scholarly journals Autologous Neutralizing Antibodies to the Transmitted/Founder Viruses Emerge Late after Simian Immunodeficiency Virus SIVmac251 Infection of Rhesus Monkeys

2010 ◽  
Vol 84 (12) ◽  
pp. 6018-6032 ◽  
Author(s):  
Wendy W. Yeh ◽  
Ishita Rahman ◽  
Peter Hraber ◽  
Rory T. Coffey ◽  
Daiva Nevidomskyte ◽  
...  
Keyword(s):  
Antibody Response ◽  
Simian Immunodeficiency Virus ◽  
Neutralizing Antibodies ◽  
Variable Region ◽  
Neutralizing Antibody ◽  
Antigenic Determinants ◽  
Sequence Evolution ◽  
Humoral Immune ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT While the simian immunodeficiency virus (SIV)-infected rhesus monkey is an important animal model for human immunodeficiency virus type 1 (HIV-1) infection of humans, much remains to be learned about the evolution of the humoral immune response in this model. In HIV-1 infection, autologous neutralizing antibodies emerge 2 to 3 months after infection. However, the ontogeny of the SIV-specific neutralizing antibody response in mucosally infected animals has not been defined. We characterized the kinetics of the autologous neutralizing antibody response to the transmitted/founder SIVmac251 using a pseudovirion-based TZM-bl cell assay and monitored env sequence evolution using single-genome amplification in four rhesus animals that were infected via intrarectal inoculations. We show that the SIVmac251 founder viruses induced neutralizing antibodies at 5 to 8 months after infection. Despite their slow emergence and low titers, these neutralizing antibodies selected for escape mutants that harbored substitutions and deletions in variable region 1 (V1), V2, and V4 of Env. The neutralizing antibody response was initially focused on V4 at 5 to 8 months after infection and then targeted V1/V2 and V4 by 16 months. These findings reveal a striking delay in the development of neutralizing antibodies in SIVmac-infected animals, thus raising questions concerning the suitability of SIVmac251 as a challenge strain to screen AIDS vaccines that elicit neutralizing antibodies as a means to prevent virus acquisition. They also illustrate the capacity of the SIVmac quasispecies to modify antigenic determinants in response to very modest titers of neutralizing antibodies.

scholarly journals Characterization and Implementation of a Diverse Simian Immunodeficiency Virus SIVsm Envelope Panel in the Assessment of Neutralizing Antibody Breadth Elicited in Rhesus Macaques by Multimodal Vaccines Expressing the SIVmac239 Envelope

2015 ◽  
Vol 89 (16) ◽  
pp. 8130-8151 ◽  
Author(s):  
Katie M. Kilgore ◽  
Megan K. Murphy ◽  
Samantha L. Burton ◽  
Katherine S. Wetzel ◽  
S. Abigail Smith ◽  
...  
Keyword(s):  
Simian Immunodeficiency Virus ◽  
Nonhuman Primate ◽  
Neutralizing Antibodies ◽  
Rhesus Macaques ◽  
Neutralizing Antibody ◽  
Cd40 Ligand ◽  
Antibody Activity ◽  
Immunodeficiency Virus ◽  
Antibody Profile ◽  
Hiv 1

