Vesicular stomatitis virus-based vaccine protects hamsters against lethal challenge with Andes virus

Andes virus (ANDV) is a highly pathogenic South American hantavirus that causes hantavirus pulmonary syndrome (HPS). A high case fatality rate, the potential for human-to-human transmission, the capacity to infect via aerosolization, and the absence of effective therapies make it imperative that a safe, fast-acting, and effective ANDV vaccine be developed. We generated and characterized a recombinant vesicular stomatitis virus (VSV) vector expressing the ANDV surface glycoprotein precursor (VSVΔG/ANDVGPC) as a possible vaccine candidate and tested its efficacy in the only lethal-disease animal model of HPS. Syrian hamsters immunized with a single injection of VSVΔG/ANDVGPC were fully protected against disease when challenged at 28, 14, 7, or 3 days postimmunization with a lethal dose of ANDV; however, the mechanism of protection seems to differ depending on when the immunization occurs. At 28 days postimmunization, a lack of detectable ANDV RNA in lung, liver, and blood tissue samples, as well as a lack of seroconversion to the ANDV nucleocapsidprotein in nearly all animals, suggested largely sterile immunity. The vaccine was able to generate high levels of neutralizing anti-ANDV G(N)/G(C) antibodies, which seem to play a role as a mechanism of vaccine protection. Administration of the vaccine at 7 or 3 days before challenge also resulted in full protection but with no specific neutralizing humoral immune response, suggesting a possible role of innate responses in protection against challenge virus replication. Administration of the vaccine 24 h postchallenge was successful in protecting 90% of hamsters and again suggested the induction of a potent antiviral state by the recombinant vector as a potential mechanism. Overall, our data suggest the potential for the use of the VSV platform as a fast-acting and effective prophylaxis/postexposure treatment against lethal hantavirus infections.     

Recombinant baculovirus-based vaccine expressing M2 protein induces protective CD8<Superscript>+</Superscript> T-cell immunity against respiratory syncytial virus infection

Respiratory syncytial virus (RSV) is an important cause of acute lower respiratory tract disease in infants, young children, immunocompromised individuals, and the elderly. However, despite ongoing efforts to develop an RSV vaccine, there is still no authorized RSV vaccine for humans. Baculovirus has attracted attention as a vaccine vector because of its ability to induce a high level of humoral and cellular immunity, low cytotoxicity against various antigens, and biological safety for humans. In this study, we constructed a recombinant baculovirus- based vaccine expressing the M2 protein of RSV under the control of cytomegalovirus promoter (Bac_RSVM2) to induce CD8⁺ T-cell responses which play an important role in viral clearance, and investigated its protective efficacy against RSV infection. Immunization with Bac_RSVM2 via intranasal or intramuscular route effectively elicited the specific CD8⁺ T-cell responses. Most notably, immunization with Bac_RSVM2 vaccine almost completely protected mice from RSV challenge without vaccine-enhanced immunopathology. In conclusion, these results suggest that Bac_RSVM2 vaccine employing the baculovirus delivery platform has promising potential to be developed as a safe and novel RSV vaccine that provides protection against RSV infection.     

Multiple alternating immunizations with DNA vaccine and replication incompetent adenovirus expressing gB of pseudorabies virus protect animals against lethal virus challenge

The prime-boost vaccination with DNA vaccine and recombinant viral vector has emerged as an effective prophylactic strategy to control infectious diseases. Here, we compared the protective immunities induced by multiple alternating immunizations with DNA vaccine (pCIgB) and replication-incompetent adenovirus (Ad-gB) expressing glycoprotein gB of pseudorabies virus (PrV). The platform of pCIgB-prime and Ad-gB-boost induced the most effective immune responses and provided protection against virulent PrV infection. However, priming with pCIgB prior to vaccinating animals by the DNA vaccine-prime and Ad-boost protocol provided neither effective immune responses nor protection against PrV. Similarly, boosting with Ad-gB following immunization with DNA vaccine-prime and Ad-boost showed no significant responses. Moreover, whereas the administration of Ad-gB for primary immunization induced Th2-type-biased immunity, priming with pCIgB induced Th1-type-biased immunity, as judged by the production of PrV-specific IgG isotypes and cytokine IFN-gamma. These results indicate that the order and injection frequency of vaccine vehicles used for heterologous prime-boost vaccination affect the magnitude and nature of the immunity. Therefore, our demonstration implies that the prime-boost protocol should be carefully considered and selected to induce the desired immune responses.     

