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Abstract Several bacterial strains have developed resistance against commercial antibiotics, and interestingly, supramolecular nanomaterials have shown considerable advantages for antibacterial applications. However, the main challenges in adopting nanotechnology for antibacterial studies are random aggregation, compromised toxicity, multi-step preparation approaches, and unclear structure-function properties. Herein, we designed the amphiphilic tripeptide that acts as a reducing and capping agent for silver metal to form silver-peptide colloidal nanohybrids with the mild assistance of UV light (254 nm) through the photochemical reduction method. The nanohybrids are characterized by different spectroscopic and microscopic techniques, and non-covalent molecular interactions between metal and peptide building blocks confirm their central role in the formation of nanohybrids. The tripeptide is biocompatible and can reduce the toxicity of silver ions (Ag+) by reducing to Ag0. These colloidal nanohybrids showed antibacterial activity against gram-negative and gram-positive bacterial strains, and the possible mechanism of killing bacterial cells could be membrane disruption. This synthetic strategy is facile and green, which helps avoid using toxic chemicals or reagents and complicated methods for colloidal nanohybrid preparation for biomedical applications.

Abstract Canine distemper virus (CDV) affects many domestic and wild animals. Variations among CDV genome linages could lead to vaccination failure. To date, there are several vaccine alternatives, such as a modified live virus and a recombinant vaccine; however, most of these alternatives are based on the ancestral strain Onderstepoort, which has not been circulating for years. Vaccine failures and the need to update vaccines have been widely discussed, and the development of new vaccine candidates is necessary to reduce circulation and mortality. Current vaccination alternatives cannot be used in wildlife animals due to the lack of safety data for most of the species, in addition to the insufficient immune response against circulating strains worldwide in domestic species. Computational tools, including peptide-based therapies, have become essential for developing new-generation vaccines for diverse models. In this work, a peptide-based vaccine candidate with a peptide library derived from CDV H and F protein consensus sequences was constructed employing computational tools. The molecular docking and dynamics of the selected peptides with canine MHC-I and MHC-II and with TLR-2 and TLR-4 were evaluated. In silico safety was assayed through determination of antigenicity, allergenicity, toxicity potential, and homologous canine peptides. Additionally, in vitro safety was also evaluated through cytotoxicity in cell lines and canine peripheral blood mononuclear cells (cPBMCs) and through a hemolysis potential assay using canine red blood cells. A multiepitope CDV polypeptide was constructed, synthetized, and evaluated in silico and in vitro by employing the most promising peptides for comparison with single CDV immunogenic peptides. Our findings suggest that predicting immunogenic CDV peptides derived from most antigenic CDV proteins could aid in the development of new vaccine candidates, such as multiple single CDV peptides and multiepitope CDV polypeptides, that are safe in vitro and optimized in silico. In vivo studies are being conducted to validate potential vaccines that may be effective in preventing CDV infection in domestic and wild animals.

Abstract 2K4L is a rationally designed analog of the short α-helical peptide temporin-1CEc, a natural peptide isolated and purified from the skin secretions of the Chinese brown frog Rana chensinensis by substituting amino acid residues. 2K4L displayed improved and broad-spectrum antibacterial activity than temporin-1CEc in vitro. Here, the antibacterial and anti-inflammatory activities of 2K4L in macrophages, C. elegans and mice were investigated. The results demonstrated that 2K4L could enter THP-1 cells to kill a multidrug-resistant Acinetobacter baumannii strain (MRAB 0227) and a sensitive A. baumannii strain (AB 22933), as well as reduce proinflammatory responses induced by MRAB 0227 by inhibiting NF-κB signaling pathway. Similarly, 2K4L exhibited strong bactericidal activity against A. baumannii uptake into C. elegans, extending the lifespan and healthspan of the nematodes. Meanwhile, 2K4L alleviated the oxidative stress response by inhibiting the expression of core genes in the p38 MAPK/PMK-1 signaling pathway and downregulating the phosphorylation level of p38, thereby protecting the nematodes from damage by A. baumannii. Finally, in an LPS-induced septic model, 2K4L enhanced the survival of septic mice and decreased the production of proinflammatory cytokines by inhibiting the signaling protein expression of the MAPK and NF-κB signaling pathways and protecting LPS-induced septic mice from a lethal inflammatory response. In conclusion, 2K4L ameliorated LPS-induced inflammation both in vitro and in vivo.

Abstract The inhibition of tumor necrosis factor (TNF)-α trimer formation renders it inactive for binding to its receptors, thus mitigating the vicious cycle of inflammation. We designed a peptide (PIYLGGVFQ) that simulates a sequence strand of human TNFα monomer using a series of in silico methods, such as active site finding (Acsite), protein–protein interaction (PPI), docking studies (GOLD and Flex-X) followed by molecular dynamics (MD) simulation studies. The MD studies confirmed the intermolecular interaction of the peptide with the TNFα. Fluorescence-activated cell sorting and fluorescence microscopy revealed that the peptide effectively inhibited the binding of TNF to the cell surface receptors. The cell culture assays showed that the peptide significantly inhibited the TNFα-mediated cell death. In addition, the nuclear translocation of the nuclear factor kappa B (NFκB) was significantly suppressed in the peptide-treated A549 cells, as observed in immunofluorescence and gel mobility-shift assays. Furthermore, the peptide protected against joint damage in the collagen-induced arthritis (CIA) mouse model, as revealed in the micro focal-CT scans. In conclusion, this TNFα antagonist would be helpful for the prevention and repair of inflammatory bone destruction and subsequent loss in the mouse model of CIA as well as human rheumatoid arthritis (RA) patients. This calls upon further clinical investigation to utilize its potential effect as an antiarthritic drug.

Abstract Sepsis represents a critical medical condition stemming from an imbalanced host immune response to infections, which is linked to a significant burden of disease. Despite substantial efforts in laboratory and clinical research, sepsis remains a prominent contributor to mortality worldwide. Nanotechnology presents innovative opportunities for the advancement of sepsis diagnosis and treatment. Due to their unique properties, including diversity, ease of synthesis, biocompatibility, high specificity, and excellent pharmacological efficacy, peptides hold great potential as part of nanotechnology approaches against sepsis. Herein, we present a comprehensive and up-to-date review of the applications of peptides in nanosystems for combating sepsis, with the potential to expedite diagnosis and enhance management outcomes. Firstly, sepsis pathophysiology, antisepsis drug targets, current modalities in management and diagnosis with their limitations, and the potential of peptides to advance the diagnosis and management of sepsis have been adequately addressed. The applications have been organized into diagnostic or managing applications, with the last one being further sub-organized into nano-delivered bioactive peptides with antimicrobial or anti-inflammatory activity, peptides as targeting moieties on the surface of nanosystems against sepsis, and peptides as nanocarriers for antisepsis agents. The studies have been grouped thematically and discussed, emphasizing the constructed nanosystem, physicochemical properties, and peptide-imparted enhancement in diagnostic and therapeutic efficacy. The strengths, limitations, and research gaps in each section have been elaborated. Finally, current challenges and potential future paths to enhance the use of peptides in nanosystems for combating sepsis have been deliberately spotlighted. This review reaffirms peptides' potential as promising biomaterials within nanotechnology strategies aimed at improving sepsis diagnosis and management. Graphical Abstract

This study explores the potential of natural bioactive peptides from animal venoms as targeted anti-cancer agents with reduced toxicity. Initially, we screened a broad collection of animal venoms for their antiproliferative activity against cancer cell lines. From this collection, we selected venoms from Naja anchietae and Naja senegalensis due to their promising activity. Utilizing reverse- phase high-performance liquid chromatography (RP HPLC), mass spectrometry (MALDI-TOF MS and MALDI-TOF TOF MSMS), and Edman degradation sequencing, we isolated and characterized three peptides named CTNanc1, CTNanc2, and CTNanc3 from Naja anchietae, and three others named CTNsen1, CTNsen2, and CTNsen3 from Naja senegalensis, each with a molecular weight of around 7 kDa. These purified peptides demonstrated inhibition of U87 glioblastoma cell proliferation, but not of U251 and T98G cells, in cell viability assays. To assess the impact of these treatments on cell viability, apoptosis, and necrosis, flow cytometry assays were conducted on U87 cells at 72 h. The results showed a decrease in cell viability and an increase in dead cells, suggesting that the treatments not only promote apoptosis, but may also lead to increased necrosis or late-stage apoptosis as the exposure time increases. These findings suggest that these peptides could be developed as leads for cancer therapy.

