Temporal and anatomic relationship between virus replication and cytokine gene expression after vaginal simian immunodeficiency virus infection

The current knowledge about early innate immune responses at mucosal sites of human immunodeficiency virus (HIV) entry is limited but likely to be important in the design of effective HIV vaccines against heterosexual transmission. This study examined the temporal and anatomic relationship between virus replication, lymphocyte depletion, and cytokine gene expression levels in mucosal and lymphoid tissues in a vaginal-transmission model of HIV in rhesus macaques. The results of the study show that the kinetics of cytokine gene expression levels in the acute phase of infection are positively correlated with virus replication in a tissue. Thus, cytokine responses after vaginal simian immunodeficiency virus (SIV) inoculation are earliest and strongest in mucosal tissues of the genital tract and lowest in systemic lymphoid tissues. Importantly, the early cytokine response was dominated by the induction of proinflammatory cytokines, while the induction of cytokines with antiviral activity, alpha/beta interferon, occurred too late to prevent virus replication and dissemination. Thus, the early cytokine response favors immune activation, resulting in the recruitment of potential target cells for SIV. Further, unique cytokine gene expression patterns were observed in distinct anatomic locations with a rapid and persistent inflammatory response in the gut that is consistent with the gut being the major site of early CD4 T-cell depletion in SIV infection.     

Protection of human immunodeficiency virus type 2-exposed seronegative macaques from mucosal simian immunodeficiency virus transmission.

At present it is not known which form of immunity would be most effective against infection with human immunodeficiency virus (HIV). To evaluate the possible role of cellular immunity, we examined whether four HIV type 2-exposed but seronegative macaques developed cellular immune responses and determined whether these exposed macaques were resistant to mucosal transmission of simian immunodeficiency virus (SIV). Following intrarectal challenge with SIV, 2 monkeys were protected against detectable SIV replication and another showed suppressed viral replication compared to 14 persistently infected controls. The two protected monkeys demonstrated SIV-specific cytotoxic T lymphocytes before as well as after SIV challenge. Here we provide evidence that activation of the cell-mediated arm of the immune system only, without antibody formation, can control SIV replication in macaques. The results imply that vaccines that stimulate a strong and broad cellular immune response could prevent mucosal HIV transmission.     

Systemic vaccination prevents the total destruction of mucosal CD4 T cells during acute SIV challenge

BACKGROUND: Acute human immunodeficiency virus (HIV)/simian immunodeficiency virus (SIV) infections are accompanied by a systemic loss of memory CD4 T cells, with mucosal sites serving as a major site for viral replication, dissemination and CD4 T cell depletion. Protecting the mucosal CD4 T cell compartment thus is critical to contain HIV, and preserve the integrity of the mucosal immune system. The primary objective of this study was to determine if systemic vaccination with DNA/rAd-5 encoding SIV-mac239-env, gag and pol could prevent the destruction of CD4 T cells in mucosal tissues. METHODS: Rhesus macaques were immunized with DNA/r-Ad-5 encoding SIV genes and compared with those immunized with sham vectors following high dose intravenous challenge with SIVmac251. SIV specific CD4 and CD8 T cell responses, cell associated viral loads and mucosal CD4 T cell dynamics were evaluated. RESULTS: Strong SIV specific immune responses were induced in mucosal tissues of vaccinated animals as compared with sham controls. These responses expanded rapidly following challenge suggesting a strong anamnestic response. Immune responses were associated with a decrease in cell associated viral loads, and a loss of fewer mucosal CD4 T cells. Approximately 25% of mucosal CD4 T cells were preserved in vaccinated animals as compared with <5% in sham controls. These results demonstrate that systemic immunization strategies can induce immune responses in mucosal tissues that can protect mucosal CD4 T cells from complete destruction following challenge. CONCLUSIONS: Preservation of mucosal CD4 T cells can contribute to maintaining immune competence in mucosal tissues and provide a substantial immune benefit to the vaccinees.     

