Author: Oleg Melnyk

Distinctive amide I and II VCD band shapes computed for a 3₁₀-helical octapeptide (red unlabeled, green, labeled with ¹³C=O on 4,5 positions) showing distinctive weak amide I couplet and strong negative amide II for the unlabeled peptide, and an additional couplet side band for the labeled one.

ABSTRACT

Optical spectroscopic techniques have long been used to determine average secondary structure in proteins and peptides, but these are primarily limited to determination of relative sheet and helix content, with the remainder often designated as “other.” Several decades ago, in cooperation with Claudio Toniolo and co-workers, our group at UIC was able to show that vibrational circular dichroism (VCD) could be used to distinguish between formation of 3₁₀-helices and α-helices in small peptides, as well as from other uniform structures. Conventional electronic CD (ECD) or infrared (IR) spectroscopy did not give unambiguous distinctions between these two right-handed, internally H-bonded, helical forms. Theoretical modeling of the VCD spectra showed that the differences between helical forms could not be attributed to just use of Aib residues or of organic solvents, both used to stabilize the 3₁₀ conformation. Later we showed that, with IR and VCD enhanced by isotopic labeling, it was possible to differentiate site-specific structures using vibrational spectra interpreted with DFT-level theoretical modeling. These advances are briefly reviewed here and results of applying them to 3₁₀-helical structures are discussed.

Edge-selective covalent grafting of poly-arginine-11 onto B60 graphene nanoparticles preserves the aromatic basal plane for π-π immobilization of compound 8, highlighting this promising NEK6 inhibitor as the key therapeutic cargo in a peptide-graphene nanoplatform for potential delivery.

ABSTRACT

Covalent peptide grafting is widely used to improve the biological performance of nanocarriers, especially through cell-penetrating peptides (CPPs) that enhance cellular uptake. However, when drug loading relies on noncovalent interactions, peptide functionalization may interfere with surface adsorption processes. This concern is particularly relevant for graphene-based nanocarriers, where π-conjugated molecules bind via π–π interactions on the basal plane. Here, we present an orthogonal functionalization strategy in which peptide conjugation occurs selectively at the edges of graphene nanoparticles (B60), which bear carboxylic acid groups, while π-conjugated cargo is adsorbed on the basal plane. Poly-arginine-11 (R11) was covalently immobilized, preserving the aromatic surface for π-π interactions. Using 1-pyrenecarboxylic acid and compound 8, a π-conjugated NEK6 inhibitor, we show that R11 grafting does not affect loading capacity or thermally induced release. Spectroscopic and microscopic analyses confirm that the basal plane remains intact and accessible after functionalization. Molecular dynamics simulations indicate that peptide chains form a flexible, charged corona at the nanoparticle periphery without perturbing molecule–graphene interactions. Overall, edge-grafted R11 preserves π-π-mediated loading and supports the design of peptide–graphene hybrid systems for delivering poorly soluble aromatic bioactive compounds.

Novel short-chain peptides targeting the KEAP1-NRF2 axis were identified through a multimodal discovery platform combining a stress-tolerant microbial library with machine learning and molecular docking. These peptides regulate collagen synthesis and reduce MMP1/9 levels in human fibroblasts, providing significant protection against photoaging.

ABSTRACT

Excess reactive oxygen species generated by ultraviolet exposure cause photoaging by degrading collagen and inhibiting its synthesis. This study presents a comprehensive strategy connecting the biological stress responses of γ-irradiated microorganisms to the discovery of novel anti-photoaging peptides. We profiled the radiation-regulated transcriptomes of Deinococcus radiodurans and Cryptococcus neoformans, focusing on DNA repair and oxidative stress responses. From these datasets, peptide libraries were generated in silico, filtered for biochemical properties, and prioritized using a seven-classifier machine-learning algorithm. Structural validity was established using Rosetta FlexPepDocking against the KEAP1–NRF2 pocket, which identified 48 docking-positive sequences. We then synthesized the top 21 peptides and subjected them to in vitro validation. Seven of these candidate peptides inhibited collagenase activity at 200 μM. Among them, four peptides dose-dependently increased the procollagen type I C-peptide level in ultraviolet B-induced fibroblasts. Furthermore, these peptides significantly elevated COL1A1 mRNA levels while simultaneously reducing MMP1 and MMP9 transcript and protein levels. In summary, this study provides an integrated strategy that combines omics, machine learning, and docking to discover promising peptide candidates, which were validated through in vitro assessments. This approach offers promising anti-photoaging candidates that can be applied to other oxidative stress pathways and biological resources.

The active ingredient of the immunoregulatory drug Jusvinza, the altered peptide ligand CIGB-814, binds apolipoprotein A-I with higher affinity than its wild-type counterpart. This study demonstrates that a single amino acid substitution is critical for apolipoprotein A-I recognition, potentially broadening the therapeutic applications of Jusvinza to lipid-related diseases.

ABSTRACT

Jusvinza is an immunomodulatory pharmaceutical drug developed for the treatment of rheumatoid arthritis. Its active ingredient is an altered peptide ligand—derived from a T-cell epitope of the human 60-kDa heat shock protein—referred to as CIGB-814. Jusvinza has also proven effective in reducing hyperinflammation in COVID-19. However, molecular targets of CIGB-814 in patients remain undefined, and this study is aimed at identifying specific plasma proteins that interact with CIGB-814. Affinity chromatography and mass spectrometry analysis revealed apolipoprotein A-I as a CIGB-814-binding partner. Interestingly, no apolipoprotein A-I bands were detected in the affinity matrix of the wild-type peptide (designated as E18-3). In this context, it was relevant to analyze how the substitution of Asp18 with Leu—used to design CIGB-814 from the wild-type peptide—confers its ability to bind apolipoprotein A-I. Docking studies predicted the N-terminal domain of apolipoprotein A-I as the most likely region for peptide interaction and suggested a strong connection between CIGB-814 and apolipoprotein A-I. Further validation by molecular dynamics simulations suggests that CIGB-814 may enhance the flexibility of structural areas in apolipoprotein A-I critical for high-density lipoproteins assembly. Additionally, an affinity enzyme-linked immunosorbent assay (ELISA) confirmed that apolipoprotein A-I has a tenfold greater affinity for CIGB-814 compared to the wild-type peptide. These integrated experimental and computational findings demonstrate that a single residue substitution is critical for apolipoprotein A-I recognition. In addition, this study provides mechanistic insight into CIGB-814 immunomodulatory and metabolic effects, expanding the therapeutic potential of Jusvinza to metabolic diseases associated with lipid homeostasis imbalance.

