{"gene":"PFKFB3","run_date":"2026-06-10T06:43:35","timeline":{"discoveries":[{"year":2013,"finding":"PFKFB3 controls endothelial cell filopodia/lamellipodia formation and directional migration by compartmentalizing with F-actin in motile protrusions, and overrides the pro-stalk activity of Notch signaling to regulate vessel branching tip/stalk cell competition.","method":"Loss-of-function (siRNA/genetic KO), mosaic in vitro and in vivo sprouting assays, subcellular co-localization with F-actin, epistasis with Notch pathway","journal":"Cell","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (genetic KO, mosaic assays, localization, epistasis), replicated in vitro and in vivo, high-citation landmark study","pmids":["23911327"],"is_preprint":false},{"year":2002,"finding":"PFKFB3 (iPFK-2) is induced by hypoxia in cancer cells and its expression and fructose-2,6-bisphosphate levels increase specifically during S phase of the cell cycle, functionally coupling glycolytic activation to cell cycle progression.","method":"In situ hybridization, immunohistochemistry, cell-cycle fractionation, hypoxia treatment of cultured cells","journal":"Cancer research","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — multiple cell-based methods in single lab; no in vitro reconstitution or mutagenesis","pmids":["12384552"],"is_preprint":false},{"year":2015,"finding":"During mitotic arrest, AMPK directly phosphorylates PFKFB3, increasing glycolysis to replace lost oxidative phosphorylation (caused by mitophagy-dependent mitochondrial loss), thereby promoting cell survival. PFKFB3 protein levels also increase due to mitotic-specific translational activation of its mRNA.","method":"AMPK kinase assay, phospho-specific immunoblotting, siRNA knockdown, autophagy induction/inhibition, breast cancer cell viability assays","journal":"Nature cell biology","confidence":"High","confidence_rationale":"Tier 2 / Strong — direct phosphorylation demonstrated, multiple orthogonal methods (kinase assay, KD, translational activation), published in Nature Cell Biology","pmids":["26322680"],"is_preprint":false},{"year":2020,"finding":"The lncRNA AGPG binds to and stabilizes PFKFB3 protein by preventing APC/C-mediated ubiquitination, thereby protecting PFKFB3 from proteasomal degradation and leading to accumulation of PFKFB3 and enhanced glycolytic flux in cancer cells.","method":"Co-immunoprecipitation, ubiquitination assay, proteasome inhibition, siRNA/overexpression, PDX tumor models","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP, ubiquitination assay, multiple orthogonal methods, in vivo PDX validation","pmids":["32198345"],"is_preprint":false},{"year":2018,"finding":"Cisplatin induces acetylation of PFKFB3 at lysine 472 (K472), which impairs its nuclear localization signal (NLS) activity, causing cytoplasmic accumulation of PFKFB3. Cytoplasmic PFKFB3 is then phosphorylated by AMPK, leading to PFKFB3 activation and enhanced glycolysis that protects cells from DNA damage-induced apoptosis.","method":"Site-directed mutagenesis (K472 acetylation-dead mutant), subcellular fractionation, AMPK phosphorylation assay, siRNA knockdown, xenograft model","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — mutagenesis of specific residue combined with fractionation and kinase assay, in vivo validation, rigorous mechanistic dissection","pmids":["29410405"],"is_preprint":false},{"year":2014,"finding":"PFKFB3 promotes cell cycle progression and suppresses apoptosis via its product F2,6BP activating Cdk1, which phosphorylates p27 (T187) leading to p27 ubiquitination and proteasomal degradation. siRNA silencing of PFKFB3 inhibits Cdk1 activity, stabilizes p27, and causes G1/S arrest; co-silencing of p27 reverses these effects.","method":"siRNA silencing, co-siRNA epistasis, Cdk1 kinase assay, p27 protein stability/ubiquitination assay, flow cytometry (cell cycle/apoptosis), HeLa cells","journal":"Cell death & disease","confidence":"High","confidence_rationale":"Tier 2 / Strong — epistasis experiment (rescue by co-KD), kinase activity assay, multiple readouts, mechanistic pathway established","pmids":["25032860"],"is_preprint":false},{"year":2012,"finding":"NMDAR activation stabilizes PFKFB3 protein in neurons (normally degraded by APC/C-Cdh1 proteasomal pathway via its KEN motif) and promotes PFKFB3 release from the nucleus to cytosol, switching neuronal metabolism from pentose-phosphate pathway (PPP) to glycolysis, causing oxidative stress and apoptotic neuronal death.","method":"NMDAR stimulation in cortical neurons, APC/C-Cdh1 KEN-motif deletion mutant, subcellular fractionation, PPP/glycolysis flux measurements, overexpression of G6PD rescue experiment","journal":"Cell death and differentiation","confidence":"High","confidence_rationale":"Tier 1-2 / Strong — mutagenesis (KEN motif deletion), pathway epistasis (G6PD rescue), subcellular fractionation with functional consequence, multiple orthogonal approaches","pmids":["22421967"],"is_preprint":false},{"year":2018,"finding":"PFKFB3 rapidly relocates into ionizing radiation-induced nuclear foci in an MRN-ATM-γH2AX-MDC1-dependent manner and is critical for recruitment of homologous recombination (HR) repair proteins and HR activity. PFKFB3 enzymatic activity is required for ribonucleotide reductase M2 (RRM2) recruitment, deoxynucleotide incorporation during DNA repair, and maintenance of dNTP levels.","method":"Immunofluorescence co-localization with DNA damage/HR foci, pharmacological inhibitor (KAN0438757), siRNA, HR reporter assay, dNTP level measurements, radiosensitization assays in transformed vs. non-transformed cells","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple orthogonal methods (co-localization, HR assay, dNTP measurement, selective inhibitor), functional mechanistic dissection","pmids":["30250201"],"is_preprint":false},{"year":2006,"finding":"siRNA silencing of PFKFB3 in HeLa cells decreases fructose-2,6-bisphosphate, lactate, and ATP, leading to reduced cell viability, cell cycle delay, increased apoptosis, and inhibition of anchorage-independent growth.","method":"siRNA knockdown, metabolite measurement (F2,6BP, lactate, ATP), flow cytometry (cell cycle, apoptosis), soft agar colony formation","journal":"FEBS letters","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — clean KD with defined metabolic and proliferative phenotype, single lab, multiple readouts","pmids":["16698023"],"is_preprint":false},{"year":2020,"finding":"CPEB4 binds cytoplasmic polyadenylation elements (CPEs) in the 3'-UTR of PFKFB3 mRNA to promote its polyadenylation and translational upregulation (not transcriptional), thereby increasing PFKFB3 protein and glycolysis during hepatic stellate cell activation and liver fibrosis.","method":"CPEB4 siRNA knockdown, CPEB4-KO mice, RIP/pulldown of PFKFB3 mRNA by CPEB4, mRNA polyadenylation assay, PFKFB3 protein levels, primary HSC and LX2 cells","journal":"Gastroenterology","confidence":"High","confidence_rationale":"Tier 2 / Strong — direct RNA binding demonstrated (RIP), CPEB4-KO in vivo validation, distinction from transcriptional regulation made explicitly, multiple cell types and models","pmids":["32169429"],"is_preprint":false},{"year":2020,"finding":"c-Src phosphorylates PFKFB3 at tyrosine 194 (Tyr194), activating PFKFB3 and stimulating glycolysis. PFKFB3-Y194F knockin mice show impaired glycolysis and attenuated spontaneous colon tumor formation when crossed with APCmin/+ mice.","method":"In vitro kinase assay (c-Src + PFKFB3), site-directed mutagenesis (Y194F), PFKFB3-Y194F knockin mice, APCmin/+ genetic cross, cell proliferation and xenograft assays, clinical tumor sample correlation","journal":"Cell reports","confidence":"High","confidence_rationale":"Tier 1 / Strong — in vitro kinase assay, mutagenesis at specific residue, knockin mouse model with in vivo epistasis, multiple orthogonal methods","pmids":["32209481"],"is_preprint":false},{"year":2014,"finding":"Estradiol (E2) promotes PFKFB3 mRNA transcription through direct estrogen receptor (ER) binding to the PFKFB3 promoter, leading to increased PFKFB3 protein, elevated F2,6BP, and enhanced glucose uptake and glycolysis in ER+ breast cancer cells.","method":"ER-chromatin binding (promoter assay), siRNA, PFKFB3 inhibitor, 14C-glucose uptake, F2,6BP measurement, apoptosis assay in MCF-7 cells","journal":"The Journal of biological chemistry","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ER-promoter binding demonstrated, siRNA + inhibitor with metabolic readouts, single lab","pmids":["24515104"],"is_preprint":false},{"year":2019,"finding":"Autophagy machinery, specifically the UBA domain of p62/sequestosome-1, physically interacts with PFKFB3; autophagy reduces PFKFB3 expression and induces cellular dormancy, whereas impairment of autophagy (knockdown of Atg3, Atg7, or p62) restores PFKFB3 expression and reactivates proliferation in dormant breast cancer stem cells.","method":"Co-immunoprecipitation (PFKFB3-p62 interaction), siRNA (Atg3, Atg7, p62), microarray, cell dormancy/proliferation assays, in vivo metastasis model","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 2 / Strong — reciprocal Co-IP identifying specific domain (UBA), genetic epistasis via multiple Atg gene KDs, in vivo validation","pmids":["31413316"],"is_preprint":false},{"year":2018,"finding":"Nuclear PFKFB3 silencing in hepatocellular carcinoma decreases AKT phosphorylation and reduces ERCC1 expression, impairing DNA repair and causing G2/M arrest and apoptosis, revealing a non-glycolytic nuclear function.","method":"siRNA knockdown, AKT phosphorylation western blot, ERCC1 expression, cell cycle analysis, xenograft model","journal":"Cell death & disease","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — KD with defined molecular pathway (AKT-ERCC1), in vivo xenograft, single lab","pmids":["29559632"],"is_preprint":false},{"year":2019,"finding":"PFKFB3-driven glycolysis in pulmonary endothelial cells promotes pulmonary hypertension; endothelial PFKFB3 knockout reduces HIF2A levels, decreases production of growth factors (PDGFB, FGF2) and proinflammatory factors (CXCL12, IL-1β), and prevents vascular smooth muscle cell proliferation and leukocyte recruitment.","method":"Conditional endothelial Pfkfb3-KO mice (constitutive and inducible), heterozygous global KO, 3PO pharmacological inhibition, Western blot, immunostaining of lung ECs","journal":"Proceedings of the National Academy of Sciences","confidence":"High","confidence_rationale":"Tier 2 / Strong — multiple genetic models (constitutive, inducible, heterozygous KO), mechanistic link to HIF2A and downstream paracrine factors, in vivo and ex vivo validation","pmids":["31213542"],"is_preprint":false},{"year":2024,"finding":"PFKFB3-mediated tubular glycolysis increases kidney lactate levels, which drives histone lactylation (particularly H4K12la) at the promoters of NF-κB pathway genes (Ikbkb, Rela, Relb), activating their transcription and promoting renal inflammation and fibrosis. PFKFB3 also directly activates IKKβ, IκBα, and p65.","method":"PTC-specific PFKFB3-KO mice, ChIP-seq (H4K12la at gene promoters), kidney IRI model, heterozygous KO and pharmacological inhibition, metabolite measurement (lactate), Western blot","journal":"Kidney international","confidence":"High","confidence_rationale":"Tier 2 / Strong — ChIP-seq with specific histone mark, cell-type-specific KO, multiple genetic models, mechanistic pathway from metabolite (lactate) to histone modification to gene expression","pmids":["38789037"],"is_preprint":false},{"year":2022,"finding":"PFKFB3-driven glycolysis in endothelial cells drives EndoMT by hijacking glucose flux from the pentose phosphate pathway, reducing cytoplasmic NADPH production. Mitochondrial NADPH efflux via the isocitrate/α-ketoglutarate shuttle replenishes cytoplasmic NADPH but impairs mitochondrial respiration by hampering iron-sulfur cluster biosynthesis.","method":"PFKFB3 haplodeficiency and overexpression, NADPH flux measurement, mitochondrial respiration assay (Seahorse), iron-sulfur cluster biosynthesis analysis, pharmacological inhibition with salvianolic acid C","journal":"Signal transduction and targeted therapy","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic haplodeficiency and overexpression with metabolic flux measurements, single lab, multiple metabolic readouts","pmids":["36045132"],"is_preprint":false},{"year":2019,"finding":"PFKFB3 inhibition by small molecule AZ67 prevents NMDAR excitotoxicity-induced NADPH oxidation, redox stress, and apoptotic neuronal death; in vivo administration of AZ67 alleviates motor discoordination and brain infarct in middle carotid artery occlusion ischemia/reperfusion model in mice.","method":"Pharmacological inhibition (AZ67), NADPH measurement, apoptosis assay in primary neurons, oxygen-glucose deprivation model, in vivo MCAO mouse model","journal":"Scientific reports","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — in vitro and in vivo validation with pharmacological inhibitor, mechanistic link to NADPH/redox, single lab","pmids":["31406177"],"is_preprint":false},{"year":2022,"finding":"In CLN7 neuronal ceroid lipofuscinosis, failure of autophagy causes mitochondrial accumulation and elevated mitochondrial ROS (mROS), which signal to stabilize PFKFB3 (normally proteasomally degraded in healthy neurons) via a mROS-dependent protein stabilization cascade, driving aberrant glycolysis in neurons and contributing to disease pathogenesis.","method":"Cln7∆ex2 mouse model, in vivo genetic mROS measurement, PFKFB3 protein stability assays, AZ67 inhibitor treatment in vivo and in CLN7 patient-derived cells","journal":"Nature communications","confidence":"High","confidence_rationale":"Tier 2 / Strong — in vivo genetic model, patient-derived cell validation, in vivo pharmacological rescue, mechanistic cascade (mROS → PFKFB3 stabilization)","pmids":["35087090"],"is_preprint":false},{"year":2022,"finding":"PFKFB3 interacts with cyclin-dependent kinase 4 (CDK4) in the nucleus of kidney tubular cells following cisplatin treatment, leading to CDK4 activation and consequent phosphorylation/inactivation of the retinoblastoma tumor suppressor (Rb), causing apoptosis; this is independent of PFKFB3's canonical glycolytic function.","method":"Co-immunoprecipitation (PFKFB3-CDK4), CDK4 kinase activity assay, Rb phosphorylation immunoblot, renal proximal tubule-specific PFKFB3-KO mice, CDK4 inhibition rescue","journal":"Translational research","confidence":"High","confidence_rationale":"Tier 2 / Strong — Co-IP of PFKFB3-CDK4 complex, kinase activity assay, cell-type-specific KO, epistasis with CDK4 inhibition, identifies non-canonical nuclear function","pmids":["36243313"],"is_preprint":false},{"year":2023,"finding":"OTUD4 deubiquitinase binds PFKFB3 and blocks its ubiquitination-mediated proteasomal degradation, stabilizing PFKFB3 protein in cardiac fibroblasts stimulated with TGF-β1 and promoting glycolysis-driven fibroblast activation.","method":"Co-immunoprecipitation (PFKFB3-OTUD4), ubiquitination assay, siRNA knockdown, cardiac fibroblast activation assays, post-MI mouse model","journal":"Journal of molecular medicine","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — Co-IP and ubiquitination assay identify deubiquitinase-substrate relationship, in vivo model, single lab","pmids":["37162556"],"is_preprint":false},{"year":2022,"finding":"ROCK2 stabilizes PFKFB3 protein in osteosarcoma cells by modifying its ubiquitination and reducing proteasomal degradation; PFKFB3 is epistatic to ROCK2 in promoting proliferation and metastasis, as PFKFB3 overexpression rescues ROCK2 knockdown-induced growth impairment.","method":"siRNA knockdown, PFKFB3 overexpression rescue, ubiquitination assay, proliferation/invasion assays, clinical tissue correlation","journal":"Experimental cell research","confidence":"Medium","confidence_rationale":"Tier 2-3 / Moderate — ubiquitination assay and epistasis experiment, single lab, no detailed mapping of modification sites","pmids":["31678169"],"is_preprint":false},{"year":2022,"finding":"CYLD stabilizes p53 and promotes its nuclear translocation by removing K63- and K48-linked ubiquitin chains from p53, enabling