Journal: Cellular & Molecular Biology Letters

Article Title: Chicoric acid targets PYGL to normalize glycogenolysis-driven glycolysis to suppress non-small cell lung cancer

doi: 10.1186/s11658-026-00930-y

Figure Lengend Snippet: CA inhibits NSCLC proliferation and glycolysis by directly binding to PYGL. A Workflow for the identification of CA target proteins. B , C Predicted binding sites of CA with PYGL visualized using PyMOL ( B ) and Discovery Studio ( C ). D CETSA validating the physical interaction between CA and PYGL. E SPR analysis of the binding affinity between CA and PYGL. F MD simulations depicting the conformational dynamics of the CA–PYGL complex in aqueous solution over 100 ns. G RMSD of CA relative to PYGL over a 100‑ns trajectory. H RMSF of residues contacting CA. I Time-dependent hydrogen bond formation between CA and catalytic residues of PYGL. J Free energy landscape projection highlighting low-energy conformations of the complex. K , L Binding free-energy components from MM/PBSA analysis (ΔGgas = ΔEvdW + ΔEele; ΔGsolv = ΔEgb + ΔEsurf; ΔGtotal = ΔGgas + ΔGsolv), where ΔEele denotes electrostatic interaction energy, ΔEsurf nonpolar solvation energy, and ΔEgb polar solvation energy. CA chicoric acid, NSCLC non-small cell lung cancer, CETSA Cellular thermal shift assay, SPR Surface plasmon resonance, MD Molecular dynamics, RMSD Root-mean-square deviation, RMSF Root-mean-square fluctuation

Article Snippet: Recombinant PYGL protein (Cat#TMPH-01404, TargetMol, USA) was immobilized onto an active CM5 sensor chip (Cytiva).

Techniques: Binding Assay, Analysis, Thermal Shift Assay, SPR Assay, RNA Sequencing, Docking Assay, Western Blot

Journal: Cellular & Molecular Biology Letters

Article Title: Chicoric acid targets PYGL to normalize glycogenolysis-driven glycolysis to suppress non-small cell lung cancer

doi: 10.1186/s11658-026-00930-y

Figure Lengend Snippet: PYGL drives NSCLC progression via glycogen mobilization and glycolysis reprogramming. A Schematic illustration of PYGL-mediated glycogen mobilization. B , C Cell proliferation assays in H1975 and H460 cells with PYGL knockdown or overexpression. D , E Colony formation assays of H1975 and H460 cells with altered PYGL expression. F GSEA identifying enriched glycolytic pathways with high PYGL expression in the TCGA-LUAD database. G Correlation analysis between PYGL expression levels and glycolysis-related GSVA scores. H , I Measurement of extracellular acidification rate in PYGL-knockdown or -overexpressing cells using the Seahorse XF HS Mini Analyzer. All ECAR values were normalized to the final cell number per well. Left: Representative kinetic profiles of ECAR in response to Rot/AA and 2-DG injections. Right: Quantitative analysis of glycolytic parameters, including basal glycolysis, compensatory glycolysis, glycolytic reserve, and nonglycolytic acidification. J Quantification of lactate production and glucose consumption in PYGL-knockdown cells. K 2-Deoxy-D-glucose reverses PYGL-driven proliferation and glycolysis. (Left) Cell proliferation; (median) lactate production; (Right) glucose consumption. L Proliferation of Vector and PYGL-overexpressing H1975 and H460 cells cultured under normal or glucose deprivation (GD) conditions. M Glycogen accumulation detected by PAS staining in H1975 and H460 cells with PYGL knockdown or overexpression (scale bar, 50 μm). Right: Quantification of PAS staining intensity. N Glycogen concentration in PYGL-knockdown or -overexpressing cells. O Glycogen accumulation in PYGL-depleted xenograft models (scale bar, 100 μm). P Cell proliferation (Left), lactate production (median), and glucose consumption (Right) in PYGL-overexpressing or vector cells treated with CP-91149 (20 μM; n = 3). Data are expressed as mean ± SD. Statistical significance levels are indicated as follows: * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001 versus respective controls. CA chicoric acid, NSCLC non-small cell lung cancer, GSEA Gene Set Enrichment Analysis, TCGA The Cancer Genome Atlas, LUAD lung adenocarcinoma, GSVA Gene Set Variation Analysis, PAS Periodic Acid-Schiff

Article Snippet: Recombinant PYGL protein (Cat#TMPH-01404, TargetMol, USA) was immobilized onto an active CM5 sensor chip (Cytiva).

