Background: An effective approach for identifying epitopes that mediate neutralization of primary HIV-1 isolates is the isolation of monoclonal antibodies with neutralizing activities. Recently, we demonstrated that transgenic mice engineered to produce human IgGs (XenoMouse from Abgenix, Inc.) produced a broad range of antibodies in response to immunization with recombinant SF162 gp120, allowing the efficient isolation of fully human monoclonal antibodies (HumAbs) against a variety of sites. In the present study the efficacy of a number of different gp120-based recombinant antigens, including rgp120s from different primary strains and a rgp120-CD4 single chain molecule, were compared in the XenoMouse system.

Methods: Soluble rgp120s from 3 different R5 primary HIV-1 isolates (SF162, JR-FL, and BaL gp120) and a BaL gp120-CD4 fusion protein (FLSC, obtained from A. DeVico) were used with Ribi adjuvant to immunize XenoMouse mice producing human IgG2/k antibodies. The antibody titers and specificities of the resulting sera were analyzed and HumAbs were isolated and characterized from selected animals. Neutralization activities of the polyclonal sera and HumAbs were tested using luciferase-based assays with viral pseudotypes.

Results: All immunized mice developed antibody responses to the corresponding immunogens. Sera of mice immunized with SF162 gp120 contained neutralizing activity against the autologous virus, while JR-FL gp120, BaL gp120 and FLSC failed to induce detectable serum neutralizing activity. Broadly reactive HumAbs isolated from JR-FL gp120 immunized mice did not possess neutralizing activities for any of the strains tested. Thirty-nine (39) HumAbs were generated from FLSC-immunized mice; only 3 recognized native BaL gp120, while 36 were directed against conformational epitopes specific for the gp120-CD4 fusion protein. None of these HumAbs possessed neutralizing activities.

Conclusions: Of the 4 gp120-based immunogens tested, only SF162 gp120 induced neutralizing antibodies in this system. The gp120-CD4 single chain fusion protein was quite immunogenic, but the great majority of HumAbs isolated were directed against non-native epitopes specific for the fusion protein. This study highlights the variability of the humoral response against different rgp120s and suggests possible limitations in the utility of the single chain gp120-CD4 protein as an effective immunogen.