p53 to bind the PFKFB3 promoter and inhibit its transcription. CYLD also interacts with FZR1 to promote APC/C-FZR1 E3 ligase activity, which ubiquitinates and degrades PFKFB3 via 26S proteasomal system.","method":"Co-immunoprecipitation (CYLD-FZR1, CYLD-p53), ubiquitination assay (K48/K63 chains on p53), ChIP (p53 binding to PFKFB3 promoter), proteasome inhibition, siRNA","journal":"Cancer letters","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — two distinct mechanisms (transcriptional repression and proteolysis), ChIP and Co-IP data, single lab","pmids":["35131382"],"is_preprint":false},{"year":2020,"finding":"Fascin promotes PFKFB3 transcription by activating YAP1 through its canonical actin-bundling activity; YAP1 binds a TEAD1/4 binding motif 30 bp upstream of the PFKFB3 transcription start site to increase PFKFB3 expression and glycolysis in lung cancer cells.","method":"ChIP (YAP1-TEAD at PFKFB3 promoter), fascin actin-bundling mutant, YAP1 siRNA, PFKFB3 promoter luciferase, xenograft metastasis models, organoid cultures","journal":"Cancer letters","confidence":"High","confidence_rationale":"Tier 2 / Strong — ChIP at specific promoter element, actin-bundling mutant epistasis, in vivo xenograft, luciferase reporter, multiple models","pmids":["34303764"],"is_preprint":false},{"year":2021,"finding":"YAP acts as a transcriptional co-activator with TEAD1 to bind the PFKFB3 promoter and increase PFKFB3 expression under hypoxia in endothelial cells, thereby promoting glycolysis and angiogenesis; YAP silencing inhibits endothelial glycolysis and can be rescued by enforced PFKFB3 expression.","method":"ChIP (YAP-TEAD1 binding to PFKFB3 promoter), siRNA, PFKFB3 overexpression rescue, Seahorse glycolysis assay, intravitreal siRNA injection in CNV/OIR mouse models","journal":"Angiogenesis","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ChIP and epistasis, in vivo animal model, single lab, independent from PMID:34303764 which provides orthogonal replication in cancer context","pmids":["33400016"],"is_preprint":false},{"year":2022,"finding":"PFKFB3-driven glycolysis in macrophages activates HIF-1α/HIF-2α and NF-κB, inducing M1/M2 polarization markers and pro-angiogenic cytokines, thereby reprogramming macrophages toward an angiogenic phenotype that promotes choroidal neovascularization; myeloid-specific PFKFB3 KO attenuates this process.","method":"Myeloid cell-specific PFKFB3 KO mice, bone marrow-derived macrophage stimulation, PFKFB3 inhibitor (AZ67), HIF-1α/HIF-2α and NF-κB pathway immunoblotting, laser-induced CNV model","journal":"British journal of pharmacology","confidence":"High","confidence_rationale":"Tier 2 / Strong — myeloid cell-specific genetic KO, pharmacological validation, defined downstream pathway (HIF-1α/2α, NF-κB), in vivo disease model","pmids":["35830274"],"is_preprint":false},{"year":2022,"finding":"PFKFB3-driven glycolysis is required for actin polymerization in macrophages; homozygous loss of Pfkfb3 impairs macrophage efferocytosis by disrupting actin-dependent cytoskeletal function, exacerbating atherosclerosis.","method":"Homozygous Pfkfb3-KO Apoe-/- mice, efferocytosis assay, actin polymerization measurement, flow cytometry","journal":"British journal of pharmacology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic KO with specific cellular phenotype (efferocytosis) linked to actin, in vivo atherosclerosis model, single study","pmids":["35834356"],"is_preprint":false},{"year":2022,"finding":"ALK fusion oncoproteins promote PFKFB3 transcription through the downstream transcription factor STAT3; pharmacological or genetic blockade of ALK reduces PFKFB3 expression and glycolysis, and PFKFB3 inhibition can overcome TKI resistance in ALK-mutant cancer cells.","method":"Quantitative proteomics, STAT3 ChIP/reporter assay, ALK TKI treatment, PFKFB3 siRNA and overexpression, correlation in clinical ALK+ NSCLC samples","journal":"Oncogene","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ChIP/reporter assay for STAT3 at PFKFB3 promoter, proteomic identification, clinical correlation, single lab","pmids":["36064579"],"is_preprint":false},{"year":2022,"finding":"The deubiquitinase UCHL1 stabilizes PFKFB3 in astrocytes by cleaving K48-linked ubiquitin chains; the UCHL1/PFKFB3 axis increases lactate production, which drives histone H4K8 lactylation (H4K8la), subsequently activating transcription of Uchl1 and glycolysis genes, forming a positive feedback loop that supports astrocytic metabolic reprogramming after spinal cord injury.","method":"Genetic deletion of Uchl1 and Pfkfb3, K48-ubiquitin chain cleavage assay, ChIP (H4K8la at gene promoters), lactate measurement, in vivo SCI model, scRNA-seq analysis","journal":"Cell death and differentiation","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — deubiquitinase assay, ChIP at specific histone mark/promoter, in vivo genetic models, single lab","pmids":["40016338"],"is_preprint":false},{"year":2023,"finding":"In pulmonary arterial smooth muscle cells, HIF-1α binds the promoter of miR-26a-5p to inhibit its expression; miR-26a-5p directly targets PFKFB3 (validated by dual-luciferase assay), and PFKFB3 enhances phosphorylation of ULK1 to promote autophagy and cell proliferation in PAH.","method":"ChIP (HIF-1α at miR-26a-5p promoter), dual-luciferase reporter assay (miR-26a-5p targeting PFKFB3 3'UTR), ULK1 phosphorylation immunoblot, tandem mRFP-GFP-LC3B autophagy assay, adeno-miR-26a-5p in vivo rat PAH model","journal":"FASEB journal","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — direct target validation by luciferase + blot, ChIP for upstream regulation, in vivo rat model, single lab","pmids":["37052859"],"is_preprint":false},{"year":2003,"finding":"The human PFKFB3 gene spans 32.5 kb on chromosome 10p15.3-p15.2, contains 19 exons (15 normally expressed), and encodes a 590 amino acid, 66.9 kDa bifunctional protein with the highest kinase/phosphatase activity ratio among all PFKFB isozymes.","method":"Genomic sequencing, exon-intron junction determination, FISH chromosomal localization, open reading frame analysis","journal":"International journal of oncology","confidence":"Medium","confidence_rationale":"Tier 1 / Weak — definitive gene structure by sequencing and FISH, but single characterization study with no functional validation of domains","pmids":["12963966"],"is_preprint":false},{"year":2018,"finding":"X-ray crystal structures of PFKFB3 were used to guide structure-based design of N-aryl 6-aminoquinoxaline inhibitors that bind the kinase active site; the most potent compound showed IC50 of 14 nM for PFKFB3 kinase and 0.49 μM for F2,6BP production in HCT116 cells.","method":"X-ray crystallography, docking, structure-activity relationship (SAR), enzymatic IC50 assay, cellular F2,6BP assay","journal":"ChemMedChem","confidence":"High","confidence_rationale":"Tier 1 / Moderate — crystal structure with direct active-site binding demonstrated, enzymatic and cellular assay validation, structure-based design","pmids":["30378281"],"is_preprint":false},{"year":2022,"finding":"LINC00930 lncRNA acts as a scaffold to recruit the RBBP5 and GCN5 complex to the PFKFB3 promoter, increasing H3K4 trimethylation and H3K9 acetylation at the PFKFB3 promoter to epigenetically activate PFKFB3 transcription and glycolysis in nasopharyngeal carcinoma.","method":"ChIP (H3K4me3, H3K9ac at PFKFB3 promoter), RNA-immunoprecipitation (RBBP5/GCN5 interaction with LINC00930), siRNA, in vivo tumor model","journal":"Journal of experimental & clinical cancer research","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — ChIP with specific histone marks, RIP for scaffold mechanism, in vivo validation, single lab","pmids":["35209949"],"is_preprint":false},{"year":2019,"finding":"PFKFB3 inhibition reduces TNF-α-induced endothelial proinflammatory responses (cytokines, ICAM-1, monocyte adhesion/transmigration) by suppressing IKKβ phosphorylation, IκBα phosphorylation/degradation, NF-κB-p65 nuclear translocation, and NF-κB DNA-binding activity.","method":"siRNA knockdown, PFKFB3 inhibitor, cytokine antibody array, monocyte adhesion/transmigration assay, NF-κB EMSA, immunofluorescence (p65 localization), western blotting","journal":"Inflammation","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — multiple readouts (EMSA, immunofluorescence, western), siRNA and inhibitor convergent, single lab","pmids":["30171427"],"is_preprint":false},{"year":2019,"finding":"PFKFB3/HIF-1α form a positive feedback loop in hepatocellular carcinoma: HIF-1α drives PFKFB3 expression, and exogenous PFKFB3 overexpression in turn upregulates HIF-1α protein levels; HIF-1α deficiency impairs PFKFB3-induced sorafenib resistance.","method":"HIF-1α blockade, PFKFB3 overexpression, HIF-1α and PFKFB3 protein immunoblotting, apoptosis assay, GEO dataset validation","journal":"Biochemical and biophysical research communications","confidence":"Low","confidence_rationale":"Tier 3 / Weak — single lab, primarily western blot-based, no direct mechanistic link between PFKFB3 and HIF-1α protein stability identified","pmids":["30981500"],"is_preprint":false},{"year":2023,"finding":"cGAS-STING-IRF3 signaling pathway promotes renal fibrosis by upregulating PFKFB3 expression under hypoxia; inhibition of STING or IRF3 reverses elevated PFKFB3, placing PFKFB3 downstream of cGAS-STING-IRF3 in hypoxia-induced fibrosis.","method":"STING and IRF3 pharmacological inhibition, hypoxia-stimulated HK-2 cells, IRI mouse model, PFKFB3 expression immunoblotting, fibrosis markers","journal":"Free radical biology & medicine","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — epistasis (inhibition of upstream pathway reverses PFKFB3 induction), both in vitro and in vivo models, single lab","pmids":["37714438"],"is_preprint":false},{"year":2022,"finding":"HIF1A directly induces PFKFB3 expression in alveolar epithelial cells; alveolar epithelium-specific Pfkfb3 deletion dramatically increases acute lung injury severity, and pyruvate (restoring metabolic substrate) rescues both Pfkfb3loxP/loxP and Hif1aloxP/loxP mice from ALI.","method":"Alveolar epithelium-specific Pfkfb3-KO and Hif1a-KO mice (SPC-ER-Cre), injurious ventilation and acid instillation ALI models, intratracheal pyruvate rescue, human lung biopsy immunostaining","journal":"JCI insight","confidence":"High","confidence_rationale":"Tier 2 / Strong — cell-type-specific genetic KO of both HIF1A and PFKFB3, in vivo rescue experiment, human tissue validation, establishes HIF1A→PFKFB3 axis in alveolar epithelia","pmids":["36326834"],"is_preprint":false},{"year":2021,"finding":"PFKFB3 inhibition in small cell lung carcinoma attenuates invasion/migration by downregulating YAP/TAZ signaling while increasing pLATS1 via activation of pMST1 and NF2, linking PFKFB3-driven glycolysis to regulation of the Hippo pathway.","method":"PFK158 inhibitor, shRNA stable knockdown, Hippo pathway component immunoblotting (pMST1, NF2, pLATS1, YAP/TAZ), xenograft model","journal":"Oncogene","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — pharmacological and genetic inhibition with Hippo pathway readout, in vivo xenograft, single lab","pmids":["35804016"],"is_preprint":false},{"year":2019,"finding":"RANKL-induced osteoclastogenesis increases PFKFB3 expression and glycolysis; L-lactate (the glycolytic product) partially reverses the suppression of osteoclastogenesis caused by PFKFB3 inhibition and abrogates the inhibitory effect on NF-κB and MAPK pathways, establishing lactate as a downstream mediator of PFKFB3's pro-osteoclast function.","method":"siRNA, PFKFB3 inhibitor (PFK15), L-lactate supplementation rescue, NF-κB and MAPK immunoblotting, lactate/glucose measurement, ovariectomy bone loss mouse model","journal":"Journal of cellular and molecular medicine","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — metabolite rescue experiment identifies lactate as mediator, pathway (NF-κB/MAPK) dissection, in vivo bone loss model, single lab","pmids":["31880389"],"is_preprint":false},{"year":2023,"finding":"PFKFB3-derived lactate in renal fibroblasts promotes fibrotic activation; the glycolytic metabolite lactate directly promotes fibrotic phenotype in NRK-49F cells, and myofibroblast-specific PFKFB3 KO mice show substantially reduced fibrosis after UUO or IRI.","method":"Myofibroblast-specific Pfkfb3-KO mice (Pfkfb3f/f/PostnMCM), exogenous lactate supplementation, TGF-β1 stimulation, α-SMA and fibronectin markers, scRNA-seq reanalysis","journal":"Cells","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — cell-type-specific KO in vivo, lactate rescue experiment, mechanistic link to downstream fibrotic activation, single lab","pmids":["37626891"],"is_preprint":false},{"year":2024,"finding":"SphK1/S1P signaling stabilizes PFKFB3 in endothelial cells to supply glycolytic energy for tumor angiogenesis; pharmacological SphK1 inhibition induces proteasomal degradation of PFKFB3, which can be reversed by S1P supplementation in an S1P receptor-dependent manner.","method":"SphK1 inhibitor (PF-543), Sphk1-KO mice, S1P supplementation rescue, PFKFB3 protein stability assay (proteasome inhibitor), DEN-induced primary HCC model, lentiviral SphK1 KD","journal":"Journal of translational medicine","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — genetic KO and S1P rescue establish pathway, proteasomal degradation mechanism, in vivo primary HCC model, single lab","pmids":["38200582"],"is_preprint":false},{"year":2023,"finding":"In myeloid cells, PFKFB3-driven glycolysis stabilizes HIF1α, which alters macrophage phenotype to contribute to renal fibrosis; myeloid-specific PFKFB3-KO reduces M1 and M2 macrophage infiltration, suppresses macrophage-to-myofibroblast transition, and decreases kidney fibrosis.","method":"Myeloid-specific Pfkfb3-KO mice (Pfkfb3ΔMφ), UUO model, HIF1α stabilization assay, macrophage phenotyping by flow cytometry, scRNA-seq reanalysis","journal":"Frontiers in immunology","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — cell-type-specific genetic KO, HIF1α mechanistic link, in vivo disease model, single lab","pmids":["38035106"],"is_preprint":false},{"year":2020,"finding":"FAT10 upregulates PFKFB3 in osteosarcoma by directly binding to EGFR and inhibiting EGFR ubiquitination and degradation, thereby stabilizing EGFR which then promotes PFKFB3 expression and glycolysis.","method":"Co-immunoprecipitation (FAT10-EGFR), ubiquitination assay, siRNA knockdown, PFKFB3 expression immunoblot, glycolysis measurement, osteosarcoma tissue correlation","journal":"American journal of cancer research","confidence":"Medium","confidence_rationale":"Tier 3 / Moderate — Co-IP and ubiquitination assay identify FAT10-EGFR-PFKFB3 cascade, multiple complementary experiments, single lab","pmids":["32775001"],"is_preprint":false},{"year":2023,"finding":"PFKFB3-driven glycolysis in vascular smooth muscle cells promotes calcification through FoxO3 expression and lactate production; silencing PFKFB3 reduces FoxO3, and pyruvate/lactate supplementation reverses PFKFB3-depletion effects on ALP activity and OPG expression, establishing lactate as an osteogenic mediator downstream of PFKFB3.","method":"RNA-seq after PFKFB3 KD, FoxO3 silencing epistasis, lactate/pyruvate supplementation rescue, miR-26a/b-5p overexpression, in vivo vitamin D3 calcification model, VSMC osteogenic transdifferentiation assays","journal":"FASEB journal","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — RNA-seq + epistasis (FoxO3 KD) + metabolite rescue, in vivo model, single lab","pmids":["37682013"],"is_preprint":false},{"year":2022,"finding":"PFKFB3 overexpression in bone marrow endothelial progenitor cells facilitates pro-apoptotic transcription factor FOXO3A and its downstream genes (p21, p27, FAS), activates NF-κB and E-selectin expression, and reduces SDF-1, impairing hematopoiesis-supporting function after chemotherapy; FOXO3A silencing rescues these effects.","method":"PFKFB3 overexpression/knockdown in BM EPCs, FOXO3A silencing rescue experiment, NF-κB activation immunoblot, E-selectin and SDF-1 measurement, BM EC-specific PFKFB3 overexpression transgenic mice, 5-FU chemotherapy model, patient BM EPC analysis","journal":"Haematologica","confidence":"Medium","confidence_rationale":"Tier 2 / Moderate — transgenic in vivo model, FOXO3A epistasis rescue, multiple pathway readouts, clinical patient validation, single lab","pmids":["35354250"],"is_preprint":false}],"current_model":"PFKFB3 is a bifunctional enzyme with the highest kinase/phosphatase ratio among PFKFB family members that synthesizes fructose-2,6-bisphosphate (F2,6BP) to allosterically activate PFK-1 and drive glycolysis; its activity and stability are regulated by post-translational modifications including AMPK-mediated phosphorylation, c-Src-mediated tyrosine phosphorylation at Y194, acetylation at K472 (causing cytoplasmic redistribution), and proteasomal degradation via APC/C-Cdh1 (KEN motif) which is regulated by deubiquitinases (UCHL1, OTUD4) and stabilizing proteins (AGPG lncRNA, ROCK2, FAT10/EGFR axis); beyond glycolysis, PFKFB3 has non-canonical nuclear roles in homologous recombination DNA repair (via RRM2 recruitment and dNTP supply), CDK4 activation (causing Rb phosphorylation), and interaction with p62/autophagy machinery, while its downstream metabolite lactate drives histone lactylation (H4K12la, H4K8la) to epigenetically activate NF-κB pathway genes and fibrotic programs; transcriptionally, PFKFB3 is induced by HIF-1α, ER/GPER1, PI3K-Akt-mTOR, STAT3 (downstream of ALK), YAP-TEAD, and cGAS-STING-IRF3 pathways, and its mRNA is regulated post-transcriptionally by CPEB4-mediated polyadenylation."