Techniques: Knockdown, Over Expression, Colony Assay, Expressing, Analysis, Plasmid Preparation, Cell Culture, Staining, Concentration Assay, Xenograft Assay, Gene Expression

Journal: Cellular & Molecular Biology Letters

Article Title: Chicoric acid targets PYGL to normalize glycogenolysis-driven glycolysis to suppress non-small cell lung cancer

doi: 10.1186/s11658-026-00930-y

Figure Lengend Snippet: CA promotes glycogen accumulation and disrupts the PYGL–LDHA axis to accelerate proteasomal degradation of LDHA. A Nonreducing gradient gels assessing PYGL polymerization following CA treatment for 48 h. B PAS staining of CA-treated PYGL-overexpressing cells for 48 h (scale bar, 50 μm; n = 3). C Volcano plot of RNA-seq data showing differentially expressed genes following PYGL knockdown in H1975 cells. D , E Cell viability of H1975 and H460 cells was measured over 3 days following PYGL depletion ( D ) or CA treatment ( E ), with or without LDHA overexpression. F Endogenous co-immunoprecipitation (Co-IP) of PYGL with LDHA. G Cycloheximide (50 μg/mL) chase of LDHA in control (shNC) and PYGL-knockdown (shPYGL) cells. H LDHA protein levels in shPYGL cells treated with or without MG132 (10 μM, 2 h). I LDHA expression in NSCLC cells treated with increasing concentrations of CA for 48 h. J LDHA levels in CA-treated cells in the presence or absence of MG132 (10 μM, 2 h). K Endogenous Co-IP assay of PYGL and LDHA in cells treated with CA (100 μM, 48 h). L LDHA abundance in PYGL-overexpressing cells treated with or without CA (100 μM, 48 h). M Computational molecular docking model illustrating the spatial binding configuration among CA, PYGL, and LDHA. B , D , and E Data are presented as mean ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001 versus controls. G – L Western blot analyses and corresponding densitometric quantifications. All bar graphs adjacent to the immunoblots represent the relative protein levels from three independent biological replicates. Data are presented as mean ± SEM. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. CA chicoric acid, LDHA lactate dehydrogenase A, PAS Periodic Acid-Schiff, Co-IP assay Co-immunoprecipitation assay

Article Snippet: Recombinant PYGL protein (Cat#TMPH-01404, TargetMol, USA) was immobilized onto an active CM5 sensor chip (Cytiva).

Techniques: Staining, RNA Sequencing, Knockdown, Over Expression, Co-Immunoprecipitation Assay, Control, Expressing, Docking Assay, Binding Assay, Western Blot, Clear Native PAGE, SDS Page

Journal: Cellular & Molecular Biology Letters

Article Title: Chicoric acid targets PYGL to normalize glycogenolysis-driven glycolysis to suppress non-small cell lung cancer

doi: 10.1186/s11658-026-00930-y

Figure Lengend Snippet: PYGL expression determines sensitivity to CA treatment. A , B Cell proliferation of PYGL knockdown of CA treatment versus controls. C , D Cell proliferation in PYGL-overexpressing cells treated with CA versus controls. E , F Time-course proliferation analysis (24, 48, 72 h) of H1975 and H460 cells treated with 100 μM CA following PYGL knockdown or overexpression. G – I Flow cytometric analysis of apoptosis in CA-treated (100 μM, 48 h) H1975 and H460 cells with PYGL knockdown or overexpression. J , K Quantification of lactate production and glucose consumption in PYGL-manipulated cells following CA treatment (100 μM CA; 48 h). Data are presented as mean ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001 versus controls. CA chicoric acid, LDHA lactate dehydrogenase A, PAS Periodic Acid-Schiff, Co-IP assay Co-immunoprecipitation assay, ECAR Extracellular acidification rate

Article Snippet: Recombinant PYGL protein (Cat#TMPH-01404, TargetMol, USA) was immobilized onto an active CM5 sensor chip (Cytiva).

