Induction of antibodies in rhesus macaques that recognize a fusion-intermediate conformation of HIV-1 gp41 - PubMed (original) (raw)

doi: 10.1371/journal.pone.0027824. Epub 2011 Nov 30.

Laura L Sutherland, Frederick H Jaeger, Kara M Anasti, Robert Parks, Shelley Stewart, Cindy Bowman, Shi-Mao Xia, Ruijun Zhang, Xiaoying Shen, Richard M Scearce, Gilad Ofek, Yongping Yang, Peter D Kwong, Sampa Santra, Hua-Xin Liao, Georgia Tomaras, Norman L Letvin, Bing Chen, S Munir Alam, Barton F Haynes

Affiliations

Induction of antibodies in rhesus macaques that recognize a fusion-intermediate conformation of HIV-1 gp41

S Moses Dennison et al. PLoS One. 2011.

Abstract

A component to the problem of inducing broad neutralizing HIV-1 gp41 membrane proximal external region (MPER) antibodies is the need to focus the antibody response to the transiently exposed MPER pre-hairpin intermediate neutralization epitope. Here we describe a HIV-1 envelope (Env) gp140 oligomer prime followed by MPER peptide-liposomes boost strategy for eliciting serum antibody responses in rhesus macaques that bind to a gp41 fusion intermediate protein. This Env-liposome immunization strategy induced antibodies to the 2F5 neutralizing epitope ⁶⁶⁴DKW residues, and these antibodies preferentially bound to a gp41 fusion intermediate construct as well as to MPER scaffolds stabilized in the 2F5-bound conformation. However, no serum lipid binding activity was observed nor was serum neutralizing activity for HIV-1 pseudoviruses present. Nonetheless, the Env-liposome prime-boost immunization strategy induced antibodies that recognized a gp41 fusion intermediate protein and was successful in focusing the antibody response to the desired epitope.

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Conflict of interest statement

Competing Interests: The authors have declared that no competing interests exist.

Figures

Figure 1

Figure 1. Comparison of gp41 MPER specific antibody responses induced in guinea pigs by JRFL gp140CF and MPER peptide-liposomes.

The immunization scheme of the study is shown in the top panel. A–B: The 2F5 epitope peptide (SP62 peptide) specific responses in guinea pigs sera (at 1∶50 dilution) determined by SPR are shown after vaccination with JRFL gp140CF (A) and MPER peptide-liposome (B). PB, pre-bleed; P1–P3, post-immune bleeds 1–3. C–D: SPR sensogram displaying the comparison of 2F5 epitope peptide specific binding of guinea pig 1402 (C) and 1703 (D) sera at different time points with 2F5 mAb is shown as representative data. The estimated dissociation rates (kd) from the sensogram are indicated for 2F5 mAb and Post-bleed 3 (P3). E: Epitope mapping of post-immune bleed 3 of guinea pigs 1402 (immunized with JRFL gp140CF alone) and 1702–1705 (immunized with MPER656 peptide-liposomes alone) is shown in comparison to 2F5 mAb. The normalized binding shown is the ratio between binding responses of sera to the alanine scanning mutant peptides and WT 2F5 epitope peptide SP62. Alanine substitution of residues that resulted in ≥50% reduction in binding are indicated adjacent to the symbols. Data is representative of at least two measurements on adjacent spots on the same sensor chip.

Figure 2

Figure 2. MPER specific immunogenicity and fine specificity of different prime-boost regimens in guinea pigs.

A–C: Immunization scheme (A), 2F5 epitope peptide (SP62 peptide) specific binding serum responses (B) and fine specificity of post immune bleed 7 (C) are shown for MPER656 liposome prime-JRFL gp140CF boost regimen are shown. PB, pre-bleed; P1–P7, post-immune bleeds 1–7. D–F: The JRFL gp140CF prime-MPER656 liposome boost regimen scheme (D), 2F5 epitope peptide specific binding serum responses € and the fine specificity of post immune bleed 4 (F) are shown. Data is representative of at least two measurements on adjacent spots on the same sensor chip.

