PCR. Monkeys were challenged with MPOV on day 0. †, fatality . VOL. 78, 2004 SMALLPOX DNA VACCINE AGAINST LETHAL MONKEYPOX 4441 FIG. 5. Antibody...01 MAY 2004 2. REPORT TYPE N/A 3. DATES COVERED - 4. TITLE AND SUBTITLE Smallpox DNA vaccine protects nonhuman primates against lethal ...vaccinia virus genes (L1R, A27L, A33R, and B5R) were protected from severe disease after an otherwise lethal challenge with monkeypox virus. Animals
Liu, Jing; Wu, Jianqi; Wang, Bing; Zeng, Sheng; Qi, Feifei; Lu, Changlong; Kimura, Yoshinobu; Liu, Beixing
It is well accepted that vaccination by oral administration has many advantages over injected parenteral immunization. The present study focuses on whether oral vaccination with a DNA vaccine could induce protective immunity against respiratory challenge infection. The M1 gene of influenza A virus was used to construct DNA vaccine using pcDNA 3.1(+) plasmid, a eukaryotic expression vector. The cationic liposomes were used to deliver the constructed DNA vaccine. In vitro and in vivo expression of M1 gene was observed in the cell line and in the intestine of orally vaccinated C57BL/6 mice, respectively. It became clear that this type of oral DNA vaccination was capable of inducing both humoral and cellular immune responses, together with an augmentation of IFN-γ production. In addition, oral vaccination with liposome-encapsulated DNA vaccine could protect the mice against respiratory challenge infection. These results suggest that gastrointestinal tract, a constituent member of the common mucosal immune system, is a potent candidate applicable as a DNA vaccine route against virus respiratory diseases.
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.
Wagemakers, A; Mason, L M K; Oei, A; de Wever, B; van der Poll, T; Bins, A D; Hovius, J W R
Borrelia afzelii is the predominant Borrelia species causing Lyme borreliosis in Europe. Currently there is no human vaccine against Lyme borreliosis, and most research focuses on recombinant protein vaccines against Borrelia burgdorferi sensu stricto. DNA tattooing is a novel vaccination method that can be applied in a rapid vaccination schedule. We vaccinated C3H/HeN mice with B. afzelii strain PKo OspC (outer-surface protein C) using a codon-optimized DNA vaccine tattoo and compared this with recombinant protein vaccination in a 0-2-4 week vaccination schedule. We also assessed protection by DNA tattoo in a 0-3-6 day schedule. DNA tattoo and recombinant OspC vaccination induced comparable total IgG responses, with a lower IgG1/IgG2a ratio after DNA tattoo. Two weeks after syringe-challenge with 5 × 10(5) B. afzelii spirochetes most vaccinated mice had negative B. afzelii tissue DNA loads and all were culture negative. Furthermore, DNA tattoo vaccination in a 0-3-6 day regimen also resulted in negative Borrelia loads and cultures after challenge. To conclude, DNA vaccination by tattoo was fully protective against B. afzelii challenge in mice in a rapid vaccination protocol, and induces a favorable humoral immunity compared to recombinant protein vaccination. Rapid DNA tattoo is a promising vaccination strategy against spirochetes.
Guo, Aijiang; Jin, Zhizhong; Zheng, Yadong; Hai, Gang; Yuan, Gailing; Li, Hailong; Cai, Xuepeng
A DNA vaccine, pcDNA3-B, was developed by using the nucleotide sequence of Taenia solium B antigen and cloning into pcDNA3.1 plasmid. The growing pigs were vaccinated by one intramuscular infection of 200 or 1000 microg pcDNA3-B. The immunization with 1000 microg of pcDNA3-B showed 92.6% protection when the pigs were challenged by T. solium eggs and four of the five pigs vaccinated had no viable cysts. The results provide encouraging information on the use of pcDNA3-B vaccination for the prevention of cysticercosis.
Emmenegger, E.J.; Kurath, G.
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 < 0.05) as compared to fish receiving a mock 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.
Wefer, Judit; Harris, Robert A; Lobell, Anna
DNA vaccines encoding encephalitogenic peptides protect against subsequent development of rat experimental autoimmune encephalomyelitis (EAE) through unknown mechanisms. We investigated immune cell phenotypes at different time points after DNA vaccination with vaccine encoding myelin oligodendrocyte glycoprotein peptide 91-108 and subsequent induction of EAE. In protected rats, we observed (i) no alterations in antigen-specific Th2 or Th3 responses, (ii) reduced MHC II expression on splenocytes early after EAE induction, (iii) antigen-specific upregulation of IFNbeta upon recall stimulation and (iv) reduced IL-12Rbeta2 on lymphocytes. We suggest that the underlying mechanism of DNA vaccination is associated with immunomodulation exerted by induced IFNbeta.
Albrecht, Mark T; Livingston, Brian D; Pesce, John T; Bell, Matt G; Hannaman, Drew; Keane-Myers, Andrea M
Electroporation of DNA vaccines represents a platform technology well positioned for the development of multivalent biodefense vaccines. To evaluate this hypothesis, three vaccine constructs were produced using codon-optimized genes encoding Bacillus anthracis Protective Antigen (PA), and the Yersinia pestis genes LcrV and F1, cloned into pVAX1. A/J mice were immunized on a prime-boost schedule with these constructs using the electroporation-based TriGrid Delivery System. Immunization with the individual pDNA vaccines elicited higher levels of antigen-specific IgG than when used in combination. DNA vaccine effectiveness was proven, the pVAX-PA titers were toxin neutralizing and fully protective against a lethal B. anthracis spore challenge when administered alone or co-formulated with the plague pDNA vaccines. LcrV and F1 pVAX vaccines against plague were synergistic, resulting in 100% survival, but less protective individually and when co-formulated with pVAX-PA. These DNA vaccine responses were Th1/Th2 balanced with high levels of IFN-γ and IL-4 in splenocyte recall assays, contrary to complimentary protein Alum vaccinations displaying a Th2 bias with increased IL-4 and low levels of IFN-γ. These results demonstrate the feasibility of electroporation to deliver and maintain the overall efficacy of an anthrax-plague DNA vaccine cocktail whose individual components have qualitative immunological differences when combined.
Chen, Yung-Yi; Hsieh, Ming Kun; Tung, Chun-Yu; Wu, Ching Ching; Lin, Tsang Long
The present study was undertaken to determine the kinetics of viral load and immune response in protection against infectious bursal disease virus (IBDV) by DNA vaccination. Chickens were DNA-vaccinated and challenged with IBDV one week after the third vaccination. Tissues were collected at 12 hours postinfection (HPI), 1 day postinfection (DPI), 3, 5, 7 and 10 DPI. The vaccinated chickens had less viral RNA, with delayed appearance and shorter duration in the bursa of Fabricius, spleen, and cecal tonsil than the challenged control chickens. Their ELISA and neutralizing antibody titers were decreased at 12 HPI and significantly lower (P < 0.05) than those in the challenged control chickens at later time points. Their spleen IFNγ expression was up-regulated compared to that in the DNA-vaccinated chickens without IBDV challenge. These results indicate that DNA vaccination confers protection against IBDV challenge by delayed appearance and rapid clearance of the invading viruses.
Xu, Yingying; Yuen, Pak-Wai; Lam, Jenny Ka-Wing
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. PMID:25014738
Song, Jae-Min; Kim, Yeu-Chun; O, Eunju; Compans, Richard W; Prausnitz, Mark R; Kang, Sang-Moo
In this study, we tested the hypothesis that DNA vaccination in the skin using microneedles improves protective immunity compared to conventional intramuscular (IM) injection of a plasmid DNA vaccine encoding the influenza hemagglutinin (HA). In vivo fluorescence imaging demonstrated the expression of a reporter gene delivered to the skin using a solid microneedle patch coated with plasmid DNA. Vaccination at a low dose (3 µg HA DNA) using microneedles generated significantly stronger humoral immune responses and better protective responses post-challenge compared to IM vaccination at either low or high (10 µg HA DNA) dose. Vaccination using microneedles at a high (10 µg) dose further generated improved post-challenge protection, as measured by survival, recall antibody-secreting cell responses in spleen and bone marrow, and interferon (IFN)-γ cytokine T-cell responses. This study demonstrates that DNA vaccination in the skin using microneedles induces higher humoral and cellular immune responses as well as improves protective immunity compared to conventional IM injection of HA DNA vaccine. PMID:22508490
Sepúlveda, Dagoberto; Lorenzen, Niels
DNA vaccines encoding viral glycoproteins have been very successful for induction of protective immunity against diseases caused by rhabdoviruses in cultured fish species. However, the vaccine concept is based on a single viral gene and since RNA viruses are known to possess high variability and adaptation capacity, this work aimed at evaluating whether viral haemorrhagic septicaemia virus (VHSV), an RNA virus and member of Rhabdoviridae family, was able to evade the protective immune response induced by the DNA vaccination of rainbow trout. The experiments comprised repeated passages of a highly pathogenic VHSV isolate in a fish cell line in the presence of neutralizing fish serum (in vitro approach), and in rainbow trout immunized with the VHS DNA vaccine (in vivo approach). For the in vitro approach, the virus collected from the last passage (passaged virus) was as sensitive as the parental virus to serum neutralization, suggesting that the passaging did not promote the selection of virus populations able to bypass the neutralization by serum antibodies. Also, in the in vivo approach, where virus was passaged several times in vaccinated fish, no increased virulence nor increased persistence in vaccinated fish was observed in comparison with the parental virus. However, some of the vaccinated fish did get infected and could transmit the infection to naïve cohabitant fish. The results demonstrated that the DNA vaccine induced a robust protection, but also that the immunity was non-sterile. It is consequently important not to consider vaccinated fish as virus free in veterinary terms.
Sepúlveda, Dagoberto; Lorenzen, Niels
DNA vaccines encoding viral glycoproteins have been very successful for induction of protective immunity against diseases caused by rhabdoviruses in cultured fish species. However, the vaccine concept is based on a single viral gene and since RNA viruses are known to possess high variability and adaptation capacity, this work aimed at evaluating whether viral haemorrhagic septicaemia virus (VHSV), an RNA virus and member of Rhabdoviridae family, was able to evade the protective immune response induced by the DNA vaccination of rainbow trout. The experiments comprised repeated passages of a highly pathogenic VHSV isolate in a fish cell line in the presence of neutralizing fish serum (in vitro approach), and in rainbow trout immunized with the VHS DNA vaccine (in vivo approach). For the in vitro approach, the virus collected from the last passage (passaged virus) was as sensitive as the parental virus to serum neutralization, suggesting that the passaging did not promote the selection of virus populations able to bypass the neutralization by serum antibodies. Also, in the in vivo approach, where virus was passaged several times in vaccinated fish, no increased virulence nor increased persistence in vaccinated fish was observed in comparison with the parental virus. However, some of the vaccinated fish did get infected and could transmit the infection to naïve cohabitant fish. The results demonstrated that the DNA vaccine induced a robust protection, but also that the immunity was non-sterile. It is consequently important not to consider vaccinated fish as virus free in veterinary terms. PMID:27054895
de las Heras, Ana I; Rodríguez Saint-Jean, S; Pérez-Prieto, Sara I
DNA vaccines and oral DNA-based immunotherapy against infectious pancreatic necrosis virus (IPNV) have scarcely been studied in salmonid fish. Here, a vector with the capsid VP2 gene inserted was encapsulated in alginate microspheres to avoid the aggressive gastrointestinal conditions experienced following oral administration. Alginate microspheres were effective to protect the pDNA encoding VP2, which was expressed early in different organs of the vaccinated trout and that persisted for at least 60 days. The vaccine induces innate immune responses, raising the expression of IFN more than 10-fold relative to the fish vaccinated with the empty plasmid, at 7 and 15 days post-vaccination. Likewise, maximal expression of the IFN-induced antiviral Mx protein was recorded 15 days post-vaccination and neutralizing antibodies were also detected after 15 days, although their titre rose further at 21 days post-vaccination. Protection was high in the immunized fish, which showed around an 80% relative survival when challenged 15 and 30 days after vaccine delivery. Very low viral load with respect to the control group was detected in the vaccinated fish that survived 45 days after challenge. Thus, this study demonstrates the potential of the encapsulation technique for IPNV-DNA vaccine delivery and the relevance of the IPNV-VP2 gene for future plasmid constructs.
Rout, Namita; Kumar, Sudhir; Jaganmohan, Shanmugam; Murugan, Vadivel
White Spot Syndrome Virus (WSSV) is a major cause of mortality in shrimp and poses a huge threat to aquaculture industry. Till now no comprehensive or individual strategy has been established to combat white spot disease. Previous efforts by other investigators have given insight of protein vaccination and its efficacy to protect shrimp against WSSV infection. In this study, we have explored the protective efficacy of DNA vaccination and tissue distribution of the immunised recombinant plasmid in black tiger shrimp (Penaeus monodon). Four recombinant constructs were generated by inserting four genes encoding the WSSV structural proteins VP15, VP28, VP35 and VP281 individually into DNA vaccine vector pVAX1. Expression of these proteins from the recombinant plasmids was confirmed in vitro in CHO cell lines. For vaccination experiments, shrimp were immunised with these DNA constructs and later challenged with WSSV. A significant level of protection was offered by the plasmids encoding VP28 or VP281 till 7 weeks whereas protein vaccination failed to protect vaccinated shrimp after 3 weeks of first immunisation. In addition, our tissue distribution study revealed the persistence of immunised DNA at least upto 2 months in the injected shrimp muscle. Thus, our results suggest that DNA vaccination strategy will have potential utility against WSSV infection in shrimp cultivation.
Lorenzen, N; Lorenzen, E; Einer-Jensen, K; LaPatra, S E
DNA vaccines based on the glycoprotein genes of the salmonid rhabdoviruses VHSV and IHNV have been demonstrated to be very efficient in inducing a protective immune response against the respective diseases in rainbow trout. Nanogram doses of plasmid DNA delivered by intramuscular injection are sufficient to induce high levels of immunity in fingerling-size fish, whereas larger fish require more vaccine for protection. The protection is long lasting and, more surprisingly, is partly established already 4 days post vaccination. The early protection involves cross-protective anti-viral defence mechanisms, while the long duration immunity is highly specific. The nature of these immune response mechanisms is discussed and it is suggested that the efficacy of the vaccines is related to their ability to activate the innate immune system as it is activated by live virus.
Porter, Kevin R; Ewing, Daniel; Chen, Lan; Wu, Shuenn-Jue; Hayes, Curtis G; Ferrari, Marilyn; Teneza-Mora, Nimfa; Raviprakash, Kanakatte
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.
Liu, Lei; Gong, Yu-Xin; Liu, Guang-Lu; Zhu, Bin; Wang, Gao-Xue
To reduce the economic losses caused by diseases in aquaculture industry, more efficient and economic prophylactic measures should be urgently investigated. In this research, the effects of a novel functionalized single-walled carbon nanotubes (SWCNTs) applied as a delivery vehicle for DNA vaccine administration in juvenile grass carp against Aeromonas hydrophila were studied. Our results showed that SWCNTs loaded with DNA vaccine induced a better protection to juvenile grass carp against A. hydrophila. Moreover, SWCNTs conjugated with DNA vaccine provided significantly protective immunity compared with free DNA vaccine. Thereby, SWCNTs may be considered as a potential efficient DNA vaccine carrier to enhance the immunological activity.
Ault, Alida; Zajac, Alyse M.; Kong, Wing-Pui; Gorres, J. Patrick; Royals, Michael; Wei, Chih-Jen; Bao, Saran; Yang, Zhi-yong; Reedy, Stephanie E.; Sturgill, Tracy L.; Page, Allen E.; Donofrio-Newman, Jennifer; Adams, Amanda A.; Balasuriya, Udeni B.R.; Horohov, David W.; Chambers, Thomas M.; Nabel, Gary J.; Rao, Srinivas S.
Equine influenza A (H3N8) virus is a leading cause of infectious respiratory disease in horses causing widespread morbidity and economic losses. As with influenza in other species, equine influenza strains continuously mutate, requiring constant re-evaluation of current vaccines and development of new vaccines. Current inactivated (killed) vaccines, while efficacious, only offer limited protection against multiple strains and require frequent boosts. Ongoing research into new vaccine technologies, including gene-based vaccines, aims to increase the neutralization potency, breadth, and duration of protective immunity of new or existing vaccines. In these hypothesis-generating experiments, we demonstrate that a DNA vaccine expressing the hemagglutinin protein of equine H3N8 influenza virus generates homologous and heterologous immune responses, and protects against clinical disease and viral replication following homologous H3N8 infection in horses. Furthermore, we demonstrate that a needle-free delivery device is as efficient and effective as conventional parenteral injection using a needle and syringe. The observed trends in this study drive the hypothesis that DNA vaccines offer a safe, effective, and promising alternative approach for veterinary vaccines against influenza, and applicable to combat equine influenza. PMID:22449425
Song, Xiaokai; Xu, Lixin; Yan, Ruofeng; Huang, Xinmei; Li, Xiangrui
The search for effective vaccines against chicken coccidiosis remains a challenge because of the complex organisms with multiple life cycle stages of Eimeria. Combination of T-cell epitopes from different stages of Eimeria life cycle could be an optimal strategy to overcome the antigen complexity of the parasite. In this study, 4 fragments with concentrated T-cell epitopes from the sporozoite antigen SO7 and the merozoite antigen MZ5-7 of Eimeria tenella were cloned into eukaryotic expression vector pVAX1 in different forms, with or without chicken cytokines IL-2 or IFN-γ genes as genetic adjuvants, to construct multistage, multi-epitope DNA vaccines against Eimeria tenella. Transcription and expression of the multi-epitope DNA vaccines in vivo were detected by reverse transcription-PCR (RT-PCR) and Western blot. On the basis of survival rate, lesion score, body weight gain, oocyst decrease ratio and the anti-coccidial index (ACI), Animal experiments were carried out to evaluate the protective efficacy against Eimeria tenella. Results showed the constructed DNA vaccines were transcribed and translated successfully in vivo. Animal experiment showed that the multi-epitopes DNA vaccines were more effective to stimulate immune response than single fragment. Compared with the DNA vaccines composed with less T-cell epitopes, DNA vaccine pVAX1-m1-m2-s1-s2 containing 4 fragments with concentrated T-epitopes provided the highest ACI of 180.39. DNA vaccines composed of antigens from two developmental stages were more effective than the single-stage ones. Especially DNA vaccine pVAX1-m1-m2-s1-s2 provided the most effective protection with the ACI of 180.39. Furthermore, cytokines IL-2 or IFN-γ could improve the efficacy of the multi-epitope DNA vaccines significantly. Overall, pVAX1-m1-m2-s1-s2-IFN-γ provided the most effective protection with the ACI of 189.92. The multi-epitope DNA vaccines revealed in this study provide new candidates for Eimeria vaccine development.
Microbiology. All Rights Reserved. Influences of Glycosylation on Antigenicity, Immunogenicity, and Protective Efficacy of Ebola Virus GP DNA Vaccines ...carbohydrates. We measured the influences of GP glycosylation on antigenicity, immu- nogenicity, and protection by testing DNA vaccines comprised of GP...may protect against filovirus infection (6, 39). Con- sistent with this, B-cell-deficient mice vaccinated with EBOV-like particles were not protected
Deliyannis, Georgia; Boyle, Jefferey S.; Brady, Jamie L.; Brown, Lorena E.; Lew, Andrew M.
Improving the immunological potency, particularly the Ab response, is a serious hurdle for the protective efficacy and hence broad application of DNA vaccines. We examined the immunogenicity and protective efficacy of a hemagglutinin-based influenza DNA vaccine that was targeted to antigen-presenting cells (APCs) by fusion to CTLA4. The targeted vaccine was shown to induce an accelerated and increased Ab response (as compared with those receiving the nontargeted control) that was predominated by IgG1 and recognized conformationally dependent viral epitopes. Moreover, mice receiving the APC-targeted DNA vaccine had significantly reduced viral titers (100-fold) after a nonlethal virus challenge. The increased protective efficacy was most likely because of increased Ab responses, as cytotoxic T lymphocyte responses were not enhanced. Targeting was demonstrated by direct binding studies of CTLA4 fusion proteins to the cognate ligand (B7; expressed on APCs in vivo). In addition, a targeted protein was detected at 4-fold higher levels in draining lymph nodes within 2-24 h of administration. Therefore, this study demonstrates that targeting DNA-encoded antigen to APCs results in enhanced immunity and strongly suggests that this approach may be useful in improving the protective efficacy of DNA vaccines.
Gorres, J Patrick; Lager, Kelly M; Kong, Wing-Pui; Royals, Michael; Todd, John-Paul; Vincent, Amy L; Wei, Chih-Jen; Loving, Crystal L; Zanella, Eraldo L; Janke, Bruce; Kehrli, Marcus E; Nabel, Gary J; Rao, Srinivas S
Swine influenza is a highly contagious viral infection in pigs that significantly impacts the pork industry due to weight loss and secondary infections. There is also the potential of a significant threat to public health, as was seen in 2009 when the pandemic H1N1 influenza virus strain emerged from reassortment events among avian, swine, and human influenza viruses within pigs. As classic and pandemic H1N1 strains now circulate in swine, an effective vaccine may be the best strategy to protect the pork industry and public health. Current inactivated-virus vaccines available for swine influenza protect only against viral strains closely related to the vaccine strain, and egg-based production of these vaccines is insufficient to respond to large outbreaks. DNA vaccines are a promising alternative since they can potentially induce broad-based protection with more efficient production methods. In this study we evaluated the potentials of monovalent and trivalent DNA vaccine constructs to (i) elicit both humoral and gamma interferon (IFN-γ) responses and (ii) protect pigs against viral shedding and lung disease after challenge with pandemic H1N1 or classic swine H1N1 influenza virus. We also compared the efficiency of a needle-free vaccine delivery method to that of a conventional needle/syringe injection. We report that DNA vaccination elicits robust serum antibody and cellular responses after three immunizations and confers significant protection against influenza virus challenge. Needle-free delivery elicited improved antibody responses with the same efficiency as conventional injection and should be considered for development as a practical alternative for vaccine administration.
Hosie, Margaret J.; Flynn, J. Norman; Rigby, Mark A.; Cannon, Celia; Dunsford, Thomas; Mackay, Nancy A.; Argyle, David; Willett, Brian J.; Miyazawa, Takayuki; Onions, David E.; Jarrett, Oswald; Neil, James C.
To test the potential of a multigene DNA vaccine against lentivirus infection, we generated a defective mutant provirus of feline immunodeficiency virus (FIV) with an in-frame deletion in pol (FIVΔRT). In a first experiment, FIVΔRT DNA was administered intramuscularly to 10 animals, half of which also received feline gamma interferon (IFN-γ) DNA. The DNA was administered in four 100-μg doses at 0, 10, and 23 weeks. Immunization with FIVΔRT elicited cytotoxic T-cell (CTL) responses to FIV Gag and Env in the absence of a serological response. After challenge with homologous virus at week 26, all 10 of the control animals became seropositive and viremic but 4 of the 10 vaccinates remained seronegative and virus free. Furthermore, quantitative virus isolation and quantitative PCR analysis of viral DNA in peripheral blood mononuclear cells revealed significantly lower virus loads in the FIVΔRT vaccinates than in the controls. Immunization with FIVΔRT in conjunction with IFN-γ gave the highest proportion of protected cats, with only two of five vaccinates showing evidence of infection following challenge. In a second experiment involving two groups (FIVΔRT plus IFN-γ and IFN-γ alone), the immunization schedule was reduced to 0, 4, and 8 weeks. Once again, CTL responses were seen prior to challenge in the absence of detectable antibodies. Two of five cats receiving the proviral DNA vaccine were protected against infection, with an overall reduction in virus load compared to the five infected controls. These findings demonstrate that DNA vaccination can elicit protection against lentivirus infection in the absence of a serological response and suggest the need to reconsider efficacy criteria for lentivirus vaccines. PMID:9696827
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
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.
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.
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
Garver, Kyle A.; Conway, Carla M.; Kurath, Gael
A highly efficacious DNA vaccine against a fish rhabdovirus, infectious hematopoietic necrosis virus (IHNV), was mutated to introduce two stop codons to prevent glycoprotein translation while maintaining the plasmid DNA integrity and RNA transcription ability. The mutated plasmid vaccine, denoted pIHNw-G2stop, when injected intramuscularly into fish at high doses, lacked detectable glycoprotein expression in the injection site muscle, and did not provide protection against lethal virus challenge 7 days post-vaccination. These results suggest that the G-protein itself is required to stimulate the early protective antiviral response observed after vaccination with the nonmutated parental DNA vaccine.
Garver, K.A.; Conway, C.M.; Kurath, G.
A highly efficacious DNA vaccine against a fish rhabdovirus, infectious hematopoietic necrosis virus (IHNV), was mutated to introduce two stop codons to prevent glycoprotein translation while maintaining the plasmid DNA integrity and RNA transcription ability. The mutated plasmid vaccine, denoted pIHNw-G2stop, when injected intramuscularly into fish at high doses, lacked detectable glycoprotein expression in the injection site muscle, and did not provide protection against lethal virus challenge 7 days post-vaccination. These results suggest that the G-protein itself is required to stimulate the early protective antiviral response observed after vaccination with the nonmutated parental DNA vaccine. ?? Springer Science+Business Media, Inc. 2006.
Tretyakova, Irina; Hearn, Jason; Wang, Eryu; Weaver, Scott; Pushko, Peter
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: firstname.lastname@example.org.
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
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.
Keane-Myers, Andrea M; Bell, Matt; Hannaman, Drew; Albrecht, Mark
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.
Chaudhari, Aparna; Pathakota, Gireesh-Babu; Annam, Pavan-Kumar
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.
LaPatra, S.E.; Corbeil, S.; Jones, G.R.; Shewmaker, W.D.; Lorenzen, N.; Anderson, E.D.; Kurath, G.
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??C. Nearly complete protection was also observed at later time points (7, 14, and 28 d) using a standardized waterborne challenge model. In a test of the specificity of this early protection, immunization of rainbow trout with a DNA vaccine against another fish rhabdovirus, viral hemorrhagic septicemia virus, provided a significant level of cross-protection against IHNV challenge for a transient period of time, whereas a rabies virus DNA vaccine was not protective. This indication of distinct early and late protective mechanisms was not dependent on DNA vaccine doses from 0.1 to 2.5 ??g. ?? 2001 Elsevier Science Ltd.
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. PMID:21933385
Millard, Elena V; Bourke, Ashley M; LaPatra, Scott E; Brenden, Travis O; Fitzgerald, Scott D; Faisal, Mohamed
A DNA vaccine containing the glycoprotein (G) gene of the North American viral hemorrhagic septicemia virus (VHSV) genotype IVb was developed to evaluate the immune response of fish following vaccination and evaluate its efficacy in protecting a susceptible species, the Muskellunge Esox masquinongy, against VHSV-IVb challenge. Seven weeks (539 degree-days) following vaccination with 10 μg of either pVHSivb-G or a control plasmid, Muskellunge were challenged by immersion with 10(5) plaque-forming units (pfu)/mL of VHSV-IVb. Fish vaccinated with pVHSivb-G had a relative percent survival (RPS) of 45%. Vaccinated fish also had significantly lower mean viral titers in tissues (4.2 × 10(2) pfu/g) and viral prevalence (4%) than fish receiving the plasmid control vaccine (3.3 × 10(5) pfu/g; 82%). Neutralizing antibodies were detected 28 d (308 degree-days) postchallenge (11 weeks postvaccination) in 100% of Muskellunge vaccinated with pVHSivb-G compared with only 12% of plasmid-control-vaccinated Muskellunge, suggesting robust induction of a secondary, adaptive immune response. In addition, pVHSivb-G-vaccinated Rainbow Trout Oncorhynchus mykiss challenged 7 d (100 degree-days) postvaccination with the heterologous novirhabdovirus, infectious hematopoietic necrosis virus (IHNV), experienced an RPS of 61%, compared to control fish, suggesting induction of an early and transient nonspecific antiviral immune response. This study provides an important starting point for VHSV-IVb vaccine development and useful information about the antiviral immune response elicited by DNA vaccination in a nondomesticated fish species. Received May 1, 2016; accepted September 1, 2016.
Kutzler, M A; Wise, M C; Hutnick, N A; Moldoveanu, Z; Hunter, M; Reuter, M A; Yuan, S; Yan, J; Ginsberg, A A; Sylvester, A; Pahar, B; Carnathan, D G; Kathuria, N; Khan, A S; Montefiori, D; Sardesai, N Y; Betts, M R; Mestecky, J; Marx, P A; Weiner, D B
There have been encouraging results for the development of an effective HIV vaccine. However, many questions remain regarding the quality of immune responses and the role of mucosal antibodies. We addressed some of these issues by using a simian immunodeficiency virus (SIV) DNA vaccine adjuvanted with plasmid-expressed mucosal chemokines combined with an intravaginal SIV challenge in rhesus macaque (RhM) model. We previously reported on the ability of CCR9 and CCR10 ligand (L) adjuvants to enhance mucosal and systemic IgA and IgG responses in small animals. In this study, RhMs were intramuscularly immunized five times with either DNA or DNA plus chemokine adjuvant delivered by electroporation followed by challenge with SIVsmE660. Sixty-eight percent of all vaccinated animals (P<0.01) remained either uninfected or had aborted infection compared with only 14% in the vaccine naïve group. The highest protection was observed in the CCR10L chemokines group, where six of nine animals had aborted infection and two remained uninfected, leading to 89% protection (P<0.001). The induction of mucosal SIV-specific antibodies and neutralization titers correlated with trends in protection. These results indicate the need to further investigate the contribution of chemokine adjuvants to modulate immune responses and the role of mucosal antibodies in SIV/HIV protection.
Hutnick, N A; Moldoveanu, Z; Hunter, M; Reuter, M; Yuan, S; Yan, J; Ginsberg, A; Sylvester, A; Pahar, B; Carnathan, D; Kathuria, N; Khan, A S; Montefiori, D; Sardesai, N Y; Betts, M R; Mestecky, J; Marx, P; Weiner, D B
There have been encouraging results for the development of an effective HIV vaccine. However, many questions remain regarding the quality of immune responses and the role of mucosal antibodies. We addressed some of these issues by using a simian immunodeficiency virus (SIV) DNA vaccine adjuvanted with plasmid-expressed mucosal chemokines combined with an intravaginal SIV challenge in rhesus macaque (RhM) model. We previously reported on the ability of CCR9 and CCR10 ligand (L) adjuvants to enhance mucosal and systemic IgA and IgG in small animals. In this study, RhMs were intramuscularly immunized five times with either DNA or DNA plus chemokine adjuvant delivered by electroporation followed by challenge with SIVsmE660. Sixty-eight percent of all vaccinated animals (P=0.0016) remained either uninfected or had aborted infection compared to only 14% in the vaccine naïve group. The highest protection was observed in the CCR10L chemokines group, where 6 of 9 animals had aborted infection and two remained uninfected, leading to 89% protection (P=0.0003). The induction of mucosal SIV-specific antibodies and neutralization titers correlated with trends in protection. These results indicate the need to further investigate the contribution of chemokine adjuvants to modulate immune responses and the role of mucosal antibodies in SIV/HIV protection. PMID:25943275
Fynan, Ellen F.; Webster, Robert G.; Fuller, Deborah H.; Haynes, Joel R.; Santoro, Joseph C.; Robinson, Harriet L.
Plasmid DNAs expressing influenza virus hemagglutinin glycoproteins have been tested for their ability to raise protective immunity against lethal influenza challenges of the same subtype. In trials using two inoculations of from 50 to 300 μg of purified DNA in saline, 67-95% of test mice and 25-63% of test chickens have been protected against a lethal influenza challenge. Parenteral routes of inoculation that achieved good protection included intramuscular and intravenous injections. Successful mucosal routes of vaccination included DNA drops administered to the nares or trachea. By far the most efficient DNA immunizations were achieved by using a gene gun to deliver DNA-coated gold beads to the epidermis. In mice, 95% protection was achieved by two immunizations with beads loaded with as little as 0.4 μg of DNA. The breadth of routes supporting successful DNA immunizations, coupled with the very small amounts of DNA required for gene-gun immunizations, highlight the potential of this remarkably simple technique for the development of subunit vaccines.
Long, Jian-Er; Huang, Li-Na; Qin, Zhi-Qiang; Wang, Wen-Yi; Qu, Di
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 co-immunized with plasmid expressing DuIFN-γ. DuIFN-γ mRNA 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 anti-DuIFN-γ antibodies neutralized the antiviral effects. DuIFN-γ in 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. PMID:16124047
Sylla, Seydou; Cong, Yan-Long; Sun, Yi-Xue; Yang, Gui-Lian; Ding, Xue-Mei; Yang, Zhan-Qing; Zhou, Yu-Long; Yang, Minnan; Wang, Chun-Feng; Ding, Zhuang
Post-weaning multisystemic wasting syndrome (PMWS) associated with porcine circovirus type 2 (PCV2) has caused the swine industry significant health challenges and economic damage. Although inactivated and subunit vaccines against PMWS have been used widely, so far no DNA vaccine is available. In this study, with the aim of exploring a new route for developing a vaccine against PCV2, the immunogenicity of a DNA vaccine was evaluated in mice. The pEGFP-N1 vector was used to construct a PCV2 Cap gene recombinant vaccine. To assess the immunogenicity of pEGFP-Cap, 80 BALB/c mice were immunized three times at 2 weekly intervals with pEGFP-Cap, LG-strain vaccine, pEGFP-N1 vector or PBS and then challenged with PCV2. IgG and cytokines were assessed by indirect ELISA and ELISA, respectively. Specimens stained with hematoxylin and eosin (HE) and immunohistochemistry (IHC) techniques were examined histopathologically. It was found that vaccination of the mice with the pEGFP-Cap induced solid protection against PCV2 infection through induction of highly specific serum IgG antibodies and cytokines (IFN-γ and IL-10), and a small PCV2 viral load. The mice treated with the pEGFP-Cap and LG-strain developed no histopathologically detectable lesions (HE stain) and IHC techniques revealed only a few positive cells. Thus, this study demonstrated that recombinant pEGFP-Cap substantially alleviates PCV2 infection in mice and provides evidence that a DNA vaccine could be an alternative to PCV2 vaccines against PMWS. © 2014 The Societies and Wiley Publishing Asia Pty Ltd.
VEEV IA/B challenge. Our results indicate that it is pos- sible to improve the immunogenicity and protective efficacy of alphavirus DNA vaccines using... alphaviruses that ause periodic epizootics in the Americas . These New World lphaviruses cause diseases in humans characterized by fever, eadache...equine encephalitis virus, VEE, alphavirus , DNA vaccine, envelope glycoproteins, directed molecular evolution, efficacy, immunogenicity, laboratory
Corbeil, S.; LaPatra, S.E.; Anderson, E.D.; Kurath, G.
The efficacy of a DNA vaccine containing the glycoprotein gene of infectious hematopoietic necrosis virus (IHNV), a rhabdovirus affecting trout and salmon, was investigated. The minimal dose of vaccine required, the protection against heterologous strains, and the titers of neutralizing antibodies produced were used to evaluate the potential of the vaccine as a control pharmaceutical. Results indicated that a single dose of as little as 1–10 ng of vaccine protected rainbow trout fry against waterborne challenge by IHNV. An optimal dose of 100 ng per fish was selected to assure strong protection under various conditions. Neutralizing antibody titers were detected in fish vaccinated with concentrations of DNA ranging from 5 to 0.01 μg. Furthermore, the DNA vaccine protected fish against a broad range of viral strains from different geographic locations, including isolates from France and Japan, suggesting that the vaccine could be used worldwide. A single dose of this DNA vaccine induced protection in fish at a lower dose than is usually reported in mammalian DNA vaccine studies.
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
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.
Pradhan, Satya Narayan; Prince, Prabhu Rajaiah; Madhumathi, Jayaprakasam; Arunkumar, Chakkaravarthy; Roy, Parimal; Narayanan, Rangarajan Badri; Antony, Usha
Infectious Bursal Disease Virus (IBDV) causes immunosuppression in young chickens by destruction of antibody producing B cells in the Bursa of Fabricius and poses a potential threat to the poultry industry. We have examined the protective efficacy of a subunit DNA vaccine against IBDV infection in chickens in this study. An immunodominant VP2 gene fragment (VP252-417) was cloned into CMV promoter based DNA vaccine vector pVAX1 and in vitro expression of the DNA encoded antigens was confirmed by transfection of CHO cells with vaccine constructs followed by RT-PCR and western blot analysis using IBDV-antiserum. Two weeks old chickens were immunized intramuscularly with pVAXVP252-417 and the in vivo transcription of the plasmid DNA was confirmed by RT-PCR analysis of DNA injected muscle tissue at different intervals of post immunization. Tissue distribution analysis revealed that the plasmid DNA was extensively distributed in muscle, spleen, kidney, liver, and bursa tissues. Chickens immunized with pVAXVP252-417 developed high titer (1:12,000) of anti-VP252-417 antibodies. Further, chicken splenocytes from pVAXVP252-417 immunized group showed a significantly high proliferation to the whole viral and recombinant antigen (P<0.01) compared to control groups, which implies that pVAXVP252-417 codes for immunogenic fragment which has epitopes capable of eliciting both B and T cell responses. This is evident by the fact that, pVAXVP252-417 immunized chicken conferred 75% protection against virulent IBDV (vIBDV) challenge compared to the control group. Thus, the present study confirms that the immunodominant VP2 fragment can be used as a potential DNA vaccine against IBDV infection in chickens. Copyright © 2014 Elsevier B.V. All rights reserved.
Liu, Xueyan; Liu, Qing; Xiao, Kangpeng; Li, Pei; Liu, Qiong; Zhao, Xinxin; Kong, Qingke
DNA vaccines are widely used to prevent and treat infectious diseases, cancer and autoimmune diseases; however, their relatively low immunogenicity is an obstacle to their use. In this study, we constructed a novel and universal DNA vaccine vector (pSS898) that can be used to build DNA vaccines against duck enteritis virus (DEV) and other viruses that require DNA vaccines to provide protection. This vaccine vector has many advantages, including innate immunogenicity, efficient nuclear trafficking and resistance to attack from nucleases. UL24 and tgB from DEV were chosen as the antigens, and the heat labile enterotoxin B subunit (LTB) from Escherichia coli and the IL-2 gene (DuIL-2) from duck were used as adjuvants for the construction of DNA vaccine plasmids. Ducklings that were orally immunized with S739 (Salmonella Typhimurium Δasd-66 Δcrp-24 Δcya-25) and harboring these DEV DNA vaccines produced strong mucosal and systemic immune responses, and they resisted an otherwise lethal DEV challenge. More importantly, S739 (UL24-LTB) provided 90% protection after a priming-boost immunization. This study shows that our novel and universal DNA vaccine vector can be used efficiently in practical applications and may provide a promising method of orally inoculating ducks with a DEV DNA vaccine delivered by attenuated Salmonella Typhimurium for prevention of DVE.
Shedlock, Devon J; Aviles, Jenna; Talbott, Kendra T; Wong, Gary; Wu, Stephan J; Villarreal, Daniel O; Myles, Devin Jf; Croyle, Maria A; Yan, Jian; Kobinger, Gary P; Weiner, David B
Marburg and Ebola hemorrhagic fevers have been described as the most virulent viral diseases known to man due to associative lethality rates of up to 90%. Death can occur within days to weeks of exposure and there is currently no licensed vaccine or therapeutic. Recent evidence suggests an important role for antiviral T cells in conferring protection, but little detailed analysis of this response as driven by a protective vaccine has been reported. We developed a synthetic polyvalent-filovirus DNA vaccine against Marburg marburgvirus (MARV), Zaire ebolavirus (ZEBOV), and Sudan ebolavirus (SUDV). Preclinical efficacy studies were performed in guinea pigs and mice using rodent-adapted viruses, whereas murine T-cell responses were extensively analyzed using a novel modified assay described herein. Vaccination was highly potent, elicited robust neutralizing antibodies, and completely protected against MARV and ZEBOV challenge. Comprehensive T-cell analysis revealed cytotoxic T lymphocytes (CTLs) of great magnitude, epitopic breadth, and Th1-type marker expression. This model provides an important preclinical tool for studying protective immune correlates that could be applied to existing platforms. Data herein support further evaluation of this enhanced gene-based approach in nonhuman primate studies for in depth analyses of T-cell epitopes in understanding protective efficacy.
Silva, Bruna Daniella de Souza; da Silva, Ediane Batista; do Nascimento, Ivan Pereira; Dos Reis, Michelle Cristina Guerreiro; Kipnis, André; Junqueira-Kipnis, Ana Paula
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.
Liu, Yingli; Zheng, Jun; Li, Jianhua; Gong, Pengtao; Zhang, Xichen
Rhomboid protein in Apicomplexa was associated with the process of host cell invasion. To evaluate the potential of the protein in eliciting protective immunity against challenge, a DNA vaccine pVAX1-Rho encoding Eimeria tenella rhomboid was constructed. Recombinant protein was expressed in Hela cells and verified by indirect immunofluorescence and western blotting analysis. In vivo experiments, 1-week-old chickens were randomly divided into three groups. Experimental group of chickens were immunized with DNA vaccines while control group of chickens were injected with pVAX1 plasmid alone or sterile water. Two weeks following the booster dose, all chickens were inoculated orally with 5 × 10(4) sporulated oocysts of E. tenella. The host immunity and protective efficacy of this vaccine against E. tenella challenge in broilers were evaluated. Results showed that specific antibody, the levels of interleukin-2 (IL-2), interferon-γ (IFN-γ), and the percentages of CD4(+) and CD8(+) T lymphocyte cells were significantly increased in the pVAX1-Rho group. Challenge experiments demonstrated that pVAX1-Rho vaccination could reduce oocyst excretion, decrease cecal lesion, increase bodyweight gains and provide chickens with oocysts decrease ratio around 75.8 %. These results suggest that the pVAX1-Rho was able to induce humoral and cellular responses and generate protective immunity against E. tenella infection.
Ramachandran, Sabarinathan; Kumar, Mishra Pankaj; Rami, Reddy Maryada Venkata; Chinnaiah, Harinath Basker; Nutman, Thomas; Kaliraj, Perumal; McCarthy, James
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.
Zhou, Feng-Juan; Hu, Zhen-Lin; Dai, Jian-Xin; Chen, Rui-Wen; Shi, Ke; Lin, Yi; Sun, Shu-Han
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.
Xu, Cheng; Mutoloki, Stephen; Evensen, Øystein
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.
Lee, Chih-Chun; Kim, Bong-Suk; Wu, Ching Ching; Lin, Tsang Long
Infectious bursal disease virus (IBDV) infection destroys the bursa of Fabricius, causing immunosuppression and rendering chickens susceptible to secondary bacterial or viral infections. IBDV large-segment-protein-expressing DNA has been shown to confer complete protection of chickens from infectious bursal disease (IBD). The purpose of the present study was to compare DNA-vaccinated chickens and unvaccinated chickens upon IBDV challenge by transcriptomic analysis of bursa regarding innate immunity, inflammation, immune cell regulation, apoptosis and glucose transport. One-day-old specific-pathogen-free chickens were vaccinated intramuscularly three times at weekly intervals with IBDV large-segment-protein-expressing DNA. Chickens were challenged orally with 8.2 × 10(2) times the egg infective dose (EID)50 of IBDV strain variant E (VE) one week after the last vaccination. Bursae collected at 0.5, 1, 3, 5, 7, and 10 days post-challenge (dpc) were subjected to real-time RT-PCR quantification of bursal transcripts related to innate immunity, inflammation, immune cell regulation, apoptosis and glucose transport. The expression levels of granzyme K and CD8 in DNA-vaccinated chickens were significantly (p < 0.05) higher than those in unvaccinated chickens upon IBDV challenge at 0.5 or 1 dpc. The expression levels of other genes involved in innate immunity, inflammation, immune cell regulation, apoptosis and glucose transport were not upregulated or downregulated in DNA-vaccinated chickens during IBDV challenge. Bursal transcripts related to innate immunity and inflammation, including TLR3, MDA5, IFN-α, IFN-β, IRF-1, IRF-10, IL-1β, IL-6, IL-8, iNOS, granzyme A, granzyme K and IL-10, were upregulated or significantly (p < 0.05) upregulated at 3 dpc and later in unvaccinated chickens challenged with IBDV. The expression levels of genes related to immune cell regulation, apoptosis and glucose transport, including CD4, CD8, IL-2, IFN-γ, IL-12(p40), IL-18, GM-CSF, GATA-3
Rodriguez-Carreno, Maria P; Nelson, Michael S; Botten, Jason; Smith-Nixon, Kim; Buchmeier, Michael J; Whitton, J Lindsay
Several viruses in the Arenavirus genus of the family Arenaviridae cause severe, often fatal, hemorrhagic fever. One such virus, Lassa virus (LV), is a frequent cause of disease in Africa, and survivors often are left with substantial neurological impairment. The feasibility of protective immunization against LV infection, and the associated disease, has been demonstrated in animal models, using recombinant vaccinia viruses to deliver Lassa proteins. Circumstantial evidence implicates cellular immunity in this Lassa-induced protection, but this has not been confirmed. Here, we describe DNA vaccines that encode LV proteins. A single inoculation of a plasmid encoding full-length Lassa nucleoprotein (LNP) can induce CD8(+) T cell responses in mice and can protect against challenge with two arenaviruses, lymphocytic choriomeningitis virus (LCMV) and Pichinde virus (PV). A DNA minigene vaccine encoding a 9 amino acid sequence from LNP also induces CD8(+) T cells and protects against arenavirus challenge, thus confirming prior speculation that protective cellular immunity is induced by LV proteins.
Kurath, Gael; Garver, Kyle A.; Corbeil, Serge; Elliott, Diane G.; Anderson, Eric D.; LaPatra, Scott E.
The DNA vaccine pIHNw-G encodes the glycoprotein of the fish rhabdovirus infectious hematopoietic necrosis virus (IHNV). Vaccine performance in rainbow trout was measured 3, 6, 13, 24, and 25 months after vaccination. At three months all fish vaccinated with 0.1 μg pIHNw-G had detectable neutralizing antibody (NAb) and they were completely protected from lethal IHNV challenge with a relative percent survival (RPS) of 100% compared to control fish. Viral challenges at 6, 13, 24, and 25 months post-vaccination showed protection with RPS values of 47–69%, while NAb seroprevalence declined to undetectable levels. Passive transfer experiments with sera from fish after two years post-vaccination were inconsistent but significant protection was observed in some cases. The long-term duration of protection observed here defined a third temporal phase in the immune response to IHNV DNA vaccination, characterized by reduced but significant levels of protection, and decline or absence of detectable NAb titers. Examination of multiple tissues showed an absence of detectable long-term histopathological damage due to DNA vaccination.
Yuan, Hu; You-En, Shi; Long-Jiang, Yu; Xiao-Hua, Zhu; Liu-Zhe, Li; Cash, Melanie; Lu, Zhu; Zhi, Liu; Deng-Xin, Song
In order to explore the high performance bivalent DNA vaccine of Schistosoma japonicum, the fatty-acid-binding protein (Sj14) and the 23 kDa transmembrane protein (Sj23) two proteins were selected to construct the DNA-based vaccine. It was successful to construct a bivalent DNA vaccine using three strategies: the co-expression of two genes, a fusion gene expression and two kinds of plasmids in combination (cocktail vaccine). The bivalent DNA was proven to express well in vitro and in vivo by indirect immunofluorescence test (IIF) and reverse transcriptase-polymerase chain reaction (RT-PCR). The protective immunity of bivalent DNA vaccine was higher than that of univalent DNA vaccine (p<0.05). There were four groups of bivalent vaccine whose protective immunity was higher than 50%. Granuloma diameter reduction rates were in the range of 18-39%. There was no significant impact on immunity protection exerted by the four factors including dosage, inoculated times, inoculated routes and challenge time after the last immunization in three levels (p>0.05).
Kaur, Sukhbir; Kaur, Tejinder; Joshi, Jyoti
Abstract The current study was designed to examine the protective efficacy of DNA vaccines based on gp63 and Hsp70 against murine visceral leishmaniasis. Inbred BALB/c mice were immunized subcutaneously twice at an interval of three weeks with pcDNA3.1(+) encoding T cell epitopes of gp63 and Hsp70 individually and in combination. Animals were challenged intracardially with 107 promastigotes of Leishmania donovani 10 days post immunization and sacrificed 1, 2 and 3 months post challenge. The immunized animals revealed a significant reduction (P < 0.05) in splenic and hepatic parasite burden as compared to the infected controls. Maximum reduction in parasite load (P < 0.05) was observed in animals treated with a combination of pcDNA/gp63 and pcDNA/Hsp70. These animals also showed heightened DTH response, increased IgG2a, elevated Th1 cytokines (IFN-γ and IL-2) and reduced IgG1 and IL-10 levels. Thus, mice immunized with the cocktail vaccine exhibited significantly greater protection in comparison to those immunized with individual antigens. PMID:27533939
Torrieri-Dramard, Lea; Lambrecht, Bénédicte; Ferreira, Helena Lage; Van den Berg, Thierry; Klatzmann, David; Bellier, Bertrand
The induction of potent virus-specific immune responses at mucosal surfaces where virus transmission occurs is a major challenge for vaccination strategies. In the case of influenza vaccination, this has been achieved only by intranasal delivery of live-attenuated vaccines that otherwise pose safety problems. Here, we demonstrate that potent mucosal and systemic immune responses, both cellular and humoral, are induced by intranasal immunization using formulated DNA. We show that formulation with the DNA carrier polyethylenimine (PEI) improved by a 1,000-fold the efficiency of gene transfer in the respiratory track following intranasal administration of luciferase-coding DNA. Using PEI formulation, intranasal vaccination with DNA-encoding hemagglutinin (HA) from influenza A H5N1 or (H1N1)2009 viruses induced high levels of HA-specific immunoglobulin A (IgA) antibodies that were detected in bronchoalveolar lavages (BALs) and the serum. No mucosal responses could be detected after parenteral or intranasal immunization with naked-DNA. Furthermore, intranasal DNA vaccination with HA from a given H5N1 virus elicited full protection against the parental strain and partial cross-protection against a distinct highly pathogenic H5N1 strain that could be improved by adding neuraminidase (NA) DNA plasmids. Our observations warrant further investigation of intranasal DNA as an effective vaccination route. PMID:20959813
Garg, Rajni; Kaur, Manpreet; Saxena, Ankur; Prasad, Rajendra; Bhatnagar, Rakesh
Rabies is a serious concern world-wide. Despite availability of rabies vaccines for long; their efficacy, safety, availability and cost effectiveness has been a tremendous issue. This calls for improvement of rabies vaccination strategies. DNA vaccination has immense potential in this regard. The DNA vaccine pgp.LAMP-1 conferred 60% protection to BALB/c mice against 20 LD50 rabies challenge virus standard (CVS) strain challenge. Upon supplementation with Emulsigen-D, the vaccine formulation conferred complete protection against lethal challenge. To assess the feasibility of this vaccine formulation for human use, it was tested along with other FDA approved adjuvants, namely, Alum, Immuvac, Montanide ISA720 VG. Enhanced immune response correlated with high IgG antibody titer, Th2 biased response with a high level of rabies virus neutralizing antibodies (RVNAs) and IgG1/IgG2a ratio >1, observed upon alum supplementation of the rabies DNA vaccine. The total IgG antibody titer was 2IU/ml and total RVNA titer was observed to be 4IU/ml which is eight times higher than the minimum protective titer recommended by WHO. Furthermore, it conferred 80% protection against challenge with 50 LD50 of the rabies CVS strain, conducted in compliance with the potency test for rabies recommended by the National Institutes of Health (NIH), USA. Previously, we have established pre-clinical safety of this vaccine as per the guidelines of Schedule Y, FDA as well as The European Agency for evaluation of Medicinal Products. The vaccine showed no observable toxicity at the site of injection as well as at systemic level in Wistar rats when administered with 10X recommended dose. Therefore, supplementation of rabies DNA vaccine, pgp.LAMP-1 with alum would lead to development of a non-toxic, efficacious, stable and affordable vaccine that can be used to combat high numbers of fatal rabies infections tormenting developing countries.
Lewis, P J; Babiuk, L A
Therapeutic and prophylactic DNA vaccine clinical trials for a variety of pathogens and cancers are underway (Chattergoon et al., 1997; Taubes, 1997). The speed with which initiation of these trials occurred is no less than astounding; clinical trials for a human immunodeficiency virus (HIV) gp160 DNA-based vaccine were underway within 36 months of the first description of "genetic immunization" (Tang et al., 1992) and within 24 months of publication of the first article describing intramuscular delivery of a DNA vaccine (Ulmer et al., 1993). Despite the relative fervor with which clinical trials have progressed, it can be safely stated that DNA-based vaccines will not be an immunological "silver bullet." In this regard, it was satisfying to see a publication entitled "DNA Vaccines--A Modern Gimmick or a Boon to Vaccinology?" (Manickan et al., 1997b). There is no doubt that this technology is well beyond the phenomenology phase of study. Research niches and models have been established and will allow the truly difficult questions of mechanism and application to target species to be studied. These two aspects of future studies are intricately interwoven and will ultimately determine the necessity for mechanistic understanding and the evolution of target species studies. The basic science of DNA vaccines has yet to be clearly defined and will ultimately determine the success or failure of this technology to find a place in the immunological arsenal against disease. In a commentary on a published study describing DNA vaccine-mediated protection against heterologous challenge with HIV-1 in chimpanzees, Ronald Kennedy (1997) states, "As someone who has been in the trenches of AIDS vaccine research for over a decade and who, together with collaborators, has attempted a number of different vaccine approaches that have not panned out, I have a relatively pessimistic view of new AIDS vaccine approaches." Kennedy then goes on to summarize a DNA-based multigene vaccine
Danko, Janine R; Beckett, Charmagne G; Porter, Kevin R
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.
Song, Xiaokai; Ren, Zhe; Yan, Ruofeng; Xu, Lixin; Li, Xiangrui
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.
Zheng Min; Jin Ningyi; Liu Qi; Huo Xiaowei; Li Yang; Hu Bo; Ma Haili; Zhu Zhanbo; Cong Yanzhao; Li Xiao; Jin Minglan; Zhu Guangze
Goatpox, caused by goatpox virus (GTPV), is an acute feverish and contagious disease in goats often associated with high morbidity and high mortality. To resolve potential safety risks and vaccination side effects of existing live attenuated goatpox vaccine (AV41), two Semliki forest virus (SFV) replicon-based bicistronic expression DNA vaccines (pCSm-AAL and pCSm-BAA) which encode GTPV structural proteins corresponding to the Vaccinia virus proteins A27, L1, A33, and B5, respectively, were constructed. Then, theirs ability to induce humoral and cellular response in mice and goats, and protect goats against virulent virus challenge were evaluated. The results showed that, vaccination with pCSm-AAL and pCSm-BAA in combination could elicit strong humoral and cellular responses in mice and goats, provide partial protection against viral challenge in goats, and reduce disease symptoms. Additionally, priming vaccination with the above-mentioned DNA vaccines could significantly reduce the goats' side reactions from boosting vaccinations with current live vaccine (AV41), which include skin lesions at the inoculation site and fevers. Data obtained in this study could not only facilitate improvement of the current goatpox vaccination strategy, but also provide valuable guidance to suitable candidates for evaluation and development of orthopoxvirus vaccines.
Zheng, Min; Jin, Ningyi; Liu, Qi; Huo, Xiaowei; Li, Yang; Hu, Bo; Ma, Haili; Zhu, Zhanbo; Cong, Yanzhao; Li, Xiao; Jin, Minglan; Zhu, Guangze
Goatpox, caused by goatpox virus (GTPV), is an acute feverish and contagious disease in goats often associated with high morbidity and high mortality. To resolve potential safety risks and vaccination side effects of existing live attenuated goatpox vaccine (AV41), two Semliki forest virus (SFV) replicon-based bicistronic expression DNA vaccines (pCSm-AAL and pCSm-BAA) which encode GTPV structural proteins corresponding to the Vaccinia virus proteins A27, L1, A33, and B5, respectively, were constructed. Then, theirs ability to induce humoral and cellular response in mice and goats, and protect goats against virulent virus challenge were evaluated. The results showed that, vaccination with pCSm-AAL and pCSm-BAA in combination could elicit strong humoral and cellular responses in mice and goats, provide partial protection against viral challenge in goats, and reduce disease symptoms. Additionally, priming vaccination with the above-mentioned DNA vaccines could significantly reduce the goats' side reactions from boosting vaccinations with current live vaccine (AV41), which include skin lesions at the inoculation site and fevers. Data obtained in this study could not only facilitate improvement of the current goatpox vaccination strategy, but also provide valuable guidance to suitable candidates for evaluation and development of orthopoxvirus vaccines.