Techniques: Expressing, Sensitivity, Knockdown, Analysis, Over Expression, Co-Immunoprecipitation Assay, Cell Viability Assay, FACS

Journal: Cellular & Molecular Biology Letters

Article Title: Chicoric acid targets PYGL to normalize glycogenolysis-driven glycolysis to suppress non-small cell lung cancer

doi: 10.1186/s11658-026-00930-y

Figure Lengend Snippet: Glu162, Arg247, and Glu273 of PYGL are required for its binding to CA. A Alanine scanning mutagenesis quantified changes in CA–PYGL binding affinity upon residue substitution. B RMSD trajectories from 100-ns MD simulations of CA bound to WT or mutant PYGL. C RMSF profiles of CA during simulations with WT versus mutant PYGL. D Time-dependent hydrogen bond formation between CA and catalytic residues in WT versus mutant PYGL. E Distribution of hydrogen bond distances between CA and catalytic residues in WT and mutant PYGL simulations. F , G Hydrophilic ( F ) and hydrophobic ( G ) interaction landscapes of CA with WT and mutant PYGL catalytic residues across the 100‑ns simulations. H Comparison of hydrogen-bonding networks between CA and catalytic residues in WT versus mutant PYGL complexes (CA, red; interacting residues, yellow). I Schematic representation of PYGL point mutations. J – L Cell proliferation and colony formation assays in cells expressing PYGL-WT, PYGL-E162A, D181A, R247A, E273A, or E276A. Data are presented as mean ± SD. * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001 versus controls. CA chicoric acid, RMSD root-mean-square deviation, RMSF root-mean-square fluctuation, WT wild type

Article Snippet: Recombinant PYGL protein (Cat#TMPH-01404, TargetMol, USA) was immobilized onto an active CM5 sensor chip (Cytiva).

Techniques: Binding Assay, Mutagenesis, Residue, Comparison, Colony Assay, Expressing, Docking Assay, Cell Viability Assay

Journal: Cellular & Molecular Biology Letters

Article Title: Chicoric acid targets PYGL to normalize glycogenolysis-driven glycolysis to suppress non-small cell lung cancer

doi: 10.1186/s11658-026-00930-y

Figure Lengend Snippet: PYGL mediates CA-induced tumor growth inhibition in vivo. A Experimental strategy for the establishment of tumor xenografts and drug treatment. NS, normal saline; i.p., intraperitoneal injection. B – F Tumor growth kinetics of H1975 xenografts with indicated PYGL levels and CA treatment. G Tumor volume response to CA treatment across PYGL-modulated xenografts. H Tumor biomarker and safety assessments. Top: Immunohistochemical staining for PYGL, Ki67, cleaved caspase-3, and LDHA in tumor sections ( n = 3). Middle: H&E staining of major organs (heart, liver, spleen, kidney) from CA-treated mice ( n = 3). Bottom: Glycogen accumulation assessed by PAS staining following PYGL overexpression and CA treatment, with quantification of staining intensity ( n = 3). All scale bars: 100 μm. Statistical significance versus controls is denoted as * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001. CA chicoric acid, H&E hematoxylin and eosin, PAS Periodic Acid-Schiff

Article Snippet: Recombinant PYGL protein (Cat#TMPH-01404, TargetMol, USA) was immobilized onto an active CM5 sensor chip (Cytiva).

