Search Results (698)

Spurred by the enormous therapeutic success of glucagon-like peptide-1 receptor (GLP-1R) agonists (GLP1-RAs) against diabetes and obesity, glucagon family receptor pharmacology has garnered a tremendous amount of interest. Glucagon family receptors, e.g., the glucagon receptor itself (GCGR), the GLP-1R, and the glucose-dependent insulinotropic peptide receptor (GIPR), belong to the incretin receptor superfamily, i.e., receptors that increase blood glucose-dependent insulin secretion. All incretin receptors are class B1 G protein-coupled receptors (GPCRs), coupling to the G_s type of heterotrimeric G proteins which activates adenylyl cyclase (AC) to produce cyclic adenosine monophosphate (cAMP). Most GPCRs undergo desensitization, i.e., uncouple from G proteins and internalize, thanks to interactions with the βarrestins (arrestin-2 and -3). Since the βarrestins can also mediate their own G protein-independent signaling, any given GPCR can theoretically signal (predominantly) either via G proteins or βarrestins, i.e., be a G protein- or βarrestin-“biased” receptor, depending on the bound ligand. A plethora of experimental evidence suggests that the GLP-1R does not undergo desensitization in physiologically relevant tissues in vivo, but rather, it produces robust and prolonged cAMP signals. A particular property of constant cycling between the cell membrane and caveolae/lipid rafts of the GLP-1R may underlie its lack of desensitization. In contrast, GIPR signaling is extensively mediated by βarrestins and the GIPR undergoes significant desensitization, internalization, and downregulation, which may explain why both agonists and antagonists of the GIPR exert the same physiological effects. Here, we discuss this evidence and make a case for the GLP-1R being a phenotypically or functionally G_s-selective receptor. We also discuss the implications of this for the development of GLP-1R poly-ligands, which are increasingly pursued for the treatment of obesity and other diseases. Full article

Gramicidin S (GS), an antimicrobial peptide (AMP), exhibits broad-spectrum activity against bacteria and cancer cells but is limited in clinical use due to its cytotoxicity toward eukaryotic cells. Lipid-based delivery systems may overcome this limitation; in this study, we proposed and tested simple and promising lipid formulations, including dipalmitoylphosphatidylcholine (DPPC), cardiolipin (CL), and cholesterol (CHOL). We evaluated the interactions of these lipid membranes with GS by assessing membrane fluidity, dielectric permittivity, dielectric losses, dielectric relaxation frequency, and static dielectric constant. Among these, membrane fluidity and dielectric permittivity were the most sensitive to GS, showing significant changes in the formulation containing 90 mol% DPPC and 10 mol% CHOL when exposed to 20 μM GS. Notably, although membrane fluidity changed in a dose-dependent manner following GS binding, the liposomes still supported relatively high GS concentrations—up to 80 μM—which is important for future high-dose GS applications. Additionally, we performed preliminary cytotoxicity tests comparing free GS with liposome-carried GS using the tested lipid compositions and observed a significant reduction in GS-associated toxicity on L929 cell line. This study provides new insights into GS–membrane interactions and supports the rational design of AMP nanocarriers for biomedical applications. Full article

The market for non-concentrated (NFC) orange juice is increasing rapidly due to consumer demand for nutrients and flavor. However, it encounters challenges in microbial safety, particularly from Salmonella enterica and Listeria monocytogenes. This study aimed to exploit a bio-preservative for NFC orange juice. Results showed that the cyclic peptide cyclo-zp80r had good antibacterial activity, with minimum inhibitory concentration values of 2–8 μM against S. enterica and L. monocytogenes. It exhibited bactericidal action against S. enterica and bacteriostatic action against L. monocytogenes at a concentration of 128 μM. This study explored the effect of cyclo-zp80r on the pathogenicity of S. enterica and L. monocytogenes. The mortality rate of Galleria mellonella exposed to these pathogens in NFC orange juice decreased from 100% to 60% after cyclo-zp80r treatment, surpassing the effectiveness of nisin. Cyclo-zp80r exhibited depolarization effects on S. enterica and L. monocytogenes. It increased outer membrane permeability and damaged the membrane structure of S. enterica. Cyclo-zp80r also caused distinct morphological changes, mainly cell collapse in S. enterica and localized bubble-like protrusions in L. monocytogenes. It induced reactive oxygen species production and DNA binding. The species diversity and abundance in NFC orange juice were also reduced by cyclo-zp80r, particularly in the genera Pantoea, Aeromonas, Pseudomonas, and Erwinia. Additionally, cyclo-zp80r exhibited excellent stability at high temperature (121 °C, 5 min) and in fresh orange juice. These results suggest that cyclo-zp80r could be developed as an effective food bio-preservative. Full article

