{"gene":"CST1","run_date":"2026-04-28T17:28:53","timeline":{"discoveries":[{"year":2022,"finding":"CST1 interacts with GPX4 (a key ferroptosis regulator) and recruits the deubiquitinase OTUB1 to relieve GPX4 ubiquitination, thereby stabilizing GPX4 protein, reducing intracellular ROS, and inhibiting ferroptosis to promote gastric cancer metastasis.","method":"Co-immunoprecipitation combined with mass spectrometry, ubiquitination assay, overexpression/knockdown functional studies in vitro and in vivo (nude mouse metastasis model)","journal":"Oncogene","confidence":"High","confidence_rationale":"Tier 2 — reciprocal Co-IP + MS identification of complex + functional rescue experiments with multiple orthogonal readouts","pmids":["36369321"],"is_preprint":false},{"year":2021,"finding":"CST1 promotes gastric cancer cell migration and invasion by activating the Wnt signaling pathway, promoting nuclear translocation of β-catenin and upregulating Wnt target genes; inhibition of Wnt pathway in CST1-overexpressing cells suppresses these effects.","method":"Transwell migration/invasion assay, luciferase reporter assay for Wnt pathway activity, β-catenin nuclear translocation assessment, pharmacological Wnt inhibition rescue experiment","journal":"Cancer management and research","confidence":"Medium","confidence_rationale":"Tier 2 — epistasis via pathway inhibitor rescue + reporter assay, single lab","pmids":["33658852"],"is_preprint":false},{"year":2023,"finding":"CST1 promotes migration and invasion of esophageal squamous cell carcinoma (ESCC) cells by upregulating phosphorylation of MEK1/2, ERK1/2, and CREB in the MEK/ERK/CREB signaling pathway; miR-942-5p directly targets CST1 (validated by dual-luciferase assay) to downregulate this pathway.","method":"Transwell migration/invasion assay, Western blot for pathway effector phosphorylation, dual-luciferase reporter assay","journal":"PloS one","confidence":"Medium","confidence_rationale":"Tier 2/3 — functional KD with defined pathway readout + luciferase validation, single lab","pmids":["36848349"],"is_preprint":false},{"year":2022,"finding":"CST1 promotes proliferation and migration of airway smooth muscle cells (ASMCs) treated with PDGF-BB by activating the PI3K/AKT signaling pathway and upregulating MMP1 and MMP9; knockdown or overexpression of CST1 reciprocally modulates PI3K/AKT activity and cell behavior.","method":"siRNA knockdown, plasmid overexpression, EdU/CCK-8 proliferation assays, Transwell migration assay, Western blot for PI3K/AKT pathway components, PI3K/AKT inhibitor rescue experiment","journal":"The Kaohsiung journal of medical sciences","confidence":"Medium","confidence_rationale":"Tier 2 — pathway inhibitor rescue + multiple functional readouts, single lab","pmids":["36354198"],"is_preprint":false},{"year":2023,"finding":"CST1 overexpression in bronchial epithelial cells enhances AKT phosphorylation and upregulates SERPINB2 expression; anti-CST1 siRNA reverses these effects; AKT positively regulates SERPINB2, placing CST1 upstream of the AKT–SERPINB2 axis in eosinophilic airway inflammation.","method":"CST1 overexpression and siRNA knockdown in bronchial epithelial cells, RNA-seq transcriptome sequencing, Western blot for p-AKT and SERPINB2, OVA-induced asthma mouse model","journal":"Allergy, asthma & immunology research","confidence":"Medium","confidence_rationale":"Tier 2 — gain/loss-of-function with pathway readout + in vivo model, single lab","pmids":["37075800"],"is_preprint":false},{"year":2018,"finding":"In nasal epithelial cells, stimulation with CST1 combined with IL-4 and dsRNA significantly elevates TSLP mRNA and protein expression; TSLP and IL-33 in turn upregulate CST1 expression, forming a positive feedback loop. CST1 also induces CCL11 and POSTN expression in nasal fibroblasts.","method":"Cytokine stimulation experiments in primary nasal epithelial cells and fibroblasts, RT-qPCR, protein level measurement","journal":"American journal of respiratory cell and molecular biology","confidence":"Medium","confidence_rationale":"Tier 3 — direct stimulation with functional cytokine readouts, single lab, moderate mechanistic depth","pmids":["29698614"],"is_preprint":false},{"year":2024,"finding":"TFAP2C transcriptionally activates CST1 expression in breast cancer cells (validated by dual-luciferase reporter and ChIP assays); CST1 in turn maintains GPX4 levels and suppresses ferroptosis; silencing TFAP2C reduces CST1 and GPX4, promoting ferroptosis, and ectopic CST1 rescues these effects.","method":"Dual-luciferase reporter assay, ChIP, siRNA knockdown, western blot for GPX4/4HNE/ferroptosis markers, xenograft tumor model","journal":"Biochemical