Cementum attachment protein-derived peptides adsorb to specific crystallographic faces of brushite and calcium oxalate, blocking calcium deposition. This phosphorylation-enhanced modulation alters crystal growth under conditions in vitro, leading to the formation of irregular brushite crystals and rosette-like calcium oxalate morphologies. Ultimately, these peptides demonstrate their potential as regulators of pathological mineral crystallization pathways.

ABSTRACT

Pathological mineralization involves the uncontrolled crystallization of calcium phosphate (brushite) and calcium oxalate, leading to renal calculi and ectopic calcifications. Peptides enriched in acidic or phosphorylated residues are potential crystal growth modulators due to their ability to interact with calcium-rich surfaces. This study investigated the in vitro effects of cementum attachment protein-derived peptides, CAP-pi and its phosphorylated analog CAP-pip, on brushite and calcium oxalate crystallization. By isolating these highly anionic motifs, this work introduces a novel biomimetic approach to investigate and modulate the physicochemical mechanisms driving pathological mineral deposition. Assays performed under physiological conditions were analyzed by scanning electron microscopy, Raman spectroscopy, and confocal microscopy, alongside molecular dynamics simulations to examine peptide–calcium oxalate interactions. Both peptides altered crystal growth patterns and lattice organization in a concentration-dependent manner. Peptide treatments induced marked morphological perturbations, promoted irregular and rosette-like habits, and modified vibrational profiles. Furthermore, confocal microscopy revealed selective adsorption onto specific crystallographic regions, whereas molecular dynamics demonstrated enhanced peptide–calcium coordination and interfacial stability for CAP-pip. Collectively, these findings establish phosphorylation as a key determinant of peptide–mineral interactions, providing mechanistic insight into how phosphopeptides regulate crystallization through surface-mediated modulation.

The self-assembly in aqueous solutions, cytocompatibility and collagen production of lipopeptide C₁₆–KTTKS (C₁₆: hexadecyl) and a variant with arginine substitution for lysine, C₁₆–RTTRS, are compared.

ABSTRACT

The self-assembly in aqueous solutions, cytocompatibility, and collagen production of lipopeptide C₁₆–KTTKS and a variant with arginine substitution for lysine, C₁₆–RTTRS, are investigated. C₁₆–KTTKS is known commercially as Matrixyl and is used in cosmetic formulations as it can stimulate collagen production. The self-assembly and conformations and collagen-stimulating effects of the two lipopeptides in two salt forms, trifluoroacetate (TFA) and acetate, are compared. Lipopeptide C₁₆-KTTKS self-assembles into nanotapes based on a multi-bilayer stacking across a pH range pH 4–7 and micelles at pH 2, with little influence of the counterion. In contrast, C₁₆-RTTRS forms a substantial population of spherical micelles for the acetate salt for pH 2–7, but mainly nanotapes for the TFA salt for pH 4–7. Conditions for hydrogel formation by C₁₆-KTTKS were identified. Both lipopeptides show good cytocompatibility to fibroblasts at sufficiently low concentration. The two lipopeptides also stimulate collagen production in Human Dermal Fibroblasts (HDFa) at low concentration (0.0062 wt%). No significant effect of the counterion was noted on cell viability or collagen production. Our results suggest that the peptide sequence influences the pH-dependent self-assembly properties and that this can be modulated for certain lipopeptides by the nature of the counterions.

No abstract is available for this article.

Replacing lysine with arginine, the standard genetic approach to eliminate amine reactivity, alters physicochemical properties and leaves the N terminus unprotected. Here, reductive methylation is characterized as a conservative chemical alternative. Complete dimethylation of human ribonucleases preserves thermostability, protein–protein interaction, cellular uptake, and intracellular persistence, while predictably diminishing enzymatic catalysis. Fully compatible with bioconjugation, this rapid modification is a practical strategy for protein and peptide engineering and developing intracellular biologics such as bioPROTACs.

ABSTRACT

Modification of lysine residues is a common strategy in protein engineering, whether to prevent posttranslational modifications, control bioconjugation, or improve crystallization. The standard genetic approach—replacement with arginine by site-directed mutagenesis—preserves positive charge but alters other physicochemical attributes and cannot address the N-terminal amino group. Here, we characterize reductive methylation as a chemical alternative. This reaction converts every primary amino group to a dimethylamino group rapidly under mild aqueous conditions. Using human ribonuclease 1 and a cytotoxic variant engineered to evade the endogenous ribonuclease inhibitor as model systems, we assess the effects of complete dimethylation on thermostability, enzymatic catalysis, protein–protein interaction, compatibility with bioconjugation, cellular uptake, and intracellular persistence. Dimethylation preserves thermostability and a protein–protein interaction. Enzymatic catalysis, in contrast, is reduced by 10²– to 10³-fold, consistent with the role of catalytic lysine residues. Dimethylation is fully compatible with bioconjugation chemistry. Dimethylated and unmodified ribonucleases show comparable uptake and persistence in human cells. These findings establish reductive methylation as a practical and conservative strategy for lysine modification in protein and peptide engineering and support its use in applications such as biological proteolysis-targeting chimeras (bioPROTACs).

A case study using the TAMARA microfluidic device (Inside Therapeutics) was performed for peptide-based nanoparticles under different operational parameters such as mixing channel geometry, total flow rate, and flow rate ratio. In this context, WRAP5 cell-penetrating peptide-based nanoparticles for siRNA or pDNA delivery demonstrated enhanced homogeneity and reproducibility compared to manually performed formulations.

