Stoichiometry of antibody neutralization of human immunodeficiency virus type 1

The human immunodeficiency virus envelope glycoproteins function as trimers on the viral surface, where they are targeted by neutralizing antibodies. Different monoclonal antibodies neutralize human immunodeficiency virus type 1 (HIV-1) infectivity by binding to structurally and functionally distinct moieties on the envelope glycoprotein trimer. By measuring antibody neutralization of viruses with mixtures of neutralization-sensitive and neutralization-resistant envelope glycoproteins, we demonstrate that the HIV-1 envelope glycoprotein trimer is inactivated by the binding of a single antibody molecule. Virus neutralization requires essentially all of the functional trimers to be occupied by at least one antibody. This model applies to antibodies differing in neutralizing potency and to virus isolates with various neutralization sensitivities. Understanding these requirements for HIV-1 neutralization by antibodies will assist in establishing goals for an effective AIDS vaccine.     

Evolution of envelope-specific antibody responses in monkeys experimentally infected or immunized with simian immunodeficiency virus and its association with the development of protective immunity.

Previous studies of attenuated simian immunodeficiency virus (SIV) vaccines in rhesus macaques have demonstrated the development of broad protection against experimental challenge, indicating the potential for the production of highly effective immune responses to SIV antigens. However, the development of this protective immune status was found to be critically dependent on the length of time postvaccination with the attenuated virus strain, suggesting a necessary maturation of immune responses. In this study, the evolution of SIV envelope-specific antibodies in monkeys experimentally infected with various attenuated strains of SIV was characterized by using a comprehensive panel of serological assays to assess the progression of antibodies in longitudinal serum samples that indicate the development of protective immunity. In parallel studies, we also used the same panel of antibody assays to characterize the properties of SIV envelope-specific antibodies elicited by inactivated whole-virus and envelope subunit vaccines previously reported to be ineffective in producing protective immunity. The results of these studies demonstrate that the evolution of protective immunity in monkeys inoculated with attenuated strains of SIV is associated with a complex and lengthy maturation of antibody responses over the first 6 to 8 months postinoculation, as reflected in progressive changes in antibody conformational dependence and avidity properties. The establishment of long-term protective immunity at this time in general parallels the absence of further detectable changes in antibody responses and a maintenance of relatively constant antibody titer, avidity, conformational dependence, and the presence of neutralizing antibody for at least 2 years postinoculation. In contrast to the mature antibody responses elicited by the attenuated SIV vaccines, the whole-virus and envelope subunit vaccines in general elicited only immature antibody responses characterized by poor reactivity with native envelope proteins, low avidity, low conformational dependence, and the absence of neutralization activity against the challenge strain. Thus, these studies establish for the first time an association between the effectiveness of experimental vaccines and the capacity of the vaccine to produce a mature antibody response to SIV envelope proteins and further indicate that a combination of several antibody parameters (including titer, avidity, conformational dependence, and virus neutralization) are superior to any single antibody parameter as prognostic indicators to evaluate candidate AIDS vaccines.     

Targeted Isolation of Antibodies Directed against Major Sites of SIV Env Vulnerability

The simian immunodeficiency virus (SIV) challenge model of lentiviral infection is often used as a model to human immunodeficiency virus type 1 (HIV-1) for studying vaccine mediated and immune correlates of protection. However, knowledge of the structure of the SIV envelope (Env) glycoprotein is limited, as is knowledge of binding specificity, function and potential efficacy of SIV antibody responses. In this study we describe the use of a competitive probe binding sort strategy as well as scaffolded probes for targeted isolation of SIV Env-specific monoclonal antibodies (mAbs). We isolated nearly 70 SIV-specific mAbs directed against major sites of SIV Env vulnerability analogous to broadly neutralizing antibody (bnAb) targets of HIV-1, namely, the CD4 binding site (CD4bs), CD4-induced (CD4i)-site, peptide epitopes in variable loops 1, 2 and 3 (V1, V2, V3) and potentially glycan targets of SIV Env. The range of SIV mAbs isolated includes those exhibiting varying degrees of neutralization breadth and potency as well as others that demonstrated binding but not neutralization. Several SIV mAbs displayed broad and potent neutralization of a diverse panel of 20 SIV viral isolates with some also neutralizing HIV-2_7312A. This extensive panel of SIV mAbs will facilitate more effective use of the SIV non-human primate (NHP) model for understanding the variables in development of a HIV vaccine or immunotherapy.     

