Production of site-selected neutralizing human monoclonal antibodies against the third variable domain of the human immunodeficiency virus type 1 envelope glycoprotein.

Abstract

Cell lines secreting IgG1 human monoclonal antibodies (mAbs) to the envelope glycoprotein, gpl2O, of human immunodeficiency virus (IRV) have been produced by transformation of peripheral blood cells front IIV-infected individuals and by fusion of transformed cells to a humanmouse heteromyeloma cell line (SHM-D33). Two human mAbs were site-selected by means of a 23-mer synthetic peptide spanning a portion of the third variable domain of gpI20 from the MN strain of HIV. The two heterohybridomas produce three times more IgG than do their parent lymphoblastoid cell lines. The specificities of these mAbs have been mapped to sequences near the tip of the disulfide loop of the gpl20 third variable domain, Lys-Arg-lle-His-Ile and His-Ie-Gly-Pro-GlyArg, respectively. The mAbs have dissociation constants of 3.7 x 10-6 M and 8.3 x 10-7 M, neutralize HIVMN in vitro at nanogram levels, and bear the characteristics of antibodies associated with protective immunity in vivo.

Toward Effective HIV Vaccination

“…To determine whether the mAb YZ23 epitope overlaps previously reported neutralizing epitopes, we measured the ability of mAb YZ23 to inhibit binding of gp120 (MN strain) by the following reference mAbs: IgG b12 directed to a CD4BS epitope not involving residues 421-433 (60, 61); mAb 2G12 to a conformational carbohydrate-dependent epitope (62,63); mAbs 17b and 48d to a CD4-inducible epitope exposed upon gp120 binding to sCD4 (64,65); and mAb 447-52D to the conserved V3 apex epitope (66,67). These mAbs neutralize some but not all strains within and across group M HIV subtypes (60, 68 -70).…”

Interaction of the HIV-1 gp120 Viral Protein V3 Loop with Bacterial Lipopolysaccharide

“…We demonstrated that Re-LPS inhibited binding of fluorescently labeled gp120 to the surface of the H9 cells (Fig. 5A) similarly to anti-gp120 monoclonal antibodies 257-D IV or 268-D IV (45), which react with the HIV-1 MN V3 epitope KRIH or HIGPGR (data not shown).…”

Selection of a novel gp41-specific HIV-1 neutralizing human antibody by competitive antigen panning

“…Many HIV-1-neutralizing human monoclonal antibodies (nhmAbs) have been identified by immortalization of B lymphocytes from HIV-infected patients eitherby EBV transformation (Gorny et al, 1989;Robinson et al, 1990) or by cell fusion (Grunow et al, 1988;Buchacher et al, 1994) (hybridomas obtained by fusion of EBV transformants with heteromyeloma cells have also been extensively used (Posner et al, 1987;Gorny et al, 1991;Posner et al, 1991)] followed by screening of their supernatants for antigen-specific antibodies. Selection of HIV-1-neutralizing antibodies from phage-displayed human antibody libraries has also been used by panning against one antigen (Burton et al, 1991) or several antigens sequentially ; it is a powerfull and versatile approach that allows modifications of the panning process for enhanced selection of antibodies with desirable properties (Zhang and Dimitrov, 2006, in press).…”