Abstract
Background The membrane proximal external region (MPER) of HIV-1 is an important target of broadly cross-reactive mAbs. CAPRISA participant, CAP206, developed antiMPER antibodies early that became cross-neutralizing at 18 months post-infection. This coincided with neutralization of the C4 HIV-2/HIV-1 chimera containing the W670 residue, suggesting changes in an antibody paratope may have resulted in the acquisition of breadth. A neutralizing mAb (CAP206-CH12) was isolated, providing an opportunity to determine which somatic hypermutations contribute to breadth. Methods The putative CAP206-CH12 reverted unmutated ancestor (RUA) was inferred using SoDA (http://www.dulci.org). Batch transient transfections were used to generate recombinant antibodies. Neutralization breadth and potency was tested against autologous, Tier 2, and HIV-2/HIV-1 MPER chimeric viruses using the TZM-bl assay. Binding to MPER peptides was assessed by ELISA and surface plasmon resonance (SPR). Results CAP206-CH12_RUA bound the MPER.03 peptide with a Kd of 120nM, 15-fold weaker than CAP206-CH12 binding, but had no neutralizing activity. Since CAP206CH12 and its RUA differed by 19 residues in the heavy chain and 9 in the light chain, we designed an intermediate precursor (IP) where changes near the CDRs in CAP206-CH12 were reverted back to the germline sequence (11 in the heavy and 5 in the light chain). This CAP206-CH12_IP did not neutralize the C4 chimera suggesting that changes responsible for the affinitymatured CAP206-CH12 neutralizing capacity were among these 16 residues. Chimeric pairs of the light chain IP with the heavy chain of CAP206-CH12 or CAP206-CH12_RUA showed binding to MPER with differences in binding kinetics. Conclusion The reduced binding and neutralizing activity of CAP206CH12_RUA and CAP206-CH12_IP compared to CAP206CH12 suggests a correlation between affinity maturation, neutralization breadth and potency. Ongoing work will assess the affinity maturation of CAP206-CH12 by determining moieties in the antibody paratope associated with effective epitope recognition and the effects of somatic mutations on the evolution of neutralization.
Highlights
The membrane proximal external region (MPER) of HIV-1 is an important target of broadly cross-reactive mAbs
CAPRISA participant, CAP206, developed antiMPER antibodies early that became cross-neutralizing at 18 months post-infection
Since CAP206CH12 and its reverted unmutated ancestor (RUA) differed by 19 residues in the heavy chain and 9 in the light chain, we designed an intermediate precursor (IP) where changes near the CDRs in CAP206-CH12 were reverted back to the germline sequence (11 in the heavy and 5 in the light chain)
Summary
The membrane proximal external region (MPER) of HIV-1 is an important target of broadly cross-reactive mAbs. CAPRISA participant, CAP206, developed antiMPER antibodies early that became cross-neutralizing at 18 months post-infection. CAPRISA participant, CAP206, developed antiMPER antibodies early that became cross-neutralizing at 18 months post-infection This coincided with neutralization of the C4 HIV-2/HIV-1 chimera containing the W670 residue, suggesting changes in an antibody paratope may have resulted in the acquisition of breadth. A neutralizing mAb (CAP206-CH12) was isolated, providing an opportunity to determine which somatic hypermutations contribute to breadth
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