ABSTRACTAntibodies that can neutralize diverse viral strains are likely to be an important component of a protective human immunodeficiency virus type 1 (HIV-1) vaccine. To this end, preclinical simian immunodeficiency virus (SIV)-based nonhuman primate immunization regimens have been designed to evaluate and enhance antibody-mediated protection. However, these trials often rely on a limited selection of SIV strains with extreme neutralization phenotypes to assess vaccine-elicited antibody activity. To mirror the viral panels used to assess HIV-1 antibody breadth, we created and characterized a novel panel of 14 genetically and phenotypically diverse SIVsm envelope (Env) glycoproteins. To assess the utility of this panel, we characterized the neutralizing activity elicited by four SIVmac239 envelope-expressing DNA/modified vaccinia virus Ankara vector- and protein-based vaccination regimens that included the immunomodulatory adjuvants granulocyte-macrophage colony-stimulating factor, Toll-like receptor (TLR) ligands, and CD40 ligand. The SIVsm Env panel exhibited a spectrum of neutralization sensitivity to SIV-infected plasma pools and monoclonal antibodies, allowing categorization into three tiers. Pooled sera from 91 rhesus macaques immunized in the four trials consistently neutralized only the highly sensitive tier 1a SIVsm Envs, regardless of the immunization regimen. The inability of vaccine-mediated antibodies to neutralize the moderately resistant tier 1b and tier 2 SIVsm Envs defined here suggests that those antibodies were directed toward epitopes that are not accessible on most SIVsm Envs. To achieve a broader and more effective neutralization profile in preclinical vaccine studies that is relevant to known features of HIV-1 neutralization, more emphasis should be placed on optimizing the Env immunogen, as the neutralization profile achieved by the addition of adjuvants does not appear to supersede the neutralizing antibody profile determined by the immunogen.IMPORTANCEMany in the HIV/AIDS vaccine field believe that the ability to elicit broadly neutralizing antibodies capable of blocking genetically diverse HIV-1 variants is a critical component of a protective vaccine. Various SIV-based nonhuman primate vaccine studies have investigated ways to improve antibody-mediated protection against a heterologous SIV challenge, including administering adjuvants that might stimulate a greater neutralization breadth. Using a novel SIV neutralization panel and samples from four rhesus macaque vaccine trials designed for cross comparison, we show that different regimens expressing the same SIV envelope immunogen consistently elicit antibodies that neutralize only the very sensitive tier 1a SIV variants. The results argue that the neutralizing antibody profile elicited by a vaccine is primarily determined by the envelope immunogen and is not substantially broadened by including adjuvants, resulting in the conclusion that the envelope immunogen itself should be the primary consideration in efforts to elicit antibodies with greater neutralization breadth.

scholarly journals Comparative Immunogenicity of Subtype A Human Immunodeficiency Virus Type 1 Envelope Exhibiting Differential Exposure of Conserved Neutralization Epitopes

2009 ◽  
Vol 84 (5) ◽  
pp. 2573-2584 ◽  
Author(s):  
Catherine A. Blish ◽  
D. Noah Sather ◽  
George Sellhorn ◽  
Leonidas Stamatatos ◽  
Yide Sun ◽  
...  
Keyword(s):  
Antibody Response ◽  
Neutralizing Antibodies ◽  
Vaccine Development ◽  
Neutralizing Antibody ◽  
Immunodeficiency Virus ◽  
Cd4 Binding Site ◽  
Variable Loops ◽  
Linear Epitopes ◽  
Subtype A ◽  
Hiv 1

ABSTRACT Development of broadly cross-reactive neutralizing antibodies (NAbs) remains a major goal of HIV-1 vaccine development, but most candidate envelope immunogens have had limited ability to cross-neutralize heterologous strains. To evaluate the immunogenicity of subtype A variants of HIV-1, rabbits were immunized with pairs of closely related subtype A envelopes from the same individual. In each immunogen pair, one variant was readily neutralized by a variety of monoclonal antibodies and plasma antibodies, while the other was neutralization resistant, suggesting differences in the exposures of key epitopes. The breadth of the antibody response was evaluated against subtype A, B, C, and D variants of HIV-1. The specificity of the immunogen-derived neutralizing antibody response was also compared to that of the infected individuals from whom these variants were cloned. None of the immunogens produced broad neutralizing antibodies in immunized animals, and most of the neutralizing antibodies were directed to the variable loops, particularly the V3 loop. No detectable antibodies to either of the potentially exposed conserved epitopes, the membrane proximal external region, or the CD4 binding site were found with immunized rabbits. In contrast, relatively little of the neutralizing activity within the plasma samples of the infected individuals was directed to linear epitopes within the variable loops. These data indicate that immunogens designed to expose conserved regions did not enhance generation of broadly neutralizing antibodies in comparison with the immunogens that failed to expose those regions using this immunization approach.

scholarly journals Recombinant Modified Vaccinia Virus Ankara Expressing the Surface gp120 of Simian Immunodeficiency Virus (SIV) Primes for a Rapid Neutralizing Antibody Response to SIV Infection in Macaques

2000 ◽  
Vol 74 (6) ◽  
pp. 2960-2965 ◽  
Author(s):  
Ilnour Ourmanov ◽  
Miroslawa Bilska ◽  
Vanessa M. Hirsch ◽  
David C. Montefiori
Keyword(s):  
Antibody Response ◽  
Vaccinia Virus ◽  
Simian Immunodeficiency Virus ◽  
Neutralizing Antibodies ◽  
Rhesus Macaques ◽  
Neutralizing Antibody ◽  
Anamnestic Response ◽  
Modified Vaccinia Virus Ankara ◽  
Immunodeficiency Virus ◽  
Before And After

