still necessary to develop safer, more effective, and less costly vaccines. In this study, a DNA vaccine candidate pVAX1-D1ME expressing the prME protein of DV1 was inoculated in BALB/c mice via intramuscular injection or electroporation, and the immunogenicity and protection were evaluated. Compared with traditional intramuscular injection, administration with 50 μg pVAX1-D1ME via electroporation with three immunizations induced persistent humoral and cellular immune responses and effectively protected mice against lethal DV1 challenge. In addition, immunization with a bivalent vaccine consisting of pVAX1-D1ME and pVAX1-D2ME via electroporation generated a balanced IgG response and neutralizing antibodies against DV1 and DV2 and could protect mice from lethal challenge with DV1 and DV2. This study sheds new light on developing a dengue tetravalent DNA vaccine.

  2. Protection of Mice with a Divalent Tuberculosis DNA Vaccine Encoding Antigens Ag85B and MPT64

    Institute of Scientific and Technical Information of China (English)

    Xia TIAN; Hong CAI; Yu-Xian ZHU

    2004-01-01

    DNA vaccine may be a promising tool for controlling tuberculosis development. However,vaccines encoding single antigens of mycobacterium did not produce protective effect as BCG did. In the present study, we evaluated the immunogenicity and protective efficacy of a divalent DNA vaccine encoding two immunodominant antigens Ag85B and MPT64 of Mycobacterium tuberculosis. We found that both humoral and Th1-type (high IFN-γ, low IL-4) cellular responses obtained from the divalent DNA vaccine group were significantly higher than that conferred by BCG. RT-PCR results showed that antigens were expressed differentially in various organs in divalent DNA vaccine group. The survival rate for mice treated with the divalent DNA vaccine after challenging with high doses of virulent M. tuberculosis H37Rv was significantly higher than that of the BCG group or any of the single DNA vaccine group. Significant differences were also found between the single and divalent DNA vaccinated mice in terms of body, spleen and lung weight. Bacterial loading decreased about 2000-fold in lungs and about 100-fold in spleens of divalent DNA vaccinated mice when compared with that of the control group. We conclude that our divalent DNA vaccine may be a better choice for controlling tuberculosis disease in animals.

  3. THE PROTECTIVE MECHANISMS INDUCED BY A FISH RHABDOVIRUS DNA-VACCINE DEPENDS ON TTEMPERATURE

    DEFF Research Database (Denmark)

    Lorenzen, Niels; Lorenzen, Ellen; Einer-Jensen, Katja;

    DNA-vaccines encoding the viral glycoproteins of viral haemorrhagic septicaemia virus (VHSV) and infectious haematopoietic necrosis virus (IHNV) have proved highly efficient in rainbow trout (Oncorhynchus mykiss) under experimental conditions. In the early phase following vaccination, innate cross-protective...... mechanisms are dominating but the protection becomes highly specific within 3-4 weeks at 12-15C. Temperature is known as an important external parameter affecting the immune response in fish and the present study aimed at characterizing temperature effects on the immune response to a VHS DNA vaccine....... Rainbow trout fingerlings acclimated at 5°C, 10°C or 15C, were given an intramuscular injection of 1g purified plasmid DNA and challenged with virulent VHSV 9 or 36-40 days later. The vaccine protected the fish well at all three temperatures, however the non-specific mechanisms lasted for a longer...

  4. The Protective Mechanisms Induced by a DNA Vaccine in Fish Depend on Temperature

    DEFF Research Database (Denmark)

    Lorenzen, Ellen; Einer-Jensen, Katja; Rasmussen, Jesper Skou;

    2011-01-01

    In veterinary vaccinology, DNA-vaccines encoding the viral glycoproteins of viral haemorrhagic septicaemia virus (VHSV) and infectious haematopoietic necrosis virus (IHNV) have proved highly efficient in fish under experimental conditions. In the early phase following vaccination, innate cross-protective...... mechanisms are dominating but the protection becomes highly specific within 3–4 weeks at 12–15 C. Temperature is known as an important external parameter affecting the immune response in fish and the present study aimed at characterizing temperature effects on the immune response to a VHS DNA vaccine....... Rainbow trout fingerlings acclimated at 5, 10 or 15 C, were given an intramuscular injection of 1 lg purified plasmid DNA and challenged with virulent VHSV 9 or 36–40 days later. The vaccine protected the fish well at all three temperatures, however the non-specific mechanisms lasted for a longer period...

  5. Increased humoral immunity by DNA vaccination using an alpha-tocopherol-based adjuvant

    DEFF Research Database (Denmark)

    Karlsson, Ingrid; Borggren, Marie; Nielsen, Jens

    2017-01-01

    DNA vaccines induce broad immunity, which involves both humoral and strong cellular immunity, and can be rapidly designed for novel or evolving pathogens such as influenza. However, the humoral immunogenicity in humans and higher animals has been suboptimal compared to that of traditional vaccine......). The animals received two intracutaneous immunizations spaced 3 weeks apart. When combined with Diluvac Forte® or the emulsion containing alpha-tocopherol, the DNA vaccine induced a more potent and balanced immunoglobulin G (IgG)1 and IgG2c response, and both IgG subclass responses were significantly enhanced...... constituent alpha-tocopherol plays an important role in this immunogenicity. This induction of a potent and balanced humoral response without impairment of cellular immunity constitutes an important advancement toward effective DNA vaccines....

  6. Plasmid DNA initiates replication of yellow fever vaccine in vitro and elicits virus-specific immune response in mice

    Energy Technology Data Exchange (ETDEWEB)

    Tretyakova, Irina; Nickols, Brian; Hidajat, Rachmat [Medigen, Inc., 8420 Gas House Pike, Suite S, Frederick, MD 21701 (United States); Jokinen, Jenny; Lukashevich, Igor S. [Department of Pharmacology and Toxicology, School of Medicine, Center for Predictive Medicine and Emerging Infectious Diseases, University of Louisville, Louisville, KY (United States); Pushko, Peter, E-mail: ppushko@medigen-usa.com [Medigen, Inc., 8420 Gas House Pike, Suite S, Frederick, MD 21701 (United States)

    2014-11-15

    Yellow fever (YF) causes an acute hemorrhagic fever disease in tropical Africa and Latin America. To develop a novel experimental YF vaccine, we applied iDNA infectious clone technology. The iDNA represents plasmid that encodes the full-length RNA genome of 17D vaccine downstream from a cytomegalovirus (CMV) promoter. The vaccine was designed to transcribe the full-length viral RNA and to launch 17D vaccine virus in vitro and in vivo. Transfection with 10 ng of iDNA plasmid was sufficient to start replication of vaccine virus in vitro. Safety of the parental 17D and iDNA-derived 17D viruses was confirmed in AG129 mice deficient in receptors for IFN-α/β/γ. Finally, direct vaccination of BALB/c mice with a single 20 μg dose of iDNA plasmid resulted in seroconversion and elicitation of virus-specific neutralizing antibodies in animals. We conclude that iDNA immunization approach combines characteristics of DNA and attenuated vaccines and represents a promising vaccination strategy for YF. - Highlights: • The iDNA{sup ®} platform combines advantages of DNA and live attenuated vaccines. • Yellow fever (YF) 17D vaccine was launched from iDNA plasmid in vitro and in vivo. • Safety of iDNA-generated 17D virus was confirmed in AG129 mice. • BALB/c mice seroconverted after a single-dose vaccination with iDNA. • YF virus-neutralizing response was elicited in iDNA-vaccinated mice.

  7. Transcriptome profiles associated to VHSV infection or DNA vaccination in turbot (Scophthalmus maximus.

    Directory of Open Access Journals (Sweden)

    Patricia Pereiro

    Full Text Available DNA vaccines encoding the viral G glycoprotein show the most successful protection capability against fish rhabdoviruses. Nowadays, the molecular mechanisms underlying the protective response remain still poorly understood. With the aim of shedding light on the protection conferred by the DNA vaccines based in the G glycoprotein of viral haemorrhagic septicaemia virus (VHSV in turbot (Scophthalmus maximus we have used a specific microarray highly enriched in antiviral sequences to carry out the transcriptomic study associated to VHSV DNA vaccination/infection. The differential gene expression pattern in response to empty plasmid (pMCV1.4 and DNA vaccine (pMCV1.4-G860 intramuscular administration with regard to non-stimulated turbot was analyzed in head kidney at 8, 24 and 72 hours post-vaccination. Moreover, the effect of VHSV infection one month after immunization was also analyzed in vaccinated and non-vaccinated fish at the same time points. Genes implicated in the Toll-like receptor signalling pathway, IFN inducible/regulatory proteins, numerous sequences implicated in apoptosis and cytotoxic pathways, MHC class I antigens, as well as complement and coagulation cascades among others were analyzed in the different experimental groups. Fish receiving the pMCV1.4-G860 vaccine showed transcriptomic patterns very different to the ones observed in pMCV1.4-injected turbot after 72 h. On the other hand, VHSV challenge in vaccinated and non-vaccinated turbot induced a highly different response at the transcriptome level, indicating a very relevant role of the acquired immunity in vaccinated fish able to alter the typical innate immune response profile observed in non-vaccinated individuals. This exhaustive transcriptome study will serve as a complete overview for a better understanding of the crosstalk between the innate and adaptive immune response in fish after viral infection/vaccination. Moreover, it provides interesting clues about molecules

  8. Interleukin-12 as a Genetic Adjuvant Enhances Hepatitis C Virus NS3 DNA Vaccine Immunogenicity

    Institute of Scientific and Technical Information of China (English)

    Malihe Naderi; Atefeh Saeedi; Abdolvahab Moradi; Mishar Kleshadi; Mohammad Reza Zolfaghari; Ali Gorji; Amir Ghaemi

    2013-01-01

    Hepatitis C virus (HCV) chronic infection is a worldwide health problem,and numerous efforts have been invested to develop novel vaccines.An efficient vaccine requires broad immune response induction against viral proteins.To achieve this goal,we constructed a DNA vaccine expressing nonstructural 3 (NS3) gene (pcDNA3.1-HCV-NS3) and assessed the immune response in C57BL/6 mice.In this study,the NS3 gene was amplified with a nested-reverse transcriptase-polymerase chain reaction (RT-PCR) method using sera of HCV-infected patients with genotype 1 a.The resulting NS3 gene was subcloned into a pcDNA3.1 eukaryotic expression vector,and gene expression was detected by western blot.The resultant DNA vaccine was co-administered with interleukin-12 (IL-12) as an adjuvant to female C57BL/6 mice.After the final immunizations,lymphocyte proliferation,cytotoxicity,and cytokine levels were assessed to measure immune responses.Our data suggest that co-administration of HCV NS3 DNA vaccine with IL-12 induces production of significant levels of both IL-4 and interferon (IFN)-γ (p<0.05).Cytotoxicity and lymphocyte proliferation responses of vaccinated mice were significantly increased compared to control (p<0.05).Collectively,our results demonstrated that co-administration of HCV NS3 and IL-12 displayed strong immunogenicity in a murine model.

  9. PCR-based detection of a rare linear DNA in cell culture

    Directory of Open Access Journals (Sweden)

    Saveliev Sergei V.

    2002-01-01

    Full Text Available The described method allows for detection of rare linear DNA fragments generated during genomic deletions. The predicted limit of the detection is one DNA molecule per 107 or more cells. The method is based on anchor PCR and involves gel separation of the linear DNA fragment and chromosomal DNA before amplification. The detailed chemical structure of the ends of the linear DNA can be defined with the use of additional PCR-based protocols. The method was applied to study the short-lived linear DNA generated during programmed genomic deletions in a ciliate. It can be useful in studies of spontaneous DNA deletions in cell culture or for tracking intracellular modifications at the ends of transfected DNA during gene therapy trials.

  10. TAA Polyepitope DNA-Based Vaccines: A Potential Tool for Cancer Therapy

    Directory of Open Access Journals (Sweden)

    Roberto Bei

    2010-01-01

    Full Text Available DNA-based cancer vaccines represent an attractive strategy for inducing immunity to tumor associated antigens (TAAs in cancer patients. The demonstration that the delivery of a recombinant plasmid encoding epitopes can lead to epitope production, processing, and presentation to CD8+ T-lymphocytes, and the advantage of using a single DNA construct encoding multiple epitopes of one or more TAAs to elicit a broad spectrum of cytotoxic T-lymphocytes has encouraged the development of a variety of strategies aimed at increasing immunogenicity of TAA polyepitope DNA-based vaccines. The polyepitope DNA-based cancer vaccine approach can (a circumvent the variability of peptide presentation by tumor cells, (b allow the introduction in the plasmid construct of multiple immunogenic epitopes including heteroclitic epitope versions, and (c permit to enroll patients with different major histocompatibility complex (MHC haplotypes. This review will discuss the rationale for using the TAA polyepitope DNA-based vaccination strategy and recent results corroborating the usefulness of DNA encoding polyepitope vaccines as a potential tool for cancer therapy.

  11. Advances in host and vector development for the production of plasmid DNA vaccines.

    Science.gov (United States)

    Mairhofer, Juergen; Lara, Alvaro R

    2014-01-01

    Recent developments in DNA vaccine research provide a new momentum for this rather young and potentially disruptive technology. Gene-based vaccines are capable of eliciting protective immunity in humans to persistent intracellular pathogens, such as HIV, malaria, and tuberculosis, for which the conventional vaccine technologies have failed so far. The recent identification and characterization of genes coding for tumor antigens has stimulated the development of DNA-based antigen-specific cancer vaccines. Although most academic researchers consider the production of reasonable amounts of plasmid DNA (pDNA) for immunological studies relatively easy to solve, problems often arise during this first phase of production. In this chapter we review the current state of the art of pDNA production at small (shake flasks) and mid-scales (lab-scale bioreactor fermentations) and address new trends in vector design and strain engineering. We will guide the reader through the different stages of process design starting from choosing the most appropriate plasmid backbone, choosing the right Escherichia coli (E. coli) strain for production, and cultivation media and scale-up issues. In addition, we will address some points concerning the safety and potency of the produced plasmids, with special focus on producing antibiotic resistance-free plasmids. The main goal of this chapter is to make immunologists aware of the fact that production of the pDNA vaccine has to be performed with as much as attention and care as the rest of their research.

  12. Impaired autoimmune T helper 17 cell responses following DNA vaccination against rat experimental autoimmune encephalomyelitis.

    Directory of Open Access Journals (Sweden)

    Asa Andersson

    Full Text Available BACKGROUND: We have previously shown that vaccination with DNA encoding the encephalitogenic peptide myelin oligodendrocyte glycoprotein (MOG(91-108 (pMOG suppresses MOG(91-108-induced rat Experimental Autoimmune Encephalomyelitis (EAE, a model for human Multiple Sclerosis (MS. The suppressive effect of pMOG is dependent on inclusion of CpG DNA in the plasmid backbone and is associated with early induction of Interferon (IFN-beta. PRINCIPAL FINDINGS: In this study we examined the mechanisms underlying pMOG-induced protection. We found that in the DNA vaccinated cohort proinflammatory Interleukin (IL-17 and IL-21 responses were dramatically reduced compared to in the control group, but that the expression of Foxp3 and Tumor Growth Factor (TGF-beta1, which are associated with regulatory T cells, was not enhanced. Moreover, genes associated with Type I IFNs were upregulated. To delineate the role of IFN-beta in the protective mechanism we employed short interfering RNA (siRNA to IFN-beta in the DNA vaccine. SiRNA to IFN-beta completely abrogated the protective effects of the vaccine, demonstrating that a local early elaboration of IFN-beta is important for EAE protection. IL-17 responses comparable to those in control rats developed in rats injected with the IFN-beta-silencing DNA vaccine. CONCLUSIONS: We herein demonstrate that DNA vaccination protects from proinflammatory Th17 cell responses during induction of EAE. The mechanism involves IFN-beta as IL-17 responses are rescued by silencing of IFN-beta during DNA vaccination.

  13. Boosting of DNA Vaccine-Elicited Gamma Interferon Responses in Humans by Exposure to Malaria Parasites

    Science.gov (United States)

    2004-12-11

    McClure, J. M. McNicholl, B. Moss, and H. L. Robinson. 2001. Control of a mucosal challenge and prevention of AIDS by a multiprotein DNA/ MVA vaccine...and H. M. McClure. 1999. Neutralizing antibody-independent con- tainment of immunodeficiency virus challenges by DNA priming and recom- binant pox virus

  14. Canine distemper virus DNA vaccination of mink can overcome interference by maternal antibodies.

    Science.gov (United States)

    Jensen, Trine Hammer; Nielsen, Line; Aasted, Bent; Pertoldi, Cino; Blixenkrone-Møller, Merete

    2015-03-10

    Canine distemper virus (CDV) is highly contagious and can cause severe disease against which conventional live vaccines are ineffective in the presence of maternal antibodies. Vaccination in the presences of maternal antibodies was challenged by vaccination of 5 days old and 3 weeks old mink kits with CDV DNA vaccines. Virus neutralising (VN) antibody responses were induced in mink kits vaccinated with a plasmid encoding the haemaglutinin protein (H) of CDV (n=5, pCDV-H) or a combination of the H, fusion (F) and nucleoprotein (N) of CDV (n=5, pCDV-HFN). These DNA vaccinated kits were protected against virulent experimental infection with field strains of CDV. The pCDV-H was more efficient in inducing protective immunity in the presence of maternal antibodies compared to the pCDV-HFN. The results show that DNA vaccination with the pCDV-H or pCDV-HFN (n=4) only given once at 5 days of age induces virus specific immune response in neonatal mink and protection against virulent CDV exposure later in life.

  15. The Effect of Bovine IFN-α on the Immune Response in Guinea Pigs Vaccinated with DNA Vaccine of Foot-and-Mouth Disease Virus

    Institute of Scientific and Technical Information of China (English)

    Hui-Chen GUO; Zai-Xin LIU; Shi-Qi SUN; Qing-Wen LENG; Dong LI; Xiang-Tao LIU; Qing-Ge XIE

    2004-01-01

    In this study, we constructed recombinant plasmid pcDNA3.1/P 12X3C3D including P 1, 2A,3C, 3D and part of 2B gene of FMDV and pcDNA3.1/IFN containing the gene encoding bovine IFN-α. We inoculated the DNA vaccine pcDNA3.1/P 12X3C3D with or without pcDNA3.1/IFN to evaluate the efficiency of this DNA vaccine and the immunogenicity of DNA vaccine enhanced by the co-delivery with pcDNA3.1/IFN. After two times of vaccination with DNA vaccine, all of guinea pigs were challenged with 103 ID50 FMDV type O. Anti-FMDV antibody levels were detected by ELISA and T lymphocyte proliferation response was tested by MTT assay. The result shows that guinea pigs inoculated by pcDNA3. 1/P 12X3C3D alone or with pcDNA3.1/IFN generated specific antibodies and induced an FMDV-specific T lymphocyte proliferation response. FMDV challenge tests showed that one in four guinea pigs immunized by pcDNA3.1/P 12X3C3D with pcDNA3.1/IFN was protected from the FMDV serotype O infection. This result indicated that the efficiency of the DNA vaccine was enhanced by co-delivery with pcDNA3.1/IFN. However, the protection rate was considerably lower than that immunized with conventional FMD vaccine.

  16. Testing the efficacy of a multi-component DNA-prime/DNA-boost vaccine against Trypanosoma cruzi infection in dogs.

    Directory of Open Access Journals (Sweden)

    José E Aparicio-Burgos

    Full Text Available BACKGROUND: Trypanosoma cruzi, the etiologic agent of Chagas Disease, is a major vector borne health problem in Latin America and an emerging infectious disease in the United States. METHODS: We tested the efficacy of a multi-component DNA-prime/DNA-boost vaccine (TcVac1 against experimental T. cruzi infection in a canine model. Dogs were immunized with antigen-encoding plasmids and cytokine adjuvants, and two weeks after the last immunization, challenged with T. cruzi trypomastigotes. We measured antibody responses by ELISA and haemagglutination assay, parasitemia and infectivity to triatomines by xenodiagnosis, and performed electrocardiography and histology to assess myocardial damage and tissue pathology. RESULTS: Vaccination with TcVac1 elicited parasite-and antigen-specific IgM and IgG (IgG2>IgG1 responses. Upon challenge infection, TcVac1-vaccinated dogs, as compared to non-vaccinated controls dogs, responded to T. cruzi with a rapid expansion of antibody response, moderately enhanced CD8(+ T cell proliferation and IFN-γ production, and suppression of phagocytes' activity evidenced by decreased myeloperoxidase and nitrite levels. Subsequently, vaccinated dogs controlled the acute parasitemia by day 37 pi (44 dpi in non-vaccinated dogs, and exhibited a moderate decline in infectivity to triatomines. TcVac1-immunized dogs did not control the myocardial parasite burden and electrocardiographic and histopatholgic cardiac alterations that are the hallmarks of acute Chagas disease. During the chronic stage, TcVac1-vaccinated dogs exhibited a moderate decline in cardiac alterations determined by EKG and anatomo-/histo-pathological analysis while chronically-infected/non-vaccinated dogs continued to exhibit severe EKG alterations. CONCLUSIONS: Overall, these results demonstrated that TcVac1 provided a partial resistance to T. cruzi infection and Chagas disease, and provide an impetus to improve the vaccination strategy against Chagas disease.

  17. Gene-gun DNA vaccination aggravates respiratory syncytial virus-induced pneumonitis

    DEFF Research Database (Denmark)

    Bartholdy, Christina; Olszewska, Wieslawa; Stryhn, Anette

    2004-01-01

    elicited with recombinant vaccinia virus expressing the complete RSV M2 protein, but stronger than those induced by a similar DNA construct without the beta2m gene. DNA vaccination led to enhanced pulmonary disease after RSV challenge, with increased weight loss and cell recruitment to the lung. Depletion......A CD8+ T-cell memory response to respiratory syncytial virus (RSV) was generated by using a DNA vaccine construct encoding the dominant Kd-restricted epitope from the viral transcription anti-terminator protein M2 (M2(82-90)), linked covalently to human beta2-microglobulin (beta2m). Cutaneous gene......+ T-cell responses were not induced. Thus, in addition to specific CD8+ T cell-mediated immunopathology, gene-gun DNA vaccination causes non-specific enhancement of RSV disease without affecting virus clearance....

  18. Codon-optimized filovirus DNA vaccines delivered by intramuscular electroporation protect cynomolgus macaques from lethal Ebola and Marburg virus challenges.

    Science.gov (United States)

    Grant-Klein, Rebecca J; Altamura, Louis A; Badger, Catherine V; Bounds, Callie E; Van Deusen, Nicole M; Kwilas, Steven A; Vu, Hong A; Warfield, Kelly L; Hooper, Jay W; Hannaman, Drew; Dupuy, Lesley C; Schmaljohn, Connie S

    2015-01-01

    Cynomolgus macaques were vaccinated by intramuscular electroporation with DNA plasmids expressing codon-optimized glycoprotein (GP) genes of Ebola virus (EBOV) or Marburg virus (MARV) or a combination of codon-optimized GP DNA vaccines for EBOV, MARV, Sudan virus and Ravn virus. When measured by ELISA, the individual vaccines elicited slightly higher IgG responses to EBOV or MARV than did the combination vaccines. No significant differences in immune responses of macaques given the individual or combination vaccines were measured by pseudovirion neutralization or IFN-γ ELISpot assays. Both the MARV and mixed vaccines were able to protect macaques from lethal MARV challenge (5/6 vs. 6/6). In contrast, a greater proportion of macaques vaccinated with the EBOV vaccine survived lethal EBOV challenge in comparison to those that received the mixed vaccine (5/6 vs. 1/6). EBOV challenge survivors had significantly higher pre-challenge neutralizing antibody titers than those that succumbed.

  19. Non-cytolytic antigen clearance in DNA-vaccinated mice with electropotation

    Institute of Scientific and Technical Information of China (English)

    Jin-liang PENG; Yong-gang ZHAO; Jun-hua MAI; Wen-ka PANG; Wei GUO; Guang-ming CHEN; Guo-yu MO; Gui-rong RAO; Yu-hong XU

    2007-01-01

    Aim: To explore the potential of electroporation (EP)-mediated hepatitis B virus (HBV) DNA vaccination for the treatment of chronic HBV infection. Methods: BALB/c mice were vaccinated with HBV DNA vaccine encoding for the HBV preS2-S antigen, combined with or without EP. HBV surface antigen expression plasmid was administered into mice liver via a hydrodynamic injection to mimic HBV infection. The clearance of antigen in the serum and liver was detected by ELISA assay and immunohistochemical staining. The histopathology of the liver tissues was examined by HE staining and serum alanine aminotransferase assay.Results: The immunogenicity ofHBV DNA vaccine encoding for the HBV preS2-S antigen can be improved by EP-mediated vaccine delivery. The elicited immune responses can indeed reduce the expression of HBV surface antigen (HBsAg) in hepatocytes of the mouse model that was transfected to express HBsAg using the hydrodynamic injection method. The antigen clearance process did not cause significant toxicity to liver tissue, suggesting a non-cytolytic mechanism. Conclusion: The EP-aided DNA vaccination may have potential in mediating viral clearance in chronic hepatitis B patients.

  20. Pilot study of p62 DNA vaccine in dogs with mammary tumors.

    Science.gov (United States)

    Gabai, Vladimir; Venanzi, Franco M; Bagashova, Elena; Rud, Oksana; Mariotti, Francesca; Vullo, Cecilia; Catone, Giuseppe; Sherman, Michael Y; Concetti, Antonio; Chursov, Andrey; Latanova, Anastasia; Shcherbinina, Vita; Shifrin, Victor; Shneider, Alexander

    2014-12-30

    Our previous data demonstrated profound anti-tumor and anti-metastatic effects of p62 (sqstm1) DNA vaccine in rodents with various types of transplantable tumors. Testing anti-cancer medicine in dogs as an intermediary step of translational research program provides two major benefits. First, clinical data collected in target animals is required for FDA/USDA approval as a veterinary anti-cancer drug or vaccine. It is noteworthy that the veterinary community is in need of novel medicine for the prevention and treatment of canine and feline cancers. The second more important benefit of testing anti-cancer vaccines in dogs is that spontaneous tumors in dogs may provide invaluable information for human trials. Here, we evaluated the effect(s) of p62 DNA vaccine on mammary tumors of dogs. We found that p62 DNA vaccine administered i.m. decreased or stabilized growth of locally advanced lesions in absence of its overall toxic effects. The observed antitumor activity was associated with lymphocyte infiltration and tumor encapsulation via fibrotic reaction. This data justifies both human clinical trials and veterinary application of p62 DNA vaccine.

  1. Research Progress of DNA Vaccine%DNA疫苗的研究进展

    Institute of Scientific and Technical Information of China (English)

    杨海; 王芳宇

    2013-01-01

    DNA疫苗是在分子生物学技术基础上发展起来的第3代新型疫苗,已体现出其竞争优势和应用潜能.同传统的疫苗相比,DNA疫苗具有免疫效果好、生产成本低、临床应用方便等优点,但同样存在安全性的担忧.文章回顾了DNA疫苗的发展简史,阐述了DNA疫苗的免疫机理,分析了DNA疫苗研究现状,并对DNA疫苗的安全性提出了自己的观点与看法,旨在为DNA疫苗研究提供参考.%DNA vaccine was the third generation of vaccine, and had reflected the competitive advantages and application potential in the past. DNA vaccine was developed on the basis of molecular biology technologies. Compared with traditional vaccines, it had more advantages,such as good immune effect, low production cost, and convenient for the clinical application, but it also could be found safety concerns. To provide the references for DNA vaccine researchers, the DNA vaccine history, immune mechanism and research status quo were summarized, and the viewpoint about its security was presented in present paper.

  2. Recent advances in design of immunogenic and effective naked DNA vaccines against cancer.

    Science.gov (United States)

    Fioretti, Daniela; Iurescia, Sandra; Rinaldi, Monica

    2014-01-01

    A variety of clinical trials for vaccines against cancer have provided evidence that DNA vaccines are well tolerated and have an excellent safety profile. DNA vaccines require much improvement to make them sufficiently effective against cancer in the clinic. Nowadays, it is clear that an increased antigen expression correlates with improved immunogenicity and it is critical to vaccine performance in large animals and humans. Similarly, additional strategies are required to activate effective immunity against poorly immunogenic tumour antigens. This review discusses very recent scientific references focused on the development of sophisticated DNA vaccines against cancer. We report a selection of novel and relevant patents employed to improve their immunogenicity through several strategies such as the use of tissue-specific transcriptional elements, nuclear localisation signalling, codon-optimisation and by targeting antigenic proteins to secretory pathway. Recent patents validating portions or splice variants of tumour antigens as candidates for cancer DNA vaccines with improved specificity, such as mesothelin and hTERT, are also discussed. Lastly, we review novel patents on the use of genetic immunomodulators, such as "universal" T helper epitopes derived from tetanus toxin, E. coli heat labile enterotoxin and vegetable proteins, as well as cytokines, chemokines or costimulatory molecules such as IL-6, IL-15, IL- 21 to amplify immunity against cancer.

  3. Improved immunogenicity of Newcastle disease virus inactivated vaccine following DNA vaccination using Newcastle disease virus hemagglutinin-neuraminidase and fusion protein genes.

    Science.gov (United States)

    Firouzamandi, Masoumeh; Moeini, Hassan; Hosseini, Davood; Bejo, Mohd Hair; Omar, Abdul Rahman; Mehrbod, Parvaneh; Ideris, Aini

    2016-03-01

    The present study describes the development of DNA vaccines using the hemagglutinin-neuraminidase (HN) and fusion (F) genes from AF2240 Newcastle disease virus strain, namely pIRES/HN, pIRES/F and pIRES-F/HN. Transient expression analysis of the constructs in Vero cells revealed the successful expression of gene inserts in vitro. Moreover, in vivo experiments showed that single vaccination with the constructed plasmid DNA (pDNA) followed by a boost with inactivated vaccine induced a significant difference in enzyme-linked immunosorbent assay antibody levels (p < 0.05) elicited by either pIRES/F, pIRES/F+ pIRES/HN or pIRES-F/HN at one week after the booster in specific pathogen free chickens when compared with the inactivated vaccine alone. Taken together, these results indicated that recombinant pDNA could be used to increase the efficacy of the inactivated vaccine immunization procedure.

  4. Magnetic Nanovectors for the Development of DNA Blood-Stage Malaria Vaccines

    OpenAIRE

    Fatin M. Nawwab Al-Deen; Xiang, Sue D.; Charles Ma; Kirsty Wilson; Ross L. Coppel; Cordelia Selomulya; Magdalena Plebanski

    2017-01-01

    DNA vaccines offer cost, flexibility, and stability advantages, but administered alone have limited immunogenicity. Previously, we identified optimal configurations of magnetic vectors comprising superparamagnetic iron oxide nanoparticles (SPIONs), polyethylenimine (PEI), and hyaluronic acid (HA) to deliver malaria DNA encoding Plasmodium yoelii (Py) merozoite surface protein MSP119 (SPIONs/PEI/DNA + HA gene complex) to dendritic cells and transfect them with high efficiency in vitro. Herein,...

  5. Treg cell resistance to apoptosis in DNA vaccination for experimental autoimmune encephalomyelitis treatment.

    Directory of Open Access Journals (Sweden)

    Youmin Kang

    Full Text Available BACKGROUND: Regulatory T (Treg cells can be induced with DNA vaccinations and protect mice from the development of experimental autoimmune encephalomyelitis (EAE, a mouse model of multiple sclerosis (MS. Tacrolimus (FK506 has been shown to have functions on inducing immunosuppression and augmenting apoptosis of pathologic T cells in autoimmune disease. Here we examined the therapeutic effect of DNA vaccine in conjunction with FK506 on EAE. METHODOLOGY/PRINCIPAL FINDINGS: After EAE induction, C57BL/6 mice were treated with DNA vaccine in conjunction with FK506. Functional Treg cells were induced in treated EAE mice and suppressed Th1 and Th17 cell responses. Infiltrated CD4 T cells were reduced while Treg cells were induced in spinal cords of treated EAE mice. Remarkably, the activated CD4 T cells augmented apoptosis, but the induced Treg cells resisted apoptosis in treated EAE mice, resulting in alleviation of clinical EAE severity. CONCLUSIONS/SIGNIFICANCE: DNA vaccine in conjunction with FK506 treatment ameliorates EAE by enhancing apoptosis of CD4 T cells and resisting apoptosis of induced Treg cells. Our findings implicate the potential of tolerogenic DNA vaccines for treating MS.

  6. Nucleic acid (DNA) immunization as a platform for dengue vaccine development.

    Science.gov (United States)

    Porter, Kevin R; Raviprakash, Kanakatte

    2015-12-10

    Since the early 1990s, DNA immunization has been used as a platform for developing a tetravalent dengue vaccine in response to the high priority need for protecting military personnel deployed to dengue endemic regions of the world. Several approaches have been explored ranging from naked DNA immunization to the use of live virus vectors to deliver the targeted genes for expression. Pre-clinical animal studies were largely successful in generating anti-dengue cellular and humoral immune responses that were protective either completely or partially against challenge with live dengue virus. However, Phase 1 clinical evaluation of a prototype monovalent dengue 1 DNA vaccine expressing prM and E genes revealed anti-dengue T cell IFNγ responses, but poor neutralizing antibody responses. These less than optimal results are thought to be due to poor uptake and expression of the DNA vaccine plasmids. Because DNA immunization as a vaccine platform has the advantages of ease of manufacture, flexible genetic manipulation and enhanced stability, efforts continue to improve the immunogenicity of these vaccines using a variety of methods.

  7. Construction of pEGFP-ChEgTrp as DNA model for multi-epitope vaccine against Echinococcus granulosus

    Directory of Open Access Journals (Sweden)

    M. Ahmadzadeh

    2015-12-01

    Full Text Available Background: Infection with Echinococcus granulosus causes hydatidosis in human and ruminants. With regards to the high prevalence of hydatidosis in Iran, dealing with this disease is important in terms of public health. Objective: The aim of this study was to construct pEGFP-ChEgTrp as DNA model for multi-epitope vaccine against Echinococcus granulosus Methods: This experimental study was conducted in the Razi Vaccine & Serum Research Institute, Karaj in 2013. Initially, epitopes stimulating the host immune response were predicted by IEDB Database and the coding sequences were made. The sequences were amplified by PCR. The PCR products were cloned into pEGFP-N1 vector after digestion with XhoI restriction enzyme. The bacteria containing recombinant plasmid were evaluated using Colony PCR, agarose gel electrophoresis and sequencing methods. Findings: Four peptides with 10 linear epitopes were predicted in EgTrp antigen. The nucleotide sequence coding ChEgTrp was amplified by PCR using specific primers and a 270 bp fragment was obtained. This fragment was cloned into pEGFP-N1 vector and the recombinant plasmid was confirmed by Colony PCR and agarose gel electrophoresis. For final confirmation, the recombinant plasmid was sequenced and the pEGFP-ChEgTrp was constructed. Conclusion: The ChEgTrp was successfully cloned into the pEGFP-N1 vector and this plasmid can be used to design DNA vaccines.

  8. DNA vaccination protects mice against Zika virus-induced damage to the testes

    Science.gov (United States)

    Griffin, Bryan D.; Muthumani, Kar; Warner, Bryce M.; Majer, Anna; Hagan, Mable; Audet, Jonathan; Stein, Derek R.; Ranadheera, Charlene; Racine, Trina; De La Vega, Marc-Antoine; Piret, Jocelyne; Kucas, Stephanie; Tran, Kaylie N.; Frost, Kathy L.; De Graff, Christine; Soule, Geoff; Scharikow, Leanne; Scott, Jennifer; McTavish, Gordon; Smid, Valerie; Park, Young K.; Maslow, Joel N.; Sardesai, Niranjan Y.; Kim, J. Joseph; Yao, Xiao-jian; Bello, Alexander; Lindsay, Robbin; Boivin, Guy; Booth, Stephanie A.; Kobasa, Darwyn; Embury-Hyatt, Carissa; Safronetz, David; Weiner, David B.; Kobinger, Gary P.

    2017-01-01

    Zika virus (ZIKV) is an emerging pathogen causally associated with serious sequelae in fetuses, inducing fetal microcephaly and other neurodevelopment defects. ZIKV is primarily transmitted by mosquitoes, but can persist in human semen and sperm, and sexual transmission has been documented. Moreover, exposure of type-I interferon knockout mice to ZIKV results in severe damage to the testes, epididymis and sperm. Candidate ZIKV vaccines have shown protective efficacy in preclinical studies carried out in animal models, and several vaccines have entered clinical trials. Here, we report that administration of a synthetic DNA vaccine encoding ZIKV pre-membrane and envelope (prME) completely protects mice against ZIKV-associated damage to the testes and sperm and prevents viral persistence in the testes following challenge with a contemporary strain of ZIKV. These data suggest that DNA vaccination merits further investigation as a potential means to reduce ZIKV persistence in the male reproductive tract. PMID:28589934

  9. Progress in recombinant DNA-derived vaccines for Lassa virus and filoviruses.

    Science.gov (United States)

    Grant-Klein, Rebecca J; Altamura, Louis A; Schmaljohn, Connie S

    2011-12-01

    Developing vaccines for highly pathogenic viruses such as those causing Lassa, Ebola, and Marburg hemorrhagic fevers is a daunting task due to both scientific and logistical constraints. Scientific hurdles to overcome include poorly defined relationships between pathogenicity and protective immune responses, genetic diversity of viruses, and safety in a target population that includes a large number of individuals with compromised immune systems. Logistical obstacles include the requirement for biosafety level-4 containment to study the authentic viruses, the poor public health infrastructure of the endemic disease areas, and the cost of developing these vaccines for use in non-lucrative markets. Recombinant DNA-based vaccine approaches offer promise of overcoming some of these issues. In this review, we consider the status of various recombinant DNA candidate vaccines against Lassa virus and filoviruses which have been tested in animals.

  10. Characterization of GD2 peptide mimotope DNA vaccines effective against spontaneous neuroblastoma metastases.

    Science.gov (United States)

    Fest, Stefan; Huebener, Nicole; Weixler, Silke; Bleeke, Matthias; Zeng, Yan; Strandsby, Anne; Volkmer-Engert, Rudolf; Landgraf, Christiane; Gaedicke, Gerhard; Riemer, Angelika B; Michalsky, Elke; Jaeger, Ines S; Preissner, Robert; Förster-Wald, Elisabeth; Jensen-Jarolim, Erika; Lode, Holger N

    2006-11-01

    Disialoganglioside GD2 is an established target for immunotherapy in neuroblastoma. We tested the hypothesis that active immunization against the glycolipid GD2 using DNA vaccines encoding for cyclic GD2-mimicking decapeptides (i.e., GD2 mimotopes) is effective against neuroblastoma. For this purpose, two GD2 peptide mimotopes (MA and MD) were selected based on docking experiments to anti-GD2 antibody ch14.18 (binding free energy: -41.23 kJ/mol for MA and -48.06 kJ/mol for MD) and Biacore analysis (K(d) = 12.3 x 10(-5) mol/L for MA and 5.3 x 10(-5) mol/L for MD), showing a higher affinity of MD over MA. These sequences were selected for DNA vaccine design based on pSecTag2-A (pSA) also including a T-cell helper epitope. GD2 mimicry was shown following transfection of CHO-1 cells with pSA-MA and pSA-MD DNA vaccines, with twice-higher signal intensity for cells expressing MD over MA. Finally, these DNA vaccines were tested for induction of tumor protective immunity in a syngeneic neuroblastoma model following oral DNA vaccine delivery with attenuated Salmonella typhimurium (SL 7207). Only mice receiving the DNA vaccines revealed a reduction of spontaneous liver metastases. The highest anti-GD2 humoral immune response and natural killer cell activation was observed in mice immunized with the pSA-MD, a finding consistent with superior calculated binding free energy, dissociation constant, and GD2 mimicry potential for GD2 mimotope MD over MA. In summary, we show that DNA immunization with pSA-MD may provide a useful strategy for active immunization against neuroblastoma.

  11. Using recombinant DNA technology for the development of live-attenuated dengue vaccines.

    Science.gov (United States)

    Lee, Hsiang-Chi; Butler, Michael; Wu, Suh-Chin

    2012-07-15

    Dramatic increases in dengue (DEN) incidence and disease severity have been reported, in great part due to the geographic expansion of Aedes aegypti and Aedes albopictus mosquitoes. One result is the expanded co-circulation of all dengue 1-4 serotype viruses (DENV) in urban areas worldwide, especially in South and South-East Asia, and South America. DEN disease severity ranges from asymptomatic infections to febrile dengue fevers (DF) to life-threatening dengue hemorrhagic fever (DHF) and dengue shock syndrome (DSS). There is an urgent need for a safe and effective tetravalent DEN vaccine. Several live attenuated, tetravalent DEN vaccine candidates have been generated by recombinant DNA technology; these candidates are capable of providing immunity to all four DENV serotypes. In this paper we review (a) recombinant live-attenuated DEN vaccine candidates in terms of deletion, antigen chimerization, and the introduction of adaptive mutations; (b) strategies for improving tetravalent vaccine attenuation; and (c) live-attenuated DENV vaccine development.

  12. DNA vaccination of American robins (Turdus migratorius) against West Nile virus.

    Science.gov (United States)

    Kilpatrick, A Marm; Dupuis, Alan P; Chang, Gwong-Jen J; Kramer, Laura D

    2010-05-01

    West Nile virus (WNV) has caused at least 1150 cases of encephalitis, 100 deaths, and an estimated 30,000-80,000 illnesses in 6 of the last 7 years. Recent evidence from several regions has implicated American robins (Turdus migratorius) as an important host for feeding by Culex mosquitoes, and, when integrated with their host competence for WNV, demonstrates that they are a key WNV amplification host. We evaluated the efficacy of a DNA plasmid vaccine at reducing the viremia and infectiousness of hatch-year American robins. We found that a single dose of vaccine injected intramuscularly resulted in more than a 400-fold (10(2.6)) decrease in average viremia. Although sample sizes were small, these results suggest that vaccinated robins exhibit viremias that are likely to be mostly noninfectious to biting Culex mosquitoes. More broadly, if an orally effective formulation of this vaccine could be developed, new control strategies based on wildlife vaccination may be possible.

  13. A rationally designed tyrosine hydroxylase DNA vaccine induces specific antineuroblastoma immunity.

    Science.gov (United States)

    Huebener, Nicole; Fest, Stefan; Strandsby, Anne; Michalsky, Elke; Preissner, Robert; Zeng, Yan; Gaedicke, Gerhard; Lode, Holger N

    2008-07-01

    Therapeutic vaccination against tumor antigens without induction of autoimmunity remains a major challenge in cancer immunotherapy. Here, we show for the first time effective therapeutic vaccination followed by suppression of established spontaneous neuroblastoma metastases using a tyrosine hydroxylase (TH) DNA minigene vaccine. We identified three novel mouse TH (mTH3) derived peptides with high predicted binding affinity to MHC class I antigen H2-K(k) according to the prediction program SYFPEITHI and computer modeling of epitopes into the MHC class I antigen binding groove. Subsequently, a DNA minigene vaccine was generated based on the expression vector pCMV-F3Ub encoding mutated ubiquitin (Gly(76) to Ala(76)) and mTH3. Prophylactic and therapeutic efficacies of this vaccine were established following oral delivery with attenuated Salmonella typhimurium SL7207. Only mice immunized with mTH3 were free of spontaneous liver metastases. This effect was clearly dependent on ubiquitin and high affinity of the mTH epitopes to MHC class I antigens. Specifically, we showed a crucial role for minigene expression as a stable ubiquitin-Ala(76) fusion peptide for vaccine efficacy. The immune response following the mTH3 DNA minigene vaccination was mediated by CD8(+) T cells as indicated by infiltration of primary tumors and TH-specific cytolytic activity in vitro. Importantly, no cell infiltration was detectable in TH-expressing adrenal medulla, indicating the absence of autoimmunity. In summary, we show effective therapeutic vaccination against neuroblastoma with a novel rationally designed TH minigene vaccine without induction of autoimmunity providing an important baseline for future clinical application of this strategy.

  14. A DNA Vaccine for Crimean Congo Hemorrhagic Fever Protects Against Disease and Death in Two Lethal Mouse Models

    Science.gov (United States)

    2017-09-18

    currently no licensed vaccines to prevent CCHFV infection. We developed a DNA vaccine expressing the M-segment glycoprotein genes of CCHFV and assessed its...immunogenicity and protective efficacy in two lethal mouse models of disease: type I interferon receptor knockout (IFNAR-/-) mice; and a novel...humoral immune responses with neutralizing titers after three vaccinations in both IFNAR-/- and IS mouse models.

  15. Immune protection duration and efficacy stability of DNA vaccine encoding Eimeria tenella TA4 and chicken IL-2 against coccidiosis.

    Science.gov (United States)

    Song, Xiaokai; Zhao, Xiaofang; Xu, Lixin; Yan, Ruofeng; Li, Xiangrui

    2017-04-01

    In our previous study, an effective DNA vaccine encoding Eimeria tenella TA4 and chicken IL-2 was constructed. In the present study, the immunization dose of the DNA vaccine pVAX1.0-TA4-IL-2 was further optimized. With the optimized dose, the dynamics of antibodies induced by the DNA vaccine was determined using indirect ELISA. To evaluate the immune protection duration of the DNA vaccine, two-week-old chickens were intramuscularly immunized twice and the induced efficacy was evaluated by challenging with E. tenella at 5, 9, 13, 17 and 21weeks post the last immunization (PLI) separately. To evaluate the efficacy stability of the DNA vaccine, two-week-old chickens were immunized with 3 batches of the DNA vaccine, and the induced efficacy was evaluated by challenging with E. tenella. The results showed that the optimal dose was 25μg. The induced antibody level persisted until 10weeks PPI. For the challenge time of 5 and 9weeks PLI, the immunization resulted in ACIs of 182.28 and 162.23 beyond 160, showing effective protection. However, for the challenge time of 13, 17 and 21weeks PLI, the immunization resulted in ACIs below 160 which means poor protection. Therefore, the immune protection duration of the DNA vaccination was at least 9weeks PLI. DNA immunization with three batches DNA vaccine resulted in ACIs of 187.52, 191.57 and 185.22, which demonstrated that efficacies of the three batches DNA vaccine were effective and stable. Overall, our results indicate that DNA vaccine pVAX1.0-TA4-IL-2 has the potential to be developed as effective vaccine against coccidiosis. Copyright © 2016 Elsevier Ltd. All rights reserved.

  16. Preparation and efficacy of Newcastle disease virus DNA vaccine encapsulated in PLGA nanoparticles.

    Directory of Open Access Journals (Sweden)

    Kai Zhao

    Full Text Available BACKGROUND: Although the Newcastle disease virus (NDV inactivated vaccines and attenuated live vaccines have been used to prevent and control Newcastle disease (ND for years, there are some disadvantages. Recently, newly developed DNA vaccines have the potential to overcome these disadvantages. The low delivery efficiency, however, hindered the application of DNA vaccines for ND in practice. METHODOLOGY/PRINCIPAL FINDINGS: The eukaryotic expression plasmid pVAX1-F (o DNA that expressed the F gene of NDV encapsulated in PLGA nanoparticles (pFNDV-PLGA-NPs were prepared by a double emulsion-solvent evaporation method and optimal preparation conditions of the pFNDV-PLGA-NPs were determined. Under the optimal conditions, the pFNDV-PLGA-NPs were produced in good morphology and had high stability with a mean diameter of 433.5 ± 7.5 nm, with encapsulation efficiency of 91.8 ± 0.3% and a Zeta potential of +2.7 mV. Release assay in vitro showed that the fusion gene plasmid DNA could be sustainably released from the pFNDV-PLGA-NPs up to 93.14% of the total amount. Cell transfection test indicated that the vaccine expressed and maintained its bioactivity. Immunization results showed that better immune responses of SPF chickens immunized with the pFNDV-PLGA-NPs were induced compared to the chickens immunized with the DNA vaccine alone. In addition, the safety of mucosal immunity delivery system of the pFNDV-PLGA-NPs was also tested in an in vitro cytotoxicity assay. CONCLUSIONS/SIGNIFICANCE: The pFNDV-PLGA-NPs could induce stronger cellular, humoral, and mucosal immune responses and reached the sustained release effect. These results laid a foundation for further development of vaccines and drugs in PLGA nanoparticles.

  17. Molecular Adjuvant Ag85A Enhances Protection against Influenza A Virus in Mice Following DNA Vaccination

    Directory of Open Access Journals (Sweden)

    Hong Li

    2012-12-01

    Full Text Available A novel DNA vaccine vector encoding the Mycobacterium tuberculosis secreted antigen Ag85A fused with the influenza A virus (IAV HA2 protein epitopes, pEGFP/Ag85A-sHA2 (pAg85A-sHA2, was designed to provide protection against influenza. The antigen encoded by the DNA vaccine vector was efficiently expressed in mammalian cells, as determined by reverse transcription polymerase chain reaction (RT-PCR and fluorescence analyses. Mice were immunized with the vaccine vector by intramuscular injection before challenge with A/Puerto Rico/8/34 virus (PR8 virus. Sera and the splenocyte culture IFN-γ levels were significantly higher in immunized mice compared with the control mice. The novel vaccine group showed a high neutralization antibody titer in vitro. The novel vaccine vector also reduced the viral loads, increased the survival rates in mice after the PR8 virus challenge and reduced the alveolar inflammatory cell numbers. Sera IL-4 concentrations were significantly increased in mice immunized with the novel vaccine vector on Day 12 after challenge with the PR8 virus. These results demonstrated that short HA2 (sHA2 protein epitopes may provide protection against the PR8 virus and that Ag85A could strengthen the immune response to HA2 epitopes, thus, Ag85A may be developed as a new adjuvant for influenza vaccines.

  18. Suppression of breast tumor growth by DNA vaccination against phosphatase of regenerating liver 3.

    Science.gov (United States)

    Lv, J; Liu, C; Huang, H; Meng, L; Jiang, B; Cao, Y; Zhou, Z; She, T; Qu, L; Wei Song, S; Shou, C

    2013-08-01

    Phosphatase of regenerating liver (PRL)-3 is highly expressed in multiple cancers and has important roles in cancer development. Some small-molecule inhibitors and antibodies targeting PRL-3 have been recently reported to inhibit tumor growth effectively. To determine whether PRL-3-targeted DNA vaccination can induce immune response to prevent or inhibit the tumor growth, we established mouse D2F2 breast cancer cells expressing PRL-3 (D2F2/PRL-3) and control cells (D2F2/NC) with lentivirus, and constructed pVAX1-Igκ-PRL-3 plasmid (named as K-P3) as DNA vaccine to immunize BALB/c mice. We found that the K-P3 vaccine delivered by gene gun significantly prevented the growth of D2F2/PRL-3 compared with pVAX1-vector (Padjuvants, such as Mycobacterium tuberculosis heat-shock protein, CTL antigen 4 and M. tuberculosis T-cell stimulatory epitope (MT), into K-P3 vaccine for expressing the fusion proteins. We found that these adjuvant molecules did not significantly improve the antitumor activity of PRL-3 vaccine, but enhanced the production of PRL-3 antibodies in immunized mice. Summarily, our findings demonstrate that PRL-3-targeted DNA vaccine can generate significantly preventive and therapeutic effects on the growth of breast cancer expressing PRL-3 through the induction of cellular immune responses to PRL-3.

  19. A DNA vaccine directed against a rainbow trout rhabdovirus induces early protection against a nodavirus challenge in turbot

    DEFF Research Database (Denmark)

    Sommerset, I.; Lorenzen, Ellen; Lorenzen, Niels;

    2003-01-01

    A DNA vaccine encoding the envelope glycoprotein from a fish rhabdovirus, viral hemorrhagic septicemia virus (VHSV), has previously been shown to induce both early and long time protection against the virus in rainbow trout. Challenge experiments have revealed that the immunity established shortly...... after vaccination is cross-protective against heterologous fish rhabdoviruses. In this study, we show that the DNA vaccine encoding the VHSV glycoprotein also induces early protection against a non-enveloped, positive-sense RNA vir-us belonging to the Nodavirus family, the Atlantic halibut nodavirus...... (AHNV). In a vaccine. efficacy test using juvenile turbot as model fish, the fish injected with the VHSV vaccine were completely protected against a nodavirus challenge performed 8 days post vaccination, while the cumulative mortality in the control group reached 54%. A DNA vaccine carrying the gene...