Infectivity and neutralization of simian immunodeficiency virus with FLAG epitope insertion in gp120 variable loops

A FLAG epitope tag was substituted within variable loop 1 (V1), 2 (V2), or 4 (V4) of the gp120 envelope glycoprotein of simian immunodeficiency virus strain 239 (SIV239) to evaluate the extent to which each variable loop may serve as a target for antibody-mediated neutralization. Two sites within each variable loop of SIV239 were chosen for individual epitope tag insertions. FLAG epitope substitutions were also made in the V1, V2, and V4 loops of a neutralization-sensitive derivative of SIV239, SIV316. Of the 10 FLAG-tagged recombinant viruses analyzed, three (SIV239FV1b, SIV239FV2b, and SIV239FV4a) replicated with kinetics similar to those of the parental strain, SIV239, in both CEMx174 cells and the immortalized rhesus monkey T-cell line 221. The SIV316FV1b and SIV316FV4a FLAG variants replicated with a substantial lag, and the five remaining recombinants did not replicate detectably. Both gp160 and gp120 from replication-competent FLAG variants could be immunoprecipitated from transfected 293T cells by the anti-gp120 rhesus monoclonal antibody (RhMAb) 3.11H, the anti-FLAG MAb M2, and CD4-immunoglobulin, whereas only unprocessed gp160 was detected in 293T cells transfected with replication-defective variants. Furthermore, gp120 was detectably incorporated only into virions that were infectious. SIV239FV1b was sensitive to neutralization by MAb M2, with a 50% inhibitory concentration of 1 mug/ml. Neither SIV239FV2b nor SIV239FV4a was sensitive to M2 neutralization. The ability of the M2 antibody to neutralize SIV239FV1b infectivity was associated with an increased ability of the M2 antibody to detect native, oligomeric SIV239FV1b envelope protein on the surfaces of cells relative to that for the other SIV FLAG variants. Furthermore, SIV239FV1b was globally more sensitive to antibody-mediated neutralization than was parental SIV239 when these strains were screened with a panel of anti-SIV MAbs of various specificities. These results indicate that the V1 loop can serve as an effective target for neutralization on SIV239FV1b. However, antibody-mediated neutralization of this variant, similar to that of other SIV239 variants that have been studied previously, was associated with a global increase in neutralization sensitivity. These results suggest that the variable loops on the neutralization-resistant SIV239 strain are difficult for antibodies to access effectively and that mutations that allow neutralization have global effects on the trimeric envelope glycoprotein structure and accessibility.     

Studies of the conformation-dependent neutralizing epitopes of simian immunodeficiency virus envelope protein.

It has been shown previously that the major neutralizing epitopes in simian immunodeficiency virus (SIV) are discontinuous and conformation dependent and that the V3 loop, in contrast to that of human immunodeficiency virus (HIV) type 1, does not by itself elicit neutralizing antibodies (K. Javaherian et al., Proc. Natl. Acad. Sci. USA 89:1418-1422, 1992). We now present data showing that on the basis of fractionation of infected macaque sera, protease digestion of the envelope, and binding properties of two neutralizing monoclonal antibodies to SIV and SIV-HIV chimeric envelope proteins, changes in V3 can disrupt the conformation-dependent neutralization region. The chimeric protein did not produce significant neutralizing antibodies against either SIV or HIV. We also report that neutralizing antibodies elicited by recombinant SIV envelope proteins of mac251 and B670 isolates cross-neutralize. Finally, we show that deglycosylation of the SIV envelope results in a molecule which binds neither soluble CD4 nor the neutralizing monoclonal antibodies being investigated here and does not elicit sera with a significant neutralizing titer.     