Immunogenicity of Live Attenuated B. pertussis BPZE1 Producing the Universal Influenza Vaccine Candidate M2e

Background

Intranasal delivery of vaccines directed against respiratory pathogens is an attractive alternative to parenteral administration. However, using this delivery route for inactivated vaccines usually requires the use of potent mucosal adjuvants, and no such adjuvant has yet been approved for human use.

Methodology/Principal Findings

We have developed a live attenuated Bordetella pertussis vaccine, called BPZE1, and show here that it can be used to present the universal influenza virus epitope M2e to the mouse respiratory tract to prime for protective immunity against viral challenge. Three copies of M2e were genetically fused to the N-terminal domain of filamentous hemagglutinin (FHA) and produced in recombinant BPZE1 derivatives in the presence or absence of endogenous full-length FHA. Only in the absence of FHA intranasal administration of the recombinant BPZE1 derivative induced antibody responses to M2e and effectively primed BALB/c mice for protection against influenza virus-induced mortality and reduced the viral load after challenge. Strong M2e-specific antibody responses and protection were observed after a single nasal administration with the recombinant BPZE1 derivative, followed by a single administration of M2e linked to a virus-like particle without adjuvant, whereas priming alone with the vaccine strain did not protect.

Conclusions/Significance

Using recombinant FHA-3M2e-producing BPZE1 derivatives for priming and the universal influenza M2e peptide linked to virus-like particles for boosting may constitute a promising approach for needle-free and adjuvant-free nasal vaccination against influenza.     

共表达H5N1流感病毒M1和HA基因的DNA疫苗与腺病毒载体疫苗的小鼠免疫评价

为评价在小鼠体内表达流感病毒M1和HA基因诱导的免疫反应,制备共表达H5N1亚型禽流感病毒 (A/Anhui/1/2005) 全长基质蛋白1 (M1) 基因和血凝素 (HA) 基因的重组DNA疫苗pStar-M1/HA和重组腺病毒载体疫苗Ad-M1/HA,将其按初免-加强程序免疫BALB/c小鼠,共免疫4次,每次间隔14 d。第1、3次用DNA疫苗,第2、4次用重组腺病毒载体疫苗,每次免疫前及末次免疫后14 d采集小鼠血清用于检测体液免疫应答,末次免疫后14 d采集小鼠脾淋巴细胞用于检测细胞免疫应答。血凝     

Potent T cell responses induced by single DNA vaccine boosted with recombinant vaccinia vaccine

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

Mucosal vaccination with a multicomponent adenovirus-vectored vaccine protects against Streptococcus pneumoniae infection in the lung

Streptococcus pneumoniae is a major bacterial respiratory pathogen. Current licensed pneumococcal polysaccharide and polysaccharide–protein conjugate vaccines are administered by an intramuscular injection. In order to develop a new-generation vaccine that can be administered in a needle-free mucosal manner, we have constructed early 1 and 3 gene regions (E1/E3) deleted, replication-defective adenoviral vectors encoding pneumococcal surface antigen A (PsaA), the N-fragment of pneumococcal surface protein A (N-PspA), and the detoxified mutant pneumolysin (PdB) from S. pneumoniae strain D39. Intranasal vaccination with the three adenoviral vectors (Ad/PsaA, Ad/N-PspA, and Ad/PdB) in mice resulted in robust antigen-specific serum immunoglobulin G responses, as demonstrated by an enzyme-linked immunosorbent assay. In addition, nasal mucosal vaccination with the combination of the three adenoviral vectors conferred protection against S. pneumoniae strain D39 colonization in mouse lungs. Taken together, these data demonstrate the feasibility of developing a mucosal vaccine against S. pneumoniae using recombinant adenoviruses for antigen delivery.     

