Developing DNA vaccines that call to dendritic cells

DNA vaccination is a novel immunization strategy that has great potential for the development of vaccines and immune therapeutics. This strategy has been highly effective in mice, while less immunogenic in nonhuman primates and humans. Enhancing DNA vaccine potency remains a challenge. It is likely that APCs, and especially DCs, play a paramount role in the presentation of vaccine antigen to the immune system. A new study reports the synergistic recruitment, expansion, and activation of DCs in vivo in a mouse model through covaccination with plasmids encoding macrophage inflammatory protein-1alpha (MIP-1alpha), fms-like tyrosine kinase 3 ligand (Flt3L), and the DNA vaccine. Such cooperative strategies delivering vaccine in a single, simple platform result in improved cellular immunity in vivo, including enhanced tetramer responses and IFN-gamma secretion by antigen-specific cells.     

Use of simian immunodeficiency viruses for AIDS research

Despite frequent statements to the contrary, there are good animal models for AIDS. In this review, we summarize the properties of one of the most useful animal models: infection of rhesus monkeys with simian immunodeficiency virus (SIV). The SIVs are an extensive group of HIV-related lentiviruses of nonhuman primates. They closely resemble the human AIDS viruses, HIV-1 and HIV-2, in both genetic sequence and biological properties. Some SIV isolates, most notably those derived from macaques and mangabeys, induce AIDS in rhesus monkeys in a time frame suitable for laboratory investigation. Rhesus monkeys are not endangered in the wild, they breed well in captivity, and they are available in reasonably large numbers. Study of SIV has already resulted in seminal contributions regarding the origins of the HIVs, AIDS pathogenesis, and vaccine and therapy research. Continued use of SIV systems will be an important weapon in our arsenal against AIDS.     

Intradermal DNA vaccination enhanced by low-current electroporation improves antigen expression and induces robust cellular and humoral immune responses

Abstract DNA represents an ideal vaccine platform for HIV and many infectious diseases because of its safety, stability, and ease of manufacture. However, the immunogenicity of DNA vaccines has traditionally been low compared with viral vectors, recombinant protein, and live attenuated vaccines. The immunogenicity of DNA vaccines has been significantly enhanced by delivery with in vivo electroporation. Further improvements now allow electroporation to be performed in the dermis, which could potentially improve patient tolerability and may further enhance immunogenicity. In this study we examined how the current of intradermal vaccination impacts antigen expression, inflammation, and the induction of both humoral and cellular immunity in guinea pigs and nonhuman primates. We observed that a lower (0.1 A) current reduced inflammation and improved antigen expression compared with a 0.2 A current. The improved antigen expression resulted in a trend toward higher cellular immune responses but no impact on HIV- and influenza-specific binding titers. This study highlights the need for optimization of electroporation conditions in vivo in order to balance enhanced plasmid transfection with a loss of expression due to tissue inflammation and necrosis. These results suggest that a lower, 0.1-A current may not only improve patient tolerability but also improve immunogenicity.     

A Cationic Nanoemulsion for the Delivery of Next-generation RNA Vaccines

Nucleic acid-based vaccines such as viral vectors, plasmid DNA, and mRNA are being developed as a means to address a number of unmet medical needs that current vaccine technologies have been unable to address. Here, we describe a cationic nanoemulsion (CNE) delivery system developed to deliver a self-amplifying mRNA vaccine. This nonviral delivery system is based on Novartis''s proprietary adjuvant MF59, which has an established clinical safety profile and is well tolerated in children, adults, and the elderly. We show that nonviral delivery of a 9 kb self-amplifying mRNA elicits potent immune responses in mice, rats, rabbits, and nonhuman primates comparable to a viral delivery technology, and demonstrate that, relatively low doses (75 µg) induce antibody and T-cell responses in primates. We also show the CNE-delivered self-amplifying mRNA enhances the local immune environment through recruitment of immune cells similar to an MF59 adjuvanted subunit vaccine. Lastly, we show that the site of protein expression within the muscle and magnitude of protein expression is similar to a viral vector. Given the demonstration that self-amplifying mRNA delivered using a CNE is well tolerated and immunogenic in a variety of animal models, we are optimistic about the prospects for this technology.     