Background/Objective: Programmed cell death ligand-1 (PD-L1), which is overexpressed in certain tumors, inhibits the body’s natural immune response by providing an “off” signal that enables cancer cells to evade the immune system. It has been demonstrated that [¹⁷⁷Lu]Lu-DOTA-iPD-L1 (PD-L1 inhibitor cyclic peptide) promotes immune responses. This study aimed to synthesize and evaluate [¹⁸F]AlF-NOTA-iPD-L1 as a novel radiotracer for PD-L1 positron emission tomography (PET) imaging and as a potential theranostic pair for [¹⁷⁷Lu]Lu-DOTA-iPD-L1. Methods: The NOTA-iPD-L1 peptide conjugate was synthesized and characterized by U.V.-vis, I.R.-FT, and UPLC-mass spectroscopies. Radiolabeling was performed using [¹⁸F]AlF as the precursor, and the radiochemical purity (HPLC), partition coefficient, and serum stability were assessed. Cellular uptake and internalization (in 4T1 triple-negative breast cancer cells), binding competition, immunofluorescence, and Western blot assays were applied for the radiotracer in vitro characterization. Biodistribution in mice bearing 4T1 tumors was performed, and molecular imaging (Cerenkov images) of [¹⁸F]AlF-NOTA-iPD-L1 and [¹⁷⁷Lu]Lu-DOTA-iPD-L1 in the same mouse was obtained. Results: [¹⁸F]AlF-NOTA-iPD-L1 was prepared with a radiochemical purity greater than 97%, and it demonstrated high in vitro and in vivo stability, as well as specific recognition by the PD-L1 protein (IC₅₀ = 9.27 ± 2.69 nM). Biodistribution studies indicated a tumor uptake of 6.4% ± 0.9% ID/g at 1-hour post-administration, and Cerenkov images showed a high tumor uptake of both [¹⁸F]AlF-NOTA-iPD-L1 and ¹⁷⁷Lu-iPD-L1 in the same mouse. Conclusions: These results warrant further studies to evaluate the clinical usefulness of [¹⁸F]AlF-NOTA-iPD-L1/[¹⁷⁷Lu]Lu-DOTA-iPD-L1 as a radiotheranostic pair in combination with anti-PD-L1/PD1 immunotherapy. Full article

Phage display has advanced the discovery of peptides that selectively bind to a wide variety of cell surface molecules, offering new modalities to modulate disease-related protein–protein interactions (PPIs). These cell-binding peptides occupy a unique pharmaceutical space between small molecules and large biologics, and their growing popularity has opened up new avenues for targeting cell surface proteins that were previously considered undruggable. This work provides an overview of methods for identifying cell-selective peptides using phage display combinatorial libraries, covering in vitro, ex vivo, and in vivo biopanning approaches. It addresses key considerations in library design, including the peptide conformation (linear vs. cyclic) and length, and highlights examples of clinically approved peptides developed through phage display. It also discusses the on-phage chemical cyclization of peptides to overcome the limitations of genetically encoded disulfide bridges and emphasizes advances in combining next-generation sequencing (NGS) with phage display to improve peptide selection and analysis workflows. Furthermore, due to the often suboptimal binding affinity of peptides identified in phage display selections, this article discusses affinity maturation techniques, including random mutagenesis and rational design through structure–activity relationship (SAR) studies to optimize initial peptide candidates. By integrating these developments, this review outlines practical strategies and future directions for harnessing phage display in targeting challenging cell surface proteins. Full article