Both mucosal and systemic routes of immunization with the live,attenuated NYVAC/simian immunodeficiency virus SIV(gpe) recombinant vaccine result in gag-specific CD8(+) T-cell responses in mucosal tissues of macaques

As most human immunodeficiency virus (HIV) infection occurs via mucosal surfaces, an important goal of vaccination may be the induction of virus-specific immune responses at mucosal sites to contain viral infection early on. Here we designed a study in macaques carrying the major histocompatibility complex class I Mamu-A(*)01 molecule to assess the capacity of the highly attenuated poxvirus NYVAC/simian immunodeficiency virus (SIV) SIV(gpe) vaccine candidate administered by the intranasal, intramuscular, or intrarectal route to induce mucosal immunity. All macaques, including one naive macaque, were exposed to SIV(mac251) by the intrarectal route and sacrificed 48 h after infection. The kinetics of immune response at various time points following immunization with NYVAC/SIV(gpe) and the anamnestic response to SIV(mac251) at 48 h after challenge were assessed in blood, in serial rectal and vaginal biopsy samples, and in tissues at euthanasia with an SIV(mac) Gag-specific tetramer. In addition, at euthanasia, antigen-specific cells producing gamma interferon or tumor necrosis factor alpha from the jejunum lamina propria were quantified in all macaques. Surprisingly, antigen-specific CD8(+) T cells were found in the mucosal tissues of all immunized macaques regardless of whether the vaccine was administered by a mucosal route (intranasal or intrarectal) or systemically. In addition, following mucosal SIV(mac251) challenge, antigen-specific responses were mainly confined to mucosal tissues, again regardless of the route of immunization. We conclude that immunization with a live vector vaccine results in the appearance of CD8(+) T-cell responses at mucosal sites even when the vaccine is delivered by nonmucosal routes.     

Oral immunization with recombinant Mycobacterium bovis BCG simian immunodeficiency virus nef induces local and systemic cytotoxic T-lymphocyte responses in mice.

Recombinant live Mycobacterium bovis BCG vectors (rBCG) induce strong cellular and humoral immune responses against various antigens after either systemic or oral immunization of mice. Cytotoxic T-lymphocyte (CTL) responses may contribute to the control of human immunodeficiency virus (HIV) or simian immunodeficiency virus (SIV) infections whose portal of entry is the gastrointestinal or genital mucosa. In this study, we immunized BALB/c mice with a recombinant BCG SIV nef and observed its behavior in oropharyngeal and target organ lymphoid tissues. The cellular immune responses, particularly the intestinal intraepithelial and systemic CTL responses, were investigated. The results showed that rBCG SIV nef translocated the oropharyngeal mucosa and intestinal epithelium. It diffused to and persisted in target lymphoid organs. Specific SIV Nef peptide proliferative responses and cytokine production were observed. Strong systemic and mucosal CTL responses were induced. In particular, we demonstrated direct specific anti-Nef CTL in intestinal intraepithelial CD8beta+ T cells. These findings provide evidence that orally administered rBCG SIV nef may contribute to local defenses against viral invasion. Therefore, rBCG SIV nef could be a candidate vaccine to protect against SIV infection and may be used to develop an oral rBCG HIV nef vaccine.     

Induction of mucosal protection against primary, heterologous simian immunodeficiency virus by a DNA vaccine

An effective vaccine against human immunodeficiency virus (HIV) should protect against mucosal transmission of genetically divergent isolates. As a safe alternative to live attenuated vaccines, the immunogenicity and protective efficacy of a DNA vaccine containing simian immunodeficiency virus (SIV) strain 17E-Fr (SIV/17E-Fr) gag-pol-env was analyzed in rhesus macaques. Significant levels of cytotoxic T lymphocytes (CTL), but low to undetectable serum antibody responses, were observed following multiple immunizations. SIV-specific mucosal antibodies and CTL were also detected in rectal washes and gut-associated lymphoid tissues, respectively. Vaccinated and naive control monkeys were challenged intrarectally with SIV strain DeltaB670 (SIV/DeltaB670), a primary isolate whose env is 15% dissimilar to that of the vaccine strain. Four of seven vaccinees were protected from infection as determined by the inability to identify viral RNA or DNA sequences in the peripheral blood and the absence of anamnestic antibody responses postchallenge. This is the first report of mucosal protection against a primary pathogenic, heterologous isolate of SIV by using a commercially viable vaccine approach. These results support further development of a DNA vaccine for protection against HIV.     