Comparative Analysis of Immune Responses Induced by Vaccination With SIV Antigens by Recombinant Ad5 Vector or Plasmid DNA in Rhesus Macaques

DNA vaccines have undergone important enhancements in their design, formulation, and delivery process. Past literature supports that DNA vaccines are not as immunogenic in nonhuman primates as live vector systems. The most potent recombinant vector system for induction of cellular immune responses in macaques and humans is adenovirus serotype 5 (Ad5), an important benchmark for new vaccine development. Here, we performed a head-to-head evaluation of the Merck Ad5 SIV vaccine and an optimized electroporation (EP) delivered SIV DNA vaccine in macaques. Animals receiving the Ad5 vaccine were immunized three times, whereas the DNA-vaccinated animals were immunized up to four times based on optimized protocols. We observed significant differences in the quantity of IFNγ responses by enzyme-linked immunosorbent spot (ELISpot), greater proliferative capacity of CD8⁺ T cells, and increased polyfunctionality of both CD4⁺ and CD8⁺ T cells in the DNA-vaccinated group. Importantly, Ad5 immunizations failed to boost following the first immunization, whereas DNA responses were continually boosted with all four immunizations demonstrating a major advantage of these improved DNA vaccines. These optimized DNA vaccines induce very different immune phenotypes than traditional Ad5 vaccines, suggesting that they could play an important role in vaccine research and development.     

Removal of gp160 induced bio-hazards for a safe AIDS vaccine candidate.

In the first AIDS vaccine trial, immunizing preparations were based on HIV-1 Env protein (gp160). Immunogenic properties of gp160 which trigger both a humoral and cellular immune response have since justified its use in various vaccine programs, both past and present. Many reports however have underlined deleterious effects on the immune system--anti-HIV-1 enhanced antibodies, anti-CD4 autoantibodies, and inhibition of T cell activation by HIV-1--particularly associated with the Env protein. The present study shows that gp160 presented in a biologically inactivated but immunogenic form, as used in our trial, could avoid these complications. Bio-hazards associated with gp160 which indeed could be removed by appropriate treatment of the native protein, should be taken into consideration in AIDS vaccine programs.     

Which comes first: the antigen or the adjuvant?

Recent attempts to develop an HIV-1 vaccine indicate that viral replication can be limited by the induction of viral-specific T cell responses; however, recent trials of vaccine candidates designed to target CD8⁺ T cell responses were unsuccessful. In this issue, Sui and colleagues used a nonhuman primate model to investigate the effect of various vaccine adjuvants on the efficacy of SIV immunization. Unexpectedly, Sui et al. discovered that animals given adjuvant alone in the absence of SIV antigen exhibited a pronounced decrease in viral load following viral challenge. Vaccination with viral antigens combined with adjuvant correlated with the expansion of a population of cells with similarity to myeloid-derived suppressor cells (MDSCs) that may have suppressed vaccine-elicited T cell responses. Together, these results suggest that both innate and adaptive vaccine-elicited immune responses will need to be considered in future HIV-1 vaccine development.The HIV-1 vaccine field has debated the importance of eliciting strong functional antibody responses to prevent viral invasion of target cells versus eliciting potent T cell responses to kill virus-infected cells. This debate has converged on the consensus that both arms of the immune response will likely be necessary to achieve effective HIV-1 vaccination (1, 2). Additionally, the field has turned toward investigating TLR agonists and other adjuvants to enhance dendritic cell antigen presentation and augment vaccine-elicited responses (3–8). Despite repeated attempts to produce an efficacious HIV-1 vaccine, our knowledge of the adjuvant-specific impact on both the humoral and cellular arms of vaccine-elicited immune responses is still nascent. Moreover, the necessity of HIV-1 vaccine candidates to engage the innate immune system is an ongoing area of investigation (9, 10).     

An underestimated lentivirus model: what can HIV-2 research contribute to the development of an effective HIV-1 vaccine?

The development of an HIV-1 vaccine that would be effective against all existing subtypes and circulating recombinant forms remains one of the great scientific and public health challenges of our generation. One of the major barriers to HIV-1 vaccine development is a lack of understanding of the correlates of protective immunity against the virus. In this context, research has focused on the rare phenomenon of spontaneous control of HIV-1 infection, in groups referred to as 'long-term nonprogressors' and 'elite controllers', together with models of nonprogressive sooty mangabey simian immunodeficiency (SIV) infection in African nonhuman primate hosts such as sooty mangabeys and African green monkeys, in which the majority of animals tolerate high levels of viral replication without development of immunodeficiency or disease. Much less attention has been given to humans infected with the nonpandemic strain HIV-2, derived from the SIV in West Africa, most of whom behave as long-term nonprogressors or viral controllers, while a minority develop disease clinically indistinguishable from AIDS caused by HIV-1. This apparent dichotomous outcome is, based on the evidence accumulated to date, more clearly related to the host immune response than the good clinical outcome of HIV-1 controllers. We propose that complementing research into HIV-1 controllers and nonpathogenic SIV models with the prioritization of HIV-2 research could enhance the HIV-1 vaccine research effort. The absence of disease progression or detectable plasma viral replication in the presence of an effective immune response in most patients living with HIV-2 represents an opportunity to unravel the virus' evolutionary adaptation in human hosts and to establish the correlates of such a protective response.     