ABSTRACT

Peptide-based nanoparticles (PBN) have emerged as a promising alternative to lipid nanoparticles (LNP) for nucleic acid delivery and efficient cellular uptake. In this study, we evaluated the formulation of WRAP5 (W- and R-rich amphipathic peptide 5)–based PBN using a microfluidic device and assessed the impact of key process parameters, flow rate ratio (FRR), total flow rate (TFR), and mixing channel design, on nanoparticle characteristics. Across 72 formulations encapsulating small interfering RNA (siRNA) or plasmid DNA (pDNA), dynamic light scattering revealed consistent mean sizes ranging from 50 to 70 nm, with a low polydispersity index (PdI < 0.22), independent of FRR, TFR, or mixer type. Stability studies demonstrated that siRNA-loaded PBN exhibited moderate size increases during storage at 4°C, whereas pDNA-loaded PBN remained highly stable for up to 70 days. Biological assays confirmed robust activity: WRAP5:siRNA PBN achieved approximately 50% CDK4 silencing in GIST-T1 cells, and WRAP5:pDNA PBN mediated efficient mCHERRY expression in HeLa cells, regardless of formulation method or storage duration. These findings highlight the robustness and scalability of WRAP5-based PBN, contrasting with LNP systems that require stringent control of FRR and TFR, and partially underscore their potential for nucleic acid delivery applications.

The developed DOTA-Angiopep-2 conjugate was well tolerated in glioma cells and enabled high-yield radiolabeling with ¹⁷⁷Lu and ¹⁶¹Tb. The ¹⁶¹Tb conjugate showed brain delivery with predominantly renal clearance, supporting a stable radiotheranostic platform for BBB-targeted glioma therapy and SPECT imaging.

ABSTRACT

Brain tumor therapy remains limited by the blood–brain barrier (BBB), which restricts drug access. BBB-penetrating peptides offer a promising strategy for delivering therapeutic and diagnostic payloads. Angiopep-2 is a well-established vector, yet novel radioconjugates based on this vector remain of interest. We report the synthesis and evaluation of DOTA-Angiopep-2 for radiolabeling with Lutetium-177 (¹⁷⁷Lu) and Terbium-161 (¹⁶¹Tb). Notably, ¹⁷⁷Lu serves as a β- and γ-emitter, whereas ¹⁶¹Tb is an Auger and β-emitter; both are utilized in therapy and SPECT imaging. Peptides were synthesized via solid-phase peptide synthesis. Cytotoxicity assays in T98 glioblastoma cells showed that Angiopep-2 is well-tolerated, maintaining ~100% viability at 20 μM and a moderate decline up to 100 μM. Radiolabeling achieved yields > 95% with excellent radiochemical stability at room temperature for up to 10 days and moderate stability in the presence of human serum. Biodistribution in healthy CFW mice showed a brain-associated radioactivity of 0.24% ± 0.05% IA/g at 5 min p.i. and a 12-fold increase in the brain-to-blood ratio (0.028–0.339) by 60 min p.i. These results support DOTA-Angiopep-2 as a versatile platform for radionuclide delivery and a potential candidate for future glioma-targeted studies. Further studies in tumor-bearing models are ongoing to evaluate therapeutic efficacy and translational potential.

The introduction of a single hydrophobic residue at the C-terminus of short cationic peptides enhances passive translocation and cellular uptake.

ABSTRACT

Cationic cell-penetrating peptides (CPPs) are a versatile platform for intracellular cargo delivery into mammalian cells but often suffer from low cellular uptake and endosomal entrapment. Here, we investigated the effect of a single cyclohexylalanine (Cha) residue at the termini of short, conformationally constrained cationic peptides composed of (4S)-guanidiniumproline (Gup). We show that this hydrophobic residue promotes internalization. Our studies also revealed that a hydrophobic residue positioned at the C-terminus enhances cellular uptake more than when at the N-terminus. Comparative studies at different temperatures are consistent with a major entry pathway via direct translocation across the plasma membrane. The findings are useful for the design of CPPs, particularly for enhancing the cellular translocation of otherwise weakly cell-penetrating peptides.

In vitro evidence from crystallization assays demonstrates that peptides inhibit mineral nucleation and crystal growth through sequence-dependent interactions (acidic residues and phosphorylation) with mineral surfaces. In cell cultures, these molecules reduce calcium deposition and ALP activity and modulate the expression of biomineralization-related markers, including RUNX2, BMP, OPN, and MGP.

ABSTRACT

Biomineralization is regulated by interactions between inorganic phases and biological macromolecules. Peptides derived from mineral-associated proteins are key modulators due to their ability to bind crystal nuclei and mineral surfaces. However, the molecular features governing peptide-mediated mineralization inhibition remain insufficiently synthesized in current research. This systematic review, conducted following PRISMA and Cochrane standards, evaluated in vitro evidence regarding the physicochemical properties and inhibitory mechanisms of protein- and peptide-based mineral modulators. Comprehensive searches in MEDLINE, LILACS, and Scopus (up to November 2025) identified fifty studies utilizing crystallization assays or mineralizing cell models. A dedicated risk-of-bias assessment tool, adapted from the RoB2 framework, was developed to evaluate methodological quality in in vitro studies. Although both proteins and peptides were investigated, many inhibitory molecules corresponded to peptides derived from proteins, particularly osteopontin and matrix Gla protein. Effective inhibitors shared common features, including enrichment in acidic residues, low isoelectric points (< 4.8), and posttranslational modifications such as phosphorylation. These properties significantly inhibit mineral nucleation and crystal growth, while reducing calcium deposition and alkaline phosphatase activity in cellular models. This review highlights sequence-dependent physicochemical determinants of peptide–mineral interactions and provides a framework for the rational design of peptide-based mineralization inhibitors.

Extrahepatic siRNA delivery to inflammatory macrophages remains limited by poor tissue selectivity and inefficient endosomal escape. A structure-guided, histidine-rich peptide platform was engineered to create pH-switchable endosomolytic carriers. Peptides were optimized for low membrane activity at physiological pH and high endosomal disruption at acidic pH. This study establishes histidine-rich, pH-responsive peptides as a promising platform for spleen-selective, extrahepatic RNA delivery.

ABSTRACT

Extrahepatic delivery of small interfering RNA (siRNA) remains a major translational challenge because most nanocarriers preferentially accumulate in the liver, while endosomal sequestration limits productive cytosolic release. Inflammatory macrophages in the spleen are attractive therapeutic targets in systemic inflammation, yet spleen-selective delivery systems with efficient endosomal escape remain underdeveloped. Here, a structure-guided peptide engineering workflow was used to generate histidine-rich, pH-switchable endosomolytic peptides for spleen-selective siRNA delivery. Sequence design integrated pH-dependent charge transition modeling, amphipathic helix prediction, membrane interaction scoring, and safety filtering. Six candidate peptides were synthesized and evaluated for pH-responsive structure, membrane disruption, hemocompatibility, siRNA complexation, serum stability, macrophage uptake, endosomal escape, biodistribution, and anti-inflammatory efficacy. The lead peptide, HSEP-6, showed a predicted net charge increase from +3.1 at pH 7.4 to +7.4 at pH 5.5, helix content increasing from 17% to 56%, and acidic calcein release increasing from 9% to 62%. In inflammatory macrophages and LPS-challenged mice, HSEP-6 enabled efficient siRNA delivery, spleen-selective accumulation, marked Irf5 silencing, reduced TNF-α and IL-6, and no measurable systemic toxicity, supporting histidine-rich pH-switchable peptides as a rational platform for extrahepatic RNA delivery.