Intravenous inoculation of replication-deficient recombinant vaccinia virus DIs expressing simian immunodeficiency virus gag controls highly pathogenic simian-human immunodeficiency virus in monkeys

To be effective, a vaccine against human immunodeficiency virus type 1 (HIV-1) must induce virus-specific T-cell responses and it must be safe for use in humans. To address these issues, we developed a recombinant vaccinia virus DIs vaccine (rDIsSIVGag), which is nonreplicative in mammalian cells and expresses the full-length gag gene of simian immunodeficiency virus (SIV). Intravenous inoculation of 10(6) PFU of rDIsSIVGag in cynomologus macaques induced significant levels of gamma interferon (IFN-gamma) spot-forming cells (SFC) specific for SIV Gag. Antigen-specific lymphocyte proliferative responses were also induced and were temporally associated with the peak of IFN-gamma SFC activity in each macaque. In contrast, macaques immunized with a vector control (rDIsLacZ) showed no significant induction of antigen-specific immune responses. After challenge with a highly pathogenic simian-human immunodeficiency virus (SHIV), CD4(+) T lymphocytes were maintained in the peripheral blood and lymphoid tissues of the immunized macaques. The viral set point in plasma was also reduced in these animals, which may be related to the enhancement of virus-specific intracellular IFN-gamma(+) CD8(+) cell numbers and increased antibody titers after SHIV challenge. These results demonstrate that recombinant DIs has potential for use as an HIV/AIDS vaccine.     

Immunization of newborn rhesus macaques with simian immunodeficiency virus (SIV) vaccines prolongs survival after oral challenge with virulent SIVmac251

There is an urgent need for active immunization strategies that, if administered shortly after birth, could protect infants in developing countries from acquiring human immunodeficiency virus (HIV) infection through breast-feeding. Better knowledge of the immunogenic properties of vaccine candidates in infants and of the effect of maternal antibodies on vaccine efficacy will aid in the development of such a neonatal HIV vaccine. Simian immunodeficiency virus (SIV) infection of infant macaques is a useful animal model of pediatric HIV infection with which to address these questions. Groups of infant macaques were immunized at birth and 3 weeks of age with either modified vaccinia virus Ankara (MVA) expressing SIV Gag, Pol, and Env (MVA-SIVgpe) or live-attenuated SIVmac1A11. One MVA-SIVgpe-immunized group had maternally derived anti-SIV antibodies prior to immunization. Animals were challenged orally at 4 weeks of age with a genetically heterogeneous stock of virulent SIVmac251. Although all animals became infected, the immunized animals mounted better antiviral antibody responses, controlled virus levels more effectively, and had a longer disease-free survival than the unvaccinated infected monkeys. Maternal antibodies did not significantly reduce the efficacy of the MVA-SIVgpe vaccine. In conclusion, although the tested vaccines delayed the onset of AIDS, further studies are warranted to determine whether a vaccine that elicits stronger early immune responses at the time of virus exposure may be able to prevent viral infection or AIDS in infants.     

Mucosal priming of simian immunodeficiency virus-specific cytotoxic T-lymphocyte responses in rhesus macaques by the Salmonella type III secretion antigen delivery system

Nearly all human immunodeficiency virus (HIV) infections are acquired mucosally, and the gut-associated lymphoid tissues are important sites for early virus replication. Thus, vaccine strategies designed to prime virus-specific cytotoxic T lymphocyte (CTL) responses that home to mucosal compartments may be particularly effective at preventing or containing HIV infection. The Salmonella type III secretion system has been shown to be an effective approach for stimulating mucosal CTL responses in mice. We therefore tested DeltaphoP-phoQ attenuated strains of Salmonella enterica serovar Typhimurium and S. enterica serovar Typhi expressing fragments of the simian immunodeficiency virus (SIV) Gag protein fused to the type III-secreted SopE protein for the ability to prime virus-specific CTL responses in rhesus macaques. Mamu-A*01(+) macaques were inoculated with three oral doses of recombinant Salmonella, followed by a peripheral boost with modified vaccinia virus Ankara expressing SIV Gag (MVA Gag). Transient low-level CTL responses to the Mamu-A*01 Gag(181-189) epitope were detected following each dose of SALMONELLA: After boosting with MVA Gag, strong Gag-specific CTL responses were consistently detected, and tetramer staining revealed the expansion of Gag(181-189)-specific CD8(+) T-cell responses in peripheral blood. A significant percentage of the Gag(181-189)-specific T-cell population in each animal also expressed the intestinal homing receptor alpha4beta7. Additionally, Gag(181-189)-specific CD8(+) T cells were detected in lymphocytes isolated from the colon. Yet, despite these responses, Salmonella-primed/MVA-boosted animals did not exhibit improved control of virus replication following a rectal challenge with SIVmac239. Nevertheless, this study demonstrates the potential of mucosal priming by the Salmonella type III secretion system to direct SIV-specific cellular immune responses to the gastrointestinal mucosa in a primate model.     