Controversies and uncertainties in managing SARS

The development of an HIV-1 vaccine that would be effective against all existing subtypes and circulating recombinant forms remains one of the great scientific and public health challenges of our generation. One of the major barriers to HIV-1 vaccine development is a lack of understanding of the correlates of protective immunity against the virus. In this context, research has focused on the rare phenomenon of spontaneous control of HIV-1 infection, in groups referred to as ‘long-term nonprogressors’ and ‘elite controllers’, together with models of nonprogressive sooty mangabey simian immunodeficiency (SIV) infection in African nonhuman primate hosts such as sooty mangabeys and African green monkeys, in which the majority of animals tolerate high levels of viral replication without development of immunodeficiency or disease. Much less attention has been given to humans infected with the nonpandemic strain HIV-2, derived from the SIV in West Africa, most of whom behave as long-term nonprogressors or viral controllers, while a minority develop disease clinically indistinguishable from AIDS caused by HIV-1. This apparent dichotomous outcome is, based on the evidence accumulated to date, more clearly related to the host immune response than the good clinical outcome of HIV-1 controllers. We propose that complementing research into HIV-1 controllers and nonpathogenic SIV models with the prioritization of HIV-2 research could enhance the HIV-1 vaccine research effort. The absence of disease progression or detectable plasma viral replication in the presence of an effective immune response in most patients living with HIV-2 represents an opportunity to unravel the virus’ evolutionary adaptation in human hosts and to establish the correlates of such a protective response.     

HIV-1 vaccines and co-infection

Vaccines are an economically efficient means of controlling viral infections, and it is likely that a vaccine against HIV-1 will be the most effective way of controlling the global AIDS crisis. However, an effective vaccine has not yet been attainable and in developing countries co-infection with protozoa and other chronic diseases adds another level of complexity to the design of an HIV-1 vaccine. Helminthic and protozoan infections can result in a constant state of immune activation that is characterised by a dominant T helper (Th)2 type of cytokine profile. Such an immune profile is likely to have an adverse impact on the efficacy of an HIV-1 vaccine CD8 cellular immune response and the corresponding Th1 cytokines that are most likely to be important for clearing viral infections.     

HIV-1 vaccines and co-infection

Vaccines are an economically efficient means of controlling viral infections, and it is likely that a vaccine against HIV-1 will be the most effective way of controlling the global AIDS crisis. However, an effective vaccine has not yet been attainable and in developing countries co-infection with protozoa and other chronic diseases adds another level of complexity to the design of an HIV-1 vaccine. Helminthic and protozoan infections can result in a constant state of immune activation that is characterised by a dominant T helper (Th)2 type of cytokine profile. Such an immune profile is likely to have an adverse impact on the efficacy of an HIV-1 vaccine CD8 cellular immune response and the corresponding Th1 cytokines that are most likely to be important for clearing viral infections.     

Recruitment and expansion of dendritic cells in vivo potentiate the immunogenicity of plasmid DNA vaccines

DCs are critical for priming adaptive immune responses to foreign antigens. However, the utility of harnessing these cells in vivo to optimize the immunogenicity of vaccines has not been fully explored. Here we investigate a novel vaccine approach that involves delivering synergistic signals that both recruit and expand DC populations at the site of antigen production. Intramuscular injection of an unadjuvanted HIV-1 envelope (env) DNA vaccine recruited few DCs to the injection site and elicited low-frequency, env-specific immune responses in mice. Coadministration of plasmids encoding the chemokine macrophage inflammatory protein-1alpha (MIP-1alpha) and the DC-specific growth factor fms-like tyrosine kinase 3 ligand with the DNA vaccine resulted in the recruitment, expansion, and activation of large numbers of DCs at the site of inoculation. Consistent with these findings, coadministration of these plasmid cytokines also markedly augmented DNA vaccine---elicited cellular and humoral immune responses and increased protective efficacy against challenge with recombinant vaccinia virus. These data suggest that the availability of mature DCs at the site of inoculation is a critical rate-limiting factor for DNA vaccine immunogenicity. Synergistic recruitment and expansion of DCs in vivo may prove a practical strategy for overcoming this limitation and potentiating immune responses to vaccines as well as other immunotherapeutic strategies.     