genetics","confidence":"Medium","confidence_rationale":"Tier 2 — ChIP + luciferase + rescue experiment + in vivo model, single lab","pmids":["38243003"],"is_preprint":false},{"year":2025,"finding":"TFAP2A transcriptionally activates CST1 in lung adenocarcinoma (validated by ChIP and dual-luciferase reporter assay); CST1 promotes proliferation and migration and suppresses ferroptosis; targeting the TFAP2A/CST1 axis recapitulates ferroptosis phenotypes in vitro and inhibits tumor growth in xenograft models.","method":"ChIP assay, dual-luciferase reporter assay, siRNA/overexpression, ferroptosis markers (Fe2+, ROS, lipid peroxidation, GPX4), xenograft tumor assay","journal":"Journal of biochemical and molecular toxicology","confidence":"Medium","confidence_rationale":"Tier 2 — ChIP + luciferase validation + functional rescue + in vivo model, single lab","pmids":["39692484"],"is_preprint":false},{"year":2018,"finding":"Let-7d inhibits colorectal cancer cell proliferation by targeting CST1, which acts through the p65 (NF-κB) pathway; CST1 siRNA knockdown and let-7d mimics both suppress proliferation, and CST1 is a direct target of let-7d (inferred from luciferase assay context in the study).","method":"siRNA knockdown, let-7d mimic transfection, Western blot, RT-qPCR, functional proliferation assays","journal":"International journal of oncology","confidence":"Low","confidence_rationale":"Tier 3 — pathway placement via KD with proliferation readout, limited mechanistic detail on CST1–p65 direct link","pmids":["29845224"],"is_preprint":false},{"year":2021,"finding":"ENO1 acts upstream of CST1 in thyroid carcinoma; the mTOR/HIF1α pathway regulates ENO1, which in turn regulates CST1 expression; knockdown of CST1 reverses the enhanced tumorigenicity caused by ENO1 overexpression, placing CST1 downstream of ENO1 in this oncogenic axis.","method":"ENO1 knockdown/overexpression, CST1 knockdown rescue experiments, Western blot, proliferation/migration assays","journal":"Frontiers in cell and developmental biology","confidence":"Low","confidence_rationale":"Tier 3 — genetic epistasis by rescue experiment, single lab, limited direct mechanistic detail for CST1 itself","pmids":["33968941"],"is_preprint":false},{"year":2024,"finding":"BRD9 cooperates with the transcription factor FOXP1 (shown by ChIP-qPCR interaction) to regulate CST1 expression in gallbladder cancer; BRD9 inhibitor I-BRD9 suppresses CST1 expression and inhibits the PI3K/AKT pathway, thereby reducing GBC cell proliferation.","method":"siRNA knockdown, CHIP-qPCR, mRNA sequencing, Western blot, BRD9 inhibitor (I-BRD9) treatment, in vivo nude mouse GBC model","journal":"Gene therapy","confidence":"Medium","confidence_rationale":"Tier 2 — ChIP-qPCR + transcriptomic + pharmacological inhibition + in vivo model, single lab","pmids":["39306629"],"is_preprint":false},{"year":2025,"finding":"UBE2D2 stabilizes CST1 through autophagy-dependent degradation suppression; CST1 in turn stabilizes GPX4 to inhibit ferroptosis in gastric cancer; UBE2D2 knockdown reduces CST1 and GPX4, increases ROS accumulation, and induces ferroptosis via the CST1–GPX4 axis.","method":"Proteomic screening, in vitro and in vivo functional experiments (proliferation, invasion, migration, EMT), ROS/ferroptosis assays, Western blot for CST1/GPX4","journal":"International journal of biological macromolecules","confidence":"Medium","confidence_rationale":"Tier 2 — proteomic identification of CST1 as UBE2D2 downstream target + functional rescue, single lab","pmids":["40912429"],"is_preprint":false},{"year":2011,"finding":"CST1 protein accumulates intracellularly within vesicular structures during cellular senescence in human fibroblasts; CST1 expression at mRNA and protein levels is strongly and consistently upregulated upon senescence induction regardless of trigger, consistent with its function as a lysosomal cysteine protease inhibitor.","method":"Immunoblotting, immunofluorescence cytochemistry, comparison across multiple senescence-inducing conditions and cell types","journal":"The journals of gerontology. Series A, Biological sciences and medical sciences","confidence":"Medium","confidence_rationale":"Tier 2 — direct subcellular localization by immunofluorescence + multiple conditions, single lab","pmids":["21636832"],"is_preprint":false},{"year":2026,"finding":"NFATC2 transcriptionally activates CST1 in cholangiocarcinoma; CST1 suppresses cellular senescence by stabilizing SOX4 protein (ectopic SOX4 rescues senescence induced by CST1 depletion); this axis promotes CCA tumor growth and metastasis.","method":"Gain- and loss-of-function experiments, senescence-associated β-galactosidase activity, SASP factor measurement, multi-omics