ALVAC-SIV-gag-pol-env-based vaccination and macaque major histocompatibility complex class I (A*01) delay simian immunodeficiency virus SIVmac-induced immunodeficiency.

T-cell-mediated immune effector mechanisms play an important role in the containment of human immunodeficiency virus/simian immunodeficiency virus (HIV/SIV) replication after infection. Both vaccination- and infection-induced T-cell responses are dependent on the host major histocompatibility complex classes I and II (MHC-I and MHC-II) antigens. Here we report that both inherent, host-dependent immune responses to SIVmac251 infection and vaccination-induced immune responses to viral antigens were able to reduce virus replication and/or CD4+ T-cell loss. Both the presence of the MHC-I Mamu-A*01 genotype and vaccination of rhesus macaques with ALVAC-SIV-gag-pol-env (ALVAC-SIV-gpe) contributed to the restriction of SIVmac251 replication during primary infection, preservation of CD4+ T cells, and delayed disease progression following intrarectal challenge exposure of the animals to SIV(mac251 (561)). ALVAC-SIV-gpe immunization induced cytotoxic T-lymphocyte (CTL) responses cumulatively in 67% of the immunized animals. Following viral challenge, a significant secondary virus-specific CD8+ T-cell response was observed in the vaccinated macaques. In the same immunized macaques, a decrease in virus load during primary infection (P = 0.0078) and protection from CD4 loss during both acute and chronic phases of infection (P = 0.0099 and P = 0.03, respectively) were observed. A trend for enhanced survival of the vaccinated macaques was also observed. Neither boosting the ALVAC-SIV-gpe with gp120 immunizations nor administering the vaccine by the combination of mucosal and systemic immunization routes increased significantly the protective effect of the ALVAC-SIV-gpe vaccine. While assessing the role of MHC-I Mamu-A*01 alone in the restriction of viremia following challenge of nonvaccinated animals with other SIV isolates, we observed that the virus load was not significantly lower in Mamu-A*01-positive macaques following intravenous challenge with either SIV(mac251 (561)) or SIV(SME660). However, a significant delay in CD4+ T-cell loss was observed in Mamu-A*01-positive macaques in each group. Of interest, in the case of intravenous or intrarectal challenge with the chimeric SIV/HIV strains SHIV(89.6P) or SHIV(KU2), respectively, MHC-I Mamu-A*01-positive macaques did not significantly restrict primary viremia. The finding of the protective effect of the Mamu-A*01 molecule parallels the protective effect of the B*5701 HLA allele in HIV-1-infected humans and needs to be accounted for in the evaluation of vaccine efficacy against SIV challenge models.     

Induction of CD8+ cells able to suppress CCR5-tropic simian immunodeficiency virus SIVmac239 replication by controlled infection of CXCR4-tropic simian-human immunodeficiency virus in vaccinated rhesus macaques

Recent recombinant viral vector-based AIDS vaccine trials inducing cellular immune responses have shown control of CXCR4-tropic simian-human immunodeficiency virus (SHIV) replication but difficulty in containment of pathogenic CCR5-tropic simian immunodeficiency virus (SIV) in rhesus macaques. In contrast, controlled infection of live attenuated SIV/SHIV can confer the ability to contain SIV superchallenge in macaques. The specific immune responses responsible for this control may be induced by live virus infection but not consistently by viral vector vaccination, although those responses have not been determined. Here, we have examined in vitro anti-SIV efficacy of CD8+ cells in rhesus macaques that showed prophylactic viral vector vaccine-based control of CXCR4-tropic SHIV89.6PD replication. Analysis of the effect of CD8+ cells obtained at several time points from these macaques on CCR5-tropic SIVmac239 replication in vitro revealed that CD8+ cells in the chronic phase after SHIV challenge suppressed SIV replication more efficiently than those before challenge. SIVmac239 superchallenge of two of these macaques at 3 or 4 years post-SHIV challenge was contained, and the following anti-CD8 antibody administration resulted in transient CD8+ T-cell depletion and appearance of plasma SIVmac239 viremia in both of them. Our results indicate that CD8+ cells acquired the ability to efficiently suppress SIV replication by controlled SHIV infection, suggesting the contribution of CD8+ cell responses induced by controlled live virus infection to containment of HIV/SIV superinfection.     