Human immunodeficiency virus type 1 neutralization by a single molecule of V3-targeted antibody

Human immunodeficiency virus type-1 (HIV-1) infection generally provokes antibody responses to the viral envelope glycoprotein. Two major regions of gp120, the third variable (V3) domain and the CD4-binding site, have been identified as neutralization targets. The precise mechanism of HIV-1 neutralization by antibodies against the V3 domain is still unknown. It is shown that by kinetic neutralization studies, one molecule of V3-targeted monoclonal antibody (0.5beta) is enough to neutralize one virion. This antibody, which neutralized more than 99% of the virus, inhibited the binding of the virus to cells by 42%. HIV-1 pseudotyped with G glycoprotein from vesicular stomatitis virus was also neutralized by 0.5beta, suggesting that the antibody did not inhibit the viral attachment but caused some alteration in the envelope. These results indicate that the antibody plays an additional role on steric change of the envelope involved in inhibition of viral entry.     

Vaccine-Induced Linear Epitope-Specific Antibodies to Simian Immunodeficiency Virus SIVmac239 Envelope Are Distinct from Those Induced to the Human Immunodeficiency Virus Type 1 Envelope in Nonhuman Primates

To evaluate antibody specificities induced by simian immunodeficiency virus (SIV) versus human immunodeficiency virus type 1 (HIV-1) envelope antigens in nonhuman primate (NHP), we profiled binding antibody responses to linear epitopes in NHP studies with HIV-1 or SIV immunogens. We found that, overall, HIV-1 Env IgG responses were dominated by V3, with the notable exception of the responses to the vaccine strain A244 Env that were dominated by V2, whereas the anti-SIVmac239 Env responses were dominated by V2 regardless of the vaccine regimen.     

Virion envelope content, infectivity, and neutralization sensitivity of simian immunodeficiency virus

A truncating E767stop mutation was introduced into the envelope glycoprotein of simian immunodeficiency virus (SIV) strain SIV239-M5 (moderately sensitive to antibody-mediated neutralization and lacking five sites for N-linked carbohydrate attachment) and strain SIV316 (very sensitive to neutralization, with eight amino acid changes from the neutralization-resistant parental molecular clone, SIV239). The truncating mutation increased Env content in virions, increased infectivity, and decreased sensitivity to antibody-mediated neutralization in both strains. However, the magnitude of the effect on infectivity and neutralization sensitivity differed considerably between the two strains. In the context of strain SIV239-M5, truncation increased Env content in virions approximately 10-fold and infectivity in a reporter cell assay 24-fold. The truncated SIV239-M5 was only slightly more resistant to neutralization by polyclonal monkey sera and by monoclonal antibodies than SIV239-M5 with a full-length envelope glycoprotein. In the context of strain SIV316, truncation increased infectivity a dramatic 480-fold, while envelope content in virions was increased only about 14-fold. This dramatic increase in infectivity cannot be simply explained by the increase in envelope content and is likely due to an increase in inherent infectivity, i.e., infectivity per spike, that results from truncation. The truncated SIV316 was extremely resistant to antibody-mediated neutralization. In fact, it was not neutralized by any of the antibodies tested. When increasing amounts of SIV316 envelope glycoprotein (full length) were provided in trans to SIV316, infectivity was increased and sensitivity to neutralization was decreased, but to nowhere near the degree that was obtained when truncated SIV316 envelope glycoprotein was used. Truncated forms of SIV239 and SIV239-M5 required higher levels of soluble CD4 for inhibition of infection than their nontruncated forms; truncated SIV316 did not. Our results suggest that envelope content in SIV virions, infectivity, and resistance to antibody-mediated neutralization can be increased not only by truncation of the cytoplasmic domain but also by provision of excess envelope in trans. The striking increase in infectivity that results from truncation in the context of SIV316 appears to be due principally to an increase in inherent infectivity per spike.     