Techniques: Inhibition, In Vivo, Saline, Injection, Biomarker Discovery, Immunohistochemical staining, Staining, Over Expression, Xenograft Assay, Immunohistochemistry, H&E Stain

Journal: Cellular & Molecular Biology Letters

Article Title: Chicoric acid targets PYGL to normalize glycogenolysis-driven glycolysis to suppress non-small cell lung cancer

doi: 10.1186/s11658-026-00930-y

Figure Lengend Snippet: PYGL is transcriptionally regulated by E2F1. A Venn diagram illustrating the overlap of predicted PYGL-regulating transcription factors from multiple databases. B KEGG pathway enrichment of PYGL-associated genes. C E2F1 ChIP-seq read density profiles at the PYGL promoter region (ENCODE dataset: ENCSR000EWX). D Prediction of E2F1-binding motifs and sites within the PYGL promoter region (JASPAR database). E PYGL and E2F1 mRNA expression levels in H1975 and H460 cells following treatment with the E2F inhibitor HLM006474 (10 μM, 24 h). Data are presented as mean ± SD. F RT-qPCR analysis of E2F1 and PYGL mRNA levels following E2F1 siRNA knockdown in H1975 and H460 cells. Data are presented as mean ± SD. G Immunoblot analysis and corresponding densitometry quantification of PYGL and E2F1 protein levels after siRNA knockdown in H1975 and H460 cells. Densitometry data are presented as mean ± SEM. H Schematic representation of the E2F1/PYGL axis in lung cancer metabolic reprogramming. Statistical significance versus controls is denoted as * p < 0.05, ** p < 0.01, *** p < 0.001, **** p < 0.0001

Article Snippet: Recombinant PYGL protein (Cat#TMPH-01404, TargetMol, USA) was immobilized onto an active CM5 sensor chip (Cytiva).

Techniques: ChIP-sequencing, Binding Assay, Expressing, Quantitative RT-PCR, Analysis, Knockdown, Western Blot, Real-time Polymerase Chain Reaction, Luciferase

Journal: Cellular & Molecular Biology Letters

Article Title: Chicoric acid targets PYGL to normalize glycogenolysis-driven glycolysis to suppress non-small cell lung cancer

doi: 10.1186/s11658-026-00930-y

Figure Lengend Snippet: Mechanism by which CA targets and inhibits PYGL to remodel metabolism and suppress tumor progression. Left: E2F1 transcriptionally upregulates PYGL expression, which accelerates the glycolytic pathway by promoting glycogenolysis, thereby driving tumor cell proliferation. Right: CA, acting as an allosteric inhibitor, directly binds to PYGL (Glu162, Arg247, Glu273), inducing a conformational change and inhibiting its enzymatic activity, while simultaneously promoting the degradation of the PYGL–LDHA complex. This blocks the abnormally active glycolytic flux, ultimately inducing tumor cell apoptosis and suppressing proliferation

Article Snippet: Recombinant PYGL protein (Cat#TMPH-01404, TargetMol, USA) was immobilized onto an active CM5 sensor chip (Cytiva).

Techniques: Expressing, Activity Assay

Journal: Journal of Cellular and Molecular Medicine

Article Title: Identification and Verification of Necroptosis‐Related Genes in Patients With Sepsis by Bioinformatic Analysis and Molecular Experiments

doi: 10.1111/jcmm.70582

Figure Lengend Snippet: Study flow diagram. This figure was created with BioRender.com. ELISA, enzyme‐linked immunosorbent assay; qPCR, quantitative real‐time polymerase chain reaction; SMC‐RoCI, Samsung Medical Center Registry of Critical Illness.

Article Snippet: The glycogen phosphorylase L (PYGL) levels were measured using the Human PYGL ELISA Kit (Novus Biologicals, Centennial CO, USA).

Techniques: Enzyme-linked Immunosorbent Assay, Real-time Polymerase Chain Reaction

Journal: Journal of Cellular and Molecular Medicine

Article Title: Identification and Verification of Necroptosis‐Related Genes in Patients With Sepsis by Bioinformatic Analysis and Molecular Experiments

doi: 10.1111/jcmm.70582

Figure Lengend Snippet: Comparison of eight differentially expressed genes, grouped into both immune system‐related and necroptosis‐related categories, plus corresponding proteins between the sepsis and control groups.

Article Snippet: The glycogen phosphorylase L (PYGL) levels were measured using the Human PYGL ELISA Kit (Novus Biologicals, Centennial CO, USA).