},"narrative":{"mechanistic_narrative":"PFKFB3 is a bifunctional 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase that synthesizes fructose-2,6-bisphosphate (F2,6BP) to drive glycolytic flux, coupling glucose metabolism to proliferation, migration, survival, and DNA repair [PMID:12963966, PMID:16698023, PMID:12384552]. Beyond sustaining metabolism, its glycolytic output directly governs cell behavior: in endothelial cells PFKFB3 compartmentalizes with F-actin in motile protrusions to control filopodia/lamellipodia formation and tip/stalk cell selection during vessel branching, overriding Notch [PMID:23911327], and in macrophages it provides the glycolytic energy required for actin-dependent efferocytosis [PMID:35834356]. PFKFB3 also executes non-canonical nuclear functions independent of bulk glycolysis: it relocates to ionizing-radiation-induced foci in an MRN-ATM-γH2AX-MDC1-dependent manner where its enzymatic activity recruits RRM2 and maintains the dNTP pool needed for homologous recombination [PMID:30250201], and it activates CDK1 (driving p27 degradation and G1/S progression) and CDK4 (driving Rb phosphorylation) [PMID:25032860, PMID:36243313]. Its terminal metabolite lactate acts as an epigenetic signal, driving histone lactylation (H4K12la, H4K8la) at NF-κB pathway and glycolytic gene promoters to amplify inflammatory and fibrotic programs [PMID:38789037, PMID:40016338]. PFKFB3 abundance and activity are tightly tuned post-translationally: AMPK and c-Src (at Tyr194) phosphorylate and activate it [PMID:26322680, PMID:29410405, PMID:32209481], cisplatin-induced K472 acetylation disrupts its nuclear localization and redistributes it to the cytoplasm [PMID:29410405], and its protein stability is set by a balance between APC/C-Cdh1-mediated proteasomal degradation via a KEN motif [PMID:22421967] and competing stabilizers including the lncRNA AGPG, deubiquitinases UCHL1 and OTUD4, ROCK2, and the FAT10/EGFR axis [PMID:32198345, PMID:40016338, PMID:37162556, PMID:31678169, PMID:32775001]. Transcriptionally and post-transcriptionally PFKFB3 is induced by HIF-1α, estrogen receptor, YAP-TEAD, STAT3, and cGAS-STING-IRF3 signaling and by CPEB4-mediated cytoplasmic polyadenylation [PMID:36326834, PMID:24515104, PMID:34303764, PMID:36064579, PMID:37714438, PMID:32169429]. Through these mechanisms PFKFB3 promotes tumor growth, angiogenesis, neuronal excitotoxic death, and organ fibrosis across multiple tissues [PMID:32209481, PMID:31213542, PMID:31406177, PMID:37626891].","teleology":[{"year":2002,"claim":"Established that PFKFB3 expression and its product F2,6BP are induced by hypoxia and peak in S phase, first linking this glycolytic activator to cell-cycle progression and the tumor microenvironment.","evidence":"in situ hybridization, IHC, cell-cycle fractionation and hypoxia treatment of cancer cells","pmids":["12384552"],"confidence":"Medium","gaps":["No direct demonstration that S-phase F2,6BP is causally required for cycle progression","Upstream hypoxia transcription factor not yet identified in this study"]},{"year":2003,"claim":"Defined the human PFKFB3 gene structure and protein, establishing it as a bifunctional enzyme with the highest kinase/phosphatase ratio among isozymes, rationalizing its role as a net glycolysis activator.","evidence":"genomic sequencing, exon-intron mapping, FISH, ORF analysis","pmids":["12963966"],"confidence":"Medium","gaps":["No functional validation of catalytic domains","Regulatory residues not mapped"]},{"year":2006,"claim":"Showed by loss of function that PFKFB3 is required for F2,6BP, lactate and ATP production and supports viability and anchorage-independent growth, cementing it as a metabolic dependency in cancer cells.","evidence":"siRNA knockdown with metabolite measurement, cell-cycle/apoptosis flow cytometry, soft agar assay in HeLa","pmids":["16698023"],"confidence":"Medium","gaps":["Single cell line","Mechanism downstream of metabolite depletion not dissected"]},{"year":2012,"claim":"Revealed that PFKFB3 is normally degraded by APC/C-Cdh1 via a KEN motif and that NMDAR signaling stabilizes it and shifts it from nucleus to cytosol, switching neuronal metabolism from PPP to glycolysis and causing oxidative death — the first non-proliferative, degradation-controlled role.","evidence":"NMDAR stimulation, KEN-motif deletion mutant, subcellular fractionation, PPP/glycolysis flux, G6PD rescue in cortical neurons","pmids":["22421967"],"confidence":"High","gaps":["E2/E3 components beyond APC/C-Cdh1 not detailed","Trigger linking NMDAR activity to Cdh1 inhibition not fully resolved"]},{"year":2013,"claim":"Demonstrated PFKFB3 compartmentalizes with F-actin in motile protrusions and controls endothelial migration and tip/stalk competition over Notch, establishing a localized glycolytic role in cell motility and angiogenesis.","evidence":"siRNA/genetic KO, mosaic in vitro and in vivo sprouting assays, F-actin co-localization, Notch epistasis","pmids":["23911327"],"confidence":"High","gaps":["Molecular basis of PFKFB3-F-actin compartmentalization unresolved","Whether ATP/F2,6BP locally fuels actin dynamics not directly shown"]},{"year":2014,"claim":"Identified the CDK1–p27 axis as the mechanism by which PFKFB3 promotes cycle progression, and that estrogen receptor directly transactivates PFKFB3, connecting hormonal signaling to glycolytic cell-cycle control.","evidence":"siRNA + co-siRNA epistasis, CDK1 kinase assay, p27 ubiquitination (HeLa); ER-promoter binding, glucose uptake, F2,6BP in MCF-7","pmids":["25032860","24515104"],"confidence":"High","gaps":["How F2,6BP activates CDK1 mechanistically not defined","ER binding site on PFKFB3 promoter not finely mapped"]},{"year":2015,"claim":"Showed AMPK directly phosphorylates PFKFB3 during mitotic arrest to raise glycolysis and compensate for mitophagy-driven loss of oxidative phosphorylation, establishing direct kinase regulation coupled to translational upregulation.","evidence":"AMPK kinase assay, phospho-immunoblot, siRNA, autophagy modulation, breast cancer viability assays","pmids":["26322680"],"confidence":"High","gaps":["Phosphosite identity and effect on kinase vs phosphatase activity not resolved here","Mechanism of mitotic translational activation not defined"]},{"year":2018,"claim":"Connected PTM-controlled localization to chemoresistance: cisplatin-induced K472 acetylation disrupts the NLS, driving cytoplasmic PFKFB3 that AMPK then activates, while a separate nuclear PFKFB3 pool supports AKT-ERCC1-dependent DNA repair.","evidence":"K472 acetylation-dead mutant, fractionation, AMPK assay, xenograft (Nat Commun); siRNA + AKT/ERCC1 readout in HCC (Cell Death Dis)","pmids":["29410405","29559632"],"confidence":"High","gaps":["Acetyltransferase/deacetylase for K472 not identified","Direct vs indirect link between nuclear PFKFB3 and AKT-ERCC1 unresolved"]},{"year":2018,"claim":"Established a non-glycolytic nuclear role in DNA repair by showing PFKFB3 enters DNA-damage foci dependent on the MRN-ATM cascade and that its enzymatic activity is required for RRM2 recruitment and dNTP supply during homologous recombination.","evidence":"IF co-localization with HR foci, selective inhibitor KAN0438757, HR reporter, dNTP measurement, radiosensitization","pmids":["30250201"],"confidence":"High","gaps":["How a glycolytic enzyme is recruited to chromatin foci is unknown","Relationship between local F2,6BP/dNTP pools and RRM2 not structurally defined"]},{"year":2019,"claim":"Defined PFKFB3 as a node in inflammation and tissue-specific disease: it sustains NF-κB activation in endothelium, supports osteoclastogenesis via lactate, drives pulmonary hypertension through HIF2A/paracrine factors, and is suppressed by autophagy via p62 to enforce dormancy.","evidence":"siRNA/inhibitor with NF-κB EMSA (Inflammation); lactate rescue + bone-loss model (osteoclast); conditional endothelial KO (PNAS); p62 UBA Co-IP + Atg KD + metastasis model (Nat Commun)","pmids":["30171427","31880389","31213542","31413316"],"confidence":"High","gaps":["Whether PFKFB3 acts on NF-κB via metabolite vs protein interaction varies across systems","Direct p62-PFKFB3 functional consequence on degradation pathway not fully mapped"]},{"year":2020,"claim":"Mapped multiple layers of PFKFB3 stabilization and activation: c-Src phosphorylation at Tyr194 (validated in knockin mice), AGPG lncRNA blocking APC/C ubiquitination, and CPEB4-driven cytoplasmic polyadenylation — each independently amplifying glycolysis and tumor/fibrotic phenotypes.","evidence":"in vitro c-Src kinase assay + Y194F knockin/APCmin cross; AGPG Co-IP/ubiquitination + PDX; CPEB4 RIP + KO mice in liver fibrosis","pmids":["32209481","32198345","32169429"],"confidence":"High","gaps":["Interplay/hierarchy among Tyr194 phosphorylation, AGPG, and AMPK regulation not integrated","Structural effect of Tyr194 phosphorylation on catalysis unresolved"]},{"year":2021,"claim":"Positioned PFKFB3 within Hippo/YAP signaling bidirectionally — YAP-TEAD transactivates PFKFB3 to fuel angiogenesis, while PFKFB3 inhibition feeds back to modulate MST1/NF2/LATS1 and YAP/TAZ — linking glycolysis to a master growth-control pathway.","evidence":"YAP-TEAD1 ChIP at PFKFB3 promoter + rescue in OIR/CNV models; PFK158/shRNA with Hippo readouts + xenograft in SCLC","pmids":["33400016","35804016"],"confidence":"Medium","gaps":["Mechanism by which PFKFB3 feeds back onto Hippo kinases unclear","Direct vs metabolite-mediated effect on YAP not distinguished"]},{"year":2022,"claim":"Consolidated PFKFB3 as a stress-stabilized driver across neurodegeneration, angiogenesis, atherosclerosis, and cancer, controlled by diverse stabilizers (ROCK2, FAT10/EGFR), upstream inducers (fascin-YAP, ALK-STAT3, HIF1A), and a CDK4 non-glycolytic axis, with metabolic crosstalk to PPP/NADPH and mitochondrial function.","evidence":"Cln7 mouse + mROS stabilization; myeloid/endothelial/alveolar KO models; PFKFB3-CDK4 Co-IP; ubiquitination assays (ROCK2, FAT10-EGFR); NADPH/Seahorse flux; ChIP for fascin-YAP","pmids":["35087090","35830274","35834356","36243313","31678169","32775001","34303764","36064579","36326834","36045132","35354250"],"confidence":"High","gaps":["How mROS chemically stabilizes PFKFB3 protein not defined","Whether CDK4 binding involves catalytic activity unresolved"]},{"year":2023,"claim":"Established lactate-driven epigenetic signaling as a PFKFB3 effector mechanism and added further upstream regulators (cGAS-STING-IRF3, SphK1/S1P, OTUD4, HIF1α-miR-26a) controlling its abundance in fibrosis and angiogenesis.","evidence":"lactate rescue + myofibroblast/VSMC KO; STING/IRF3 inhibition; SphK1 KO + S1P rescue; OTUD4 Co-IP/ubiquitination; HIF-1α-miR-26a-5p luciferase + ULK1 phosphorylation","pmids":["37626891","37682013","37714438","38200582","37162556","37052859"],"confidence":"Medium","gaps":["Tissue specificity of competing stabilizers/regulators not reconciled","Direct enzymatic targets vs lactate-mediated effects sometimes conflated"]},{"year":2024,"claim":"Demonstrated directly that PFKFB3-derived lactate writes histone lactylation marks (H4K12la, H4K8la) at NF-κB and glycolytic gene promoters, providing a concrete metabolite-to-chromatin mechanism for PFKFB3-driven inflammation, fibrosis, and feed-forward metabolic reprogramming.","evidence":"PTC-specific and Uchl1/Pfkfb3 KO mice, ChIP-seq for H4K12la/H4K8la, injury models, lactate measurement","pmids":["38789037","40016338"],"confidence":"High","gaps":["Lactylation writer/eraser enzymes acting downstream of PFKFB3 lactate not identified","Promoter selectivity of lactylation not mechanistically explained"]},{"year":null,"claim":"How PFKFB3's distinct cytosolic glycolytic, F-actin-associated, and nuclear (HR repair, CDK activation) functions are coordinately partitioned and regulated by its layered PTMs and competing stabilizers/degraders remains unresolved.","evidence":"","pmids":[],"confidence":"Low","gaps":["No integrated model of how localization, PTM state, and binding partners dictate which function PFKFB3 executes","Structural basis for non-catalytic nuclear interactions (CDK4, p62) unknown","Whether kinase vs phosphatase activity is differentially regulated by the various PTMs not resolved"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0016740","term_label":"transferase activity","supporting_discovery_ids":[30,31,8]},{"term_id":"GO:0016787","term_label":"hydrolase activity","supporting_discovery_ids":[30]}],"localization":[{"term_id":"GO:0005829","term_label":"cytosol","supporting_discovery_ids":[4,6,0]},{"term_id":"GO:0005634","term_label":"nucleus","supporting_discovery_ids":[6,7,13,19]},{"term_id":"GO:0005856","term_label":"cytoskeleton","supporting_discovery_ids":[0,26]}],"pathway":[{"term_id":"R-HSA-1430728","term_label":"Metabolism","supporting_discovery_ids":[8,30,1]},{"term_id":"R-HSA-1640170","term_label":"Cell Cycle","supporting_discovery_ids":[5,1,19]},{"term_id":"R-HSA-73894","term_label":"DNA Repair","supporting_discovery_ids":[7,13]},{"term_id":"R-HSA-4839726","term_label":"Chromatin organization","supporting_discovery_ids":[15,28]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[25,33,41]}],"complexes":[],"partners":["RRM2","CDK4","AMPK","SRC","AGPG","OTUD4","UCHL1","SQSTM1"],"other_free_text":[]}},"prefetch_data":{"uniprot":{"accession":"Q16875","full_name":"6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 3","aliases":["6PF-2-K/Fru-2,6-P2ase brain/placenta-type isozyme","Renal carcinoma antigen NY-REN-56","iPFK-2"],"length_aa":520,"mass_kda":59.6,"function":"Catalyzes both the synthesis and degradation of fructose 2,6-bisphosphate","subcellular_location":"","url":"https://www.uniprot.org/uniprotkb/Q16875/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/PFKFB3","classification":"Not Classified","n_dependent_lines":2,"n_total_lines":1208,"dependency_fraction":0.0016556291390728477},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[],"url":"https://opencell.sf.czbiohub.org/search/PFKFB3","total_profiled":1310},"omim":[{"mim_id":"610182","title":"PALMDELPHIN; PALMD","url":"https://www.omim.org/entry/610182"},{"mim_id":"605319","title":"6-@PHOSPHOFRUCTO-2-KINASE/FRUCTOSE-2,6-BISPHOSPHATASE 3; PFKFB3","url":"https://www.omim.org/entry/605319"},{"mim_id":"605213","title":"3-@PHOSPHOINOSITIDE-DEPENDENT PROTEIN KINASE 1; PDPK1","url":"https://www.omim.org/entry/605213"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"Enhanced","locations":[{"location":"Nucleoplasm","reliability":"Enhanced"}],"tissue_specificity":"Tissue enhanced","tissue_distribution":"Detected in all","driving_tissues":[{"tissue":"adipose tissue","ntpm":314.1},{"tissue":"skeletal muscle","ntpm":534.6}],"url":"https://www.proteinatlas.org/search/PFKFB3"},"hgnc":{"alias_symbol":[],"prev_symbol":[]},"alphafold":{"accession":"Q16875","domains":[{"cath_id":"3.40.50.300","chopping":"35-239","consensus_level":"high","plddt":94.7126,"start":35,"end":239},{"cath_id":"3.40.50.1240","chopping":"248-446","consensus_level":"high","plddt":97.0052,"start":248,"end":446}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/Q16875","model_url":"https://alphafold.ebi.ac.uk/files/AF-Q16875-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-Q16875-F1-predicted_aligned_error_v6.png","plddt_mean":86.62},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=PFKFB3","jax_strain_url":"https://www.jax.org/strain/search?query=PFKFB3"},"sequence":{"accession":"Q16875","fasta_url":"https://rest.uniprot.org/uniprotkb/Q16875.