ABSTRACT Neutralizing antibodies were assessed before and after intravenous challenge with pathogenic SIVsmE660 in rhesus macaques that had been immunized with recombinant modified vaccinia virus Ankara expressing one or more simian immunodeficiency virus gene products (MVA-SIV). Animals received either MVA-gag-pol, MVA-env, MVA-gag-pol-env, or nonrecombinant MVA. Although no animals were completely protected from infection with SIV, animals immunized with recombinant MVA-SIV vaccines had lower virus loads and prolonged survival relative to control animals that received nonrecombinant MVA (I. Ourmanov et al., J. Virol. 74:2740–2751, 2000). Titers of neutralizing antibodies measured with the vaccine strain SIVsmH-4 were low in the MVA-env and MVA-gag-pol-env groups of animals and were undetectable in the MVA-gag-pol and nonrecombinant MVA groups of animals on the day of challenge (4 weeks after final immunization). Titers of SIVsmH-4-neutralizing antibodies remained unchanged 1 week later but increased approximately 100-fold 2 weeks postchallenge in the MVA-env and MVA-gag-pol-env groups while the titers remained low or undetectable in the MVA-gag-pol and nonrecombinant MVA groups. This anamnestic neutralizing antibody response was also detected with T-cell-line-adapted stocks of SIVmac251 and SIV/DeltaB670 but not with SIVmac239, as this latter virus resisted neutralization. Most animals in each group had high titers of SIVsmH-4-neutralizing antibodies 8 weeks postchallenge. Titers of neutralizing antibodies were low or undetectable until about 12 weeks of infection in all groups of animals and showed little or no evidence of an anamnestic response when measured with SIVsmE660. The results indicate that recombinant MVA is a promising vector to use to prime for an anamnestic neutralizing antibody response following infection with primate lentiviruses that cause AIDS. However, the Env component of the present vaccine needs improvement in order to target a broad spectrum of viral variants, including those that resemble primary isolates.

scholarly journals Envelope Vaccination Shapes Viral Envelope Evolution following Simian Immunodeficiency Virus Infection in Rhesus Monkeys

2009 ◽  
Vol 84 (2) ◽  
pp. 953-963 ◽  
Author(s):  
Aravind Basavapathruni ◽  
Wendy W. Yeh ◽  
Rory T. Coffey ◽  
James B. Whitney ◽  
Peter T. Hraber ◽  
...  
Keyword(s):  
Antibody Response ◽  
Simian Immunodeficiency Virus ◽  
Rhesus Monkeys ◽  
Neutralizing Antibodies ◽  
Neutralizing Antibody ◽  
Neutralization Assay ◽  
Viral Envelope ◽  
Variable Regions ◽  
Virus Challenge ◽  
Immunodeficiency Virus

ABSTRACT The evolution of envelope mutations by replicating primate immunodeficiency viruses allows these viruses to escape from the immune pressure mediated by neutralizing antibodies. Vaccine-induced anti-envelope antibody responses may accelerate and/or alter the specificity of the antibodies, thus shaping the evolution of envelope mutations in the replicating virus. To explore this possibility, we studied the neutralizing antibody response and the envelope sequences in rhesus monkeys vaccinated with either gag-pol-nef immunogens or gag-pol-nef immunogens in combination with env and then infected with simian immunodeficiency virus (SIV). Using a pseudovirion neutralization assay, we demonstrate that envelope vaccination primed for an accelerated neutralizing antibody response following virus challenge. To monitor viral envelope evolution in these two cohorts of monkeys, full-length envelopes from plasma virus isolated at weeks 37 and 62 postchallenge were sequenced by single genome amplification to identify sites of envelope mutations. We show that env vaccination was associated with a change in the pattern of envelope mutations. Prevalent mutations in sequences from gag-pol-nef vaccinees included deletions in both variable regions 1 and 4 (V1 and V4), whereas deletions in the env vaccinees occurred only in V1. These data show that env vaccination altered the focus of the antibody-mediated selection pressure on the evolution of envelope following SIV challenge.

scholarly journals Neutralizing Antibodies Elicited by Immunization of Monkeys with DNA Plasmids and Recombinant Adenoviral Vectors Expressing Human Immunodeficiency Virus Type 1 Proteins