Wang, Xiaoting; Dai, Yang; Zhao, Song; Tang, Jianxia; Li, Hongjun; Xing, Yuntian; Qu, Guoli; Li, Xinsong; Dai, Jianrong; Zhu, Yinchang; Zhang, Xueguang
Background Schistosomiasis japonica remains a major public-health concern in China. Praziquantel-based chemotherapy effectively reduces both infections and intensity; however, it can not prevent re-infection. Furthermore, there is an increasing concern about praziquantel resistance following long-term repeated use of the drug in endemic areas. Therefore, development of a schistosomiasis vaccine, as a strategy to prevent and control schistosomiasis japonica, has been given high priority. The present study was conducted to develop PAMAM dendrimers as a novel vaccine delivery vector for a schistosomiasis japonica DNA vaccine and evaluate its ability to enhance protective effects against Schistosoma japonicum infection. Methodology/Principal Findings Lysine was used to modify 4.0G PAMAM, and the modified product PAMAM-Lys was synthesized. PAMAM-Lys showed both high transfection and low cytotocity for gene delivery in vitro. DNA vaccines combined with PAMAM-Lys produced higher level of protection compare with naked DNA vaccines against S. japonicum infection in a mouse model. Futhermore,antibodies from mice immunized with PAMAM-Lys combined DNA vaccines were significantly higher than those of mice immunized with the naked DNA vaccines. The PAMAM-Lys vector elicited a predominantly IgG2a antibody response and a tremendously increase in the production of IL-2 and IFN-γ. Conclusion/Significance Lysine-modified PAMAM-Lys is an excellent vector. PAMAM-Lys may enhance the immunoreactivity of DNA vaccine and increase the protective effect of the SjC23 DNA vaccine against S. japonicum infection. PMID:24497955
Yonezawa, Hideo; Ishihara, Kazuyuki; Okuda, Katsuji
Arginine-specific cysteine proteinases (RgpA and RgpB) produced by the periodontal pathogen Porphyromonas gingivalis are suspected virulence factors and are involved in interrupting host defense mechanisms as well as in penetrating and destroying periodontal connective tissues. To induce a protective immune response against P. gingivalis, we constructed an rgpA DNA vaccine. BALB/c mice were immunized intradermally by Gene Gun with plasmid DNA carrying rgpA. Antibody responses against P. gingivalis were determined by an enzyme-linked immunosorbent assay. The rgpA DNA vaccine induced high levels of serum antibodies against P. gingivalis. Sera from the rgpA DNA vaccine-immunized mice diminished the proteolytic activity of RgpA and RgpB and inhibited the binding of P. gingivalis to a type I collagen sponge. Moreover, the sera effectively reduced the hemagglutination of P. gingivalis, indicating that the hemagglutinin activity of the organism is associated with RgpA. We found with a murine abscess model that mice immunized with the rgpA DNA vaccine were resistant to an invasive P. gingivalis W50 challenge. These results suggest that the rgpA DNA vaccine induced specific antibodies against the enzyme and that this vaccine could confer protective immunity against P. gingivalis infection. PMID:11292699
Hwang, Hye Suk; Lee, Young-Tae; Kim, Ki-Hye; Park, Soojin; Kwon, Young-Man; Lee, Youri; Ko, Eun-Ju; Jung, Yu-Jin; Lee, Jong Seok; Kim, Yu-Jin; Lee, Yu-Na; Kim, Min-Chul; Cho, Minkyoung; Kang, Sang-Moo
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.
Gorres, J. Patrick; Lager, Kelly M.; Kong, Wing-Pui; Royals, Michael; Todd, John-Paul; Vincent, Amy L.; Wei, Chih-Jen; Loving, Crystal L.; Zanella, Eraldo L.; Janke, Bruce; Kehrli, Marcus E.; Nabel, Gary J.; Rao, Srinivas S.
Swine influenza is a highly contagious viral infection in pigs that significantly impacts the pork industry due to weight loss and secondary infections. There is also the potential of a significant threat to public health, as was seen in 2009 when the pandemic H1N1 influenza virus strain emerged from reassortment events among avian, swine, and human influenza viruses within pigs. As classic and pandemic H1N1 strains now circulate in swine, an effective vaccine may be the best strategy to protect the pork industry and public health. Current inactivated-virus vaccines available for swine influenza protect only against viral strains closely related to the vaccine strain, and egg-based production of these vaccines is insufficient to respond to large outbreaks. DNA vaccines are a promising alternative since they can potentially induce broad-based protection with more efficient production methods. In this study we evaluated the potentials of monovalent and trivalent DNA vaccine constructs to (i) elicit both humoral and gamma interferon (IFN-γ) responses and (ii) protect pigs against viral shedding and lung disease after challenge with pandemic H1N1 or classic swine H1N1 influenza virus. We also compared the efficiency of a needle-free vaccine delivery method to that of a conventional needle/syringe injection. We report that DNA vaccination elicits robust serum antibody and cellular responses after three immunizations and confers significant protection against influenza virus challenge. Needle-free delivery elicited improved antibody responses with the same efficiency as conventional injection and should be considered for development as a practical alternative for vaccine administration. PMID:21918118
Einer-Jensen, Katja; Delgado, Lourdes; Lorenzen, Ellen; Bovo, Giuseppe; Evensen, Øystein; Lapatra, Scott; Lorenzen, Niels
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-vaccination against the two diseases would be a preferable option. In the present study we demonstrated that a single injection of mixed DNA vaccines induced long-lasting protection against both individual and a simultaneous virus challenge 80 days post vaccination. Transfected muscle cells at the injection site expressed both G proteins. This study confirms the applied potential of using a combined DNA vaccination for protection of fish against two different rhabdoviral diseases.
Rodríguez-Cortés, Alhelí; Ojeda, Ana; López-Fuertes, Laura; Timón, Marcos; Altet, Laura; Solano-Gallego, Laia; Sánchez-Robert, Elisenda; Francino, Olga; Alberola, Jordi
Vaccination of dogs, the domestic reservoir of Leishmania infantum, is the best method for controlling zoonotic visceral leishmaniasis. This strategy would reduce the incidence of disease in both the canine and, indirectly, the human population. Different vaccination approaches have been investigated against canine leishmaniasis (CaL) but to date there is only one licensed vaccine against this disease in dogs, in Brazil. DNA immunization is a promising method for inducing both humoral and cellular immune responses against this parasitic disease. Here, we report the results of a multiantigenic plasmid DNA vaccine encoding KMPII, TRYP, LACK and GP63 L. infantum antigens against experimentally induced CaL. Twelve dogs were randomly assigned to two groups receiving, at a 15 days interval, either four doses of plasmid DNA or similar injections of PBS. After vaccination, dogs were intravenously challenged with 5 x 10(7) promastigotes of L. infantum. The vaccine showed to be safe and well-tolerated. Neither cellular immune response nor antibodies directed against whole Leishmania antigen were detected after immunization in vaccinated dogs, although anti-LACK-specific antibodies were sporadically detected in two vaccinated dogs before challenge, thus suggesting that antigens were indeed expressed. A delay in the development of detectable specific immune response and parasite multiplication in vaccinated dogs was observed after challenge. Nevertheless, the multiantigenic Leishmania DNA vaccine was unable to induce protection against parasite dissemination or disease. This study emphasizes the need to strengthen DNA vaccines in order to obtain effective immune responses in models other than the murine.
Laddy, Dominick J; Weiner, David B
The field of DNA vaccine development began over 16 years ago with the observation that plasmid DNA could be injected into and expressed in vivo and drive adaptive immune responses. Since then, there has been great interest in developing this technology to create a new generation of vaccines with the ability to elicit both humoral and cellular immune responses from an inherently innocuous injection. However, DNA vaccines have yet to proceed past phase I/II clinical trials in humans--primarily due to a desire to induce more potent immune responses. This review will examine how DNA vaccines function to induce an immune response and how this information might be useful in future vaccine design.
Zhou, Jian; Wang, Lin; Lu, Gang; Zhou, Aihua; Zhu, Meiyan; Li, Qihang; Wang, Zhilin; Arken, Miradel; Wang, Ao; He, Shenyi
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. PMID:27055564
Garrod, Tamsin J; Gargett, Tessa; Yu, Wenbo; Major, Lee; Burrell, Christopher J; Wesselingh, Steven; Suhrbier, Andreas; Grubor-Bauk, Branka; Gowans, Eric J
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.
Jensen, Trine Hammer; Nielsen, Line; Aasted, Bent; Blixenkrone-Møller, Merete
Young mink kits (n=8) were vaccinated with DNA plasmids encoding the viral haemagglutinin protein (H) of a vaccine strain of Canine distemper virus (CDV). Virus neutralising (VN) antibodies were induced after 2 immunisations and after the third immunisation all kits had high VN antibody titres. The VN antibody titres remained high for more than 4 months and the mink were protected against viraemia, lymphopenia, clinical disease and changes in the percentage of IFN-gamma producing peripheral blood leucocytes after challenge inoculation with a recent wild type strain of CDV. Essentially, these results 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.
Chi, Hang; Zheng, Xuexing; Wang, Xiwen; Wang, Chong; Wang, Hualei; Gai, Weiwei; Perlman, Stanley; Yang, Songtao; Zhao, Jincun; Xia, Xianzhu
The Middle East respiratory syndrome coronavirus (MERS-CoV), is an emerging pathogen that continues to cause outbreaks in the Arabian peninsula and in travelers from this region, raising the concern that a global pandemic could occur. Here, we show that a DNA vaccine encoding the first 725 amino acids (S1) of MERS-CoV spike (S) protein induces antigen-specific humoral and cellular immune responses in mice. With three immunizations, high titers of neutralizing antibodies (up to 1: 10(4)) were generated without adjuvant. DNA vaccination with the MERS-CoV S1 gene markedly increased the frequencies of antigen-specific CD4(+) and CD8(+) T cells secreting IFN-γ and other cytokines. Both pcDNA3.1-S1 DNA vaccine immunization and passive transfer of immune serum from pcDNA3.1-S1 vaccinated mice protected Ad5-hDPP4-transduced mice from MERS-CoV challenge. These results demonstrate that a DNA vaccine encoding MERS-CoV S1 protein induces strong protective immune responses against MERS-CoV infection. Copyright © 2017 Elsevier Ltd. All rights reserved.
Ebola virus (ZEBOV). Numerous strains of these viruses have been identified. One species of Marburg-like viruses has been officially designated... viruses to guinea pigs challenged with ebola virus . In: Vaccines 97. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, New York, pp. 87/92. Hevey...Short communication Comparison of the protective efficacy of DNA and baculovirus- derived protein vaccines for EBOLA virus in guinea pigs Jenny L
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.
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.
Quattrocchi, Valeria; Soria, Ivana; Langellotti, Cecilia Ana; Gnazzo, Victoria; Gammella, Mariela; Moore, Dadin P; Zamorano, Patricia I
Bovine herpesvirus-1 (BoHV-1) is the causative agent of bovine infectious rhinotracheitis, an important disease worldwide. Although conventional BoHV-1 vaccines, including those based on the use of modified live virus and also inactivated vaccines, are currently used in many countries, they have several disadvantages. DNA vaccines have emerged as an attractive approach since they have the potential to induce both humoral and cellular immune response; nevertheless, it is largely known that potency of naked DNA vaccines is limited. We demonstrated previously, in the murine model, that the use of adjuvants in combination with a DNA vaccine against BoHV-1 is immunologically beneficial. In this study, we evaluate the immune response and protection against challenge elicited in bovines, by a DNA vaccine carrying the sequence of secreted version of glycoprotein D (gD) of BoHV-1 formulated with chemical adjuvants. Bovines were vaccinated with formulations containing the sequence of gD alone or in combination with adjuvants ESSAI 903110 or Montanide™ 1113101PR. After prime vaccination and two boosters, animals were challenged with infectious BoHV-1. Formulations containing adjuvants Montanide™ 1113101PR and ESSAI 903110 were both, capable of increasing humoral immune response against the virus and diminishing clinical symptoms. Nevertheless, only formulations containing adjuvant Montanide™ 1113101PR was capable of improving cellular immune response and diminishing viral excretion. To our knowledge, it is the first time that a BoHV-1 DNA vaccine is combined with adjuvants and tested in cattle. These results could be useful to design a vaccine for the control of bovine rhinotracheitis.
Quattrocchi, Valeria; Soria, Ivana; Langellotti, Cecilia Ana; Gnazzo, Victoria; Gammella, Mariela; Moore, Dadin P.; Zamorano, Patricia I.
Bovine herpesvirus-1 (BoHV-1) is the causative agent of bovine infectious rhinotracheitis, an important disease worldwide. Although conventional BoHV-1 vaccines, including those based on the use of modified live virus and also inactivated vaccines, are currently used in many countries, they have several disadvantages. DNA vaccines have emerged as an attractive approach since they have the potential to induce both humoral and cellular immune response; nevertheless, it is largely known that potency of naked DNA vaccines is limited. We demonstrated previously, in the murine model, that the use of adjuvants in combination with a DNA vaccine against BoHV-1 is immunologically beneficial. In this study, we evaluate the immune response and protection against challenge elicited in bovines, by a DNA vaccine carrying the sequence of secreted version of glycoprotein D (gD) of BoHV-1 formulated with chemical adjuvants. Bovines were vaccinated with formulations containing the sequence of gD alone or in combination with adjuvants ESSAI 903110 or Montanide™ 1113101PR. After prime vaccination and two boosters, animals were challenged with infectious BoHV-1. Formulations containing adjuvants Montanide™ 1113101PR and ESSAI 903110 were both, capable of increasing humoral immune response against the virus and diminishing clinical symptoms. Nevertheless, only formulations containing adjuvant Montanide™ 1113101PR was capable of improving cellular immune response and diminishing viral excretion. To our knowledge, it is the first time that a BoHV-1 DNA vaccine is combined with adjuvants and tested in cattle. These results could be useful to design a vaccine for the control of bovine rhinotracheitis. PMID:28179907
González, Belén; Reina, Ramsés; García, Iker; Andrés, Sara; Glaria, Idoia; Alzueta, María; Mora, María Isabel; Jugo, Begoña M; Arrieta-Aguirre, Inés; de la Lastra, José M Pérez; Rodríguez, Dolores; Rodríguez, Juan Ramón; Esteban, Mariano; Grilló, María Jesús; Blacklaws, Barbara A; Harkiss, Gordon D; Chebloune, Yahia; Luján, Lluís; de Andrés, Damián; Amorena, Beatriz
Gene gun mucosal DNA immunization of sheep with a plasmid expressing the env gene of Maedi-Visna virus (MVV) was used to examine the protection against MVV infection in sheep from a naturally infected flock. For immunization, sheep were primed with a pcDNA plasmid (pcDNA-env) encoding the Env glycoproteins of MVV and boosted with combined pcDNA-env and pCR3.1-IFN-gamma plasmid inoculations. The pcDNA plasmid used in the control group contained the lacZ coding sequences instead of the env gene. Within a month post-challenge, the viral load in the vaccinated group was lower (p < or = 0.05) and virus was only detected transiently compared with the control group. Furthermore, 2 months later, neutralizing antibodies (NtAb) were detected in all the control animals and none of the vaccinated animals (p < or = 0.01). These results demonstrated a significant early protective effect of this immunization strategy against MVV infection that restricts the virus replication following challenge in the absence of NtAb production. This vaccine protective effect against MVV infection disappeared after two years post-challenge, when active replication of MVV challenge strain was observed. Protection conferred by the vaccine could not be explained by OLA DRB1 allele or genotype differences. Most of the individuals were DRB1 heterozygous and none was totally resistant to infection.
involved topical dministration of DNA followed by electroporation [21–23] r subcutaneous injection of DNA followed by electropora - ion [24–26]. Others have...demonstrated improved delivery f DNA vaccines to the skin by micromechanical disruption 27]; however this procedure did not involve electropora - ion
Jiang, Mingzi; Yao, Jing; Feng, Ganzhu
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.
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
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. PMID:25996997
Ferreira, Daniela M; Miyaji, Eliane N; Oliveira, Maria Leonor S; Darrieux, Michelle; Arêas, Ana Paula M; Ho, Paulo L; Leite, Luciana C C
Pneumococcal surface protein A (PspA) is a promising candidate for the development of cost-effective vaccines against Streptococcus pneumoniae. In the present study, BALB/c mice were immunized with DNA vaccine vectors expressing the N-terminal region of PspA. Animals immunized with a vector expressing secreted PspA developed higher levels of antibody than mice immunized with the vector expressing the antigen in the cytosol. However, both immunogens elicited similar levels of protection against intraperitoneal challenge. Furthermore, immunization with exactly the same fragment in the form of a recombinant protein, with aluminium hydroxide as an adjuvant, elicited even higher antibody levels, but this increased humoral response did not correlate with enhanced protection. These results show that DNA vaccines expressing PspA are able to elicit protection levels comparable to recombinant protein, even though total anti-PspA IgG response is considerably lower.
Liu, Shan; Shi, Lin; Cheng, Yan-bin; Fan, Gui-xiang; Ren, Hui-xun; Yuan, Yu-kang
To investigate the vaccine potential of multi-epitope vaccines against toxoplasmosis, a multi-epitope DNA vaccine, eukaryotic plasmid pcDNA3.1/T-ME expressing six antigen segments (SAG1(238-256), SAG1(281-320), GRA1(170-193), GRA4(331-345), GRA4(229-245), and GRA2(171-185)) of Toxoplasma gondii was constructed. We investigated the efficacy of pcDNA3.1/T-ME with or without co-administration of a CpG-oligodeoxynucleotide (CpG-ODN) as an adjuvant to protect mice (BALB/c and C57BL/6) against toxoplasmosis. High survival rates were observed in mice immunized with pcDNA3.1/T-ME when challenged with T. gondii RH strain. Lymphocyte proliferation assays, cytokine, and antibody determinations show that mice immunized with pcDNA3.1/T-ME produced stronger humoral and Th1-type cellular immune responses compared to untreated mice or those immunized with empty plasmids. However, co-immunization with CpG-ODN resulted in impaired immune responses. Our data demonstrates that multi-epitope DNA vaccination is a potential strategy for the control of toxoplasmosis and paves the way for further investigations into producing a multi-epitope anti-T. gondii DNA vaccine.
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
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.
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
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. PMID:23049728
Yang, Benjamin; Jeang, Jessica; Yang, Andrew; Wu, T C; Hung, Chien-Fu
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. PMID:25625927
Yang, Benjamin; Jeang, Jessica; Yang, Andrew; Wu, T C; Hung, Chien-Fu
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.
Hong, David K.; Chang, Stella; Botham, Crystal M.; Giffon, Thierry D.; Fairman, Jeffery; Lewis, David B.
Influenza A virus is a negative-strand segmented RNA virus in which antigenically distinct viral subtypes are defined by the hemagglutinin (HA) and neuraminidase (NA) major viral surface proteins. An ideal inactivated vaccine for influenza A virus would induce not only highly robust strain-specific humoral and T-cell immune responses but also cross-protective immunity in which an immune response to antigens from a particular viral subtype (e.g., H3N2) would protect against other viral subtypes (e.g., H1N1). Cross-protective immunity would help limit outbreaks from newly emerging antigenically novel strains. Here, we show in mice that the addition of cationic lipid/noncoding DNA complexes (CLDC) as adjuvant to whole inactivated influenza A virus vaccine induces significantly more robust adaptive immune responses both in quantity and quality than aluminum hydroxide (alum), which is currently the most widely used adjuvant in clinical human vaccination. CLDC-adjuvanted vaccine induced higher total influenza virus-specific IgG, particularly for the IgG2a/c subclass. Higher levels of multicytokine-producing influenza virus-specific CD4 and CD8 T cells were induced by CLDC-adjuvanted vaccine than with alum-adjuvanted vaccine. Importantly, CLDC-adjuvanted vaccine provided significant cross-protection from either a sublethal or lethal influenza A viral challenge with a different subtype than that used for vaccination. This superior cross-protection afforded by the CLDC adjuvant required CD8 T-cell recognition of viral peptides presented by classical major histocompatibility complex class I proteins. Together, these results suggest that CLDC has particular promise for vaccine strategies in which T cells play an important role and may offer new opportunities for more effective control of human influenza epidemics and pandemics by inactivated influenza virus vaccine. PMID:20943978
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
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.
Wang, Haiyang; Zhu, Fan; Huang, Yucong; Ding, Yu; Jian, Jichang; Wu, Zaohe
The main aims of this study were to construct glutathione peroxidase (GPx) DNA vaccine of Vibrio harveyi ZJ0603 and to investigate its immune protective efficiency as a vaccine candidate on the orange-spotted grouper (Epinephelus coioides) treated with V. harveyi. Base on the cloning of ZJ0603 GPx gene, a DNA vaccine, named as pcDNA-GPx, was constructed by inserting GPx gene into pcDNA3.1 (+) plasmid. Orange-spotted groupers were immunized with the pcDNA-GPx plasmid by injection intramuscularly. The relative percent of survival (RPS) of fish vaccinated with the DNA vaccine against pathogenic V. harveyi infection was 77.5%. The expression of DNA vaccine was analyzed in the tissues of orange-spotted grouper by PCR and RT-PCR. The results indicated that pcDNA-GPx distributed and expressed in the head kidney, liver, spleen, gill and injected muscle at 7 and 28 days after vaccination. Significant specific antibody responses were also detected in the vaccinated orange-spotted groupers by indirect ELISA method. In a conclusion, DNA vaccine pcDNA-GPx showed an effective immune protection to the orange-spotted grouper treated with V. harveyi. The GPx can be used as a candidate DNA vaccine for the control of vibriosis.
Cao, Yan; Zhao, Bin; Han, Yanhui; Zhang, Juan; Li, Xuezhen; Qiu, Chunhui; Wu, Xiujuan; Hong, Yang; Ai, Dezhou; Lin, Jiaojiao; Fu, Zhiqiang
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% (P < 0.01) of worm reduction and 40.38–44.51% (P < 0.01) 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. PMID:23509820
Bragstad, Karoline; Vinner, Lasse; Hansen, Mette Sif; Nielsen, Jens; Fomsgaard, Anders
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 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 vaccine components. We found that pigs challenged with a virus homologous to the HA and NA DNA vaccine components were well protected from infection. In addition, heterologous challenge virus was cleared rapidly compared to the unvaccinated control pigs. Immunisation by electroporation induced HI antibodies >40 HAU/ml seven days after second vaccination. Heterologous virus challenge as long as ten weeks after last immunisation was able to trigger a vaccine antibody HI response 26 times higher than in the control pigs. The H3N2 DNA vaccine HA and NA genes also triggered an effective vaccine response with protective antibody titres towards heterologous H3N2 virus. The described influenza DNA vaccine is able to induce broadly protective immune responses even in a larger animal, like the pig, against both heterologous and homologous virus challenges despite relatively low HI titres after vaccination. The ability of this DNA vaccine to limit virus shedding may have an impact on virus spread among pigs which could possibly extend to humans as well, thereby diminishing the
Scott, Veronica L; Patel, Ami; Villarreal, Daniel O; Hensley, Scott E; Ragwan, Edwin; Yan, Jian; Sardesai, Niranjan Y; Rothwell, Paul J; Extance, Jonathan P; Caproni, Lisa J; Weiner, David B
Nucleic acid-based vaccines (NAVs) are a promising alternative to conventional influenza vaccines with the potential to increase influenza vaccine availability due to their simplicity in design and rapid speed of production. NAVs can also target multiple influenza antigens and control flu variants. Traditionally NAVs have been DNA plasmids however, we are continuing to explore new methods that may enhance vaccine efficacy. Recently new focus has been on RNA cassettes as NAVs. RNA vaccines combine conceptual advantages in that they focus on delivery of only the coding cassette. However, RNA vaccines have a short half-life and cause interferon-induced fevers. Here we describe a new NAV approach where we study delivery of a linear DNA cassette [Doggybone™ linear closed DNA [(dbDNA™)] produced by an enzymatic process that yields an antigen expression cassette comprising a promoter, DNA antigen, poly A tail, and telomeric ends. This focused approach has many of the advantages of plasmid DNA as well as a minimal cassette size similar to RNA strategies. For this study, we characterized the specific CD4+ and CD8+ T cell responses and determined the hemagglutination inhibition (HI) titers induced by dbDNA™ and compared the responses with those of an optimized plasmid DNA (pDNA) vaccine encoding the same H1N1 influenza A/PR/8/34 HA gene. Immunizations with the constructs resulted in similar humoral and cellular immune responses. Both constructs induced high-titer HI antibodies and fully protected animals from lethal viral challenge. The data obtained from this study provides important validation for further development of novel vector approaches. PMID:26091432
Rosas, Gabriela; Fragoso, Gladis; Garate, Teresa; Hernández, Beatriz; Ferrero, Patricia; Foster-Cuevas, Mildred; Parkhouse, R Michael E; Harrison, Leslie J S; Briones, Sergio López; González, Luis Miguel; Sciutto, Edda
This study investigated the protective capacity of the recombinant Taenia saginata Tso18 antigen administered as a DNA vaccine in the Taenia crassiceps murine model of cysticercosis. This Tso18 DNA sequence, isolated from a T. saginata oncosphere cDNA library, has homologies with Taenia solium and Echinococcus sp. It was cloned in the pcDNA3.1 plasmid and injected once intramuscularly into mice. Compared to saline-vaccinated control mice, immunization reduced the parasite burden by 57.3-81.4%, while lower levels of non-specific protection were induced in control mice injected with the plasmid pcDNA3.1 (18.8-33.1%) or a plasmid with irrelevant construct, pcDNA3.1/3D15 (33.4-38.8%). Importantly, significant levels of protection were observed between the pcDNA3.1/Tso18 plasmid and pcDNA3.1/3D15 plasmid immunized mice. Mice immunized with pTso18 synthesized low levels of, primarily IgG1 sub-class, antibodies. These antibodies were shown to recognize a 66 kDa antigen fraction of T. crassiceps and T. solium. Splenocytes enriched in both CD4+CD8- and CD4-CD8+ T cells from these vaccinated mice proliferated in vitro when exposed to antigens from both T. solium and T. crassiceps cestodes. Immunolocalization studies revealed the Tso18 antigen in oncospheres of T. saginata and T. solium, in the adult tapeworm and in the tegument of T. solium cysticerci. The protective capacity of this antigen and its extensive distribution in different stages, species and genera of cestodes points to the potential of Tso18 antigen for the possible design of a vaccine against cestodes.
Grant-Klein, Rebecca J; Van Deusen, Nicole M; Badger, Catherine V; Hannaman, Drew; Dupuy, Lesley C; Schmaljohn, Connie S
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.
Gartner, Tatiana; Romano, Marta; Suin, Vanessa; Kalai, Michaël; Korf, Hannelie; De Baetselier, Patrick; Huygen, Kris
DNA vaccination is a potent means for inducing strong cell-mediated immune responses and protective immunity against viral, bacterial and parasite pathogens in rodents. In an attempt to increase cross-presentation through apoptosis, the DNA-encoding caspase-2 prodomain followed by wild-type or catalytically inactive mutated caspase-3 was inserted into a plasmid encoding the 32 kDa mycolyl transferase (Ag85A) from Mycobacterium tuberculosis. Transient transfection showed that the mutated caspase induced slow apoptosis, normal protein expression and NF-kappaB activation while wild-type caspase induced rapid apoptosis, lower protein expression and no NF-kappaB activation. Ag85A specific antibody production was increased by co-expressing the mutated and decreased by co-expressing the wild-type caspase. Vaccination with pro-apoptotic plasmids triggered more Ag85A specific IFN-gamma producing spleen cells, and more efficient IL-2 and IFN-gamma producing memory cells in spleen and lungs after M. tuberculosis challenge. Compared to DNA-encoding secreted Ag85A, vaccination with DNA co-expressing wild-type caspase increased protection after infection with M. tuberculosis, while vaccination with plasmid co-expressing mutated caspase was not protective, possibly due to the stimulation of IL-6, IL-10 and IL-17A production.
Zheng, Lina; Hu, Yue; Hua, Qianqian; Luo, Fangjun; Xie, Guizhen; Li, Xiangzhi; Lin, Jiaxin; Wan, Yujing; Ren, Shoufeng; Pan, Changwang; Tan, Feng
DNA-based alphaviral RNA replicon vectors, also called suicidal DNA vectors, have been employed to alleviate biosafety concerns attribution to its ability to induce apoptotic cell death of the transfected cells. Toxoplasma gondii nucleoside triphosphate hydrolase-II (TgNTPase-II), which facilitates the parasite to salvage purines from the host cell for survival and replication, have been demonstrated to be a potential vaccine candidate for toxoplasmosis. Herein, we evaluated the immunogenic potential of a suicidal DNA vaccine encoding TgNTPase-II gene, pDREP-TgNTPase-II, delivered intramuscularly in combination with electroporation. Immunization of mice with pDREP-TgNTPase-II elicited specific humoral responses, with high IgG antibody titers and a mixed IgG1/IgG2a response. The cellular immune response was associated with high level production of IFN-γ, IL-2, IL-10 cytokines and low level IL-4 production as well as the increase of the percentage of CD8+ T cells, indicating that a Th1 predominant response was elicited. Furthermore, mice vaccinated with this suicidal DNA vaccine displayed partial protection against acute infection with the virulent RH strain as well as chronic infection with PRU cyst, which shows 77.7% and 71.4% reduction in brain cyst burden in comparison to PBS and pDREP-eGFP control group, respectively. Based on the cellular and antibody responses, the suicidal DNA vaccine elicited a Th1-predominant immune response against T. gondii challenge.
Dowling, William; Thompson, Elizabeth; Badger, Catherine; Mellquist, Jenny L; Garrison, Aura R; Smith, Jeffery M; Paragas, Jason; Hogan, Robert J; Schmaljohn, Connie
The Ebola virus (EBOV) envelope glycoprotein (GP) is the primary target of protective immunity. Mature GP consists of two disulfide-linked subunits, GP1 and membrane-bound GP2. GP is highly glycosylated with both N- and O-linked carbohydrates. We measured the influences of GP glycosylation on antigenicity, immunogenicity, and protection by testing DNA vaccines comprised of GP genes with deleted N-linked glycosylation sites or with deletions in the central hypervariable mucin region. We showed that mutation of one of the two N-linked GP2 glycosylation sites was highly detrimental to the antigenicity and immunogenicity of GP. Our data indicate that this is likely due to the inability of GP2 and GP1 to dimerize at the cell surface and suggest that glycosylation at this site is required for achieving the conformational integrity of GP2 and GP1. In contrast, mutation of two N-linked sites on GP1, which flank previously defined protective antibody epitopes on GP, may enhance immunogenicity, possibly by unmasking epitopes. We further showed that although deleting the mucin region apparently had no effect on antigenicity in vitro, it negatively impacted the elicitation of protective immunity in mice. In addition, we confirmed the presence of previously identified B-cell and T-cell epitopes in GP but show that when analyzed individually none of them were neither absolutely required nor sufficient for protective immunity to EBOV. Finally, we identified other potential regions of GP that may contain relevant antibody or T-cell epitopes.
Gupta, Shishir Kumar; Dey, Sohini; Chellappa, Madhan Mohan
Robust and sustainable development of poultry industry requires prevention of deadly infectious diseases. Vigorous vaccination of the birds is a routine practice; however, the live and inactivated vaccines that are used have inherent disadvantages. New-generation vaccines such as DNA vaccines offer several advantages over conventional vaccines. DNA vaccines, which encode an antigen of interest or multiple antigens in the target host, are stable, easy to produce and administer, do not require cold chain maintenance, and are not affected by the maternal antibodies. In addition, DNA vaccines can also be administered in ovo, and thus, mass vaccination and early induction of immune response can effectively be achieved. In this chapter, we focus on the development of DNA vaccines against important infectious viral as well as parasitic diseases of poultry.
Wang, Hai-Long; Wang, Yu-Jing; Pei, Yan-Jiang; Bai, Ji-Zhong; Yin, Li-Tian; Guo, Rui; Yin, Guo-Rong
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. PMID:26842927
Martín-Acebes, Miguel A; Blázquez, Ana-Belén; Cañas-Arranz, Rodrigo; Vázquez-Calvo, Ángela; Merino-Ramos, Teresa; Escribano-Romero, Estela; Sobrino, Francisco; Saiz, Juan-Carlos
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.
Arce-Fonseca, Minerva; Ramos-Ligonio, Angel; López-Monteón, Aracely; Salgado-Jiménez, Berenice; Talamás-Rohana, Patricia; Rosales-Encina, José Luis
Immunization of mice with plasmids containing genes of Trypanosoma cruzi induces protective immunity in the murine model of Chagas disease. A cDNA clone that codes for an amastigote-specific surface protein (TcSSP4) was used as a candidate to develop a DNA vaccine. Mice were immunized with the recombinant protein rTcSSP4 and with cDNA for TcSSP4, and challenged with bloodstream trypomastigotes. Immunization with rTcSSP4 protein makes mice more susceptible to trypomastigote infection, with high mortality rates, whereas mice immunized with a eukaryotic expression plasmid containing the TcSSP4 cDNA were able to control the acute phase of infection. Heart tissue of gene-vaccinated animals did not show myocarditis and tissue damage at 365 days following infection, as compared with control animals. INF-γ was detected in sera of DNA vaccinated mice shortly after immunization, suggesting the development of a Th1 response. The TcSSP4 gene is a promising candidate for the development of an anti-T. cruzi DNA vaccine. PMID:22110377
Al-Mariri, Ayman; Abbady, Abdul Qader
Brucella melitensis is a facultative intracellular Gram-negative bacterial pathogen that may enter the host via ingestion or inhalation, or through conjunctiva or skin abrasions. Some Brucella spp surface proteins (SPs) play an important role in bacterial adhesion and invasion and thus represent targets for the host immune system. Brucella spp surface protein with apparent molecular mass of 41 kDa interacts selectively with HeLa cells. To evaluate the role of SP41 (41 kDa) as a DNA vaccine against Brucella spp., pCISP41, a plasmid construct for protein expression in mammalian cells, was established. Exogenous SP41 was detected in pCISP41-transfected Vero cell line by immune blotting using specific polyclonal antibody. The protective role of pCISP41 against B. melitensis 16M in mice was evaluated by measuring B and T cell responses in comparison to those achieved with attenuated B. melitensis Rev. 1 vaccine. BALB/c mice injected with pCISP41 were able to develop SP41-specific serum immunoglobulin G (IgG) antibodies. In addition, splenocytes from DNA-SP41-vaccinated mice elicited a T-cell-proliferative response and also induced gamma interferon (IFN-γ) production, but not interleukin-5 (IL-5), suggesting the induction of a T-helper-1-dominated immune response. Vaccination with attenuated B. melitensis Rev.1 strain induced better protection levels than DNA vaccination with SP41 against B. melitensis 16M in mice. Such responses play an important role against intracellular infecting agents such as Brucella spp. Altogether, our data suggest that SP41 may represent a promising candidate for DNA vaccination against brucellosis, but more investigation to increase its protective efficacy should be done.
Roh, Ha Jung; Sung, Haan Woo
This study examined the adjuvant effects of dimethyl dioctadecyl ammonium bromide (DDA), CpG oligodeoxynucleotides (CpG-ODN), and chicken interferon-γ (ChIFN-γ) on a DNA vaccine (pcDNA-VP243) against the infectious bursal disease virus (IBDV). A plasmid encoding chicken IFN-ã was constructed. Twice at 2-week intervals, two-week-old chickens were injected intramuscularly and intraperitoneally with either a DNA vaccine alone or a DNA vaccine together with the respective adjuvants. On week 2 after the second immunization, the chickens were orally challenged with the highly virulent IBDV. The groups that received the DNA vaccines plus either DDA or CpG-ODN showed significantly lower survival rates than the group that received the DNA vaccine alone. However, the survival rates for the DNA vaccine alone and for the DNA vaccine plus ChIFN-γ were similar. The chickens had no detectable antibodies to the IBDV before the challenge but all the surviving chickens in all groups except for the normal control group showed the induction of antibodies to the IBDV at day 10 after the challenge. As judged by the lymphocyte proliferation assays using the a WST-8 solution performed on the peripheral blood and splenic lymphocytes, the stimulation indices (SI) of the peripheral blood lymphocytes in all groups except for the normal control group were similar immediately before the challenge. At 10 days post-challenge, the SI for DNA vaccine plus either CpG-ODN or ChIFN-γ was similar to that of the DNA vaccine control group. For splenic lymphocytes, the SI in the DNA vaccine plus CpG-ODN and DNA vaccine plus ChIFN-γ groups were higher than for the DNA vaccine control. These results suggest that DDA actually compromises the protection against the IBDV by DNA vaccine, and CpG-ODN and IFN-γ had no significant effect. PMID:17106228
Lorenzen, N; LaPatra, S E
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.
Wang, Lei; Wang, Xiaohuan; Bi, Kuo; Sun, Ximeng; Yang, Jing; Gu, Yuan; Huang, Jingjing; Zhan, Bin; Zhu, Xinping
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
Costa, Simone M; Yorio, Anna Paula; Gonçalves, Antônio J S; Vidale, Mariana M; Costa, Emmerson C B; Mohana-Borges, Ronaldo; Motta, Marcia A; Freire, Marcos S; Alves, Ada M B
The dengue non-structural 3 (NS3) is a multifunctional protein, containing a serino-protease domain, located at the N-terminal portion, and helicase, NTPase and RTPase domains present in the C-terminal region. This protein is considered the main target for CD4+ and CD8+ T cell responses during dengue infection, which may be involved in protection. However, few studies have been undertaken evaluating the use of this protein as a protective antigen against dengue, as well as other flavivirus. In the present work, we investigate the protective efficacy of DNA vaccines based on the NS3 protein from DENV2. Different recombinant plasmids were constructed, encoding either the full-length NS3 protein or only its functional domains (protease and helicase), fused or not to a signal peptide (t-PA). The recombinant proteins were successfully expressed in transfected BHK-21 cells, and only plasmids encoding the t-PA signal sequence mediated protein secretion. Balb/c mice were immunized with the different DNA vaccines and challenged with a lethal dose of DENV2. Most animals immunized with plasmids encoding the full-length NS3 or the helicase domain survived challenge, regardless of the presence of the t-PA. However, some mice presented clinical signs of infection with high morbidity (hind leg paralysis and hunched posture), mainly in animal groups immunized with the DNA vaccines based on the helicase domain. On the other hand, inoculation with plasmids encoding the protease domain did not induce any protection, since mortality and morbidity rates in these mouse groups were similar to those detected in the control animals. The cellular immune response was analyzed by ELISPOT with a specific-CD8+ T cell NS3 peptide. Results revealed that the DNA vaccines based on the full-length protein induced the production of INF-γ, thus suggesting the involvement of this branch of the immune system in the protection.
Raadsma, H W; O'Meara, T J; Egerton, J R; Lehrbach, P R; Schwartzkoff, C L
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
Zhou, Jian; Wang, Lin
Toxoplasma gondii can lead to congenital infections in human. Surface antigen protein 4 (SAG4) of T. gondii is a potential stimulator for humoral and cellular immune responses. In the present study, a DNA vaccine encoding SAG4 from T. gondii was constructed and used to immunize BALB/c mice with peptide to evaluate the protective efficacy of the vaccine. The productions of IgG antibodies and cytokines (gamma interferon) from the vaccine (pSAG4/peptide) group were significantly higher than pSAG4 or peptide groups. After a lethal challenge by 1 × 10(4) tachyzoites from the I strain (RH), the survival time of mice immunized by pSAG4/peptide was longer than that of pSAG4 or peptide immunized mice or control mice. Moreover, after challenging by 20 cysts of the II strain (PRU) of T. gondii, the number of brain cysts from pSAG4/peptide vaccinated mice was only 31% of the number in PBS injected mice. The findings suggested the SAG4 DNA vaccine with peptide led significant immune responses and improved the protection against T. gondii challenges.
Zhou, Jian; Wang, Lin
Toxoplasma gondii can lead to congenital infections in human. Surface antigen protein 4 (SAG4) of T. gondii is a potential stimulator for humoral and cellular immune responses. In the present study, a DNA vaccine encoding SAG4 from T. gondii was constructed and used to immunize BALB/c mice with peptide to evaluate the protective efficacy of the vaccine. The productions of IgG antibodies and cytokines (gamma interferon) from the vaccine (pSAG4/peptide) group were significantly higher than pSAG4 or peptide groups. After a lethal challenge by 1 × 104 tachyzoites from the I strain (RH), the survival time of mice immunized by pSAG4/peptide was longer than that of pSAG4 or peptide immunized mice or control mice. Moreover, after challenging by 20 cysts of the II strain (PRU) of T. gondii, the number of brain cysts from pSAG4/peptide vaccinated mice was only 31% of the number in PBS injected mice. The findings suggested the SAG4 DNA vaccine with peptide led significant immune responses and improved the protection against T. gondii challenges. PMID:28936207
Jin, Ke; Wang, Shixia; Zhang, Chunhua; Xiao, Yanling; Lu, Shan; Huang, Zuhu
A DNA vaccination approach was used in the current study to screen for the immunogenicity of different fragments of toxin A and toxin B from Clostridium difficile. With this approach, protein antigens do not need to be produced in vitro and the immunogenicity of candidate C. difficile antigens can be identified directly in animals. Codon optimized toxin gene fragments were individually cloned into the DNA vaccine vector and tested in mice and rabbits for their ability to elicit C. difficile toxin-specific antibody responses. Only a subset of the C. difficile toxin fragments, including the C-terminal receptor binding domain of toxin A and a novel N-terminal enzymatic domain of toxin B, were able to elicit protective antibody responses as determined by protection of target cells in a cytotoxicity assay or by preventing death of mice in a passive antibody protection study. Significantly, antibodies elicited by the novel N-terminus of the toxin B DNA vaccine were able to increase the level of protection when used in combination with anti-toxin A antibodies in a toxin challenge model in mice.
Chattopadhyay, Suchismita; Jiang, Ju; Nawtaisong, Pruksa; Lee, John S.; Tan, Esterlina; Dela Cruz, Eduardo; Burgos, Jasmin; Abalos, Rodolfo; Blacksell, Stuart D.; Lombardini, Eric; Turner, Gareth D.; Day, Nicholas P. J.; Richards, Allen L.
We developed an intradermal (ID) challenge cynomolgus macaque (Macaca fascicularis) model of scrub typhus, the leading cause of treatable undifferentiated febrile illness in tropical Asia, caused by the obligate intracellular bacterium, Orientia tsutsugamushi. A well-characterized animal model is required for the development of clinically relevant diagnostic assays and evaluation of therapeutic agents and candidate vaccines. We investigated scrub typhus disease pathophysiology and evaluated two O. tsutsugamushi 47-kDa, Ag-based candidate vaccines, a DNA plasmid vaccine (pKarp47), and a virus-vectored vaccine (Kp47/47-Venezuelan equine encephalitis virus replicon particle) for safety, immunogenicity, and efficacy against homologous ID challenge with O. tsutsugamushi Karp. Control cynomolgus macaques developed fever, classic eschars, lymphadenopathy, bacteremia, altered liver function, increased WBC counts, pathogen-specific Ab (IgM and IgG), and cell-mediated immune responses. Vaccinated macaques receiving the DNA plasmid pKarp47 vaccine had significantly increased O. tsutsugamushi–specific, IFN-γ–producing PBMCs (p = 0.04), reduced eschar frequency and bacteremia duration (p ≤ 0.01), delayed bacteremia onset (p < 0.05), reduced circulating bacterial biomass (p = 0.01), and greater reduction of liver transaminase levels (p < 0.03) than controls. This study demonstrates a vaccine-induced immune response capable of conferring sterile immunity against high-dose homologous ID challenge of O. tsutsugamushi in a nonhuman primate model, and it provides insight into cell-mediated immune control of O. tsutsugamushi and dissemination dynamics, highlights the importance of bacteremia indices for evaluation of both natural and vaccine-induced immune responses, and importantly, to our knowledge, has determined the first phenotypic correlates of immune protection in scrub typhus. We conclude that this model is suitable for detailed investigations into vaccine
Grosfeld, Haim; Cohen, Sara; Bino, Tamar; Flashner, Yehuda; Ber, Raphael; Mamroud, Emanuelle; Kronman, Chanoch; Shafferman, Avigdor; Velan, Baruch
Three plasmids expressing derivatives of the Yersinia pestis capsular F1 antigen were evaluated for their potential as DNA vaccines. These included plasmids expressing the full-length F1, F1 devoid of its putative signal peptide (deF1), and F1 fused to the signal-bearing E3 polypeptide of Semliki Forest virus (E3/F1). Expression of these derivatives in transfected HEK293 cells revealed that deF1 is expressed in the cytosol, E3/F1 is targeted to the secretory cisternae, and the nonmodified F1 is rapidly eliminated from the cell. Intramuscular vaccination of mice with these plasmids revealed that the vector expressing deF1 was the most effective in eliciting anti-F1 antibodies. This response was not limited to specific mouse strains or to the mode of DNA administration, though gene gun-mediated vaccination was by far more effective than intramuscular needle injection. Vaccination of mice with deF1 DNA conferred protection against subcutaneous infection with the virulent Y. pestis Kimberley53 strain, even at challenge amounts as high as 4,000 50% lethal doses. Antibodies appear to play a major role in mediating this protection, as demonstrated by passive transfer of anti-deF1 DNA antiserum. Taken together, these observations indicate that a tailored genetic vaccine based on a bacterial protein can be used to confer protection against plague in mice without resorting to regimens involving the use of purified proteins. PMID:12496187
Bruffaerts, Nicolas; Huygen, Kris; Romano, Marta
Tuberculosis (TB) remains a major health problem and novel vaccination regimens are urgently needed. DNA vaccines against TB have been tested in various preclinical models and strategies have been developed to increase their immunogenicity in large animal species. DNA vaccines are able to induce a wide variety of immune responses, including CD8(+) T-cell-mediated cytolytic and IFN-γ responses. DNA vaccination may be valuable in heterologous prime-boost strategies with the currently used bacillus Calmette-Guérin (BCG) vaccine. This approach could broaden the antigenic repertoire of BCG and enhance its weak induction of MHC class I-restricted immune responses. DNA vaccines offer a number of advantages over certain other types of vaccines, such as the induction of robust MHC class I-restricted cytotoxic T lymphocyte (CTL), their generic manufacturing platform and their relatively low manufacturing costs. Because of their strong potential for inducing memory responses, DNA vaccines are particularly suited for priming immune responses. Furthermore, DNA vaccine technology may help antigen discovery by facilitating screening of candidate vaccines. Co-administration of BCG with plasmid DNA coding for immunodominant, subdominant and phase-specific antigens, poorly expressed by BCG, may lead to the development of improved TB vaccines.
Hwang, Hye Suk; Lee, Young-Tae; Kim, Ki-Hye; Park, Soojin; Kwon, Young-Man; Lee, Youri; Ko, Eun-Ju; Jung, Yu-Jin; Lee, Jong Seok; Kim, Yu-Jin; Lee, Yu-Na; Kim, Min-Chul; Cho, Minkyoung; Kang, Sang-Moo
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. Copyright © 2016. Published by Elsevier Inc.
Zhao, Kai; Rong, Guangyu; Hao, Yan; Yu, Lu; Kang, Hong; Wang, Xin; Wang, Xiaohua; Jin, Zheng; Ren, Zhiyu; Li, Zejun
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. PMID:27170532
Zhao, Kai; Rong, Guangyu; Hao, Yan; Yu, Lu; Kang, Hong; Wang, Xin; Wang, Xiaohua; Jin, Zheng; Ren, Zhiyu; Li, Zejun
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.
Zhao, Jia; Deng, Shu; Liang, Jiayuan; Cao, Yaming; Liu, Jun; Du, Feng; Shang, Hong; Cui, Liwang; Luo, Enjie
Although great efforts have been undertaken for the development of malaria vaccines, no completely effective malaria vaccines are available yet. Despite being clinically silent, the pre-erythrocytic stage is considered an ideal target for the development of malaria vaccines. Sporozoite asparagine-rich protein 1 (SAP1) is a sporozoite-localized protein that regulates the expression of UIS (upregulated in infectious sporozoites) genes, which are essential for the infectivity of sporozoites. In this study, a recombinant DNA vaccine encoding a predicted antigenic determinant region of Plasmodium yoelii SAP1 (PySAP1) was constructed. Immunization of mice with this DNA vaccine construct resulted in significant elevation of cytokines such as IFN-γ, IL-2, IL-4 and IL-10, and total IgG as compared with control groups immunized with either the empty DNA vector or saline. After challenge with sporozoites, the group receiving the DNA vaccine showed delayed development of parasitemia and prolonged survival time compared with the control group. The DNA vaccine provided partial protection against P. yoelii 17XL infection, with an overall protection rate of 20%. In addition, the DNA vaccine did not show integration into the host genome. Further studies of SAP1 are needed to test whether it can be used as subunit vaccine candidate.
Sixt, N; Cardoso, A; Vallier, A; Fayolle, J; Buckland, R; Wild, T F
We have studied the immune responses to the two glycoproteins of the Morbillivirus canine distemper virus (CDV) after DNA vaccination of BALB/c mice. The plasmids coding for both CDV hemagglutinin (H) and fusion protein (F) induce high levels of antibodies which persist for more than 6 months. Intramuscular inoculation of the CDV DNA induces a predominantly immunoglobulin G2a (IgG2a) response (Th1 response), whereas gene gun immunization with CDV H evokes exclusively an IgG1 response (Th2 response). In contrast, the CDV F gene elicited a mixed, IgG1 and IgG2a response. Mice vaccinated (by gene gun) with either the CDV H or F DNA showed a class I-restricted cytotoxic lymphocyte response. Immunized mice challenged intracerebrally with a lethal dose of a neurovirulent strain of CDV were protected. However, approximately 30% of the mice vaccinated with the CDV F DNA became obese in the first 2 months following the challenge. This was not correlated with the serum antibody levels.
Anand, Setty Balakrishnan; Murugan, Vadivel; Prabhu, Prince Rajaiah; Anandharaman, Veerabhadran; Reddy, Maryada Venkata Rami; Kaliraj, Perumal
Although DNA vaccines have several advantages over conventional vaccines, antibody production and protection are often not adequate, particularly in single plasmid vaccine formulation. In the present study we evaluated the efficacy of a cocktail vaccine based on plasmids encoding larval (L3) stage-specific Brugia malayi abundant larval transcript (BmALT-2) and antioxidant detoxification enzyme B. malayi thioredoxin peroxidase (BmTPX) to induce antibodies, protective efficacy and cell-mediated immune response in mice. Mice immunized with cocktail DNA vaccines containing the pVAX ALT-2+TPX developed higher titers of anti-BmALT-2+TPX (1:5000) antibodies, compared to the mice immunized with single DNA vaccine of pVAX ALT-2 or pVAX TPX (1:2000). Correlating with this, the mice administered with cocktail vaccine induced up to 78% of cytotoxicity against B. malayi mf. This cytotoxicity was high compared to 34% induced by the pVAX-ALT2 or 37% by pVAX-TPX. Moreover, cocktail vaccination of mice resulted in significantly higher level of cellular proliferative response associated with raised levels of IFN-gamma that skewed towards Th1 type of response compared to vaccination using either of the components. Taken together, these data suggest that the combination of two or more antigens maybe an effective vaccine development strategy to improve protection and immunogenicity against human lymphatic filariasis.
Luo, Deyan; Ni, Bing; Li, Peng; Shi, Wei; Zhang, Songle; Han, Yue; Mao, Liwei; He, Yangdong; Wu, Yuzhang; Wang, Xiliang
This study was designed to evaluate the immunogenicity and the protective efficacy of a divalent fusion DNA vaccine encoding both the Brucella abortus L7/L12 protein (ribosomal protein) and Omp16 protein (outer membrane lipoprotein), designated pcDNA3.1-L7/L12-Omp16. Intramuscular injection of this divalent DNA vaccine into BALB/c mice elicited markedly both humoral and cellular immune responses. The specific antibodies exhibited a dominance of immunoglobulin G2a (IgG2a) over IgG1. In addition, the dual-gene DNA vaccine elicited a strong T-cell proliferative response and induced a large amount of gamma interferon-producing T cells upon restimulation in vitro with recombinant fusion protein L7/L12-Omp16, suggesting the induction of a typical T-helper-1-dominated immune response in vivo. This divalent DNA vaccine could also induce a significant level of protection against challenge with the virulent strain B. abortus 544 in BALB/c mice. Furthermore, the protection level induced by the divalent DNA vaccine was significantly higher than that induced by the univalent DNA vaccines pcDNA3.1-L7/L12 or pcDNA3.1-Omp16. Taken together, the results of this study verify for the first time that the Omp16 gene can be a candidate target for a DNA vaccine against brucellosis. Additionally, a divalent genetic vaccine based on the L7/L12 and Omp16 genes can elicit a stronger cellular immune response and better immunoprotection than the relevant univalent vaccines can.
Chen, Lan; Ewing, Dan; Subramanian, Hemavathy; Block, Karla; Rayner, Jonathan; Alterson, Kimberly D; Sedegah, Martha; Hayes, Curtis; Porter, Kevin; Raviprakash, Kanakatte
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.
da Silva, Robson A.A.; Tavares, Natália M.; Costa, Dirceu; Pitombo, Maiana; Barbosa, Larissa; Fukutani, Kyioshi; Miranda, Jose C.; de Oliveira, Camila I.; Valenzuela, Jesus G.; Barral, Aldina; Soto, Manuel; Barral-Netto, Manoel; Brodskyn, Cláudia
It was recently shown that immunization of hamsters with DNA plasmids coding LJM19, a sand fly salivary protein, partially protected against a challenge with Leishmania chagasi, whereas immunization with KMP11 DNA plasmid, a Leishmania antigen, induced protection against L. donovani infection. In the present study, we evaluated the protective effect of immunization with both LJM19 and KMP11 DNA plasmid together. Concerning the protection against an infection by L. chagasi, immunization with DNA plasmids coding LJM19 or KMP11, as well as with both plasmids combined, induced IFN-γ production in draining lymph nodes at 7, 14 and 21 days post-immunization. Immunized hamsters challenged with L. chagasi plus Salivary Gland Sonicate (SGS) from Lutzomyia longipalpis showed an enhancement of IFN-γ/IL-10 and IFN-γ/TGF-β in draining lymph nodes after 7 and 14 days of infection. Two and five months after challenge, immunized animals showed reduced parasite load in the liver and spleen, as well as increased IFN-γ/IL-10 and IFN-γ/TGF-β ratios in the spleen. Furthermore, immunized animals remained with a normal hematological profile even five months after the challenge, whereas L. chagasi in unimmunized hamsters lead to a significant anemia. The protection observed with LJM19 or KMP11 DNA plasmids used alone was very similar to the protection obtained by the combination of both plasmids. PMID:21875567
Golden, Joseph W.; Maes, Piet; Kwilas, Steven A.; Ballantyne, John
ABSTRACT Several members of the Arenaviridae can cause acute febrile diseases in humans, often resulting in lethality. The use of convalescent-phase human plasma is an effective treatment in humans infected with arenaviruses, particularly species found in South America. Despite this, little work has focused on developing potent and defined immunotherapeutics against arenaviruses. In the present study, we produced arenavirus neutralizing antibodies by DNA vaccination of rabbits with plasmids encoding the full-length glycoprotein precursors of Junín virus (JUNV), Machupo virus (MACV), and Guanarito virus (GTOV). Geometric mean neutralizing antibody titers, as measured by the 50% plaque reduction neutralization test (PRNT50), exceeded 5,000 against homologous viruses. Antisera against each targeted virus exhibited limited cross-species binding and, to a lesser extent, cross-neutralization. Anti-JUNV glycoprotein rabbit antiserum protected Hartley guinea pigs from lethal intraperitoneal infection with JUNV strain Romero when the antiserum was administered 2 days after challenge and provided some protection (∼30%) when administered 4 days after challenge. Treatment starting on day 6 did not protect animals. We further formulated an IgG antibody cocktail by combining anti-JUNV, -MACV, and -GTOV antibodies produced in DNA-vaccinated rabbits. This cocktail protected 100% of guinea pigs against JUNV and GTOV lethal disease. We then expanded on this cocktail approach by simultaneously vaccinating rabbits with a combination of plasmids encoding glycoproteins from JUNV, MACV, GTOV, and Sabia virus (SABV). Sera collected from rabbits vaccinated with the combination vaccine neutralized all four targets. These findings support the concept of using a DNA vaccine approach to generate a potent pan-arenavirus immunotherapeutic. IMPORTANCE Arenaviruses are an important family of emerging viruses. In infected humans, convalescent-phase plasma containing neutralizing antibodies can
Golden, Joseph W; Maes, Piet; Kwilas, Steven A; Ballantyne, John; Hooper, Jay W
Several members of the Arenaviridae can cause acute febrile diseases in humans, often resulting in lethality. The use of convalescent-phase human plasma is an effective treatment in humans infected with arenaviruses, particularly species found in South America. Despite this, little work has focused on developing potent and defined immunotherapeutics against arenaviruses. In the present study, we produced arenavirus neutralizing antibodies by DNA vaccination of rabbits with plasmids encoding the full-length glycoprotein precursors of Junín virus (JUNV), Machupo virus (MACV), and Guanarito virus (GTOV). Geometric mean neutralizing antibody titers, as measured by the 50% plaque reduction neutralization test (PRNT(50)), exceeded 5,000 against homologous viruses. Antisera against each targeted virus exhibited limited cross-species binding and, to a lesser extent, cross-neutralization. Anti-JUNV glycoprotein rabbit antiserum protected Hartley guinea pigs from lethal intraperitoneal infection with JUNV strain Romero when the antiserum was administered 2 days after challenge and provided some protection (∼30%) when administered 4 days after challenge. Treatment starting on day 6 did not protect animals. We further formulated an IgG antibody cocktail by combining anti-JUNV, -MACV, and -GTOV antibodies produced in DNA-vaccinated rabbits. This cocktail protected 100% of guinea pigs against JUNV and GTOV lethal disease. We then expanded on this cocktail approach by simultaneously vaccinating rabbits with a combination of plasmids encoding glycoproteins from JUNV, MACV, GTOV, and Sabia virus (SABV). Sera collected from rabbits vaccinated with the combination vaccine neutralized all four targets. These findings support the concept of using a DNA vaccine approach to generate a potent pan-arenavirus immunotherapeutic. Arenaviruses are an important family of emerging viruses. In infected humans, convalescent-phase plasma containing neutralizing antibodies can mitigate the
Zhang, Weidong; Ahmad1, Gul; Torben, Workineh; Noor, Zahid; Le, Loc; Damian, Raymond T.; Wolf, Roman F.; White, Gary L.; Chavez-Suarez, Maria; Podesta, Ronald B.; Kennedy, Ronald C.; Siddiqui, Afzal A.