CD4 independence of simian immunodeficiency virus Envs is associated with macrophage tropism, neutralization sensitivity, and attenuated pathogenicity

To investigate the basis for envelope (Env) determinants influencing simian immunodeficiency virus (SIV) tropism, we studied a number of Envs that are closely related to that of SIVmac239, a pathogenic, T-tropic virus that is neutralization resistant. The Envs from macrophage-tropic (M-tropic) virus strains SIVmac316, 1A11, 17E-Fr, and 1100 facilitated infection of CCR5-positive, CD4-negative cells. In contrast, the SIVmac239 Env was strictly dependent upon the presence of CD4 for membrane fusion. We also found that the Envs from M-tropic virus strains, which are less pathogenic in vivo, were very sensitive to antibody-mediated neutralization. Antibodies to the V3-loop, as well as antibodies that block SIV gp120 binding to CCR5, efficiently neutralized CD4-independent, M-tropic Envs but not the 239 Env. However, triggering the 239 Env with soluble CD4, presumably resulting in exposure of the CCR5 binding site, made it as neutralization sensitive as the M-tropic Envs. In addition, mutations of N-linked glycosylation sites in the V1/V2 region, previously shown to enhance antigenicity and immunogenicity, made the 239 Env partially CD4 independent. These findings indicate that Env-based determinants of M tropism of these strains are generally associated with decreased dependence on CD4 for entry into cells. Furthermore, CD4 independence and M tropism are also associated with neutralization sensitivity and reduced pathogenicity, suggesting that the humoral immune response may exert strong selective pressure against CD4-independent M-tropic SIVmac strains. Finally, genetic modification of viral Envs to enhance CD4 independence may also result in improved humoral immune responses.     

Whole inactivated SIV virion vaccines with functional envelope glycoproteins: safety,immunogenicity, and activity against intrarectal challenge

A novel type of whole inactivated simian immunodeficiency virus (SIV) virion vaccine immunogen with functional envelope glycoproteins was evaluated, without adjuvant, in rhesus macaques. Immunogens included purified inactivated virions of SIVmac239, a designed mutant of SIVmac239 with gp120 carbohydrate attachment sites deleted (SIVmac239 g4,5), and SIVmneE11S. The vaccines were noninfectious, safe, and immunogenic, inducing antibody responses and cellular responses, including responses by CD8+ lymphocytes. Interpretation of protective efficacy following intrarectal challenge was complicated by incomplete take of the challenge in some SIV na?ve controls.     

Neutralization of the human immunodeficiency virus type 1 primary isolate JR-FL by human monoclonal antibodies correlates with antibody binding to the oligomeric form of the envelope glycoprotein complex.

To test whether antibodies that are neutralizing or nonneutralizing for human immunodeficiency virus type 1 (HIV-1) primary isolates can be distinguished by their affinities for the oligomeric envelope glycoproteins, we selected HIV-1(JR-FL) as a model primary virus and a panel of 13 human monoclonal antibodies (MAbs) and evaluated three parameters: (i) half-maximal binding to recombinant monomeric envelope, gp120(JR-FL); (ii) half-maximal binding to oligomeric envelope of HIV-1(JR-FL) expressed on the surface of transfected 293 cells; and (iii) neutralization of HIV-1(JR-FL) in a peripheral blood mononuclear cell-based neutralization assay. Two conclusions can be drawn from these experiments. First, we confirm that antibody interactions with monomeric gp120 do not predict primary virus neutralization. Second, we show that neutralization correlates qualitatively with the relative affinity of an antibody for the oligomeric envelope glycoproteins, at least for HIV-1(JR-FL).     