  20. Tumor antigens for cancer immunotherapy: therapeutic potential of xenogeneic DNA vaccines

    Directory of Open Access Journals (Sweden)

    Srinivasan Roopa

    2004-04-01

    Full Text Available Abstract Preclinical animal studies have convincingly demonstrated that tumor immunity to self antigens can be actively induced and can translate into an effective anti-tumor response. Several of these observations are being tested in clinical trials. Immunization with xenogeneic DNA is an attractive approach to treat cancer since it generates T cell and antibody responses. When working in concert, these mechanisms may improve the efficacy of vaccines. The use of xenogeneic DNA in overcoming immune tolerance has been promising not only in inbred mice with transplanted tumors but also in outbred canines, which present with spontaneous tumors, as in the case of human. Use of this strategy also overcomes limitations seen in other types of cancer vaccines. Immunization against defined tumor antigens using a xenogeneic DNA vaccine is currently being tested in early phase clinical trials for the treatment of melanoma and prostate cancers, with proposed trials for breast cancer and Non-Hodgkin's Lymphoma.

  1. The protective mechanisms induced by a fish rhabdovirus DNA vaccine depend on temperature

    DEFF Research Database (Denmark)

    Lorenzen, Ellen; Einer-Jensen, Katja; Rasmussen, Jesper Skou

    2009-01-01

    an intramuscular injection of 1 mu g purified plasmid DNA and challenged with virulent VHSV 8 or 36-40 days later. The vaccine protected the fish well at all three temperatures, but the involvement of innate and adaptive mechanisms differed: at low temperature. non-specific protection lasted longer and at 36 dpv......DNA vaccines encoding the viral glycoproteins of viral haemorrhagic septicaemia virus (VHSV) and infectious haematopoietic necrosis Virus (IHNV) have proved highly efficient in rainbow trout (Oncorhynchus mykiss) under experimental conditions. Non-specific as well as specific immune mechanisms seem...... to be activated. Temperature is an important external parameter affecting the immune response in fish. The present study aimed at determining the effectiveness of a DNA vaccine against VHS at different temperatures. Rainbow trout fingerlings acclimated at 5 degrees C, 10 degrees C or 15 degrees C, were given...

  2. A simple method encoding linear single strain DNA sequence with natural numbers

    Institute of Scientific and Technical Information of China (English)

    LI Jiye; XU Yuan; ZHANG Wang

    2008-01-01

    A simple method presenting linear single strain DNA (LssDNA) sequence with natural numbers is introduced in this paper. The method presents LssDNA correspondingly with the numerals 1, 2, 3 and 4. After calculation, the sequence can be coded in natural numbers which can also be decoded into the DNA sequence. Thus, an LssDNA sequence can be expressed in a natural number and a dot at coordinate axes. In the future, a new LssDNA sequences database termed "DotBank" would be realized in which each LssDNA sequence is determined as a dot.

  3. Evaluation of a prototype dengue-1 DNA vaccine in a Phase 1 clinical trial.

    Science.gov (United States)

    Beckett, Charmagne G; Tjaden, Jeffrey; Burgess, Timothy; Danko, Janine R; Tamminga, Cindy; Simmons, Monika; Wu, Shuenn-Jue; Sun, Peifang; Kochel, Tadeusz; Raviprakash, Kanakatte; Hayes, Curtis G; Porter, Kevin R

    2011-01-29

    Candidate dengue DNA vaccine constructs for each dengue serotype were developed by incorporating pre-membrane and envelope genes into a plasmid vector. A Phase 1 clinical trial was performed using the dengue virus serotype-1 (DENV-1) vaccine construct (D1ME(100)). The study was an open-label, dose-escalation, safety and immunogenicity trial involving 22 healthy flavivirus-naïve adults assigned to one of two groups. Each group received three intramuscular injections (0, 1, and 5 months) of either a high dose (5.0mg, n=12) or a low dose (1.0mg, n=10) DNA vaccine using the needle-free Biojector(®) 2000. The most commonly reported solicited signs and symptoms were local mild pain or tenderness (10/22, 45%), local mild swelling (6/22, 27%), muscle pain (6/22, 27%) and fatigue (6/22, 27%). Five subjects (41.6%) in the high dose group and none in the low dose group developed detectable anti-dengue neutralizing antibodies. T-cell IFN gamma responses were detected in 50% (4/8) and 83.3% (10/12) of subjects in the low and high dose groups, respectively. The safety profile of the DENV-1 DNA vaccine is acceptable at both doses administered in the study. These results demonstrate a favorable reactogenicity and safety profile of the first in human evaluation of a DENV-1 DNA vaccine.

  4. Detection of Progeny Immune Responses after Intravenous Administration of DNA Vaccine to Pregnant Mice

    Directory of Open Access Journals (Sweden)

    Xin Ke-Qin

    2002-01-01

    Full Text Available A number of factors influence the development of tolerance, including the nature, concentration and mode of antigen presentation to the immune system, as well as the age of the host. The studies were conducted to determine whether immunizing pregnant mice with liposome-encapsulated DNA vaccines had an effect on the immune status of their offspring. Two different plasmids (encoding antigens from HIV-1 and influenza virus were administered intravenously to pregnant mice. At 9.5 days post conception with cationic liposomes, injected plasmid was present in the tissues of the fetus, consistent with trans-placental transfer. When the offspring of vaccinated dams were immunized with DNA vaccine, they mounted stronger antigen-specific immune responses than controls and were protected against challenge by homologous influenza virus after vaccination. Moreover, such immune responses were strong in the offspring of mothers injected with DNA plasmid 9.5 days after coitus. These results suggest that DNA vaccinated mothers confer the antigen-specific immunity to their progeny. Here we describe the methods in detail as they relate to our previously published work.

  5. Restriction endonuclease mapping of linear unintegrated proviral DNA of bovine leukemia virus.

    OpenAIRE

    Kettmann, R; Couez, D; Burny, A

    1981-01-01

    A detailed restriction map was deduced for the genome of the exogenous bovine leukemia virus. The cleavage sites for nine restriction enzymes were mapped. The unintegrated linear viral DNA intermediate that is produced by infection of permissive cells with bovine leukemia virus was isolated. The linear viral DNA had a unique restriction map, indicating that it is not a set of random circular permutations of the RNA genome. From hybridization with a 3'-enriched probe, the DNA restriction map w...

  6. Plasmid DNA Initiates Replication of Yellow Fever Vaccine In Vitro and Elicits Virus-Specific Immune Response in Mice

    OpenAIRE

    Tretyakova, Irina; Nickols, Brian; Hidajat, Rachmat; Jokinen, Jenny; Lukashevich, Igor S.; Pushko, Peter

    2014-01-01

    Yellow fever (YF) causes an acute hemorrhagic fever disease in tropical Africa and Latin America. To develop a novel experimental YF vaccine, we applied iDNA infectious clone technology. The iDNA represents plasmid that encodes the full-length RNA genome of 17D vaccine downstream from a cytomegalovirus (CMV) promoter. The vaccine was designed to transcribe the full-length viral RNA and to launch 17D vaccine virus in vitro and in vivo. Transfection with 10ng of iDNA plasmid was sufficient to s...

  7. DNA electroporation of multi-agent vaccines conferring protection against select agent challenge: TriGrid delivery system.

    Science.gov (United States)

    Keane-Myers, Andrea M; Bell, Matt; Hannaman, Drew; Albrecht, Mark

    2014-01-01

    Effective multi-agent/multivalent vaccines that confer protection against more than one disease are highly desirable to the patient and to health-care professionals. Electroporation of DNA vaccines, whereby tissues injected with DNA are subjected to localized electrical currents, is an ideal platform technology that achieves protective immune responses to multivalent vaccination. Here, we describe an electroporation-based immunization technique capable of administering a cocktail of DNA vaccinations in vivo. Immune response measurements, including protection from pathogen challenge and induction of antigen-specific antibody responses and cell-mediated immune responses, are also discussed.

  8. Single-tube linear DNA amplification (LinDA) for robust ChIP-seq

    NARCIS (Netherlands)

    Shankaranarayanan, P.; Mendoza-Parra, M.A.; Walia, M.; Wang, L.; Li, N.; Trindade, L.M.; Gronemeyer, H.

    2011-01-01

    Genome-wide profiling of transcription factors based on massive parallel sequencing of immunoprecipitated chromatin (ChIP-seq) requires nanogram amounts of DNA. Here we describe a high-fidelity, single-tube linear DNA amplification method (LinDA) for ChIP-seq and reChIP-seq with picogram DNA amounts

  9. Immunogenicity of two different dosages (10 and 5 μg) of recombinant DNA hepatitis B vaccine in healthy neonates

    NARCIS (Netherlands)

    R. Del Cancho (R.); P.M. Grosheie (P.); M. Voogd-Schotanus (M.); W. Huisman (Willem); R.A. Heijtink; S.W. Schalm (Solko)

    1994-01-01

    textabstractThe immunogenicity of a half (5 μg) and a full (10 μg) dosage of recombinant DNA yeast-derived hepatitis B vaccine (HB-Vax-DNA) in healthy neonates was assessed in order to compare two candidate dosages of vaccine. After randomization 174 newborns of HBsAg-negative mothers entered the st

  10. [Problems and prospects of gene therapeutics and DNA vaccines development and application].

    Science.gov (United States)

    Kibirev, Ia A; Drobkov, B I; Marakulin, I V

    2010-01-01

    The review is summarized foreign publications devoted to different aspects of DNA vaccines and gene therapeutics' biological safety. In spite of incomprehension in their action, numerous prototype DNA-based biopharmaceuticals are in advanced stages of human clinical trials. This review is focused on some safety concerns of gene formulations vaccines relate to toxic effects, vertical transmission possibility, genome integration complications, immunologic and immunopathologic effects and environmental spread. It is noted that necessity of national regulatory documents development related to gene therapy medicinal products is significant condition of their application to medical practice.

  11. Anti-tumor effects of a human VEGFR-2-based DNA vaccine in mouse models

    OpenAIRE

    XIE, KE; Bai, Rui-Zhen; Wu, Yang; Liu, Quan; Liu,Kang; Wei, Yu-Quan

    2009-01-01

    Background Vascular endothelial growth factor (VEGF) and its receptor, VEGFR-2 (Flk-1/KDR), play a key role in tumor angiogenesis. Blocking the VEGF-VEGFR-2 pathway may inhibit tumor growth. Here, we used human VEGFR-2 as a model antigen to explore the feasibility of immunotherapy with a plasmid DNA vaccine based on a xenogeneic homologue of this receptor. Methods The protective effects and therapeutic anti-tumor immunity mediated by the DNA vaccine were investigated in mouse models. Anti-ang...

  12. Directed Molecular Evolution Improves the Immunogenicity and Protective Efficacy of a Venezuelan Equine Encephalitis Virus DNA Vaccine

    Science.gov (United States)

    2009-05-01

    developed by serial passage of the virulent Trinidad donkey strain in cultures of guinea pig heart cells [5]. Although TC-83 is gen- erally safe and...vaccinated with plasmid DNA 18]. In addition, improvements in the immunogenicity and cross- eactivity of DNA vaccine candidates for HIV -1 and the malaria...antibodies to the VEEV E2 protein. Consequently, we used pools of the day 63 pre-challenge sera from each vaccination group of the pathogen challenge

  13. The immune response induced by DNA vaccine expressing nfa1 gene against Naegleria fowleri.

    Science.gov (United States)

    Kim, Jong-Hyun; Lee, Sang-Hee; Sohn, Hae-Jin; Lee, Jinyoung; Chwae, Yong-Joon; Park, Sun; Kim, Kyongmin; Shin, Ho-Joon

    2012-12-01

    The pathogenic free-living amoeba, Naegleria fowleri, causes fatal primary amoebic meningoencephalitis in experimental animals and in humans. The nfa1 gene that was cloned from N. fowleri is located on pseudopodia, especially amoebic food cups and plays an important role in the pathogenesis of N. fowleri. In this study, we constructed and characterized retroviral vector and lentiviral vector systems for nfa1 DNA vaccination in mice. We constructed the retroviral vector (pQCXIN) and the lentiviral vector (pCDH) cloned with the egfp-nfa1 gene. The expression of nfa1 gene in Chinese hamster ovary cell and human primary nasal epithelial cell transfected with the pQCXIN/egfp-nfa1 vector or pCDH/egfp-nfa1 vector was observed by fluorescent microscopy and Western blotting analysis. Our viral vector systems effectively delivered the nfa1 gene to the target cells and expressed the Nfa1 protein within the target cells. To evaluate immune responses of nfa1-vaccinated mice, BALB/c mice were intranasally vaccinated with viral particles of each retro- or lentiviral vector expressing nfa1 gene. DNA vaccination using viral vectors expressing nfa1 significantly stimulated the production of Nfa1-specific IgG subclass, as well as IgG levels. In particular, both levels of IgG2a (Th1) and IgG1 (Th2) were significantly increased in mice vaccinated with viral vectors. These results show the nfa1-vaccination induce efficiently Th1 type, as well as Th2 type immune responses. This is the first report to construct viral vector systems and to evaluate immune responses as DNA vaccination in N. fowleri infection. Furthermore, these results suggest that nfal vaccination may be an effective method for treatment of N. fowleri infection.

  14. Protective immunity to H7N9 influenza viruses elicited by synthetic DNA vaccine.

    Science.gov (United States)

    Yan, Jian; Villarreal, Daniel O; Racine, Trina; Chu, Jaemi S; Walters, Jewell N; Morrow, Matthew P; Khan, Amir S; Sardesai, Niranjan Y; Kim, J Joseph; Kobinger, Gary P; Weiner, David B

    2014-05-19

    Despite an intensive vaccine program influenza infections remain a major health problem, due to the viruses' ability to change its envelope glycoprotein hemagglutinin (HA), through shift and drift, permitting influenza to escape protection induced by current vaccines or natural immunity. Recently a new variant, H7N9, has emerged in China causing global concern. First, there have been more than 130 laboratory-confirmed human infections resulting in an alarmingly high death rate (32.3%). Second, genetic changes found in H7N9 appear to be associated with enabling avian influenza viruses to spread more effectively in mammals, thus transmitting infections on a larger scale. Currently, no vaccines or drugs are effectively able to target H7N9. Here, we report the rapid development of a synthetic consensus DNA vaccine (pH7HA) to elicit potent protective immunity against the H7N9 viruses. We show that pH7HA induces broad antibody responses that bind to divergent HAs from multiple new members of the H7N9 family. These antibody responses result in high-titer HAI against H7N9. Simultaneously, this vaccine induces potent polyfunctional effector CD4 and CD8T cell memory responses. Animals vaccinated with pH7HA are completely protected from H7N9 virus infection and any morbidity associated with lethal challenge. This study establishes that this synthetic consensus DNA vaccine represents a new tool for targeting emerging infection, and more importantly, its design, testing and development into seed stock for vaccine production in a few days in the pandemic setting has significant implications for the rapid deployment of vaccines protecting against emerging infectious diseases.

  15. Construction of a recombinant attenuated Salmonella typhimurium DNA vaccine carrying Helicobacter pylori hpaA.

    Science.gov (United States)

    Xu, Can; Li, Zhao-Shen; Du, Yi-Qi; Tu, Zhen-Xing; Gong, Yan-Fang; Jin, Jing; Wu, Hong-Yu; Xu, Guo-Ming

    2005-01-07

    To construct a recombinant attenuated Salmonella typhimurium DNA vaccine carrying Helicobacter pylori hpaA gene and to detect its immunogenicity. Genomic DNA of the standard H pylori strain 17 874 was isolated as the template, hpaA gene fragment was amplified by polymerase chain reaction (PCR) and cloned into pUCmT vector. DNA sequence of the amplified hpaA gene was assayed, then cloned into the eukaryotic expression vector pIRES through enzyme digestion and ligation reactions. The recombinant plasmid was used to transform competent Escherichia coli DH5alpha, and the positive clones were screened by PCR and restriction enzyme digestion. Then, the recombinant pIRES-hpaA was used to transform LB5000 and the recombinant plasmid isolated from LB5000 was finally used to transform SL7207. After that, the recombinant strain was grown in vitro repeatedly. In order to identify the immunogenicity of the vaccine in vitro, the recombinant pIRES-hpaA was transfected to COS-7 cells using Lipofectamine2000, the immunogenicity of expressed HpaA protein was detected with SDS-PAGE and Western blot. The 750-base pair hpaA gene fragment was amplified from the genomic DNA and was consistent with the sequence of H pylori hpaA by sequence analysis. It was confirmed by PCR and restriction enzyme digestion that H pylori hpaA gene was inserted into the eukaryotic expression vector pIRES and a stable recombinant live attenuated Salmonella typhimurium DNA vaccine carrying H pylori hpaA gene was successfully constructed and the specific strip of HpaA expressed by pIRES-hpaA was detected through Western blot. The recombinant attenuated Salmonella typhimurium DNA vaccine strain expressing HpaA protein with immunogenicity can be constructed and it may be helpful for further investigating the immune action of DNA vaccine in vivo.

  16. Nano-Delivery Vehicles/Adjuvants for DNA Vaccination Against HIV.

    Science.gov (United States)

    Dong, Yaqiong; Yang, Jun; Zhang, Jinchao; Zhang, Xin

    2016-03-01

    More than 75 million people has been infected HIV and it is responsible for nearly 36 million deaths on a global scale. As one of the deadliest infectious diseases, HIV is becoming the urgent issue of the global epidemic to tackle. In order to settle this problem from the source, some effective prevention strategies should be developed to control the pandemic of HIV. Vaccines, especially DNA vaccines, could be the optimal way to control the spread of HIV due to the unparalleled superiority that DNA vaccines could generate long-term humoral and cellular immune responses which could provide protective immunity for HIV. But the naked DNA could hardly enter into cells and is easily degraded by DNases and lysosomes, so designing effective delivery system is a promising strategy. Since delivery system could be constructed to promote efficient delivery of DNA into mammalian cells, protect them from degradation, and also could be established to be a target system to arrive at certain position of expectation. The current review discusses the potential of various nano-delivery vehicles/adjuvants such as polymer, lipid, liposome, peptide and inorganic material in improving efficiency of diverse modalities available for HIV DNA vaccines.

  17. Construction of multivalent DNA vaccines for Mycobacte-rium tuberculosis and its immunogenicity

    Institute of Scientific and Technical Information of China (English)

    2002-01-01

    The coding regions of Ag85B MPT-64, and ESAT-6 secreted proteins were cloned initially into the eukaryotic expression vector pJW4303, then transformed to E. coli Top 10 strain for plasmid DNA extraction and further analysis. Plasmids containing the right insertion were sequenced to confirm their identity. COS7 cells were transfected with a mixture containing serially diluted plasmid DNA encoding three secreted proteins and Lipofectin (Gibco). The supernatants and pellets prepared from various cell lines were run on SDS-PAGE gel and the expression of these proteins in COS7 cells were demonstrated by immunoblot using polyclonal or monoclonal antiserum of M.TBH37Rv. 21 days after first vaccination of C57BL-6 mice by all three recombinant eukaryotic expressing vectors, antibody titer for Ag85B reached 1∶3200. 21 days after second vaccination, the antibody titer reached 1∶102400. The highest antibody levels induced by multivalent vaccines after the second injection were equal to or even greater than the highest antibody levels of single DNA vaccine reported in literature after third injections. Antibody titer of MPT-64 was 1∶50 after the first injection and it reached 1∶200 after the second injection. No antigen-specific antibody against ESAT-6 was detected in sera harvested from immunized mice 21 days after both injections. Antigen-specific IFN-g level of Ag85B was 110 pg/mL while no antigen-specific IFN- g level of ESAT-6 and MPT-64 was detected even after third injections. To our knowledge, it is the first time that studies of polyvalent recombinant DNA vaccines against TB were carried out in C57BL-6 mice. Our results indicated that multiple DNA vaccines could be used to enhance protective responses against M.TB.

  18. IFN-γ increases efficiency of DNA vaccine in protecting ducks against infection

    Institute of Scientific and Technical Information of China (English)

    Jian-Er Long; Li-Na Huang; Zhi-Qiang Qin; Wen-Yi Wang; Di Qu

    2005-01-01

    AIM: To detect the effects of DNA vaccines in combination with duck IFN-γ gene on the protection of ducks against duck hepatitis B virus (DHBV) infection.METHODS: DuIFN-γ cDNA was cloned and expressed in COS-7 cells, and the antiviral activity of DuIFN-γ was detected and neutralized by specific antibodies. Ducks were vaccinated with DHBpreS/S DNA alone or coimmunized with plasmid expressing DuIFN-γ. DuIFN-γmRNA in peripheral blood mononuclear cells (PBMCs) from immunized ducks was detected by semi-quantitative competitive RT-PCR. Anti-DHBpreS was titrated by enzyme-linked immunosorbent assay (ELISA). DHBV DNA in sera and liver was detected by Southern blot hybridization, after ducks were challenged with high doses of DHBV.RESULTS: DuIFN-γ expressed by COS-7 was able to protect duck fibroblasts against vesicular stomatitis virus (VSV) infection in a dose-dependent fashion, and antiDuIFN-γ antibodies neutralized the antiviral effects. DuIFN-γin the supernatant also inhibited the release of DHBV DNA from LMH-D2 cells. When ducks were co-immunized with DNA vaccine expressing DHBpreS/S and DuIFN-γ gene as an adjuvant, the level of DuIFN-γ mRNA in PBMCs was higher than that in ducks vaccinated with DHBpreS/S DNA alone. However, the titer of anti-DHBpreS elicited by DHBpreS/S DNA alone was higher than that co-immunized with DuIFN-γ gene and DHBpreS/S DNA. After being challenged with DHBV at high doses, the load of DHBV in sera dropped faster, and the amount of total DNA and cccDNA in the liver decreased more significantly in the group of ducks co-immunized with DuIFN-γ gene and DHBpreS/S DNA than in other groups.CONCLUSION: DHBV preS/S DNA vaccine can protect ducks against DHBV infection, DuIFN-γ gene as an immune adjuvant enhances its efficacy.

  19. Protection of mice against Chlamydophila abortus infection with a bacteriophage-mediated DNA vaccine expressing the major outer membrane protein.

    Science.gov (United States)

    Ling, Yong; Liu, Wei; Clark, Jason R; March, John B; Yang, Junjing; He, Cheng

    2011-12-15

    A bacteriophage-delivered DNA vaccine against Chlamydophila abortus was constructed by cloning a eukaryotic cassette containing the ompA gene (which expresses the Major Outer Membrane Protein) into a bacteriophage lambda vector. Four groups, each of 20 BALB/c mice were inoculated separately with the phage vaccine, a conventional DNA vaccine based on the same ompA expression cassette, a live attenuated vaccine (strain 1B) or the empty phage vector. The phage and DNA vaccines and empty phage vector were administered intramuscularly on days 0, 14 and 28; the attenuated vaccine was given once on day 0. Half the animals in each group were challenged on day 42 by intraperitoneal injection of live C. abortus and sacrificed on day 49. Phage-vaccinated mice developed moderate antibody levels against C. abortus and yielded higher levels of IFN-γ and IL-2 compared with the attenuated live vaccine group. Clearance of chlamydiae from spleens was significantly better in the attenuated vaccine group compared with the phage vaccine group, while both groups were significantly superior to the DNA vaccine and control groups (p<0.01). Although levels of protection in the mouse model were lower in phage-vaccinated animals, than in 1B vaccinated animals, phage vaccines offer several other advantages, such as easier handling and safety, potentially cheaper production and no chance of reversion to virulence. Although these are preliminary results in a model system, it is possible that with further optimisation immunization with phage vaccines may provide a novel way to improve protection against C. abortus infection and trials in large animals are currently being initiated.

  20. Plasmid DNA initiates replication of yellow fever vaccine in vitro and elicits virus-specific immune response in mice.

    Science.gov (United States)

    Tretyakova, Irina; Nickols, Brian; Hidajat, Rachmat; Jokinen, Jenny; Lukashevich, Igor S; Pushko, Peter

    2014-11-01

    Yellow fever (YF) causes an acute hemorrhagic fever disease in tropical Africa and Latin America. To develop a novel experimental YF vaccine, we applied iDNA infectious clone technology. The iDNA represents plasmid that encodes the full-length RNA genome of 17D vaccine downstream from a cytomegalovirus (CMV) promoter. The vaccine was designed to transcribe the full-length viral RNA and to launch 17D vaccine virus in vitro and in vivo. Transfection with 10 ng of iDNA plasmid was sufficient to start replication of vaccine virus in vitro. Safety of the parental 17D and iDNA-derived 17D viruses was confirmed in AG129 mice deficient in receptors for IFN-α/β/γ. Finally, direct vaccination of BALB/c mice with a single 20 μg dose of iDNA plasmid resulted in seroconversion and elicitation of virus-specific neutralizing antibodies in animals. We conclude that iDNA immunization approach combines characteristics of DNA and attenuated vaccines and represents a promising vaccination strategy for YF.

  1. Immunogenicity of RSV F DNA Vaccine in BALB/c Mice

    Directory of Open Access Journals (Sweden)

    Erdal Eroglu

    2016-01-01

    Full Text Available Respiratory syncytial virus (RSV causes severe acute lower respiratory tract disease leading to numerous hospitalizations and deaths among the infant and elderly populations worldwide. There is no vaccine or a less effective drug available against RSV infections. Natural RSV infection stimulates the Th1 immune response and activates the production of neutralizing antibodies, while earlier vaccine trials that used UV-inactivated RSV exacerbated the disease due to the activation of the allergic Th2 response. With a focus on Th1 immunity, we developed a DNA vaccine containing the native RSV fusion (RSV F protein and studied its immune response in BALB/c mice. High levels of RSV specific antibodies were induced during subsequent immunizations. The serum antibodies were able to neutralize RSV in vitro. The RSV inhibition by sera was also shown by immunofluorescence analyses. Antibody response of the RSV F DNA vaccine showed a strong Th1 response. Also, sera from RSV F immunized and RSV infected mice reduced the RSV infection by 50% and 80%, respectively. Our data evidently showed that the RSV F DNA vaccine activated the Th1 biased immune response and led to the production of neutralizing antibodies, which is the desired immune response required for protection from RSV infections.

  2. In vivo electroporation enhances the immunogenicity of an HIV-1 DNA vaccine candidate in healthy volunteers.

    Directory of Open Access Journals (Sweden)

    Sandhya Vasan

    Full Text Available BACKGROUND: DNA-based vaccines have been safe but weakly immunogenic in humans to date. METHODS AND FINDINGS: We sought to determine the safety, tolerability, and immunogenicity of ADVAX, a multigenic HIV-1 DNA vaccine candidate, injected intramuscularly by in vivo electroporation (EP in a Phase-1, double-blind, randomized placebo-controlled trial in healthy volunteers. Eight volunteers each received 0.2 mg, 1 mg, or 4 mg ADVAX or saline placebo via EP, or 4 mg ADVAX via standard intramuscular injection at weeks 0 and 8. A third vaccination was administered to eleven volunteers at week 36. EP was safe, well-tolerated and considered acceptable for a prophylactic vaccine. EP delivery of ADVAX increased the magnitude of HIV-1-specific cell mediated immunity by up to 70-fold over IM injection, as measured by gamma interferon ELISpot. The number of antigens to which the response was detected improved with EP and increasing dosage. Intracellular cytokine staining analysis of ELISpot responders revealed both CD4+ and CD8+ T cell responses, with co-secretion of multiple cytokines. CONCLUSIONS: This is the first demonstration in healthy volunteers that EP is safe, tolerable, and effective in improving the magnitude, breadth and durability of cellular immune responses to a DNA vaccine candidate. TRIAL REGISTRATION: ClinicalTrials.gov NCT00545987.

  3. Immunotherapy with an HIV-DNA Vaccine in Children and Adults

    Directory of Open Access Journals (Sweden)

    Paolo Palma

    2014-07-01

    Full Text Available Therapeutic HIV immunization is intended to induce new HIV-specific cellular immune responses and to reduce viral load, possibly permitting extended periods without antiretroviral drugs. A multigene, multi-subtype A, B, C HIV-DNA vaccine (HIVIS has been used in clinical trials in both children and adults with the aim of improving and broadening the infected individuals’ immune responses. Despite the different country locations, different regimens and the necessary variations in assays performed, this is, to our knowledge, the first attempt to compare children’s and adults’ responses to a particular HIV vaccine. Ten vertically HIV-infected children aged 4–16 years were immunized during antiretroviral therapy (ART. Another ten children were blindly recruited as controls. Both groups continued their antiretroviral treatment during and after vaccinations. Twelve chronically HIV-infected adults were vaccinated, followed by repeated structured therapy interruptions (STI of their antiretroviral treatment. The adult group included four controls, receiving placebo vaccinations. The HIV-DNA vaccine was generally well tolerated, and no serious adverse events were registered in any group. In the HIV-infected children, an increased specific immune response to Gag and RT proteins was detected by antigen-specific lymphoproliferation. Moreover, the frequency of HIV-specific CD8+ T-cell lymphocytes releasing perforin was significantly higher in the vaccinees than the controls. In the HIV-infected adults, increased CD8+ T-cell responses to Gag, RT and viral protease peptides were detected. No augmentation of HIV-specific lymphoproliferative responses were detected in adults after vaccination. In conclusion, the HIV-DNA vaccine can elicit new HIV-specific cellular immune responses, particularly to Gag antigens, in both HIV-infected children and adults. Vaccinated children mounted transient new HIV-specific immune responses, including both CD4+ T

  4. Prospects and progress of DNA vaccines for treating hepatitis B

    NARCIS (Netherlands)

    Chen, Margaret; Jagya, Neetu; Bansal, Ruchi; Frelin, Lars; Sällberg, Matti

    2015-01-01

    The hepatitis B virus (HBV) is a global cause of liver disease. The preventive HBV vaccine has effectively reduced the disease burden. However, an estimated 340 million chronic HBV cases are in need of treatment. Current standard therapy for chronic HBV blocks reversed transcription. As this therapy

  5. Superior protection conferred by inactivated whole virus vaccine over subunit and DNA vaccines against salmonid alphavirus infection in Atlantic salmon (Salmo salar L.).

    Science.gov (United States)

    Xu, Cheng; Mutoloki, Stephen; Evensen, Øystein

    2012-06-06

    Salmonid alphavirus 3 (SAV-3) is an emerging pathogen in Norwegian salmon farming and causes severe annual losses. We studied the immunogenicity and protective ability of subunit and DNA vaccines based on E1 and E2 spike proteins of salmonid alphavirus subtype 3 (SAV-3), and compared these to an experimental inactivated, whole virus (IWV) vaccine in Atlantic salmon. The antigens were delivered as water-in-oil emulsions for the subunit and inactivated vaccines and non-formulated for the DNA vaccines. The IWV and the E2 subunit prime-boost groups had circulating neutralizing antibodies at challenge, correlating with high protection against lethal challenge and 3-log(10) reduction of virus titer in heart for the IWV group. Prime-boost with E1 subunit vaccine also conferred significant protection against mortality, but did not correlate with neutralizing antibody levels. Protection against pathology in internal organs was only seen for the IWV group. Prime-boost with E1 and E2 DNA vaccines showed marginal protection in terms of reduction of viral replication in target organs and protection against mortality was not different from controls. The IWV group showed significant upregulation of IFNγ and IL2 mRNA expression at 4 weeks post challenge possibly indicating that other mechanisms in addition to antibody responses play a role in mediating protection against infection. This is the first report comparing the immunogenicity and protection against mortality for IWV vaccines and spike protein subunit and DNA vaccines against salmonid alphavirus infection in Atlantic salmon. The IWV vaccine has superior immunogenicity over sub-unit and DNA vaccines.

  6. Recombinant DNA technology for melanoma immunotherapy: anti-Id DNA vaccines targeting high molecular weight melanoma-associated antigen.

    Science.gov (United States)

    Barucca, A; Capitani, M; Cesca, M; Tomassoni, D; Kazmi, U; Concetti, F; Vincenzetti, L; Concetti, A; Venanzi, F M

    2014-11-01

    Anti-idiotypic MK2-23 monoclonal antibody (anti-Id MK2-23 mAb), which mimics the high molecular weight melanoma-associated antigen (HMW-MAA), has been used to implement active immunotherapy against melanoma. However, due to safety and standardization issues, this approach never entered extensive clinical trials. In the present study, we investigated the usage of DNA vaccines as an alternative to MK2-23 mAb immunization. MK2-23 DNA plasmids coding for single chain (scFv) MK2-23 antibody were constructed via the insertion of variable heavy (V H) and light (V L) chains of MK2-23 into the pVAC-1mcs plasmids. Two alternative MK2-23 plasmids format V H/V L, and V L/V H were assembled. We demonstrate that both polypeptides expressed by scFv plasmids in vitro retained the ability to mimic HMW-MAA antigen, and to elicit specific anti-HMW-MAA humoral and cellular immunoresponses in immunized mice. Notably, MK2-23 scFv DNA vaccines impaired the onset and growth of transplantable B16 melanoma cells not engineered to express HMW-MAA. This pilot study suggests that optimized MK2-23 scFv DNA vaccines could potentially provide a safer and cost-effective alternative to anti-Id antibody immunization, for melanoma immunotherapy.

  7. Comparative evaluation of therapeutic DNA vaccines against Trypanosoma cruzi in mice

    OpenAIRE

    2007-01-01

    Chagas disease, caused by the protozoan parasite Trypanosoma cruzi, is a major public health problem in most of Latin America. A key priority is the development of new treatments, due to the poor efficacy of current ones. We report here the comparative evaluation of therapeutic DNA vaccines encoding various T. cruzi antigens. ICR mice infected with 500 parasites intraperitoneally were treated at 5 and 12 days postinfection with 20 mu g of plasmid DNA encoding T. cruzi antigens TSA-1, TS, ASP-...

  8. Electroporation mediated DNA vaccination directly to a mucosal surface results in improved immune responses

    OpenAIRE

    Kichaev, Gleb; Mendoza, Janess M; Amante, Dinah; Smith, Trevor RF; McCoy, Jay R; Sardesai, Niranjan Y.; Kate E. Broderick

    2013-01-01

    In vivo electroporation (EP) has been shown to be a highly efficient non-viral method for enhancing DNA vaccine delivery and immunogenicity, when the site of immunization is the skin or muscle of animals and humans. However, the route of entry for many microbial pathogens is via the mucosal surfaces of the human body. We have previously reported on minimally invasive, surface and contactless EP devices for enhanced DNA delivery to dermal tissue. Robust antibody responses were induced followin...

  9. Construction and immunogenicity of a codon-optimized Entamoeba histolytica Gal-lectin-based DNA vaccine.

    Science.gov (United States)

    Gaucher, Denis; Chadee, Kris

    2002-09-10

    Invasive amebiasis caused by Entamoeba histolytica is the third leading parasitic cause of mortality, and there are no vaccines available to help control the disease. The galactose-adherence lectin (Gal-lectin) is the parasite's major molecule allowing it to adhere to colonic mucin for colonization and to target cells for tissue destruction. It is immunodominant and is regarded as the most promising candidate molecule to be included in a subunit vaccine against amebiasis. In this study, we are reporting the construction of a codon-optimized DNA vaccine encoding a portion of the Gal-lectin heavy subunit that includes the carbohydrate recognition domain (CRD), and its in vivo testing in mice. The vaccine stimulated a Th1-type Gal-lectin-specific cellular immune response as well as the development of serum antibodies that recognized a recombinant portion of the heavy subunit, and that inhibited the adherence of trophozoites to target cells in vitro.

  10. The role of peptide and DNA vaccines in myeloid leukemia immunotherapy

    Directory of Open Access Journals (Sweden)

    Lin Chen

    2013-02-01

    Full Text Available Abstract While chemotherapy and targeted therapy are successful in inducing the remission of myeloid leukemia as acute myeloid leukemia (AML and chronic myeloid leukemia (CML, the disease remains largely incurable. This observation is likely due to the drug resistance of leukemic cells, which are responsible for disease relapse. Myeloid leukemia vaccines may most likely be beneficial for eradicating minimal residual disease after treatment with chemotherapy or targeted therapy. Several targeted immunotherapies using leukemia vaccines have been heavily investigated in clinical and preclinical trials. This review will focus on peptides and DNA vaccines in the context of myeloid leukemias, and optimal strategies for enhancing the efficacy of vaccines based on myeloid leukemia immunization are also summarized.

  11. Vaccinations

    Science.gov (United States)

    ... vaccinated? For many years, a set of annual vaccinations was considered normal and necessary for dogs and ... to protect for a full year. Consequently, one vaccination schedule will not work well for all pets. ...

  12. Efficacy of an autophagy-targeted DNA vaccine against avian leukosis virus subgroup J.

    Science.gov (United States)

    Dai, Zhenkai; Huang, Jianfei; Lei, Xiaoya; Yan, Yiming; Lu, Piaopiao; Zhang, Huanmin; Lin, Wencheng; Chen, Weiguo; Ma, Jingyun; Xie, Qingmei

    2017-02-01

    Infection with the avian leukosis virus subgroup J (ALV-J) can lead to neoplastic disease in chickens, inflicting significant economic losses to the poultry industry. Recent reports have identified inhibitory effects of ALV-J on autophagy, a process involving in innate and adaptive immunity. Inspired by this connection between autophagy and immunity, we developed a novel DNA vaccine against ALV-J which includes co-administration of rapamycin to stimulate autophagy. To measure the efficacy of the developed prototype vaccine, five experimental groups of seven-day-old chickens was immunized three times at three-week intervals respectively with vector, pVAX1-gp85, pVAX1-gp85-LC3, pVAX1-gp85+rapamycin and pVAX1-gp85-LC3+rapamycin through electroporation. We then tested their antibody titers, cytokine levels and cellular immune responses. The immunoprotective efficacy of the prototype vaccines against the challenge of the ALV-J GD1109 strain was also examined. The results showed that the combination of pVAX1-gp85-LC3 and rapamycin was able to induce the highest antibody titers, and enhance interleukin(IL)-2, IL-10 and interferon (IFN)-γ expression, and the chickens immunized with the combination of pVAX1-gp85-LC3 and rapamycin showed the highest percentage of CD3+CD8+T lymphocytes. Based on our results, we suggest that stimulating autophagy can improve the efficacy of DNA vaccines and that our DNA vaccine shows the potential of being a candidate vaccine against ALV-J. This study provides a novel strategy for developing vaccines against ALV-J.

  13. Combined virus-like particle and fusion protein-encoding DNA vaccination of cotton rats induces protection against respiratory syncytial virus without causing vaccine-enhanced disease

    Energy Technology Data Exchange (ETDEWEB)

    Hwang, Hye Suk; Lee, Young-Tae; Kim, Ki-Hye; Park, Soojin; Kwon, Young-Man; Lee, Youri; Ko, Eun-Ju; Jung, Yu-Jin [Center for Inflammation, Immunity & Infection, Institute for Biomedical Sciences and Department of Biology, Georgia State University, Atlanta, GA (United States); Lee, Jong Seok [Center for Inflammation, Immunity & Infection, Institute for Biomedical Sciences and Department of Biology, Georgia State University, Atlanta, GA (United States); National Institute of Biological Resources, Incheon (Korea, Republic of); Kim, Yu-Jin [Center for Inflammation, Immunity & Infection, Institute for Biomedical Sciences and Department of Biology, Georgia State University, Atlanta, GA (United States); Lee, Yu-Na; Kim, Min-Chul [Center for Inflammation, Immunity & Infection, Institute for Biomedical Sciences and Department of Biology, Georgia State University, Atlanta, GA (United States); Animal and Plant Quarantine Agency, Gyeonggi-do, Gimcheon, Gyeongsangbukdo (Korea, Republic of); Cho, Minkyoung [Center for Inflammation, Immunity & Infection, Institute for Biomedical Sciences and Department of Biology, Georgia State University, Atlanta, GA (United States); Kang, Sang-Moo, E-mail: skang24@gsu.edu [Center for Inflammation, Immunity & Infection, Institute for Biomedical Sciences and Department of Biology, Georgia State University, Atlanta, GA (United States)

    2016-07-15

    A safe and effective vaccine against respiratory syncytial virus (RSV) should confer protection without causing vaccine-enhanced disease. Here, using a cotton rat model, we investigated the protective efficacy and safety of an RSV combination vaccine composed of F-encoding plasmid DNA and virus-like particles containing RSV fusion (F) and attachment (G) glycoproteins (FFG-VLP). Cotton rats with FFG-VLP vaccination controlled lung viral replication below the detection limit, and effectively induced neutralizing activity and antibody-secreting cell responses. In comparison with formalin inactivated RSV (FI-RSV) causing severe RSV disease after challenge, FFG-VLP vaccination did not cause weight loss, airway hyper-responsiveness, IL-4 cytokines, histopathology, and infiltrates of proinflammatory cells such as eosinophils. FFG-VLP was even more effective in preventing RSV-induced pulmonary inflammation than live RSV infections. This study provides evidence that FFG-VLP can be developed into a safe and effective RSV vaccine candidate. - Highlights: • Combined RSV FFG VLP vaccine is effective in inducing F specific responses. • FFG VLP vaccine confers RSV neutralizing activity and viral control in cotton rats. • Cotton rats with RSV FFG VLP vaccination do not show vaccine-enhanced disease. • Cotton rats with FFG VLP vaccine induce F specific antibody secreting cell responses. • Cotton rats with FFG VLP do not induce lung cellular infiltrates and Th2 cytokine.

  14. Effectiveness of DNA-recombinant anti-hepatitis B vaccines in blood donors: a cohort study

    Directory of Open Access Journals (Sweden)

    Petry Andrea

    2007-11-01

    Full Text Available Abstract Background Although various studies have demonstrated efficacy of DNA-recombinant anti-hepatitis B vaccines, their effectiveness in health care settings has not been researched adequately. This gap is particularly visible for blood donors, a group of significant importance in the reduction of transfusion-transmitted hepatitis B. Methods This is a double cohort study of 1411 repeat blood donors during the period 1998–2002, involving a vaccinated and an unvaccinated cohort, with matching of the two in terms of sex, age and residence. Average follow-up was 3.17 person-years. The outcome measure was infection with hepatitis B virus (HBV, defined by testing positive on serologic markers HBsAg or anti-HBC. All blood donors were from the blood bank in Joaçaba, federal state of Santa Catarina, Brazil. Results The cohorts did not differ significantly regarding sex, age and marital status but the vaccinated cohort had higher mean number of blood donations and higher proportion of those residing in the county capital Joaçaba. Hepatitis B incidences per 1000 person-years were zero among vaccinated and 2,33 among non-vaccinated, resulting in 100% vaccine effectiveness with 95% confidence interval from 30,1% to 100%. The number of vaccinated persons necessary to avoid one HBV infection in blood donors was estimated at 429 with 95% confidence interval from 217 to 21422. Conclusion The results showed very high effectiveness of DNA-recombinant anti-HBV vaccines in blood donors. Its considerable variation in this study is likely due to the limited follow-up and the influence of confounding factors normally balanced out in efficacy clinical trials.

  15. Evaluation of attenuated Salmonella choleraesuis-mediated inhibin recombinant DNA vaccine in rats.

    Science.gov (United States)

    Hui, F M; Meng, C L; Guo, N N; Yang, L G; Shi, F X; Mao, D G

    2014-08-07

    DNA vaccination has been studied intensively as a potential vaccine technology. We evaluated the effect of an attenuated Salmonella choleraesuis-mediated inhibin DNA vaccine in rats. First, 15 rats were treated with different doses of an inhibin vaccine to evaluate vaccine safety. Next, 30 rats were divided into 3 groups and injected intramuscularly with the inhibin vaccine two (T1) or three times (T2) or with control bacteria (Con) at 4-week intervals. The inhibin antibody levels increased [positive/negative well (P/N) value: T1 vs Con = 2.39 ± 0.01 vs 1.08 ± 0.1; T2 vs Con = 2.36 ± 0.1 vs 1.08 ± 0.1, P < 0.05] at week 2 and were maintained at a high level in T1 and T2 until week 8, although a small decrease in T2 was observed at week 10. Rats in the T1 group showed more corpora lutea compared with the Con group (10.50 ± 0.87 vs 7.4 ± 0.51, P < 0.05). Estradiol (0.439 ± 0.052 vs 0.719 ± 0.063 ng/mL, P < 0.05) and progesterone (1.315 ± 0.2 vs 0.737 ± 0.11 ng/mL, P < 0.05) levels differed significantly at metestrus after week 10 between rats in the T1 and Con groups. However, there were no significant differences in body, ovary, uterus weights, or pathological signs in the ovaries after immunization, indicating that this vaccine is safe. In conclusion, the attenuated S. choleraesuis-mediated inhibin vaccine may be an alternative to naked inhibin plasmids for stimulating ovarian follicular development to increase the ovulation rate in rats.

  16. Comparative evaluation of therapeutic DNA vaccines against Trypanosoma cruzi in mice.

    Science.gov (United States)

    Sanchez-Burgos, Gilma; Mezquita-Vega, R Gabino; Escobedo-Ortegon, Javier; Ramirez-Sierra, Maria Jesus; Arjona-Torres, Arletty; Ouaissi, Ali; Rodrigues, Mauricio M; Dumonteil, Eric

    2007-08-01

    Chagas disease, caused by the protozoan parasite Trypanosoma cruzi, is a major public health problem in most of Latin America. A key priority is the development of new treatments, due to the poor efficacy of current ones. We report here the comparative evaluation of therapeutic DNA vaccines encoding various T. cruzi antigens. ICR mice infected with 500 parasites intraperitoneally were treated at 5 and 12 days postinfection with 20 microg of plasmid DNA encoding T. cruzi antigens TSA-1, TS, ASP-2-like, Tc52 or Tc24. Treatment with plasmid encoding TS and/or ASP-2-like antigens had no significant effect on parasitemia or survival. Treatment with Tc52 DNA significantly reduced parasitemia, as well as cardiac parasite burden, and improved survival, although myocarditis was not significantly affected. Finally, treatment with plasmids encoding Tc24 and TSA-1 induced the most complete control of disease as evidenced by significant reductions in parasitemia, mortality, myocarditis and heart parasite burden. These data demonstrate that therapeutic vaccine efficacy is dependent on the antigen and suggest that DNA vaccines encoding Tc24, TSA-1, and Tc52 represent the best candidates for further studies of a therapeutic vaccine against Chagas disease.

  17. Adverse effects of feline IL-12 during DNA vaccination against feline infectious peritonitis virus

    NARCIS (Netherlands)

    Horzinek, M.C.; Haagmans, B.L.; Lintelo, E.G. te; Egberink, H.F.; Duquesne, V.; Aubert, A.; Rottier, P.J.M.

    2002-01-01

    Cell-mediated immunity is thought to play a decisive role in protecting cats against feline infectious peritonitis (FIP), a progressive and lethal coronavirus disease. In view of the potential of DNA vaccines to induce cell-mediated responses, their efficacy to induce protective immunity in cats was

  18. 78 FR 29698 - Availability of an Environmental Assessment for Field Testing a Canine Lymphoma Vaccine, DNA

    Science.gov (United States)

    2013-05-21

    ... Service has prepared an environmental assessment concerning authorization to ship for the purpose of field testing, and then to field test, an unlicensed Canine Lymphoma Vaccine, DNA. The environmental assessment... Animal and Plant Health Inspection Service Availability of an Environmental Assessment for Field...

  19. Characterization of immune responses induced by inactivated, live attenuated and DNA vaccines against Japanese encephalitis virus in mice.

    Science.gov (United States)

    Li, Jieqiong; Chen, Hui; Wu, Na; Fan, Dongying; Liang, Guodong; Gao, Na; An, Jing

    2013-08-28

    Vaccination is the most effective countermeasure for protecting individuals from Japanese encephalitis virus (JEV) infection. There are two types of JEV vaccines currently used in China: the Vero cell-derived inactivated vaccine and the live attenuated vaccine. In this study, we characterized the immune response and protective efficacy induced in mice by the inactivated vaccine, live attenuated vaccine and the DNA vaccine candidate pCAG-JME, which expresses JEV prM-E proteins. We found that the live attenuated vaccine conferred 100% protection and resulted in the generation of high levels of specific anti-JEV antibodies and cytokines. The pCAG-JME vaccine induced protective immunity as well as the live attenuated vaccine. Unexpectedly, immunization with the inactivated vaccine only induced a limited immune response and partial protection, which may be due to the decreased activity of dendritic cells and the expansion of CD4+CD25+Foxp3+ regulatory T cells observed in these mice. Altogether, our results suggest that the live attenuated vaccine is more effective in providing protection against JEV infection than the inactivated vaccine and that pCAG-JME will be a potential JEV vaccine candidate. Copyright © 2013 Elsevier Ltd. All rights reserved.

  20. Polyclonal antibody cocktails generated using DNA vaccine technology protect in murine models of orthopoxvirus disease

    Directory of Open Access Journals (Sweden)

    Ballantyne John

    2011-09-01

    Full Text Available Abstract Background Previously we demonstrated that DNA vaccination of nonhuman primates (NHP with a small subset of vaccinia virus (VACV immunogens (L1, A27, A33, B5 protects against lethal monkeypox virus challenge. The L1 and A27 components of this vaccine target the mature virion (MV whereas A33 and B5 target the enveloped virion (EV. Results Here, we demonstrated that the antibodies produced in vaccinated NHPs were sufficient to confer protection in a murine model of lethal Orthopoxvirus infection. We further explored the concept of using DNA vaccine technology to produce immunogen-specific polyclonal antibodies that could then be combined into cocktails as potential immunoprophylactic/therapeutics. Specifically, we used DNA vaccines delivered by muscle electroporation to produce polyclonal antibodies against the L1, A27, A33, and B5 in New Zealand white rabbits. The polyclonal antibodies neutralized both MV and EV in cell culture. The ability of antibody cocktails consisting of anti-MV, anti-EV, or a combination of anti-MV/EV to protect BALB/c mice was evaluated as was the efficacy of the anti-MV/EV mixture in a mouse model of progressive vaccinia. In addition to evaluating weight loss and lethality, bioimaging technology was used to characterize the spread of the VACV infections in mice. We found that the anti-EV cocktail, but not the anti-MV cocktail, limited virus spread and lethality. Conclusions A combination of anti-MV/EV antibodies was significantly more protective than anti-EV antibodies alone. These data suggest that DNA vaccine technology could be used to produce a polyclonal antibody cocktail as a possible product to replace vaccinia immune globulin.

  1. Immunity induced shortly after DNA vaccination of rainbow trout against rhabdoviruses protects against heterologous virus but not against bacterial pathogens

    DEFF Research Database (Denmark)

    Lorenzen, Niels; Lorenzen, Ellen; Einer-Jensen, Katja;

    2002-01-01

    It was recently reported that DNA vaccination of rainbow trout fingerlings against viral hemorrhagic septicaemia virus (VHSV) induced protection within 8 days after intramuscular injection of plasmid DNA. In order to analyse the specificity of this early immunity, fish were vaccinated with plasmid...... DNA encoding the VHSV or the infectious haematopoietic necrosis virus (IHNV) glycoprotein genes and later challenged with homologous or heterologous pathogens. Challenge experiments revealed that immunity established shortly after vaccination was cross-protective between the two viral pathogens...... whereas no increased survival was found upon challenge with bacterial pathogens. Within two months after vaccination, the cross-protection disappeared while the specific immunity to homologous virus remained high. The early immunity induced by the DNA vaccines thus appeared to involve short-lived non...

  2. Individual and bivalent vaccines against botulinum neurotoxin serotypes A and B using DNA-based Semliki Forest virus vectors.

    Science.gov (United States)

    Yu, Yunzhou; Yu, Jiyu; Li, Na; Wang, Shuang; Yu, Weiyuan; Sun, Zhiwei

    2009-10-19

    We evaluated individual and bivalent replicon vaccines against Clostridiumbotulinum neurotoxin serotypes A (BoNT/A) or B (BoNT/B). The DNA replicon vaccine (pSCARSBHc) encoding the Hc domain of BoNT/B (BHc) induced better responses and protection against BoNT/B mouse challenge than conventional DNA vaccine. The dual-expressing DNA vaccine (pSCARSA/BHc) protected similarly to a DNA replicon vaccine mixture (pSCARSAHc+pSCARSBHc). Additionally, recombinant SFV particles, VRP-AHc or VRP-BHc, protected mice from high-dose BoNT/A or BoNT/B challenge, respectively. Mice given either dual-expressing VRP-A/BHc or mixture of VRP-AHc and VRP-BHc were protected from challenge with serotype A/B mixtures. These data justify further testing in other animals or humans.

