Development of Broadly Neutralizing Antibodies and Their Mapping by Monomeric gp120 in Human Immunodeficiency Virus Type 1-Infected Humans and Simian-Human Immunodeficiency Virus SHIVSF162P3N-Infected Macaques

Abstract

To improve our understanding of the similarities and differences between neutralizing antibodies elicited by simian-human immunodeficiency virus (SHIV)-infected rhesus macaques and human immunodeficiency virus type 1 (HIV-1)-infected humans, we examined the plasma of 13 viremic macaques infected with SHIVSF162P3Nand 85 HIV-1-infected humans with known times of infection. We identified 5 macaques (38%) from 1 to 2 years postinfection (p.i.) with broadly neutralizing antibodies (bnAbs) against tier 2 HIV-1. In comparison, only 2 out of 42 (5%) human plasma samples collected in a similar time frame of 1 to 3 years p.i. exhibited comparable neutralizing breadths and potencies, with the number increasing to 7 out of 21 (30%) after 3 years p.i. Plasma mapping with monomeric gp120 identified only 2 out of 9 humans and 2 out of 4 macaques that contained gp120-reactive neutralizing antibodies, indicating distinct specificities in these plasma samples, with most of them recognizing the envelope trimer (including gp41) rather than the gp120 monomer. Indeed, a total of 20 gp120-directed monoclonal antibodies (MAbs) isolated from a human subject (AD358) and a Chinese rhesus macaque (GB40) displayed no or limited neutralizing activity against tier 2 strains. These isolated MAbs, mapped to the CD4-binding site, the V3 loop, the inner domain, and the C5 region of gp120, revealed genetic similarity between the human and macaque immunoglobulin genes used to encode some V3-directed MAbs. These results also support the use of envelope trimer probes for efficient isolation of HIV-1 bnAbs.

Importance: HIV-1 vaccine research can benefit from understanding the development of broadly neutralizing antibodies (bnAbs) in rhesus macaques, commonly used to assess vaccine immunogenicity and efficacy. Here, we examined 85 HIV-1-infected humans and 13 SHIVSF162P3N-infected macaques for bnAbs and found that, similar to HIV-1-infected humans, bnAbs in SHIV-infected macaques are also rare, but their development might have been faster in some of the studied macaques. Plasma mapping with monomeric gp120 indicated that most bnAbs bind to the envelope trimer rather than the gp120 monomer. In support of this, none of the isolated gp120-reactive monoclonal antibodies (MAbs) displayed the neutralization breadth observed in the corresponding plasma. However, the MAb sequences revealed similarity between human and macaque genes used to encode some V3-directed MAbs. Our study sheds light on the timing and development of bnAbs in SHIV-infected macaques in comparison to HIV-1-infected humans and highlights the use of envelope trimer probes for efficient recovery of bnAbs.

FIG 1

Neutralization ID₅₀ titers of plasma (reciprocal dilutions) from 85 HIV-1-infected human subjects from the ADARC cohort against 10 HIV-1 Env pseudoviruses as indicated, with clades in parentheses. The dashed horizontal lines indicate the tested limit of 1:25 dilution; resistant values (<25) are plotted between 11 and 20. The study subjects are grouped based on the estimated lengths of their infection times. mpi, months postinfection; YPI, year(s) postinfection.

FIG 2

Neutralization ID₅₀ titers of plasma (reciprocal dilutions) from 13 viremic SHIV_SF162P3N-infected rhesus macaques against 4 homologous SHIV Env pseudoviruses (top) and 10 HIV-1 Env pseudoviruses (bottom), as indicated. The dashed horizontal lines indicate the tested limit of 1:25 dilution; resistant values (<25) are plotted between 11 and 20. The blue symbols in the top diagram indicate autologous responses, as animals EE29 and FF59 were infected with the SHIV_SF162P3N inoculum-derived molecular clones 8 and 11 and animals FD83 and FF94 were infected with clone 8. Each animal was tested at two time points. wpi, weeks postinfection.

FIG 3

Characterization of recombinant gp120s for ELISA binding with CD4-Ig (top) and VRC01 (middle) and for neutralization competition with VRC01 (bottom). The gp120 sequences were based on JR-FL, Yu2, and AC10.29, as indicated. Each strain was tested as the WT or with a point mutation (D368R or W479A). (Bottom) For neutralization competition assays, the D368R and W479A gp120 mutants were tested at 25 μg/ml to adsorb VRC01 neutralizing activity against the indicated gp120-matched strains.

FIG 4

Plasma-mapping analysis using recombinant gp120 proteins. Broadly neutralizing plasma samples from 9 HIV-1-infected humans (A) and 4 SHIV_SF162P3N-infected macaques (B) were tested for antibody neutralization against HIV-1 strain JR-FL, Yu2, or AC10.29, as indicated, in the presence of strain-matched gp120 recombinant proteins with a D368R or W479A mutation. The gp120s were tested at 25 μg/ml to adsorb plasma neutralizing activity. Compared with mock controls, reduced or ablated activities indicate the presence of neutralizing antibodies directed to the corresponding gp120s.

FIG 5

Isolation and characterization of AD358 MAbs. (A) B-cell staining and sorting from the PBMCs of AD358_66mpi by FACS. SSC, side scatter; FSC, forward scatter. (B) ELISA binding curves of AD358 MAbs to gp120 and RSC3 proteins, as indicated. (C) Competition ELISA analysis of AD358 MAbs.

FIG 6

Isolation and characterization of GB40 MAbs. (A) B-cell staining and sorting from the PBMCs of GB40_75wpi by FACS. (B) ELISA and competition ELISA analyses of 8 GB40 MAbs directed to the V3 loop of gp120. (C) ELISA and competition ELISA analyses of 5 non-V3-loop-directed GB40 MAbs; the dashed lines in the top left graph show MAb binding curves to endo H-treated gp120.