Peptide mimetics of immunoglobulin A (IgA) and FcαRI block IgA-induced human neutrophil activation and migration

The cross-linking of the IgA Fc receptor (FcαRI) by IgA induces release of the chemoattractant LTB4, thereby recruiting neutrophils in a positive feedback loop. IgA autoantibodies of patients with autoimmune blistering skin diseases therefore induce massive recruitment of neutrophils, resulting in severe tissue damage. To interfere with neutrophil mobilization and reduce disease morbidity, we developed a panel of specific peptides mimicking either IgA or FcαRI sequences. CLIPS technology was used to stabilize three-dimensional structures and to increase peptides’ half-life. IgA and FcαRI peptides reduced phagocytosis of IgA-coated beads, as well as IgA-induced ROS production and neutrophil migration in in vitro and ex vivo (human skin) experiments. Since topical application would be the preferential route of administration, Cetomacrogol cream containing an IgA CLIPS peptide was developed. In the presence of a skin permeation enhancer, peptides in this cream were shown to penetrate the skin, while not diffusing systemically. Finally, epitope mapping was used to discover sequences important for binding between IgA and FcαRI. In conclusion, a cream containing IgA or FcαRI peptide mimetics, which block IgA-induced neutrophil activation and migration in the skin may have therapeutic potential for patients with IgA-mediated blistering skin diseases

The principal neutralization determinant of simian immunodeficiency virus differs from that of human immunodeficiency virus type 1.

To identify the principal neutralization determinant (PND) of simian immunodeficiency virus (SIV), antisera were generated using recombinant gp110 [the SIV analog of the human immunodeficiency virus type 1 (HIV-1) external envelope glycoprotein, gp120], gp140, several large recombinant and proteolytic envelope fragments, and synthetic peptides of the SIVmac251 isolate. When purified under conditions that retain its native structure, gp110 bound CD4 and elicited antisera that neutralized SIVmac251 with high titer. Native gp110 also completely inhibited neutralizing antibody in sera from SIVmac251-infected macaques. In contrast, denatured gp110 and gp140, large envelope fragments, and synthetic peptides (including peptides analogous to the HIV-1 PND) elicited very low or undetectable neutralizing antibody titers and did not inhibit neutralizing antibody in infected macaque sera. Enzymatically deglycosylated gp110 efficiently absorbed neutralizing antibodies from macaque sera, showing that neutralizing antibodies primarily bind the protein backbone. A 45-kDa protease digest product, mapping to the carboxyl-terminal third of gp110, also completely absorbed neutralizing antibodies from infected macaque sera. These results show that the PND(s) of this SIV isolate depends on the native conformation and that linear peptides corresponding to the V3 loop of SIV envelope, in contrast to that of HIV-1, do not elicit neutralizing antibody. This may affect the usefulness of SIVmac for evaluating HIV-1 envelope vaccine approaches that rely on eliciting neutralizing antibody

The Principal Neutralization Determinant of Simian Immunodeficiency Virus Differs from that of Human Immunodeficiency Virus Type 1

To identify the principal neutralization determinant (PND) of simian immunodeficiency virus (SIV), antisera were generated using recombinant gp110 [the SIV analog of the human immunodeficiency virus type 1 (HIV-1) external envelope glycoprotein, gp120], gp140, several large recombinant and proteolytic envelope fragments, and synthetic peptides of the SIVmac251 isolate. When purified under conditions that retain its native structure, gp110 bound CD4 and elicited antisera that neutralized SIVmac251 with high titer. Native gp110 also completely inhibited neutralizing antibody in sera from SIVmac251-infected macaques. In contrast, denatured gp110 and gp140, large envelope fragments, and synthetic peptides (including peptides analogous to the HIV-1 PND) elicited very low or undetectable neutralizing antibody titers and did not inhibit neutralizing antibody in infected macaque sera. Enzymatically deglycosylated gp110 efficiently absorbed neutralizing antibodies from macaque sera, showing that neutralizing antibodies primarily bind the protein backbone. A 45-kDa protease digest product, mapping to the carboxyl-terminal third of gp110, also completely absorbed neutralizing antibodies from infected macaque sera. These results show that the PND(s) of this SIV isolate depends on the native conformation and that linear peptides corresponding to the V3 loop of SIV envelope, in contrast to that of HIV-1, do not elicit neutralizing antibody. This may affect the usefulness of SIVmac for evaluating HIV-1 envelope vaccine approaches that rely on eliciting neutralizing antibody

A Universal Approach to Optimize the Folding and Stability of Prefusion-Closed HIV-1 Envelope Trimers

