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. 2013 Jan 24;38(1):176-86.
doi: 10.1016/j.immuni.2012.11.011. Epub 2013 Jan 11.

Vaccine induction of antibodies against a structurally heterogeneous site of immune pressure within HIV-1 envelope protein variable regions 1 and 2

Hua-Xin Liao  1 , Mattia Bonsignori, S Munir Alam, Jason S McLellan, Georgia D Tomaras, M Anthony Moody, Daniel M Kozink, Kwan-Ki Hwang, Xi Chen, Chun-Yen Tsao, Pinghuang Liu, Xiaozhi Lu, Robert J Parks, David C Montefiori, Guido Ferrari, Justin Pollara, Mangala Rao, Kristina K Peachman, Sampa Santra, Norman L Letvin, Nicos Karasavvas, Zhi-Yong Yang, Kaifan Dai, Marie Pancera, Jason Gorman, Kevin Wiehe, Nathan I Nicely, Supachai Rerks-Ngarm, Sorachai Nitayaphan, Jaranit Kaewkungwal, Punnee Pitisuttithum, James Tartaglia, Faruk Sinangil, Jerome H Kim, Nelson L Michael, Thomas B Kepler, Peter D Kwong, John R Mascola, Gary J Nabel, Abraham Pinter, Susan Zolla-Pazner, Barton F Haynes
Affiliations

Vaccine induction of antibodies against a structurally heterogeneous site of immune pressure within HIV-1 envelope protein variable regions 1 and 2

Hua-Xin Liao et al. Immunity. .

Abstract

The RV144 HIV-1 trial of the canary pox vector (ALVAC-HIV) plus the gp120 AIDSVAX B/E vaccine demonstrated an estimated efficacy of 31%, which correlated directly with antibodies to HIV-1 envelope variable regions 1 and 2 (V1-V2). Genetic analysis of trial viruses revealed increased vaccine efficacy against viruses matching the vaccine strain at V2 residue 169. Here, we isolated four V2 monoclonal antibodies from RV144 vaccinees that recognize residue 169, neutralize laboratory-adapted HIV-1, and mediate killing of field-isolate HIV-1-infected CD4(+) T cells. Crystal structures of two of the V2 antibodies demonstrated that residue 169 can exist within divergent helical and loop conformations, which contrasted dramatically with the β strand conformation previously observed with a broadly neutralizing antibody PG9. Thus, RV144 vaccine-induced immune pressure appears to target a region that may be both sequence variable and structurally polymorphic. Variation may signal sites of HIV-1 envelope vulnerability, providing vaccine designers with new options.