Parkinson’s disease (PD) is the second most common neurodegenerative disorder globally. Recognizing the potential of velvet antler in the nervous system, as shown in numerous studies, this research was aimed at evaluating the neuroprotective effects of Sika Deer velvet antler peptide (VAP), along with the underlying mechanisms in neurotoxin-induced PD models. Initially, a peptidomic analysis of the VAP, which comprised 189 varieties of peptides, was conducted using LC-MS. Nine sequences were identified as significant using Proteome Discoverer 2.5 software. In a cellular model of PD, where PC12 cells are treated with the neurotoxin 1-methyl-4-phenylpyridinium (MPP+), the administration of the VAP reduced the cell damage and apoptosis induced by MPP+. This protective effect was associated with a decrease in oxidative stress. This protective mechanism was found to be mediated through the activation of the SIRT1-dependent Akt/Nrf2/HO-1-signaling pathway. In animal models, specifically in mice with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced PD, the administration of the VAP effectively reduced the dopaminergic neuron damage and reversed the neurobehavioral deficits. They also diminished microglia activation and apoptosis, all without any noticeable adverse effects. Additionally, the VAP was observed to beneficially alter the gut microbiota, as marked by an increase in the abundances of Prevotellaceae, Helicobacteraceae, and Prevotella. These findings suggest that VAP exerts its neuroprotective effect against neurodegeneration by inhibiting oxidative stress and modulating gut microbiota.

The discovery of new peptides and their derivatives is an outcome of ongoing efforts to identify a peptide with significant biological activity for effective usage as a possible therapeutic agent. Spinorphin peptides have been documented to exhibit numerous applications and features. In this study, biologically active peptide derivatives based on novel peptide analogues of spinorphin conjugated with 5,5′-dimethyl (Dm) and 5,5′-diphenyl (Ph) hydantoin derivatives have been successfully synthesized and characterized. Scanning electron microscopy (SEM) and spectral methods such as UV-Vis, FT-IR (Fourier Transform Infrared Spectroscopy), CD (Circular Dichroism), and fluorimetry were used to characterize the microstructure of the resulting compounds. The results revealed changes in peptide morphology as a result of the restructuring of the aminoacidic sequences and aromatic bonds, which is related to the formation of intermolecular hydrogen bonds between tyrosyl groups and the hydantoin moiety. Electrochemical and fluorescence approaches were used to determine some physicochemical parameters related to the biological behavior of the compounds. The biological properties of the spinorphin derivatives were evaluated in vivo for anticonvulsant activity against the psychomotor seizures at different doses of the studied peptides. Both spinorphin analog peptides with Ph and Dm groups showed activity against all three phases of the seizure in the intravenous Pentylenetetrazole Seizure (ivPTZ) test. This suggests that hydantoin residues do not play a crucial role in the structure of spinorphin compounds and in determining the potency to raise the seizure threshold. On the other hand, analogs with a phenytoin residue are active against the drug-resistant epilepsy test (6-Hz test). In addition, bioactivity analyses revealed that the new peptide analogues have the potential to be used as antimicrobial and antioxidant compounds. These findings suggest promising avenues for further research that may lead to the development of alternative medicines or applications in various fields beyond epilepsy treatment.

Chronic subdural hemorrhage (CSDH) refers to a hematoma with an envelope between the dura mater and the arachnoid membrane and is more common among the elderly. It was reported that the dura mater, which is highly vascularized with capillary beds, precapillary arterioles and postcapillary venules play an important role in the protection of the central nervous system (CNS). Numerous evidences suggests that peptides play an important role in neuroprotection of CNS. However, whether dura mater derived endogenous peptides participate in the pathogenesis of CSDH remains undetermined. In the current study, the peptidomic profiles were performed in human dura of CSDH (three patients) and the relative control group (three non-CSDH samples) by LC-MS (liquid chromatography–mass spectrometry). The results suggested that a total of 569 peptides were differentially expressed in the dura matter of CSDH compared with relative controls, including 217 up-regulated peptides and 352 down-regulated peptides. Gene Ontology (GO) analysis demonstrated that the precursor proteins of those differentially expressed peptides were involved in the various biological processes. Interestingly, Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis suggested that NETs participated in the pathogenies of CSDH. Further investigate showed that H3Cit was significantly elevated in the dural and hematoma membranes of patients with CSDH compared to patients without CSDH. Taken together, our results showed the differentially expressed peptides in human dura mater of CSDH and demonstrated that NETs formation in the dural and hematoma membranes might be involved in the pathogenesis of CSDH. It is worth noting that pharmacological inhibition of NETs may have potential therapeutic implications for CSDH.

The T cell receptor (TCR) is a complex molecular machine that directs the activation of T cells, allowing the immune system to fight pathogens and cancer cells. Despite decades of investigation, the molecular mechanism of TCR activation is still controversial. One of the leading activation hypotheses is the allosteric model. This model posits that binding of pMHC at the extracellular domain triggers a dynamic change in the transmembrane (TM) domain of the TCR subunits, which leads to signaling at the cytoplasmic side. We sought to test this hypothesis by creating a TM ligand for TCR. Previously we described a method to create a soluble peptide capable of inserting into membranes and binding to the TM domain of the receptor tyrosine kinase EphA2 (Alves et al., eLife, 2018). Here, we show that the approach is generalizable to complex membrane receptors, by designing a TM ligand for TCR. We observed that the designed peptide caused a reduction of Lck phosphorylation of TCR at the CD3ζ subunit in T cells. As a result, in the presence of this peptide inhibitor of TCR (PITCR), the proximal signaling cascade downstream of TCR activation was significantly dampened. Co-localization and co-immunoprecipitation in diisobutylene maleic acid (DIBMA) native nanodiscs confirmed that PITCR was able to bind to the TCR. AlphaFold-Multimer predicted that PITCR binds to the TM region of TCR, where it interacts with the two CD3ζ subunits. Our results additionally indicate that PITCR disrupts the allosteric changes in the compactness of the TM bundle that occur upon TCR activation, lending support to the allosteric TCR activation model. The TCR inhibition achieved by PITCR might be useful to treat inflammatory and autoimmune diseases and to prevent organ transplant rejection, as in these conditions aberrant activation of TCR contributes to disease.

Condensed coacervate phases are now understood to be important features of modern cell biology, as well as valuable protocellular models in origin-of-life studies and synthetic biology. In each of these fields, the development of model systems with varied and tuneable material properties is of great importance for replicating properties of life. Here, we develop a ligase ribozyme system capable of concatenating short RNA fragments into long chains. Our results show that the formation of coacervate microdroplets with the ligase ribozyme and poly(L-lysine) enhances ribozyme rate and yield, which in turn increases the length of the anionic polymer component of the system and imparts specific physical properties to the droplets. Droplets containing active ribozyme sequences resist growth, do not wet or spread on unpassivated surfaces, and exhibit reduced transfer of RNA between droplets when compared to controls containing inactive sequences. These altered behaviours, which stem from RNA sequence and catalytic activity, constitute a specific phenotype and potential fitness advantage, opening the door to selection and evolution experiments based on a genotype–phenotype linkage.