Techniques: Comparison, Control

Journal: Journal of Cellular and Molecular Medicine

Article Title: Identification and Verification of Necroptosis‐Related Genes in Patients With Sepsis by Bioinformatic Analysis and Molecular Experiments

doi: 10.1111/jcmm.70582

Figure Lengend Snippet: Comparison of the expression levels of necroptosis‐related differentially expressed genes between the sepsis and control groups and between the deceased and surviving patients in the sepsis group (quantitative real‐time polymerase chain reaction analysis).

Article Snippet: The glycogen phosphorylase L (PYGL) levels were measured using the Human PYGL ELISA Kit (Novus Biologicals, Centennial CO, USA).

Techniques: Comparison, Expressing, Control, Real-time Polymerase Chain Reaction

Journal: Journal of Cellular and Molecular Medicine

Article Title: Identification and Verification of Necroptosis‐Related Genes in Patients With Sepsis by Bioinformatic Analysis and Molecular Experiments

doi: 10.1111/jcmm.70582

Figure Lengend Snippet: Comparison of the levels of proteins related to necroptosis‐related differentially expressed genes between the sepsis and control groups and between the deceased and surviving patients in the sepsis group (ELISA analysis).

Article Snippet: The glycogen phosphorylase L (PYGL) levels were measured using the Human PYGL ELISA Kit (Novus Biologicals, Centennial CO, USA).

Techniques: Comparison, Control, Enzyme-linked Immunosorbent Assay

Journal: Journal of Cellular and Molecular Medicine

Article Title: Identification and Verification of Necroptosis‐Related Genes in Patients With Sepsis by Bioinformatic Analysis and Molecular Experiments

doi: 10.1111/jcmm.70582

Figure Lengend Snippet: Graphs showing receiver operating characteristic curves of proteins corresponding to the upregulated and downregulated necroptosis‐related genes for diagnosing sepsis in all study participants ( n = 145) (A) and predicting mortality in patients with sepsis ( n = 133) (B). Protein levels were measured using an enzyme‐linked immunosorbent assay. AUC, area under the curve; CYLD, Cylindromatosis Lysine 63 deubiquitinase; FADD, Fas associated via death domain; FasL, Fas ligand; NLRP6, NLR family pyrin domain containing 6; PYGL, glycogen phosphorylase L; TLR3, toll‐like receptor 3; TNF, tumour necrosis factor.

Article Snippet: The glycogen phosphorylase L (PYGL) levels were measured using the Human PYGL ELISA Kit (Novus Biologicals, Centennial CO, USA).

Techniques: Enzyme-linked Immunosorbent Assay

Journal: Journal of Cellular and Molecular Medicine

Article Title: Identification and Verification of Necroptosis‐Related Genes in Patients With Sepsis by Bioinformatic Analysis and Molecular Experiments

doi: 10.1111/jcmm.70582

Figure Lengend Snippet: Quantification of corresponding proteins to the eight necroptosis‐related differentially expressed genes between patients with sepsis and healthy volunteers. The protein expression levels were measured in three cells, comprising HUVEC, Jurkat T and THP‐1 cells, with media transfected with plasma from sepsis patients and healthy volunteers for 24 h. Statistical significance was determined by comparing the protein expression level in each cell medium between patients with sepsis and healthy volunteers (* p < 0.05 and ** p < 0.01). CYLD, Cylindromatosis Lysine 63 deubiquitinase; FADD, Fas associated via death domain; FasL, Fas ligand; NLRP6, NLR family pyrin domain containing 6; PYGL, glycogen phosphorylase L; TLR3, toll‐like receptor 3; TNF, tumour necrosis factor.

Article Snippet: The glycogen phosphorylase L (PYGL) levels were measured using the Human PYGL ELISA Kit (Novus Biologicals, Centennial CO, USA).

