Wild-type Simian Immunodeficiency Virus Research Articles

Multivariate profiling of African green monkey and rhesus macaque T lymphocytes

The complexity of immune responses limits the usefulness of univariate methods in answering complex immunology questions. To demonstrate the utility of a multivariate approach, we employ such approach to compare T cells of African green monkeys (AGMs) and rhesus macaques (RMs). Among the most prominent distinguishing features we found were lower CD3 and higher CD28 surface expression in AGMs compared to RMs. After in vitro stimulation, a larger proportion of AGM T cells secreted cytokines, especially those producing more than one cytokine (i.e. multifunctional cells). To find out whether multifunctional responses associate with protection in other species, we compared T cells of cynomolgus macaques (CMs) infected with wild-type Simian Immunodeficiency Virus (SIV) to those of CMs infected (vaccinated) with a replication-defective virus. Wild-type SIV infection in macaques leads to simian Acquired Immunodeficiency Syndrome (AIDS), which does not happen in animals previously vaccinated with a replication-defective virus. Interestingly, after in vitro stimulation, multifunctional cells were more abundant among T cells of vaccinated CMs. Our results propose T-cell multifunctionality as a potentially useful marker of immunity, although additional verification is needed. Finally, we hope our multivariate model and its associated validation methods will inform future studies in the field of immunology.

Reproducing SIVΔnef vaccine correlates of protection: trimeric gp41 antibody concentrated at mucosal front lines.

Vaccination with SIVmac239Δnef provides robust protection against subsequent challenge with wild-type simian immunodeficiency virus (SIV), but safety issues have precluded designing an HIV-1 vaccine based on a live-attenuated virus concept. Safe immunogens and adjuvants that could reproduce identified immune correlates of SIVmac239Δnef protection therefore offer an alternative path for development of an HIV vaccine. Here we describe SIV envelope trimeric gp41 (gp41t) immunogens based on a protective correlate of antibodies to gp41t concentrated on the path of virus entry by the neonatal Fc receptor (FcRn) in cervical vaginal epithelium. We developed a gp41t immunogen-monophosphoryl lipid A adjuvant liposomal nanoparticle for intramuscular (i.m.) immunization and a gp41t-Fc immunogen for intranasal immunization for pilot studies in mice, rabbits, and rhesus macaques. Repeated immunizations to mimic persistent antigen exposure in infection elicited gp41t antibodies in rhesus macaques that were detectable in FcRn+ cervical vaginal epithelium, thus recapitulating one key feature of SIVmac239Δnef vaccinated and protected animals. Although this strategy did not reproduce the system of local production of antibody in SIVmac239Δnef-vaccinated animals, passive immunization experiments supported the concept that sufficiently high levels of antibody can be concentrated by the FcRn at mucosal frontlines, thus setting the stage for assessing protection against vaginal challenge by gp41t immunization.

Live attenuated simian immunodeficiency virus vaccination confers superinfection resistance against macrophage-tropic and neurovirulent wild-type SIV challenge.

Vaccination with live attenuated simian immunodeficiency virus (SIV) in non-human primate species provides a means of characterizing the protective processes of retroviral superinfection and may lead to novel advances of human immunodeficiency virus (HIV)/AIDS vaccine design. The minimally attenuated SIVmacC8 vaccine has been demonstrated to elicit early potent protection against pathogenic rechallenge with genetically diverse viral isolates in cynomolgus macaques (Macaca fascicularis). In this study, we have characterized further the biological breadth of this vaccine protection by assessing the ability of both the nef-disrupted SIVmacC8 and its nef-intact counterpart SIVmacJ5 viruses to prevent superinfection with the macrophage/neurotropic SIVmac239/17E-Fr (SIVmac17E-Fr) isolate. Inoculation with either SIVmacC8 or SIVmacJ5 and subsequent detailed characterization of the viral replication kinetics revealed a wide range of virus–host outcomes. Both nef-disrupted and nef-intact immunizing viruses were able to prevent establishment of SIVmac17E-Fr in peripheral blood and secondary lymphoid tissues. Differences in virus kinetics, indicative of an active process, identified uncontrolled replication in one macaque which although able to prevent SIVmac17E-Fr superinfection led to extensive neuropathological complications. The ability to prevent a biologically heterologous, CD4-independent/CCR5+ viral isolate and the macrophage-tropic SIVmac316 strain from establishing infection supports the hypothesis that direct target cell blocking is unlikely to be a central feature of live lentivirus vaccination. These data provide further evidence to demonstrate that inoculation of a live retroviral vaccine can deliver broad spectrum protection against both macrophage-tropic as well as lymphocytotropic viruses. These data add to our knowledge of live attenuated SIV vaccines but further highlight potential safety concerns of vaccinating with a live retrovirus.