  3. Construction of a recombinant attenuated Salmonella typhimurium DNA vaccine carrying Helicobacter pylori hpaA

    Institute of Scientific and Technical Information of China (English)

    Can Xu; Zhao-Shen Li; Yi-Qi Du; Zhen-Xing Tu; Yan-Fang Gong; Jing Jin; Hong-Yu Wu; Guo-Ming Xu

    2005-01-01

    AIM: To construct a recombinant attenuated Salmonella typhimurium DNA vaccine carrying Helicobacter pylori hpaA gene and to detect its immunogenicity.METHODS: Genomic DNA of the standard H pylori strain 17 874 was isolated as the template, hpaA gene fragment was amplified by polymerase chain reaction (PCR) and cloned into pUCmT vector. DNA sequence of the amplified hpaA gene was assayed, then cloned into the eukaryotic expression vector pIRES through enzyme digestion and ligation reactions. The recombinant plasmid was used to transform competent Escherichia coliDH5α, and the positive clones were screened by PCR and restriction enzyme digestion. Then, the recombinant pIRES-hpaA was used to transform LB5000 and the recombinant plasmid isolated from LB5000 was finally used to transform SL7207. After that, the recombinant strain was grown in vitrorepeatedly. In order to iclentify the immunogenicity of the vaccinein vitro, the recombinant pIRES-hpaA was transfected to COS-7 cells using LipofectamineTM2000, the immunogenicity of expressed HpaA protein was detected with SDS-PAGE and Western blot.RESULTS: The 750-base pair hpaA gene fragment was amplified from the genomic DNA and was consistent with the sequence of H pylori hpaA by sequence analysis. It was confirmed by PCR and restriction enzyme digestion that H pylori hpaA gene was inserted into the eukaryotic expression vector pIRES and a stable recombinant live attenuated Salmonella typhimurium DNA vaccine carrying H pylori hpaA gene was successfully constructed and the specific strip of HpaA expressed by pIRES-hpaA was detected through Western blot.CONCLUSION: The recombinant attenuated Salmonella typhimurium DNA vaccine strain expressing HpaA protein with immunogenicity can be constructed and it may be helpful for further investigating the immune action of DNA vaccine in vivo.

  4. Enhancement of DNA vaccine potency through linkage of antigen to filamentous bacteriophage coat protein III domain I

    DEFF Research Database (Denmark)

    Cuesta, Àngel M; Suárez, Eduardo; Larsen, Martin

    2006-01-01

    Although DNA-based cancer vaccines have been successfully tested in mouse models, a major drawback of cancer vaccination still remains, namely that tumour antigens are weak and fail to generate a vigorous immune response in tumour-bearing patients. Genetic technology offers strategies for promoting...... immune pathways by adding immune-activating genes to the tumour antigen sequence. In this work, we converted a model non-immunogenic antigen into a vaccine by fusing it to domain I of the filamentous bacteriophage coat protein III gene. Vaccination with a DNA construct encoding the domain I fusion...

  5. Enhancing the Immunogenicity of a Tetravalent Dengue DNA Vaccine

    Science.gov (United States)

    2016-08-01

    season’s influenza vaccine. There is no overlap with the proposed project. Title: Serological survey for Zika virus and other vector -borne pathogen...measures during future deployments to the region. Aim1: Determine the seroprevalence and seroincidence of antibodies against a range of vector -borne...potentially high burden for vector -borne diseases. Aim 2: Determine the pathogens associated with overt febrile illness. There is no overlap with the

  6. Linear synthesis and immunological properties of a fully synthetic vaccine candidate containing a sialylated MUC1 glycopeptide

    NARCIS (Netherlands)

    Thompson, Pamela; Lakshminarayanan, Vani; Supekar, Nitin T.; Bradley, Judy M.; Cohen, Peter A.; Wolfert, Margreet A.; Gendler, Sandra J.; Boons, Geert Jan

    2015-01-01

    A strategy for the linear synthesis of a sialylated glycolipopeptide cancer vaccine candidate has been developed using a strategically designed sialyl-Tn building block and microwave-assisted solid-phase peptide synthesis. The glycolipopeptide elicited potent humoral and cellular immune responses. T

  7. Oriented and vectorial immobilization of linear M13 dsDNA between interdigitated electrodes--towards single molecule DNA nanostructures.

    Science.gov (United States)

    Hölzel, Ralph; Gajovic-Eichelmann, Nenad; Bier, Frank F

    2003-05-01

    The ability to control molecules at a resolution well below that offered by photolithography has gained much interest recently. DNA is a promising candidate for this task since it offers excellent specificity in base-pairing combined with addressability at the nanometer scale. New applications in biosensing, e.g. interaction analysis at the single molecule level, or nanobiotechnology, e.g. ultradense DNA microarrays, have been devised that rely on stretched DNA bridges. The basic technology required is the ability to deposit spatially defined, stretched DNA-bridges between anchoring structures on surfaces. In this paper we present two techniques for spanning 2 microm long dsDNA bridges between neighboring interdigitated electrodes (IDEs). The extended DNA used was linearized M13 dsDNA (M13mp18 7231 bp, ca. 2.5 microm length), either unmodified, or with chemical modifications at both ends. The first approach is based on the dielectrophoretic (DEP) concentration and alignment of linearized wild-type dsDNA. IDEs with 1.7 microm spacing are driven with an AC voltage around 1 MHz leading to field strengths in the order of 1 MV m(-1). The dsDNA is polarized and linearized by the force field and accumulates in the gap between two neighboring electrodes. This process is reversible and was visualized by fluorescence staining of M13 DNA using PicoGreen, as intercalating dye. The resulting dsDNA bridges and their orientation are discernible under the fluorescence microscope using fluorescent particles of different color. The particles are tagged with sequence specific peptide nucleic acid (PNA) probes that bind to the DNA double strand at specific sites. The second approach is based on asymmetric electrochemical modification of a gold IDE with 2.0 microm spacings followed by spontaneous or stimulated deposition of a chemically modified M13-DNA. One side of the IDE was selectively coated with streptavidin by electropolymerization of a novel hydrophilic conductive polymer in

  8. Antitumor immunity induced by DNA vaccine encoding alpha-fetoprotein/heat shock protein 70

    Institute of Scientific and Technical Information of China (English)

    Xiao-Ping Wang; Guo-Zhen Liu; Ai-Li Song; Hai-Yan Li; Yu Liu

    2004-01-01

    AIM: To construct a DNA vaccine encoding human alphafetoprotein (hAFP)/heat shock protein 70 (HSP70), and to study its ability to induce specific CTL response and its protective effect against AFP-expressing tumor.METHODS: A DNA vaccine was constructed by combining hAFP gene with HSP70 gene. SP2/0 cells were stably transfected with pBBS212-hAFP and pBBS212-hAFP/HSP70eukaryotic expression vectors. Mice were primed and boosted with DNA vaccine hAFP/HSP70 by intramuscular injection, whereas plasmid with hAFP or HSP70 was used as controls. ELISPOT and ELISA were used to detect IFN-γ-producing splenocytes and the level of serum anti-AFP antibody from immunized mice respectively. In vivo tumor challenge was measured to assess the immune effect of the DNA vaccine.RESULTS: By DNA vaccine immunization, the results of ELISPOT and ELISA showed that the number of IFN-γ-producing splenocytes and the level of serum anti-AFP antibody were significantly higher in rhAFP/HSP70 group than in hAFP and empty plasmid groups (95.50±10.90IFN-γ spots/106 cells vs 23.60±11.80 IFN-γ spots/106 cells,7.17±4.24 IFN-γ spots/106 cells, P<0.01; 126.50±8.22 μg/mL vs 51.72±3.40 μg/mL, 5.83±3.79 μg/mL, P<0.01). The tumor volume in rhAFP/HSP70 group was significantly smaller than that in pBBS212-hAFP and empty plasmid groups (37.41±7.34 mm3 vs381.13±15.48 mm3, 817.51±16.25 mm3,P<0.01).CONCLUSION: Sequential immunization with a recombinant DNA vaccine encoding AFP and heat shock protein70 could generate effective AFP-specific T cell responses and induce definite antitumor effects on AFP-producing tumors, which may be suitable for some clinical testing as a vaccine for HCC.

  9. Tumor prevention in HPV8 transgenic mice by HPV8-E6 DNA vaccination.

    Science.gov (United States)

    Marcuzzi, Gian Paolo; Awerkiew, Sabine; Hufbauer, Martin; Schädlich, Lysann; Gissmann, Lutz; Eming, Sabine; Pfister, Herbert

    2014-06-01

    The genus beta human papillomavirus 8 (HPV8) is involved in the development of cutaneous squamous cell carcinomas (SCCs) in individuals with epidermodysplasia verruciformis. Immunosuppressed transplant recipients are prone to harbor particularly high betapapillomavirus DNA loads, which may contribute to their highly increased risk of SCC. Tumor induction in HPV8 transgenic mice correlates with increased expression of viral oncogenes E6 and E2. In an attempt to prevent skin tumor development, we evaluated an HPV8-E6-DNA vaccine, which was able to stimulate a detectable HPV8-E6-specific cell-mediated immune response in 8/15 immunized mice. When skin of HPV8 transgenic mice was grafted onto non-transgenic littermates, the grafted HPV8 transgenic tissue was not rejected and papillomas started to grow within 14 days all over the transplant of 9/9 non-vaccinated and 7/15 not successfully vaccinated mice. In contrast, no papillomas developed in 6/8 successfully vaccinated mice. In the other two of these eight mice, a large ulcerative lesion developed within the initial papilloma growth or papilloma development was highly delayed. As the vaccine completely or partially prevented papilloma development without rejecting the transplanted HPV8 positive skin, the immune system appears to attack only keratinocytes with increased levels of E6 protein, which would give rise to papillomas.

  10. Antiangiogenic immunotherapy targeting Flk-1, DNA vaccine and adoptive T cell transfer, inhibits ocular neovascularization

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Han [Department of Ophthalmology, Graduate School of Medical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-Ku, Fukuoka 812-8582 (Japan); Sonoda, Koh-Hei, E-mail: sonodak@med.kyushu-u.ac.jp [Department of Ophthalmology, Graduate School of Medical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-Ku, Fukuoka 812-8582 (Japan); Hijioka, Kuniaki; Qiao, Hong; Oshima, Yuji; Ishibashi, Tatsuro [Department of Ophthalmology, Graduate School of Medical Sciences, Kyushu University, 3-1-1 Maidashi, Higashi-Ku, Fukuoka 812-8582 (Japan)

    2009-04-17

    Ocular neovascularization (NV) is the primary cause of blindness in a wide range of ocular diseases. The exact mechanism underlying the pathogenesis of ocular NV is not yet well understood, and so there is no satisfactory therapy for ocular NV. Here, we describe a strategy targeting Flk-1, a self-antigen overexpressed on proliferating endothelial cells in ocular NV, by antiangiogenic immunotherapy-DNA vaccine and adoptive T cell therapy. An oral DNA vaccine encoding Flk-1 carried by attenuated Salmonella typhimurium markedly suppressed development of laser-induced choroidal NV. We further demonstrated that adoptive transfer of vaccine-induced CD8{sup +} T cells reduced pathological preretinal NV, with a concomitant facilitation of physiological revascularization after oxygen-induced retinal vessel obliteration. However, physiological retinal vascular development was unaffected in neonatal mice transferred with vaccine-induced CD8{sup +} T cells. These findings suggested that antiangiogenic immunotherapy targeting Flk-1 such as vaccination and adoptive immunotherapy may contribute to future therapies for ocular NV.

  11. Efficacy of a DNA vaccine carrying Eimeria maxima Gam56 antigen gene against coccidiosis in chickens.

    Science.gov (United States)

    Xu, Jinjun; Zhang, Yan; Tao, Jianping

    2013-04-01

    To control coccidiosis without using prophylactic medications, a DNA vaccine targeting the gametophyte antigen Gam56 from Eimeria maxima in chickens was constructed, and the immunogenicity and protective effects were evaluated. The ORF of Gam56 gene was cloned into an eukaryotic expression vector pcDNA3.1(zeo)+. Expression of Gam56 protein in COS-7 cells transfected with recombinant plasmid pcDNA-Gam56 was confirmed by indirect immunofluorescence assay. The DNA vaccine was injected intramuscularly to yellow feathered broilers of 1-week old at 3 dosages (25, 50, and 100 µg/chick). Injection was repeated once 1 week later. One week after the second injection, birds were challenged orally with 5×10(4) sporulated oocysts of E. maxima, then weighed and killed at day 8 post challenge. Blood samples were collected and examined for specific peripheral blood lymphocyte proliferation activity and serum antibody levels. Compared with control groups, the administration of pcDNA-Gam56 vaccine markedly increased the lymphocyte proliferation activity (Pcoccidiosis control.

  12. Multivalent HA DNA vaccination protects against highly pathogenic H5N1 avian influenza infection in chickens and mice.

    Directory of Open Access Journals (Sweden)

    Srinivas Rao

    Full Text Available BACKGROUND: Sustained outbreaks of highly pathogenic avian influenza (HPAI H5N1 in avian species increase the risk of reassortment and adaptation to humans. The ability to contain its spread in chickens would reduce this threat and help maintain the capacity for egg-based vaccine production. While vaccines offer the potential to control avian disease, a major concern of current vaccines is their potency and inability to protect against evolving avian influenza viruses. METHODOLOGY / PRINCIPAL FINDINGS: The ability of DNA vaccines encoding hemagglutinin (HA proteins from different HPAI H5N1 serotypes was evaluated for its ability to elicit neutralizing antibodies and to protect against homologous and heterologous HPAI H5N1 strain challenge in mice and chickens after DNA immunization by needle and syringe or with a pressure injection device. These vaccines elicited antibodies that neutralized multiple strains of HPAI H5N1 when given in combinations containing up to 10 HAs. The response was dose-dependent, and breadth was determined by the choice of the influenza virus HA in the vaccine. Monovalent and trivalent HA vaccines were tested first in mice and conferred protection against lethal H5N1 A/Vietnam/1203/2004 challenge 68 weeks after vaccination. In chickens, protection was observed against heterologous strains of HPAI H5N1 after vaccination with a trivalent H5 serotype DNA vaccine with doses as low as 5 microg DNA given twice either by intramuscular needle injection or with a needle-free device. CONCLUSIONS/SIGNIFICANCE: DNA vaccines offer a generic approach to influenza virus immunization applicable to multiple animal species. In addition, the ability to substitute plasmids encoding different strains enables rapid adaptation of the vaccine to newly evolving field isolates.

  13. Two doses of bovine viral diarrhea virus DNA vaccine delivered by electroporation induce long-term protective immune responses.

    Science.gov (United States)

    van Drunen Littel-van den Hurk, Sylvia; Lawman, Zoe; Snider, Marlene; Wilson, Don; van den Hurk, Jan V; Ellefsen, Barry; Hannaman, Drew

    2013-02-01

    Bovine viral diarrhea virus (BVDV) is a pathogen of major importance in cattle, so there is a need for new effective vaccines. DNA vaccines induce balanced immune responses and are relatively inexpensive and thus promising for both human and veterinary applications. In this study, newborn calves with maternal antibodies were vaccinated intramuscularly (i.m.) with a BVDV E2 DNA vaccine with the TriGrid Delivery System for i.m. delivery (TDS-IM). Two doses of this vaccine spaced 6 or 12 weeks apart were sufficient to induce significant virus-neutralizing antibody titers, numbers of activated T cells, and reduction in viral shedding and clinical presentations after BVDV-2 challenge. In contrast to the placebo-treated animals, the vaccinated calves did not lose any weight, which is an excellent indicator of the well-being of an animal and has a significant economic impact. Furthermore, the interval between the two vaccinations did not influence the magnitude of the immune responses or degree of clinical protection, and a third immunization was not necessary or beneficial. Since electroporation may enhance not only the magnitude but also the duration of immunity after DNA immunization, the interval between vaccination and challenge was extended in a second trial, which showed that two doses of this E2 DNA vaccine again significantly reduced clinical disease against BVDV for several months. These results are promising and support this technology for use against infectious diseases in cattle and large species, including humans, in general.

  14. Elicitation of strong immune responses by a DNA vaccine expressing a secreted form of hepatitis C virus envelope protein E2 in murine and porcine animal models

    Institute of Scientific and Technical Information of China (English)

    Yi-Ping Li; Hye Na Kang; Lorne A Babiuk; Qiang Liu

    2006-01-01

    AIM: To characterize the immunogenicity of a hepatitis C virus (HCV) E2 DNA vaccine alone or with a protein vaccine boost in murine and porcine animal models.METHODS: A DNA vaccine expressing a secreted form of HCV E2 protein was constructed and used to vaccinate mice and piglets with or without boosting with a recombinant E2 protein vaccine formulated with CpG ODN and 10% Emulsigen. The immunogenicity of HCV E2 vaccines was analyzed by ELISA for antibody responses, MTT assay for lymphocyte proliferation,ELISPOT for the number of interferon-γ secreting cells,and cytotoxic T lymphocyte assays.RESULTS: Intradermal injection of E2 DNA vaccine induced strong Th1-like immune responses in mice. In piglets, E2 DNA vaccine elicited moderate and more balanced immune responses. A DNA vaccine prime and protein boost vaccination strategy induced significantly higher E2-specific antibody levels and shifted the immune response towards Th2-like ones in piglets.CONCLUSION: A DNA vaccine expressing a secreted form of HCV E2 protein elicited E2-specific immune responses in mice and piglets. Recombinant E2 protein vaccination following DNA immunization significantly increased the antibody response in piglets. These HCV E2 vaccines may represent promising hepatitis C vaccine candidates for further investigations.

  15. Immunogenicity and efficacy of codon optimized DNA vaccines encoding the F-protein of respiratory syncytial virus.

    Science.gov (United States)

    Ternette, Nicola; Tippler, Bettina; Uberla, Klaus; Grunwald, Thomas

    2007-10-10

    Respiratory syncytial virus F-protein (RSV-F) is poorly expressed from DNA expression plasmids containing the wild type RSV-F open reading frame. By codon optimization, premature polyadenylation signals were deleted and a striking enhancement of RSV-F expression levels was achieved. Therefore, the immunogenicity and efficacy of wild type DNA vaccines were compared to codon optimized expression plasmids encoding full-length RSV-F or its ectodomain. Mice were immunized twice with the different DNA vaccines followed by an RSV challenge. Only codon optimized DNA vaccines and in particular the one encoding the ectodomain of RSV-F induced substantial antibody levels and reduced viral load 13-170-fold. Thus, codon optimization enhances the immunogenicity and efficacy of RSV encoding DNA vaccines.

  16. Evaluation of a novel non-penetrating electrode for use in DNA vaccination.

    Directory of Open Access Journals (Sweden)

    Amy Donate

    Full Text Available Current progress in the development of vaccines has decreased the incidence of fatal and non-fatal infections and increased longevity. However, new technologies need to be developed to combat an emerging generation of infectious diseases. DNA vaccination has been demonstrated to have great potential for use with a wide variety of diseases. Alone, this technology does not generate a significant immune response for vaccination, but combined with delivery by electroporation (EP, can enhance plasmid expression and immunity. Most EP systems, while effective, can be invasive and painful making them less desirable for use in vaccination. Our lab recently developed a non-invasive electrode known as the multi-electrode array (MEA, which lies flat on the surface of the skin without penetrating the tissue. In this study we evaluated the MEA for its use in DNA vaccination using Hepatitis B virus as the infectious model. We utilized the guinea pig model because their skin is similar in thickness and morphology to humans. The plasmid encoding Hepatitis B surface antigen (HBsAg was delivered intradermally with the MEA to guinea pig skin. The results show increased protein expression resulting from plasmid delivery using the MEA as compared to injection alone. Within 48 hours of treatment, there was an influx of cellular infiltrate in experimental groups. Humoral responses were also increased significantly in both duration and intensity as compared to injection only groups. While this electrode requires further study, our results suggest that the MEA has potential for use in electrically mediated intradermal DNA vaccination.

  17. Single-tube linear DNA amplification for genome-wide studies using a few thousand cells

    NARCIS (Netherlands)

    Shankaranarayanan, P.; Mendoza-Parra, M.A.; Gool, van W.; Trindade, L.M.; Gronemeyer, H.

    2012-01-01

    Linear amplification of DNA (LinDA) by T7 polymerase is a versatile and robust method for generating sufficient amounts of DNA for genome-wide studies with minute amounts of cells. LinDA can be coupled to a great number of global profiling technologies. Indeed, chromatin immunoprecipitation coupled

  18. Complete DNA sequence of the linear mitochondrial genome of the pathogenic yeast Candida parapsilosis

    DEFF Research Database (Denmark)

    Nosek, J.; Novotna, M.; Hlavatovicova, Z.

    2004-01-01

    The complete sequence of the mitochondrial DNA of the opportunistic yeast pathogen Candida parapsilosis was determined. The mitochondrial genome is represented by linear DNA molecules terminating with tandem repeats of a 738-bp unit. The number of repeats varies, thus generating a population...

  19. Complete DNA sequence of the linear mitochondrial genome of the pathogenic yeast Candida parapsilosis

    DEFF Research Database (Denmark)

    Nosek, J.; Novotna, M.; Hlavatovicova, Z.

    2004-01-01

    The complete sequence of the mitochondrial DNA of the opportunistic yeast pathogen Candida parapsilosis was determined. The mitochondrial genome is represented by linear DNA molecules terminating with tandem repeats of a 738-bp unit. The number of repeats varies, thus generating a population...

  20. Incomplete effector/memory differentiation of antigen-primed CD8+ T cells in gene gun DNA-vaccinated mice

    DEFF Research Database (Denmark)

    Bartholdy, Christina; Stryhn, Anette; Hansen, Nils Jacob Vest

    2003-01-01

    DNA vaccination is an efficient way to induce CD8+ T cell memory, but it is still unclear to what extent such memory responses afford protection in vivo. To study this, we induced CD8+ memory responses directed towards defined viral epitopes, using DNA vaccines encoding immunodominant MHC class I...... sites. Thus, our DNA vaccine induces a long-lived memory CD8+ T cell population that provides efficient protection against high-dose systemic infection. However, viral replication in solid non-lymphoid organs is not curtailed sufficiently fast to prevent significant virus-induced inflammation. Our...

  1. A combination DNA vaccine encoding nucleoside hydrolase 36 and glycoproteine 63 protects female but not male hamsters against Leishmania mexicana

    Directory of Open Access Journals (Sweden)

    Chalé-balboa W.G.

    2009-09-01

    Full Text Available Leishmaniasis is a group of diseases caused by protozoan parasites of the Leishmania genus. Previous studies have shown that a DNA vaccine encoding Leishmania donovani antigen nucleoside hydrolase 36 and L. mexicana glycoprotein 63 is protective in mice. We investigated here the efficacy of this DNA vaccine to induce protection in golden hamsters. Male hamsters were more susceptible to infection by Leishmania mexicana than females. Following immunization with two doses of the DNA vaccine, only females resulted protected while males developed normal lesions.

  2. Bacterial antigen expression is an important component in inducing an immune response to orally administered Salmonella-delivered DNA vaccines.

    Directory of Open Access Journals (Sweden)

    Michelle E Gahan

    Full Text Available BACKGROUND: The use of Salmonella to deliver heterologous antigens from DNA vaccines is a well-accepted extension of the success of oral Salmonella vaccines in animal models. Attenuated S. typhimurium and S. typhi strains are safe and efficacious, and their use to deliver DNA vaccines combines the advantages of both vaccine approaches, while complementing the limitations of each technology. An important aspect of the basic biology of the Salmonella/DNA vaccine platform is the relative contributions of prokaryotic and eukaryotic expression in production of the vaccine antigen. Gene expression in DNA vaccines is commonly under the control of the eukaryotic cytomegalovirus (CMV promoter. The aim of this study was to identify and disable putative bacterial promoters within the CMV promoter and evaluate the immunogenicity of the resulting DNA vaccine delivered orally by S. typhimurium. METHODOLOGY/PRINCIPAL FINDINGS: The results reported here clearly demonstrate the presence of bacterial promoters within the CMV promoter. These promoters have homology to the bacterial consensus sequence and functional activity. To disable prokaryotic expression from the CMV promoter a series of genetic manipulations were performed to remove the two major bacterial promoters and add a bacteria transcription terminator downstream of the CMV promoter. S. typhimurium was used to immunise BALB/c mice orally with a DNA vaccine encoding the C-fragment of tetanus toxin (TT under control of the original or the modified CMV promoter. Although both promoters functioned equally well in eukaryotic cells, as indicated by equivalent immune responses following intramuscular delivery, only the original CMV promoter was able to induce an anti-TT specific response following oral delivery by S. typhimurium. CONCLUSIONS: These findings suggest that prokaryotic expression of the antigen and co-delivery of this protein by Salmonella are at least partially responsible for the successful

  3. A Novel DNA-Based Vaccine Methodology for Aids

    Science.gov (United States)

    1998-11-01

    delivery of DNA to the tongue suggests the tongue may be an inductive site for mucosal immunity . 18 To evaluate the immune effects of IL-6, we delivered DNA...indicate the enhanced protection seen with IL-6 and tongue delivery is not due to induction of localized mucosal immunity . These results further suggest...that the tongue is not an inductive site for mucosal immunity . However, these results do not account for potential differences between the mouse and

  4. Enhancement of DNA vaccine-induced immune responses by a 72-bp element from SV40 enhancer

    Institute of Scientific and Technical Information of China (English)

    LI Hai-shan; XU Jian-qing; HONG Kun-xue; SHAO Yi-ming; LIU Yong; LI Ding-feng; ZHANG Ran-ran; TANG Hai-li; ZHANG Yu-wei; HUANG Wei; LIU Ying; PENG Hong

    2007-01-01

    Background Although DNA vaccine is considered as the next generation of vaccine, most DNA vaccine candidates are still suffering from the relatively weak immunogenicity despite the increased dosage of plasmid DNA administered. In order to enhance the immune responses elicited by a codon-optimized HIV gag DNA vaccine, a modified plasmid vector pDRVI1.0 and a booster immunization with replicating Tiantan vaccinia (RTV) strain expressing the same gene were employed.Methods Vector pDRVI1.0 was constructed through inserting the 72-bp element from the SV40 enhancer, which was reported promoting nuclear transport of plasmid DNA, to the upstream of cytomegalovirus enhancer/promoter region of the plasmid vector pVR1012. Gene expression levels from expression plasmids based on pDRVI1.0 and pVR1012 were tested. Humoral and cellular immune responses induced by DNA vaccine alone or DNA prime-RTV boost regimen were determined in mice.Results It was shown that the 72-bp element significantly enhanced the gene expression level in non-dividing cells.gag-specific humoral and cellular immune responses induced by DNA vaccination were both significantly improved, while the Th1/Th2 balance was not obviously affected by the 72-bp element. RTV boosting further significantly enhanced DNA vaccine-primed antibody and T cell responses in a Th1-biased manner.Conclusions The 72-bp SV40 enhancer element should be included in the DNA vaccine vector and RTV strain is a very efficient live vector for boosting immunization.

  5. Construction of an oral recombinant DNA vaccine from H pylori neutrophil activating protein and its immunogenicity

    Institute of Scientific and Technical Information of China (English)

    Bo Sun; Zhao-Shen Li; Zhen-Xing Tu; Guo-Ming Xu; Yi-Qi Du

    2006-01-01

    AIM: To construct a live attenuated Salmonella typhimurium (S.typhimurium) strain harboring the H pylori neutrophil activating protein (HP-NAP) gene as an oral recombinant DNA vaccine, and to evaluate its immunogenicity.METHODS: By genetic engineering methods, the genomic DNA of H pylori was extracted as a template. The total length of the HP-NAP gene was amplified by polymerase chain reaction (PCR) and cloned into pBT vector for sequencing and BLAST analysis, then subcloned into a eukaryotic expression vector pIRES followed by PCR identification and restriction enzyme digestion. The identified recombinant plasmid pIRES-NAP was transfected into COS-7 cells for target fusion protein expression, and its antigenicity was detected by Western blotting. Then the recombinant plasmid was transformed into a live attenuated S. typhimurium strain SL7207 as an oral vaccine strain, and its immunogenicity was evaluated with animal experiments.RESULTS: A 435 bp product was cloned using high homology with HP-NAP gene in GenBank (more than 98%). With identification by PCR and restriction enzyme digestion, a recompinant eukaryotic expression plasmid pIRES-NAP containing the HP-NAP gene of H pylori was successfully constructed. The expressed target protein had a specific reaction with H pylor(i) whole cell antibody and showed a single strip result detected by Western blotting. Oral immunization of mice with recombinant DNA vaccine strain SL7207 (pIRES-NAP) also induced a specific immune response.CONCLUSION: The successful construction of HP-NAP oral DNA vaccine with good immunogenicity may help to further investigate its immunoprotection effects and develop vaccine against H pylori infection.

  6. DNA vaccination of poultry%禽核酸疫苗的研究进展

    Institute of Scientific and Technical Information of China (English)

    唐静静; 周巧丽; 殷光文; 江和基; 黄志坚

    2016-01-01

    本文对禽用(鸡、鸭、火鸡)核酸疫苗的研究进行综述。首先描述禽用核酸疫苗的进展:病原,质粒以及免疫途径。其次,描述提高核酸疫苗免疫效果的方式:接种途径,疫苗剂量以及首免时间,增加宿主细胞对质粒的摄入,添加免疫增强分子,优化质粒骨架和密码子,疫苗抗原的选择,异源性的首免-加强免疫策略。最后,描述禽用核酸疫苗的其他特点:接种后质粒的去向,免疫反应的特点以及核酸疫苗的其他用途。%This review describes studies in this field performed exclusively on birds (chickens, ducks and turkeys). No evaluations of avian DNA vaccine efficacy performed on mice as preliminary tests have been taken into consideration. The review first describes the state of the art for DNA vaccination in poultry: pathogens targeted, plasmids used and different routes of vaccine administration. Sec-ond, it presents strategies designed to improve DNA vaccine efficacy:influence of the route of administration, plasmid dose and age of birds on their first inoculation; increasing plasmid uptake by host cells; addition of immunomodulators; optimization of plasmid back-bones and codon usage;association of vaccine antigens and finally, heterologous prime-boost regimens. The final part will indicate ad-ditional properties of DNA vaccines in poultry: fate of the plasmids upon inoculation, immunological considerations and the use of DNA vaccines for purposes other than preventing infectious diseases.

  7. Use of Staby(®) technology for development and production of DNA vaccines free of antibiotic resistance gene.

    Science.gov (United States)

    Reschner, Anca; Scohy, Sophie; Vandermeulen, Gaëlle; Daukandt, Marc; Jacques, Céline; Michel, Benjamin; Nauwynck, Hans; Xhonneux, Florence; Préat, Véronique; Vanderplasschen, Alain; Szpirer, Cédric

    2013-10-01

    The appearance of new viruses and the cost of developing certain vaccines require that new vaccination strategies now have to be developed. DNA vaccination seems to be a particularly promising method. For this application, plasmid DNA is injected into the subject (man or animal). This plasmid DNA encodes an antigen that will be expressed by the cells of the subject. In addition to the antigen, the plasmid also encodes a resistance to an antibiotic, which is used during the construction and production steps of the plasmid. However, regulatory agencies (FDA, USDA and EMA) recommend to avoid the use of antibiotics resistance genes. Delphi Genetics developed the Staby(®) technology to replace the antibiotic-resistance gene by a selection system that relies on two bacterial genes. These genes are small in size (approximately 200 to 300 bases each) and consequently encode two small proteins. They are naturally present in the genomes of bacteria and on plasmids. The technology is already used successfully for production of recombinant proteins to achieve higher yields and without the need of antibiotics. In the field of DNA vaccines, we have now the first data validating the innocuousness of this Staby(®) technology for eukaryotic cells and the feasibility of an industrial production of an antibiotic-free DNA vaccine. Moreover, as a proof of concept, mice have been successfully vaccinated with our antibiotic-free DNA vaccine against a deadly disease, pseudorabies (induced by Suid herpesvirus-1).

  8. Evaluation of the persistence and gene expression of an anti-Chlamydophila psittaci DNA vaccine in turkey muscle

    Directory of Open Access Journals (Sweden)

    Vanrompay Daisy

    2006-06-01

    Full Text Available Abstract Background DNA vaccination has been shown to elicit specific cellular and humoral immune responses to many different agents in a broad variety of species. However, looking at a commercial use, the duration of the immune response against the vaccine is critical. Therefore the persistence of the DNA vaccine, as well as its expression, should be investigated. We conducted these investigations on a DNA vaccine against Chlamydophila psittaci, a Gram-negative intracellular bacterium which causes respiratory disease in turkeys and humans. Previous studies showed that the DNA vaccine confers partial protection against C. psittaci infection in turkeys. Turkeys were injected intramuscularly with the DNA vaccine : a eukaryotic expression vector (pcDNA1::MOMP expressing the major outer membrane protein (MOMP of an avian C. psittaci serovar D strain. Over a period of 11 weeks, cellular uptake of the DNA vaccine was examined by PCR, transcription of the insert by reverse transcript-PCR (RT-PCR and mRNA translation by immunofluorescence staining of muscle biopsies. Results The results indicate that the DNA vaccine persists in turkey muscle for at least 10 weeks. Moreover, during this period of time MOMP was continuously expressed, as evidenced by the immunofluorescence staining and RT-PCR. Conclusion Since C. psittaci infections occur at the age of 3 to 6 and 8 to 12 weeks, a vaccine persistence of 10 weeks seems adequate. Therefore, further research should concentrate on improving the elicited immune response, more specifically the cell-mediated immune response, rather than prolonging the lifespan of the plasmid.

  9. Characterization of a linear epitope on Chlamydia trachomatis serovar L2 DnaK-like protein

    DEFF Research Database (Denmark)

    Ozkokmen, D; Birkelund, Svend; Christiansen, Gunna

    1994-01-01

    A cytoplasmic 75-kDa immunogen from Chlamydia trachomatis serovar L2 has previously been characterized as being similar to the Escherichia coli heat shock protein DnaK. We have localized a linear epitope for one monoclonal antibody specific for C. trachomatis DnaK. By use of a recombinant DNA...... technique, the epitope was limited to 14 amino acids. With synthetic peptides, the epitope was further limited to eight amino acids. Six of these amino acids are conserved in bovine HSP70, which has a known three-dimensional structure. The amino acid sequence homologous to the epitope is located in a linear...

  10. THE CONSTRUCTION AND PRELIMINARY APPRAISEMENT OF HSV-2gD GENE DNA VACCINE

    Institute of Scientific and Technical Information of China (English)

    王军阳; 范桂香; 胜利; 袁育康

    2002-01-01

    Objective To prevent infection from herpes simplex virus type 2(HSV-2),and make the foundation for the construction of multi-valent DNA vaccine. Methods The complete DNA sequence,which encoded the amino acid sequence of the viral glycoprotein D(gD),was obtained from the HSV-2 genome by polymerase chain reaction(PCR).The fragment was inserted into the lower stream of Cytomegalovirus(CMV) promoter in the eukaryotic expression plasmid pcDNA3.1(+), then immunized mice by bilateral intramuscular injection into the rear legs with this recombinant plasmid and tested the specific antibodies against glycoprotein D by ELISA. Results Animal experiment have demonstrated that the recombinant plasmid(pcDNA-gD2)inoculated into mice could induce the production of specific antibodies against glycoprotein D. Conclusion The eukaryotic plasmid pcDNA-gD2 constructed by us could correctly express gD gene and induce the production of specific antibodies.

  11. Magnetic Nanovectors for the Development of DNA Blood-Stage Malaria Vaccines

    Directory of Open Access Journals (Sweden)

    Fatin M. Nawwab Al-Deen

    2017-02-01

    Full Text Available DNA vaccines offer cost, flexibility, and stability advantages, but administered alone have limited immunogenicity. Previously, we identified optimal configurations of magnetic vectors comprising superparamagnetic iron oxide nanoparticles (SPIONs, polyethylenimine (PEI, and hyaluronic acid (HA to deliver malaria DNA encoding Plasmodium yoelii (Py merozoite surface protein MSP119 (SPIONs/PEI/DNA + HA gene complex to dendritic cells and transfect them with high efficiency in vitro. Herein, we evaluate their immunogenicity in vivo by administering these potential vaccine complexes into BALB/c mice. The complexes induced antibodies against PyMSP119, with higher responses induced intraperitoneally than intramuscularly, and antibody levels further enhanced by applying an external magnetic field. The predominant IgG subclasses induced were IgG2a followed by IgG1 and IgG2b. The complexes further elicited high levels of interferon gamma (IFN-γ, and moderate levels of interleukin (IL-4 and IL-17 antigen-specific splenocytes, indicating induction of T helper 1 (Th1, Th2, and Th17 cell mediated immunity. The ability of such DNA/nanoparticle complexes to induce cytophilic antibodies together with broad spectrum cellular immunity may benefit malaria vaccines.

  12. Magnetic Nanovectors for the Development of DNA Blood-Stage Malaria Vaccines

    Science.gov (United States)

    Al-Deen, Fatin M. Nawwab; Xiang, Sue D.; Ma, Charles; Wilson, Kirsty; Coppel, Ross L.; Selomulya, Cordelia; Plebanski, Magdalena

    2017-01-01

    DNA vaccines offer cost, flexibility, and stability advantages, but administered alone have limited immunogenicity. Previously, we identified optimal configurations of magnetic vectors comprising superparamagnetic iron oxide nanoparticles (SPIONs), polyethylenimine (PEI), and hyaluronic acid (HA) to deliver malaria DNA encoding Plasmodium yoelii (Py) merozoite surface protein MSP119 (SPIONs/PEI/DNA + HA gene complex) to dendritic cells and transfect them with high efficiency in vitro. Herein, we evaluate their immunogenicity in vivo by administering these potential vaccine complexes into BALB/c mice. The complexes induced antibodies against PyMSP119, with higher responses induced intraperitoneally than intramuscularly, and antibody levels further enhanced by applying an external magnetic field. The predominant IgG subclasses induced were IgG2a followed by IgG1 and IgG2b. The complexes further elicited high levels of interferon gamma (IFN-γ), and moderate levels of interleukin (IL)-4 and IL-17 antigen-specific splenocytes, indicating induction of T helper 1 (Th1), Th2, and Th17 cell mediated immunity. The ability of such DNA/nanoparticle complexes to induce cytophilic antibodies together with broad spectrum cellular immunity may benefit malaria vaccines.

  13. Do uncertainty analyses reveal uncertainties? Using the introduction of DNA vaccines to aquaculture as a case.

    Science.gov (United States)

    Gillund, Frøydis; Kjølberg, Kamilla A; von Krauss, Martin Krayer; Myhr, Anne I

    2008-12-15

    The Walker and Harremoës (W&H) uncertainty framework is a tool to systematically identify scientific uncertainty. We applied the W&H uncertainty framework to elicit scientists' judgements of potential sources of uncertainty associated with the use of DNA vaccination in aquaculture. DNA vaccination is considered a promising solution to combat pathological fish diseases. There is, however, lack of knowledge regarding its ecological and social implications. Our findings indicate that scientists are open and aware of a number of uncertainties associated with DNA vaccination e.g. with regard to immune response, degradation and distribution of the DNA plasmid after injection and environmental release, and consider most of these uncertainties to be adequately reduced through more research. We proceed to discuss our experience of using the W&H uncertainty framework. Some challenges related to the application of the framework were recognised. This was especially related to the respondents' unfamiliarity with the concepts used and their lack of experience in discussing qualitative aspects of uncertainties. As we see it, the W&H framework should be considered as a useful tool to stimulate reflection on uncertainty and an important first step in a more extensive process of including and properly dealing with uncertainties in science and policymaking.

  14. Vaccination with trypomastigote surface antigen 1-encoding plasmid DNA confers protection against lethal Trypanosoma cruzi infection.

    Science.gov (United States)

    Wizel, B; Garg, N; Tarleton, R L

    1998-11-01

    DNA vaccination was evaluated with the experimental murine model of Trypanosoma cruzi infection as a means to induce antiparasite protective immunity, and the trypomastigote surface antigen 1 (TSA-1), a target of anti-T. cruzi antibody and major histocompatibility complex (MHC) class I-restricted CD8(+) cytotoxic T-lymphocyte (CTL) responses, was used as the model antigen. Following the intramuscular immunization of H-2(b) and H-2(d) mice with a plasmid DNA encoding an N-terminally truncated TSA-1 lacking or containing the C-terminal nonapeptide tandem repeats, the antibody level, CTL response, and protection against challenge with T. cruzi were assessed. In H-2(b) mice, antiparasite antibodies were induced only by immunization with the DNA construct encoding TSA-1 containing the C-terminal repeats. However, both DNA constructs were efficient in eliciting long-lasting CTL responses against the protective H-2Kb-restricted TSA-1515-522 epitope. In H-2(d) mice, inoculation with either of the two TSA-1-expressing vectors effectively generated antiparasite antibodies and primed CTLs that lysed T. cruzi-infected cells in an antigen-specific, MHC class I-restricted, and CD8(+)-T-cell-dependent manner. When TSA-1 DNA-vaccinated animals were challenged with T. cruzi, 14 of 22 (64%) H-2(b) and 16 of 18 (89%) H-2(d) mice survived the infection. The ability to induce significant murine anti-T. cruzi protective immunity by immunization with plasmid DNA expressing TSA-1 provides the basis for the application of this technology in the design of optimal DNA multicomponent anti-T. cruzi vaccines which may ultimately be used for the prevention or treatment of Chagas' disease.

  15. Synthesis of biodegradable polymer-mesoporous silica composite microspheres for DNA prime-protein boost vaccination.

    Science.gov (United States)

    Ho, Jenny; Huang, Yi; Danquah, Michael K; Wang, Huanting; Forde, Gareth M

    2010-03-18

    DNA vaccines or proteins are capable of inducing specific immunity; however, the translation to the clinic has generally been problematic, primarily due to the reduced magnitude of immune response and poor pharmacokinetics. Herein we demonstrate a composite microsphere formulation, composed of mesoporous silica spheres (MPS) and poly(D,L-lactide-co-glycolide) (PLGA), enables the controlled delivery of a prime-boost vaccine via the encapsulation of plasmid DNA (pDNA) and protein in different compartments. Method with modified dual-concentric-feeding needles attached to a 40 kHz ultrasonic atomizer was studied. These needles focus the flow of two different solutions, which passed through the ultrasonic atomizer. The process synthesis parameters, which are important to the scale-up of composite microspheres, were also studied. These parameters include polymer concentration, feed flowrate, and volumetric ratio of polymer and pDNA-PEI/MPS-BSA. This fabrication technique produced composite microspheres with mean D[4,3] ranging from 6 to 34 microm, depending upon the microsphere preparation. The resultant physical morphology of composite microspheres was largely influenced by the volumetric ratio of pDNA-PEI/MPS-BSA to polymer, and this was due to the precipitation of MPS at the surface of the microspheres. The encapsulation efficiencies were predominantly in the range of 93-98% for pDNA and 46-68% for MPS. In the in vitro studies, the pDNA and protein showed different release kinetics in a 40 day time frame. The dual-concentric-feeding in ultrasonic atomization was shown to have excellent reproducibility. It was concluded that this fabrication technique is an effective method to prepare formulations containing a heterologous prime-boost vaccine in a single delivery system.

  16. Neutralizing antibodies respond to a bivalent dengue DNA vaccine or/and a recombinant bivalent antigen.

    Science.gov (United States)

    Zhang, Zhi-Shan; Weng, Yu-Wei; Huang, Hai-Long; Zhang, Jian-Ming; Yan, Yan-Sheng

    2015-02-01

    There is currently no effective vaccine to prevent dengue infection, despite the existence of multiple studies on potential methods of immunization. The aim of the present study was to explore the effect of DNA and/or recombinant protein on levels of neutralizing antibodies. For this purpose, envelope domain IIIs of dengue serotypes 1 and 2 (DEN-1/2)were spliced by a linker (Gly‑Gly‑Ser‑Gly‑Ser)3 and cloned into the prokaryotic expression plasmid pET30a (+) and eukaryotic vector pcDNA3.1 (+). The chimeric bivalent protein was expressed in Escherichia coli, and one‑step purification by high‑performance liquid chromatography was conducted. Protein expression levels of the DNA plasmid were tested in BHK‑21 cells by indirect immunofluorescent assay. In order to explore a more effective immunization strategy and to develop neutralizing antibodies against the two serotypes, mice were inoculated with recombinant bivalent protein, the DNA vaccine, or the two given simultaneously. Presence of the specific antibodies was tested by ELISA and the presence of the neutralizing antibodies was determined by plaque reduction neutralization test. Results of the analysis indicated that the use of a combination of DNA and protein induced significantly higher titers of neutralizing antibodies against either DEN‑1 or DEN‑2 (1:64.0 and 1:76.1, respectively) compared with the DNA (1:24.7 and 1:26.9, DEN‑1 and DEN‑2, respectively) or the recombinant protein (1:34.9 and 1:45.3 in DEN‑1 and DEN‑2, respectively). The present study demonstrated that the combination of recombinant protein and DNA as an immunization strategy may be an effective method for the development of a vaccine to prevent dengue virus infection.

  17. Anti-Idiotypic Regulatory Responses Induced by Vaccination with DNA Encoding Murine TCR Vo5 and Vβ2

    Institute of Scientific and Technical Information of China (English)

    Ying Wu; Yan Zhang; Xiaojun Xu; Ping Lv; Xiaoming Gao

    2007-01-01

    There is evidence suggesting that anti-idiotypic regulation against T cells plays a role in maintaining homeostasis in the immune system, although its mechanism is not fully understood. By using DNA constructs encoding the TCR Vα5.2 and Vβ2.1 chains derived from an ovalbumin (OVA)-specific T cell clone (OVA-T), we herein demonstrated that vaccination with TCR-DNA effectively induced anti-idiotypic cellular as well as humoral responses. Serum samples from the TCR-DNA-vaccinated BALB/c mice were able to stain T cells in an idiotype-specific manner.CD4+ T cells from the TCR-DNA-vaccinated mice proliferated in response to stimulation with irradiated syngeneic OVA-T cells and secreted interferon-γ but very little IL-4. Splenocytes from the TCR-DNA-vaccinated mice showed strong idiotype-specific CTL activity against the OVA-T cells. Furthermore, adoptive transfer of the CD4+ or CD8+ T cells from the TCR-DNA-vaccinated mice resulted in hyporesponsiveness of syngeneic recipients. These results demonstrated that vaccination with DNA encoding TCR can effectively activate anti-idiotypic regulatory responses in vivo and thus providing a useful way for immunological intervention.

  18. DNA Virus Vectors for Vaccine Production in Plants: Spotlight on Geminiviruses

    Directory of Open Access Journals (Sweden)

    Kathleen L. Hefferon

    2014-08-01

    Full Text Available Plants represent a safe, efficacious and inexpensive production platform by which to provide vaccines and other therapeutic proteins to the world’s poor. Plant virus expression vector technology has rapidly become one of the most popular methods to express pharmaceutical proteins in plants. This review discusses several of the state-of-the-art plant expression systems based upon geminiviruses that have been engineered for vaccine production. An overview of the advantages of these small, single-stranded DNA viruses is provided and comparisons are made with other virus expression systems. Advances in the design of several different geminivirus vectors are presented in this review, and examples of vaccines and other biologics generated from each are described.

  19. Temperature influences the expression profiling of immune response genes in rainbow trout following DNA vaccination and VHS virus infection

    DEFF Research Database (Denmark)

    Einer-Jensen, Katja; Gautier, Laurent; Rasmussen, Jesper Skou

    A DNA vaccine encoding the glycoprotein (G) genes of the salmonid rhabdovirus viral haemorrhagic septicaemia virus (VHSV) has proven highly efficient against the disease caused by this virus in rainbow trout (Oncorhynchus mykiss). Several studies have demonstrated that this vaccine induces both...

  20. Protection of rainbow trout against infectious hematopoietic necrosis virus four days after specific or semi-specific DNA vaccination

    DEFF Research Database (Denmark)

    LaPatra, S.E.; Corbeil, S.; Jones, G.R.;

    2001-01-01

    A DNA vaccine against a fish rhabdovirus, infectious hematopoietic necrosis virus (IHNV), was shown to provide significant protection as soon as 4 d after intramuscular vaccination in 2 g rainbow trout (Oncorhynchus mykiss) held at 15 degreesC. Nearly complete protection was also observed at late...

  1. Efficacy of an infectious hematopoietic necrosis (IHN) virus DNA vaccine in Chinook Oncorhynchus tshawytscha and sockeye O. nerka salmon.

    Science.gov (United States)

    Garver, Kyle A; LaPatra, Scott E; Kurath, Gael

    2005-04-06

    The level of protective immunity was determined for Chinook Oncorhynchus tshawytscha and sockeye/kokanee salmon (anadromous and landlocked) O. nerka following intramuscular vaccination with a DNA vaccine against the aquatic rhabdovirus, infectious hematopoietic necrosis virus (IHNV). A DNA vaccine containing the glycoprotein gene of IHNV protected Chinook and sockeye/kokanee salmon against waterborne or injection challenge with IHNV, and relative percent survival (RPS) values of 23 to 86% were obtained under a variety of lethal challenge conditions. Although this is significant protection, it is less than RPS values obtained in previous studies with rainbow trout (O. mykiss). In addition to the variability in the severity of the challenge and inherent host susceptibility differences, it appears that use of a cross-genogroup challenge virus strain may lead to reduced efficacy of the DNA vaccine. Neutralizing antibody titers were detected in both Chinook and sockeye that had been vaccinated with 1.0 and 0.1 pg doses of the DNA vaccine, and vaccinated fish responded to viral challenges with higher antibody titers than mock-vaccinated control fish.

  2. Induction of partial protection against infection with Toxoplasma gondii genotype II by DNA vaccination with recombinant chimeric tachyzoite antigens

    DEFF Research Database (Denmark)

    Rosenberg, Carina Agerbo; De Craeye, S.; Jongert, E.

    2009-01-01

    complications. Although several strategies have been suggested for making a vaccine, none is currently available. Here, we investigate the protection conferred by DNA vaccination with two constructs, pcEC2 (MIC2-MIC3-SAG1) and pcEC3 (GRA3-GRA7-M2AP), encoding chimeric proteins containing multiple antigenic...

  3. Plasmid DNA Vaccine Co-Immunisation Modulates Cellular and Humoral Immune Responses Induced by Intranasal Inoculation in Mice.

    Directory of Open Access Journals (Sweden)

    Deborah F L King

    Full Text Available An effective HIV vaccine will likely require induction of both mucosal and systemic cellular and humoral immune responses. We investigated whether intramuscular (IM delivery of electroporated plasmid DNA vaccine and simultaneous protein vaccinations by intranasal (IN and IM routes could be combined to induce mucosal and systemic cellular and humoral immune responses to a model HIV-1 CN54 gp140 antigen in mice.Co-immunisation of DNA with intranasal protein successfully elicited both serum and vaginal IgG and IgA responses, whereas DNA and IM protein co-delivery did not induce systemic or mucosal IgA responses. Cellular IFNγ responses were preserved in co-immunisation protocols compared to protein-only vaccination groups. The addition of DNA to IN protein vaccination reduced the strong Th2 bias observed with IN protein vaccination alone. Luminex analysis also revealed that co-immunisation with DNA and IN protein induced expression of cytokines that promote B-cell function, generation of TFH cells and CCR5 ligands that can reduce HIV infectivity.These data suggest that while IN inoculation alone elicits both cellular and humoral responses, co-administration with homologous DNA vaccination can tailor these towards a more balanced Th1/Th2 phenotype modulating the cellular cytokine profile while eliciting high-levels of antigen-specific antibody. This work provides insights on how to generate differential immune responses within the same vaccination visit, and supports co-immunisation with DNA and protein by a mucosal route as a potential delivery strategy for HIV vaccines.

  4. Approaches towards DNA vaccination against a skin ciliate parasite in fish.

    Directory of Open Access Journals (Sweden)

    Louise von Gersdorff Jørgensen

    Full Text Available Rainbow trout (Oncorhynchus mykiss were immunized with plasmid DNA vaccine constructs encoding selected antigens from the parasite Ichthyophthirius multifiliis. Two immobilization antigens (I-ags and one cysteine protease were tested as genetic vaccine antigen candidates. Antigenicity was evaluated by immunostaining of transfected fish cells using I-ag specific mono- and polyclonal antibodies. I. multifiliis specific antibody production, regulation of immune-relevant genes and/or protection in terms of parasite burden or mortality was measured to evaluate the induced immune response in vaccinated fish. Apart from intramuscular injection, needle free injection and gene gun delivery were tested as alternative administration techniques. For the I-ags the complement protein fragment C3d and the termini of the viral haemorrhagic septicaemia virus glyco(Gprotein (VHSV G were tested as opsonisation and cellular localisation mediators, respectively, while the full length viral G protein was tested as molecular adjuvant. Expression of I-ags in transfected fish cells was demonstrated for several constructs and by immunohistochemistry it was possible to detect expression of a secreted form of the Iag52B in the muscle cells of injected fish. Up-regulations of mRNA coding for IgM, MHC I, MHC II and TCR β, respectively, were observed in muscle tissue at the injection site in selected trials. In the spleen up-regulations were found for IFN-γ and IL-10. The highest up-regulations were seen following co-administration of I-ag and cysteine protease plasmid constructs. This correlated with a slight elevation of an I. multifiliis specific antibody response. However, in spite of detectable antigen expression and immune reactions, none of the tested vaccination strategies provided significant protection. This might suggest an insufficiency of DNA vaccination alone to trigger protective mechanisms against I. multifiliis or that other or additional parasite antigens

  5. Pharmaceutical development of the plasmid DNA vaccine pDERMATT

    NARCIS (Netherlands)

    Quaak, S.G.L.

