Mitigation of Equine Recurrent Uveitis Through Topical Suppressor of Cytokine Signaling-1 Mimetic Peptide: Open Label Safety and Efficacy Pilot Study

Equine recurrent uveitis (ERU) is a painful and debilitating autoimmune disease, and represents the only spontaneous model of human recurrent uveitis (RU). Despite the efficacy of existing treatments, RU remains a leading cause of visual handicap in horses and humans. Cytokines, which utilize Janus kinase 2 (Jak2) for signaling, drive the inflammatory processes in ERU that promote blindness. Notably, suppressor of signaling-1 (SOCS1), which naturally limits the activation of Jak2 through binding interactions, is often deficient in autoimmune disease patients. Significantly, we previously showed that topical administration of a SOCS1 peptide mimic (SOCS1-KIR) mitigated induced rodent uveitis. In this pilot study, we test the potential to translate the therapeutic efficacy observed in experimental rodent uveitis to equine patient disease. Through bioinformatics and peptide binding assays we demonstrate putative binding of the SOCS1-KIR peptide to equine Jak2. We also show that topical, or intravitreal injection of SOCS1-KIR was well tolerated within the equine eye through physical and ophthalmic examinations. Finally, we show that topical SOCS1-KIR administration was associated with significant clinical ERU improvement. Together, these results provide a scientific rationale, and supporting experimental evidence for the therapeutic use of a SOCS1 mimetic peptide in RU.

Anti-inflammatory effects of DhHP-6 in LPS-induced RAW264.7 macrophages and carrageenan-induced air pouch model in rat

DhHP-6 (Deuterohemin-Ala-His-Thr-Val-Glu-Lys) is a novel peptide mimic of peroxidases that was previously designed in our laboratory. Here, we explored the anti-inflammatory potential of DhHP-6 against lipopolysaccharide(LPS)stimulated inflammatory response in RAW264.7 cells and carrageenan-induced air pouch model rats. DhHP-6 treatment dramatically attenuated the production of nitric oxide (NO), IL-6, andTNF-α in LPS induced RAW264.7 cells. Also, it blocked phosphorylation and degradation of IκBα and suppressed the nuclear translocation of p65. DhHP-6 (0.2, 0.6, and 2.0 mg/kg) significantly reduced the levels of total proteins and WBC counts in the exudates of the air pouch model rats. Moreover, MDA contents in the plasma of rats were reduced and SOD activities were enhanced in the DhHP-6-treatment group. Our results strongly show the effectiveness of DhHP-6 as an anti-inflammatory agent. The mechanism could be related to the reduction of Reactive oxygen species(ROS), inhibition of NF-κB nuclear translocation, and reduction of pro-inflammatory cytokines.

Rapid characterization of spike variants via mammalian cell surface display

The SARS-CoV-2 spike (S) protein is a critical component of subunit vaccines and a target for neutralizing antibodies. Spike is also undergoing immunogenic selection with clinical variants that increase infectivity and partially escape convalescent plasma. Here, we describe spike display, a high-throughput platform to rapidly characterize glycosylated spike ectodomains across multiple coronavirus-family proteins. We assayed ~200 variant SARS-CoV-2 spikes for their expression, ACE2 binding, and recognition by thirteen neutralizing antibodies (nAbs). An alanine scan of the N-terminal domain (NTD) highlights a public class of epitopes in the N3 and N5 loops that are recognized by most of the NTD-binding nAbs assayed in this study. Some clinical NTD substitutions abrogate binding to these epitopes but are circulating at low frequencies around the globe. NTD mutations in variants of concern B.1.1.7 (United Kingdom), B.1.351 (South Africa), B.1.1.248 (Brazil), and B.1.427/B.1.429 (California) impact spike expression and escape most NTD-targeting nAbs. However, two classes of NTD nAbs still bind B.1.1.7 spikes and neutralize in pseudoviral assays. B.1.1351 and B.1.1.248 include compensatory mutations that either increase spike expression or increase ACE2 binding affinity. Finally, B.1.351 and B.1.1.248 completely escape a potent ACE2 peptide mimic. We anticipate that spike display will be useful for rapid antigen design, deep scanning mutagenesis, and epitope mapping of antibody interactions for SARS-CoV-2 and other emerging viral threats.

Phage-Display Based Discovery and Characterization of Peptide Ligands against WDR5

WD40 is a ubiquitous domain presented in at least 361 human proteins and acts as scaffold to form protein complexes. Among them, WDR5 protein is an important mediator in several protein complexes to exert its functions in histone modification and chromatin remodeling. Therefore, it was considered as a promising epigenetic target involving in anti-cancer drug development. In view of the protein–protein interaction nature of WDR5, we initialized a campaign to discover new peptide-mimic inhibitors of WDR5. In current study, we utilized the phage display technique and screened with a disulfide-based cyclic peptide phage library. Five rounds of biopanning were performed and isolated clones were sequenced. By analyzing the sequences, total five peptides were synthesized for binding assay. The four peptides are shown to have the moderate binding affinity. Finally, the detailed binding interactions were revealed by solving a WDR5-peptide cocrystal structure.

A Versatile Surface Bioengineering Strategy Based on Mussel-Inspired and Bioclickable Peptide Mimic

In this work, we present a versatile surface engineering strategy by the combination of mussel adhesive peptide mimicking and bioorthogonal click chemistry. The main idea reflected in this work derived from a novel mussel-inspired peptide mimic with a bioclickable azide group (i.e., DOPA4-azide). Similar to the adhesion mechanism of the mussel foot protein (i.e., covalent/noncovalent comediated surface adhesion), the bioinspired and bioclickable peptide mimic DOPA4-azide enables stable binding on a broad range of materials, such as metallic, inorganic, and organic polymer substrates. In addition to the material universality, the azide residues of DOPA4-azide are also capable of a specific conjugation of dibenzylcyclooctyne- (DBCO-) modified bioactive ligands through bioorthogonal click reaction in a second step. To demonstrate the applicability of this strategy for diversified biofunctionalization, we bioorthogonally conjugated several typical bioactive molecules with DBCO functionalization on different substrates to fabricate functional surfaces which fulfil essential requirements of biomedically used implants. For instance, antibiofouling, antibacterial, and antithrombogenic properties could be easily applied to the relevant biomaterial surfaces, by grafting antifouling polymer, antibacterial peptide, and NO-generating catalyst, respectively. Overall, the novel surface bioengineering strategy has shown broad applicability for both the types of substrate materials and the expected biofunctionalities. Conceivably, the “clean” molecular modification of bioorthogonal chemistry and the universality of mussel-inspired surface adhesion may synergically provide a versatile surface bioengineering strategy for a wide range of biomedical materials.

A Modified ACE2 peptide mimic to block SARS-CoV2 entry

A 23-residue peptide fragment that forms a part of the α-1 helix of the ACE2 peptidase domain, the recognition domain for SARS-CoV2 on the ACE2 receptor, holds the potential as a drug to block the viral receptor binding domain (RBD) from forming a complex with ACE2. The peptide has recently been shown to bind the viral RBD with good efficiency. Here, we present a detailed analysis of the energetics of binding of the peptide to the SARS-CoV2 RBD. We use equilibrium molecular dynamics simulation to study the dynamics of the complex. We perform end-state binding energy calculations to gain a residue-level insight into the binding process and use the information to incorporate point mutations into the peptide. We demonstrate using binding energy calculations that the peptide with certain point mutations, especially E17L, shows a stronger binding to the RBD as compared to the wild type peptide. We propose that the modified peptide will thus be more efficient in blocking RBD-ACE2 binding.