Phage display is a powerful technology that has demonstrated great potential in identifying peptides with high binding affinity and specificity toward a broad spectrum of biological targets. The undesired enrichment of nonspecific binders remains a major challenge in phage display selection. The integration of next-generation sequencing (NGS) into phage display has expanded the horizons of ligand discovery by significantly enhancing our ability to interrogate the massive sequence space of combinatorial phage display libraries and providing quantitative information about their composition and evolution during biopanning. NGS findings have provided strong support for the notion that the selection output still contains a large number of nonspecifically enriched peptide sequences that could not be removed or identified by traditional strategies for biopanning optimization. Despite its great potential for increasing the strength of peptide discovery, the routine NGS-based phage display workflow, which relies on analyzing the biopanning output, fails to effectively distinguish thousands of nonspecific peptides from specific target-binding sequences. By incorporating precise control experiments—including the NGS characterization of the unamplified and amplified naïve libraries and the outputs of targetless and replicate selections—alongside the thoughtful data analysis and interpretation, we propose an optimized workflow of NGS-based phage display that would be capable of distinguishing many target-specific peptides from the overwhelming background of nonspecific binders. Applying such a systematic approach will not only advance fundamental research for peptide discovery but also hold promise for the clinic, where these peptides can serve as the foundation for next-generation diagnostic and therapeutic platforms in precision medicine.

From January 2000 to December 2019, the SEER Plus Database collecting human epidermal growth factor receptor 2 (HER2) and hormone receptor (HR; including estrogen receptor [ER] and progesterone receptor [PR]) status for breast cancer (BC) cases. HER2-positive (HER2+) BC is an aggressive type, and HER2-targeted therapies have significantly improved the therapeutic outcome of patients. However, not every HER2+ BC patient achieves optimal benefits from current HER2-targeted therapies. Here, we conducted a detailed analysis to compare the demographic and clinic-pathological characteristics, survival, differential genes and mutations between HR+ and HR– in HER2+ BC patients. In this retrospective cohort study, Joint HR and HER2 status distributions by more than ten specific clinic-pathological characteristics were evaluated by using Pearson’s chi-squared (χ2) test. The transciptome RNA-seqencing (RNA-seq) expression data together with detailed clinic-pathological information of HER2+ BC were from The Cancer Genome Atlas (TCGA) and UCSC Xena. 30,482 (71.02%) patients were identified as HR+ /HER2+ BC and 12,440 (28.98%) patients were identified as HR−/HER2+ BC. HR+ /HER2+ BC patients were more likely to be younger than 50-year-old, white and infiltrating lobular carcinoma history than patients with HR−/HER2+. Patients with HR+ /HER2+ BC had lower risks of breast cancer-specific death and higher overall survival rates. Mast cells were enriched in the HR+ /HER2+ BC group, while plasma cells were more abundant in the HR–/HER2+ BC group.In conclusion, HER2+ BC patients benefit differently from current HER2-directed therapies, maybe partly due to the HR status and gene mutations, and they may provide potentially prognostic and predictive value and new treatment strategies for clinicians.Supplementary InformationThe online version contains supplementary material available at 10.1007/s00726-025-03494-x.

Antibiotic resistance is a growing problem in the treatment of life-threatening diseases. Recently, a variety of covalent drugs have emerged. Cysteine ​​is one of the least abundant amino acids in the proteins of many organisms, and the thiol group in its structure makes it unique and has become a common covalent amino acid residue in covalent drug development. Therefore, it is important to conduct research on cysteine ​​derivatives. In this study, the in vitro antibacterial activity of L-cysteine esters were tested against Gram-positive Staphylococcus aureus and Gram-negative Escherichia coli using the modified microdilution broth method. In vitro cytotoxic activities of the esters were carried out against the healthy HEK293T cell line and well-differentiated liver cancer cell lines PLC/PRF/5 and HEP3B at different concentrations by using an MTS assay. In addition, molecular docking studies, ADMET properties, and drug-likeness were also reported. The results obtained are new and it is thought that these results of the study will contribute to the development of new synthesizable cysteine-based drugs. In conclusion, a thorough examination of the frontier orbital (HOMO and LUMO) and MEP studies was conducted using quantum chemistry techniques to determine the molecule’s reactivity, electrophilic and nucleophilic sites.