Right heart failure (RHF) is a major cause of morbidity and mortality, often resulting from pulmonary arterial hypertension and characterized by impaired calcium (Ca²⁺) handling and maladaptive remodeling. Phosphodiesterase 9A (PDE9A), a cGMP-specific phosphodiesterase, has been proposed as a potential contributor to RHF pathogenesis by suppressing the cardioprotective cGMP–PKG signaling pathway—a conclusion largely extrapolated from left-sided heart failure models. This review examines existing evidence regarding PDE9A’s role in RHF, focusing on its effects on intracellular calcium cycling, fibrosis, hypertrophy, and contractile dysfunction. Data from preclinical models demonstrate that pathological stress upregulates PDE9A expression in cardiomyocytes, leading to diminished PKG activation, impaired SERCA2a function, RyR2 instability, and increased arrhythmogenic Ca²⁺ leak. Pharmacological or genetic inhibition of PDE9A restores cGMP signaling, improves calcium handling, attenuates hypertrophic and fibrotic remodeling, and enhances ventricular compliance. Early-phase clinical studies in heart failure populations suggest that PDE9A inhibitors are well tolerated and effectively augment cGMP levels, although dedicated trials in RHF are still needed. Overall, these findings indicate that targeting PDE9A may represent a promising therapeutic strategy to improve outcomes in RHF by directly addressing the molecular mechanisms underlying calcium mishandling and myocardial remodeling. Full article

Background: Metastatic prostate cancer (mPrC), with a median survival of under 2 years, represents an important unmet medical need which may benefit from the development of more effective targeted drug delivery systems. Several cell surface receptors have been identified as candidates for targeted drug delivery to mPrC cells; however, these receptors were selected for their overabundance on PrC cells rather than for their suitability for targeted delivery and uptake of cytotoxic drug payloads. Methods: We describe a novel, unbiased strategy to isolate peptides that fulfill functional criteria required for effective intracellular drug delivery and the specific cytotoxicity of PrC cells without prior knowledge of the targeted receptor. Phage clones displaying 7-mer cyclic peptides were negatively selected in vivo and then positively biopanned through a series of parent and drug-resistant mPrC cells. Peptides from the internalized clones were then subjected to a panel of biochemical and functional tests that led to the selection of several peptide candidates. Results: The selected peptides do not bind PSMA. Peptide-drug conjugates (PDCs) incorporating one of the peptides selectively killed wild-type and drug-resistant PrC cell lines and patient PrC cells but not normal prostate tissue cells in vitro. The PDC also halted the growth of PC3 tumors in a xenograft model. Conclusions: Our study demonstrates that adding unbiased, functional criteria into drug carrier selection protocols can lead to the discovery of novel peptides with appropriate properties required for effective targeted drug delivery into target cancer cells. Full article

This research represents the first application of the MOLS method to characterize the conformational energy landscape of an antimicrobial peptide within a solvent environment, providing a novel approach to understanding peptide behavior in solution. This method’s exhaustive nature ensures that all minimum-energy conformations for a given amino acid sequence are sampled. In this work, we employed a combination of MOLS and VMD software to generate structural models of a cyclic peptide, both solvated and non-solvated, and then utilized the CHARMM force field to conduct energy calculations throughout the sampling process. In the presence of a solvent, this method predicted a structure close to the experimental crystal structure. A significant reduction was observed in gamma turn motifs in the presence of water. The solvent molecules also favored different hydrogen bonding patterns in the peptide by orchestrating an intermolecular interaction with the peptide atoms. This intermolecular interaction involves an ARG side chain and further stabilizes the backbone. It is evident that solvent interactions are key in designing antimicrobial peptides. This study will help in designing and understanding peptides for use as therapeutic agents like antibacterial or antimicrobial peptides. Each conformer obtained from the MOLS method would be one of the best starting points for molecular dynamic simulation to further explore the landscape. Full article

Background: Cell-penetrating peptides cross cell membrane barriers while carrying cargoes in a functional form. Our work identified two novel lung-targeting peptides, S7A and R11A. Here, we present studies on biodistribution, the cell types targeted, and an in vitro proof of application. Methods: Studies were performed in human bronchial epithelial cells (HBECs) with and without various endocytic inhibitors, and coincubation with fluorescently labeled transferrin or endocytic markers. Cyclic R11A (cR11A) was conjugated to siRNA duplexes and anti-viral activity against SARS-CoV-2 was tested. Biodistribution studies were performed by injecting wild-type mice with fluorescently labeled peptides, and various circulation times were allowed for, as well as cross-staining of lung sections or isolated single cells with various cellular markers, followed by fluorescence-activated cell sorting or confocal microscopy. Results: cR11A showed peak uptake in 15 min, with the highest uptake in airway epithelial type II (ATII) cells, followed by p63+ basal cells and ionocytes. Cyclization increased transduction efficiencies ~100-fold. Endocytosis studies showed a decrease in peptide uptake by pre-treatment with Pitstop2 but not Amiloride or Nystatin. Endocytic marker Lamp1 showed colocalization at the earliest time point, with the escape of the peptide from endocytic vesicles later. cR11A conjugated to ant-spike and anti-envelop proteins showed anti-viral effects with an EC₉₀ of 0.6 μM and 1.0 µM, respectively. Conclusions: We have identified a novel peptide, cR11A, that targets ATII, basal cells, and ionocytes, the cyclization of which increased transduction efficiency in vitro and in vivo. The uptake mechanism appears to be via clathrin-mediated endocytosis with escape from endocytic vesicles. cR11A can act as a vector to deliver anti-viral siRNA to epithelial cells. Full article