Localization of productively infected cells in the spleen and Peyer's patches of rhesus macaques during acute infection with SIVmac239Δnef-enhanced green fluorescent protein.

Improved knowledge of the early targets of productive simian immunodeficiency virus (SIV) infection in the context of tissue architecture could provide important information for our understanding of AIDS pathogenesis and vaccine development. While immunohistochemistry and in situ hybridization have been used to detect SIV in situ, in this article we describe results using an SIV engineered to express green fluorescent protein (GFP). SIVmac239Δnef-enhanced green fluorescent protein (EGFP) has been shown to productively infect cells of the same phenotype as wild-type SIVmac239.1 As shown in Fig. 1, large numbers of productively infected cells expressing GFP can be seen in spleen and Peyer's patches of rhesus macaques 7 days after infection with SIVmac239Δnef-EGFP. FIG. 1. Localization of enhanced green fluorescent protein (EGFP+) cells in spleen (a, c, d) and Peyer's patches (b). EGFP+ cells (green) were present in lymphoid nodules in spleen (a, c, d) and Peyer's patches (b) labeled with PNA (a, b: red) and Ki67 (c, d ... In the spleen and Peyer's patches, most of these productively infected cells were localized to secondary lymphoid follicles. The infected cells were mostly distributed at the edge of germinal centers. As shown in Fig. 1a and b, most of the infected cells (green) were colocalized in the peanut agglutinin (PNA)+ area, which is indicative of germinal centers.2 Since most of the cells in germinal centers should be undergoing proliferation or differentiation, cells were labeled for Ki67 anitbody to further delineate the germinal centers. As shown in Fig. 1c and d, the infected cells (green) were distributed in an area positive for Ki67 (red), which confirmed the above results. Of note, although rare, a few infected cells expressing Ki67 were observed (arrowhead). Germinal centers are the areas in which antigen-specific B lymphocytes proliferate, interact with T helper cells (mostly Tfh) and follicular dendritic cells, and finally differentiate into effector cells.2 As shown above, during acute infection with SIVmac239Δnef-EGFP we find numerous infected cells within germinal centers that could be Tfh cells. Infection and loss of Tfh cells would be expected to have negative impacts on humoral immunity. Infection of Tfh cells might also alter the function of these cells, which might result in abnormal differentiation due to lack of normal T cell help. In infections with wild-type SIV, large numbers of infected cells are detected in the interfollicular areas with lesser numbers in organized lymphoid nodules.3 In contrast, with SIVmac239Δnef, which is attenuated, infection of lymphoid follicles and particularly germinal centers is more prominent. While SIVmac239Δnef-EGFP is attenuated it is likely that the same cell types in germinal centers are early targets for virulent SIV infection of rhesus macaques and HIV-1 infection of humans.

Broadening of CD8+ cell responses in vaccine-based simian immunodeficiency virus controllers

In our prior study on a prophylactic T-cell-based vaccine, some vaccinated macaques controlled a simian immunodeficiency virus (SIV) challenge. These animals allowed viremia in the acute phase but showed persistent viral control after the setpoint. Here, we examined the breadth of postchallenge virus-specific cellular immune responses in these SIV controllers. We previously reported that in a group of Burmese rhesus macaques possessing the MHC haplotype 90-120-Ia, immunization with a Gag-expressing vaccine results in nonsterile control of a challenge with SIVmac239 but not a mutant SIV carrying multiple cytotoxic T lymphocyte (CTL) escape gag mutations. In the present study, we investigated whether broader cellular immune responses effective against the mutant SIV replication are induced after challenge in those vaccinees that maintained wild-type SIVmac239 control. We analyzed cellular immune responses in these SIV controllers (n = 8). These controllers elicited CTL responses directed against SIV non-Gag antigens as well as Gag in the chronic phase. Postvaccinated, prechallenge CD8(+) cells obtained from these animals suppressed wild-type SIV replication in vitro, but mostly had no suppressive effect on the mutant SIV replication, whereas CD8(+) cells in the chronic phase after challenge showed efficient antimutant SIV efficacy. The levels of in-vitro antimutant SIV efficacy of CD8(+) cells correlated with Vif-specific CD8(+) T-cell frequencies. Plasma viremia was kept undetectable even after the mutant SIV superchallenge in the chronic phase. These results suggest that vaccine-based wild-type SIV controllers can acquire CD8(+) cells with the potential to suppress replication of SIV variants carrying CTL escape mutations.