    2009-01-01

    The discovery of tumor specific antigens and self tolerance mechanisms against these antigens led to the assumption that antigens circulating at sufficient concentration levels could break this self tolerance mechanism and evoke immunological antitumor effects. pDERMATT (plasmid DNA encoding recombi

  6. Pharmaceutical development of the plasmid DNA vaccine pDERMATT

    NARCIS (Netherlands)

    Quaak, S.G.L.

    2009-01-01

    The discovery of tumor specific antigens and self tolerance mechanisms against these antigens led to the assumption that antigens circulating at sufficient concentration levels could break this self tolerance mechanism and evoke immunological antitumor effects. pDERMATT (plasmid DNA encoding

  7. The larval specific lymphatic filarial ALT-2: induction of protection using protein or DNA vaccination.

    Science.gov (United States)

    Ramachandran, Sabarinathan; Kumar, Mishra Pankaj; Rami, Reddy Maryada Venkata; Chinnaiah, Harinath Basker; Nutman, Thomas; Kaliraj, Perumal; McCarthy, James

    2004-01-01

    Genes from the infective stage of lymphatic filarial parasites expressed at the time of host invasion have been identified as potential vaccine candidates. By screening an L3 cDNA library with sera from uninfected longstanding residents of an area endemic for onchocerciasis, so-called "endemic normals" (EN), we have cloned and characterized one such gene termed the abundant larval transcript two (ALT-2). The stage specificity of ALT-2 gene transcription and protein synthesis was confirmed by PCR using genespecific primers, and by western blot analysis of protein extracts from various stages of the parasite life cycle using specific antisera. Significant differences in antibody response to the recombinant ALT-2 were observed in endemic populations with differing clinical manifestations of lymphatic filariasis with an antibody response present in sera from 18 of 25 (72%) EN subjects compared to 9 of 25 (36%) with subclinical microfilaracmia (MF) and 14 of 25 (52%) of those with chronic lymphatic obstruction (CP) (P=0.01 for comparison of EN to CP or to MF). This differential responsiveness suggests that the protective immunity postulated to account for their uninfected status might be associated with a response to this protein. When the utility of ALT-2 as a vaccine candidate was tested in a murine model using either recombinant protein or a DNA vaccine construct, statistically significant protection was observed when compared to a control filarial gene product expressed across all stages of the parasite lifecycle (SXP-1; P=0.02 for protein and P=0.01 for the DNA vaccine) or compared to adjuvant alone. This level of protection indicates that this vaccine is a promising candidate for further development.

  8. BTX AgilePulse(TM) system is an effective electroporation device for intramuscular and intradermal delivery of DNA vaccine.

    Science.gov (United States)

    Davtyan, Hayk; Hovakimyan, Armine; Zagorski, Karen; Davtyan, Arpine; Petrushina, Irina; Agdashian, David; Murthy, Vidya; Cribbs, David H; Agadjanyan, Michael G; Ghochikyan, Anahit

    2014-01-01

    DNA vaccines promote immune system activation in small animals and exhibit certain advantages when compared to conventional recombinant protein vaccines. However in clinical trials DNA vaccines are less effective in inducing potent immune responses due to the low delivery efficiency and expression of antigens. Currently, various delivery devices such as gene-guns, bioinjectors and electroporation systems are being used in order to increase the potency of DNA vaccines. However, the optimal delivery parameters are required and must be carefully set to obtain the highest levels of gene expression and strong immune responses in humans. The focus of this study was to optimize electroporation settings (voltage, pulse length, pulse intervals, and number of pulses), as well as the route of administration (intradermal vs. intramuscular) and dosage of the DNA epitope vaccine, AV-1959D, delivered by the BTX AgilePulse(TM) system. As a result, we have chosen the optimal settings for electroporation delivery using different routes of immunization with this vaccine, generating (i) robust antibody production to the B cell epitope (a small peptide, derived from β-amyloid), and (ii) strong cellular immune responses to Th epitopes (a small synthetic peptide and eleven peptides from various pathogens) incorporated into DNA vaccine platform.

  9. Separation of topological forms of plasmid DNA by anion-exchange HPLC: shifts in elution order of linear DNA.

    Science.gov (United States)

    Smith, Clara R; DePrince, Randolph B; Dackor, Jennifer; Weigl, Debra; Griffith, Jack; Persmark, Magnus

    2007-07-01

    We sought to establish a single anion-exchange HPLC method for the separation of linear, open circular and supercoiled plasmid topoisomers using purified topoisomeric forms of three plasmids (3.0, 5.5 and 7.6 kb). However, finding one condition proved elusive as the topoisomer elution order was determined to depend on salt gradient slope. The observed change in selectivity increased with plasmid size and was most pronounced for the linear form. Indeed, the elution order of the linear 7.6 kb plasmid was reversed relative to the supercoiled form. This observation may have implications for methods used in quality control of plasmid DNA.

  10. MPT-51/CpG DNA vaccine protects mice against Mycobacterium tuberculosis.

    Science.gov (United States)

    Silva, Bruna Daniella de Souza; da Silva, Ediane Batista; do Nascimento, Ivan Pereira; Dos Reis, Michelle Cristina Guerreiro; Kipnis, André; Junqueira-Kipnis, Ana Paula

    2009-07-16

    Tuberculosis (TB) is a severe infectious disease that kills approximately two million people worldwide every year. Because BCG protection is variable and does not protects adults, there is a great need for a new vaccine against TB that does not represent a risk for immunocompromised patients and that is also capable of protecting adult individuals. MPT-51 is a protein found in the genome of mycobacteria and binds to the fibronectin of the extracellular matrix, which may have a role in host tissue attachment and virulence. In order to test the usefulness of MPT-51 as a subunit vaccine, BALB/c were vaccinated and challenged with Mycobacterium tuberculosis. The infection of BALB/c with M. tuberculosis increased the number of IFN-gamma(+) T lymphocytes specific to MPT-51 in the spleen and lungs. Inoculation with rMPT-51/FIA and with rMPT-51/CpG DNA in non-infected BALB/c increased the amounts of IFN-gamma(+) T lymphocytes. Inoculation with rMPT-51/FIA also induced a humoral response specific to MPT-51. CFU counts of lung tissues done 60 days after infection showed a reduction of about 2 log in the bacteria load in the group of animals inoculated with rMPT-51/CpG DNA. These results make MPT-51 a valuable component to be further evaluated in the development of other subunit vaccines.

  11. A novel DNA vaccine technology conveying protection against a lethal herpes simplex viral challenge in mice.

    Directory of Open Access Journals (Sweden)

    Julie L Dutton

    Full Text Available While there are a number of licensed veterinary DNA vaccines, to date, none have been licensed for use in humans. Here, we demonstrate that a novel technology designed to enhance the immunogenicity of DNA vaccines protects against lethal herpes simplex virus 2 (HSV-2 challenge in a murine model. Polynucleotides were modified by use of a codon optimization algorithm designed to enhance immune responses, and the addition of an ubiquitin-encoding sequence to target the antigen to the proteasome for processing and to enhance cytotoxic T cell responses. We show that a mixture of these codon-optimized ubiquitinated and non-ubiquitinated constructs encoding the same viral envelope protein, glycoprotein D, induced both B and T cell responses, and could protect against lethal viral challenge and reduce ganglionic latency. The optimized vaccines, subcloned into a vector suitable for use in humans, also provided a high level of protection against the establishment of ganglionic latency, an important correlate of HSV reactivation and candidate endpoint for vaccines to proceed to clinical trials.

  12. DNA Vaccine of SARS-Cov S Gene Induces Antibody Response in Mice

    Institute of Scientific and Technical Information of China (English)

    PingZHAO; Jin-ShanKE; Zhao-LinQIN; HaoREN; Lan-JuanZHAO; Jian-GuoYU

    2004-01-01

    The spike (S) protein, a main surface antigen of SARS-coronavirus (SARS-CoV), is one of the most important antigen candidates for vaccine design. In the present study, three fragments of the truncated S protein were expressed in E.coli, and analyzed with pooled sera of convalescence phase of SARS patients.The full length S gene DNA vaccine was constructed and used to immunize BALB/c mice. The mouse serum IgG antibody against SARS-CoV was measured by ELISA with E.coli expressed truncated S protein or SARS-CoV lysate as diagnostic antigen. The results showed that all the three fragments of S protein expressed by E.coli was able to react with sera of SARS patients and the S gene DNA candidate vaccine could induce the production of specific IgG antibody against SARS-CoV efficiently in mice with seroconversion ratio of 75% after 3 times of immunization. These findings lay some foundations for further understanding the immunology of SARS-CoV and developing SARS vaccines.

  13. DNA Vaccine of SARS-Cov S Gene Induces Antibody Response in Mice

    Institute of Scientific and Technical Information of China (English)

    Ping ZHAO; Jin-Shan KE; Zhao-Lin QIN; Hao REN; Lan-Juan ZHAO; Jian-Guo YU; Jun GAO; Shi-Ying ZHU; Zhong-Tian QI

    2004-01-01

    The spike (S) protein, a main surface antigen of SARS-coronavirus (SARS-CoV), is one of the most important antigen candidates for vaccine design. In the present study, three fragments of the truncated S protein were expressed in E. Coli, and analyzed with pooled sera of convalescence phase of SARS patients.The full length S gene DNA vaccine was constructed and used to immunize BALB/c mice. The mouse serum IgG antibody against SARS-CoV was measured by ELISA with E. Coli expressed truncated S protein or SARS-CoV lysate as diagnostic antigen. The results showed that all the three fragments of S protein expressed by E. Coli was able to react with sera of SARS patients and the S gene DNA candidate vaccine could induce the production of specific IgG antibody against SARS-CoV efficiently in mice with seroconversion ratio of 75% after 3 times of immunization. These findings lay some foundations for further understanding the immunology of SARS-CoV and developing SARS vaccines.

  14. The protective efficacy of chimeric SO7/IL-2 DNA vaccine against coccidiosis in chickens.

    Science.gov (United States)

    Song, Hongyan; Qiu, Baofeng; Yan, Ruofeng; Xu, Lixin; Song, Xiaokai; Li, Xiangrui

    2013-06-01

    The protective efficacy of recombinant vaccines encoding an Eimeria refractile body antigen SO7 was assessed in broiler chickens following oral infection with Eimeria tenella. The SO7 and chicken IL-2 genes were cloned into the expression vector pVAX1 consecutively to construct DNA vaccines pVAX-SO7 and pVAX-SO7-IL-2. Expression of SO7 and IL-2 gene transcripts and proteins encoded by the plasmid DNAs in vivo was detected by reverse transcription-polymerase chain reaction and Western blot. Chickens were inoculated with 100 μg of plasmids pVAX-SO7 or pVAX-SO7-IL-2, or 200 μg of recombinant SO7 protein or chicken IL-2 protein by leg intramuscular injection. At 28days of age, all chickens except the unchallenged control group were challenged orally with 5×10(4) sporulated oocysts of E. tenella. All chickens were euthanized to determine the effects of immunization on the 7th day post-challenge. The results showed that both DNA vaccines containing the SO7 gene and the recombinant SO7 protein could obviously alleviate body weight loss and cecal lesions compared with unvaccinated and challenged control. These findings also suggested that chicken IL-2 could effectively enhance the immunity of SO7 against E. tenella challenge compared with vaccination using pVAX-SO7 alone. Copyright © 2012 Elsevier Ltd. All rights reserved.

  15. Protection of tree shrews by pVAX-PS DNA vaccine against HBV infection.

    Science.gov (United States)

    Zhou, Feng-Juan; Hu, Zhen-Lin; Dai, Jian-Xin; Chen, Rui-Wen; Shi, Ke; Lin, Yi; Sun, Shu-Han

    2003-07-01

    The immunological protection of pVAX-PS, a DNA vaccine, was assessed in the tree shrews model. pVAX-PS was constructed by inserting the gene encoding the middle (pre-S2 plus S) envelope protein of HBV into a plasmid vector pVAX1. Tree shrews (Tupaia belangeri chinenesis) were experimentally infected with human HBV by inoculation with human serum positive for HBV markers. DNA vaccination-induced seroconversion and antibody to HBV surface antigen (anti-HBs) were analyzed by ELISA, and protective effects elicited by pVAX-PS vaccination against subsequent HBV challenge were evaluated by detection of HBV seromarkers and observation of hepatic lesions in HBV-infected tree shrews. The results shown that anti-HBs were detectable in serum at week 2 after pVAX-PS vaccination and peaked at week 4, and immunization with pVAX-PS decreased the positive conversion rate of HBV seromarkers and relieved hepatic lesions in tree shrews challenged with HBV. These results indicated that pVAX-PS immunization could induce remarkable humoral immune response and prevent the experimental tree shrews from infection of HBV.

  16. A candidate DNA vaccine elicits HCV specific humoral and cellular immune responses

    Institute of Scientific and Technical Information of China (English)

    Li-Xin Zhu; Jing Liu; Ye Ye; You-Hua Xie; Yu-Ying Kong; Guang-Di Li; Yuan Wang

    2004-01-01

    AIM: To investigate the immunogenicity of candidate DNA vaccine against hepatitis C virus (HCV) delivered by two plasmids expressing HCV envelope protein 1 (E1) and envelope protein 2 (E2) antigens respectively and to study the effect of CpG adjuvant on this candidate vaccine.METHODS: Recombinant plasmids expressing HCV E1 and E2 antigens respectively were used to simultaneously inoculate mice with or without CpG adjuvant. Antisera were then collected and titers of anti-HCV antibodies were analyzed by ELISA. One month after the last injection, animals were sacrificed to prepare single-cell suspension of splenocytes.These cells were subjected to HCVantigen specific proliferation assays and cytokine secretion assays to evaluate the cellular immune responses of the vaccinated animals.RESULTS: Antibody responses to HCV E1 and E2 antigens were detected in vaccinated animals. Animals receiving CpG adjuvant had slightly lower titers of anti-HCV antibodies in the sera, while the splenocytes from these animals showed higher HCV-antigen specific proliferation. Analysis of cytokine secretion from the splenocytes was consistent with the above results. While no antigen-specific IL-4 secretion was detected for all vaccinated animals, HCV antigen-specific INF-γ secretion was detected for the splenocytes of vaccinated animals. CpG adjuvant enhanced the secretion of INF-γ but did not change the profile of IL-4 secretion.CONCLUSION: Vaccination of mice with plasmids encoding HCV E1 and E2 antigens induces humoral and cellular immune responses. CpG adjuvant significantly enhances the cellular immune response.

  17. Expanded breadth of the T-cell response to mosaic HIV-1 envelope DNA vaccination

    Energy Technology Data Exchange (ETDEWEB)

    Korber, Bette [Los Alamos National Laboratory; Fischer, William [Los Alamos National Laboratory; Wallstrom, Timothy [Los Alamos National Laboratory

    2009-01-01

    An effective AIDS vaccine must control highly diverse circulating strains of HIV-1. Among HIV -I gene products, the envelope (Env) protein contains variable as well as conserved regions. In this report, an informatic approach to the design of T-cell vaccines directed to HIV -I Env M group global sequences was tested. Synthetic Env antigens were designed to express mosaics that maximize the inclusion of common potential Tcell epitope (PTE) 9-mers and minimize the inclusion of rare epitopes likely to elicit strain-specific responses. DNA vaccines were evaluated using intracellular cytokine staining (ICS) in inbred mice with a standardized panel of highly conserved 15-mer PTE peptides. I, 2 and 3 mosaic sets were developed that increased theoretical epitope coverage. The breadth and magnitude ofT-cell immunity stimulated by these vaccines were compared to natural strain Env's; additional comparisons were performed on mutant Env's, including gpl60 or gpl45 with or without V regions and gp41 deletions. Among them, the 2 or 3 mosaic Env sets elicited the optimal CD4 and CD8 responses. These responses were most evident in CD8 T cells; the 3 mosaic set elicited responses to an average of 8 peptide pools compared to 2 pools for a set of3 natural Env's. Synthetic mosaic HIV -I antigens can therefore induce T-cell responses with expanded breadth and may facilitate the development of effective T -cell-based HIV -1 vaccines.

  18. Granulocyte-macrophage colony-stimulating factor DNA prime-protein boost strategy to enhance efficacy of a recombinant pertussis DNA vaccine

    Institute of Scientific and Technical Information of China (English)

    Qing-tian LI; Yong-zhang ZHU; Jia-you CHU; Ke DONG; Ping HE; Chun-yan FENG; Bao-yu HU; Shu-min ZHANG; Xiao-kui GUO

    2006-01-01

    Aim: To investigate a new strategy to enhance the efficacy of a recombinant pertussis DNA vaccine. The strategy is co-injection with cytokine plasmids as prime, and boosted with purified homologous proteins. Method: A recombinant pertussis DNA vaccine containing the pertussis toxin subunit 1 (PTS1), fragments of the filamentous hemagglutinin (FHA) gene and pertactin (PRN) gene encoding filamentous hemagglutinin and pertactin were constructed. Balb/c mice were immunized with several DNA vaccines and antigen-specific antibodies anti-PTSl, anti-PRN, anti-FHA, cytokines interleukin (IL)-10, IL-4, IFN-γ, TNF-oc, and spleno-cyte-proliferation assay were used to describe immune responses. Results: The recombinant DNA vaccine could elicit similar immune responses in mice as that of separate plasmids encoding the 3 fragments, respectively. Mice immunized with DNA and boosted with the corresponding protein elicited more antibodies than those that received DNA as boost. In particular, when the mice were co-immunized with murine granulocyte-macrophage colony-stimulating factor plasmids and boosted with proteins, all 4 cytokines and the 3 antigen-specific antibodies were significantly increased compared to the pVAXl group. Anti-PTSl, anti-FHA, IL-4 and TNF-α elicited in the colony stimulating factor (CSF) prime-protein boost group showed significant increase compared to all the other groups. Conclusion: This prime and boost strategy has proven to be very useful in improving the immunogenicity of DNA vaccines against pertussis.

  19. DNA vaccines used for biodefense%生物防御用DNA疫苗

    Institute of Scientific and Technical Information of China (English)

    刘文宇; 朱晓斐; 戚中田

    2011-01-01

    Most of the recent biodefense work has been focused on developing DNA vaccines,which are safe to both human and environment and cost-effective,and delivery of the combined DNA vaccines can offer the potential for multiple protection.Consequently,the DNA vaccines have good prospects for research,development and applications,but their immunogenicity and delivery methods remain to be improved.This review summarizes the research status of the DNA vaccines against several biodefense pathogens listed on both the NIH priority pathogen and CDC bioterrorism registries:Bacillus anthracis,Ebola and Marburg viruses,monkeypox virus,smallpox virus,and Venezuelan equine encephalitis virus.%近年的生物防御疫苗研究集中于DNA疫苗.DNA疫苗对人以及环境相对安全、经济,并且多价联合DNA疫苗还可以提供多重保护,因此有良好的研发前景和潜在的实用价值,但也存在免疫原性不强和接种方式有待改进等问题.此文就炭疽杆菌、埃博拉病毒、马尔堡病毒、猴痘病毒、天花病毒和委内瑞拉马脑炎病毒等几种被美国NIH和CDC列为重要生物战剂的DNA疫苗研发现状作一综述.

  20. Contamination of DNase Preparations Confounds Analysis of the Role of DNA in Alum-Adjuvanted Vaccines

    OpenAIRE

    Noges, Laura E.; White, Janice; Cambier, John C.; Kappler, John W.; Marrack, Philippa

    2016-01-01

    Aluminum salt (alum) adjuvants have been used for many years as adjuvants for human vaccines because they are safe and effective. Despite its widespread use, the means by which alum acts as an adjuvant remains poorly understood. Recently, it was shown that injected alum is rapidly coated with host chromatin within mice. Experiments suggested that the host DNA in the coating chromatin contributed to alum’s adjuvant activity. Some of the experiments used commercially purchased DNase and showed ...

  1. Immunization of olive flounder (Paralichthys olivaceus) with an auxotrophic Edwardsiella tarda mutant harboring the VHSV DNA vaccine.

    Science.gov (United States)

    Choi, Seung Hyuk; Kim, Min Sun; Kim, Ki Hong

    2012-09-01

    The aims of the present study were to find more powerful promoter for DNA vaccines in olive flounder (Paralichthys olivaceus) and to evaluate the availability of the auxotrophic Edwardsiella tarda mutant (Δalr Δasd E. tarda) as a delivery vehicle for DNA vaccine against VHSV in olive flounder. The marine medaka (Oryzias dancena) β-actin promoter was clearly stronger than cytomegalovirus (CMV) promoter when the vectors were transfected to Epithelioma papulosum cyprini (EPC) cells or injected into the muscle of olive flounder, suggesting that marine medaka β-actin promoter would be more appropriate promoter for DNA vaccines in olive flounder than CMV promoter. Olive flounder immunized with the Δalr Δasd E. tarda harboring viral hemorrhagic septicemia virus (VHSV) DNA vaccine vector driven by the marine medaka β-actin promoter showed significantly higher serum neutralization titer and higher survival rates against challenge with VHSV than fish immunized with the bacteria carrying VHSV DNA vaccine vector driven by CMV promoter. These results indicate that auxotrophic E. tarda mutant harboring marine medaka β-actin promoter-driven DNA vaccine vectors would be a potential system for prophylactics of infectious diseases in olive flounder.

  2. Poor immune responses of newborn rhesus macaques to measles virus DNA vaccines expressing the hemagglutinin and fusion glycoproteins.

    Science.gov (United States)

    Polack, Fernando P; Lydy, Shari L; Lee, Sok-Hyong; Rota, Paul A; Bellini, William J; Adams, Robert J; Robinson, Harriet L; Griffin, Diane E

    2013-02-01

    A vaccine that would protect young infants against measles could facilitate elimination efforts and decrease morbidity and mortality in developing countries. However, immaturity of the immune system is an important obstacle to the development of such a vaccine. In this study, DNA vaccines expressing the measles virus (MeV) hemagglutinin (H) protein or H and fusion (F) proteins, previously shown to protect juvenile macaques, were used to immunize groups of 4 newborn rhesus macaques. Monkeys were inoculated intradermally with 200 μg of each DNA at birth and at 10 months of age. As controls, 2 newborn macaques were similarly vaccinated with DNA encoding the influenza virus H5, and 4 received one dose of the current live attenuated MeV vaccine (LAV) intramuscularly. All monkeys were monitored for development of MeV-specific neutralizing and binding IgG antibody and cytotoxic T lymphocyte (CTL) responses. These responses were poor compared to the responses induced by LAV. At 18 months of age, all monkeys were challenged intratracheally with a wild-type strain of MeV. Monkeys that received the DNA vaccine encoding H and F, but not H alone, were primed for an MeV-specific CD8(+) CTL response but not for production of antibody. LAV-vaccinated monkeys were protected from rash and viremia, while DNA-vaccinated monkeys developed rashes, similar to control monkeys, but had 10-fold lower levels of viremia. We conclude that vaccination of infant macaques with DNA encoding MeV H and F provided only partial protection from MeV infection.

  3. Vaccination with Trypomastigote Surface Antigen 1-Encoding Plasmid DNA Confers Protection against Lethal Trypanosoma cruzi Infection

    OpenAIRE

    1998-01-01

    DNA vaccination was evaluated with the experimental murine model of Trypanosoma cruzi infection as a means to induce antiparasite protective immunity, and the trypomastigote surface antigen 1 (TSA-1), a target of anti-T. cruzi antibody and major histocompatibility complex (MHC) class I-restricted CD8+ cytotoxic T-lymphocyte (CTL) responses, was used as the model antigen. Following the intramuscular immunization of H-2b and H-2d mice with a plasmid DNA encoding an N-terminally truncated TSA-1 ...

  4. Linearly programmed DNA-based molecular computer operated on magnetic particle surface in test-tube

    Institute of Scientific and Technical Information of China (English)

    ZHAO Jian; ZHANG Zhizhou; SHI Yongyong; Li Xiuxia; HE Lin

    2004-01-01

    The postgenomic era has seen an emergence of new applications of DNA manipulation technologies, including DNA-based molecular computing. Surface DNA computing has already been reported in a number of studies that, however, all employ different mechanisms other than automaton functions. Here we describe a programmable DNA surface-computing device as a Turing machine-like finite automaton. The laboratory automaton is primarily composed of DNA (inputs, output-detectors, transition molecules as software), DNA manipulating enzymes and buffer system that solve artificial computational problems autonomously. When fluoresceins were labeled in the 5′ end of (-) strand of the input molecule, direct observation of all reaction intermediates along the time scale was made so that the dynamic process of DNA computing could be conveniently visualized. The features of this study are: (i) achievement of finite automaton functions by linearly programmed DNA computer operated on magnetic particle surface and (ii) direct detection of all DNA computing intermediates by capillary electrophoresis. Since DNA computing has the massive parallelism and feasibility for automation, this achievement sets a basis for large-scale implications of DNA computing for functional genomics in the near future.

  5. Adjuvant effect of polysaccharide from fruits of Physalis alkekengi L. in DNA vaccine against systemic candidiasis.

    Science.gov (United States)

    Yang, Huimin; Han, Shuying; Zhao, Danyang; Wang, Guiyun

    2014-08-30

    Adjuvant effect mediated by polysaccharide (PPSB) isolated from the fruits of Physalis alkekengi L. in DNA vaccine was evaluated in mice. Recombinant plasmid containing epitope C (LKVIRK) from heat shock protein 90 (HSP90) of Candida albicans (C. albican) was used as DNA vaccine (pD-HSP90C). The results indicated that PPSB significantly enhanced specific antibody titers IgG, IgG1, IgG2b, and concentration of IL-2 and IL-4 in sera of mice immunized with pD-HSP90C (p<0.05). More importantly, it was found that the mice immunized with pD-HSP90C/PPSB not only had fewer CFU (colony forming unites) in the kidneys than mice immunized with pD-HSP90C, but also a statistically significant higher survival rate over PBS-injected group (p<0.05) when the immunized mice were challenged with living C. albican cells. However, no statistically significant difference in survival rate was observed between pD-HSP90C-immunized group and PBS-injected group. Therefore, PPSB can be considered as a promising adjuvant eliciting both Th1 and Th2 responses to enhance the efficacy of DNA vaccines.

  6. On the efficacy of malaria DNA vaccination with magnetic gene vectors.

    Science.gov (United States)

    Nawwab Al-Deen, Fatin; Ma, Charles; Xiang, Sue D; Selomulya, Cordelia; Plebanski, Magdalena; Coppel, Ross L

    2013-05-28

    We investigated the efficacy and types of immune responses from plasmid malaria DNA vaccine encoding VR1020-PyMSP119 condensed on the surface of polyethyleneimine (PEI)-coated SPIONs. In vivo mouse studies were done firstly to determine the optimum magnetic vector composition, and then to observe immune responses elicited when magnetic vectors were introduced via different administration routes. Higher serum antibody titers against PyMSP119 were observed with intraperitoneal and intramuscular injections than subcutaneous and intradermal injections. Robust IgG2a and IgG1 responses were observed for intraperitoneal administration, which could be due to the physiology of peritoneum as a major reservoir of macrophages and dendritic cells. Heterologous DNA prime followed by single protein boost vaccination regime also enhanced IgG2a, IgG1, and IgG2b responses, indicating the induction of appropriate memory immunity that can be elicited by protein on recall. These outcomes support the possibility to design superparamagnetic nanoparticle-based DNA vaccines to optimally evoke desired antibody responses, useful for a variety of diseases including malaria.

  7. Preparation and efficacy of Newcastle disease virus DNA vaccine encapsulated in chitosan nanoparticles.

    Science.gov (United States)

    Zhao, Kai; Zhang, Yang; Zhang, Xiaoyan; Li, Wei; Shi, Ci; Guo, Chen; Dai, Chunxiao; Chen, Qian; Jin, Zheng; Zhao, Yan; Cui, Hongyu; Wang, Yunfeng

    2014-01-01

    Optimal preparation conditions of Newcastle disease virus (NDV) F gene deoxyribonucleic acid (DNA) vaccine encapsulated in chitosan nanoparticles (pFNDV-CS-NPs) were determined. The pFNDV-CS-NPs were prepared according to a complex coacervation method. The pFNDV-CS-NPs were produced with good morphology, high stability, a mean diameter of 199.5 nm, encapsulation efficiency of 98.37% ± 0.87%, loading capacity of 36.12% ± 0.19%, and a zeta potential of +12.11 mV. The in vitro release assay showed that the plasmid DNA was sustainably released from the pFNDV-CS-NPs, up to 82.9% ± 2.9% of the total amount. Cell transfection test indicated that the vaccine expressed the F gene in cells and maintained good bioactivity. Additionally, the safety of mucosal immunity delivery system of the pFNDV-CS-NPs was also tested in vitro by cell cytotoxicity and in vivo by safety test in chickens. In vivo immunization showed that better immune responses of specific pathogen-free chickens immunized with the pFNDV-CS-NPs were induced, and prolonged release of the plasmid DNA was achieved compared to the chickens immunized with the control plasmid. This study lays the foundation for the further development of mucosal vaccines and drugs encapsulated in chitosan nanoparticles.

  8. A recoding method to improve the humoral immune response to an HIV DNA vaccine.

    Directory of Open Access Journals (Sweden)

    Yaoxing Huang

    Full Text Available This manuscript describes a novel strategy to improve HIV DNA vaccine design. Employing a new information theory based bioinformatic algorithm, we identify a set of nucleotide motifs which are common in the coding region of HIV, but are under-represented in genes that are highly expressed in the human genome. We hypothesize that these motifs contribute to the poor protein expression of gag, pol, and env genes from the c-DNAs of HIV clinical isolates. Using this approach and beginning with a codon optimized consensus gag gene, we recode the nucleotide sequence so as to remove these motifs without modifying the amino acid sequence. Transfecting the recoded DNA sequence into a human kidney cell line results in doubling the gag protein expression level compared to the codon optimized version. We then turn both sequences into DNA vaccines and compare induced antibody response in a murine model. Our sequence, which has the motifs removed, induces a five-fold increase in gag antibody response compared to the codon optimized vaccine.

  9. Attenuated Salmonella typhimurium SV4089 as a potential carrier of oral DNA vaccine in chickens.

    Science.gov (United States)

    Jazayeri, Seyed Davoud; Ideris, Aini; Zakaria, Zunita; Omar, Abdul Rahman

    2012-01-01

    Attenuated Salmonella has been used as a carrier for DNA vaccine. However, in vitro and in vivo studies on the bacteria following transfection of plasmid DNA were poorly studied. In this paper, eukaryotic expression plasmids encoding avian influenza virus (AIV) subtype H5N1 genes, pcDNA3.1/HA, NA, and NP, were transfected into an attenuated Salmonella enteric typhimurium SV4089. In vitro stability of the transfected plasmids into Salmonella were over 90% after 100 generations. The attenuated Salmonella were able to invade MCF-7 (1.2%) and MCF-10A (0.5%) human breast cancer cells. Newly hatched specific-pathogen-free (SPF) chicks were inoculated once by oral gavage with 10(9) colony-forming unit (CFU) of the attenuated Salmonella. No abnormal clinical signs or deaths were recorded after inoculation. Viable bacteria were detected 3 days after inoculation by plating from spleen, liver, and cecum. Fluorescent in situ hybridization (FISH) and polymerase chain reaction (PCR) were carried out for confirmation. Salmonella was not detected in blood cultures although serum antibody immune responses to Salmonella O antiserum group D1 factor 1, 9, and 12 antigens were observed in all the inoculated chickens after 7 days up to 35 days. Our results showed that live attenuated S. typhimurium SV4089 harboring pcDNA3.1/HA, NA, and NP may provide a unique alternative as a carrier for DNA oral vaccine in chickens.

  10. Chitosan Microparticles Intended for Anti-caries DNA Vaccine Mucosal Delivery: Synthesis, Characterization and Transfection

    Institute of Scientific and Technical Information of China (English)

    LI Yuhong; FAN Mingwen; BIAN Zhuan; CHEN Zhi; Zhang Qi

    2005-01-01

    In order to enhance the mucosal immunity of anti-caries DNA vaccine, chitosan-DNA microparticles for musocal vaccination were prepared by a coacervation method. The physicochemical structure of microparticles was investigated by a scanning electron microscope (SEM) and a cofocal laser scanning microscope (CLSM). For in-vitro studies, Hela cell was transfected by chitosan-DNA microparticles.The expression of proteins was measured by the immunohistochemical methods, and the cytotocity of chitosan in Hela cell line was determined by the MTT assay. The experimental results show that the microparticles are about 2-6 μm in size and spherical in shape. The encapsulation efficiency is 99%, and the DNA is almost captured in the micropraticles. Plasmid loaded into chitosan microparticles is distributed throughout these particles. The number of positive staining cells of chitosan-pGJA-P transfected cell is more than that of naked plasmid transfect cells, but less than that of Lipofect-DNA complex group. Chitosan was found to be less cytotoxic compared with lipofectin (p<0.01).

  11. Induction of protective immunity against Eimeria tenella, Eimeria necatrix, Eimeria maxima and Eimeria acervulina infections using multivalent epitope DNA vaccines.

    Science.gov (United States)

    Song, Xiaokai; Ren, Zhe; Yan, Ruofeng; Xu, Lixin; Li, Xiangrui

    2015-06-04

    Avian coccidiosis is mostly caused by mixed infection of several Eimeria species under natural conditions and immunity to avian coccidiosis is largely dependent on T-cell immune response. In this study, 14 T-cell epitope fragments from eight antigens of Eimeria tenella (E. tenella), Eimeria necatrix (E. necatrix), Eimeria maxima (E. maxima) and Eimeria acervulina (E. acervulina) were ligated with pVAX1 producing 14 monovalent DNA vaccines, respectively. Protective immunity of the monovalent DNA vaccines was assessed by in vivo challenge experiments and then four most protective fragments of each species were chosen to construct multivalent epitope DNA vaccines with or without chicken IL-2 as genetic adjuvant. Protective efficacies of the epitope DNA vaccines on chickens against E. tenella, E. necatrix, E. maxima and E. acervulina were evaluated. The results showed that the constructed multivalent epitope DNA vaccines significantly increased body weight gain, alleviated enteric lesions and reduced oocyst output of the infected birds. Especially, the multivalent epitope DNA vaccines of pVAX1-NA4-1-TA4-1-LDH-2-EMCDPK-1 and pVAX1-NA4-1-TA4-1-LDH-2-EMCDPK-1-IL-2 not only significantly increased body weight gain, alleviated enteric lesions and reduced oocyst output of the infected birds, but also resulted in anti-coccidial index (ACI) more than 170 against E. tenella, E. necatrix, E. maxima and E. acervulina, which indicated they could induce protective immunity against E. tenella, E. necatrix, E. maxima and E. acervulina. Our findings suggest the constructed multivalent epitope DNA vaccines are the potential candidate multivalent vaccines against mixed infection of Eimeria.

  12. Involvement of two microRNAs in the early immune response to DNA vaccination against a fish rhabdovirus

    DEFF Research Database (Denmark)

    Bela-ong, Dennis Berbulla; Schyth, Brian Dall; Zou, Jun

    2015-01-01

    Mechanisms that account for the high protective efficacy in teleost fish of a DNA vaccine expressing the glycoprotein (G) of Viral hemorrhagic septicemia virus (VHSV) are thought to involve early innate immune responses mediated by interferons (IFNs). Microribonucleic acids (miRNAs) are a diverse...... trout infected with VHSV. In this study, we analyzed the expression of these miRNAs in fish following administration of the DNA vaccine and their potential functions. Quantitative RT-PCR analysis revealed the increased levels of miR-462, and miR-731 in the skeletal muscle tissue at the site of vaccine...... administration and in the liver of vaccinated fish relative to empty plasmid backbone-injected controls. The increased expression of these miRNAs in the skeletal muscle correlated with the increased levels of the type I interferon (IFN)-inducible gene Mx, type I IFN and IFN-γ genes at the vaccination site...

  13. Therapeutic efficacy of a tuberculosis DNA vaccine encoding heat shock protein 65 of Mycobacterium tuberculosis and the human interleukin 2 fusion gene.

    Science.gov (United States)

    Changhong, Shi; Hai, Zhang; Limei, Wang; Jiaze, An; Li, Xi; Tingfen, Zhang; Zhikai, Xu; Yong, Zhao

    2009-01-01

    Use of therapeutic DNA vaccines is a promising strategy against tuberculosis (TB), however, their immunogenicity still needs to be improved. In this study, a plasmid DNA vaccine expressing heat shock protein 65 (HSP65) and the human interleukin 2 (IL-2) fusion gene was constructed. Immune responses induced by the vaccine in the mice and protection against Mycobacterium tuberculosis (MTB) were investigated, along with the therapeutic effect of the DNA vaccine on tuberculosis in mice. Administration of the HSP65-IL-2-DNA vaccine enhanced Th1-type cellular responses by producing greater amounts of interferon-gamma (IFN-gamma) and IL-2 with a higher titer of antigen-specific anti-Hsp65 IgG2a. Compared with the Bacille Calmette-Guérin (BCG) vaccine, the DNA vaccine was able to evoke both CD4 and CD8 T-cell responses, with an especially high percentage of CD8 T-cells. The DNA vaccine was also able to induce high antigen-specific cytotoxicity activity against target cells. When the mice were challenged with virulent MTB H37Rv, a dramatic decrease in the numbers of MTB colony forming units in the spleen and lungs was observed in the mice immunized with HSP65-IL-2-DNA (P<0.05). Meanwhile, the bacterial numbers in TB infected mice treated with the DNA vaccine were also significantly reduced. The protective and therapeutic effects of the HSP65-IL-2-DNA vaccine in the spleen and lungs were superior to that of the HSP65-DNA vaccine (P<0.05). These results suggest that the DNA vaccine expression of IL-2 and the HSP65 fusion gene enhances the immunogenicity and protective as well as therapeutic effects of the HSP65-DNA vaccine against TB in mice by improving the Th1-type response.

  14. Protective effect of DNA vaccine encoding pseudomonas exotoxin A and PcrV against acute pulmonary P. aeruginosa Infection.

    Directory of Open Access Journals (Sweden)

    Mingzi Jiang

    Full Text Available Infections with Pseudomonas aeruginosa have been a long-standing challenge for clinical therapy because of complex pathogenesis and resistance to antibiotics, thus attaching importance to explore effective vaccines for prevention and treatment. In the present study, we constructed a novel DNA vaccine by inserting mutated gene toxAm encoding Pseudomonas Exotoxin A and gene pcrV encoding tip protein of the type III secretion system into respective sites of a eukaryotic plasmid pIRES, named pIRES-toxAm-pcrV, and next evaluated the efficacy of the vaccine in murine acute Pseudomonas pneumonia models. Compared to DNA vaccines encoding single antigen, mice vaccinated with pIRES-toxAm-pcrV elicited higher levels of antigen-specific serum immunoglobulin G (IgG, enhanced splenic cell proliferation and cytokine secretion in response to Pseudomonas aeruginosa antigens, additionally PAO1 challenge in mice airway resulted in reduced bacteria burden and milder pathologic changes in lungs. Besides, it was observed that immunogenicity and protection could be promoted by the CpG ODN 1826 adjuvant. Taken together, it's revealed that recombinant DNA vaccine pIRES-toxAm-pcrV was a potential candidate for immunotherapy of Pseudomonas aeruginosa infection and the CpG ODN 1826 a potent stimulatory adjuvant for DNA vaccination.

  15. Recombinant Saccharomyces cerevisiae serves as novel carrier for oral DNA vaccines in Carassius auratus.

    Science.gov (United States)

    Yan, Nana; Xu, Kun; Li, Xinyi; Liu, Yuwan; Bai, Yichun; Zhang, Xiaohan; Han, Baoquan; Chen, Zhilong; Zhang, Zhiying

    2015-12-01

    Oral delivery of DNA vaccines represents a promising vaccinating method for fish. Recombinant yeast has been proved to be a safe carrier for delivering antigen proteins and DNAs to some species in vivo. However, whether recombinant yeast can be used to deliver functional DNAs for vaccination to fish is still unknown. In this study, red crucian carp (Carassius auratus) was orally administrated with recombinant Saccharomyces cerevisiae harboring CMV-EGFP expression cassette. On day 5 post the first vaccination, EGFP expression in the hindgut was detected under fluorescence microscope. To further study whether the delivered gene could induce specific immune responses, the model antigen ovalbumin (OVA) was used as immunogen, and oral administrations were conducted with recombinant S. cerevisiae harboring pCMV-OVA mammalian gene expression cassette as gene delivery or pADH1-OVA yeast gene expression cassette as protein delivery. Each administration was performed with three different doses, and the OVA-specific serum antibody was detected in all the experimental groups by western blotting and enzyme-linked immunosorbent assay (ELISA). ELISA assay also revealed that pCMV-OVA group with lower dose (pCMV-OVA-L) and pADH1-OVA group with moderate dose (pADH1-OVA-M) triggered relatively stronger antibody response than the other two doses. Moreover, the antibody level induced by pCMV-OVA-L group was significantly higher than pADH1-OVA-M group at the same serum dilutions. All the results suggested that recombinant yeast can be used as a potential carrier for oral DNA vaccines and would help to develop more practical strategies to control infectious diseases in aquaculture.

  16. Membrane and envelope virus proteins co-expressed as lysosome associated membrane protein (LAMP) fused antigens: a potential tool to develop DNA vaccines against flaviviruses

    OpenAIRE

    Rafael Dhalia; Milton Maciel Jr.; Cruz,Fábia S.P.; Viana,Isabelle F.T.; Palma,Mariana L.; Thomas August; Ernesto T. A. Marques Jr.

    2009-01-01

    Vaccination is the most practical and cost-effective strategy to prevent the majority of the flavivirus infection to which there is an available vaccine. However, vaccines based on attenuated virus can potentially promote collateral side effects and even rare fatal reactions. Given this scenario, the developent of alternative vaccination strategies such as DNA-based vaccines encoding specific flavivirus sequences are being considered. Endogenous cytoplasmic antigens, characteristically plasmi...

  17. Immunization with chlamydial plasmid protein pORF5 DNA vaccine induces protective immunity against genital chlamydial infection in mice

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    To validate the immune protective efficacy of pORF5 DNA vaccine and to analyze potential mechanisms related to this protection. In this study, pORF5 DNA vaccine was constructed and evaluated for its protective immunity in a mouse model of genital chlamydial infection. Groups of BALB/c mice were immunized intranasally with pORF5 DNA vaccine. Humoral and cell mediated immune responses were evaluated. The clearance ability of chlamydial challenge from the genital tract and the chlamy- dia-induced upper genital tract gross pathology and histopathological characterization were also de- tected. The results showed that the total and the IgG2a anti-pORF5 antibody levels in serum were sig- nificantly elevated after pcDNA3.1-pORF5 vaccination, as were the total antibody and IgA levels in vaginal fluids. pcDNA3.1-pORF5 induced a significantly high level of Th1 response as measured by robust gamma interferon (IFN-γ). Minimal IL-4 was produced by immune T cells in response to the re-stimulation with pORF5 protein or the inactive elementary body in vitro. pcDNA3.1-pORF5-vacci- nated mice displayed significantly reduced bacterial shedding upon a chlamydial challenge and an accelerated resolution of infection. 100% of pcDNA3.1-pORF5 vaccinated mice successfully resolved the infection by day 24. pcDNA3.1-pORF5-immunized mice also exhibited protection against patho- logical consequences of chlamydial infection. The stimulated index was significantly higher than that of mice immunized with pcDNA3.1 and PBS (P<0.05). Together, these results demonstrated that immu- nization with pORF5 DNA vaccine is a promising approach for eliciting a protective immunity against a genital chlamydial challenge.

  18. Elicitation of strong immune responses by a DNA vaccine expressing a secreted form of hepatitis C virus envelope protein E2 in murine and porcine animal models

    DEFF Research Database (Denmark)

    Li, Yiping; Kang, H.N.; Babiuk, L.A.

    2006-01-01

    AIM: To characterize the immunogenicity of a hepatitis C virus (HCV) E2 DNA vaccine alone or with a protein vaccine boost in murine and porcine animal models. METHODS: A DNA vaccine expressing a secreted form of HCV E2 protein was constructed and used to vaccinate mice and piglets with or without...... boosting with a recombinant E2 protein vaccine formulated with CpG ODN and 10% Emulsigen. The immunogenicity of HCV E2 vaccines was analyzed by ELISA for antibody responses, MTT assay for lymphocyte proliferation, ELISPOT for the number of interferon-gamma secreting cells, and cytotoxic T lymphocyte assays...

  19. A recombinant DNA vaccine encoding C. andersoni oocyst wall protein induces immunity against experimental C. parvum infection.

    Science.gov (United States)

    Zheng, Jun; Ren, Wenzhi; Pan, Qingshan; Wang, Qiuyue; Elhag, I A Elfaki; Li, Jianhua; Li, Mingying; Gong, Pengtao; Liu, Yingli; Zhang, Xichen

    2011-06-30

    Cryptosporidium andersoni parasited in the abomasum has been demonstrated as a cause of reduction of milk production in dairy cow. In this study, a novel chimeric DNA vaccine pVAX1-AB was constructed and the efficacy against Cryptosporidium parvum was determined. BALB/c mice were divided into 3 groups and immunized with DNA vaccine expressing the oocyst wall protein, AB protein of C. andersoni, the recombinant plasmid containing the AB gene, respectively. After inoculation of 1 × 10(6) oocysts of C. parvum, the humoral and cellular immune responses were detected. Experimental results showed that the recombinant plasmid can induce corresponding specific antibody response, simultaneously influenced cellular immune responses, and provided greater protection rate (48.6%) than the other groups. These results indicated that chimeric DNA vaccine has a potential in Cryptosporidium vaccine development.

  20. Antitumor Response to a Codon-Optimized HPV-16 E7/HSP70 Fusion Antigen DNA Vaccine.

    Science.gov (United States)

    Soleimanjahi, Hoorieh; Razavinikoo, Hadi; Fotouhi, Fatemeh; Ardebili, Abdollah

    2017-09-01

    Vaccines based on virus-like particles are effective against Human Papilloma Virus (HPV) infection; however, they have not shown a therapeutic effect against HPV-associated diseases. New immunotherapy strategies based on immune responses against tumor antigens can positively affect the clearance of HPV-associated lesions. To generate two therapeutic fusion DNA vaccines (optimizedE7/mouseHSP70 and wildE7/mouseHSP70) to induce antitumor specific responses in mice models. Mice were immunized with recombinant DNA vaccines. The splenocytes of immunized mice were collected and lactate dehydrogenase and IFN-γ productions were measured after three injections in order to evaluate cytotoxic T lymphocytes (CTLs) activity. MTT assay was carried out for lymphocyte stimulation. The fusion DNA vaccines, specifically uE7-HSP70, elicited varying levels of IFN-γ and CTLs responses compared to the control group (P<0.05). Furthermore, antitumor response and tumor size reduction in fusion DNA vaccines groups were significantly higher than in the negative control group (P<0.05). It is concluded that our fusion DNA vaccines considerably enhanced specific cellular responses against HPV tumor model. In addition, optimized E7 showed a notable immunogenicity and inhibitory effect on the reduction of tumor size.

  1. Good Manufacturing Practices production and analysis of a DNA vaccine against dental caries

    Institute of Scientific and Technical Information of China (English)

    Ya-ping YANG; Yu-hong LI; Ai-hua ZHANG; Lan BI; Ming-wen FAN

    2009-01-01

    Aim: To prepare a clinical-grade anti-caries DNA vaccine pGJA-P/VAX and explore its immune effect and protective efficacy against a cariogenic bacterial challenge.Methods: A large-scale industrial production process was developed under Good Manufacturing Practices (GMP) by combining and optimizing common unit operations such as alkaline lysis, precipitation, endotoxin removal and column chromatography. Quality controls of the purified bulk and final lyophilized vaccine were conducted according to authoritative guidelines. Mice and gnotobiotic rats were intranasally immunized with clinical-grade pGJA-P/VAX with chitosan. Antibody levels of serum IgG and salivary SlgA were assessed by an enzyme-linked immunosorbent assay (ELISA), and caries activity was evaluated by the Keyes method. pGJA-P/VAX and pVAX1 prepared by a laboratory-scale commercial kit were used as controls.Results: The production process proved to be scalable and reproducible. Impurities including host protein, residual RNA, genomic DNA and endotoxin in the purified plasmid were all under the limits of set specifications. Intranasal vaccination with clinical-grade pGJA-P/ VAX induced higher serum IgG and salivary SlgA in both mice and gnotobiotic rats. While in the experimental caries model, the enamel (E), dentinal slight (Ds), and dentinal moderate (Dm) caries lesions were reduced by 21.1%, 33.0%, and 40.9%, respectively. Conclusion: The production process under GMP was efficient in preparing clinical-grade pGJA-P/VAX with high purity and intended effectiveness, thus facilitating future clinical trials for the anti-caries DNA vaccine.

  2. Oral Vaccination with Attenuated Salmonella typhimurium-Delivered TsPmy DNA Vaccine Elicits Protective Immunity against Trichinella spiralis in BALB/c Mice

    Science.gov (United States)

    Wang, Lei; Wang, Xiaohuan; Bi, Kuo; Sun, Ximeng; Yang, Jing; Gu, Yuan; Huang, Jingjing; Zhan, Bin; Zhu, Xinping

    2016-01-01

    Background Our previous studies showed that Trichinella spiralis paramyosin (TsPmy) is an immunomodulatory protein that inhibits complement C1q and C8/C9 to evade host complement attack. Vaccination with recombinant TsPmy protein induced protective immunity against T. spiralis larval challenge. Due to the difficulty in producing TsPmy as a soluble recombinant protein, we prepared a DNA vaccine as an alternative approach in order to elicit a robust immunity against Trichinella infection. Methods and Findings The full-length TsPmy coding DNA was cloned into the eukaryotic expression plasmid pVAX1, and the recombinant pVAX1/TsPmy was transformed into attenuated Salmonella typhimurium strain SL7207. Oral vaccination of mice with this attenuated Salmonella-delivered TsPmy DNA vaccine elicited a significant mucosal sIgA response in the intestine and a systemic IgG antibody response with IgG2a as the predominant subclass. Cytokine analysis also showed a significant increase in the Th1 (IFN-γ, IL-2) and Th2 (IL-4, 5, 6, 10) responses in lymphocytes from the spleen and MLNs of immunized mice upon stimulation with TsPmy protein. The expression of the homing receptors CCR9/CCR10 on antibody secreting B cells may be related to the translocation of IgA-secreted B cells to local intestinal mucosa. The mice immunized with Salmonella-delivered TsPmy DNA vaccine produced a significant 44.8% reduction in adult worm and a 46.6% reduction in muscle larvae after challenge with T. spiralis larvae. Conclusion Our results demonstrated that oral vaccination with TsPmy DNA delivered by live attenuated S. typhimurium elicited a significant local IgA response and a mixed Th1/Th2 immune response that elicited a significant protection against T. spiralis infection in mice. PMID:27589591

  3. Enhanced efficacy of CTLA-4 fusion anti-caries DNA vaccines in gnotobiotic hamsters

    Institute of Scientific and Technical Information of China (English)

    Feng ZHANG; Yu-hong LI; Ming-wen FAN; Rong JIA; Qing-an XU; Ji-hua GUO; Fei YU; Qi-wei TIAN

    2007-01-01

    Aim:To evaluate the comparative immunogenicity and protective efficacy of the cytotoxic T-lymphocyte.associated antigen 4(CTLA-4)fusion anti-caries DNA vaccines pGJA-P/VAX1,pGJA-P,and non-fusion anti-caries DNA construct pGLUA-P in hamsters.In addition,the ability of CTLA-4 to target pGJA-P/VAX1-encoding antigen to dendritic cells was tested in vitro.Methods:All DNA constructs contain genes encoding the A-P regions of a cell surface protein(PAc) and the glucan binding(GLU) domain of glucosyltransferases(GTFs)of cari-ogenic organism Streptococcus mutans.Human dendritic cells were mixed with the CTLA-4-Ig-GLU-A-P protein expressed by pGJA-P/VAX1-transfected cells and analyzed by flow cytometry.Gnotobiotic hamsters were immunized with anti-caries DNA vaccines by intramuscular injection or intranasal administration.Antibody responses to a representative antigen PAc were assayed by ELlSA,and caries protection was evaluated by Keyes caries scores.Results:A flow cytometric analysis demonstrated that CTLA-4-Ig-GLU-A-P protein was capable of bind-ing to human dendritic cells.pGJA-P/VAX1 and pGJA-P induced significantly higher specific salivary and serum anti-PAc antibody responses than pGLUA-R.Significantly fewet caries lesions were alSO observed in hamsters immunized with pGJA-P/VAX1 and pGJA-p There was no significant difference in the anti-PAC antibody level or caries scores between pGJA-P/VAX1 and pGJA-P-immunized groups.Conclusion:Antigen encoded by CTLA-4 fusion anti-caries DNA vac-cine pGJA-P/VAX1 could specifically bind to human dendritic cells through the interaction of CTLA-4 and B7 molecules.Fusing antigen to CTLA-4 has been proven to greatly enhance the immunogenicity and protective efficacy of anti-caries DNA vaccines.

