Cyclic Peptide Research Articles

Covalent Peptide Evolution: Redefining Protein–Protein Interaction Inhibition Through Phage Display

Covalent cyclic peptides represent a transformative approach for targeting challenging protein-protein interactions (PPIs) characterized by flat, extensive binding surfaces. Recent advances in electrophilic phage display now enable the evolution of these peptides through integrating sulfur(VI) fluoride exchange (SuFEx) chemistry with functional selection strategies. This innovative platform combines genetic encoding with site-specific cyclization and warhead incorporation to generate high-affinity, irreversible binders. When targeting the SARS-CoV-2 Spike-ACE2 interface, the approach produced sub-100 nM inhibitors with >10-fold improved potency over non-covalent analogues. The methodology's success against this clinically relevant target underscores its potential to address longstanding challenges in PPI modulation, particularly for high-value targets in oncology and neurodegeneration. By combining covalent engagement with phage display's evolutionary power, this technology establishes a new paradigm for developing mechanistically validated peptide therapeutics against previously intractable interactions. Keywords: covalent peptide inhibitors, cyclic peptide therapeutics, phage display evolution, protein-protein interaction inhibition, SuFEx chemistry, irreversible binders, SARS-CoV-2 inhibitors, Spike-ACE2 disruption, electrophilic warheads, undruggable targets, functional selection, peptide macrocycles, PPI drug discovery, covalent phage display

Targeting amyloidogenic proteins through cyclic peptides - A medicinal chemistry perspective.

Targeting amyloidogenic proteins through cyclic peptides - A medicinal chemistry perspective.

Cellular uptake and transport mechanism of flaxseed cyclic peptide CLB via clathrin-dependent endocytosis.

Cellular uptake and transport mechanism of flaxseed cyclic peptide CLB via clathrin-dependent endocytosis.

Intestinal anti-inflammatory, histopathologic and anti-oxidative regulatory effects of total alkaloids extract from Linum usitatissimum L. (flaxseed) in vivo.

Intestinal anti-inflammatory, histopathologic and anti-oxidative regulatory effects of total alkaloids extract from Linum usitatissimum L. (flaxseed) in vivo.

Rational design of cyclic peptides, with an emphasis on bicyclic peptides.

Rational design of cyclic peptides, with an emphasis on bicyclic peptides.

From Rat Tails to Glycoproteostasis: Motivated by Biology, Enabled by Biophysics, and Lucky.

From Rat Tails to Glycoproteostasis: Motivated by Biology, Enabled by Biophysics, and Lucky.

Staphylococcus aureus: Current perspectives on molecular pathogenesis and virulence.

Staphylococcus aureus: Current perspectives on molecular pathogenesis and virulence.

Identification and characterization of oncogenic KRAS G12V inhibitory peptides by phage display, molecular docking and molecular dynamic simulation.

Identification and characterization of oncogenic KRAS G12V inhibitory peptides by phage display, molecular docking and molecular dynamic simulation.

Prevalence and significance of anti-citrullinated cyclic peptide antibodies in relapsing polychondritis

Prevalence and significance of anti-citrullinated cyclic peptide antibodies in relapsing polychondritis

Controllable nickel ions release from deferoxamine mesylate-triggered nickel-iron layered double hydroxide for eliciting apoptotic cell death in prostate cancer