No vaccine is available to prevent human schistosomiasis to date. We have targeted a protein of Schistosoma mansoni that plays an important role in the surface membrane renewal process, a mechanism widely believed to be utilized by the parasite as an immune evasion strategy. Sm-p80 antigen is a promising vaccine target because of its documented immunogenicity, protective efficacy and antifecundity effects observed in both experimental murine and nonhuman primate models of this infectious disease. In this study we report that a Sm-p80-based DNA vaccine formulation, in a human use approved vector (VR1020), confers 46% reduction in worm burden in the baboon (Papio anubis) model. Baboons vaccinated with Sm-p80-VR1020 showed 28% decrease in egg production following challenge with the infectious parasite. Sm-p80-VR1020 vaccine elicited robust antigen specific immune responses that included IgG; its subtypes IgG1 and IgG2; IgA and IgM in vaccinated animals. Peripheral blood mononuclear cells (PMBCs) and splenocytes from baboons vaccinated with Sm-p80-VR1020 when stimulated in vitro with recombinant Sm-p80 produced considerably higher levels of Th1 response enhancing cytokines (IL-2, IFN-γ) than Th2 response enhancing cytokines (IL-4, IL-10). PBMCs produced significantly higher number of spot forming units (SFU) for INF-γ than for IL-4 in enzyme-linked immunosorbent spot (ELISPOT) assays. A mixed Th1/Th2 type of humoral and T cell responses were generated following immunization with Sm-p80-VR1020. These findings again highlight the potential of Sm-p80 as a promising vaccine candidate for schistosomiasis. PMID:20187746
Schneeweiss, Anne; Chabierski, Stefan; Salomo, Mathias; Delaroque, Nicolas; Al-Robaiy, Samiya; Grunwald, Thomas; Bürki, Kurt; Liebert, Uwe G; Ulbert, Sebastian
West Nile Virus (WNV) is an emerging pathogenic flavivirus with increasing distribution worldwide. Birds are the natural host of the virus, but also mammals, including humans, can be infected. In some cases, a WNV infection can be associated with severe neurological symptoms. All currently available WNV vaccines are in the veterinary sector, and there is a need to develop safe and effective immunization technologies, which can also be used in humans. An alternative to current vaccination methods is DNA immunization. Most current DNA vaccine candidates against flaviviruses simultaneously express the viral envelope (E) and membrane (prM) proteins, which leads to the formation of virus-like particles. Here we generated a DNA plasmid, which expresses only the E-protein ectodomain. Vaccination of mice stimulated anti-WNV T-cell responses and neutralizing antibodies that were higher than those obtained after immunizing with a recombinant protein previously shown to be a protective WNV vaccine. A single dose of the plasmid was sufficient to protect animals from a lethal challenge with the virus. Moreover, immunogenicity could be boosted when DNA injection was followed by immunization with recombinant domain DIII of the E-protein. This resulted in significantly enhanced neutralizing antibody titers and a more prominent cellular immune response. The results suggest that the WNV E-protein is sufficient as a protective antigen in DNA vaccines and that protection can be significantly improved by adding a recombinant protein boost to the DNA prime.
LaPatra, Scott E.; Corbeil, Serge; Jones, Gerald R.; Shewmaker, William D.; Kurath, Gael
A dose–response study that used the DNA vaccine pIHNw-G against infectious hematopoietic necrosis virus (IHNV) showed that complete and highly significant (P < 0.001) protection against a virus injection challenge can be attained in subyearling rainbow trout Oncorhynchus mykiss (145–160 g, 8- to 10-months-old) 6 weeks after a single intramuscular injection with doses as low as 1 μg. Complete protection was also reproducibly demonstrated at higher vaccine doses; however, no protection was observed with a 0.1-μg dose. Virus-neutralizing antibody titers were detected in fish that had been vaccinated with different doses of the DNA vaccine and then sham-infected; there appeared to be a dose-dependent effect, with higher titers obtained with higher doses of vaccine. The DNA-vaccinated animals that survived virus challenge had significantly (P < 0.05) higher neutralizing antibody titers than sham-infected, DNA-vaccinated control fish. Additionally, the titers detected in the IHN survivors exhibited a significant (P < 0.05) dose-dependent effect, with the highest titers being present in fish that received the highest vaccine doses.
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
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.
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.
Xu, Ying; Yang, Enzhuo; Wang, Jianguang; Li, Rui; Li, Guanghua; Liu, Guoyuan; Song, Na; Huang, Qi; Kong, Cong; Wang, Honghai
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.
Fact Sheet Hepatitis B Vaccination Protection Hepatitis B virus (HBV) is a pathogenic microorganism that can cause potentially life- threatening disease in humans. HBV infection is transmitted through exposure ...
Singh, Dinesh K; Liu, Zhenqian; Sheffer, Darlene; Mackay, Glenn A; Smith, Marilyn; Dhillon, Sukhbir; Hegde, Ramakrishna; Jia, Fenglan; Adany, Istvan; Narayan, Opendra
Simian/human immunodeficiency virus SHIV(KU2) replicates with extremely high titers in macaques. In order to determine whether the DNA of the viral genome could be used as a vaccine if the DNA were rendered noninfectious, we deleted the reverse transcriptase gene from SHIVKU2 and inserted this DNA (DeltartSHIVKU2) into a plasmid that was then used to test gene expression and immunogenicity. Transfection of Jurkat and human embryonic kidney epithelial (HEK 293) cells with the DNA resulted in production of all of the major viral proteins and their precursors and transient export of a large quantity of the Gag p27 into the supernatant fluid. As expected, no infectious virus was produced in these cultures. Four macaques were injected intradermally with 2 mg of the DNA at 0, 8, and 18 weeks. The animals developed neutralizing antibodies and low enzyme-linked immunospot assay (E-SPOT) titers against SHIVKU2. These four animals and two unvaccinated control animals were then challenged with heterologous SHIV89.6P administered into their rectums. The two control animals developed viral RNA titers exceeding 10(6) copies/ml of plasma, and these titers were accompanied by the loss of CD4+ T cells by 2 weeks after challenge. The two control animals died at weeks 8 and 16, respectively. All four of the immunized animals became infected with the challenge virus but developed lower titers of viral RNA in plasma than the control animals, and the titers decreased over time in three of the four macaques. The fourth animal remained viremic and died at week 47. Whereas the control animals failed to develop E-SPOT responses, all four of the immunized animals developed anamnestic E-SPOT responses after challenge. The animal that died developed the highest E-SPOT response and was the only one that produced neutralizing antibodies against the challenge virus. These results established that noninfectious DNA of pathogenic SHIV could be used as a vaccine to prevent AIDS, even though the
Rivas-Santiago, Bruno; Cervantes-Villagrana, Alberto R
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.
Borggren, Marie; Nielsen, Jens; Bragstad, Karoline; Karlsson, Ingrid; Krog, Jesper S; Williams, James A; Fomsgaard, Anders
The threat posed by the 2009 pandemic H1N1 virus emphasized the need for new influenza A virus vaccines inducing a broad cross-protective immune response for use in both humans and pigs. An effective and broad influenza vaccine for pigs would greatly benefit the pork industry and contribute to public health by diminishing the risk of emerging highly pathogenic reassortants. Current inactivated protein vaccines against swine influenza produce only short-lived immunity and have no efficacy against heterologous strains. DNA vaccines are a potential alternative with advantages such as the induction of cellular and humoral immunity, inherent safety and rapid production time. We have previously developed a DNA vaccine encoding selected influenza proteins of pandemic origin and demonstrated broad protective immune responses in ferrets and pigs. In this study, we evaluated our DNA vaccine expressed by next-generation vectors. These new vectors can improve gene expression, but they are also efficiently produced on large scales and comply with regulatory guidelines by avoiding antibiotic resistance genes. In addition, a new needle-free delivery of the vaccine, convenient for mass vaccinations, was compared with intradermal needle injection followed by electroporation. We report that when our DNA vaccine is expressed by the new vectors and delivered to the skin with the needle-free device in the rabbit model, it can elicit an antibody response with the same titers as a conventional vector with intradermal electroporation. The needle-free delivery is already in use for traditional protein vaccines in pigs but should be considered as a practical alternative for the mass administration of broadly protective influenza DNA vaccines.
Kato, Goshi; Yamashita, Kozue; Kondo, Hidehiro; Hirono, Ikuo
Photobacterium damselae subsp. piscicida (Pdp) kills many cultured marine fish. As it evolves resistance to existing vaccines, new vaccines are needed. PPA1 is a major antigenic protein of Pdp. Here, DNA vaccines encoding wild-type PPA1 (pPPA1(wt)) and codon-optimized PPA1 (pPPA1(opt)) were constructed and tested against Pdp in Japanese flounder. The mRNA levels of the two antigenic genes at the vaccination site were not different, but the protein level was significantly higher in the pPPA1(opt)-vaccinated fish. In addition, after a bacterial challenge, the levels of interleukin (IL)-1β, IL-6 and IFN-γ mRNA significantly increased in the pPPA1(opt)-vaccinated fish but not in the pPPA1(wt)-vaccinated fish. The relative percent survival (RPS) after the challenge was higher in the pPPA1(opt)-vaccinated fish (90.9) than in the pPPA1(wt)-vaccinated fish (69.2). At the early stage of the infection after the challenge, the number of viable Pdp in the spleen was significantly lower in the pPPA1(opt)-vaccinated fish than in the pPPA1(wt)-vaccinated fish. These data show that codon-optimized DNA vaccine pPPA1(opt) had a strong immunogenicity and conferred protective efficacy against Pdp infection in Japanese flounder.
Haese, Nicole; Brocato, Rebecca L; Henderson, Thomas; Nilles, Matthew L; Kwilas, Steve A; Josleyn, Matthew D; Hammerbeck, Christopher D; Schiltz, James; Royals, Michael; Ballantyne, John; Hooper, Jay W; Bradley, David S
Andes virus (ANDV) and ANDV-like viruses are responsible for most hantavirus pulmonary syndrome (HPS) cases in South America. Recent studies in Chile indicate that passive transfer of convalescent human plasma shows promise as a possible treatment for HPS. Unfortunately, availability of convalescent plasma from survivors of this lethal disease is very limited. We are interested in exploring the concept of using DNA vaccine technology to produce antiviral biologics, including polyclonal neutralizing antibodies for use in humans. Geese produce IgY and an alternatively spliced form, IgYΔFc, that can be purified at high concentrations from egg yolks. IgY lacks the properties of mammalian Fc that make antibodies produced in horses, sheep, and rabbits reactogenic in humans. Geese were vaccinated with an ANDV DNA vaccine encoding the virus envelope glycoproteins. All geese developed high-titer neutralizing antibodies after the second vaccination, and maintained high-levels of neutralizing antibodies as measured by a pseudovirion neutralization assay (PsVNA) for over 1 year. A booster vaccination resulted in extraordinarily high levels of neutralizing antibodies (i.e., PsVNA80 titers >100,000). Analysis of IgY and IgYΔFc by epitope mapping show these antibodies to be highly reactive to specific amino acid sequences of ANDV envelope glycoproteins. We examined the protective efficacy of the goose-derived antibody in the hamster model of lethal HPS. α-ANDV immune sera, or IgY/IgYΔFc purified from eggs, were passively transferred to hamsters subcutaneously starting 5 days after an IM challenge with ANDV (25 LD50). Both immune sera, and egg-derived purified IgY/IgYΔFc, protected 8 of 8 and 7 of 8 hamsters, respectively. In contrast, all hamsters receiving IgY/IgYΔFc purified from normal geese (n=8), or no-treatment (n=8), developed lethal HPS. These findings demonstrate that the DNA vaccine/goose platform can be used to produce a candidate antiviral biological product
Henderson, Thomas; Nilles, Matthew L.; Kwilas, Steve A.; Josleyn, Matthew D.; Hammerbeck, Christopher D.; Schiltz, James; Royals, Michael; Ballantyne, John; Hooper, Jay W.; Bradley, David S.
Andes virus (ANDV) and ANDV-like viruses are responsible for most hantavirus pulmonary syndrome (HPS) cases in South America. Recent studies in Chile indicate that passive transfer of convalescent human plasma shows promise as a possible treatment for HPS. Unfortunately, availability of convalescent plasma from survivors of this lethal disease is very limited. We are interested in exploring the concept of using DNA vaccine technology to produce antiviral biologics, including polyclonal neutralizing antibodies for use in humans. Geese produce IgY and an alternatively spliced form, IgYΔFc, that can be purified at high concentrations from egg yolks. IgY lacks the properties of mammalian Fc that make antibodies produced in horses, sheep, and rabbits reactogenic in humans. Geese were vaccinated with an ANDV DNA vaccine encoding the virus envelope glycoproteins. All geese developed high-titer neutralizing antibodies after the second vaccination, and maintained high-levels of neutralizing antibodies as measured by a pseudovirion neutralization assay (PsVNA) for over 1 year. A booster vaccination resulted in extraordinarily high levels of neutralizing antibodies (i.e., PsVNA80 titers >100,000). Analysis of IgY and IgYΔFc by epitope mapping show these antibodies to be highly reactive to specific amino acid sequences of ANDV envelope glycoproteins. We examined the protective efficacy of the goose-derived antibody in the hamster model of lethal HPS. α-ANDV immune sera, or IgY/IgYΔFc purified from eggs, were passively transferred to hamsters subcutaneously starting 5 days after an IM challenge with ANDV (25 LD50). Both immune sera, and egg-derived purified IgY/IgYΔFc, protected 8 of 8 and 7 of 8 hamsters, respectively. In contrast, all hamsters receiving IgY/IgYΔFc purified from normal geese (n=8), or no-treatment (n=8), developed lethal HPS. These findings demonstrate that the DNA vaccine/goose platform can be used to produce a candidate antiviral biological product
Hartl, A; Hochreiter, R; Stepanoska, T; Ferreira, F; Thalhamer, J
An estimated 100 million individuals suffer from birch pollen allergy. More than 95% of birch pollen-allergic subjects react with the major birch pollen allergen Bet v 1a, and almost 60% of them are sensitized exclusively to this allergen. DNA immunization using the Bet v 1a gene was evaluated with respect to its prophylactic and therapeutic efficacy. A DNA vaccine containing the entire Bet v 1a cDNA under the control of a CMV-promoter was constructed. In order to estimate the protective efficiency, animals received three injections of this vaccine prior to sensitization with recombinant Bet v 1a. Vice versa, in a therapeutic approach, sensitization was followed by treatment with the DNA vaccine. The Bet v 1a DNA vaccine induced strong Bet v 1-specific antibody responses with a Th1-biased response type. Animals which received the DNA vaccine were protected against a following allergic sensitization with Bet v 1a. The protective effect was characterized by suppression of Bet v 1-specific immunoglobulin (Ig)E production, lack of basophil activation and enhanced interferon (IFN)-gamma expression. In a therapeutic situation, treatment of sensitized animals with DNA vaccines decreased IgE production, IgE-mediated basophil release and drastically reduced anaphylactic activity as measured by passive cutaneous anaphylaxis assays. Concerning the cellular immune response, DNA immunization induced a sustaining and dominant shift from a Th2 type response towards a balanced Th1/Th2 type response as indicated by increased IFN-gamma but unchanged IL-5 levels in lymphoproliferation assays. The results demonstrate the allergen-specific protective and therapeutic efficacy of a DNA vaccine encoding the clinically highly relevant allergen Bet v 1a indicating the suitability of this concept for the treatment of allergic diseases.
Lu, Yu-Chun; Li, Min-Chen; Chen, Yi-Min; Chu, Chun-Yen; Lin, Shuen-Fuh; Yang, Wen-Jen
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.
Farrag, Mohamed A; Amer, Haitham M; Öhlschläger, Peter; Hamad, Maaweya E; Almajhdi, Fahad N
The development of safe and potent vaccines for human respiratory syncytial virus (HRSV) is still a challenge for researchers worldwide. DNA-based immunization is currently a promising approach that has been used to generate human vaccines for different age groups. In this study, novel HRSV DNA vaccine candidates were generated and preclinically tested in BALB/c mice. Three different versions of the codon-optimized HRSV fusion (F) gene were individually cloned into the pPOE vector. The new recombinant vectors either express full-length (pPOE-F), secretory (pPOE-TF), or M282-90 linked (pPOE-FM2) forms of the F protein. Distinctive expression of the F protein was identified in HEp-2 cells transfected with the different recombinant vectors using ELISA and immunofluorescence. Mice immunization verified the potential for recombinant vectors to elicit significant levels of neutralizing antibodies and CD8(+) T-cell lymphocytes. pPOE-TF showed higher levels of gene expression in cell culture and better induction of the humoral and cellular immune responses. Following virus challenge, mice that had been immunized with the recombinant vectors were able to control virus replication and displayed lower inflammation compared with mice immunized with empty pPOE vector or formalin-inactivated HRSV vaccine. Moreover, pulmonary cytokine profiles of mice immunized with the 3 recombinant vectors were similar to those of the mock infected group. In conclusion, recombinant pPOE vectors are promising HRSV vaccine candidates in terms of their safety, immunogenicity and protective efficiency. These data encourage further evaluation in phase I clinical trials.
Cervantes-Villagrana, Alberto R; Hernández-Pando, Rogelio; Biragyn, Arya; Castañeda-Delgado, Julio; Bodogai, Monica; Martínez-Fierro, Margarita; Sada, Eduardo; Trujillo, Valentin; Enciso-Moreno, Antonio; Rivas-Santiago, Bruno
The World Health Organization (WHO) has estimated that there are about 8 million new cases annually of active Tuberculosis (TB). Despite its irregular effectiveness (0-89%), the Bacillus Calmette-Guérin) BCG is the only vaccine available worldwide for prevention of TB; thus, the design is important of novel and more efficient vaccination strategies. Considering that β-defensin-2 is an antimicrobial peptide that induces dendritic cell maturation through the TLR-4 receptor and that both ESAT-6 and Ag85B are immunodominant mycobacterial antigens and efficient activators of the protective immune response, we constructed two DNA vaccines by the fusion of the gene encoding β-defensin-2 and antigens ESAT6 (pDE) and 85B (pDA). After confirming efficient local antigen expression that induced high and stable Interferon gamma (IFN-γ) production in intramuscular (i.m.) vaccinated Balb/c mice, groups of mice were vaccinated with DNA vaccines in a prime-boost regimen with BCG and with BCG alone, and 2 months later were challenged with the mild virulence reference strain H37Rv and the highly virulent clinical isolate LAM 5186. The level of protection was evaluated by survival, lung bacilli burdens, and extension of tissue damage (pneumonia). Vaccination with both DNA vaccines showed similar protection to that of BCG. After the challenge with the highly virulent Mycobacterium tuberculosis strain, animals that were prime-boosted with BCG and then boosted with both DNA vaccines showed significant higher survival and less tissue damage than mice vaccinated only with BCG. These results suggest that improvement of BCG vaccination, such as the prime-boost DNA vaccine, represents a more efficient vaccination scheme against TB.
Maksiutov, R A; Shchelkunov, S N
Efficacy of candidate DNA-vaccines based on the variola virus natural gene A30L and artificial gene A30Lopt with modified codon usage, optimized for expression in mammalian cells, was tested. The groups of mice were intracutaneously immunized three times with three-week intervals with candidate DNA-vaccines: pcDNA_A30L or pcDNA_A30Lopt, and in three weeks after the last immunization all mice in the groups were intraperitoneally infected by the ectromelia virus K1 strain in 10 LD50 dose for the estimation of protection. It was shown that the DNA-vaccines based on natural gene A30L and codon-optimized gene A30Lopt elicited virus, thereby neutralizing the antibody response and protected mice from lethal intraperitoneal challenge with the ectromelia virus with lack of statistically significant difference.
Corbeil, S.; LaPatra, S.E.; Anderson, E.D.; Jones, J.; Vincent, B.; Hsu, Ya Li; Kurath, G.
The protective immunogenicity of the nucleoprotein (N), phosphoprotein (P), matrix protein (M), non-virion protein (NV) and glycoprotein (G) of the rhabdovirus infectious hematopoietic necrosis virus (IHNV) was assessed in rainbow trout using DNA vaccine technology. DNA vaccines were produced by amplifying and cloning the viral genes in the plasmid pCDNA 3.1. The protective immunity elicited by each vaccine was evaluated through survival of immunized fry after challenge with live virus. Neutralizing antibody titers were also determined in vaccinated rainbow troutOncorhynchus mykiss fry (mean weight 2 g) and 150 g sockeye salmon Oncorhynchus nerka. The serum from the 150 g fish was also used in passive immunization studies with naïve fry. Our results showed that neither the internal structural proteins (N, P and M) nor the NV protein of IHNV induced protective immunity in fry or neutralizing antibodies in fry and 150 g fish when expressed by a DNA vaccine construct. The G protein, however, did confer significant protection in fry up to 80 d post-immunization and induced protective neutralizing antibodies. We are currently investigating the role of different arms of the fish immune system that contribute to the high level of protection against IHNV seen in vaccinated fish.
Cherif, Mahamoud Sama; Shuaibu, Mohammed Nasir; Kodama, Yukinobu; Kurosaki, Tomoaki; Helegbe, Gideon Kofi; Kikuchi, Mihoko; Ichinose, Akitoyo; Yanagi, Tetsuo; Sasaki, Hitoshi; Yui, Katsuyuki; Tien, Nguyen Huy; Karbwang, Juntra; Hirayama, Kenji
We have previously reported the new formulation of polyethylimine (PEI) with gamma polyglutamic acid (γ-PGA) nanoparticle (NP) to have provided Plasmodium yoelii merozoite surface protein-1 (PyMSP-1) plasmid DNA vaccine with enhanced protective cellular and humoral immunity in the lethal mouse malaria model. PyGPI8p-transamidase-related protein (PyTAM) was selected as a possible candidate vaccine antigen by using DNA vaccination screening from 29 GPI anchor and signal sequence motif positive genes picked up using web-based bioinformatics tools; though the observed protection was not complete. Here, we observed augmented protective effect of PyTAM DNA vaccine by using PEI and γ-PGA complex as delivery system. NP-coated PyTAM plasmid DNA immunized mice showed a significant survival rate from lethal P. yoelii challenge infection compared with naked PyTAM plasmid or with NP-coated empty plasmid DNA group. Antigen-specific IgG1 and IgG2b subclass antibody levels, proportion of CD4 and CD8T cells producing IFN-γ in the splenocytes and IL-4, IFN-γ, IL-12 and TNF-α levels in the sera and in the supernatants from ex vivo splenocytes culture were all enhanced by the NP-coated PyTAM DNA vaccine. These data indicates that NP augments PyTAM protective immune response, and this enhancement was associated with increased DC activation and concomitant IL-12 production.
Jain, Shikha; Afley, Prachiti; Dohre, Sudhir K; Saxena, Nandita; Kumar, Subodh
Brucellosis is a worldwide zoonotic disease. No Brucella vaccine is available for use in humans and existing animal vaccines have limitations. We have previously described the ribosomal protein L9 to have the vaccine potential. In this study, L9 based DNA vaccine (pVaxL9) was generated and evaluated in mouse model. Intramuscular immunisation of pVaxL9 was able to elicit the anti-L9 IgG antibody response of both IgG1 and IgG2a isotypes when compared with PBS and pVax immunised control animals. Heightened antibody response was observed in mice groups immunised with pVaxL9 priming and recombinant L9 boosting (PB) and where pDNA immunisation was carried out by in vivo electroporation (EP). The vaccine groups proliferated splenocytes and released Th1 type cytokines e.g. IFN-γ, TNF-α, IL-2. Further, flow cytometric analysis revealed that IFN-γ was released by both by CD4+ and CD8+ T cells particularly in PB and EP groups when compared with mice immunised with empty control vector. The L9 based pDNA vaccine was able to confer significant protection in mice against challenge with virulent B. abortus with PB and EP groups offering better protection. Taken together, it can be concluded that L9 based DNA vaccine is immunogenic and confer protection in mouse model.
An oral DNA vaccine against infectious haematopoietic necrosis virus (IHNV) encapsulated in alginate microspheres induces dose-dependent immune responses and significant protection in rainbow trout (Oncorrhynchus mykiss).
Ballesteros, Natalia A; Alonso, Marta; Saint-Jean, Sylvia Rodríguez; Perez-Prieto, Sara I
Administered by intramuscular injection, a DNA vaccine (pIRF1A-G) containing the promoter regions upstream of the rainbow trout interferon regulatory factor 1A gene (IRF1A) driven the expression of the infectious hematopoietic necrosis virus (IHNV) glycoprotein (G) elicited protective immune responses in rainbow trout (Oncorhynchus mykiss). However, less laborious and cost-effective routes of DNA vaccine delivery are required to vaccinate large numbers of susceptible farmed fish. In this study, the pIRF1A-G vaccine was encapsulated into alginate microspheres and orally administered to rainbow trout. At 1, 3, 5, and 7 d post-vaccination, IHNV G transcripts were detected by quantitative real-time PCR in gills, spleen, kidney and intestinal tissues of vaccinated fish. This result suggested that the encapsulation of pIRF1A-G in alginate microparticles protected the DNA vaccine from degradation in the fish stomach and ensured vaccine early delivery to the hindgut, vaccine passage through the intestinal mucosa and its distribution thought internal and external organs of vaccinated fish. We also observed that the oral route required approximately 20-fold more plasmid DNA than the injection route to induce the expression of significant levels of IHNV G transcripts in kidney and spleen of vaccinated fish. Despite this limitation, increased IFN-1, TLR-7 and IgM gene expression was detected by qRT-PCR in kidney of vaccinated fish when a 10 μg dose of the oral pIRF1A-G vaccine was administered. In contrast, significant Mx-1, Vig-1, Vig-2, TLR-3 and TLR-8 gene expression was only detected when higher doses of pIRF1A-G (50 and 100 μg) were orally administered. The pIRF1A-G vaccine also induced the expression of several markers of the adaptive immune response (CD4, CD8, IgM and IgT) in kidney and spleen of immunized fish in a dose-dependent manner. When vaccinated fish were challenged by immersion with live IHNV, evidence of a dose-response effect of the oral vaccine could also
Nielsen, Line; Jensen, Trine Hammer; Kristensen, Birte; Jensen, Tove Dannemann; Karlskov-Mortensen, Peter; Lund, Morten; Aasted, Bent; Blixenkrone-Møller, Merete
Immunity induced by DNA vaccines containing the hemagglutinin (H) and nucleoprotein (N) genes of wild-type and attenuated canine distemper virus (CDV) was investigated in mink (Mustela vison), a highly susceptible natural host of CDV. All DNA-immunized mink seroconverted, and significant levels of virus-neutralizing (VN) antibodies were present on the day of challenge with wild-type CDV. The DNA vaccines also primed the cell-mediated memory responses, as indicated by an early increase in the number of interferon-gamma (IFN-γ)-producing lymphocytes after challenge. Importantly, the wild-type and attenuated CDV DNA vaccines had a long-term protective effect against wild-type CDV challenge. The vaccine-induced immunity induced by the H and N genes from wild-type CDV and those from attenuated CDV was comparable. Because these two DNA vaccines were shown to protect equally well against wild-type virus challenge, it is suggested that the genetic/antigenic heterogeneity between vaccine strains and contemporary wild-type strains are unlikely to cause vaccine failure.
Wang, Shixia; Goguen, Jon D; Li, Fusheng; Lu, Shan
Yersinia pestis (Y. pestis) is the causative pathogen of plague, a highly fatal disease for which an effective vaccine, especially against mucosal transmission, is still not available. Like many bacterial infections, antigen-specific antibody responses have been traditionally considered critical, if not solely responsible, for vaccine-induced protection against Y. pestis. Studies in recent years have suggested the importance of T cell immune responses against Y. pestis infection but information is still limited about the details of Y. pestis antigen-specific T cell immune responses. In current report, studies are conducted to identify the presence of CD8+ T cell epitopes in LcrV protein, the leading antigen of plague vaccine development. Furthermore, depletion of CD8+ T cells in LcrV DNA vaccinated Balb/C mice led to reduced protection against lethal intranasal challenge of Y. pestis. These findings establish that an LcrV DNA vaccine is able to elicit CD8+ T cell immune responses against specific epitopes of this key plague antigen and that a CD8+ T cell immune response is involved in LcrV DNA vaccine-elicited protection. Future studies in plague vaccine development will need to examine if the presence of detectable T cell immune responses, in particular CD8+ T-cell immune responses, will enhance the protection against Y. pestis in higher animal species or humans. Copyright © 2010 Elsevier Ltd. All rights reserved.
Hwang, Hye Suk; Kwon, Young-Man; Lee, Jong Seok; Yoo, Si-Eun; Lee, Yu-Na; Ko, Eun-Ju; Kim, Min-Chul; Cho, Min-Kyoung; Lee, Young-Tae; Jung, Yu-Jin; Lee, Ji-Yun; Li, Jian Dong; Kang, Sang-Moo
This study demonstrates that immunization with non-replicating virus-like particle (FFG VLP) containing RSV F and G glycoproteins together with RSV F DNA induced T helper type 1 antibody responses to RSV F similar to live RSV infection. Upon RSV challenge 21 weeks after immunization, FFG VLP vaccination induced protection against RSV infection as shown by clearance of lung viral loads, and the absence of eosinophil infiltrates, and did not cause lung pathology. In contrast, formalin-inactivated RSV (FI-RSV) vaccination showed significant pulmonary eosinophilia, severe mucus production, and extensive histopathology resulting in a hallmark of pulmonary pathology. Substantial lung pathology was also observed in mice with RSV re-infections. High levels of systemic and local inflammatory cytokine-secreting cells were induced in mice with FI-RSV but not with FFG VLP immunization after RSV challenge. Therefore, the results provide evidence that recombinant RSV FFG VLP vaccine can confer long-term protection against RSV without causing lung pathology. PMID:25110201
Hwang, Hye Suk; Kwon, Young-Man; Lee, Jong Seok; Yoo, Si-Eun; Lee, Yu-Na; Ko, Eun-Ju; Kim, Min-Chul; Cho, Min-Kyoung; Lee, Young-Tae; Jung, Yu-Jin; Lee, Ji-Yun; Li, Jian-Dong; Kang, Sang-Moo
This study demonstrates that immunization with non-replicating virus-like particle (FFG VLP) containing RSV F and G glycoproteins together with RSV F DNA induced T helper type 1 antibody responses to RSV F similar to live RSV infection. Upon RSV challenge 21weeks after immunization, FFG VLP vaccination induced protection against RSV infection as shown by clearance of lung viral loads, and the absence of eosinophil infiltrates, and did not cause lung pathology. In contrast, formalin-inactivated RSV (FI-RSV) vaccination showed significant pulmonary eosinophilia, severe mucus production, and extensive histopathology resulting in a hallmark of pulmonary pathology. Substantial lung pathology was also observed in mice with RSV re-infections. High levels of systemic and local inflammatory cytokine-secreting cells were induced in mice with FI-RSV but not with FFG VLP immunization after RSV challenge. Therefore, the results provide evidence that recombinant RSV FFG VLP vaccine can confer long-term protection against RSV without causing lung pathology.
Background Pigs have been implicated as mixing reservoir for the generation of new pandemic influenza strains, control of swine influenza has both veterinary and public health significance. Unlike human influenza vaccines, strains used for commercially available swine influenza vaccines are not regularly replaced, making the vaccines provide limited protection against antigenically diverse viruses. It is therefore necessary to develop broadly protective swine influenza vaccines that are efficacious to both homologous and heterologous virus infections. In this study, two forms of DNA vaccines were constructed, one was made by fusing M2e to consensus H3HA (MHa), which represents the majority of the HA sequences of H3N2 swine influenza viruses. Another was made by fusing M2e and a conserved CTL epitope (NP147-155) to consensus H3HA (MNHa). Their protective efficacies against homologous and heterologous challenges were tested. Results BALB/c mice were immunized twice by particle-mediated epidermal delivery (gene gun) with the two DNA vaccines. It was shown that the two vaccines elicited substantial antibody responses, and MNHa induced more significant T cell-mediated immune response than MHa did. Then two H3N2 strains representative of different evolutional and antigenic clusters were used to challenge the vaccine-immunized mice (homosubtypic challenge). Results indicated that both of the DNA vaccines prevented homosubtypic virus infections completely. The vaccines’ heterologous protective efficacies were further tested by challenging with a H1N1 swine influenza virus and a reassortant 2009 pandemic strain. It was found that MNHa reduced the lung viral titers significantly in both challenge groups, histopathological observation showed obvious reduction of lung pathogenesis as compared to MHa and control groups. Conclusions The combined utility of the consensus HA and the conserved M2e and CTL epitope can confer complete and partial protection against homologous and
Kaur, Manpreet; Saxena, Ankur; Rai, Anant; Bhatnagar, Rakesh
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 an improved rabies vaccine. Thus, DNA vaccine based on lysosome-targeted glycoprotein of the rabies virus was evaluated in BALB/c mice. It imparted partial protection (60%) against challenge with 20 LD(50) of the challenge virus standard (CVS) strain of rabies virus. To improve the outcome of vaccination, to ultimately enhance the immune response, we investigated different routes for DNA vaccine delivery, varied doses of DNA, and the influence of adjuvant supplementation. The highest immune response pertaining to IgG antibody titer, with a predominantly IgG1/IgG2a subclass distribution, effective cellular immunity, and a high level of rabies virus neutralizing antibodies (RVNAs) was attained by the optimized DNA vaccine formulation comprising intramuscular administration of 100 microg of DNA vaccine supplemented with Emulsigen-D. In preexposure prophylaxis, a 3-dose regimen of this formulation generated a high RVNA titer (32 IU/ml) and conferred complete protection against challenge with 20 LD(50) of CVS. For postexposure efficacy analysis, rabies was experimentally induced with 50 LD(50) of CVS. Subsequent therapy with 5 doses of the formulation completely prevented rabies in BALB/c mice, which maintained protective RVNA titers of 4 IU/ml. The World Health Organization recommended rabies protective titer threshold is 0.5 IU/ml. Thus, this optimized DNA vaccine formulation provides an avenue for preventing and controlling rabies.
Wang, Wenling; Li, Renqing; Deng, Yao; Lu, Ning; Chen, Hong; Meng, Xin; Wang, Wen; Wang, Xiuping; Yan, Kexia; Qi, Xiangrong; Zhang, Xiangmin; Xin, Wei; Lu, Zhenhua; Li, Xueren; Bian, Tao; Gao, Yingying; Tan, Wenjie; Ruan, Li
The conventional hemagglutinin (HA)- and neuraminidase (NA)-based influenza vaccines need to be updated most years and are ineffective if the glycoprotein HA of the vaccine strains is a mismatch with that of the epidemic strain. Universal vaccines targeting conserved viral components might provide cross-protection and thus complement and improve conventional vaccines. In this study, we generated DNA plasmids and recombinant vaccinia viruses expressing the conserved proteins nucleoprotein (NP), polymerase basic 1 (PB1), and matrix 1 (M1) from influenza virus strain A/Beijing/30/95 (H3N2). BALB/c mice were immunized intramuscularly with a single vaccine based on NP, PB1, or M1 alone or a combination vaccine based on all three antigens and were then challenged with lethal doses of the heterologous influenza virus strain A/PR/8/34 (H1N1). Vaccines based on NP, PB1, and M1 provided complete or partial protection against challenge with 1.7 50% lethal dose (LD50) of PR8 in mice. Of the three antigens, NP-based vaccines induced protection against 5 LD50 and 10 LD50 and thus exhibited the greatest protective effect. Universal influenza vaccines based on the combination of NP, PB1, and M1 induced a strong immune response and thus might be an alternative approach to addressing future influenza virus pandemics.
Lin, Ken; Roosinovich, Elena; Ma, Barbara; Hung, Chien-Fu
It is now well established that most cervical cancers are causally associated with HPV infection. This realization has led to efforts to control HPV-associated malignancy through prevention or treatment of HPV infection. Currently, commercially available HPV vaccines are not designed to control established HPV infection and associated premalignant and malignant lesions. To treat and eradicate pre-existing HPV infections and associated lesions which remain prevalent in the U.S. and worldwide, effective therapeutic HPV vaccines are needed. DNA vaccination has emerged as a particularly promising form of therapeutic HPV vaccines due to its safety, stability and ability to induce antigen-specific immunity. This review focuses on improving the potency of therapeutic HPV vaccines through modification of dendritic cells (DCs) by  increasing the number of antigen-expressing/antigen-loaded DCs,  improving HPV antigen expression, processing and presentation in DCs, and  enhancing DC and T cell interaction. Continued improvement in therapeutic HPV DNA vaccines may ultimately lead to an effective DNA vaccine for the treatment of HPV-associated malignancies. PMID:20066511
Li, Wenfang; Ye, Ling; Carrion, Ricardo; Mohan, Gopi S; Nunneley, Jerritt; Staples, Hilary; Ticer, Anysha; Patterson, Jean L; Compans, Richard W; Yang, Chinglai
In addition to its surface glycoprotein (GP), Ebola virus directs the production of large quantities of a truncated glycoprotein isoform (sGP) that is secreted into the extracellular space. We recently reported that sGP actively diverts host antibody responses against the epitopes that it shares with GP and thereby allows itself to absorb anti-GP antibodies, a phenomenon we termed "antigenic subversion." To investigate the effect of antigenic subversion by sGP on protection against virus infection, we compared immune responses induced by different prime-boost immunization regimens with GP and sGP DNA vaccines in mice and their efficacy against lethal Ebola virus challenge. Similar levels of anti-GP antibodies were induced by 2 immunizations with sGP and GP DNA vaccines. However, 2 immunizations with GP but not sGP DNA vaccine fully protected mice from lethal challenge. Boosting with sGP or GP DNA vaccine in mice that had been primed by GP or sGP DNA vaccine augmented the levels of anti-GP antibody responses and further improved protective efficacy against Ebola virus infection. These results show that both the quality and the levels of anti-GP antibody responses affect the efficacy of protection against Ebola virus infection.
Li, Wenfang; Ye, Ling; Carrion, Ricardo; Mohan, Gopi S.; Nunneley, Jerritt; Staples, Hilary; Ticer, Anysha; Patterson, Jean L.; Compans, Richard W.; Yang, Chinglai
In addition to its surface glycoprotein (GP), Ebola virus directs the production of large quantities of a truncated glycoprotein isoform (sGP) that is secreted into the extracellular space. We recently reported that sGP actively diverts host antibody responses against the epitopes that it shares with GP and thereby allows itself to absorb anti-GP antibodies, a phenomenon we termed “antigenic subversion.” To investigate the effect of antigenic subversion by sGP on protection against virus infection, we compared immune responses induced by different prime-boost immunization regimens with GP and sGP DNA vaccines in mice and their efficacy against lethal Ebola virus challenge. Similar levels of anti-GP antibodies were induced by 2 immunizations with sGP and GP DNA vaccines. However, 2 immunizations with GP but not sGP DNA vaccine fully protected mice from lethal challenge. Boosting with sGP or GP DNA vaccine in mice that had been primed by GP or sGP DNA vaccine augmented the levels of anti-GP antibody responses and further improved protective efficacy against Ebola virus infection. These results show that both the quality and the levels of anti-GP antibody responses affect the efficacy of protection against Ebola virus infection. PMID:25877553
Negash, Tamiru; Liman, Martin; Rautenschlein, Silke
Infectious bursal disease virus (IBDV) is an immunosuppressive virus of chickens. The virus protein (VP) 2 induces neutralizing antibodies, which protect chickens against the disease. The aim of this study was to develop a cationic poly(d,l-lactide-co-glycolide) (PLGA) microparticle (MP) based IBDV-VP2 DNA vaccine (MP-IBDV-DNA) for chickens to be delivered orally and by eye drop route. The tested IBDV-VP2 DNA vaccines were immunogenic for specific-pathogen-free chickens and induced an antibody response after intramuscular application. Co-inoculation with a plasmid encoding chicken IL-2 (chIL-2) or CpG-ODN did not significantly improve protection against IBDV challenge. However, the application of a MP-IBDV-DNA vaccine alone or in combination with a delayed oral and eye drop application of cationic MP loaded with CpG-ODN or chIL-2 improved protection against challenge. The MP-IBDV-DNA-vaccinated chickens showed less pathological and histopathological bursal lesions, a reduced IBDV antigen load as well as T-cell influx into the bursa of Fabricius (BF) compared to the other groups (p<0.05). The addition of chIL-2 loaded MP improved challenge virus clearance from the BF as demonstrated by lower neutralizing antibody titers and reduced IL-4 and IFN-α mRNA expression in the bursa at 7 days postchallenge compared to the other challenged groups. Overall, the efficacy of the IBDV-DNA vaccine was improved by adsorption of the DNA vaccine onto cationic PLGA-MP, which also allowed mucosal application of the DNA vaccine.
Jiang, Wenbo; Wang, Shuangshuang; Chen, Honglin; Ren, Huanhuan; Huang, Xun; Wang, Guiqin; Chen, Ze; Chen, Ling; Chen, Zhiwei; Zhou, Paul
Current seasonal influenza vaccines are efficacious when vaccine strains are matched with circulating strains. However, they do not protect antigenic variants and newly emerging pandemic and outbreak strains. Thus, there is a critical need for developing so-called "universal" vaccines that protect against all influenza viruses. In the present study, we developed a bivalent heterologous DNA virus-like particle prime-boost vaccine strategy. We show that mice immunized with this vaccine were broadly protected against lethal challenge from group 1 (H1, H5, and H9) and group 2 (H3 and H7) viruses, with 94% aggregate survival. To determine the immune correlates of protection, we performed passive immunizations and in vitro assays. We show that this vaccine elicited antibody responses that bound HA from group 1 (H1, H2, H5, H6, H8, H9, H11, and H12) and group 2 (H3, H4, H7, H10, H14, and H15) and neutralized homologous and intrasubtypic H5 and H7 and heterosubtypic H1 viruses and hemagglutinin-specific CD4 and CD8 T cell responses. As a result, passive immunization with immune sera fully protected mice against H5, H7, and H1 challenge, whereas with both immune sera and T cells the mice survived heterosubtypic H3 and H9 challenge. Thus, it appears that (i) neutralizing antibodies alone fully protect against homologous and intrasubtypic H5 and H7 and (ii) neutralizing and binding antibodies are sufficient to protect against heterosubtypic H1, (iii) but against heterosubtypic H3 and H9, binding antibodies and T cells are required for complete survival. We believe that this vaccine regimen could potentially be a candidate for a "universal" influenza vaccine.IMPORTANCE Influenza virus infection is global health problem. Current seasonal influenza vaccines are efficacious only when vaccine strains are matched with circulating strains. However, these vaccines do not protect antigenic variants and newly emerging pandemic and outbreak strains. Because of this, there is an urgent
Pan, Chien-Hsiung; Nair, Nitya; Adams, Robert J.; Zink, M. Christine; Lee, Eun-Young; Polack, Fernando P.; Singh, Manmohan; O'Hagan, Derek T.; Griffin, Diane E.
Measles remains an important cause of vaccine-preventable child mortality. Development of a low-cost, heat-stable vaccine for infants under the age of 6 months could improve measles control by facilitating delivery at the time of other vaccines and by closing a window of susceptibility prior to immunization at 9 months of age. DNA vaccines hold promise for development, but achieving protective levels of antibody has been difficult and there is an incomplete understanding of protective immunity. In the current study, we evaluated the use of a layered alphavirus DNA/RNA vector encoding measles virus H (SINCP-H) adsorbed onto polylactide glycolide (PLG) microparticles. In mice, antibody and T-cell responses to PLG-formulated DNA were substantially improved compared to those to naked DNA. Rhesus macaques received two doses of PLG/SINCP-H delivered either intramuscularly (0.5 mg) or intradermally (0.5 or 0.1 mg). Antibody and T-cell responses were induced but not sustained. On challenge, the intramuscularly vaccinated monkeys did not develop rashes and had lower viremias than vector-treated control monkeys. Monkeys vaccinated with the same dose intradermally developed rashes and viremia. Monkeys vaccinated intradermally with the low dose developed more severe rashes, with histopathologic evidence of syncytia and intense dermal and epidermal inflammation, eosinophilia, and higher viremia compared to vector-treated control monkeys. Protection after challenge correlated with gamma interferon-producing T cells and with early production of high-avidity antibody that bound wild-type H protein. We conclude that PLG/SINCP-H is most efficacious when delivered intramuscularly but does not provide an advantage over standard DNA vaccines for protection against measles. PMID:18287579
Guan, Jie; Deng, Yao; Chen, Hong; Yin, Xiao; Yang, Yang; Tan, Wenjie
Development an effective vaccine may offer an alternative preventive and therapeutic strategy against HCV infection. DNA vaccination has been shown to induce robust humoral and cellular immunity and overcome many problems associated with conventional vaccines. In this study, mice were primed with either conventional pVRC-based or suicidal pSC-based DNA vaccines carrying DEC-205-targeted NS3 antigen (DEC-NS3) and boosted with type 5 adenoviral vectors encoding the partial NS3 and core antigens (C44P). The prime boost regimen induced a marked increase in antigen-specific humoral and T-cell responses in comparison with either rAd5-based vaccines or DEC-205-targeted DNA immunization in isolation. The protective effect against heterogeneous challenge was correlated with high levels of anti-NS3 IgG and T-cell-mediated immunity against NS3 peptides. Moreover, priming with a suicidal DNA vaccine (pSC-DEC-NS3), which elicited increased TNF-α-producing CD4+ and CD8+ T-cells against NS3-2 peptides (aa 1245-1461), after boosting, showed increased heterogeneous protective potential compared with priming with a conventional DNA vaccine (pVRC-DEC-NS3). In conclusion, a suicidal DNA vector (pSC-DEC-NS3) expressing DEC-205-targeted NS3 combined with boosting using an rAd5-based HCV vaccine (rAd5-C44P) is a good candidate for a safe and effective vaccine against HCV infection.
Feng, Ganzhu; Jiang, Qingtao; Xia, Mei; Lu, Yanlai; Qiu, Wen; Zhao, Dan; Lu, Liwei; Peng, Guangyong; Wang, Yingwei
Development of a novel and effective vaccine against Mycobacterium tuberculosis (M.tb) is a challenging for preventing TB infection. In this study, a novel nanoparticle-based recombinant DNA vaccine was developed, which contains Esat-6 three T cell epitopes (Esat-6/3e) and fms-like tyrosine kinase 3 ligand (FL) genes (termed Esat-6/3e-FL), and was enveloped with chitosan (CS) nanoparticles (nano-chitosan). The immunologic and protective efficacy of the nano-chitosan-based DNA vaccine (termed nano-Esat-6/3e-FL) was assessed in C57BL/6 mice after intramuscular prime vaccination with the plasmids DNA and nasal boost with the Esat-6/3e peptides. The results showed that the immunized mice remarkably elicited enhanced T cell responses and protection against M.tb H37Rv challenge. These findings indicate that the nano-chitosan can significantly elevate the immunologic and protective effects of the DNA vaccine, and the nano-Esat-6/3e-FL is a useful vaccine for preventing M.tb infection in mice. PMID:23637790
Borrego, Belén; Argilaguet, Jordi M; Pérez-Martín, Eva; Dominguez, Javier; Pérez-Filgueira, Mariano; Escribano, José M; Sobrino, Francisco; Rodriguez, Fernando
Development of efficient and safer vaccines against foot-and-mouth disease virus (FMDV) is a must. Previous results obtained in our laboratory have demonstrated that DNA vaccines encoding B and T cell epitopes from type C FMDV, efficiently controlled virus replication in mice, while they did not protect against FMDV challenge in pigs, one of the FMDV natural hosts. The main finding of this work is the ability to improve the protection afforded in swine using a new DNA-vaccine prototype (pCMV-APCH1BTT), encoding FMDV B and T-cell epitopes fused to the single-chain variable fragment of the 1F12 mouse monoclonal antibody that recognizes Class-II Swine Leukocyte antigens. Half of the DNA-immunized pigs were fully protected upon viral challenge, while the remaining animals were partially protected, showing a delayed, shorter and milder disease than control pigs. Full protection in a given vaccinated-pig correlated with the induction of specific IFNγ-secreting T-cells, detectable prior to FMDV-challenge, together with a rapid development of neutralizing antibodies after viral challenge, pointing towards the relevance that both arms of the immune response can play in protection. Our results open new avenues for developing future FMDV subunit vaccines.
Chen, Austen Y; Fry, Scott R; Daggard, Grant E; Mukkur, Trilochan K S
Intramuscular immunization of mice with DNA cocktail vaccines, comprising potential protective antigens P36, P46, NrdF, and P97or P97R1 of Mycoplasma hyopneumoniae, induced strong Th1-polarized immune responses against each antigen, with only P46 eliciting a serum IgG response. Subcutaneous immunization with protein cocktail vaccines, surprisingly, induced both Th1-polarized immune response as well as antibody response whereas mice immunized with DNA cocktail vaccines followed by boosting with protein cocktail vaccines generated strong Th1-polarized and humoral immune responses. P97 was not recognized by serum antibodies from commercial bacterin-immunized mice indicating potential lack of expression of this important antigen in inactivated whole-cell vaccines.
Yang, Wen-Bin; Zhou, Dong-Hui; Zou, Yang; Chen, Kai; Liu, Qing; Wang, Jin-Lei; Zhu, Xing-Quan; Zhao, Guang-Hui
Toxoplasma gondii is an obligatory intracellular protozoan, which infects most of the warm-blooded animals, causing serious public health problems and enormous economic losses worldwide. The rhoptry effector protein 54 (ROP54) has been indicated as a virulence factor that promotes Toxoplasma infection by modulating GBP2 loading onto parasite-containing vacuoles, which can modulate some aspects of the host immune response. In order to evaluate the immuno-protective value of ROP54, we constructed a eukaryotic recombinant plasmid expressing T. gondii ROP54 and intramuscularly immunized Kunming mice with this recombinant plasmid against acute and chronic toxoplasmosis. All mice immunized with pVAX-ROP54 elicited a high level of specific antibody responses, a significant increase of lymphocyte proliferation, and a significant level of Th1-type cytokines (IFN-γ, IL-2 and IL-12p70), in addition to an increased production of Th2-type cytokines (IL-4 and IL-10). These results demonstrated that pVAX-ROP54 induced significant cellular and humoral (Th1/Th2) immune responses, which extended the survival time (13.0±1.15days for pVAX-ROP54 vs 6.7±0.48days for pVAX I, 6.8±0.42days for PBS and 6.5±0.53 for blank control) and significantly reduced cyst burden (35.9% for pVAX-ROP54, 1% for pVAX I and 2% for PBS, compared with blank control) of immunized mice. These results indicate that the recombinant ROP54 plasmid can provide partial protection and might be a potential vaccine candidate against acute and chronic toxoplasmosis. Copyright © 2017 Elsevier B.V. All rights reserved.
Fissolo, Nicolás; Montalban, Xavier; Comabella, Manuel
Multiple sclerosis (MS) is the most common inflammatory, demyelinating, and neurodegenerative disorder of the central nervous system (CNS) in humans. Although the etiology of MS remains unknown, several lines of evidence support the notion that autoimmunity against components of the myelin sheath plays a major role in susceptibility to and development of the disease. At present, there are no approved MS therapies aimed specifically toward downregulating antigen-specific autoreactive immune cells. One antigen-specific approach that appears promising for the treatment of MS is DNA vaccination. This technique has demonstrated efficacy in clinical trials while maintaining safety.Here, we describe the generation of DNA vaccines containing immunologically relevant antigens of MS. Moreover, we present a detailed protocol for the prophylactic and therapeutic administration of DNA vaccines via intramuscular injection targeting on the development of experimental autoimmune encephalomyelitis (EAE), an animal model resembling MS.
Han, Yali; Zhou, Aihua; Lu, Gang; Zhao, Guanghui; Wang, Lin; Guo, Jingjing; Song, Pengxia; Zhou, Jian; Zhou, Huaiyu; Cong, Hua; He, Shenyi
Toxoplasma gondii (T. gondii) is an obligate intracellular protozoan parasite with a broad host range including most warm-blooded animals, including humans. T. gondii surface antigen 1 (SAG1) is a well-characterized T. gondii antigen. T. gondii expresses five nonmitochondrial rhomboid intramembrane proteases, TgROM1-5. TgROM4 is uniformly distributed on the surface of T. gondii and involved in regulating MIC2, MIC3, MIC6, and AMA1 during T. gondii invasion of host cells. Bioinformatics have predicted ROM4 B-cell and T-cell epitopes. Immunization strategy is also a key factor in determining the effectiveness of the immune response and has gained increasing attention in T. gondii vaccine research. In this study, we used a DNA prime-peptide boost vaccination regimen to assess the protective efficacy of various vaccination strategies using TgROM4. We identified a polypeptide (YALLGALIPYCVEYWKSIPR) using a bioinformatics approach, and immunized mice using a DNA-prime and polypeptide-boost regimen. BALB/c mice were randomly divided into six groups, including three experimental groups (peptide, pROM4 and pROM4/peptide) and three control groups (PBS, pEGFP-C1 and pSAG1). Mice were then immunized intramuscularly four times. After immunization, IgG and cytokine productions were determined using enzyme-linked immunosorbent assays. The survival time of mice was evaluated after challenge with tachyzoites of T. gondii RH strain. Additionally, the number of cysts in the brain was determined after intragastric challenge with cysts of T. gondii PRU strain. Mice vaccinated with different immunization regimens (peptide, pROM4 and pROM4/peptide) elicited specific humoral and cellular responses, with high levels of IgG, IgG2a, and interferon (IFN)-γ. Moreover, IgG, IgG2a and IFN-γ levels were highest in the pROM4/peptide group. Immunized mice, especially those in the pROM4/peptide group, had prolonged survival times after challenge with tachyzoites and reduced numbers of brain cysts
Ko, Eun-Ju; Kwon, Young-Man; Lee, Jong Seok; Hwang, Hye Suk; Yoo, Si-Eun; Lee, Yu-Na; Lee, Young-Tae; Kim, Min-Chul; Cho, Min Kyoung; Lee, You Ri; Quan, Fu-Shi; Song, Jae-Min; Lee, Sujin; Moore, Martin L.; Kang, Sang-Moo
Respiratory syncytial virus (RSV) is an important human pathogen. Expression of virus structural proteins produces self-assembled virus-like nanoparticles (VLP). We investigated immune phenotypes after RSV challenge of immunized mice with VLP containing RSV F and G glycoproteins mixed with F-DNA (FdFG VLP). In contrast to formalin-inactivated RSV (FIRSV) causing vaccination-associated eosinophilia, FdFG VLP immunization induced low bronchoalveolar cellularity, higher ratios of CD11c+ versus CD11b+ phenotypic cells and CD8+ T versus CD4+ T cells secreting interferon (IFN)-γ, T helper type-1 immune responses, and no sign of eosinophilia upon RSV challenge. Furthermore, RSV neutralizing activity, lung viral clearance, and histology results suggest that FdFG VLP can be comparable to live RSV in conferring protection against RSV and in preventing RSV disease. This study provides evidence that a combination of recombinant RSV VLP and plasmid DNA may have a potential anti-RSV prophylactic vaccine inducing balanced innate and adaptive immune responses. PMID:25109662
Haddad, Diana; Bilcikova, Erika; Witney, Adam A; Carlton, Jane M; White, Charles E; Blair, Peter L; Chattopadhyay, Rana; Russell, Joshua; Abot, Esteban; Charoenvit, Yupin; Aguiar, Joao C; Carucci, Daniel J; Weiss, Walter R
We describe a novel approach for identifying target antigens for preerythrocytic malaria vaccines. Our strategy is to rapidly test hundreds of DNA vaccines encoding exons from the Plasmodium yoelii yoelii genomic sequence. In this antigen identification method, we measure reduction in parasite burden in the liver after sporozoite challenge in mice. Orthologs of protective P. y. yoelii genes can then be identified in the genomic databases of Plasmodium falciparum and Plasmodium vivax and investigated as candidate antigens for a human vaccine. A pilot study to develop the antigen identification method approach used 192 P. y. yoelii exons from genes expressed during the sporozoite stage of the life cycle. A total of 182 (94%) exons were successfully cloned into a DNA immunization vector with the Gateway cloning technology. To assess immunization strategies, mice were vaccinated with 19 of the new DNA plasmids in addition to the well-characterized protective plasmid encoding P. y. yoelii circumsporozoite protein. Single plasmid immunization by gene gun identified a novel vaccine target antigen which decreased liver parasite burden by 95% and which has orthologs in P. vivax and P. knowlesi but not P. falciparum. Intramuscular injection of DNA plasmids produced a different pattern of protective responses from those seen with gene gun immunization. Intramuscular immunization with plasmid pools could reduce liver parasite burden in mice despite the fact that none of the plasmids was protective when given individually. We conclude that high-throughput cloning of exons into DNA vaccines and their screening is feasible and can rapidly identify new malaria vaccine candidate antigens.