profiling, ectopic SOX4 rescue experiments, murine orthotopic liver implantation and pulmonary metastasis models","journal":"Cell death discovery","confidence":"Medium","confidence_rationale":"Tier 2 — genetic epistasis rescue + multi-omics + in vivo models, single lab","pmids":["41881961"],"is_preprint":false},{"year":2024,"finding":"ENO1 silencing in breast cancer cells promotes autophagy-dependent ferroptosis and inhibits glycolysis; CST1 overexpression reverses these effects of ENO1 silencing, placing CST1 downstream of ENO1 in regulating the mTOR-autophagy-ferroptosis axis.","method":"siRNA knockdown, overexpression rescue, ECAR/glycolysis assays, iron/lipid peroxidation assays, autophagy markers, mTOR pathway inhibitor/activator experiments, in vivo xenograft","journal":"Drug development research","confidence":"Low","confidence_rationale":"Tier 3 — rescue epistasis experiment for CST1, mechanism primarily attributed to ENO1/mTOR axis","pmids":["39503165"],"is_preprint":false},{"year":2002,"finding":"CST1 mRNA expression in adult human tissues is tissue-specific, with predominant expression in submandibular and parotid glands, lacrimal gland, gallbladder epithelium, seminal vesicle, and tracheal glands; immunohistochemistry localized expression to serous-type secretory end pieces, consistent with a secretory/protective function of CST1 as a cysteine protease inhibitor.","method":"RNase protection assay (gene-specific), immunohistochemistry across 23 adult human tissues","journal":"DNA and cell biology","confidence":"Medium","confidence_rationale":"Tier 2 — direct localization by IHC + quantitative tissue expression profiling with multiple tissues","pmids":["11879580"],"is_preprint":false}],"current_model":"CST1 (Cystatin SN) is a secreted cysteine protease inhibitor that, in cancer contexts, promotes tumor progression by: (1) interacting with GPX4 and recruiting the deubiquitinase OTUB1 to stabilize GPX4 and thereby suppress ferroptosis; (2) activating Wnt/β-catenin, PI3K/AKT, and MEK/ERK/CREB signaling pathways to drive migration and invasion; (3) being transcriptionally activated by upstream regulators including TFAP2C, TFAP2A, NFATC2, and BRD9/FOXP1 complexes; and (4) acting downstream of ENO1 to modulate glycolysis and autophagy-dependent ferroptosis, while in inflammatory airway disease it amplifies eosinophilic inflammation via AKT-SERPINB2 signaling and cytokine crosstalk with epithelial cells."},"narrative":{"teleology":[{"year":2002,"claim":"Mapping CST1 tissue expression established that it is a gland-restricted secretory cysteine protease inhibitor rather than a ubiquitously expressed housekeeping gene, framing its later study in mucosal biology and cancer.","evidence":"RNase protection assay and immunohistochemistry across 23 human tissues","pmids":["11879580"],"confidence":"Medium","gaps":["No direct demonstration of protease-inhibitory activity in the studied tissues","Protein-level quantitation limited to IHC"]},{"year":2011,"claim":"Discovery that CST1 accumulates in intracellular vesicular compartments during senescence broadened its biology beyond secretory protease inhibition and suggested an intracellular role linked to lysosomal proteostasis.","evidence":"Immunofluorescence and immunoblotting in human fibroblasts across multiple senescence triggers","pmids":["21636832"],"confidence":"Medium","gaps":["No functional test of whether vesicular CST1 is required for senescence phenotype","Identity of the vesicular compartment not resolved (lysosomal vs. other)"]},{"year":2018,"claim":"Identification of a CST1–TSLP/IL-33 positive feedback loop in nasal epithelial cells and fibroblasts revealed how CST1 amplifies type-2 airway inflammation, establishing its first defined signaling circuit outside of cancer.","evidence":"Cytokine stimulation of primary nasal epithelial cells and fibroblasts with RT-qPCR and protein measurement","pmids":["29845224"],"confidence":"Medium","gaps":["Mechanism by which CST1 induces TSLP not defined at the receptor level","In vivo airway model not included in this study"]},{"year":2021,"claim":"Demonstration that CST1 activates Wnt/β-catenin signaling to promote gastric cancer migration and invasion provided the first defined oncogenic pathway downstream of CST1, while ENO1 was placed upstream of CST1 in thyroid carcinoma, indicating CST1 acts as a pathway relay node.","evidence":"Luciferase reporter, β-catenin nuclear translocation, Wnt inhibitor rescue (gastric cancer); ENO1 OE/KD with CST1 rescue epistasis (thyroid carcinoma)","pmids":["33658852","33968941"],"confidence":"Medium","gaps":["No direct binding partner identified for Wnt pathway activation by CST1","ENO1–CST1 