Despite their unique advantages and vast potential, nanomaterials employed in cancer therapy still encounter challenges such as uneven biodistribution, unintended drug leakage, and especially potential tissue damage caused by off-target toxicity. Bioinert nanomaterials, known for their excellent chemical stability, and minimal biological reactivity, can exert localized tumoricidal effects in response to specific external stimuli. However, the lack of precise control or poor penetration depth largely limits the therapeutic efficacy, necessitating the development of innovative stimuli-responsive therapeutic strategies. This study presents an alternative drug-responsive cancer therapeutic approach based on nickel-iron layered double hydroxide (NiFe-LDH), which exhibited negligible toxicity to both normal cells and cancer cells. By conjugating a platelet-derived growth factor receptor (PDGFR)-β-targeting cyclic peptide, NiFe-LDH achieved high specificity for prostate cancer cells, significantly enhancing tumor targeting and accumulation. Upon administration of deferoxamine mesylate (DFOM), an FDA-approved iron chelator, NiFe-LDH transitioned from a “bioinert” state to a “bioactive” nanotherapeutic through structural disassembly and robust release of nickel ions (Ni²⁺). The released ions disrupted mitochondrial function, upregulated insulin-like growth factor binding protein 3 (IGFBP3), and further inhibited the PI3K/AKT/mTOR signaling pathway, consequently leading to potent and selective induction of apoptosis in prostate cancer cells. Unlike conventional therapies, which often cause varying degrees of toxicity in non-target organs, this stimuli-responsive nanoplatform could minimize off-target effects and systemic toxicity by combining the non-toxic LDH with the clinically used DFOM. Our findings demonstrate that DFOM-responsive NiFe-LDH can effectively inhibit tumor growth in both cultured cells and tumor xenografts, suggesting a rational and clinically translatable platform for precision cancer therapy.Graphical

Mechanisms of aureobasidin A inhibition and drug resistance in a fungal IPC synthase complex

The enzyme inositol phosphorylceramide (IPC) synthase is essential for survival and virulence in fungi, while absent in mammals, thus representing a potential target for antifungal treatments. Aureobasidin A (AbA), a natural cyclic peptide, displays antifungal activity and inhibits IPC synthase, but the precise molecular mechanism remains unclear. Here, we present the cryo-EM structure of the Saccharomyces cerevisiae IPC synthase, composed of catalytic subunit Aur1 and regulatory subunit Kei1, in its AbA-bound state. The complex is resolved as a dimer of Aur1-Kei1 heterodimers, with Aur1 mediating homodimerization. AbA occupies a predominantly hydrophobic pocket in the catalytic core domain of each Aur1 subunit, blocking the entry of both substrates. Mutations conferring AbA resistance cluster near the AbA-binding site, thus interfering with AbA binding. Our study lays a foundation for the development of therapeutic drugs targeting fungal IPC synthase.

The Isolation, Structural Characterization, and Biosynthetic Pathway of Unguisin from the Marine-Derived Fungus Aspergillus candidus.

Unguisins, a class of structurally complex cyclic peptides featuring a γ-aminobutyric acid residue embedded in the skeleton, exhibit diverse biological activities. Here, a new unguisin K, along with three known congeners, was isolated from the marine-derived fungus Aspergillus candidus MEFC1001. The biosynthetic pathway was elucidated through gene disruption coupled with in vitro enzymatic characterization. The ugs biosynthetic gene cluster (BGC) containing ugsA and ugsB, in conjunction with an extra-clustered gene ugsC, collaborates to synthesize these unguisins. The alanine racemase (AR) UgsC catalyzes the isomerization of Ala and provides d-Ala as the starter unit for the non-ribosomal peptide synthetase (NRPS). The unique localization of ugsC outside the ugs BGC is different from previously reported unguisin-producing systems where AR genes reside within BGCs. The methyltransferase UgsB mediates a key pre-modification step by methylating phenylpyruvic acid to yield β-methylphenylpyruvate, which is subsequently incorporated as β-methylphenylalanine during NRPS assembly. This represents the first experimental evidence of the β-carbon methylation of Phe residue occurring at the precursor level rather than through post-assembly modification. The NRPS UgsA recruits a variety of amino acids for assembly and cyclization to form mature unguisins. Additionally, genome mining utilizing UgsA as a query identified homologous NRPSs in diverse fungal species, highlighting the potential for unguisin production in fungi. This study enriches the biosynthetic diversity of cyclic peptides and provides guidance for exploring unguisin-like natural products derived from fungi.

Role of Peptide Molecular Conformation in Improving the Gene Delivery Efficiency of Liposomal Formulations.