Contribution of Cyclophilin A to determination of simian immunodeficiency virus tropism: A progress update

An understanding of cellular factors that affect viral replication contributes to elucidation of the mechanism for the determination of viral tropism. Cyclophilin A (CypA), a peptidyl-prolyl cis- trans isomerase (PPIase), is an essential host factor for the efficient replication of human immunodeficiency virus type 1 (HIV-1) in human cells. However, its role in simian immunodeficiency virus (SIV) replication has not been determined. In the 2008 US–Japan AIDS panel meeting, I have presented the effect of cyclosporine A (CsA), a PPIase inhibitor, on replication of wild-type SIV. Interestingly, CsA treatment enhanced SIV replication in human cells but abrogated SIV replication in macaque cells, implying a species-specific effect of CsA on SIV replication. After this meeting, analysis using CypA knocked-down human cells indicated that CypA was considered inhibitory for SIV replication. These results suggest possible involvement of CypA in the determination of SIV tropism.

Variability in a dominant block to SIV early reverse transcription in rhesus monkey cells predicts in vivo viral replication and time to death

While it has long been appreciated that there is considerable variability in host containment of HIV/SIV replication, the determinants of that variability are not fully understood. Previous studies demonstrated that the degree of permissivity of a macaque's peripheral blood mononuclear cells (PBMC) for infection with simian immunodeficiency virus (SIV) in vitro predicted that animal's peak plasma virus RNA levels following SIV infection in vivo. The present study was conducted to define the mechanisms underlying the variable intrinsic susceptibility of rhesus monkey PBMC to SIVsmE660 infection. In a cohort of 15 unrelated Indian-origin rhesus monkeys, infectability of PBMC of individual animals with SIVsmE660, as defined by tissue culture infectious dose (TCID50), varied by more than 3 logs and was a stable phenotype over time. Susceptibility of a monkey's PBMC to wild type SIVsmE660 infection correlated with the susceptibility of that monkey's PBMC to infection with VSV-G pseudotyped SIVsm543-GFP. Moreover, the permissivity of an individual monkey's PBMC for infection with this construct correlated with the permissivity of a B-lymphoblastoid cell line (B-LCL) generated from PBMC of the same animal. We found that the degree of intrinsic resistance of monkey B-LCL correlated with the copy number of early reverse transcription (ERT) SIV DNA. The resistance of monkey B-LCL to SIVsmE660 replication could be abrogated by preincubation of cells with the SIV virus-like particles (VLPs) and SIV resistance phenotype could be transferred to a SIV susceptible B-LCL through cell fusion. Finally, we observed a positive correlation between susceptibility of monkey B-LCL to SIV infection with a VSV-G pseudotyped SIV-GFP construct in vitro and both the peak plasma virus RNA levels in vivo and time to death following wild type SIV infection. These findings suggest that a dominant early RT restricting factor that can be saturated by SIV capsid may contribute to the variable resistance to SIV infection in rhesus monkey B-LCL and that this differential intrinsic susceptibility contributes to the clinical outcome of an SIV infection.