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/Q16875/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/Q16875"}},"corpus_meta":[{"pmid":"23911327","id":"PMC_23911327","title":"Role of PFKFB3-driven glycolysis in vessel sprouting.","date":"2013","source":"Cell","url":"https://pubmed.ncbi.nlm.nih.gov/23911327","citation_count":1279,"is_preprint":false},{"pmid":"12384552","id":"PMC_12384552","title":"High expression of inducible 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase (iPFK-2; PFKFB3) in human cancers.","date":"2002","source":"Cancer research","url":"https://pubmed.ncbi.nlm.nih.gov/12384552","citation_count":268,"is_preprint":false},{"pmid":"29263928","id":"PMC_29263928","title":"Roles of PFKFB3 in cancer.","date":"2017","source":"Signal transduction and targeted therapy","url":"https://pubmed.ncbi.nlm.nih.gov/29263928","citation_count":251,"is_preprint":false},{"pmid":"26322680","id":"PMC_26322680","title":"AMPK and PFKFB3 mediate glycolysis and survival in response to mitophagy during mitotic arrest.","date":"2015","source":"Nature cell biology","url":"https://pubmed.ncbi.nlm.nih.gov/26322680","citation_count":223,"is_preprint":false},{"pmid":"32051533","id":"PMC_32051533","title":"PI3K-Akt-mTOR/PFKFB3 pathway mediated lung fibroblast aerobic glycolysis and collagen synthesis in lipopolysaccharide-induced pulmonary fibrosis.","date":"2020","source":"Laboratory investigation; a journal of technical methods and pathology","url":"https://pubmed.ncbi.nlm.nih.gov/32051533","citation_count":207,"is_preprint":false},{"pmid":"31213542","id":"PMC_31213542","title":"PFKFB3-mediated endothelial glycolysis promotes pulmonary hypertension.","date":"2019","source":"Proceedings of the National Academy of Sciences of the United States of America","url":"https://pubmed.ncbi.nlm.nih.gov/31213542","citation_count":182,"is_preprint":false},{"pmid":"32198345","id":"PMC_32198345","title":"Long noncoding RNA AGPG regulates PFKFB3-mediated tumor glycolytic reprogramming.","date":"2020","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/32198345","citation_count":173,"is_preprint":false},{"pmid":"29410405","id":"PMC_29410405","title":"Acetylation accumulates PFKFB3 in cytoplasm to promote glycolysis and protects cells from cisplatin-induced apoptosis.","date":"2018","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/29410405","citation_count":165,"is_preprint":false},{"pmid":"25032860","id":"PMC_25032860","title":"6-Phosphofructo-2-kinase (PFKFB3) promotes cell cycle progression and suppresses apoptosis via Cdk1-mediated phosphorylation of p27.","date":"2014","source":"Cell death & disease","url":"https://pubmed.ncbi.nlm.nih.gov/25032860","citation_count":161,"is_preprint":false},{"pmid":"38789037","id":"PMC_38789037","title":"The glycolytic enzyme PFKFB3 drives kidney fibrosis through promoting histone lactylation-mediated NF-κB family activation.","date":"2024","source":"Kidney international","url":"https://pubmed.ncbi.nlm.nih.gov/38789037","citation_count":153,"is_preprint":false},{"pmid":"31413316","id":"PMC_31413316","title":"Autophagy inhibition elicits emergence from metastatic dormancy by inducing and stabilizing Pfkfb3 expression.","date":"2019","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/31413316","citation_count":146,"is_preprint":false},{"pmid":"32169429","id":"PMC_32169429","title":"CPEB4 Increases Expression of PFKFB3 to Induce Glycolysis and Activate Mouse and Human Hepatic Stellate Cells, Promoting Liver Fibrosis.","date":"2020","source":"Gastroenterology","url":"https://pubmed.ncbi.nlm.nih.gov/32169429","citation_count":130,"is_preprint":false},{"pmid":"33671514","id":"PMC_33671514","title":"Role of PFKFB3 and PFKFB4 in Cancer: Genetic Basis, Impact on Disease Development/Progression, and Potential as Therapeutic Targets.","date":"2021","source":"Cancers","url":"https://pubmed.ncbi.nlm.nih.gov/33671514","citation_count":120,"is_preprint":false},{"pmid":"22421967","id":"PMC_22421967","title":"Excitotoxic stimulus stabilizes PFKFB3 causing pentose-phosphate pathway to glycolysis switch and neurodegeneration.","date":"2012","source":"Cell death and differentiation","url":"https://pubmed.ncbi.nlm.nih.gov/22421967","citation_count":119,"is_preprint":false},{"pmid":"30250201","id":"PMC_30250201","title":"Targeting PFKFB3 radiosensitizes cancer cells and suppresses homologous recombination.","date":"2018","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/30250201","citation_count":106,"is_preprint":false},{"pmid":"36045132","id":"PMC_36045132","title":"Suppression of PFKFB3-driven glycolysis restrains endothelial-to-mesenchymal transition and fibrotic response.","date":"2022","source":"Signal transduction and targeted therapy","url":"https://pubmed.ncbi.nlm.nih.gov/36045132","citation_count":99,"is_preprint":false},{"pmid":"16698023","id":"PMC_16698023","title":"PFKFB3 gene silencing decreases glycolysis, induces cell-cycle delay and inhibits anchorage-independent growth in HeLa cells.","date":"2006","source":"FEBS letters","url":"https://pubmed.ncbi.nlm.nih.gov/16698023","citation_count":96,"is_preprint":false},{"pmid":"28857200","id":"PMC_28857200","title":"By inhibiting PFKFB3, aspirin overcomes sorafenib resistance in hepatocellular carcinoma.","date":"2017","source":"International journal of cancer","url":"https://pubmed.ncbi.nlm.nih.gov/28857200","citation_count":92,"is_preprint":false},{"pmid":"27613609","id":"PMC_27613609","title":"Inhibition of 6-phosphofructo-2-kinase (PFKFB3) suppresses glucose metabolism and the growth of HER2+ breast cancer.","date":"2016","source":"Breast cancer research and treatment","url":"https://pubmed.ncbi.nlm.nih.gov/27613609","citation_count":89,"is_preprint":false},{"pmid":"34303764","id":"PMC_34303764","title":"Fascin promotes lung cancer growth and metastasis by enhancing glycolysis and PFKFB3 expression.","date":"2021","source":"Cancer letters","url":"https://pubmed.ncbi.nlm.nih.gov/34303764","citation_count":83,"is_preprint":false},{"pmid":"24515104","id":"PMC_24515104","title":"Estradiol stimulates glucose metabolism via 6-phosphofructo-2-kinase (PFKFB3).","date":"2014","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/24515104","citation_count":81,"is_preprint":false},{"pmid":"34019671","id":"PMC_34019671","title":"The Role of HIF1α-PFKFB3 Pathway in Diabetic Retinopathy.","date":"2021","source":"The Journal of clinical endocrinology and metabolism","url":"https://pubmed.ncbi.nlm.nih.gov/34019671","citation_count":80,"is_preprint":false},{"pmid":"35418167","id":"PMC_35418167","title":"IGFBP5 promotes diabetic kidney disease progression by enhancing PFKFB3-mediated endothelial glycolysis.","date":"2022","source":"Cell death & disease","url":"https://pubmed.ncbi.nlm.nih.gov/35418167","citation_count":77,"is_preprint":false},{"pmid":"29985086","id":"PMC_29985086","title":"The potential utility of PFKFB3 as a therapeutic target.","date":"2018","source":"Expert opinion on therapeutic targets","url":"https://pubmed.ncbi.nlm.nih.gov/29985086","citation_count":71,"is_preprint":false},{"pmid":"29559632","id":"PMC_29559632","title":"PFKFB3 blockade inhibits hepatocellular carcinoma growth by impairing DNA repair through AKT.","date":"2018","source":"Cell death & disease","url":"https://pubmed.ncbi.nlm.nih.gov/29559632","citation_count":68,"is_preprint":false},{"pmid":"31406177","id":"PMC_31406177","title":"Targeting PFKFB3 alleviates cerebral ischemia-reperfusion injury in mice.","date":"2019","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/31406177","citation_count":67,"is_preprint":false},{"pmid":"29393396","id":"PMC_29393396","title":"PFKFB3 is involved in breast cancer proliferation, migration, invasion and angiogenesis.","date":"2018","source":"International journal of oncology","url":"https://pubmed.ncbi.nlm.nih.gov/29393396","citation_count":64,"is_preprint":false},{"pmid":"35804016","id":"PMC_35804016","title":"PFKFB3 regulates cancer stemness through the hippo pathway in small cell lung carcinoma.","date":"2022","source":"Oncogene","url":"https://pubmed.ncbi.nlm.nih.gov/35804016","citation_count":62,"is_preprint":false},{"pmid":"33282864","id":"PMC_33282864","title":"Inhibition of PFKFB3 Hampers the Progression of Atherosclerosis and Promotes Plaque Stability.","date":"2020","source":"Frontiers in cell and developmental biology","url":"https://pubmed.ncbi.nlm.nih.gov/33282864","citation_count":57,"is_preprint":false},{"pmid":"28348059","id":"PMC_28348059","title":"The Glycolytic Enzyme PFKFB3 Is Involved in Estrogen-Mediated Angiogenesis via GPER1.","date":"2017","source":"The Journal of pharmacology and experimental therapeutics","url":"https://pubmed.ncbi.nlm.nih.gov/28348059","citation_count":56,"is_preprint":false},{"pmid":"32679452","id":"PMC_32679452","title":"PFKFB3 inhibitors as potential anticancer agents: Mechanisms of action, current developments, and structure-activity relationships.","date":"2020","source":"European journal of medicinal chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/32679452","citation_count":55,"is_preprint":false},{"pmid":"33400016","id":"PMC_33400016","title":"YAP promotes ocular neovascularization by modifying PFKFB3-driven endothelial glycolysis.","date":"2021","source":"Angiogenesis","url":"https://pubmed.ncbi.nlm.nih.gov/33400016","citation_count":55,"is_preprint":false},{"pmid":"32209481","id":"PMC_32209481","title":"c-Src Promotes Tumorigenesis and Tumor Progression by Activating PFKFB3.","date":"2020","source":"Cell reports","url":"https://pubmed.ncbi.nlm.nih.gov/32209481","citation_count":54,"is_preprint":false},{"pmid":"30171427","id":"PMC_30171427","title":"The Glycolytic Enzyme PFKFB3 Controls TNF-α-Induced Endothelial Proinflammatory Responses.","date":"2019","source":"Inflammation","url":"https://pubmed.ncbi.nlm.nih.gov/30171427","citation_count":53,"is_preprint":false},{"pmid":"36577299","id":"PMC_36577299","title":"Enhancement of glycolysis-dependent DNA repair regulated by FOXO1 knockdown via PFKFB3 attenuates hyperglycemia-induced endothelial oxidative stress injury.","date":"2022","source":"Redox biology","url":"https://pubmed.ncbi.nlm.nih.gov/36577299","citation_count":52,"is_preprint":false},{"pmid":"37199341","id":"PMC_37199341","title":"Role of PFKFB3-driven glycolysis in sepsis.","date":"2023","source":"Annals of medicine","url":"https://pubmed.ncbi.nlm.nih.gov/37199341","citation_count":51,"is_preprint":false},{"pmid":"24351650","id":"PMC_24351650","title":"The glycolytic enzyme PFKFB3/phosphofructokinase regulates autophagy.","date":"2013","source":"Autophagy","url":"https://pubmed.ncbi.nlm.nih.gov/24351650","citation_count":51,"is_preprint":false},{"pmid":"31671668","id":"PMC_31671668","title":"PFKFB3 Inhibition Attenuates Oxaliplatin-Induced Autophagy and Enhances Its Cytotoxicity in Colon Cancer Cells.","date":"2019","source":"International journal of molecular sciences","url":"https://pubmed.ncbi.nlm.nih.gov/31671668","citation_count":51,"is_preprint":false},{"pmid":"35419769","id":"PMC_35419769","title":"Treatment against glucose-dependent cancers through metabolic PFKFB3 targeting of glycolytic flux.","date":"2022","source":"Cancer metastasis reviews","url":"https://pubmed.ncbi.nlm.nih.gov/35419769","citation_count":49,"is_preprint":false},{"pmid":"29151977","id":"PMC_29151977","title":"PFKFB3 promotes proliferation, migration and angiogenesis in nasopharyngeal carcinoma.","date":"2017","source":"Journal of Cancer","url":"https://pubmed.ncbi.nlm.nih.gov/29151977","citation_count":48,"is_preprint":false},{"pmid":"33420377","id":"PMC_33420377","title":"Inhibition of PFKFB3 induces cell death and synergistically enhances chemosensitivity in endometrial cancer.","date":"2021","source":"Oncogene","url":"https://pubmed.ncbi.nlm.nih.gov/33420377","citation_count":47,"is_preprint":false},{"pmid":"37714438","id":"PMC_37714438","title":"cGAS-STING signaling pathway promotes hypoxia-induced renal fibrosis by regulating PFKFB3-mediated glycolysis.","date":"2023","source":"Free radical biology & medicine","url":"https://pubmed.ncbi.nlm.nih.gov/37714438","citation_count":46,"is_preprint":false},{"pmid":"39294445","id":"PMC_39294445","title":"SGLT2 inhibitors ameliorate NAFLD in mice via downregulating PFKFB3, suppressing glycolysis and modulating macrophage polarization.","date":"2024","source":"Acta pharmacologica Sinica","url":"https://pubmed.ncbi.nlm.nih.gov/39294445","citation_count":44,"is_preprint":false},{"pmid":"39434134","id":"PMC_39434134","title":"GLP-1R activation attenuates the progression of pulmonary fibrosis via disrupting NLRP3 inflammasome/PFKFB3-driven glycolysis interaction and histone lactylation.","date":"2024","source":"Journal of translational medicine","url":"https://pubmed.ncbi.nlm.nih.gov/39434134","citation_count":42,"is_preprint":false},{"pmid":"26337471","id":"PMC_26337471","title":"The expression pattern of PFKFB3 enzyme distinguishes between induced-pluripotent stem cells and cancer stem cells.","date":"2015","source":"Oncotarget","url":"https://pubmed.ncbi.nlm.nih.gov/26337471","citation_count":40,"is_preprint":false},{"pmid":"35830274","id":"PMC_35830274","title":"Suppression of myeloid PFKFB3-driven glycolysis protects mice from choroidal neovascularization.","date":"2022","source":"British journal of pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/35830274","citation_count":40,"is_preprint":false},{"pmid":"35155224","id":"PMC_35155224","title":"PFKFB3 Regulates Chemoresistance, Metastasis and Stemness via IAP Proteins and the NF-κB Signaling Pathway in Ovarian Cancer.","date":"2022","source":"Frontiers in oncology","url":"https://pubmed.ncbi.nlm.nih.gov/35155224","citation_count":36,"is_preprint":false},{"pmid":"28559290","id":"PMC_28559290","title":"A role for PFKFB3/iPFK2 in metformin suppression of adipocyte inflammatory responses.","date":"2017","source":"Journal of molecular endocrinology","url":"https://pubmed.ncbi.nlm.nih.gov/28559290","citation_count":36,"is_preprint":false},{"pmid":"30981500","id":"PMC_30981500","title":"PFKFB3/HIF-1α feedback loop modulates sorafenib resistance in hepatocellular carcinoma cells.","date":"2019","source":"Biochemical and biophysical research communications","url":"https://pubmed.ncbi.nlm.nih.gov/30981500","citation_count":35,"is_preprint":false},{"pmid":"35834356","id":"PMC_35834356","title":"Gene-dosage effect of Pfkfb3 on monocyte/macrophage biology in atherosclerosis.","date":"2022","source":"British journal of pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/35834356","citation_count":34,"is_preprint":false},{"pmid":"35057732","id":"PMC_35057732","title":"Overexpression of PFKFB3 promotes cell glycolysis and proliferation in renal cell carcinoma.","date":"2022","source":"BMC cancer","url":"https://pubmed.ncbi.nlm.nih.gov/35057732","citation_count":33,"is_preprint":false},{"pmid":"35209949","id":"PMC_35209949","title":"Long noncoding RNA LINC00930 promotes PFKFB3-mediated tumor glycolysis and cell proliferation in nasopharyngeal carcinoma.","date":"2022","source":"Journal of experimental & clinical cancer research : CR","url":"https://pubmed.ncbi.nlm.nih.gov/35209949","citation_count":33,"is_preprint":false},{"pmid":"36740704","id":"PMC_36740704","title":"Blockage of glycolysis by targeting PFKFB3 suppresses the development of infantile hemangioma.","date":"2023","source":"Journal of translational medicine","url":"https://pubmed.ncbi.nlm.nih.gov/36740704","citation_count":33,"is_preprint":false},{"pmid":"35087090","id":"PMC_35087090","title":"Aberrant