µ-Conotoxins are small, potent pore-blocker inhibitors of voltage-gated sodium (NaV) channels, which have been identified as pharmacological probes and putative leads for analgesic development. A limiting factor in their therapeutic development has been their promiscuity for different NaV channel subtypes, which can lead to undesirable side-effects. This review will focus on four areas of µ-conotoxin research: (1) mapping the interactions of µ-conotoxins with different NaV channel subtypes, (2) µ-conotoxin structure–activity relationship studies, (3) observed species selectivity of µ-conotoxins and (4) the effects of µ-conotoxin disulfide connectivity on activity. Our aim is to provide a clear overview of the current status of µ-conotoxin research.

The subcutaneous administration of therapeutic peptides would provide significant benefits to patients. However, subcutaneous injections are limited in dosing volume, potentially resulting in high peptide concentrations that can incur significant challenges with solubility limitations, high viscosity, and stability liabilities. Herein, we report on the discovery that low-shear resonant acoustic mixing can be used as a general method to prepare stable nanoparticles of a number of peptides of diverse molecular weights and structures in water without the need for extensive amounts of organic solvents or lipid excipients. This approach avoids the stability issues observed with typical high-shear, high-intensity milling methods. The resultant peptide nanosuspensions exhibit low viscosity even at high concentrations of >100 mg/mL while remaining chemically and physically stable. An example nanosuspension of cyclosporine nanoparticles was dosed in rats via a subcutaneous injection and exhibited sustained release behavior. This suggests that peptide nanosuspension formulations can be one approach to overcome the challenges with high-concentration peptide formulations.

When the skin is damaged, accelerating the repair of skin trauma and promoting the recovery of tissue function are crucial considerations in clinical treatment. Previously, we isolated and identified an active peptide (livisin) from the skin secretion of the frog Odorrana livida. Livisin exhibited strong protease inhibitory activity, water solubility, and stability, yet its wound-healing properties have not yet been studied. In this study, we assessed the impact of livisin on wound healing and investigated the underlying mechanism contributing to its effect. Our findings revealed livisin effectively stimulated the migration of keratinocytes, with the underlying mechanisms involved the activation of CaSR as a peptide calcium mimetic. This activation resulted in the stimulation of the CaSR/E-cadherin/EGFR/ERK signaling pathways. Moreover, the therapeutic effects of livisin were partially reduced by blocking the CaSR/E-cadherin/EGFR/ERK signaling pathway. The interaction between livisin and CaSR was further investigated by molecular docking. Additionally, studies using a mouse full-thickness wound model demonstrated livisin could accelerate skin wound healing by promoting re-epithelialization and collagen deposition. In conclusion, our study provides experimental evidence supporting the use of livisin in skin wound healing, highlighting its potential as an effective therapeutic option.

We previously reported a protective antibody response in mice immunized with synthetic microparticle vaccines made using layer-by-layer fabrication (LbL-MP) and containing the conserved T1BT* epitopes from the P. falciparum circumsporozoite protein. To further optimize the vaccine candidate, a benchtop tangential flow filtration method (LbL-by-TFF) was developed and utilized to produce vaccine candidates that differed in the status of base layer crosslinking, inclusion of a TLR2 ligand in the antigenic peptide, and substitution of serine or alanine for an unpaired cysteine residue in the T* epitope. Studies in mice revealed consistent superiority of the Pam3Cys-modified candidates and a modest benefit of base layer crosslinking, as evidenced by higher and more persistent antibody titers (up to 18 months post-immunization), a qualitative improvement of T-cell responses toward a Th1 phenotype, and greater protection from live parasite challenges compared to the unmodified prototype candidate. Immunogenicity was also tested in a non-human primate model, the rhesus macaque. Base layer-crosslinked LbL-MP loaded with T1BT* peptide with or without covalently linked Pam3Cys elicited T1B-specific antibody responses and T1BT*-specific T-cell responses dominated by IFNγ secretion with lower levels of IL-5 secretion. The Pam3Cys-modified construct was more potent, generating antibody responses that neutralized wild-type P. falciparum in an in vitro hepatocyte invasion assay. IgG purified from individual macaques immunized with Pam3Cys.T1BT* LbL-MP protected naïve mice from challenges with transgenic P. berghei sporozoites that expressed the full-length PfCS protein, with 50–88% of passively immunized mice parasite-free for ≥15 days. Substitution of serine for an unpaired cysteine in the T* region of the T1BT* subunit did not adversely impact immune potency in the mouse while simplifying the manufacture of the antigenic peptide. In a Good Laboratory Practices compliant rabbit toxicology study, the base layer-crosslinked, Pam3Cys-modified, serine-substituted candidate was shown to be safe and immunogenic, eliciting parasite-neutralizing antibody responses and establishing the dose/route/regimen for a clinical evaluation of this novel synthetic microparticle pre-erythrocytic malaria vaccine candidate.

Osteoarthritis (OA) is the most common joint disorder, with a social and financial burden that is expected to increase in the coming years. Currently, there are no effective medications to treat it. Due to limited treatment options, patients often resort to supplements, such as collagen hydrolysates (CHs). CHs are products with low molecular weight (MW) peptides, often between 3 and 6 kDa, and are a result of industrialized processed collagen. Collagen extraction is often a by-product of the meat industry, with the main source for collagen-based products being bovine, although it can also be obtained from porcine and piscine sources. CHs have demonstrated positive results in clinical trials related to joint health, such as decreased joint pain, increased mobility, and structural joint improvements. The bioactivity of CHs is primarily attributed to their bioactive peptide (BAP) content. However, there are significant knowledge gaps regarding the digestion, bioavailability, and bioactivity of CH-derived BAPs, and how different CH products compare in that regard. The present review discusses CHs and their BAP content as potential treatments for OA.

Since the first news about the detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) appeared, a large amount of data on the variability of the virus genome have accumulated. Most of the mutations entrenched in the viral genome are aimed at improving the mechanisms of host cell penetration, altering the degree of binding to protein receptors, evading the immune system, and suppressing the antiviral immune response. Knowledge of the functional role of mutations will allow improving diagnostic methods, treatment, and vaccine prophylaxis regimens, as well as predicting the further spread and evolution of the virus. In this study, we analyzed a number of SARS-CoV-2 virus mutations in the context of viral epitope affinity for most common in Russia HLA class I and class II alleles (according to Allele Frequency Net Database). This study examined clade-forming mutations of viral clades that are classified as variants of concern according to the WHO classification. We found that some mutations reduce the number of predicted epitopes, the number of HLA alleles that bind them, or the number of both epitopes and HLA alleles simultaneously. Mutations of the viral clade B.1.1.7 (S:Y144del, S:H69-V70del, and S:A570D), mutations of the viral clade B.1.617.2 (S:T19R, S:G12D, S:F157del, and S:R158del), and mutation N:R203K related to all clades except B.1.617.2 have pronounced effects on reducing epitope affinity for HLA alleles, with some of them affecting on epitopes of all the studied strong and weak binders lengths in both HLA class I and class II.