Single mucosal, but not parenteral, immunization with recombinant adenoviral-based vaccine provides potent protection from pulmonary tuberculosis

Bacillus Calmette-Guerin (BCG) vaccine has failed to control the global tuberculosis (TB) epidemic, and there is a lack of safe and effective mucosal vaccines capable of potent protection against pulmonary TB. A recombinant replication-deficient adenoviral-based vaccine expressing an immunogenic Mycobacterium tuberculosis Ag Ag85A (AdAg85A) was engineered and evaluated for its potential to be used as a respiratory mucosal TB vaccine in a murine model of pulmonary TB. A single intranasal, but not i.m., immunization with AdAg85A provided potent protection against airway Mycobacterium tuberculosis challenge at an improved level over that by cutaneous BCG vaccination. Systemic priming with an Ag85A DNA vaccine and mucosal boosting with AdAg85A conferred a further enhanced immune protection which was remarkably better than BCG vaccination. Such superior protection triggered by AdAg85 mucosal immunization was correlated with much greater retention of Ag-specific T cells, particularly CD4 T cells, in the lung and was shown to be mediated by both CD4 and CD8 T cells. Thus, adenoviral TB vaccine represents a promising novel vaccine platform capable of potent mucosal immune protection against TB. Our study also lends strong evidence that respiratory mucosal vaccination is critically advantageous over systemic routes of vaccination against TB.     

Codelivery of the chemokine CCL3 by an adenovirus-based vaccine improves protection from retrovirus infection

Processing and presentation of vaccine antigens by professional antigen-presenting cells (APCs) is of great importance for the efficient induction of protective immunity. We analyzed whether the efficacy of an adenovirus-based retroviral vaccine can be enhanced by coadministration of adenovirus-encoded chemokines that attract and stimulate APCs. In the Friend retrovirus (FV) mouse model we coexpressed CCL3, CCL20, CCL21, or CXCL14 from adenoviral vectors, together with FV Gag and Env antigens, and then analyzed immune responses and protection from pathogenic FV infection. Although most tested chemokines did not improve protection against FV challenge, mice that received adenoviral vectors encoding CCL3 together with FV antigens showed significantly better control over viral loads and FV-induced disease than mice immunized with the viral antigens only. Improved protection correlated with enhanced virus-specific CD4+ T cell responses and higher neutralizing antibody titers. To apply these results to an HIV vaccine, mice were immunized with adenoviral vectors encoding the HIV antigens Env and Gag-Pol and coadministered vectors encoding CCL3. Again, this combination vaccine induced higher virus-specific antibody titers and CD4+ T cell responses than did the HIV antigens alone. These results indicate that coexpression of the chemokine CCL3 by adenovirus-based vectors may be a promising tool to improve antiretroviral vaccination strategies.     

A novel chimpanzee adenovirus vector with low human seroprevalence: improved systems for vector derivation and comparative immunogenicity

Recombinant adenoviruses are among the most promising tools for vaccine antigen delivery. Recently, the development of new vectors has focused on serotypes to which the human population is less exposed in order to circumvent pre-existing anti vector immunity. This study describes the derivation of a new vaccine vector based on a chimpanzee adenovirus, Y25, together with a comparative assessment of its potential to elicit transgene product specific immune responses in mice. The vector was constructed in a bacterial artificial chromosome to facilitate genetic manipulation of genomic clones. In order to conduct a fair head-to-head immunological comparison of multiple adenoviral vectors, we optimised a method for accurate determination of infectious titre, since this parameter exhibits profound natural variability and can confound immunogenicity studies when doses are based on viral particle estimation. Cellular immunogenicity of recombinant E1 E3-deleted vector ChAdY25 was comparable to that of other species E derived chimpanzee adenovirus vectors including ChAd63, the first simian adenovirus vector to enter clinical trials in humans. Furthermore, the prevalence of virus neutralizing antibodies (titre >1:200) against ChAdY25 in serum samples collected from two human populations in the UK and Gambia was particularly low compared to published data for other chimpanzee adenoviruses. These findings support the continued development of new chimpanzee adenovirus vectors, including ChAdY25, for clinical use.     