regulatory mechanism not resolved at the promoter/protein level","Single-lab findings in each cancer type"]},{"year":2022,"claim":"The critical mechanistic advance was the identification of a CST1–GPX4–OTUB1 ternary complex: CST1 physically binds GPX4 and recruits OTUB1 to deubiquitinate and stabilize GPX4, directly suppressing ferroptosis and promoting metastasis — the first molecular mechanism explaining how a secreted cystatin inhibits cell death.","evidence":"Reciprocal co-IP with mass spectrometry, ubiquitination assays, overexpression/knockdown in vitro and nude mouse metastasis model","pmids":["36369321"],"confidence":"High","gaps":["Structural basis of CST1–GPX4 interaction unknown","Whether catalytic cystatin activity is required for GPX4 stabilization not tested"]},{"year":2022,"claim":"CST1 was shown to activate PI3K/AKT in airway smooth muscle cells to promote proliferation/migration, paralleling its oncogenic PI3K/AKT activation and extending its signaling repertoire to non-malignant disease.","evidence":"siRNA/overexpression with PI3K/AKT inhibitor rescue, EdU/CCK-8, Transwell assays in PDGF-BB-treated ASMCs","pmids":["36354198"],"confidence":"Medium","gaps":["Receptor or cell-surface partner through which secreted CST1 engages PI3K/AKT not identified"]},{"year":2023,"claim":"Identification of MEK/ERK/CREB activation downstream of CST1 in ESCC and AKT–SERPINB2 axis in asthmatic epithelial cells expanded the signaling portfolio of CST1 and placed it upstream of multiple MAPK and AKT effector branches.","evidence":"Western blot for phospho-MEK/ERK/CREB, dual-luciferase for miR-942-5p targeting CST1 (ESCC); CST1 OE/KD with RNA-seq and p-AKT/SERPINB2 readout plus OVA mouse asthma model (airway)","pmids":["36848349","37075800"],"confidence":"Medium","gaps":["Direct target/receptor mediating CST1-to-MAPK signal remains unknown","SERPINB2 functional role downstream of AKT not fully dissected"]},{"year":2024,"claim":"Upstream transcriptional control of CST1 was resolved: TFAP2C in breast cancer and BRD9/FOXP1 in gallbladder cancer each directly activate the CST1 promoter, linking epigenetic/transcription factor programs to the CST1–GPX4 ferroptosis suppression axis.","evidence":"ChIP + dual-luciferase reporter + siRNA rescue + xenograft models (TFAP2C); ChIP-qPCR + I-BRD9 inhibitor + mRNA-seq + in vivo GBC model (BRD9/FOXP1)","pmids":["38243003","39306629"],"confidence":"Medium","gaps":["Whether TFAP2C and BRD9/FOXP1 act in the same or distinct cancer contexts is unexplored","Promoter elements mediating each TF's binding not mapped at nucleotide resolution"]},{"year":2025,"claim":"TFAP2A was validated as a second AP-2 family transcriptional activator of CST1 (in lung adenocarcinoma), and UBE2D2 was found to stabilize CST1 protein via suppression of autophagy-dependent degradation, together revealing both transcriptional and post-translational regulatory layers controlling CST1 abundance.","evidence":"ChIP + dual-luciferase + ferroptosis markers + xenograft (TFAP2A); proteomic screening + autophagy/ferroptosis assays + in vivo functional studies (UBE2D2)","pmids":["39692484","40912429"],"confidence":"Medium","gaps":["Autophagy receptor or E3 ligase targeting CST1 for autophagic degradation not identified","Whether UBE2D2 ubiquitinates CST1 directly or acts indirectly is unresolved"]},{"year":2026,"claim":"NFATC2 was identified as a transcriptional activator of CST1 in cholangiocarcinoma, and CST1 was shown to suppress cellular senescence by stabilizing SOX4, revealing a new anti-senescence function beyond ferroptosis suppression.","evidence":"Multi-omics profiling, gain/loss-of-function, SOX4 rescue, SA-β-gal assay, orthotopic liver and pulmonary metastasis models","pmids":["41881961"],"confidence":"Medium","gaps":["Mechanism by which CST1 stabilizes SOX4 protein not defined (direct binding vs. intermediate)","Relationship between senescence suppression and ferroptosis suppression by CST1 not tested"]},{"year":null,"claim":"The cell-surface receptor or binding partner through which extracellular CST1 activates intracellular signaling cascades (Wnt, PI3K/AKT, MEK/ERK) remains unidentified, and whether its classical cysteine protease inhibitory activity is required for any of its oncogenic or anti-ferroptotic functions has not been tested.","evidence":"","pmids":[],"confidence":"High","gaps":["No receptor identified for CST1 signal transduction","Protease-inhibitory activity not dissected from signaling functions using catalytic-dead mutants","Structural basis of CST1–GPX4 interaction unknown"]}],"mechanism_profile":{"molecular_activity":[{"term_id":"GO:0098772","term_label":"molecular function regulator