Azevedo, Adriana S.; Gonçalves, Antônio J. S.; Archer, Marcia; Freire, Marcos S.; Galler, Ricardo; Alves, Ada M. B.
The dengue envelope glycoprotein (E) is the major component of virion surface and its ectodomain is composed of domains I, II and III. This protein is the main target for the development of a dengue vaccine with induction of neutralizing antibodies. In the present work, we tested two different vaccination strategies, with combined immunizations in a prime/booster regimen or simultaneous inoculation with a DNA vaccine (pE1D2) and a chimeric yellow fever/dengue 2 virus (YF17D-D2). The pE1D2 DNA vaccine encodes the ectodomain of the envelope DENV2 protein fused to t-PA signal peptide, while the YF17D-D2 was constructed by replacing the prM and E genes from the 17D yellow fever vaccine virus by those from DENV2. Balb/c mice were inoculated with these two vaccines by different prime/booster or simultaneous immunization protocols and most of them induced a synergistic effect on the elicited immune response, mainly in neutralizing antibody production. Furthermore, combined immunization remarkably increased protection against a lethal dose of DENV2, when compared to each vaccine administered alone. Results also revealed that immunization with the DNA vaccine, regardless of the combination with the chimeric virus, induced a robust cell immune response, with production of IFN-γ by CD8+ T lymphocytes. PMID:23472186
Azevedo, Adriana S; Gonçalves, Antônio J S; Archer, Marcia; Freire, Marcos S; Galler, Ricardo; Alves, Ada M B
The dengue envelope glycoprotein (E) is the major component of virion surface and its ectodomain is composed of domains I, II and III. This protein is the main target for the development of a dengue vaccine with induction of neutralizing antibodies. In the present work, we tested two different vaccination strategies, with combined immunizations in a prime/booster regimen or simultaneous inoculation with a DNA vaccine (pE1D2) and a chimeric yellow fever/dengue 2 virus (YF17D-D2). The pE1D2 DNA vaccine encodes the ectodomain of the envelope DENV2 protein fused to t-PA signal peptide, while the YF17D-D2 was constructed by replacing the prM and E genes from the 17D yellow fever vaccine virus by those from DENV2. Balb/c mice were inoculated with these two vaccines by different prime/booster or simultaneous immunization protocols and most of them induced a synergistic effect on the elicited immune response, mainly in neutralizing antibody production. Furthermore, combined immunization remarkably increased protection against a lethal dose of DENV2, when compared to each vaccine administered alone. Results also revealed that immunization with the DNA vaccine, regardless of the combination with the chimeric virus, induced a robust cell immune response, with production of IFN-γ by CD8+ T lymphocytes.
Maity, Hemanta Kumar; Dey, Sohini; Mohan, C Madhan; Khulape, Sagar A; Pathak, Dinesh C; Vakharia, Vikram N
Infectious bursal disease (IBD) is an acute, infectious, immunosuppressive disease affecting young chicken worldwide. The etiological agent IBD virus (IBDV) is a double stranded RNA virus with outer capsid protein VP2 of IBDV is the major antigenic determinant capable of inducing neutralizing antibody. DNA vaccines encoding VP2 has been extensively studied achieving only partial protection. However, the efficacy of DNA vaccines against IBDV can be augmented by choosing a potential molecular adjuvant. The goal of the present study is to evaluate the immune response and protective efficacy of a DNA vaccine encoding the C-terminal domain of the heat shock protein 70 (cHSP70) of Mycobacterium tuberculosis gene genetically fused with the full length VP2 gene of IBDV (pCIVP2-cHSP70) in comparison to a 'DNA prime-protein boost' approach and a DNA vaccine encoding the VP2 gene (pCIVP2) alone. The results indicate that both pCIVP2-cHSP70 and 'DNA prime-protein boost' elicited humoral as well as cellular immune responses. Chickens in the pCIVP2-cHSP70 and 'DNA prime-protein boost' groups developed significantly higher levels of ELISA titer to IBDV antigen compared to the group immunized with pCIVP2 alone (p<0.01). However, significantly higher levels of lymphocyte proliferative response, IL-12 and IFN-γ production were found in the pCIVP2-cHSP70 group compared to 'DNA prime-protein boost' group. Additionally, chickens immunized with pCIVP2-cHSP70 and 'DNA prime-protein boost' vaccines were completely protected against the vvIBDV whereas pCIVP2 DNA vaccine alone was able to protect only 70%. These findings suggest that the truncated C-terminal HSP70 mediated DNA vaccine genetically fused with the VP2 gene construct stimulated both humoral and cell mediated immune responses and conferred complete protection against IBDV. This novel strategy is perhaps a seminal concept in utilizing HSP70 as an adjuvant molecule to elicit an immune response against IBD affecting chickens.
Song, Pengxia; He, Shenyi; Zhou, Aihua; Lv, Gang; Guo, Jingjing; Zhou, Jian; Han, Yali; Zhou, Huaiyu; Hao, Zhen; Cong, Hua
A widely prevalent disease, toxoplasmosis poses serious health threats to both humans and animals; therefore, development of an ideal DNA vaccine against Toxoplasma gondii is needed eagerly. The purpose of the present study is to assess the protective efficacy of a DNA vaccine encoding the T. gondii toxofilin gene (pEGFP-toxofilin). In addition, toxofilin DNA vaccine combined with the individual adjuvants, alum or monophosphoryl lipid A (MPLA), or a mixture of alum-MPLA adjuvant were screened for their ability to enhance antibody responses. Using bioinformatics, we analyzed the gene and amino acid sequences of the toxofilin protein, recognizing and identifying several potential linear B and T helper (Th)-1 cell epitopes. BALB/c mice were immunized three times with either toxofilin DNA vaccine alone or in combination with the adjuvants such as alum, MPLA or an alum-MPLA mixture. The systemic immune response was evaluated by cytokine, the percentage of CD4 (+) and CD8 (+) T cells and specific antibody measurement. Two weeks after the last immunization, protective efficacy was evaluated by challenging intraperitoneally with 1 × 10(4) tachyzoites of T. gondii or intragastrically with 20 cysts of T. gondii PRU strain. All experimentally immunized mice developed strong humoral and cellular immune responses compared with the control groups. Moreover, by comparison with the non-adjuvant toxofilin DNA vaccine group, adding alum adjuvant to toxofilin DNA vaccine resulted in an increase in humoral response and a skewed Th2 response. However, the MPLA adjuvant with toxofilin DNA vaccine induced significantly enhanced humoral and Th1-biased immune responses. Importantly, the co-administration of alum-MPLA adjuvant in combination with the toxofilin DNA vaccine shifted the Th2 immune response to a Th1 response compared with the alum-toxofilin group, and elicited the strongest humoral and Th1 responses among all the groups. At the same time, a longer survival time and less
Watts, A M; Kennedy, R C
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.
T-cell-mediated autoimmune diseases such as multiple sclerosis, rheumatoid arthritis or type 1 diabetes result from an aggressive attack of self-components by autoimmune T-cells. Pro-inflammatory mediators, particularly cytokines and chemokines, direct the homing and effectorfunction of these cells. It has recently been demonstrated that the immune system, which can attack self-components, also generates 'beneficial' autoimmunity against pro-inflammatory mediators. During the course of an autoimmune condition, and to a much lesser extent in response to microbial inflammation, the immune system produces auto-antibodies to pro-inflammatory mediators. This reduces the harm from these diseases. We also discovered that targeted DNA vaccines could effectively amplify these responses to provide protective immunity. The underlying mechanism is partially understood. At the site of immunization, the relevant gene product is produced and then presented by dendritic cells/macrophages, which undergo activation due to an interaction of plasmid CpG with toll-like receptor 9 on the dendritic cell. This then activates CD4+ T-cells, which help the production of T-cell-dependent antibodies against the gene product of the vaccines. These antibodies neutralize their target product and suppress inflammation. This review explores this interesting concept and its therapeutic implications.
Luo, Kun; Zhang, Hong; Zavala, Fidel; Biragyn, Arya; Espinosa, Diego A; Markham, Richard B
Although sterilizing immunity to malaria can be elicited by irradiated sporozoite vaccination, no clinically practical subunit vaccine has been shown to be capable of preventing the approximately 600,000 annual deaths attributed to this infection. DNA vaccines offer several potential advantages for a disease that primarily affects the developing world, but new approaches are needed to improve the immunogenicity of these vaccines. By using a novel, lipid-based adjuvant, Vaxfectin, to attract immune cells to the immunization site, in combination with an antigen-chemokine DNA construct designed to target antigen to immature dendritic cells, we elicited a humoral immune response that provided sterilizing immunity to malaria challenge in a mouse model system. The chemokine, MIP3αCCL20, did not significantly enhance the cellular infiltrate or levels of cytokine or chemokine expression at the immunization site but acted with Vaxfectin to reduce liver stage malaria infection by orders of magnitude compared to vaccine constructs lacking the chemokine component. The levels of protection achieved were equivalent to those observed with irradiated sporozoites, a candidate vaccine undergoing development for further large scale clinical trial. Only vaccination with the combined regimen of adjuvant and chemokine provided 80-100% protection against the development of bloodstream infection. Treating the immunization process as requiring the independent steps of 1) attracting antigen-presenting cells to the site of immunization and 2) specifically directing vaccine antigen to the immature dendritic cells that initiate the adaptive immune response may provide a rational strategy for the development of a clinically applicable malaria DNA vaccine.
Luo, Kun; Zhang, Hong; Zavala, Fidel; Biragyn, Arya; Espinosa, Diego A.; Markham, Richard B.
Although sterilizing immunity to malaria can be elicited by irradiated sporozoite vaccination, no clinically practical subunit vaccine has been shown to be capable of preventing the approximately 600,000 annual deaths attributed to this infection. DNA vaccines offer several potential advantages for a disease that primarily affects the developing world, but new approaches are needed to improve the immunogenicity of these vaccines. By using a novel, lipid-based adjuvant, Vaxfectin, to attract immune cells to the immunization site, in combination with an antigen-chemokine DNA construct designed to target antigen to immature dendritic cells, we elicited a humoral immune response that provided sterilizing immunity to malaria challenge in a mouse model system. The chemokine, MIP3αCCL20, did not significantly enhance the cellular infiltrate or levels of cytokine or chemokine expression at the immunization site but acted with Vaxfectin to reduce liver stage malaria infection by orders of magnitude compared to vaccine constructs lacking the chemokine component. The levels of protection achieved were equivalent to those observed with irradiated sporozoites, a candidate vaccine undergoing development for further large scale clinical trial. Only vaccination with the combined regimen of adjuvant and chemokine provided 80–100% protection against the development of bloodstream infection. Treating the immunization process as requiring the independent steps of 1) attracting antigen-presenting cells to the site of immunization and 2) specifically directing vaccine antigen to the immature dendritic cells that initiate the adaptive immune response may provide a rational strategy for the development of a clinically applicable malaria DNA vaccine. PMID:24599116
Golshani, Maryam; Rafati, Sima; Siadat, Seyed Davar; Nejati-Moheimani, Mehdi; Shahcheraghi, Fereshteh; Arsang, Amin; Bouzari, Saeid
Brucellosis is one of the most common zoonotic diseases caused by species of Brucella. At present, there is no commercially available vaccine for the human brucellosis. Brucella melitensis and Brucella abortus are the main causes of human brucellosis, worldwide. The outer membrane protein 31 (Omp31) and L7/L12 are immunodominant and protective antigens conserved among human Brucella pathogens. The purpose of the current study was to evaluate and compare the immunogenicity and protective efficacy of the L7/L12-TOmp31 construct administered as DNA/DNA and DNA/Pro vaccine regimens. Vaccination of BALB/c mice with the DNA/Pro regimen provided more protection levels against B. melitenisis and B. abortus challenge than did the DNA/DNA regimen. IgG1 and IgG2a titers were higher in the sera from DNA/Pro-immunized mice than in those from mice immunized with DNA alone. Moreover, splenocytes from DNA/Pro-immunized mice produced significantly higher levels of IFN-γ than did those from mice given DNA alone. The pcDNA-L7/L12-TOmp31 priming followed by rL7/L12-TOmp31 boosting led to improved protection against B. abortus or B. melitensis infection.
Khan, Kishwar Hayat
Vaccination is the most successful application of immunological principles to human health. Vaccine efficacy needs to be reviewed from time to time and its safety is an overriding consideration. DNA vaccines offer simple yet effective means of inducing broad-based immunity. These vaccines work by allowing the expression of the microbial antigen inside host cells that take up the plasmid. These vaccines function by generating the desired antigen inside the cells, with the advantage that this may facilitate presentation through the major histocompatibility complex. This review article is based on a literature survey and it describes the working and designing strategies of DNA vaccines. Advantages and disadvantages for this type of vaccines have also been explained, together with applications of DNA vaccines. DNA vaccines against cancer, tuberculosis, Edwardsiella tarda, HIV, anthrax, influenza, malaria, dengue, typhoid and other diseases were explored. PMID:24432284
Redding, Laurel; Weiner, David B
DNA vaccines represent a new frontier in vaccine technology. One important application of this technology is in the veterinary arena. DNA vaccines have already gained a foothold in certain fields of veterinary medicine. However, several important questions must be addressed when developing DNA vaccines for animals, including whether or not the vaccine is efficacious and cost effective compared with currently available options. Another important question to consider is how to apply this developing technology in a wide range of different situations, from the domestic pet to individual fish in fisheries with several thousand animals, to wildlife programs for disease control. In some cases, DNA vaccines represent an interesting option for vaccination, while in others, currently available options are sufficient. This review will examine a number of diseases of veterinary importance and the progress being made in DNA vaccine technology relevant to these diseases, and we compare these with the conventional treatment options available.
Redding, Laurel; Werner, David B
DNA vaccines represent a new frontier in vaccine technology. One important application of this technology is in the veterinary arena. DNA vaccines have already gained a foothold in certain fields of veterinary medicine. However, several important questions must be addressed when developing DNA vaccines for animals, including whether or not the vaccine is efficacious and cost effective compared with currently available options. Another important question to consider is how to apply this developing technology in a wide range of different situations, from the domestic pet to individual fish in fisheries with several thousand animals, to wildlife programs for disease control. In some cases, DNA vaccines represent an interesting option for vaccination, while in others, currently available options are sufficient. This review will examine a number of diseases of veterinary importance and the progress being made in DNA vaccine technology relevant to these diseases, and we compare these with the conventional treatment options available. PMID:19722897
Larocca, Rafael A; Abbink, Peter; Peron, Jean Pierre S; Zanotto, Paolo M de A; Iampietro, M Justin; Badamchi-Zadeh, Alexander; Boyd, Michael; Ng'ang'a, David; Kirilova, Marinela; Nityanandam, Ramya; Mercado, Noe B; Li, Zhenfeng; Moseley, Edward T; Bricault, Christine A; Borducchi, Erica N; Giglio, Patricia B; Jetton, David; Neubauer, George; Nkolola, Joseph P; Maxfield, Lori F; De La Barrera, Rafael A; Jarman, Richard G; Eckels, Kenneth H; Michael, Nelson L; Thomas, Stephen J; Barouch, Dan H
Zika virus (ZIKV) is a flavivirus that is responsible for the current epidemic in Brazil and the Americas. ZIKV has been causally associated with fetal microcephaly, intrauterine growth restriction, and other birth defects in both humans and mice. The rapid development of a safe and effective ZIKV vaccine is a global health priority, but very little is currently known about ZIKV immunology and mechanisms of immune protection. Here we show that a single immunization with a plasmid DNA vaccine or a purified inactivated virus vaccine provides complete protection in susceptible mice against challenge with a strain of ZIKV involved in the outbreak in northeast Brazil. This ZIKV strain has recently been shown to cross the placenta and to induce fetal microcephaly and other congenital malformations in mice. We produced DNA vaccines expressing ZIKV pre-membrane and envelope (prM-Env), as well as a series of deletion mutants. The prM-Env DNA vaccine, but not the deletion mutants, afforded complete protection against ZIKV, as measured by absence of detectable viraemia following challenge, and protective efficacy correlated with Env-specific antibody titers. Adoptive transfer of purified IgG from vaccinated mice conferred passive protection, and depletion of CD4 and CD8 T lymphocytes in vaccinated mice did not abrogate this protection. These data demonstrate that protection against ZIKV challenge can be achieved by single-shot subunit and inactivated virus vaccines in mice and that Env-specific antibody titers represent key immunologic correlates of protection. Our findings suggest that the development of a ZIKV vaccine for humans is likely to be achievable.
Larocca, Rafael A.; Abbink, Peter; Peron, Jean Pierre S.; de A. Zanotto, Paolo M.; Iampietro, M. Justin; Badamchi-Zadeh, Alexander; Boyd, Michael; Ng’ang’a, David; Kirilova, Marinela; Nityanandam, Ramya; Mercado, Noe B.; Li, Zhenfeng; Moseley, Edward T.; Bricault, Christine A.; Borducchi, Erica N.; Giglio, Patricia B.; Jetton, David; Neubauer, George; Nkolola, Joseph P.; Maxfield, Lori F.; De La Barrera, Rafael A.; Jarman, Richard G.; Eckels, Kenneth H.; Michael, Nelson L.; Thomas, Stephen J.; Barouch, Dan H.
Zika virus (ZIKV) is a flavivirus that is responsible for an unprecedented current epidemic in Brazil and the Americas1,2. ZIKV has been causally associated with fetal microcephaly, intrauterine growth restriction, and other birth defects in both humans3–8 and mice9–11. The rapid development of a safe and effective ZIKV vaccine is a global health priority1,2, but very little is currently known about ZIKV immunology and mechanisms of immune protection. Here we show that a single immunization of a plasmid DNA vaccine or a purified inactivated virus vaccine provides complete protection in susceptible mice against challenge with a ZIKV outbreak strain from northeast Brazil. This ZIKV strain has recently been shown to cross the placenta and to induce fetal microcephaly and other congenital malformations in mice11. We produced DNA vaccines expressing full-length ZIKV pre-membrane and envelope (prM-Env) as well as a series of deletion mutants. The full-length prM-Env DNA vaccine, but not the deletion mutants, afforded complete protection against ZIKV as measured by absence of detectable viremia following challenge, and protective efficacy correlated with Env-specific antibody titers. Adoptive transfer of purified IgG from vaccinated mice conferred passive protection, and CD4 and CD8 T lymphocyte depletion in vaccinated mice did not abrogate protective efficacy. These data demonstrate that protection against ZIKV challenge can be achieved by single-shot subunit and inactivated virus vaccines in mice and that Env-specific antibody titers represent key immunologic correlates of protection. Our findings suggest that the development of a ZIKV vaccine for humans will likely be readily achievable. PMID:27355570
An important new approach to vaccination is plasmid DNA injection in vivo that can elicit an immune response against protein(s) encoded. Antigen that is expressed from the in vivo transfected cells induces both humoral and cellular immune response. DNA immunization is generally applicable for a wide range of proteins. It can provide an organism with immunity against viruses, bacteria, parasites, and tumors. DNA vaccines can overcome the disadvantages of vaccines presently used as well as provide various new vaccines that are currently not available. This minireview provides an overview of evaluated DNA vaccine candidates against infectious agents and certain cancers.
Iezzi, Manuela; Quaglino, Elena; Amici, Augusto; Lollini, Pier-Luigi; Forni, Guido; Cavallo, Federica
The emerging evidence that DNA vaccines elicit a protective immune response in rodents, dogs and cancer patients, coupled with the US Food and Drug Administration (FDA) approval of an initial DNA vaccine to treat canine tumors is beginning to close the gap between the optimistic experimental data and their difficult application in a clinical setting. Here we review a series of conceptual and biotechnological advances that are working together to make DNA vaccines targeting molecules that play important roles during cancer progression (oncoantigens) a promise with near-term clinical impact.
Farris, Eric; Brown, Deborah M; Ramer-Tait, Amanda E; Pannier, Angela K
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. © 2016 by the Society for Experimental Biology and Medicine.
Farris, Eric; Brown, Deborah M; Ramer-Tait, Amanda E
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. PMID:27048557
Al-Quraishy, Saleh; Dkhil, Mohamed A; Abdel-Baki, Abdel-Azeem S; Ghanjati, Foued; Erichsen, Lars; Santourlidis, Simeon; Wunderlich, Frank; Araúzo-Bravo, Marcos J
Epigenetic mechanisms such as DNA methylation are increasingly recognized to be critical for vaccination efficacy and outcome of different infectious diseases, but corresponding information is scarcely available for host defense against malaria. In the experimental blood-stage malaria Plasmodium chabaudi, we investigate the possible effects of a blood-stage vaccine on DNA methylation of gene promoters in the liver, known as effector against blood-stage malaria, using DNA methylation microarrays. Naturally susceptible Balb/c mice acquire, by protective vaccination, the potency to survive P. chabaudi malaria and, concomitantly, modifications of constitutive DNA methylation of promoters of numerous genes in the liver; specifically, promoters of 256 genes are hyper(=up)- and 345 genes are hypo(=down)-methylated (p < 0.05). Protective vaccination also leads to changes in promoter DNA methylation upon challenge with P. chabaudi at peak parasitemia on day 8 post infection (p.i.), when 571 and 1013 gene promoters are up- and down-methylated, respectively, in relation to constitutive DNA methylation (p < 0.05). Gene set enrichment analyses reveal that both vaccination and P. chabaudi infections mainly modify promoters of those genes which are most statistically enriched with functions relating to regulation of transcription. Genes with down-methylated promoters encompass those encoding CX3CL1, GP130, and GATA2, known to be involved in monocyte recruitment, IL-6 trans-signaling, and onset of erythropoiesis, respectively. Our data suggest that vaccination may epigenetically improve parts of several effector functions of the liver against blood-stage malaria, as, e.g., recruitment of monocyte/macrophage to the liver accelerated liver regeneration and extramedullary hepatic erythropoiesis, thus leading to self-healing of otherwise lethal P. chabaudi blood-stage malaria.
Rosada, Rogério S; Torre, Lucimara Gaziola de la; Frantz, Fabiani G; Trombone, Ana PF; Zárate-Bladés, Carlos R; Fonseca, Denise M; Souza, Patrícia RM; Brandão, Izaíra T; Masson, Ana P; Soares, Édson G; Ramos, Simone G; Faccioli, Lúcia H; Silva, Célio L; Santana, Maria HA; Coelho-Castelo, Arlete AM
Background The greatest challenges in vaccine development include optimization of DNA vaccines for use in humans, creation of effective single-dose vaccines, development of delivery systems that do not involve live viruses, and the identification of effective new adjuvants. Herein, we describe a novel, simple technique for efficiently vaccinating mice against tuberculosis (TB). Our technique consists of a single-dose, genetic vaccine formulation of DNA-hsp65 complexed with cationic liposomes and administered intranasally. Results We developed a novel and non-toxic formulation of cationic liposomes, in which the DNA-hsp65 vaccine was entrapped (ENTR-hsp65) or complexed (COMP-hsp65), and used to immunize mice by intramuscular or intranasal routes. Although both liposome formulations induced a typical Th1 pattern of immune response, the intramuscular route of delivery did not reduce the number of bacilli. However, a single intranasal immunization with COMP-hsp65, carrying as few as 25 μg of plasmid DNA, leads to a remarkable reduction of the amount of bacilli in lungs. These effects were accompanied by increasing levels of IFN-γ and lung parenchyma preservation, results similar to those found in mice vaccinated intramuscularly four times with naked DNA-hsp65 (total of 400 μg). Conclusion Our objective was to overcome the significant obstacles currently facing DNA vaccine development. Our results in the mouse TB model showed that a single intranasal dose of COMP-hsp65 elicited a cellular immune response that was as strong as that induced by four intramuscular doses of naked-DNA. This formulation allowed a 16-fold reduction in the amount of DNA administered. Moreover, we demonstrated that this vaccine is safe, biocompatible, stable, and easily manufactured at a low cost. We believe that this strategy can be applied to human vaccines to TB in a single dose or in prime-boost protocols, leading to a tremendous impact on the control of this infectious disease. PMID
Bolesta, Elizabeth; Kowalczyk, Aleksandra; Wierzbicki, Andrzej; Rotkiewicz, Piotr; Bambach, Barbara; Tsao, Chun-Yen; Horwacik, Irena; Kolinski, Andrzej; Rokita, Hanna; Brecher, Martin; Wang, Xinhui; Ferrone, Soldano; Kozbor, Danuta
The GD2 ganglioside expressed on neuroectodermally derived tumors, including neuroblastoma and melanoma, is weakly immunogenic in tumor-bearing patients and induces predominantly immunoglobulin (Ig)-M antibody responses in the immunized host. Here, we investigated whether interconversion of GD2 into a peptide mimetic form would induce GD2 cross-reactive IgG antibody responses in mice. Screening of the X(15) phage display peptide library with the anti-GD2 monoclonal antibody (mAb) 14G2a led to isolation of mimetic peptide 47, which inhibited the binding of 14G2a antibody to GD2-positive tumor cells. The peptide was also recognized by GD2-specific serum antibodies from a patient with neuroblastoma, suggesting that it bears an internal image of GD2 ganglioside expressed on the tumor cells. The molecular basis for antigenicity of the GD2 mimetic peptide, established by molecular modeling and mutagenesis studies, led to the generation of a 47-LDA mutant with an increased mimicry to GD2. Immunization of mice with peptide 47-LDA-encoded plasmid DNA elicited GD2 cross-reactive IgG antibody responses, which were increased on subsequent boost with GD2 ganglioside. The vaccine-induced antibodies recognized GD2-positive tumor cells, mediated complement-dependent cytotoxicity, and exhibited protection against s.c. human GD2-positive melanoma growth in the severe combined immunodeficient mouse xenograft model. The results from our studies provide insights into approaches for boosting GD2 cross-reactive IgG antibody responses by minigene vaccination with a protective epitope of GD2 ganglioside.
Lee, Jong Seok; Kwon, Young-Man; Hwang, Hye Suk; Lee, Yu-Na; Ko, Eun-Ju; Yoo, Si-Eun; Kim, Min-Chul; Kim, Ki-Hye; Cho, Min-Kyoung; Lee, Young-Tae; Lee, You Ri; Quan, Fu-Shi; Kang, Sang-Moo
Respiratory syncytial virus (RSV) is a major viral agent causing significant morbidity and mortality in young infants and the elderly. There is no licensed vaccine against RSV and it is a high priority to develop a safe RSV vaccine. We determined the immunogenicity and protective efficacy of combined virus-like particle and DNA vaccines presenting RSV glycoproteins (Fd.VLP) in comparison with formalin inactivated RSV (FI-RSV). Immunization of mice with Fd.VLP induced higher ratios of IgG2a/IgG1 antibody responses compared to those with FI-RSV. Upon live RSV challenge, Fd.VLP and FI-RSV vaccines were similarly effective in clearing lung viral loads. However, FI-RSV immunized mice showed a substantial weight loss and high levels of T helper type 2 (Th2) cytokines as well as extensive lung histopathology and eosinophil infiltration. In contrast, Fd.VLP immunized mice did not exhibit Th2 type cytokines locally and systemically, which might contribute to preventing vaccine-associated RSV lung disease. These results indicate that virus-like particles in combination with DNA vaccines represent a potential approach for developing a safe and effective RSV vaccine. PMID:25173478
Lee, Jong Seok; Kwon, Young-Man; Hwang, Hye Suk; Lee, Yu-Na; Ko, Eun-Ju; Yoo, Si-Eun; Kim, Min-Chul; Kim, Ki-Hye; Cho, Min Kyoung; Lee, Young-Tae; Lee, You Ri; Quan, Fu-Shi; Kang, Sang-Moo
Respiratory syncytial virus (RSV) is a major viral agent causing significant morbidity and mortality in young infants and the elderly. There is no licensed vaccine against RSV and it is a high priority to develop a safe RSV vaccine. We determined the immunogenicity and protective efficacy of combined virus-like particle and DNA vaccines presenting RSV glycoproteins (Fd.VLP) in comparison with formalin inactivated RSV (FI-RSV). Immunization of mice with Fd.VLP induced higher ratios of IgG2a/IgG1 antibody responses compared to those with FI-RSV. Upon live RSV challenge, Fd.VLP and FI-RSV vaccines were similarly effective in clearing lung viral loads. However, FI-RSV immunized mice showed a substantial weight loss and high levels of T helper type 2 (Th2) cytokines as well as extensive lung histopathology and eosinophil infiltration. In contrast, Fd.VLP immunized mice did not exhibit Th2 type cytokines locally and systemically, which might contribute to preventing vaccine-associated RSV lung disease. These results indicate that virus-like particles in combination with DNA vaccines represent a potential approach for developing a safe and effective RSV vaccine.
Sullivan, Nicole L.; Blazevic, Azra; Bruna-Romero, Oscar; Rodrigues, Mauricio M.; Hoft, Daniel F.
Background Immunization of mice with the Trypanosoma cruzi trans-sialidase (TS) gene using plasmid DNA, adenoviral vector, and CpG-adjuvanted protein delivery has proven highly immunogenic and provides protection against acute lethal challenge. However, long-term protection induced by TS DNA vaccines has not been reported. The goal of the present work was to test whether the co-administration of a plasmid encoding IL-15 (pIL-15) could improve the duration of protection achieved through genetic vaccination with plasmid encoding TS (pTS) alone. Methodology We immunized BALB/c mice with pTS in the presence or absence of pIL-15 and studied immune responses [with TS-specific IFN-γ ELISPOT, serum IgG ELISAs, intracellular cytokine staining (IFN-γ, TNF-α, and IL-2), tetramer staining, and CFSE dilution assays] and protection against lethal systemic challenge at 1 to 6 months post vaccination. Mice receiving pTS alone developed robust TS-specific IFN-γ responses and survived a lethal challenge given within the first 3 months following immunization. The addition of pIL-15 to pTS vaccination did not significantly alter T cell responses or protection during this early post-vaccination period. However, mice vaccinated with both pTS and pIL-15 challenged 6 months post-vaccination were significantly more protected against lethal T. cruzi challenges than mice vaccinated with pTS alone (P<0.05). Improved protection correlated with significantly higher numbers of TS-specific IFN-γ producing total and CD8+ T cells detected>6 months post immunization. Also, these TS-specific T cells were better able to expand after in vitro re-stimulation. Conclusion Addition of pIL-15 during genetic vaccination greatly improved long-term T cell survival, memory T cell expansion, and long-term protection against the important human parasite, T. cruzi. PMID:21408124
Kazanji, Mirdad; Tartaglia, James; Franchini, Genoveffa; de Thoisy, Benoit; Talarmin, Antoine; Contamin, Hugues; Gessain, Antoine; de Thé, Guy
We assessed the immunogenicities and efficacies of two highly attenuated vaccinia virus-derived NYVAC vaccine candidates encoding the human T-cell leukemia/lymphoma virus type 1 (HTLV-1) env gene or both the env and gag genes in prime-boost pilot regimens in combination with naked DNA expressing the HTLV-1 envelope. Three inoculations of NYVAC HTLV-1 env at 0, 1, and 3 months followed by a single inoculation of DNA env at 9 months protected against intravenous challenge with HTLV-1-infected cells in one of three immunized squirrel monkeys. Furthermore, humoral and cell-mediated immune responses against HTLV-1 Env could be detected in this protected animal. However, priming the animal with a single dose of env DNA, followed by immunization with the NYVAC HTLV-1 gag and env vaccine at 6, 7, and 8 months, protected all three animals against challenge with HTLV-1-infected cells. With this protocol, antibodies against HTLV-1 Env and cell-mediated responses against Env and Gag could also be detected in the protected animals. Although the relative superiority of a DNA prime-NYVAC boost regimen over addition of the Gag component as an immunogen cannot be assessed directly, our findings nevertheless show that an HTLV-1 vaccine approach is feasible and deserves further study. PMID:11390595
Gong, Pengtao; Cao, Lili; Guo, Yanbing; Dong, Hang; Yuan, Shuxian; Yao, Xinhua; Ren, Wenzhi; Yao, Lin; Xu, Zhilin; Sun, Qiang; Zhang, Xichen
The intracellular parasite Toxoplasma gondii is a major cause of abortion and neonatal loss in livestock, and can cause severe illness to human with weakened immune system. The heavy incidence and severe consequence indicate the development of vaccines against T. gondii is required. In this study, DNA vaccines encoding GRA1 and MIC3 antigens were developed. The parasite-specific immune responses and protection efficiency against toxoplasmosis by these DNA vaccines were evaluated in BALB/c mice. The results demonstrated that the IgG antibody production was significantly increased in multi-antigenic vaccine encoding GRA1 and MIC3 immunized group, as well as the IFN-γ level, when compared with single-gene vaccines and controls groups (p < 0.05). Two weeks after the final vaccination, the mice were challenged with either 1 × 10(4) or 1 × 10(2) RH strain tachyzoites, and the mortality and parasite reduction were observed. The multi-antigenic vaccine encoding GRA1 and MIC3 lead to the longest survival time as well as the less parasite-loads in brain and liver of immunized mice (p < 0.01). The present study indicates that the GRA1 and MIC3 showed the potential as target for vaccine investigation against toxoplasmosis. And the immune efficacy induced by multi-antigenic vaccine encoding GRA1 and MIC3 was better than that induced by single-antigenic vaccines alone.
Bounds, Callie E.; Kwilas, Steven A.; Kuehne, Ana I.; Brannan, Jennifer M.; Bakken, Russell R.; Dye, John M.; Hooper, Jay W.; Dupuy, Lesley C.; Ellefsen, Barry; Hannaman, Drew; Wu, Hua; Jiao, Jin-an; Sullivan, Eddie J.; Schmaljohn, Connie S.
DNA vaccination of transchromosomal bovines (TcBs) with DNA vaccines expressing the codon-optimized (co) glycoprotein (GP) genes of Ebola virus (EBOV) and Sudan virus (SUDV) produce fully human polyclonal antibodies (pAbs) that recognize both viruses and demonstrate robust neutralizing activity. Each TcB was vaccinated by intramuscular electroporation (IM-EP) a total of four times and at each administration received 10 mg of the EBOV-GPco DNA vaccine and 10 mg of the SUDV-GPco DNA vaccine at two sites on the left and right sides, respectively. After two vaccinations, robust antibody responses (titers > 1000) were detected by ELISA against whole irradiated EBOV or SUDV and recombinant EBOV-GP or SUDV-GP (rGP) antigens, with higher titers observed for the rGP antigens. Strong, virus neutralizing antibody responses (titers >1000) were detected after three vaccinations when measured by vesicular stomatitis virus-based pseudovirion neutralization assay (PsVNA). Maximal neutralizing antibody responses were identified by traditional plaque reduction neutralization tests (PRNT) after four vaccinations. Neutralizing activity of human immunoglobulins (IgG) purified from TcB plasma collected after three vaccinations and injected intraperitoneally (IP) into mice at a 100 mg/kg dose was detected in the serum by PsVNA up to 14 days after administration. Passive transfer by IP injection of the purified IgG (100 mg/kg) to groups of BALB/c mice one day after IP challenge with mouse adapted (ma) EBOV resulted in 80% protection while all mice treated with non-specific pAbs succumbed. Similarly, interferon receptor 1 knockout (IFNAR -/-) mice receiving the purified IgG (100 mg/kg) by IP injection one day after IP challenge with wild type SUDV resulted in 89% survival. These results are the first to demonstrate that filovirus GP DNA vaccines administered to TcBs by IM-EP can elicit neutralizing antibodies that provide post-exposure protection. Additionally, these data describe
Bounds, Callie E; Kwilas, Steven A; Kuehne, Ana I; Brannan, Jennifer M; Bakken, Russell R; Dye, John M; Hooper, Jay W; Dupuy, Lesley C; Ellefsen, Barry; Hannaman, Drew; Wu, Hua; Jiao, Jin-an; Sullivan, Eddie J; Schmaljohn, Connie S
DNA vaccination of transchromosomal bovines (TcBs) with DNA vaccines expressing the codon-optimized (co) glycoprotein (GP) genes of Ebola virus (EBOV) and Sudan virus (SUDV) produce fully human polyclonal antibodies (pAbs) that recognize both viruses and demonstrate robust neutralizing activity. Each TcB was vaccinated by intramuscular electroporation (IM-EP) a total of four times and at each administration received 10 mg of the EBOV-GPco DNA vaccine and 10 mg of the SUDV-GPco DNA vaccine at two sites on the left and right sides, respectively. After two vaccinations, robust antibody responses (titers > 1000) were detected by ELISA against whole irradiated EBOV or SUDV and recombinant EBOV-GP or SUDV-GP (rGP) antigens, with higher titers observed for the rGP antigens. Strong, virus neutralizing antibody responses (titers >1000) were detected after three vaccinations when measured by vesicular stomatitis virus-based pseudovirion neutralization assay (PsVNA). Maximal neutralizing antibody responses were identified by traditional plaque reduction neutralization tests (PRNT) after four vaccinations. Neutralizing activity of human immunoglobulins (IgG) purified from TcB plasma collected after three vaccinations and injected intraperitoneally (IP) into mice at a 100 mg/kg dose was detected in the serum by PsVNA up to 14 days after administration. Passive transfer by IP injection of the purified IgG (100 mg/kg) to groups of BALB/c mice one day after IP challenge with mouse adapted (ma) EBOV resulted in 80% protection while all mice treated with non-specific pAbs succumbed. Similarly, interferon receptor 1 knockout (IFNAR(-/-)) mice receiving the purified IgG (100 mg/kg) by IP injection one day after IP challenge with wild type SUDV resulted in 89% survival. These results are the first to demonstrate that filovirus GP DNA vaccines administered to TcBs by IM-EP can elicit neutralizing antibodies that provide post-exposure protection. Additionally, these data describe
Nguyen-Hoai, Tam; Kobelt, Dennis; Hohn, Oliver; Vu, Minh D; Schlag, Peter M; Dörken, Bernd; Norley, Steven; Lipp, Martin; Walther, Wolfgang; Pezzutto, Antonio; Westermann, Jörg
DNA vaccines are potential tools for the induction of immune responses against both infectious disease and cancer. The dermal application of DNA vaccines is of particular interest since the epidermal and dermal layers of the skin are characterized by an abundance of antigen-presenting cells (APCs). The aim of our study was to compare tumor protection as obtained by two different methods of intradermal DNA delivery (gene gun and jet injector) in a well-established HER2/neu mouse tumor model. BALB/c mice were immunized twice with a HER2/neu-coding plasmid by gene gun or jet injector. Mice were then subcutaneously challenged with HER2/neu(+) syngeneic D2F2/E2 tumor cells. Protection against subsequent challenges with tumor cells as well as humoral and T-cell immune responses induced by the vaccine were monitored. Gene gun immunization was far superior to jet injector both in terms of tumor protection and induction of HER2/neu-specific immune responses. After gene gun immunization, 60% of the mice remained tumor-free until day 140 as compared with 25% after jet injector immunization. Furthermore, gene gun vaccination was able to induce both a strong T(H)1-polarized T-cell response with detectable cytotoxic T-lymphocyte (CTL) activity and a humoral immune response against HER2/neu, whereas the jet injector was not. Although the disadvantages that were associated with the use of the jet injector in our model may be overcome with methodological modifications and/or in larger animals, which exhibit a thicker skin and/or subcutaneous muscle tissue, we conclude that gene gun delivery constitutes the method of choice for intradermal DNA delivery in preclinical mouse models and possibly also for the clinical development of DNA-based vaccines.
Yu, Yun-Zhou; Ma, Yao; Xu, Wen-Hui; Wang, Shuang; Sun, Zhi-Wei
DNA vaccines are generally weak stimulators of the immune system. Fortunately, their efficacy can be improved using a viral replicon vector or by the addition of immunostimulatory CpG motifs, although the design of these engineered DNA vectors requires optimization. Our results clearly suggest that multiple copies of three types of CpG motifs or combinations of various types of CpG motifs cloned into a viral replicon vector backbone with strong immunostimulatory activities on human PBMC are efficient adjuvants for these DNA vaccines to modulate and enhance protective immunity against anthrax, although modifications with these different CpG forms in vivo elicited inconsistent immune response profiles. Modification with more copies of CpG motifs elicited more potent adjuvant effects leading to the generation of enhanced immunity, which indicated a CpG motif dose-dependent enhancement of antigen-specific immune responses. Notably, the enhanced and/or synchronous adjuvant effects were observed in modification with combinations of two different types of CpG motifs, which provides not only a contribution to the knowledge base on the adjuvant activities of CpG motifs combinations but also implications for the rational design of optimal DNA vaccines with combinations of CpG motifs as "built-in" adjuvants. We describe an efficient strategy to design and optimize DNA vaccines by the addition of combined immunostimulatory CpG motifs in a viral replicon DNA plasmid to produce strong immune responses, which indicates that the CpG-modified viral replicon DNA plasmid may be desirable for use as vector of DNA vaccines.
Tu, Yaqin; Hu, Yang; Fan, Guorun; Chen, Zhihao; Liu, Lin; Man, Dandan; Liu, Shuojie; Tang, Chengwu; Zhang, Yin; Dai, Wuxing
In order to explore the high performance bivalent DNA-based vaccine against schistosomes, SjFABP and Sj26GST were selected and used to construct a vaccine. Two strategies were used to construct the bivalent DNA vaccine. In the first strategy, a plasmid encoding antigen in the secreted form was used, while in the other, a plasmid encoding a truncated form of SjFABP and Sj26GST targeted to the cell surface was used. Various parameters, including antibody and cytokine response, proliferation, histopathological examination, and characterization of T cell subsets were used to evaluate the type of immune response and the level of protection against challenge infection. Injection with secreted pIRES-sjFABP-sj26GST significantly increased the levels of antibody, splenocyte proliferation, and production of IFN-γ, compared with membrane-anchored groups. Analysis of splenic T cell subsets showed that the secreted vaccine significantly increased the percentage of CD3+CD4+ and CD3+CD8+ T cells. Liver immunopathology (size of liver granulomas) was significantly reduced in the secreted group compared with the membrane-anchored groups. Moreover, challenge experiments showed that the worm and egg burdens were significantly reduced in animals immunized with recombinant vaccines. Most importantly, secreted Sj26GST-SjFABP markedly enhanced protection, by reducing worm and egg burdens by 31.8% and 24.78%, respectively, while the membrane-anchored group decreased worm and egg burdens by 24.80% and 18.80%, respectively. Taken together, these findings suggest that the secretory vaccine is more promising than the membrane-anchored vaccine, and provides support for the development and application of this vaccine. PMID:24466157
Wu, Yunpeng; Wang, Xinhai; Csencsits, Keri L.; Haddad, Asmahan; Walters, Nancy; Pascual, David W.
DNA immunization, although attractive, is poor for inducing mucosal immunity, thus limiting its protective value against most infectious agents. To surmount this shortcoming, we devised a method for mucosal transgene vaccination by using an M cell ligand to direct the DNA vaccine to mucosal inductive tissues and the respiratory epithelium. This ligand, reovirus protein 1, when conjugated to polylysine (PL), can bind the apical surface of M cells from nasal-associated lymphoid tissues. Intranasal immunizations with protein 1-PL-DNA complexes produced antigen-specific serum IgG and prolonged mucosal IgA, as well as enhanced cell-mediated immunity, made evident by elevated pulmonary cytotoxic T lymphocyte responses. Therefore, targeted transgene vaccination represents an approach for enabling DNA vaccination of the mucosa.
Dory, Daniel; Torché, Anne-Marie; Béven, Véronique; Cariolet, Roland; Jestin, André
No pseudorabies virus (PRV)-specific neutralizing or immunoglobulin G1-type antibodies were detected in sera 5 days after injection of a DNA vaccine against PRV infection in pigs. PRV-stimulated peripheral blood mononuclear cells produced gamma interferon mRNA in vitro. Two out of five pigs recovered from lethal PRV infection without attenuation of nasal viral excretion. PMID:17301217
Bissa, Massimiliano; Quaglino, Elena; Zanotto, Carlo; Illiano, Elena; Rolih, Valeria; Pacchioni, Sole; Cavallo, Federica; De Giuli Morghen, Carlo; Radaelli, Antonia
The control of smallpox was achieved using live vaccinia virus (VV) vaccine, which successfully eradicated the disease worldwide. As the variola virus no longer exists as a natural infection agent, mass vaccination was discontinued after 1980. However, emergence of smallpox outbreaks caused by accidental or deliberate release of variola virus has stimulated new research for second-generation vaccine development based on attenuated VV strains. Considering the closely related animal poxviruses that also arise as zoonoses, and the increasing number of unvaccinated or immunocompromised people, a safer and more effective vaccine is still required. With this aim, new vectors based on avian poxviruses that cannot replicate in mammals should improve the safety of conventional vaccines, and protect from zoonotic orthopoxvirus diseases, such as cowpox and monkeypox. In this study, DNA and fowlpox (FP) recombinants that expressed the VV L1R, A27L, A33R, and B5R genes were generated (4DNAmix, 4FPmix, respectively) and tested in mice using novel administration routes. Mice were primed with 4DNAmix by electroporation, and boosted with 4FPmix applied intranasally. The lethal VVIHD-J strain was then administered by intranasal challenge. All of the mice receiving 4DNAmix followed by 4FPmix, and 20% of the mice immunized only with 4FPmix, were protected. The induction of specific humoral and cellular immune responses directly correlated with this protection. In particular, higher anti-A27 antibodies and IFNγ-producing T lymphocytes were measured in the blood and spleen of the protected mice, as compared to controls. VVIHD-J neutralizing antibodies in sera from the protected mice suggest that the prime/boost vaccination regimen with 4DNAmix plus 4FPmix may be an effective and safe mode to induce protection against smallpox and poxvirus zoonotic infections. The electroporation/intranasal administration routes contributed to effective immune responses and mouse survival.
Song, Xiaokai; Zhang, Ruirui; Xu, Lixin; Yan, Ruofeng; Li, Xiangrui
Chimeric DNA vaccines co-expressing Eimeria acervulina macrophage migration inhibitory factor (E.MIF) and chicken IL-2 (IL-2) or interferon-γ (IFN-γ) were constructed and their efficacies against E. acervulina were evaluated. The open reading frame (ORF) of E.MIF was cloned from E. acervulina merozoites and subcloned into the eukaryotic expression vector pVAX1 with chicken cytokine gene IFN-γ or IL-2 to construct the DNA vaccines pVAX-E.MIF-IFN-γ, pVAX-E.MIF-IL-2 and pVAX-E.MIF. The in vivo transfection of the target genes was detected by use of reverse transcription polymerase chain reaction (RT-PCR) and Western blot. Immunizations were carried out by vaccinating chickens twice with a dose rate of 100 μg intramuscularly. Seven days post second immunization, all chickens except the unchallenged control group were challenged orally with 1 × 105 sporulated oocysts of E. acervulina. Seven days later, the duodenum was collected. The results showed that the target genes were expressed effectively in vivo. DNA vaccines and the recombinant E.MIF protein could alleviate body weight loss and duodenal lesions significantly compared to the control groups. Furthermore, pVAX-E.MIF-IL-2 and pVAX-E.MIF-IFN-γ induced anticoccidial indexs (ACIs) of 179.12 and 170, respectively, which were significantly higher than that of pVAX-E.MIF (ACI = 162.31). Our results demonstrated that E.MIF is a potential vaccine candidate against E. acervulina and chicken IFN-γ or IL- 2 may be used as genetic adjuvants to improve the efficacies of DNA vaccines against avian coccidiosis.
Suppression of antitumour protective cytotoxic T lymphocyte responses to a human papillomavirus 16 E7 DNA vaccine by coinjection of interleukin-12 complementary DNA: involvement of nitric oxide in immune suppression
Interleukin-12 (IL-12) has been shown to enhance cellular immunity in vitro and in vivo. The beneficial roles of IL-12 as a DNA vaccine adjuvant have been commonly observed. Here the impact of IL-12 complementary DNA (cDNA) as an adjuvant for a human papillomavirus (HPV) type 16 E7 DNA vaccine is investigated in a mouse tumour model. Coinjection of E7 DNA vaccine with IL-12 cDNA completely suppressed antigen-specific cytotoxic T-lymphocyte (CTL) responses, leading to a complete loss of antitumour protection from a tumour cell challenge. In addition, antigen-specific antibody and T helper cell proliferative responses were also suppressed by IL-12 cDNA coinjection. This inhibition was observed over different IL-12 cDNA doses. Furthermore, separate leg injections of IL-12 and E7 cDNAs suppressed antigen-specific CTL and tumour protective responses, but not antibody and T helper cell proliferative responses, suggesting different pathways for suppression of these two separate responses. Further knockout animal studies demonstrated that interferon-γ and nitric oxide are not directly associated with suppression of antigen-specific antibody responses by IL-12 cDNA coinjection. However, nitric oxide was found to be involved in suppression of antigen-specific CTL and tumour protective responses by IL-12 cDNA coinjection. These data suggest that coinjection of IL-12 cDNA results in suppression of E7-specific CTL responses through nitric oxide, leading to a loss of antitumour resistance in this DNA vaccine model. This study further shows that the adjuvant effect of IL-12 is dependent on the antigen types tested. PMID:19740332
Suppression of antitumour protective cytotoxic T lymphocyte responses to a human papillomavirus 16 E7 DNA vaccine by coinjection of interleukin-12 complementary DNA: involvement of nitric oxide in immune suppression.
Interleukin-12 (IL-12) has been shown to enhance cellular immunity in vitro and in vivo. The beneficial roles of IL-12 as a DNA vaccine adjuvant have been commonly observed. Here the impact of IL-12 complementary DNA (cDNA) as an adjuvant for a human papillomavirus (HPV) type 16 E7 DNA vaccine is investigated in a mouse tumour model. Coinjection of E7 DNA vaccine with IL-12 cDNA completely suppressed antigen-specific cytotoxic T-lymphocyte (CTL) responses, leading to a complete loss of antitumour protection from a tumour cell challenge. In addition, antigen-specific antibody and T helper cell proliferative responses were also suppressed by IL-12 cDNA coinjection. This inhibition was observed over different IL-12 cDNA doses. Furthermore, separate leg injections of IL-12 and E7 cDNAs suppressed antigen-specific CTL and tumour protective responses, but not antibody and T helper cell proliferative responses, suggesting different pathways for suppression of these two separate responses. Further knockout animal studies demonstrated that interferon-gamma and nitric oxide are not directly associated with suppression of antigen-specific antibody responses by IL-12 cDNA coinjection. However, nitric oxide was found to be involved in suppression of antigen-specific CTL and tumour protective responses by IL-12 cDNA coinjection. These data suggest that coinjection of IL-12 cDNA results in suppression of E7-specific CTL responses through nitric oxide, leading to a loss of antitumour resistance in this DNA vaccine model. This study further shows that the adjuvant effect of IL-12 is dependent on the antigen types tested.
Palmer, John; Bell, Matt; Darko, Christian; Barnewall, Roy; Keane-Myers, Andrea
In the past decade, several Bacillus cereus strains have been isolated from otherwise healthy individuals who succumbed to bacterial pneumonia presenting symptoms resembling inhalational anthrax. One strain was indistinguishable from B. cereus G9241, previously cultured from an individual who survived a similar pneumonia-like illness and which was shown to possess a complete set of plasmid-borne anthrax toxin-encoding homologs. The finding that B. cereus G9241 pathogenesis in mice is dependent on pagA1-derived protective antigen (PA) synthesis suggests that an anthrax toxin-based vaccine may be effective against this toxin-encoding B. cereus strain. Dunkin Hartley guinea pigs were immunized with protein- and DNA-based anthrax toxin-based vaccines, immune responses were evaluated and survival rates were calculated after lethal aerosol exposure with B. cereus G9241 spores. Each vaccine induced seroconversion with the protein immunization regimen eliciting significantly higher serum levels of antigen-specific antibodies at the prechallenge time-point compared with the DNA-protein prime-boost immunization schedule. Complete protection against lethal challenge was observed in all groups with a detectable prechallenge serum titer of toxin neutralizing antibodies. For the first time, we demonstrated that the efficacy of fully defined anthrax toxin-based vaccines was protective against lethal B. cereus G9241 aerosol challenge in the guinea pig animal model.
Protective immunity conferred by a DNA adenine methylase deficient Salmonella enterica serovar Typhimurium vaccine when delivered in-water to sheep challenged with Salmonella enterica serovar Typhimurium.
Mohler, V L; Heithoff, D M; Mahan, M J; Walker, K H; Hornitzky, M A; Gabor, L; Thomson, P C; Thompson, A; House, J K
Stimulation of acquired immunity to Salmonella in livestock is not feasible in neonates (which can be infected within 24h of birth) and is challenging in feedlots, which typically source animals from diverse locations and vendors. Induction of innate immune mechanisms through mass vaccination of animals upon arrival to feedlots is an alternative approach. Transport, environmental conditions, changes in social grouping, and further handling during feedlot assembly are significant stressors. These factors, as well as concurrent exposure to a diversity of pathogens, contribute to the risk of disease. We have shown that oral immunization of calves with a modified live Salmonella enterica serovar Typhimurium vaccine strain, which lacks the DNA adenine methylase gene (S. Typhimurium dam), attenuates the severity of clinical disease, reduces fecal shedding, and promotes clearance of salmonellae following virulent homologous and heterologous challenge. This study examines the safety and efficacy of a S. Typhimurium dam vaccine in sheep via oral delivery in drinking water (ad libitum), as a means to effectively vaccinate large groups of animals. Adult merino sheep were vaccinated in drinking water -28 days, -7 days and 24h pre and 24h post-virulent Salmonella Typhimurium challenge which was administered via the oral route. Significant attenuation of clinical disease (temperature, appetite, and attitude) and reduction in mortality and virulent Salmonella Typhimurium fecal shedding and tissue colonization was observed in animals that received the vaccine 28 and 7 days pre-challenge. Further, vaccination did not pose a risk to stock previously infected with virulent salmonellae as mortalities and clinical disease in sheep vaccinated prior to or following virulent challenge did not differ significantly from the non-vaccinated controls. The capacity of S. Typhimurium dam vaccines delivered in drinking water to protect livestock from virulent Salmonella challenge offers an
Santana, Vinicius C; Diniz, Mariana O; Cariri, Francisco A M O; Ventura, Armando M; Cunha-Neto, Edécio; Almeida, Rafael R; Campos, Marco A; Lima, Graciela K; Ferreira, Luís C S
Millions of people worldwide are currently infected with human papillomavirus (HPV), herpes simplex virus (HSV) or human immunodeficiency virus (HIV). For this enormous contingent of people, the search for preventive and therapeutic immunological approaches represents a hope for the eradication of latent infection and/or virus-associated cancer. To date, attempts to develop vaccines against these viruses have been mainly based on a monovalent concept, in which one or more antigens of a virus are incorporated into a vaccine formulation. In the present report, we designed and tested an immunization strategy based on DNA vaccines that simultaneously encode antigens for HIV, HSV and HPV. With this purpose in mind, we tested two bicistronic DNA vaccines (pIRES I and pIRES II) that encode the HPV-16 oncoprotein E7 and the HIV protein p24 both genetically fused to the HSV-1 gD envelope protein. Mice i.m. immunized with the DNA vaccines mounted antigen-specific CD8⁺ T cell responses, including in vivo cytotoxic responses, against the three antigens. Under experimental conditions, the vaccines conferred protective immunity against challenges with a vaccinia virus expressing the HIV-derived protein Gag, an HSV-1 virus strain and implantation of tumor cells expressing the HPV-16 oncoproteins. Altogether, our results show that the concept of a trivalent HIV, HSV, and HPV vaccine capable to induce CD8⁺ T cell-dependent responses is feasible and may aid in the development of preventive and/or therapeutic approaches for the control of diseases associated with these viruses.