upregulation of the glycolytic enzyme PFKFB3 in CLN7 neuronal ceroid lipofuscinosis.","date":"2022","source":"Nature communications","url":"https://pubmed.ncbi.nlm.nih.gov/35087090","citation_count":31,"is_preprint":false},{"pmid":"35131382","id":"PMC_35131382","title":"CYLD deficiency enhances metabolic reprogramming and tumor progression in nasopharyngeal carcinoma via PFKFB3.","date":"2022","source":"Cancer letters","url":"https://pubmed.ncbi.nlm.nih.gov/35131382","citation_count":31,"is_preprint":false},{"pmid":"37626891","id":"PMC_37626891","title":"PFKFB3-Mediated Glycolysis Boosts Fibroblast Activation and Subsequent Kidney Fibrosis.","date":"2023","source":"Cells","url":"https://pubmed.ncbi.nlm.nih.gov/37626891","citation_count":30,"is_preprint":false},{"pmid":"29327288","id":"PMC_29327288","title":"Expression of PFKFB3 and Ki67 in lung adenocarcinomas and targeting PFKFB3 as a therapeutic strategy.","date":"2018","source":"Molecular and cellular biochemistry","url":"https://pubmed.ncbi.nlm.nih.gov/29327288","citation_count":30,"is_preprint":false},{"pmid":"36405760","id":"PMC_36405760","title":"Increased stromal PFKFB3-mediated glycolysis in inflammatory bowel disease contributes to intestinal inflammation.","date":"2022","source":"Frontiers in immunology","url":"https://pubmed.ncbi.nlm.nih.gov/36405760","citation_count":30,"is_preprint":false},{"pmid":"40016338","id":"PMC_40016338","title":"Metabolic reprogramming in astrocytes prevents neuronal death through a UCHL1/PFKFB3/H4K8la positive feedback loop.","date":"2025","source":"Cell death and differentiation","url":"https://pubmed.ncbi.nlm.nih.gov/40016338","citation_count":28,"is_preprint":false},{"pmid":"34107992","id":"PMC_34107992","title":"HSF1 promotes endometriosis development and glycolysis by up-regulating PFKFB3 expression.","date":"2021","source":"Reproductive biology and endocrinology : RB&E","url":"https://pubmed.ncbi.nlm.nih.gov/34107992","citation_count":28,"is_preprint":false},{"pmid":"29620138","id":"PMC_29620138","title":"PFK15, a PFKFB3 antagonist, inhibits autophagy and proliferation in rhabdomyosarcoma cells.","date":"2018","source":"International journal of molecular medicine","url":"https://pubmed.ncbi.nlm.nih.gov/29620138","citation_count":28,"is_preprint":false},{"pmid":"38200582","id":"PMC_38200582","title":"Targeting the SphK1/S1P/PFKFB3 axis suppresses hepatocellular carcinoma progression by disrupting glycolytic energy supply that drives tumor angiogenesis.","date":"2024","source":"Journal of translational medicine","url":"https://pubmed.ncbi.nlm.nih.gov/38200582","citation_count":26,"is_preprint":false},{"pmid":"36326834","id":"PMC_36326834","title":"HIF1A-dependent induction of alveolar epithelial PFKFB3 dampens acute lung injury.","date":"2022","source":"JCI insight","url":"https://pubmed.ncbi.nlm.nih.gov/36326834","citation_count":26,"is_preprint":false},{"pmid":"29511345","id":"PMC_29511345","title":"Targeting PFKFB3 sensitizes chronic myelogenous leukemia cells to tyrosine kinase inhibitor.","date":"2018","source":"Oncogene","url":"https://pubmed.ncbi.nlm.nih.gov/29511345","citation_count":26,"is_preprint":false},{"pmid":"32775001","id":"PMC_32775001","title":"Ubiquitin-like protein FAT10 promotes osteosarcoma glycolysis and growth by upregulating PFKFB3 via stabilization of EGFR.","date":"2020","source":"American journal of cancer research","url":"https://pubmed.ncbi.nlm.nih.gov/32775001","citation_count":26,"is_preprint":false},{"pmid":"31678169","id":"PMC_31678169","title":"ROCK2 promotes osteosarcoma growth and metastasis by modifying PFKFB3 ubiquitination and degradation.","date":"2019","source":"Experimental cell research","url":"https://pubmed.ncbi.nlm.nih.gov/31678169","citation_count":25,"is_preprint":false},{"pmid":"34432198","id":"PMC_34432198","title":"PFKFB3 gene deletion in endothelial cells inhibits intraplaque angiogenesis and lesion formation in a murine model of venous bypass grafting.","date":"2021","source":"Angiogenesis","url":"https://pubmed.ncbi.nlm.nih.gov/34432198","citation_count":25,"is_preprint":false},{"pmid":"34193800","id":"PMC_34193800","title":"A Potential Oncogenic Role for PFKFB3 Overexpression in Gastric Cancer Progression.","date":"2021","source":"Clinical and translational gastroenterology","url":"https://pubmed.ncbi.nlm.nih.gov/34193800","citation_count":25,"is_preprint":false},{"pmid":"29435871","id":"PMC_29435871","title":"PI3K-Akt signaling controls PFKFB3 expression during human T-lymphocyte activation.","date":"2018","source":"Molecular and cellular biochemistry","url":"https://pubmed.ncbi.nlm.nih.gov/29435871","citation_count":23,"is_preprint":false},{"pmid":"38035106","id":"PMC_38035106","title":"Myeloid PFKFB3-mediated glycolysis promotes kidney fibrosis.","date":"2023","source":"Frontiers in immunology","url":"https://pubmed.ncbi.nlm.nih.gov/38035106","citation_count":22,"is_preprint":false},{"pmid":"35094634","id":"PMC_35094634","title":"Hyperglycemia induces miR-26-5p down-regulation to overexpress PFKFB3 and accelerate epithelial-mesenchymal transition in gastric cancer.","date":"2022","source":"Bioengineered","url":"https://pubmed.ncbi.nlm.nih.gov/35094634","citation_count":22,"is_preprint":false},{"pmid":"39096131","id":"PMC_39096131","title":"Hypoxia activates macrophage-NLRP3 inflammasome promoting atherosclerosis via PFKFB3-driven glycolysis.","date":"2024","source":"FASEB journal : official publication of the Federation of American Societies for Experimental Biology","url":"https://pubmed.ncbi.nlm.nih.gov/39096131","citation_count":21,"is_preprint":false},{"pmid":"34831136","id":"PMC_34831136","title":"Canonical and Non-Canonical Roles of PFKFB3 in Brain Tumors.","date":"2021","source":"Cells","url":"https://pubmed.ncbi.nlm.nih.gov/34831136","citation_count":21,"is_preprint":false},{"pmid":"31209811","id":"PMC_31209811","title":"Role of PFKFB3 and CD163 in Oral Squamous Cell Carcinoma Angiogenesis.","date":"2019","source":"Current medical science","url":"https://pubmed.ncbi.nlm.nih.gov/31209811","citation_count":21,"is_preprint":false},{"pmid":"35354250","id":"PMC_35354250","title":"The glycolytic enzyme PFKFB3 determines bone marrow endothelial progenitor cell damage after chemotherapy and irradiation.","date":"2022","source":"Haematologica","url":"https://pubmed.ncbi.nlm.nih.gov/35354250","citation_count":21,"is_preprint":false},{"pmid":"26681033","id":"PMC_26681033","title":"MicroRNA-26b inhibits osteosarcoma cell migration and invasion by down-regulating PFKFB3 expression.","date":"2015","source":"Genetics and molecular research : GMR","url":"https://pubmed.ncbi.nlm.nih.gov/26681033","citation_count":21,"is_preprint":false},{"pmid":"34094669","id":"PMC_34094669","title":"Necroptosis pathway blockage attenuates PFKFB3 inhibitor-induced cell viability loss and genome instability in colorectal cancer cells.","date":"2021","source":"American journal of cancer research","url":"https://pubmed.ncbi.nlm.nih.gov/34094669","citation_count":21,"is_preprint":false},{"pmid":"36162723","id":"PMC_36162723","title":"PFKFB3-mediated Pro-glycolytic Shift in Hepatocellular Carcinoma Proliferation.","date":"2022","source":"Cellular and molecular gastroenterology and hepatology","url":"https://pubmed.ncbi.nlm.nih.gov/36162723","citation_count":20,"is_preprint":false},{"pmid":"36990998","id":"PMC_36990998","title":"Targeting Aurora-A inhibits tumor progression and sensitizes thyroid carcinoma to Sorafenib by decreasing PFKFB3-mediated glycolysis.","date":"2023","source":"Cell death & disease","url":"https://pubmed.ncbi.nlm.nih.gov/36990998","citation_count":20,"is_preprint":false},{"pmid":"37162556","id":"PMC_37162556","title":"Upregulation of glycolytic enzyme PFKFB3 by deubiquitinase OTUD4 promotes cardiac fibrosis post myocardial infarction.","date":"2023","source":"Journal of molecular medicine (Berlin, Germany)","url":"https://pubmed.ncbi.nlm.nih.gov/37162556","citation_count":19,"is_preprint":false},{"pmid":"37052859","id":"PMC_37052859","title":"The HIF-1α/miR-26a-5p/PFKFB3/ULK1/2 axis regulates vascular remodeling in hypoxia-induced pulmonary hypertension by modulation of autophagy.","date":"2023","source":"FASEB journal : official publication of the Federation of American Societies for Experimental Biology","url":"https://pubmed.ncbi.nlm.nih.gov/37052859","citation_count":19,"is_preprint":false},{"pmid":"37682013","id":"PMC_37682013","title":"PFKFB3-driven vascular smooth muscle cell glycolysis promotes vascular calcification via the altered FoxO3 and lactate production.","date":"2023","source":"FASEB journal : official publication of the Federation of American Societies for Experimental Biology","url":"https://pubmed.ncbi.nlm.nih.gov/37682013","citation_count":18,"is_preprint":false},{"pmid":"35553342","id":"PMC_35553342","title":"The role of PFKFB3 in maintaining colorectal cancer cell proliferation and stemness.","date":"2022","source":"Molecular biology reports","url":"https://pubmed.ncbi.nlm.nih.gov/35553342","citation_count":18,"is_preprint":false},{"pmid":"34016793","id":"PMC_34016793","title":"κ-opioid receptor stimulation alleviates rat vascular smooth muscle cell calcification via PFKFB3-lactate signaling.","date":"2021","source":"Aging","url":"https://pubmed.ncbi.nlm.nih.gov/34016793","citation_count":18,"is_preprint":false},{"pmid":"36064579","id":"PMC_36064579","title":"ALK fusion promotes metabolic reprogramming of cancer cells by transcriptionally upregulating PFKFB3.","date":"2022","source":"Oncogene","url":"https://pubmed.ncbi.nlm.nih.gov/36064579","citation_count":17,"is_preprint":false},{"pmid":"35107852","id":"PMC_35107852","title":"Targeting of PFKFB3 with miR-206 but not mir-26b inhibits ovarian cancer cell proliferation and migration involving FAK downregulation.","date":"2022","source":"FASEB journal : official publication of the Federation of American Societies for Experimental Biology","url":"https://pubmed.ncbi.nlm.nih.gov/35107852","citation_count":17,"is_preprint":false},{"pmid":"38341583","id":"PMC_38341583","title":"PFKFB3 in neovascular eye disease: unraveling mechanisms and exploring therapeutic strategies.","date":"2024","source":"Cell & bioscience","url":"https://pubmed.ncbi.nlm.nih.gov/38341583","citation_count":16,"is_preprint":false},{"pmid":"34359849","id":"PMC_34359849","title":"PFKFB3 Inhibition Impairs Erlotinib-Induced Autophagy in NSCLCs.","date":"2021","source":"Cells","url":"https://pubmed.ncbi.nlm.nih.gov/34359849","citation_count":16,"is_preprint":false},{"pmid":"31880389","id":"PMC_31880389","title":"Inhibition of PFKFB3 suppresses osteoclastogenesis and prevents ovariectomy-induced bone loss.","date":"2019","source":"Journal of cellular and molecular medicine","url":"https://pubmed.ncbi.nlm.nih.gov/31880389","citation_count":16,"is_preprint":false},{"pmid":"30378281","id":"PMC_30378281","title":"Discovery and Structure-Activity Relationships of N-Aryl 6-Aminoquinoxalines as Potent PFKFB3 Kinase Inhibitors.","date":"2018","source":"ChemMedChem","url":"https://pubmed.ncbi.nlm.nih.gov/30378281","citation_count":16,"is_preprint":false},{"pmid":"39301662","id":"PMC_39301662","title":"Advances in the understanding of the role and mechanism of action of PFKFB3‑mediated glycolysis in liver fibrosis (Review).","date":"2024","source":"International journal of molecular medicine","url":"https://pubmed.ncbi.nlm.nih.gov/39301662","citation_count":15,"is_preprint":false},{"pmid":"33777937","id":"PMC_33777937","title":"PFKFB3: A Potential Key to Ocular Angiogenesis.","date":"2021","source":"Frontiers in cell and developmental biology","url":"https://pubmed.ncbi.nlm.nih.gov/33777937","citation_count":15,"is_preprint":false},{"pmid":"30776389","id":"PMC_30776389","title":"PFKFB3 promotes endotoxemia-induced myocardial dysfunction through inflammatory signaling and apoptotic induction.","date":"2019","source":"Toxicology and applied pharmacology","url":"https://pubmed.ncbi.nlm.nih.gov/30776389","citation_count":15,"is_preprint":false},{"pmid":"12963966","id":"PMC_12963966","title":"Cloning and chromosomal characterization of the 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase-3 gene (PFKFB3, iPFK2).","date":"2003","source":"International journal of oncology","url":"https://pubmed.ncbi.nlm.nih.gov/12963966","citation_count":15,"is_preprint":false},{"pmid":"39953547","id":"PMC_39953547","title":"Micheliolide attenuates sepsis-induced acute lung injury by suppressing mitochondrial oxidative stress and PFKFB3-driven glycolysis.","date":"2025","source":"Journal of translational medicine","url":"https://pubmed.ncbi.nlm.nih.gov/39953547","citation_count":14,"is_preprint":false},{"pmid":"35135735","id":"PMC_35135735","title":"Nuances of PFKFB3 Signaling in Breast Cancer.","date":"2022","source":"Clinical breast cancer","url":"https://pubmed.ncbi.nlm.nih.gov/35135735","citation_count":14,"is_preprint":false},{"pmid":"36243313","id":"PMC_36243313","title":"PFKFB3 mediates tubular cell death in cisplatin nephrotoxicity by activating CDK4.","date":"2022","source":"Translational research : the journal of laboratory and clinical medicine","url":"https://pubmed.ncbi.nlm.nih.gov/36243313","citation_count":14,"is_preprint":false},{"pmid":"38705396","id":"PMC_38705396","title":"MiR-106a-5p targets PFKFB3 and improves sepsis through regulating macrophage pyroptosis and inflammatory response.","date":"2024","source":"The Journal of biological chemistry","url":"https://pubmed.ncbi.nlm.nih.gov/38705396","citation_count":14,"is_preprint":false},{"pmid":"27387960","id":"PMC_27387960","title":"Glucose and Palmitate Differentially Regulate PFKFB3/iPFK2 and Inflammatory Responses in Mouse Intestinal Epithelial Cells.","date":"2016","source":"Scientific reports","url":"https://pubmed.ncbi.nlm.nih.gov/27387960","citation_count":14,"is_preprint":false},{"pmid":"34346780","id":"PMC_34346780","title":"Endothelial cell PHD2-HIF1α-PFKFB3 contributes to right ventricle vascular adaptation in pulmonary hypertension.","date":"2021","source":"American journal of physiology. Lung cellular and molecular physiology","url":"https://pubmed.ncbi.nlm.nih.gov/34346780","citation_count":14,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":55635,"output_tokens":12110,"usd":0.174278,"stage1_stop_reason":"end_turn"},"stage2":{"model":"claude-opus-4-8","input_tokens":23214,"output_tokens":6189,"usd":0.135398,"stage2_stop_reason":"end_turn"},"total_usd":0.309676,"stage1_batch_id":"msgbatch_01CNoDbkChHEkgrJnL7BwtGJ","stage2_batch_id":"msgbatch_016Y4x9E9Wg5VQDANj9quSQD","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2013,\n      \"finding\": \"PFKFB3 controls endothelial cell filopodia/lamellipodia formation and directional migration by compartmentalizing with F-actin in motile protrusions, and overrides the pro-stalk activity of Notch signaling to regulate vessel branching tip/stalk cell competition.\",\n      \"method\": \"Loss-of-function (siRNA/genetic KO), mosaic in vitro and in vivo sprouting assays, subcellular co-localization with F-actin, epistasis with Notch pathway\",\n      \"journal\": \"Cell\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (genetic KO, mosaic assays, localization, epistasis), replicated in vitro and in vivo, high-citation landmark study\",\n      \"pmids\": [\"23911327\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"PFKFB3 (iPFK-2) is induced by hypoxia in cancer cells and its expression and fructose-2,6-bisphosphate levels increase specifically during S phase of the cell cycle, functionally coupling glycolytic activation to cell cycle progression.\",\n      \"method\": \"In situ hybridization, immunohistochemistry, cell-cycle fractionation, hypoxia treatment of cultured cells\",\n      \"journal\": \"Cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — multiple cell-based methods in single lab; no in vitro reconstitution or mutagenesis\",\n      \"pmids\": [\"12384552\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2015,\n      \"finding\": \"During mitotic arrest, AMPK directly phosphorylates PFKFB3, increasing glycolysis to replace lost oxidative phosphorylation (caused by mitophagy-dependent mitochondrial loss), thereby promoting cell survival. PFKFB3 protein levels also increase due to mitotic-specific translational activation of its mRNA.