  4. Novel cyclen-based linear polymer as a high-affinity binding material for DNA condensation

    Institute of Scientific and Technical Information of China (English)

    XIANG YongZhe; WANG Na; ZHANG Ji; LI Kun; ZHANG ZhongWei; LIN HongHui; YU XiaoQi

    2009-01-01

    A novel cyclen-based linear polyamine (POGEC) was designed and synthesized from the reaction be-tween 1,3-propanediol diglycidyl ether and 1,7-bis(diethoxyphosphory)-1,4,7,10-tetraazacyclod- odecane.High-affinity binding between POGEC and DNA was demonstrated by agarose gel electrophoresis and scanning electron microscopy (SEM). Moreover, the formed POGEC/DNA complex (termed polyplex) could be disassociated to release the free DNA through addition of the physiological concentration of NaCl solution. Fluorescence spectrum was used to measure the high-affinity binding and DNA con-densation capability of POGEC. Circular dichroism (CD) spectrum indicates that the DNA conformation did not change after binding to POEGC.

  5. Novel cyclen-based linear polymer as a high-affinity binding material for DNA condensation

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    A novel cyclen-based linear polyamine (POGEC) was designed and synthesized from the reaction between 1,3-propanediol diglycidyl ether and 1,7-bis(diethoxyphosphory)-1,4,7,10-tetraazacyclod-odecane. High-affinity binding between POGEC and DNA was demonstrated by agarose gel electrophoresis and scanning electron microscopy (SEM). Moreover,the formed POGEC/DNA complex (termed polyplex) could be disassociated to release the free DNA through addition of the physiological concentration of NaCl solution. Fluorescence spectrum was used to measure the high-affinity binding and DNA condensation capability of POGEC. Circular dichroism (CD) spectrum indicates that the DNA conformation did not change after binding to POEGC.

  6. The Protection Efficacity of DNA Vaccine Encoding Hemagglutinin of H5 Subtype Avian Influenza Virus

    Institute of Scientific and Technical Information of China (English)

    JIANG Yong-ping; YU Kang-zhen; DENG Guo-hua; TIAN Guo-bin; QIAO Chuan-ling; CHEN Hua-lan

    2004-01-01

    The DNA vaccine pCIHA5 encoding hemagglutinin can protect SPF chicken against lethal H5N1 avian influenza virus challenge. The more characters about its protection efficacity were studied. The protective rates in 10, 40, 70, 100 and 150μg groups immunized with pCIHA5 were 12.5 (1/8), 58.3 (7/12), 72.7 (8/11), 50.0 (6/12) and 66.7% (8/12), respectively. The protective rates in 5, 20, 35 and 50μg groups were 145.5 (5/11), 58.3 (7/12), 58.3 (7/12) and 91.7% (11/12), respectively. The 70, 100 and 5μg groups have virus shedding of 1/8, 2/6 and 1/5. Though the inactived oil-emulsion vaccine has high HI antibody titers and 100% protective rate, the AGP antibody could be detected after vaccination. Results show that the pCIHA5 is fit to boost by intramuscular injection. This would be useful to the study on gene engineering vaccine of avian influenza virus.

  7. DNA vaccination in fish promotes an early chemokine-related recruitment of B cells to the muscle

    DEFF Research Database (Denmark)

    Castro, R.; Martínez-Alonso, S.; Fischer, U.

    2013-01-01

    might explain the recruitment of immune cells to the site of DNA injection. Our results suggest that B cells are involved in the initial phase of the immune response to intramuscular DNA vaccination against VHSV. This appears to be a major difference to what we know from mammalian models where T cells...

  8. Safety and immunogenicity of a novel therapeutic DNA vaccine encoding chicken type II collagen for rheumatoid arthritis in normal rats.

    Science.gov (United States)

    Juan, Long; Xiao, Zhao; Song, Yun; Zhijian, Zhang; Jing, Jin; Kun, Yu; Yuna, Hao; Dongfa, Dai; Lili, Ding; Liuxin, Tan; Fei, Liang; Nan, Liu; Fang, Yuan; Yuying, Sun; Yongzhi, Xi

    2015-01-01

    Current clinically available treatments for rheumatoid arthritis (RA) fail to cure the disease or unsatisfactorily halt disease progression. To overcome these limitations, the development of therapeutic DNA vaccines and boosters may offer new promising strategies. Because type II collagen (CII) as a critical autoantigen in RA and native chicken type II collagen (nCCII) has been used to effectively treat RA, we previously developed a novel therapeutic DNA vaccine encoding CCII (pcDNA-CCOL2A1) with efficacy comparable to that of the current "gold standard", methotrexate(MTX). Here, we systemically evaluated the safety and immunogenicity of the pcDNA-CCOL2A1 vaccine in normal Wistar rats. Group 1 received only a single intramuscular injection into the hind leg with pcDNA-CCOL2A1 at the maximum dosage of 3 mg/kg on day 0; Group 2 was injected with normal saline (NS) as a negative control. All rats were monitored daily for any systemic adverse events, reactions at the injection site, and changes in body weights. Plasma and tissues from all experimental rats were collected on day 14 for routine examinations of hematology and biochemistry parameters, anti-CII IgG antibody reactivity, and histopathology. Our results indicated clearly that at the maximum dosage of 3 mg/kg, the pcDNA-CCOL2A1 vaccine was safe and well-tolerated. No abnormal clinical signs or deaths occurred in the pcDNA-CCOL2A1 group compared with the NS group. Furthermore, no major alterations were observed in hematology, biochemistry, and histopathology, even at the maximum dose. In particularly, no anti-CII IgG antibodies were detected in vaccinated normal rats at 14 d after vaccination; this was relevant because we previously demonstrated that the pcDNA-CCOL2A1 vaccine, when administered at the therapeutic dosage of 300 μg/kg alone, did not induce anti-CII IgG antibody production and significantly reduced levels of anti-CII IgG antibodies in the plasma of rats with established collagen-induced arthritis

  9. HPV16E7-HSP70 Hybrid DNA Vaccine Induces E7-Specific Cytotoxic T Cells and Antitumor Immunity

    Institute of Scientific and Technical Information of China (English)

    ZHU Liqin; LI Hui; XIONG Jinhu; WANG Tongxiang; OU Xuan; WEI Yun; WU Xinxing

    2006-01-01

    Using human papillomavirus type 16 (HPV16) E7 as an antigen and Heat Shock Protein 70 as adjuvant, we constructed a DNA vaccine by linking HSP70 gene to E7C91G gene. Mice, after being immunized with E7C91G-HSP70, E7C91G/HSP70, E7C91G, and wild E7 DNA vaccines respectively, produced E7 specific CD8+ T-cell precursor frequencies oF280. 33±2.52, 144.34±4. 04, 164.34±5.13 and 82.33± 3.51 respectively within every 1 × 105 mouse splenocytes. This proves that E7C91G-HSP70 fusion vaccine can significantly enhance the E7 specific cellular immunity within the mice body(p<0.01). After being immunized with E7C91G-HSP70 fusion vaccine, tumor-bearing mice of the group being treated have significantly longer latency and survival periods, comparing with other three categories of E7 vaccines. Experiment shows that this vaccine has a significant effect on enhancing E7 positive tumor-treatment within mice body. After being immunized with E7C91G-HSP70 vaccine, there were no pathological changes found in livers, kidneys and spleens of the mice, which proves that the vaccine is quite safe. After all,E7C91G-HSP70 fusion vaccine has a much stronger tumor- treatment effect than that of wild type E7 DNA vaccine.

  10. Linear superclusters of colloidal gold particles by electrostatic assembly on DNA templates

    Energy Technology Data Exchange (ETDEWEB)

    Kumar, A.; Bhadbhade, M.; Mandale, A.B.; Sastry, M. [National Chemical Lab., Pune (India). Materials Chemistry Div.; Pattarkine, M.; Ganesh, K.N. [National Chemical Lab., Pune (IN). Organic Chemistry (Synthesis) Div.; Datar, S.S.; Dharmadhikari, C.V. [Pune Univ. (India). Dept. of Physics

    2001-03-02

    The organization of nanoparticles into superstructures of predefined geometry is an important challenge in the area of nanoscale architecture. Attractive Coulombic interaction between positively charged amine groups on gold particle surfaces and negatively charged phosphate backbones of DNA molecules drives the self-assembly of gold nanoparticles into linear supercluster structures. (orig.)

  11. Control of Trypanosoma cruzi infection and changes in T-cell populations induced by a therapeutic DNA vaccine in mice.

    Science.gov (United States)

    Zapata-Estrella, Hiatzy; Hummel-Newell, Caroline; Sanchez-Burgos, Gilma; Escobedo-Ortegon, Javier; Ramirez-Sierra, Maria Jesus; Arjona-Torres, Arletty; Dumonteil, Eric

    2006-03-15

    Previous work showed that immunotherapy with a DNA vaccine encoding Trypanosoma cruzi antigen TSA-1 reduced cardiac tissue damage and improved survival in mice when administered during the acute or chronic phases of T. cruzi infection. In the present study, we investigated changes in T-cell populations induced by DNA vaccine immunotherapy. ICR mice were infected with 500 T. cruzi blood trypomastigotes and treated during the acute or chronic phases with two 100 microg doses of DNA vaccine. Analysis of stained splenocytes by flow cytometry indicated that the therapeutic vaccine induced a rapid increase in the number of CD4+ and CD8+ T cells in both the acute and chronic phases. Also, there was a rapid increase in T. cruzi-specific IFNgamma-producing CD8+ T cells following treatment during the chronic phase. The effects of these changes on the control of infection required longer time periods to be detectable but resulted in a reduction in myocarditis and T. cruzi parasite burden in both phases of the infection, as assessed by histopathologic analysis and semi-quantitative PCR detection of T. cruzi in cardiac tissue. These results suggest that DNA vaccines that induce CD8+ T-cells activity and IFNgamma production, would be good candidates for effective therapeutic vaccination against T. cruzi infection.

  12. Long-Term Reduction of High Blood Pressure by Angiotensin II DNA Vaccine in Spontaneously Hypertensive Rats.

    Science.gov (United States)

    Koriyama, Hiroshi; Nakagami, Hironori; Nakagami, Futoshi; Osako, Mariana Kiomy; Kyutoku, Mariko; Shimamura, Munehisa; Kurinami, Hitomi; Katsuya, Tomohiro; Rakugi, Hiromi; Morishita, Ryuichi

    2015-07-01

    Recent research on vaccination has extended its scope from infectious diseases to chronic diseases, including Alzheimer disease, dyslipidemia, and hypertension. The aim of this study was to design DNA vaccines for high blood pressure and eventually develop human vaccine therapy to treat hypertension. Plasmid vector encoding hepatitis B core-angiotensin II (Ang II) fusion protein was injected into spontaneously hypertensive rats using needleless injection system. Anti-Ang II antibody was successfully produced in hepatitis B core-Ang II group, and antibody response against Ang II was sustained for at least 6 months. Systolic blood pressure was consistently lower in hepatitis B core-Ang II group after immunization, whereas blood pressure reduction was continued for at least 6 months. Perivascular fibrosis in heart tissue was also significantly decreased in hepatitis B core-Ang II group. Survival rate was significantly improved in hepatitis B core-Ang II group. This study demonstrated that Ang II DNA vaccine to spontaneously hypertensive rats significantly lowered high blood pressure for at least 6 months. In addition, Ang II DNA vaccines induced an adequate humoral immune response while avoiding the activation of self-reactive T cells, assessed by ELISPOT assay. Future development of DNA vaccine to treat hypertension may provide a new therapeutic option to treat hypertension.

  13. Progress of research on DNA vaccines against parasitosis%寄生虫病DNA疫苗研究进展

    Institute of Scientific and Technical Information of China (English)

    齐文娟; 方强

    2011-01-01

    寄生虫病防治策略之一是研制安全有效的疫苗,DNA疫苗是近10多年发展起来的新型疫苗.近年来寄生虫病DNA疫苗研究取得了很大进展.本文就DNA疫苗的免疫机理、构建与优化、佐剂、递送途径,以及疟疾、血吸虫病、囊尾蚴病、弓形虫病等重要寄生虫病DNA疫苗的研究进展作一综述.%One of the effective prevention and treatment strategies to parasitosis is to develop safe and effective vaccines. The DNA vaccine is a new kind of vaccine developed in last 10 years. In recent years, many advances in DNA vaccines against parasitosis have been made. This article reviews the advances in the mechanism, construction , optimization , adjuvants and delivery ways of DNA vaccines and the advances in the study of DNA vaccines against some parasitosis including malaria, schistosomiasis, cysti-cercosis and toxoplasmosis in recent years.

  14. Cholera toxin B subunit acts as a potent systemic adjuvant for HIV-1 DNA vaccination intramuscularly in mice.

    Science.gov (United States)

    Hou, Jue; Liu, Ying; Hsi, Jenny; Wang, Hongzhi; Tao, Ran; Shao, Yiming

    2014-01-01

    Cholera toxin B subunit (CTB) was investigated as a classical mucosal adjuvant that can increase vaccine immunogenicity. In this study, we found out the in vitro efficacy of cholera toxin B subunit (CTB) in activating mice bone marrow-derived dendritic cells (BMDCs) through Toll-like receptor signaling pathways. In vitro RNA and transcriptional level profiling arrays revealed that CTB guides high levels of Th1 and Th2 type cytokines, inflammatory cytokines, and chemokines. Based on the robustness of these profiling results, we examined the induction of HIV Env-specific immunity by CTB co-inoculated with HIV Env DNA vaccine intramuscularly in vivo. CTB enhanced HIV-Env specific cellular immune responses in Env-specific IFN-γ ELISPOT, compared with DNA vaccine alone. Moreover, CTB induced high levels of Env specific humoral response and promoted antibody maturation after the third round of vaccination. This combination immunization strategy induced a Th2-type bias response which is indicative of a high ratio of IgG1/IgG2a. This study reports that CTB as a classical mucosal adjuvant could enhance HIV-1 DNA-based vaccine immunogenicity intramuscularly; therefore, these findings suggest that CTB could serve as an effective candidate adjuvant for DNA vaccination.

  15. Immunologic Evaluation of DNA Vaccine Encoding Influenza Virus M2 Gene in Type A- Influenza Mice Model

    Directory of Open Access Journals (Sweden)

    Shaffifar, M. (MSc

    2015-05-01

    Full Text Available Background and Objective: The M2 gene expressing the conserved protein in influenza virus can be used to make a single-dose vaccine with permanent immunity. Material and Methods: The mice were allocated to one case group immunized with pcDNA3-M2 and two control groups with pcDNA and PBS, in three dozes with interval of two weeks. Two weeks after the last injection, Cellular immunity was analyzed by MTT lymphocyte proliferation, interferon gamma (IFN-gamma and interleukin 4 (IL-4 ratio assays. The remaining animals were challenged with PR8 virus. Results: The production rate of IFN8 and IL4 in pcDNA - M2 group was higher than that of control groups (P >0.0001. Given the results of lymphocyte proliferation, Stimulation index (SI in vaccinated mice was significantly higher than that of control groups (P<0.05. In comparison with mortality rate of 100% in control groups , the animals Challenged with PR8 vaccine had a 50% fatal rate implying a high protection level for this vaccine. Conclusion: The pcDNA3-M2 Vaccine can be considered as a promising vaccine against influenza infections.

  16. Highly immunogenic prime–boost DNA vaccination protects chickens against challenge with homologous and heterologous H5N1 virus

    Directory of Open Access Journals (Sweden)

    Anna Stachyra

    2014-01-01

    Full Text Available Highly pathogenic avian influenza viruses (HPAIVs cause huge economic losses in the poultry industry because of high mortality rate in infected flocks and trade restrictions. Protective antibodies, directed mainly against hemagglutinin (HA, are the primary means of protection against influenza outbreaks. A recombinant DNA vaccine based on the sequence of H5 HA from the H5N1/A/swan/Poland/305-135V08/2006 strain of HPAIV was prepared. Sequence manipulation included deletion of the proteolytic cleavage site to improve protein stability, codon usage optimization to improve translation and stability of RNA in host cells, and cloning into a commercially available vector to enable expression in animal cells. Naked plasmid DNA was complexed with a liposomal carrier and the immunization followed the prime–boost strategy. The immunogenic potential of the DNA vaccine was first proved in broilers in near-to-field conditions resembling a commercial farm. Next, the protective activity of the vaccine was confirmed in SPF layer-type chickens. Experimental infections (challenge experiments indicated that 100% of vaccinated chickens were protected against H5N1 of the same clade and that 70% of them were protected against H5N1 influenza virus of a different clade. Moreover, the DNA vaccine significantly limited (or even eliminated transmission of the virus to contact control chickens. Two intramuscular doses of DNA vaccine encoding H5 HA induced a strong protective response in immunized chicken. The effective protection lasted for a minimum 8 weeks after the second dose of the vaccine and was not limited to the homologous H5N1 virus. In addition, the vaccine reduced shedding of the virus.

  17. IgA response and protection following nasal vaccination of chickens with Newcastle disease virus DNA vaccine nanoencapsulated with Ag@SiO2 hollow nanoparticles

    Science.gov (United States)

    Zhao, Kai; Rong, Guangyu; Hao, Yan; Yu, Lu; Kang, Hong; Wang, Xin; Wang, Xiaohua; Jin, Zheng; Ren, Zhiyu; Li, Zejun

    2016-05-01

    Newcastle disease caused by ND virus (NDV) is a highly contagious disease of birds. Vaccine for effective protection of poultry animals from NDV infection is urgently needed. Mucosal immunity plays a very important role in the antiviral immune response. In this study, a NDV F gene-containing DNA vaccine encapsulated in Ag@SiO2 hollow nanoparticles (pFDNA-Ag@SiO2-NPs) with an average diameter of 500 nm were prepared to assess the mucosal immune response. These nanoparticles exhibited low cytotoxicity and did not destroy the bioactivity of plasmid DNA, which could be expressed in vitro. The plasmid DNA was sustainably released after an initial burst release. In vivo immunization showed that the intranasal immunization of chickens with pFDNA-Ag@SiO2-NPs induced high titers of serum antibody, significantly promoted lymphocyte proliferation and induced higher expression levels of IL-2 and IFN-γ in a dose-dependent manner. These results indicated that the Ag@SiO2 hollow nanoparticles could serve as an efficient and safe delivery carrier for NDV DNA vaccine to induce mucosal immunity. This study has provided promising results for the further development of mucosal vaccines encapsulated in inorganic nanoparticles.

  18. Scientific advice on the suitability of data for the assessment of DNA integration into the fish genome of a genetically modified DNA plasmid-based veterinary vaccine

    Directory of Open Access Journals (Sweden)

    European Food Safety Authority

    2013-05-01

    Full Text Available Pancreas disease caused by salmonid alphavirus in farmed Atlantic salmon (Salmo salar leads to high mortality rates post infection and histopathological lesions in several organs. As protection against pancreas disease, Novartis developed a prophylactic DNA plasmid-based vaccine to be administered to salmon as naked plasmid in a single intramuscular injection. In order to assess the legal status of the fish vaccinated with this new vaccine with regard to the legislation on genetically modified organisms, the European Commission suggested that the company carry out a scientific study on the integration/non-integration of the plasmid DNA into the fish genome. Subsequently, the European Commission requested EFSA to give scientific advice on the study design and the conclusions drawn by the company. PCR based analysis of genomic DNA from muscle samples, taken from at or around the injection site 436 days post vaccination, led the company to conclude that integration of plasmid DNA into the fish genome is extremely unlikely. After an assessment of the study, EFSA considers that the study presented by Novartis Animal Health on the integration/non-integration of DNA plasmid-based vaccine into the salmon genomic DNA provides insufficient information on the potential integration of plasmid DNA fragments into the fish genome due to a limited coverage of the plasmid DNA by the detection method provided, the limited number of samples analysed and an insufficient limit of detection and method validation. Therefore, EFSA is of the opinion that the results from the integration/non-integration study submitted by Novartis Animal Health are not sufficient to support the conclusion of non-integration of plasmid DNA into the fish genome drawn by the company.

  19. Soluble multi-trimeric TNF superfamily ligand adjuvants enhance immune responses to a HIV-1 Gag DNA vaccine.

    Science.gov (United States)

    Kanagavelu, Saravana K; Snarsky, Victoria; Termini, James M; Gupta, Sachin; Barzee, Suzanne; Wright, Jacqueline A; Khan, Wasif N; Kornbluth, Richard S; Stone, Geoffrey W

    2012-01-17

    DNA vaccines remain an important component of HIV vaccination strategies, typically as part of a prime/boost vaccination strategy with viral vector or protein boost. A number of DNA prime/viral vector boost vaccines are currently being evaluated for both preclinical studies and in Phase I and Phase II clinical trials. These vaccines would benefit from molecular adjuvants that increase correlates of immunity during the DNA prime. While HIV vaccine immune correlates are still not well defined, there are a number of immune assays that have been shown to correlate with protection from viral challenge including CD8+ T cell avidity, antigen-specific proliferation, and polyfunctional cytokine secretion. Recombinant DNA vaccine adjuvants composed of a fusion between Surfactant Protein D (SP-D) and either CD40 Ligand (CD40L) or GITR Ligand (GITRL) were previously shown to enhance HIV-1 Gag DNA vaccines. Here we show that similar fusion constructs composed of the TNF superfamily ligands (TNFSFL) 4-1BBL, OX40L, RANKL, LIGHT, CD70, and BAFF can also enhanced immune responses to a HIV-1 Gag DNA vaccine. BALB/c mice were vaccinated intramuscularly with plasmids expressing secreted Gag and SP-D-TNFSFL fusions. Initially, mice were analyzed 2 weeks or 7 weeks following vaccination to evaluate the relative efficacy of each SP-D-TNFSFL construct. All SP-D-TNFSFL constructs enhanced at least one Gag-specific immune response compared to the parent vaccine. Importantly, the constructs SP-D-4-1BBL, SP-D-OX40L, and SP-D-LIGHT enhanced CD8+ T cell avidity and CD8+/CD4+ T cell proliferation 7 weeks post vaccination. These avidity and proliferation data suggest that 4-1BBL, OX40L, and LIGHT fusion constructs may be particularly effective as vaccine adjuvants. Constructs SP-D-OX40L, SP-D-LIGHT, and SP-D-BAFF enhanced Gag-specific IL-2 secretion in memory T cells, suggesting these adjuvants can increase the number of self-renewing Gag-specific CD8+ and/or CD4+ T cells. Finally adjuvants SP

  20. New vaccine strategies against enterotoxigenic Escherichia coli: II: Enhanced systemic and secreted antibody responses against the CFA/I fimbriae by priming with DNA and boosting with a live recombinant Salmonella vaccine

    Directory of Open Access Journals (Sweden)

    M.O. Lásaro

    1999-02-01

    Full Text Available The induction of systemic (IgG and mucosal (IgA antibody responses against the colonization factor I antigen (CFA/I of enterotoxigenic Escherichia coli (ETEC was evaluated in mice primed with an intramuscularly delivered CFA/I-encoding DNA vaccine followed by two oral immunizations with a live recombinant Salmonella typhimurium vaccine strain expressing the ETEC antigen. The booster effect induced by the oral immunization was detected two weeks and one year after the administration of the DNA vaccine. The DNA-primed/Salmonella-boosted vaccination regime showed a synergistic effect on the induced CFA/I-specific systemic and secreted antibody levels which could not be attained by either immunization strategy alone. These results suggest that the combined use of DNA vaccines and recombinant Salmonella vaccine strains can be a useful immunization strategy against enteric pathogens.

  1. Modulation of cellular and humoral immune responses to anHIV-1 DNA vaccine by interleukin-12 and interleukin-18 DNA immunization

    Institute of Scientific and Technical Information of China (English)

    孙永涛; 王福祥; 孙永年; 徐哲; 王临旭; 刘娟; 白雪帆; 黄长形

    2004-01-01

    Objective: To investigate the effect of interleukin-12 (IL-12) and interleukin-18 (IL-18)DNA immunization on immune response induced by HIV-1 DNA vaccine and to explore new strategies for therapeutic HIV DNA vaccine.Methods: The recombinant expression vector pCI-neoGAG was constructed by inserting HIV Gag gene into the eukaryotic expression vector pCI-neo. Balb/c mice were immunized with pCI-neoGAG alone or co-immunized with the DNA encoding for IL-12 or IL-18. Anti-HIV antibody and IFN-γ were tested by ELISA, and splenocytes were isolated for detecting antigen-specific lymphoproliferative responses and specific CTL response by MTT assay and LDH assay respectively. Results: The antiHIV antibody titers of mice co-immunized with pCI-neoGAG and the DNA encoding for IL-12 or IL-18 were lower than that of mice immunized with pCI-neoGAG alone( P < 0.01). In contrast, the IFN-γ level of mice co-immunized with pCI-neoGAG and the DNA encoding for IL-12 or IL-18 was higher than that of mice immunized with pCI-neoGAG alone ( P <0.01). Furthermore, compared with mice injected with pCI-neoGAG alone, the specific CTL cytotoxity activity and antigenspecific lymphoproliferative responses of mice immunized with pCI-neoGAG and the DNA encoding for IL-12 or IL-18 were significantly enhanced respectively ( P < 0.01). Conclusion: The DNA encoding for IL-12 or IL-18 together with HIV DNA vaccine may enhance specific Th-1 responses and cellular immune response elicited in mice. Hence, the DNA encoding for IL-12 or IL-18 are promising immune adjuvants for HIV-1 DNA vaccine.

  2. Mitochondrial genome of the moon jelly Aurelia aurita (Cnidaria, Scyphozoa): A linear DNA molecule encoding a putative DNA-dependent DNA polymerase.

    Science.gov (United States)

    Shao, Zhiyong; Graf, Shannon; Chaga, Oleg Y; Lavrov, Dennis V

    2006-10-15

    The 16,937-nuceotide sequence of the linear mitochondrial DNA (mt-DNA) molecule of the moon jelly Aurelia aurita (Cnidaria, Scyphozoa) - the first mtDNA sequence from the class Scypozoa and the first sequence of a linear mtDNA from Metazoa - has been determined. This sequence contains genes for 13 energy pathway proteins, small and large subunit rRNAs, and methionine and tryptophan tRNAs. In addition, two open reading frames of 324 and 969 base pairs in length have been found. The deduced amino-acid sequence of one of them, ORF969, displays extensive sequence similarity with the polymerase [but not the exonuclease] domain of family B DNA polymerases, and this ORF has been tentatively identified as dnab. This is the first report of dnab in animal mtDNA. The genes in A. aurita mtDNA are arranged in two clusters with opposite transcriptional polarities; transcription proceeding toward the ends of the molecule. The determined sequences at the ends of the molecule are nearly identical but inverted and lack any obvious potential secondary structures or telomere-like repeat elements. The acquisition of mitochondrial genomic data for the second class of Cnidaria allows us to reconstruct characteristic features of mitochondrial evolution in this animal phylum.

  3. DAI (DLM-1/ZBP1) as a genetic adjuvant for DNA vaccines that promotes effective antitumor CTL immunity.

    Science.gov (United States)

    Lladser, Alvaro; Mougiakakos, Dimitrios; Tufvesson, Helena; Ligtenberg, Maarten A; Quest, Andrew Fg; Kiessling, Rolf; Ljungberg, Karl

    2011-03-01

    DNA vaccination is an attractive approach to induce antigen-specific cytotoxic CD8(+) T lymphocytes (CTLs), which can mediate protective antitumor immunity. The potency of DNA vaccines encoding weakly immunogenic tumor-associated antigens (TAAs) can be enhanced by codelivering gene-encoded adjuvants. Pattern recognition receptors (PRRs) that sense intracellular DNA could potentially be used to harness intrinsic immune-stimulating properties of plasmid DNA vaccines. Consequently, the cytosolic DNA sensor, DNA-dependent activator of interferon (IFN) regulatory factors (DAI), was used as a genetic adjuvant. In vivo electroporation (EP) of mice with a DAI-encoding plasmid (pDAI) promoted transcription of genes encoding type I IFNs, proinflammatory cytokines, and costimulatory molecules. Coimmunization with pDAI and antigen-encoding plasmids enhanced in vivo antigen-specific proliferation, and induction of effector and memory CTLs. Moreover, codelivery of pDAI effectively promoted CTL and CD4(+) Th1 responses to the TAA survivin. The DAI-enhanced CTL induction required nuclear factor κB (NF-κB) activation and type I IFN signaling, but did not involve the IFN regulatory factor 3 (IRF3). Codelivery of pDAI also increased CTL responses to the melanoma-associated antigen tyrosinase-related protein-2 (TRP2), enhanced tumor rejection and conferred long-term protection against B16 melanoma challenge. This study constitutes "proof-of-principle" validating the use of intracellular PRRs as genetic adjuvants to enhance DNA vaccine potency.

  4. Gene Gun Bombardment with DNA-Coated Golden Particles Enhanced the Protective Effect of a DNA Vaccine Based on Thioredoxin Glutathione Reductase of Schistosoma japonicum

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    Yan Cao

    2013-01-01

    Full Text Available Schistosomiasis, caused by infection with Schistosoma species, remains an important parasitic zoonosis. Thioredoxin glutathione reductase of Schistosoma japonicum (SjTGR plays an important role in the development of the parasite and for its survival. Here we present a recombinant plasmid DNA vaccine, pVAX1/SjTGR, to estimate its protection against S. japonicum in BALB/c mice. The DNA vaccine administrated by particle bombardment induced higher protection than by intramuscular injection. All animals vaccinated with pVAX1/SjTGR developed significant specific anti-SjTGR antibodies than control groups. Moreover, animals immunized by gene gun exhibited a splenocyte proliferative response, with an increase in IFN-γ and IL-4. The recombinant plasmid administrated by gene gun achieved a medium protective efficacy of 27.83–38.83% ( of worm reduction and 40.38–44.51% ( of liver egg count reduction. It suggests that different modes of administering a DNA vaccine can influence the protective efficacy induced by the vaccine. Interestingly, from the enzymatic activity results, we found that worms obtained from pVAX1/SjTGR-vaccinated animals expressed lower enzymatic activity than the control group and the antibodies weakened the enzymatic activity of SjTGR in vitro, too. It implies that the high-level antibodies may contribute to the protective effects.

  5. Contamination of DNase Preparations Confounds Analysis of the Role of DNA in Alum-Adjuvanted Vaccines.

    Science.gov (United States)

    Noges, Laura E; White, Janice; Cambier, John C; Kappler, John W; Marrack, Philippa

    2016-08-15

    Aluminum salt (alum) adjuvants have been used for many years as adjuvants for human vaccines because they are safe and effective. Despite its widespread use, the means by which alum acts as an adjuvant remains poorly understood. Recently, it was shown that injected alum is rapidly coated with host chromatin within mice. Experiments suggested that the host DNA in the coating chromatin contributed to alum's adjuvant activity. Some of the experiments used commercially purchased DNase and showed that coinjection of these DNase preparations with alum and Ag reduced the host's immune response to the vaccine. In this study, we report that some commercial DNase preparations are contaminated with proteases. These proteases are responsible for most of the ability of DNase preparations to inhibit alum's adjuvant activity. Nevertheless, DNase somewhat reduces responses to some Ags with alum. The effect of DNase is independent of its ability to cleave DNA, suggesting that alum improves CD4 responses to Ag via a pathway other than host DNA sensing. Copyright © 2016 by The American Association of Immunologists, Inc.

  6. A Moraxella bovis pili vaccine produced by recombinant DNA technology for the prevention of infectious bovine keratoconjunctivitis.

    Science.gov (United States)

    Lepper, A W; Elleman, T C; Hoyne, P A; Lehrbach, P R; Atwell, J L; Schwartzkoff, C L; Egerton, J R; Tennent, J M

    1993-07-01

    Pili (fimbriae) were prepared from Moraxella bovis strain Dalton 2d (Dal2d) and from a derivative of Pseudomonas aeruginosa K/2PfS that contained a plasmid-borne Dal2d pilin gene and produced pili having serogroup-specific identity to Dal2d. Nine calves were vaccinated with two doses each of 30 micrograms authentic M. bovis Dal2d pili in oil adjuvant and 10 calves were vaccinated with a similar dose of P. aeruginosa-derived Dal2d pili in the same formulation. All 19 calves and 10 non-vaccinated controls were challenged by instillation of 1 x 10(9) virulent M. bovis Dal2d cells into both conjunctival sacs 19 days after the second vaccine dose. The serological response to vaccination and the degree of protection against experimentally induced infectious bovine keratoconjunctivitis (IBK) were assessed. None of the nine calves vaccinated with authentic M. bovis Dal2d pili developed IBK while two of those vaccinated with P. aeruginosa-derived Dal2d pili developed lesions which accounted for a mean group lesion score of 0.3. In contrast, 9 of the 10 non-vaccinated calves developed IBK lesions, the majority of which were progressive, required early treatment and accounted for a mean group lesion score of 1.5. These results demonstrate the potential of a relatively low dose of pili produced by recombinant DNA technology for development of an effective vaccine against IBK.

  7. Loss of long term protection with the inclusion of HIV pol to a DNA vaccine encoding gag.

    Science.gov (United States)

    Garrod, Tamsin J; Gargett, Tessa; Yu, Wenbo; Major, Lee; Burrell, Christopher J; Wesselingh, Steven; Suhrbier, Andreas; Grubor-Bauk, Branka; Gowans, Eric J

    2014-11-04

    Traditional vaccine strategies that induce antibody responses have failed to protect against HIV infection in clinical trials, and thus cell-mediated immunity is now an additional criterion. Recent clinical trials that aimed to induce strong T cell responses failed to do so. Therefore, to enhance induction of protective T cell responses, it is crucial that the optimum antigen combination is chosen. Limited research has been performed into the number of antigens selected for an HIV vaccine. This study aimed to compare DNA vaccines encoding either a single HIV antigen or a combination of two antigens, using intradermal vaccination of C57BL/6 mice. Immune assays were performed on splenocytes, and in vivo protection was examined by challenge with a chimeric virus, EcoHIV, able to infect mouse but not human leukocytes, at 10 days (short term) and 60 days (long term) post final vaccination. At 60 days there was significantly lower frequency of induced antigen-specific CD8(+) T cells in the spleens of pCMVgag-pol-vaccinated mice compared with mice which received pCMVgag only. Most importantly, short term viral control of EcoHIV was similar for pCMVgag and pCMVgag-pol-vaccinated mice at day 10, but only the pCMVgag-vaccinated significantly controlled EcoHIV at day 60 compared with pCMV-vaccinated mice, showing that control was reduced with the inclusion of the HIV pol gene. Copyright © 2014 Elsevier B.V. All rights reserved.

  8. CpG DNA enhances the immune responses elicited by the DNA vaccine against foot-and-mouth disease virus in guinea pigs

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    CpG DNA is DNA sequence that has immune stimulatory effects. Several lines of investigation over the past few years indicate that CpG DNA plays an important role in the induction of immune responses to DNA vaccines.In this study, CpG DNA-containing synthetic oligodeoxynucleotide (CpG-ODN) was cloned into the eukaryotic expression plasmid encoding a fusion protein containing β- galactosidase from E. coli and immunogenic epitopes of footand-mouth disease virus (FMDV) type O, and the immune responses induced by the plasmid were assayed. The results showed that guinea pigs immunized with the recombinant plasmid containing CpG-ODN generated a higher level of FMDV-neutralizing antibody and a stronger T cell proliferative response and protection against viral challenge than those receiving the plasmid containing no CpG-ODN. Our study demonstrated that it is an effective route to enhance the efficacy of DNA vaccines by inserting exogenous CpG DNA into the plasmids, and the DNA vaccine developed here is a promising candidate to prevent FMDV infection.``

  9. Preparation and efficacy of Newcastle disease virus DNA vaccine encapsulated in chitosan nanoparticles

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    Zhao K

    2014-01-01

    Full Text Available Kai Zhao,1,2,* Yang Zhang,1,2,* Xiaoyan Zhang,1,* Wei Li,1 Ci Shi,1,2 Chen Guo,1 Chunxiao Dai,3 Qian Chen,1 Zheng Jin,3 Yan Zhao,2 Hongyu Cui,2 Yunfeng Wang2 1College of Life Science, Heilongjiang University, 2Division of Avian Infectious Diseases, State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute, 3Key Laboratory of Chemical Engineering Process and Technology for High-Efficiency Conversion, Heilongjiang University, Harbin, People's Republic of China *These authors contributed equally to this work Abstract: Optimal preparation conditions of Newcastle disease virus (NDV F gene deoxyribonucleic acid (DNA vaccine encapsulated in chitosan nanoparticles (pFNDV-CS-NPs were determined. The pFNDV-CS-NPs were prepared according to a complex coacervation method. The pFNDV-CS-NPs were produced with good morphology, high stability, a mean diameter of 199.5 nm, encapsulation efficiency of 98.37%±0.87%, loading capacity of 36.12%±0.19%, and a zeta potential of +12.11 mV. The in vitro release assay showed that the plasmid DNA was sustainably released from the pFNDV-CS-NPs, up to 82.9%±2.9% of the total amount. Cell transfection test indicated that the vaccine expressed the F gene in cells and maintained good bioactivity. Additionally, the safety of mucosal immunity delivery system of the pFNDV-CS-NPs was also tested in vitro by cell cytotoxicity and in vivo by safety test in chickens. In vivo immunization showed that better immune responses of specific pathogen-free chickens immunized with the pFNDV-CS-NPs were induced, and prolonged release of the plasmid DNA was achieved compared to the chickens immunized with the control plasmid. This study lays the foundation for the further development of mucosal vaccines and drugs encapsulated in chitosan nanoparticles. Keywords: Newcastle disease, DNA vaccine, chitosan nanoparticles, mucosal immunity delivery system, immune effectiveness

  10. Replication of hepatitis B virus in primary duck hepatocytes transfected with linear viral DNA

    Institute of Scientific and Technical Information of China (English)

    Yun-Qing Yao; Wei-Ping Zhou; Hong Ren; Qi Liu; Shu-Hua Guo; Ding-Feng Zhang; Ni Tang; Ai-Long Huang; Xiao-Yi Zou; Jiang-Feng Xiao; Yun Luo; Da-Zhi Zhang; Bo Wang

    2005-01-01

    AIM: To explore the expression and replication of hepatitis B virus (HBV) DNA in primary duck hepatocytes (PDHs).METHODS: Complete HBV genome was transfected into PDHs by electroporation (transfected group, 1.19×1012copies of linear HBV DNA/1×107 PDHs). After 1-5 d of transfection, HBsAg and HBeAg in the supernatant and lysate of PDHs were measured with the IMX System.Meanwhile, replicative intermediates of HBV DNA were analyzed by Southern blotting and Dot blotting. PDHs electroporated were used as control group.RESULTS: HBsAg in the hepatocyte lysates of transfected group was 15.24 (1 d), 14.55 (3 d) and 5.13 (5 d; P/N values, positive≥2.1) respectively. HBeAg was negative (<2.1). Both HBsAg and HBeAg were negative in the supernatant of transfected group. Dot blotting revealed that HBV DNA was strongly positive in the transfected group and negative in the control group. Southern blot analysis of intracellular total DNA indicated that there were relaxed circular (rc DNA), covalently closed circular (ccc DNA), and single-stranded (ss DNA) HBV DNA replicative intermediates in the transfected group, there was no integrated HBV DNA in the cellular genome. These parameters were negative in control group.CONCLUSION: Expression and replication of HBV genes can occur in hepatocytes from non-mammalian species.HBV replication has no critical species-specificity, and yet hepatic-specific regulating factors in hepatocytes may be essential for viral replication.

  11. Cellular immune responses of BALB/c mice induced by intramuscular injection of PRRSV-ORF5 DNA vaccine with different doses

    Institute of Scientific and Technical Information of China (English)

    CHENG Anchun; WANG Mingshu; CHEN Xiwen; XINI Nigen; DOU Wenbo; LI Xuemei; LIU Wumei; WANG Gang; ZHANG Pingying

    2007-01-01

    BALB/c mice were immunized with 50 μg,100 μg,200 μg of pcDNA-PRRSV-ORF5 DNA vaccine respectively by intramuscular injection,with PBS and pcDNA3.1(+)as controls.Fluorescence activated cell Sorter (FACS)was used to detect the number of CD4+ and CD8+T-lymphocytes.T-lymphocyte proliferation test was used to detect proliferation of the T-lymphocyte cells in peripheral blood lymphocytes of mice vaccinated with pcDNA-PRRSV-ORF5 DNA vaccine.The results showed that the difference in ConA response to T-lymphocytes in blood was highly significant between all experimental groups and the control group(P<0.01).The number of CD4+T-lymphocytes in experimental groups was significantly higher than that of the control group 7d after vaccination.The number of CD8+ T-lymphocytes in the experimental groups was higher than that of the control group 28 d after vaccination.Mice immunized with a higher dose(200 μg)of DNA vaccine demonstrated higher cellular immune response than those immunized with a lower dose(100 μg,50 μg)of DNA vaccine.The results demonstrated that pcDNA-PRRSV-ORF5 DNA vaccine could induce a good cellular immune response which may be dose-dependent.

  12. Design of different strategies of multivalent DNA-based vaccination against rabies and canine distemper in mice and dogs

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    Touihri Leila

    2012-12-01

    Full Text Available Abstract Background During the vaccination campaigns, puppies younger than 3 months old are not targeted and remain unvaccinated for at least the first year of their lives. Almost half of the reported rabid dogs are 6 months or younger. Hence, we should recommend the vaccination against rabies of young puppies. Unfortunately, owing to the exposure of puppies to infections with either canine parvovirus (CPV or distemper virus (CDV after the intervention of the vaccinators, owners are reluctant to vaccinate puppies against rabies. Therefore, it is necessary to include the CPV and CDV valences in the vaccine against rabies. Multivalent DNA-based vaccination in dogs, including rabies and distemper valences, could help in raising vaccine coverage. Methods We have designed monovalent and multivalent DNA-based vaccine candidates for in vitro and in vivo assays. These plasmids encode to the rabies virus glycoprotein and/or the canine distemper virus hemagglutinin. The first strategy of multivalent DNA-based vaccination is by mixing plasmids encoding to a single antigen each. The second is by simply fusing the genes of the antigens together. The third is by adding the foot and mouth disease virus (FMDV 2A oligopeptide gene into the antigen genes. The last strategy is by the design and use of a bicistronic plasmid with an “Internal Ribosome Entry Site” (IRES domain. Results The monovalent construct against canine distemper was efficiently validated by inducing higher humoral immune responses compared to cell-culture-derived vaccine both in mice and dogs. All multivalent plasmids efficiently expressed both valences after in vitro transfection of BHK-21 cells. In BALB/c mice, the bicistronic IRES-dependant construct was the most efficient inducer of virus-neutralizing antibodies against both valences. It was able to induce better humoral immune responses compared to the administration of either cell-culture-derived vaccines or monovalent plasmids. The

  13. Poly(lactic-co-glycolic acid) nanoparticles as candidate DNA vaccine carrier for oral immunization of Japanese flounder (Paralichthys olivaceus) against lymphocystis disease virus.

    Science.gov (United States)

    Tian, Jiyuan; Yu, Juan

    2011-01-01

    In order to protect DNA vaccine against degradation in alimentary tract of fish, poly(lactic-co-glycolic acid) (PLGA) nanoparticles encapsulating vaccine were prepared using W/O/W emulsification combined with spray drying technique in our laboratory. The characteristics of PLGA nanoparticles were described as follows: (1) shape, spherical; (2) size, 0.05). Pearson's correlation displayed that correlation of immune factors mentioned above (not including serum complement) were all positive for fish vaccinated. The data in this study suggested that PLGA nanoparticles were promising carriers for plasmid DNA vaccine and might be used to vaccinate fish by oral approach.

  14. Differential effects of IL-15 on the generation, maintenance and cytotoxic potential of adaptive cellular responses induced by DNA vaccination.

    Science.gov (United States)

    Li, Jinyao; Valentin, Antonio; Ng, Sinnie; Beach, Rachel Kelly; Alicea, Candido; Bergamaschi, Cristina; Felber, Barbara K; Pavlakis, George N

    2015-02-25

    IL-15 is an important cytokine for the regulation of lymphocyte homeostasis. However, the role of IL-15 in the generation, maintenance and cytotoxic potential of antigen specific T cells is not fully understood. Because the route of antigenic delivery and the vaccine modality could influence the IL-15 requirement for mounting and preserving cytotoxic T cell responses, we have investigated the immunogenicity of DNA-based vaccines in IL-15 KO mice. DNA vaccination with SIV Gag induced antigen-specific CD4(+) and CD8(+) T cells in the absence of IL-15. However, the absolute number of antigen-specific CD8(+) T cells was decreased in IL-15 KO mice compared to WT animals, suggesting that IL-15 is important for the generation of maximal number of antigen-specific CD8(+) T cells. Interestingly, antigen-specific memory CD8 cells could be efficiently boosted 8 months after the final vaccination in both WT and KO strains of mice, suggesting that the maintenance of antigen-specific long-term memory T cells induced by DNA vaccination is comparable in the absence and presence of IL-15. Importantly, boosting by DNA 8-months after vaccination revealed severely reduced granzyme B content in CD8(+) T cells of IL-15 KO mice compared to WT mice. This suggests that the cytotoxic potential of the long-term memory CD8(+) T cells is impaired. These results suggest that IL-15 is not essential for the generation and maintenance of adaptive cellular responses upon DNA vaccination, but it is critical for the preservation of maximal numbers and for the activity of cytotoxic CD8(+) T cells. Published by Elsevier Ltd.

  15. Construction of sperm-specific lactate dehydrogenase DNA vaccine and experimental study of its immuno-contraceptive effect on mice

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    Lactate dehydrogenase C4 (LDHC4) is a key enzyme for sperm metabolism. It is distributed specifically in testis and is highly immunogenic. In this study, two DNA vaccines pVAX1-hLDHC and pVAX1-mLDHC were constructed by inserting coding sequences of human and mice LDHC4 into the eukaryotic ex-pression vector pVAX1. The production of LDHC4 specific antibodies was induced in the sera of vac-cinated mice and the reproductive tract secretions of vaccinated female mice through immunization by mucosal surface instillation. Furthermore, the antibody titer increased with the times of immunization. In the mating experiment, the number of newborns of the vaccinated mice reduced significantly and some immunized female mice even lost the ability to bear any offsprings, suggesting that the difference between the immunized and control mice was statistically significant. Sperm agglutination analysis indicated that both the antisera from immunized mice and the reproductive tract secretions of vacci-nated female mice could agglutinate normal sperms. Results of immunohistochemistry showed that the antibodies present in the sera of immunized mice and the reproductive tract secretions of vaccinated female mice could specifically react with LDHC4 antigen, which mainly locates in the cytoplasm, acrosome membrane externa and acrosome capsule of the sperm. Taken together, our results indicated that the constructed contraceptive DNA vaccines did yield immunocontraceptive effects on mice and this would enable clinical trials in near future.

  16. Construction of sperm-specific lactate dehydrogenase DNA vaccine and experimental study of its immunocontraceptive effect on mice

    Institute of Scientific and Technical Information of China (English)

    CHEN Yong; ZHANG Duo; XIN Na; XIONG YongZhong; CHEN Ping; LI Bo; TU XiangDong; LAN FengHua

    2008-01-01

    Lactate dehydrogenase C4 (LDHC4) is a key enzyme for sperm metabolism. It is distributed specifically in testis and is highly Immunogenic. In this study, two DNA vaccines pVAX1-hLDHC and pVAX1-mLDHC were constructed by inserting coding sequences of human and mice LDHC4 into the eukaryotic expression vector pVAX1. The production of LDHC4 specific antibodies was induced in the sere of vaccinated mice and the reproductive tract secretions of vaccinated female mice through Immunization by mucosal surface instillation. Furthermore, the antibody titer increased with the times of immunization.In the mating experiment, the number of newborns of the vaccinated mice reduced significantly and some immunized female mice even lost the ability to bear any offsprings, suggesting that the difference between the immunized and control mice was statistically significant. Sperm agglutination analysis indicated that both the antisera from immunized mice end the reproductive tract secretions of vaccinated female mice could agglutinate normal sperms. Results of immunohistochemistry showed that the antibodies present in the sere of immunized mice and the reproductive tract secretions of vaccinated female mice could specifically react with LDHC4 antigen, which mainly locates in the cytoplasm,acrosome membrane externa and acrosome capsule of the sperm. Taken together, our results indicated that the constructed contraceptive DNA vaccines did yield immunocontreceptive effects on mice and this would enable clinical trials in near future.

  17. A recombinant DNA vaccine protects mice deficient in the alpha/beta interferon receptor against lethal challenge with Usutu virus.

    Science.gov (United States)

    Martín-Acebes, Miguel A; Blázquez, Ana-Belén; Cañas-Arranz, Rodrigo; Vázquez-Calvo, Ángela; Merino-Ramos, Teresa; Escribano-Romero, Estela; Sobrino, Francisco; Saiz, Juan-Carlos

    2016-04-19

    Usutu virus (USUV) is a mosquito-borne flavivirus whose circulation had been confined to Africa since it was first detected in 1959. However, in the last decade USUV has emerged in Europe causing episodes of avian mortality and sporadic severe neuroinvasive infections in humans. Remarkably, adult laboratory mice exhibit limited susceptibility to USUV infection, which has impaired the analysis of the immune responses, thus complicating the evaluation of virus-host interactions and of vaccine candidates against this pathogen. In this work, we showed that mice deficient in the alpha/beta interferon receptor (IFNAR (-/-) mice) were highly susceptible to USUV infection and provided a lethal challenge model for vaccine testing. To validate this infection model, a plasmid DNA vaccine candidate encoding the precursor of membrane (prM) and envelope (E) proteins of USUV was engineered. Transfection of cultured cells with this plasmid resulted in expression of USUV antigens and the assembly and secretion of small virus-like particles also known as recombinant subviral particles (RSPs). A single intramuscular immunization with this plasmid was sufficient to elicit a significant level of protection against challenge with USUV in IFNAR (-/-) mice. The characterization of the humoral response induced revealed that DNA vaccination primed anti-USUV antibodies, including neutralizing antibodies. Overall, these results probe the suitability of IFNAR (-/-) mice as an amenable small animal model for the study of USUV host virus interactions and vaccine testing, as well as the feasibility of DNA-based vaccine strategies for the control of this pathogen.

  18. Fusion of CTLA-4 with HPV16 E7 and E6 enhanced the potency of therapeutic HPV DNA vaccine.

    Directory of Open Access Journals (Sweden)

    Lili Gan

    Full Text Available Preventive anti-HPV vaccines are effective against HPV infection but not against existing HPV-associated diseases, including cervical cancer and other malignant diseases. Therefore, the development of therapeutic vaccines is urgently needed. To improve anti-tumor effects of therapeutic vaccine, we fused cytotoxic T-lymphocyte antigen 4 (CTLA-4 with HPV16 E7 and E6 as a fusion therapeutic DNA vaccine (pCTLA4-E7E6. pCTLA4-E7E6 induced significantly higher anti-E7E6 specific antibodies and relatively stronger specific CTL responses than the nonfusion DNA vaccine pE7E6 in C57BL/6 mice bearing with TC-1 tumors. pCTLA4-E7E6 showed relatively stronger anti-tumor effects than pE7E6 in therapeutic immunization. These results suggest that fusing CTLA-4 with E7E6 is a useful strategy to develop therapeutic HPV DNA vaccines. In addition, fusing the C-terminal of E7 with the N-terminal of E6 impaired the functions of both E7 and E6.

  19. Inhibition of hepatitis B virus replication with linear DNA sequences expressing antiviral micro-RNA shuttles

    Energy Technology Data Exchange (ETDEWEB)

    Chattopadhyay, Saket; Ely, Abdullah; Bloom, Kristie; Weinberg, Marc S. [Antiviral Gene Therapy Research Unit, University of the Witwatersrand (South Africa); Arbuthnot, Patrick, E-mail: Patrick.Arbuthnot@wits.ac.za [Antiviral Gene Therapy Research Unit, University of the Witwatersrand (South Africa)

    2009-11-20

    RNA interference (RNAi) may be harnessed to inhibit viral gene expression and this approach is being developed to counter chronic infection with hepatitis B virus (HBV). Compared to synthetic RNAi activators, DNA expression cassettes that generate silencing sequences have advantages of sustained efficacy and ease of propagation in plasmid DNA (pDNA). However, the large size of pDNAs and inclusion of sequences conferring antibiotic resistance and immunostimulation limit delivery efficiency and safety. To develop use of alternative DNA templates that may be applied for therapeutic gene silencing, we assessed the usefulness of PCR-generated linear expression cassettes that produce anti-HBV micro-RNA (miR) shuttles. We found that silencing of HBV markers of replication was efficient (>75%) in cell culture and in vivo. miR shuttles were processed to form anti-HBV guide strands and there was no evidence of induction of the interferon response. Modification of terminal sequences to include flanking human adenoviral type-5 inverted terminal repeats was easily achieved and did not compromise silencing efficacy. These linear DNA sequences should have utility in the development of gene silencing applications where modifications of terminal elements with elimination of potentially harmful and non-essential sequences are required.