Heraud, Jean-Michel; Edghill-Smith, Yvette; Ayala, Victor; Kalisz, Irene; Parrino, Janie; Kalyanaraman, Vaniambadi S; Manischewitz, Jody; King, Lisa R; Hryniewicz, Anna; Trindade, Christopher J; Hassett, Meredith; Tsai, Wen-Po; Venzon, David; Nalca, Aysegul; Vaccari, Monica; Silvera, Peter; Bray, Mike; Graham, Barney S; Golding, Hana; Hooper, Jay W; Franchini, Genoveffa
The smallpox vaccine Dryvax, a live vaccinia virus (VACV), protects against smallpox and monkeypox, but is contraindicated in immunocompromised individuals. Because Abs to VACV mediate protection, a live virus vaccine could be substituted by a safe subunit protein-based vaccine able to induce a protective Ab response. We immunized rhesus macaques with plasmid DNA encoding the monkeypox orthologs of the VACV L1R, A27L, A33R, and B5R proteins by the intradermal and i.m. routes, either alone or in combination with the equivalent recombinant proteins produced in Escherichia coli. Animals that received only DNA failed to produce high titer Abs, developed innumerable skin lesions after challenge, and died in a manner similar to placebo controls. By contrast, the animals vaccinated with proteins developed moderate to severe disease (20-155 skin lesions) but survived. Importantly, those immunized with DNA and boosted with proteins had mild disease with 15 or fewer lesions that resolved within days. DNA/protein immunization elicited Th responses and binding Ab titers to all four proteins that correlated negatively with the total lesion number. The sera of the immunized macaques recognized a limited number of linear B cell epitopes that are highly conserved among orthopoxviruses. Their identification may guide future efforts to develop simpler, safer, and more effective vaccines for monkeypox and smallpox.
Coban, Cevayir; Kobiyama, Kouji; Jounai, Nao; Tozuka, Miyuki; Ishii, Ken J
Since the introduction of DNA vaccines two decades ago, this attractive strategy has been hampered by its low immunogenicity in humans. Studies conducted to improve the immunogenicity of DNA vaccines have shown that understanding the mechanism of action of DNA vaccines might be the key to successfully improving their immunogenicity. Our current understanding is that DNA vaccines induce innate and adaptive immune responses in two ways: (1) encoded protein (or polypeptide) antigen(s) by the DNA plasmid can be expressed in stromal cells (i.e., muscle cells) as well as DCs, where these antigens are processed and presented to naïve CD4 or CD8 T cells either by direct or cross presentation, respectively; and (2) the transfected DNA plasmid itself may bind to an un-identified cytosolic DNA sensor and activate the TBK1-STING pathway and the production of type I interferons (IFNs) which function as an adjuvant. Recent studies investigating double-stranded cytosolic DNA sensor(s) have highlighted new mechanisms in which cytosolic DNA may release secondary metabolites, which are in turn recognized by a novel DNA sensing machinery. Here, we discuss these new metabolites and the possibilities of translating this knowledge into improved immunogenicity for DNA vaccines.
Cao, Aiping; Liu, Yuan; Wang, Jingjing; Li, Xun; Wang, Shuai; Zhao, Qunli; Cong, Hua; He, Shenyi; Zhou, Huaiyu
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. Copyright © 2015 Elsevier Ltd. All rights reserved.
Steisslinger, Vera; Korten, Simone; Brattig, Norbert W; Erttmann, Klaus D
River blindness, caused by the filarial parasite Onchocerca volvulus, is a major socio-economic and public health problem in Sub-Saharan Africa. In January 2015, The Onchocerciasis Vaccine for Africa (TOVA) Initiative has been launched with the aim of providing new tools to complement mass drug administration (MDA) of ivermectin, thereby promoting elimination of onchocerciasis in Africa. In this context we here present Onchocerca volvulus glyceraldehyde-3-phosphate dehydrogenase (Ov-GAPDH) as a possible DNA vaccine candidate. We report that in a laboratory model for filariasis, immunization with Ov-GAPDH led to a significant reduction of adult worm load and microfilaraemia in BALB/c mice after challenge infection with the filarial parasite Litomosoides sigmodontis. Mice were either vaccinated with Ov-GAPDH.DNA plasmid (Ov-pGAPDH.DNA) alone or in combination with recombinantly expressed Ov-GAPDH protein (Ov-rGAPDH). During the following challenge infection of immunized and control mice with L. sigmodontis, those formulations which included the DNA plasmid, led to a significant reduction of adult worm loads (up to 57% median reduction) and microfilaraemia (up to 94% reduction) in immunized animals. In a further experiment, immunization with a mixture of four overlapping, synthetic Ov-GAPDH peptides (Ov-GAPDHpept), with alum as adjuvant, did not significantly reduce worm loads. Our results indicate that DNA vaccination with Ov-GAPDH has protective potential against filarial challenge infection in the mouse model. This suggests a transfer of the approach into the cattle Onchocerca ochengi model, where it is possible to investigate the effects of this vaccination in the context of a natural host-parasite relationship.
Chuang, Ilin; Sedegah, Martha; Cicatelli, Susan; Spring, Michele; Polhemus, Mark; Tamminga, Cindy; Patterson, Noelle; Guerrero, Melanie; Bennett, Jason W.; McGrath, Shannon; Ganeshan, Harini; Belmonte, Maria; Farooq, Fouzia; Abot, Esteban; Banania, Jo Glenna; Huang, Jun; Newcomer, Rhonda; Rein, Lisa; Litilit, Dianne; Richie, Nancy O.; Wood, Chloe; Murphy, Jittawadee; Sauerwein, Robert; Hermsen, Cornelus C.; McCoy, Andrea J.; Kamau, Edwin; Cummings, James; Komisar, Jack; Sutamihardja, Awalludin; Shi, Meng; Epstein, Judith E.; Maiolatesi, Santina; Tosh, Donna; Limbach, Keith; Angov, Evelina; Bergmann-Leitner, Elke; Bruder, Joseph T.; Doolan, Denise L.; King, C. Richter; Carucci, Daniel; Dutta, Sheetij; Soisson, Lorraine; Diggs, Carter; Hollingdale, Michael R.; Ockenhouse, Christian F.; Richie, Thomas L.
Background Gene-based vaccination using prime/boost regimens protects animals and humans against malaria, inducing cell-mediated responses that in animal models target liver stage malaria parasites. We tested a DNA prime/adenovirus boost malaria vaccine in a Phase 1 clinical trial with controlled human malaria infection. Methodology/Principal Findings The vaccine regimen was three monthly doses of two DNA plasmids (DNA) followed four months later by a single boost with two non-replicating human serotype 5 adenovirus vectors (Ad). The constructs encoded genes expressing P. falciparum circumsporozoite protein (CSP) and apical membrane antigen-1 (AMA1). The regimen was safe and well-tolerated, with mostly mild adverse events that occurred at the site of injection. Only one AE (diarrhea), possibly related to immunization, was severe (Grade 3), preventing daily activities. Four weeks after the Ad boost, 15 study subjects were challenged with P. falciparum sporozoites by mosquito bite, and four (27%) were sterilely protected. Antibody responses by ELISA rose after Ad boost but were low (CSP geometric mean titer 210, range 44–817; AMA1 geometric mean micrograms/milliliter 11.9, range 1.5–102) and were not associated with protection. Ex vivo IFN-γ ELISpot responses after Ad boost were modest (CSP geometric mean spot forming cells/million peripheral blood mononuclear cells 86, range 13–408; AMA1 348, range 88–1270) and were highest in three protected subjects. ELISpot responses to AMA1 were significantly associated with protection (p = 0.019). Flow cytometry identified predominant IFN-γ mono-secreting CD8+ T cell responses in three protected subjects. No subjects with high pre-existing anti-Ad5 neutralizing antibodies were protected but the association was not statistically significant. Significance The DNA/Ad regimen provided the highest sterile immunity achieved against malaria following immunization with a gene-based subunit vaccine (27%). Protection was
Cheng, Yingjie; Chen, Zongyan; Li, Chuanfeng; Meng, Chun; Wu, Run; Liu, Guangqing
A suicidal DNA vaccine based on a Semliki Forest virus (SFV) replicon was evaluated for the development of a vaccine against rabbit hemorrhagic disease virus (RHDV). The VP60 gene of RHDV was cloned and inserted into pSCA1, an SFV DNA-based replicon vector. The resultant plasmid, pSCA/VP60, was transfected into BHK-21 cells, and the antigenicity of the expressed protein was confirmed using indirect immunofluorescence and a western blot assay. In addition, immunogenicity was studied in rabbits. Fifteen rabbits were injected intramuscularly twice with pSCA/VP60 at 2-week intervals. They were challenged with an RHDV isolate 2weeks after the second immunization. In all cases, anti-RHDV antibodies were detected by ELISA. Additionally, the lymphocyte proliferation response was tested by the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide method, and neutralizing antibodies were measured by microneutralization tests. Our results showed that RHDV-specific antibodies and an RHDV-specific cell-mediated immune response were strongly induced in rabbits. Furthermore, all of the rabbits were protected against challenge with wild type RHDV. In conclusion, we demonstrated that the suicidal DNA vaccine is a promising vaccine candidate that facilitates the prevention of rabbit hemorrhagic disease caused by RHDV.
Sun, Shi-Qi; Liu, Xiang-Tao; Guo, Hui-Chen; Yin, Shuang-Hui; Shang, You-Jun; Feng, Xia; Liu, Zai-Xin; Xie, Qing-Ge
A suicidal DNA vaccine based on a Semliki Forest virus (SFV) replicon was evaluated for the development of a vaccine against swine vesicular disease virus (SVDV). The 1BCD gene of SVDV was cloned and inserted into pSCA1, an SFV DNA-based replicon vector. The resultant plasmid, pSCA/1BCD, was transfected into BHK-21 cells and the antigenicity of the expressed protein was confirmed using an indirect immunofluorescence assay. Immunogenicity was studied in guinea pigs and swine. Animals were injected intramuscularly three times with pSCA/1BCD at regular intervals. Anti-SVDV antibodies were detected by ELISA, the lymphocyte proliferation response was tested by the 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide method and neutralizing antibodies were measured by microneutralization tests. The data showed that SVDV-specific antibodies, neutralizing antibodies and lymphocyte proliferation were induced in both guinea pigs and swine. Furthermore, after three successive vaccinations with pSCA/1BCD, half of the pigs were protected against challenge with SVDV. These results should encourage further work towards the development of a DNA vaccine against SVDV.
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. PMID:24552235
Dory, Daniel; Fischer, Timo; Béven, Véronique; Cariolet, Roland; Rziha, Hanns-Joachim; Jestin, André
The present study demonstrates the protective potential of a novel prime-boost vaccination strategy of pigs against lethal Pseudorabies virus (PRV; Herpes suid 1) infection. Animals were primed with Sindbis virus-derived plasmids that express viral glycoproteins gC and gD (gC- and gD-pSIN) and subsequently booster immunized with Orf virus (ORFV; Parapoxvirus) recombinants expressing gC and gD (D1701-VrVgC and -VrVgD). The prime-boost vaccination induced strong humoral and cellular-like PRV-specific immune responses. All prime-boost vaccinated pigs survived the lethal challenge infection without PRV-specific clinical symptoms and presented excellent body weight loss attenuation. Most notably, nasal shedding of challenge virus was reduced by more than about 3log(10), clearly reducing the risk of infection of non-immunized pigs.
Chen, Hui; Gao, Na; Fan, Dongying; Wu, Jiangman; Zhu, Junping; Li, Jieqiong; Wang, Juan; Chen, Yanlei; An, Jing
As a potential cytokine adjuvant of DNA vaccines, granulocyte-macrophage colony–stimulating factor (GM-CSF) has received considerable attention due to its essential role in the recruitment of antigen-presenting cells, differentiation and maturation of dendritic cells. However, in our recent study of a Japanese encephalitis virus (JEV) DNA vaccine, co-inoculation of a GM-CSF plasmid dramatically suppressed the specific IgG response and resulted in decreased protection against JEV challenge. It is known that GM-CSF has been used in clinic to treat neutropenia for repopulating myeloid cells, and as an adjuvant in vaccine studies; it has shown various effects on the immune response. Therefore, in this study, we characterized the suppressive effects on the immune response to a JEV DNA vaccine by the co-administration of the GM-CSF-expressing plasmid and clarified the underlying mechanisms of the suppression in mice. Our results demonstrated that co-immunization with GM-CSF caused a substantial dampening of the vaccine-induced antibody responses. The suppressive effect was dose- and timing-dependent and likely related to the immunogenicity of the antigen. The suppression was associated with the induction of immature dendritic cells and the expansion of regulatory T cells but not myeloid-derived suppressor cells. Collectively, our findings not only provide valuable information for the application of GM-CSF in clinic and using as a vaccine adjuvant but also offer further insight into the understanding of the complex roles of GM-CSF. PMID:22493704
Soto, Manuel; Corvo, Laura; Garde, Esther; Ramírez, Laura; Iniesta, Virginia; Bonay, Pedro; Gómez-Nieto, Carlos; González, Víctor M; Martín, M Elena; Alonso, Carlos; Coelho, Eduardo A F; Barral, Aldina; Barral-Netto, Manoel; Iborra, Salvador
Highly conserved intracellular proteins from Leishmania have been described as antigens in natural and experimental infected mammals. The present study aimed to evaluate the antigenicity and prophylactic properties of the Leishmania infantum Poly (A) binding proteins (LiPABPs). Three different members of the LiPABP family have been described. Recombinant tools based on these proteins were constructed: recombinant proteins and DNA vaccines. The three recombinant proteins were employed for coating ELISA plates. Sera from human and canine patients of visceral leishmaniasis and human patients of mucosal leishmaniasis recognized the three LiPABPs. In addition, the protective efficacy of a DNA vaccine based on the combination of the three Leishmania PABPs has been tested in a model of progressive murine leishmaniasis: BALB/c mice infected with Leishmania major. The induction of a Th1-like response against the LiPABP family by genetic vaccination was able to down-regulate the IL-10 predominant responses elicited by parasite LiPABPs after infection in this murine model. This modulation resulted in a partial protection against L. major infection. LiPABP vaccinated mice showed a reduction on the pathology that was accompanied by a decrease in parasite burdens, in antibody titers against Leishmania antigens and in the IL-4 and IL-10 parasite-specific mediated responses in comparison to control mice groups immunized with saline or with the non-recombinant plasmid. The results presented here demonstrate for the first time the prophylactic properties of a new family of Leishmania antigenic intracellular proteins, the LiPABPs. The redirection of the immune response elicited against the LiPABP family (from IL-10 towards IFN-γ mediated responses) by genetic vaccination was able to induce a partial protection against the development of the disease in a highly susceptible murine model of leishmaniasis.
Corvo, Laura; Garde, Esther; Ramírez, Laura; Iniesta, Virginia; Bonay, Pedro; Gómez-Nieto, Carlos; González, Víctor M.; Martín, M. Elena; Alonso, Carlos; Coelho, Eduardo A. F.; Barral, Aldina; Barral-Netto, Manoel
Background Highly conserved intracellular proteins from Leishmania have been described as antigens in natural and experimental infected mammals. The present study aimed to evaluate the antigenicity and prophylactic properties of the Leishmania infantum Poly (A) binding proteins (LiPABPs). Methodology/Principal Findings Three different members of the LiPABP family have been described. Recombinant tools based on these proteins were constructed: recombinant proteins and DNA vaccines. The three recombinant proteins were employed for coating ELISA plates. Sera from human and canine patients of visceral leishmaniasis and human patients of mucosal leishmaniasis recognized the three LiPABPs. In addition, the protective efficacy of a DNA vaccine based on the combination of the three Leishmania PABPs has been tested in a model of progressive murine leishmaniasis: BALB/c mice infected with Leishmania major. The induction of a Th1-like response against the LiPABP family by genetic vaccination was able to down-regulate the IL-10 predominant responses elicited by parasite LiPABPs after infection in this murine model. This modulation resulted in a partial protection against L. major infection. LiPABP vaccinated mice showed a reduction on the pathology that was accompanied by a decrease in parasite burdens, in antibody titers against Leishmania antigens and in the IL-4 and IL-10 parasite-specific mediated responses in comparison to control mice groups immunized with saline or with the non-recombinant plasmid. Conclusion/Significance The results presented here demonstrate for the first time the prophylactic properties of a new family of Leishmania antigenic intracellular proteins, the LiPABPs. The redirection of the immune response elicited against the LiPABP family (from IL-10 towards IFN-γ mediated responses) by genetic vaccination was able to induce a partial protection against the development of the disease in a highly susceptible murine model of leishmaniasis. PMID:25955652
Gupta, Shivali; Garg, Nisha J.
In this study, we evaluated the long-term efficacy of a two-component subunit vaccine against Trypanosoma cruzi infection. C57BL/6 mice were immunized with TcG2/TcG4 vaccine delivered by a DNA-prime/Protein-boost (D/P) approach and challenged with T. cruzi at 120 or 180 days post-vaccination (dpv). We examined whether vaccine-primed T cell immunity was capable of rapid expansion and intercepting the infecting T. cruzi. Our data showed that D/P vaccine elicited CD4+ (30-38%) and CD8+ (22-42%) T cells maintained an effector phenotype up to 180 dpv, and were capable of responding to antigenic stimulus or challenge infection by a rapid expansion (CD8>CD4) with type 1 cytokine (IFNγ+ and TFNα+) production and cytolytic T lymphocyte (CTL) activity. Subsequently, challenge infection at 120 or 180 dpv, resulted in 2-3-fold lower parasite burden in vaccinated mice than was noted in unvaccinated/infected mice. Co-delivery of IL-12- and GMCSF-encoding expression plasmids provided no significant benefits in enhancing the anti-parasite efficacy of the vaccine-induced T cell immunity. Booster immunization (bi) with recombinant TcG2/TcG4 proteins 3-months after primary vaccine enhanced the protective efficacy, evidenced by an enhanced expansion (1.2-2.8-fold increase) of parasite-specific, type 1 CD4+ and CD8+ T cells and a potent CTL response capable of providing significantly improved (3-4.5-fold) control of infecting T. cruzi. Further, CD8+T cells in vaccinated/bi mice were predominantly of central memory phenotype, and capable of responding to challenge infection 4-6-months post bi by a rapid expansion to a poly-functional effector phenotype, and providing a 1.5-2.3-fold reduction in tissue parasite replication. We conclude that the TcG2/TcG4 D/P vaccine provided long-term anti-T. cruzi T cell immunity, and bi would be an effective strategy to maintain or enhance the vaccine-induced protective immunity against T. cruzi infection and Chagas disease. PMID:25951312
Méndez, Susana; Belkaid, Yasmine; Seder, Robert A; Sacks, David
The present studies were designed to examine the requirements of dose, route of inoculation and constituent antigens for the maintenance of complete and long lasting protection against cutaneous leishmaniasis due to Leishmania major conferred by a cocktail DNA vaccine encoding the Leishmania antigens LACK, LmST11 and TSA. Vaccination of C57Bl/6 mice with LACK DNA alone resulted in partial protection, whereas the combination of LmST11 and TSA provided stronger, though still incomplete protection compared to the combination of all three Ag DNAs. When intradermal (i.d), intramuscular (i.m.), and subcutaneous (s.c.) vaccination routes were compared, i.d. immunization reduced by five-fold the dose necessary to maintain complete protection. In vivo depletion of CD4+ or CD8+ T cells provided direct evidence that both populations are necessary to mediate complete protection. These results establish intradermal vaccination using DNA encoding multiple Leishmania antigens as a way to optimize priming of CD4+ and CD8+ T cells necessary for potent and durable protection against cutaneous leishmaniasis.
Zhang, Min; Zhao, Lingxiao; Song, Jing; Li, Ying; Zhao, Qunli; He, Shenyi; Cong, Hua
The surface antigens SAG2C, SAG2D, and SAG2X, which expressed specifically on bradyzoite stage of Toxoplasma gondii, have been demonstrated to be important for persistence of cyst in the brain. In this study, DNA vaccines expressing SAG2C, SAG2D, and SAG2X of T. gondii were constructed and their protective efficacy were evaluated in BALB/c mice. Mice vaccinated with pVAX1-SAG2C (pSAG2C), pVAX1-2D (pSAG2D) or pVAX1-2X (pSAG2C) showed higher levels of serum IgG antibodies and lymphocyte proliferation response compared to PBS and pVAX1 treated mice (p<0.05). The immune response was characterized by a strong Th1 response and increased cytokine production of IL-2 and IFN-γ. Vaccinated mice displayed significant protection against the challenge with the cyst of T. gondii genotype II strain of PRU (cyst-forming in mouse). A significant reduction in the brain cyst burden was detected in the mice immunized with pSAG2C (72%), pSAG2D (23%), pSAG2X (69%) alone and even more reduction rate, 77%, was achieved in the combination group compared to PBS treated mice. The results implied that immunization with DNA vaccines expressing SAG2C, SAG2D, and SAG2X, and, in particular, a combination of all three DNA plasmids, could effectively protect the mice against T. gondii chronic infection.
unlimited. Abstract 32 33 Crimean-Congo hemorrhagic fever virus (CCHFV) is a tick-borne virus capable of causing a 34 severe hemorrhagic fever...the 50 glycoprotein genes of CCHFV elicits protective immunity against CCHFV. 51 52 Author summary 53 Crimean-Congo Hemorrhagic Fever Virus ...CCHFV) is a tick-borne virus capable of causing 54 lethal human disease against which there are currently no approved vaccines. In this study, we 55 TR
Rashid, Imran; Hedhli, Dorsaf; Moiré, Nathalie; Pierre, Josette; Debierre-Grockiego, Françoise; Dimier-Poisson, Isabelle; Mévélec, Marie Noëlle
The development of an effective vaccine against Toxoplasma gondii infection is an important issue due to the seriousness of the related public health problems, and the economic importance of this parasitic disease worldwide. Rhoptry neck proteins (RONs) are components of the moving junction macromolecular complex formed during invasion. The aim of this study was to evaluate the vaccine potential of RON4 using two vaccination strategies: DNA vaccination by the intramuscular route, and recombinant protein vaccination by the nasal route. We produced recombinant RON4 protein (RON4S2) using the Schneider insect cells expression system, and validated its antigenicity and immunogenicity. We also constructed optimized plasmids encoding full length RON4 (pRON4), or only the N-terminal (pNRON4), or the C-terminal part (pCRON4) of RON4. CBA/J mice immunized with pRON4, pNRON4 or pCRON4 plus a plasmid encoding the granulocyte-macrophage-colony-stimulating factor showed high IgG titers against rRON4S2. Mice immunized by the nasal route with rRON4S2 plus cholera toxin exhibited low levels of anti-RON4S2 IgG antibodies, and no intestinal IgA antibodies specific to RON4 were detected. Both DNA and protein vaccination generated a mixed Th1/Th2 response polarized towards the IgG1 antibody isotype. Both DNA and protein vaccination primed CD4+ T cells in vivo. In addition to the production of IFN-γ, and IL-2, Il-10 and IL-5 were also produced by the spleen cells of the immunized mice stimulated with RON4S2, suggesting that a mixed Th1/Th2 type immune response occurred in all the immunized groups. No cytokine was detectable in stimulated mesenteric lymph nodes from mice immunized by the nasal route. Immune responses were induced by both DNA and protein vaccination, but failed to protect the mice against a subsequent oral challenge with T. gondii cysts. In conclusion, strategies designed to enhance the immunogenicity and to redirect the cellular response towards a Th1 type response
Grødeland, Gunnveig; Bogen, Bjarne
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.
Liu, Margaret A
This review provides a detailed look at the attributes and immunologic mechanisms of plasmid DNA vaccines and their utility as laboratory tools as well as potential human vaccines. The immunogenicity and efficacy of DNA vaccines in a variety of preclinical models is used to illustrate how they differ from traditional vaccines in novel ways due to the in situ antigen production and the ease with which they are constructed. The ability to make new DNA vaccines without needing to handle a virulent pathogen or to adapt the pathogen for manufacturing purposes demonstrates the potential value of this vaccine technology for use against emerging and epidemic pathogens. Similarly, personalized anti-tumor DNA vaccines can also readily be made from a biopsy. Because DNA vaccines bias the T-helper (Th) cell response to a Th1 phenotype, DNA vaccines are also under development for vaccines against allergy and autoimmune diseases. The licensure of four animal health products, including two prophylactic vaccines against infectious diseases, one immunotherapy for cancer, and one gene therapy delivery of a hormone for a food animal, provides evidence of the efficacy of DNA vaccines in multiple species including horses and pigs. The size of these target animals provides evidence that the somewhat disappointing immunogenicity of DNA vaccines in a number of human clinical trials is not due simply to the larger mass of humans compared with most laboratory animals. The insights gained from the mechanisms of protection in the animal vaccines, the advances in the delivery and expression technologies for increasing the potency of DNA vaccines, and encouragingly potent human immune responses in certain clinical trials, provide insights for future efforts to develop DNA vaccines into a broadly useful vaccine and immunotherapy platform with applications for human and animal health.
Bao, Huihui; Ramanathan, Aarti A; Kawalakar, Omkar; Sundaram, Senthil G; Tingey, Colleen; Bian, Charoran B; Muruganandam, Nagarajan; Vijayachari, Paluru; Sardesai, Niranjan Y; Weiner, David B; Ugen, Kenneth E; Muthumani, Karuppiah
Chikungunya virus (CHIKV) is an important emerging mosquito-borne alphavirus, indigenous to tropical Africa and Asia. It can cause epidemic fever and acute illness characterized by fever and arthralgias. The epidemic cycle of this infection is similar to dengue and urban yellow fever viral infections. The generation of an efficient vaccine against CHIKV is necessary to prevent and/or control the disease manifestations of the infection. In this report, we studied immune response against a CHIKV-envelope DNA vaccine (pEnv) and the role of the CHIKV nonstructural gene 2 (nsP2) as an adjuvant for the induction of protective immune responses in a relevant mouse challenge model. When injected with the CHIKV pEnv alone, 70% of the immunized mice survived CHIKV challenge, whereas when co-injected with pEnv+pnsP2, 90% of the mice survived viral challenge. Mice also exhibited a delayed onset signs of illness, and a marked decrease in morbidity, suggesting a nsP2 mediated adjuvant effect. Co-injection of the pnsP2 adjuvant with pEnv also qualitatively and quantitatively increased antigen specific neutralizing antibody responses compared to vaccination with pEnv alone. In sum, these novel data imply that the addition of nsP2 to the pEnv vaccine enhances anti-CHIKV-Env immune responses and maybe useful to include in future CHIKV clinical vaccination strategies.
McNeel, Douglas G.; Becker, Jordan T.; Johnson, Laura E.; Olson, Brian M.
Delivery of plasmid DNA encoding an antigen of interest has been demonstrated to be an effective means of immunization, capable of eliciting antigen-specific T cells. Plasmid DNA vaccines offer advantages over other anti-tumor vaccine approaches in terms of simplicity, manufacturing, and possibly safety. The primary disadvantage is their poor transfection efficiency and subsequent lower immunogenicity relative to other genetic vaccine approaches. However, multiple preclinical models demonstrate anti-tumor efficacy, and many efforts are underway to improve the immunogenicity and anti-tumor effect of these vaccines. Clinical trials using DNA vaccines as treatments for prostate cancer have begun, and to date have demonstrated safety and immunological effect. This review will focus on DNA vaccines as a specific means of antigen delivery, advantages and disadvantages of this type of immunization, previous experience in preclinical models and human trials specifically conducted for the treatment of prostate cancer, and future directions for the application of DNA vaccines to prostate cancer immunotherapy. PMID:24587772
Gonçalves, Antônio J. S.; Oliveira, Edson R. A.; Costa, Simone M.; Paes, Marciano V.; Silva, Juliana F. A.; Azevedo, Adriana S.; Mantuano-Barradas, Marcio; Nogueira, Ana Cristina M. A.; Almeida, Cecília J.; Alves, Ada M. B.
Dengue virus (DENV) is spread through most tropical and subtropical areas of the world and represents a serious public health problem. At present, the control of dengue disease is mainly hampered by the absence of antivirals or a vaccine, which results in an estimated half worldwide population at risk of infection. The immune response against DENV is not yet fully understood and a better knowledge of it is now recognized as one of the main challenge for vaccine development. In previous studies, we reported that a DNA vaccine containing the signal peptide sequence from the human tissue plasminogen activator (t-PA) fused to the DENV2 NS1 gene (pcTPANS1) induced protection against dengue in mice. In the present work, we aimed to elucidate the contribution of cellular and humoral responses elicited by this vaccine candidate for protective immunity. We observed that pcTPANS1 exerts a robust protection against dengue, inducing considerable levels of anti-NS1 antibodies and T cell responses. Passive immunization with anti-NS1 antibodies conferred partial protection in mice infected with low virus load (4 LD50), which was abrogated with the increase of viral dose (40 LD50). The pcTPANS1 also induced activation of CD4+ and CD8+ T cells. We detected production of IFN-γ and a cytotoxic activity by CD8+ T lymphocytes induced by this vaccine, although its contribution in the protection was not so evident when compared to CD4+ cells. Depletion of CD4+ cells in immunized mice completely abolished protection. Furthermore, transfer experiments revealed that animals receiving CD4+ T cells combined with anti-NS1 antiserum, both obtained from vaccinated mice, survived virus infection with survival rates not significantly different from pcTPANS1-immunized animals. Taken together, results showed that the protective immune response induced by the expression of NS1 antigen mediated by the pcTPANS1 requires a cooperation between CD4+ T cells and the humoral immunity. PMID:26650916
Gonçalves, Antônio J S; Oliveira, Edson R A; Costa, Simone M; Paes, Marciano V; Silva, Juliana F A; Azevedo, Adriana S; Mantuano-Barradas, Marcio; Nogueira, Ana Cristina M A; Almeida, Cecília J; Alves, Ada M B
Dengue virus (DENV) is spread through most tropical and subtropical areas of the world and represents a serious public health problem. At present, the control of dengue disease is mainly hampered by the absence of antivirals or a vaccine, which results in an estimated half worldwide population at risk of infection. The immune response against DENV is not yet fully understood and a better knowledge of it is now recognized as one of the main challenge for vaccine development. In previous studies, we reported that a DNA vaccine containing the signal peptide sequence from the human tissue plasminogen activator (t-PA) fused to the DENV2 NS1 gene (pcTPANS1) induced protection against dengue in mice. In the present work, we aimed to elucidate the contribution of cellular and humoral responses elicited by this vaccine candidate for protective immunity. We observed that pcTPANS1 exerts a robust protection against dengue, inducing considerable levels of anti-NS1 antibodies and T cell responses. Passive immunization with anti-NS1 antibodies conferred partial protection in mice infected with low virus load (4 LD50), which was abrogated with the increase of viral dose (40 LD50). The pcTPANS1 also induced activation of CD4+ and CD8+ T cells. We detected production of IFN-γ and a cytotoxic activity by CD8+ T lymphocytes induced by this vaccine, although its contribution in the protection was not so evident when compared to CD4+ cells. Depletion of CD4+ cells in immunized mice completely abolished protection. Furthermore, transfer experiments revealed that animals receiving CD4+ T cells combined with anti-NS1 antiserum, both obtained from vaccinated mice, survived virus infection with survival rates not significantly different from pcTPANS1-immunized animals. Taken together, results showed that the protective immune response induced by the expression of NS1 antigen mediated by the pcTPANS1 requires a cooperation between CD4+ T cells and the humoral immunity.
Alexander, Jeff; Bilsel, Pamuk; del Guercio, Marie-France; Stewart, Stephani; Marinkovic-Petrovic, Aleksandra; Southwood, Scott; Crimi, Claire; Vang, Lo; Walker, Les; Ishioka, Glenn; Chitnis, Vivek; Sette, Alessandro; Assarsson, Erika; Hannaman, Drew; Botten, Jason; Newman, Mark J
The goal of the present study was to design a vaccine that would provide universal protection against infection of humans with diverse influenza A viruses. Accordingly, protein sequences from influenza A virus strains currently in circulation (H1N1, H3N2), agents of past pandemics (H1N1, H2N2, H3N2) and zoonotic infections of man (H1N1, H5N1, H7N2, H7N3, H7N7, H9N2) were evaluated for the presence of amino acid sequences, motifs, that are predicted to mediate peptide epitope binding with high affinity to the most frequent HLA-DR allelic products. Peptides conserved among diverse influenza strains were then synthesized, evaluated for binding to purified HLA-DR molecules and for their capacity to induce influenza-specific immune recall responses using human donor peripheral blood mononuclear cells (PBMC). Accordingly, 20 epitopes were selected for further investigation based on their conservancy among diverse influenza strains, predicted population coverage in diverse ethnic groups and capacity to recall influenza-specific responses. A DNA plasmid encoding the epitopes was constructed using amino acid spacers between epitopes to promote optimum processing and presentation. Immunogenicity of the DNA vaccine was measured using HLA-DR4 transgenic mice and the TriGrid™ in vivo electroporation device. Vaccination resulted in peptide-specific immune responses, augmented HA-specific antibody responses and protection of HLA-DR4 transgenic mice from lethal PR8 influenza virus challenge. These studies demonstrate the utility of this vaccine format and the contribution of CD4+ T cell responses to protection against influenza infection. PMID:19895924
Pereiro, P; Martinez-Lopez, A; Falco, A; Dios, S; Figueras, A; Coll, J M; Novoa, B; Estepa, A
Turbot (Scophthalmus maximus) is a high-value farmed marine flatfish with growing demand and production levels in Europe susceptible to turbot-specific viral haemorrhagic septicaemia virus (VHSV) strains. To evaluate the possibility of controlling the outbreaks of this infectious disease by means of DNA vaccination, the gpG of a VHSV isolated from farmed turbot (VHSV(860)) was cloned into an expression plasmid containing the human cytomegalovirus (CMV) promoter (pMCV1.4-G(860)). In our experimental conditions, DNA immunised turbots were more than 85% protected against VHSV(860) lethal challenge and showed both VHSV-gpG specific and neutralizing antibodies. To our knowledge this is the first report showing the efficacy of turbot genetic immunisation against a VHSV. Work is in progress to determine the contribution of innate and adaptive immunity to the protective response elicited by the immunization.
Alonso, Marta; Leong, Jo-Ann C
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.
Heppell, J; Davis, H L
The aquaculture industry needs to augment its global production and efficiency to meet the increasing consumer needs for fish and shellfish products. Unfortunately, infectious diseases have been a major impediment to the development and profitability of fish farms. While vaccines offer the most efficient way to control infectious pathogens, current products have only been successful against some diseases. These are mostly bacterial, and there are still several important diseases, mainly of viral and parasitic origin, for which no prophylactic treatment exists. DNA vaccines, compared to traditional antigen vaccines, have several practical and immunological advantages that make them very attractive for the aquaculture industry. The early success of DNA vaccines in animal models was very encouraging, but fish are unique in many aspects, and findings with other classes of vertebrate, namely mammals and birds, do not necessarily apply to aquatic animals. However, more recent studies with reporter genes showed that fish cells efficiently express foreign proteins encoded by eukaryotic expression vectors. A piscine-specific backbone vector might eventually improve immune responses to DNA vaccines, but there is already strong direct evidence for the induction of protective immunity with currently available plasmids. Immune responses to plasmid DNA injected intramuscularly (IM) into fish are characterized by the production of antibodies, which have been shown to be neutralizing in two different viral disease models. There is also indirect evidence suggesting the induction of cell-mediated immunity. Despite this evidence, immune responses to DNA vaccines have only been poorly characterized in fish because of the limited knowledge of the piscine immune system, and the small number of studies on the subject. Apart from optimizing the efficiency of DNA vaccines, other important issues, such as safety and production cost will be determinants for the potential application of this
Comparative assessment of a DNA and protein Leishmania donovani gamma glutamyl cysteine synthetase vaccine to cross-protect against murine cutaneous leishmaniasis caused by L. major or L. mexicana infection.
Campbell, S A; Alawa, J; Doro, B; Henriquez, F L; Roberts, C W; Nok, A; Alawa, C B I; Alsaadi, M; Mullen, A B; Carter, K C
Leishmaniasis is a major health problem and it is estimated that 12 million people are currently infected. A vaccine which could cross-protect people against different Leishmania spp. would facilitate control of this disease as more than one species of Leishmania may be present. In this study the ability of a DNA vaccine, using the full gene sequence for L. donovani gamma glutamyl cysteine synthetase (γGCS) incorporated in the pVAX vector (pVAXγGCS), and a protein vaccine, using the corresponding recombinant L. donovani γGCS protein (LdγGCS), to protect against L. major or L. mexicana infection was evaluated. DNA vaccination gave transient protection against L. major and no protection against L. mexicana despite significantly enhancing specific antibody titres in vaccinated infected mice compared to infected controls. Vaccination with the LdγGCS protected against both species but only if the protein was incorporated into non-ionic surfactant vesicles for L. mexicana. The results of this study indicate that a L. donovani γGCS vaccine could be used to vaccinate against more than one Leishmania species but only if the recombinant protein is used.
Tregoning, John S; Kinnear, Ekaterina
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.
Liu, Feng; Sun, Xiangjie; Fairman, Jeffery; Lewis, David B; Katz, Jacqueline M; Levine, Min; Tumpey, Terrence M; Lu, Xiuhua
Influenza A (H5N1) viruses continue to pose a public health threat. As inactivated H5N1 vaccines are poorly immunogenic, adjuvants are needed to improve the immunogenicity of H5N1 vaccine in humans. Here, we investigated the immunogenicity and cross-protective efficacy in ferrets of a clade 2.2-derived vaccine with addition of JVRS-100, an adjuvant consisting of cationic liposome-DNA complexes (CLDC). After the first vaccination, significantly higher levels of hemagglutination-inhibition (HAI) and neutralizing antibody titers were detected in ferrets immunized with adjuvanted vaccine compared to unadjuvanted vaccine. Following a second dose of adjuvanted vaccine, HAI antibody titers of ≥ 40 were detected against viruses from multiple H5N1 clades. HAI antibodies against newly isolated H5N2 and H5N8 viruses were also augmented by JVRS-100. Ferrets were challenged with a heterologous H5N1 virus. All ferrets that received two doses of adjuvanted vaccine exhibited mild illness, significantly reduced nasal wash virus titers and protection from lethal challenge. In contrast, ferrets that received unadjuvanted vaccine showed greater weight loss, high viral titers and 3 of 6 animals succumbed to the lethal challenge. Our results indicate that the addition of JVRS-100 to H5N1 vaccine enhanced immunogenicity and cross-protection against lethal H5N1 virus disease in ferrets. JVRS-100 warrants further investigation as a potential adjuvant for influenza vaccines.
Wang, Yuan; Liu, Guang-Lu; Li, Dong-Liang; Ling, Fei; Zhu, Bin; Wang, Gao-Xue
To reduce the lethal hemorrhagic disease caused by grass carp reovirus (GCRV) and improve the production of grass carp, efficient and economic prophylactic measure against GCRV is the most pressing desired for the grass carp farming industry. In this work, a novel SWCNTs-pEGFP-vp5 DNA vaccine linked vp5 recombinant in the form of plasmid pEGFP-vp5 and ammonium-functionalized SWCNTs by a chemical modification method was prepared to enhance the efficacy of a vp5 DNA vaccine against GCRV in juvenile grass carp. After intramuscular injection (1, 2.5 and 5 μg) and bath administration (1, 10, and 20 mg/L), the ability of the different immune treatments to induce transgene expression was analyzed. The results showed that higher levels of transcription and expression of vp5 gene could be detected in muscle tissues of grass carp in SWCNTs-pEGFP-vp5 treatment groups compare with naked pEGFP-vp5 treatment groups. Moreover, antibody levels, immune-related genes, and relative percentage survival were significantly enhanced in fish immunized with SWCNTs-pEGFP-vp5 vaccine. In addition, we found that a good immune protective effect was observed in bath immunization group; which at a concentration of 20 mg/L could reach the similar relative percentage survival (approximately 100%) in injection group at a dose of 5 μg. All these results indicated that ammonium-functionalized SWCNTs could provide extensive application prospect to aquatic vaccine and might be used to vaccinate fish by intramuscular injection or bath administration method.
Baliban, Scott M; Michael, Amanda; Shammassian, Berje; Mudakha, Shikata; Khan, Amir S; Cocklin, Simon; Zentner, Isaac; Latimer, Brian P; Bouillaut, Laurent; Hunter, Meredith; Marx, Preston; Sardesai, Niranjan Y; Welles, Seth L; Jacobson, Jeffrey M; Weiner, David B; Kutzler, Michele A
Clostridium difficile-associated disease (CDAD) constitutes a large majority of nosocomial diarrhea cases in industrialized nations and is mediated by the effects of two secreted toxins, toxin A (TcdA) and toxin B (TcdB). Patients who develop strong antitoxin antibody responses can clear C. difficile infection and remain disease free. Key toxin-neutralizing epitopes have been found within the carboxy-terminal receptor binding domains (RBDs) of TcdA and TcdB, which has generated interest in developing the RBD as a viable vaccine target. While numerous platforms have been studied, very little data describes the potential of DNA vaccination against CDAD. Therefore, we created highly optimized plasmids encoding the RBDs from TcdA and TcdB in which any putative N-linked glycosylation sites were altered. Mice and nonhuman primates were immunized intramuscularly, followed by in vivo electroporation, and in these animal models, vaccination induced significant levels of both anti-RBD antibodies (blood and stool) and RBD-specific antibody-secreting cells. Further characterization revealed that sera from immunized mice and nonhuman primates could detect RBD protein from transfected cells, as well as neutralize purified toxins in an in vitro cytotoxicity assay. Mice that were immunized with plasmids or given nonhuman-primate sera were protected from a lethal challenge with purified TcdA and/or TcdB. Moreover, immunized mice were significantly protected when challenged with C. difficile spores from homologous (VPI 10463) and heterologous, epidemic (UK1) strains. These data demonstrate the robust immunogenicity and efficacy of a TcdA/B RBD-based DNA vaccine in preclinical models of acute toxin-associated and intragastric, spore-induced colonic disease.
Tretyakova, Irina; Lukashevich, Igor S; Glass, Pamela; Wang, Eryu; Weaver, Scott; Pushko, Peter
DNA vaccines combine remarkable genetic and chemical stability with proven safety and efficacy in animal models, while remaining less immunogenic in humans. In contrast, live-attenuated vaccines have the advantage of inducing rapid, robust, long-term immunity after a single-dose vaccination. Here we describe novel iDNA vaccine technology that is based on an infectious DNA platform and combines advantages of DNA and live attenuated vaccines. We applied this technology for vaccination against infection with Venezuelan equine encephalitis virus (VEEV), an alphavirus from the Togaviridae family. The iDNA vaccine is based on transcription of the full-length genomic RNA of the TC-83 live-attenuated virus from plasmid DNA in vivo. The in vivo-generated viral RNA initiates limited replication of the vaccine virus, which in turn leads to efficient immunization. This technology allows the plasmid DNA to launch a live-attenuated vaccine in vitro or in vivo. Less than 10 ng of pTC83 iDNA encoding the full-length genomic RNA of the TC-83 vaccine strain initiated replication of the vaccine virus in vitro. In order to evaluate this approach in vivo, BALB/c mice were vaccinated with a single dose of pTC83 iDNA. After vaccination, all mice seroconverted with no adverse reactions. Four weeks after immunization, animals were challenged with the lethal epidemic strain of VEEV. All iDNA-vaccinated mice were protected from fatal disease, while all unvaccinated controls succumbed to infection and died. To our knowledge, this is the first example of launching a clinical live-attenuated vaccine from recombinant plasmid DNA in vivo.
Meng, Min; He, Shenyi; Zhao, Guanghui; Bai, Yang; Zhou, Huaiyu; Cong, Hua; Lu, Gang; Zhao, Qunli; Zhu, Xing-Quan
Toxoplasmosis, caused by an obligate intracellular protozoan parasite Toxoplasma gondii, has been a serious clinical and veterinary problem. Effective DNA vaccines against T. gondii can prevent and control the spread of toxoplasmosis, which is important for both human health and the farming industry. The T. gondii 14-3-3 protein has been proved to be antigenic and immunogenic and was a potential vaccine candidate against toxoplasmosis. In this study, we evaluated the immune responses induced by recombinant plasmids encoding T. gondii surface antigen 1 (SAG1) and 14-3-3 protein by immunizing BALB/c mice intramuscularly. In the present study, BALB/c mice were randomly divided into five groups, including three experimental groups (pSAG1, p14-3-3 and pSAG1/14-3-3) and two control groups (PBS and pBudCE4.1), and were immunized intramuscularly three times. The levels of IgG antibodies and cytokine production in mouse sera were determined by enzyme-linked immunosorbent assays (ELISA). Two weeks after the last immunization, all mice were challenged intraperitoneally (i.p.) with 1×10(4) tachyzoites of T. gondii and the survival time of mice was observed and recorded every day. Mice vaccinated with pSAG1, p14-3-3 or pSAG1/14-3-3 developed high levels of IgG2a and gamma interferon (IFN-γ) and low levels of interleukin-4 (IL-4) and interleukin-10 (IL-10) compared to control groups (PBS or pBudCE4.1), which suggested a modulated Th1 type immune response (P<0.05). After intraperitoneal challenge with 1×10(4) tachyzoites of T. gondii (RH strain), the survival time of mice in experimental groups was longer than control groups (P<0.05). Mouse immunized with pSAG1/14-3-3 induced a higher level of IgG antibody response and significantly prolonged the survival time when compared with pSAG1 or p14-3-3 (P<0.05). The study suggested that T. gondii 14-3-3 protein can induce effective immune responses in BALB/c mice and was a novel DNA vaccine candidate against toxoplasmosis, and the
De Filette, Marina; Soehle, Silke; Ulbert, Sebastian; Richner, Justin; Diamond, Michael S.; Sinigaglia, Alessandro; Barzon, Luisa; Roels, Stefan; Lisziewicz, Julianna; Lorincz, Orsolya; Sanders, Niek N.
West Nile virus (WNV) is a mosquito-borne flavivirus that is endemic in Africa, the Middle East, Europe and the United States. There is currently no antiviral treatment or human vaccine available to treat or prevent WNV infection. DNA plasmid-based vaccines represent a new approach for controlling infectious diseases. In rodents, DNA vaccines have been shown to induce B cell and cytotoxic T cell responses and protect against a wide range of infections. In this study, we formulated a plasmid DNA vector expressing the ectodomain of the E-protein of WNV into nanoparticles by using linear polyethyleneimine (lPEI) covalently bound to mannose and examined the potential of this vaccine to protect against lethal WNV infection in mice. Mice were immunized twice (prime – boost regime) with the WNV DNA vaccine formulated with lPEI-mannose using different administration routes (intramuscular, intradermal and topical). In parallel a heterologous boost with purified recombinant WNV envelope (E) protein was evaluated. While no significant E-protein specific humoral response was generated after DNA immunization, protein boosting of DNA-primed mice resulted in a marked increase in total neutralizing antibody titer. In addition, E-specific IL-4 T-cell immune responses were detected by ELISPOT after protein boost and CD8+ specific IFN-γ expression was observed by flow cytometry. Challenge experiments using the heterologous immunization regime revealed protective immunity to homologous and virulent WNV infection. PMID:24503579
cetacean antibody titers to E . rhusiopathiae ; and to test the DNA from the recombinant protein in vitro in cetacean specific cell lines for its ability...to transfect the cells. APPROACH: The gene for the protective antigen of E . rhusiopathiae will be inserted into a eukaryotic vector both for the...cetacean serum samples to E . rhusiopathiae . ACCOMPLISHMENTS: The identity of the U.S. Navy Space and Naval Warfare System Center (SPAWARSYSTENS) isolate of
Bicho, Diana; Queiroz, João António; Tomaz, Cândida Teixeira
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.
Ye, Ting; Yue, Yan; Fan, Xiangmei; Dong, Chunsheng; Xu, Wei; Xiong, Sidong
Efficient delivery of antigen to mucosal associated lymphoid tissue is a first and critical step for successful induction of mucosal immunity by vaccines. Considering its potential transcytotic capability, M cell has become a more and more attractive target for mucosal vaccines. In this research, we designed an M cell-targeting strategy by which mucosal delivery system chitosan (CS) was endowed with M cell-targeting ability via conjugating with a CPE30 peptide, C terminal 30 amino acids of clostridium perfringens enterotoxin (CPE), and then evaluated its immune-enhancing ability in the context of coxsackievirus B3 (CVB3)-specific mucosal vaccine consisting of CS and a plasmid encoding CVB3 predominant antigen VP1. It had shown that similar to CS-pVP1, M cell-targeting CPE30-CS-pVP1 vaccine appeared a uniform spherical shape with about 300 nm diameter and +22 mV zeta potential, and could efficiently protect DNA from DNase I digestion. Mice were orally immunized with 4 doses of CPE30-CS-pVP1 containing 50 μg pVP1 at 2-week intervals and challenged with CVB3 4 weeks after the last immunization. Compared with CS-pVP1 vaccine, CPE30-CS-pVP1 vaccine had no obvious impact on CVB3-specific serum IgG level and splenic T cell immune responses, but significantly increased specific fecal SIgA level and augmented mucosal T cell immune responses. Consequently, much milder myocarditis and lower viral load were witnessed in CPE30-CS-pVP1 immunized group. The enhanced immunogenicity and immunoprotection were associated with the M cell-targeting ability of CPE30-CS-pVP1 which improved its mucosal uptake and transcytosis. Our findings indicated that CPE30-CS-pVP1 may represent a novel prophylactic vaccine against CVB3-induced myocarditis, and this M cell-targeting strategy indeed could be applied as a promising and universal platform for mucosal vaccine development.
Lever, Melissa; Silveira, Eduardo L; Nakaya, Helder I
DNA vaccination represents a new milestone in our technological efforts to avoid infectious diseases. Although this method of vaccination has had success in providing protection in animals, these vaccines suffer from low immunogenicity in humans. Questions remain over the molecular mechanism of DNA vaccination, the best ways in which to safely increase vaccine reactogenecity, and what biomarkers can be used as correlates of protection. Systems vaccinology, which utilizes modern experimental and computational approaches to provide an integrated view of the vaccination process, offers the potential to answer these questions. In this review we discuss the current tools utilized in systems vaccinology, the ways in which they have and can be applied to DNA vaccinology, and challenges faced in the field.
Hussein, H A; Ahmed, B M; Aly, S M; El-Deeb, A H; El-Sanousi, A A; Rohaim, M A; Arafa, A A; Gadalla, M R
In this study, a recombinant DNA plasmid was constructed, encoding for HA1 of a selected Egyptian H5N1 virus (isolated during the 2012 outbreaks). In the immunization and challenge experiments, SPF chickens received 1 or 2 doses of H5-DNA plasmid prime, and boosted with the inactivated H5N2 vaccine. Haemagglutination inhibition (HI) titers, protection levels, and the magnitude of virus shedding were compared to that of the chickens that received either DNA plasmid or inactivated H5N2 vaccine alone. H5N1 virus A/chicken/Egypt/128s/2012 (H5N1) highly pathogenic avian influenza (HPAI) clade 2.2.1/C was used for the challenge. Chickens immunized with 1 or 2 doses of H5-DNA vaccine failed to overcome the challenge with 0% and 10% protection, respectively. Quantitative real-time reverse transcription-PCR revealed virus shedding of 2.2 x 104 PCR copies/ml 3 days post challenge (dpc) in the only surviving bird from the group that received 2 doses of plasmid. However, chickens immunized with 1 or 2 doses of H5-DNA plasmid as prime and inactivated H5N2 vaccine as booster, showed 80% protection after challenge, with a viral shedding of 1.2 x 104 PCR copies/ml (1 dose) and 1.6 x 104 PCR copies/ml (2 doses) 3 dpc. The surviving birds in both groups did not shed the virus at 5 and 7 dpc. In H5N2-vaccinated chickens, protection levels were 70% with relatively high virus shedding (1.8 x 104 PCR copies/ml) 3 dpc. HI titers were protective to the surviving chickens. This study reports the efficacy of H5-DNA plasmid to augment reduction in viral shedding and to provide better protection when applied in a prime-boost program with the inactivated AI vaccine.
... https://medlineplus.gov/news/fullstory_163384.html New Zika Vaccine Candidate Provides Powerful Protection Made without live ... HealthDay News) -- A single dose of an experimental Zika vaccine protected mice and monkeys from the virus, ...
DNA vaccines targeting heavy chain C-terminal fragments of Clostridium botulinum neurotoxin serotypes A, B, and E induce potent humoral and cellular immunity and provide protection from lethal toxin challenge.
Scott, Veronica L; Villarreal, Daniel O; Hutnick, Natalie A; Walters, Jewell N; Ragwan, Edwin; Bdeir, Khalil; Yan, Jian; Sardesai, Niranjan Y; Finnefrock, Adam C; Casimiro, Danilo R; Weiner, David B
Botulinum neurotoxins (BoNTs) are deadly, toxic proteins produced by the bacterium Clostridium botulinum that can cause significant diseases in humans. The use of the toxic substances as potential bioweapons has raised concerns by the Centers for Disease Control and Prevention and the United States Military. Currently, there is no licensed vaccine to prevent botulinum intoxication. Here we present an immunogenicity study to evaluate the efficacy of novel monovalent vaccines and a trivalent cocktail DNA vaccine targeting the heavy chain C-terminal fragments of Clostridium botulinum neurotoxin serotypes A, B, and E. These synthetic DNA vaccines induced robust humoral and polyfunctional CD4(+) T-cell responses which fully protected animals against lethal challenge after just 2 immunizations. In addition, naïve animals administered immunized sera mixed with the lethal neurotoxin were 100% protected against intoxication. The data demonstrate the protective efficacy induced by a combinative synthetic DNA vaccine approach. This study has importance for the development of vaccines that provide protective immunity against C. botulinum neurotoxins and other toxins.
Borhani, Kiandokht; Ajorloo, Mehdi; Bamdad, Taravat; Mozhgani, Sayed Hamid Reza; Ghaderi, Mostafa; Gholami, Ali Reza
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.
Gao, Shanshan; Zhao, Na; Amer, Said; Qian, Mingming; Lv, Mengxi; Zhao, Yuliang; Su, Xin; Cao, Jieying; He, Hongxuan; Zhao, Baohua
In the present study, poly (lactic-co-glycolic) acid (PLGA) was used as a carrier for a divalent fusion DNA vaccine encoding the Aeromonas veronii outer membrane protein A (ompA) and Aeromonas hydrophila hemolysins (hly) protein. The recombinant pET-28a-ompA-hly was constructed by inserting the ompA gene and hly gene into a pET-28a expression vector. Loading of ompA-hly antigen module on PLGA microspheres were accomplished by water-in-oil-in-water (W/O/W) encapsulation. The molecular weight and specificity of pET-28a-ompA-hly were detected by dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and western blotting. The microspheres showed an average particle size of 100-150 μm and a loading efficiency (LE) of 68.8%. Mice received ompA-hly antigen-loaded PLGA microspheres by intraperitoneal or intragastric administration mounted strong and sustained IgG response, which was significantly higher (p<0.05) than those achieved by pET-28a-ompA-hly antigen alone. OmpA-hly antigen-loaded PLGA microsphere vaccine uniquely conferred a long lasting (30 days) sterile immunity against challenge infection. Results indicated that ompA-hly antigen-loaded PLGA microsphere vaccine is a qualified candidate vector system for sterile protective immunity against A. hydrophila and A. veronii infections.