\",\n      \"method\": \"AMPK kinase assay, phospho-specific immunoblotting, siRNA knockdown, autophagy induction/inhibition, breast cancer cell viability assays\",\n      \"journal\": \"Nature cell biology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — direct phosphorylation demonstrated, multiple orthogonal methods (kinase assay, KD, translational activation), published in Nature Cell Biology\",\n      \"pmids\": [\"26322680\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"The lncRNA AGPG binds to and stabilizes PFKFB3 protein by preventing APC/C-mediated ubiquitination, thereby protecting PFKFB3 from proteasomal degradation and leading to accumulation of PFKFB3 and enhanced glycolytic flux in cancer cells.\",\n      \"method\": \"Co-immunoprecipitation, ubiquitination assay, proteasome inhibition, siRNA/overexpression, PDX tumor models\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP, ubiquitination assay, multiple orthogonal methods, in vivo PDX validation\",\n      \"pmids\": [\"32198345\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Cisplatin induces acetylation of PFKFB3 at lysine 472 (K472), which impairs its nuclear localization signal (NLS) activity, causing cytoplasmic accumulation of PFKFB3. Cytoplasmic PFKFB3 is then phosphorylated by AMPK, leading to PFKFB3 activation and enhanced glycolysis that protects cells from DNA damage-induced apoptosis.\",\n      \"method\": \"Site-directed mutagenesis (K472 acetylation-dead mutant), subcellular fractionation, AMPK phosphorylation assay, siRNA knockdown, xenograft model\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — mutagenesis of specific residue combined with fractionation and kinase assay, in vivo validation, rigorous mechanistic dissection\",\n      \"pmids\": [\"29410405\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"PFKFB3 promotes cell cycle progression and suppresses apoptosis via its product F2,6BP activating Cdk1, which phosphorylates p27 (T187) leading to p27 ubiquitination and proteasomal degradation. siRNA silencing of PFKFB3 inhibits Cdk1 activity, stabilizes p27, and causes G1/S arrest; co-silencing of p27 reverses these effects.\",\n      \"method\": \"siRNA silencing, co-siRNA epistasis, Cdk1 kinase assay, p27 protein stability/ubiquitination assay, flow cytometry (cell cycle/apoptosis), HeLa cells\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — epistasis experiment (rescue by co-KD), kinase activity assay, multiple readouts, mechanistic pathway established\",\n      \"pmids\": [\"25032860\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2012,\n      \"finding\": \"NMDAR activation stabilizes PFKFB3 protein in neurons (normally degraded by APC/C-Cdh1 proteasomal pathway via its KEN motif) and promotes PFKFB3 release from the nucleus to cytosol, switching neuronal metabolism from pentose-phosphate pathway (PPP) to glycolysis, causing oxidative stress and apoptotic neuronal death.\",\n      \"method\": \"NMDAR stimulation in cortical neurons, APC/C-Cdh1 KEN-motif deletion mutant, subcellular fractionation, PPP/glycolysis flux measurements, overexpression of G6PD rescue experiment\",\n      \"journal\": \"Cell death and differentiation\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1-2 / Strong — mutagenesis (KEN motif deletion), pathway epistasis (G6PD rescue), subcellular fractionation with functional consequence, multiple orthogonal approaches\",\n      \"pmids\": [\"22421967\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"PFKFB3 rapidly relocates into ionizing radiation-induced nuclear foci in an MRN-ATM-γH2AX-MDC1-dependent manner and is critical for recruitment of homologous recombination (HR) repair proteins and HR activity. PFKFB3 enzymatic activity is required for ribonucleotide reductase M2 (RRM2) recruitment, deoxynucleotide incorporation during DNA repair, and maintenance of dNTP levels.\",\n      \"method\": \"Immunofluorescence co-localization with DNA damage/HR foci, pharmacological inhibitor (KAN0438757), siRNA, HR reporter assay, dNTP level measurements, radiosensitization assays in transformed vs. non-transformed cells\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple orthogonal methods (co-localization, HR assay, dNTP measurement, selective inhibitor), functional mechanistic dissection\",\n      \"pmids\": [\"30250201\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2006,\n      \"finding\": \"siRNA silencing of PFKFB3 in HeLa cells decreases fructose-2,6-bisphosphate, lactate, and ATP, leading to reduced cell viability, cell cycle delay, increased apoptosis, and inhibition of anchorage-independent growth.\",\n      \"method\": \"siRNA knockdown, metabolite measurement (F2,6BP, lactate, ATP), flow cytometry (cell cycle, apoptosis), soft agar colony formation\",\n      \"journal\": \"FEBS letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — clean KD with defined metabolic and proliferative phenotype, single lab, multiple readouts\",\n      \"pmids\": [\"16698023\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"CPEB4 binds cytoplasmic polyadenylation elements (CPEs) in the 3'-UTR of PFKFB3 mRNA to promote its polyadenylation and translational upregulation (not transcriptional), thereby increasing PFKFB3 protein and glycolysis during hepatic stellate cell activation and liver fibrosis.\",\n      \"method\": \"CPEB4 siRNA knockdown, CPEB4-KO mice, RIP/pulldown of PFKFB3 mRNA by CPEB4, mRNA polyadenylation assay, PFKFB3 protein levels, primary HSC and LX2 cells\",\n      \"journal\": \"Gastroenterology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — direct RNA binding demonstrated (RIP), CPEB4-KO in vivo validation, distinction from transcriptional regulation made explicitly, multiple cell types and models\",\n      \"pmids\": [\"32169429\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"c-Src phosphorylates PFKFB3 at tyrosine 194 (Tyr194), activating PFKFB3 and stimulating glycolysis. PFKFB3-Y194F knockin mice show impaired glycolysis and attenuated spontaneous colon tumor formation when crossed with APCmin/+ mice.\",\n      \"method\": \"In vitro kinase assay (c-Src + PFKFB3), site-directed mutagenesis (Y194F), PFKFB3-Y194F knockin mice, APCmin/+ genetic cross, cell proliferation and xenograft assays, clinical tumor sample correlation\",\n      \"journal\": \"Cell reports\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Strong — in vitro kinase assay, mutagenesis at specific residue, knockin mouse model with in vivo epistasis, multiple orthogonal methods\",\n      \"pmids\": [\"32209481\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2014,\n      \"finding\": \"Estradiol (E2) promotes PFKFB3 mRNA transcription through direct estrogen receptor (ER) binding to the PFKFB3 promoter, leading to increased PFKFB3 protein, elevated F2,6BP, and enhanced glucose uptake and glycolysis in ER+ breast cancer cells.\",\n      \"method\": \"ER-chromatin binding (promoter assay), siRNA, PFKFB3 inhibitor, 14C-glucose uptake, F2,6BP measurement, apoptosis assay in MCF-7 cells\",\n      \"journal\": \"The Journal of biological chemistry\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ER-promoter binding demonstrated, siRNA + inhibitor with metabolic readouts, single lab\",\n      \"pmids\": [\"24515104\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"Autophagy machinery, specifically the UBA domain of p62/sequestosome-1, physically interacts with PFKFB3; autophagy reduces PFKFB3 expression and induces cellular dormancy, whereas impairment of autophagy (knockdown of Atg3, Atg7, or p62) restores PFKFB3 expression and reactivates proliferation in dormant breast cancer stem cells.\",\n      \"method\": \"Co-immunoprecipitation (PFKFB3-p62 interaction), siRNA (Atg3, Atg7, p62), microarray, cell dormancy/proliferation assays, in vivo metastasis model\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — reciprocal Co-IP identifying specific domain (UBA), genetic epistasis via multiple Atg gene KDs, in vivo validation\",\n      \"pmids\": [\"31413316\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Nuclear PFKFB3 silencing in hepatocellular carcinoma decreases AKT phosphorylation and reduces ERCC1 expression, impairing DNA repair and causing G2/M arrest and apoptosis, revealing a non-glycolytic nuclear function.\",\n      \"method\": \"siRNA knockdown, AKT phosphorylation western blot, ERCC1 expression, cell cycle analysis, xenograft model\",\n      \"journal\": \"Cell death & disease\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — KD with defined molecular pathway (AKT-ERCC1), in vivo xenograft, single lab\",\n      \"pmids\": [\"29559632\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"PFKFB3-driven glycolysis in pulmonary endothelial cells promotes pulmonary hypertension; endothelial PFKFB3 knockout reduces HIF2A levels, decreases production of growth factors (PDGFB, FGF2) and proinflammatory factors (CXCL12, IL-1β), and prevents vascular smooth muscle cell proliferation and leukocyte recruitment.\",\n      \"method\": \"Conditional endothelial Pfkfb3-KO mice (constitutive and inducible), heterozygous global KO, 3PO pharmacological inhibition, Western blot, immunostaining of lung ECs\",\n      \"journal\": \"Proceedings of the National Academy of Sciences\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — multiple genetic models (constitutive, inducible, heterozygous KO), mechanistic link to HIF2A and downstream paracrine factors, in vivo and ex vivo validation\",\n      \"pmids\": [\"31213542\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"PFKFB3-mediated tubular glycolysis increases kidney lactate levels, which drives histone lactylation (particularly H4K12la) at the promoters of NF-κB pathway genes (Ikbkb, Rela, Relb), activating their transcription and promoting renal inflammation and fibrosis. PFKFB3 also directly activates IKKβ, IκBα, and p65.\",\n      \"method\": \"PTC-specific PFKFB3-KO mice, ChIP-seq (H4K12la at gene promoters), kidney IRI model, heterozygous KO and pharmacological inhibition, metabolite measurement (lactate), Western blot\",\n      \"journal\": \"Kidney international\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — ChIP-seq with specific histone mark, cell-type-specific KO, multiple genetic models, mechanistic pathway from metabolite (lactate) to histone modification to gene expression\",\n      \"pmids\": [\"38789037\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"PFKFB3-driven glycolysis in endothelial cells drives EndoMT by hijacking glucose flux from the pentose phosphate pathway, reducing cytoplasmic NADPH production. Mitochondrial NADPH efflux via the isocitrate/α-ketoglutarate shuttle replenishes cytoplasmic NADPH but impairs mitochondrial respiration by hampering iron-sulfur cluster biosynthesis.\",\n      \"method\": \"PFKFB3 haplodeficiency and overexpression, NADPH flux measurement, mitochondrial respiration assay (Seahorse), iron-sulfur cluster biosynthesis analysis, pharmacological inhibition with salvianolic acid C\",\n      \"journal\": \"Signal transduction and targeted therapy\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic haplodeficiency and overexpression with metabolic flux measurements, single lab, multiple metabolic readouts\",\n      \"pmids\": [\"36045132\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"PFKFB3 inhibition by small molecule AZ67 prevents NMDAR excitotoxicity-induced NADPH oxidation, redox stress, and apoptotic neuronal death; in vivo administration of AZ67 alleviates motor discoordination and brain infarct in middle carotid artery occlusion ischemia/reperfusion model in mice.\",\n      \"method\": \"Pharmacological inhibition (AZ67), NADPH measurement, apoptosis assay in primary neurons, oxygen-glucose deprivation model, in vivo MCAO mouse model\",\n      \"journal\": \"Scientific reports\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — in vitro and in vivo validation with pharmacological inhibitor, mechanistic link to NADPH/redox, single lab\",\n      \"pmids\": [\"31406177\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"In CLN7 neuronal ceroid lipofuscinosis, failure of autophagy causes mitochondrial accumulation and elevated mitochondrial ROS (mROS), which signal to stabilize PFKFB3 (normally proteasomally degraded in healthy neurons) via a mROS-dependent protein stabilization cascade, driving aberrant glycolysis in neurons and contributing to disease pathogenesis.\",\n      \"method\": \"Cln7∆ex2 mouse model, in vivo genetic mROS measurement, PFKFB3 protein stability assays, AZ67 inhibitor treatment in vivo and in CLN7 patient-derived cells\",\n      \"journal\": \"Nature communications\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — in vivo genetic model, patient-derived cell validation, in vivo pharmacological rescue, mechanistic cascade (mROS → PFKFB3 stabilization)\",\n      \"pmids\": [\"35087090\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"PFKFB3 interacts with cyclin-dependent kinase 4 (CDK4) in the nucleus of kidney tubular cells following cisplatin treatment, leading to CDK4 activation and consequent phosphorylation/inactivation of the retinoblastoma tumor suppressor (Rb), causing apoptosis; this is independent of PFKFB3's canonical glycolytic function.\",\n      \"method\": \"Co-immunoprecipitation (PFKFB3-CDK4), CDK4 kinase activity assay, Rb phosphorylation immunoblot, renal proximal tubule-specific PFKFB3-KO mice, CDK4 inhibition rescue\",\n      \"journal\": \"Translational research\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — Co-IP of PFKFB3-CDK4 complex, kinase activity assay, cell-type-specific KO, epistasis with CDK4 inhibition, identifies non-canonical nuclear function\",\n      \"pmids\": [\"36243313\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"OTUD4 deubiquitinase binds PFKFB3 and blocks its ubiquitination-mediated proteasomal degradation, stabilizing PFKFB3 protein in cardiac fibroblasts stimulated with TGF-β1 and promoting glycolysis-driven fibroblast activation.\",\n      \"method\": \"Co-immunoprecipitation (PFKFB3-OTUD4), ubiquitination assay, siRNA knockdown, cardiac fibroblast activation assays, post-MI mouse model\",\n      \"journal\": \"Journal of molecular medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — Co-IP and ubiquitination assay identify deubiquitinase-substrate relationship, in vivo model, single lab\",\n      \"pmids\": [\"37162556\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"ROCK2 stabilizes PFKFB3 protein in osteosarcoma cells by modifying its ubiquitination and reducing proteasomal degradation; PFKFB3 is epistatic to ROCK2 in promoting proliferation and metastasis, as PFKFB3 overexpression rescues ROCK2 knockdown-induced growth impairment.\",\n      \"method\": \"siRNA knockdown, PFKFB3 overexpression rescue, ubiquitination assay, proliferation/invasion assays, clinical tissue correlation\",\n      \"journal\": \"Experimental cell research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2-3 / Moderate — ubiquitination assay and epistasis experiment, single lab, no detailed mapping of modification sites\",\n      \"pmids\": [\"31678169\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"CYLD stabilizes p53 and promotes its nuclear translocation by removing K63- and K48-linked ubiquitin chains from p53, enabling p53 to bind the PFKFB3 promoter and inhibit its transcription. CYLD also interacts with FZR1 to promote APC/C-FZR1 E3 ligase activity, which ubiquitinates and degrades PFKFB3 via 26S proteasomal system.\",\n      \"method\": \"Co-immunoprecipitation (CYLD-FZR1, CYLD-p53), ubiquitination assay (K48/K63 chains on p53), ChIP (p53 binding to PFKFB3 promoter), proteasome inhibition, siRNA\",\n      \"journal\": \"Cancer letters\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — two distinct mechanisms (transcriptional repression and proteolysis), ChIP and Co-IP data, single lab\",\n      \"pmids\": [\"35131382\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"Fascin promotes PFKFB3 transcription by activating YAP1 through its canonical actin-bundling activity; YAP1 binds a TEAD1/4 binding motif 30 bp upstream of the PFKFB3 transcription start site to increase PFKFB3 expression and glycolysis in lung cancer cells.