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

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 (Man 4 ) that corresponds to the D1 arm of Man 9 GlcNAc 2 inhibits 2G12 binding to gp120 as efficiently as Man 9 GlcNAc 2 itself, indicating the potential use of Man 4 as a building block for creating immunogens. Here, we describe the development of neoglycoconjugates displaying variable copy numbers of Man 4 on bovine serum albumin (BSA) molecules by conjugation to Lys residues. The increased valency enhances the apparent affinity of 2G12 for Man 4 up to a limit which is achieved at ϳ10 copies per BSA molecule, beyond which no further enhancement is observed. Immunization of rabbits with BSA-(Man 4 ) 14 elicits significant serum Ab titers to Man 4 . 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 Man 9 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 Man 4 on neoglycoconjugates, compared to glycoproteins, poses challenges for eliciting anti-mannose Abs capable of crossreacting with gp120 and HIV-1.

In-Solution Virus Capture Assay Helps Deconstruct Heterogeneous Antibody Recognition of Human Immunodeficiency Virus Type 1

“…Most likely, MPER MAbs capture Env(Ϫ) virions through weak membrane interactions. 2G12 specifically targets a cluster of high mannose glycans on gp120, but it has also been shown to bind to clusters of synthetic oligomannose (4,40,77), a yeast glycoprotein (46,47), and 293T cells treated with oligomannose-modifying drugs (75). Thus, Env-independent virus capture by 2G12 is most likely mediated through weak interactions with virion-incorporated host glycoproteins.…”

High-Mannose Glycan-Dependent Epitopes Are Frequently Targeted in Broad Neutralizing Antibody Responses during Human Immunodeficiency Virus Type 1 Infection

“…Glycan-specific antibody responses can be an attractive line of vaccine development, since some glycan-specific MAbs demonstrate great neutralizing breadth and potency (51,54). It has also been shown that glycan-binding antibodies can be induced by synthetic glycans (2,3,17,19,26,57) or HM-glycan on non-HIV-1 proteins (1, 25, 34, 35). Unfortunately, a glycan-binding antibody capable of neutralizing diverse strains of primary HIV-1 has yet to be achieved.…”

Nonneutralizing HIV-1 gp41 Envelope Cluster II Human Monoclonal Antibodies Show Polyreactivity for Binding to Phospholipids and Protein Autoantigens

“…Although a third broadly neutralizing MAb, 2G12, did not show any reactivity for the tested autoantigens (24), the epitope recognized by this gp120 MAb, 2G12, is composed primarily of a cluster of high-mannose oligosaccharides that are similar to "self" carbohydrates (4,42,50). Thus, the rarity of broadly neutralizing MAbs after either natural infection or immunization (7, 30) raises the question of whether expression of these specificities for gp41 or carbohydrates may be immunoregulated and whether crossreactivity with self antigens, such as phospholipids, is essential for anti-gp41 MAbs to neutralize HIV-1 (1, 2).…”