Dong, Libo; Liu, Feng; Fairman, Jeffery; Hong, David K; Lewis, David B; Monath, Thomas; Warner, John F; Belser, Jessica A; Patel, Jenish; Hancock, Kathy; Katz, Jacqueline M; Lu, Xiuhua
The development of pre-pandemic influenza A H5N1 vaccines that confer both antigen-sparing and cross-clade protection are a high priority given the limited worldwide capacity for influenza vaccine production, and the antigenic and genetic heterogeneity of circulating H5N1 viruses. The inclusion of potent adjuvants in vaccine formulations may achieve both of these aims. Here we show that the addition of JVRS-100, an adjuvant consisting of cationic liposome-DNA complexes (CLDC) to a clade 1-derived H5N1 split vaccine induced significantly higher virus-specific antibody than unadjuvanted formulations, with a >30-fold dose-sparing effect and induction of increased antigen-specific CD4(+) T-cell responses in mice. All mice that received one dose of adjuvanted vaccine and subsequent H5N1 viral challenges exhibited mild illness, lower lung viral titers, undetectable spleen and brain viral titers, and 100% survival after either homologous clade 1 or heterologous clade 2 H5N1 viral challenges, whereas unadjuvanted vaccine recipients showed significantly increased weight loss, viral titers, and mortality. The protective immunity induced by JVRS-100 adjuvanted H5N1 vaccine was shown to last for over one year without significant waning. Thus, JVRS-100 adjuvanted H5N1 vaccine elicited enhanced humoral and T-cell responses, dose-sparing, and cross-clade protection in mice. CLDC holds promise as an adjuvant for human pre-pandemic inactivated H5N1 vaccines.
Iborra, Salvador; Soto, Manuel; Carrión, Javier; Nieto, Ana; Fernández, Edgar; Alonso, Carlos; Requena, Jose M
In this study, we examined the immunogenic properties of the Leishmania infantum acidic ribosomal protein P0 (LiP0) in the BALB/c mouse model. The humoral and cellular responses induced by the administration of the LiP0 antigen, either as soluble recombinant LiP0 (rLiP0) or as a plasmid DNA formulation (pcDNA3-LiP0), were determined. Also, the immunological response associated with a prime-boost strategy, consisting of immunization with pcDNA3-LiP0 followed by a boost with rLiP0, was assayed. Immunization with rLiP0 induced a predominant Th2-like humoral response, but no anti-LiP0 antibodies were induced after immunization with pcDNA3-LiP0, whereas a strong humoral response consisting of a mixed immunoglobulin G2a (IgG2a)-IgG1 isotype profile was induced in mice immunized with the prime-boost regime. For all three immunization protocols, rLiP0-stimulated production of gamma interferon (IFN-gamma) in both splenocytes and lymph node cells from immunized mice was observed. However, it was only when mice were immunized with pcDNA3-LiP0 that noticeable protection against L. major infection was achieved, as determined by both lesion development and parasite burden. Immunization of mice with LiP0-DNA primes both CD4(+) and CD8(+) T cells, which, with the L. major challenge, were boosted to produce significant levels of IL-12-dependent, antigen-specific IFN-gamma. Taken together, these data indicate that genetic vaccination with LiP0 induces protective immunological effector mechanisms, yet the immunological response elicited by LiP0 is not sufficient to keep the infection from progressing.
Li, Chang; Shen, Zhenwei; Li, Xiao; Bai, Jieying; Zeng, Lin; Tian, Mingyao; Song, Ying Jin; Ye, Ming; Du, Shouwen; Ren, Dayong; Liu, Cunxia; Zhu, Na; Sun, Dandan; Li, Yi; Jin, Ningyi
Developing an effective vaccine against HIV infection remains an urgent goal. We used a DNA prime/fowlpox virus boost regimen to immunize Chinese rhesus macaques. The animals were challenged intramuscularly with pathogenic molecularly cloned SHIV-KB9. Immunogenicity and protective efficacy of vaccines were investigated by measuring IFN-γ levels, monitoring HIV-specific binding antibodies, examining viral load, and analyzing CD4/CD8 ratio. Results show that, upon challenge, the vaccine group can induce a strong immune response in the body, represented by increased expression of IFN-γ, slow and steady elevated antibody production, reduced peak value of acute viral load, and increase in the average CD4/CD8 ratio. The current research suggests that rapid reaction speed, appropriate response strength, and long-lasting immune response time may be key protection factors for AIDS vaccine. The present study contributes significantly to AIDS vaccine and preclinical research.
Donnelly, J J; Martinez, D; Jansen, K U; Ellis, R W; Montgomery, D L; Liu, M A
Genital infections with human papillomavirus (HPV) are increasingly recognized as a significant source of human disease; HPV is now implicated in up to 90% of cervical carcinomas. Neutralizing antibodies against papillomaviruses recognize conformational epitopes formed when viral capsid proteins assemble into virions or virus-like particles. Immunization with plasmid DNA encoding the major viral capsid protein L1 was studied as a means of inducing neutralizing antibodies and protection against virus challenge. In a cottontail rabbit papillomavirus (CRPV) model, immunization with plasmid DNA encoding L1 elicited conformationally specific neutralizing antibodies and provided immunity against papilloma formation upon challenge with CRPV. Immunization with DNA encoding the capsid protein may provide a means of protecting humans against HPV and would simplify the production of multivalent vaccines by combining plasmids that encode the viral capsid proteins of different strains. This may be of importance given the multiplicity of HPV types capable of causing disease.
Silveira, Marcelle Moura; Oliveira, Thaís Larré; Schuch, Rodrigo Andrade; McBride, Alan John Alexander; Dellagostin, Odir Antônio; Hartwig, Daiane Drawanz
Leptospirosis is an infectious disease caused by pathogenic Leptospira species. The vaccines that are currently available for leptospirosis are composed of whole-cell preparations and suffer from limitations such as low efficacy, multiple side-effects, poor immunological memory and lack of cross-protection against different serovars of Leptospira spp. In light of the global prevalence of this disease, the development of a more effective vaccine against leptospirosis is of paramount importance. Genetic immunization is a promising alternative to conventional vaccine development. In the last 25years, several novel strategies have been developed for increasing the efficacy of DNA vaccines. Examples of such strategies include the introduction of novel plasmid vectors, adjuvants, alternate delivery routes, and prime-boost regimens. Herein we discuss the latest and most promising advances that have been made in developing DNA vaccines against leptospirosis. We also deliberate over the future directions that must be undertaken in order to improve results in this field. Copyright © 2017 Elsevier Ltd. All rights reserved.
Riquelme-Neira, Roberto; Retamal-Díaz, Angello; Acuña, Francisca; Riquelme, Pablo; Rivera, Alejandra; Sáez, Darwin; Oñate, Angel
The immunogenicity of a DNA vaccine containing an open reading frame (ORF) of genomic island 3 (GI-3), specific for Brucella abortus and Brucella melitensis, has been examined. Intramuscular injection of plasmid DNA carrying the open reading frame with homology to an ABC-type transporter (pV278a) into BALB/c mice elicited both humoral and cellular immune responses. Mice injected with pV278a had a dominant immunoglobulin G2a (IgG2a) response. This DNA vaccine elicited a T-cell-proliferative response and induced significant levels of interferon gamma (INF-γ) upon restimulation with recombinant 278a protein. Upon stimulation with an appropriate recombinant protein or crude Brucella protein, the vaccine did not induce IL-4, suggesting a typical T-helper (TH1) response. Furthermore, the vaccine induced protection in BALB/c mice when challenged with the virulent strain Brucella abortus 2308. Taken together, these data suggest that DNA vaccination offers an improved delivery of the homologous of an ABC-type transporter antigen, and provides the first evidence of a protective effect of this antigen in the construction of vaccines against B. abortus.
Zhang, Han; El Zowalaty, Mohamed E
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.
Li, Lei; Petrovsky, Nikolai
Summary In the two decades since their initial discovery, DNA vaccines technologies have come a long way. Unfortunately, when applied to human subjects inadequate immunogenicity is still the biggest challenge for practical DNA vaccine use. Many different strategies have been tested in preclinical models to address this problem, including novel plasmid vectors and codon optimization to enhance antigen expression, new gene transfection systems or electroporation to increase delivery efficiency, protein or live virus vector boosting regimens to maximise immune stimulation, and formulation of DNA vaccines with traditional or molecular adjuvants. Better understanding of the mechanisms of action of DNA vaccines has also enabled better use of the intrinsic host response to DNA to improve vaccine immunogenicity. This review summarizes recent advances in DNA vaccine technologies and related intracellular events and how these might impact on future directions of DNA vaccine development. PMID:26707950
Li, Lei; Petrovsky, Nikolai
In the two decades since their initial discovery, DNA vaccines technologies have come a long way. Unfortunately, when applied to human subjects inadequate immunogenicity is still the biggest challenge for practical DNA vaccine use. Many different strategies have been tested in preclinical models to address this problem, including novel plasmid vectors and codon optimization to enhance antigen expression, new gene transfection systems or electroporation to increase delivery efficiency, protein or live virus vector boosting regimens to maximise immune stimulation, and formulation of DNA vaccines with traditional or molecular adjuvants. Better understanding of the mechanisms of action of DNA vaccines has also enabled better use of the intrinsic host response to DNA to improve vaccine immunogenicity. This review summarizes recent advances in DNA vaccine technologies and related intracellular events and how these might impact on future directions of DNA vaccine development.
Kubicek-Sutherland, Jessica Z; Heithoff, Douglas M; Ersoy, Selvi C; Shimp, William R; Mahan, Michael J
Yersinia pseudotuberculosis is a foodborne pathogen that can cause serious human illness. Although the source and route of transmission often remain obscure, livestock have been implicated in some cases. The diversity of yersiniae present on farms and their widespread distribution in animal and environmental reservoirs necessitates the use of broad prophylactic strategies that are efficacious against many serotypes simultaneously. Herein, immunization of mice with a modified, live attenuated Y. pseudotuberculosis vaccine that overproduces the DNA adenine methylase (Dam(OP)) conferred robust protection against virulent challenge (150-fold LD50) with homologous and heterologous serotypes that have been associated with human disease (O:1, O:1a, O:3). Further, the dam gene was shown to be essential for cell viability in all (7 of 7) Y. pseudotuberculosis strains tested. Direct selection for the inheritance of dam mutant alleles in Y. pseudotuberculosis resulted in dam strain variants that contained compensatory (second-site suppressor) mutations in genes encoding methyl-directed mismatch repair proteins (mutHLS) that are involved in suppression of the non-viable cell phenotype in all (19/19) strains tested. Such dam mutH variants exhibited a significant increase in virulence and spontaneous mutation frequency relative to that of a Dam(OP) vaccine strain. These studies indicate that Y. pseudotuberculosis Dam(OP) strains conferred potent cross-protective efficacy as well as decreased virulence and spontaneous mutation frequency relative to those that lack Dam, which have compensatory mutations in mutHLS loci. These data suggest that development of yersiniae livestock vaccines based on Dam overproduction is a viable mitigation strategy to reduce these potential foodborne contaminants.
Pushko, Peter; Lukashevich, Igor S; Weaver, Scott C; Tretyakova, Irina
A novel vaccine platform uses DNA immunization to launch live-attenuated virus vaccines in vivo. This technology has been applied for vaccine development against positive-strand RNA viruses with global public health impact including alphaviruses and flaviviruses. The DNA-launched vaccine represents the recombinant plasmid that encodes the full-length genomic RNA of live-attenuated virus downstream from a eukaryotic promoter. When administered in vivo, the genomic RNA of live-attenuated virus is transcribed. The RNA initiates limited replication of a genetically defined, live-attenuated vaccine virus in the tissues of the vaccine recipient, thereby inducing a protective immune response. This platform combines the strengths of reverse genetics, DNA immunization and the advantages of live-attenuated vaccines, resulting in a reduced chance of genetic reversions, increased safety, and improved immunization. With this vaccine technology, the field of DNA vaccines is expanded from those that express subunit antigens to include a novel type of DNA vaccines that launch live-attenuated viruses.
Pushko, Peter; Lukashevich, Igor S.; Weaver, Scott C.; Tretyakova, Irina
Summary A novel vaccine platform uses DNA immunization to launch live-attenuated virus vaccines in vivo. This technology has been applied for vaccine development against positive-strand RNA viruses with global public health impact including alphaviruses and flaviviruses. The DNA-launched vaccine represents the recombinant plasmid that encodes the full-length genomic RNA of live-attenuated virus downstream from a eukaryotic promoter. When administered in vivo, the genomic RNA of live-attenuated virus is transcribed. The RNA initiates limited replication of a genetically defined, live-attenuated vaccine virus in the tissues of the vaccine recipient, thereby inducing a protective immune response. This platform combines the strengths of reverse genetics, DNA immunization and the advantages of live-attenuated vaccines, resulting in a reduced chance of genetic reversions, increased safety, and improved immunization. With this vaccine technology, the field of DNA vaccines is expanded from those that express subunit antigens to include a novel type of DNA vaccines that launch live-attenuated viruses. PMID:27055100
Lu, Hui; Xu, Xiao-Feng; Gao, Na; Fan, Dong-Ying; Wang, Juan; An, Jing
Public health is still seriously threatened by dengue virus (DENV) and no vaccine against DENV is yet available for clinical use till now. In this study, DNA vaccine candidates encoding DENV serotype 2 (DENV-2) prM/E (premembrane and envelope proteins) and NS1 (non-structural 1 protein) with or without a gene adjuvant, granulocyte-macrophage colony-stimulating factor (GM-CSF), were evaluated in the aspects of immunity and protective efficacy in mice. We constructed three plasmids, pCAG-prM/E (which only expressed DENV2 prM/E), pCAG-prM/E/NS1 (which only expressed DENV2 prM/E/NS1) and pCAG-DG (which co-expressed DENV2 prM/E/NS1 and GM-CSF). The expressions of the recombined plasmids were analyzed by immuno-staining in Vero cells. Antibody responses and neutralization activity of the sera from the mice were assayed by ELISA and plaque reduction neutralization test after immunization with the plasmids. Immunized BALB/c mice were intracerebrally challenged with DENV2 to evaluate protective efficacy of the plasmids. The recombinant plasmids could be efficiently expressed in Vero cells and induced different levels of specific anti-DENV2 immune responses. The immunized mice were partially protected. The highest survival rate was observed in the pCAG-DG group although the anti-DENV2 titer and neutralization antibody titer were not the highest among the three groups. Our data suggested that pCAG-DG offered better protection against DENV2 infection.
Herrada, Andrés A.; Rojas-Colonelli, Nicole; González-Figueroa, Paula; Roco, Jonathan; Oyarce, César; Ligtenberg, Maarten A.; Lladser, Alvaro
DNA vaccines have emerged as an attractive strategy to promote protective cellular and humoral immunity against the encoded antigen. DNA vaccines are easy to generate, inexpensive to produce and purify at large-scale, highly stable and safe. In addition, plasmids used for DNA vaccines act as powerful “danger signals” by stimulating several DNA-sensing innate immune receptors that promote the induction of protective adaptive immunity. The induction of tumor-specific immune responses represents a major challenge for DNA vaccines because most of tumor-associated antigens are normal non-mutated self-antigens. As a consequence, induction of potentially self-reactive T cell responses against such poorly immunogenic antigens is controlled by mechanisms of central and peripheral tolerance as well as tumor-induced immunosuppression. Although several DNA vaccines against cancer have reached clinical testing, disappointing results have been observed. Therefore, the development of new adjuvants that strongly stimulate the induction of antitumor T cell immunity and counteract immune-suppressive regulation is an attractive approach to enhance the potency of DNA vaccines and overcome tumor-associated tolerance. Understanding the DNA-sensing signaling pathways of innate immunity that mediate the induction of T cell responses elicited by DNA vaccines represents a unique opportunity to develop novel adjuvants that enhance vaccine potency. The advance of DNA adjuvants needs to be complemented with the development of potent delivery systems, in order to step toward successful clinical application. Here, we briefly discuss recent evidence showing how to harness DNA-induced immune response to improve the potency of cancer vaccines and counteract tumor-associated tolerance. PMID:23111166
Kotla, Sivareddy; Sanghratna Vishanath, Bahire; H J, Dechamma; K, Ganesh; V V S, Suryanarayana; Reddy, G R
Foot-and-mouth disease (FMD) is a highly contagious disease of cloven-hoofed animals causing considerable economic loss in the affected countries. Presently used tissue culture inactivated vaccine protects the vaccinated animals for a short duration. DNA vaccines along with appropriate adjutants is one of the approach for the development of alternative vaccine. In the present study, we constructed P1-2A-3CpCDNA (containing P1-2A-3C coding sequences of FMDV Asia-1 Ind 63/72) and bovine IL-18 pCDNA plasmids and evaluated in cattle. Four groups of calves each group containing six calves were vaccinated with 200μg of plasmid DNA vaccine P1-2A-3CpCDNA, P1-2A-3CpCDNA+ bIL-18pCDNA and inactivated vaccine respectively where as fourth group was unvaccinated. P1-2A-3CpCDNA+bIL-18pCDNA vaccinated animals have shown higher levels of neutralizing antibodies and specific T-cell proliferation responses. Higher levels of CD4(+) and CD8(+) cells were observed in these animals. Similarly, IL-18 adjuvanted group has shown increased Th1 and Th2 cytokine responses. All the vaccinated animals were challenged with cattle adapted FMD homologous Asia1 virus two weeks after the booster dose. IL18 co administered DNA vaccine construct has protected four out of six animals challenged with homologous virus. Copyright © 2016 Elsevier B.V. All rights reserved.
Carvalho, Joana A; Rodgers, Jean; Atouguia, Jorge; Prazeres, Duarte M F; Monteiro, Gabriel A
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.
Tomusange, Khamis; Wijesundara, Danushka; Gummow, Jason; Wesselingh, Steve; Suhrbier, Andreas; Gowans, Eric J.; Grubor-Bauk, Branka
Mucosal immunity is deemed crucial to control sexual transmission of human immunodeficiency virus (HIV). Herein we report the efficacy of a mucosal HIV vaccine strategy comprising intranasal (IN) vaccination with a cocktail of live recombinant human rhinoviruses (HRVs) encoding overlapping fragments of HIV Gag and full length Tat (rHRV-Gag/Tat) followed by intradermal (ID) vaccination with DNA vaccines encoding HIV Gag and Tat (pVAX-Gag-Tat). This heterologous prime-boost strategy will be referred to hereafter as rHRV-DNA. As a control, IN vaccination with wild type (wt)-HRV-A1 followed by a single ID dose of pVAX (wt-HRV-A1/pVAX vaccination) was included. rHRV-DNA vaccination elicited superior multi-functional CD8+T cell responses in lymphocytes harvested from mesenteric lymph nodes and spleens, and higher titres of Tat-specific antibodies in blood and vaginal lavages, and reduced the viral load more effectively after challenge with EcoHIV, a murine HIV challenge model, in peritoneal macrophages, splenocytes and blood compared compared with wt-HRV-A1/pVAX vaccination or administration of 3 ID doses of pVAX-Gag-Tat (3X pVAX-Gag-Tat vaccination). These data provide the first evidence that a rHRV-DNA vaccination regimen can induce HIV-specific immune responses in the gut, vaginal mucosa and systemically, and supports further testing of this regimen in the development of an effective mucosally-targeted HIV-1 vaccine. PMID:27853256
Greenland, John R; Letvin, Norman L
Plasmid DNA vaccines are a promising modality for immunization against a variety of human pathogens. Immunization via multiple routes with plasmid DNA can elicit potent cellular immune responses, and these immunogens can be administered repeatedly without inducing anti-vector immunity. Nonetheless, the immunogenicity of plasmid DNA vaccines has been limited by problems associated with delivery. A number of adjuvants have been designed to improve plasmid DNA immunogenicity, either by directly stimulating the immune system or by enhancing plasmid DNA expression. Chemical adjuvants for enhancing plasmid DNA expression include liposomes, polymers, and microparticles, all of which have shown promise for enhancing the expression and immunogenicity of plasmid DNA vaccines in animal models. Micro- and nanoparticles have not been shown to enhance immune responses to plasmid DNA vaccines. However, formulation of plasmid DNA with some non-particulate polymeric adjuvants has led to a statistically significant enhancement of immune responses. Further development of these technologies will significantly improve the utility of plasmid DNA vaccination.
Callendret, Benoît; Lorin, Valérie; Charneau, Pierre; Marianneau, Philippe; Contamin, Hugues; Betton, Jean-Michel; van der Werf, Sylvie; Escriou, Nicolas
The SARS-CoV spike glycoprotein (S) is the main target of the protective immune response in humans and animal models of SARS. Here, we demonstrated that efficient expression of S from the wild-type spike gene in cultured cells required the use of improved plasmid vectors containing donor and acceptor splice sites, as well as heterologous viral RNA export elements, such as the CTE of Mazon-Pfizer monkey virus or the PRE of Woodchuck hepatitis virus (WPRE). The presence of both splice sites and WPRE markedly improved the immunogenicity of S-based DNA vaccines against SARS. Upon immunization of mice with low doses (2 microg) of naked DNA, only intron and WPRE-containing vectors could induce neutralizing anti-S antibodies and provide protection against challenge with SARS-CoV. Our observations are likely to be useful for the construction of plasmid and viral vectors designed for optimal expression of intronless genes derived from cytoplasmic RNA viruses.
Chen, Jiansen; Li, Neng; She, Feifei
Helicobacter pylori (H. pylori) infection is associated with incidents of gastrointestinal diseases in half of the human population. However, management of its infection remains a challenge. Hence, it is necessary to develop an efficient vaccine to fight against this pathogen. In the present study, a novel vaccine based on the production of attenuated Salmonella typhimurium bacterial ghost (SL7207-BG), delivering H. pylori outer inflammatory protein gene (oipA) encoded DNA vaccine was developed, and the efficiency was evaluated in C57BL/6 mice. Significant higher levels of IgG2a/IgG1 antibodies and IFN-γ/IL-4 cytokines were detected after mice were oral administered with oipA DNA vaccine loaded SL7207-BG, indicating that a mixed Th1/Th2 immune response was elicited. When challenged with infective doses H. pylori strain SS1, the ghost based vaccine was capable of reducing bacterium colonization in the vaccinated mice. In addition, codon-optimized oipA plasmid loaded SL7207-BG significantly eliminates H. pylori colonization density in mice model. Thus, it has been demonstrated that this novel bacterial ghost based DNA vaccine could be used as a promising vaccine candidate for the control of H. pylori infection.
Protection against dengue virus by non-replicating and live attenuated vaccines used together in a prime boost vaccination strategy Monika Simmons a...Dengue DNA Punfied inacdvared virus Uvc artenuatcd virus Jlnmc boost A new vaccination strategy for dengue virus (DENV) was eval uated in rhesus...region (TDNA) then boosting 2 months l,ltcr with a tetravalent live aucnuated virus (TLAV) vaccine . Both vaccine combinations elicited virus
Fuller, Deborah Heydenburg; Rajakumar, Premeela; Che, Jenny W.; Narendran, Amithi; Nyaundi, Julia; Michael, Heather; Yager, Eric J.; Stagnar, Cristy; Wahlberg, Brendon; Taber, Rachel; Haynes, Joel R.; Cook, Fiona C.; Ertl, Peter; Tite, John; Amedee, Angela M.; Murphey-Corb, Michael
Immunotherapies that induce durable immune control of chronic HIV infection may eliminate the need for life-long dependence on drugs. We investigated a DNA vaccine formulated with a novel genetic adjuvant that stimulates immune responses in the blood and gut for the ability to improve therapy in rhesus macaques chronically infected with SIV. Using the SIV-macaque model for AIDS, we show that epidermal co-delivery of plasmids expressing SIV Gag, RT, Nef and Env, and the mucosal adjuvant, heat-labile E. coli enterotoxin (LT), during antiretroviral therapy (ART) induced a substantial 2–4-log fold reduction in mean virus burden in both the gut and blood when compared to unvaccinated controls and provided durable protection from viral rebound and disease progression after the drug was discontinued. This effect was associated with significant increases in IFN-γ T cell responses in both the blood and gut and SIV-specific CD8+ T cells with dual TNF-α and cytolytic effector functions in the blood. Importantly, a broader specificity in the T cell response seen in the gut, but not the blood, significantly correlated with a reduction in virus production in mucosal tissues and a lower virus burden in plasma. We conclude that immunizing with vaccines that induce immune responses in mucosal gut tissue could reduce residual viral reservoirs during drug therapy and improve long-term treatment of HIV infection in humans. PMID:22442716
Lai, Lilin; Vödrös, Dalma; Kozlowski, Pamela A.; Montefiori, David C.; Wilson, Robert L.; Akerstrom, Vicki L.; Chennareddi, Lakshmi; Yu, Tianwei; Kannanganat, Sunil; Ofielu, Lazarus; Villinger, Francois; Wyatt, Linda S.; Moss, Bernard; Amara, Rama Rao; Robinson, Harriet L.
Single intradermal or intramuscular inoculations of GM-CSF DNA with the DNA prime for a simian-human immunodeficiency virus (SHIV)-89.6 vaccine, which consists of DNA priming followed by modified vaccinia Ankara (MVA) boosting, increased protection of both the blood and intestines against the acute phase of an intrarectal SHIV-89.6P challenge. GM-CSF appeared to contribute to protection by enhancing two antibody responses: the avidity maturation of anti-Env IgG in blood (p=<0.01) and the presence of long lasting anti-viral IgA in rectal secretions (p<0.01). The avidity of anti-Env IgG showed strong correlations with protection both pre and post challenge. Animals with the highest avidity anti-Env Ab had 1000-fold reductions in peak viremia over those with the lowest avidity anti-Env Ab. The enhanced IgA response was associated with the best protection, but did not achieve significance. PMID:17698160
Fioretti, Daniela; Iurescia, Sandra; Fazio, Vito Michele; Rinaldi, Monica
Due to their rapid and widespread development, DNA vaccines have entered into a variety of human clinical trials for vaccines against various diseases including cancer. Evidence that DNA vaccines are well tolerated and have an excellent safety profile proved to be of advantage as many clinical trials combines the first phase with the second, saving both time and money. It is clear from the results obtained in clinical trials that such DNA vaccines require much improvement in antigen expression and delivery methods to make them sufficiently effective in the clinic. Similarly, it is clear that additional strategies are required to activate effective immunity against poorly immunogenic tumor antigens. Engineering vaccine design for manipulating antigen presentation and processing pathways is one of the most important aspects that can be easily handled in the DNA vaccine technology. Several approaches have been investigated including DNA vaccine engineering, co-delivery of immunomodulatory molecules, safe routes of administration, prime-boost regimen and strategies to break the immunosuppressive networks mechanisms adopted by malignant cells to prevent immune cell function. Combined or single strategies to enhance the efficacy and immunogenicity of DNA vaccines are applied in completed and ongoing clinical trials, where the safety and tolerability of the DNA platform are substantiated. In this review on DNA vaccines, salient aspects on this topic going from basic research to the clinic are evaluated. Some representative DNA cancer vaccine studies are also discussed. PMID:20368780
Fioretti, Daniela; Iurescia, Sandra; Fazio, Vito Michele; Rinaldi, Monica
Due to their rapid and widespread development, DNA vaccines have entered into a variety of human clinical trials for vaccines against various diseases including cancer. Evidence that DNA vaccines are well tolerated and have an excellent safety profile proved to be of advantage as many clinical trials combines the first phase with the second, saving both time and money. It is clear from the results obtained in clinical trials that such DNA vaccines require much improvement in antigen expression and delivery methods to make them sufficiently effective in the clinic. Similarly, it is clear that additional strategies are required to activate effective immunity against poorly immunogenic tumor antigens. Engineering vaccine design for manipulating antigen presentation and processing pathways is one of the most important aspects that can be easily handled in the DNA vaccine technology. Several approaches have been investigated including DNA vaccine engineering, co-delivery of immunomodulatory molecules, safe routes of administration, prime-boost regimen and strategies to break the immunosuppressive networks mechanisms adopted by malignant cells to prevent immune cell function. Combined or single strategies to enhance the efficacy and immunogenicity of DNA vaccines are applied in completed and ongoing clinical trials, where the safety and tolerability of the DNA platform are substantiated. In this review on DNA vaccines, salient aspects on this topic going from basic research to the clinic are evaluated. Some representative DNA cancer vaccine studies are also discussed.
DNA vaccines expressing the duck hepatitis B virus surface proteins lead to reduced numbers of infected hepatocytes and protect ducks against the development of chronic infection in a virus dose-dependent manner.
Miller, Darren S; Kotlarski, Ieva; Jilbert, Allison R
We tested the efficacy of DNA vaccines expressing the duck hepatitis B virus (DHBV) pre-surface (pre-S/S) and surface (S) proteins in modifying the outcome of infection in 14-day-old ducks. In two experiments, Pekin Aylesbury ducks were vaccinated on days 4 and 14 of age with plasmid DNA vaccines expressing either the DHBV pre-S/S or S proteins, or the control plasmid vector, pcDNA1.1Amp. All ducks were then challenged intravenously on day 14 of age with 5 x 10(7) or 5 x 10(8) DHBV genomes. Levels of initial DHBV infection were assessed using liver biopsy tissue collected at day 4 post-challenge (p.c.) followed and immunostained for DHBV surface antigen to determine the percentage of infected hepatocytes. All vector vaccinated ducks challenged with 5 x 10(7) and 5 x 10(8) DHBV genomes had an average of 3.21% and 20.1% of DHBV-positive hepatocytes respectively at day 4 p.c. and 16 out of 16 ducks developed chronic DHBV infection. In contrast, pre-S/S and S vaccinated ducks challenged with 5 x 10(7) DHBV genomes had reduced levels of initial infection with an average of 1.38% and 1.93% of DHBV-positive hepatocytes at day 4 p.c. respectively and 10 of 18 ducks were protected against chronic infection. The pre-S/S and the S DNA vaccinated ducks challenged with 5 x 10(8) DHBV genomes had an average of 31.5% and 9.2% of DHBV-positive hepatocytes on day 4 p.c. respectively and only 4 of the 18 vaccinated ducks were protected against chronic infection. There was no statistically significant difference in the efficacy of the DHBV pre-S/S or S DNA vaccines. In conclusion, vaccination of young ducks with DNA vaccines expressing the DHBV pre-S/S and S proteins induced rapid immune responses that reduced the extent of initial DHBV infection in the liver and prevented the development of chronic infection in a virus dose-dependent manner.
Cao, Shinuo; Mousa, Ahmed Abdelmoniem; Aboge, Gabriel Oluga; Kamyingkird, Ketsarin; Zhou, Mo; Moumouni, Paul Franck Adjou; Terkawi, Mohamad Alaa; Masatani, Tatsunori; Nishikawa, Yoshifumi; Suzuki, Hiroshi; Fukumoto, Shinya; Xuan, Xuenan
A heterologous prime-boost vaccination regime with DNA and recombinant vaccinia virus (rvv) vectors expressing relevant antigens has been shown to induce effective immune responses against several infectious pathogens. In this study, we describe the effectiveness of the prime-boost strategy by immunizing dogs with a recombinant plasmid followed by vaccinia virus, both of which expressed the glutamic acid-rich protein (BgGARP) of Babesia gibsoni. The dogs immunized with the prime-boost regime developed a significantly high level of specific antibodies against BgGARP when compared with the control groups. The antibody level was strongly increased after a booster immunization with a recombinant vaccinia virus. Two weeks after the booster immunization with a recombinant vaccinia virus expressing BgGARP, the dogs were challenged with B. gibsoni parasite. The dogs immunized with the prime-boost regime showed partial protection, manifested as a significantly low level of parasitemia. These results indicated that this type of DNA/rvv prime-boost immunization approach may have use against B. gibsoni infection in dogs.
Development of a DNA vaccine for chicken infectious anemia and its immunogenicity studies using high mobility group box 1 protein as a novel immunoadjuvant indicated induction of promising protective immune responses.
Sawant, Pradeep Mahadev; Dhama, Kuldeep; Rawool, Deepak Bhiva; Wani, Mohd Yaqoob; Tiwari, Ruchi; Singh, Shambhu Dayal; Singh, Raj Kumar
Chicken infectious anaemia (CIA) is an economically important and emerging poultry disease reported worldwide. Current CIA vaccines have limitations like, the inability of the virus to grow to high titres in embryos/cell cultures, possession of residual pathogenicity and a risk of reversion to virulence. In the present study, a DNA vaccine, encoding chicken infectious anaemia virus (CIAV) VP1 and VP2 genes, was developed and co-administered with truncated chicken high mobility group box 1 (HMGB1ΔC) protein in young chicks for the evaluation of vaccine immune response. CIAV VP1 and VP2 genes were cloned in pTARGET while HMGB1ΔC in PET32b vector. In vitro expression of these gene constructs was evaluated by Western blotting. Further, recombinant HMGB1ΔC was evaluated for its biological activity. The CIAV DNA vaccine administration in specific pathogen free chicks resulted in moderately protective ELISA antibody titres in the range of 4322.87 ± 359.72 to 8288.19 ± 136.38, increased CD8(+) cells, and a higher titre was observed by co-administration of novel adjuvant (HMGB1ΔC) and booster immunizations. The use of vaccine with adjuvant showed achieving antibody titres nearly 8500, titre considered as highly protective, which indicates that co-immunization of HMGB1ΔC may have a strong adjuvant activity on CIAV DNA vaccine induced immune responses. The able potential of HMGB1 protein holding strong adjuvant activity could be exploited further with trials with vaccines for other important pathogens for achieving the required protective immune responses.
Jones, Bart G; Sealy, Robert E; Zhan, Xiaoyan; Freiden, Pamela J; Surman, Sherri L; Blanchard, James L; Hurwitz, Julia L
The pandemic of HIV-1 has continued for decades, yet there remains no licensed vaccine. Previous research has demonstrated the effectiveness of a multi-envelope, multi-vectored HIV-1 vaccine in a macaque-SHIV model, illustrating a potential means of combating HIV-1. Specifically, recombinant DNA, vaccinia virus (VV) and purified protein (DVP) delivery systems were used to vaccinate animals with dozens of antigenically distinct HIV-1 envelopes for induction of immune breadth. The vaccinated animals controlled disease following challenge with a heterologous SHIV. This demonstration suggested that the antigenic cocktail vaccine strategy, which has succeeded in several other vaccine fields (e.g. pneumococcus), might also succeed against HIV-1. The strategy remains untested in an advanced clinical study, in part due to safety concerns associated with the use of replication-competent VV. To address this concern, we designed a macaque study in which psoralen/ultraviolet light-inactivated VV (UV VV) was substituted for replication-competent VV in the multi-envelope DVP protocol. Control animals received a vaccine encompassing no VV, or no vaccine. All VV vaccinated animals generated an immune response toward VV, and all vaccinated animals generated an immune response toward HIV-1 envelope. After challenge with heterologous SHIV 89.6P, animals that received replication-competent VV or UV VV experienced similar outcomes. They exhibited reduced peak viral loads, maintenance of CD4+ T cell counts and improved survival compared to control animals that received no VV or no vaccine; there were 0/15 deaths among all animals that received VV and 5/9 deaths among controls. Results define a practical means of improving VV safety, and encourage advancement of a promising multi-envelope DVP HIV-1 vaccine candidate. Copyright © 2012 Elsevier Ltd. All rights reserved.
Yu, Yun-Zhou; Guo, Jin-Peng; An, Huai-Jie; Zhang, Shu-Ming; Wang, Shuang; Yu, Wei-Yuan; Sun, Zhi-Wei
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.
He, J; Hayes, C G; Binn, L N; Seriwatana, J; Vaughn, D W; Kuschner, R A; Innis, B L
Injection of an expression vector pJHEV containing hepatitis E virus (HEV) structural protein open reading frame 2 gene generates a strong antibody response in BALB/c mice that can bind to and agglutinate HEV. In this study, we tested for immunologic memory in immunized mice whose current levels of IgG to HEV were low or undetectable despite 3 doses of HEV DNA vaccine 18 months earlier. Mice previously vaccinated with vector alone were controls. All mice were administered a dose of HEV DNA vaccine to simulate an infectious challenge with HEV. The endpoint was IgG to HEV determined by ELISA. Ten days after the vaccine dose, 5 of 9 mice previously immunized with HEV DNA vaccine had a slight increase in IgG to HEV. By 40 days after the vaccine dose, the level of IgG to HEV had increased dramatically in all 9 mice (108-fold increase in geometric mean titer). In contrast, no control mice became seropositive. These results indicate that mice vaccinated with 3 doses of HEV DNA vaccine retain immunologic memory. In response to a small antigenic challenge delivered as DNA, possibly less than delivered by a human infective dose of virus, mice with memory were able to generate high levels of antibody in less time than the usual incubation period of hepatitis E. We speculate that this type of response could protect a human from overt disease.
Dowd, Kimberly A; Ko, Sung-Youl; Morabito, Kaitlyn M; Yang, Eun Sung; Pelc, Rebecca S; DeMaso, Christina R; Castilho, Leda R; Abbink, Peter; Boyd, Michael; Nityanandam, Ramya; Gordon, David N; Gallagher, John Robert; Chen, Xuejun; Todd, John-Paul; Tsybovsky, Yaroslav; Harris, Audray; Huang, Yan-Jang S; Higgs, Stephen; Vanlandingham, Dana L; Andersen, Hanne; Lewis, Mark G; De La Barrera, Rafael; Eckels, Kenneth H; Jarman, Richard G; Nason, Martha C; Barouch, Dan H; Roederer, Mario; Kong, Wing-Pui; Mascola, John R; Pierson, Theodore C; Graham, Barney S
Zika virus (ZIKV) was identified as a cause of congenital disease during the explosive outbreak in the Americas and Caribbean that began in 2015. Because of the ongoing fetal risk from endemic disease and travel-related exposures, a vaccine to prevent viremia in women of childbearing age and their partners is imperative. We found that vaccination with DNA expressing the premembrane and envelope proteins of ZIKV was immunogenic in mice and nonhuman primates, and protection against viremia after ZIKV challenge correlated with serum neutralizing activity. These data not only indicate that DNA vaccination could be a successful approach to protect against ZIKV infection, but also suggest a protective threshold of vaccine-induced neutralizing activity that prevents viremia after acute infection. Copyright © 2016, American Association for the Advancement of Science.
Cheng, Po-Ching; Lin, Ching-Nan; Peng, Shih-Yi; Kang, Tsung-Fu; Lee, Kin-Mu
Schistosomiasis is listed as one of most important tropical diseases and more than 200 million people are estimated to be infected. Development of a vaccine is thought to be the most effective way to control this disease. Recombinant 26-kDa glutathione S-transferase (rSjGST) has previously been reported to achieve a worm reduction rate of 42-44%. To improve the efficiency of the vaccine against Schistosoma japonicum, we immunized mice with a combination of pcDNA vector-encoded 26-kDa SjGST (pcDNA/SjGST), IL-12 expressing-plasmid (pIL-12), and rSjGST. Co-vaccination with pcDNA/SjGST, pIL-12, and rSjGST led to a reduction in worm burden, hepatic egg burden, and the size of liver tissue granulomas than that in the untreated infection controls. In addition, we detected high levels of specific IgG, IgG1, and IgG2a against the rSjGST antigen in infected mice vaccinated with this combination of pcDNA/SjGST, pIL-12, and rSjGST. Moreover, high expression levels of Th2 cytokines, including IL-4 and IL-10, were also detected in this group, without diminished levels of IL-12, INF-γ, and TNF-α cytokines that are related to parasite killing. In conclusion, we have developed a new vaccination regimen against S. japonicum infection and shown that co-immunization with pcDNA/SjGST vaccine, pIL-12, and rSjGST has significant anti-parasite, anti-hepatic egg and anti-pathology effects in mice. The efficacy of this vaccination method should be further validated in large animals such as water buffalo. This method may help to reduce the transmission of zoonotic schistosomiasis japonica.
Cheng, Po-Ching; Lin, Ching-Nan; Peng, Shih-Yi; Kang, Tsung-Fu; Lee, Kin-Mu
Schistosomiasis is listed as one of most important tropical diseases and more than 200 million people are estimated to be infected. Development of a vaccine is thought to be the most effective way to control this disease. Recombinant 26-kDa glutathione S-transferase (rSjGST) has previously been reported to achieve a worm reduction rate of 42–44%. To improve the efficiency of the vaccine against Schistosoma japonicum, we immunized mice with a combination of pcDNA vector-encoded 26-kDa SjGST (pcDNA/SjGST), IL-12 expressing-plasmid (pIL-12), and rSjGST. Co-vaccination with pcDNA/SjGST, pIL-12, and rSjGST led to a reduction in worm burden, hepatic egg burden, and the size of liver tissue granulomas than that in the untreated infection controls. In addition, we detected high levels of specific IgG, IgG1, and IgG2a against the rSjGST antigen in infected mice vaccinated with this combination of pcDNA/SjGST, pIL-12, and rSjGST. Moreover, high expression levels of Th2 cytokines, including IL-4 and IL-10, were also detected in this group, without diminished levels of IL-12, INF-γ, and TNF-α cytokines that are related to parasite killing. In conclusion, we have developed a new vaccination regimen against S. japonicum infection and shown that co-immunization with pcDNA/SjGST vaccine, pIL-12, and rSjGST has significant anti-parasite, anti-hepatic egg and anti-pathology effects in mice. The efficacy of this vaccination method should be further validated in large animals such as water buffalo. This method may help to reduce the transmission of zoonotic schistosomiasis japonica. PMID:26891172
Several novel antigens of Mycobacterium avium subsp. paratuberculosis have been studied as vaccine components and their immunogenicity has been evaluated. Previously, we reported that 85 antigen complex (85A, 85B, and 85C), superoxide dismutase (SOD), and 35kDa protein could induce significant lymph...
Crill, Wayne D.; Hughes, Holly R.; Trainor, Nicole B.; Davis, Brent S.; Whitney, Matt T.; Chang, Gwong-Jen J.
Dengue viruses (DENV) are the most important mosquito transmitted viral pathogens infecting humans. DENV infection produces a spectrum of disease, most commonly causing a self-limiting flu-like illness known as dengue fever; yet with increased frequency, manifesting as life-threatening dengue hemorrhagic fever (DHF). Waning cross-protective immunity from any of the four dengue serotypes may enhance subsequent infection with another heterologous serotype to increase the probability of DHF. Decades of effort to develop dengue vaccines are reaching the finishing line with multiple candidates in clinical trials. Nevertheless, concerns remain that imbalanced immunity, due to the prolonged prime-boost schedules currently used in clinical trials, could leave some vaccinees temporarily unprotected or with increased susceptibility to enhanced disease. Here we develop a DENV serotype 1 (DENV-1) DNA vaccine with the immunodominant cross-reactive B cell epitopes associated with immune enhancement removed. We compare wild-type (WT) with this cross-reactivity reduced (CRR) vaccine and demonstrate that both vaccines are equally protective against lethal homologous DENV-1 challenge. Under conditions mimicking natural exposure prior to acquiring protective immunity, WT vaccinated mice enhanced a normally sub-lethal heterologous DENV-2 infection resulting in DHF-like disease and 95% mortality in AG129 mice. However, CRR vaccinated mice exhibited redirected serotype-specific and protective immunity, and significantly reduced morbidity and mortality not differing from naїve mice. Thus, we demonstrate in an in vivo DENV disease model, that non-protective vaccine-induced immunity can prime vaccinees for enhanced DHF-like disease and that CRR DNA immunization significantly reduces this potential vaccine safety concern. The sculpting of immune memory by the modified vaccine and resulting redirection of humoral immunity provide insight into DENV vaccine-induced immune responses. PMID
Characterization of the antibody response elicited by immunization with pneumococcal surface protein A (PspA) as recombinant protein or DNA vaccine and analysis of protection against an intranasal lethal challenge with Streptococcus pneumoniae.
Vadesilho, Cintia F M; Ferreira, Daniela M; Moreno, Adriana T; Chavez-Olortegui, Carlos; Machado de Avila, Ricardo A; Oliveira, Maria Leonor S; Ho, Paulo L; Miyaji, Eliane N
Pneumococcal surface protein A (PspA) is an important candidate for a vaccine against pneumococcal infections. DNA vaccines expressing PspA were shown to protect mice against intraperitoneal and colonization challenge models in mice. We now show that a DNA vaccine expressing PspA from clade 4 (pSec-pspA4Pro) is also able to elicit protection against an intranasal lethal challenge model at levels similar to the recombinant protein PspA4Pro adjuvanted with alum. PspA4Pro + alum induced an IgG response characterized by a high IgG1/IgG2a ratio, leading to a lack of binding of anti-PspA IgG2a antibodies to intact pneumococci in vitro, which is in contrast to the response elicited by pSec-pspA4Pro. Epitopes recognized by the sera were mapped and antibodies induced by immunization with PspA4Pro + alum showed positive reaction with several synthetic peptides, mostly located in the first half of the protein. On the other hand, antibodies induced by the DNA vaccine showed reactivity with only two peptides. Though both strategies were protective against the intranasal lethal challenge model, the elicited humoral responses differ significantly, with the detection of important differences in the Fc (IgG1/IgG2a ratios) and Fab (recognized epitopes) regions of the induced antibodies.
Holmgren, Jan; Parashar, Umesh D; Plotkin, Stanley; Louis, Jacques; Ng, Su-Peing; Desauziers, Eric; Picot, Valentina; Saadatian-Elahi, Mitra
An immunological Correlate of Protection (CoP) is an immune response that is statistically interrelated with protection. Identification of CoPs for enteric vaccines would help design studies to improve vaccine performance of licensed vaccines in low income settings, and would facilitate the testing of future vaccines in development that might be more affordable. CoPs are lacking today for most existing and investigational enteric vaccines. In order to share the latest information on CoPs for enteric vaccines and to discuss novel approaches to correlate mucosal immune responses in humans with protection, the Foundation Mérieux organized an international conference of experts where potential CoPs for vaccines were examined using case-studies for both bacterial and viral enteric pathogens. Experts on the panel concluded that to date, all established enteric vaccine CoPs, such as those for hepatitis A, Vi typhoid and poliovirus vaccines, are based on serological immune responses even though these may poorly reflect the relevant gut immune responses or predict protective efficacy. Known CoPs for cholera, norovirus and rotavirus could be considered as acceptable for comparisons of similarly composed vaccines while more work is still needed to establish CoPs for the remaining enteric pathogens and their candidate vaccines. Novel approaches to correlate human mucosal immune responses with protection include the investigation of gut-originating antibody-secreting cells (ASCs), B memory cells and follicular helper T cells from samples of peripheral blood during their recirculation. Copyright © 2017.
DNA vaccines encoding the envelope protein of West Nile virus lineages 1 or 2 administered intramuscularly, via electroporation and with recombinant virus protein induce partial protection in large falcons (Falco spp.).
Fischer, Dominik; Angenvoort, Joke; Ziegler, Ute; Fast, Christine; Maier, Kristina; Chabierski, Stefan; Eiden, Martin; Ulbert, Sebastian; Groschup, Martin H; Lierz, Michael
As West Nile virus (WNV) can cause lethal diseases in raptors, a vaccination prophylaxis of free-living and captive populations is desirable. In the absence of vaccines approved for birds, equine vaccines have been used in falcons, but full protection against WNV infection was not achieved. Therefore, two DNA vaccines encoding the ectodomain of the envelope protein of WNV lineages 1 and 2, respectively, were evaluated in 28 large falcons. Four different vaccination protocols were used, including electroporation and booster-injections of recombinant WNV domain III protein, before challenge with the live WNV lineage 1 strain NY99. Drug safety, plasmid shedding and antibody production were monitored during the vaccination period. Serological, virological, histological, immunohistochemical and molecular biological investigations were performed during the challenge trials. Antibody response following vaccination was low overall and lasted for a maximum of three weeks. Plasmid shedding was not detected at any time. Viremia, mortality and levels, but not duration, of oral virus shedding were reduced in all of the groups during the challenge trial compared to the non-vaccinated control group. Likewise, clinical scoring, levels of cloacal virus shedding and viral load in organs were significantly reduced in three vaccination groups. Histopathological findings associated with WNV infections (meningo-encephalitis, myocarditis, and arteritis) were present in all groups, but immunohistochemical detection of the viral antigen was reduced. In conclusion, the vaccines can be used safely in falcons to reduce mortality and clinical signs and to lower the risk of virus transmission due to decreased levels of virus shedding and viremia, but full protection was not achieved in all groups.
Rosado-Vallado, Miguel; Mut-Martin, Mirza; García-Miss, Maria del Rosario; Dumonteil, Eric
DNA vaccines have been able to induce partial protection against infection with Leishmania in mice, but it is still necessary to increase their efficacy. In the present study we evaluated aluminium phosphate as an adjuvant of different formulations and doses of DNA vaccines against L. mexicana in BALB/c mice. Aluminium phosphate had no effect on the humoral response induced by a high dose (100 microg) DNA vaccine, but slightly increased the cellular response and the protection against infection. It also allowed a non-immunoprotective low dose (20 microg) DNA vaccine encoding L. mexicana GP63 and Leishmania donovani NH36 to become protective. Aluminium phosphate may thus be an effective, low cost and safe adjuvant for DNA vaccines, and the vaccine formulation described here may be an excellent candidate for further vaccine development against Leishmania.
Enhanced protective efficacy against Mycobacterium tuberculosis afforded by BCG prime-DNA boost regimen in an early challenge mouse model is associated with increased splenic interleukin-2-producing CD4 T-cell frequency post-vaccination.
Kang, Han; Yuan, Qin; Ma, Hui; Hu, Zhi-Dong; Han, De-Ping; Wu, Kang; Lowrie, Douglas B; Fan, Xiao-Yong
The development of improved vaccines and vaccination strategies against Mycobacterium tuberculosis has been hindered by a limited understanding of the immune correlates of anti-tuberculosis protective immunity. Simple measurement of interferon-γ frequency or production per se does not provide adequate prediction of immune protection. In this study, we examined the relationship between T-cell immune responses and protective efficacy conferred by the heterologous vaccination strategy, bacillus Calmette-Guérin (BCG) prime-Ag85A DNA boost (B/D), in an early challenge mouse model of pulmonary tuberculosis. The results demonstrated that mice vaccinated with the B/D regimen had a significantly reduced bacillary load compared with BCG-vaccinated mice, and the reduction in colony-forming units was associated with decreased pathology and lower levels of inflammatory cytokines in the infected lungs. Further analysis of immunogenicity showed that the superior protection afforded by the B/D regimen was associated with significantly increased frequency of splenic interleukin-2 (IL-2) -producing CD4 T cells and increased IL-2 production when measured as integrated mean fluorescence intensity post-vaccination as well. These data suggest that measurement of elevated frequency of IL-2-producing CD4 T cells or IL-2 production in the spleens of vaccinated mice can predict vaccine efficacy, at least in the B/D strategy, and add to the accumulating body of evidence suggesting that BCG prime-boost strategies may be a useful approach to the control of M. tuberculosis infection.
evaluation of a DN A vaccine for E . rhusiopathiae ). The research represents the most feasible, as well as the most useful, expenditure of funds and...controlling DoD office). • DISTRIBUTION STATEMENT E . Distribution authorized to DoD Components only (fill in reason) (date of determination). Other... rhusiopathiae AWARD PERIOD: December 1, 2004 through December 31, 2006 20090325244 OBJECTIVES: The work described in this report is based on discussion with
Gupta, Jyoti; Pathak, Manisha; Misra, Sweta; Misra-Bhattacharya, Shailja
We earlier demonstrated the immunoprophylactic efficacy of recombinant heavy chain myosin (Bm-Myo) of Brugia malayi (B. malayi) in rodent models. In the current study, further attempts have been made to improve this efficacy by employing alternate approaches such as homologous DNA (pcD-Myo) and heterologous DNA/protein prime boost (pcD-Myo+Bm-Myo) in BALB/c mouse model. The gene bm-myo was cloned in a mammalian expression vector pcDNA 3.1(+) and protein expression was confirmed in mammalian Vero cell line. A significant degree of protection (79.2%±2.32) against L3 challenge in pcD-Myo+Bm-Myo immunized group was observed which was much higher than that exerted by Bm-Myo (66.6%±2.23) and pcD-Myo (41.6%±2.45). In the heterologous immunized group, the percentage of peritoneal leukocytes such as macrophages, neutrophils, B cells and T cells marginally increased and their population augmented further significantly following L3 challenge. pcD-Myo+Bm-Myo immunization elicited robust cellular and humoral immune responses as compared to pcD-Myo and Bm-Myo groups as evidenced by an increased accumulation of CD4+, CD8+ T cells and CD19+ B cells in the mouse spleen and activation of peritoneal macrophages. Though immunized animals produced antigen-specific IgG antibodies and isotypes, sera of mice receiving pcD-Myo+Bm-Myo or Bm-Myo developed much higher antibody levels than other groups and there was profound antibody-dependent cellular adhesion and cytotoxicity (ADCC) to B. malayi infective larvae (L3). pcD-Myo+Bm-Myo as well as Bm-Myo mice generated a mixed T helper cell phenotype as evidenced by the production of both pro-inflammatory (IL-2, IFN-γ) and anti-inflammatory (IL-4, IL-10) cytokines. Mice receiving pcD-Myo on contrary displayed a polarized pro-inflammatory immune response. The findings suggest that the priming of animals with DNA followed by protein booster generates heightened and mixed pro- and anti-inflammatory immune responses that are capable of providing
Myhr, Anne Ingeborg
DNA vaccines have great potential as preventive or therapeutic vaccines against viral, bacterial, or parasitic diseases as well as cancer, and may also be used as gene therapy products. Although many human and veterinary DNA vaccines have been investigated in laboratory trials, only four of these have been approved for commercial use. In this paper an overview of the regulatory requirements for the development of DNA vaccines is given. The regulatory process in EU and USA is described. A discussion concerning the relevance of national regulations on gene technology is included. In addition the main safety concerns associated with DNA vaccines, relating to unwanted side effects in the vaccinated mammal or fish, are presented. Finally, the need for greater openness regarding the assessment information is discussed.
Zhu, Wei-Ning; Wang, Jin-Lei; Chen, Kai; Yue, Dong-Mei; Zhang, Xiao-Xuan; Huang, Si-Yang; Zhu, Xing-Quan
Toxoplasma gondii, an obligatory intracellular protozoan, can cause serious public health problems and economic losses worldwide. Two novel dense granule proteins (GRA17 and GRA23) were recently identified as T. gondii-secreted proteins which are localized to the parasitophorous vacuole membrane (PVM) and can mediate the movement of small molecules between the host cell and parasitophorous vacuole (PV). In the present study, we evaluated the protective immunity induced by DNA vaccination with genes encoding GRA17 and GRA23 against acute toxoplasmosis in mice. Eukaryotic expressing plasmids pVAX-TgGRA17 and pVAX-TgGRA23 were constructed. Then, BALB/c mice were intramuscularly immunized with pVAX-TgGRA17, pVAX-TgGRA23, or pVAX-TgGRA17 + pVAX-TgGRA23 followed by challenge infection with the highly virulent RH strain of T. gondii. The specific immune responses and protective efficacy against T. gondii were examined by cytokine and serum antibody measurements, lymphocyte proliferation assays, flow cytometry of lymphocytes and the survival time after challenge. Our results showed that mice immunized with pVAX-TgGRA17, pVAX-TgGRA23, or pVAX-TgGRA17 + pVAX-TgGRA23 induced specific humoral and cellular responses, with higher level of IgG antibody, increased levels of Th1-type cytokines IFN-γ and IL-12 (p70), and CD3(+)CD4(+)CD8(-) and CD3(+)CD8(+)CD4(-) T cells, as well as prolonged survival time (9.1 ± 0.32 days for pVAX-TgGRA17, 10.8 ± 0.79 days for pVAX-TgGRA23, and 12.6 ± 2.55 days for pVAX-TgGRA17 + pVAX-TgGRA23) compared to the blank control (7.11 ± 0.33 days), PBS control (7.22 ± 0.44 days), and pVAX I control (7.11 ± 0.33 days). These results demonstrated that both TgGRA17 and TgGRA23 are potential vaccine candidates, TgGRA23 has a better immunogenicity, and co-immunization of pVAX-TgGRA17 and pVAX-TgGRA23 induces better protective efficacy. Copyright © 2017. Published by Elsevier Inc.
de Souza, J B
A good understanding of the immunological correlates of protective immunity is an important requirement for the development of effective vaccines against malaria. However, this concern has received little attention even in the face of two decades of intensive vaccine research. Here, we review the immune response to blood-stage malaria, with a particular focus on the type of vaccine most likely to induce the kind of response required to give strong protection against infection.