Summary: Background: Successful embryo implantation requires the attachment of a blastocyst to the receptive endometrial epithelium, which was disturbed in the women with recurrent implantation failure (RIF). Endometrial β3-integrin was the most important adhesion molecule contributing to endometrial receptivity in both humans and mice. Nur77 has been proven indispensable for fertility in mice, here we explore the role of Nur77 on embryo-epithelial adhesion and potential treatment to embryo implantation failure. Methods: The expression and location of Mst1 and Nur77 in endometrium from fertile women and RIF patients were examined by IHC, qRT-PCR and Western blotting. In vitro kinase assay following with LC-MS/MS were used to identify the phosphorylation site of Nur77 activated by Mst1. The phosphorylated Nur77 was detected by phos-tag SDS-PAGE assay and specific antibody against phospho-Nur77-Thr366. The effect of embryo-epithelium interaction was determined in the BeWo spheroid or mouse embryo adhesion assay, and delayed implantation mouse model. RNA-seq was used to explore the mechanism by which Nur77 derived peptide promotes endometrial receptivity. Findings: Endometrial Mammalian sterile 20 (STE20)-like kinase 1 (Mst1) expression level was decreased in the women with RIF than that in the fertile control group, while Mst1 activation in the epithelial cells promoted trophoblast–uterine epithelium adhesion. The effect of Nur77 mediated trophoblast–uterine epithelium adhesion was facilitated by active Mst1. Mechanistically, mst1 promotes the transcription activity of Nur77 by phosphorylating Nur77 at threonine 366 (T366), and consequently increased downstream target β3-integrin expression. Furthermore, a Nur77-derived peptide containing phosphorylated T366 markedly promoted mouse embryo attachment to Ishikawa cells ([4 (2–4)] vs [3 (2–4)]) and increased the embryo implantation rate (4 vs 1.4) in a delayed implantation mouse model by regulating integrin signalling. Finally, it is observed that the endometrial phospho-Nur77 (T366) level is decreased by 80% in the women with RIF. Interpretation: In addition to uncovering a potential regulatory mechanism of Mst1/Nur77/β3-integrin signal axis involved in the regulation of embryo-epithelium interaction, our finding provides a novel marker of endometrial receptivity and a potential therapeutic agent for embryo implantation failure. Funding: National Key Research and Development Program of China (2018YFC1004400), the National Natural Science Foundation of China (82171653, 82271698, 82030040, 81971387 and 30900727), and National Institutes of Health grants (R01HL103869).

Peptidases generate bioactive peptides that can regulate cell signaling and mediate intercellular communication. While the processing of peptide precursors is initiated intracellularly, some modifications by peptidases may be conducted extracellularly. Thimet oligopeptidase (TOP) is a peptidase that processes neuroendocrine peptides with roles in mood, metabolism, and immune responses, among other functions. TOP also hydrolyzes angiotensin I to angiotensin 1–7, which may be involved in the pathophysiology of COVID-19 infection. Although TOP is primarily cytosolic, it can also be associated with the cell plasma membrane or secreted to the extracellular space. Recent work indicates that membrane-associated TOP can be released with extracellular vesicles (EVs) to the extracellular space. Here we briefly summarize the enzyme’s classical function in extracellular processing of neuroendocrine peptides, as well as its more recently understood role in intracellular processing of various peptides that impact human diseases. Finally, we discuss new findings of EV-associated TOP in the extracellular space.

Naturally produced peptides (

The highly toxic plant toxin ricin is one of the most known threatening toxins. Accurate and sensitive biosensing methods for the first emergency response and intoxication treatment, are always pursued in the biodefense field. Screening affinity molecules is the fundamental mainstream approach for developing biosensing methods. Compared with common affinity molecules such as antibodies and oligonucleotide aptamers, peptides have great potential as biosensing modules with more accessible chemical synthesis capability and better batch-to-batch stability than antibodies, more abundant interaction sites, and robust sensing performance towards complex environments. However, anti-ricin peptides are so scant to be screened and discovered, and an advanced screening strategy is the utmost to tackle this issue. Here, we present a new in silico-in vitro iteration-assisted affinity maturation strategy of anti-ricin peptides. We first obtained affinity peptides targeting ricin through phage display with five panning rounds of “coating-elution-amplification-enrichment” procedures. The binding affinity and kinetic parameters characterized by surface plasmon resonance (SPR) showed that we had obtained four peptides owning dissociation constants (KD) around 2~35 μM, in which peptide PD-2-R5 has the lower KD of 4.7 μM and higher stable posture to interact with ricin. We then constructed a new strategy for affinity maturity, composing two rounds of in silico-in vitro iterations. Firstly, towards the single-site alanine scanning mutation peptide library, the molecular docking predictions match the SPR evaluation results well, laying a solid foundation for designing a full saturation mutated peptide library. Secondly, plenty of in silico saturation mutation prediction results guided the discovery of peptides PD2-R5-T3 and PD-2-R5-T4 with higher affinity from only a limited number of SPR evaluation experiments. Both evolved peptides had increased affinity by about 5~20 times, i.e., KD of 230 nM and 900 nM. A primary cellular toxicity assay indicated that both peptides could protect cells against ricin damage. We further established an SPR assay based on PD-2-R5-T3 and PD-2-R5-T4 elongated with an antifouling peptide linkage and achieved good linearity with a sensitivity of 1 nM and 0.5 nM, respectively. We hope this new affinity-mature strategy will find its favorable position in relevant peptide evolution, biosensing, and medical countermeasures for biotoxins to protect society’s security and human life better.

Vascular endothelial growth factor (VEGF) and its receptors (VEGFRs) play a main role in the regulation of angiogenesis and lymphangiogenesis. Furthermore, they are implicated in the onset of several diseases such as rheumatoid arthritis, degenerative eye conditions, tumor growth, ulcers and ischemia. Therefore, molecules able to target the VEGF and its receptors are of great pharmaceutical interest. Several types of molecules have been reported so far. In this review, we focus on the structure-based design of peptides mimicking VEGF/VEGFR binding epitopes. The binding interface of the complex has been dissected and the different regions challenged for peptide design. All these trials furnished a better understanding of the molecular recognition process and provide us with a wealth of molecules that could be optimized to be exploited for pharmaceutical applications.

Cancer neoantigens that arise from somatic mutations have emerged as important targets for personalized immunization. Here, we report an improved overall survival of a HER2-positive metastatic breast cancer patient using a bioinformatic-based personalized peptide immunization called BITAP (BioInformatic Tumor Address Peptides). The epitopes were predicted using our in-house bioinformatic pipeline, and the immunogenicity was tested by IFN-γ ELISPOT and intracellular cytokine staining assays. In total, a significant peptide-specific T-cell response was detected against 18 out of the 76 (≈24%) tested peptides. The patient’s follow-up by measuring serologic markers showed a significant reduction in the tumor marker levels following BITAP immunization. Along with standard treatment, the patient treated with the BITAP showed stable disease with a remarkably improved overall survival, and no serious treatment-related adverse effects. In conclusion, our findings suggest that BITAP immunization is feasible, and safe, and may induce tumor regressions in patients with HER2-positive subsets of breast cancer.

Nanotechnology is a new, interdisciplinary field of science, the products of which are nanoparticles, i.e. structures that, thanks to their unique physical and chemical properties conditioned by their nanometric dimensions, have found wide application in medicine, pharmacy and biotechnology. Currently, they play a particularly important role as drug carriers, antibacterial and antiviral substances. They are also used in diagnostics, genetic tests, and increasing the accuracy of biological tests. The enormous possibilities of nanoparticles concern their potential use as drug delivery systems, factors improving the quality of anticancer radiotherapy and supporting molecular imaging, as well as compounds with highly bactericidal, fungicidal and antiviral properties. Therefore, it is crucial to implement therapies based on nanoparticles into clinical trials, as they can be an excellent diagnostic and therapeutic tool for many diseases, including cancer. However, it is necessary to overcome certain barriers due to the nature of some nanoparticles, such as problems with biodegradability and porosity. Despite the benefits of using nanostructures, the possibility of side effects caused by the action of metal nanoparticles absorbed by organisms has not been ruled out.