activity","supporting_discovery_ids":[0,6,7,11]},{"term_id":"GO:0140096","term_label":"catalytic activity, acting on a protein","supporting_discovery_ids":[0,12]}],"localization":[{"term_id":"GO:0005576","term_label":"extracellular region","supporting_discovery_ids":[15]},{"term_id":"GO:0031410","term_label":"cytoplasmic vesicle","supporting_discovery_ids":[12]}],"pathway":[{"term_id":"R-HSA-5357801","term_label":"Programmed Cell Death","supporting_discovery_ids":[0,6,7,11,14]},{"term_id":"R-HSA-162582","term_label":"Signal Transduction","supporting_discovery_ids":[1,2,3,4,10]},{"term_id":"R-HSA-1643685","term_label":"Disease","supporting_discovery_ids":[0,1,2,6,7,10,13]},{"term_id":"R-HSA-168256","term_label":"Immune System","supporting_discovery_ids":[4,5]}],"complexes":[],"partners":["GPX4","OTUB1","TFAP2C","TFAP2A","NFATC2","BRD9","SOX4","UBE2D2"],"other_free_text":[]},"mechanistic_narrative":"CST1 (Cystatin SN) is a secreted cysteine protease inhibitor predominantly expressed in exocrine glands and mucosal epithelia that has been co-opted in multiple cancer types to suppress ferroptosis and promote tumor progression [PMID:11879580, PMID:36369321]. CST1 physically interacts with GPX4 and recruits the deubiquitinase OTUB1 to stabilize GPX4 protein, thereby reducing lipid peroxidation and blocking ferroptotic cell death; this axis is transcriptionally activated by TFAP2C, TFAP2A, NFATC2, and BRD9/FOXP1 complexes in a tissue-dependent manner [PMID:36369321, PMID:38243003, PMID:39692484, PMID:41881961, PMID:39306629]. Beyond ferroptosis suppression, CST1 activates Wnt/β-catenin, PI3K/AKT, and MEK/ERK/CREB signaling to drive migration and invasion, and in airway epithelial cells it participates in a TSLP/IL-33 positive feedback loop that amplifies eosinophilic inflammation via AKT–SERPINB2 signaling [PMID:33658852, PMID:36354198, PMID:36848349, PMID:29845224, PMID:37075800]. CST1 also accumulates in intracellular vesicular structures during cellular senescence and suppresses senescence in cholangiocarcinoma by stabilizing SOX4 [PMID:21636832, PMID:41881961]."},"prefetch_data":{"uniprot":{"accession":"P01037","full_name":"Cystatin-SN","aliases":["Cystain-SA-I","Cystatin-1","Salivary cystatin-SA-1"],"length_aa":141,"mass_kda":16.4,"function":"Human saliva appears to contain several cysteine proteinase inhibitors that are immunologically related to cystatin S but that differ in their specificity due to amino acid sequence differences. Cystatin SN, with a pI of 7.5, is a much better inhibitor of papain and dipeptidyl peptidase I than is cystatin S, although both inhibit ficin equally well","subcellular_location":"Secreted","url":"https://www.uniprot.org/uniprotkb/P01037/entry"},"depmap":{"release":"DepMap","has_data":true,"is_common_essential":false,"resolved_as":"","url":"https://depmap.org/portal/gene/CST1","classification":"Not Classified","n_dependent_lines":0,"n_total_lines":1208,"dependency_fraction":0.0},"opencell":{"profiled":false,"resolved_as":"","ensg_id":"","cell_line_id":"","localizations":[],"interactors":[{"gene":"DNAJC8","stoichiometry":0.2}],"url":"https://opencell.sf.czbiohub.org/search/CST1","total_profiled":1310},"omim":[{"mim_id":"604965","title":"SERINE/THREONINE PROTEIN KINASE 4; STK4","url":"https://www.omim.org/entry/604965"},{"mim_id":"604312","title":"CYSTATIN 3; CST3","url":"https://www.omim.org/entry/604312"},{"mim_id":"602681","title":"FORKHEAD BOX O3; FOXO3","url":"https://www.omim.org/entry/602681"},{"mim_id":"123858","title":"CYSTATIN 5; CST5","url":"https://www.omim.org/entry/123858"},{"mim_id":"123857","title":"CYSTATIN 4; CST4","url":"https://www.omim.org/entry/123857"}],"hpa":{"profiled":true,"resolved_as":"","reliability":"","locations":[],"tissue_specificity":"Tissue enriched","tissue_distribution":"Detected in some","driving_tissues":[{"tissue":"salivary gland","ntpm":26681.9}],"url":"https://www.proteinatlas.org/search/CST1"},"hgnc":{"alias_symbol":[],"prev_symbol":[]},"alphafold":{"accession":"P01037","domains":[{"cath_id":"3.10.450.10","chopping":"33-141","consensus_level":"high","plddt":96.1864,"start":33,"end":141}],"viewer_url":"https://alphafold.ebi.ac.uk/entry/P01037","model_url":"https://alphafold.ebi.ac.uk/files/AF-P01037-F1-model_v6.cif","pae_url":"https://alphafold.ebi.ac.uk/files/AF-P01037-F1-predicted_aligned_error_v6.png","plddt_mean":92.56},"mouse_models":{"mgi_url":"https://www.informatics.jax.org/marker/summary?nomen=CST1","jax_strain_url":"https://www.jax.org/strain/search?query=CST1"},"sequence":{"accession":"P01037","fasta_url":"https://rest.uniprot.org/uniprotkb/P01037.fasta","uniprot_url":"https://www.uniprot.org/uniprotkb/P01037/entry","alphafold_viewer_url":"https://alphafold.ebi.ac.uk/entry/P01037"}},"corpus_meta":[{"pmid":"36369321","id":"PMC_36369321","title":"CST1 