G-protein-coupled receptors (GPCRs) are a group of various membrane proteins that mediate essential physiological processes by translating extracellular signals into intracellular responses. The β2-Adrenergic Receptor (β2-AR), a key GPCR, plays a critical role in smooth muscle relaxation, bronchodilation, and cardiovascular function, making it an important therapeutic target for diseases such as asthma and hypertension. Diketopiperazines (DPKs), as cyclic peptides, have shown promise as scaffolds for inhibiting protein interactions and modulating receptor activity, offering a potential alternative to traditional small-molecule inhibitors with reduced side effects. In this study, five DPK derivatives were selected from the PubChem database and evaluated for their binding affinity to the 3D structure of β2-AR (PDB ID = 2RH1) through molecular docking studies using AutoDock 4.6 and MGLTools. The binding energy and hydrogen bond formation of each compound were evaluated to determine their interaction efficiency. Among the compounds, tryptophan-proline diketopiperazine (compound 3) exhibited the highest binding affinity with a binding energy of −5.89 kcal/mol. This enhanced interaction is attributed to the aromatic nature of tryptophan, which promotes strong π-π stacking interactions, and the rigidity of proline, which optimally fits within the receptor’s binding pocket. Hydrophobic interactions further stabilized the complex. These findings highlight compound 3 as a promising β2-AR modulator, providing valuable insights for the design of peptide-based inhibitors targeting β2-AR-related pathologies. Full article

Objectives: This study compared the frequencies of circulating CD4+ T helper (Th) cell subsets in rheumatoid arthritis (RA) patients and healthy controls (HCs) and investigated their relationship with RA disease activity. Methods: Peripheral blood samples and demographic/clinical data were collected from 75 RA patients and 28 HCs from the A3BC Biobank. Flow cytometry was utilized to identify cell subsets. Data were analyzed using FlowJo and GraphPad Prism. Results: RA patients displayed altered Th cell subset frequencies compared with HCs, including a higher overall proportion of Th cells (p = 0.02) but lower proportions of memory Th (p = 0.03), Th1 (p < 0.0001), and Th17.1 (p = 0.004) cells. In DMARD-naïve RA patients (n = 16), lower proportions of Th1 (p = 0.0005), Th9 (p = 0.04), and Th17.1 (p = 0.003) cells, alongside a higher Th17 cell proportion (p = 0.017), were observed compared with those in HCs. Further analysis of matched treatment-naïve, new-onset RA patients and HCs confirmed these findings. Within the RA cohort, lower proportions of Th1 (p = 0.002) and Th17.1 (p = 0.025) cells and a higher proportion of Th2 cells (p = 0.015) were correlated with increased disease activity. Inverse correlations were also found between the proportions of Th1 (p = 0.002), Th9 (p = 0.024), and Th17.1 (p = 0.00017) cells and CRP levels in RA patients. Conclusions: This study demonstrates an imbalance in circulating Th cell subset frequencies in RA patients compared with those in HCs, with notably lower Th1 and Th17.1 cell proportions in RA patients. Decreased frequencies of these cell subsets were linked to increased disease activity, indicating that restoring the balance of Th cell subsets could be a potential therapeutic strategy for RA. Full article

Developing appropriate methods for real-time in situ investigation of how drugs influence signaling pathways and related biomolecules holds enormous potential for helping to provide an understanding of how anticancer drugs exert their effects. Herein, we report a smart nanoprobe, PDA-MB (DOX)-Pep, constructed on the basis of polydopamine nanoparticles (PDA NPs) modified with a dense shell of molecular beacon (MB) with embedded doxorubicin (DOX) and peptide, which can respond specifically to miRNA-34a and Caspase-3 targets. Intracellular experiments demonstrated that, in comparison to the control nanoprobe PDA-MB-Pep, the smart nanoprobe could selectively respond to miRNA-34a, facilitating the release of the embedded DOX. The released DOX subsequently activated the p53 pathway, which further upregulated miRNA-34a expression, leading to additional DOX release. This initiated a cyclical process involving the probe’s response to miRNA-34a, DOX release, p53 activation, and miRNA-34a upregulation, ultimately enhancing cell apoptosis and increasing Caspase-3 expression. The designed smart nanoprobe offers a visual approach to explore how anticancer drugs influence signaling pathways and related molecules at the cellular level. Full article