Mucosal model of genital immunization in male rhesus macaques with a recombinant simian immunodeficiency virus p27 antigen.

Human immunodeficiency virus (HIV) can be transmitted through infected seminal fluid or vaginal or rectal secretions during heterosexual or homosexual intercourse. To prevent mucosal transmission and spread to the regional lymph nodes, an effective vaccine may need to stimulate immune responses at the genitourinary mucosa. In this study, we have developed a mucosal model of genital immunization in male rhesus macaques, by topical urethral immunization with recombinant simian immunodeficiency virus p27gag, expressed as a hybrid Ty virus-like particle (Ty-VLP) and covalently linked to cholera toxin B subunit. This treatment was augmented by oral immunization with the same vaccine but with added killed cholera vibrios. Polymeric secretory immunoglobulin A (sIgA) and IgG antibodies to p27 were induced in urethral secretions, urine, and seminal fluid. This raises the possibility that the antibodies may function as a primary mucosal defense barrier against SIV (HIV) infection. The regional lymph nodes which constitute the genital-associated lymphoid tissue contained p27-specific CD4+ proliferative and helper T cells for antibody synthesis by B cells, which may function as a secondary immune barrier to infection. Blood and splenic lymphocytes also showed p27-sensitized CD4+ T cells and B cells in addition to serum IgG and IgA p27-specific antibodies; this constitutes a third level of immunity against dissemination of the virus. A comparison of genito-oral with recto-oral and intramuscular routes of immunization suggests that only genito-oral immunization elicits specific sIgA and IgG antibodies in the urine, urethra, and seminal fluid. Both genito-oral and recto-oral immunizations induced T-cell and B-cell immune responses in regional lymph nodes, with preferential IgA antibody synthesis. The mucosal route of immunization may prevent not only virus transmission through the genital mucosa but also dissemination and latency of the virus in the draining lymph nodes.     

The relationship between simian immunodeficiency virus RNA levels and the mRNA levels of alpha/beta interferons (IFN-alpha/beta) and IFN-alpha/beta-inducible Mx in lymphoid tissues of rhesus macaques during acute and chronic infection

To define the role of alpha/beta interferons (IFN-alpha/beta) in simian immunodeficiency virus (SIV) infection, IFN-alpha and IFN-beta mRNA levels and mRNA levels of Mx, an antiviral effector molecule, were determined in lymphoid tissues of rhesus macaques infected with pathogenic SIV. IFN-alpha/beta responses were induced during the acute phase and persisted in various lymphoid tissues throughout the chronic phase of infection. IFN-alpha/beta responses were most consistent in tissues with high viral RNA levels; thus, IFN-alpha/beta responses were not generally associated with effective control of SIV replication. IFN-alpha/beta responses were differentially regulated in different lymphoid tissues and at different stages of infection. The most consistent IFN-alpha/beta responses in acute and chronic SIV infection were observed in peripheral lymph nodes. In the spleen, only a transient increase in IFN-alpha/beta mRNA levels during acute SIV infection was observed. Further, IFN-alpha and IFN-beta mRNA levels showed a tissue-specific expression pattern during the chronic, but not the acute, phase of infection. In the acute phase of infection, SIV RNA levels in lymphoid tissues of rhesus macaques correlated with mRNA levels of both IFN-alpha and IFN-beta, whereas during chronic SIV infection only increased IFN-alpha mRNA levels correlated with the level of virus replication in the same tissues. In lymphoid tissues of all SIV-infected monkeys, higher viral RNA levels were associated with increased Mx mRNA levels. We found no evidence that monkeys with increased Mx mRNA levels in lymphoid tissues had enhanced control of virus replication. In fact, Mx mRNA levels were associated with high viral RNA levels in lymphoid tissues of chronically infected animals.     

