Predicting the broadly neutralizing antibody susceptibility of the HIV reservoir

Abstract

Broadly neutralizing antibodies (bNAbs) against HIV-1 are under evaluation for both prevention and therapy. HIV-1 sequence diversity observed in most HIV-infected individuals and archived variations in critical bNAb epitopes present a major challenge for the clinical application of bNAbs, as preexistent resistant viral strains can emerge, resulting in bNAb failure to control HIV. In order to identify viral resistance in patients prior to antibody therapy and to guide the selection of effective bNAb combination regimens, we developed what we believe to be a novel Bayesian machine-learning model that uses HIV-1 envelope protein sequences and foremost approximated glycan occupancy information as variables to quantitatively predict the half-maximal inhibitory concentrations (IC50) of 126 neutralizing antibodies against a variety of cross clade viruses. We then applied this model to peripheral blood mononuclear cell-derived proviral Env sequences from 25 HIV-1-infected individuals mapping the landscape of neutralization resistance within each individual's reservoir and determined the predicted ideal bNAb combination to achieve 100% neutralization at IC50 values <1 μg/ml. Furthermore, predicted cellular viral reservoir neutralization signatures of individuals before an analytical antiretroviral treatment interruption were consistent with the measured neutralization susceptibilities of the respective plasma rebound viruses, validating our model as a potentially novel tool to facilitate the advancement of bNAbs into the clinic.

Keywords: AIDS/HIV; Bioinformatics; Diagnostics; Immunotherapy; Therapeutics.

Figure 1. Prediction model performance for selected mAb neutralization sensitivities (log₂ IC50) representing relevant envelope broadly neutralizing antibody target sites (V2 apex, V3 loop, CD4-binding site, gp120/gp41 interface, and MPER).

Scatter plots illustrate the correlation between measured log₂(IC50) using neutralization data from the Los Alamos “Compile, Analyze and Tally NAb Panels” (CATNAP) database and log₂(IC50) values predicted by our Bayesian machine-learning model. For each antibody, log₂(IC50) prediction based on HIV-1 envelope (Env) sequence alone or using sequence and approximated glycan occupancy information is shown. Overall, 71–717 available paired env sequence-neutralization values per mAb were available. The Spearman’s rho and its 2-sided P value are labeled.

Figure 2. IC50 values (μg/mL) for outgrown viruses, as determined by TZM-bl neutralization assay.

PBMCs from 25 HIV-infected individuals were stimulated and infectious culture supernatants were tested for neutralization sensitivity against the broadly neutralizing antibodies (bNAb) VRC01, 3BNC117, PGT121, and PGDM1400 in a TZM-bl neutralization assay using increasing amounts of antibody. IC50 values ≤0.1 μg/mL are highlighted in red, values between 0.1 and 1 μg/mL are highlighted in orange, values between 1 and 10 μg/mL are highlighted in yellow, values between 10 and 25 μg/mL are highlighted in green, and values >25 μg/mL are highlighted gray.

Figure 3. Predicted log₂(IC50) values for proviral HIV-1 envelope sequences from all 25 HIV-infected study participants.

In total, 727 single-genome full-length envelope (Env) sequences were obtained from DNA isolated from primary PBMCs with a mean of 29 sequences per individual (range 8–83). Shown are box-and-whisker plots demonstrating the breadth of neutralization sensitivities against multiple class broadly neutralizing antibodies (bNAb) and for each participant’s proviral sequences. Each dot represents the predicted IC50 values for an Env sequence.

Figure 4. Maximum likelihood phylogenetic trees of proviral HIV-1 envelope sequences from 4 randomly selected study participants.

For participant 239667, proviral sequence from 2 time points (blue and green boxes) are shown. For some sequences (identified by numbers), predicted IC50 values (in μg/mL) for the bNAbs 3BNC117, VRC01, PGT121, and PGDM1400 are shown in the chart to demonstrate the heterogeneity in neutralization susceptibility across sequences and broadly neutralizing antibodies (bNAb). IC50 values ≤0.1 μg/mL are highlighted in red, values between 0.1 and 1 μg/mL are highlighted in orange, values between 1 and 10 μg/mL are highlighted in yellow, values between 10 and 50 μg/mL are highlighted in green, and values >50 μg/mL are highlighted white. Genetic distance scale bars are shown for each tree.

Figure 5. Predicted IC50 values (μg/mL) based on proviral HIV-1 envelope sequences from multiple time points per individual.

For 3 of the 25 study participants, proviral sequences from a second time point (3 months to 1 year after the first time point) were available and for 2 study participants 3 time points up to 8 years apart were available. Shown are predicted IC50 values for the broadly neutralizing antibodies (bNAb) 3BNC117, VRC01, PGT121, and PGDM1400. Each dot represents the predicted IC50 values for an envelope (Env) sequence. P values were calculated by 2-sided Wilcoxon rank-sum test and multiple testing corrected by the Benjamini and Hochberg method. *P < 0.05; **P < 0.01; ***P < 0.001.

Figure 6. Predicted log₂(IC50) values for proviral HIV-1 envelope sequences from 9 participants of the NIH 15-I-0140 trial.

A total of 199 single-genome envelope (Env) sequences were obtained from isolated CD4⁺ T cells from 9 participants of the NIH 15-I-0140 trial prior to VRC01 administration and prior to analytical antiretroviral treatment interruption (ATI). Shown are box-and-whisker plots demonstrating the breadth of predicted neutralization sensitivities against multiple class broadly neutralizing antibodies (bNAb) and for each participant’s proviral sequences. Each dot represents the predicted IC50 values for an Env sequence.

Figure 7. Comparison between IC50 prediction and measured IC50 values using isolated plasma HIV-1 envelope sequences from 6 participants of the NIH 15-I-0140 trial following viral rebound during ATI.

Twenty-four pseudoviruses were generated based on plasma virus envelope (Env) sequences. IC50 values were predicted based on the Env sequences, and neutralization of the respective pseudoviruses against the broadly neutralizing antibodies (bNAb) 3BNC117, VRC01, 10-1074, and PGT121 was measured using a TZM/bl neutralization assay. The regression lines with 95% confidence interval boundaries are indicated in black and blue dotted lines. The Spearman’s rho and its 2-sided P value are labeled.