Removal of N-linked glycosylation sites in the V1 region of simian immunodeficiency virus gp120 results in redirection of B-cell responses to V3

One mechanism of immune evasion utilized by human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) envelope glycoproteins is the presence of a dense carbohydrate shield. Accumulating evidence from in vitro and in vivo experiments suggests that alterations in N-linked glycosylation of SIV gp120 can enhance host humoral immune responses that may be involved in immune control. The present study was designed to determine the ability of glycosylation mutant viruses to redirect antibody responses to shielded envelope epitopes. The influence of glycosylation on the maturation and specificity of antibody responses elicited by glycosylation mutant viruses containing mutations of specific N-linked sites in and near the V1 and V2 regions of SIVmac239 gp120 was determined. Results from these studies demonstrated a remarkably similar maturation of antibody responses to native, fully glycosylated envelope proteins. However, analyses of antibodies to defined envelope domains revealed that mutation of glycosylation sites in V1 resulted in increased antibody recognition to epitopes in V1. In addition, we demonstrated for the first time that mutation of glycosylation sites in V1 resulted in a redirection of antibody responses to the V3 loop. Taken together, these results demonstrate that N-linked glycosylation is a determinant of SIV envelope B-cell immunogenicity in addition to in vitro antigenicity. In addition, our results demonstrate that the absence of N-linked carbohydrates at specific sites can influence the exposure of epitopes quite distant in the linear sequence.     

Evidence against extracellular exposure of a highly immunogenic region in the C-terminal domain of the simian immunodeficiency virus gp41 transmembrane protein

The generally accepted model for human immunodeficiency virus type 1 (HIV-1) envelope glycoprotein topology includes a single membrane-spanning domain. An alternate model has been proposed which features multiple membrane-spanning domains. Consistent with the alternate model, a high percentage of HIV-1-infected individuals produce unusually robust antibody responses to a region of envelope, the so-called "Kennedy epitope," that in the conventional model should be in the cytoplasm. Here we show analogous, robust antibody responses in simian immunodeficiency virus SIVmac239-infected rhesus macaques to a region of SIVmac239 envelope located in the C-terminal domain, which in the conventional model should be inside the cell. Sera from SIV-infected rhesus macaques consistently reacted with overlapping oligopeptides corresponding to a region located within the cytoplasmic domain of gp41 by the generally accepted model, at intensities comparable to those observed for immunodominant areas of the surface component gp120. Rabbit serum raised against this highly immunogenic region (HIR) reacted with SIV envelope in cell surface-staining experiments, as did monoclonal anti-HIR antibodies isolated from an SIVmac239-infected rhesus macaque. However, control experiments demonstrated that this surface staining could be explained in whole or in part by the release of envelope protein from expressing cells into the supernatant and the subsequent attachment to the surfaces of cells in the culture. Serum and monoclonal antibodies directed against the HIR failed to neutralize even the highly neutralization-sensitive strain SIVmac316. Furthermore, a potential N-linked glycosylation site located close to the HIR and postulated to be outside the cell in the alternate model was not glycosylated. An artificially introduced glycosylation site within the HIR was also not utilized for glycosylation. Together, these data support the conventional model of SIV envelope as a type Ia transmembrane protein with a single membrane-spanning domain and without any extracellular loops.     

Assorted mutations in the envelope gene of simian immunodeficiency virus lead to loss of neutralization resistance against antibodies representing a broad spectrum of specificities