Antiviral therapy during primary simian immunodeficiency virus infection fails to prevent acute loss of CD4+ T cells in gut mucosa but enhances their rapid restoration through central memory T cells

Gut-associated lymphoid tissue (GALT) is an early target of human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) and a site for severe CD4+ T-cell depletion. Although antiretroviral therapy (ART) is effective in suppressing HIV replication and restoring CD4+ T cells in peripheral blood, restoration in GALT is delayed. The role of restored CD4+ T-cell help in GALT during ART and its impact on antiviral CD8+ T-cell responses have not been investigated. Using the SIV model, we investigated gut CD4+ T-cell restoration in infected macaques, initiating ART during either the primary stage (1 week postinfection), prior to acute CD4+ cell loss (PSI), or during the chronic stage at 10 weeks postinfection (CSI). ART led to viral suppression in GALT and peripheral blood mononuclear cells of PSI and CSI animals at comparable levels. CSI animals had incomplete CD4+ T-cell restoration in GALT. In PSI animals, ART did not prevent acute CD4+ T-cell loss by 2 weeks postinfection in GALT but supported rapid and complete CD4+ T-cell restoration thereafter. This correlated with an accumulation of central memory CD4+ T cells and better suppression of inflammation. Restoration of CD4+ T cells in GALT correlated with qualitative changes in SIV gag-specific CD8+ T-cell responses, with a dominance of interleukin-2-producing responses in PSI animals, while both CSI macaques and untreated SIV-infected controls were dominated by gamma interferon responses. Thus, central memory CD4+ T-cell levels and qualitative antiviral CD8+ T-cell responses, independent of viral suppression, were the immune correlates of gut mucosal immune restoration during ART.     

Dominant induction of vaccine antigen-specific cytotoxic T lymphocyte responses after simian immunodeficiency virus challenge

Cytotoxic T lymphocyte (CTL) responses are crucial for the control of human and simian immunodeficiency virus (HIV and SIV) replication. A promising AIDS vaccine strategy is to induce CTL memory resulting in more effective CTL responses post-viral exposure compared to those in natural HIV infections. We previously developed a CTL-inducing vaccine and showed SIV control in some vaccinated rhesus macaques. These vaccine-based SIV controllers elicited vaccine antigen-specific CTL responses dominantly in the acute phase post-challenge. Here, we examined CTL responses post-challenge in those vaccinated animals that failed to control SIV replication. Unvaccinated rhesus macaques possessing the major histocompatibility complex class I haplotype 90-088-Ij dominantly elicited SIV non-Gag antigen-specific CTL responses after SIV challenge, while those induced with Gag-specific CTL memory by prophylactic vaccination failed to control SIV replication with dominant Gag-specific CTL responses in the acute phase, indicating dominant induction of vaccine antigen-specific CTL responses post-challenge even in non-controllers. Further analysis suggested that prophylactic vaccination results in dominant induction of vaccine antigen-specific CTL responses post-viral exposure but delays SIV non-vaccine antigen-specific CTL responses. These results imply a significant influence of prophylactic vaccination on CTL immunodominance post-viral exposure, providing insights into antigen design in development of a CTL-inducing AIDS vaccine.     

Control of simian/human immunodeficiency virus viremia and disease progression after IL-2-augmented DNA-modified vaccinia virus Ankara nasal vaccination in nonhuman primates

A successful HIV vaccine may need to stimulate antiviral immunity in mucosal and systemic immune compartments, because HIV transmission occurs predominantly at mucosal sites. We report here the results of a combined DNA-modified vaccinia virus Ankara (MVA) vaccine approach that stimulated simian/human immunodeficiency virus (SHIV)-specific immune responses by vaccination at the nasal mucosa. Fifteen male rhesus macaques, divided into three groups, received three nasal vaccinations on day 1, wk 9, and wk 25 with a SHIV DNA plasmid producing noninfectious viral particles (group 1), or SHIV DNA plus IL-2/Ig DNA (group 2), or SHIV DNA plus IL-12 DNA (group 3). On wk 33, all macaques were boosted with rMVA expressing SIV Gag-Pol and HIV Env 89.6P, administered nasally. Humoral responses were evaluated by measuring SHIV-specific IgG and neutralizing Abs in plasma, and SHIV-specific IgA in rectal secretions. Cellular responses were monitored by evaluating blood-derived virus-specific IFN-gamma-secreting cells and TNF-alpha-expressing CD8+ T cells, and blood- and rectally derived p11C tetramer-positive T cells. Many of the vaccinated animals developed both mucosal and systemic humoral and cell-mediated anti-SHIV immune responses, although the responses were not homogenous among animals in the different groups. After rectal challenge of vaccinated and naive animals with SHIV89.6P, all animals became infected. However a subset, including all group 2 animals, were protected from CD4+ T cell loss and AIDS development. Taken together, these data indicate that nasal vaccination with SHIV-DNA plus IL-2/Ig DNA and rMVA can provide significant protection from disease progression.     