Undecapeptide derived from the interacting interface of SARS-CoV-2 spike protein and ACE2.

ABSTRACT

The envelope-anchored trimeric spike protein of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) mediates the attachment and entry of the virus within human cells by utilizing the angiotensin converting enzyme 2 (ACE2) as a receptor, present in the epithelial cells of the upper respiratory tract and lungs. The ACE2 interaction interface with the SARS-CoV-2 spike protein was utilized to design an ACE2-derived peptide sequence (FD11) that targeted the viral spike protein. It was found to be non-cell penetrating, while in vivo studies revealed the non-cytotoxic nature of the peptide molecule. The minimum effective concentration of the peptide was found to be 10 μM with 95% cell viability in Vero E6 cell line. The IC₅₀ of the peptide was found to be around 6.4 μM. Treatment of the peptide in SARS-CoV-2 infected Vero E6 cells was found to decrease the infectivity as compared to control. This presents a simple approach where host-pathogen protein interaction-interface derived peptides can be used for binding with substrate macromolecules to target and modulate pathogen infectivity.

No abstract is available for this article.

The intrinsic reactivity of [Au(bnpy)Cl₂] toward the 12-mer C-terminal peptide of human thioredoxin reductase revealed mechanistic insights into the chemoselective arylation of the Cys-Sec pair of the enzyme. The reductive elimination and liberation of gold(I) is the key step of the process occurring more rapidly at selenocysteine than at cysteine.

ABSTRACT

Thioredoxin reductase (TrxR) is a well-known target for gold-based drugs, which typically behave as strong inhibitors of this selenoenzyme. Despite recent evidence that cyclometalated Au(III) complexes inhibit TrxR1 and arylate cysteine (Cys)- and selenocysteine (Sec)-containing systems, there is still limited direct mechanistic evidence at the level of the TrxR C-terminal motif containing a Cys-Sec pair that is essential for catalytic activity. In this study, we investigated the intrinsic reactivity of a cyclometalated Au(III) compound, namely [Au(bnpy)Cl₂], toward the 12-mer C-terminal peptide of human TrxR1, at a molecular level, to gain mechanistic insights into the chemoselective arylation of TrxR1. Time-dependent and reagent-ratio studies showed that reductive elimination and liberation of gold(I) is the key step in the process that occurs more rapidly at selenocysteine than at cysteine. Furthermore, comparable extents of arylation were observed under organic and biocompatible conditions, highlighting the efficiency of this transformation in physiologically relevant media. Overall, these findings provide direct mechanistic support for exploiting cyclometalated gold(III) complexes as covalent, Sec-directed agents for targeting thioredoxin reductase and related redox-active proteins. The TrxR C-terminal peptide offers a minimal but informative model to rationalize how scaffold-dependent reactivity can translate into selective and potent inhibition of the full enzyme.

Short arthropod-derived antimicrobial peptides can play pivotal roles in tackling MDR infections in the future.

ABSTRACT

The global surge in antimicrobial resistance has intensified the search for novel therapeutic strategies that can overcome the limitations of conventional antibiotics. Antimicrobial peptides (AMPs), particularly those derived from arthropod venoms, have emerged as promising alternatives due to their unique ability to target conserved microbial structures and minimize resistance development. Among these, Lasioglossin-III (LL-III) stands out for its remarkable potency and versatility. LL-III exhibits a broad spectrum of activity encompassing antibacterial, antifungal, and anticancer properties, attributed to its structural features that facilitate selective interactions with microbial and cancer cell membranes. Despite its promising therapeutic profile, comprehensive reviews focusing specifically on LL-III remain scarce, with most existing literature addressing lasioglossins only in general terms. This review provides the first in-depth and consolidated discussion on LL-III by examining its structural characteristics, mechanisms of action, and pharmacological applications, alongside insights from computational modeling and experimental studies. By integrating current knowledge on the structure–function relationship of LL-III, this article underscores its translational potential as a next-generation therapeutic. Notably, the peptide’s dual mechanism of action, combining membrane disruption with intracellular targeting, together with its multifunctional properties, including antibacterial, antifungal, anticancer, and immunomodulatory activities, positions LL-III as a promising, versatile peptide-based therapeutic candidate.

No abstract is available for this article.

Rationally designed β-hairpin peptide mimetics selectively detect ELABELA for preeclampsia diagnosis in pregnant women. The OP27 probe enables sensitive, concentration-dependent electrochemical detection of ELABELA in serum.

ABSTRACT

Preeclampsia (PE) poses risks to mothers and infants, especially in low-resource settings. Diagnosis of PE is difficult due to nonspecific symptoms and limited biomarker screening. ELABELA, a circulating peptide hormone and endogenous ligand of the apelin receptor (APJ), has emerged as a promising biomarker for earlier detection of PE, alone or in combination with other biomarkers. Here, we present β-hairpin protein epitope mimetics (BH-PEMs) as a tool for ELABELA detection. Employing an epitope mimetics approach, OP27 from a BH-PEM peptide library emerged as a high-affinity candidate for ELABELA following rational design and screening of a focused peptide library. Experimental assays verified the OP27-ELABELA complex’s stability and binding affinity. We successfully detected ELABELA in clinical donor serum samples using the OP27-ELABELA complex–based electrochemical method integrated with electrochemical impedance spectroscopy (EIS), enabling sensitive detection in a complex biological matrix. Our findings establish a proof-of-concept biomarker-driven electrochemical detection strategy for PE, supporting the development of simplified electrochemical diagnostic approaches while highlighting the broader applicability of BH-PEM probes in biomarker detection. The BH-PEM probe demonstrates feasibility for biomarker-driven PE detection and warrants further validation in expanded clinical cohorts, thereby contributing to improved maternal risk assessment and mitigation of health risks for mothers and infants.