Objective: To test cell penetrating and interfering peptide Mut3DPT-PP2A/SET in interaction between serine threonine phosphatase PP2A and its physiological inhibitor, the oncoprotein SET. Materials and methods: Adult male C3H/S-strain mice, 60 days old, were given a graft of breast adenocarcinoma cells (TN60) into subcutaneous tissue. Mut3DPT-PP2A/SET peptide was used to block PP2A and SET oncoprotein interaction. The graft-bearing animals were divided into a control group (injected with saline buffer), and an intervention group injected intraperitoneally with Mut3DPT-PP2A/SET peptide (5 mg/kg) every day from day 5 to day 37. The variables we used to compare the outcome in both groups were tumor size in mm (length×width) and histological changes. In the statistical analysis we used ANOVA and Student-Keuls multiple comparisons test and Tuckey for the post-test analysis. Results: 48 mice were grafted at day 0 with breast UNLP-C3H/S tumor cells, and after randomization, they were assigned to one of the two study groups. At day 5 all mice were injected either with placebo or with the peptide. The treated group showed significant tumor reduction (p

Breast cancer is a tumor entity that is one of the leading causes of mortality among women worldwide. Although numerous treatment options are available, current explorations of personalized vaccines have shown potential as promising new treatment options to prevent the recurrence of cancer. Here we present a small proof of concept study using a prophylactic peptide vaccination approach in four female breast cancer patients who achieved remission after standard treatment. The patients were initially analyzed for somatic tumor mutations and then treated with personalized neoantigen-derived peptide vaccines. These vaccines consisted of HLA class I and class II peptides and were administered intracutaneously followed by subcutaneous application of sargramostim and/or topical imiquimod as an immunological adjuvant. After an initial priming phase of four vaccinations within two weeks, patients received monthly boosting/maintenance vaccinations. Chemotherapy or checkpoint inhibition was not performed during vaccination. One patient received hormone therapy. The vaccines were well tolerated with no serious adverse events. All patients displayed vaccine-induced CD4+ and/or CD8+ T-cell responses against various neoantigens. Furthermore, all patients remained tumor-free and had persistent T-cell responses, even several months after the last vaccination, suggesting the potential of peptide vaccines as an immunosurveillance and long term prophylaxis option.

It is known that schistosome-derived antigens induce innate and adaptive immune responses that are essential for the formation of hepatic immunopathology. Here, we screened and synthesized four peptides derived from Schistosoma japonicum (S. japonicum) heat shock protein 90α (Sjp90α-1, -2, -3, and -4), which is widely expressed in adults and eggs of the genus S. japonicum and induces remarkable immune reactions. To define the antigenicity of these peptides, we stimulated splenocytes with peptides, and the results showed that only the Sjp90α-1 peptide could predominately induce the activation of dendritic cells (DCs) and macrophages as well as alter the proportion of follicular helper T (Tfh) cells. Next, CD4+ T cells were purified and cocultured with mouse bone-marrow-derived DCs (BMDCs) with or without Sjp90α-1 peptide stimulation in vitro, and the results showed that Sjp90α-1-stimulated BMDCs can significantly induce CD4+ T-cell differentiation into Tfh cells, while the direct stimulation of CD4+ T cells with Sjp90α-1 did not induce Tfh cells, indicating that the Sjp90α-1 peptide promotes Tfh cell differentiation depending on the presence of DCs. Furthermore, we selected and prepared an Sjp90α-1-peptide-based antibody and illustrated that it has excellent reactivity with the immunizing peptide and detects a single band of 29 kDa corresponding to the Sjp90α protein. The immunolocalization results showed that the protein recognized by this Sjp90α-1-peptide-based antibody is present in the mature eggs and the tegument of adults, implying that the parasite-derived peptide has a potential interaction with the host immune system. Finally, we evaluated antipeptide IgG antibodies and revealed a significantly higher level of anti-Sjp90α-1 peptide IgG antibodies in mice 3 weeks after S. japonicum infection. In conclusion, we illustrate that these synthetic peptides warrant further investigation by evaluating their antigen-specific immune response and their ability to efficiently induce Tfh cells. Moreover, they may constitute a potentially helpful method for the laboratory diagnosis of schistosomiasis japonica.

Willardiine is a nonprotein amino acid containing uracil, and thus classified as nucleobase amino acid or nucleoamino acid, that together with isowillardiine forms the family of uracilylalanines isolated more than six decades ago in higher plants. Willardiine acts as a partial agonist of ionotropic glutamate receptors and more in particular it agonizes the non-N-methyl-D-aspartate (non-NMDA) receptors of L-glutamate: ie. the α-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA) and kainate receptors. Several analogues and derivatives of willardiine have been synthesised in the laboratory in the last decades and these compounds show different binding affinities for the non-NMDA receptors. More in detail, the willardiine analogues have been employed not only in the investigation of the structure of AMPA and kainate receptors, but also to evaluate the effects of receptor activation in the various brain regions. Remarkably, there are a number of neurological diseases determined by alterations in glutamate signaling, and thus, ligands for AMPA and kainate receptors deserve attention as potential neurodrugs. In fact, similar to willardiine its analogues often act as agonists of AMPA and kainate receptors. A particular importance should be recognized to willardiine and its thymine-based analogue AlaT also in the peptide chemistry field. In fact, besides the naturally-occurring short nucleopeptides isolated from plant sources, there are different examples in which this class of nucleoamino acids was investigated for nucleopeptide development. The applications are various ranging from the realization of nucleopeptide/DNA chimeras for diagnostic applications, and nucleoamino acid derivatization of proteins for facilitating protein-nucleic acid interaction, to nucleopeptide-nucleopeptide molecular recognition for nanotechnological applications. All the above aspects on both chemistry and biotechnological applications of willardine/willardine-analogues and nucleopeptide will be reviewed in this work.

Acid-sensing ion channels (ASICs) are voltage-independent H+-gated cation channels largely expressed in the nervous system of rodents and humans. At least six isoforms (ASIC1a, 1b, 2a, 2b, 3 and 4) associate into homotrimers or heterotrimers to form functional channels with highly pH-dependent gating properties. This review provides an update on the pharmacological profiles of animal peptide toxins targeting ASICs, including PcTx1 from tarantula and related spider toxins, APETx2 and APETx-like peptides from sea anemone, and mambalgin from snake, as well as the dimeric protein snake toxin MitTx that have all been instrumental to understanding the structure and the pH-dependent gating of rodent and human cloned ASICs and to study the physiological and pathological roles of native ASICs in vitro and in vivo. ASICs are expressed all along the pain pathways and the pharmacological data clearly support a role for these channels in pain. ASIC-targeting peptide toxins interfere with ASIC gating by complex and pH-dependent mechanisms sometimes leading to opposite effects. However, these dual pH-dependent effects of ASIC-inhibiting toxins (PcTx1, mambalgin and APETx2) are fully compatible with, and even support, their analgesic effects in vivo, both in the central and the peripheral nervous system, as well as potential effects in humans.