A new multi-clade DNA prime/recombinant MVA boost vaccine induces broad and high levels of HIV-1-specific CD8(+) T-cell and humoral responses in mice.

The results presented here are from the preclinical evaluation in BALB/c mice of a DNA prime/modified vaccinia virus Ankara (MVA) boost multi-gene multi-subtype human immunodeficiency virus-1 (HIV-1) vaccine intended for use in humans. The plasmid DNA vaccine was delivered intradermally using a Biojector, and the MVA was delivered intramuscularly by needle. This combination of recombinant DNA and MVA proved to induce extraordinarily strong cellular responses, with more than 80% of the CD8(+) T cells specific for HIV-1 antigens. Furthermore, we show that the DNA priming increases the number of T-cell epitopes recognized after the MVA boost. In the prime/boost-immunized animals, a significant proportion of CD8(+) T cells were stained positive for both interferon-gamma (IFN-gamma) and interleukin-2 (IL-2), a feature that has been associated with control of HIV-1 infection in long-term non-progressors. The HIV-1-specific antibody levels were moderate after the plasmid DNA immunizations but increased dramatically after the MVA boost. Although the initial injection of MVA induced significant levels of vaccinia-neutralizing antibodies, the HIV-specific responses were still significantly boosted by the second MVA immunization. The results from this study demonstrate the potency of this combination of DNA plasmids and MVA construct to induce broad and high levels of immune responses against several HIV-1 proteins of different subtypes.     

Multigene/multisubtype HIV-1 vaccine induces potent cellular and humoral immune responses by needle-free intradermal delivery.

Gene vaccination encounters problems different from those of gene therapy since both a short half-life of the gene and a strong immune response to the gene product are desirable. We have evaluated a DNA vaccine consisting of seven plasmids encoding nine HIV-1 proteins. Using a needle-free delivery device, the Biojector, together with recombinant mouse GM-CSF, this vaccine induced strong gp160 Env- and p24 Gag-specific cellular and humoral immune responses in mice. The rGM-CSF was crucial for inducing both antibodies and antigen-specific CD8(+) T cell responses against both gp160 and p24. A GMP-produced lot of this vaccine, intended for human use, was delivered intradermally or intramuscularly into BALB/c mice at a GLP-accredited animal facility. This vaccine induced strong cellular responses independent of the route of immunization; moreover, no signs of toxicity were detected after histopathological examination of various tissues. Overall, the results indicate that the intradermal delivery of multigene/multisubtype HIV DNA in combination with recombinant GM-CSF is a safe and efficacious strategy for inducing high levels of specific CD8(+) T cells and unusually high titers of antibodies. This vaccine has been approved by the Swedish Medicinal Products Agency and is currently in a Phase I clinical trial.     

8 Br-cAMP enhances both humoral and cell-mediated immune responses induced by an HIV-1 DNA vaccine

From a series of preclinical studies and animal experiments, we have been able to demonstrate that DNA vaccines are a promising tool in strategies for protecting hosts from a variety of infectious diseases. Since the promoter activity of the human cytomegalovirus immediate-early promoter/ enhancer (CMV promoter) is known to be responsive to an elevation in the level of intracellular cAMP, we hypothesized that use of cAMP analogue (8-Bromo adenosine 3'5'-cyclic monophosphate, 8 Br-cAMP) would increase the level of transgene expression supported by the CMV, and enhance the ability of DNA vaccines to evoke an immune response against the transgene product in vivo. To evaluate this hypothesis, immune responses against HIV-1 envelope protein, gp160, an immunogenic HIV-1 component expressed under the control of the CMV promoter, were evaluated in BALB/c mice with or without stimulation by 8 Br-cAMP. DNA vaccine with 8 Br-cAMP was intramuscularly (i.m.) or intranasally (i.n.) administered to BALB/c mice twice on days 0 and 14. Regardless of which route was used, the combination increased the serum IgG antibody (Ab) titer, HIV-1-specific cytotoxic T lymphocyte (CTL) activity and the delayed-type hypersensitivity (DTH) response, compared with the effect of using the vaccine alone. When administered via the i.n. route, the combination also remarkably increased the titer of secretory IgA (sIgA). Moreover, it induced increased production of interferon-gamma with reduction in IL-4 synthesis, and decreased the ratio of serum IgG1/IgG2a. However, these enhancements were not observed when 8 Br-cAMP was coadministered with peptide vaccine or protein antigen. These data suggest that 8 Br-cAMP is able to enhance both humoral and cellular immune responses induced by the DNA vaccine. The induction of T helper type 1 (Th1) immunity against HIV-1 was also enhanced by coadministration of 8 Br-cAMP. A CAT assay study demonstrated that the adjuvant effect of 8 Br-cAMP may be due to the activation of the CMV promoter in the DNA vaccine. The virus challenge experiment in a mouse influenza model also proved our hypothesis.     

