Background:
The membrane proximal external region (MPER) is a conserved site of vulnerability on the HIV-1 gp41 envelope, targeted by highly potent broadly neutralizing monoclonal antibodies (bNAbs) such as 4E10, VRC42 and PGZL1. Although elicited by unrelated persons living with HIV (PLWH), from multiple different HIV-1 subtypes, these bNAbs have similar features. We report the isolation of 4E10-class bNAb, MHRP.01, from an untreated person living with subtype B HIV infection that had 100% neutralization breadth against a 34-pseudovirus (pSV) panel and predicted specificity for MPER binding mAbs. Using bNAbs 4E10, VRC42, and MHRP.01, we provide evidence of an amino acid signature pertaining to 4E10-class bNAbs.
Methods:
CD19+/IgD-/IgM- B cells were sorted from donor PBMCs by flow cytometry and cultured in 384-well plates. Cultured supernatants were screened via microneutralization assays against HIV-1 pSVs. B cell receptors from wells with neutralization >70% were sequenced, followed by cloning and expression of heavy and light chain genes to produce monoclonal antibodies (mAbs). mAbs were characterized for binding, epitope mapping and neutralization assays against panels of diverse HIV-1 strains. Based on crystal diffraction analysis, mutations were introduced in the heavy chain of the MHRP.01 to generate a 4E10-like bNAb signature, and the resulting mutant mAbs were further characterized.
Results:
MHRP.01 neutralized 99.5% of a globally diverse 208-pSV panel with IC50 of 1.49 µg/ml and demonstrated striking similarities to 4E10, VRC42, and PGZL1, including utilization of VH1-69 and VK3-20 gene usage and similar critical residue epitopes. Crystallization of MHRP.01 revealed residues important for MPER binding, which bound in the same conformation as 4E10 and VRC42. Mutation of glycine at 50 or proline at 108 locations within CDRH2 and CDRH3 respectively abolished binding to the MPER peptide and neutralization of autologous pSV (Fig 1), with further paratope residues contributing to a definable sequence signature.
Conclusions:
MPER targeting bNAb, MHRP.01 contained identical germline usage, binding angle of approach and neutralization breadth as 4E10-class bNAbs. Structural analysis reveals a 4E10-like bNAb signature important for MPER binding and neutralization. MPER represents a common epitope which has the potential to elicit similar B cell responses from several PLWH, making it desirable for vaccine design.