Abstract Objective Coronary artery disease contributes to noncommunicable disease deaths worldwide. In order to make preventive methods more accurate, we need to know more about the development and progress of this pathology, including the genetic aspects. Humanin is a small peptide known for its cytoprotective and anti-apoptotic properties. Our study looked for genomic associations between humanin-like nuclear isoform genes and coronary artery disease using CARDIoGRAMplusC4D Consortium data. Results Lookup from meta-analysis datasets gave single nucleotide polymorphisms in all 13 humanin-like nuclear isoform genes with the lowest P value for rs6151662 from the MTRNR2L2 gene including the 50 kb flanking region in both directions (P-value = 0.0037). Within the gene region alone the top variant was rs78083998 from the MTRNR2L13 region (meta-analysis P-value = 0.042). None of the found associations were statistically significant after correction for multiple testing. Lookup for expression trait loci in these gene regions gave no statistically significant variants.

Over the last two decades, due to the advances in the techniques used in the design and synthesis of peptides as well as the understanding of the underlying molecular mechanisms involved in their therapeutic action on skin disorders, novel applications have been emerged for peptides as cosmeceuticals. There are several advantages in the use of bioactive and biomimetic peptides such as availability, diverse and selective mode of action in skin physiology, and lack of immunogenic reactions that have attracted skin specialists’ attention for their potential use as anti-wrinkle agents. These peptides are known to have various biological roles against skin ageing, most prominently as extracellular matrix signal modulators and collagen synthesis stimulants. In addition, they may act as carrier peptides, antioxidants, neurotransmitter or enzyme inhibitors. However, they are facing a few obstacles for widespread applications in topical anti-ageing skin products: a) clinical evidence for the therapeutic efficacy is still limited; b) skin penetration may be weak due to their hydrophilicity, high molar mass, and binding to other ingredients; and c) high cost. Peptide transport through skin can be effectively boosted by penetration enhancers, lipophilic modifications, peptide microencapsulation, and skin microporation. There are currently more than 30 therapeutic peptides that have been occasionally introduced into the market of skin care products. This review summarizes different classes of topical peptides with proven effectiveness against skin ageing in-vivo. Moreover, the challenges of and approaches to peptide transport through skin OR (the delivery challenges and approaches) are discussed.

Abstract Background Angiogenesis inhibition research is a cutting edge area in angiogenesis-dependent disease therapy, especially in cancer therapy. Recently, studies on anti-angiogenic peptides have provided promising results in the field of cancer treatment. Methods A non-redundant dataset of 135 anti-angiogenic peptides (positive instances) and 135 non anti-angiogenic peptides (negative instances) was used in this study. Also, 20% of each class were selected to construct an independent test dataset (see Additional files 1, 2). We proposed an effective machine learning based R package (AntAngioCOOL) to predict anti-angiogenic peptides. We have examined more than 200 different classifiers to build an efficient predictor. Also, more than 17,000 features were extracted to encode the peptides. Results Finally, more than 2000 informative features were selected to train the classifiers for detecting anti-angiogenic peptides. AntAngioCOOL includes three different models that can be selected by the user for different purposes; it is the most sensitive, most specific and most accurate. According to the obtained results AntAngioCOOL can effectively suggest anti-angiogenic peptides; this tool achieved sensitivity of 88%, specificity of 77% and accuracy of 75% on the independent test set. AntAngioCOOL can be accessed at https://cran.r-project.org/. Conclusions Only 2% of the extracted descriptors were used to build the predictor models. The results revealed that physico-chemical profile is the most important feature type in predicting anti-angiogenic peptides. Also, atomic profile and PseAAC are the other important features.

Dengue is a viral mosquito-borne disease that rapidly spreads in tropical and subtropical countries, including the Philippines. One of its most distinguishing characteristics is the ability of the Dengue Virus (DENV) to easily surpass the innate responses of the body, thus activating B cells of the adaptive immunity to produce virus-specific antibodies. Moreover, Dengvaxia® is the only licensed vaccine for DENV, but recent studies showed that seronegative individuals become prone to increased disease severity and hospitalization. Owing to this limitation of the dengue vaccine, this study determined and compared consensus and unique B cell epitopes among each DENV (1–4) Philippine isolate to identify potential areas of interest for future vaccine studies and therapeutic developments. An in silico-based epitope prediction of forty (40) DENV 1–4 strains, each serotype represented by ten (10) sequences from The National Center for Biotechnology Information (NCBI), was conducted using Kolaskar and Tongaonkar antigenicity, Emini surface accessibility, and Parker hydrophilicity prediction in Immune Epitope Database (IEDB). Results showed that five (5) epitopes were consensus for DENV-1 with no detected unique epitope, one (1) consensus epitope for DENV-2 with two (2) unique epitopes, one (1) consensus epitope for DENV-3 plus two (2) unique epitopes, and two (2) consensus epitopes and one (1) unique epitope for DENV-4. The findings of this study would contribute to determining potential vaccine and diagnostic marker candidates for further research studies and immunological applications against DENV (1–4) Philippine isolates.

In pursuit of therapeutics for human polyomaviruses, we identified a peptide derived from the BK polyomavirus (BKV) minor structural proteins VP2/3 that is a potent inhibitor of BKV infection with no observable cellular toxicity. The thirteen-residue peptide binds to major structural protein VP1 with single-digit nanomolar affinity. Alanine-scanning of the peptide identified three key residues, substitution of each of which results in ~1000 fold loss of binding affinity with a concomitant reduction in antiviral activity. Structural studies demonstrate specific binding of the peptide to the pore of pentameric VP1. Cell-based assays demonstrate nanomolar inhibition (EC50) of BKV infection and suggest that the peptide acts early in the viral entry pathway. Homologous peptide exhibits similar binding to JC polyomavirus VP1 and inhibits infection with similar potency to BKV in a model cell line. Lastly, these studies validate targeting the VP1 pore as a novel strategy for the development of anti-polyomavirus agents.

Endotoxemia induces lung injury. We assessed the therapeutic efficacy between triple cytokine (tumor necrosis factor-α [TNF-α], interleukin-1β [IL-1β], and IL-6) inhibition (mediated by KCF18 peptide) and single cytokine (TNF-α) inhibition (mediated by SEM18 peptide) on alleviating lung injury in the early phase of endotoxemia. Mice receiving endotoxin (Endo group), endotoxin plus KCF18 (EKCF group), or endotoxin plus SEM18 (ESEM) were monitored and euthanized at 24 h after endotoxin. Our data demonstrated altered lung function (decreases in tidal volume, minute ventilation, and dynamic compliance; and by contrast, increases in airway resistance and end expiration work) and histology (increases in injury scores, leukocyte infiltration, vascular permeability, and tissue water content) in the Endo group with significant protection observed in the EKCF and ESEM groups (all p < 0.05). Levels of inflammation (macrophage activation and cytokine upregulations), oxidation (lipid peroxidation), necroptosis, pyroptosis, and apoptosis in EKCF and ESEM groups were comparable and all were significantly lower than in the Endo group (all p < 0.05). These data demonstrate that single cytokine TNF-α inhibition can achieve therapeutic effects similar to triple cytokines TNF-α, IL-1β, and IL-6 inhibition on alleviating endotoxin-induced lung injury, indicating that TNF-α is the major cytokine in mediating lung injury in the early phase of endotoxemia.

Methods used to control the quality of peptide products depend on the level of development of analytical and bioorganic chemistry, and the level of instrumentation. Peptide identification is a difficult task and largely depends on the complexity of its structure. There does not exist a comprehensive and simple test, except for NMR, which, however, is rather expensive and time-consuming and involves complex data interpretations. Moreover, it does not allow for unambiguous determination of the peptide purity and formula (amino acid composition, sequence, chirality of amino acid residues). For this reason, a combination of methods is often used, including amino acid analysis, TLC/HPLC and mass spectrometry, and, less frequently, sequencing. Current international practice of peptide analysis is to use HPLC in combination with mass spectrometric, mainly tandem (HPLC-MS/MS), detection. According to literature sources the amino acid sequence of linear peptides can be analysed using various enzymes and subsequent identification of proteolysis products by mass spectrometry. This article presents approaches to the development of test methods for analysis of purity and identification testing of a small interfering RNA-based novel medicinal product, which will help standardise and control the quality of the production process.