Peptide-based chelator consisting of Cys-Gly-Gly-Gly incorporated into HER2-targeting rL-A9 peptide for ^99mTc-labeling. [^99mTc]Tc-CGGG-rL-A9 exhibited high binding affinity and tumor uptake and retention.

ABSTRACT

In our pursuit of developing HER2-targeting radiolabeled peptides for detection and therapy of HER2-positive breast cancer, the previously reported rL-A9 peptide had shown promising potential. In the present work, a different variant of rL-A9 was designed by incorporation of peptide-based chelators for ^99mTc-labeling. Cysteine-based amino acids (CGGG, CSSG, CEEG) were conjugated at the N-terminus of the peptide and were radiolabeled with [^99mTc]NaTcO₄. Of the three radiopeptides, [^99mTc]Tc-CGGG-rL-A9 exhibited high stability, whereas [^99mTc]Tc-CSSG-rL-A9 and [^99mTc]Tc-CEEG-rL-A9 decomposed rapidly. Hence, only CGGG-incorporated radiopeptide was further investigated in vitro in human breast carcinoma SKBR3 cells and in vivo in tumor xenografted mice. Cell-binding studies demonstrated high binding affinity [K_d: 5.82 ± 0.92 nM] and specificity (63% reduction in blocking study) of [^99mTc]Tc-CGGG-rL-A9 toward HER2-expressing cells. In vivo studies showed moderate tumor uptake (3.56 ± 0.81% ID/g, 1 h) with 82.8% retention at 3 h (3.16 ± 0.22% ID/g). Moderately high tumor-to-background ratios, along with washout from the other non-target organs, were observed. Inhibition studies revealed 52.6% reduction in the tumor activity, signifying high HER2-receptor specificity of the presently studied radiopeptide, [^99mTc]Tc-CGGG-rL-A9.

Residues with backbone N-amination have been introduced into a region of parallel β-sheet secondary structure. Contrary to predictions, the N-amination of Trp9 or Phe33 did not give an increase in the thermal stability of DS119. MD simulations suggest this is due to the highly fluctuating nature of the β-sheet region.

ABSTRACT

The miniprotein DS119 has been used as a model system to probe the effects of introducing residues with backbone N-amination into a region of parallel β-sheet secondary structure. Derivatives featuring backbone N-methylation have also been synthesized for comparison. As expected, N-methylation of Trp9 or Phe33, positioned on the outer edge of the β-strands, led to a reduction in the thermal stability of the protein. However, contrary to predictions, the N-amination of Trp9 or Phe33 did not lead to an increase in thermal stability of DS119. Refinement of the DS119 structure, using NOE restrained molecular dynamics simulations, shows that the β-sheet region is highly fluctuating in nature. Key interstrand hydrogen bonds have populations of 24%–77%, whereas others have populations of less than 10%. In this disordered β-sheet region, the altered conformational preferences arising from backbone N-amination therefore have only minimal effects. This study demonstrates how MD simulation refinement can identify important dynamical features in a protein structure that might be overlooked in standard protein structure determination protocols.

No abstract is available for this article.

No abstract is available for this article.

Strategic shift to oral delivery: Oral peptide nano-formulations address the critical need for non-invasive, chronic administration. Enhancer-centric design: The review advocates for using permeation enhancers with established regulatory precedence. Focus on reproducibility: Viability in oncology is defined by controlled pharmacokinetic variability and reproducible exposure. Minimalist architecture: Lipid-based nano-platforms that facilitate peptide–enhancer co-localization over complex, novel structures. Translational priorities: Development pathways must prioritize CMC robustness and scale-up feasibility, and for successful commercialization.

ABSTRACT

Breast cancer is increasingly managed as a long-term condition, intensifying the need for therapies that are effective, tolerable, and suitable for chronic administration. Peptide-based therapeutics offer high molecular specificity and favorable safety profiles; however, their clinical utility remains largely limited by reliance on parenteral delivery. Oral administration is strategically attractive for improving patient adherence, quality of life, and healthcare efficiency, yet peptide drugs face formidable gastrointestinal barriers, including enzymatic degradation, restricted epithelial permeability, and substantial pharmacokinetic variability. These challenges may be further exacerbated by treatment-related gastrointestinal dysfunction in oncology populations. This review critically evaluates oral peptide nano-formulations that incorporate molecular permeation enhancers with established regulatory precedence as a pragmatic approach to achieving clinically meaningful oral absorption in breast cancer. Rather than emphasizing formulation novelty, the article adopts a translational, industry-oriented perspective that prioritizes regulatory alignment, chronic-use safety, manufacturability, and CMC robustness. Mechanistic and translational aspects of key enhancer classes—including medium-chain fatty acids, bile-derived agents, and surfactant-based enhancers—are examined, with particular attention to epithelial reversibility, dose–response nonlinearity, gastrointestinal tolerability, and exposure consistency. Evidence supports an enhancer-centric paradigm which primarily serves to protect peptides and co-localize peptide–enhancer exposure at the intestinal absorption site.

Exenatide and H-GLP-1 belong to the GLP-1 agonist class of potential therapeutic agents. A robust multienzyme peptide-mapping workflow combined with LC-HRMS/MS enables high-resolution sequence coverage and monitoring of post-translational modifications, providing confident structural verification and comprehensive quality evaluation.

ABSTRACT

Peptide therapeutics are becoming an increasingly important class of pharmaceuticals due to their attractive therapeutic properties. However, their development remains challenging because of inherent structural complexity, requiring advanced analytical techniques. Peptide mapping is a key method for comprehensive identification of a peptide’s primary structure, and FDA and EMA recommend its use to demonstrate structural “sameness.” In this study, we describe a multienzyme peptide-mapping workflow designed to improve sequence coverage and structural verification. The protocol integrates multienzymatic digestion with complementary specificity, enabling generation of alternative peptide maps. Waters XBridge Peptide BEH C18 column (300 Å, 2.5 μm, 4.6 × 150 mm) with gradient separation program was used for separation. Further, LC-HRMS (Orbitrap) was employed to obtain high-resolution MS/MS data, achieving precise peptide fragment identification with mass accuracy within < 5 ppm. Two antidiabetic peptides, exenatide and H-GLP-1, were selected as model systems. Full sequence coverage of both peptides was achieved by combining peptide maps produced with trypsin, Glu-C, and chymotrypsin. Application of the workflow to a GLP-1 analog provided > 95% sequence coverage and accurate intact-mass confirmation. Overall, the described method offers a reliable and stepwise approach for peptide-mapping-based structural verification, enabling confident assessment of the primary structure of exenatide and H-GLP-1.