The study explored the impact of peptide conformations on the efficiency of nucleic acids transfection by designing two sets of linear (liKK and liRR) and cyclic peptides (cyKK and cyRR) with various sequences (KK: KKKKCRGDSCKKKK and RR: RRRRCLLLLCRRRR). Incorporating peptides in the liposome formulation improved gene transfection efficiency compared to the peptide and 1,2-dioleoyl-3-trimethylammonium-propane (DOTAP)/1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE) alone. The results revealed that adding linear peptides demonstrated a higher delivery efficiency in comparison to cyclic peptides in transfection formulations. Additionally, two effective groups of transfection formulations were established. The liKK/DOTAP/DOPE formulation proved exceptional at delivering mRNA, achieving a transfection efficiency of 93.3%, comparable to that of Lipofectamine 2000 (91.7%). In contrast, the liRR/DOTAP/DOPE formulation effectively delivered both pDNA and siRNA. The pDNA transfection efficiency reached 42.3%, closely matching Lipofectamine 2000 (43.9%), while the siRNA transfection efficiency hit 90.7%, surpassing Lipofectamine 2000's performance (85.4%). The transfection efficiency of linear conformational formulations is superior to that of cyclic ones. Our research provides new options for liposome formulations and offers valuable insights for designing peptide-based gene delivery vectors with different conformations.

Development of the "Phase Separation" Strategy: Addressing Dilution Effects in Macrocyclization

The phase separation strategy exploits aggregated solvent mixtures of poly(ethylene)glycol (PEG) solvents and hydrophilic organic solvents to control the effective molarity of substrates and catalysts in macrocyclization reactions. The slow diffusion of acyclic precursors from the PEG aggregate “phase” to the organic solvent/catalyst “phase” mimics the slow addition processes typically employed in the synthesis of macrocycles. With the aid of mechanistic studies, the strategy has been applied to copper-catalyzed Glaser-Hay couplings, copper-catalyzed azide-alkyne cycloadditions, copper-catalyzed azide-iodoalkyne cycloadditions and ruthenium-catalyzed olefin metathesis processes. “Phase separation” has been used to form macrocycles of interest in aromachemicals, lipid-based macrocycles, bioactive natural products, mac-rocyclic peptides and bioactive pharmaceuticals. The “phase separation” strategy has been proven to promote transition metal-catalyzed macrocyclization processes with increases in concentration up to 3 orders of magnitude larger than traditional methods like high dilution or pseudo high dilution/slow addition.

CABS-flex 3.0: an online tool for simulating protein structural flexibility and peptide modeling.

Simulating protein structure flexibility using classical methods is computationally demanding, especially for large proteins. To address this challenge, we have been developing the CABS-flex method, which enables fast simulations of protein structural flexibility by combining a coarse-grained simulation approach with all-atom detail. Previously available as the CABS-flex 2.0 web server, the method has now undergone a major upgrade with the release of CABS-flex 3.0. Key improvements include the introduction of intuitive flexibility modes that simplify the control of distance restraints and allow users to reflect known or expected dynamic regions; improved all-atom reconstruction for higher-quality model generation; a new feature for de novo peptide structure prediction, supporting both linear and cyclic peptides along with their conformational flexibility; and new tools for result analysis and visualization, facilitating deeper insights into structural flexibility. Additionally, AlphaFold pLDDT-derived restraints can be used as optional input for guiding simulations. The method accepts input as either a PDB/mmCIF structure or a sequence (for peptide modeling). Advanced options allow users to incorporate experimental or computational restraints. The CABS-flex 3.0 web server is available at https://lcbio.pl/cabsflex3. This website is free and open to all users, with no login requirement.

Natural products for anti-fibrotic therapy in idiopathic pulmonary fibrosis: marine and terrestrial insights.