Immunization with Single-Cycle SIV Significantly Reduces Viral Loads After an Intravenous Challenge with SIVmac239

Strains of simian immunodeficiency virus (SIV) that are limited to a single cycle of infection were evaluated for the ability to elicit protective immunity against wild-type SIVmac239 infection of rhesus macaques by two different vaccine regimens. Six animals were inoculated at 8-week intervals with 6 identical doses consisting of a mixture of three different envelope variants of single-cycle SIV (scSIV). Six additional animals were primed with a mixture of cytoplasmic domain-truncated envelope variants of scSIV and boosted with two doses of vesicular stomatitis virus glycoprotein (VSV G) trans-complemented scSIV. While both regimens elicited detectable virus-specific T cell responses, SIV-specific T cell frequencies were more than 10-fold higher after boosting with VSV G trans-complemented scSIV (VSV G scSIV). Broad T cell recognition of multiple viral antigens and Gag-specific CD4+ T cell responses were also observed after boosting with VSV G scSIV. With the exception of a single animal in the repeated immunization group, all of the animals became infected following an intravenous challenge with SIVmac239. However, significantly lower viral loads and higher memory CD4+ T cell counts were observed in both immunized groups relative to an unvaccinated control group. Indeed, both scSIV immunization regimens resulted in containment of SIVmac239 replication after challenge that was as good as, if not better than, what has been achieved by other non-persisting vaccine vectors that have been evaluated in this challenge model. Nevertheless, the extent of protection afforded by scSIV was not as good as typically conferred by persistent infection with live, attenuated SIV. These observations have potentially important implications to the design of an effective AIDS vaccine, since they suggest that ongoing stimulation of virus-specific immune responses may be essential to achieving the degree of protection afforded by live, attenuated SIV.

Functional replacement of the R region of simian immunodeficiency virus-based vectors by heterologous elements

Substitution of lentiviral cis-acting elements by heterologous sequences might allow the safety of lentiviral vectors to be enhanced by reducing the risk of homologous recombination and vector mobilization. Therefore, a substitution and deletion analysis of the R region of simian immunodeficiency virus (SIV)-based vectors was performed and the effect of the modifications on packaging and transfer by SIV and human immunodeficiency virus type 1 (HIV-1) particles was analysed. Deletion of the first 7 nt of R reduced vector titres by 10- to 20-fold, whilst deletion of the entire R region led to vector titres that were 1500-fold lower. Replacement of the R region of SIV-based vectors by HIV-1 or Moloney murine sarcoma virus R regions partially restored vector titres. A non-retroviral cellular sequence was also functional, although to a lesser extent. In the absence of tat, modification of the R region had only minor effects on cytoplasmic RNA stability, steady-state levels of vector RNA and packaging, consistent with the known primary function of R during reverse transcription. Although the SIV R region of SIV-based vectors could be replaced functionally by heterologous sequences, the same modifications of R led to a severe replication defect in the context of a replication-competent SIV. As SIV-based vectors containing the HIV-1 R region were transferred less efficiently by HIV-1 particles than wild-type SIV vectors, a match between R and cis-acting elements of the vector construct seems to be more important than a match between R and the Gag or Pol proteins of the vector particle.

Impact of Viral Factors on Very Early In Vivo Replication Profiles in Simian Immunodeficiency Virus SIVagm-Infected African Green Monkeys

To better understand which factors govern the levels of viral loads in early lentiviral infections of primates, we developed a model that allows distinguishing between the influences of host and viral factors on viremia. Herein we report that two species of African green monkeys (Chlorocebus sabaeus and C. pygerythrus) infected with their respective wild-type simian immunodeficiency virus SIVagm viruses (SIVagm.sab92018 and SIVagm.ver644) consistently showed reproducible differences in viremia during primary infection but not at later stages of infection. Cross-infections of SIVagm.sab92018 and SIVagm.ver644 into, respectively, C. pygerythrus and C. sabaeus revealed that the dynamics of viral replication during primary infection were dependent on the viral strain used for the infection but not on the host. Hence, the kinetics of SIVagm.sab92018 and SIVagm.ver644 were similar in both sabaeus and vervet animals, indicating that the difference in viremia levels between the two groups during the early phase of infection was not associated with the host. Coreceptor usage for these two strains showed a larger coreceptor repertoire for SIVagm.sab92018, which is able to efficiently use CXCR4 in addition to CCR5, than for SIVagm.ver644, which showed a classical CCR5 coreceptor usage pattern. These differences could not be explained by different charges of the V3 loop for SIVagm.sab92018 and for SIVagm.ver644. In conclusion, our study showed that the extent of virus replication during the primary infection is primarily dependent on viral determinants.

