Heterologous Epitope-Scaffold Prime∶Boosting Immuno-Focuses B Cell Responses to the HIV-1 gp41 2F5 Neutralization Determinant

Abstract

The HIV-1 envelope glycoproteins (Env) gp120 and gp41 mediate entry and are the targets for neutralizing antibodies. Within gp41, a continuous epitope defined by the broadly neutralizing antibody 2F5, is one of the few conserved sites accessible to antibodies on the functional HIV Env spike. Recently, as an initial attempt at structure-guided design, we transplanted the 2F5 epitope onto several non-HIV acceptor scaffold proteins that we termed epitope scaffolds (ES). As immunogens, these ES proteins elicited antibodies with exquisite binding specificity matching that of the 2F5 antibody. These novel 2F5 epitope scaffolds presented us with the opportunity to test heterologous prime∶boost immunization strategies to selectively boost antibody responses against the engrafted gp41 2F5 epitope. Such strategies might be employed to target conserved but poorly immunogenic sites on the HIV-1 Env, and, more generally, other structurally defined pathogen targets. Here, we assessed ES prime∶boosting by measuring epitope specific serum antibody titers by ELISA and B cell responses by ELISpot analysis using both free 2F5 peptide and an unrelated ES protein as probes. We found that the heterologous ES prime∶boosting immunization regimen elicits cross-reactive humoral responses to the structurally constrained 2F5 epitope target, and that incorporating a promiscuous T cell helper epitope in the immunogens resulted in higher antibody titers against the 2F5 graft, but did not result in virus neutralization. Interestingly, two epitope scaffolds (ES1 and ES2), which did not elicit a detectable 2F5 epitope-specific response on their own, boosted such responses when primed with the ES5. Together, these results indicate that heterologous ES prime∶boost immunization regimens effectively focus the humoral immune response on the structurally defined and immunogen-conserved HIV-1 2F5 epitope.

Structure and Immunogenicity of a Peptide Vaccine, Including the Complete HIV-1 gp41 2F5 Epitope

“…Recently reported compelling mutagenesis of the CDR-H3 loop by Güenaga and Wyatt (25) sequence structurally constrained into a protein scaffold (30). Moreover, L669A, W670A, N671A, W672A, and F673A substitutions, in residues immediately C-terminal to the core epitope, resulted in an affinity decrease.…”

“…2F5 was isolated in mAb form by Katinger and co-workers (21,22) from a panel of sera from naturally infected asymptomatic individuals. Given the neutralization breath and potency shown by the bNAb 2F5 (13,21,(23)(24)(25)(26), development of peptide-based vaccines targeting the 2F5 epitope has since been pursued (6,22,(27)(28)(29)(30)(31)(32)(33).…”

Structure of an HIV-1–neutralizing antibody target, the lipid-bound gp41 envelope membrane proximal region trimer

“…This observation has motivated efforts to develop vaccines designed to induce antibodies specific to this region. Vaccine candidates based on linear peptides from the MPER (19), trimeric gp41 constructs (20,21), and conformationally constrained peptides have been previously reported (22,23). In animal models, many of these vaccine designs have elicited antibodies that recognize epitopes in the MPER (19,22,23).…”

“…Vaccine candidates based on linear peptides from the MPER (19), trimeric gp41 constructs (20,21), and conformationally constrained peptides have been previously reported (22,23). In animal models, many of these vaccine designs have elicited antibodies that recognize epitopes in the MPER (19,22,23). However, none of the induced plasma antibodies strongly neutralize HIV-1 (19,20,23,24), either because the trial vaccines do not present the epitope residues in a native conformation or in the presence of the correct molecular environment, or because of the limitation of induction of MPER antibodies by host tolerance mechanisms (25)(26)(27)(28).…”

Immunisation with foamy virus Bet fusion proteins as novel strategy for HIV-1 epitope delivery

“…Additionally, some Bet hybrids contained the FPPR derived E1 domain and the E2 domain linked by a loop to stabilise a MPER conformation that increases 2F5 binding [56,53,57,54]. Introducing the MPER domain alone resulted in high affinity recognition by 2F5, with values comparable to or better than previously reported for MPER peptides or MPER epitope scaffolds [68,47,69]. Unexpectedly, the affinity of 2F5 to its epitope decreased to some extent when the FPPR residues were present ( Table 1).…”