The GFPP is administered by a scientific committee composed of members elected at the general assembly held at each congress, as well as the delegate from France to the scientific council of the European Peptide Society (EPS), which is elected by the French members of the EPS. GFPP board members 2025-2027   Maud Larregola President …

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A predictive workflow enables accurate transfer from analytical HPLC to preparative FPLC for peptide purification. Systematic evaluation of key chromatographic parameters identified particle size, flow optimization and modifier choice as critical for selectivity and resolution. A correction model minimized elution transfer errors from −17% to less than −5%, achieving first-pass purifications with purities higher than > 90%.

ABSTRACT

Optimal purification, high purity and robust purity control for synthetic peptides are critical, as even minor impurities can alter biological activity, distort analytical results and compromise downstream applications. Reliable translation of analytical HPLC results to preparative FPLC conditions remains challenging due to system-specific differences in column geometry, gradient formation and mobile-phase chemistry. This study addresses these limitations through a systematic evaluation of chromatographic parameters influencing peptide selectivity, resolution and transferability. Using a defined peptide impurity library, the effects of gradient steepness, flow rate, temperature and mobile-phase modifier were quantified, revealing that flow-rate optimization and modifier choice have the greatest impact on separation quality. A correction equation was developed to compensate for system-dependent deviations, reducing transfer errors in elution percentage from approximately 17% to less than 5%. The optimized workflow enabled initial preparative purifications with purities above 90% and yields exceeding 30%. Additionally, substitution of trifluoroacetic acid with formic acid was explored as a greener modifier, providing selective improvements in separation performance. The approach establishes a practical and sustainable workflow for the transfer of HPLC-to-FPLC methods for peptide purification.

N-Myc-derived peptidomimetics bearing aryl sulfonyl fluoride warheads were shown to act as effective N-Myc/Aurora-A PPI inhibitors, selectively labelling Aurora-A in a recognition-directed manner.

ABSTRACT

Orthosteric inhibition of the N-Myc/Aurora-A protein–protein interaction (PPI) represents a potential mechanism by which degradation of N-Myc can be induced, given its interaction with Aurora-A competes with the factors that tag it for proteasomal degradation. As such, this would constitute an approach for the development of drugs to treat neuroblastoma, a childhood cancer that depends upon N-Myc. Reactive electrophiles have proven useful in the context of targeted covalent inhibitors, and in this work, we sought to improve the potency of a previously identified N-Myc-derived peptide by introducing a sulfonyl fluoride warhead. We successfully demonstrated selective labelling of Aurora-A using the resultant peptidomimetics and established this labelling as recognition-directed, providing valuable insight for further future development of N-Myc peptidomimetics and further broadening the use of aryl sulfonyl fluoride warheads in the context of peptidomimetic PPI inhibitors.

Discovery, evolution and potential future applications of bismuth bicycle molecules—a novel class of therapeutics with unique properties.

ABSTRACT

Bicyclic peptides are emerging as next generation therapeutics by combining the affinity and specificity of antibodies with the synthetic convenience of small molecules. Phage-encoded libraries of bicyclic peptides enable the discovery of high-affinity molecules against virtually any protein target. The generation of bicyclic peptides that advanced into clinical development involves the reaction of three cysteines in a peptide to a C₃-symmetric alkylating agent. In phage display, this chemical modification transforms a pool of conformationally flexible peptides into a library of structurally unique protein mimetics that are able to bind traditionally challenging protein surfaces like those with limited structural definition. In recent years, a new class of bicyclic peptides has emerged using a single atom—bismuth—in place of C₃-symmetric organic scaffolds, thus expanding into an unexplored chemical space at the intersection of inorganic chemistry and biology. This mini-review aims to reflect on the discovery, evolution and potential future applications of bismuth bicycle molecules.

As an alternative to the conventional approach, which combines amino acid monomers in a one-by-one fashion to peptide derivatives, the chemical modification of existing peptides has attracted significant attention in recent years. However, such approaches generally target the reactive functional groups in cysteine and lysine residues, particularly in the case of larger peptides, and the modification of peptides that do not feature these functionalities is more difficult. This review focuses on recent attempts to achieve such more ambitious chemical modifications.

ABSTRACT

Very minor structural modifications in peptides can result in significant changes in their function. To obtain analogues with such small structural but big functional differences compared to the original peptides, amino acid monomers are usually combined one-by-one from scratch, which is a simple and reliable but painfully laborious strategy. One alternative and very fascinating approach is the chemical modification of existing peptides, which allows for the rapid production of derivatives with fewer synthetic steps. However, such approaches generally target the reactive functional groups in cysteine and lysine residues, particularly in the case of larger peptides, and the modification of peptides that do not feature these functionalities is more difficult. Nevertheless, chemists have also been exploring methods that can be applied even to such chemically inert peptides based on recent advances in C–H activation and hydrogen atom transfer (HAT) chemistry. If successful, these strategies would represent a breakthrough in terms of obtaining unusual peptide structures in a time- and cost-effective manner. This review focuses on recent attempts to achieve such ambitious chemical modifications, albeit that these are currently limited to relatively small peptides.

An elbow epitope is identified as the specific binding site of BMP2 coreceptor, in addition to the previously reported wrist and knuckle epitopes serving as the recognition sites of BMP2 type-I and type-II receptors, respectively. Several bioactive peptides are successfully derived from the elbow epitope segment, which exhibit good binding potency to specifically target BMP2 coreceptor.