APOBEC3B and APOBEC3C Are Potent Inhibitors of Simian Immunodeficiency Virus Replication

In the human genome the apolipoprotein B mRNA-editing enzyme catalytic polypeptide (APOBEC)3 gene has expanded into a tandem array of genes termed APOBEC3A-G. Two members of this family, APOBEC3G and APOBEC3F, have been found to have potent activity against virion infectivity factor deficient (Deltavif) human immunodeficiency virus 1 (HIV-1). These enzymes become encapsidated in Deltavif HIV-1 virions and in the next round of infection deaminate the newly synthesized reverse transcripts. The lentiviral Vif protein prevents the deamination by inducing the degradation of APOBEC3G and APOBEC3F. We report here that two additional APOBEC3 family members, APOBEC3B and APOBEC3C, have potent antiviral activity against simian immuno-deficiency virus (SIV), but not HIV-1. Both enzymes were encapsidated in HIV-1 and SIV virions and were active against Deltavif SIV(mac) and SIV(agm). SIV Vif neutralized the antiviral activity of APOBEC3C, but not that of APOBEC3B. APOBEC3B induced abundant G --> A mutations in both wild-type and Deltavif SIV reverse transcripts. APOBEC3C induced substantially fewer mutations. APOBEC3F was found to be active against SIV and sensitive to SIV(mac) Vif. These findings raise the possibility that the different APOBEC3 family members function to neutralize specific lentiviruses.

Vaccination with live attenuated simian immunodeficiency virus for 21 days protects against superinfection

The identification of mechanisms that prevent infection with human immunodeficiency virus (HIV) or simian immunodeficiency virus (SIV) would facilitate the development of an effective AIDS vaccine. In time-course experiments, protection against detectable superinfection with homologous wild-type SIV was achieved within 21 days of inoculation with live attenuated SIV, prior to the development of detectable anti-SIV humoral immunity. Partial protection against superinfection was achieved within 10 days of inoculation with live attenuated SIV, prior to the development of detectable anti-SIV humoral and cellular immunity. Furthermore, co-inoculation of live attenuated SIV with wild-type SIV resulted in a significant reduction in peak virus loads compared to controls that received wild-type SIV alone. These findings imply that innate immunity or non-immune mechanisms are a significant component of early protection against superinfection conferred by inoculation with live attenuated SIV.

Live attenuated HIV vaccines: pitfalls and prospects.

When simian immunodeficiency virus (SIV) deleted in the nef gene caused no disease in macaques and provided protection against wild-type SIV challenge, hopes were high that the removal of nef would convert a pathogenic immunodeficiency virus into a live attenuated vaccine. We seek to highlight recent studies focused on several major issues regarding live attenuated AIDS viruses as vaccine candidates: (1). safety, (2). efficacy, (3). the correlates of immune protection, and (4) the molecular determinants for lentiviral virulence or attenuation. Nef-deletion mutants have retained virulence; compared with wild-type SIV, disease progression was slowed but not abrogated. After long-term observation, all adult macaques given SIVmac239delta3 exhibited immune dysfunction; over 50% had T-cell depletion, and 18% developed AIDS. Vaccine efficacy has been disappointing, with limited or no cross-protection and no protection against homologous virus challenge years after initial vaccination. To date, the correlates of protective immunity have defied precise definition; no dominant mechanism has yet emerged. Data from passive serum transfer and CD8+ T-cell depletion studies have raised the possibility that alternate mechanism of protection may be operative. Due to relentless viral replication and continuous selective pressure, initially benign viruses can generate virulent progeny with unpredictable genotypes. Neither safety nor efficacy of the current live attenuated primate immunodeficiency virus vaccines has withstood the test of time. However, such viruses are invaluable tools to address two key questions: (1). what are the correlates of protection, and (2). what are the molecular determinants of viral immunopathogenesis?

Decreased neurotropism of nef long terminal repeat (nef/LTR)-deleted simian immunodeficiency virus.