inhibits ferroptosis and promotes gastric cancer metastasis by regulating GPX4 protein stability via OTUB1.","date":"2022","source":"Oncogene","url":"https://pubmed.ncbi.nlm.nih.gov/36369321","citation_count":256,"is_preprint":false},{"pmid":"24385904","id":"PMC_24385904","title":"The Toxoplasma gondii cyst wall protein CST1 is critical for cyst wall integrity and promotes bradyzoite persistence.","date":"2013","source":"PLoS pathogens","url":"https://pubmed.ncbi.nlm.nih.gov/24385904","citation_count":134,"is_preprint":false},{"pmid":"12744460","id":"PMC_12744460","title":"The Colletotrichum lagenariu Ste12-like gene CST1 is essential for appressorium penetration.","date":"2003","source":"Molecular plant-microbe interactions : MPMI","url":"https://pubmed.ncbi.nlm.nih.gov/12744460","citation_count":74,"is_preprint":false},{"pmid":"29698614","id":"PMC_29698614","title":"Expression and Functional Analysis of CST1 in Intractable Nasal Polyps.","date":"2018","source":"American journal of respiratory cell and molecular biology","url":"https://pubmed.ncbi.nlm.nih.gov/29698614","citation_count":64,"is_preprint":false},{"pmid":"11879580","id":"PMC_11879580","title":"Expression of type 2 cystatin genes CST1-CST5 in adult human tissues and the developing submandibular gland.","date":"2002","source":"DNA and cell biology","url":"https://pubmed.ncbi.nlm.nih.gov/11879580","citation_count":42,"is_preprint":false},{"pmid":"29845224","id":"PMC_29845224","title":"Let‑7d inhibits colorectal cancer cell proliferation through the CST1/p65 pathway.","date":"2018","source":"International journal of oncology","url":"https://pubmed.ncbi.nlm.nih.gov/29845224","citation_count":36,"is_preprint":false},{"pmid":"36733482","id":"PMC_36733482","title":"Transcriptomic analysis of asthma and allergic rhinitis reveals CST1 as a biomarker of unified airways.","date":"2023","source":"Frontiers in immunology","url":"https://pubmed.ncbi.nlm.nih.gov/36733482","citation_count":32,"is_preprint":false},{"pmid":"33658852","id":"PMC_33658852","title":"CST1 Promoted Gastric Cancer Migration and Invasion Through Activating Wnt Pathway.","date":"2021","source":"Cancer management and research","url":"https://pubmed.ncbi.nlm.nih.gov/33658852","citation_count":28,"is_preprint":false},{"pmid":"34764696","id":"PMC_34764696","title":"Development and Evaluation of Serum CST1 Detection for Early Diagnosis of Esophageal Squamous Cell Carcinoma.","date":"2021","source":"Cancer management and research","url":"https://pubmed.ncbi.nlm.nih.gov/34764696","citation_count":13,"is_preprint":false},{"pmid":"33968941","id":"PMC_33968941","title":"α-Enolase Lies Downstream of mTOR/HIF1α and Promotes Thyroid Carcinoma Progression by Regulating CST1.","date":"2021","source":"Frontiers in cell and developmental biology","url":"https://pubmed.ncbi.nlm.nih.gov/33968941","citation_count":13,"is_preprint":false},{"pmid":"36848349","id":"PMC_36848349","title":"MiR-942-5p inhibits tumor migration and invasion through targeting CST1 in esophageal squamous cell carcinoma.","date":"2023","source":"PloS one","url":"https://pubmed.ncbi.nlm.nih.gov/36848349","citation_count":12,"is_preprint":false},{"pmid":"38243003","id":"PMC_38243003","title":"TFAP2C Activates CST1 Transcription to Facilitate Breast Cancer Progression and Suppress Ferroptosis.","date":"2024","source":"Biochemical genetics","url":"https://pubmed.ncbi.nlm.nih.gov/38243003","citation_count":12,"is_preprint":false},{"pmid":"35653624","id":"PMC_35653624","title":"Identification and Validation of Serum CST1 as a Diagnostic Marker for Differentiating Early-Stage Non-Small Cell Lung Cancer from Pulmonary Benign Nodules.","date":"2022","source":"Cancer control : journal of the Moffitt Cancer 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CST1.","date":"2020","source":"mSphere","url":"https://pubmed.ncbi.nlm.nih.gov/32132158","citation_count":8,"is_preprint":false},{"pmid":"38077439","id":"PMC_38077439","title":"Combination of serum CST1 and HE4 for early diagnosis of endometrial cancer.","date":"2023","source":"PeerJ","url":"https://pubmed.ncbi.nlm.nih.gov/38077439","citation_count":7,"is_preprint":false},{"pmid":"38270326","id":"PMC_38270326","title":"The functional role of CST1 and CCL26 in asthma development.","date":"2024","source":"Immunity, inflammation and disease","url":"https://pubmed.ncbi.nlm.nih.gov/38270326","citation_count":6,"is_preprint":false},{"pmid":"39692484","id":"PMC_39692484","title":"TFAP2A Promotes Cell Progression and Suppresses Ferroptosis in Lung Adenocarcinoma via Activating Transcription of CST1.","date":"2025","source":"Journal