The development of CD4 binding site antibodies during HIV-1 infection

Abstract

Broadly neutralizing antibodies to the CD4 binding site (CD4bs) of gp120 are generated by some HIV-1-infected individuals, but little is known about the prevalence and evolution of this antibody response during the course of HIV-1 infection. We analyzed the sera of 113 HIV-1 seroconverters from three cohorts for binding to a panel of gp120 core proteins and their corresponding CD4bs knockout mutants. Among sera collected between 99 and 258 weeks post-HIV-1 infection, 88% contained antibodies to the CD4bs and 47% contained antibodies to resurfaced stabilized core (RSC) probes that react preferentially with broadly neutralizing CD4bs antibodies (BNCD4), such as monoclonal antibodies (MAbs) VRC01 and VRC-CH31. Analysis of longitudinal serum samples from a subset of 18 subjects revealed that CD4bs antibodies to gp120 arose within the first 4 to 16 weeks of infection, while the development of RSC-reactive antibodies was more varied, occurring between 10 and 152 weeks post-HIV-1 infection. Despite the presence of these antibodies, serum neutralization mediated by RSC-reactive antibodies was detected in sera from only a few donors infected for more than 3 years. Thus, CD4bs antibodies that bind a VRC01-like epitope are often induced during HIV-1 infection, but the level and potency required to mediate serum neutralization may take years to develop. An improved understanding of the immunological factors associated with the development and maturation of neutralizing CD4bs antibodies during HIV-1 infection may provide insights into the requirements for eliciting this response by vaccination.

Figures

Fig 1

Prevalence of CD4 binding site antibodies among donors in three HIV-1 seroconverter cohorts. (A) The percentages of donors reacting by ELISA with the indicated protein probes are shown by the colored bars. A 2.5-fold or greater difference in serum binding to the wild type compared to the corresponding CD4bs knockout mutant probe was considered positive. For each cohort, the number of subjects analyzed (N) and the median time post-HIV-1 infection are indicated. (B) Summary of percent positive serum ELISA data for each core protein probe or the combination of both RSC3 and RSC3/G367R. Samples reactive with both RSC3 and RSC3/G367R have binding profiles similar to those of BNCD4 MAbs. The last column indicates the percentages of sera reactive with gp120 core that also reacted with both RSC3 and RSC3/G367R.

Fig 2

Lack of association between plasma viral load and RSC3-reactive CD4bs antibodies. RSC3-positive sera were those reactive with both RSC3 and RSC3/G367R. The viral load (VL) was calculated as the geometric mean of VL values between 24 weeks postinfection and the time of screening (set point), VL values 52 weeks prior to screening (prior), or the peak viral load in a subset of donors with early sampling time points (peak). The box plot shows the median values and the 25% and 75% quartiles. The range and outliers are shown by the bars and circles, respectively. neg., negative; pos., positive.

Fig 3

Kinetics of the CD4 binding site antibody responses in 18 selected subjects. Serial longitudinal serum samples from six subjects from each cohort were analyzed. The ELISA endpoint titers (y axis) are graphed for multiple weeks after HIV-1 infection (x axis). The line colors correspond to the protein probes shown in Fig. 1. The solid lines show reactivity to the probe, while the dashed lines indicate the corresponding CD4bs knockout mutant. The first time point reactive with RSC3 is indicated by the vertical dotted lines.

Fig 4

Neutralization breadth of RSC3-reactive serum binding. Selected sera from the CHAVI and CAPRISA cohorts (rows) that were ELISA reactive with both RSC3 and RSC3/G367R were screened for neutralization against six Env pseudoviruses (columns) representing three genetic subtypes. All viruses are tier 2 primary Envs, except for the tier 1 HXBc2, which was used to allow subsequent neutralization competition assays. Reciprocal serum ID50 values of >100 are highlighted in yellow, >400 in orange, and >1,000 in red. SIV, simian immunodeficiency virus.

Fig 5

Analysis of serum CD4bs-directed neutralization. Sera that were ELISA reactive with both RSC3 and RSC3/G367R (BNCD4-like probes) were assayed for inhibition of neutralization, using both RSC3 and RSC3/Δ371I/P363N as inhibitors. The net percent reduction in neutralization of HXBc2 attributed to RSC3 compared to RSC3/Δ371I/P363N is plotted (y axis), with each dot representing one subject. The number of subjects analyzed in each cohort is indicated on the x axis. A greater than 30% reduction is considered positive. The mean and standard error of the mean (SEM) for each cohort are indicated by the long and short horizontal lines.