Structure-based Design of Cyclically Permuted HIV-1 gp120 Trimers That Elicit Neutralizing Antibodies

Abstract

A major goal for HIV-1 vaccine development is an ability to elicit strong and durable broadly neutralizing antibody (bNAb) responses. The trimeric envelope glycoprotein (Env) spikes on HIV-1 are known to contain multiple epitopes that are susceptible to bNAbs isolated from infected individuals. Nonetheless, all trimeric and monomeric Env immunogens designed to date have failed to elicit such antibodies. We report the structure-guided design of HIV-1 cyclically permuted gp120 that forms homogeneous, stable trimers, and displays enhanced binding to multiple bNAbs, including VRC01, VRC03, VRC-PG04, PGT128, and the quaternary epitope-specific bNAbs PGT145 and PGDM1400. Constructs that were cyclically permuted in the V1 loop region and contained an N-terminal trimerization domain to stabilize V1V2-mediated quaternary interactions, showed the highest homogeneity and the best antigenic characteristics. In guinea pigs, a DNA prime-protein boost regimen with these new gp120 trimer immunogens elicited potent neutralizing antibody responses against highly sensitive Tier 1A isolates and weaker neutralizing antibody responses with an average titer of about 115 against a panel of heterologous Tier 2 isolates. A modest fraction of the Tier 2 virus neutralizing activity appeared to target the CD4 binding site on gp120. These results suggest that cyclically permuted HIV-1 gp120 trimers represent a viable platform in which further modifications may be made to eventually achieve protective bNAb responses.The HIV-1 Env displayed on the virion surface mediates entry of the virion into target CD4 ϩ T cells to establish infection. Due to its surface accessibility, Env is the predominant target for neutralizing antibody responses (1, 2). Env is synthesized as a gp160 precursor protein, which is further cleaved into surface proximal gp120 and membrane anchored gp41 subunits. A functional HIV-1 Env spike consists of a trimer of gp120 and gp41 heterodimers. The base of the trimer is proximal to the viral membrane (3) and is held together by trimerization motifs within the N-heptad repeat of gp41 (4 -6). The trimer apex is stabilized by interactions between the V1V2 loops of adjacent gp120 monomers (3). Binding of Env to the CD4 receptor on T cells causes extensive conformational changes in the Env trimer and leads to the formation of an open CD4-bound conformation in which the cryptic, co-receptor binding site becomes accessible. This facilitates interaction of the co-receptor (CCR5 or CXCR4) with Env, further driving the fusion of viral and host cell membranes (3, 7).In natural HIV-1 infection, most of the B cell response is elicited against the Env protein, due to its location on the virion surface and relatively high immunogenicity. The antibody response induced during natural HIV-1 infection is predominantly non-neutralizing and strain specific, due to the shed gp120 (immunodominant decoys) and various other immune evasive mechanisms (8). However, ϳ20% of HIV-1-infected patients develop bNAbs during the course of 1-3 years of infectio...

Bacterially expressed HIV-1 gp120 outer-domain fragment immunogens with improved stability and affinity for CD4-binding site neutralizing antibodies

“…By comparing all the groups boosted with monomeric gp120 protein, the OD priming immunogens can be arranged in increasing order of their ability to induce anti-gp120 titers (week 22) as follows: ⌬BS-OD EC Ͻ OD EC Ͻ OD EC C1 Ͻ OD EC Consensus Ͻ OD EC C2 Ͻ OD EC CycV4. Similarly, a comparison of week 22 anti-gp120 titers for the three groups primed with ⌬BS-OD EC but boosted with different Env derivatives indicated that the V1cycP gp120 previously designed by us (30,35,36) provides the best boost for inducing high anti-gp120 titers. In summary, the ELISA results indicated that the OD EC CycV4 and OD EC C2 are the best primes for eliciting sera with high anti-gp120 titers.…”

“…V1cycP gp120 molecule is a trimeric cyclic permutant of gp120, made by introducing new N and C termini at the V1 loop (at amino acids 144 -142) while connecting the original N and C termini with a short linker. A human cartilage matrix protein trimerization domain was also added at the newly created N and C termini to generate this cyclically permuted trimeric gp120 variant (human cartilage matrix protein(144 -142) V1cyc-JRCSF gp120), hereafter referred to as V1cycP gp120 (30,35). All four rabbits in group 1 showed undetectable gp120-specific titers after two primes with OD EC protein (at weeks 0 and 4).…”

A Trimeric HIV-1 Envelope gp120 Immunogen Induces Potent and Broad Anti-V1V2 Loop Antibodies against HIV-1 in Rabbits and Rhesus Macaques

“…We recently described the design and immunogenicity of a novel trimeric gp120 immunogen in guinea pigs ( 19 ). This immunogen, hCMP-v1-cyc-gp120 (referred to as cycP-gp120 here), is based on a cyclically permuted gp120 in which a de novo N terminus is generated within the V1 loop region with the native N and C termini joined via an amino acid linker chain ( 19 ).…”

“…We recently described the design and immunogenicity of a novel trimeric gp120 immunogen in guinea pigs ( 19 ). This immunogen, hCMP-v1-cyc-gp120 (referred to as cycP-gp120 here), is based on a cyclically permuted gp120 in which a de novo N terminus is generated within the V1 loop region with the native N and C termini joined via an amino acid linker chain ( 19 ). To induce a trimeric complex, the human matrix cartilage protein (hCMP) coiled-coil trimerization domain was fused to the de novo N termini, resulting in a stabilized, disulfide bond-linked trimeric gp120 ( 20 ).…”

A facile method of mapping HIV-1 neutralizing epitopes using chemically masked cysteines and deep sequencing

“…The NAB059 elite neutralizer exhibited extraordinarily high titers against a panel of Tier-3 pseudoviruses (26,27), which are the hardest to neutralize. Epitope mapping was also carried out using sera from guinea pigs immunized with an HIV-1 candidate vaccine, a trimeric cyclic permutant (CycP) of gp120 (28) which elicited Tier-2 neutralization activity against a global panel of HIV-1 isolates (29).…”