Adenovirus-vectored T cell vaccine for hepacivirus shows reduced effectiveness against a CD8 T cell escape variant in rats

There is an urgent need for a vaccine to prevent chronic infection by hepatitis C virus (HCV) and its many genetic variants. The first human vaccine trial, using recombinant viral vectors that stimulate pan-genotypic T cell responses against HCV non-structural proteins, failed to demonstrate efficacy despite significant preclinical promise. Understanding the factors that govern HCV T cell vaccine success is necessary for design of improved immunization strategies. Using a rat model of chronic rodent hepacivirus (RHV) infection, we assessed the impact of antigenic variation and immune escape upon success of a conceptually analogous RHV T cell vaccine. Naïve Lewis rats were vaccinated with a recombinant human adenovirus expressing RHV non-structural proteins (NS)3-5B and later challenged with a viral variant containing immune escape mutations within major histocompatibility complex (MHC) class I-restricted epitopes (escape virus). Whereas 7 of 11 (64%) rats cleared infection caused by wild-type RHV, only 3 of 12 (25%) were protected against heterologous challenge with escape virus. Uncontrolled replication of escape virus was associated with durable CD8 T cell responses targeting escaped epitopes alone. In contrast, clearance of escape virus correlated with CD4 T cell helper immunity and maintenance of CD8 T cell responses against intact viral epitopes. Interestingly, clearance of wild-type RHV infection after vaccination conferred enhanced protection against secondary challenge with escape virus. These results demonstrate that the efficacy of an RHV T cell vaccine is reduced when challenge virus contains escape mutations within MHC class I-restricted epitopes and that failure to sustain CD8 T cell responses against intact epitopes likely underlies immune failure in this setting. Further investigation of the immune responses that yield protection against diverse RHV challenges in this model may facilitate design of broadly effective HCV vaccines.     

Virus-like particles: designing an effective AIDS vaccine

Viruses that infect eukaryotic organisms have the unique characteristic of self-assembling into particles. The mammalian immune system is highly attuned to recognizing and attacking these viral particles following infection. The use of particle-based immunogens, often delivered as live-attenuated viruses, has been an effective vaccination strategy for a variety of viruses. The development of an effective vaccine against the human immunodeficiency virus (HIV) has proven to be a challenge, since HIV infects cells of the immune system causing severe immunodeficiency resulting in the syndrome known as AIDS. In addition, the ability of the virus to adapt to immune pressure and reside in an integrated form in host cells presents hurdles for vaccinologists to overcome. A particle-based vaccine strategy has promise for eliciting high titer, long-lived, immune responses to a diverse number of viral epitopes against different HIV antigens. Live-attenuated viruses are effective at generating both cellular and humoral immune responses. However, while these vaccines stimulate immunity, challenged animals rarely clear the viral infection and the degree of attenuation directly correlates with protection from disease. Further, a live-attenuated vaccine has the potential to revert to a pathogenic form. Alternatively, virus-like particles (VLPs) mimic the viral particle without causing an immunodeficiency disease. VLPs are self-assembling, non-replicating, non-pathogenic particles that are similar in size and conformation to intact virions. A variety of VLPs for lentiviruses are currently in preclinical and clinical trials. This review focuses on our current status of VLP-based AIDS vaccines, regarding issues of purification and immune design for animal and clinical trials.     