Background: Due to the increasing in antibiotic resistance in bacteria, especially the infections which their treatment is very difficult, there is a need for producing new drugs. The aim of this study was identification and purification of quorum sensing peptides causing apoptosis in Staphylococcus aureus as new treatment antibiotics. Methods: The supernatant from Staphylococcus aureus, Enterococcus faecalis, and Enterococcus faecium were collected after centrifugation. Then, the supernatant was isolated from the specimens that had the greatest effect on the growth of bacteria. Liquid chromatography was used to purify it. In next step, for detection of protein concentration, Bradford test and for confirmation, two dimensional electrophoresis were used. Finally, to determine the antimicrobial activity of purified peptides, minimum inhibitory concentration (MIC) and minimum inhibitory concentration (MBC) of peptides were investigated. Findings: The obtained effective ingredient was a polypeptide that was effective against multi-drug resistant bacteria. The results of MIC for methicillin-resistant Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, Salmonella paratyphi A and B, Klebsiella oxytoka, Acinetobacter Bumanni, and Shigella dysenteria were 3.2, 7.0, 5.0, 5.0, 3.0, 6.0, 1.0, and 7.0 µg/ml, respectively. Conclusion: The polypeptide derived from this study showed a vast antimicrobial property against gram-positive and gram-negative bacteria.

Background: Nexvax2® is a novel, peptide-based, epitope-specific immunotherapy intended to be administered by regular injections at dose levels that increase the threshold for clinical reactivity to natural exposure to gluten and ultimately restore tolerance to gluten in patients with celiac disease. Celiac disease patients administered fixed intradermal doses of Nexvax2 become unresponsive to the HLA-DQ2·5-restricted gluten epitopes in Nexvax2, but gastrointestinal symptoms and cytokine release mimicking gluten exposure, that accompany the first dose, limit the maximum tolerated dose to 150 μg. Our aim was to test whether stepwise dose escalation attenuated the first dose effect of Nexvax2 in celiac disease patients. Methods: We conducted a randomized, double-blind, placebo-controlled trial at four community sites in Australia (3) and New Zealand (1) in HLA-DQ2·5 genotype positive adults with celiac disease who were on a gluten-free diet. Participants were assigned to cohort 1 if they were HLA-DQ2·5 homozygotes; other participants were assigned to cohort 2, or to cohort 3 subsequent to completion of cohort 2. Manual central randomization without blocking was used to assign treatment for each cohort. Initially, Nexvax2-treated participants in cohorts 1 and 2 received an intradermal dose of 30 μg (consisting of 10 μg of each constituent peptide), followed by 60 μg, 90 μg, 150 μg, and then eight doses of 300 μg over six weeks, but this was amended to include doses of 3 μg and 9 μg and extended over a total of seven weeks. Nexvax2-treated participants in cohort 3 received doses of 3 μg, 9 μg, 30 μg, 60 μg, 90 μg, 150 μg, 300 μg, 450 μg, 600 μg, 750 μg, and then eight of 900 μg over nine weeks. The dose interval was 3 or 4 days. Participants, care providers, data managers, sponsor personnel, and study site personnel were blinded to treatment assignment. The primary outcome was the number of adverse events and percentage of participants with adverse events during the treatment period. This completed trial is registered with ClinicalTrials.gov, number NCT02528799. Findings: From the 73 participants who we screened from 19 August 2015 to 31 October 2016, 24 did not meet eligibility criteria, and 36 were ultimately randomized and received study drug. For cohort 1, seven participants received Nexvax2 (two with the starting dose of 30 μg and then five at 3 μg) and three received placebo. For cohort 2, 10 participants received Nexvax2 (four with starting dose of 30 μg and then six at 3 μg) and four received placebo. For cohort 3, 10 participants received Nexvax2 and two received placebo. All 36 participants were included in safety and immune analyses, and 33 participants completed treatment and follow-up; in cohort 3, 11 participants were assessed and included in pharmacokinetics and duodenal histology analyses. Whereas the maximum dose of Nexvax2 had previously been limited by adverse events and cytokine release, no such effect was observed when dosing escalated from 3 μg up to 300 μg in HLA-DQ2·5 homozygotes or to 900 μg in HLA-DQ2.5 non-homozygotes. Adverse events with Nexvax2 treatment were less common in cohorts 1 and 2 with the starting dose of 3 μg (72 for 11 participants) than with the starting dose of 30 μg (91 for six participants). Adverse events during the treatment period in placebo-treated participants (46 for nine participants) were similar to those in Nexvax2-treated participants when the starting dose was 3 μg in cohort 1 (16 for five participants), cohort 2 (56 for six participants), and cohort 3 (44 for 10 participants). Two participants in cohort 2 and one in cohort 3 who received Nexvax2 starting at 3 μg did not report any adverse event, while the other 33 participants experienced at least one adverse event. One participant, who was in cohort 1, withdrew from the study due to adverse events, which included abdominal pain graded moderate or severe and associated with nausea after receiving the starting dose of 30 μg and one 60 μg dose. The most common treatment-emergent adverse events in the Nexvax2 participants were headache (52%), diarrhoea (48%), nausea (37%), abdominal pain (26%), and abdominal discomfort (19%). Administration of Nexvax2 at dose levels from 150 μg to 900 μg preceded by dose escalation was not associated with elevations in plasma cytokines at 4 h. Nexvax2 treatment was associated with trends towards improved duodenal histology. Plasma concentrations of Nexvax2 peptides were dose-dependent. Interpretation: We show that antigenic peptides recognized by CD4-positive T cells in an autoimmune disease can be safely administered to patients at high maintenance dose levels without immune activation if preceded by gradual dose escalation. These findings facilitate efficacy studies that test high-dose epitope-specific immunotherapy in celiac disease.

Preferred methods for generating mouse dendritic cells (DC) would encompass qualities of consistency, high yield, and potent function. Serum-free culture is also highly desirable, since this is the standard for cell-based therapies used in humans. We report here a serum-free modification of a culture method generating mature, activated DCs from bone marrow precursors. This is achieved through a two-stage culture comprised of 6-day expansion in Flt3 ligand and IL-6 followed by brief differentiation in a medium containing GM-CSF and IL-4, with subsequent activation using TLR ligands ODN1826 and LPS. The serum-free DCs achieve yields and surface phenotype including IL-12p70 secretion similar to standard serum-replete cultures, display a capacity to sensitize in vivo against both MHC class I- and Class II-restricted antigens, and exhibit some aspects of “killer DC” function against tumor cells. We used these DCs to help identify novel CD4pos Th epitopes on the rat ErbB2/HER-2 protein and demonstrated a subset of these as effective immunogens in a DC-based therapeutic model of HER-2pos breast cancer in Balb/c mice, where they induced powerful Th1-polarized immune responses. This method represents a useful way to efficiently produce large numbers of murine dendritic cells with excellent in vivo function well-suited for use in experimental vaccine studies.