Idiopathic Pulmonary Fibrosis (IPF) is a chronic fibrotic interstitial lung disease (ILD) of unknown etiology, characterized by increasing incidence and intricate pathogenesis. Current FDA-approved drugs suffer from significant side effects and limited efficacy, highlighting the urgent need for innovative therapeutic agents for IPF. Natural products (NPs), with their multi-target and multifaceted properties, present promising candidates for new drug development. This review delineates the anti-fibrotic pathways and targets of various natural products based on the established pathological mechanisms of IPF. It encompasses over 20 compounds, including flavonoids, saponins, polyphenols, terpenoids, natural polysaccharides, cyclic peptides, deep-sea fungal alkaloids, and algal proteins, sourced from both terrestrial and marine environments. The review explores their potential roles in mitigating pulmonary fibrosis, such as inhibiting inflammatory responses, protecting against lipid peroxidation damage, suppressing mesenchymal cell activation and proliferation, inhibiting fibroblast migration, influencing the synthesis and secretion of pro-fibrotic factors, and regulating extracellular matrix (ECM) synthesis and degradation. Additionally, it covers various in vivo and in vitro disease models, methodologies for analyzing marker expression and signaling pathways, and identifies potential new therapeutic targets informed by the latest research on IPF pathogenesis, as well as challenges in bioavailability and clinical translation. This review aims to provide essential theoretical and technical insights for the advancement of novel anti-pulmonary fibrosis drugs.

Design, Synthesis, and Biological Evaluation of a Novel Long-Acting Human Complement C3 Inhibitor Synthesized via the PASylation-Lipidation Modular (PLM) Platform.

The complement system is essential for immune defense, but its dysregulation contributes to various complement-mediated disorders, including paroxysmal nocturnal hemoglobinuria (PNH). CP40 (a cyclic peptide also known as AMY101), effectively inhibits complement activation by preventing the initial binding of the C3 substrate to convertase. Despite its potency, CP40 has a very short plasma half-life when unbound to human C3, necessitating frequent dosing. We developed a novel PASylation-Lipidation Modular (PLM) platform. This platform incorporates a solubilizing PAS module and a half-life-extending lipid moiety into CP40 via a chemical linker. Systematic optimization of the spacer and lipid components in PLM-modified CP40 analogues identified 6C1 as a lead compound. Compared to CP40, 6C1 exhibited a 5-fold increase in antihemolytic potency in the classical complement pathway and a 6.3-fold improvement in solubility. In vivo studies demonstrated that PLM-CP40 analogues possess superior pharmacokinetic properties, with a 15.6-fold extension in half-life relative to unmodified CP40. Mechanistic studies revealed that the PLM platform extends half-life by interacting with albumin, which serves as a circulating depot for the compound. Surface plasmon resonance analysis and hemolysis assays postalbumin incubation demonstrated that PLM modifications maintain receptor affinity by strategically positioning the albumin-binding moiety away from the peptide region, preserving its biological activity. In a clinically relevant in vitro model of complement-mediated hemolysis in PNH, 6C1 effectively reduced erythrocyte lysis. The PLM platform thus offers a versatile strategy for enhancing peptide therapeutics by improving solubility, extending circulation time, and increasing efficacy, broadening their therapeutic potential.

Molecular Self-Assembly of Peptides into Supramolecular Nanoarchitectures for Target-Specific Drug Delivery.

Self-assembled fluorescent peptides are promising drug-delivery vehicles targeting cancer cells and enhancing the precision of therapeutic agents. Several systems have been developed including fluorescent peptides as cysteine-core peptides, cyclic peptides, nanostructures and peptide polymer conjugates specifically designed for targeted drug delivery. Further, these supramolecular carriers aid in targeted drug transport by using different cargos like doxorubicin (Dox), paclitaxel (PTX), etc. Additionally, dipeptides such as tryptophan-phenylalanine self-assemble via zinc ion chelation, facilitating the endosomal escape thereby enhancing the drug efficacy within multifunctional nanoparticle systems. Furthermore, pH-activatable and enzyme-responsive peptide nanostructures have been engineered to exhibit potential for controlled drug release. These self-assembled peptide systems not only enable targeted drug delivery but also provide controlled release, with applications extending to ocular drug delivery and the treatment of retinal diseases. These systems possess intrinsic fluorescence properties that allow real-time tracking of drug release and cellular uptake, making them highly useful for theranostic applications. Moreover, fluorescently tagged cell-penetrating peptides (CPPs) are widely used to explore how these systems enter cells, revealing multiple ways they are taken up, like endocytosis, micropinocytosis, direct membrane crossing, and counterion-assisted transport. This versatility adds real value to peptide-based approaches in cancer therapy. Further research advancements should enhance stability, explore combination therapies, and improve clinical translation for broader therapeutic applications.