Celiac disease (CD) is a chronic autoimmune disorder characterized by gluten-induced intestinal inflammation, epithelial barrier dysfunction, and malabsorption of nutrients, including amino acids (AAs). While essential AAs have been extensively studied, the roles of conditionally essential amino acids (CEAAs), cysteine (Cys), and tyrosine (Tyr) in CD pathogenesis remain comparatively underexplored. This review synthesizes current evidence on the contributions of these AAs to intestinal barrier integrity, immune regulation, oxidative stress mitigation, and gut microbiota modulation in CD. Key findings indicate that asparagine (Asn), glycine (Gly), Cys, glutamate (Glu), aspartate (Asp), and glutamine (Gln) contribute to the reinforcement of intestinal barrier integrity. In parallel, Cys and Gly, along with Gln and Asp, play important roles in suppressing oxidative stress, while Gln and Asp have been shown to positively influence the composition and function of the gut microbiota. Serine (Ser) may contribute to gliadin-induced epithelial damage but could also have potential protective effects during treatment. Arginine (Arg), through its metabolism via the inducible nitric oxide synthase (iNOS) and arginase pathways, contributes to immune regulation. This review underscores the therapeutic potential of AAs as adjunctive targets to gluten-free diets (GFD), offering avenues for nutritional interventions, enzyme therapies, and biomarker development. A deeper understanding of AA-mediated mechanisms may pave the way for personalized management strategies to improve clinical outcomes in CD.

ABCA1 is a key protein in maintaining cholesterol homeostasis, and its abnormal expression is associated with the progression of many cancers. Nonetheless, the specific molecular mechanisms by which ABCA1 facilitates the development of LUAD remain largely unexplored, necessitating further in-depth investigation. The TCGA-LUAD database was used to analyze the expression of ABCA1 in LUAD tissues. Subsequently, a cell model with overexpressed ABCA1 was constructed for verification through cell experiments. Cell function was evaluated using the Transwell assay and the colony formation assay. Intracellular cholesterol levels were detected using a kit. At the same time, the online database RM2 Target was employed to predict upstream factors that may have a methylation regulatory relationship with ABCA1. On this basis, Dot blot and MeRIP-qPCR techniques were employed to determine the degree of m6A modification. To clarify the mechanism of IGF2BP1 regulating ABCA1 through the m6A pathway, RNA pull-down binding experiments were carried out, and changes in mRNA stability were assessed using actinomycin D treatment. Finally, the biological function of the IGF2BP1/ABCA1 signaling axis during the growth and metastasis of LUAD in vivo was evaluated by establishing a xenograft animal model. Bioinformatics analysis and cell experimental results confirmed the low expression of ABCA1 in LUAD tissues and cells. ABCA1 significantly inhibited cell proliferation, migration, and invasion capabilities, promoted apoptosis, and reduced intracellular cholesterol levels. From a molecular perspective, IGF2BP1 recognized and bound to methylation sites on ABCA1 mRNA, thereby accelerating its degradation process, resulting in a substantial decrease in the stability of ABCA1 mRNA. Moreover, in vivo and in vitro experiments further confirmed that IGF2BP1 affected cholesterol metabolism by regulating the expression of ABCA1, thereby facilitating the malignant progression of LUAD. Overall, our research revealed that IGF2BP1 affects cholesterol metabolism by reducing the stability of ABCA1 mRNA through m6A modification, thereby boosting the malignant progression of LUAD and formulating a theoretical basis for subsequent LUAD treatment.

Silk glands are modified labial glands that produce silk which has immense commercial importance. Silk is extruded out in liquid form after which the glands undergo autophagy and apoptosis during larval to pupal transition. Biogenic amines, specially spermidine and γ-aminobutyric acid (GABA) are known to play an important role in autophagy. Yet, GABA is not identified in the silk glands till now and therefore its role in autophagy remains unknown. Current study aimed to evaluate role of biogenic amines in the autophagy of silk glands. Fifth instar silkworms were fed with control and spermidine supplemented mulberry leaves under controlled conditions. Qualitative and quantitative analysis of biogenic amines were analyzed in silk glands of control and spermidine fed groups at the end of feeding stage, spinning and pre-pupal stages. Biogenic amines were significantly decreased in the silk glands from feeding stage to non-feeding prepupal stages. Elevated levels of biogenic amines; putrescine, spermidine, and spermine were observed in silk glands at pre-pupal stage in the spermidine fed group. The unknown biogenic amine whose levels were significantly elevated during silk gland degeneration in both control and spermidine fed groups was identified as GABA by spectroscopic techniques. This is the first report of the identification of GABA in the silk glands of Bombyx mori which increased significantly following spermidine supplementation, resulting in elevated levels of calcium deposits, contributing to the early degeneration of the silk glands.