Simian immunodeficiency virus (SIV) of macaques isolate SIVmac239 is highly resistant to neutralization by polyclonal antisera or monoclonal antibodies, a property that it shares with most primary isolates of human immunodeficiency virus type 1 (HIV-1). This resistance is important for the ability of the virus to persist at high levels in vivo. To explore the physical features of the viral envelope complex that contribute to the neutralization-resistant phenotype, we examined a panel of SIVmac239 derivatives for sensitivity to neutralization by a large collection of monoclonal antibodies (MAbs). These MAbs recognize both linear and conformational epitopes throughout the viral envelope proteins. The variant viruses included three derivatives of SIVmac239 with substitutions in specific N-linked glycosylation sites of gp120 and a fourth variant that lacked the 100 amino acids that encompass the V1 and V2 loops. Also included in this study was SIVmac316, a variant of SIVmac239 with distributed mutations in env that confer significantly increased replicative capacity in tissue macrophages. These viruses were chosen to represent a broad range of neutralization sensitivities based on susceptibility to pooled, SIV-positive plasma. All three of these very different kinds of mutations (amino acid substitutions, elimination of N-glycan attachment sites, and a 100-amino-acid deletion spanning variable loops V1 and V2) dramatically increased sensitivity to neutralization by MAbs from multiple competition groups. Thus, the mutations did not simply expose localized epitopes but rather conferred global increases in neutralization sensitivity. The removal of specific N-glycan attachment sites from V1 and V2 led to increased sensitivity to neutralization by antibodies recognizing epitopes from both within and outside of the V1-V2 sequence. Surprisingly, while most of the mutations that gave rise to increased sensitivity were located in the N-terminal half of gp120 (surface subunit [SU]), the greatest increases in sensitivity were to MAbs recognizing the C-terminal half of gp120 or the ectodomain of gp41 (transmembrane subunit [TM]). This reagent set and information should now be useful for defining the physical, structural, thermodynamic, and kinetic factors that influence relative sensitivity to antibody-mediated neutralization.     

Methods for analysis of biologically functional antibodies to the HIV-1 gp120 principal neutralizing domain.

The humoral response to HIV-1 infection has been demonstrated by a variety of immunoassays utilizing viral proteins. While several assays detect HIV-1 infection with high sensitivity and great specificity, little progress has been made to develop immunoassays correlative with disease progression and viral transmission. Antibodies toward the V3 domain of HIV-1 envelope can prevent virus infection and block virus-mediated cell fusion in vitro. Such properties may be critical to the course of the disease. Furthermore, understanding the role of neutralizing antibodies against HIV-1 during infection in humans and generating biologically relevant neutralizing antibodies are paramount to developing an efficacious AIDS vaccine. In this study we explored peptide binding and neutralization assays and their relation to predicting disease progression and viral transmission. Biologically relevant polyclonal and monoclonal neutralizing antibodies that were derived from natural HIV-1 infection of humans, experimental infections of chimpanzees, and viral envelope protein peptide immunizations were characterized. Comparison of V3-specific monoclonal antibodies by antigen-limited ELISA and a quantitative HIV-1 neutralization assay demonstrated a less than optimal predictive relationship between binding and neutralization potency. On the other hand, polyclonal sera from goats immunized with V3-specific peptides derived from three different HIV-1 strains, as well as sera from other HIV-1-infected individuals demonstrated correlation between binding affinity and neutralization.     

Role of CD8(+) lymphocytes in control of simian immunodeficiency virus infection and resistance to rechallenge after transient early antiretroviral treatment

Transient antiretroviral treatment with tenofovir, (R)-9-(2-phosphonylmethoxypropyl)adenine, begun shortly after inoculation of rhesus macaques with the highly pathogenic simian immunodeficiency virus (SIV) isolate SIVsmE660, facilitated the development of SIV-specific lymphoproliferative responses and sustained effective control of the infection following drug discontinuation. Animals that controlled plasma viremia following transient postinoculation treatment showed substantial resistance to subsequent intravenous rechallenge with homologous (SIVsmE660) and highly heterologous (SIVmac239) SIV isolates, up to more than 1 year later, despite the absence of measurable neutralizing antibody. In some instances, resistance to rechallenge was observed despite the absence of detectable SIV-specific binding antibody and in the face of SIV lymphoproliferative responses that were low or undetectable at the time of challenge. In vivo monoclonal antibody depletion experiments demonstrated a critical role for CD8(+) lymphocytes in the control of viral replication; plasma viremia rose by as much as five log units after depletion of CD8(+) cells and returned to predepletion levels (as low as <100 copy Eq/ml) as circulating CD8(+) cells were restored. The extent of host control of replication of highly pathogenic SIV strains and the level of resistance to heterologous rechallenge achieved following transient postinoculation treatment compared favorably to the results seen after SIVsmE660 and SIVmac239 challenge with many vaccine strategies. This impressive control of viral replication was observed despite comparatively modest measured immune responses, less than those often achieved with vaccination regimens. The results help establish the underlying feasibility of efforts to develop vaccines for the prevention of AIDS, although the exact nature of the protective host responses involved remains to be elucidated.     

