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. 1998 Dec;72(12):9656-67.
doi: 10.1128/JVI.72.12.9656-9667.1998.

Neutralizing antibodies from the sera of human immunodeficiency virus type 1-infected individuals bind to monomeric gp120 and oligomeric gp140

N M Stamatos  1 , J R Mascola, V S Kalyanaraman, M K Louder, L M Frampton, D L Birx, T C VanCott
Affiliations

Neutralizing antibodies from the sera of human immunodeficiency virus type 1-infected individuals bind to monomeric gp120 and oligomeric gp140

N M Stamatos et al. J Virol. 1998 Dec.

Abstract

Antibodies that neutralize primary isolates of human immunodeficiency virus type 1 (HIV-1) appear during HIV-1 infection but are difficult to elicit by immunization with current vaccine products comprised of monomeric forms of HIV-1 envelope glycoprotein gp120. The limited neutralizing antibody response generated by gp120 vaccine products could be due to the absence or inaccessibility of the relevant epitopes. To determine whether neutralizing antibodies from HIV-1-infected patients bind to epitopes accessible on monomeric gp120 and/or oligomeric gp140 (ogp140), purified total immunoglobulin from the sera of two HIV-1-infected patients as well as pooled HIV immune globulin were selectively depleted of antibodies which bound to immobilized gp120 or ogp140. After passage of each immunoglobulin preparation through the respective columns, antibody titers against gp120 and ogp140 were specifically reduced at least 128-fold. The gp120- and gp140-depleted antibody fraction from each serum displayed reduced neutralization activity against three primary and two T-cell line-adapted (TCLA) HIV-1 isolates. Significant residual neutralizing activity, however, persisted in the depleted sera, indicating additional neutralizing antibody specificities. gp120- and ogp140-specific antibodies eluted from each column neutralized both primary and TCLA viruses. These data demonstrate the presence and accessibility of epitopes on both monomeric gp120 and ogp140 that are specific for antibodies that are capable of neutralizing primary isolates of HIV-1. Thus, the difficulties associated with eliciting neutralizing antibodies by using current monomeric gp120 subunit vaccines may be related less to improper protein structure and more to ineffective immunogen formulation and/or presentation.

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Figures

FIG. 1
FIG. 1
Preparation of purified Ig and depleted samples, design of viral neutralization assays, and evaluation of p24 antigen were as described in Materials and Methods. (A) Results for gp120- and ogp140-depleted material from US20; (B) results for HIVIG. Each point represents the average from triplicate wells from one experiment of four that gave similar results. Depleted fractions were standardized to the purified Ig and serum by equating reactivity to p24 such that equivalent amounts of each were added. Diluted samples were evaluated by ELISA for titers of Abs to p24 after the neutralization assay to confirm that equivalent amounts were added. The amount of viral growth in comparable dilutions of NHS was used as the standard for 100% virus growth. Purified Ig and depleted fractions were assessed for neutralizing capacity against primary (US1, CM237, and 056) and TCLA (MN and IIIB) HIV-1 isolates. gp120-depleted and ogp140-depleted fractions correspond to HIV-1 serum US20 or HIVIG depleted over the gp120 or ogp140 affinity columns, respectively.
FIG. 1
FIG. 1
Preparation of purified Ig and depleted samples, design of viral neutralization assays, and evaluation of p24 antigen were as described in Materials and Methods. (A) Results for gp120- and ogp140-depleted material from US20; (B) results for HIVIG. Each point represents the average from triplicate wells from one experiment of four that gave similar results. Depleted fractions were standardized to the purified Ig and serum by equating reactivity to p24 such that equivalent amounts of each were added. Diluted samples were evaluated by ELISA for titers of Abs to p24 after the neutralization assay to confirm that equivalent amounts were added. The amount of viral growth in comparable dilutions of NHS was used as the standard for 100% virus growth. Purified Ig and depleted fractions were assessed for neutralizing capacity against primary (US1, CM237, and 056) and TCLA (MN and IIIB) HIV-1 isolates. gp120-depleted and ogp140-depleted fractions correspond to HIV-1 serum US20 or HIVIG depleted over the gp120 or ogp140 affinity columns, respectively.
FIG. 2
FIG. 2
Affinity-purified antibodies from the gp120451 and ogp140451 affinity columns for US20 (A) and from HIVIG (B) were compared with the respective unfractionated Ig for neutralization activity against primary (US1, CM237, and 056) and TCLA (MN and IIIB) HIV-1 isolates. Abs affinity purified from the monomeric gp120 column (gp120 Abs) or ogp140 column (ogp140 Abs) were standardized to the unfractionated serum and/or purified Ig by adding equivalent amounts of reactivity to gp120 or ogp140, respectively. Dilutions were evaluated by ELISA for titers to gp120 and/or ogp140 after the assay to confirm that equivalent amounts of reactivity had been added.
FIG. 2
FIG. 2
Affinity-purified antibodies from the gp120451 and ogp140451 affinity columns for US20 (A) and from HIVIG (B) were compared with the respective unfractionated Ig for neutralization activity against primary (US1, CM237, and 056) and TCLA (MN and IIIB) HIV-1 isolates. Abs affinity purified from the monomeric gp120 column (gp120 Abs) or ogp140 column (ogp140 Abs) were standardized to the unfractionated serum and/or purified Ig by adding equivalent amounts of reactivity to gp120 or ogp140, respectively. Dilutions were evaluated by ELISA for titers to gp120 and/or ogp140 after the assay to confirm that equivalent amounts of reactivity had been added.
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