\",\n      \"method\": \"ChIP (YAP1-TEAD at PFKFB3 promoter), fascin actin-bundling mutant, YAP1 siRNA, PFKFB3 promoter luciferase, xenograft metastasis models, organoid cultures\",\n      \"journal\": \"Cancer letters\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — ChIP at specific promoter element, actin-bundling mutant epistasis, in vivo xenograft, luciferase reporter, multiple models\",\n      \"pmids\": [\"34303764\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"YAP acts as a transcriptional co-activator with TEAD1 to bind the PFKFB3 promoter and increase PFKFB3 expression under hypoxia in endothelial cells, thereby promoting glycolysis and angiogenesis; YAP silencing inhibits endothelial glycolysis and can be rescued by enforced PFKFB3 expression.\",\n      \"method\": \"ChIP (YAP-TEAD1 binding to PFKFB3 promoter), siRNA, PFKFB3 overexpression rescue, Seahorse glycolysis assay, intravitreal siRNA injection in CNV/OIR mouse models\",\n      \"journal\": \"Angiogenesis\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP and epistasis, in vivo animal model, single lab, independent from PMID:34303764 which provides orthogonal replication in cancer context\",\n      \"pmids\": [\"33400016\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"PFKFB3-driven glycolysis in macrophages activates HIF-1α/HIF-2α and NF-κB, inducing M1/M2 polarization markers and pro-angiogenic cytokines, thereby reprogramming macrophages toward an angiogenic phenotype that promotes choroidal neovascularization; myeloid-specific PFKFB3 KO attenuates this process.\",\n      \"method\": \"Myeloid cell-specific PFKFB3 KO mice, bone marrow-derived macrophage stimulation, PFKFB3 inhibitor (AZ67), HIF-1α/HIF-2α and NF-κB pathway immunoblotting, laser-induced CNV model\",\n      \"journal\": \"British journal of pharmacology\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — myeloid cell-specific genetic KO, pharmacological validation, defined downstream pathway (HIF-1α/2α, NF-κB), in vivo disease model\",\n      \"pmids\": [\"35830274\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"PFKFB3-driven glycolysis is required for actin polymerization in macrophages; homozygous loss of Pfkfb3 impairs macrophage efferocytosis by disrupting actin-dependent cytoskeletal function, exacerbating atherosclerosis.\",\n      \"method\": \"Homozygous Pfkfb3-KO Apoe-/- mice, efferocytosis assay, actin polymerization measurement, flow cytometry\",\n      \"journal\": \"British journal of pharmacology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic KO with specific cellular phenotype (efferocytosis) linked to actin, in vivo atherosclerosis model, single study\",\n      \"pmids\": [\"35834356\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"ALK fusion oncoproteins promote PFKFB3 transcription through the downstream transcription factor STAT3; pharmacological or genetic blockade of ALK reduces PFKFB3 expression and glycolysis, and PFKFB3 inhibition can overcome TKI resistance in ALK-mutant cancer cells.\",\n      \"method\": \"Quantitative proteomics, STAT3 ChIP/reporter assay, ALK TKI treatment, PFKFB3 siRNA and overexpression, correlation in clinical ALK+ NSCLC samples\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP/reporter assay for STAT3 at PFKFB3 promoter, proteomic identification, clinical correlation, single lab\",\n      \"pmids\": [\"36064579\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"The deubiquitinase UCHL1 stabilizes PFKFB3 in astrocytes by cleaving K48-linked ubiquitin chains; the UCHL1/PFKFB3 axis increases lactate production, which drives histone H4K8 lactylation (H4K8la), subsequently activating transcription of Uchl1 and glycolysis genes, forming a positive feedback loop that supports astrocytic metabolic reprogramming after spinal cord injury.\",\n      \"method\": \"Genetic deletion of Uchl1 and Pfkfb3, K48-ubiquitin chain cleavage assay, ChIP (H4K8la at gene promoters), lactate measurement, in vivo SCI model, scRNA-seq analysis\",\n      \"journal\": \"Cell death and differentiation\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — deubiquitinase assay, ChIP at specific histone mark/promoter, in vivo genetic models, single lab\",\n      \"pmids\": [\"40016338\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"In pulmonary arterial smooth muscle cells, HIF-1α binds the promoter of miR-26a-5p to inhibit its expression; miR-26a-5p directly targets PFKFB3 (validated by dual-luciferase assay), and PFKFB3 enhances phosphorylation of ULK1 to promote autophagy and cell proliferation in PAH.\",\n      \"method\": \"ChIP (HIF-1α at miR-26a-5p promoter), dual-luciferase reporter assay (miR-26a-5p targeting PFKFB3 3'UTR), ULK1 phosphorylation immunoblot, tandem mRFP-GFP-LC3B autophagy assay, adeno-miR-26a-5p in vivo rat PAH model\",\n      \"journal\": \"FASEB journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — direct target validation by luciferase + blot, ChIP for upstream regulation, in vivo rat model, single lab\",\n      \"pmids\": [\"37052859\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2003,\n      \"finding\": \"The human PFKFB3 gene spans 32.5 kb on chromosome 10p15.3-p15.2, contains 19 exons (15 normally expressed), and encodes a 590 amino acid, 66.9 kDa bifunctional protein with the highest kinase/phosphatase activity ratio among all PFKFB isozymes.\",\n      \"method\": \"Genomic sequencing, exon-intron junction determination, FISH chromosomal localization, open reading frame analysis\",\n      \"journal\": \"International journal of oncology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 1 / Weak — definitive gene structure by sequencing and FISH, but single characterization study with no functional validation of domains\",\n      \"pmids\": [\"12963966\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"X-ray crystal structures of PFKFB3 were used to guide structure-based design of N-aryl 6-aminoquinoxaline inhibitors that bind the kinase active site; the most potent compound showed IC50 of 14 nM for PFKFB3 kinase and 0.49 μM for F2,6BP production in HCT116 cells.\",\n      \"method\": \"X-ray crystallography, docking, structure-activity relationship (SAR), enzymatic IC50 assay, cellular F2,6BP assay\",\n      \"journal\": \"ChemMedChem\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 1 / Moderate — crystal structure with direct active-site binding demonstrated, enzymatic and cellular assay validation, structure-based design\",\n      \"pmids\": [\"30378281\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"LINC00930 lncRNA acts as a scaffold to recruit the RBBP5 and GCN5 complex to the PFKFB3 promoter, increasing H3K4 trimethylation and H3K9 acetylation at the PFKFB3 promoter to epigenetically activate PFKFB3 transcription and glycolysis in nasopharyngeal carcinoma.\",\n      \"method\": \"ChIP (H3K4me3, H3K9ac at PFKFB3 promoter), RNA-immunoprecipitation (RBBP5/GCN5 interaction with LINC00930), siRNA, in vivo tumor model\",\n      \"journal\": \"Journal of experimental & clinical cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — ChIP with specific histone marks, RIP for scaffold mechanism, in vivo validation, single lab\",\n      \"pmids\": [\"35209949\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"PFKFB3 inhibition reduces TNF-α-induced endothelial proinflammatory responses (cytokines, ICAM-1, monocyte adhesion/transmigration) by suppressing IKKβ phosphorylation, IκBα phosphorylation/degradation, NF-κB-p65 nuclear translocation, and NF-κB DNA-binding activity.\",\n      \"method\": \"siRNA knockdown, PFKFB3 inhibitor, cytokine antibody array, monocyte adhesion/transmigration assay, NF-κB EMSA, immunofluorescence (p65 localization), western blotting\",\n      \"journal\": \"Inflammation\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — multiple readouts (EMSA, immunofluorescence, western), siRNA and inhibitor convergent, single lab\",\n      \"pmids\": [\"30171427\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"PFKFB3/HIF-1α form a positive feedback loop in hepatocellular carcinoma: HIF-1α drives PFKFB3 expression, and exogenous PFKFB3 overexpression in turn upregulates HIF-1α protein levels; HIF-1α deficiency impairs PFKFB3-induced sorafenib resistance.\",\n      \"method\": \"HIF-1α blockade, PFKFB3 overexpression, HIF-1α and PFKFB3 protein immunoblotting, apoptosis assay, GEO dataset validation\",\n      \"journal\": \"Biochemical and biophysical research communications\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 / Weak — single lab, primarily western blot-based, no direct mechanistic link between PFKFB3 and HIF-1α protein stability identified\",\n      \"pmids\": [\"30981500\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"cGAS-STING-IRF3 signaling pathway promotes renal fibrosis by upregulating PFKFB3 expression under hypoxia; inhibition of STING or IRF3 reverses elevated PFKFB3, placing PFKFB3 downstream of cGAS-STING-IRF3 in hypoxia-induced fibrosis.\",\n      \"method\": \"STING and IRF3 pharmacological inhibition, hypoxia-stimulated HK-2 cells, IRI mouse model, PFKFB3 expression immunoblotting, fibrosis markers\",\n      \"journal\": \"Free radical biology & medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — epistasis (inhibition of upstream pathway reverses PFKFB3 induction), both in vitro and in vivo models, single lab\",\n      \"pmids\": [\"37714438\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"HIF1A directly induces PFKFB3 expression in alveolar epithelial cells; alveolar epithelium-specific Pfkfb3 deletion dramatically increases acute lung injury severity, and pyruvate (restoring metabolic substrate) rescues both Pfkfb3loxP/loxP and Hif1aloxP/loxP mice from ALI.\",\n      \"command\": \"\",\n      \"method\": \"Alveolar epithelium-specific Pfkfb3-KO and Hif1a-KO mice (SPC-ER-Cre), injurious ventilation and acid instillation ALI models, intratracheal pyruvate rescue, human lung biopsy immunostaining\",\n      \"journal\": \"JCI insight\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 / Strong — cell-type-specific genetic KO of both HIF1A and PFKFB3, in vivo rescue experiment, human tissue validation, establishes HIF1A→PFKFB3 axis in alveolar epithelia\",\n      \"pmids\": [\"36326834\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"PFKFB3 inhibition in small cell lung carcinoma attenuates invasion/migration by downregulating YAP/TAZ signaling while increasing pLATS1 via activation of pMST1 and NF2, linking PFKFB3-driven glycolysis to regulation of the Hippo pathway.\",\n      \"method\": \"PFK158 inhibitor, shRNA stable knockdown, Hippo pathway component immunoblotting (pMST1, NF2, pLATS1, YAP/TAZ), xenograft model\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — pharmacological and genetic inhibition with Hippo pathway readout, in vivo xenograft, single lab\",\n      \"pmids\": [\"35804016\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2019,\n      \"finding\": \"RANKL-induced osteoclastogenesis increases PFKFB3 expression and glycolysis; L-lactate (the glycolytic product) partially reverses the suppression of osteoclastogenesis caused by PFKFB3 inhibition and abrogates the inhibitory effect on NF-κB and MAPK pathways, establishing lactate as a downstream mediator of PFKFB3's pro-osteoclast function.\",\n      \"method\": \"siRNA, PFKFB3 inhibitor (PFK15), L-lactate supplementation rescue, NF-κB and MAPK immunoblotting, lactate/glucose measurement, ovariectomy bone loss mouse model\",\n      \"journal\": \"Journal of cellular and molecular medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — metabolite rescue experiment identifies lactate as mediator, pathway (NF-κB/MAPK) dissection, in vivo bone loss model, single lab\",\n      \"pmids\": [\"31880389\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"PFKFB3-derived lactate in renal fibroblasts promotes fibrotic activation; the glycolytic metabolite lactate directly promotes fibrotic phenotype in NRK-49F cells, and myofibroblast-specific PFKFB3 KO mice show substantially reduced fibrosis after UUO or IRI.\",\n      \"method\": \"Myofibroblast-specific Pfkfb3-KO mice (Pfkfb3f/f/PostnMCM), exogenous lactate supplementation, TGF-β1 stimulation, α-SMA and fibronectin markers, scRNA-seq reanalysis\",\n      \"journal\": \"Cells\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — cell-type-specific KO in vivo, lactate rescue experiment, mechanistic link to downstream fibrotic activation, single lab\",\n      \"pmids\": [\"37626891\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"SphK1/S1P signaling stabilizes PFKFB3 in endothelial cells to supply glycolytic energy for tumor angiogenesis; pharmacological SphK1 inhibition induces proteasomal degradation of PFKFB3, which can be reversed by S1P supplementation in an S1P receptor-dependent manner.\",\n      \"method\": \"SphK1 inhibitor (PF-543), Sphk1-KO mice, S1P supplementation rescue, PFKFB3 protein stability assay (proteasome inhibitor), DEN-induced primary HCC model, lentiviral SphK1 KD\",\n      \"journal\": \"Journal of translational medicine\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — genetic KO and S1P rescue establish pathway, proteasomal degradation mechanism, in vivo primary HCC model, single lab\",\n      \"pmids\": [\"38200582\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"In myeloid cells, PFKFB3-driven glycolysis stabilizes HIF1α, which alters macrophage phenotype to contribute to renal fibrosis; myeloid-specific PFKFB3-KO reduces M1 and M2 macrophage infiltration, suppresses macrophage-to-myofibroblast transition, and decreases kidney fibrosis.\",\n      \"method\": \"Myeloid-specific Pfkfb3-KO mice (Pfkfb3ΔMφ), UUO model, HIF1α stabilization assay, macrophage phenotyping by flow cytometry, scRNA-seq reanalysis\",\n      \"journal\": \"Frontiers in immunology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — cell-type-specific genetic KO, HIF1α mechanistic link, in vivo disease model, single lab\",\n      \"pmids\": [\"38035106\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2020,\n      \"finding\": \"FAT10 upregulates PFKFB3 in osteosarcoma by directly binding to EGFR and inhibiting EGFR ubiquitination and degradation, thereby stabilizing EGFR which then promotes PFKFB3 expression and glycolysis.\",\n      \"method\": \"Co-immunoprecipitation (FAT10-EGFR), ubiquitination assay, siRNA knockdown, PFKFB3 expression immunoblot, glycolysis measurement, osteosarcoma tissue correlation\",\n      \"journal\": \"American journal of cancer research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 / Moderate — Co-IP and ubiquitination assay identify FAT10-EGFR-PFKFB3 cascade, multiple complementary experiments, single lab\",\n      \"pmids\": [\"32775001\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"PFKFB3-driven glycolysis in vascular smooth muscle cells promotes calcification through FoxO3 expression and lactate production; silencing PFKFB3 reduces FoxO3, and pyruvate/lactate supplementation reverses PFKFB3-depletion effects on ALP activity and OPG expression, establishing lactate as an osteogenic mediator downstream of PFKFB3.\",\n      \"method\": \"RNA-seq after PFKFB3 KD, FoxO3 silencing epistasis, lactate/pyruvate supplementation rescue, miR-26a/b-5p overexpression, in vivo vitamin D3 calcification model, VSMC osteogenic transdifferentiation assays\",\n      \"journal\": \"FASEB journal\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — RNA-seq + epistasis (FoxO3 KD) + metabolite rescue, in vivo model, single lab\",\n      \"pmids\": [\"37682013\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"PFKFB3 overexpression in bone marrow endothelial progenitor cells facilitates pro-apoptotic transcription factor FOXO3A and its downstream genes (p21, p27, FAS), activates NF-κB and E-selectin expression, and reduces SDF-1, impairing hematopoiesis-supporting function after chemotherapy; FOXO3A silencing rescues these effects.