  20. Leptospirosis vaccines

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    Jin Li

    2007-12-01

    Full Text Available Abstract Leptospirosis is a serious infection disease caused by pathogenic strains of the Leptospira spirochetes, which affects not only humans but also animals. It has long been expected to find an effective vaccine to prevent leptospirosis through immunization of high risk humans or animals. Although some leptospirosis vaccines have been obtained, the vaccination is relatively unsuccessful in clinical application despite decades of research and millions of dollars spent. In this review, the recent advancements of recombinant outer membrane protein (OMP vaccines, lipopolysaccharide (LPS vaccines, inactivated vaccines, attenuated vaccines and DNA vaccines against leptospirosis are reviewed. A comparison of these vaccines may lead to development of new potential methods to combat leptospirosis and facilitate the leptospirosis vaccine research. Moreover, a vaccine ontology database was built for the scientists working on the leptospirosis vaccines as a starting tool.

  1. Development of new plasmid DNA vaccine vectors with R1-based replicons

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    Bower Diana M

    2012-08-01

    Full Text Available Abstract Background There has been renewed interest in biopharmaceuticals based on plasmid DNA (pDNA in recent years due to the approval of several veterinary DNA vaccines, on-going clinical trials of human pDNA-based therapies, and significant advances in adjuvants and delivery vehicles that have helped overcome earlier efficacy deficits. With this interest comes the need for high-yield, cost-effective manufacturing processes. To this end, vector engineering is one promising strategy to improve plasmid production. Results In this work, we have constructed a new DNA vaccine vector, pDMB02-GFP, containing the runaway R1 origin of replication. The runaway replication phenotype should result in plasmid copy number amplification after a temperature shift from 30°C to 42°C. However, using Escherichia coli DH5α as a host, we observed that the highest yields of pDMB02-GFP were achieved during constant-temperature culture at 30°C, with a maximum yield of approximately 19 mg pDNA/g DCW being observed. By measuring mRNA and protein levels of the R1 replication initiator protein, RepA, we determined that RepA may be limiting pDMB02-GFP yield at 42°C. A mutant plasmid, pDMB-ATG, was constructed by changing the repA start codon from the sub-optimal GTG to ATG. In cultures of DH5α[pDMB-ATG], temperature-induced plasmid amplification was more dramatic than that observed with pDMB02-GFP, and RepA protein was detectable for several hours longer than in cultures of pDMB02-GFP at 42°C. Conclusions Overall, we have demonstrated that R1-based plasmids can produce high yields of high-quality pDNA without the need for a temperature shift, and have laid the groundwork for further investigation of this class of vectors in the context of plasmid DNA production.

  2. Preclinical evaluation of multi antigenic HCV DNA vaccine for the prevention of Hepatitis C virus infection.

    Science.gov (United States)

    Lee, Hyojin; Jeong, Moonsup; Oh, Jooyeon; Cho, Youngran; Shen, Xuefei; Stone, John; Yan, Jian; Rothkopf, Zachary; Khan, Amir S; Cho, Byung Mun; Park, Young K; Weiner, David B; Son, Woo-Chan; Maslow, Joel N

    2017-03-07

    Direct-acting antiviral treatment for hepatitis C virus (HCV) infection is costly and does not protect from re-infection. For human and chimpanzees, recovery from acute HCV infection correlates with host CD4+ and CD8+ T cell responses. DNA plasmids targeting the HCV non-structural antigens NS3, NS4, and NS5, were previously reported to induce robust and sustained T cell responses in mice and primates. These plasmids were combined with a plasmid encoding cytokine IL-28B, together named as VGX-6150. The dose-dependent T cell response and safety of VGX-6150 administered intramuscularly and followed by electroporation was assessed in mice. Immune responses plateaued at 20 μg/dose with IL-28B demonstrating significant immunoadjuvant activity. Mice administered VGX-6150 at 40, 400, and 800 μg given either as a single injection or as 14 injections given bi-weekly over 26 weeks showed no vaccine related changes in any clinical parameter compared to placebo recipients. There was no evidence of VGX-6150 accumulation at the injection site or in any organ 1 month following the 14(th) vaccination. Based on these studies, the approximate lethal dose (ALD) exceeds 800 μg/dose and the NOAEL was 800 μg/dose in mouse. In conclusion, VGX-6150 appears safe and a promising preventive vaccine candidate for HCV infection.

  3. Epitope analysis and protection by a ROP19 DNA vaccine against Toxoplasma gondii

    Directory of Open Access Journals (Sweden)

    Zhou Jian

    2016-01-01

    Full Text Available We used bioinformatics approaches to identify B-cell and T-cell epitopes on the ROP19 protein of Toxoplasma gondii. Then, we constructed plasmids with ROP19 (pEGFP-C1-ROP19 and injected them into BALB/c mice to test the immunoprotection induced by this vaccine candidate. The results showed that immunization with pEGFP-C1-ROP19 induced effective cellular and humoral immune responses in mice; specifically, high serum levels of T. gondii-specific IgG and increased interferon-gamma production by splenocytes. Furthermore, the mice vaccinated with pROP19 had significantly fewer brain cysts (583 ± 160 than the mice injected with phosphate-buffered saline (1350 ± 243 or with the control plasmid, pEGFP-C1 (1300 ± 167. Compared with PBS-treated mice, those immunized with pROP19 had only 43% of the number of brain cysts. These results suggest that the DNA vaccine encoding ROP19 induced a significant immune response and provided protection against a challenge with T. gondii strain PRU cysts.

  4. Preclinical evaluation of multi antigenic HCV DNA vaccine for the prevention of Hepatitis C virus infection

    Science.gov (United States)

    Lee, Hyojin; Jeong, Moonsup; Oh, Jooyeon; Cho, Youngran; Shen, Xuefei; Stone, John; Yan, Jian; Rothkopf, Zachary; Khan, Amir S.; Cho, Byung Mun; Park, Young K.; Weiner, David B.; Son, Woo-Chan; Maslow, Joel N.

    2017-01-01

    Direct-acting antiviral treatment for hepatitis C virus (HCV) infection is costly and does not protect from re-infection. For human and chimpanzees, recovery from acute HCV infection correlates with host CD4+ and CD8+ T cell responses. DNA plasmids targeting the HCV non-structural antigens NS3, NS4, and NS5, were previously reported to induce robust and sustained T cell responses in mice and primates. These plasmids were combined with a plasmid encoding cytokine IL-28B, together named as VGX-6150. The dose-dependent T cell response and safety of VGX-6150 administered intramuscularly and followed by electroporation was assessed in mice. Immune responses plateaued at 20 μg/dose with IL-28B demonstrating significant immunoadjuvant activity. Mice administered VGX-6150 at 40, 400, and 800 μg given either as a single injection or as 14 injections given bi-weekly over 26 weeks showed no vaccine related changes in any clinical parameter compared to placebo recipients. There was no evidence of VGX-6150 accumulation at the injection site or in any organ 1 month following the 14th vaccination. Based on these studies, the approximate lethal dose (ALD) exceeds 800 μg/dose and the NOAEL was 800 μg/dose in mouse. In conclusion, VGX-6150 appears safe and a promising preventive vaccine candidate for HCV infection. PMID:28266565

  5. DermaVir: a plasmid DNA-based nanomedicine therapeutic vaccine for the treatment of HIV/AIDS.

    Science.gov (United States)

    Lori, Franco

    2011-10-01

    The HIV global pandemic continues to rage with over 33 million people living with the disease. Although multidrug therapy has improved the prognosis for those infected by the virus, it has not eradicated the infection. Immunological therapies, including therapeutic vaccines, are needed to supplement drug therapy in the search for a 'functional cure' for HIV. DermaVir (Genetic Immunity Kft, Budapest, Hungary and McLean, Virginia, USA), an experimental HIV/AIDS therapeutic vaccine, combines three key elements of rational therapeutic vaccine design: a single plasmid DNA (pDNA) immunogen expressing 15 HIV antigens, a synthetic pDNA nanomedicine formulation and a dendritic cell-targeting topical-vaccine administration. DermaVir's novel mechanism of action, natural transport by epidermal Langerhans cells to the lymph nodes to express the pDNA-encoded HIV antigens and induce precursor/memory T cells with high proliferation capacity, has been consistently demonstrated in mouse, rabbit, primate and human subjects. Safety, immunogenicity and preliminary efficacy of DermaVir have been clinically demonstrated in HIV-infected human subjects. The DermaVir technology platform for dendritic cell-based therapeutic vaccination might offer a new treatment paradigm for cancer and infectious diseases.

  6. A DNA vaccine targeting TcdA and TcdB induces protective immunity against Clostridium difficile.

    Science.gov (United States)

    Zhang, Bao-Zhong; Cai, Jianpiao; Yu, Bin; Hua, Yanhong; Lau, Candy Choiyi; Kao, Richard Yi-Tsun Tsun; Sze, Kong-Hung; Yuen, Kwok-Yung; Huang, Jian-Dong

    2016-10-22

    Clostridium difficile-associated disease (CDAD) constitutes a great majority of hospital diarrhea cases in industrialized countries and is induced by two types of large toxin molecules: toxin A (TcdA) and toxin B (TcdB). Development of immunotherapeutic approaches, either active or passive, has seen a resurgence in recent years. Studies have described vaccine plasmids that express either TcdA and/or TcdB receptor binding domain (RBD). However, the effectiveness of one vector encoding both toxin RBDs against CDAD has not been evaluated. In the study, we constructed highly optimized plasmids to express the receptor binding domains of both TcdA and TcdB from a single vector. The DNA vaccine was evaluated in two animal models for its immunogenicity and protective effects. The DNA vaccine induced high levels of serum antibodies to toxin A and/or B and demonstrated neutralizing activity in both in vitro and in vivo systems. In a C. difficile hamster infection model, immunization with the DNA vaccine reduced infection severity and conferred significant protection against a lethal C. difficile strain. This study has demonstrated a single plasmid encoding the RBD domains of C. difficile TcdA and TcdB as a DNA vaccine that could provide protection from C. difficile disease.

  7. The site of administration influences both the type and the magnitude of the immune response induced by DNA vaccine electroporation.

    Science.gov (United States)

    Vandermeulen, Gaëlle; Vanvarenberg, Kevin; De Beuckelaer, Ans; De Koker, Stefaan; Lambricht, Laure; Uyttenhove, Catherine; Reschner, Anca; Vanderplasschen, Alain; Grooten, Johan; Préat, Véronique

    2015-06-22

    We investigated the influence of the site of administration of DNA vaccine on the induced immune response. DNA vaccines were administered by electroporation at three different sites: tibial cranial muscle, abdominal skin and ear pinna. Aiming to draw general conclusions about DNA vaccine delivery, we successively used several plasmids encoding either luciferase and ovalbumin as models or gp160 and P1A as vaccines against HIV and P815 mastocytoma, respectively. Low levels and duration of luciferase transgene expression were observed after electroporation of the abdominal skin, partly explaining its lower immunogenic performance as compared to the other sites of administration. Analyses of OT-I CD8+ and OT-II CD4+ T cell responses highlighted the differential impact of the delivery site on the elicited immune response. Muscle electroporation induced the strongest humoral immune response and both muscle and ear pinna sites induced cellular immunity against gp160. Ear pinna delivery generated the highest level of CTL responses against P1A but electroporation of muscle and ear pinna were equally efficient in delaying P815 growth and improving mice survival. The present study demonstrated that the site of administration is a key factor to be tested in the development of DNA vaccine.

  8. Ex vivo transfection of trout pronephros leukocytes, a model for cell culture screening of fish DNA vaccine candidates.

    Science.gov (United States)

    Ortega-Villaizan, M; Martinez-Lopez, A; Garcia-Valtanen, P; Chico, V; Perez, L; Coll, J M; Estepa, A

    2012-09-07

    DNA vaccination opened a new era in controlling and preventing viral diseases since DNA vaccines have shown to be very efficacious where some conventional vaccines have failed, as it occurs in the case of the vaccines against fish novirhabdoviruses. However, there is a big lack of in vitro model assays with immune-related cells for preliminary screening of in vivo DNA vaccine candidates. In an attempt to solve this problem, rainbow trout pronephros cells in early primary culture were transfected with two plasmid DNA constructions, one encoding the green fluorescent protein (GFP) and another encoding the viral haemorrhagic septicaemia virus (VHSV) glycoprotein G (G(VHSV)) - the only viral antigen which has conferred in vivo protection. After assessing the presence of GFP- and G(VHSV)-expressing cells, at transcription and protein levels, the immune response in transfected pronephros cells was evaluated. At 24h post-transfection, G(VHSV) up-regulated migm and tcr transcripts expression, suggesting activation of B and T cells, as well, a high up-regulation of tnfα gene was observed. Seventy-two hours post-transfection, we detected the up-regulation of mx and tnfα genes transcripts and Mx protein which correlated with the induction of an anti-VHSV state. All together we have gathered evidence for successful transfection of pronephros cells with pAE6G, which correlates with in vivo protection results, and is less time-consuming and more rapid than in vivo assays. Therefore, this outcome opens the possibility to use pronephros cells in early primary culture for preliminary screening fish DNA vaccines as well as to further investigate the function that these cells perform in fish immune response orchestration after DNA immunisation.

  9. Comparison of Immune Responses against FMD by a DNA Vaccine Encoding the FMDV/O/IRN/2007 VP1 Gene and the Conventional Inactivated Vaccine in an Animal Model

    Institute of Scientific and Technical Information of China (English)

    Farahnaz Motamedi Sedeh; Hoorieh Soleimanjahi; AmirReza Jalilian; Homayoon Mahravani

    2012-01-01

    Foot-and-mouth disease virus (FMDV) is highly contagious and responsible for huge outbreaks among cloven hoofed animals.The aim of the present study is to evaluate a plasmid DNA immunization system that expresses the FMDV/O/IRN/2007 VP1 gene and compare it with the conventional inactivated vaccine in an animal model.The VP1 gene was sub-cloned into the unique Kpn I and BamH I cloning sites of the pcDNA3.1+ and pEGFP-N1 vectors to construct the VP1 gene cassettes.The transfected BHKT7 cells with sub-cloned pEGFP-N1-VP1 vector expressed GFP-VP1 fusion protein and displayed more green fluorescence spots than the transfected BHKT7 cells with pEGFP-N1 vector,which solely expressed the GFP protein.Six mice groups were respectively immunized by the sub-cloned pcDNA3.1+-VP1 gene cassette as the DNA vaccine,DNA vaccine and PCMV-SPORT-GMCSF vector (as molecular adjuvant) together,conventional vaccine,PBS (as negative control),pcDNA3.1+ vector (as control group) and PCMV-SPORT vector that contained the GMCSF gene (as control group).Significant neutralizing antibody responses were induced in the mice which were immunized using plasmid vectors expressing the VP1 and GMCSF genes together,the DNA vaccine alone and the conventional inactivated vaccine (P<0.05).Co-administration of DNA vaccine and GMCSF gene improved neutralizing antibody response in comparison with administration of the DNA vaccine alone,but this response was the most for the conventional vaccine group.However,induction of humeral immunity response in the conventional vaccine group was more protective than for the DNA vaccine,but T-cell proliferation and IFN-γ concentration were the most in DNA vaccine with the GMCSF gene.Therefore the group that was vaccinated by DNA vaccine with the GMCSF gene,showed protective neutralizing antibody response and the most Th1 cellular immunity.

  10. Protective efficacy and immunogenicity of a combinatory DNA vaccine against Influenza A Virus and the Respiratory Syncytial Virus.

    Directory of Open Access Journals (Sweden)

    Viktoria Stab

    Full Text Available The Respiratory Syncytial Virus (RSV and Influenza A Virus (IAV are both two major causative agents of severe respiratory tract infections in humans leading to hospitalization and thousands of deaths each year. In this study, we evaluated the immunogenicity and efficacy of a combinatory DNA vaccine in comparison to the single component vaccines against both diseases in a mouse model. Intramuscular electroporation with plasmids expressing the hemagglutinin (HA of IAV and the F protein of RSV induced strong humoral immune responses regardless if they were delivered in combination or alone. In consequence, high neutralizing antibody titers were detected, which conferred protection against a lethal challenge with IAV. Furthermore, the viral load in the lungs after a RSV infection could be dramatically reduced in vaccinated mice. Concurrently, substantial amounts of antigen-specific, polyfunctional CD8⁺ T-cells were measured after vaccination. Interestingly, the cellular response to the hemagglutinin was significantly reduced in the presence of the RSV-F encoding plasmid, but not vice versa. Although these results indicate a suppressive effect of the RSV-F protein, the protective efficacy of the combinatory vaccine was comparable to the efficacy of both single-component vaccines. In conclusion, the novel combinatory vaccine against RSV and IAV may have great potential to reduce the rate of severe respiratory tract infections in humans without increasing the number of necessary vaccinations.

  11. Protective efficacy and immunogenicity of a combinatory DNA vaccine against Influenza A Virus and the Respiratory Syncytial Virus.

    Science.gov (United States)

    Stab, Viktoria; Nitsche, Sandra; Niezold, Thomas; Storcksdieck Genannt Bonsmann, Michael; Wiechers, Andrea; Tippler, Bettina; Hannaman, Drew; Ehrhardt, Christina; Uberla, Klaus; Grunwald, Thomas; Tenbusch, Matthias

    2013-01-01

    The Respiratory Syncytial Virus (RSV) and Influenza A Virus (IAV) are both two major causative agents of severe respiratory tract infections in humans leading to hospitalization and thousands of deaths each year. In this study, we evaluated the immunogenicity and efficacy of a combinatory DNA vaccine in comparison to the single component vaccines against both diseases in a mouse model. Intramuscular electroporation with plasmids expressing the hemagglutinin (HA) of IAV and the F protein of RSV induced strong humoral immune responses regardless if they were delivered in combination or alone. In consequence, high neutralizing antibody titers were detected, which conferred protection against a lethal challenge with IAV. Furthermore, the viral load in the lungs after a RSV infection could be dramatically reduced in vaccinated mice. Concurrently, substantial amounts of antigen-specific, polyfunctional CD8⁺ T-cells were measured after vaccination. Interestingly, the cellular response to the hemagglutinin was significantly reduced in the presence of the RSV-F encoding plasmid, but not vice versa. Although these results indicate a suppressive effect of the RSV-F protein, the protective efficacy of the combinatory vaccine was comparable to the efficacy of both single-component vaccines. In conclusion, the novel combinatory vaccine against RSV and IAV may have great potential to reduce the rate of severe respiratory tract infections in humans without increasing the number of necessary vaccinations.

  12. The immunogenicity of tetravalent dengue DNA vaccine in mice pre-exposed to Japanese encephalitis or Dengue virus antigens.

    Science.gov (United States)

    Prompetchara, Eakachai; Ketloy, Chutitorn; Keelapang, Poonsook; Sittisombut, Nopporn; Ruxrungtham, Kiat

    2015-09-01

    Asian countries are an endemic area for both dengue (DENV) and Japanese encephalitis viruses (JEV). While JEV vaccines have been used extensively in this region, DENV vaccines remains under development. Whether preexisting naturally acquired or vaccination-induced immunity against JEV may affect the immune response to dengue vaccine candidate is unclear. In this study we used mice previously immunized with JEV vaccines to evaluate the impact on dengue-specific neutralizing antibody responses to a tetravalent dengue DNA vaccine candidate (TDNA). A tetravalent cocktail of plasmids encoding pre-membrane and envelope proteins from each dengue serotype was administered into mice which had been previously primed with inactivated or live-attenuated JEV vaccines, or dengue serotype2 virus (DENV-2). Neutralizing antibody response was measured employing a plaque reduction neutralization test at two weeks after the priming and at four weeks after the second dose of the dengue tetravalent plasmids. Inactivated or live-attenuated JEV vaccines, or DENV-2 induced low levels of neutralizing antibodies against the homologous viruses (JE and dengue virus, respectively). DENV-2 injection induced also low levels of cross-reactive antibodies against DENV-1, -3 and -4. JEV vaccines have no effect on the dengue-specific neutralizing antibody responses to the subsequent TDNA immunization. Pre-exposure to DENV-2 infection increased DENV-2 specific response neutralizing antibody to two doses of TDNA plasmids by six folds, but did not affect antibody response to other serotypes. Priming with JEV vaccines did not impact on dengue virus-specific neutralizing antibody response to a dengue TDNA vaccine candidate in mice.

  13. Linear induction of DNA double-strand breakage with X-ray dose, as determined from DNA fragment size distribution

    Energy Technology Data Exchange (ETDEWEB)

    Erixon, K.; Cedervall, B. [Karolinksa Institutet, Stockholm (Sweden)

    1995-05-01

    Pulsed-field gel electrophoresis has been applied to separate DNA from mouse L1210 cells exposed to X-ray doses of 1 to 50 Gy. Simultaneous separation of marker chromosomes in the range 0.1 to 12.6 Mbp allowed calculation of the size distribution of the radiation-induced fragments. The distribution was consistent with a random induction of double-strand breaks (DSBs). A theoretical relationship between the size distribution of such fragments and the average number of induced breaks was used to calculate the yield and dose response. The DNA distribution was determined by both radiolabeling and fluorescence staining. Two independent methods were use to evaluate the radiation-induced yield of DSBs, both assuming that all DNA is broken at random. In the first method we compared the theoretical and experimental fraction of DNA that is below a given size limit. By this method we estimated the yield to be 0.006-0.007 DSB/GY per million base pairs using the radiolabel and 0.004-0.008 DSB/Gy per million base pairs by fluorescence staining. The dose response was linear in both cases. In the second method we looked only at the size distribution in the resolving part of the gel and compared it to the theoretical distribution. By this method a value of approximately 0.012 DSB/Gy/Mb was found, using fluorescence as a measure of DNA distribution. In a normal diploid mammalian genome of size 60000 Mbp, this is equivalent to a yield of 25-50 DSBs/Gy or 70 DSBs/GY, respectively. The second approach, which looks only at the smaller fragments, may overestimate the yield, while the first approach suffers from uncertainties about the fraction of DNA irreversibly trapped in the well. The assay has the capacity to detect a dose of less than 1 Gy. 58 refs., 10 figs.

  14. Prime-boost bacillus Calmette-Guérin vaccination with lentivirus-vectored and DNA-based vaccines expressing antigens Ag85B and Rv3425 improves protective efficacy against Mycobacterium tuberculosis in mice.

    Science.gov (United States)

    Xu, Ying; Yang, Enzhuo; Wang, Jianguang; Li, Rui; Li, Guanghua; Liu, Guoyuan; Song, Na; Huang, Qi; Kong, Cong; Wang, Honghai

    2014-10-01

    To prevent the global spread of tuberculosis (TB), more effective vaccines and vaccination strategies are urgently needed. As a result of the success of bacillus Calmette-Guérin (BCG) in protecting children against miliary and meningeal TB, the majority of individuals will have been vaccinated with BCG; hence, boosting BCG-primed immunity will probably be a key component of future vaccine strategies. In this study, we compared the ability of DNA-, protein- and lentiviral vector-based vaccines that express the antigens Ag85B and Rv3425 to boost the effects of BCG in the context of immunity and protection against Mycobacterium tuberculosis in C57BL/6 mice. Our results demonstrated that prime-boost BCG vaccination with a lentiviral vector expressing the antigens Ag85B and Rv3425 significantly enhanced immune responses, including T helper type 1 and CD8(+) cytotoxic T lymphocyte responses, compared with DNA- and protein-based vaccines. However, lentivirus-vectored and DNA-based vaccines greatly improved the protective efficacy of BCG against M. tuberculosis, as indicated by a lack of weight loss and significantly reduced bacterial loads and histological damage in the lung. Our study suggests that the use of lentiviral or DNA vaccines containing the antigens Ag85B and Rv3425 to boost BCG is a good choice for the rational design of an efficient vaccination strategy against TB.

  15. Protection of Guinea Pigs against Leptospira interrogans Serovar Lai by LipL21 DNA Vaccine

    Institute of Scientific and Technical Information of China (English)

    Hanjiang He; Wenyu Wang; Zhongdao Wu; Zhiyue Lv; Jun Li; Lizhi Tan

    2008-01-01

    In this study,the full lipL21 gene fragment encoding outer membrane protein LipL21 was cloned from L. Interrogans serovar Lai and inserted into eukaryotic expression vector pcDNA3.1(+).The guinea pigs were immunized with pcDNA3.1(+)-lipL21,pcDNA3.1(+) or PBS.Six weeks after the second immunization,the splenocytes were isolated to detect their proliferative ability by lymphocyte transformation experiments.In addition,microscopic agglutination test was used for quantitative detection of specific antibodies.The rest guinea pigs were challenged intraperitoneally with L.interogans sorevar Lai.Then,protective effect was evaluated on the basis of survival and histopathological lesions in the kidneys,lungs,and liver.The lipL21 gene Was successfully expressed in COS-7 cells through recombinant pcDNA3.1(+)-lipL21.The titer of specific antibodies substantially increased,and the stimulation index of splenocytes increased significantly.Hence,the pcDNA3.1(+)-lipL21 could protect the immunized guinea pigs from homotypic Leptospira infection.Furthermore,no obvious pathologic changes were observed in the pcDNA3.1(+)-lipL21 immunized guinea pigs.The results showed that the protective effect with pathogenic strains of Leptospira was shared by LipL21 mediated through a plasmid vector. Consequently,these results indicated that the lipL21 DNA vaccine Was a promising candidate for the prevention of leptospirosis.

  16. DNA Prime-Boost Vaccine Regimen To Increase Breadth, Magnitude, and Cytotoxicity of the Cellular Immune Responses to Subdominant Gag Epitopes of Simian Immunodeficiency Virus and HIV.

    Science.gov (United States)

    Hu, Xintao; Valentin, Antonio; Dayton, Frances; Kulkarni, Viraj; Alicea, Candido; Rosati, Margherita; Chowdhury, Bhabadeb; Gautam, Rajeev; Broderick, Kate E; Sardesai, Niranjan Y; Martin, Malcolm A; Mullins, James I; Pavlakis, George N; Felber, Barbara K

    2016-11-15

    HIV sequence diversity and the propensity of eliciting immunodominant responses targeting variable regions of the HIV proteome are hurdles in the development of an effective AIDS vaccine. An HIV-derived conserved element (CE) p24(gag) plasmid DNA (pDNA) vaccine is able to redirect immunodominant responses to otherwise subdominant and often more vulnerable viral targets. By homology to the HIV immunogen, seven CE were identified in SIV p27(Gag) Analysis of 31 rhesus macaques vaccinated with full-length SIV gag pDNA showed inefficient induction (58% response rate) of cellular responses targeting these CE. In contrast, all 14 macaques immunized with SIV p27CE pDNA developed robust T cell responses recognizing CE. Vaccination with p27CE pDNA was also critical for the efficient induction and increased the frequency of Ag-specific T cells with cytotoxic potential (granzyme B(+) CD107a(+)) targeting subdominant CE epitopes, compared with the responses elicited by the p57(gag) pDNA vaccine. Following p27CE pDNA priming, two booster regimens, gag pDNA or codelivery of p27CE+gag pDNA, significantly increased the levels of CE-specific T cells. However, the CE+gag pDNA booster vaccination elicited significantly broader CE epitope recognition, and thus, a more profound alteration of the immunodominance hierarchy. Vaccination with HIV molecules showed that CE+gag pDNA booster regimen further expanded the breadth of HIV CE responses. Hence, SIV/HIV vaccine regimens comprising CE pDNA prime and CE+gag pDNA booster vaccination significantly increased cytotoxic T cell responses to subdominant highly conserved Gag epitopes and maximized response breadth. Copyright © 2016 by The American Association of Immunologists, Inc.

  17. Formulation, characterization, and expression of a recombinant MOMP Chlamydia trachomatis DNA vaccine encapsulated in chitosan nanoparticles

    Directory of Open Access Journals (Sweden)

    Cambridge CD

    2013-05-01

    Full Text Available Chino D Cambridge, Shree R Singh, Alain B Waffo, Stacie J Fairley, Vida A DennisCenter for NanoBiotechnology Research (CNBR, Alabama State University, Montgomery, AL, USAAbstract: Chlamydia trachomatis is a bacterial sexually transmitted infection affecting millions of people worldwide. Previous vaccination attempts have employed the recombinant major outer membrane protein (MOMP of C. trachomatis nonetheless, with limited success, perhaps, due to stability, degradation, and delivery issues. In this study we cloned C. trachomatis recombinant MOMP DNA (DMOMP and encapsulated it in chitosan nanoparticles (DMCNP using the complex coacervation technique. Physiochemical characterizations of DMCNP included transmission and scanning electron microcopy, Fourier transform infrared and ultraviolet-visible spectroscopy, and zeta potential. Encapsulated DMOMP was 167–250 nm, with a uniform spherical shape and homogenous morphology, and an encapsulation efficiency > 90%. A slow release pattern of encapsulated DMOMP, especially in acidic solution, was observed over 7 days. The zeta potential of DMCNP was ~8.80 mV, which indicated that it was highly stable. Toxicity studies of DMCNP (25–400 µg/mL to Cos-7 cells using the MTT assay revealed minimal toxicity over 24–72 hours with >90% viable cells. Ultra-violet visible (UV-vis spectra indicated encapsulated DMOMP protection by chitosan, whereas agarose gel electrophoresis verified its protection from enzymatic degradation. Expression of MOMP protein in DMCNP-transfected Cos-7 cells was demonstrated via Western blotting and immunofluorescence microscopy. Significantly, intramuscular injection of BALB/c mice with DMCNP confirmed the delivery of encapsulated DMOMP, and expression of the MOMP gene transcript in thigh muscles and spleens. Our data show that encapsulation of DMOMP in biodegradable chitosan nanoparticles imparts stability and protection from enzymatic digestion, and enhances delivery and

  18. Effect of Different Human Papillomavirus Serological and DNA Criteria on Vaccine Efficacy Estimates

    Science.gov (United States)

    Lang Kuhs, Krystle A.; Porras, Carolina; Schiller, John T.; Rodriguez, Ana Cecilia; Schiffman, Mark; Gonzalez, Paula; Wacholder, Sholom; Ghosh, Arpita; Li, Yan; Lowy, Douglas R.; Kreimer, Aimée R.; Poncelet, Sylviane; Schussler, John; Quint, Wim; van Doorn, Leen-Jan; Sherman, Mark E.; Sidawy, Mary; Herrero, Rolando; Hildesheim, Allan; Safaeian, Mahboobeh; Lang Kuhs, Krystle A.; Schiller, John T.; Schiffman, Mark; Wacholder, Sholom; Lowy, Douglas R.; Kreimer, Aimée R.; Sherman, Mark E.; Hildesheim, Allan; Safaeian, Mahboobeh; Porras, Carolina; Rodriguez, Ana Cecilia; Gonzalez, Paula; Herrero, Rolando; Gonzalez, Paula; Herrero, Rolando; Ghosh, Arpita; Li, Yan; Poncelet, Sylviane; Schussler, John; Quint, Wim; van Doorn, Leen-Jan; Sidawy, Mary; Self, Steve; Benavides, Adriana; Calzada, Luis Diego; Karron, Ruth; Nayar, Ritu; Roach, Nancy; Cain, Joanna; Davey, Diane; DeMets, David; Fuster, Francisco; Gershon, Ann; Holly, Elizabeth; Raventós, Henriette; Rida, Wasima; Rosero-Bixby, Luis; Suthers, Kristen; Lara, Silvia; Thomas, Sarah; Alfaro, Mario; Barrantes, Manuel; Concepción Bratti, M.; Cárdenas, Fernando; Cortés, Bernal; Espinoza, Albert; Estrada, Yenory; González, Paula; Guillén, Diego; Herrero, Roland; Jiménez, Silvia E.; Morales, Jorge; Villegas, Luis; Morera, Lidia Ana; Pérez, Elmer; Porras, Carolina; Rodríguez, Ana Cecilia; Rivas, Libia; Freer, Enrique; Bonilla, José; García-Piñeres, Alfanso; Silva, Sandra; Atmella, Ivannia; Ramírez, Margarita; Hildesheim, Allan; Kreimer, Aimée R.; Lowy, Douglas R.; Macklin, Nora; Schiffman, Mark; Schiller, John T.; Sherman, Mark; Solomon, Diane; Wacholder, Sholom; Pinto, Ligia; Kemp, Troy; Eklund, Claire; Hutchinson, Martha; Sidawy, Mary; Quint, Wim; van Doorn, Leen-Jan

    2014-01-01

    Two trials of clinically approved human papillomavirus (HPV) vaccines, Females United to Unilaterally Reduce Endo/Ectocervical Disease (FUTURE I/II) and the Papilloma Trial Against Cancer in Young Adults (PATRICIA), reported a 22% difference in vaccine efficacy (VE) against cervical intraepithelial neoplasia grade 2 or worse in HPV-naïve subcohorts; however, serological testing methods and the HPV DNA criteria used to define HPV-unexposed women differed between the studies. We applied previously described methods to simulate these HPV-naïve subcohorts within the Costa Rica HPV16/18 Vaccine Trial and assessed how these criteria affect the estimation of VE. We applied 2 enzyme-linked immunosorbent assay (ELISA) thresholds for HPV16 and HPV18 seropositivity (8 and 7 ELISA units/mL, respectively, for PATRICIA; 54 and 65 ELISA units/mL, respectively, for FUTURE I/II (to approximate the competitive Luminex immunoassay)) and 2 criteria for HPV DNA positivity (12 oncogenic HPV types, plus HPV66 and 68/73 for PATRICIA; or plus HPV6 and 11 for FUTURE I/II). VE was computed in the 2 naïve subcohorts. Using the FUTURE I/II and PATRICIA criteria, VE estimates against cervical intraepithelial neoplasia grade 2 or worse, regardless of HPV type, were 69.0% (95% confidence interval: 40.3%, 84.9%) and 80.8% (95% confidence interval: 52.6%, 93.5%), respectively (P = 0.1). Although the application of FUTURE I/II criteria to our cohort resulted in the inclusion of more sexually experienced women, methodological differences did not fully explain the VE differences. PMID:25139208

  19. DNA Vaccines delivered by dermal electroporation elicit durable protective immunity against individual or simultaneous infections with lassa and ebola viruses in guinea pigs.

    Science.gov (United States)

    2017-08-22

    DNA vaccines elicit durable protective immunity against individual or simultaneous 1  infections with Lassa and Ebola viruses in guinea pigs 2  3...previously developed optimized DNA vaccines against both Lassa fever and Ebola 15  hemorrhagic fever viruses and demonstrated that they were protective...individually in 16  guinea pig and nonhuman primate models. In this study, we vaccinated groups of strain 17  13 guinea pigs two times, four weeks apart

  20. Phase IIb trial of in vivo electroporation mediated dual-plasmid hepatitis B virus DNA vaccine in chronic hepatitis B patients under lamivudine therapy

    Science.gov (United States)

    Yang, Fu-Qiang; Rao, Gui-Rong; Wang, Gui-Qiang; Li, Yue-Qi; Xie, Yao; Zhang, Zhan-Qing; Deng, Cun-Liang; Mao, Qing; Li, Jun; Zhao, Wei; Wang, Mao-Rong; Han, Tao; Chen, Shi-Jun; Pan, Chen; Tan, De-Ming; Shang, Jia; Zhang, Ming-Xiang; Zhang, Yue-Xin; Yang, Ji-Ming; Chen, Guang-Ming

    2017-01-01

    AIM To assess the efficacy and safety of in vivo electroporation (EP)-mediated dual-plasmid hepatitis B virus (HBV) DNA vaccine vs placebo for sequential combination therapy with lamivudine (LAM) in patients with chronic hepatitis B. METHODS Two hundred and twenty-five patients were randomized to receive either LAM + vaccine (vaccine group, n = 109) or LAM + placebo (control group, n = 116). LAM treatment lasted 72 wk. Patients received the DNA vaccine or placebo by intramuscular injection mediated by EP at weeks 12 (start of treatment with vaccine or placebo, SOT), 16, 24, and 36 (end of treatment with vaccine or placebo, EOT). RESULTS In the modified intent-to-treat population, more patients had a decrease in HBV DNA > 2 log10 IU/mL in the vaccine group at week 12 after EOT compared with the control group. A trend toward a difference in the number of patients with undetectable HBV DNA at week 28 after EOT was obtained. Adverse events were similar. In the dynamic per-protocol set, which excluded adefovir (ADV) add-on cases at each time point instantly after ADV administration due to LAM antiviral failure, more patients had a decrease in HBV DNA > 2 log10 IU/mL in the vaccine group at week 12 and 28 after EOT compared with the control group. More patients with undetectable HBV DNA at week 28 after EOT in the vaccine group were also observed. Among patients with a viral load < 1000 copies/mL at week 12, more patients achieved HBeAg seroconversion in the vaccine group than among controls at week 36 after EOT, as well as less virological breakthrough and YMDD mutations. CONCLUSION The primary endpoint was not achieved using the HBV DNA vaccine. The HBV DNA vaccine could only be beneficial in subjects that have achieved initial virological response under LAM chemotherapy. PMID:28127204

  1. A HIV-Tat/C4-binding protein chimera encoded by a DNA vaccine is highly immunogenic and contains acute EcoHIV infection in mice

    Science.gov (United States)

    Tomusange, Khamis; Wijesundara, Danushka; Gummow, Jason; Garrod, Tamsin; Li, Yanrui; Gray, Lachlan; Churchill, Melissa; Grubor-Bauk, Branka; Gowans, Eric J.

    2016-01-01

    DNA vaccines are cost-effective to manufacture on a global scale and Tat-based DNA vaccines have yielded protective outcomes in preclinical and clinical models of human immunodeficiency virus (HIV), highlighting the potential of such vaccines. However, Tat-based DNA vaccines have been poorly immunogenic, and despite the administration of multiple doses and/or the addition of adjuvants, these vaccines are not in general use. In this study, we improved Tat immunogenicity by fusing it with the oligomerisation domain of a chimeric C4-binding protein (C4b-p), termed IMX313, resulting in Tat heptamerisation and linked Tat to the leader sequence of tissue plasminogen activator (TPA) to ensure that the bulk of heptamerised Tat is secreted. Mice vaccinated with secreted Tat fused to IMX313 (pVAX-sTat-IMX313) developed higher titres of Tat-specific serum IgG, mucosal sIgA and cell-mediated immune (CMI) responses, and showed superior control of EcoHIV infection, a surrogate murine HIV challenge model, compared with animals vaccinated with other test vaccines. Given the crucial contribution of Tat to HIV-1 pathogenesis and the precedent of Tat-based DNA vaccines in conferring some level of protection in animal models, we believe that the virologic control demonstrated with this novel multimerised Tat vaccine highlights the promise of this vaccine candidate for humans. PMID:27358023

  2. Isolation and Characterization of Linear DNA Elements from the Mitochondria of Gaeumannomyces graminis†

    OpenAIRE

    Honeyman, Allen L.; Currier, Thomas C.

    1986-01-01

    Different Gaeumannomyces graminis strains of diverse geographic origin contain one or two small DNAs ranging in size from 7.2 to 10 kilobases. These DNAs exhibit different degrees of homology with each other. We have characterized these low-molecular-weight DNAs from one strain, Ha-01. These small DNAs, E1 and E2, are mitochondrial in origin and were isolated as linear molecules which exhibited an intrinsic difference in density from the high-molecular-weight DNA.

  3. Isolation and Characterization of Linear DNA Elements from the Mitochondria of Gaeumannomyces graminis.

    Science.gov (United States)

    Honeyman, A L; Currier, T C

    1986-10-01

    Different Gaeumannomyces graminis strains of diverse geographic origin contain one or two small DNAs ranging in size from 7.2 to 10 kilobases. These DNAs exhibit different degrees of homology with each other. We have characterized these low-molecular-weight DNAs from one strain, Ha-01. These small DNAs, E1 and E2, are mitochondrial in origin and were isolated as linear molecules which exhibited an intrinsic difference in density from the high-molecular-weight DNA.

  4. DNA vaccination with a gene encoding Toxoplasma gondii Rhoptry Protein 17 induces partial protective immunity against lethal challenge in mice

    Directory of Open Access Journals (Sweden)

    Wang Hai-Long

    2016-01-01

    Full Text Available Toxoplasma gondii is an obligate intracellular apicomplexan parasite that affects humans and various vertebrate livestock and causes serious economic losses. To develop an effective vaccine against T. gondii infection, we constructed a DNA vaccine encoding the T. gondii rhoptry protein 17 (TgROP17 and evaluated its immune protective efficacy against acute T. gondii infection in mice. The DNA vaccine (p3×Flag-CMV-14-ROP17 was intramuscularly injected to BALB/c mice and the immune responses of the vaccinated mice were determined. Compared to control mice treated with empty vector or PBS, mice immunized with the ROP17 vaccine showed a relatively high level of specific anti-T. gondii antibodies, and a mixed IgG1/IgG2a response with predominance of IgG2a production. The immunized mice also displayed a specific lymphocyte proliferative response, a Th1-type cellular immune response with production of IFN-γ and interleukin-2, and increased number of CD8+ T cells. Immunization with the ROP17 DNA significantly prolonged the survival time (15.6 ± 5.4 days, P < 0.05 of mice after challenge infection with the virulent T. gondii RH strain (Type I, compared with the control groups which died within 8 days. Therefore, our data suggest that DNA vaccination with TgROP17 triggers significant humoral and cellular responses and induces effective protection in mice against acute T. gondii infection, indicating that TgROP17 is a promising vaccine candidate against acute toxoplasmosis.

  5. DNA vaccine-derived human IgG produced in transchromosomal bovines protect in lethal models of hantavirus pulmonary syndrome.

    Science.gov (United States)

    Hooper, Jay W; Brocato, Rebecca L; Kwilas, Steven A; Hammerbeck, Christopher D; Josleyn, Matthew D; Royals, Michael; Ballantyne, John; Wu, Hua; Jiao, Jin-an; Matsushita, Hiroaki; Sullivan, Eddie J

    2014-11-26

    Polyclonal immunoglobulin-based medical products have been used successfully to treat diseases caused by viruses for more than a century. We demonstrate the use of DNA vaccine technology and transchromosomal bovines (TcBs) to produce fully human polyclonal immunoglobulins (IgG) with potent antiviral neutralizing activity. Specifically, two hantavirus DNA vaccines [Andes virus (ANDV) DNA vaccine and Sin Nombre virus (SNV) DNA vaccine] were used to produce a candidate immunoglobulin product for the prevention and treatment of hantavirus pulmonary syndrome (HPS). A needle-free jet injection device was used to vaccinate TcB, and high-titer neutralizing antibodies (titers >1000) against both viruses were produced within 1 month. Plasma collected at day 10 after the fourth vaccination was used to produce purified α-HPS TcB human IgG. Treatment with 20,000 neutralizing antibody units (NAU)/kg starting 5 days after challenge with ANDV protected seven of eight animals, whereas zero of eight animals treated with the same dose of normal TcB human IgG survived. Likewise, treatment with 20,000 NAU/kg starting 5 days after challenge with SNV protected immunocompromised hamsters from lethal HPS, protecting five of eight animals. Our findings that the α-HPS TcB human IgG is capable of protecting in animal models of lethal HPS when administered after exposure provides proof of concept that this approach can be used to develop candidate next-generation polyclonal immunoglobulin-based medical products without the need for human donors, despeciation protocols, or inactivated/attenuated vaccine antigen. Copyright © 2014, American Association for the Advancement of Science.

  6. [Serologic response to a DNA recombinant vaccine against hepatitis B in natives of the Peruvian Amazonian jungle].

    Science.gov (United States)

    Colichón, A; Vildósola, H; Sjogren, M; Cantella, R; Rojas, C

    1990-01-01

    Large areas of the Amazon basin in Brazil, Colombia, Ecuador, and in the nonoriental region of the peruvian jungle have been found to be hyperendemic to Hepatitis B with high prevalence of asymptomatic carriers (11 to 25%) and, in more selected areas, Hepatitis Delta has been also reported. In the present report, we have studied 108 volunteers from six different Jivaroes communities living in a hyperendemic Hepatitis B area. They received 2 doses of DNA recombinant yeast derivated HBV vaccine. All the selected persons were HBsAb negatives, but many (80%) had antibodies to HBc. Following immunization schedule, 80% responded with the formation of HBsAb; a better seroconversion was achieved in those negatives to anticore IgG compared with those having HBcAb. We obtained 90% of seroconversion in spite of the fact that our vaccination schedule was prolonged up to 10 months from the one recommended by the manufacturer. The vaccination schedule 0,4, 14 months, and the schedule 0,4 months, had 76 and 29% of seroconversion, respectively. We want to point out three observations: 1) It is quite possible that many of the Anti-core positives, that did not respond to vaccination were carriers of HBsAg undetectable by the conventional EIA test carried out; 2) The seroconversion rate in these natives was low (up to six months after the vaccination schedule); and 3) Many of the HBcAb were false positives and many of them were recently infected. We conclude: A) It is highly important to assess the anti-HBs hyperendemic areas before attempting vaccinations; B) All persons negative to anti-HBs should be vaccinated in spite to anticore antibodies; C) Areas with difficult access could be vaccinated even until 10 months without affecting good results, and D) DNA recombinant vaccine (ENGERIX B) was well tolerated. No side effects were observed.

  7. Increasing a Robust Antigen-Specific Cytotoxic T Lymphocyte Response by FMDV DNA Vaccination with IL-9 Expressing Construct

    Directory of Open Access Journals (Sweden)

    Qiang Zou

    2010-01-01

    Full Text Available Various chemokines and cytokines as adjuvants can be used to improve efficacy of DNA vaccination. In this study, we sought to investigate if a DNA construct expressing IL-9 (designed as proV-IL9 as a molecular adjuvant enhance antigen specific immune responses elicited by the pcD-VP1 DNA vaccination. Mice immunized with pcD-VP1 combined with proV-IL9 developed a strong humoral response. In addition, the coinoculation induced significant higher level of antigen-specific cell proliferation and cytotoxic response. This agreed well with higher expression level of IFN-γ and perforin in CD8+ T cells, but not with IL-17 in these T cells. The results indicate that IL-9 induces the development of IFN-γ-producing CD8+ T cells (Tc1, but not the IL-17-producing CD8+ T cells (Tc17. Up-regulated expressions of BCL-2 and BCL-XL were exhibited in these Tc1 cells, suggesting that IL-9 may trigger antiapoptosis mechanism in these cells. Together, these results demonstrated that IL-9 used as molecular adjuvant could enhance the immunogenicity of DNA vaccination, in augmenting humoral and cellular responses and particularly promoting Tc1 activations. Thus, the IL-9 may be utilized as a potent Tc1 adjuvant for DNA vaccines.

  8. Antitumor immunopreventive effect in mice induced by DNA vaccine encoding a fusion protein of α-fetoprotein and CTLA4

    Institute of Scientific and Technical Information of China (English)

    Geng Tian; Ji-Lin Yi; Ping Xiong

    2004-01-01

    AIM: To develop a tumor DNA vaccine encoding a fusion protein of murine AFP and CTLA4, and to study its ability to induce specific CTL response and its protective effect against AFP-producing tumor.METHODS: Murine α-fetoprotein (mAFP) gene was cloned from total RNA of Hepa1-6 cells by RT-PCR. A DNA vaccine was constructed by fusion murine α-fetoprotein gene and extramembrane domain of murine CTLA4 gene. The DNA vaccine was identified by restriction enzyme analysis,sequencing and expression. EL-4 (mAFP) was developed by stable transfection of EL-4 cells with pmAFP. The frequency of cells produdng IFN-γ in splenocytes harvested from the immunized mice was measured by ELISPOT. Mice immunized with DNA vaccine were inoculated with EL-4 (mAFP) cells in back to observe the protective effect of immunization on tumor. On the other hand, blood samples were collected from the immunized mice to check the functions of liver and kidney.RESULTS: 1.8 kb mAFP cDNA was cloned from total RNA of Hepa1-6 cells by RT-PCR. The DNA vaccine encoding a fusion protein of mAFP-CTLA4 was constructed and confirmed by restriction enzyme analysis, sequencing and expression. The expression of mAFP mRNA in EL-4 (mAFP) was confirmed by RT-PCR. The ELISPOT results showed that the number of IFN-γ-producing cells in pmAFP-CTLA4 group was significantly higher than that in pmAFP, pcDNA3.1 and PBS group. The tumor volume in pmAFP-CTLA4 group was significantly smaller than that in pmAFP, pcDNA3.1 and PBS group, respectively. The hepatic and kidney functions in each group were not altered.CONCLUSION: AFP-CTLA4 DNA vaccine can stimulate potent specific CTL responses and has distinctive antitumor effect on AFP-producing tumor. The vaccine has no impact on the function of mouse liver and kidney.

  9. Intravaginal HPV DNA vaccination with electroporation induces local CD8+ T-cell immune responses and antitumor effects against cervicovaginal tumors.

    Science.gov (United States)

    Sun, Y; Peng, S; Qiu, J; Miao, J; Yang, B; Jeang, J; Hung, C-F; Wu, T-C

    2015-07-01

    Therapeutic human papillomavirus (HPV) vaccines have the potential to inhibit the progression of an established HPV infection to precancer and cancer lesions by targeting HPV oncoproteins. We have previously developed a therapeutic DNA vaccine encoding calreticulin (CRT) linked to E7, CRT/E7 DNA vaccine, for use in the treatment of HPV-associated lesions. Since the transfection efficiency of DNA vaccines administered in vivo is typically low, we examined the use of electroporation as well as different routes of administration to enhance antigen-specific tumor control. We tested the effects of the CRT/E7 DNA vaccine administered intramuscularly or intravaginally, with or without electroporation, on the generation of CD8+ T-cell immunity and therapeutic antitumor effects in HPV16 E7-expressing cervicovaginal tumor-bearing mice. We found that intravaginal vaccination of CRT/E7 DNA followed by electroporation-induced potent E7-specific CD8(+) T-cell responses in the cervicovaginal tract, compared with intramuscular injection followed by electroporation. Furthermore, tumor-bearing mice vaccinated intravaginally followed by electroporation had an enhanced survival, antitumor effects and local production of IFN-γ+CD8+ T cells compared with those vaccinated intramuscularly with electroporation. Thus, we show that intravaginal CRT/E7 DNA vaccination followed by electroporation generates the most potent therapeutic antitumor effects against an orthotopic E7-expressing tumor model. The current study will have significant clinical implications once a clinically applicable electroporation device for intravaginal use becomes available.

  10. Construction and Characterization of an in-vivo Linear Covalently Closed DNA Vector Production System

    Science.gov (United States)

    2012-01-01

    Background While safer than their viral counterparts, conventional non-viral gene delivery DNA vectors offer a limited safety profile. They often result in the delivery of unwanted prokaryotic sequences, antibiotic resistance genes, and the bacterial origins of replication to the target, which may lead to the stimulation of unwanted immunological responses due to their chimeric DNA composition. Such vectors may also impart the potential for chromosomal integration, thus potentiating oncogenesis. We sought to engineer an in vivo system for the quick and simple production of safer DNA vector alternatives that were devoid of non-transgene bacterial sequences and would lethally disrupt the host chromosome in the event of an unwanted vector integration event. Results We constructed a parent eukaryotic expression vector possessing a specialized manufactured multi-target site called “Super Sequence”, and engineered E. coli cells (R-cell) that conditionally produce phage-derived recombinase Tel (PY54), TelN (N15), or Cre (P1). Passage of the parent plasmid vector through R-cells under optimized conditions, resulted in rapid, efficient, and one step in vivo generation of mini lcc—linear covalently closed (Tel/TelN-cell), or mini ccc—circular covalently closed (Cre-cell), DNA constructs, separated from the backbone plasmid DNA. Site-specific integration of lcc plasmids into the host chromosome resulted in chromosomal disruption and 105 fold lower viability than that seen with the ccc counterpart. Conclusion We offer a high efficiency mini DNA vector production system that confers simple, rapid and scalable in vivo production of mini lcc DNA vectors that possess all the benefits of “minicircle” DNA vectors and virtually eliminate the potential for undesirable vector integration events. PMID:23216697

  11. DNA疫苗与DC疫苗抗肿瘤作用的研究进展%Development of antitumor DNA vaccine and DC vaccine

    Institute of Scientific and Technical Information of China (English)

    何耀明; 葛海燕

    2010-01-01

    Tumor is one of the most serious problems threatening people's life in the 21 st century. Antitumor vaccine becomes a hot research spot of tumor therapy for low toxicity, specificity and durability. Efficient recognization and presentation of tumor antigen contribute to the foundation of an powerful anti-tumor immunologic response. Along with the deepen apprehension of the immunity mechanism and high development of the biochemical technology, a great number of new vaccines emerge and show us some favors. And DNA vaccine and DC vaccine attract most sights for their advantage in antigens expression and presentation.They become the strong weapon for tumor immunity therapy. This is a review about the mechanism, development and current problem of DNA vaccine and DC vaccine.%肿瘤是21世纪医学界面临的难题之一,肿瘤疫苗以其低毒、持久和特异性强等特点成为肿瘤治疗研究的热点,有效的肿瘤抗原被识别、递呈是建立抗肿瘤免疫应答的基础,在这个基础的不断理解和深化研究下,新型肿瘤疫苗不断涌现,并展现出一定的抗肿瘤效果,其中DNA疫苗以其独特的肿瘤抗原提供方式及DC在肿瘤抗原呈递中的重要作用,成为了肿瘤免疫治疗的有力工具,本文就目前这两类疫苗作用的机制、进展及存在问题进行综述.