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Figures

Figure 1
Figure 1. Binding of RV44 mAbs CH58 and CH59 to HIV-1-infected cells and to HIV-1 V2 peptides
A. Effect of alanine point substituted mutations on the binding of CH58 (in blue) and CH59 (in red) to the HIV-1 V2 peptide. For each mutation (y axis), results were normalized as EC50 relative to wild-type V2 peptide. B. Shown is the flow cytometric analysis of binding of mAbs CH58 (upper left), CH59 (middle left), and A32 (lower left) to the activated PB CD4+ T cells infected IMCCM235. Synagis (anti-respiratory syncytial virus mAb) and mAb HIV-1 A32 were used as negative and positive controls, respectively. Mean fluorescence intensity (MFI) and % of positive cells are indicated next to the histograms. Data shown are representative of 3 independent experiments. See also Figure S1 and Table S1.
Figure 2
Figure 2. Binding of RV144 V2 and PG9 bnAbs to AE.A244 V1–V2 tags protein and AE.A244 gp120Δ11
Each of the mAbs was captured on an anti-Fc antibody immobilized sensor surface to about 100–125 RU. For binding to A244gp120 Δ11, 2–10 ug/mL (CH58, A), 2–10 ug/mL (CH59, B), 10–50 ug per mL (PG9, C) of monomeric gp120 were injected over each of the mAbs. AE.A244 V1–V2 tags protein was injected at concentrations ranging from 0.5 – 5ug/mL (CH58, D), 0.1 – 5 ug/mL (CH59, E), 10–100 ug/mL (PG9, F). A negative control mAb (Synagis) was used to subtract non-specific binding. Each plot shows binding curves with increasing concentrations of gp120 or V1–V2 proteins (shown in different colors) injected over two independent flow cells immobilized with the same mAbs. For binding to CH58 and CH59 mAbs, A244 gp120Δ11 protein was injected at 2, 4, 6, 8, and 10ug/mL and AE.A244 V1–V2 protein at 0.2, 0.5, 1, 2, 3, 4 and 5 ug/mL. For PG9 mAb, A244 gp120Δ11 and AE.A244 V1–V2 proteins were injected at 10, 20, 30, 40 and 50ug/mL and 10, 25, 50, 75 and 100 ug/mL respectively. Global Curve fitting (shown in black) to a 1:1 Langmuir model was used to derive rate constants and Kd values following simultaneous fitting to binding data from two independent flow cells with the same mAb captured. A third flow cell with each of the mAbs gave similar rate constant values. See also Figure S2 and Tables S2.
Figure 3
Figure 3. Structures of antibodies CH58 and CH59 bound to an HIV-1 gp120 V2 peptide
Vaccine-elicited antibodies CH58 and CH59 recognize alternative conformations of V2 compared to bnAb PG9. A. top: Ribbon representation of the CH58 antigen-binding fragment in complex with an A244 V2 peptide. Heavy chain is colored orange, light chain is blue, and peptide is green. The sequence of the peptide is shown, with modeled residues in green; bottom: Close-up of the top panel rotated 90° about a horizontal axis. The side-chains of residues involved in hydrogen bonds or salt bridges are shown as sticks, with the interactions depicted as dashed lines. B, Structure of CH59 in complex with peptide, depicted as in A. The heavy chain is tan, and the light chain is light blue. C, Structure of bnAb PG9 in complex with the V1–V2 domain from HIV-1 strain CAP45 (PDB ID: 3U4E)(McLellan et al., 2011). The PG9 structure is shown as ribbons with heavy and light chains (colored yellow and blue, respectively) in the same orientation as in A and B. The V1–V2 domain is shown as a grey ribbon with residues 168–176 colored green, and N-linked glycans attached to residues Asn156 and Asn160 shown as sticks. See also Figures S3 and Table S3.
Figure 4
Figure 4. See also Figures S4 and S6. Effect of V2 mAbs CH58, CH59, HG107, HG120 footprint mutations in HIV-1 vaccine AE.A244 Env and RV144 breakthrough AE.427299 and AE.703357 Envs on ability of V2 mAbs to mediate ADCC
Panels show the ability of mAb A32 and RV144 V2 mAbs CH58, CH59, HG107, HG120 to mediate ADCC against gp120-coated CD4 cell (CEMCCR5) target T cells. Data shown is maximum percent granzyme B activity from ADCC. Top panel shows that CH58, CH59, HG107 and HG120 mAbs all mediate high levels of ADCC against WT AE.244 Env coated CD4 T cell targets (white bars), and this killing is mitigated by a single K169V mutation (grey bars), and is abrogated by the full V2 mAb footprint set of mutations (black bars). Middle and lower panels show, in contrast, that none of the CH58, CH59, HG107 and HG120 mAbs mediated ADCC against RV144 breakthrough Env AE.427299 and AE.703357 WT CD4 T cell targets (with V2s that did not match the RV144 vaccine) (white bars), and that ADCC was restored minimally with AE.703357 targets with the Q169K mutation (grey bars), and restored in a pronounced manner in both breakthrough Env targets with the full set of mutations that include Q169K that restored the V2 mAb footprint mutations (black bars). Purified NK cells isolated from a normal donor with Fc-gamma receptor IIIα FF phenotype were used as effector cells. The effector to target ratio was 10:1. Error bars show mean +/− SEM. Each antibody was tested in a wide dose curve starting at 40ug/ml with 4-fold dilutions. See also Figures S6 and Table S5.
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