\",\n      \"method\": \"PFKFB3 overexpression/knockdown in BM EPCs, FOXO3A silencing rescue experiment, NF-κB activation immunoblot, E-selectin and SDF-1 measurement, BM EC-specific PFKFB3 overexpression transgenic mice, 5-FU chemotherapy model, patient BM EPC analysis\",\n      \"journal\": \"Haematologica\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 / Moderate — transgenic in vivo model, FOXO3A epistasis rescue, multiple pathway readouts, clinical patient validation, single lab\",\n      \"pmids\": [\"35354250\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"PFKFB3 is a bifunctional enzyme with the highest kinase/phosphatase ratio among PFKFB family members that synthesizes fructose-2,6-bisphosphate (F2,6BP) to allosterically activate PFK-1 and drive glycolysis; its activity and stability are regulated by post-translational modifications including AMPK-mediated phosphorylation, c-Src-mediated tyrosine phosphorylation at Y194, acetylation at K472 (causing cytoplasmic redistribution), and proteasomal degradation via APC/C-Cdh1 (KEN motif) which is regulated by deubiquitinases (UCHL1, OTUD4) and stabilizing proteins (AGPG lncRNA, ROCK2, FAT10/EGFR axis); beyond glycolysis, PFKFB3 has non-canonical nuclear roles in homologous recombination DNA repair (via RRM2 recruitment and dNTP supply), CDK4 activation (causing Rb phosphorylation), and interaction with p62/autophagy machinery, while its downstream metabolite lactate drives histone lactylation (H4K12la, H4K8la) to epigenetically activate NF-κB pathway genes and fibrotic programs; transcriptionally, PFKFB3 is induced by HIF-1α, ER/GPER1, PI3K-Akt-mTOR, STAT3 (downstream of ALK), YAP-TEAD, and cGAS-STING-IRF3 pathways, and its mRNA is regulated post-transcriptionally by CPEB4-mediated polyadenylation.\"\n}\n```","stage2_raw":"{\n  \"mechanistic_narrative\": \"PFKFB3 is a bifunctional 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase that synthesizes fructose-2,6-bisphosphate (F2,6BP) to drive glycolytic flux, coupling glucose metabolism to proliferation, migration, survival, and DNA repair [#30, #8, #1]. Beyond sustaining metabolism, its glycolytic output directly governs cell behavior: in endothelial cells PFKFB3 compartmentalizes with F-actin in motile protrusions to control filopodia/lamellipodia formation and tip/stalk cell selection during vessel branching, overriding Notch [#0], and in macrophages it provides the glycolytic energy required for actin-dependent efferocytosis [#26]. PFKFB3 also executes non-canonical nuclear functions independent of bulk glycolysis: it relocates to ionizing-radiation-induced foci in an MRN-ATM-\\u03b3H2AX-MDC1-dependent manner where its enzymatic activity recruits RRM2 and maintains the dNTP pool needed for homologous recombination [#7], and it activates CDK1 (driving p27 degradation and G1/S progression) and CDK4 (driving Rb phosphorylation) [#5, #19]. Its terminal metabolite lactate acts as an epigenetic signal, driving histone lactylation (H4K12la, H4K8la) at NF-\\u03baB pathway and glycolytic gene promoters to amplify inflammatory and fibrotic programs [#15, #28]. PFKFB3 abundance and activity are tightly tuned post-translationally: AMPK and c-Src (at Tyr194) phosphorylate and activate it [#2, #4, #10], cisplatin-induced K472 acetylation disrupts its nuclear localization and redistributes it to the cytoplasm [#4], and its protein stability is set by a balance between APC/C-Cdh1-mediated proteasomal degradation via a KEN motif [#6] and competing stabilizers including the lncRNA AGPG, deubiquitinases UCHL1 and OTUD4, ROCK2, and the FAT10/EGFR axis [#3, #28, #20, #21, #42]. Transcriptionally and post-transcriptionally PFKFB3 is induced by HIF-1\\u03b1, estrogen receptor, YAP-TEAD, STAT3, and cGAS-STING-IRF3 signaling and by CPEB4-mediated cytoplasmic polyadenylation [#36, #11, #23, #27, #35, #9]. Through these mechanisms PFKFB3 promotes tumor growth, angiogenesis, neuronal excitotoxic death, and organ fibrosis across multiple tissues [#10, #14, #17, #39].\",\n  \"teleology\": [\n    {\n      \"year\": 2002,\n      \"claim\": \"Established that PFKFB3 expression and its product F2,6BP are induced by hypoxia and peak in S phase, first linking this glycolytic activator to cell-cycle progression and the tumor microenvironment.\",\n      \"evidence\": \"in situ hybridization, IHC, cell-cycle fractionation and hypoxia treatment of cancer cells\",\n      \"pmids\": [\"12384552\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No direct demonstration that S-phase F2,6BP is causally required for cycle progression\", \"Upstream hypoxia transcription factor not yet identified in this study\"]\n    },\n    {\n      \"year\": 2003,\n      \"claim\": \"Defined the human PFKFB3 gene structure and protein, establishing it as a bifunctional enzyme with the highest kinase/phosphatase ratio among isozymes, rationalizing its role as a net glycolysis activator.\",\n      \"evidence\": \"genomic sequencing, exon-intron mapping, FISH, ORF analysis\",\n      \"pmids\": [\"12963966\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No functional validation of catalytic domains\", \"Regulatory residues not mapped\"]\n    },\n    {\n      \"year\": 2006,\n      \"claim\": \"Showed by loss of function that PFKFB3 is required for F2,6BP, lactate and ATP production and supports viability and anchorage-independent growth, cementing it as a metabolic dependency in cancer cells.\",\n      \"evidence\": \"siRNA knockdown with metabolite measurement, cell-cycle/apoptosis flow cytometry, soft agar assay in HeLa\",\n      \"pmids\": [\"16698023\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Single cell line\", \"Mechanism downstream of metabolite depletion not dissected\"]\n    },\n    {\n      \"year\": 2012,\n      \"claim\": \"Revealed that PFKFB3 is normally degraded by APC/C-Cdh1 via a KEN motif and that NMDAR signaling stabilizes it and shifts it from nucleus to cytosol, switching neuronal metabolism from PPP to glycolysis and causing oxidative death \\u2014 the first non-proliferative, degradation-controlled role.\",\n      \"evidence\": \"NMDAR stimulation, KEN-motif deletion mutant, subcellular fractionation, PPP/glycolysis flux, G6PD rescue in cortical neurons\",\n      \"pmids\": [\"22421967\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"E2/E3 components beyond APC/C-Cdh1 not detailed\", \"Trigger linking NMDAR activity to Cdh1 inhibition not fully resolved\"]\n    },\n    {\n      \"year\": 2013,\n      \"claim\": \"Demonstrated PFKFB3 compartmentalizes with F-actin in motile protrusions and controls endothelial migration and tip/stalk competition over Notch, establishing a localized glycolytic role in cell motility and angiogenesis.\",\n      \"evidence\": \"siRNA/genetic KO, mosaic in vitro and in vivo sprouting assays, F-actin co-localization, Notch epistasis\",\n      \"pmids\": [\"23911327\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Molecular basis of PFKFB3-F-actin compartmentalization unresolved\", \"Whether ATP/F2,6BP locally fuels actin dynamics not directly shown\"]\n    },\n    {\n      \"year\": 2014,\n      \"claim\": \"Identified the CDK1\\u2013p27 axis as the mechanism by which PFKFB3 promotes cycle progression, and that estrogen receptor directly transactivates PFKFB3, connecting hormonal signaling to glycolytic cell-cycle control.\",\n      \"evidence\": \"siRNA + co-siRNA epistasis, CDK1 kinase assay, p27 ubiquitination (HeLa); ER-promoter binding, glucose uptake, F2,6BP in MCF-7\",\n      \"pmids\": [\"25032860\", \"24515104\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How F2,6BP activates CDK1 mechanistically not defined\", \"ER binding site on PFKFB3 promoter not finely mapped\"]\n    },\n    {\n      \"year\": 2015,\n      \"claim\": \"Showed AMPK directly phosphorylates PFKFB3 during mitotic arrest to raise glycolysis and compensate for mitophagy-driven loss of oxidative phosphorylation, establishing direct kinase regulation coupled to translational upregulation.\",\n      \"evidence\": \"AMPK kinase assay, phospho-immunoblot, siRNA, autophagy modulation, breast cancer viability assays\",\n      \"pmids\": [\"26322680\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Phosphosite identity and effect on kinase vs phosphatase activity not resolved here\", \"Mechanism of mitotic translational activation not defined\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Connected PTM-controlled localization to chemoresistance: cisplatin-induced K472 acetylation disrupts the NLS, driving cytoplasmic PFKFB3 that AMPK then activates, while a separate nuclear PFKFB3 pool supports AKT-ERCC1-dependent DNA repair.\",\n      \"evidence\": \"K472 acetylation-dead mutant, fractionation, AMPK assay, xenograft (Nat Commun); siRNA + AKT/ERCC1 readout in HCC (Cell Death Dis)\",\n      \"pmids\": [\"29410405\", \"29559632\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Acetyltransferase/deacetylase for K472 not identified\", \"Direct vs indirect link between nuclear PFKFB3 and AKT-ERCC1 unresolved\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Established a non-glycolytic nuclear role in DNA repair by showing PFKFB3 enters DNA-damage foci dependent on the MRN-ATM cascade and that its enzymatic activity is required for RRM2 recruitment and dNTP supply during homologous recombination.\",\n      \"evidence\": \"IF co-localization with HR foci, selective inhibitor KAN0438757, HR reporter, dNTP measurement, radiosensitization\",\n      \"pmids\": [\"30250201\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How a glycolytic enzyme is recruited to chromatin foci is unknown\", \"Relationship between local F2,6BP/dNTP pools and RRM2 not structurally defined\"]\n    },\n    {\n      \"year\": 2019,\n      \"claim\": \"Defined PFKFB3 as a node in inflammation and tissue-specific disease: it sustains NF-\\u03baB activation in endothelium, supports osteoclastogenesis via lactate, drives pulmonary hypertension through HIF2A/paracrine factors, and is suppressed by autophagy via p62 to enforce dormancy.\",\n      \"evidence\": \"siRNA/inhibitor with NF-\\u03baB EMSA (Inflammation); lactate rescue + bone-loss model (osteoclast); conditional endothelial KO (PNAS); p62 UBA Co-IP + Atg KD + metastasis model (Nat Commun)\",\n      \"pmids\": [\"30171427\", \"31880389\", \"31213542\", \"31413316\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Whether PFKFB3 acts on NF-\\u03baB via metabolite vs protein interaction varies across systems\", \"Direct p62-PFKFB3 functional consequence on degradation pathway not fully mapped\"]\n    },\n    {\n      \"year\": 2020,\n      \"claim\": \"Mapped multiple layers of PFKFB3 stabilization and activation: c-Src phosphorylation at Tyr194 (validated in knockin mice), AGPG lncRNA blocking APC/C ubiquitination, and CPEB4-driven cytoplasmic polyadenylation \\u2014 each independently amplifying glycolysis and tumor/fibrotic phenotypes.\",\n      \"evidence\": \"in vitro c-Src kinase assay + Y194F knockin/APCmin cross; AGPG Co-IP/ubiquitination + PDX; CPEB4 RIP + KO mice in liver fibrosis\",\n      \"pmids\": [\"32209481\", \"32198345\", \"32169429\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Interplay/hierarchy among Tyr194 phosphorylation, AGPG, and AMPK regulation not integrated\", \"Structural effect of Tyr194 phosphorylation on catalysis unresolved\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Positioned PFKFB3 within Hippo/YAP signaling bidirectionally \\u2014 YAP-TEAD transactivates PFKFB3 to fuel angiogenesis, while PFKFB3 inhibition feeds back to modulate MST1/NF2/LATS1 and YAP/TAZ \\u2014 linking glycolysis to a master growth-control pathway.\",\n      \"evidence\": \"YAP-TEAD1 ChIP at PFKFB3 promoter + rescue in OIR/CNV models; PFK158/shRNA with Hippo readouts + xenograft in SCLC\",\n      \"pmids\": [\"33400016\", \"35804016\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism by which PFKFB3 feeds back onto Hippo kinases unclear\", \"Direct vs metabolite-mediated effect on YAP not distinguished\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"Consolidated PFKFB3 as a stress-stabilized driver across neurodegeneration, angiogenesis, atherosclerosis, and cancer, controlled by diverse stabilizers (ROCK2, FAT10/EGFR), upstream inducers (fascin-YAP, ALK-STAT3, HIF1A), and a CDK4 non-glycolytic axis, with metabolic crosstalk to PPP/NADPH and mitochondrial function.\",\n      \"evidence\": \"Cln7 mouse + mROS stabilization; myeloid/endothelial/alveolar KO models; PFKFB3-CDK4 Co-IP; ubiquitination assays (ROCK2, FAT10-EGFR); NADPH/Seahorse flux; ChIP for fascin-YAP\",\n      \"pmids\": [\"35087090\", \"35830274\", \"35834356\", \"36243313\", \"31678169\", \"32775001\", \"34303764\", \"36064579\", \"36326834\", \"36045132\", \"35354250\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"How mROS chemically stabilizes PFKFB3 protein not defined\", \"Whether CDK4 binding involves catalytic activity unresolved\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Established lactate-driven epigenetic signaling as a PFKFB3 effector mechanism and added further upstream regulators (cGAS-STING-IRF3, SphK1/S1P, OTUD4, HIF1\\u03b1-miR-26a) controlling its abundance in fibrosis and angiogenesis.\",\n      \"evidence\": \"lactate rescue + myofibroblast/VSMC KO; STING/IRF3 inhibition; SphK1 KO + S1P rescue; OTUD4 Co-IP/ubiquitination; HIF-1\\u03b1-miR-26a-5p luciferase + ULK1 phosphorylation\",\n      \"pmids\": [\"37626891\", \"37682013\", \"37714438\", \"38200582\", \"37162556\", \"37052859\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Tissue specificity of competing stabilizers/regulators not reconciled\", \"Direct enzymatic targets vs lactate-mediated effects sometimes conflated\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Demonstrated directly that PFKFB3-derived lactate writes histone lactylation marks (H4K12la, H4K8la) at NF-\\u03baB and glycolytic gene promoters, providing a concrete metabolite-to-chromatin mechanism for PFKFB3-driven inflammation, fibrosis, and feed-forward metabolic reprogramming.\",\n      \"evidence\": \"PTC-specific and Uchl1/Pfkfb3 KO mice, ChIP-seq for H4K12la/H4K8la, injury models, lactate measurement\",\n      \"pmids\": [\"38789037\", \"40016338\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Lactylation writer/eraser enzymes acting downstream of PFKFB3 lactate not identified\", \"Promoter selectivity of lactylation not mechanistically explained\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"How PFKFB3's distinct cytosolic glycolytic, F-actin-associated, and nuclear (HR repair, CDK activation) functions are coordinately partitioned and regulated by its layered PTMs and competing stabilizers/degraders remains unresolved.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"Low\",\n      \"gaps\": [\"No integrated model of how localization, PTM state, and binding partners dictate which function PFKFB3 executes\", \"Structural basis for non-catalytic nuclear interactions (CDK4, p62) unknown\", \"Whether kinase vs phosphatase activity is differentially regulated by the various PTMs not resolved\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0016740\", \"supporting_discovery_ids\": [30, 31, 8]},\n      {\"term_id\": \"GO:0016787\", \"supporting_discovery_ids\": [30]},\n      {\"term_id\": \"GO:0016301\", \"supporting_discovery_ids\": [31]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005829\", \"supporting_discovery_ids\": [4, 6, 0]},\n      {\"term_id\": \"GO:0005634\", \"supporting_discovery_ids\": [6, 7, 13, 19]},\n      {\"term_id\": \"GO:0005856\", \"supporting_discovery_ids\": [0, 26]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-1430728\", \"supporting_discovery_ids\": [8, 30, 1]},\n      {\"term_id\": \"R-HSA-1640170\", \"supporting_discovery_ids\": [5, 1, 19]},\n      {\"term_id\": \"R-HSA-73894\", \"supporting_discovery_ids\": [7, 13]},\n      {\"term_id\": \"R-HSA-4839726\", \"supporting_discovery_ids\": [15, 28]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [25, 33, 41]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"RRM2\", \"CDK4\", \"AMPK\", \"SRC\", \"AGPG\", \"OTUD4\", \"UCHL1\", \"SQSTM1\"],\n    \"other_free_text\": []\n  }\n}","audit_flag":null,"evaluation":{"pairwise":"win","faith_supported":7,"faith_total":7,"faith_pct":100.0}}