  12. Influence of temperature on the efficacy of homologous and heterologous DNA vaccines against viral hemorrhagic septicemia in Pacific Herring

    Science.gov (United States)

    Hart, Lucas; Lorenzen, Niels; Einer-Jensen, Katja; Purcell, Maureen; Hershberger, Paul

    2017-01-01

    Homologous and heterologous (genogroup Ia) DNA vaccines against viral hemorrhagic septicemia virus (genogroup IVa) conferred partial protection in Pacific Herring Clupea pallasii. Early protection at 2 weeks postvaccination (PV) was low and occurred only at an elevated temperature (12.6°C, 189 degree days), where the relative percent survival following viral exposure was similar for the two vaccines (IVa and Ia) and higher than that of negative controls at the same temperature. Late protection at 10 weeks PV was induced by both vaccines but was higher with the homologous vaccine at both 9.0°C and 12.6°C. Virus neutralization titers were detected among 55% of all vaccinated fish at 10 weeks PV. The results suggest that the immune response profile triggered by DNA vaccination of herring was similar to that reported for Rainbow Trout Oncorhynchus mykiss by Lorenzen and LaPatra in 2005, who found interferon responses in the early days PV and the transition to adaptive response later. However, the protective effect was far less prominent in herring, possibly reflecting different physiologies or adaptations of the two fish species.

  13. DNA-based vaccination against hepatitis B virus using dissolving microneedle arrays adjuvanted by cationic liposomes and CpG ODN.

    Science.gov (United States)

    Qiu, Yuqin; Guo, Lei; Zhang, Suohui; Xu, Bai; Gao, Yunhua; Hu, Yan; Hou, Jun; Bai, Bingke; Shen, Honghui; Mao, Panyong

    2016-09-01

    DNA vaccines are simple to produce and can generate strong cellular and humoral immune response, making them attractive vaccine candidates. However, a major shortcoming of DNA vaccines is their poor immunogenicity when administered intramuscularly. Transcutaneous immunization (TCI) via microneedles is a promising alternative delivery route to enhance the vaccination efficacy. A novel dissolving microneedle array (DMA)-based TCI system loaded with cationic liposomes encapsulated with hepatitis B DNA vaccine and adjuvant CpG ODN was developed and characterized. The pGFP expression in mouse skin using DMA was imaged over time. In vivo immunity tests in mice were performed to observe the capability of DMA to induce immune response after delivery of DNA. The results showed that pGFP could be delivered into skin by DMA and expressed in skin. Further, the amount of expressed GFP was likely to peak at day 4. The immunity tests showed that the DMA-based DNA vaccination could induce effective immune response. CpG ODN significantly improved the immune response and achieved the shift of immune type from predominate Th2 type to a balance Th1/Th2 type. The cationic liposomes could further improve the immunogenicity of DNA vaccine. In conclusion, the novel DMA-based TCI system can effectively deliver hepatitis B DNA vaccine into skin, inducing effective immune response and change the immune type by adjuvant CpG ODN.

  14. The immunogenicity of viral haemorragic septicaemia rhabdovirus (VHSV) DNA vaccines can depend on plasmid regulatory sequences.

    Science.gov (United States)

    Chico, V; Ortega-Villaizan, M; Falco, A; Tafalla, C; Perez, L; Coll, J M; Estepa, A

    2009-03-18

    A plasmid DNA encoding the viral hemorrhagic septicaemia virus (VHSV)-G glycoprotein under the control of 5' sequences (enhancer/promoter sequence plus both non-coding 1st exon and 1st intron sequences) from carp beta-actin gene (pAE6-G(VHSV)) was compared to the vaccine plasmid usually described the gene expression is regulated by the human cytomegalovirus (CMV) immediate-early promoter (pMCV1.4-G(VHSV)). We observed that these two plasmids produced a markedly different profile in the level and time of expression of the encoded-antigen, and this may have a direct effect upon the intensity and suitability of the in vivo immune response. Thus, fish genetic immunisation assays were carried out to study the immune response of both plasmids. A significantly enhanced specific-antibody response against the viral glycoprotein was found in the fish immunised with pAE6-G(VHSV). However, the protective efficacy against VHSV challenge conferred by both plasmids was similar. Later analysis of the transcription profile of a set of representative immune-related genes in the DNA immunized fish suggested that depending on the plasmid-related regulatory sequences controlling its expression, the plasmid might activate distinct patterns of the immune system. All together, the results from this study mainly point out that the selection of a determinate encoded-antigen/vector combination for genetic immunisation is of extraordinary importance in designing optimised DNA vaccines that, when required for inducing protective immune response, could elicit responses biased to antigen-specific antibodies or cytotoxic T cells generation.

  15. Vaccination with Bivalent DNA Vaccine of α1-Giardin and CWP2 Delivered by Attenuated Salmonella typhimurium Reduces Trophozoites and Cysts in the Feces of Mice Infected with Giardia lamblia.

    Directory of Open Access Journals (Sweden)

    Xian-Min Feng

    Full Text Available Giardia lamblia is one of the most common infectious protozoans in human that may cause diarrhea in travelers. Searching for antigens that induced effectively protective immunity has become a key point in the development of vaccine against giardiasis.Mice vaccinated with G. lamblia trophozozite-specific α1-giardin DNA vaccine delivered orally by attenuated Salmonella typhimurium SL7027 elicited 74.2% trophozoite reduction, but only 28% reduction in cyst shedding compared with PBS buffer control. Oral vaccination with Salmonella-delivered cyst-specific CWP2 DNA produced 89% reduction in cysts shedding in feces of vaccinated mice. Significantly, the mice vaccinated with Salmonella-delivered bivalent α1-giardin and CWP2 DNA vaccines produced significant reduction in both trophozoite (79% and cyst (93% in feces of vaccinated mice. This parasite reduction is associated with the strong local mucosal IgA secretion and the IgG2a-dominant systemic immune responses in vaccinated mice.The results demonstrate that bivalent vaccines targeting α1-giardin and CWP2 can protect mice against the colonization of Giardia trophozoite and block the transformation of cyst in host at the same time, and can be used to prevent Giardia infection and block the transmission of giardiasis.

  16. Vaccination with Bivalent DNA Vaccine of α1-Giardin and CWP2 Delivered by Attenuated Salmonella typhimurium Reduces Trophozoites and Cysts in the Feces of Mice Infected with Giardia lamblia.

    Science.gov (United States)

    Feng, Xian-Min; Zheng, Wen-Yu; Zhang, Hong-Mei; Shi, Wen-Yan; Li, Yao; Cui, Bai-Ji; Wang, Hui-Yan

    2016-01-01

    Giardia lamblia is one of the most common infectious protozoans in human that may cause diarrhea in travelers. Searching for antigens that induced effectively protective immunity has become a key point in the development of vaccine against giardiasis. Mice vaccinated with G. lamblia trophozozite-specific α1-giardin DNA vaccine delivered orally by attenuated Salmonella typhimurium SL7027 elicited 74.2% trophozoite reduction, but only 28% reduction in cyst shedding compared with PBS buffer control. Oral vaccination with Salmonella-delivered cyst-specific CWP2 DNA produced 89% reduction in cysts shedding in feces of vaccinated mice. Significantly, the mice vaccinated with Salmonella-delivered bivalent α1-giardin and CWP2 DNA vaccines produced significant reduction in both trophozoite (79%) and cyst (93%) in feces of vaccinated mice. This parasite reduction is associated with the strong local mucosal IgA secretion and the IgG2a-dominant systemic immune responses in vaccinated mice. The results demonstrate that bivalent vaccines targeting α1-giardin and CWP2 can protect mice against the colonization of Giardia trophozoite and block the transformation of cyst in host at the same time, and can be used to prevent Giardia infection and block the transmission of giardiasis.

  17. Physical Characterization of Gemini Surfactant-Based Synthetic Vectors for the Delivery of Linear Covalently Closed (LCC DNA Ministrings.

    Directory of Open Access Journals (Sweden)

    Chi Hong Sum

    Full Text Available In combination with novel linear covalently closed (LCC DNA minivectors, referred to as DNA ministrings, a gemini surfactant-based synthetic vector for gene delivery has been shown to exhibit enhanced delivery and bioavailability while offering a heightened safety profile. Due to topological differences from conventional circular covalently closed (CCC plasmid DNA vectors, the linear topology of LCC DNA ministrings may present differences with regards to DNA interaction and the physicochemical properties influencing DNA-surfactant interactions in the formulation of lipoplexed particles. In this study, N,N-bis(dimethylhexadecyl-α,ω-propanediammonium(16-3-16gemini-based synthetic vectors, incorporating either CCC plasmid or LCC DNA ministrings, were characterized and compared with respect to particle size, zeta potential, DNA encapsulation, DNase sensitivity, and in vitro transgene delivery efficacy. Through comparative analysis, differences between CCC plasmid DNA and LCC DNA ministrings led to variations in the physical properties of the resulting lipoplexes after complexation with 16-3-16 gemini surfactants. Despite the size disparities between the plasmid DNA vectors (CCC and DNA ministrings (LCC, differences in DNA topology resulted in the generation of lipoplexes of comparable particle sizes. The capacity for ministring (LCC derived lipoplexes to undergo complete counterion release during lipoplex formation contributed to improved DNA encapsulation, protection from DNase degradation, and in vitro transgene delivery.

  18. 1918 pandemic H1N1 DNA vaccine protects ferrets against 2007 H1N1 virus infection

    DEFF Research Database (Denmark)

    Bragstad, Karoline; Martel, Cyril Jean-Marie; Aasted, Bent

    of the H1N1 pandemic virus from 1918 induce protection in ferrets against infection with a H1N1 (A/New Caledonia/20/99(H1N1)) virus which was included in the conventional vaccine for the 2006-2007 season. The viruses are separated by a time interval of 89 years and differ by 21.2% in the HA1 protein......Influenza vaccines with the ability to induce immune responses cross-reacting with drifted virus variants would be of great advantage for vaccine development against seasonal and emerging new strains. We demonstrate that gene gun administrated DNA vaccine encoding HA and NA and/or NP and M proteins...

  19. A polyvalent influenza DNA vaccine applied by needle-free intradermal delivery induces cross-reactive humoral and cellular immune responses in pigs

    DEFF Research Database (Denmark)

    Borggren, Marie; Nielsen, Jens; Karlsson, Ingrid

    2016-01-01

    BACKGROUND: Pigs are natural hosts for influenza A viruses, and the infection is widely prevalent in swine herds throughout the world. Current commercial influenza vaccines for pigs induce a narrow immune response and are not very effective against antigenically diverse viruses. To control...... influenza in pigs, the development of more effective swine influenza vaccines inducing broader cross-protective immune responses is needed. Previously, we have shown that a polyvalent influenza DNA vaccine using vectors containing antibiotic resistance genes induced a broadly protective immune response...... of the optimized DNA vaccine were evaluated in groups of five to six pigs. The DNA vaccine consisted of six selected influenza genes of pandemic origin, including internally expressed matrix and nucleoprotein and externally expressed hemagglutinin and neuraminidase. RESULTS: Needle-free vaccination of growing pigs...

  20. DNA-based vaccines activate innate and adaptive antitumor immunity by engaging the NKG2D receptor.

    Science.gov (United States)

    Zhou, He; Luo, Yunping; Lo, Jeng-fan; Kaplan, Charles D; Mizutani, Masato; Mizutani, Noriko; Lee, Jiing-Dwan; Primus, F James; Becker, Jürgen C; Xiang, Rong; Reisfeld, Ralph A

    2005-08-02

    The interaction of NKG2D, a stimulatory receptor expressed on natural killer (NK) cells and activated CD8(+) T cells, and its ligands mediates stimulatory and costimulatory signals to these cells. Here, we demonstrate that DNA-based vaccines, encoding syngeneic or allogeneic NKG2D ligands together with tumor antigens such as survivin or carcinoembryonic antigen, markedly activate both innate and adaptive antitumor immunity. Such vaccines result in highly effective, NK- and CD8(+) T cell-mediated protection against either breast or colon carcinoma cells in prophylactic and therapeutic settings. Notably, this protection was irrespective of the NKG2D ligand expression level of the tumor cells. Hence, this strategy has the potential to lead to widely applicable and possibly clinically useful DNA-based cancer vaccines.

  1. A multiagent filovirus DNA vaccine delivered by intramuscular electroporation completely protects mice from ebola and Marburg virus challenge.

    Science.gov (United States)

    Grant-Klein, Rebecca J; Van Deusen, Nicole M; Badger, Catherine V; Hannaman, Drew; Dupuy, Lesley C; Schmaljohn, Connie S

    2012-11-01

    We evaluated the immunogenicity and protective efficacy of DNA vaccines expressing the codon-optimized envelope glycoprotein genes of Zaire ebolavirus, Sudan ebolavirus, and Marburg marburgvirus (Musoke and Ravn). Intramuscular or intradermal delivery of the vaccines in BALB/c mice was performed using the TriGrid™ electroporation device. Mice that received DNA vaccines against the individual viruses developed robust glycoprotein-specific antibody titers as determined by ELISA and survived lethal viral challenge with no display of clinical signs of infection. Survival curve analysis revealed there was a statistically significant increase in survival compared to the control groups for both the Ebola and Ravn virus challenges. These data suggest that further analysis of the immune responses generated in the mice and additional protection studies in nonhuman primates are warranted.

  2. A heterologous DNA prime-Venezuelan equine encephalitis virus replicon particle boost dengue vaccine regimen affords complete protection from virus challenge in cynomolgus macaques.

    Science.gov (United States)

    Chen, Lan; Ewing, Dan; Subramanian, Hemavathy; Block, Karla; Rayner, Jonathan; Alterson, Kimberly D; Sedegah, Martha; Hayes, Curtis; Porter, Kevin; Raviprakash, Kanakatte

    2007-11-01

    A candidate vaccine (D1ME-VRP) expressing dengue virus type 1 premembrane and envelope proteins in a Venezuelan equine encephalitis (VEE) virus replicon particle (VRP) system was constructed and tested in conjunction with a plasmid DNA vaccine (D1ME-DNA) expressing identical dengue virus sequences. Cynomolgus macaques were vaccinated with three doses of DNA (DDD), three doses of VRP (VVV group), or a heterologous DNA prime-VRP boost regimen (DDV) using two doses of DNA vaccine and a third dose of VRP vaccine. Four weeks after the final immunization, the DDV group produced the highest dengue virus type 1-specific immunoglobulin G antibody responses and virus-neutralizing antibody titers. Moderate T-cell responses were demonstrated only in DDD- and DDV-vaccinated animals. When vaccinated animals were challenged with live virus, all vaccination regimens showed significant protection from viremia. DDV-immunized animals were completely protected from viremia (mean time of viremia = 0 days), whereas DDD- and VVV-vaccinated animals had mean times of viremia of 0.66 and 0.75 day, respectively, compared to 6.33 days for the control group of animals.

  3. Assessment of delivery parameters with the multi-electrode array for development of a DNA vaccine against Bacillus anthracis.

    Science.gov (United States)

    Donate, Amy; Heller, Richard

    2013-12-01

    Gene electrotransfer (GET) enhances delivery of DNA vaccines by increasing both gene expression and immune responses. Our lab has developed the multi-electrode array (MEA) for DNA delivery to skin. The MEA was used at constant pulse duration (150 ms) and frequency (6.67 Hz). In this study, delivery parameters including applied voltage (5-45 V), amount of plasmid (100-300 μg), and number of treatments (2-3) were evaluated for delivery of a DNA vaccine. Mice were intradermally injected with plasmid expressing Bacillus anthracis protective antigen with or without GET and αPA serum titers measured. Within this experiment no significant differences were noted in antibody levels from varying dose or treatment number. However, significant differences were measured from applied voltages of 25 and 35 V. These voltages generated antibody levels between 20,000 and 25,000. Serum from animals vaccinated with these conditions also resulted in toxin neutralization in 40-60% of animals. Visual damage was noted at MEA conditions of 40 V. No damage was noted either visually or histologically from conditions of 35 V or below. These results reflect the importance of establishing appropriate electrical parameters and the potential for the MEA in non-invasive DNA vaccination against B. anthracis.

  4. Systemic Administration of CpG Oligodeoxynucleotide and Levamisole as Adjuvants for Gene-Gun-Delivered Antitumor DNA Vaccines

    Science.gov (United States)

    Šmahel, Michal; Poláková, Ingrid; Sobotková, Eva; Vajdová, Eva

    2011-01-01

    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. In this study, we tested systemic administration of two conventional adjuvants, synthetic oligodeoxynucleotide carrying immunostimulatory CpG motifs (CpG-ODN) and levamisole (LMS), and evaluated their effect on immune reactions induced by DNA vaccines delivered by a gene gun. DNA vaccination was directed either against the E7 oncoprotein of human papillomavirus type 16 or against the BCR-ABL1 oncoprotein characteristic for chronic myeloid leukemia. High doses of both adjuvants reduced activation of mouse splenic CD8+ T lymphocytes, but the overall antitumor effect was enhanced in both tumor models. High-dose CpG-ODN exhibited a superior adjuvant effect in comparison with any combination of CpG-ODN with LMS. In summary, our results demonstrate the benefit of combined therapy with gene-gun-delivered antitumor DNA vaccines and systemic administration of CpG-ODN or LMS. PMID:22028727

  5. Systemic Administration of CpG Oligodeoxynucleotide and Levamisole as Adjuvants for Gene-Gun-Delivered Antitumor DNA Vaccines

    Directory of Open Access Journals (Sweden)

    Michal Šmahel

    2011-01-01

    Full Text Available 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. In this study, we tested systemic administration of two conventional adjuvants, synthetic oligodeoxynucleotide carrying immunostimulatory CpG motifs (CpG-ODN and levamisole (LMS, and evaluated their effect on immune reactions induced by DNA vaccines delivered by a gene gun. DNA vaccination was directed either against the E7 oncoprotein of human papillomavirus type 16 or against the BCR-ABL1 oncoprotein characteristic for chronic myeloid leukemia. High doses of both adjuvants reduced activation of mouse splenic CD8+ T lymphocytes, but the overall antitumor effect was enhanced in both tumor models. High-dose CpG-ODN exhibited a superior adjuvant effect in comparison with any combination of CpG-ODN with LMS. In summary, our results demonstrate the benefit of combined therapy with gene-gun-delivered antitumor DNA vaccines and systemic administration of CpG-ODN or LMS.

  6. Recombinant invasive Lactococcus lactis can transfer DNA vaccines either directly to dendritic cells or across an epithelial cell monolayer

    NARCIS (Netherlands)

    Azevedo, de Marcela; Meijerink, Marjolein; Taverne, Nico; Pereira, Vanessa Bastos; LeBlanc, Jean Guy; Azevedo, Vasco; Miyoshi, Anderson; Langella, Philippe; Wells, J.M.; Chatel, Jean Marc

    2015-01-01

    Lactococcus lactis (L. lactis), a generally regarded as safe (GRAS) bacterium has recently been investigated as a mucosal delivery vehicle for DNA vaccines. Because of its GRAS status, L. lactis represents an attractive alternative to attenuated pathogens. Previous studies showed that eukaryotic

  7. Immunity to viral haemorrhagic septicaemia (VHS) following DNA vaccination of rainbow trout at an early life-stage

    DEFF Research Database (Denmark)

    Lorenzen, Niels; Lorenzen, Ellen; Einer-Jensen, Katja

    2001-01-01

    Rainbow trout fry of average weight 0.5 g were vaccinated against viral haemorrhagic septicaemia (VHS) by intramuscular injection of 1 mug of plasmid DNA encoding the VHS virus glycoprotein gene. Challenge with a lethal dose of virus at two different time points, 9 and 71 days post...

  8. Novel DNA vaccine based on hepatitis B virus core gene induces specific immune responses in Balb/c mice

    Institute of Scientific and Technical Information of China (English)

    Yi-Ping Xing; Zu-Hu Huang; Shi-Xia Wang; Jie Cai; Jun Li; Te-Hui W Chou; Shan Lu

    2005-01-01

    AIM: To investigate the immunogenicity of a novel DNA vaccine,pSW3891/HBc, based on HBV core gene in Balb/c mice.METHODS: A novel DNA vaccine, pSW3891/HBc, encoding HBV core gene was constructed using a vector plasmid pSW3891. Balb/c mice were immunized with either pSW3891/HBc or empty vector DNA via gene gun. IgG anti-HBc responses in mouse sera were demonstrated by ELISA. Specific cytotoxicity of cytotoxic T lymphocytes (CTLs) of mice was quantitatively measured by lactate dehydrogenase release assay.RESULTS: HBcAg was expressed effectively in 293T cell line transiently transfected with pSW3891/HBc. Strong IgG anti-HBc responses were elicited in mice immunized with pSW3891/HBc. The end-point titers of anti-HBc reached the highest 1:97 200, 4 wk after the third immunization. The specific CTL killing with the highest specific lysis reached 73.25% at effector:target ratio of 20:1 in mice that received pSW3891/HBc DNA vaccine.CONCLUSION: pSW3891/HBc vaccination elicits specific anti-HBc response and induces HBc-specific CTL response in immunized Balb/c mice.

  9. Enhancing DNA vaccine potency against hantavirus by co-administration of interleukin-12 expression vector as a genetic adjuvant

    Institute of Scientific and Technical Information of China (English)

    ZHENG Lan-yan; MOU Ling; LIN Song; LU Run-ming; LUO En-jie

    2005-01-01

    Background The heavy incidence and mortality of hemorrhagic fever with renal syndrome, as well as no specific drugs in curing the disease,clearly indicate the need for development of the more effective hantavirus vaccine. Refining the DNA vaccination strategy to elicit more clinically efficacious immune responses is now under intensive investigation. In the present study, we examined the effects of using an interleukin-12 expression plasmid as a genetic adjuvant to enhance the immune responses induced by a DNA vaccine based on the S gene encoding nucleocapsid protein against hantavirus. Methods BALB/c mice were immunized three times by intramuscular inoculations of DNA vaccine encoding of hantanvirus nucleocapsid protein alone or in combination with a plasmid expressing murine interleukin-12 (pcIL-12). Booster immunizations were employed 2 times at 2-week interval. To evaluate the humoral and cellular immune responses, antigen-specific lymphocyte proliferation and antibody production were assayed by MTT method and ELISA respectively. The level of interleukin-4 and interferon-γ in the splenic lymphocytic cultured supernatant were detected with ELISA kit at day 5, 10, 17, 35 and 42 after primary immunization.Conclusion Humoral and cytokine responses elicited by pcDNA3.1+S inoculation can be modulated by co-inoculation with pcIL-12 and efficiently induced Th1-dominant immune responses.

  10. Xenogeneic human p53 DNA vaccination by electroporation breaks immune tolerance to control murine tumors expressing mouse p53.

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    Ruey-Shyang Soong

    Full Text Available The pivotal role of p53 as a tumor suppressor protein is illustrated by the fact that this protein is found mutated in more than 50% of human cancers. In most cases, mutations in p53 greatly increase the otherwise short half-life of this protein in normal tissue and cause it to accumulate in the cytoplasm of tumors. The overexpression of mutated p53 in tumor cells makes p53 a potentially desirable target for the development of cancer immunotherapy. However, p53 protein represents an endogenous tumor-associated antigen (TAA. Immunization against a self-antigen is challenging because an antigen-specific immune response likely generates only low affinity antigen-specific CD8(+ T-cells. This represents a bottleneck of tumor immunotherapy when targeting endogenous TAAs expressed by tumors. The objective of the current study is to develop a safe cancer immunotherapy using a naked DNA vaccine. The vaccine employs a xenogeneic p53 gene to break immune tolerance resulting in a potent therapeutic antitumor effect against tumors expressing mutated p53. Our study assessed the therapeutic antitumor effect after immunization with DNA encoding human p53 (hp53 or mouse p53 (mp53. Mice immunized with xenogeneic full length hp53 DNA plasmid intramuscularly followed by electroporation were protected against challenge with murine colon cancer MC38 while those immunized with mp53 DNA were not. In a therapeutic model, established MC38 tumors were also well controlled by treatment with hp53 DNA therapy in tumor bearing mice compared to mp53 DNA. Mice vaccinated with hp53 DNA plasmid also exhibited an increase in mp53-specific CD8(+ T-cell precursors compared to vaccination with mp53 DNA. Antibody depletion experiments also demonstrated that CD8(+ T-cells play crucial roles in the antitumor effects. This study showed intramuscular vaccination with xenogeneic p53 DNA vaccine followed by electroporation is capable of inducing potent antitumor effects against tumors

  11. Quantitative expression profiling of immune response genes in rainbow trout following infectious haematopoietic necrosis virus (IHNV) infection or DNA vaccination

    Science.gov (United States)

    Purcell, Maureen K.; Kurath, Gael; Garver, Kyle A.; Herwig, Russell P.; Winton, James R.

    2004-01-01

    Infectious haematopoietic necrosis virus (IHNV) is a well-studied virus of salmonid fishes. A highly efficacious DNA vaccine has been developed against this virus and studies have demonstrated that this vaccine induces both an early and transient non-specific anti-viral phase as well as long-term specific protection. The mechanisms of the early anti-viral phase are not known, but previous studies noted changes in Mx gene expression, suggesting a role for type I interferon. This study used quantitative real-time reverse transcriptase PCR methodology to compare expression changes over time of a number of cytokine or cytokine-related genes in the spleen of rainbow trout following injection with poly I:C, live IHNV, the IHNV DNA vaccine or a control plasmid encoding the non-antigenic luciferase gene. The target genes included Mx-1, viral haemorrhagic septicaemia virus induced gene 8 (Vig-8), TNF-α1, TNF-α2, IL-1β1, IL-8, TGF-β1 and Hsp70. Poly I:C stimulation induced several genes but the strongest and significant response was observed in the Mx-1 and Vig-8 genes. The live IHN virus induced a significant response in all genes examined except TGF-β1. The control plasmid construct and the IHNV DNA vaccine marginally induced a number of genes, but the main difference between these two groups was a statistically significant induction of the Mx-1 and Vig-8 genes by the IHNV vaccine only. The gene expression profiles elicited by the live virus and the IHNV DNA vaccine differed in a number of aspects but this study confirms the clear role for a type I interferon-like response in early anti-viral defence.

  12. A polyvalent influenza A DNA vaccine induces heterologous immunity and protects pigs against pandemic A(H1N1)pdm09 virus infection

    DEFF Research Database (Denmark)

    Bragstad, Karoline; Vinner, Lasse; Hansen, Mette Sif

    2013-01-01

    intradermally with a combination of influenza DNA vaccine components based on the pandemic 1918 H1N1 (M and NP genes), pandemic 2009 H1N1pdm09 (HA and NA genes) and seasonal 2005 H3N2 genes (HA and NA genes) and investigated the protection against infection with virus both homologous and heterologous to the DNA......The composition of current influenza protein vaccines has to be reconsidered every season to match the circulating influenza viruses, continuously changing antigenicity. Thus, influenza vaccines inducing a broad cross-reactive immune response would be a great advantage for protection against both...... seasonal and emerging influenza viruses. We have developed an alternative influenza vaccine based on DNA expressing selected influenza proteins of pandemic and seasonal origin. In the current study, we investigated the protection of a polyvalent influenza DNA vaccine approach in pigs. We immunised pigs...

  13. Prime-boost therapeutic vaccination in mice with DNA/DNA or DNA/Fowlpox virus recombinants expressing the Human Papilloma Virus type 16 E6 and E7 mutated proteins fused to the coat protein of Potato virus X.

    Science.gov (United States)

    Illiano, Elena; Bissa, Massimiliano; Paolini, Francesca; Zanotto, Carlo; De Giuli Morghen, Carlo; Franconi, Rosella; Radaelli, Antonia; Venuti, Aldo

    2016-10-02

    The therapeutic antitumor potency of a prime-boost vaccination strategy was explored, based on the mutated, nontransforming forms of the E6 (E6F47R) and E7 (E7GGG) oncogenes of Human Papilloma Virus type 16 (HPV16), fused to the Potato virus X (PVX) coat protein (CP) sequence. Previous data showed that CP fusion improves the immunogenicity of tumor-associated antigens and may thus increase their efficacy. After verifying the correct expression of E6F47RCP and E7GGGCP inserted into DNA and Fowlpox virus recombinants by Western blotting and immunofluorescence, their combined use was evaluated for therapy in a pre-clinical mouse model of HPV16-related tumorigenicity. Immunization protocols were applied using homologous (DNA/DNA) or heterologous (DNA/Fowlpox) prime-boost vaccine regimens. The humoral immune responses were determined by ELISA, and the therapeutic efficacy evaluated by the delay in tumor appearance and reduced tumor volume after inoculation of syngeneic TC-1* tumor cells. Homologous DNA/DNA genetic vaccines were able to better delay tumor appearance and inhibit tumor growth when DNAE6F47RCP and DNAE7GGGCP were administered in combination. However, the heterologous DNA/Fowlpox vaccination strategy was able to delay tumor appearance in a higher number of animals when E6F47RCP and in particular E7GGGCP were administered alone. Copyright © 2016 Elsevier B.V. All rights reserved.

  14. Construction of a trivalent candidate vaccine against Shigella species with DNA recombination

    Institute of Scientific and Technical Information of China (English)

    王恒樑; 冯尔玲; 林云; 廖翔; 金明; 黄留玉; 苏国富; 黄翠芬

    2002-01-01

    In this work asd gene of Shigella flexneri 2a strain T32 was replaced by Vibrio cholerae toxin B subunit (ctxB) gene with DNA recombination in vivo and in vitro. The resulting derivative of T32, designed as FWL01, could stably express CtxB, but its growth in LB medium depended on the presence of diaminopimelic acid (DAP). Then form I plasmid of Shigella sonnei strain S7 was labeled with strain T32 asd gene and mobilized into FWL01. Thus a trivalent candidate oral vaccine strain, designed as FSW01, was constructed. In this candidate strain, a balanced-lethal system was constituted between the host strain and the form I plasmid expressing S. sonnei O antigen. Therefore the candidate strain can express stably not only its own O antigen but also CtxB and O antigen of S. sonnei in the absence of any antibiotic. Experiments showed that FSW01 did not invade HeLa cells or cause keratoconjunctivitis in guinea pigs. However, rabbits immunized FSW01 can elicit significant immune responses. In mice and rhesus monkey models, vaccinated animals were protected against the challenges of wild S. flexneri 2a strain 2457T and S. sonnei strain S9.

  15. Chimeric DNA Vaccines against ErbB2{sup +} Carcinomas: From Mice to Humans

    Energy Technology Data Exchange (ETDEWEB)

    Quaglino, Elena; Riccardo, Federica; Macagno, Marco; Bandini, Silvio; Cojoca, Rodica; Ercole, Elisabetta [Molecular Biotechnology Center, Department of Clinical and Biological Sciences, University of Turin, 10126 Turin (Italy); Amici, Augusto [Department of Molecular Cellular and Animal Biology, University of Camerino, 62032 Camerino (Italy); Cavallo, Federica, E-mail: federica.cavallo@unito.it [2 Department of Molecular Cellular and Animal Biology, University of Camerino, 62032 Camerino (Italy)

    2011-08-10

    DNA vaccination exploits a relatively simple and flexible technique to generate an immune response against microbial and tumor-associated antigens (TAAs). Its effectiveness is enhanced by the application of an electrical shock in the area of plasmid injection (electroporation). In our studies we exploited a sophisticated electroporation device approved for clinical use (Cliniporator, IGEA, Carpi, Italy). As the target antigen is an additional factor that dramatically modulates the efficacy of a vaccine, we selected ErbB2 receptor as a target since it is an ideal oncoantigen. It is overexpressed on the cell membrane by several carcinomas for which it plays an essential role in driving their progression. Most oncoantigens are self-tolerated molecules. To circumvent immune tolerance we generated two plasmids (RHuT and HuRT) coding for chimeric rat/human ErbB2 proteins. Their immunogenicity was compared in wild type mice naturally tolerant for mouse ErbB2, and in transgenic mice that are also tolerant for rat or human ErbB2. In several of these mice, RHuT and HuRT elicited a stronger anti-tumor response than plasmids coding for fully human or fully rat ErbB2. The ability of heterologous moiety to blunt immune tolerance could be exploited to elicit a significant immune response in patients. A clinical trial to delay the recurrence of ErbB2{sup +} carcinomas of the oral cavity, oropharynx and hypopharynx is awaiting the approval of the Italian authorities.

  16. Clitocybe nuda Activates Dendritic Cells and Acts as a DNA Vaccine Adjuvant

    Directory of Open Access Journals (Sweden)

    Mei-Hsing Chen

    2013-01-01

    Full Text Available This work represents the first evaluation of the effects of water extract of C. nuda (WE-CN, an edible mushroom, on murine bone marrow-derived dendritic cells (BMDCs and the potential pathway through which the effects are mediated. Our experimental results show that WE-CN could induce phenotypic maturation of DCs, as shown by the increased expression of MHC and costimulatory molecules. In addition, it also induced the proinflammatory cytokines expression on DCs and enhanced both the proliferation and IFN-γ secretion of allogenic T cells. Therefore, since WE-CN did not induce maturation of DCs generated from mice with mutated TLR-4 or TLR-2, suggesting that TLR4 and TLR2 might function as membrane receptors for WE-CN. Moreover, the mechanism of action of WE-CN may be mediated by increased phosphorylation of ERK, p38, and JNK mitogen-activated protein kinase (MAPK and increased NF-κB p65 activity, which are important signaling molecules downstream of TLR-4 and TLR-2. Finally, coimmunization of mice with WE-CN and a HER-2/neu DNA vaccine induced a HER-2/neu-specific Th1 response that resulted in significant inhibition of HER-2/neu overexpressing mouse bladder tumor (MBT-2 growth. These data suggest that WE-CN induces DC maturation through TLR-4 and/or TLR-2 and that WE-CN can be used as an adjuvant in cancer vaccine immunotherapy.

  17. CONSTRUCTION OF HUMAN INTERLUEKIN-18 DNA VACCINE AND IT'S EXPRESSION IN MAMMALIAN CELLS

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Objective To construct the human interleukin-18(hIL-18) DNA plasmids vaccine and to express the eukaryotic plasmids vaccine in mammalian cell lines Cos-7 and D5.Methods Gene recombinant technique was used to construct hIL-18 eukaryotic expression vectors.Calcium phosphate method was performed to transect recombinant hIL-18 eukaryotic expression vectors into Cos-7 and D5 cells.In situ hybridization and Western Blot were implemented to verify the transient expression of recombinant hIL-18 in Cos-7 and D5.Results The eukaryotic expression plasmid pVAX1-IL 18 was constructed successfully.hIL-18 was transiently expressed in Cos-7 and D5.Conclusion The eukaryotic expression plasmid pVAX1-IL 18 was constructed.In situ hybridization and Western Blot results proved the successful transient expression of pVAX1-IL 18 in Cos-7 and D5.Therefore,the work has settled the foundation for further biological research on hIL-18,including immunogene therapy through hIL-18.

  18. Immunogenicity of novel nanoparticle-coated MSP-1 C-terminus malaria DNA vaccine using different routes of administration.

    Science.gov (United States)

    Cherif, Mahamoud Sama; Shuaibu, Mohammed Nasir; Kurosaki, Tomoaki; Helegbe, Gideon Kofi; Kikuchi, Mihoko; Yanagi, Tetsuo; Tsuboi, Takafumi; Sasaki, Hitoshi; Hirayama, Kenji

    2011-11-08

    An important aspect in optimizing DNA vaccination is antigen delivery to the site of action. In this way, any alternative delivery system having higher transfection efficiency and eventual superior antibody production needs to be further explored. The novel nanoparticle, pDNA/PEI/γ-PGA complex, is one of a promising delivery system, which is taken up by cells and is shown to have high transfection efficiency. The immunostimulatory effect of this novel nanoparticle (NP) coated plasmid encoding Plasmodium yoelii MSP1-C-terminus was examined. Groups of C57BL/6 mice were immunized either with NP-coated MSP-1 plasmid, naked plasmid or NP-coated blank plasmid, by three different routes of administration; intravenous (i.v.), intraperitoneal (i.p.) and subcutaneous (s.c). Mice were primed and boosted twice at 3-week intervals, then challenged 2 weeks after; and 100%, 100% and 50% mean of survival was observed in immunized mice with coated DNA vaccine by i.p., i.v. and s.c., respectively. Coated DNA vaccine showed significant immunogenicity and elicited protective levels of antigen specific IgG and its subclass antibody, an increased proportion of CD4(+) and CD8(+) T cells and INF-γ and IL-12 levels in the serum and cultured splenocyte supernatant, as well as INF-γ producing cells in the spleen. We demonstrate that, NP-coated MSP-1 DNA-based vaccine confers protection against lethal P. yoelii challenge in murine model across the various route of administration and may therefore, be considered a promising delivery system for vaccination.

  19. Development of an oral DNA vaccine against MG7-Ag of gastric cancer using attenuated salmonella typhimurium as carrier

    Institute of Scientific and Technical Information of China (English)

    Chang-Cun Guo; Jie Ding; Bo-Rong Pan; Zhao-Cai Yu; Quan-Li Han; Fan-Ping Meng; Na Liu; Dai-Ming Fan

    2003-01-01

    AIM: To develop an oral DNA vaccine against gastric cancer and evaluate its efficacy in mice.METHODS: The genes of the MG7-Ag mimotope and a universal Th epitope (Pan-DR epitope, PADRE) were included in the PCR primers. By PCR, the fusion gene of the two epitopes was amplified. The fusion gene was confirmed by sequencing and was then cloned into pcDNA3.1(+) plasmid. The pcDNA3.1 (+)-MG7/PADRE was used to transfect an attenuated Salrmonella typhimuriurm.C57BL/6 mice were orally immunized with 1x108 cfu Salrmonella transfectants. Salmonella harboring the empty pcDNA3.1(+) plasmid and phosphate buffer saline (PBS)were used as negative controls. At the 6th week, serum titer of MG7-Ag specific antibody was detected by ELtSA.At the 8th week cellular immunity was detected by an unprimed proliferation test of the spleenocytes by using a [3H]-thymidine incorporation assay. Ehrlich ascites carcinoma cells expressing MG7-Ag were used as a model in tumor challenge assay to evaluate the protective effect of the vaccine.RESULTS: Serum titer of antibody against MG7-Ag was significantly higher in mice immunized with the vaccine than that in control groups (0.841 vs 0.347, P<0.01; 0.841 vs 0.298,P<0.01), while in vitro unprimed proliferation assay of the spleenocytes showed no statistical difference between those three groups. Two weeks after tumor challenge, 2 in 7 immunized mice were tumor free, while all the mice in the control groups showed tumor formation. CONCLUSION: Oral DNA vaccine against the MG7-Ag momitope of gastric cancer is immunogenic. It can induce significant humoral immunity against tumor in mice, and the vaccine has partially protective effects.

  20. A novel Sin Nombre virus DNA vaccine and its inclusion in a candidate pan-hantavirus vaccine against hantavirus pulmonary syndrome (HPS) and hemorrhagic fever with renal syndrome (HFRS).

    Science.gov (United States)

    Hooper, Jay W; Josleyn, Matthew; Ballantyne, John; Brocato, Rebecca

    2013-09-13

    Sin Nombre virus (SNV; family Bunyaviridae, genus Hantavirus) causes a hemorrhagic fever known as hantavirus pulmonary syndrome (HPS) in North America. There have been approximately 200 fatal cases of HPS in the United States since 1993, predominantly in healthy working-age males (case fatality rate 35%). There are no FDA-approved vaccines or drugs to prevent or treat HPS. Previously, we reported that hantavirus vaccines based on the full-length M gene segment of Andes virus (ANDV) for HPS in South America, and Hantaan virus (HTNV) and Puumala virus (PUUV) for hemorrhagic fever with renal syndrome (HFRS) in Eurasia, all elicited high-titer neutralizing antibodies in animal models. HFRS is more prevalent than HPS (>20,000 cases per year) but less pathogenic (case fatality rate 1-15%). Here, we report the construction and testing of a SNV full-length M gene-based DNA vaccine to prevent HPS. Rabbits vaccinated with the SNV DNA vaccine by muscle electroporation (mEP) developed high titers of neutralizing antibodies. Furthermore, hamsters vaccinated three times with the SNV DNA vaccine using a gene gun were completely protected against SNV infection. This is the first vaccine of any kind that specifically elicits high-titer neutralizing antibodies against SNV. To test the possibility of producing a pan-hantavirus vaccine, rabbits were vaccinated by mEP with an HPS mix (ANDV and SNV plasmids), or HFRS mix (HTNV and PUUV plasmids), or HPS/HFRS mix (all four plasmids). The HPS mix and HFRS mix elicited neutralizing antibodies predominantly against ANDV/SNV and HTNV/PUUV, respectively. Furthermore, the HPS/HFRS mix elicited neutralizing antibodies against all four viruses. These findings demonstrate a pan-hantavirus vaccine using a mixed-plasmid DNA vaccine approach is feasible and warrants further development. Copyright © 2013 The Authors. Published by Elsevier Ltd.. All rights reserved.

  1. A novel Sin Nombre virus DNA vaccine and its inclusion in a candidate pan-hantavirus vaccine against hantavirus pulmonary syndrome (HPS) and hemorrhagic fever with renal syndrome (HFRS)☆

    Science.gov (United States)

    Hooper, Jay W.; Josleyn, Matthew; Ballantyne, John; Brocato, Rebecca

    2014-01-01

    Sin Nombre virus (SNV; family Bunyaviridae, genus Hantavirus) causes a hemorrhagic fever known as hantavirus pulmonary syndrome (HPS) in North America. There have been approximately 200 fatal cases of HPS in the United States since 1993, predominantly in healthy working-age males (case fatality rate 35%). There are no FDA-approved vaccines or drugs to prevent or treat HPS. Previously, we reported that hantavirus vaccines based on the full-length M gene segment of Andes virus (ANDV) for HPS in South America, and Hantaan virus (HTNV) and Puumala virus (PUUV) for hemorrhagic fever with renal syndrome (HFRS) in Eurasia, all elicited high-titer neutralizing antibodies in animal models. HFRS is more prevalent than HPS (>20,000 cases per year) but less pathogenic (case fatality rate 1–15%). Here, we report the construction and testing of a SNV full-length M gene-based DNA vaccine to prevent HPS. Rabbits vaccinated with the SNV DNA vaccine by muscle electroporation (mEP) developed high titers of neutralizing antibodies. Furthermore, hamsters vaccinated three times with the SNV DNA vaccine using a gene gun were completely protected against SNV infection. This is the first vaccine of any kind that specifically elicits high-titer neutralizing antibodies against SNV. To test the possibility of producing a pan-hantavirus vaccine, rabbits were vaccinated by mEP with an HPS mix (ANDV and SNV plasmids), or HFRS mix (HTNV and PUUV plasmids), or HPS/HFRS mix (all four plasmids). The HPS mix and HFRS mix elicited neutralizing antibodies predominantly against ANDV/SNV and HTNV/PUUV, respectively. Furthermore, the HPS/HFRS mix elicited neutralizing antibodies against all four viruses. These findings demonstrate a pan-hantavirus vaccine using a mixed-plasmid DNA vaccine approach is feasible and warrants further development. PMID:23892100

  2. Linear and branched glyco-lipopeptide vaccines follow distinct cross-presentation pathways and generate different magnitudes of antitumor immunity.

    Directory of Open Access Journals (Sweden)

    Olivier Renaudet

    Full Text Available BACKGROUND: Glyco-lipopeptides, a form of lipid-tailed glyco-peptide, are currently under intense investigation as B- and T-cell based vaccine immunotherapy for many cancers. However, the cellular and molecular mechanisms of glyco-lipopeptides (GLPs immunogenicity and the position of the lipid moiety on immunogenicity and protective efficacy of GLPs remain to be determined. METHODS/PRINCIPAL FINDINGS: We have constructed two structural analogues of HER-2 glyco-lipopeptide (HER-GLP by synthesizing a chimeric peptide made of one universal CD4(+ epitope (PADRE and one HER-2 CD8(+ T-cell epitope (HER(420-429. The C-terminal end of the resulting CD4-CD8 chimeric peptide was coupled to a tumor carbohydrate B-cell epitope, based on a regioselectively addressable functionalized templates (RAFT, made of four alpha-GalNAc molecules. The resulting HER glyco-peptide (HER-GP was then linked to a palmitic acid moiety, attached either at the N-terminal end (linear HER-GLP-1 or in the middle between the CD4+ and CD8+ T cell epitopes (branched HER-GLP-2. We have investigated the uptake, processing and cross-presentation pathways of the two HER-GLP vaccine constructs, and assessed whether the position of linkage of the lipid moiety would affect the B- and T-cell immunogenicity and protective efficacy. Immunization of mice revealed that the linear HER-GLP-1 induced a stronger and longer lasting HER(420-429-specific IFN-gamma producing CD8(+ T cell response, while the branched HER-GLP-2 induced a stronger tumor-specific IgG response. The linear HER-GLP-1 was taken up easily by dendritic cells (DCs, induced stronger DCs maturation and produced a potent TLR- 2-dependent T-cell activation. The linear and branched HER-GLP molecules appeared to follow two different cross-presentation pathways. While regression of established tumors was induced by both linear HER-GLP-1 and branched HER-GLP-2, the inhibition of tumor growth was significantly higher in HER-GLP-1 immunized

  3. A dengue DNA vaccine formulated with Vaxfectin® is well tolerated, and elicits strong neutralizing antibody responses to all four dengue serotypes in New Zealand white rabbits.

    Science.gov (United States)

    Raviprakash, Kanakatte; Luke, Thomas; Doukas, John; Danko, Janine; Porter, Kevin; Burgess, Timothy; Kochel, Tadeusz

    2012-12-01

    A tetravalent DNA vaccine formulated with Vaxfectin adjuvant was shown to elicit high levels of neutralizing antibody against all four dengue virus serotypes (Porter et al., ( 16) ), warranting further testing in humans. In preparation for a phase 1 clinical testing, the vaccine and the adjuvant were manufactured under current good manufacturing practice guidelines. The formulated vaccine and the adjuvant were tested for safety and/or immunogenicity in New Zealand white rabbits using a repeat dose toxicology study. The formulated vaccine and the adjuvant were found to be well tolerated by the animals. Animals injected with formulated vaccine produced strong neutralizing antibody response to all four dengue serotypes.

  4. A Versatile Multiple Target Detection System Based on DNA Nano-assembled Linear FRET Arrays.

    Science.gov (United States)

    Li, Yansheng; Du, Hongwu; Wang, Wenqian; Zhang, Peixun; Xu, Liping; Wen, Yongqiang; Zhang, Xueji

    2016-05-27

    DNA molecules have been utilized both as powerful synthetic building blocks to create nanoscale architectures and as inconstant programmable templates for assembly of biosensors. In this paper, a versatile, scalable and multiplex detection system is reported based on an extending fluorescent resonance energy transfer (FRET) cascades on a linear DNA assemblies. Seven combinations of three kinds of targets are successfully detected through the changes of fluorescence spectra because of the three-steps FRET or non-FRET continuity mechanisms. This nano-assembled FRET-based nanowire is extremely significant for the development of rapid, simple and sensitive detection system. The method used here could be extended to a general platform for multiplex detection through more-step FRET process.

  5. Systemically administered gp100 encoding DNA vaccine for melanoma using water-in-oil-in-water multiple emulsion delivery systems.

    Science.gov (United States)

    Kalariya, Mayurkumar; Amiji, Mansoor M

    2013-09-10

    The purpose of this study was to develop a water-in-oil-in-water (W/O/W) multiple emulsions-based vaccine delivery system for plasmid DNA encoding the gp100 peptide antigen for melanoma immunotherapy. The gp100 encoding plasmid DNA was encapsulated in the inner-most aqueous phase of squalane oil containing W/O/W multiple emulsions using a two-step emulsification method. In vitro transfection ability of the encapsulated plasmid DNA was investigated in murine dendritic cells by transgene expression analysis using fluorescence microscopy and ELISA methods. Prophylactic immunization using the W/O/W multiple emulsions encapsulated the gp100 encoding plasmid DNA vaccine significantly reduced tumor volume in C57BL/6 mice during subsequent B16-F10 tumor challenge. In addition, serum Th1 cytokine levels and immuno-histochemistry of excised tumor tissues indicated activation of cytotoxic T-lymphocytes mediated anti-tumor immunity causing tumor growth suppression. The W/O/W multiple emulsions-based vaccine delivery system efficiently delivers the gp100 plasmid DNA to induce cell-mediated anti-tumor immunity.

  6. Cationic Lipid-Formulated DNA Vaccine against Hepatitis B Virus : Immunogenicity of MIDGE-Th1 Vectors Encoding Small and Large Surface Antigen in Comparison to a Licensed Protein Vaccine

    NARCIS (Netherlands)

    Endmann, Anne; Klunder, Katharina; Kapp, Kerstin; Riede, Oliver; Oswald, Detlef; Talman, Eduard G.; Schroff, Matthias; Kleuss, Christiane; Ruiters, Marcel H. J.; Juhls, Christiane

    2014-01-01

    Currently marketed vaccines against hepatitis B virus (HBV) based on the small (S) hepatitis B surface antigen (HBsAg) fail to induce a protective immune response in about 10% of vaccinees. DNA vaccination and the inclusion of PreS1 and PreS2 domains of HBsAg have been reported to represent feasible

  7. Self-entanglement of long linear DNA vectors using transient non-B-DNA attachment points: a new concept for improvement of non-viral therapeutic gene delivery.

    Science.gov (United States)

    Tolmachov, Oleg E

    2012-05-01

    The cell-specific and long-term expression of therapeutic transgenes often requires a full array of native gene control elements including distal enhancers, regulatory introns and chromatin organisation sequences. The delivery of such extended gene expression modules to human cells can be accomplished with non-viral high-molecular-weight DNA vectors, in particular with several classes of linear DNA vectors. All high-molecular-weight DNA vectors are susceptible to damage by shear stress, and while for some of the vectors the harmful impact of shear stress can be minimised through the transformation of the vectors to compact topological configurations by supercoiling and/or knotting, linear DNA vectors with terminal loops or covalently attached terminal proteins cannot be self-compacted in this way. In this case, the only available self-compacting option is self-entangling, which can be defined as the folding of single DNA molecules into a configuration with mutual restriction of molecular motion by the individual segments of bent DNA. A negatively charged phosphate backbone makes DNA self-repulsive, so it is reasonable to assume that a certain number of 'sticky points' dispersed within DNA could facilitate the entangling by bringing DNA segments into proximity and by interfering with the DNA slipping away from the entanglement. I propose that the spontaneous entanglement of vector DNA can be enhanced by the interlacing of the DNA with sites capable of mutual transient attachment through the formation of non-B-DNA forms, such as interacting cruciform structures, inter-segment triplexes, slipped-strand DNA, left-handed duplexes (Z-forms) or G-quadruplexes. It is expected that the non-B-DNA based entanglement of the linear DNA vectors would consist of the initial transient and co-operative non-B-DNA mediated binding events followed by tight self-ensnarement of the vector DNA. Once in the nucleoplasm of the target human cells, the DNA can be disentangled by type II

  8. Head-to-head comparison of three vaccination strategies based on DNA and raw insect-derived recombinant proteins against Leishmania.