2005 ◽  
Vol 79 (2) ◽  
pp. 771-779 ◽  
Author(s):  
John R. Mascola ◽  
Anna Sambor ◽  
Kristin Beaudry ◽  
Sampa Santra ◽  
Brent Welcher ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Immune Responses ◽  
Antibody Response ◽  
Human Immunodeficiency Virus Type ◽  
Neutralizing Antibodies ◽  
Neutralizing Antibody ◽  
Immunodeficiency Virus ◽  
Anti Hiv ◽  
Hiv 1

ABSTRACT Immunization with recombinant serotype 5 adenoviral (rAd5) vectors or a combination of DNA plasmid priming and rAd5 boosting is known to elicit potent immune responses. However, little data exist regarding these immunization strategies and the development of anti-human immunodeficiency virus type 1 (HIV-1) neutralizing antibodies. We used DNA plasmids and rAd5 vectors encoding the HIV-1 89.6P or chimeric HxB2/BaL envelope glycoprotein to immunize macaque monkeys. A single rAd5 immunization elicited anti-Env antibody responses, but there was little boosting with subsequent rAd5 immunizations. In contrast, rAd5 boosting of DNA-primed monkeys resulted in a rapid rise in antibody titers, including the development of anti-HIV-1 neutralizing antibodies. The potency and breadth of neutralization were evaluated by testing plasma against a panel of 14 clade B primary isolates. Moderate levels of plasma neutralizing activity were detected against about one-third of the viruses tested, and immunoglobulin G fractionation demonstrated that virus neutralization was antibody mediated. After a challenge with a chimeric simian-human immunodeficiency virus (SHIV89.6P), an anamnestic neutralizing antibody response was observed, although the breadth of the response was limited to the subset of viruses that were neutralized after the primary immunization. These data are the first detailed description of the anti-HIV-1 neutralizing antibody response in nonhuman primates elicited by DNA and rAd5 immunization. In addition to the well-established ability of DNA priming and rAd5 boosting to elicit potent anti-HIV-1 cellular immune responses, this immunization strategy elicits anti-HIV-1 neutralizing antibodies and therefore can be used to study novel Env immunogens designed to elicit more potent neutralizing antibodies.

scholarly journals Neutralizing Antibody Responses in Acute Human Immunodeficiency Virus Type 1 Subtype C Infection

2007 ◽  
Vol 81 (12) ◽  
pp. 6187-6196 ◽  
Author(s):  
E. S. Gray ◽  
P. L. Moore ◽  
I. A. Choge ◽  
J. M. Decker ◽  
F. Bibollet-Ruche ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Human Immunodeficiency Virus Type ◽  
Virus Type ◽  
Neutralizing Antibodies ◽  
Neutralizing Antibody ◽  
Antibody Responses ◽  
Subtype C ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT The study of the evolution and specificities of neutralizing antibodies during the course of human immunodeficiency virus type 1 (HIV-1) infection may be important in the discovery of possible targets for vaccine design. In this study, we assessed the autologous and heterologous neutralization responses of 14 HIV-1 subtype C-infected individuals, using envelope clones obtained within the first 2 months postinfection. Our data show that potent but relatively strain-specific neutralizing antibodies develop within 3 to 12 months of HIV-1 infection. The magnitude of this response was associated with shorter V1-to-V5 envelope lengths and fewer glycosylation sites, particularly in the V1-V2 region. Anti-MPER antibodies were detected in 4 of 14 individuals within a year of infection, while antibodies to CD4-induced (CD4i) epitopes developed to high titers in 12 participants, in most cases before the development of autologous neutralizing antibodies. However, neither anti-MPER nor anti-CD4i antibody specificity conferred neutralization breadth. These data provide insights into the kinetics, potency, breadth, and epitope specificity of neutralizing antibody responses in acute HIV-1 subtype C infection.

scholarly journals Removal of a Single N-Linked Glycan in Human Immunodeficiency Virus Type 1 gp120 Results in an Enhanced Ability To Induce Neutralizing Antibody Responses

2007 ◽  
Vol 82 (2) ◽  
pp. 638-651 ◽  
Author(s):  
Yun Li ◽  
Bradley Cleveland ◽  
Igor Klots ◽  
Bruce Travis ◽  
Barbra A. Richardson ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Neutralizing Antibodies ◽  
Neutralizing Antibody ◽  
Pronounced Increase ◽  
Enhanced Sensitivity ◽  
Subtype B ◽  
Immunodeficiency Virus ◽  
Hiv 1