New prospects for the development of a vaccine against human immunodeficiency virus type 1. An overview

During the past few years, definite progress has been made in the field of human immunodeficiency virus type 1 (HIV-1) vaccines. Initial attempts using envelope gp120 or gp140 from T-cell line-adapted (TCLA) HIV-1 strains to vaccinate chimpanzees showed that neutralizing antibody-based immune responses were protective against challenge with homologous TCLA virus strains or strains with low replicative capacity, but these neutralizing antibodies remained inactive when tested on primary HIV-1 isolates, casting doubts on the efficacy of gp120-based vaccines in the natural setting. Development of a live attenuated simian immunodeficiency virus (SIV) vaccine was undertaken in the macaque model using whole live SIV bearing multiple deletions in the nef, vpr and vpx genes. This vaccine provided remarkable protective efficacy against wild-type SIV challenge, but the deletion mutants remain pathogenic, notably in neonate monkeys. Study of the mechanisms of protection in the SIV model unravelled the importance of the T-cell responses, whether in the form of cytotoxic T-lymphocyte (CTL) killing activity, or in that of antiviral factor secretion of cytokines, beta-chemokines and other unidentified antiviral factors by CD8+ T-cells. Induction of such a response is being sought at this time using various live recombinant virus vaccines, either poxvirus or alphavirus vectors or DNA vectors, which can be combined together or with a gp120/gp140 boost in various prime-boost combination strategies. New vectors include attenuated vaccinia virus NYVAC, modified vaccinia strain Ankara (MVA), Semliki Forest virus, Venezuelan equine encephalitis virus, and Salmonellas. Recent DNA prime-poxvirus boost combination regimens have generated promising protection results against SIV or SIV/HIV (SHIV) challenge in macaque models. Emphasis is also put on the induction of a mucosal immune response, involving both a secretory IgA response and a mucosal CTL response which could constitute a 'first line of defence' in the vaccinated host. Finally, a totally novel vaccine approach based on the use of Tat or Tat and Rev antigens has been shown to induce efficient protection from challenge with pathogenic SIV or SHIV in vaccinated macaques. The only vaccine in phase 3 clinical trials in human volunteers is a gp120-based vaccine, AIDSVAX. A prime-boost combination of a recombinant canarypoxvirus and a subunit gp120 vaccine is in phase 2. Emphasis has been put recently on the necessity of testing prototype vaccines in developing countries using immunogens derived from local virus strains. Trial sites have thus been identified in Kenya, Uganda, Thailand and South Africa where phase I trials have begun or are expected to start presently.     

Novel adeno-associated virus vector vaccine restricts replication of simian immunodeficiency virus in macaques

Gene transfer vectors based on recombinant adeno-associated virus (rAAV) are simple, versatile, and safe. While the conventional applications for rAAV vectors have focused on delivery of therapeutic genes, we have developed the system for delivery of vaccine antigens. In particular, we are interested in generating rAAV vectors for use as a prophylactic human immunodeficiency virus type 1 (HIV-1) vaccine. To that end, we constructed vaccine vectors that expressed genes from the simian immunodeficiency virus (SIV) for evaluation in the monkey SIV model. After a single intramuscular dose, rAAV/SIV vaccines elicited SIV-specific T cells and antibodies in macaques. Furthermore, immunized animals were able to significantly restrict replication of a live, virulent SIV challenge. These data suggest that rAAV vaccine vectors induced biologically relevant immune responses, and thus, warrant continued development as a viable HIV-1 vaccine candidate.