Techniques: Expressing, Transfection, Clinical Proteomics

Journal: Journal of Cellular and Molecular Medicine

Article Title: Identification and Verification of Necroptosis‐Related Genes in Patients With Sepsis by Bioinformatic Analysis and Molecular Experiments

doi: 10.1111/jcmm.70582

Figure Lengend Snippet: Schematic representation of the necroptosis pathway. Five upregulated necroptosis‐related differentially expressed genes that were identified and verified in this study are presented in grey colour. TNFR1 activation by TNF results in the complex formation between RIPK1 and TRADD. Under conditions of caspase‐8 inhibition, necroptosis occurs through the formation of the necrosome, which is a three‐protein complex (RIPK1, RIPK3 and FADD). The necrosome activates MLKL and phospho‐MLKL oligomerizes and translocates into the plasma membrane, where it causes membrane rupture and release of damage‐associated molecular patterns. Alternatively, the activation of Fas or receptor/ligand interaction, such as TLR3/LPS, also results in necroptosis. Casp8, caspase‐8; cFLIP, cellular FLICE (FADD‐like IL‐1β‐converting enzyme)‐inhibitory protein; FADD, Fas‐associated death domain; FasL, Fas ligand; LPS, lipopolysaccharide; MLKL, mixed‐lineage kinase domain‐like; PYGL, glycogen phosphorylase L; RIPK1, receptor‐interacting protein kinase 1; RIPK3, receptor‐interacting protein kinase 3; TLR3, toll‐like receptor 3; TLR4, toll‐like receptor 4; TNF, tumour necrosis factor; TNFR1, TNF receptor 1; TRADD, TNFR1‐associated death domain; TRAM, TRIF‐related adaptor molecule; TRIF, TIR domain‐containing adaptor protein.

Article Snippet: The glycogen phosphorylase L (PYGL) levels were measured using the Human PYGL ELISA Kit (Novus Biologicals, Centennial CO, USA).

Techniques: Activation Assay, Inhibition, Clinical Proteomics, Membrane

Journal: Nature metabolism

Article Title: Spatial hepatocyte plasticity of gluconeogenesis during the metabolic transitions between fed, fasted and starvation states

doi: 10.1038/s42255-025-01269-y

Figure Lengend Snippet: ( a, b ) Compass-score differential activity analysis of the Vitamin A pathway between 0 h and 8 h pericentral/periportal hepatocytes ( A ) and between 0 h and 24 h pericentral/periportal hepatocytes ( B ) based on Cohen’s D analysis. BCDO (BCMO1) is highlighted. ( c, d ) Compass-score differential activity analysis of the Vitamin B6 pathway between 0 h and 8 h pericentral/periportal hepatocytes ( C ) and between 0 h and 24 h pericentral/periportal hepatocytes ( D ) based on Cohen’s D analysis. PNPO is highlighted. ( e, f ) Compass-score differential activity analysis of the Starch and sucrose (glycogen) pathway between 0 h and 8 h pericentral/periportal hepatocytes ( E ) and between 0 h and 24 h pericentral/periportal hepatocytes ( F ) based on Cohen’s D analysis. GLPASE (PYGL) is highlighted. ( g ) Representative immunoblotting of BCMO1, PNPO, PYGL, CYP2E1, E-Cadherin, and Vinculin from pericentral and periportal isolated hepatocytes.

Article Snippet: Commercially available primary antibodies were purchased for the detection of PEPCK (Abcam, no. ab70358; 1:2,000), CYP2E1 (Invitrogen, no. PA5–52652; 1:1,000), glutamine synthetase (Invitrogen, no. MA5–27749; 1:8,000), E-cadherin (Cell Signaling Technology, no. 3195; 1:2,000), GLS2 (Abcam, no. ab113509; 1:1,500), BCMO1 (Invitrogen, no. PA5–90759; 1:1,000), PNPO (Proteintech, no. 15552–1-AP; 1:1,000), PYGL (Santa Cruz, no. sc-517597; 1:1,000) and vinculin (Santa Cruz, no. sc-73614; 1:1,000).

Techniques: Activity Assay, Starch, Western Blot, Isolation

Journal:

Article Title: Dynamics of intra-follicular glucose during luteinization of macaque ovarian follicles

doi: 10.1016/j.mce.2010.10.011

Figure Lengend Snippet: Gene detection probe / primer sequences

Article Snippet: PYGL , Assay on Demand , Hs00161132_m1.

Techniques: Sequencing, Thrombin Time Assay, G6PD Assay