Dataset Information

Developmental Pathway of the MPER-Directed HIV-1-Neutralizing Antibody 10E8.

ABSTRACT: Antibody 10E8 targets the membrane-proximal external region (MPER) of HIV-1 gp41, neutralizes >97% of HIV-1 isolates, and lacks the auto-reactivity often associated with MPER-directed antibodies. The developmental pathway of 10E8 might therefore serve as a promising template for vaccine design, but samples from time-of-infection-often used to infer the B cell record-are unavailable. In this study, we used crystallography, next-generation sequencing (NGS), and functional assessments to infer the 10E8 developmental pathway from a single time point. Mutational analysis indicated somatic hypermutation of the 2nd-heavy chain-complementarity determining region (CDR H2) to be critical for neutralization, and structures of 10E8 variants with V-gene regions reverted to genomic origin for heavy-and-light chains or heavy chain-only showed structural differences >2 Å relative to mature 10E8 in the CDR H2 and H3. To understand these developmental changes, we used bioinformatic sieving, maximum likelihood, and parsimony analyses of immunoglobulin transcripts to identify 10E8-lineage members, to infer the 10E8-unmutated common ancestor (UCA), and to calculate 10E8-developmental intermediates. We were assisted in this analysis by the preservation of a critical D-gene segment, which was unmutated in most 10E8-lineage sequences. UCA and early intermediates weakly bound a 26-residue-MPER peptide, whereas HIV-1 neutralization and epitope recognition in liposomes were only observed with late intermediates. Antibody 10E8 thus develops from a UCA with weak MPER affinity and substantial differences in CDR H2 and H3 from the mature 10E8; only after extensive somatic hypermutation do 10E8-lineage members gain recognition in the context of membrane and HIV-1 neutralization.

SUBMITTER: Soto C

PROVIDER: S-EPMC4907498 | biostudies-literature | 2016

REPOSITORIES: biostudies-literature

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Developmental Pathway of the MPER-Directed HIV-1-Neutralizing Antibody 10E8.

Soto Cinque C Ofek Gilad G Joyce M Gordon MG Zhang Baoshan B McKee Krisha K Longo Nancy S NS Yang Yongping Y Huang Jinghe J Parks Robert R Eudailey Joshua J Lloyd Krissey E KE Alam S Munir SM Haynes Barton F BF Mullikin James C JC Connors Mark M Mascola John R JR Shapiro Lawrence L Kwong Peter D PD

PloS one 20160614 6

Antibody 10E8 targets the membrane-proximal external region (MPER) of HIV-1 gp41, neutralizes >97% of HIV-1 isolates, and lacks the auto-reactivity often associated with MPER-directed antibodies. The developmental pathway of 10E8 might therefore serve as a promising template for vaccine design, but samples from time-of-infection-often used to infer the B cell record-are unavailable. In this study, we used crystallography, next-generation sequencing (NGS), and functional assessments to infer the ...[more]