Simian immunodeficiency virus (SIV) infection of macaques results in neurological abnormalities similar to those of human immunodeficiency virus (HIV)-associated dementia in humans and is a valuable system for the identification of viral neurotropic and neurovirulence factors. The authors recently established an SIV-macaque model where macaques can be infected with wild-type or nef/LTR-deleted SIVmac239 via administration of purified proviral DNA. In this study, the ability of wild-type and nef/LTR-deleted SIV infections to enter the cerebral spinal fluid (CSF) and brain was analyzed. In situ polymerase chain reaction (PCR) readily detected SIV gag DNA-positive cells in the mid-frontal gyrus and basal ganglia of the wild-type SIV-infected macaques, but not in nef/LTR-deleted SIV-infected or SIV-uninfected macaques. PCR on extracted DNA confirmed the in situ results, with multiple brain regions of the wild-type SIV-infected macaques positive for both gag and wild-type nef, whereas in the nef/LTR-deleted SIV-infected macaques, nef/LTR and gag DNA were undetectable. Further, macaques infected with nef/LTR-deleted SIV, which later became superinfected with wild-type SIV, also remained negative for SIV DNA in the brain by both in situ and extracted DNA techniques, despite having high levels of SIV RNA both in the CSF and plasma. This study provides evidence of the inability of nef/LTR-deleted SIV to initiate central nervous system (CNS) infection and suggests that, in the brain regions examined, nef/LTR-deleted viruses have either diminished neurotropism or insufficient systemic viral replication for entry into the CNS.

Comparative Analysis of the Roles of Simian Immunodeficiency and Bovine Leukemia Virus Matrix Proteins in Gag Assembly in Insect Cells

The role of the matrix (MA) domain of simian immunodeficiency virus (SIV) and bovine leukaemia virus (BLV) Gag in the assembly of virus-like particles (VLP) in insect cells has been investigated. Wild-type SIV and BLV Gag assembled to form discrete VLP structures typical of many retroviruses analysed by similar systems. When amino acids predicated by the three-dimensional structure to be at the interface of SIV MA monomers were deleted, VLP assembly was abolished consistent with a role for MA multimerization in assembly. When amino acids predicted to be in the analogous positions in BLV MA were mutated, however, VLP assembly was not affected. These data indicate that the models of assembly derived from one model retrovirus may not necessarily apply to more distantly related viruses despite the structural similarity present in equivalent Gag domains.

Vaccination with attenuated simian immunodeficiency virus by DNA inoculation.

Delivering attenuated lentivirus vaccines as proviral DNA would be simple and inexpensive. Inoculation of macaques with wild-type simian immunodeficiency virus strain mac239 (SIV(mac239)) DNA or SIV(mac239) DNA containing a single deletion in the 3' nef-long terminal repeat overlap region (nef/LTR) led to sustained SIV infections and AIDS. Injection of SIV(mac239) DNA containing identical deletions in both the 5' LTR and 3' nef/LTR resulted in attenuated SIV infections and substantial protection against subsequent mucosal SIV(mac251) challenge.

In situ hybridization and immunolabelling study of the early replication of simian immunodeficiency virus (SIVmacJ5) in vivo.

The distribution of virus-infected cells in cynomolgus macaques was determined at 4, 7, 14 and 28 days following intravenous challenge with 1000 TCID(50) of the wild-type simian immunodeficiency virus SIVmacJ5 (stock J5C). At each time-point, pairs of macaques were killed humanely and the presence of SIV was determined and quantified in blood, spleen, peripheral and mesenteric lymph nodes, thymus, lung and ileum by virus co-cultivation with C8166 cells, by quantitative DNA PCR or by in situ hybridization (ISH). At day 4 post-infection (p.i.), detection of the virus was sporadic. By day 7 p.i., however, significant SIV loads were detected in the blood and lymphoid tissues by DNA PCR and virus co-cultivation. Large numbers of cells expressing SIV RNA were detected in mesenteric lymph nodes by ISH and significantly fewer (P<0.05) in the spleen. Significant numbers of ISH-positive cells were also observed in sections of ileum. By day 14 p.i., the distribution of SIV was more even in all lymphoid tissues analysed. By day 28, most of the tissues were negative by ISH, but all remained positive by virus isolation and DNA PCR. Immunolabelling of sections of mesenteric lymph node with monoclonal antibodies specific for SIV envelope and Nef largely confirmed the observations from ISH. These results indicate that, even following intravenous challenge, a major site of the initial replication of SIV is gut-associated lymphoid tissue. Vaccines that induce protection at this site may therefore be superior, even against parenteral challenge.