Radix Pseudostellariae (Tai-Zi-Shen), the dried tuberous root of the Caryophyllaceae plant Pseudostellaria heterophylla (Miq.) Pax (P. heterophylla), has been widely used in traditional Chinese medicine (TCM) for thousands of years. It is slightly bitter, neutral in nature, associated with the spleen and lung meridians, and used for nourishing qi, invigorating the spleen, as well as promoting body fluid production and moistening the lungs. In recent years, with the development in natural product chemistry, cyclic peptides, as some of the active constituents derived from P. heterophylla, have gained increasing attention. These cyclic peptides demonstrate a broad range of biological activities, including anticancer, antioxidant, and immunomodulatory effects, as well as cognitive benefits. This review provides an overview of the chemical characteristics and preparation strategies of cyclic peptides from P. heterophylla, and their biological activities and potential mechanisms are also described. The presented work establishes a scientific framework to facilitate the future research and development of P. heterophylla cyclic peptides as potential therapeutic agents. Full article

Background: Addison’s disease (AD) is a rare disorder that often develops in the context of autoimmune polyglandular syndromes. However, the prevalence of rheumatological autoimmune diseases and corresponding autoimmune markers in AD is poorly investigated. Therefore, the present study aims to explore systemic and organ-specific immune markers in a cohort of AD patients from a single tertiary endocrine center. Material and methods: In total, 43 adult AD patients and 31 controls were included in the study. 21-hydroxylase autoantibodies (21OHAb), glutamic acid decarboxylase autoantibodies (GADAbs), zinc transporter-8 autoantibodies (ZnT8Abs), antibodies against nuclear antigens (ANAs), autoantibodies against cyclic citrullinated peptides (CCPAbs), rheumatoid factors (RFs), IgG autoantibodies against cardiolipin (ACLAbs), and autoantibodies against beta-2-Glycoprotein I (β2-GPIAbs) were measured in all participants. Results: An increased prevalence of antibodies against RFs (27.91% vs. 0%, p < 0.001) and ANAs (13.95% vs. 0%, p = 0.037) was found in AD patients compared to controls. Moreover, the titers of 21-hydroxylase and RF antibodies correlated positively (r = +0.269, p = 0.020). The AD patients tended to show an increased prevalence of subthreshold ACL antibody reactivity compared to controls. All patients diagnosed with type 1 diabetes mellitus were GADAb- but not ZnT8Ab-positive. Conclusions: The results show an increased prevalence of ANA and RF positivity in AD patients compared to controls and a significant association between 21-OHAb and RF positivity. ZnT8Ab positivity was not typical for adult AD patients from our ethnic group, while GADAbs were an essential marker for autoimmune diabetes mellitus. Extensive studies in different ethnic groups are needed to establish the clinical significance of various immunological markers for AD comorbidity and the appropriate follow-up protocols for patients with different antibody positivity. Full article

Antimicrobial peptides (AMPs) have emerged as promising candidates in the fight against multidrug-resistant pathogens due to their broad-spectrum antimicrobial activity and low potential for resistance development. However, their clinical application is limited by poor stability and susceptibility to enzymatic degradation. This study aims to address these limitations by synthesizing a series of cyclic hexapeptides using the hydrazide method and evaluating their antimicrobial activity and stability. The hydrazide method facilitated the synthesis of 11 cyclic peptides through a reaction between C-terminal hydrazides and cysteine-containing peptides. Antimicrobial assays showed that Cy-f2 and Cy-f4 exhibited potent inhibitory effects against different kinds of bacteria, including E. coli, Staphylococcus aureus, and S. aureus. Hemolysis assays revealed minimal red blood cell lysis at effective antimicrobial concentrations, indicating good biocompatibility. Stability tests demonstrated improved stability of the cyclic peptides compared to linear counterparts in SGF and 80 °C. In conclusion, the cyclic hexapeptides synthesized in this study demonstrate excellent antimicrobial activity, enhanced stability, and low toxicity, suggesting their potential as new candidates for treating drug-resistant bacterial infections. Full article