Figure 3

Figure 3. Comparison of MPER specific immunogenicity of different immunogens in rhesus macaques.

The immunization scheme is shown at the top. A–B: MPER specific binding responses of sera from immunized rhesus macaques at different time points are shown for (A) JRFL gp140CF (ASO1B adjuvant) and (B) MPER peptide liposome immunogens (in emulsigen+oCpG). C–E: The fine specificities of MPER specific responses elicited by MPER liposomes in rhesus macaques are shown for post immunizations 2–5 sera. F: The alanine substituted peptides used in epitope mapping of MPER responses. The dotted ellipses in C, D and E highlight the predominance of 664D specific MPER responses. Data is representative of at least two measurements on adjacent spots on the same sensor chip.

Figure 4

Figure 4. MPER specific binding responses of Rhesus macaques sera in gp140 prime-MPER liposome boost regimen.

A: The immunization scheme involving JRFL gp140CF prime (ASO1B adjuvant) and MPER656/MPL-A/R848 (emulsigen+oCpG) liposome boost regimen is shown. The arrows indicate the weeks at which immunization was done. B: ELISA end point titer of the 2F5 epitope peptide specific responses in rhesus macaques sera at different time points. C: The 2F5 epitope peptide specific binding of monkeys sera determined by SPR is shown post vaccination at different time points. D: Representative data of SPR sensogram of rhesus macaque #41546 sera at different time points binding to 2F5 epitope peptide as compared to 2F5 mAb (10 µg/ml) is shown. The estimated dissociation rates (kd) are indicated for samples at week 51 and 56 as well as for the control 2F5 mAb. Data is representative of at least two measurements on adjacent spots on the same sensor chip.

Figure 5

Figure 5. Mapping of critical residues involved in MPER specific Rhesus macaques serum responses.

Normalized binding responses of immune sera from rhesus macaques primed with JRFL gp140CF and boosted with MPER656 liposomes. Panels A–D are for animals 03525, 41546, 51384 and 51902. The dotted circle highlights the mapping of MPER specific responses to D664KW residues as did 2F5 mAb. Data is representative of at least two measurements on adjacent spots on the same sensor chip.

Figure 6

Figure 6. The MPER specific serum responses in rhesus macaques recognize the mimic of fusion-intermediate gp41 structure.

A: Comparison of binding of 2F5 and 13H11 Fabs to GCN4-gp41-inter, a mimic of pre-hairpin intermediate structure of gp41. B–E: SPR sensogram of rhesus macaques sera (1∶50 diluted) binding to GCN4-gp41-inter. Each panel in B thru E compares the binding of sera obtained prior to MPER656 liposomes boost (week 48) and post-MPER656 liposomes boost (weeks 52–56). Data is representative of two measurements.

Figure 7

Figure 7. Comparison of JRFL gp140CF primed and MPER656 liposomes boosted rhesus macaque serum IgG binding to different conformational states of gp41.

A–D: SPR sensogram of rhesus macaques serum IgG (100 µg/ml) binding to GCN4-gp41 inter (solid lines), gp41-inter (dotted lines) and recombinant gp41 (broken lines). The IgGs purified from Week 56 (for animals 03525 and 51902), Week 52 (animal 41546) and Week 52 (animal 51834) sera were used. Data is representative of two measurements.

Figure 8

Figure 8. The JRFL gp140CF primed and MPER656 liposomes boosted rhesus macaque serum IgG bind scaffold proteins containing engrafted MPER in the 2F5 bound conformation.

A–C: SPR sensogram of 2F5, 11F10 and 13H11 mAbs at the indicated concentrations binding to ES2 (A), ES4 (B), and ES5 (C) scaffolds respectively are shown. D–F: The post MPER656 liposome immune serum IgG of rhesus macaques (150 µg/ml) binding to ES2 (D), E€(E), ES5 (F) are shown. For the sake of clarity, the scaffold binding responses of pre-MPER656 liposome serum IgG of only one animal (# 41546) is shown. The responses from other animal serum IgG were similar. Data is representative of at least two measurements.

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