Quintuple deglycosylation mutant of simian immunodeficiency virus SIVmac239 in rhesus macaques: robust primary replication, tightly contained chronic infection, and elicitation of potent immunity against the parental wild-type strain.

We previously generated a mutant of simian immunodeficiency virus (SIV) lacking 5 of a total of 22 N-glycans in its external envelope protein gp120 with no impairment in viral replication capability and infectivity in tissue culture cells. Here, we infected rhesus macaques with this mutant and found that it also replicated robustly in the acute phase but was tightly, though not completely, contained in the chronic phase. Thus, a critical requirement for the N-glycans for the full extent of chronic infection was demonstrated. No evidence indicating reversion to a wild type was obtained during the observation period of more than 40 weeks. Monkeys infected with the mutant were found to tolerate a challenge infection with wild-type SIV very well. Analyses of host responses following challenge revealed no neutralizing antibodies against the challenge virus but strong secondary responses of cytotoxic T lymphocytes against multiple antigens, including Gag-Pol, Nef, and Env. Thus, the quintuple deglycosylation mutant appeared to represent a novel class of SIV live attenuated vaccine.

Vaccins contre les virus de l'immunodéficience humaine

Definite progress has been registered in the field of human immunodeficiency virus type 1 (HIV1) vaccines following the observation that live attenuated simian immunodeficiency virus (SIV) vaccines can provide remarkable protection against wild type SIV or S/HIV challenge in rhesus macaques. Studies of the immune correlates of protection have unravelled the role of the CD8+ T-cell response, both in the form of cytotoxic T cells (CTL) and through the secretion of soluble antiviral factors including β-chemokines RANTES, MIP-1α and MIP-1β. In humans, it has been found that a few persons remain HIV seronegative and PCR negative in PBMC despite frequent HIV exposure by the mucosal route. These uninfected, highly exposed individuals show polyclonal HIV-specific CTL responses together with a secretory IgA (sIgA) response in the genital tract. Therefore, the induction of sIgA at mucosal surfaces and that of systemic and, most likely, mucosal CTL could be the highmark of an efficacious HIV vaccine. Induction of cellular immune responses is being sought at this time with various live recombinant vaccines using as a vector either a poxvirus, an alphavirus or a Salmonella strain. These vaccines have been combined with lipopeptides, naked DNA vectors and/or purified gp 120/gp 140 in various prime-boost combination strategies. Recently, a totally novel vaccine approach based on the use of the non-structural Tat or Tat and Rev proteins has also been shown to induce efficient protection from challenge with pathogenic S/HIV or SIV in vaccinated macaques. There is also hope that HIV vaccines could be used in a therapeutic approach in combination with antiviral chemotherapy to help control viral loads after therapy is stopped. However, we are still far from a practical, efficacious HIV vaccine that would be cheap, efficacious, and easy to administer on a global scale.

Immunization with live attenuated simian immunodeficiency virus induces strong type 1 T helper responses and beta-chemokine production.

Immunization with live attenuated simian immunodeficiency virus (SIV) strains has proved to be one of the most effective strategies to induce protective immunity in the SIV/macaque model. To better understand the role that CD4(+) T helper responses may play in mediating protection in this model, we characterized SIV-specific proliferative and cytokine responses in macaques immunized with live attenuated SIV strains. Macaques chronically infected with live attenuated SIV had strong proliferative responses to SIV proteins, with stimulation indices of up to 74. The magnitude of the proliferative response to SIV Gag varied inversely with the degree of attenuation; Gag-specific but not envelope-specific responses were lower in animals infected with more highly attenuated SIV strains. SIV-specific stimulation of lymphocytes from vaccinated macaques resulted in secretion of interferon-gamma, IL-2, regulated-upon-activation, normal T cells expressed and secreted (RANTES), macrophage inflammatory protein (MIP)-1alpha, and MIP-1beta but not IL-4 or IL-10. Intracellular flow cytometric analysis documented that, in macaques vaccinated with SIVmac239Deltanef, up to 2% of all CD4(+)T cells were specific for SIV p55. The ability of live attenuated SIV to induce a strong, sustained type 1 T helper response may play a role in the success of this vaccination approach to generate protection against challenge with wild-type SIV.