ABSTRACT

Human bone morphogenetic protein-2 (BMP2) is an effective osteoinductor in bone formation and growth via binding to its cognate receptors and coreceptors. Traditionally, the protein is identified to possess a conformational wrist epitope and a linear knuckle epitope that can be recognized by type I and type II receptors, respectively. In this study, we defined an additional epitope termed elbow as the specific binding site of its coreceptor, which is concentrated into a small helical hairpin region of BMP2 monomer 1 and partially overlaps with wrist but is far away from knuckle. A linear lEEP peptide is derived from the elbow epitope, which, however, is intrinsically disordered and cannot be maintained in native helical hairpin conformation due to lack of BMP2 protein context support, thus only exhibiting a low affinity to coreceptor. Disulfide stapling strategy is then used to stabilize the helical hairpin conformation of linear lEEP, which results in its two cyclic counterparts. The stapling is demonstrated to effectively improve peptide affinity and also exhibits a high selection for coreceptor over type-I receptor. The cyclic peptides are found to maintain a well-ordered, native-like conformation, which can be readily recognized by coreceptor through a conformational selection mechanism.

We present a simple method for isolating the novel amino acid 2-thiohistidine and its Fmoc-derivative through the addition of sodium chloride to the isolation step. The introduction of sodium chloride to this step greatly improves the yield and purity of the product. The metal binding utility of this amino acid should be further explored.

ABSTRACT

Here, we report on an improved synthesis of the novel amino acid 2-thiohistidine and its Fmoc-derivative through the addition of sodium chloride to the isolation steps of each synthetic procedure. These compounds must be synthesized in either water or a mixture of water/organic solvent and have proven difficult to isolate by traditional methods. We serendipitously discovered that the addition of sodium chloride caused each compound to precipitate from the solution during the workup. Since 2-thiohistidine is the amino acid analogue of the natural product ergothioneine, we hypothesize that sodium cations mimic the binding of other biologically relevant metal cations to the thioimidazole moiety. Ergothioneine is known to bind Cu⁺, Zn²⁺, and Hg²⁺, and it may bind other metals. Our fortuitous discovery is not only useful for the isolation and purification of 2-thiohistidine and its derivatives; peptides that contain this unique amino acid may be designed to take advantage of binding to Group I cations and other metals.

Novel chiral cyrene-derived Cyr residues were incorporated into peptides. The α,α-disubstituted amino acid residue was introduced using the Ugi reaction. X-ray crystallographic analysis of R– and S-Cyr residues indicated preference to adopt left- and right-handed α-helical conformations, respectively. Propensity to adopt positions in β- and γ-turn hydrogen-bonded conformers was also indicated in the solid state and by NMR studies in solution, highlighting potential in peptide design and conformational control.

ABSTRACT

Cyrene-derived α,α-disubstituted amino acids (H-Cyr-OH) are introduced into peptides using the multiple component Ugi reaction. Conformational analysis of the Cyr monomer and dimer was performed by X-ray crystallography and NMR spectroscopy. In the solid state, the backbone torsion angle values of (4R)- and (4S)-Cyr were respectively characteristic of left- and right-handed α-helical conformers; however, the dimer appeared to adopt a 10-membered hydrogen bond in a distorted type III β-turn. In solution, solvent shielded amide NH hydrogens suggested that the Cyr residue could adopt the central position of γ-turn conformations. With conformational properties indicative of α-, β-, and γ-turns, the Cyr residue offers interesting potential as a novel chiral α,α-disubstituted amino acid for exploring chemical space.

A lipopeptide is designed that contains an epitope from simian virus T-antigen (SV40T, PKKKRKV) conjugated to an N-terminal palmitoyl (C₁₆-) moiety, which acts as a cell-penetrating lipopeptide, with additional aggregation propensity conferred by the lipid chain. The lipopeptide enables DNA delivery into the cytoplasm and nucleus.

ABSTRACT

A lipopeptide is designed that contains an epitope from simian virus T-antigen (SV40T, PKKKRKV) conjugated to an N-terminal palmitoyl (C₁₆-) moiety, with the aim to act as an effective cell-penetrating lipopeptide, with additional aggregation propensity conferred by the lipid chain. A combination of cryo-TEM and small-angle X-ray scattering (SAXS) is used to show that the lipopeptide forms micelles, but mixtures with DNA lead to formation of fractal cluster-like co-assemblies due to intercalation of the DNA and peptide. Spectroscopic studies using fluorescence and circular dichroism (along with fiber X-ray diffraction) show that the peptide interacts with DNA and inserts into the groove. Confocal microscopy along with flow cytometry confirms delivery of DNA into both HeLa and mouse embryonic stem cells (mESCs) in pluripotent state, and the system shows excellent cytocompatibility as confirmed by MTT assays. Our data indicate that the lipopeptide may outperform the DNA transfection agent lipofectamine in DNA delivery into these stem cells and it enables DNA delivery into the cytoplasm and nucleus.

No abstract is available for this article.

This review talks about the development of incretin agonists (glucagon-like peptide-1 [GLP-1], GIP [glucose-dependent insulinotropic polypeptide], glucagon), oral formulation development approaches, and advanced drug delivery platforms. It provides details about clinical trials on incretin agonists, focussing on efficacy, safety, and innovations in clinical research.

ABSTRACT

Incretin-based therapies have become central to type 2 diabetes (T2D) management, offering benefits beyond glycemic control, including weight reduction, cardiovascular protection, and emerging roles in renal and neurological health. This review addresses the question: What are the recent advances in GLP-1, GIP, and glucagon receptor agonists, and how do formulation strategies overcome biopharmaceutical challenges? We systematically analyzed late-stage clinical trials and formulation approaches for peptide-based therapies. The review focuses on therapeutic efficacy, structural and physicochemical properties influencing absorption, distribution, and metabolic stability, and strategies to mitigate degradation pathways such as enzymatic hydrolysis and peptide aggregation. Additionally, innovative delivery systems such as oral peptide formulations and long-acting injectables demonstrate promise in addressing inherent challenges of peptide drug delivery. By integrating clinical outcomes with mechanistic and formulation insights, this review highlights the evolving landscape of incretin-based therapies and underscores innovative solutions for peptide stabilization and delivery. These findings provide a forward-looking perspective for clinicians, researchers, and pharmaceutical scientists engaged in T2D management and drug development.

Les journées scientifiques « peptides antimicrobiens et toxines peptidiques » (PAMTOX) auront lieu les 20 et 21 mai 2026 au Muséum National d’Histoire Naturelle (MNHN, Paris). Ces journées sont co-organisées par le réseau Multifonction des Peptides AntiMicrobiens (MuFoPAM, https://www.mufopam.fr/) et le Groupe Français des Peptides et Protéines (GFPP, www.gfpp.fr). Les inscriptions sont en ligne, à …

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No abstract is available for this article.