Summary: The heavily glycosylated native-like envelope (Env) trimer of HIV-1 is expected to have low immunogenicity, whereas misfolded forms are often highly immunogenic. High-quality correctly folded Envs may therefore be critical for developing a vaccine that induces broadly neutralizing antibodies. Moreover, the high variability of Env may require immunizations with multiple Envs. Here, we report a universal strategy that provides for correctly folded Env trimers of high quality and yield through a repair-and-stabilize approach. In the repair stage, we utilized a consensus strategy that substituted rare strain-specific residues with more prevalent ones. The stabilization stage involved structure-based design and experimental assessment confirmed by crystallographic feedback. Regions important for the refolding of Env were targeted for stabilization. Notably, the α9-helix and an intersubunit β sheet proved to be critical for trimer stability. Our approach provides a means to produce prefusion-closed Env trimers from diverse HIV-1 strains, a substantial advance for vaccine development. : Rutten et al. describe a universal repair and stabilize approach that corrects rare mutations and stabilizes refolding regions to obtain high-quality HIV Envs with high yields. The crystal structure shows how the optimization of the trimer interface between α9, α6, and the intersubunit β-sheet stabilizes the membrane-proximal base. Keywords: envelope protein, chronic, ConC_base, HIV, SOSIP, stabilization, transmitted/founder, vaccine, X-ray structure, hybrid shee

Heterologous Stacking of Prion Protein Peptides Reveals Structural Details of Fibrils and Facilitates Complete Inhibition of Fibril GrowthS⃞

Fibrils play an important role in the pathogenesis of amyloidosis; however, the underlying mechanisms of the growth process and the structural details of fibrils are poorly understood. Crucial in the fibril formation of prion proteins is the stacking of PrP monomers. We previously proposed that the structure of the prion protein fibril may be similar as a parallel left-handed β-helix. The β-helix is composed of spiraling rungs of parallel β-strands, and in the PrP model residues 105–143 of each PrP monomer can contribute two β-helical rungs to the growing fibril. Here we report data to support this model. We show that two cyclized human PrP peptides corresponding to residues 105–124 and 125–143, based on two single rungs of the left-handed β-helical core of the human PrPSc fibril, show spontaneous cooperative fibril growth in vitro by heterologous stacking. Because the structural model must have predictive value, peptides were designed based on the structure rules of the left-handed β-helical fold that could stack with prion protein peptides to stimulate or to block fibril growth. The stimulator peptide was designed as an optimal left-handed β-helical fold that can serve as a template for fibril growth initiation. The inhibiting peptide was designed to bind to the exposed rung but frustrate the propagation of the fibril growth. The single inhibitory peptide hardly shows inhibition, but the combination of the inhibitory with the stimulatory peptide showed complete inhibition of the fibril growth of peptide huPrP-(106–126). Moreover, the unique strategy based on stimulatory and inhibitory peptides seems a powerful new approach to study amyloidogenic fibril structures in general and could prove useful for the development of therapeutics

A neutralizing antibody selected from plasma cells that binds to group 1 and group 2 influenza A hemagglutinins

The isolation of broadly neutralizing antibodies against influenza A viruses has been a long-sought goal for therapeutic approaches and vaccine design. Using a single-cell culture method for screening large numbers of human plasma cells, we isolated a neutralizing monoclonal antibody that recognized the hemagglutinin (HA) glycoprotein of all 16 subtypes and neutralized both group 1 and group 2 influenza A viruses. Passive transfer of this antibody conferred protection to mice and ferrets. Complexes with HAs from the group 1 H1 and the group 2 H3 subtypes analyzed by x-ray crystallography showed that the antibody bound to a conserved epitope in the F subdomain. This antibody may be used for passive protection and to inform vaccine design because of its broad specificity and neutralization potency

Ad26.COV2.S protects Syrian hamsters against G614 spike variant SARS-CoV-2 and does not enhance respiratory disease

Previously we have shown that a single dose of recombinant adenovirus serotype 26 (Ad26) vaccine expressing a prefusion stabilized SARS-CoV-2 spike antigen (Ad26.COV2.S) is immunogenic and provides protection in Syrian hamster and non-human primate SARS-CoV-2 infection models. Here, we investigated the immunogenicity, protective efficacy, and potential for vaccine-associated enhanced respiratory disease (VAERD) mediated by Ad26.COV2.S in a moderate disease Syrian hamster challenge model, using the currently most prevalent G614 spike SARS-CoV-2 variant. Vaccine doses of 1 × 109 and 1 × 1010 VP elicited substantial neutralizing antibodies titers and completely protected over 80% of SARS-CoV-2 inoculated Syrian hamsters from lung infection and pneumonia but not upper respiratory tract infection. A second vaccine dose further increased neutralizing antibody titers that was associated with decreased infectious viral load in the upper respiratory tract after SARS-CoV-2 challenge. Suboptimal non-protective immune responses elicited by low-dose A26.COV2.S vaccination did not exacerbate respiratory disease in SARS-CoV-2-inoculated Syrian hamsters with breakthrough infection. In addition, dosing down the vaccine allowed to establish that binding and neutralizing antibody titers correlate with lower respiratory tract protection probability. Overall, these preclinical data confirm efficacy of a one-dose vaccine regimen with Ad26.COV2.S in this G614 spike SARS-CoV-2 virus variant Syrian hamster model, show the added benefit of a second vaccine dose, and demonstrate that there are no signs of VAERD under conditions of suboptimal immunity