PMID: 27299673

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Project description:The 10E8 antibody targets a helical epitope in the membrane-proximal external region (MPER) and transmembrane domain (TMD) of the envelope glycoprotein (Env) subunit gp41 and is among the broadest known neutralizing antibodies against HIV-1. Accordingly, this antibody and its mechanism of action valuably inform the design of effective vaccines and immunotherapies. 10E8 exhibits unusual adaptations to attain specific, high-affinity binding to the MPER at the viral membrane interface. Reversing the charge of the basic paratope surface (from net positive to net negative) reportedly lowered its neutralization potency. Here, we hypothesized that by increasing the net positive charge in similar polar surface patches, the neutralization potency of the antibody may be enhanced. We found that an increased positive charge at this paratope surface strengthened an electrostatic interaction between the antibody and lipid bilayers, enabling 10E8 to interact spontaneously with membranes. Notably, the modified 10E8 antibody did not gain any apparent polyreactivity and neutralized virus with a significantly greater potency. Binding analyses indicated that the optimized 10E8 antibody bound with a higher affinity to the epitope peptide anchored in lipid bilayers and to Env spikes on virions. Overall, our data provide a proof of principle for the rational optimization of 10E8 via manipulation of its interaction with the membrane element of its epitope. However, the observation that a similar mutation strategy did not affect the potency of the first-generation anti-MPER antibody 4E10 shows possible limitations of this principle. Altogether, our results emphasize the crucial role played by the viral membrane in the antigenicity of the MPER-TMD of HIV-1.IMPORTANCE The broadly neutralizing antibody 10E8 blocks infection by nearly all HIV-1 isolates, a capacity which vaccine design seeks to reproduce. Engineered versions of this antibody also represent a promising treatment for HIV infection by passive immunization. Understanding its mechanism of action is therefore important to help in developing effective vaccines and biologics to combat HIV/AIDS. 10E8 engages its helical MPER epitope where the base of the envelope spike submerges into the viral membrane. To enable this interaction, this antibody evolved an unusual property: the ability to interact with the membrane surface. Here, we provide evidence that 10E8 can be made more effective by enhancing its interactions with membranes. Our findings strengthen the idea that to elicit antibodies similar to 10E8, vaccines must reproduce the membrane environment where these antibodies perform their function.

Project description:UNLABELLED:Extraordinary antibodies capable of near pan-neutralization of HIV-1 have been identified. One of the broadest is antibody 10E8, which recognizes the membrane-proximal external region (MPER) of the HIV-1 envelope and neutralizes >95% of circulating HIV-1 strains. If delivered passively, 10E8 might serve to prevent or treat HIV-1 infection. Antibody 10E8, however, is markedly less soluble than other antibodies. Here, we describe the use of both structural biology and somatic variation to develop optimized versions of 10E8 with increased solubility. From the structure of 10E8, we identified a prominent hydrophobic patch; reversion of four hydrophobic residues in this patch to their hydrophilic germ line counterparts resulted in an ?10-fold decrease in turbidity. We also used somatic variants of 10E8, identified previously by next-generation sequencing, to optimize heavy and light chains; this process yielded several improved variants. Of these, variant 10E8v4 with 26 changes versus the parent 10E8 was the most soluble, with a paratope we showed crystallographically to be virtually identical to that of 10E8, a potency on a panel of 200 HIV-1 isolates also similar to that of 10E8, and a half-life in rhesus macaques of ?10 days. An anomaly in 10E8v4 size exclusion chromatography that appeared to be related to conformational isomerization was resolved by engineering an interchain disulfide. Thus, by combining a structure-based approach with natural variation in potency and solubility from the 10E8 lineage, we successfully created variants of 10E8 which retained the potency and extraordinary neutralization breadth of the parent 10E8 but with substantially increased solubility. IMPORTANCE:Antibody 10E8 could be used to prevent HIV-1 infection, if manufactured and delivered economically. It suffers, however, from issues of solubility, which impede manufacturing. We hypothesized that the physical characteristic of 10E8 could be improved through rational design, without compromising breadth and potency. We used structural biology to identify hydrophobic patches on 10E8, which did not appear to be involved in 10E8 function. Reversion of hydrophobic residues in these patches to their hydrophilic germ line counterparts increased solubility. Next, clues from somatic variants of 10E8, identified by next-generation sequencing, were incorporated. A combination of structure-based design and somatic variant optimization led to 10E8v4, with substantially improved solubility and similar potency compared to the parent 10E8. The cocrystal structure of antibody 10E8v4 with its HIV-1 epitope was highly similar to that with the parent 10E8, despite 26 alterations in sequence and substantially improved solubility. Antibody 10E8v4 may be suitable for manufacturing.

Dataset Information

Developmental Pathway of the MPER-Directed HIV-1-Neutralizing Antibody 10E8.

Publications

Developmental Pathway of the MPER-Directed HIV-1-Neutralizing Antibody 10E8.

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