A scientific overview of the development of AIDS vaccines.

This article explores the development of AIDS vaccines. It briefly describes the differences between cellular and humoral immune systems and their role in HIV disease. It also considers the use of animal models in vaccine research to simulate human immune system responses to HIV. In addition, various vaccine strategies are explained. These strategies include live attenuated vaccines, subunit vaccines, live vector-based vaccines, DNA vaccines, pseudovirions, whole inactivated virus, and combination products. Advantages and disadvantages for each strategy will be described as well as the current state of progress in vaccine development. The purpose of this article is to educate nurses and other health care providers about the recent developments in vaccine technology as well as to encourage all health care providers to advocate for increased research and development of HIV-1 vaccines.     

AIDS vaccines: an enigma in vaccine development]

The principles of viral vaccine development and the immune responses to vaccination are described. An effective vaccine should stimulate both cellular and humoral immune responses to provide lasting protection against infection and disease. We describe specific problems associated with the development of an AIDS vaccine i.e. lack of experience with human retroviruses, the integration of the HIV genome in cellular DNA, HIV infection of critical cells in the immune system, persistence of HIV in the brain, large variations in the virus envelope gene, immune interactions with the CD-4 protein and the generation of infection enhancing antibodies. The strategies of AIDS vaccine development are discussed and the candidate vaccines are described with a report on the status of each vaccine trial in progress.     

Therapeutic vaccination against HIV: current progress and future possibilities

Although effective in reducing mortality, current antiretroviral therapy for HIV infection involves complex and expensive drug regimens that are toxic and difficult to take. Eradication of HIV reservoirs is not possible with existing therapies. The concept of therapeutic vaccination has been investigated to increase the potency and breadth of anti-HIV immune responses in order to delay or reduce antiretroviral therapy use. A variety of approaches targeted to both cell- and antibody-mediated immunity have been developed, including whole inactivated HIV-1, protein subunits and synthetic peptides, DNA vaccines and a number of viral vectors expressing HIV-1. These investigations have occurred in the absence of a clear understanding of disease pathogenesis or the correlates of protective immunity. At this time, there is no licensed therapeutic vaccine for any viral disease, including HIV; however, this review will consider recent progress in the field and summarize the challenges faced in the development of a therapeutic HIV vaccine.     

Potent immune response against HIV-1 and protection from virus challenge in hu-PBL-SCID mice immunized with inactivated virus-pulsed dendritic cells generated in the presence of IFN-alpha

A major challenge of AIDS research is the development of therapeutic vaccine strategies capable of inducing the humoral and cellular arms of the immune responses against HIV-1. In this work, we evaluated the capability of DCs pulsed with aldrithiol-2-inactivated HIV-1 in inducing a protective antiviral human immune response in SCID mice reconstituted with human PBL (hu-PBL-SCID mice). Immunization of hu-PBL-SCID mice with DCs generated after exposure of monocytes to GM-CSF/IFN-alpha (IFN-DCs) and pulsed with inactivated HIV-1 resulted in a marked induction of human anti-HIV-1 antibodies, which was associated with the detection of anti-HIV neutralizing activity in the serum. This vaccination schedule also promoted the generation of a human CD8+ T cell response against HIV-1, as measured by IFN-gamma Elispot analysis. Notably, when the hu-PBL-SCID mice immunized with antigen-pulsed IFN-DCs were infected with HIV-1, inhibition of virus infection was observed as compared with control animals. These results suggest that IFN-DCs pulsed with inactivated HIV-1 can represent a valuable approach of immune intervention in HIV-1-infected patients.