ABSTRACT Glycans on human immunodeficiency virus (HIV) envelope protein play an important role in infection and evasion from host immune responses. To examine the role of specific glycans, we introduced single or multiple mutations into potential N-linked glycosylation sites in hypervariable regions (V1 to V3) of the env gene of HIV type 1 (HIV-1) 89.6. Three mutants tested showed enhanced sensitivity to soluble CD4. Mutant N7 (N197Q) in the carboxy-terminal stem of the V2 loop showed the most pronounced increase in sensitivity to broadly neutralizing antibodies (NtAbs), including those targeting the CD4-binding site (IgG1b12) and the V3 loop (447-52D). This mutant is also sensitive to CD4-induced NtAb 17b in the absence of CD4. Unlike the wild-type (WT) Env, mutant N7 mediates CD4-independent infection in U87-CXCR4 cells. To study the immunogenicity of mutant Env, we immunized pig-tailed macaques with recombinant vaccinia viruses, one expressing SIVmac239 Gag-Pol and the other expressing HIV-1 89.6 Env gp160 in WT or mutant forms. Animals were boosted 14 to 16 months later with simian immunodeficiency virus gag DNA and the cognate gp140 protein before intrarectal challenge with SHIV89.6P-MN. Day-of-challenge sera from animals immunized with mutant N7 Env had significantly higher and broader neutralizing activities than sera from WT Env-immunized animals. Neutralizing activity was observed against SHIV89.6, SHIV89.6P-MN, HIV-1 SF162, and a panel of subtype B primary isolates. Compared to control animals, immunized animals showed significant reduction of plasma viral load and increased survival after challenge, which correlated with prechallenge NtAb titers. These results indicate the potential advantages for glycan modification in vaccine design, although the role of specific glycans requires further examination.

scholarly journals Nanoparticle Vaccines for Inducing HIV-1 Neutralizing Antibodies

Vaccines ◽  
2019 ◽  
Vol 7 (3) ◽  
pp. 76 ◽  
Author(s):  
Mitch Brinkkemper ◽  
Kwinten Sliepen
Keyword(s):  
Envelope Glycoprotein ◽  
Neutralizing Antibodies ◽  
Critical Role ◽  
Neutralizing Antibody ◽  
Antibody Responses ◽  
Broadly Neutralizing Antibodies ◽  
Viral Membrane ◽  
Immunodeficiency Virus ◽  
Hiv 1

The enormous sequence diversity between human immunodeficiency virus type 1 (HIV-1) strains poses a major roadblock for generating a broadly protective vaccine. Many experimental HIV-1 vaccine efforts are therefore aimed at eliciting broadly neutralizing antibodies (bNAbs) that are capable of neutralizing the majority of circulating HIV-1 strains. The envelope glycoprotein (Env) trimer on the viral membrane is the sole target of bNAbs and the key component of vaccination approaches aimed at eliciting bNAbs. Multimeric presentation of Env on nanoparticles often plays a critical role in these strategies. Here, we will discuss the different aspects of nanoparticles in Env vaccination, including recent insights in immunological processes underlying their perceived advantages, the different nanoparticle platforms and the various immunogenicity studies that employed nanoparticles to improve (neutralizing) antibody responses against Env.

scholarly journals Truncating the gp41 Cytoplasmic Tail of Simian Immunodeficiency Virus Decreases Sensitivity to Neutralizing Antibodies without Increasing the Envelope Content of Virions

2017 ◽  
Vol 92 (3) ◽  
Author(s):  
Ellen White ◽  
Fan Wu ◽  
Elena Chertova ◽  
Julian Bess ◽  
James D. Roser ◽  
...  
Keyword(s):  
Monoclonal Antibodies ◽  
Simian Immunodeficiency Virus ◽  
Neutralizing Antibodies ◽  
Cytoplasmic Tail ◽  
Neutralizing Antibody ◽  
Global Changes ◽  
Imaging Studies ◽  
Stop Codons ◽  
Immunodeficiency Virus ◽  
Tier 3