Antibody binding is a dominant determinant of the efficiency of human immunodeficiency virus type 1 neutralization

Primary and laboratory-adapted variants of human immunodeficiency virus type 1 (HIV-1) exhibit a wide range of sensitivities to neutralization by antibodies directed against the viral envelope glycoproteins. An antibody directed against an artificial FLAG epitope inserted into the envelope glycoproteins of three HIV-1 isolates with vastly different neutralization sensitivities inhibited all three viruses equivalently. Thus, naturally occurring HIV-1 isolates that are neutralization resistant are not necessarily more impervious to the inhibitory consequences of bound antibody. Moreover, the binding affinity of the anti-FLAG antibody correlated with neutralizing potency, underscoring the dominant impact on neutralization of antibody binding to the envelope glycoproteins.     

Antibody-dependent cell-mediated viral inhibition emerges after simian immunodeficiency virus SIVmac251 infection of rhesus monkeys coincident with gp140-binding antibodies and is effective against neutralization-resistant viruses

Antibody-dependent cell-mediated viral inhibition (ADCVI) is an attractive target for vaccination because it takes advantage of both the anamnestic properties of an adaptive immune response and the rapid early response characteristics of an innate immune response. Effective utilization of ADCVI in vaccine strategies will depend on an understanding of the natural history of ADCVI during acute and chronic human immunodeficiency virus type 1 (HIV-1) infection. We used the simian immunodeficiency virus (SIV)-infected rhesus monkey as a model to study the kinetics of ADCVI in early infection, the durability of ADCVI through the course of infection, and the effectiveness of ADCVI against viruses with envelope mutations that are known to confer escape from antibody neutralization. We demonstrate the development of ADCVI, capable of inhibiting viral replication 100-fold, within 3 weeks of infection, preceding the development of a comparable-titer neutralizing antibody response by weeks to months. The emergence of ADCVI was temporally associated with the emergence of gp140-binding antibodies, and in most animals, ADCVI persisted through the course of infection. Highly evolved viral envelopes from viruses isolated at late time points following infection that were resistant to plasma neutralization remained susceptible to ADCVI, suggesting that the epitope determinants of neutralization escape are not shared by antibodies that mediate ADCVI. These findings suggest that despite the ability of SIV to mutate and adapt to multiple immunologic pressures during the course of infection, SIV envelope may not escape the binding of autologous antibodies that mediate ADCVI.     

Prevention of disease induced by a partially heterologous AIDS virus in rhesus monkeys by using an adjuvanted multicomponent protein vaccine

Recombinant protein subunit AIDS vaccines have been based predominantly on the virus envelope protein. Such vaccines elicit neutralizing antibody responses that can provide type-specific sterilizing immunity, but in most cases do not confer protection against divergent viruses. In this report we demonstrate that a multiantigen subunit protein vaccine was able to prevent the development of disease induced in rhesus monkeys by a partially heterologous AIDS virus. The vaccine was composed of recombinant human immunodeficiency virus type 1 (HIV-1) gp120, NefTat fusion protein, and simian immunodeficiency virus (SIV) Nef formulated in the clinically tested adjuvant AS02A. Upon challenge of genetically unselected rhesus monkeys with the highly pathogenic and partially heterologous SIV/HIV strain SHIV(89.6p) the vaccine was able to reduce virus load and protect the animals from a decline in CD4-positive cells. Furthermore, vaccination prevented the development of AIDS for more than 2.5 years. The combination of the regulatory proteins Nef and Tat together with the structural protein gp120 was required for vaccine efficacy.