Attenuated Bordetella pertussis BPZE1 as a live vehicle for heterologous vaccine antigens delivery through the nasal route

Whereas the great majority of the current vaccines are delivered through the parenteral route, mucosal administration has been increasingly considered for controlling infection and preventing disease. Mucosal vaccination can trigger both humoral and cell-mediated protection, not only at the targeted mucosal surface, but also systemically. In this regard, nasal vaccination has shown great potential. The live attenuated strain of Bordetella pertussis, BPZE1, is particularly attractive and promising as a nasal vaccine delivery vector of heterologous antigen vaccine candidates. BPZE1 was originally developed as a live nasal pertussis vaccine candidate, and is currently undergoing phase I clinical trial in human (http://www.child-innovac.org). Highly adapted to the human respiratory tract and offering several potential protein carriers for presentation of the heterologous antigen vaccine candidates, BPZE1 represents an appealing platform for the development of live recombinant vaccines delivered via the nasal route that would confer simultaneous protection against pertussis and the targeted infectious disease(s).     

T-cell-mediated protective efficacy of a systemic vaccine approach in cynomolgus monkeys after SIV mucosal challenge

The immunogenicity and the protective efficacy of a new polyvalent triple vector (DNA/SFV/MVA) based vaccine against mucosal challenge with pathogenic SIVmac251 were investigated. Cynomolgus monkeys (Macaca fascicularis) were primed intradermally with DNA, boosted twice subcutaneously with recombinant Semliki Forest virus (rSFV) and finally intramuscularly with recombinant Modified Vaccinia Virus Ankara strain (rMVA). Both DNA and recombinant viral vectors expressed SIV proteins (Gag, Pol, Tat, Rev, Nef and Env). The vaccinated monkeys developed T helper proliferative responses to viral antigens after the second immunization while interferon (IFN)-gamma enzyme-linked immunosorbent spot-forming cell assay (ELISPOT) specific responses appeared only after the last boost with rMVA. Upon intrarectal challenge with pathogenic SIVmac251, three of four vaccinated monkeys were either fully protected or exhibited a dramatic reduction of virus replication up to undetectable level. A major contribution to this protective effect appeared to be the anamnestic T-cell IFN-gamma ELISPOT responses to vaccine antigens (Gag, Rev, Tat, Nef) that mirrored the viral clearance. These results underline the efficacy of a multiprotein approach in combination with a triple vector system of antigen delivery.     

Recombinant adenovirus serotype 26 (Ad26) and Ad35 vaccine vectors bypass immunity to Ad5 and protect nonhuman primates against ebolavirus challenge

The use of adenoviruses (Ad) as vaccine vectors against a variety of pathogens has demonstrated their capacity to elicit strong antibody and cell-mediated immune responses. Adenovirus serotype C vectors, such as Ad serotype 5 (Ad5), expressing Ebolavirus (EBOV) glycoprotein (GP), protect completely after a single inoculation at a dose of 10(10) viral particles. However, the clinical application of a vaccine based on Ad5 vectors may be hampered, since impairment of Ad5 vaccine efficacy has been demonstrated for humans and nonhuman primates with high levels of preexisting immunity to the vector. Ad26 and Ad35 segregate genetically from Ad5 and exhibit lower seroprevalence in humans, making them attractive vaccine vector alternatives. In the series of studies presented, we show that Ad26 and Ad35 vectors generate robust antigen-specific cell-mediated and humoral immune responses against EBOV GP and that Ad5 immune status does not affect the generation of GP-specific immune responses by these vaccines. We demonstrate partial protection against EBOV by a single-shot Ad26 vaccine and complete protection when this vaccine is boosted by Ad35 1 month later. Increases in efficacy are paralleled by substantial increases in T- and B-cell responses to EBOV GP. These results suggest that Ad26 and Ad35 vectors warrant further development as candidate vaccines for EBOV.     

Enhanced mucosal immunoglobulin A response of intranasal adenoviral vector human immunodeficiency virus vaccine and localization in the central nervous system