The anti-microbial peptide (AMP) pleurocidin is found in winter flounder (Pseudopleuronectes americanus), an Atlantic flounder species. There is promising evidence for clinical, aquaculture, and veterinary applications of pleurocidin. This review provides an overview of the current literature available on pleurocidin to guide future research directions. By fully elucidating pleurocidin’s mechanism of action and developing novel treatments against pathogenic microbes, populations of flatfish and humans can be protected. This review consulted publications from PubMed and Environment Complete with search terms such as “pleurocidin”, “winter flounder”, and “antimicrobial”. The fish immune system includes AMPs as a component of the innate immune system. Pleurocidin, one of these AMPs, has been found to be effective against various Gram-positive and Gram-negative bacteria. More investigations are required to determine pleurocidin’s suitability as a treatment against antibiotic-resistant pathogens. There is promising evidence for pleurocidin as a novel anti-cancer therapy. The peptide has been found to display potent anti-cancer effects against human cancer cells. Research efforts focused on pleurocidin may result in novel treatment strategies against antibiotic-resistant bacteria and cancer. More research is required to determine if the peptide is a suitable candidate to be developed into a novel anti-microbial treatment. Some of the microbes susceptible to the peptide are also pathogens of fish, suggesting its suitability as a therapeutic treatment for fish species.

Pancreatic cancer has a low survival rate and has limited therapeutic options due to the peculiarity of the tumor tissue. Cancer nanotechnology provides several opportunities to resolve such difficulties as a result of the high surface-to-volume ratio of nanostructures. Peptide-drug nanocomplexes have proved to have immense potential in anticancer activity against pancreatic cancer cells. Thus, in the present study apamin (APA) and alendronate sodium (ALS) were combined to form nanocomplexes (APA-ALS-NC) against pancreatic cancer cells. Optimization of ALS, incubation time, and sonication time in terms of particle size of the nanocomplex was carried out. The optimized formulation was evaluated for anticancer activities in pancreatic cancer cells (PANC-1 cells). A Box-Behnken design using ALS, incubation time, and sonication time as independent factors and particle size as the response was chosen to optimize the APA-ALS-NC formulation. The optimized APA-ALS-NC had a particle size of 161.52 ± 8.4 nm. The evaluation of APA-ALS-NC in PANC-1 cells was carried out using various in vitro tests. The IC50 values were determined by MTT assay and found to be 37.6 ± 1.65, 13.4 ± 0.59, and 1.01 ± 0.04 µg/mL for ALS, APA, and APA-ALS-NC, respectively. The higher cytotoxicity activity of APA-ALS-NC was confirmed from the higher percentage of cells in the necrosis phase (apoptosis study) and the G2-M phase (cell cycle study) compared to that of ALS and APA. While the loss of mitochondrial membrane potential was less for APA-ALS-NC, the levels of IL-1β, TNF-α, caspase-3, ROS, IL-6, and NF-kB showed that APA-ALS-NC can significantly enhance apoptosis and cytotoxicity in PANC-1 cells. Moreover, Bax (10.87 ± 1.36), Bcl-2 (0.27 ± 0.02), and p53 (9.16 ± 1.22) gene expressions confirmed that APA-ALS-NC had a significant apoptotic effect compared to ALS and APA. In summary, the APA-ALS-NC had a more significant cytotoxic effect than ALS and APA. The results of the present study are promising for further evaluation in pre-clinical and clinical trials for arriving at a successful therapeutic strategy against pancreatic cancer.

Cyanobacteria are microorganisms with photosynthetic mechanisms capable of colonizing several distinct environments worldwide. They can produce a vast spectrum of bioactive compounds with different properties, resulting in an improved adaptative capacity. Their richness in secondary metabolites is related to their unique and diverse metabolic apparatus, such as Non-Ribosomal Peptide Synthetases (NRPSs). One important class of peptides produced by the non-ribosomal pathway is anabaenopeptins. These cyclic hexapeptides demonstrated inhibitory activity towards phosphatases and proteases, which could be related to their toxicity and adaptiveness against zooplankters and crustaceans. Thus, this review aims to identify key features related to anabaenopeptins, including the diversity of their structure, occurrence, the biosynthetic steps for their production, ecological roles, and biotechnological applications.

Coronavirus disease (COVID-19) pandemic caused by the coronavirus SARS-CoV-2 represents an enormous challenge to global public health, with thousands of infections and deaths in over 200 countries worldwide. The purpose of this study was to identify SARS-CoV-2 epitopes with potential to interact in silico with the alleles of the human leukocyte antigen class I (HLA I) and class II (HLA II) commonly found in the Colombian population to promote both CD4 and CD8 immune responses against this virus. The generation and evaluation of the peptides in terms of HLA I and HLA II binding, immune response, toxicity and allergenicity were performed by using computer-aided tools, such as NetMHCpan 4.1, NetMHCIIpan 4.0, VaxiJem, ToxinPred and AllerTop. Furthermore, the interaction between the predicted epitopes with HLA I and HLA II proteins frequently found in the Colombian population was studied through molecular docking simulations in AutoDock Vina and interaction analysis in LigPlot+. One of the promising peptides proposed in this study is the HLA I epitope YQPYRVVVL, which displayed an estimated coverage of over 82% and 96% for the Colombian and worldwide population, respectively. These findings could be useful for the design of new epitope-vaccines that include Colombia among their population target.

Through 4 June 2021, COVID-19 has caused over 172.84 million cases of infection and 3.71 million deaths worldwide. Due to its rapid dissemination and high mutation rate, it is essential to develop a vaccine harboring multiple epitopes and efficacious against multiple variants to prevent the immune escape of SARS-CoV-2. An in silico approach based on the viral genome was applied to identify 19 high-immunogenic B-cell epitopes and 499 human leukocyte antigen (HLA)-restricted T-cell epitopes. Thirty multi-epitope peptide vaccines were designed by iNeo-Suite and manufactured by solid-phase synthesis. Docking analysis confirmed stable hydrogen bonds of epitopes with their corresponding HLA alleles. When four peptide candidates derived from the spike protein of SARS-CoV-2 were selected to immunize mice, a significantly larger amount of total IgG in serum, as well as an increase of CD19+ cells in the inguinal lymph nodes, were observed in the peptide-immunized mice compared to the control. The ratios of IFN-γ-secreting lymphocytes in CD4+ or CD8+ T-cells in the peptide-immunized mice were higher than those in the control mice. There were also a larger number of IFN-γ-secreting T-cells in the spleens of peptide-immunized mice. The peptide vaccines in this study successfully elicited antigen-specific humoral and cellular immune responses in mice. To further validate the safety and efficacy of this vaccine, animal studies using a primate model, as well as clinical trials in humans, are required.

Stereo-selectivity is an important feature in the development of the synthesis of biologically active organic compounds. In this process, (bio) catalysts exhibit substrate specificity that allows high levels of chemo- and regio-selectivity. Over the past decade, several peptides have been developed as effective bio-catalysts for a range of synthetically valuable reactions. In comparison with proteins owing a large number of amino acids and high molecular weights, peptide-catalysts possess only a few amino acid residues, which may adopt a secondary structure suitable for synthesis of desired chiral products. In addition, the flexible nature of peptides consents for tuning of reactivity and selectivity by replacing amino acid residues. These unique aspects provide attractive biocatalysts platform for asymmetric syntheses. Highlights • Asymmetric catalysis has an impressive progression in the manufacture of pharmaceuticals. • Enzymes and natural/synthetic peptides are attractive biocatalysts of the chiral reactions. • Peptides show unique features compared with other catalysts in asymmetric catalysis.

Background: Apelin is a potent endogenous inotropic peptide with a major role in counteracting the aldosterone and angiotensin II and their negative effects on the cardiovascular system. The exact role of apelin in the patho-physiology of this disease is not well understood. We aimed to investigate the possible associations of apelin-13 with clinical and paraclinical characteristics in HF patients as well as studying its dynamics during the course of the heart failure.