    Science.gov (United States)

    Todolí, Felicitat; Rodríguez-Cortés, Alhelí; Núñez, María Del Carmen; Laurenti, Márcia D; Gómez-Sebastián, Silvia; Rodríguez, Fernando; Pérez-Martín, Eva; Escribano, José M; Alberola, Jordi

    2012-01-01

    Parasitic diseases plague billions of people among the poorest, killing millions annually, and causing additional millions of disability-adjusted life years lost. Leishmaniases affect more than 12 million people, with over 350 million people at risk. There is an urgent need for efficacious and cheap vaccines and treatments against visceral leishmaniasis (VL), its most severe form. Several vaccination strategies have been proposed but to date no head-to-head comparison was undertaken to assess which is the best in a clinical model of the disease. We simultaneously assayed three vaccination strategies against VL in the hamster model, using KMPII, TRYP, LACK, and PAPLE22 vaccine candidate antigens. Four groups of hamsters were immunized using the following approaches: 1) raw extracts of baculovirus-infected Trichoplusia ni larvae expressing individually one of the four recombinant proteins (PROT); 2) naked pVAX1 plasmids carrying the four genes individually (DNA); 3) a heterologous prime-boost (HPB) strategy involving DNA followed by PROT (DNA-PROT); and 4) a Control including empty pVAX1 plasmid followed by raw extract of wild-type baculovirus-infected T. ni larvae. Hamsters were challenged with L. infantum promastigotes and maintained for 20 weeks. While PROT vaccine was not protective, DNA vaccination achieved protection in spleen. Only DNA-PROT vaccination induced significant NO production by macrophages, accompanied by a significant parasitological protection in spleen and blood. Thus, the DNA-PROT strategy elicits strong immune responses and high parasitological protection in the clinical model of VL, better than its corresponding naked DNA or protein versions. Furthermore, we show that naked DNA coupled with raw recombinant proteins produced in insect larvae biofactories -the cheapest way of producing DNA-PROT vaccines- is a practical and cost-effective way for potential "off the shelf" supplying vaccines at very low prices for the protection against

  9. Effect of phylogenetic diversity of velogenic Newcastle disease virus challenge on virus shedding post homologous and heterologous DNA vaccination in chickens.

    Science.gov (United States)

    Mohamed, Mahmoud H A; Abdelaziz, Adel M; Kumar, Sachin; Al-Habib, Malik A; Megahed, Mohamed M

    2016-01-01

    Newcastle disease (ND) is a highly devastating disease for the poultry industry as it causes high economic losses. In this present study, a DNA vaccine containing the F and HN surface antigens of a highly virulent Newcastle disease virus (NDV), NDV/1/Chicken/2005 (FJ939313), was successfully generated. Cell transfection test indicated that the vaccine expressed the F and HN genes in Hep-2 cells. The main objective of this study was to compare the extent of protection induced by DNA vaccination after homologous and heterologous NDV-challenge as determined by the amount of NDV shedding after challenge. NDV-antibody-negative chickens were vaccinated either once, twice or thrice intramuscularly at 7, 14 and 21 days old and were challenged 14 days post vaccination with either homologous virus (vaccine-matched velogenic viscerotropic Newcastle disease virus (vvNDV) strain, FJ939313), phylogenetically related to group VII, or a phylogenetically divergent heterologous virus (unmatched vvNDV strain, AY968809), which belongs to genogroup VI and shows 84.1% nucleotide similarity to the NDV-sequences of the DNA vaccine. Our data indicate that birds, which received a single dose of the DNA vaccine were poorly protected, and only 30-40% of these birds survived after challenge with high virus shedding titre. Multiple administration of the DNA vaccine induced high protection rates of 70-90% with reduced virus shedding compared to the non-vaccinated and challenged birds. Generally, homologous challenge led to reduced tracheal and cloacal shedding compared to the heterologous vvNDV strain. This study provides a promising approach for the control of ND in chickens using DNA vaccines, which are phylogenetically closely related to the circulating field strains.

  10. Targeting Carcinoembryonic Antigen with DNA Vaccination: On-Target Adverse Events Link with Immunological and Clinical Outcomes

    Science.gov (United States)

    Chudley, Lindsey; Stasakova, Jana; Thirdborough, Stephen; King, Andrew; Lloyd-Evans, Paul; Buxton, Emily; Edwards, Ceri; Halford, Sarah; Bateman, Andrew; O’Callaghan, Ann; Clive, Sally; Anthoney, Alan; Jodrell, Duncan I.; Weinschenk, Toni; Simon, Petra; Sahin, Ugur; Thomas, Gareth J.; Stevenson, Freda K.; Ottensmeier, Christian H.

    2017-01-01

    Purpose We have clinically evaluated a DNA fusion vaccine to target the HLA-A*0201 binding peptide CAP-1 from carcinoembryonic antigen (CEA605–613) linked to an immunostimulatory domain (DOM) from fragment C of tetanus toxin. Experimental Design Twenty-seven patients with CEA-expressing carcinomas were recruited: 15 patients with measurable disease (Arm-I) and 12 patients without radiological evidence of disease (Arm-II). Six intramuscular vaccinations of naked DNA (1mg/dose) were administered up to week 12. Clinical and immunological follow-up was to week 64 or clinical/radiological disease. Results DOM-specific immune responses demonstrated successful vaccine delivery. All patients without measurable disease compared to 60% with advanced disease responded immunologically, while 58% and 20% expanded anti-CAP-1 CD8+ T-cells, respectively. CAP-1-specific T-cells were only detectable in the blood post-vaccination, but could also be identified in previously resected cancer tissue. The gastrointestinal adverse event diarrhea was reported by 48% of patients and linked to more frequent decreases in CEA (pdiarrhea. In advanced disease patients, decreases in CEA were associated with better overall survival (HR=0.14, p=0.017). CAP-1 peptide was detectable on MHC class I of normal bowel mucosa and primary colorectal cancer tissue by mass-spectrometry, offering a mechanistic explanation for diarrhea through CD8+ T-cell attack. Conclusions Our data suggest that DNA vaccination is able to overcome peripheral tolerance in normal and tumor tissue and warrants testing in combination studies, for example, by vaccinating in parallel to treatment with an anti-PD1 antibody. PMID:27091407

  11. Advances in the research of adjuvants for plasmid DNA vaccines%DNA疫苗佐剂的研究进展

    Institute of Scientific and Technical Information of China (English)

    蒋丽明; 叶琳

    2009-01-01

    DNA疫苗是一种很有希望的免疫方法,经多途径接种质粒DNA能引起有效的免疫应答,重复给予不会产生抗载体免疫.然而,质粒DNA疫苗在小型实验动物中诱导的免疫应答远强于在人类和其他非人灵长类动物中.已设计多种佐剂通过直接刺激免疫系统或增强DNA表达来提高疫苗的免疫原性,这些佐剂包括免疫协同刺激分子、细胞因子、补体分子、脂质体、核酸、聚合物、纳米粒和微粒类佐剂.此文对DNA疫苗佐剂的研究进展作一综述.%Plasmid DNA vaccine is a promising modality for immunization. Immunization with plasmid DNA by various routes can trigger effective iimnune responses. The immunogens can be administered repeatedly without inducing anti-vector immunity. However, the immune responses induced by plasmid DNA vaccines are much stronger in small laboratory animal models than in non-human primates and humans. A number of adjuvants, including immune co-stimulatory molecules, cytokines, complement molecules, liposomes, nucleic acids, polymers, micro-and nano-particles, have been designed to improve the immunogenicity of DNA vaccines by directly stimulating the immune system or by enhancing plasmid DNA expression. This review introduces the progress in development of these adjuvants for plasmid DNA vaccines.

  12. Eimeria maxima microneme protein 2 delivered as DNA vaccine and recombinant protein induces immunity against experimental homogenous challenge.

    Science.gov (United States)

    Huang, Jingwei; Zhang, Zhenchao; Li, Menghui; Song, Xiaokai; Yan, Ruofeng; Xu, Lixin; Li, Xiangrui

    2015-10-01

    E. maxima is one of the seven species of Eimeria that infects chicken. Until now, only a few antigenic genes of E. maxima have been reported. In the present study, the immune protective effects against E. maxima challenge of recombinant protein and DNA vaccine encoding EmMIC2 were evaluated. Two-week-old chickens were randomly divided into five groups. The experimental group of chickens was immunized with 100 μg DNA vaccine pVAX1-MIC2 or 200 μg rEmMIC2 protein while the control group of chickens was injected with pVAX1 plasmid or sterile PBS. The results showed that the anti-EmMIC2 antibody titers of both rEmMIC2 protein and pVAX1-MIC2 groups were significantly higher as compared to PBS and pVAX1 control (Precombinant protein and the DNA vaccine could obviously alleviate jejunum lesions, body weight loss, increase oocyst, decrease ratio and provide ACIs of more than 165. All the above results suggested that immunization with EmMIC2 was effective in imparting partial protection against E. maxima challenge and it could be an effective antigen candidate for the development of new vaccines against E. maxima.

  13. Multivalent immunity targeting tumor-associated antigens by intra-lymph node DNA-prime, peptide-boost vaccination.

    Science.gov (United States)

    Smith, K A; Qiu, Z; Wong, R; Tam, V L; Tam, B L; Joea, D K; Quach, A; Liu, X; Pold, M; Malyankar, U M; Bot, A

    2011-01-01

    Active immunotherapy of cancer has yet to yield effective therapies in the clinic. To evaluate the translatability of DNA-based vaccines we analyzed the profile of T-cell immunity by plasmid vaccination in a murine model, using transcriptome microarray analysis and flow cytometry. DNA vaccination resulted in specific T cells expressing low levels of co-inhibitory molecules (most notably PD-1), strikingly different from the expression profile elicited by peptide immunization. In addition, the T-cell response primed through this dual-antigen-expressing plasmid (MART-1/Melan-A and tyrosinase) translated into a substantial proliferation capacity and functional conversion to antitumor effector cells after tyrosinase and MART-1/Melan-A peptide analog boost. Furthermore, peptide boost rescued the immune response against the subdominant tyrosinase epitope. This immunization approach could be adapted to elicit potent immunity against multiple tumor antigens, resulting in a broader immune response that was more effective in targeting human tumor cells. Finally, this study sheds light on a novel mechanism of immune homeostasis through synchronous regulation of co-inhibitory molecules on T cells, highly relevant to heterologous prime boost approaches involving DNA vaccines as priming agents.

  14. Toxoplasma gondii: Vaccination with a DNA vaccine encoding T- and B-cell epitopes of SAG1, GRA2, GRA7 and ROP16 elicits protection against acute toxoplasmosis in mice.

    Science.gov (United States)

    Cao, Aiping; Liu, Yuan; Wang, Jingjing; Li, Xun; Wang, Shuai; Zhao, Qunli; Cong, Hua; He, Shenyi; Zhou, Huaiyu

    2015-11-27

    Toxoplasma gondii (T. gondii) is an obligate, intracellular, protozoan parasite that infects large variety of warm-blooded animals including humans, livestock, and marine mammals, and causes the disease toxoplasmosis. Although T. gondii infection rates differ significantly from country to country, it still has a high morbidity and mortality. In these circumstances, developing an effective vaccine against T. gondii is urgently needed for preventing and treating toxoplasmosis. The aim of this study was to construct a multi-epitopes DNA vaccine and evaluate the immune protective efficacy against acute toxoplasmosis in mice. Therefore, twelve T- and B-cell epitopes from SAG1, GRA2, GRA7 and ROP16 of T. gondii were predicted by bioinformatics analysis, and then a multi-epitopes DNA vaccine was constructed. Mice immunized with the multi-epitopes DNA vaccine gained higher levels of IgG titers and IgG2a subclass titers, significant production of gamma interferon (IFN-γ), percentage of T lymphocyte subsets, and longer survival times against the acute infection of T. gondii compared with those of mice administered with empty plasmid and those in control groups. Furthermore, a genetic adjuvant pEGFP-RANTES (pRANTES) could enhance the efficacy of the multi-epitopes DNA vaccine associating with humoral and cellular (Th1, CD8(+) T cell) immune responses. Above all, the DNA vaccine and the genetic adjuvant revealed in this study might be new candidates for further vaccine development against T. gondii infection.

  15. Targeting hepatitis B virus antigens to dendritic cells by heat shock protein to improve DNA vaccine potency

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    AIM: To investigate a novel DNA vaccination based upon expression of the HBV e antigen fused to a heat shock protein (HSP) as a strategy to enhance DNA vaccine potency.METHODS: A pCMV-HBeAg-HSP DNA vaccine and a control DNA vaccine were generated. Mice were immunized with these different construct. Immune responses were measured 2 wk after a second immunization by a T cell response assay, CTL cytotoxicity assay, and an antibody assay in C57BL/6 and BALB/c mice. CT26-HBeAg tumor cell challenge test in vivo was performed in BALB/c mice to monitor anti-tumor immune responses.RESULTS: In the mice immunized with pCMV-HBe-HSP DNA, superior CTL activity to target HBV-positive target cells was observed in comparison with mice immunized with pCMV-HBeAg (44% ± 5% vs 30% ± 6% in E: T > 50:1, P < 0.05). ELISPOT assays showed a stronger T-cell response from mice immunized with pCMV-HBe-HSP than that from pCMV-HBeAg immunized animals when stimulated either with MHC class Ⅰ or class Ⅱ epitopes derived from HBeAg (74% ± 9% vs 31% ± 6%, P < 0.01). ELISA assays revealed an enhanced HBeAg antibody response from mice immunized with pCMV-HBe-HSP than from those immunized with pCMV-HBeAg. The lowest tumor incidence and the slowest tumor growth were observed in mice immunized with pCMV-HBe-HSP when challenged with CT26-HBeAg.CONCLUSION: The results of this study demonstrate a broad enhancement of antigen-specific CD4+ helper,CD8+ cytotoxic T-cell, and B-cell responses by a novel DNA vaccination strategy. They also proved a stronger antigen-specific immune memory, which may be superior to currently described HBV DNA vaccination strategies for the treatment of chronic HBV infection.

  16. GM-CSF increases mucosal and systemic immunogenicity of an H1N1 influenza DNA vaccine administered into the epidermis of non-human primates.

    Directory of Open Access Journals (Sweden)

    Peter T Loudon

    Full Text Available BACKGROUND: The recent H5N1 avian and H1N1 swine-origin influenza virus outbreaks reaffirm that the threat of a world-wide influenza pandemic is both real and ever-present. Vaccination is still considered the best strategy for protection against influenza virus infection but a significant challenge is to identify new vaccine approaches that offer accelerated production, broader protection against drifted and shifted strains, and the capacity to elicit anti-viral immune responses in the respiratory tract at the site of viral entry. As a safe alternative to live attenuated vaccines, the mucosal and systemic immunogenicity of an H1N1 influenza (A/New Caledonia/20/99 HA DNA vaccine administered by particle-mediated epidermal delivery (PMED or gene gun was analyzed in rhesus macaques. METHODOLOGY/PRINCIPAL FINDINGS: Macaques were immunized at weeks 0, 8, and 16 using a disposable single-shot particle-mediated delivery device designed for clinical use that delivers plasmid DNA directly into cells of the epidermis. Significant levels of hemagglutination inhibiting (HI antibodies and cytokine-secreting HA-specific T cells were observed in the periphery of macaques following 1-3 doses of the PMED HA DNA vaccine. In addition, HA DNA vaccination induced detectable levels of HA-specific mucosal antibodies and T cells in the lung and gut-associated lymphoid tissues of vaccinated macaques. Importantly, co-delivery of a DNA encoding the rhesus macaque GM-CSF gene was found to significantly enhance both the systemic and mucosal immunogenicity of the HA DNA vaccine. CONCLUSIONS/SIGNIFICANCE: These results provide strong support for the development of a particle-mediated epidermal DNA vaccine for protection against respiratory pathogens such as influenza and demonstrate, for the first time, the ability of skin-delivered GM-CSF to serve as an effective mucosal adjuvant for vaccine induction of immune responses in the gut and respiratory tract.

  17. Immune responses in mice vaccinated with a suicidal DNA vaccine expressing the hemagglutinin glycoprotein from the peste des petits ruminants virus.

    Science.gov (United States)

    Wang, Yong; Liu, Guangqing; Shi, Lijun; Li, Wenchao; Li, Chuanfeng; Chen, Zongyan; Jin, Hongyan; Xu, Binrui; Li, Gang

    2013-11-01

    Peste des petits ruminants (PPR), an acute and highly contagious disease, affects sheep, goats, and some small ruminants. The hemagglutinin (H) glycoprotein of the PPR virus (PPRV) is considered important for inducing protective immune responses. In this study, a suicidal DNA vaccine based on the Semliki Forest virus (SFV) replicon was constructed and tested for its ability to induce immunogenicity in a mouse model. For this, the H gene of PPRV was cloned and inserted into pSCA1, an SFV replicon vector. The resultant plasmid named pSCA1-H was then transfected into BHK-21 cells following which the antigenicity of the expressed protein was confirmed by Western blotting and immunofluorescence. The pSCA1-H plasmid was then injected intramuscularly into BALB/c mice thrice at 2-week intervals. To evaluate the immunogenicity of pSCA1-H, specific antibodies and neutralizing antibodies against PPRV-H were measured using an indirect enzyme-linked immunosorbent assay and a microneutralization test, respectively. Cell-mediated immune responses were also examined using a lymphocyte proliferation assay. The results showed that pSCA1-H could express the H protein in BHK-21 cells. Specific antibodies, neutralizing antibodies, and lymphocyte proliferation responses were all induced in mice. Thus, this suicidal DNA vaccine could be a promising new approach for vaccine development against PPR. Copyright © 2013 Elsevier B.V. All rights reserved.

  18. Dual DNA vaccination of rainbow trout (Oncorhynchus mykiss) against two different rhabdoviruses, VHSV and IHNV, induces specific divalent protection

    DEFF Research Database (Denmark)

    Einer-Jensen, Katja; Delgado, L.; Lorenzen, Ellen;

    2009-01-01

    DNA vaccines encoding the glycoprotein genes of the salmonid rhabdoviruses VHSV and IHNV are very efficient in eliciting protective immune responses against their respective diseases in rainbow trout (Oncorhynchus mykiss). The early anti-viral response (EAVR) provides Protection by 4 days post...... vaccination and is non-specific and transient while the specific anti-viral response (SAVR) is long lasting and highly specific. Since both VHSV and IHNV are endemic in rainbow trout in several geographical regions of Europe and Atlantic salmon (Salmo salar) on the Pacific coast of North America, co...

  19. Co-administration of the polysaccharide of Lycium barbarum with DNA vaccine of Chlamydophila abortus augments protection.

    Science.gov (United States)

    Ling, Yong; Li, Shaowen; Yang, Junjing; Yuan, Jilei; He, Cheng

    2011-01-01

    Lycium barbarum polysaccharides (LBP) can stimulate moderate immune responses therefore could potentially be used as a substitute for oil adjuvants in veterinary vaccines. In the present study, it was shown that the isolated active component of LBP3a, combined with a DNA vaccine encoding the major outer membrane protein (MOMP) of Chlamydophila abortus, induced protection in mice against challenge. Sixty BALB/c mice were randomly assigned to 5 groups. Sub-fractions of polysaccharide LBP3a, at 12.5, 25 and 50 mg/kg concentrations, respectively, were mixed with a pCI-neo::MOMP (pMOMP) vaccine. Mice administrated with pCI-neo + LBP3a were served as a control. All mice were inoculated at day 0, 14, and 28, and challenged on day 44. The effects of LBp3a on serum antibody levels, in vitro lymphocyte proliferation, the activity of interleaukin-2 (IL-2), interferon-γ (IFN-γ), tumor necrosis factor α(TNF-α)and chlamydia clearance were determined. A combination of DNA vaccine and LBP3a induced significantly higher antibody levels in mice, higher T cell proliferation and higher levels of IFN-γ and IL-2. Mice immunized with DNA and LBP3a also showed significantly higher levels of chlamydia clearance in mice spleens and a greater Th1 immune response. The immunoenhancement induced by 25 mg/kg LBP3a is more effective than that induced by a 12.5 and 50 mg/kg. This implies that LBP3a at 25 mg/kg has a high potential to be used as an effective adjuvant with a DNA vaccine against swine Chlamydophila abortus.

  20. DNA vaccine-generated duck polyclonal antibodies as a postexposure prophylactic to prevent hantavirus pulmonary syndrome (HPS.

    Directory of Open Access Journals (Sweden)

    Rebecca Brocato

    Full Text Available Andes virus (ANDV is the predominant cause of hantavirus pulmonary syndrome (HPS in South America and the only hantavirus known to be transmitted person-to-person. There are no vaccines, prophylactics, or therapeutics to prevent or treat this highly pathogenic disease (case-fatality 35-40%. Infection of Syrian hamsters with ANDV results in a disease that closely mimics human HPS in incubation time, symptoms of respiratory distress, and disease pathology. Here, we evaluated the feasibility of two postexposure prophylaxis strategies in the ANDV/hamster lethal disease model. First, we evaluated a natural product, human polyclonal antibody, obtained as fresh frozen plasma (FFP from a HPS survivor. Second, we used DNA vaccine technology to manufacture a polyclonal immunoglobulin-based product that could be purified from the eggs of vaccinated ducks (Anas platyrhynchos. The natural "despeciation" of the duck IgY (i.e., Fc removed results in an immunoglobulin predicted to be minimally reactogenic in humans. Administration of ≥ 5,000 neutralizing antibody units (NAU/kg of FFP-protected hamsters from lethal disease when given up to 8 days after intranasal ANDV challenge. IgY/IgYΔFc antibodies purified from the eggs of DNA-vaccinated ducks effectively neutralized ANDV in vitro as measured by plaque reduction neutralization tests (PRNT. Administration of 12,000 NAU/kg of duck egg-derived IgY/IgYΔFc protected hamsters when administered up to 8 days after intranasal challenge and 5 days after intramuscular challenge. These experiments demonstrate that convalescent FFP shows promise as a postexposure HPS prophylactic. Moreover, these data demonstrate the feasibility of using DNA vaccine technology coupled with the duck/egg system to manufacture a product that could supplement or replace FFP. The DNA vaccine-duck/egg system can be scaled as needed and obviates the necessity of using limited blood products obtained from a small number of HPS survivors. This

  1. DNA Vaccine-Generated Duck Polyclonal Antibodies as a Postexposure Prophylactic to Prevent Hantavirus Pulmonary Syndrome (HPS)

    Science.gov (United States)

    Brocato, Rebecca; Josleyn, Matthew; Ballantyne, John; Vial, Pablo; Hooper, Jay W.

    2012-01-01

    Andes virus (ANDV) is the predominant cause of hantavirus pulmonary syndrome (HPS) in South America and the only hantavirus known to be transmitted person-to-person. There are no vaccines, prophylactics, or therapeutics to prevent or treat this highly pathogenic disease (case-fatality 35–40%). Infection of Syrian hamsters with ANDV results in a disease that closely mimics human HPS in incubation time, symptoms of respiratory distress, and disease pathology. Here, we evaluated the feasibility of two postexposure prophylaxis strategies in the ANDV/hamster lethal disease model. First, we evaluated a natural product, human polyclonal antibody, obtained as fresh frozen plasma (FFP) from a HPS survivor. Second, we used DNA vaccine technology to manufacture a polyclonal immunoglobulin-based product that could be purified from the eggs of vaccinated ducks (Anas platyrhynchos). The natural “despeciation" of the duck IgY (i.e., Fc removed) results in an immunoglobulin predicted to be minimally reactogenic in humans. Administration of ≥5,000 neutralizing antibody units (NAU)/kg of FFP-protected hamsters from lethal disease when given up to 8 days after intranasal ANDV challenge. IgY/IgYΔFc antibodies purified from the eggs of DNA-vaccinated ducks effectively neutralized ANDV in vitro as measured by plaque reduction neutralization tests (PRNT). Administration of 12,000 NAU/kg of duck egg-derived IgY/IgYΔFc protected hamsters when administered up to 8 days after intranasal challenge and 5 days after intramuscular challenge. These experiments demonstrate that convalescent FFP shows promise as a postexposure HPS prophylactic. Moreover, these data demonstrate the feasibility of using DNA vaccine technology coupled with the duck/egg system to manufacture a product that could supplement or replace FFP. The DNA vaccine-duck/egg system can be scaled as needed and obviates the necessity of using limited blood products obtained from a small number of HPS survivors. This is the

  2. Co-administration of Interleukin-2 Enhances Cellular and Humoral Immune Responses to HIV Vaccine DNA Prime/MVA Boost Regime

    Institute of Scientific and Technical Information of China (English)

    JIANG Chun-lai; YU Xiang-hui; WU Yong-ge; LI Wei; KONG Wei

    2005-01-01

    Interleukine-2(IL-2) is a growth factor for antigen-stimulated T lymphocytes and is responsible for T-cell clonal expansion after antigen recognition. It has been demonstrated that DNA vaccine-elicited immune responses in mice could be augmented substantially by using either an IL-2 protein or a plasmid expressing IL-2. Twenty mice, divided into four experimental groups, were immunized with: (1) sham plasmid; (2) HIV-1 DNA vaccine alone; (3) HIV-1 DNA vaccine and IL-2 protein; or (4) HIV-1 DNA vaccine and IL-2 plasmid, separately. All the groups were immunized 3 times at a 2-week interval. Fourteen days after the last DNA vaccine injection, recombinant MVA was injected into all the mice except those in group 1. ELISA and ELISPOT were employed to investigate the effect of IL-2 on DNA vaccine immune responses. The obtained results strongly indicate that the efficacy of HIV vaccine can be enhanced by co-administration of a plasmid encoding IL-2.

  3. Multiple linear B-cell epitopes of classical swine fever virus glycoprotein E2 expressed in E.coli as multiple epitope vaccine induces a protective immune response

    Directory of Open Access Journals (Sweden)

    Wei Jian-Chao

    2011-07-01

    Full Text Available Abstract Classical swine fever is a highly contagious disease of swine caused by classical swine fever virus, an OIE list A pathogen. Epitope-based vaccines is one of the current focuses in the development of new vaccines against classical swine fever virus (CSFV. Two B-cell linear epitopes rE2-ba from the E2 glycoprotein of CSFV, rE2-a (CFRREKPFPHRMDCVTTTVENED, aa844-865 and rE2-b (CKEDYRYAISSTNEIGLLGAGGLT, aa693-716, were constructed and heterologously expressed in Escherichia coli as multiple epitope vaccine. Fifteen 6-week-old specified-pathogen-free (SPF piglets were intramuscularly immunized with epitopes twice at 2-week intervals. All epitope-vaccinated pigs could mount an anamnestic response after booster vaccination with neutralizing antibody titers ranging from 1:16 to 1:256. At this time, the pigs were subjected to challenge infection with a dose of 1 × 106 TCID50 virulent CSFV strain. After challenge infection, all of the rE2-ba-immunized pigs were alive and without symptoms or signs of CSF. In contrast, the control pigs continuously exhibited signs of CSF and had to be euthanized because of severe clinical symptoms at 5 days post challenge infection. The data from in vivo experiments shown that the multiple epitope rE2-ba shown a greater protection (similar to that of HCLV vaccine than that of mono-epitope peptide(rE2-a or rE2-b. Therefore, The results demonstrated that this multiple epitope peptide expressed in a prokaryotic system can be used as a potential DIVA (differentiating infected from vaccinated animals vaccine. The E.coli-expressed E2 multiple B-cell linear epitopes retains correct immunogenicity and is able to induce a protective immune response against CSFV infection.

  4. Immunogenicity and protective efficacy of Vibrio harveyi pcFlaA DNA vaccine in Epinephelus awoara

    Institute of Scientific and Technical Information of China (English)

    QIN Yingxue; SU Yongquan; WANG Shifeng; YAN Qingpi

    2009-01-01

    The FlaA gene from Vibrio harveyi, with a short nucleotide sequence encoding the Flag marker, was cloned into the eukaryotic expression vector pcDNA3.1(+) (designated as pcFlaA). Ninety grouper (Epinephelus awoara) were separated into three equal size groups. An experimental group was immunized with pcFlaA, Control I group was immunized with the vector pcDNA3.1(+), and Control II group was immunized with PBS. The expression of pcFlaA mRNA and protein was examined using reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemistry. We also evaluated the immunogenicity and protective efficacy of pcFlaA against V. harveyi by measuring the lymphocyte proliferation response and serum levels of specific antibody and conducting a bacterial challenge test. We successfully transfected the fish muscle with pcFlaA. The pcFlaA mRNA and protein was expressed in the muscle cells for up to one month following injection. The proliferation response of lymphocytes in fish immunized with pcFlaA was significantly higher than in control group II. Furthermore, the immunized fish generated specific antibody. The vaccination also resulted in significantly higher survival during the bacterial challenge test.

  5. DENGUE VACCINES.

    Science.gov (United States)

    Thisyakorn, Usa; Thisyakorn, Chule

    2015-01-01

    The uniqueness of the dengue viruses (DENVs) and the spectrum of disease resulting from infection have made dengue vaccine development difficult. Several vaccine candidates are currently being evaluated in clinical studies. The candidate currently at the most advanced clinical development stage, a live-attenuated tetravalent vaccine based on the chimeric yellow fever-dengue virus (CYD-TDV), has progressed to Phase 3 efficacy studies. Several other live-attenuated vaccines, as well as subunit, DNA, and purified inactivated vaccine candidates are at earlier stages of clinical development. Additional technological approaches, such as virus-vectored and Virus-Like Particles (VLP)-based vaccines are under evaluation in preclinical studies.

  6. A DNA vaccine against extracellular domains 1-3 of flk-1 and its immune preventive and therapeutic effects against H22 tumor cell in vivo

    Institute of Scientific and Technical Information of China (English)

    Fan Lü; Zhao-Yin Qin; Wen-Bin Yang; Yin-Xin Qi; Yi-Min Li

    2004-01-01

    AIM: To construct a DNA vaccine against extracellular domains 1-3 of fetal liver kinase-1 (flk-1), and to investigate its preventive and therapeutic effect against H22 cellin vivo.METHODS: Flk-1 DNA vaccine was produced by cloning extracellular domains 1-3 of flk-1 and by inserting the cloned gene into pcDNA3.1 (+). Fifteen mice were divided into 3 groups and inoculated by vaccine, plasmid and saline respectively to detect specific T lymphocyte response. Thirty Mice were equally divided into preventive group and therapeutic group. Preventive group was further divided into V, P, and S subgroups, namely immunized by vaccine,pcDNA3.1 (+) and saline, respectively, and attacked by H22 cell. Therapeutical group was divided into 3 subgroups of V, P and S, and attacked by H22, then treated with vaccine, pcDNA3.1 (+) and saline, respectively. The tumor size, tumor weight, mice survival time and tumor latency period were compared within these groups. Furthermore,intratumoral microvessel density (MVD) was assessed by immunohistochemistry.RESULTS: DNA vaccine pcDNA3.1 (+) flk-1-domains 1-3 was successfully constructed and could raise specific CTL activity. In the preventive group and therapeutic group,tumor latency period and survival time were significantly longer in vaccine subgroup than that in P and S subgroups (P<0.05); the tumor size, weight and MVD were significantly less in vaccine subgroup than that in P and S subgroups (P<0.05). The survival time of therapeutic vaccine subgroup was significantly shorter than that of preventive vaccine subgroup (P<0.05); the tumor size, and MVD of therapeutic vaccine subgroup were significantly greater than that of preventive vaccine subgroup (P<0.05).CONCLUSION: DNA vaccine against flk-1 domains 1-3 can stimulate potent specific CTL activity; and has distinctive prophylactic effect on tumor H22; and also can inhibit the tumor growthin vivo. This vaccine may be used as an adjuvant therapy because it is less effective on

  7. Altering an Artificial Gagpolnef Polyprotein and Mode of ENV Co-Administration Affects the Immunogenicity of a Clade C HIV DNA Vaccine

    OpenAIRE

    Katharina Böckl; Jens Wild; Simon Bredl; Kathrin Kindsmüller; Josef Köstler; Ralf Wagner

    2012-01-01

    HIV-1 candidate vaccines expressing an artificial polyprotein comprising Gag, Pol and Nef (GPN) and a secreted envelope protein (Env) were shown in recent Phase I/II clinical trials to induce high levels of polyfunctional T cell responses; however, Env-specific responses clearly exceeded those against Gag. Here, we assess the impact of the GPN immunogen design and variations in the formulation and vaccination regimen of a combined GPN/Env DNA vaccine on the T cell responses against the variou...

  8. A cDNA Clone-Launched Platform for High-Yield Production of Inactivated Zika Vaccine

    Directory of Open Access Journals (Sweden)

    Yujiao Yang

    2017-03-01

    Full Text Available A purified inactivated vaccine (PIV using the Zika virus (ZIKV Puerto Rico strain PRVABC59 showed efficacy in monkeys, and is currently in a phase I clinical trial. High-yield manufacture of this PIV is essential for its development and vaccine access. Here we report an infectious cDNA clone-launched platform to maximize its yield. A single NS1 protein substitution (K265E was identified to increase ZIKV replication on Vero cells (a cell line approved for vaccine production for both Cambodian FSS13025 and Puerto Rico PRVABC59 strains. The NS1 mutation did not affect viral RNA synthesis, but significantly increased virion assembly through an increased interaction between NS1 and NS2A (a known regulator of flavivirus assembly. The NS1 mutant virus retained wild-type virulence in the A129 mouse model, but decreased its competence to infect Aedes aegypti mosquitoes. To further increase virus yield, we constructed an infectious cDNA clone of the clinical trial PIV strain PRVABC59 containing three viral replication-enhancing mutations (NS1 K265E, prM H83R, and NS3 S356F. The mutant cDNA clone produced >25-fold more ZIKV than the wild-type parent on Vero cells. This cDNA clone-launched manufacture platform has the advantages of higher virus yield, shortened manufacture time, and minimized chance of contamination.

  9. A Mage3/Heat Shock Protein70 DNA vaccine induces both innate and adaptive immune responses for the antitumor activity.

    Science.gov (United States)

    Wang, Lifeng; Rollins, Lisa; Gu, Qinlong; Chen, Si-Yi; Huang, Xue F

    2009-12-11

    Heat shock proteins (HSPs) are highly effective and versatile molecules in promoting antitumor immune responses. We tested whether a HSP-based DNA vaccine can induce effective immune response against Mage3, a cancer testis (CT) antigen frequently expressed in many human tumors, thereby controlling the Mage3-expressing tumor. The vaccine was constructed by linking human inducible HSP70 to the C-terminus of a modified Mage3 gene (sMage3) that was attached at its N-terminus with the signal leader sequence of the human RANTES for releasing the expressed fusion protein from the transduced cells. Intramuscular injection of sMage3Hsp DNA induced CD4(+)/CD8(+) T cell and antibody responses. Vaccination with sMage3Hsp DNA was more effective in inhibiting Mage3-expressing TC-1 tumors. When we dissected the antitumor activity of CD4(+) and CD8(+) T cells by immunizing CD4(+) and CD8(+) knockout mice with sMage3Hsp DNA, we found that both CD8(+) T and CD4(+) T cells played a role in control of inoculated tumor, but did not constitute the whole of immune protection in the prophylactic immunization. Instead, depletion of natural killer (NK) cells led to a major loss of antitumor activity in the immunized mice. These results indicate that the HSP-based Mage3 DNA vaccine can more effectively inhibit tumor growth by inducing both the innate immune responses and Mage3-specific adaptive immune responses via the Hsp-associated adjuvant function.

  10. Efficacy of a glycoprotein DNA vaccine against viral haemorrhagic septicaemia (VHS) in Pacific herring, Clupea pallasii Valenciennes

    Science.gov (United States)

    Hart, L.M.; Lorenzen, Niels; LaPatra, S.E.; Grady, C.A.; Roon, S.E.; O’Reilly, J.; Gregg, J.L.; Hershberger, P.K.

    2012-01-01

    Viral haemorrhagic septicaemia virus (VHSV) and its associated disease state, viral haemorrhagic septicaemia (VHS), is hypothesized to be a proximate factor accounting for the decline and failed recovery of Pacific herring populations in Prince William Sound, AK (Marty et al. 1998, 2003, 2010). Survivors of laboratory-induced VHSV epizootics develop resistance to subsequent viral exposure (Kocan et al. 2001; Hershberger et al. 2007, 2010), which is likely the result of immune system recognition of the viral glycoprotein (G) (Lecocq-Xhonneux et al. 1994), a surface antigen that contains neutralizing epitopes (Lorenzen, Olesen & Jorgensen 1990; Jørgensen et al. 1995) and cell attachment domains (Lecocq-Xhonneux et al. 1994; Estepa & Coll 1996). These properties have proven useful in the development of G-gene-based DNA vaccines for VHSV and a related rhabdovirus, infectious haematopoietic necrosis virus (IHNV) (Anderson et al. 1996; Heppell et al. 1998; Corbeil et al. 1999; Einer-Jensen et al. 2009). Rainbow trout fingerlings, Oncorhynchus mykiss (Walbaum), vaccinated with 1 µg of either the VHS or IHN vaccine are protected from VHS when exposed to virus as early as 4 days (44 degree days) post-vaccination (p.v.) (Lorenzen et al. 2002). At later time points (80 days p.v.; 880 degree days), the level of cross-protection against VHS by IHN vaccination is either completely lost (60 days p.v.; 660 degree days) (3 g rainbow trout; 1 µg vaccine dose) (Lorenzen et al. 2002) or present at intermediate levels (6.5 g rainbow trout; 1 µg vaccine dose) (Einer-Jensen et al. 2009). Comparatively, VHS vaccination remains effective as long as 9 months (2520 degree days) p.v. (100 g rainbow trout; 0.5 µg vaccine dose) (McLauchlan et al. 2003). These results suggest that IHN and VHS vaccination activate a rapid transitory innate immune response against VHSV that is followed by long-term adaptive immunity in VHS-vaccinated trout (Lorenzen et al. 2002).

  11. Head-to-Head Comparison of Three Vaccination Strategies Based on DNA and Raw Insect-Derived Recombinant Proteins against Leishmania

    Science.gov (United States)

    Núñez, María del Carmen; Laurenti, Márcia D.; Gómez-Sebastián, Silvia; Rodríguez, Fernando; Pérez-Martín, Eva; Escribano, José M.

    2012-01-01

    Parasitic diseases plague billions of people among the poorest, killing millions annually, and causing additional millions of disability-adjusted life years lost. Leishmaniases affect more than 12 million people, with over 350 million people at risk. There is an urgent need for efficacious and cheap vaccines and treatments against visceral leishmaniasis (VL), its most severe form. Several vaccination strategies have been proposed but to date no head-to-head comparison was undertaken to assess which is the best in a clinical model of the disease. We simultaneously assayed three vaccination strategies against VL in the hamster model, using KMPII, TRYP, LACK, and PAPLE22 vaccine candidate antigens. Four groups of hamsters were immunized using the following approaches: 1) raw extracts of baculovirus-infected Trichoplusia ni larvae expressing individually one of the four recombinant proteins (PROT); 2) naked pVAX1 plasmids carrying the four genes individually (DNA); 3) a heterologous prime-boost (HPB) strategy involving DNA followed by PROT (DNA-PROT); and 4) a Control including empty pVAX1 plasmid followed by raw extract of wild-type baculovirus-infected T. ni larvae. Hamsters were challenged with L. infantum promastigotes and maintained for 20 weeks. While PROT vaccine was not protective, DNA vaccination achieved protection in spleen. Only DNA-PROT vaccination induced significant NO production by macrophages, accompanied by a significant parasitological protection in spleen and blood. Thus, the DNA-PROT strategy elicits strong immune responses and high parasitological protection in the clinical model of VL, better than its corresponding naked DNA or protein versions. Furthermore, we show that naked DNA coupled with raw recombinant proteins produced in insect larvae biofactories –the cheapest way of producing DNA-PROT vaccines– is a practical and cost-effective way for potential “off the shelf” supplying vaccines at very low prices for the protection against

  12. Development and characterization of an infectious cDNA clone of the modified live virus vaccine strain of equine arteritis virus.

    Science.gov (United States)

    Zhang, Jianqiang; Go, Yun Young; Huang, Chengjin M; Meade, Barry J; Lu, Zhengchun; Snijder, Eric J; Timoney, Peter J; Balasuriya, Udeni B R

    2012-08-01

    A stable full-length cDNA clone of the modified live virus (MLV) vaccine strain of equine arteritis virus (EAV) was developed. RNA transcripts generated from this plasmid (pEAVrMLV) were infectious upon transfection into mammalian cells, and the resultant recombinant virus (rMLV) had 100% nucleotide identity to the parental MLV vaccine strain of EAV. A single silent nucleotide substitution was introduced into the nucleocapsid gene (pEAVrMLVB), enabling the cloned vaccine virus (rMLVB) to be distinguished from parental MLV vaccine as well as other field and laboratory strains of EAV by using an allelic discrimination real-time reverse transcription (RT)-PCR assay. In vitro studies revealed that the cloned vaccine virus rMLVB and the parental MLV vaccine virus had identical growth kinetics and plaque morphologies in equine endothelial cells. In vivo studies confirmed that the cloned vaccine virus was very safe and induced high titers of neutralizing antibodies against EAV in experimentally immunized horses. When challenged with the heterologous EAV KY84 strain, the rMLVB vaccine virus protected immunized horses in regard to reducing the magnitude and duration of viremia and virus shedding but did not suppress the development of signs of EVA, although these were reduced in clinical severity. The vaccine clone pEAVrMLVB could be further manipulated to improve the vaccine efficacy as well as to develop a marker vaccine for serological differentiation of EAV naturally infected from vaccinated animals.

  13. Boosting BCG-primed mice with chimeric DNA vaccine HG856A induces potent multifunctional T cell responses and enhanced protection against Mycobacterium tuberculosis.

    Science.gov (United States)

    Ji, Ping; Hu, Zhi-Dong; Kang, Han; Yuan, Qin; Ma, Hui; Wen, Han-Li; Wu, Juan; Li, Zhong-Ming; Lowrie, Douglas B; Fan, Xiao-Yong

    2016-02-01

    The tuberculosis pandemic continues to rampage despite widespread use of the current Bacillus Calmette-Guerin (BCG) vaccine. Because DNA vaccines can elicit effective antigen-specific immune responses, including potent T cell-mediated immunity, they are promising vehicles for antigen delivery. In a prime-boost approach, they can supplement the inadequate anti-TB immunological memory induced by BCG. Based on this, a chimeric DNA vaccine HG856A encoding Mycobacterium tuberculosis (M. tuberculosis) immunodominant antigen Ag85A plus two copies of ESAT-6 was constructed. Potent humoral immune responses, as well as therapeutic effects induced by this DNA vaccine, were observed previously in M. tuberculosis-infected mice. In this study, we further evaluated the antigen-specific T cell immune responses and showed that repeated immunization with HG856A gave modest protection against M. tuberculosis challenge infection and significantly boosted the immune protection primed by BCG vaccination. Enhanced protection was accompanied by increased multifunctional Th1 CD4(+) T cell responses, most notably by an elevated frequency of M. tuberculosis antigen-specific IL-2-producing CD4(+) T cells post-vaccination. These data confirm the potential of chimeric DNA vaccine HG856A as an anti-TB vaccine candidate.

  14. Comparison of different prime-boost regimes with DNA and recombinant Orf virus based vaccines expressing glycoprotein D of pseudorabies virus in pigs

    NARCIS (Netherlands)

    Rooij, van E.M.A.; Rijsewijk, F.A.M.; Moonen-Leusen, H.W.; Bianchi, A.T.J.; Rziha, H.J.

    2010-01-01

    Both DNA and Orf virus (ORFV; Parapox virus) based vaccines have shown promise as alternatives for conventional vaccines in pigs against pseudorabies virus (PRV) infection causing Aujeszky's disease. In the present study we evaluated the efficacy of different prime-boost regimes in pigs in terms of

  15. DNA vaccination partially protects against African swine fever virus lethal challenge in the absence of antibodies.

    Directory of Open Access Journals (Sweden)

    Jordi M Argilaguet

    Full Text Available The lack of available vaccines against African swine fever virus (ASFV means that the evaluation of new immunization strategies is required. Here we show that fusion of the extracellular domain of the ASFV Hemagglutinin (sHA to p54 and p30, two immunodominant structural viral antigens, exponentially improved both the humoral and the cellular responses induced in pigs after DNA immunization. However, immunization with the resulting plasmid (pCMV-sHAPQ did not confer protection against lethal challenge with the virulent E75 ASFV-strain. Due to the fact that CD8(+ T-cell responses are emerging as key components for ASFV protection, we designed a new plasmid construct, pCMV-UbsHAPQ, encoding the three viral determinants above mentioned (sHA, p54 and p30 fused to ubiquitin, aiming to improve Class I antigen presentation and to enhance the CTL responses induced. As expected, immunization with pCMV-UbsHAPQ induced specific T-cell responses in the absence of antibodies and, more important, protected a proportion of immunized-pigs from lethal challenge with ASFV. In contrast with control pigs, survivor animals showed a peak of CD8(+ T-cells at day 3 post-infection, coinciding with the absence of viremia at this time point. Finally, an in silico prediction of CTL peptides has allowed the identification of two SLA I-restricted 9-mer peptides within the hemagglutinin of the virus, capable of in vitro stimulating the specific secretion of IFNγ when using PBMCs from survivor pigs. Our results confirm the relevance of T-cell responses in protection against ASF and open new expectations for the future development of more efficient recombinant vaccines against this disease.

  16. DNA vaccination partially protects against African swine fever virus lethal challenge in the absence of antibodies.

    Science.gov (United States)

    Argilaguet, Jordi M; Pérez-Martín, Eva; Nofrarías, Miquel; Gallardo, Carmina; Accensi, Francesc; Lacasta, Anna; Mora, Mercedes; Ballester, Maria; Galindo-Cardiel, Ivan; López-Soria, Sergio; Escribano, José M; Reche, Pedro A; Rodríguez, Fernando

    2012-01-01

    The lack of available vaccines against African swine fever virus (ASFV) means that the evaluation of new immunization strategies is required. Here we show that fusion of the extracellular domain of the ASFV Hemagglutinin (sHA) to p54 and p30, two immunodominant structural viral antigens, exponentially improved both the humoral and the cellular responses induced in pigs after DNA immunization. However, immunization with the resulting plasmid (pCMV-sHAPQ) did not confer protection against lethal challenge with the virulent E75 ASFV-strain. Due to the fact that CD8(+) T-cell responses are emerging as key components for ASFV protection, we designed a new plasmid construct, pCMV-UbsHAPQ, encoding the three viral determinants above mentioned (sHA, p54 and p30) fused to ubiquitin, aiming to improve Class I antigen presentation and to enhance the CTL responses induced. As expected, immunization with pCMV-UbsHAPQ induced specific T-cell responses in the absence of antibodies and, more important, protected a proportion of immunized-pigs from lethal challenge with ASFV. In contrast with control pigs, survivor animals showed a peak of CD8(+) T-cells at day 3 post-infection, coinciding with the absence of viremia at this time point. Finally, an in silico prediction of CTL peptides has allowed the identification of two SLA I-restricted 9-mer peptides within the hemagglutinin of the virus, capable of in vitro stimulating the specific secretion of IFNγ when using PBMCs from survivor pigs. Our results confirm the relevance of T-cell responses in protection against ASF and open new expectations for the future development of more efficient recombinant vaccines against this disease.

  17. An Epitope-Substituted DNA Vaccine Improves Safety and Immunogenicity against Dengue Virus Type 2.

    Directory of Open Access Journals (Sweden)

    Chung-Tao Tang

    Full Text Available Dengue virus (DENV, a global disease, is divided into four serotypes (DENV1-4. Cross-reactive and non-neutralizing antibodies against envelope (E protein of DENV bind to the Fcγ receptors (FcγR of cells, and thereby exacerbate viral infection by heterologous serotypes via antibody-dependent enhancement (ADE. Identification and modification of enhancing epitopes may mitigate enhancement of DENV infection. In this study, we characterized the cross-reactive DB21-6 and DB39-2 monoclonal antibodies (mAbs against domain I-II of DENV; these antibodies poorly neutralized and potently enhanced DENV infection both in vitro and in vivo. In addition, two enhancing mAbs, DB21-6 and DB39-2, were observed to compete with sera antibodies from patients infected with dengue. The epitopes of these enhancing mAbs were identified using phage display, structural prediction, and mapping of virus-like particle (VLP mutants. N8, R9, V12, and E13 are the reactive residues of DB21-6, while N8, R9, and E13 are the reactive residues of DB39-2. N8 substitution tends to maintain VLP secretion, and decreases the binding activity of DB21-6 and DB39-2. The immunized sera from N8 substitution (N8R DNA vaccine exerted greater neutralizing and protective activity than wild-type (WT-immunized sera, both in vitro and in vivo. Furthermore, treatment with N8R-immunized sera reduced the enhancement of mortality in AG129 mice. These results support identification and substitution of enhancing epitope as a novel strategy for developing safe dengue vaccines.

  18. Formulation, characterization, and expression of a recombinant MOMP Chlamydia trachomatis DNA vaccine encapsulated in chitosan nanoparticles.

    Science.gov (United States)

    Cambridge, Chino D; Singh, Shree R; Waffo, Alain B; Fairley, Stacie J; Dennis, Vida A

    2013-01-01

    Chlamydia trachomatis is a bacterial sexually transmitted infection affecting millions of people worldwide. Previous vaccination attempts have employed the recombinant major outer membrane protein (MOMP) of C. trachomatis nonetheless, with limited success, perhaps, due to stability, degradation, and delivery issues. In this study we cloned C. trachomatis recombinant MOMP DNA (DMOMP) and encapsulated it in chitosan nanoparticles (DMCNP) using the complex coacervation technique. Physiochemical characterizations of DMCNP included transmission and scanning electron microcopy, Fourier transform infrared and ultraviolet-visible spectroscopy, and zeta potential. Encapsulated DMOMP was 167-250 nm, with a uniform spherical shape and homogenous morphology, and an encapsulation efficiency > 90%. A slow release pattern of encapsulated DMOMP, especially in acidic solution, was observed over 7 days. The zeta potential of DMCNP was ~8.80 mV, which indicated that it was highly stable. Toxicity studies of DMCNP (25-400 μg/mL) to Cos-7 cells using the MTT assay revealed minimal toxicity over 24-72 hours with >90% viable cells. Ultra-violet visible (UV-vis) spectra indicated encapsulated DMOMP protection by chitosan, whereas agarose gel electrophoresis verified its protection from enzymatic degradation. Expression of MOMP protein in DMCNP-transfected Cos-7 cells was demonstrated via Western blotting and immunofluorescence microscopy. Significantly, intramuscular injection of BALB/c mice with DMCNP confirmed the delivery of encapsulated DMOMP, and expression of the MOMP gene transcript in thigh muscles and spleens. Our data show that encapsulation of DMOMP in biodegradable chitosan nanoparticles imparts stability and protection from enzymatic digestion, and enhances delivery and expression of DMOMP in vitro and in mice. Further investigations of the nanoencapsulated DMCNP vaccine formulation against C. trachomatis in mice are warranted.

  19. DNA vaccine encoding type IV pilin of Actinobacillus pleuropneumoniae induces strong immune response but confers limited protective efficacy against serotype 2 challenge.

    Science.gov (United States)

    Lu, Yu-Chun; Li, Min-Chen; Chen, Yi-Min; Chu, Chun-Yen; Lin, Shuen-Fuh; Yang, Wen-Jen

    2011-10-13

    Actinobacillus pleuropneumoniae is a gram-negative bacterial pathogen that causes swine pleuropneumonia, a highly contagious and often fatal disease that occurs worldwide. Our previous study showed that DNA vaccines encoding Apx exotoxin structural proteins ApxIA and/or ApxIIA, are a promising novel approach for immunization against the lethal challenge of A. pleuropneumoniae serotype 1. Vaccination against A. pleuropneumoniae is impeded by the lack of vaccines inducing reliable cross-serotype protection. Type IV fimbrial protein ApfA has been shown to be present and highly conserved in various serotypes of A. pleuropneumoniae. A novel DNA vaccine encoding ApfA (pcDNA-apfA) was constructed to evaluate the protective efficacy against infection with A. pleuropneumoniae serotype 2. A significant antibody response against pilin was generated following pcDNA-apfA immunization, suggesting that it was expressed in vivo. The IgG subclass (IgG1 and IgG2a) analysis indicates that the pcDNA-apfA vaccine induces both Th1 and Th2 immune responses. The IgA analysis shows that mucosal immunity could be enhanced by this DNA vaccine. Nevertheless, the strong antibody response induced by pcDNA-apfA vaccine only provided limited 30% protective efficacy against the serotype 2 challenge. These results in this study do not coincide with that the utility of type IV pilin is a good vaccine candidate against other infectious pathogens. It indicates that pilin should play a limited role in the development of a vaccine against A. pleuropneumoniae infection. Copyright © 2011 Elsevier Ltd. All rights reserved.

  20. Optimized codon usage enhances the expression and immunogenicity of DNA vaccine encoding Taenia solium oncosphere TSOL18 gene.