of biochemical and molecular 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England)","url":"https://pubmed.ncbi.nlm.nih.gov/39320318","citation_count":4,"is_preprint":false},{"pmid":"37762137","id":"PMC_37762137","title":"Cystatin SN (CST1) as a Novel Salivary Biomarker of Periodontitis.","date":"2023","source":"International journal of molecular sciences","url":"https://pubmed.ncbi.nlm.nih.gov/37762137","citation_count":4,"is_preprint":false},{"pmid":"27550161","id":"PMC_27550161","title":"Transgenic Expression of a Viral Cystatin Gene CpBV-CST1 in Tobacco Confers Insect Resistance.","date":"2016","source":"Environmental entomology","url":"https://pubmed.ncbi.nlm.nih.gov/27550161","citation_count":4,"is_preprint":false},{"pmid":"39721372","id":"PMC_39721372","title":"CST1 promoted gastric cancer development by activating the AKT pathway.","date":"2024","source":"Clinics (Sao Paulo, Brazil)","url":"https://pubmed.ncbi.nlm.nih.gov/39721372","citation_count":2,"is_preprint":false},{"pmid":"40912429","id":"PMC_40912429","title":"UBE2D2 promotes gastric cancer progression by inhibiting ferroptosis through autophagy-dependent stabilization of CST1.","date":"2025","source":"International journal of biological macromolecules","url":"https://pubmed.ncbi.nlm.nih.gov/40912429","citation_count":1,"is_preprint":false},{"pmid":"38814219","id":"PMC_38814219","title":"Clinical implications and mechanism of CST1 in gastric carcinoma based on database screening.","date":"2024","source":"Cellular and molecular biology (Noisy-le-Grand, France)","url":"https://pubmed.ncbi.nlm.nih.gov/38814219","citation_count":1,"is_preprint":false},{"pmid":"40274175","id":"PMC_40274175","title":"Platelet-rich plasma inhibits ferroptosis and inflammation to alleviate frozen shoulder via activating the CST1/GPX4 signaling pathway.","date":"2025","source":"Archives of biochemistry and biophysics","url":"https://pubmed.ncbi.nlm.nih.gov/40274175","citation_count":1,"is_preprint":false},{"pmid":"39707108","id":"PMC_39707108","title":"Investigation of antimicrobial susceptibility and resistance gene prevalence in Capnocytophaga spp. isolated from dogs and cats and characterization of novel class A β-lactamase CST-1.","date":"2024","source":"European journal of clinical microbiology & infectious diseases : official publication of the European Society of Clinical Microbiology","url":"https://pubmed.ncbi.nlm.nih.gov/39707108","citation_count":1,"is_preprint":false},{"pmid":"41881961","id":"PMC_41881961","title":"NFATC2-mediated CST1 upregulation drives cholangiocarcinoma growth and metastasis.","date":"2026","source":"Cell death discovery","url":"https://pubmed.ncbi.nlm.nih.gov/41881961","citation_count":0,"is_preprint":false},{"pmid":"40999193","id":"PMC_40999193","title":"Genomic insights of Pseudomonas nicosulfuronedens CST1-L11, a plant growth-promoting and stress-tolerating endophyte associated with native Musa spp. in Vietnam.","date":"2025","source":"World journal of microbiology & biotechnology","url":"https://pubmed.ncbi.nlm.nih.gov/40999193","citation_count":0,"is_preprint":false},{"pmid":"41670526","id":"PMC_41670526","title":"Expression and Clinical Significance of FOXQ1, MMP11, and CST1 in Colorectal Cancer.","date":"2026","source":"Clinical laboratory","url":"https://pubmed.ncbi.nlm.nih.gov/41670526","citation_count":0,"is_preprint":false}],"cost":{"stage1":{"model":"claude-sonnet-4-6","input_tokens":20750,"output_tokens":3955,"usd":0.060788},"stage2":{"model":"claude-opus-4-6","input_tokens":7402,"output_tokens":3053,"usd":0.170003},"total_usd":0.230791,"stage1_batch_id":"msgbatch_011kfMfE81o6CTcyhVUqumuW","stage2_batch_id":"msgbatch_01DTputS6WNkH745x1nVSoBo","note":"batch pricing = 50% of standard"},"stage1_raw":"```json\n{\n  \"discoveries\": [\n    {\n      \"year\": 2022,\n      \"finding\": \"CST1 interacts with GPX4 (a key ferroptosis regulator) and recruits the deubiquitinase OTUB1 to relieve GPX4 ubiquitination, thereby stabilizing GPX4 protein, reducing intracellular ROS, and inhibiting ferroptosis to promote gastric cancer metastasis.\",\n      \"method\": \"Co-immunoprecipitation combined with mass spectrometry, ubiquitination assay, overexpression/knockdown functional studies in vitro and in vivo (nude mouse metastasis model)\",\n      \"journal\": \"Oncogene\",\n      \"confidence\": \"High\",\n      \"confidence_rationale\": \"Tier 2 — reciprocal Co-IP + MS identification of complex + functional rescue experiments with multiple orthogonal readouts\",\n      \"pmids\": [\"36369321\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"CST1 promotes gastric cancer cell migration and invasion by activating the Wnt signaling pathway, promoting nuclear translocation of β-catenin and upregulating Wnt target genes; inhibition of Wnt pathway in CST1-overexpressing cells suppresses these effects.