Chandra, Janin; Dutton, Julie L.; Li, Bo; Woo, Wai-Ping; Xu, Yan; Tolley, Lynn K.; Yong, Michelle; Wells, James W.; R. Leggatt, Graham; Finlayson, Neil
We have previously shown that a novel DNA vaccine technology of codon optimization and the addition of ubiquitin sequences enhanced immunogenicity of a herpes simplex virus 2 polynucleotide vaccine in mice, and induced cell-mediated immunity when administered in humans at relatively low doses of naked DNA. We here show that a new polynucleotide vaccine using the same technology and encoding a fusion protein of the E6 and E7 oncogenes of high-risk human papillomavirus type 16 (HPV16) is immunogenic in mice. This vaccine induces long-lasting humoral and cell-mediated immunity and protects mice from establishment of HPV16-E7-expressing tumors. In addition, it suppresses growth of readily established tumors and shows enhanced efficacy when combined with immune checkpoint blockade targeted at PD-L1. This vaccine also facilitates rejection of HPV16-E7-expressing skin grafts that demonstrate epidermal hyperplasia with characteristics of cervical and vulvar intraepithelial neoplasia. Clinical studies evaluating the efficacy of this vaccine in patients with HPV16+ premalignancies are planned. PMID:28166181
A bicistronic DNA vaccine containing apical membrane antigen 1 and merozoite surface protein 4/5 can prime humoral and cellular immune responses and partially protect mice against virulent Plasmodium chabaudi adami DS malaria.
Rainczuk, A; Scorza, T; Spithill, T W; Smooker, P M
The ultimate malaria vaccine will require the delivery of multiple antigens from different stages of the complex malaria life cycle. In order to efficiently deliver multiple antigens with use of DNA vaccine technology, new antigen delivery systems must be assessed. This study utilized a bicistronic vector construct, containing an internal ribosome entry site, expressing a combination of malarial candidate antigens: merozoite surface protein 4/5 (MSP4/5) (fused to a monocyte chemotactic protein 3 chemoattractant sequence) and apical membrane antigen 1 (AMA-1) (fused to a tissue plasminogen activator secretion signal). Transfection of COS 7 cells with bicistronic plasmids resulted in production and secretion of both AMA-1 and MSP4/5 in vitro. Vaccination of BALB/c mice via intraepidermal gene gun and intramuscular routes against AMA-1 and MSP4/5 resulted in antibody production and significant in vitro proliferation of splenocytes stimulated by both AMA-1 and MSP4/5. Survival of BALB/c mice vaccinated with bicistronic constructs after lethal Plasmodium chabaudi adami DS erythrocytic-stage challenge was variable, although significant increases in survival and reductions in peak parasitemia were observed in several challenge trials when the vaccine was delivered by the intramuscular route. This study using a murine model demonstrates that the delivery of malarial antigens via bicistronic vectors is feasible. Further experimentation with bicistronic delivery systems is required for the optimization and refinement of DNA vaccines to effectively prime protective immune responses against malaria.
A DNA vaccine encoding the viral hemorrhagic septicemia virus genotype IVb glycoprotein confers protection in muskellunge (Esox masquinongy), rainbow trout (Oncorhynchus mykiss), brown trout (Salmo trutta), and lake trout (Salvelinus namaycush).
Standish, Isaac F; Millard, Elena V; Brenden, Travis O; Faisal, Mohamed
The viral hemorrhagic septicemia virus (VHSV) is one of the most serious fish pathogens. In 2003, a novel sublineage (genotype IVb) of this deadly virus emerged in the Great Lakes basin causing serious fish kills. We have previously demonstrated that a DNA plasmid (pcDNA), containing a cytomegalovirus (CMV) promoter and the viral hemorrhagic septicemia virus (VHSV) genotype IVb glycoprotein (G) gene insert (designated pVHSivb-G) confers moderate protection in muskellunge (Esox masquinongy), a highly susceptible species upon challenge. In order to achieve optimal protection, we investigated a number of factors including the incubation time [i.e. the number of degree days (° days)] before challenge, and viral challenge dose and route. Additionally, we tested if pVHSivb-G provides protection against VHSV-IVb to less susceptible salmonids such as rainbow trout (Oncorhynchus mykiss), brown trout (Salmo trutta) and lake trout (Salvelinus namaycush). An increase in the period lapsed between vaccination and challenge to 1880° days resulted in 95% relative percent protection (RPS) in muskellunge following a single administration of the pVHSivb-G plasmid and viral challenge. An RPS of 100% for muskellunge was achieved with a longer incubation period (2400° days) and in conjunction with a booster dose of the plasmid. The pVHSivb-G vaccine also elicited significant protection in all three salmonid species, reaching 100% RPS in lake trout following an incubation period of 1001° days prior to viral challenge. Vaccination with pVHSivb-G was also associated with the development of significant levels of circulating VHSV-binding antibodies in muskellunge as measured by indirect ELISA, which reached peak levels 6-7 weeks post-vaccination. Viral shedding in vaccinated survivors was minimal and of transient nature. The study shows that the pVHSivb-G plasmid can elicit a protective response against the wild virus strain in a range of species important in recreational and commercial
Riquelme-Neira, Roberto; Rivera, Alejandra; Sáez, Darwin; Fernández, Pablo; Osorio, Gonzalo; del Canto, Felipe; Salazar, Juan C.; Vidal, Roberto M.; Oñate, Angel
Enterohemorrhagic Escherichia coli (EHEC) O157:H7 is the predominant causative agent of hemorrhagic colitis in humans and is the cause of haemolytic uraemic syndrome and other illnesses. Cattle have been implicated as the main reservoir of this organism. Here, we evaluated the immunogenicity and protective efficacy of a DNA vaccine encoding conserved sequences of truncated EHEC factor for adherence-1 (efa-1′) in a mouse model. Intranasal administration of plasmid DNA carrying the efa-1′ gene (pVAXefa-1′) into C57BL/6 mice elicited both humoral and cellular immune responses. In animals immunized with pVAXefa-1′, EHEC-secreted protein-specific IgM and IgG antibodies were detected in sera at day 45. Anti-EHEC-secreted protein sIgA was also detected in nasal and bronchoalveolar lavages. In addition, antigen-specific T-cell-proliferation, IL-10, and IFN-γ were observed upon re-stimulation with either heat-killed bacteria or EHEC-secreted proteins. Vaccinated animals were also protected against challenge with E. coli O157:H7 strain EDL933. These results suggest that DNA vaccine encoding efa-1′ have therapeutic potential in interventions against EHEC infections. This approach could lead to a new strategy in the production of vaccines that prevent infections in cattle. PMID:26835434
Riquelme-Neira, Roberto; Rivera, Alejandra; Sáez, Darwin; Fernández, Pablo; Osorio, Gonzalo; del Canto, Felipe; Salazar, Juan C; Vidal, Roberto M; Oñate, Angel
Enterohemorrhagic Escherichia coli (EHEC) O157:H7 is the predominant causative agent of hemorrhagic colitis in humans and is the cause of haemolytic uraemic syndrome and other illnesses. Cattle have been implicated as the main reservoir of this organism. Here, we evaluated the immunogenicity and protective efficacy of a DNA vaccine encoding conserved sequences of truncated EHEC factor for adherence-1 (efa-1') in a mouse model. Intranasal administration of plasmid DNA carrying the efa-1' gene (pVAXefa-1') into C57BL/6 mice elicited both humoral and cellular immune responses. In animals immunized with pVAXefa-1', EHEC-secreted protein-specific IgM and IgG antibodies were detected in sera at day 45. Anti-EHEC-secreted protein sIgA was also detected in nasal and bronchoalveolar lavages. In addition, antigen-specific T-cell-proliferation, IL-10, and IFN-γ were observed upon re-stimulation with either heat-killed bacteria or EHEC-secreted proteins. Vaccinated animals were also protected against challenge with E. coli O157:H7 strain EDL933. These results suggest that DNA vaccine encoding efa-1' have therapeutic potential in interventions against EHEC infections. This approach could lead to a new strategy in the production of vaccines that prevent infections in cattle.
Montalvo-Álvarez, Ana M.; Folgueira, Cristina; Carrión, Javier; Monzote-Fidalgo, Lianet; Cañavate, Carmen; Requena, Jose M.
Protozoa of the genus Leishmania are causative agents of leishmaniasis, an important health problem in both human and veterinary medicine. Here, we describe a new heat shock protein (HSP) in Leishmania, belonging to the small HSP (sHSP) family in kinetoplastids. The protein is highly conserved in different Leishmania species, showing instead significant divergence with sHSP's from other organisms. The humoral response elicited against this protein during Leishmania infection has been investigated in natural infected humans and dogs, and in experimentally infected hamsters. Leishmania HSP20 is a prominent antigen for canine hosts; on the contrary, the protein seems to be a poor antigen for human immune system. Time-course analysis of appearance of anti-HSP20 antibodies in golden hamsters indicated that these antibodies are produced at late stages of the infection, when clinical symptoms of disease are patent. Finally, the protective efficacy of HSP20 was assessed in mice using a DNA vaccine approach prior to challenge with Leishmania amazonensis. PMID:18401455
Bertholet, Sylvie; Goto, Yasuyuki; Carter, Lauren; Bhatia, Ajay; Howard, Randall F.; Carter, Darrick; Coler, Rhea N.; Vedvick, Thomas S.; Reed, Steven G.
Development of a protective subunit vaccine against Leishmania spp. depends on antigens and adjuvants that induce appropriate immune responses. We evaluated a second generation polyprotein antigen (Leish-110f) in different adjuvant formulations for immunogenicity and protective efficacy against Leishmania spp. challenges. Vaccine-induced protection was associated with antibody and T cell responses to Leish-110f. CD4 T cells were the source of IFN-γ, TNF, and IL-2 double and triple positive populations. This study establishes the immunogenicity and protective efficacy of the improved Leish-110f subunit vaccine antigen adjuvanted with natural (MPL-SE) or synthetic (EM005) Toll-like receptor 4 agonists. PMID:19786136
Bertholet, Sylvie; Goto, Yasuyuki; Carter, Lauren; Bhatia, Ajay; Howard, Randall F; Carter, Darrick; Coler, Rhea N; Vedvick, Thomas S; Reed, Steven G
Development of a protective subunit vaccine against Leishmania spp. depends on antigens and adjuvants that induce appropriate immune responses. We evaluated a second generation polyprotein antigen (Leish-110f) in different adjuvant formulations for immunogenicity and protective efficacy against Leishmania spp. challenges. Vaccine-induced protection was associated with antibody and T cell responses to Leish-110f. CD4 T cells were the source of IFN-gamma, TNF, and IL-2 double- and triple-positive populations. This study establishes the immunogenicity and protective efficacy of the improved Leish-110f subunit vaccine antigen adjuvanted with natural (MPL-SE) or synthetic (EM005) Toll-like receptor 4 agonists.
Kuboniwa, Masae; Amano, Atsuo; Shizukuishi, Satoshi; Nakagawa, Ichiro; Hamada, Shigeyuki
Lys-gingipain (KGP), a lysine-specific cysteine proteinase, is one of the major virulence factors of Porphyromonas gingivalis. Here we examined the involvement of the catalytic domain of KGP (KGPcd) in hemoglobin binding by P. gingivalis, using a specific immunoglobulin G (IgG) elicited by the administration of plasmid DNA encoding KGPcd or the catalytic domain of Arg-gingipain (RGPcd). The pSeq2A/kgpcd and pSeq2B/rgpcd plasmids were constructed by the ligation of kgpcd and rgpcd DNA fragments, respectively. Female BALB/c mice were immunized with each of these plasmids. pSeq2A/kgpcd elicited a strong response to recombinant KGPcd (rKGPcd), as well as to comparably produced rRGPcd-reactive antibodies. The serum antibodies elicited by pSecTag2B/rgpcd also cross-reacted with rKGPcd as well as rRGPcd. Anti-KGPcd IgG significantly inhibited hemoglobin binding by P. gingivalis. Furthermore, the inhibition of hemoglobin binding was markedly enhanced by a combination of anti-KGPcd and anti-fimbriae. Anti-RGPcd IgG showed a negligible inhibitory effect, while both anti-KGPcd and anti-RGPcd IgGs showed significant inhibitory effects on Lys- and Arg-specific proteolytic activities and on the growth of P. gingivalis under iron-restricted conditions where supplemented hemoglobin was the sole iron source. Immunized mice were challenged by intraperitoneal inoculation with P. gingivalis. All nonimmunized mice died within 72 h; however, vaccination with pSeq2A/kgpcd and pSeq2B/rgpcd prevented inflammatory responses and prolonged the survival rate of immunized mice by 43 and 27%, respectively. These results suggest that KGPcd acts as a hemoglobin-binding protein and can also be useful as an immunogen inducing a protective response to P. gingivalis infection. PMID:11292714
Van Der Ryst, E
According to animal studies, DNA vaccines employ the genes encoding proteins of pathogens or tumors, in contrast to the more conventional vaccine approaches. In addition, DNA vaccinations do not involve infectious agents, proteins are expressed in their natural form resulting to better recognition of viral proteins by the antibodies, and both strong and durable cellular immune responses as well as neutralizing antibodies are induced. Altogether, this makes DNA vaccinations one of the most promising future candidates in the field of HIV vaccines. However, safety of DNA vaccines should be examined before these vaccines can be considered for large-scale clinical trials in humans. The question of a possible induction of anti-DNA antibodies, with the consequent development of autoimmune manifestations is emphasized. Another is the possible integration of DNA with insertional mutagenesis, which could lead to tumor formation and development of immunologic tolerance of antigen production persists.
Kurath, G.; ,
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.
Vaccination of monkeys with recombinant Plasmodium falciparum apical membrane antigen 1 confers protection against blood-stage malaria . Infect Immun...circumsporozoite protein partially protects healthy malaria -naive adults against Plasmodium falciparum sporozoite challenge. Infect Immun 74: 5933–5942...Ballou WR, et al. (1986) Malaria transmitted to humans by mosquitoes infected from cultured Plasmodium falciparum . Am J Trop Med Hyg 35: 66–68. 37
Evaluation of a DNA vaccine candidate expressing prM-E-NS1 antigens of dengue virus serotype 1 with or without granulocyte-macrophage colony-stimulating factor (GM-CSF) in immunogenicity and protection.
Zheng, Qun; Fan, Dongying; Gao, Na; Chen, Hui; Wang, Juan; Ming, Ying; Li, Jieqiong; An, Jing
Dengue is one of the most important mosquito-borne viral diseases. In past years, although considerable effort has been put into the development of a vaccine, there is currently no licensed dengue vaccine. In this study, we constructed DNA vaccines that carried the prM-E-NS1 genes of dengue virus serotype 1 (DV1) with or without the granulocyte-macrophage colony-stimulating factor (GM-CSF) gene, an attractive DNA vaccine adjuvant. Immunization with the plasmid pCAG-DV1/E/NS1, which expresses viral prM-E-NS1, or the bicistronic plasmid pCAG-DV1-GM, which co-expresses viral prM-E-NS1 and GM-CSF, resulted in long-term IgG response, high levels of splenocyte-secreted interferon-γ and interleukin-2, strong cytotoxic T lymphocyte activity and sufficient protection in the DV1-challenged mice. This suggested that both humoral and cellular immune responses were induced by the immunizations and that they played important roles in protection against the DV1 challenge. Interestingly, the magnitude, quality and protective capacity of the immune responses induced by immunization with pCAG-DV1/E/NS1 or pCAG-DV1-GM seemed stronger than those induced by pCAG-DV1/E (expressing viral prM-E alone). Taken together, we demonstrated that prM/E plus NS1 would be a suitable solution for the development of a DNA vaccine against DV.
Ye, Chunting; Choi, Jang Gi; Abraham, Sojan; Shankar, Premlata; Manjunath, N.
Targeting DNA vaccines to dendritic cells (DCs) greatly enhances immunity. Although several approaches have been used to target protein antigens 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, that 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 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 antigen 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 WN neutralizing antibodies 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. PMID:25270431
Ye, Chunting; Choi, Jang Gi; Abraham, Sojan; Shankar, Premlata; Manjunath, N
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.
Wang, Hai-Chao; An, Huai-Jie; Yu, Yun-Zhou; Xu, Qing
DNA vaccines require improvement for human use because they are generally weak stimulators of the immune system in humans. The efficacy of DNA vaccines can be improved using a viral replicon as vector to administer antigen of pathogen. In this study, we comprehensively evaluated the conventional non-viral DNA, viral replicon DNA or viral replicon particles (VRP) vaccines encoding different forms of anthrax protective antigen (PA) for specific immunity and protective potency against anthrax. Our current results clearly suggested that these viral replicon DNA or VRP vaccines derived from Semliki Forest virus (SFV) induced stronger PA-specific immune responses than the conventional non-viral DNA vaccines when encoding the same antigen forms, which resulted in potent protection against challenge with the Bacillus anthracis strain A16R. Additionally, the naked PA-expressing SFV replicon DNA or VRP vaccines without the need for high doses or demanding particular delivery regimens elicited robust immune responses and afforded completely protective potencies, which indicated the potential of the SFV replicon as vector of anthrax vaccines for use in clinical application. Therefore, our results suggest that these PA-expressing SFV replicon DNA or VRP vaccines may be suitable as candidate vaccines against anthrax.
against an aerosol spore challenge with the Ames isolate of Bacillus anthracis at 6 and 12 months. At 6 months after the primary injection, survival...vaccine was examined against an aerosol spore challenge with the Ames isolate of Bacillus anthracis at 6 and 12 months. At 6 months after the...Vaccine 24 (2006) 2530–2536 Duration of protection of rabbits after vaccination with Bacillus anthracis recombinant protective antigen vaccine S.F
against an aerosol spore challenge with the Ames isolate of Bacillus anthracis at 6 and 12 months. At 6 months after the primary injection, survival...rPA) vaccine was examined against an aerosol spore challenge with the Ames isolate of Bacillus anthracis at 6 and 12 months. At 6 months after the...Vaccine 24 (2006) 2530–2536 Duration of protection of rabbits after vaccination with Bacillus anthracis recombinant protective antigen vaccine S.F
Craigo, Jodi K.; Ezzelarab, Corin; Cook, Sheila J.; Chong, Liu; Horohov, David; Issel, Charles J.; Montelaro, Ronald C.
Lentiviral envelope (Env) antigenic variation and associated immune evasion present major obstacles to vaccine development. The concept that Env is a critical determinant for vaccine efficacy is well accepted, however defined correlates of protection associated with Env variation have yet to be determined. We reported an attenuated equine infectious anemia virus (EIAV) vaccine study that directly examined the effect of lentiviral Env sequence variation on vaccine efficacy. The study identified a significant, inverse, linear correlation between vaccine efficacy and increasing divergence of the challenge virus Env gp90 protein compared to the vaccine virus gp90. The report demonstrated approximately 100% protection of immunized ponies from disease after challenge by virus with a homologous gp90 (EV0), and roughly 40% protection against challenge by virus (EV13) with a gp90 13% divergent from the vaccine strain. In the current study we examine whether the protection observed when challenging with the EV0 strain could be conferred to animals via chimeric challenge viruses between the EV0 and EV13 strains, allowing for mapping of protection to specific Env sequences. Viruses containing the EV13 proviral backbone and selected domains of the EV0 gp90 were constructed and in vitro and in vivo infectivity examined. Vaccine efficacy studies indicated that homology between the vaccine strain gp90 and the N-terminus of the challenge strain gp90 was capable of inducing immunity that resulted in significantly lower levels of post-challenge virus and significantly delayed the onset of disease. However, a homologous N-terminal region alone inserted in the EV13 backbone could not impart the 100% protection observed with the EV0 strain. Data presented here denote the complicated and potentially contradictory relationship between in vitro virulence and in vivo pathogenicity. The study highlights the importance of structural conformation for immunogens and emphasizes the need for
Griffiths, Kristin L.; Villarreal, Daniel O.; Weiner, David B.; Khader, Shabaana A.
ABSTRACT Mycobacterium tuberculosis infects one third of the world's population. Due to variable efficacy of the Bacille Calmette Guerin (BCG) vaccine, development of novel TB vaccines remains a priority. Here, we demonstrate the protective efficacy of a novel multivalent DNA vaccine, which contains 15 synthetic antigens targeting the Mtb ESX secretion system. PMID:27322875
Carlétti, Dyego; Morais da Fonseca, Denise; Gembre, Ana Flávia; Masson, Ana Paula; Weijenborg Campos, Lívia; Leite, Luciana C C; Rodrigues Pires, Andréa; Lannes-Vieira, Joseli; Lopes Silva, Célio; Bonato, Vânia Luiza Deperon; Horn, Cynthia
Mycobacterium bovis BCG prime DNA (Mycobacterium tuberculosis genes)-booster vaccinations have been shown to induce greater protection against tuberculosis (TB) than BCG alone. This heterologous prime-boost strategy is perhaps the most realistic vaccination for the future of TB infection control, especially in countries where TB is endemic. Moreover, a prime-boost regimen using biodegradable microspheres seems to be a promising immunization to stimulate a long-lasting immune response. The alanine proline antigen (Apa) is a highly immunogenic glycoprotein secreted by M. tuberculosis. This study investigated the immune protection of Apa DNA vaccine against intratracheal M. tuberculosis challenge in mice on the basis of a heterologous prime-boost regimen. BALB/c mice were subcutaneously primed with BCG and intramuscularly boosted with a single dose of plasmid carrying apa and 6,6'-trehalose dimycolate (TDM) adjuvant, coencapsulated in microspheres (BCG-APA), and were evaluated 30 and 70 days after challenge. This prime-boost strategy (BCG-APA) resulted in a significant reduction in the bacterial load in the lungs, thus leading to better preservation of the lung parenchyma, 70 days postinfection compared to BCG vaccinated mice. The profound effect of this heterologous prime-boost regimen in the experimental model supports its development as a feasible strategy for prevention of TB.
Carlétti, Dyego; Morais da Fonseca, Denise; Gembre, Ana Flávia; Masson, Ana Paula; Weijenborg Campos, Lívia; Leite, Luciana C. C.; Rodrigues Pires, Andréa; Lannes-Vieira, Joseli; Lopes Silva, Célio; Bonato, Vânia Luiza Deperon
Mycobacterium bovis BCG prime DNA (Mycobacterium tuberculosis genes)-booster vaccinations have been shown to induce greater protection against tuberculosis (TB) than BCG alone. This heterologous prime-boost strategy is perhaps the most realistic vaccination for the future of TB infection control, especially in countries where TB is endemic. Moreover, a prime-boost regimen using biodegradable microspheres seems to be a promising immunization to stimulate a long-lasting immune response. The alanine proline antigen (Apa) is a highly immunogenic glycoprotein secreted by M. tuberculosis. This study investigated the immune protection of Apa DNA vaccine against intratracheal M. tuberculosis challenge in mice on the basis of a heterologous prime-boost regimen. BALB/c mice were subcutaneously primed with BCG and intramuscularly boosted with a single dose of plasmid carrying apa and 6,6′-trehalose dimycolate (TDM) adjuvant, coencapsulated in microspheres (BCG-APA), and were evaluated 30 and 70 days after challenge. This prime-boost strategy (BCG-APA) resulted in a significant reduction in the bacterial load in the lungs, thus leading to better preservation of the lung parenchyma, 70 days postinfection compared to BCG vaccinated mice. The profound effect of this heterologous prime-boost regimen in the experimental model supports its development as a feasible strategy for prevention of TB. PMID:23740922
Qi, Wen-Juan; Fang, Qiang
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, cysticercosis and toxoplasmosis in recent years.
Ho, Peggy P; Higgins, John P; Kidd, Brian A; Tomooka, Beren; Digennaro, Carla; Lee, Lowen Y; de Vegvar, Henry E Neuman; Steinman, Lawrence; Robinson, William H
Current therapies for rheumatoid arthritis (RA) and other autoimmune diseases non-specifically suppress immune function, and there is great need for fundamental approaches such as antigen-specific tolerizing therapy. In this paper we describe development of antigen-specific tolerizing DNA vaccines to treat collagen-induced arthritis (CIA) in mice, and use of protein microarrays to monitor response to therapy and to identify potential additional autoimmune targets for next generation vaccines. We demonstrate that tolerizing DNA vaccines encoding type II collagen (CII) reduced the incidence and severity of CIA. Atorvastatin, a statin drug found to reduce the severity of autoimmunity, potentiated the effect of DNA vaccines encoding CII. Analysis of cytokines produced by collagen-reactive T cells derived from mice receiving tolerizing DNA encoding CII, as compared to control vaccines, revealed reduced production of the pro-inflammatory cytokines IFN-gamma and TNF-alpha. Arthritis microarray analysis demonstrated reduced spreading of autoantibody responses in mice treated with DNA encoding CII. The development of tolerizing DNA vaccines, and the use of antibody profiling to guide design of and to monitor therapeutic responses to such vaccines, represents a promising approach for the treatment of RA and other autoimmune diseases.
Yero, Daniel; Pajón, Rolando; Pérez, Yusleydis; Fariñas, Mildrey; Cobas, Karem; Diaz, Daiyana; Solis, Rosa L; Acosta, Armando; Brookes, Charlotte; Taylor, Stephen; Gorringe, Andrew
We have shown previously that expression library immunization is viable alternative approach to induce protective immunity against Neisseria meningitidis serogroup B. In this study we report that few rounds of library screening allow identification of protective pools of defined antigens. A previously reported protective meningococcal library (L8, with 600 clones) was screened and two sub-libraries of 95 clones each were selected based on the induction of bactericidal and protective antibodies in BALB/c mice. After sequence analysis of each clone within these sub-libraries, we identified a pool of 20 individual antigens that induced protective immune responses in mice against N. meningitidis infection, and the observed protection was associated with the induction of bactericidal antibodies. Our studies demonstrate for the first time that ELI combined with sequence analysis is a powerful and efficient tool for identification of candidate antigens for use in a meningococcal vaccine.
Akhoon, Bashir Akhlaq; Slathia, Parvez Singh; Sharma, Preeti; Gupta, Shishir Kumar; Verma, Vijeshwar
African trypanosomiasis continues to be a major health problem, with more adults dying from this disease world-wide. As the sequence diversity of Trypanosoma brucei is extreme, with VSGs having 15-25% identity with most other VSGs, hence it displays a huge diversity of adaptations and host specificities. Therefore the need for an improved vaccine has become an international priority. The highly conserved and specific epitopes acting as both CD8+ and CD4+ T-cell epitopes (FLINKKPAL and FTALCTLAA) were predicted from large bunch of VSGs of T. brucei. Besides, some other potential epitopes with very high affinity for MHC I and II molecules were also determined while taking consideration on the most common HLA in the general population which accounts for major ethnicities. The vaccine candidates were found to be effective even for non-african populations as predicted by population coverage analysis. Hence the migrating travelers acting as a spread means of the infection can probably also be treated successfully after injection of such a multiepitopic vaccine. Exploiting the immunoinformatics approaches, we designed a potential vaccine by using the consensus epitopic sequence of 388 VSG proteins of T. brucei and performed in silico cloning of multiepitopic antigenic DNA sequence in pBI-CMV1 vector. Moreover, various techniques like codon adaptation, CpG optimization, removal of self recognized epitopes, use of adjuvant and co-injection with plasmids expressing immune-stimulatory molecules were implemented to enhance the immunogenicity of the proposed in silico vaccine.
Pharmaceuticals , Inc., Blue Bell, PA., 4Office of the Chief Scientists, Headquarters, 11 USAMRIID, Fort Detrick, MD. 12 13 ABSTRACT 14 We...Approved for public release; distribution is unlimited. an additional cross-challenge experiment in which the guinea pigs vaccinated with both 70 DNA...exposures were separated temporally. To our 74 knowledge, this study is the first report of a LASV/EBOV coinfection model, so in 75 addition to the
Needle-free jet injection of small doses of Japanese encephalitis DNA and inactivated vaccine mixture induces neutralizing antibodies in miniature pigs and protects against fetal death and mummification in pregnant sows.
Imoto, Jun-ichi; Ishikawa, Tomohiro; Yamanaka, Atsushi; Konishi, Misako; Murakami, Kenji; Shibahara, Tomoyuki; Kubo, Masanori; Lim, Chang-Kweng; Hamano, Masataka; Takasaki, Tomohiko; Kurane, Ichiro; Udagawa, Haruhide; Mukuta, Yoshihiro; Konishi, Eiji
Japanese encephalitis (JE) virus causes abortion and stillbirth in swine, and encephalitis in humans and horses. We have previously reported that immunogenicity of a DNA vaccine against JE was synergistically enhanced in mice by co-immunization with a commercial inactivated JE vaccine (JEVAX) under a needle-free injection system. Here, we found that this immunization strategy was also effective in miniature pigs. Because of the synergism, miniature pigs immunized twice with a mixture of 10 μg of DNA and a 1/100 dose of JEVAX developed a high neutralizing antibody titer (1:190 at 90% plaque reduction assay). Even using 1 μg of DNA, 3 of 4 pigs developed neutralizing antibodies. Following challenge, all miniature pigs with detectable neutralizing antibodies were protected against viremia. Pregnant sows inoculated with 10 or 1 μg of DNA mixed with JEVAX (1/100 dose) developed antibody titers of 1:40-1:320. Following challenge, fetal death and mummification were protected against in DNA/JEVAX-immunized sows. Copyright © 2010 Elsevier Ltd. All rights reserved.
Traxler, G S; Anderson, E; LaPatra, S E; Richard, J; Shewmaker, B; Kurath, G
A naked plasmid DNA encoding the glycoprotein (pCMV4-G) of a 1976 isolate of infectious hematopoietic necrosis virus (IHNV) obtained from steelhead Oncorhynchus mykiss was used to vaccinate Atlantic salmon Salmo salar against IHNV. Eight weeks post-vaccination the fish were challenged with a strain of IHNV originally isolated from farmed Atlantic salmon undergoing an epizootic. Fish injected with the glycoprotein-encoding plasmid were significantly (p < 0.05) protected against IHNV by both immersion and cohabitation challenge. Survivors of the first challenges were pooled and re-challenged by immersion 12 wk after the initial challenge. Significant (p < 0.05) protection was observed in all of the previously challenged groups including those receiving the complete vaccine. Fish injected with the glycoprotein-encoding plasmid produced low levels of virus-neutralizing antibodies prior to the first challenge. Neutralizing antibodies increased in all groups after exposure to the IHNV. Passive transfer of pooled sera from pCMV4-G vaccinates and IHN survivors provided relative survivals of 40 to 100% compared to fish injected with sera collected from fish immunized with control vaccines or left unhandled. In this study, DNA vaccination effectively protected Atlantic salmon smolts against challenges with IHNV.
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...
Kikumura, Akitoshi; Fang, Hao; Mun, Hye-Seong; Uemura, Noriko; Makino, Masayuki; Sayama, Yusuke; Norose, Kazumi; Aosai, Fumie
Toxoplasma gondii-derived heat shock protein 70 (T.g.HSP70) was proven to induce lethal anaphylactic reaction in T. gondii-infected mice through platelet-activating factor (PAF)-mediated, but not classical IgE-dependent, pathway via TLR4/MyD88 signal pathway. The effector cells generating PAF and causing T.g.HSP70-induced anaphylactic reaction were CD11b(+) and CD11c(+) cells, although the reaction was enhanced by marked IFN-gamma production by CD11b(+), CD11c(+), CD4(+) and CD8(+) splenocytes. In the present study, the effects of T.g.HSP70 gene vaccine targeting peripheral dendritic cells were evaluated against T.g.HSP70-induced anaphylactic reaction in T. gondii-infected mice. C57BL/6 mice receiving T.g.HSP70 gene vaccine showed prolonged survival. Platelets of peripheral blood, which completely disappeared during the T.g.HSP70-induced anaphylactic reaction, were partially restored with the T.g.HSP70 gene vaccination. The T.g.HSP70-induced marked production of PAF and IFN-gamma from splenocytes of infected mice during the T.g.HSP70-induced anaphylactic reaction was shown to decrease after the T.g.HSP70 gene vaccination. Thus, T.g.HSP70 gene vaccine induced protective immunity against T.g.HSP70-induced PAF-mediated lethal anaphylactic reaction in T. gondii-infected mice.
Bentancor, Leticia V.; Bilen, Marcos; Brando, Romina J. Fernández; Ramos, María Victoria; Ferreira, Luis C. S.; Ghiringhelli, Pablo D.; Palermo, Marina S.
Production of verocytotoxin or Shiga-like toxin (Stx), particularly Stx2, is the basis of hemolytic uremic syndrome, a frequently lethal outcome for subjects infected with Stx2-producing enterohemorrhagic Escherichia coli (EHEC) strains. The toxin is formed by a single A subunit, which promotes protein synthesis inhibition in eukaryotic cells, and five B subunits, which bind to globotriaosylceramide at the surface of host cells. Host enzymes cleave the A subunit into the A1 peptide, endowed with N-glycosidase activity to the 28S rRNA, and the A2 peptide, which confers stability to the B pentamer. We report the construction of a DNA vaccine (pStx2ΔAB) that expresses a nontoxic Stx2 mutated form consisting of the last 32 amino acids of the A2 sequence and the complete B subunit as two nonfused polypeptides. Immunization trials carried out with the DNA vaccine in BALB/c mice, alone or in combination with another DNA vaccine encoding granulocyte-macrophage colony-stimulating factor, resulted in systemic Stx-specific antibody responses targeting both A and B subunits of the native Stx2. Moreover, anti-Stx2 antibodies raised in mice immunized with pStx2ΔAB showed toxin neutralization activity in vitro and, more importantly, conferred partial protection to Stx2 challenge in vivo. The present vector represents the second DNA vaccine so far reported to induce protective immunity to Stx2 and may contribute, either alone or in combination with other procedures, to the development of prophylactic or therapeutic interventions aiming to ameliorate EHEC infection-associated sequelae. PMID:19176691
Ingolotti, Mariana; Kawalekar, Omkar; Shedlock, Devon J; Muthumani, Karuppiah; Weiner, David B
DNA vaccination has been of great interest since its discovery in the 1990s due to its ability to elicit both humoral and cellular immune responses. DNA vaccines consist of a DNA plasmid containing a transgene that encodes the sequence of a target protein from a pathogen under the control of a eukaryotic promoter. This revolutionary technology has proven to be effective in animal models and four DNA vaccine products have recently been approved for veterinary use. Although few DNA vaccines against bacterial infections have been tested, the results are encouraging. Because of their versatility, safety and simplicity a wider range of organisms can be targeted by these vaccines, which shows their potential advantages to public health. This article describes the mechanism of action of DNA vaccines and their potential use for targeting bacterial infections. In addition, it provides an updated summary of the methods used to enhance immunogenicity from codon optimization and adjuvants to delivery techniques including electroporation and use of nanoparticles. PMID:20624048
Tian, S; Liu, Z; Donahue, C; Noh, HS; Falo, LD; You, Z
DC engineered in vitro by DNA encoding OVAhsp70 and IL-15 up-regulated their expressions of CD80, CD86, CCR7 and IL-15Rα and promoted their productions of IL-6, IL-12 and TNF-α. Transcriptional IL-15-directed in vivo DC targeting DNA vaccine encoding OVAhsp70 elicited long-lasting Th1 and CTL responses and anti-B16OVA activity. CD8 T cell-mediated primary tumor protection was abrogated by DC or CD4 T cell depletion during the induction phase of immune responses. However, CD4 T cell depletion during immunization did not impair CD8 T cell-dependent long-lasting tumor protection. Furthermore, in vivo DC-derived IL-15 exerted the enhancements of cellular and humoral immune responses and antitumor immunity elicited by OVAhsp70 DNA vaccine. Importantly, the potency of this novel DNA vaccine strategy was proven using a self/tumor Ag (TRP2) in a clinically relevant B16 melanoma model. These findings have implications for developing next generation DNA vaccines against cancers and infectious diseases in both healthy and CD4 deficient individuals. PMID:19727134
Ahmed, Sami Ben Hadj; Bahloul, Chokri; Robbana, Cyrine; Askri, Souhir; Dellagi, Koussay
Over the past few years, several reports of DNA vaccines against murine cutaneous experimental leishmaniasis came out with promising but sometimes discordant results. The present studies were designed to compare, under similar conditions, the protective effects in the highly susceptible BALB/c mice of DNA vaccine candidates encoding to various Leishmania major antigens. The candidate DNA vaccines encode to the following antigens: LACK, PSA2, Gp63, LeIF and two newly identified p20 and Ribosomal like protein, in addition to different truncated portions of the LACK antigen. The most promising gene was LACK and it is more protective when it is used as a p24 truncated form. Furthermore, the presence of a tandem repeats of immunostimulating sequences (ISS) in the plasmid backbone played an important adjuvant effect in the observed protective effect induced by the DNA vaccine encoding to the LACKp24. Nevertheless, neither of the DNA vaccine candidates was able to mount a full protection in BALB/c mice challenged with a highly virulent L. major strain. Further improvements of the DNA vaccination approach are still needed to design a fully protective vaccine against leishmaniasis. Three directions of investigations are currently explored: DNA vaccines using a cocktail of antigens; Prime/Boost approach; and association of immune modulators with the candidate antigens.
Demuth, Peter C.; Min, Younjin; Huang, Bonnie; Kramer, Joshua A.; Miller, Andrew D.; Barouch, Dan H.; Hammond, Paula T.; Irvine, Darrell J.
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.
Use of S-[2,3-bispalmitoyiloxy-(2R)-propyl]-R-cysteinyl-amido-monomethoxy polyethylene glycol as an adjuvant improved protective immunity associated with a DNA vaccine encoding Cu,Zn superoxide dismutase of Brucella abortus in mice.
Retamal-Díaz, Angello; Riquelme-Neira, Roberto; Sáez, Darwin; Rivera, Alejandra; Fernández, Pablo; Cabrera, Alex; Guzmán, Carlos A; Oñate, Angel
This study was conducted to evaluate the immunogenicity and protective efficacy of a DNA vaccine encoding Brucella abortus Cu,Zn superoxide dismutase (SOD) using the Toll-like receptor 2/6 agonist S-[2,3-bispalmitoyiloxy-(2R)-propyl]-R-cysteinyl-amido-monomethoxy polyethylene glycol (BPPcysMPEG) as an adjuvant. Intranasal coadministration of BPPcysMPEG with a plasmid carrying the SOD-encoding gene (pcDNA-SOD) into BALB/c mice elicited antigen-specific humoral and cellular immune responses. Humoral responses were characterized by the stimulation of IgG2a and IgG1 and by the presence of SOD-specific secretory IgA in nasal and bronchoalveolar lavage fluids. Furthermore, T-cell proliferative responses and increased production of gamma interferon were also observed upon splenocyte restimulation with recombinant SOD. Cytotoxic responses were also stimulated, as demonstrated by the lysis of RB51-SOD-infected J774.A1 macrophages by cells recovered from immunized mice. The pcDNA-SOD/BPPcysMPEG formulation induced improved protection against challenge with the virulent strain B. abortus 2308 in BALB/c mice over that provided by pcDNA-SOD, suggesting the potential of this vaccination strategy against Brucella infection.
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.
Wang, Shuai; Wang, Yujian; Sun, Xiaoni; Zhang, Zhenchao; Liu, Tingqi; Gadahi, Javaid Ali; Xu, Lixin; Yan, Ruofeng; Song, Xiaokai; Li, Xiangrui
Toxoplasma gondii 10 kDa excretory-secretory antigen (TgESA10) is involved in the early stages of host invasion. The aim of this study was to evaluate the immune protective efficacy of a DNA vaccine encoding TgESA10 gene against acute T. gondii infection in mice. The gene sequence encoding TgESA10 was inserted into the eukaryotic expression vector pVAX I, and the efficacy of intramuscular vaccination of BALB/c mice with pVAX-ESA10 was analyzed. Mice immunized with pVAX-ESA10 elicited high titers of total IgG, IgG1, IgG2a, IgA and IgM antibodies, while IgE showed no changes. Analysis of cytokine profiles revealed significant increases of IFN-γ, IL-4 and IL-17, while no significant changes were detected in TGF-β1. Additionally, we found that pVAX-ESA10 enhanced the activation of CD4(+) and CD8(+) T cells and the expression of MHC-I and MHC-II molecules in spleen in mice. Immunization with pVAX-ESA10 significantly prolonged survival time (14.3 ± 1.7 days) after challenge infection with the virulent T. gondii RH strain, compared with the control groups which died within 8 days. These results suggested that TgESA10 DNA vaccine could trigger strong humoral and cellular responses and induce partial protection against acute toxoplasmosis.
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
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. PMID:25875109
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
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.
In general, vaccinations can be divided into standard and indication-related vaccinations. The former, which are mandatory for adults too, include tetanus and diphtheria, and those against influenza and pneumococci in people older than 60. The term indication vaccinations is used to describe those that are recommended in certain situations, for example, in the presence of a particular occupational risk, in certain underlying diseases, or particular situations, e.g. women of childbearing age, people living in regions endemic for meningoencephalitis, and drug addicts. Since the vaccination rate in the case of adults leaves much to be desired, all physicians should be required to ensure that "gaps" are closed and immunization protection is maintained.
Abbink, Peter; Larocca, Rafael A; De La Barrera, Rafael A; Bricault, Christine A; Moseley, Edward T; Boyd, Michael; Kirilova, Marinela; Li, Zhenfeng; Ng'ang'a, David; Nanayakkara, Ovini; Nityanandam, Ramya; Mercado, Noe B; Borducchi, Erica N; Agarwal, Arshi; Brinkman, Amanda L; Cabral, Crystal; Chandrashekar, Abishek; Giglio, Patricia B; Jetton, David; Jimenez, Jessica; Lee, Benjamin C; Mojta, Shanell; Molloy, Katherine; Shetty, Mayuri; Neubauer, George H; Stephenson, Kathryn E; Peron, Jean Pierre S; Zanotto, Paolo M de A; Misamore, Johnathan; Finneyfrock, Brad; Lewis, Mark G; Alter, Galit; Modjarrad, Kayvon; Jarman, Richard G; Eckels, Kenneth H; Michael, Nelson L; Thomas, Stephen J; Barouch, Dan H
Zika virus (ZIKV) is responsible for a major ongoing epidemic in the Americas and has been causally associated with fetal microcephaly. The development of a safe and effective ZIKV vaccine is therefore an urgent global health priority. Here we demonstrate that three different vaccine platforms protect against ZIKV challenge in rhesus monkeys. A purified inactivated virus vaccine induced ZIKV-specific neutralizing antibodies and completely protected monkeys against ZIKV strains from both Brazil and Puerto Rico. Purified immunoglobulin from vaccinated monkeys also conferred passive protection in adoptive transfer studies. A plasmid DNA vaccine and a single-shot recombinant rhesus adenovirus serotype 52 vector vaccine, both expressing ZIKV premembrane and envelope, also elicited neutralizing antibodies and completely protected monkeys against ZIKV challenge. These data support the rapid clinical development of ZIKV vaccines for humans. Copyright © 2016, American Association for the Advancement of Science.
Immunity at mucosal sites can prevent pathogen infection of the host. A) oral poliovirus vaccine B) inhaled influenza vaccine C) kennel cough & Newcastle...Utility of respiratory vaccination with recombinant subunit vaccines for protection against pneumonic plague. Douglas S. Reed & Jennifer Smoll...2. REPORT TYPE N/A 3. DATES COVERED - 4. TITLE AND SUBTITLE Utility of respiratory vaccination with recombinant subunit vaccines for
A Plasmodium vivax plasmid DNA- and adenovirus-vectored malaria vaccine encoding blood stage antigens AMA1 and MSP142 in a prime/boost heterologous immunization regimen partially protects Aotus monkeys against blood stage challenge.
Obaldia, Nicanor; Stockelman, Michael G; Otero, William; Cockrill, Jennifer A; Ganeshan, Harini; Abot, Esteban N; Zhang, Jianfeng; Limbach, Keith; Charoenvit, Yupin; Doolan, Denise L; Tang, De-Chu C; Richie, Thomas L
Malaria is caused by parasites of the genus Plasmodium that are transmitted to humans by the bites of Anopheles mosquitoes. After the elimination of P. falciparum it is predicted that Plasmodium vivax will remain an important cause of morbidity and mortality outside of Africa, stressing the importance of developing a vaccine against malaria. In this study we assess the immunogenicity and protective efficacy of two P. vivax antigens, AMA1 and MSP142 in a recombinant DNA plasmid prime/adenoviral vector (Ad) boost regimen in Aotus monkeys. Groups of 4 to 5 monkeys were immunized with DNA alone, Ad alone, prime/boost regimens of each antigen, prime/boost with both antigens, and empty vector controls, and then subjected to blood stage challenge. The heterologous immunization regimen with the antigen pair was more protective than either antigen alone or both antigens delivered with a single vaccine platform, based on their ability to induced the longest pre-patent period and time to peak parasitemia; the lowest peak and mean parasitemia; the smallest area under the parasitemia curve and the highest self-cured rate. Overall, pre-challenge MSP1 antibody titers strongly correlated with decreased parasite burden. Nevertheless, a significant proportion of immunized animals developed anemia. In conclusion, P. vivax plasmid DNA/Ad5 vaccine encoding blood stage parasite antigens AMA1 and MSP142 in a heterologous prime/boost immunization regimen, provided significant protection against blood-stage challenge in Aotus monkeys, indicating the suitability of these antigens and regimen for further development.
Okwor, Ifeoma; Mou, Zhirong; Liu, Dong; Uzonna, Jude
Although a great deal of knowledge has been gained from studies on the immunobiology of leishmaniasis, there is still no universally acceptable, safe, and effective vaccine against the disease. This strongly suggests that we still do not completely understand the factors that control and/or regulate the development and sustenance of anti-Leishmania immunity, particularly those associated with secondary (memory) immunity. Such an understanding is critically important for designing safe, effective, and universally acceptable vaccine against the disease. Here we review the literature on the correlate of protective anti-Leishmania immunity and vaccination strategies against leishmaniasis with a bias emphasis on experimental cutaneous leishmaniasis. PMID:22661975
Yu, Yun-Zhou; Li, Na; Wang, Wen-Bin; Wang, Shuang; Ma, Yao; Yu, Wei-Yuan; Sun, Zhi-Wei
Here, DNA replicon vaccines encoding the Hc domain of botulinum neurotoxin serotype A (AHc) or the receptor binding domain of anthrax protective antigen (PA4) with or without signal sequences were evaluated in mice. Strong antibody and protective responses were elicited only from AHc DNA vaccines with an Ig κ signal sequence or tissue plasminogen activator signal sequence. Meanwhile, there were no differences in total antibody responses or isotypes, lymphocyte proliferative responses, cytokine profiles and protective immune responses with the PA4 DNA vaccines with or without a signal sequence. Therefore, use of targeting sequences in designing DNA replicon vaccines depends on the specific antigen.
Plasmid DNA and protein vaccination of mice to the outer surface protein A of Borrelia burgdorferi leads to induction of T helper cells with specificity for a major epitope and augmentation of protective IgG antibodies in vivo.
Zhong, W; Wiesmüller, K H; Kramer, M D; Wallich, R; Simon, M M
Plasmid DNA-based vaccination is an efficient way to evoke various forms of protective immunity in laboratory animals. Our previous experiments have shown that mice immunized with either plasmid DNA encoding the outer surface lipoprotein A (pOspA) of Borrelia burgdorferi or the respective lipoprotein (Lip-OspA) produce protective antibodies against subsequent challenge with virulent spirochetes. In the present study, we compared the specificity and function of T cells generated in AKR/N mice previously immunized to either pOspA or Lip-OspA. T cell populations derived by either of the two protocols consistently responded by proliferation in vitro to one (residues 186-203; B4) out of a panel of 27 overlapping 20-mer peptides spanning the entire OspA molecule of strain ZS7. B4 was shown to express allele-specific ligand motifs for I-Ek. Most of the other peptides produced variable and much less pronounced or marginal proliferative T cell responses. T cells reactive to B4 as well as to some minor epitopes were CD4+CD8- T cells which produced IFN-gamma but no detectable IL-4 upon antigen stimulation in vitro. Priming of AKR/N mice with B4 but not with inactive peptides of OspA led to an enhanced production of IgG antibodies, mainly of the IgG1 isotype, including those to a prominent protective epitope (LA-2) upon subsequent challenge with Lip-OspA or intact spirochetes. The data demonstrate that both plasmid DNA and protein immunization with OspA results in T cell responses with specificity for a dominant OspA epitope and suggest that priming of mice with immunodominant peptides accelerates the appearance of protective antibodies in vivo. The identification of T helper cell epitopes relevant for the induction of protective antibodies will also facilitate the design of more potent vaccines against Lyme disease.
Gardiner, David F.; Rosenberg, Talia; Zaharatos, Jerry; Franco, David; Ho, David D
Clostridium difficile is a pathogen with increasing severity for which host antibody responses provide protection from disease. DNA vaccination has several advantages compared to traditional vaccine methods, however no study has examined this platform against C. difficile toxins. A synthetic gene was created encoding the receptor-binding domain (RBD) of C. difficile toxin A, optimized for expression in human cells. Gene expression was examined in vitro. Mice were inoculated then challenged with parenteral toxin A. Vaccination provided high titer antibodies and protected mice from death. This represents the first report of DNA vaccine inducing neutralizing antibodies to Clostridium difficile toxin A. PMID:19464540
Gardiner, David F; Rosenberg, Talia; Zaharatos, Jerry; Franco, David; Ho, David D
Clostridium difficile is a pathogen with increasing severity for which host antibody responses provide protection from disease. DNA vaccination has several advantages compared to traditional vaccine methods, however no study has examined this platform against C. difficile toxins. A synthetic gene was created encoding the receptor-binding domain (RBD) of C. difficile toxin A, optimized for expression in human cells. Gene expression was examined in vitro. Mice were inoculated and then challenged with parenteral toxin A. Vaccination provided high titer antibodies and protected mice from death. This represents the first report of DNA vaccine inducing neutralizing antibodies to C. difficile toxin A.
Vaccine protective efficacy is determined by multiple factors including host genetics, the type of vaccine, vaccine dosage, the virulence and dose of challenging viruses, and the interval between vaccination and viral challenge. Studies on human immune responses to vaccinations suggest host genetic...
Wang, Chun; Ge, Qing; Ting, David; Nguyen, David; Shen, Hui-Rong; Chen, Jianzhu; Eisen, Herman N.; Heller, Jorge; Langer, Robert; Putnam, David
Genetic vaccination using plasmid DNA presents a unique opportunity for achieving potent immune responses without the potential limitations of many conventional vaccines. Here we report the design of synthetic biodegradable polymers specifically for enhancing DNA vaccine efficacy in vivo. We molecularly engineered poly(ortho ester) microspheres that are non-toxic to cells, protect DNA from degradation, enable uptake by antigen-presenting cells, and release DNA rapidly in response to phagosomal pH. One type of microsphere of poly(ortho esters) that releases DNA vaccines in synchrony with the natural development of adaptive immunity, elicited distinct primary and secondary humoral and cellular immune responses in mice, and suppressed the growth of tumour cells bearing a model antigen. This polymer microparticulate system could, with further study, have implications for advancing the clinical utility of DNA vaccines as well as other nucleic-acid-based therapeutics against viral infections and cancer.
Kutzler, Michele A.; Weiner, David B.
Since the discovery, over a decade and a half ago, that genetically engineered DNA can be delivered in vaccine form and elicit an immune response, there has been much progress in understanding the basic biology of this platform. A large amount of data has been generated in preclinical model systems, and more sustained cellular responses and more consistent antibody responses are being observed in the clinic. Four DNA vaccine products have recently been approved, all in the area of veterinary medicine. These results suggest a productive future for this technology as more optimized constructs, better trial designs and improved platforms are being brought into the clinic. PMID:18781156
The eradication of smallpox, one of the great triumphs of medicine, was accomplished through the prophylactic administration of live vaccinia virus, a comparatively benign relative of variola virus, the causative agent of smallpox. Nevertheless, recent fears that variola virus may be used as a biological weapon together with the present susceptibility of unimmunized populations have spurred the development of new-generation vaccines that are safer than the original and can be produced by modern methods. Predicting the efficacy of such vaccines in the absence of human smallpox, however, depends on understanding the correlates of protection. This review outlines the biology of poxviruses with particular relevance to vaccine development, describes protein targets of humoral and cellular immunity, compares animal models of orthopoxvirus disease with human smallpox, and considers the status of second- and third-generation smallpox vaccines. Published 2010. This article is a US Government work and is in the public domain in the USA.
Zakhartchouk, Alexander N; Viswanathan, Sathiyanarayanan; Moshynskyy, Igor; Petric, Martin; Babiuk, Lorne A
Severe acute respiratory syndrome coronavirus (SARS-CoV) first appeared in Southern China in November 2002, and then quickly spread to 33 countries on five continents along international air travel routes. Although the SARS epidemic has been contained, there is a clear need for a safe and effective vaccine should an outbreak of a SARS-CoV infection reappear in human population. In this study, we tested four DNA-vaccine constructs: (1) pLL70, containing cDNA for the SARS-CoV spike (S) gene; (2) pcDNA-SS, containing codon-optimized S gene for SARS-CoV S protein (residues 12-1255) fused with a leader sequence derived from the human CD5 gene; (3) pcDNA-St, containing the gene encoding the N-portion of the codon-optimized S gene (residues 12-532) with the CD5 leader sequence; (4) pcDNA-St-VP22C, containing the gene encoding the N-portion of the codon-optimized S protein with the CD5 leader sequence fused with the C-terminal 138 amino acids of the bovine herpesvirus-1 (BHV-1) major tegument protein VP22. Each of these plasmids was intradermally administered to C57BL/6 mice in three separate immunizations. Analysis of humoral and cellular immune responses in immunized mice demonstrated that pcDNA-SS and pcDNA-St-VP22C are the most immunogenic SARS vaccine candidates.
Aps, Luana R M M; Diniz, Mariana O; Porchia, Bruna F M M; Sales, Natiely S; Moreno, Ana Carolina R; Ferreira, Luís C S
Recently, Bacillus subtilis spores were shown to be endowed with strong adjuvant capacity when co-administered with purified antigenic proteins. In the present study we assessed whether spores possess adjuvant properties when combined with DNA vaccines. We showed that B. subtilis spores promoted the activation of dendritic cells in vitro and induced migration of pro-inflammatory cells after parenteral administration to mice. Likewise, co-administration of spores with a DNA vaccine encoding the human papillomavirus type 16 (HPV-16) E7 protein enhanced the activation of antigen-specific CD8(+) T cell responses in vivo. Mice immunized with the DNA vaccine admixed with spores presented a protective immunity increase to previously implanted tumor cells, capable of expressing HPV-16 oncoproteins. Finally, we observed that the adjuvant effect can vary accordingly to the number of co-administered spores which may be ascribed with the ability to induce. Collectively, the present results demonstrate for the first time that B. subtilis spores can also confer adjuvant effects to DNA vaccines.
Yang, Yaping; Yang, Xiaodi; Gu, Yuan; Wang, Yunyun; Zhao, Xi; Zhu, Xinping
Ts87 is an immunodominant antigen that induces protective immunity against Trichinella spiralis larval challenge in mice. To determine if a combination of recombinant Ts87 protein and its coding DNA induces a stronger immune response in female C57BL/6 mice were immunized with 100 μg of recombinant Ts87 protein plus its coding DNA cloned in vector pVAX1, or the same amount of recombinant protein or DNA only. Mouse subclass IgG responses showed that both co-immunized and single-immunized mice produced a balanced IgG2a/IgG1 (Th1/Th2) response. T-cell proliferation in co-immunized animals was significantly higher than in single-immunized mice. Cytokine profiling in the co-immunization group showed a significant increase in the levels of IL-2, IL-4, IL-6 and IFN-γ in the splenocytes of mice upon stimulation with the recombinant Ts87 protein; however, the expression of IL-17 was down-regulated. Challenge results showed that mice immunized with the recombinant Ts87 protein and its coding DNA produced reduced the muscle larval burden to a greater extent (43.8%) than the groups immunized with only the protein (39.7%) or the DNA (9.7%). A better Th1/Th2 immune response and consequent protection induced by co-immunization with the recombinant Ts87 protein and its coding DNA may result from an adjuvant effect of DNA and a specific persistent expression of Ts87.
Pereiro, Patricia; Dios, Sonia; Boltaña, Sebastián; Coll, Julio; Estepa, Amparo; Mackenzie, Simon; Novoa, Beatriz; Figueras, Antonio
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
Jathoul, Amit P; Holley, Jane L; Garmory, Helen S
DNA vaccines which expressed the Hc fragment of the Clostridium botulinum type F neurotoxin (BoNT/F Hc) fused to a signal peptide downstream of four different eukaryotic promoters were prepared. Subsequently, the immunogenicity of the DNA vaccines and protection afforded in mice against challenge with 10(4) MLD of type F botulinum toxin was evaluated. The DNA vaccine containing the human ubiquitin gene (UbC) promoter induced the highest BoNT/F Hc-specific antibody concentration following two intramuscular immunisations and afforded 90% protection against challenge. The results from this study indicate that the selection of promoter used in DNA vaccination studies may be of importance in designing optimised vaccines.