Replication-defective adenovirus (ADV) vectors represent a promising potential platform for the development of a vaccine for AIDS. Although this vector is typically administered intramuscularly, it would be desirable to induce mucosal immunity by delivery through alternative routes. In this study, the immune response and biodistribution of ADV vectors delivered by different routes were evaluated. ADV vectors expressing human immunodeficiency virus type 1 (HIV-1) Gag, Pol, and Env were delivered intramuscularly or intranasally into mice. Intranasal immunization induced greater HIV-specific immunoglobulin A (IgA) responses in mucosal secretions and sera than in animals with intramuscular injection, which showed stronger systemic cellular and IgG responses. Administration of the vaccine through an intranasal route failed to overcome prior ADV immunity. Animals exposed to ADV prior to vaccination displayed substantially reduced cellular and humoral immune responses to HIV antigens in both groups, though the reduction was greater in animals immunized intranasally. This inhibition was partially overcome by priming with a DNA expression vector expressing HIV-1 Gag, Pol, and Env before boosting with the viral vector. Biodistribution of recombinant adenovirus (rADV) vectors administered intranasally revealed infection of the central nervous system, specifically in the olfactory bulb, possibly via retrograde transport by olfactory neurons in the nasal epithelium, which may limit the utility of this route of delivery of ADV vector-based vaccines.     

Successful topical respiratory tract immunization of primates against Ebola virus

Ebola virus causes outbreaks of severe viral hemorrhagic fever with high mortality in humans. The virus is highly contagious and can be transmitted by contact and by the aerosol route. These features make Ebola virus a potential weapon for bioterrorism and biological warfare. Therefore, a vaccine that induces both systemic and local immune responses in the respiratory tract would be highly beneficial. We evaluated a common pediatric respiratory pathogen, human parainfluenza virus type 3 (HPIV3), as a vaccine vector against Ebola virus. HPIV3 recombinants expressing the Ebola virus (Zaire species) surface glycoprotein (GP) alone or in combination with the nucleocapsid protein NP or with the cytokine adjuvant granulocyte-macrophage colony-stimulating factor were administered by the respiratory route to rhesus monkeys--in which HPIV3 infection is mild and asymptomatic--and were evaluated for immunogenicity and protective efficacy against a highly lethal intraperitoneal challenge with Ebola virus. A single immunization with any construct expressing GP was moderately immunogenic against Ebola virus and protected 88% of the animals against severe hemorrhagic fever and death caused by Ebola virus. Two doses were highly immunogenic, and all of the animals survived challenge and were free of signs of disease and of detectable Ebola virus challenge virus. These data illustrate the feasibility of immunization via the respiratory tract against the hemorrhagic fever caused by Ebola virus. To our knowledge, this is the first study in which topical immunization through respiratory tract achieved prevention of a viral hemorrhagic fever infection in a primate model.     

A vaccine strategy utilizing a combination of three different chimeric vectors which share specific vaccine antigens

A large number of recombinant of viral and bacterial systems have been engineered as vectors to express foreign genes for vaccination and/or gene therapy. A common problem is the immune response to the vector itself. The presence of anti-vector immune responses may preclude sufficient 'priming' or immunogenicity if pre-existing immune responses are present, or they may impair optimal 'boosting' upon repeated immunization or delivery with the same vector. To circumvent this problem we developed a strategy using different chimeric vectors which share only the expression of common specific antigens desired for immunization. This approach not only has the advantage of avoiding increased anti-vector responses, but allows the use of combinations of vectors which could subsequently present the same or related antigen differently to the immune system as well as at alternative sites to induce the optimal type of immunity against the pathogen of interest.     

Vector Choice Determines Immunogenicity and Potency of Genetic Vaccines against Angola Marburg Virus in Nonhuman Primates

