Characterization of antibodies that inhibit HIV gp120 antigen processing and presentation

Abstract

Antibodies to the CD4-binding site (CD4bs) of HIV-1 envelope gp120 have been shown to inhibit MHC class II presentation of this antigen, but the mechanism is not fully understood. To define the key determinants contributing to the inhibitory activity of these antibodies, a panel of anti-CD4bs monoclonal antibodies with different affinities was studied and compared to antibodies specific for the chemokine receptor-binding site or other gp120 regions. Anti-CD4bs antibodies that completely obstruct gp120 presentation exhibit three common properties: relatively high affinity for gp120, acidstable interaction with gp120, and the capacity to slow the kinetics of gp120 proteolytic processing. None of these antibodies prevents gp120 internalization into APC. Notably, the broadly virus-neutralizing anti-CD4bs IgG1b12 does not block gp120 presentation as strongly, because although IgG1b12 has a relatively high affinity, it dissociates from gp120 more readily at acidic pH and only moderately retards gp120 proteolysis. Other anti-gp120 antibodies, regardless of their affinities, do not affect gp120 presentation. Hence, high-affinity anti-CD4bs antibodies that do not dissociate from gp120 at endolysosomal pH obstruct gp120 processing and prevent MHC class II presentation of this antigen. The presence of such antibodies could contribute to the dearth of antigp120 T helper responses in chronically HIV-1-infected patients.

Detection of HIV gp120 in Plasma During Early HIV Infection Is Associated with Increased Proinflammatory and Immunoregulatory Cytokines

“…This mixture of antibodies is known to cross-react with envelope glycoproteins from multiple Clade B isolates and likely binds to monomeric gp120. [13][14][15][16] Plasma was added to the plate in duplicate and gp120 was detected using a 1/2000 dilution of the same mixture of biotinylated antibodies. The plate was then developed using standard methods.…”

Identification of an N-Linked Glycosylation in the C4 Region of HIV-1 Envelope gp120 That Is Critical for Recognition of Neighboring CD4 T Cell Epitopes

“…ELISA to detect mAb binding to gp120 proteins was done as previously described (49). Gp120 proteins were captured onto the wells by sheep anti-C5 Abs (Cliniqa) and reacted with different human anti-gp120 mAbs.…”

“…T cell proliferation was assessed by the standard [ 3 H]thymidine incorporation assay as described previously (49). Autologous PBMCs or mouse splenocytes were irradiated (12,000 rad) and treated with Ags for 18 -22 h before use as APCs in the assay.…”

“…Gp120 proteins were incubated with lysosomal fraction (0.75 million cell equivalents per g of gp120) overnight at 37°C in 0.08 M of sodium acetate buffer at pH 5. The kinetics of gp120 digestion were determined by measuring intact gp120 remaining over time in the reaction using a sandwich ELISA with Abs specific for the C and N termini of gp120 (49).…”

Influence of Disulfide-Stabilized Structure on the Specificity of Helper T-Cell and Antibody Responses to HIV Envelope Glycoprotein gp120

“…The globally reduced T-cell responses suggest that the destabilized protein was more sensitive to proteolytic processing in the antigen-presenting cell and that the resulting lower yield of major histocompatibility complex class II (MHC II) ligands reduced presentation. However, alternative explanations for the disulfide-bond variant's poor T-cell immunogenicity cannot be ruled out, such as that unique processing intermediates interfered with trafficking of MHC II-peptide complexes to the cell surface (7) or that unique antibodies interfered with antigen processing (46).…”