Human Monoclonal Antibodies Specific for Conformation-Sensitive Epitopes of V3 Neutralize Human Immunodeficiency Virus Type 1 Primary Isolates from Various Clades

Abstract

The epitopes of the V3 domain of the human immunodeficiency virus type 1 (HIV-1) gp120 glycoprotein have complex structures consisting of linear and conformational antigenic determinants. Anti-V3 antibodies (Abs) recognize both types of elements, but Abs which preferentially react to the conformational aspect of the epitopes may have more potent neutralizing activity against HIV-1, as recently suggested. To test this hypothesis, human anti-V3 monoclonal Abs (MAbs) were selected using a V3 fusion protein (V3-FP) which retains the conformation of the third variable region. The V3-FP consists of the V3 JR-CSF sequence inserted into a truncated form of murine leukemia virus gp70. Six human MAbs which recognize epitopes at the crown of the V3 loop were selected with the V3-FP. They were found to react more strongly with molecules displaying conformationally intact V3 than with linear V3 peptides. In a virus capture assay, these MAbs showed cross-clade binding to native, intact virions of clades A, B, C, D, and F. No binding was found to isolates from subtype E. The neutralizing activity of MAbs against primary isolates was determined in three assays: the GHOST cell assay, a phytohemagglutinin-stimulated peripheral blood mononuclear cell assay, and a luciferase assay. While these new MAbs displayed various degrees of activity, the pattern of cross-clade neutralization of clades A, B, and F was most pronounced. The neutralization of clades C and D viruses was weak and sporadic, and neutralization of clade E by these MAbs was not detected. Analysis by linear regression showed a highly significant correlation (P < 0.0001) between the strength of binding of these anti-V3 MAbs to intact virions and the percent neutralization. These studies demonstrate that human MAbs to conformation-sensitive epitopes of V3 display cross-clade reactivity in both binding to native, intact virions and neutralization of primary isolates.The third variable region (V3) of the gp120 envelope glycoprotein of human immunodeficiency virus type 1 (HIV-1) is critical for the infectivity of the virus, and certain single mutations in this region render the virus inactive (26,35). The sequence of the V3 loop plays a primary role in determining the cell tropism of the virus (8, 25), which is a function of its ability to differentially bind to the CCR5 and/or CXCR4 coreceptors on the target cell (22, 55). As a consequence of the key functions attributable to the V3 loop, anti-V3 antibodies (Abs) can be expected to block HIV-1 infectivity. Indeed, serum neutralization of T-cell line-adapted (TCLA) strains was found to be mediated by anti-V3 Abs (50, 65) and could be abolished by absorption with V3 peptides (2, 51) or by depletion of anti-V3 Abs (57). Moreover, monoclonal Abs (MAbs) against the V3 loop were shown to have potent neutralizing activity against TCLA strains, and this neutralization sensitivity to anti-V3 Abs led to the designation of the V3 loop as the principal neutralizing domain of the virus (27,47).Several studies, however, have weakened th...

Llama Antibody Fragments with Cross-Subtype Human Immunodeficiency Virus Type 1 (HIV-1)-Neutralizing Properties and High Affinity for HIV-1 gp120

“…MAb b12 (10) was kindly provided by D. Burton, Scripps Institute, La Jolla, CA. MAb GP68 (74) (29,37,40) and 447-52D (30,31) were obtained through the CFAR, NIBSC (original source, S. Zolla-Pazner). MAb 17b (79) was obtained from the NIH AIDS Research and Reference Reagent Program, Division of AIDS, NIAID, NIH (original source, J. E. Robinson).…”

“…5B). Included in the study was the broadly neutralizing MAb 2G12, which recognizes a carbohydrate motif on gp120 (9,70,72,80), the anti-V3 MAb 447-52D, which neutralizes primary isolates of subtype B (30,31), and MAb 17b, which recognizes a CD4-induced epitope and which neutralizes mainly T-cell-line-adapted isolates (79). In addition, anti-CD4bs MAb b12 and the anti-gp41 MAb 4E10 (9, 56) were included.…”

“…Two broadly neutralizing MAbs, 4E10 and 2F5, recognize gp41 (9,56,77,96). MAbs X5 (55), which recognizes an epitope on gp120 that is better-exposed after CD4 binding, and m14 (92), which competes with CD4 for binding to gp120, also display some neutralizing activity across HIV-1 subtypes, as do a minority of MAbs to the V3 region of gp120 (30,31).…”

Multi-subtype gp160 DNA immunization induces broadly neutralizing anti-HIV antibodies

“…The peptides corresponded to the gp120 constant regions 2 to 5 (C2-5) and variable regions 3 to 5 (V3-5). 28 We show that multi-subtype immunization with HIV-1 gp160 genes of subtypes A, B and C together with rGM-CSF results in the serum IgG recognition of four major gp120 sites (V3, C3, V4 and V5). The serum IgG from ABC DNA primed animals recognized 65% of all peptides in the constant and variable regions (Figure 2a).…”

Protein conformational changes induced by adsorption onto material surfaces: an important issue for biomedical applications of material science

“…Remodeling protein conformation therefore exposes different structural determinants that can be recognized by different antibodies. Such antibodies have been used to discriminate between the inactive and active conformations of a protein [72], to neutralize viruses [73] and to differentiate between the aggregated or monomeric form of proteins [74][75][76]. When proteins are adsorbed on material surfaces, some epitopes are masked, which reduces the binding of the corresponding antibodies [77][78][79].…”