The immunogenicity and durability of genetic vaccines are influenced by the composition of gene inserts and choice of delivery vector. DNA vectors are a promising vaccine approach showing efficacy when combined in prime-boost regimens with recombinant protein or viral vectors, but they have shown limited comparative efficacy as a stand-alone platform in primates, due possibly to suboptimal gene expression or cell targeting. Here, regimens using DNA plasmids modified for optimal antigen expression and recombinant adenovirus (rAd) vectors, all encoding the glycoprotein (GP) gene from Angola Marburg virus (MARV), were compared for their ability to provide immune protection against lethal MARV Angola infection. Heterologous DNA-GP/rAd5-GP prime-boost and single-modality rAd5-GP, as well as the DNA-GP-only vaccine, prevented death in all vaccinated subjects after challenge with a lethal dose of MARV Angola. The DNA/DNA vaccine induced humoral responses comparable to those induced by a single inoculation with rAd5-GP, as well as CD4⁺ and CD8⁺ cellular immune responses, with skewing toward CD4⁺ T-cell activity against MARV GP. Vaccine regimens containing rAd-GP, alone or as a boost, exhibited cellular responses with CD8⁺ T-cell dominance. Across vaccine groups, CD8⁺ T-cell subset dominance comprising cells exhibiting a tumor necrosis factor alpha (TNF-α) and gamma interferon (IFN-γ) double-positive functional phenotype was associated with an absence or low frequency of clinical symptoms, suggesting that both the magnitude and functional phenotype of CD8⁺ T cells may determine vaccine efficacy against infection by MARV Angola.The filoviruses Marburgvirus (MARV) and Ebolavirus (EBOV) are endemic primarily to central Africa and cause a severe form of viral hemorrhagic fever. Of all the filovirus strains or species, the Angola strain of MARV is associated with the highest mortality rate (90%) in humans observed to date (26). An increase in natural filovirus outbreak frequency over the past decade and the potential for use to cause deliberate human mortality have focused attention on the need for therapeutics and vaccines against filoviruses. While regulatory pathways have been proposed to facilitate licensing of a preventive vaccine against potently lethal pathogens such as these, there is as yet no licensed vaccine for use in humans, and efforts remain targeted to the optimization of vaccine performance in nonhuman primates (NHP) since this animal model recapitulates many aspects of disease pathogenesis observed in humans.Genetic vaccines are a promising approach for immunization against pathogens that are rapidly changing due to natural evolution, cross-species transmission, or intentional modification. Gene-based vaccines are produced rapidly and can be delivered by a variety of vectors. DNA vectors are advantageous because they are inherently safe and stable and can be used repeatedly without inducing antivector immune responses. However, while filovirus DNA vaccines have demonstrated efficacy in small animal models, efforts to induce protective immunity by injection of plasmid DNA alone into NHP have yielded less encouraging results. EBOV DNA vectors generate immune protection in mice and guinea pigs, but this has not been demonstrated in NHP unless DNA immunization is boosted with a viral vector vaccine (23). MARV DNA fully protects mice and guinea pigs but provides only partial protection in NHP (17). The discordant results between rodent and primate species may be due to the use of slightly modified infectious challenge viruses in rodent models or may reflect underlying differences in vaccine performance and the mechanisms of immune protection between rodents and NHP.In the current study, we examined whether DNA plasmid-based vaccines could be improved to increase potency in NHP and compared immunogenicity of this vaccine modality with those of viral vector and prime-boost approaches. DNA-vectored vaccines were modified by codon optimizing gene target inserts for enhanced expression in primates. These vectors induced antigen-specific cellular and humoral immune responses similar to immunization using a recombinant adenoviral vector and provided protection after lethal challenge with MARV Angola. However, macaques vaccinated with DNA vectors exhibited clinical symptoms associated with MARV hemorrhagic fever (MHF) that were absent in NHP receiving a single inoculation with recombinant adenovirus (rAd) vectors, suggesting qualitative differences in the immune responses elicited by the different modalities.     

A vaccine strategy utilizing a combination of three different chimeric vectors which share specific vaccine antigens

A large number of recombinant of viral and bacterial systems have been engineered as vectors to express foreign genes for vaccination and/or gene therapy. A common problem is the immune response to the vector itself. The presence of anti-vector immune responses may preclude sufficient 'priming' or immunogenicity if pre-existing immune responses are present, or they may impair optimal 'boosting' upon repeated immunization or delivery with the same vector. To circumvent this problem we developed a strategy using different chimeric vectors which share only the expression of common specific antigens desired for immunization. This approach not only has the advantage of avoiding increased anti-vector responses, but allows the use of combinations of vectors which could subsequently present the same or related antigen differently to the immune system as well as at alternative sites to induce the optimal type of immunity against the pathogen of interest.