ABSTRACTAn incomplete understanding of native human immunodeficiency virus (HIV) and simian immunodeficiency virus (SIV) envelope glycoproteins (Envs) impedes the development of structural models of Env and vaccine design. This shortcoming is due in part to the low number of Env trimers on virus particles. For SIV, this low expression level can be counteracted by truncating the cytoplasmic tail (CT) of Env. CT truncation has been shown to increase Env incorporation into the virion and is commonly used in vaccine and imaging studies, but its effects on viral antigenicity have not been fully elucidated. To study the effects of a CT truncation of Env in viruses in similar genetic contexts, we introduced stop codons into the CT of a SIVsmE660 molecular clone and two neutralizing antibody (NAb) escape variants. These viruses shared 98% sequence identity in Env but were characterized as either tier 1 (sensitive to neutralization), tier 2 (moderately resistant to neutralization), or tier 3 (resistant to neutralization). However, the introduction of premature stop codons in Env at position Q741/Q742 converted all three transfection-derived viruses to a tier 3-like phenotype, and these viruses were uniformly resistant to neutralization by sera from infected macaques and monoclonal antibodies (MAbs). These changes in neutralization sensitivity were not accompanied by an increase in either the virion Env content of infection-derived viruses or the infectivity of transfection-derived viruses in human cells, suggesting that CT mutations may result in global changes to the Env conformation. Our results demonstrate that some CT truncations can affect viral antigenicity and, as such, may not be suitable surrogate models of native HIV/SIV Env.IMPORTANCEModifications to the SIV envelope protein (Env) are commonly used in structural and vaccine studies to stabilize and increase the expression of Env, often without consideration of effects on antigenicity. One such widespread modification is the truncation of the Env C-terminal tail. Here, we studied the effects of a particular cytoplasmic tail truncation in three SIVsm strains that have highly similar Env sequences but exhibit different sensitivities to neutralizing antibodies. After truncation of the Env CT, these viruses were all very resistant to neutralization by sera from infected macaques and monoclonal antibodies. The viruses with a truncated Env CT also did not exhibit the desired and typical increase in Env expression. These results underscore the importance of carefully evaluating the use of truncated Env as a model in HIV/SIV vaccine and imaging studies and of the continued need to find better models of native Env that contain fewer modifications.

scholarly journals A Glycoconjugate Antigen Based on the Recognition Motif of a Broadly Neutralizing Human Immunodeficiency Virus Antibody, 2G12, Is Immunogenic but Elicits Antibodies Unable To Bind to the Self Glycans of gp120

2008 ◽  
Vol 82 (13) ◽  
pp. 6359-6368 ◽  
Author(s):  
Rena D. Astronomo ◽  
Hing-Ken Lee ◽  
Christopher N. Scanlan ◽  
Ralph Pantophlet ◽  
Cheng-Yuan Huang ◽  
...  
Keyword(s):  
Human Immunodeficiency Virus ◽  
Neutralizing Antibody ◽  
Humoral Immune ◽  
Aids Vaccine ◽  
Humoral Immune Responses ◽  
Copy Numbers ◽  
Immunodeficiency Virus ◽  
Broadly Neutralizing Antibody ◽  
Hiv 1

ABSTRACT The glycan shield of human immunodeficiency virus type 1 (HIV-1) gp120 contributes to viral evasion from humoral immune responses. However, the shield is recognized by the HIV-1 broadly neutralizing antibody (Ab), 2G12, at a relatively conserved cluster of oligomannose glycans. The discovery of 2G12 raises the possibility that a carbohydrate immunogen may be developed that could elicit 2G12-like neutralizing Abs and contribute to an AIDS vaccine. We have previously dissected the fine specificity of 2G12 and reported that the synthetic tetramannoside (Man4) that corresponds to the D1 arm of Man9GlcNAc2 inhibits 2G12 binding to gp120 as efficiently as Man9GlcNAc2 itself, indicating the potential use of Man4 as a building block for creating immunogens. Here, we describe the development of neoglycoconjugates displaying variable copy numbers of Man4 on bovine serum albumin (BSA) molecules by conjugation to Lys residues. The increased valency enhances the apparent affinity of 2G12 for Man4 up to a limit which is achieved at ∼10 copies per BSA molecule, beyond which no further enhancement is observed. Immunization of rabbits with BSA-(Man4)14 elicits significant serum Ab titers to Man4. However, these Abs are unable to bind gp120. Further analysis reveals that the elicited Abs bind a variety of unbranched and, to a lesser extent, branched Man9 derivatives but not natural N-linked oligomannose containing the chitobiose core. These results suggest that Abs can be readily elicited against the D1 arm; however, potential differences in the presentation of Man4 on neoglycoconjugates, compared to glycoproteins, poses challenges for eliciting anti-mannose Abs capable of cross-reacting with gp120 and HIV-1.

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