Bioinformatics-Assisted Discovery of Antioxidant Cyclic Peptides from Corn Gluten Meal.

Using a multidisciplinary approach, this paper was designed to prepare, identify, and characterize novel maize antioxidant cyclic peptides from protein hydrolysate of corn gluten meal (CGM). A bioinformatics approach was used to identify the best protease, and the results showed that papain+subtilisin was most likely to produce antioxidant cyclic peptides. The result of the enzymatic hydrolysis validation experiment showed that hydrolysate by papain+subtilisin yielded the highest concentration of cyclic peptide (67.14 ± 1.88%) and remarkable DPPH, ABTS, and hydroxyl radical scavenging rates (81.06 ± 2.23%, 82.82 ± 1.83%, and 47.44 ± 2.43%, respectively) compared to other hydrolysates. Eleven antioxidant cyclic peptides were identified in the protein hydrolysate of CGM through sequential purification and mass spectrometry analysis. The results of molecular docking analysis indicated that the cyclic peptides can form stable hydrogen bonds and hydrophobic interactions with the key amino acid residues of Kelch-like ECH-associated protein 1 (Keap1). Cyclic peptides may regulate the Keap1-Nrf2 pathway by occupying the Kelch domain of Keap1, inhibiting the ubiquitination degradation of Nrf2 (nuclear factor erythroid 2-related factor 2), thereby stabilizing the Nrf2 protein and activating the antioxidant gene network. This study underlined the bioinformatics approach for antioxidant cyclic peptide discovery, which is time- and cost-effective and promotes new cyclic peptide drugs or functional food development.

Metal-Directed Self-Assembly of Minimal Heterochiral Peptides into Metallo-Supramolecular β-Helical Tubules for Artificial Transmembrane Water Channels.

Transmembrane selective transport of metabolites controls essential biological functions. During the last two decades, artificial channels have been developed and cyclic peptides have emerged as ideal platforms for efficient ion, sugar, and nucleic acid channel translocation. Despite these tremendous developments, cyclic peptides have eluded selective water transport. Herein, we report the formation of narrow artificial β-helical tubules with diameters ranging from 2.80 to 3.25 Å that selectively control the water translocation, akin to natural aquaporin channels. The tubular assemblies resulted from the metal-driven folding and assembly of minimal heterochiral metal-binding 3-pyridyl-terminated peptides. The bent ultrashort peptide ligand coordinates with Ag+ metal ions in a head-to-tail manner, which undergoes subsequent polymerization into a β-helical tubular structure stabilized by interstrand hydrogen bonds (H-bonds) between the β-strands and π-π staking interactions between terminal pyridyl moieties. Furthermore, sequence engineering of the heterochiral peptide and subsequent Ag+ ion coordination of the tailored peptides enabled the formation of distinct synthetic double β-barrel and artificial β-helical tubular assemblies, with water molecules encapsulated in the hydrophilic core of the tubes. These water-encapsulated tubes were further explored as artificial water channels in lipid bilayers. Our findings suggest that such β-helical tubular channels achieve a single-channel permeability of 106 water molecules/second/channel, which is within 1-2 orders of magnitude lower than that of aquaporins, with a rather good ability to sterically reject ions and prevent proton transport. These assemblies present significant potential for engineering efficient membranes for water purification and separation sciences.