The constrained tryptophan (Trp) analog 1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid (Tcc) is used to restrict peptide conformation in structure–activity relationship studies. Although Pictet–Spengler cyclization of Trp and aldehydes with mineral acid gives Tcc analogs, such harsh conditions do not tolerate acid-labile functionality. To enable broader use of Tcc residues, mild conditions are now reported for the on-resin Pictet–Spengler cyclisation in the presence of acid-labile side chain protection and linker strategies.

ABSTRACT

The constrained tryptophan (Trp) analog 1,2,3,4-tetrahydro-β-carboline-3-carboxylic acid (Tcc) is used to restrict peptide conformation in structure–activity relationship studies. Although Pictet–Spengler cyclization of Trp and aldehydes with mineral acid gives Tcc analogs, such harsh conditions do not tolerate acid-labile functionality. To enable broader use of Tcc residues, mild conditions are now reported for the on-resin Pictet–Spengler cyclisation in the presence of acid-labile side chain protection and linker strategies.

Le prochain congrès du GFPP aura lieu à l’Île d’Oléron du 13 au 17 juin 2027

L’article GFPP26 est apparu en premier sur Groupe Français des Peptides et des Protéines.

Le prochain congrès du GFPP aura lieu à l’Île d’Oléron du 13 au 17 juin 2027

L’article GFPP26 est apparu en premier sur Groupe Français des Peptides et des Protéines.

Le prochain congrès du GFPP aura lieu à l’Île d’Oléron du 13 au 17 juin 2027

L’article GFPP26 est apparu en premier sur Groupe Français des Peptides et des Protéines.

New functionalized analogues of 1-nitroacridine/4-methyl-1-nitroacridine and 4-nitroacridone connected to tuftsin/retro-tuftsin derivatives as a potential anticancer therapeutics.

ABSTRACT

The aim of our work was to analyze new functionalized analogues of 1-nitroacridine/4-methyl-1-nitroacridine and 4-nitroacridone connected to tuftsin/retro-tuftsin derivatives as a potential anticancer therapeutics. Using the MTT assay, we have evaluated the cytotoxic activity of synthesized analogues against the Jurkat cell line and healthy donor–isolated peripheral blood mononuclear cells (PBMCs). Jurkat cells were susceptible to several of the acridine derivatives, whereas PBMCs were sensitive to two compounds only. 4-Methyl-1-nitro-acridine analogue, classified as compound 28, has exhibited the greatest anticancer potential with 4-fold higher cytotoxicity toward Jurkat cells and 12-fold lower cytotoxicity against PBMCs when compared with control therapeutics. Subsequent molecular docking analysis suggested that binding to epidermal growth factor receptor (EGFR) and proto-oncogene tyrosine-protein kinase Src molecules could be, at least partially, responsible for observed biological effects of created acridine/acridone analogues.

Journal of Peptide Science, Volume 32, Issue 1, January 2026.

In the Fmoc-SPPS of the Glu-Asp-Tyr motifs, Asi, Agl, and Pgl by-products are formed, amplified by microwave assistance. Their levels depend on the sequence, protecting groups, deprotection conditions, and the conformation/steric hindrance of the growing chain. Optimal minimization: 6% piperazine with 0.1 M oxime.

ABSTRACT

Peptides are important tools in biological, medical, and pharmaceutical research. Solid-phase peptide synthesis (SPPS) is the primary method for their preparation in high yields and purity. However, SPPS often encounters difficulties due to the occurrence of side reactions, which can generate by-products that are difficult to remove. Side reactions can occur under both basic and acidic conditions, resulting in reduced yields, costly purification processes, and sometimes potential inaccessibility of the target peptides. By-products include the formation of aspartimide, glutarimide, and pyroglutamic acid. The problem of unwanted intramolecular cyclizations is well known and unavoidable, in some cases minimized, as it depends on the intrinsic thermodynamics of the final structures. In this context, we report a systematic analysis of the side reactions that occur during SPPS in peptides containing the Glu-Asp-Tyr motif. This study is the first to investigate the formation of three different by-products within the same peptide sequence and to provide valuable insights into the experimental conditions that favor one reaction over the others. The study aims to identify the optimal experimental conditions to mitigate these by-products’ formation. Our results highlight that the steric and conformational effects of the growing protected peptide chain play a role that is often overlooked or misinterpreted.

ASSOCIATION GROUPE FRANÇAIS DES PEPTIDES ET DES PROTEINES  STATUTS MIS A JOUR EN DATE DU 24/06/2025   ARTICLE 1. DÉNOMINATION Il a été fondé en 1992 une association régie par la loi du 1er juillet 1901 et le décret du 16 août 1901, sous la dénomination de GROUPE FRANÇAIS DES PEPTIDES ET DES PROTEINES (GFPP). …

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Ten novel sNPF analogs were designed and synthesized based on the structure of lead I-3 by molecular docking and peptidomimetic methods. Compound A-1 exhibits better binding affinity with the receptor, possesses excellent aphidicidal activity, and shows low toxicity to bees, making it a promising candidate for environmentally friendly insecticides.

ABSTRACT

Short neuropeptide F (sNPF) is a peptide unique to insects, characterized by a C-terminal phenylalanine and a conserved RLRFa motif, and plays key roles in controlling feeding behavior, growth, circadian rhythms, and water–salt homeostasis. We previously identified an sNPF analog, I-3, with aphidicidal activity. In this study, 10 sNPF analogs with aromatic or nonaromatic modifications at the N-terminus were designed based on I-3, using molecular docking and peptidomimetic strategies to investigate the role of N-terminal residues. Aphicidal activity showed that A-1 has stronger activity than I-3 and pymetrozine. Structure–activity analysis indicated that a benzene ring with electronegative and lipophilic groups at the N-terminus is key for aphicidal activity. Molecular docking and molecular dynamics simulations showed A-1 binds more stably to the receptor than I-3. Toxicity tests on honeybees (Apis mellifera) confirm that compound A-1, which exhibits strong aphidicidal activity, is safe for nontarget organisms. Additionally, Admetsar3 evaluations indicate low toxicity risks for all compounds. Therefore, A-1 represents a promising, selective, and eco-friendly insecticide for controlling pea aphids, and this study validates the feasibility of developing novel green pesticides based on sNPF.

No abstract is available for this article.