Abbink, Peter; Larocca, Rafael A.; De La Barrera, Rafael A.; Bricault, Christine A.; Moseley, Edward T.; Boyd, Michael; Kirilova, Marinela; Li, Zhenfeng; Ng’ang’a, David; Nanayakkara, Ovini; Nityanandam, Ramya; Mercado, Noe B.; Borducchi, Erica N.; Agarwal, Arshi; Brinkman, Amanda L.; Cabral, Crystal; Chandrashekar, Abishek; Giglio, Patricia B.; Jetton, David; Jimenez, Jessica; Lee, Benjamin C.; Mojta, Shanell; Molloy, Katherine; Shetty, Mayuri; Neubauer, George H.; Stephenson, Kathryn E.; Peron, Jean Pierre S.; Zanotto, Paolo M. de A.; Misamore, Johnathan; Finneyfrock, Brad; Lewis, Mark G.; Alter, Galit; Modjarrad, Kayvon; Jarman, Richard G.; Eckels, Kenneth H.; Michael, Nelson L.; Thomas, Stephen J.; Barouch, Dan H.
Zika virus (ZIKV) is responsible for a major ongoing epidemic in the Americas and has been causally associated with fetal microcephaly. The development of a safe and effective ZIKV vaccine is therefore an urgent global health priority. Here we demonstrate that three different vaccine platforms protect against ZIKV challenge in rhesus monkeys. A purified inactivated virus vaccine induced ZIKV-specific neutralizing antibodies and completely protected monkeys against ZIKV strains from both Brazil and Puerto Rico. Purified immunoglobulin from vaccinated monkeys conferred passive protection in adoptive transfer studies. A plasmid DNA vaccine and a single-shot recombinant rhesus adenovirus serotype 52 vector expressing ZIKV prM-Env also elicited neutralizing antibodies and completely protected monkeys against ZIKV challenge. These data support the rapid clinical development of ZIKV vaccines for humans. PMID:27492477
Zhu, Fujie; Liu, Xiao; Sun, Zhenhong; Yu, Cuilian; Liu, Liping; Yang, Shifa; Li, Bing; Wei, Kai; Zhu, Ruiliang
Bordetella avium is the causative agent of bordetellosis, which remains to be the cause of severe losses in the turkey industry. Given the lack of vaccines that can provide good protection, developing a novel vaccine against B. avium infection is crucial. In this study, we constructed a eukaryotic expression plasmid, which expressed the outer membrane protein A (ompA) of B. avium, to prepare a B. avium recombinant ompA-DNA vaccine. Three concentrations (low, middle, and high) of Taishan Pinus massoniana pollen polysaccharides (TPPPS), a known immunomodulator, were used as adjuvants, and their immune conditioning effects on the developed DNA vaccine were examined. The pure ompA-DNA vaccine, Freund’s incomplete adjuvant ompA-DNA vaccine, and the empty plasmid served as the controls. The chickens in each group were separately inoculated with these vaccines three times at 1, 7, and 14 days old. Dynamic changes in antibody production, cytokine secretion, and lymphocyte count were then determined from 7 to 49 days after the first inoculation. Protective rates of the vaccines were also determined after the third inoculation. Results showed that the pure DNA vaccine obviously induced the production of antibodies, the secretion of cytokines, and the increase in CD4+ and CD8+ T lymphocyte counts in peripheral blood, as well as provided a protective rate of 50% to the B. avium-challenged chickens. The chickens inoculated with the TPPPS adjuvant ompA-DNA vaccine and Freund’s adjuvant ompA-DNA vaccine demonstrated higher levels of immune responses than those inoculated with pure ompA-DNA vaccine, whereas only the ompA-DNA vaccine with 200 mg/mL TPPPS completely protected the chickens against B. avium infection. These findings indicate that the B. avium ompA-DNA vaccine combined with TPPPS is a potentially effective B. avium vaccine. PMID:26870023
Shuaibu, M N; Kikuchi, M; Cherif, M S; Helegbe, G K; Yanagi, T; Hirayama, K
Following a genome-wide search for a blood stage malaria DNA-based vaccine using web-based bioinformatic tools, 29 genes from the annotated Plasmodium yoelii genome sequence (www.PlasmoDB.org and www.tigr.org) were identified as encoding GPI-anchored proteins. Target genes were those with orthologues in P. falciparum, containing an N-terminal signal sequence containing hydrophobic amino acid stretch and signal P criteria, a transmembrane-like domain and GPI anchor motif. Focusing on the blood stage, we extracted mRNA from pRBCs, PCR-amplified 22 out of the 29 selected genes, and eventually cloned nine of these into a DNA vaccine plasmid, pVAX 200-DEST. Biojector-mediated delivery of the nine DNA vaccines was conducted using ShimaJET to C57BL/6 mice at a dose of 4 μg/mouse three times at an interval of 3 weeks. Two weeks after the second booster, immunized mice were challenged with P. y. yoelii 17XL-parasitized RBCs and the level of parasitaemia, protection and survival was assessed. Immunization with one gene (PY03470) resulted in 2-4 days of delayed onset and level of parasitaemia and was associated with increased survival compared to non-immunized mice. Antibody production was, however, low following DNA vaccination, as determined by immunofluorescence assay. Recombinant protein from this gene, GPI8p transamidase-related protein (rPyTAM) in PBS or emulsified with GERBU adjuvant was also used to immunize another set of C57BL/6 mice with 10-20 μg/mouse three times at 3-week interval. Higher antibody response was obtained as determined by ELISA with similar protective effects as observed after DNA vaccination.
Racz, Rebecca; He, Yongqun
A DNA vaccine is a vaccine that uses a mammalian expression vector to express one or more protein antigens and is administered in vivo to induce an adaptive immune response. Since the 1990s, a significant amount of research has been performed on DNA vaccines and the mechanisms behind them. To meet the needs of the DNA vaccine research community, we created DNAVaxDB ( http://www.violinet.org/dnavaxdb ), the first Web-based database and analysis resource of experimentally verified DNA vaccines. All the data in DNAVaxDB, which includes plasmids, antigens, vaccines, and sources, is manually curated and experimentally verified. This chapter goes over the detail of DNAVaxDB system and shows how the DNA vaccine database, combined with the Vaxign vaccine design tool, can be used for rational design of a DNA vaccine against a pathogen, such as Mycobacterium bovis.
Kumar, A; Samant, M
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.
Niborski, V; Li, Y; Brennan, F; Lane, M; Torché, A M; Remond, M; Bonneau, M; Riffault, S; Stirling, C; Hutchings, G; Takamatsu, H; Barnett, P; Charley, B; Schwartz-Cornil, I
As an alternative strategy to classical inactivated viral vaccine against FMDV, naked DNA vaccine is attractive because of safety, flexibility and low cost. However DNA vaccination is usually poorly efficient in target species. Indeed we found that naked DNA plasmids encoding for P1-2A3C3D and GM-CSF proteins did not induce any detectable immunity against FMDV in sheep. Interestingly, we demonstrate herein that formulations of DNA on poly(D,L-lactide-co-glycolide) (PLG) or in lipofectin triggered divergent types of immune responses: PLG stimulated a T cell response and could elicit significant neutralising antibody titers, whereas lipofectin generated even higher antibody titers but no significant T cell response. The DNA/PLG regimen used in five sheep protected against clinical symptoms and viraemia and prevented the carrier state in four of them. Thus formulated DNA can be remarkably efficient against FMDV in a ruminant species that is usually refractory to DNA vaccination.
Hainz, Ursula; Aigner, Kurt; Asch, Esther; Berger, Peter; Böhmer, Franz; Feldkircher, Bruno; Horwath, Brigitte; Jenewein, Brigitte; Kassal, Hermann; Kistner, Otfried; Mack, Hans; Pfeiffer, Karl-Peter; Pils, Katharina; Plank, Johannes; Renner, Doris; Saurwein-Teissl, Maria; Schwanzer, Erhard; Trieb, Klemens; Grubeck-Loebenstein, Beatrix
The aim of the study was to evaluate, if elderly persons are sufficiently protected against infectious diseases by vaccination. 300 elderly (> 60 years) and 300 young (< 35 years) persons from five Austrian cities were recruited according to the criteria of a field study. Antibody concentrations against tetanus, diphtheria, tickborne encephalitis and influenza were assessed by ELISA or by haemagglutination inhibition test. Disease and vaccination histories were recorded. The results of the study demonstrate that protection against infectious diseases was frequently insufficient in the elderly. This was partly due to the fact that old persons were not vaccinated according to recommended strategies. However, low antibody concentration and a short duration of protective humoral immunity were also observed in many elderly persons in spite of regular vaccination. This was not only the case in frail, but also in healthy elderlies. The data demonstrate that vaccination has a relatively weak and short-lasting effect in old age. The results of the study should stimulate discussions about strategies how vaccinations can be made more effective in old age. Improved campaigns, shortened vaccination intervals as well as the design of novel vaccines tailored to fulfill the specific demands of the aging immune system are imaginable.
Perforin and gamma interferon expression are required for CD4+ and CD8+ T-cell-dependent protective immunity against a human parasite, Trypanosoma cruzi, elicited by heterologous plasmid DNA prime-recombinant adenovirus 5 boost vaccination.
de Alencar, Bruna C G; Persechini, Pedro M; Haolla, Filipe A; de Oliveira, Gabriel; Silverio, Jaline C; Lannes-Vieira, Joseli; Machado, Alexandre V; Gazzinelli, Ricardo T; Bruna-Romero, Oscar; Rodrigues, Mauricio M
A heterologous prime-boost strategy using plasmid DNA, followed by replication-defective recombinant adenovirus 5, is being proposed as a powerful way to elicit CD4(+) and CD8(+) T-cell-mediated protective immunity against intracellular pathogens. We confirmed this concept and furthered existing research by providing evidence that the heterologous prime-boost regimen using the gene encoding amastigote surface protein 2 elicited CD4(+) and CD8(+) T-cell-mediated protective immunity (reduction of acute parasitemia and prolonged survival) against experimental infection with Trypanosoma cruzi. Protective immunity correlated with the presence of in vivo antigen-specific cytotoxic activity prior to challenge. Based on this, our second goal was to determine the outcome of infection after heterologous prime-boost immunization of perforin-deficient mice. These mice were highly susceptible to infection. A detailed analysis of the cell-mediated immune responses in immunized perforin-deficient mice showed an impaired gamma interferon (IFN-gamma) secretion by immune spleen cells upon restimulation in vitro with soluble recombinant antigen. In spite of a normal numeric expansion, specific CD8(+) T cells presented several functional defects detected in vivo (cytotoxicity) and in vitro (simultaneous expression of CD107a/IFN-gamma or IFN-gamma/tumor necrosis factor alpha) paralleled by a decreased expression of CD44 and KLRG-1. Our final goal was to determine the importance of IFN-gamma in the presence of highly cytotoxic T cells. Vaccinated IFN-gamma-deficient mice developed highly cytotoxic cells but failed to develop any protective immunity. Our study thus demonstrated a role for perforin and IFN-gamma in a number of T-cell-mediated effector functions and in the antiparasitic immunity generated by a heterologous plasmid DNA prime-adenovirus boost vaccination strategy.
Dale, C Jane; De Rose, Robert; Wilson, Kim M; Croom, Hayley A; Thomson, Scott; Coupar, Barbara E H; Ramsay, Alistair; Purcell, Damian F J; Ffrench, Rosemary; Law, Matthew; Emery, Sean; Cooper, David A; Ramshaw, Ian A; Boyle, David B; Kent, Stephen J
Induction of HIV-specific T-cell responses by vaccines may facilitate efficient control of HIV. Plasmid DNA vaccines and recombinant fowlpoxvirus (rFPV) vaccines are promising HIV-1 vaccine candidates, although either vaccine alone may be insufficient to protect against HIV-1. A consecutive immunisation strategy involving priming with DNA and boosting with rFPV vaccines encoding multiple common HIV-1 antigens was further evaluated in 30 macaques. The DNA vaccine vector included CpG immunostimulatory molecules, and rFPV vaccines were compared with rFPV vaccines co-expressing the pro-T cell cytokines IFNgamma or IL-12. Vaccines expressed multiple HIV-1 genes, mutated to remove active sites of the HIV proteins. The vaccines were well tolerated, and a significant enhancement of DNA-vaccine primed HIV-1 specific T lymphocyte responses was observed following rFPV boosting. Co-expression of IFNgamma or IL-12 by the rFPV vaccines did not further enhance immune responses. Non-sterilising protection from a non-pathogenic HIV-1 challenge was observed. This study provides evidence of a safe, optimised, strategy for the generation of T-cell mediated immunity to HIV-1.
Zheng, Mingquan; Ramsay, Alistair J.; Robichaux, Myles B.; Norris, Karen A.; Kliment, Corrine; Crowe, Christopher; Rapaka, Rekha R.; Steele, Chad; McAllister, Florencia; Shellito, Judd E.; Marrero, Luis; Schwarzenberger, Paul; Zhong, Qiu; Kolls, Jay K.
Depletion or dysfunction of CD4+ T lymphocytes profoundly perturbs host defenses and impairs immunogenicity of vaccines. Here, we show that plasmid DNA vaccination with a cassette encoding antigen (OVA) and a second cassette encoding full-length CD40 ligand (CD40L), a molecule expressed on activated CD4+ T lymphocytes and critical for T cell helper function, can elicit significant titers of antigen-specific immunoglobulins in serum and Tc1 CD8+ T cell responses in CD4-deficient mice. To investigate whether this approach leads to CD4+ T cell–independent vaccine protection against a prototypic AIDS-defining infection, Pneumocystis (PC) pneumonia, we used serum from mice vaccinated with PC-pulsed, CD40L-modifed DCs to immunoprecipitate PC antigens. Kexin, a PC antigen identified by this approach, was used in a similar DNA vaccine strategy with or without CD40L. CD4-deficient mice receiving DNA vaccines encoding Kexin and CD40L showed significantly higher anti-PC IgG titers as well as opsonic killing of PC compared with those vaccinated with Kexin alone. Moreover, CD4-depleted, Kexin-vaccinated mice showed a 3-log greater protection in a PC challenge model. Adoptive transfer of CD19+ cells or IgG to SCID mice conferred protection against PC challenge, indicating a role of humoral immunity in the protection. The results of these studies show promise for CD4-independent vaccination against HIV-related or other opportunistic pathogens. PMID:16308571
Kim, C H; Johnson, M C; Drennan, J D; Simon, B E; Thomann, E; Leong, J A
Protective immunity by vaccination with plasmid DNA encoding a viral glycoprotein (G) has long been assumed to result from the induction of a specific immune response. We report here that the initial protection may be due to the induction of alpha/beta interferon, with long-term protection due to a specific response to the encoded viral G. DNA vaccines encoding the Gs of three serologically unrelated fish rhabdoviruses were used to vaccinate rainbow trout against a lethal challenge with infectious hematopoietic necrosis virus (IHNV). All three vaccines, each encoding the G gene of either IHNV (IHNV-G), snakehead rhabdovirus (SHRV) (SHRV-G), or spring viremia of carp virus (SVCV) (SVCV-G), elicited protective immunity against IHNV. Vaccinated fish were challenged at 30 or 70 days postvaccination with lethal doses of IHNV. At 30 days postvaccination, only 5% of fish that had received any of the G vaccines died, whereas more than 50% of the control fish succumbed to virus challenge. When fish were vaccinated and challenged at 70 days postvaccination, only 12% of the IHNV-G-vaccinated fish died compared to 68% for the SHRV-G- and 76% for the SVCV-G-vaccinated fish. Assays for trout Mx protein, an indicator of alpha/beta interferon induction, showed that only fish vaccinated with a G-containing plasmid produced high levels of Mx protein in the kidneys and liver. Interestingly, at day 7 after virus challenge, all of the fish vaccinated with the IHNV-G plasmid were negative for Mx, but the SHRV-G- and SVCV-G-vaccinated fish still showed detectable levels of Mx. These results suggest that DNA vaccines in fish induce an early, nonspecific antiviral protection mediated by an alpha/beta interferon and, later, a specific immune response.
Meunier, Marine; Chemaly, Marianne; Dory, Daniel
DNA vaccination is a promising alternative strategy for developing new human and animal vaccines. The massive efforts made these past 25 years to increase the immunizing potential of this kind of vaccine are still ongoing. A relatively small number of studies concerning poultry have been published. Even though there is a need for new poultry vaccines, five parameters must nevertheless be taken into account for their development: the vaccine has to be very effective, safe, inexpensive, suitable for mass vaccination and able to induce immune responses in the presence of maternal antibodies (when appropriate). DNA vaccination should meet these requirements. 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. Second, 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 backbones and codon usage; association of vaccine antigens and finally, heterologous prime-boost regimens. The final part will indicate additional 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.
Yu, Longzheng; Yamagishi, Junya; Zhang, Shoufa; Jin, Chunmei; Aboge, Gabriel Oluga; Zhang, Houshuang; Zhang, Guohong; Tanaka, Tetsuya; Fujisaki, Kozo; Nishikawa, Yoshifumi; Xuan, Xuenan
A heterologous prime-boost strategy with priming plasmid DNA followed by recombinant virus expressing relevant antigens is known to stimulate protective immunity against intracellular parasites. In this study, we have evaluated a heterologous prime-boost strategy for immunizing mice against Toxoplasma gondii infection. Our results revealed that the prime-boost strategy using both plasmid DNA and adenoviral vector encoding TgAMA1 may stimulate both humoral and Th1/Th2 cellular immune responses specific for TgAMA1. Moreover, C57BL/6 mice immunized with the pAMA1/Ad5Null, pNull/Ad5AMA1, and pAMA1/Ad5AMA1 constructs showed survival rates of 12.5%, 37.5%, and 50%, respectively. In contrast, all the pNull/Ad5Null immunized mice died after infection with the PLK-GFP strain of T. gondii. Brain cyst burden was reduced by 23% in mice immunized with pAMA1/Ad5AMA1 compared with the pNull/Ad5AMA1 immunized mice. These results demonstrate that the heterologous DNA priming and recombinant adenovirus boost strategy may provide protective immunity against T. gondii infection.
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
Petrini, Stefano; Ramadori, Giorgio; Villa, Riccardo; Borghetti, Paolo; de Angelis, Elena; Cantoni, Anna Maria; Corradi, Attilio; Amici, Augusto; Ferrari, Maura
In veterinary medicine, there have been different experiences with the plasmid DNA vaccination. In this area and with the hypothesis to demonstrate the effectiveness of different plasmids encoding porcine respiratory and reproductive syndrome (PRRS), five DNA vaccines against PRRS were evaluated for their innocuity and efficacy in pigs. Eighteen animals were divided into five groups which were injected with five (A, B, C, D, E) different DNA vaccines. Albeit, none of the proposed vaccines were able to protect the animals against PRRS virus. Only vaccines A and B were able to reduce the clinical signs of the infection. ELISA IgM were detected 30 days after the first vaccination in the pigs injected by Vaccine A or B. ELISA IgG were detected 90 days after the first vaccination in the pigs injected by Vaccine B or C. Neutralizing antibody were detected Post Challenge Days 61 (PCD) in all groups. In the pigs inoculated with Vaccine C, IFN-γ were detected 90 days after first vaccination, and after challenge exposure they increased. In the other groups, the IFN-γ were detected after challenge infection. Pigs injected with each of the vaccines A, B, C, D and E showed a significantly higher level of CD4−CD8+ lymphocytes (p < 0.001) after infection in comparison with their controls. PMID:26344342
Schautteet, Katelijn; De Clercq, Evelien; Jönsson, Yannick; Lagae, Stefanie; Chiers, Koen; Cox, Eric; Vanrompay, Daisy
The current study evaluates combined aerosol-vaginal delivery of a MOMP-based Chlamydia trachomatis (serovar E) DNA vaccine in a pig genital challenge model. Most non-replicating antigens are rather poor mucosal immunogens in comparison to replicating antigens. Therefore, a mucosal administered DNA vaccine, which actually mimics a live vaccine, could be promising. Protection was promoted by plasmids encoding the porcine granulocyte macrophage-colony stimulating factor (pcDNA3.1zeo::GM-CSF), the Escherichia coli thermo-labile enterotoxin (LT) subunit A (plasmid PJV2004::LTa) and subunit B (plasmid PJV2005::LTb). Mucosal C. trachomatis DNA vaccination induced significant protection against genital C. trachomatis challenge although the infection could not be eradicated. Intradermal immunization was significantly less efficient in protecting experimentally infected pigs. Protection was correlated with efficient T cell priming and significantly higher serum IgA titers following primo vaccination. Copyright © 2012 Elsevier Ltd. All rights reserved.
Heterologous Plasmid DNA Prime-Recombinant Human Adenovirus 5 Boost Vaccination Generates a Stable Pool of Protective Long-Lived CD8+ T Effector Memory Cells Specific for a Human Parasite, Trypanosoma cruzi▿†
Rigato, Paula Ordonhez; de Alencar, Bruna C.; de Vasconcelos, José Ronnie C.; Dominguez, Mariana R.; Araújo, Adriano F.; Machado, Alexandre V.; Gazzinelli, Ricardo T.; Bruna-Romero, Oscar; Rodrigues, Mauricio M.
Recently, we described a heterologous prime-boost strategy using plasmid DNA followed by replication-defective human recombinant adenovirus type 5 as a powerful strategy to elicit long-lived CD8+ T-cell-mediated protective immunity against experimental systemic infection of mice with a human intracellular protozoan parasite, Trypanosoma cruzi. In the present study, we further characterized the protective long-lived CD8+ T cells. We compared several functional and phenotypic aspects of specific CD8+ T cells present 14 or 98 days after the last immunizing dose and found the following: (i) the numbers of specific cells were similar, as determined by multimer staining or by determining the number of gamma interferon (IFN-γ)-secreting cells by enzyme-linked immunospot (ELISPOT) assay; (ii) these cells were equally cytotoxic in vivo; (iii) following in vitro stimulation, a slight decline in the frequency of multifunctional cells (CD107a+ IFN-γ+ or CD107a+ IFN-γ+ tumor necrosis factor alpha positive [TNF-α+]) was paralleled by a significant increase of CD107a singly positive cells after 98 days; (iv) the expression of several surface markers was identical, except for the reexpression of CD127 after 98 days; (v) the use of genetically deficient mice revealed a role for interleukin-12 (IL-12)/IL-23, but not IFN-γ, in the maintenance of these memory cells; and (vi) subsequent immunizations with an unrelated virus or a plasmid vaccine or the depletion of CD4+ T cells did not significantly erode the number or function of these CD8+ T cells during the 15-week period. From these results, we concluded that heterologous plasmid DNA prime-adenovirus boost vaccination generated a stable pool of functional protective long-lived CD8+ T cells with an effector memory phenotype. PMID:21357719
Firouzamandi, Masoumeh; Moeini, Hassan; Hosseini, Seyed Davood; Bejo, Mohd Hair; Omar, Abdul Rahman; Mehrbod, Parvaneh; El Zowalaty, Mohamed E; Webster, Thomas J; Ideris, Aini
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 (P<0.05) in specific pathogen-free (SPF) chickens at 3 and 4 weeks postvaccination compared to 2 weeks postvaccination. Hemagglutination inhibition (HI) titer was not significantly different between groups injected with 40 μg pDNA + 64 μg D-SPM and 40 μg pDNA at 4 weeks postvaccination (P>0.05). 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.
Ligtenberg, M A; Witt, K; Galvez-Cancino, F; Sette, A; Lundqvist, A; Lladser, A; Kiessling, R
Metastatic melanoma is a fatal disease that responds poorly to classical treatments but can be targeted by T cell-based immunotherapy. Cancer vaccines have the potential to generate long-lasting cytotoxic CD8(+) T cell responses able to eradicate established and disseminated tumors. Vaccination against antigens expressed by tumor cells with enhanced metastatic potential represents a highly attractive strategy to efficiently target deadly metastatic disease. Cripto-1 is frequently over-expressed in human carcinomas and melanomas, but is expressed only at low levels on normal differentiated tissues. Cripto-1 is particularly upregulated in cancer-initiating cells and is involved in cellular processes such as cell migration, invasion and epithelial-mesenchymal transition, which are hallmarks of aggressive cancer cells able to initiate metastatic disease. Here, we explored the potential of Cripto-1 vaccination to target metastatic melanoma in a preclinical model. Cripto-1 was overexpressed in highly metastatic B16F10 cells as compared to poorly metastatic B16F1 cells. Moreover, B16F10 cells grown in sphere conditions to enrich for cancer stem cells (CSC) progressively upregulated cripto1 expression. Vaccination of C57Bl/6 mice with a DNA vaccine encoding mouse Cripto-1 elicited a readily detectable/strong cytotoxic CD8(+) T cell response specific for a H-2 Kb-restricted epitope identified based on its ability to bind H-2(b) molecules. Remarkably, Cripto-1 vaccination elicited a protective response against lung metastasis and subcutaneous challenges with highly metastatic B16F10 melanoma cells. Our data indicate that vaccination against Cripto-1 represents a novel strategy to be tested in the clinic.
Ligtenberg, M. A.; Witt, K.; Galvez-Cancino, F.; Sette, A.; Lundqvist, A.; Lladser, A.; Kiessling, R.
ABSTRACT Metastatic melanoma is a fatal disease that responds poorly to classical treatments but can be targeted by T cell-based immunotherapy. Cancer vaccines have the potential to generate long-lasting cytotoxic CD8+ T cell responses able to eradicate established and disseminated tumors. Vaccination against antigens expressed by tumor cells with enhanced metastatic potential represents a highly attractive strategy to efficiently target deadly metastatic disease. Cripto-1 is frequently over-expressed in human carcinomas and melanomas, but is expressed only at low levels on normal differentiated tissues. Cripto-1 is particularly upregulated in cancer-initiating cells and is involved in cellular processes such as cell migration, invasion and epithelial–mesenchymal transition, which are hallmarks of aggressive cancer cells able to initiate metastatic disease. Here, we explored the potential of Cripto-1 vaccination to target metastatic melanoma in a preclinical model. Cripto-1 was overexpressed in highly metastatic B16F10 cells as compared to poorly metastatic B16F1 cells. Moreover, B16F10 cells grown in sphere conditions to enrich for cancer stem cells (CSC) progressively upregulated cripto1 expression. Vaccination of C57Bl/6 mice with a DNA vaccine encoding mouse Cripto-1 elicited a readily detectable/strong cytotoxic CD8+ T cell response specific for a H-2 Kb-restricted epitope identified based on its ability to bind H-2b molecules. Remarkably, Cripto-1 vaccination elicited a protective response against lung metastasis and subcutaneous challenges with highly metastatic B16F10 melanoma cells. Our data indicate that vaccination against Cripto-1 represents a novel strategy to be tested in the clinic. PMID:27467944
Heithoff, Douglas M; House, John K; Thomson, Peter C; Mahan, Michael J
Intensive livestock production is associated with increased Salmonella exposure, transmission, animal disease, and contamination of food and water supplies. Modified live Salmonella enterica vaccines that lack a functional DNA adenine methylase (Dam) confer cross-protection to a diversity of salmonellae in experimental models of murine, avian, ovine, and bovine models of salmonellosis. However, the commercial success of any vaccine is dependent upon the therapeutic index, the ratio of safety/efficacy. Herein, secondary virulence-attenuating mutations targeted to genes involved in intracellular and/or systemic survival were introduced into Salmonella dam vaccines to screen for vaccine candidates that were safe in the animal and the environment, while maintaining the capacity to confer cross-protective immunity to pathogenic salmonellae serotypes. Salmonella dam mgtC, dam sifA, and dam spvB vaccine strains exhibited significantly improved vaccine safety as evidenced by the failure to give rise to virulent revertants during the infective process, contrary to the parental Salmonella dam vaccine. Further, these vaccines exhibited a low grade persistence in host tissues that was associated with reduced vaccine shedding, reduced environmental persistence, and induction of cross-protective immunity to pathogenic serotypes derived from infected livestock. These data indicate that Salmonella dam double mutant vaccines are suitable for commercial applications against salmonellosis in livestock production systems. Reducing pre-harvest salmonellae load through vaccination will promote the health and productivity of livestock and reduce contamination of livestock-derived food products, while enhancing overall food safety.
Thomas, Sunil; Luxon, Bruce A
Vaccines elicit immune responses, provide protection against microorganisms and are considered as one of the most successful medical interventions against infectious diseases. Vaccines can be produced using attenuated virus or bacteria, recombinant proteins, bacterial polysaccharides, carbohydrates or plasmid DNA. Conventional vaccines rely on the induction of immune responses against antigenic proteins to be effective. The genetic diversity of microorganisms, coupled with the high degree of sequence variability in antigenic proteins, presents a challenge to developing broadly effective conventional vaccines. The observation that whole protein antigens are not necessarily essential for inducing immunity has led to the emergence of a new branch of vaccine design termed 'structural vaccinology'. Structure-based vaccines are designed on the rationale that protective epitopes should be sufficient to induce immune responses and provide protection against pathogens. Recent studies demonstrated that designing structure-based vaccine candidates with multiple epitopes induce a higher immune response. As yet there are no commercial vaccines available based on structure-based design and most of the structure-based vaccine candidates are in the preclinical stages of development. This review focuses on recent advances in structure-based vaccine candidates and their application in providing protection against infectious diseases.
Meng, Qinglai; Lin, Yuezhi; Ma, Jian; Ma, Yan; Zhao, Liping; Li, Shenwei; Yang, Kai; Zhou, Jianhua; Shen, Rongxian; Zhang, Xiaoyan; Shao, Yiming
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.
Meng, Qinglai; Lin, Yuezhi; Ma, Jian; Ma, Yan; Zhao, Liping; Li, Shenwei; Yang, Kai; Zhou, Jianhua; Shen, Rongxian
Abstract 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 EIAVFDDV. 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. PMID:23171359
Zheng, Fengrong; Sun, Xiuqin; Liu, Hongzhan; Wu, Xingan; Zhong, Nan; Wang, Bo; Zhou, Guodong
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 analyzed in tissues of the vaccinated Japanese flounder by PCR, RT-PCR and fluorescent microscopy. Results from PCR analysis indicated that the vaccine-containing plasmids were distributed in injected muscle, the muscle opposite the injection site, the hind intestine, gill, spleen, head, kidney and liver, 6 and 25 days after vaccination. The vaccine plasmids disappeared 100 d post-vaccination. Fluorescent microscopy revealed green fluorescence in the injected muscle, the muscle opposite the injection site, the hind intestine, gill, spleen, head, kidney and liver of fish 48 h post-vaccination, green fluorescence did not appear in the control treated tissue. Green fluorescence became weak at 60 days post-vaccination. RT-PCR analysis indicated that the mcp gene was expressed in all tested tissues of vaccinated fish 6-50 days post-vaccination. These results demonstrate that the antigen encoded by the DNA vaccine is distributed and expressed in all of the tissues analyzed in the vaccinated fish. The antigen would therefore potentially initiate a specific immune response. the plasmid DNA was injected into Japanese flounder ( Paralichthys olivaceus) intramuscularly and antibodies against LCDV were evaluated. The results indicate that the plasmid encoded DNA vaccine could induce an immune response to LCDV and would therefore offer immune protection against LCD. Further studies are required for the development and application of this promising DNA vaccine.
Background Studies have shown that DNA vaccines can induce protective immunity, which demonstrated the high potential of DNA vaccines as an alternative to inactivated vaccines. Vaccines are frequently formulated with adjuvants to improve their release, delivery and presentation to the host immune system. Methods The H5 gene of H5N1 virus (A/Ck/Malaysia/5858/04) was cloned separately into pcDNA3.1 + vector. The immunogenicity of the cloned H5 DNA vaccine was tested on SPF chickens using two different approaches. First approach was using H5 DNA vaccine (pcDNA3.1/H5) and the second was using H5 DNA vaccine in addition to the pcDNA3.1/MDP1 vaccine. Ten days old chickens inoculated three times with two weeks intervals. The spleen and muscle samples from chickens immunized with H5 (pcDNA3.1/H5) and H5 + MDP1 (pcDNA3.1/H5 + pcDNA3.1/MDP1) vaccines were collected after sacrificing the chickens and successfully expressed H5 and MDP1 RNA transcripts. The sera of immunized chickens were collected prior to first immunization and every week after immunization; and analyzed using enzyme-linked immunosorbent assay (ELISA) and hemagglutination inhibition (HI) test. Results Results of competitive ELISA showed successful antibody responses two weeks post immunization. The HI test showed an increased in antibody titers during the course of experiment in group immunized with H5 and H5 + MDP1 vaccines. The result showed that the constructed DNA vaccines were able to produce detectable antibody titer in which the group immunized with H5 + MDP1 vaccine produced higher antibody comparing to H5 vaccine alone. Conclusions This study shows for the first time the usefulness of MDP1 as a genetic adjuvant for H5 DNA vaccine. PMID:20497569
Bergmann-Leitner, Elke S; Leitner, Wolfgang W; Duncan, Elizabeth H; Savranskaya, Tatyana; Angov, Evelina
Malaria represents a major global health problem but despite extensive efforts, no effective vaccine is available. Various vaccine candidates have been developed that provide protection in animal models, such as a gene gun-delivered DNA vaccine encoding the circumsporozoite protein (CSP) of Plasmodium berghei. A common shortcoming of most malaria vaccines is the requirement for multiple immunizations leaving room for improvement even for established vaccine candidates such as the CSP-DNA vaccine. In this study, we explored whether regulating apoptosis in DNA vaccine transfected host cells could accelerate the onset of protective immunity and provide significant protection after a single immunization. A pro-apoptotic gene (Bax) was used as a molecular adjuvant in an attempt to mimic the immunostimulatory apoptosis triggered by viral or virus-derived vaccines, while anti-apoptotic genes such as Bcl-XL may increase the life span of transfected cells thus prolonging antigen production. Surprisingly, co-delivery of either Bax or Bcl-XL greatly reduced CSP-DNA vaccine efficacy after a single immunization. Co-delivery of Bax for three immunizations still had a detrimental effect on protective immunity, while repeated co-delivery of Bcl-XL had no negative impact. The fine characterization of humoral and cellular immune response modulated by these two molecular adjuvants revealed a previously unknown effect, i.e., a shift in the Th-profile. These results demonstrate that pro- or anti-apoptotic molecules should not be used as molecular adjuvants without careful evaluation of the resulting immune response. This finding represents yet another example that strategies to enhance vaccine efficacy developed for other model systems such as viral diseases cannot easily be applied to any vaccine.
Li, Jieqiong; Chen, Hui; Wu, Na; Fan, Dongying; Liang, Guodong; Gao, Na; An, Jing
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.
... html Vaccinating Pregnant Moms Protects Babies From Whooping Cough Benefit is dramatic for newborns who are too ... Babies are far less likely to develop whooping cough if their mother was vaccinated while pregnant, a ...
The term "herd immunity" for the indirect effect of meningococcal conjugate vaccines is inaccurate. A more appropriate term is "herd protection," because this term correctly describes the public effects imparted by vaccination campaigns against the meningococcus.
Yu, Yunzhou; Yu, Jiyu; Li, Na; Wang, Shuang; Yu, Weiyuan; Sun, Zhiwei
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.
Minke, J M; Fischer, L; Baudu, Ph; Guigal, P M; Sindle, T; Mumford, J A; Audonnet, J C
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.
Masih, Shet; Arora, Sunil K; Vasishta, Rakesh K
The acidic ribosomal proteins of the protozoan parasites have been described as prominent antigens during human disease. We present here data showing the molecular cloning and protective efficacy of P1 gene of Leishmania donovani as DNA vaccine. The PCR amplified complete ORF cloned in either pQE or pVAX vector was used either as peptide or DNA vaccine against experimentally induced visceral leishmaniasis in hamsters. The recombinant protein rLdP1 was given along with Freund's adjuvant and the plasmid DNA vaccine, pVAX-P1 was used alone either as single dose or double dose (prime and boost) in different groups of hamsters which were subsequently challenged with a virulent dose of 1×10(7) L. donovani (MHOM/IN/DD8/1968 strain) promastigotes by intra-cardiac route. While the recombinant protein rLdP1 or DNA vaccine pVAX-P1 in single dose format were not found to be protective, DNA vaccine in a prime-boost mode was able to induce protection with reduced mortality, a significant (75.68%) decrease in splenic parasite burden and increased expression of Th1 type cytokines in immunized hamsters. Histopathology of livers and spleens from these animals showed formation of mature granulomas with compact arrangement of lymphocytes and histiocytes, indicating its protective potential as vaccine candidate.
Wallace, D B; Ellis, C E; Espach, A; Smith, S J; Greyling, R R; Viljoen, G J
The glycoprotein (GP) and nucleocapsid (NC) genes of Rift Valley fever virus (RVFV) were expressed in different expression systems and were evaluated for their ability to protect mice from virulent challenge using a prime-boost regime. Mice vaccinated with a lumpy skin disease virus-vectored recombinant vaccine (rLSDV-RVFV) expressing the two RVFV glycoproteins (G1 and G2) developed neutralising antibodies and were fully protected when challenged, as were those vaccinated with a crude extract of truncated G2 glycoprotein (tG2). By contrast mice vaccinated with a DNA vaccine expressing G1 and G2 did not sero-convert with only 20% of them surviving challenge. Mice vaccinated with the DNA vaccine and boosted with rLSDV-RVFV also failed to sero-convert but 40% survived challenge. Surprisingly, although none of the mice immunised with the purified NC protein sero-converted, 60% of them survived virulent challenge. The rLSDV-RVFV construct was then further evaluated in sheep for its dual protective abilities against RVFV and sheeppox virus (SPV). Vaccinated sheep sero-converted for both viruses and were protected against RVFV challenge, however, neither the immunised or negative control animals showed any significant reactions to the virulent SPV challenge.
Wang, Yuan Min; Alexander, Stephen I
Chronic kidney disease is one of the major health problems worldwide. DNA vaccination delivers plasmid DNA encoding the target gene to induce both humoral and cellular immune responses. Here, we describe the methods of CD40 DNA vaccine enhanced by dendritic cell (DC) targeting on the development of Heymann nephritis (HN), a rat model of human membranous nephropathy.
Chen, Zhong-Yuan; Lei, Xiao-Ying; Zhang, Qi-Ya
Plasmid DNAs containing Siniperca chuatsi rhabdovirus (SCRV) glycoprotein gene (pcDNA-G) and nucleoprotein gene (pcDNA-N) were constructed, and used to determine the antiviral immune response elicited by DNA vaccination in mandarin fish. In vitro and in vivo expression of the plasmid constructs was confirmed in transfected cells and muscle tissues of vaccinated fish by Western blot, indirect immunofluorescence or RT-PCR analysis. Fish injected with pcDNA-G exhibited protective effect against SCRV challenge with a relative percent survival (RPS) of 77.5%, but no significant protection (RPS of 2.5%) was observed in fish vaccinated with pcDNA-N. Immunohistochemical analysis showed that vaccination with pcDNA-G decreased histological lesions and suppressed the virus replication in fish target organs, e.g. kidney, liver, spleen, gill and heart. Transcriptional analysis further revealed that the expression levels of type I IFN system genes including interferon regulation factor-7 (IRF-7) gene, myxovirus resistance (Mx) gene and virus inhibitory protein (Viperin) gene were strongly up-regulated after injection with pcDNA-G, whereas the level of transcription of immunoglobulin M (IgM) gene did not show a statistically significant change. These results reveal that type I IFN antiviral immune response is rapidly triggered by the plasmid DNA containing rhabdovirus glycoprotein gene in fish, which offers an explanation of molecular mechanisms for DNA vaccination inducing mandarin fish resist to SCRV disease. Copyright © 2011 Elsevier B.V. All rights reserved.
Whalen, R. G.
A novel and powerful method for vaccine research, colloquially known as DNA vaccines, involves the deliberate introduction into tissues of a DNA plasmid carrying an antigen-coding gene that transfects cells in vivo and results in an immune response. DNA vaccines have several distinct advantages, which include ease of manipulation, use of a generic technology, simplicity of manufacture, and chemical and biological stability. In addition, DNA vaccines are a great leveler among re-searchers around the world because they provide unprecedented ease of experi-mentation. To facilitate diffusion of information, an Internet site has been established called THE DNA VACCINE WEB (URL:http://www.genweb.com/dnavax/dnavax.html). In this review, a brief survey is undertaken of the experimental models and preclinical work on DNA vaccines to contribute to a greater awareness of the possibilities for emerging infectious diseases. PMID:8903226
Clapp, Beata; Walters, Nancy; Thornburg, Theresa; Hoyt, Teri; Yang, Xinghong; Pascual, David W
Brucella abortus remains a threat to the health and well-being of livestock in states bordering the Greater Yellowstone Area. During the past several years, cohabitation of infected wildlife with cattle has jeopardized the brucellosis-free status of Idaho, USA; Wyoming, USA; and Montana, USA. Current livestock B. abortus vaccines have not proven to be efficacious in bison (Bison bison) or elk (Cervus elaphus nelsoni). One problem with the lack of vaccine efficacy may stem from the failure to understand wildlife immune responses to vaccines. In an attempt to understand their immune responses, bison were vaccinated with eukaryotic DNA expression vectors encoding the Brucella periplasmic protein, bp26, and the chaperone protein, trigger factor (TF). These DNA vaccines have previously been shown to be protective against Brucella infection in mice. Bison were immunized intramuscularly at weeks 0, 2, and 4 with bp26 and TF DNA vaccines plus CpG adjuvant or empty vector (control) plus CpG. Blood samples were collected before vaccination and at 8, 10, and 12 wk after primary vaccination. The results showed that bison immunized with bp26 and TF DNA vaccines developed enhanced antibody, proliferative T cell, and interferon-gamma (IFN-γ) responses upon in vitro restimulation with purified recombinant bp26 or TF antigens, unlike bison immunized with empty vector. Flow cytometric analysis revealed that the percentages of CD4(+) and CD8(+) T lymphocytes from the DNA-vaccinated groups were significantly greater than they were for those bison given empty vector. These data suggest that DNA vaccination of bison may elicit strong cellular immune responses and serve as an alternative for vaccination of bison for brucellosis.
Ding, Xicheng; Lillehoj, Hyun S; Dalloul, Rami A; Min, Wongi; Sato, Takanori; Yasuda, Atsushi; Lillehoj, Erik P
An Eimeria tenella microneme recombinant gene (EtMIC2) and encoded protein were evaluated as potential vaccines against avian coccidiosis. In ovo inoculation with the EtMIC2 gene increased anti-EtMIC2 antibody titers at days 10 and 17 following E. tenella infection. In addition, vaccinated birds developed protective immunity against infection by E. tenella as assessed by significantly increased body weight gain and decreased fecal oocyst shedding compared with non-vaccinated controls. Vaccination with the EtMIC2 gene also led to protective immunity against infection by E. acervulina, but not E. maxima. Combined in ovo DNA vaccination plus post-hatch boosting with EtMIC2 DNA or protein did not improve antibody titers or protective immunity beyond that achieved with in ovo vaccination alone. These results provide evidence that in ovo immunization with a recombinant Eimeria microneme gene stimulates protective intestinal immunity against coccidiosis.
Kumar, Rajesh; Nyakundi, Ruth; Kariuki, Thomas; Ozwara, Hastings; Nyamongo, Onkoba; Mlambo, Godfree; Ellefsen, Barry; Hannaman, Drew; Kumar, Nirbhay
Plasmodium falciparum Pfs25 antigen, expressed on the surface of zygotes and ookinetes, is one of the leading targets for the development of a malaria transmission-blocking vaccine (TBV). Our laboratory has been evaluating DNA plasmid based Pfs25 vaccine in mice and non-human primates. Previously, we established that in vivo electroporation (EP) delivery is an effective method to improve the immunogenicity of DNA vaccine encoding Pfs25 in mice. In order to optimize the in vivo EP procedure and test for its efficacy in more clinically relevant larger animal models, we employed in vivo EP to evaluate the immune response and protective efficacy of Pfs25 encoding DNA vaccine in nonhuman primates (olive baboons, Papio anubis). The results showed that at a dose of 2.5mg DNA vaccine, antibody responses were significantly enhanced with EP as compared to without EP resulting in effective transmission blocking efficiency. Similar immunogenicity enhancing effect of EP was also observed with lower doses (0.5mg and 1mg) of DNA plasmids. Further, final boosting with a single dose of recombinant Pfs25 protein resulted in dramatically enhanced antibody titers and significantly increased functional transmission blocking efficiency. Our study suggests priming with DNA vaccine via EP along with protein boost regimen as an effective method to elicit potent immunogenicity of malaria DNA vaccines in nonhuman primates and provides the basis for further evaluation in human volunteers.
Ljungberg, Karl; Kümmerer, Beate M.; Gosse, Leslie; Dereuddre-Bosquet, Nathalie; Tchitchek, Nicolas; Hallengärd, David; García-Arriaza, Juan; Meinke, Andreas; Esteban, Mariano; Merits, Andres
Chikungunya virus (CHIKV) is rapidly spreading across the globe, and millions are infected. Morbidity due to this virus is a serious threat to public health, but at present, there is no vaccine against this debilitating disease. We have recently developed a number of vaccine candidates, and here we have evaluated 3 of them in a nonhuman primate model. A single immunization with an attenuated strain of CHIKV (Δ5nsP3), a homologous prime-boost immunization with a DNA-launched RNA replicon encoding CHIKV envelope proteins (DREP-E), and a DREP-E prime followed by a recombinant modified vaccinia virus Ankara encoding CHIKV capsid and envelope (MVA-CE) boost all induced protection against WT CHIKV infection. The attenuated Δ5nsP3 virus proved to be safe and did not show any clinical signs typically associated with WT CHIKV infections such as fever, skin rash, lymphopenia, or joint swelling. These vaccines are based on an East/Central/South African strain of Indian Ocean lineage, but they also generated neutralizing antibodies against an isolate of the Asian genotype that now is rapidly spreading across the Americas. These results form the basis for clinical development of an efficacious CHIKV vaccine that generates both humoral and cellular immunity with long-term immunological memory. PMID:28352649
AWARD NUMBER: W81XWH-15-2-0029 TITLE: Enhancing the Immunogenicity of a Tetravalent Dengue DNA Vaccine PRINCIPAL INVESTIGATOR: Maya...TITLE AND SUBTITLE Enhancing the Immunogenicity of a Tetravalent Dengue DNA 5a. CONTRACT NUMBER Vaccine 5b. GRANT NUMBER 5c. PROGRAM ELEMENT...personnel is priority research area for the US DoD. Phase 1 clinical trials demonstrated that the Naval Medical Research Center’s DNA based dengue vaccine
Kong, Wei; Brovold, Matthew; Koeneman, Brian A; Clark-Curtiss, Josephine; Curtiss, Roy
We previously developed a biological containment system using recombinant Salmonella Typhimurium strains that are attenuated yet capable of synthesizing protective antigens. The regulated delayed attenuation and programmed self-destructing features designed into these S. Typhimurium strains enable them to efficiently colonize host tissues and allow release of the bacterial cell contents after lysis. To turn such a recombinant attenuated Salmonella vaccine (RASV) strain into a universal DNA vaccine-delivery vehicle, our approach was to genetically modify RASV strains to display a hyperinvasive phenotype to maximize Salmonella host entry and host cell internalization, to enable Salmonella endosomal escape to release a DNA vaccine into the cytosol, and to decrease Salmonella-induced pyroptosis/apoptosis that allows the DNA vaccine time to traffic to the nucleus for efficient synthesis of encoded protective antigens. A DNA vaccine vector that encodes a domain that contributes to the arabinose-regulated lysis phenotype but has a eukaryotic promoter was constructed. The vector was then improved by insertion of multiple DNA nuclear-targeting sequences for efficient nuclear trafficking and gene expression, and by increasing nuclease resistance to protect the plasmid from host degradation. A DNA vaccine encoding influenza WSN virus HA antigen delivered by the RASV strain with the best genetic attributes induced complete protection to mice against a lethal influenza virus challenge. Adoption of these technological improvements will revolutionize means for effective delivery of DNA vaccines to stimulate mucosal, systemic, and cellular protective immunities, and lead to a paradigm shift in cost-effective control and prevention of a diversity of diseases.
Hitzeroth, Inga I.; Passmore, Jo-Ann S.; Shephard, Enid; Stewart, Debbie; Müller, Martin; Williamson, Anna-Lise; Rybicki, Edward P.; Kast, W. Martin
The ability to elicit cross-neutralizing antibodies makes human papillomavirus (HPV) L2 capsid protein a possible HPV vaccine. We examined and compared the humoral response of mice immunised with a HPV-16 L2 DNA vaccine or with HPV-16 L2 protein. The L2 DNA vaccine elicited a non-neutralising antibody response unlike the L2 protein. L2 DNA vaccination suppressed the growth of L2-expressing C3 tumor cells, which is a T cell mediated effect, demonstrating that the lack of non-neutralizing antibody induction by L2 DNA was not caused by lack of T cell immunogenicity of the construct. PMID:19559114
Iurescia, Sandra; Fioretti, Daniela; Pierimarchi, Pasquale; Signori, Emanuela; Zonfrillo, Manuela; Tonon, Giancarlo; Fazio, Vito M.; Rinaldi, Monica
Therapeutic vaccination against idiotype is a promising strategy for immunotherapy of B-cell malignancies. We have previously shown that CDR3-based DNA immunization can induce immune response against lymphoma and explored this strategy to provide protection in a murine B-cell lymphoma model. Here we performed vaccination employing as immunogen a naked DNA fusion product. The DNA vaccine was generated following fusion of a sequence derived from tetanus toxin fragment C to the VHCDR3109−116 epitope. Induction of tumor-specific immunity as well as ability to inhibit growth of the aggressive 38C13 lymphoma and to prolong survival of vaccinated mice has been tested. We determined that DNA fusion vaccine induced immune response, elicited a strong protective antitumor immunity, and ensured almost complete long-term tumor-free survival of vaccinated mice. Our results show that CDR3-based DNA fusion vaccines hold promise for vaccination against lymphoma. PMID:20445751
Stevenson, Freda K; Ottensmeier, Christian H; Rice, Jason
Genetic technology allows construction of DNA vaccines encoding selected tumor antigens together with molecules to direct and amplify the desired effector pathways. Their enormous promise has been marred by a problem of scaling up to human subjects. This is now largely overcome by electroporation, which increases both antigen expression and the inflammatory milieu. While the principles of vaccine design can be developed in mouse models, the real operative test is in the clinic, using patients in temporary remission. Monitoring of induced immunity, although commonly limited to blood, is providing objective qualitative and quantitative data on T-cell and antibody responses. Prolongation of remission is the goal and an activated immune system should achieve this.
Eblé, P L; Geurts, Y; Quak, S; Moonen-Leusen, H W; Blome, S; Hofmann, M A; Koenen, F; Beer, M; Loeffen, W L A
Currently no live DIVA (Differentiating Infected from Vaccinated Animals) vaccines against classical swine fever (CSF) are available. The aim of this study was to investigate whether chimeric pestivirus vaccine candidates (CP7_E2alf, Flc11 and Flc9) are able to protect pigs against clinical signs, and to reduce virus shedding and virus transmission, after a challenge with CSF virus (CSFV), 7 or 14 days after a single intramuscular vaccination. In these vaccine candidates, either the E2 or the E(rns) encoding genome region of a bovine viral diarrhoea virus strain were combined with a cDNA copy of CSFV or vice versa. Furthermore, currently available serological DIVA tests were evaluated. The vaccine candidates were compared to the C-strain. All vaccine candidates protected against clinical signs. No transmission to contact pigs was detected in the groups vaccinated with C-strain, CP7_E2alf and Flc11. Limited transmission occurred in the groups vaccinated with Flc9. All vaccine candidates would be suitable to stop on-going transmission of CSFV. For Flc11, no reliable differentiation was possible with the current E(rns)-based DIVA test. For CP7_E2alf, the distribution of the inhibition percentages was such that up to 5% false positive results may be obtained in a large vaccinated population. For Flc9 vaccinated pigs, the E2 ELISA performed very well, with an expected 0.04% false positive results in a large vaccinated population. Both CP7_E2alf and Flc9 are promising candidates to be used as live attenuated marker vaccines against CSF, with protection the best feature of CP7_E2alf, and the DIVA principle the best feature of Flc9.
Zhao, Chen; Sun, Ying; Zhao, Yujie; Wang, Si; Yu, Tongtong; Du, Feng; Yang, X Frank; Luo, Enjie
Hemorrhagic fever with renal syndrome (HFRS) is a severe epidemic disease caused by hantaviruses including Hantaan virus (HTNV), Seoul virus (SEOV), Dobrava virus (DOBV) and Puumala virus. Three of the four HFRS hantaviruses, HTNV, SEOV, and PUUV are found in China. Currently, there is no effective strategy available to reduce infection risk. In this study, we constructed a multi-epitope chimeric DNA vaccine that encodes expressing 25 glycoprotein epitopes from SEOV, HTNV and PUUV (designated as SHP chimeric gene). Vaccination of BALb/c mice with SHP multi-epitope chimeric DNA vaccine led to a dramatic augmentation of humoral and cellular responses. The SHP vaccine DNA was detected in many organs but not for more than 60 d. There was no risk of mutation due to integration. Thus, the SHP multi-epitope chimeric DNA vaccine is a potential effective and safe DNA vaccine against infection by SEOV, HTNV, and PUUV.
Li, Si-Ming; Bai, Fu-Liang; Xu, Wen-Juan; Yang, Yong-Bi; An, Ying; Li, Tian-He; Yu, Yin-Hang; Li, De-Shan; Wang, Wen-Fei
The clearance of host cell DNA is a critical indicator for Vero-cell culture-derived rabies vaccine. In this study, we evaluated the clearance of DNA in Vero-cell culture-derived rabies vaccine by purification process utilizing ultrafiltration, nuclease digestion, and gel filtration chromatography. The results showed that the bioprocess of using nuclease decreased residual DNA. Dot-blot hybridization analysis showed that the residual host cell DNA was <100 pg/ml in the final product. The residual nuclease in rabies vaccine was less than 0.1 ng/ml protein. The residual nuclease could not paly the biologically active role of digestion of DNA. Experiments of stability showed that the freeze-drying rabies virus vaccine was stable and titers were >5.0 IU/ml. Immunogenicity test and protection experiments indicated mice were greatly induced generation of neutralizing antibodies and invoked protective effects immunized with intraperitoneal injections of the rabies vaccine. These results demonstrated that the residual DNA was removed from virus particles and nuclease was removed by gel filtration chromatography. The date indicated that technology was an efficient method to produce rabies vaccine for human use by using nuclease.
Shibui, Akiko; Nakae, Susumu; Watanabe, Junichi; Sato, Yoshitaka; Tolba, Mohammed E M; Doi, Junko; Shiibashi, Takashi; Nogami, Sadao; Sugano, Sumio; Hozumi, Nobumichi
No licensed malaria vaccine exists, in spite of intensive development efforts. We have been investigating development of a DNA vaccine to prevent malaria infection. To date, we have established a full-length cDNA expression library from the erythrocytic-stage murine malaria parasite, Plasmodium berghei. We found that immunization of mice with combined 2000 clones significantly prolonged survival after challenge infection and that splenocytes from the immunized mice showed parasite-specific cytokine production. We determined the 5'-end one-pass sequence of these clones and mapped a draft genomic sequence for P. berghei for use in screening vaccine candidates for efficacy. In this study, we annotated these cDNA clones by comparing them with the genomic sequence of Plasmodium falciparum. We then divided them into several subsets based on their characteristics and examined their protective effects against malaria infection. Consequently, we selected 104 clones that strongly induced specific IgG production and decreased the mortality rate in the early phase. Most of these 104 clones coded for unknown proteins. The results suggest that these clones represent potential novel malaria vaccine candidates.
Bins, Adriaan D; Jorritsma, Annelies; Wolkers, Monika C; Hung, Chien-Fu; Wu, T-C; Schumacher, Ton N M; Haanen, John B A G
Induction of immunity after DNA vaccination is generally considered a slow process. Here we show that DNA delivery to the skin results in a highly transient pulse of antigen expression. Based on this information, we developed a new rapid and potent intradermal DNA vaccination method. By short-interval intradermal DNA delivery, robust T-cell responses, of a magnitude sufficient to reject established subcutaneous tumors, are generated within 12 d. Moreover, this vaccination strategy confers protecting humoral immunity against influenza A infection within 2 weeks after the start of vaccination. The strength and speed of this newly developed strategy will be beneficial in situations in which immunity is required in the shortest possible time.