Pseudo-peptides are an important category of biologically active artificial small molecules. To access these important molecules, a novel series of bisazlactones was synthesized via the Erlenmeyer-Plöchl reaction, using glycine- and terephthaloyl-based diacid with aldehydes. These bisazlactones were then utilized as efficient intermediates in reactions with primary and secondary amines, providing novel pseudo-peptides containing enamide groups in high to excellent yields. The selected pseudo-peptide enamides exhibited selective cytotoxicity against hepatocarcinoma cells, while exhibiting negligible impact on normal mammalian cells. Notably, compound 6y displayed superior anti-cancer activity compared to the others.

BackgroundCreatine is a semi-essential nutrient that plays a critical role in energy metabolism, with dietary intake and endogenous synthesis contributing to overall creatine availability. While dietary creatine intake has been studied extensively, limited data exist on the dietary exposure to its precursor amino acids—glycine, arginine, and methionine—and their contribution to endogenous creatine synthesis. This study aimed to assess the dietary intake of these precursors in U.S. children and adults using data from the Third National Health and Nutrition Examination Survey (NHANES III) and to compare endogenous creatine synthesis with direct dietary creatine intake.MethodsWe analyzed NHANES III dietary recall data from 29,945 individuals aged 2 years and older. Intakes of glycine, arginine, methionine, and creatine were calculated per kilogram of body weight. The contribution of precursor amino acids to endogenous creatine synthesis was estimated using established metabolic conversion factors.ResultsThe mean daily intakes of glycine, arginine, methionine, and creatine were 59.6 ± 0.4 mg/kg, 77.2 ± 0.5 mg/kg, 31.9 ± 0.2 mg/kg, and 15.5 ± 0.1 mg/kg, respectively. Estimated endogenous creatine synthesis from precursor amino acids was significantly greater than dietary creatine intake across all age groups (P < 0.01), with precursor-derived creatine production averaging 41.9 ± 0.3 mg/kg body weight per day, approximately 2.7 times higher than dietary creatine intake. Creatine precursor availability declined with age, with the lowest values observed in individuals aged ≥ 65 years.ConclusionThis study provides the first comprehensive evaluation of total creatine availability in a representative U.S. population, highlighting the predominance of endogenous synthesis over direct dietary intake. These findings suggest that creatine metabolism is largely dependent on precursor amino acid intake and that certain populations, particularly older adults, may be at higher risk for reduced creatine availability. Future research should explore the physiological implications of these findings and potential dietary interventions to optimize creatine status across the lifespan.

To evaluate the potential of antimicrobial peptide CC34 for use as therapeutic agents for gastric cancer SGC-7901 and hepatocellular carcinoma HepG-2. In this study, the antibacterial activity and antibacterial mechanism were tested by the minimum inhibitory concentration (MIC) analysis, minimal bactericidal concentration (MBC) analysis, bacterial biofilm and NaCl permeability assays. Then, we assessed the hemolytic activity and cytotoxicity of CC34 for red blood cells and cancer cells, respectively. Apoptosis assay, cell cycle analysis, determination of intracellular ROS, western blot analysis caspase activity assay and ATP assay were further performed to investigate the mechanism of CC34 affected cancer cells. The novel peptide could inhibit Gram-negative and Gram-positive bacteria, with low hemolytic activity against mouse and chicken erythrocytes. Moreover, CC34 exhibited higher inhibitory activity against biofilm formation. In addition, our data showed that CC34 significantly suppressed cell proliferation, in a dose dependent manner. CC34 induced apoptosis, induced reactive oxygen species (ROS) generation, inhibited B-cell lymphoma-2 (Bcl-2) expression, increase B-cell lymphoma protein 2 associated X protein (Bax) expression, release of cytochrome c (Cyt C), promoted caspase-3 and − 9 activities and reduced cellular ATP levels in cancer cells. Our results indicate that CC34 with antimicrobial activity have a highly potent ability to induced apoptosis via mitochondrial-mediated apoptotic pathway in cancer cells.