\",\n      \"method\": \"Transwell migration/invasion assay, luciferase reporter assay for Wnt pathway activity, β-catenin nuclear translocation assessment, pharmacological Wnt inhibition rescue experiment\",\n      \"journal\": \"Cancer management and research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — epistasis via pathway inhibitor rescue + reporter assay, single lab\",\n      \"pmids\": [\"33658852\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"CST1 promotes migration and invasion of esophageal squamous cell carcinoma (ESCC) cells by upregulating phosphorylation of MEK1/2, ERK1/2, and CREB in the MEK/ERK/CREB signaling pathway; miR-942-5p directly targets CST1 (validated by dual-luciferase assay) to downregulate this pathway.\",\n      \"method\": \"Transwell migration/invasion assay, Western blot for pathway effector phosphorylation, dual-luciferase reporter assay\",\n      \"journal\": \"PloS one\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2/3 — functional KD with defined pathway readout + luciferase validation, single lab\",\n      \"pmids\": [\"36848349\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2022,\n      \"finding\": \"CST1 promotes proliferation and migration of airway smooth muscle cells (ASMCs) treated with PDGF-BB by activating the PI3K/AKT signaling pathway and upregulating MMP1 and MMP9; knockdown or overexpression of CST1 reciprocally modulates PI3K/AKT activity and cell behavior.\",\n      \"method\": \"siRNA knockdown, plasmid overexpression, EdU/CCK-8 proliferation assays, Transwell migration assay, Western blot for PI3K/AKT pathway components, PI3K/AKT inhibitor rescue experiment\",\n      \"journal\": \"The Kaohsiung journal of medical sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — pathway inhibitor rescue + multiple functional readouts, single lab\",\n      \"pmids\": [\"36354198\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2023,\n      \"finding\": \"CST1 overexpression in bronchial epithelial cells enhances AKT phosphorylation and upregulates SERPINB2 expression; anti-CST1 siRNA reverses these effects; AKT positively regulates SERPINB2, placing CST1 upstream of the AKT–SERPINB2 axis in eosinophilic airway inflammation.\",\n      \"method\": \"CST1 overexpression and siRNA knockdown in bronchial epithelial cells, RNA-seq transcriptome sequencing, Western blot for p-AKT and SERPINB2, OVA-induced asthma mouse model\",\n      \"journal\": \"Allergy, asthma & immunology research\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — gain/loss-of-function with pathway readout + in vivo model, single lab\",\n      \"pmids\": [\"37075800\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"In nasal epithelial cells, stimulation with CST1 combined with IL-4 and dsRNA significantly elevates TSLP mRNA and protein expression; TSLP and IL-33 in turn upregulate CST1 expression, forming a positive feedback loop. CST1 also induces CCL11 and POSTN expression in nasal fibroblasts.\",\n      \"method\": \"Cytokine stimulation experiments in primary nasal epithelial cells and fibroblasts, RT-qPCR, protein level measurement\",\n      \"journal\": \"American journal of respiratory cell and molecular biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 3 — direct stimulation with functional cytokine readouts, single lab, moderate mechanistic depth\",\n      \"pmids\": [\"29698614\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"TFAP2C transcriptionally activates CST1 expression in breast cancer cells (validated by dual-luciferase reporter and ChIP assays); CST1 in turn maintains GPX4 levels and suppresses ferroptosis; silencing TFAP2C reduces CST1 and GPX4, promoting ferroptosis, and ectopic CST1 rescues these effects.\",\n      \"method\": \"Dual-luciferase reporter assay, ChIP, siRNA knockdown, western blot for GPX4/4HNE/ferroptosis markers, xenograft tumor model\",\n      \"journal\": \"Biochemical genetics\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — ChIP + luciferase + rescue experiment + in vivo model, single lab\",\n      \"pmids\": [\"38243003\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"TFAP2A transcriptionally activates CST1 in lung adenocarcinoma (validated by ChIP and dual-luciferase reporter assay); CST1 promotes proliferation and migration and suppresses ferroptosis; targeting the TFAP2A/CST1 axis recapitulates ferroptosis phenotypes in vitro and inhibits tumor growth in xenograft models.\",\n      \"method\": \"ChIP assay, dual-luciferase reporter assay, siRNA/overexpression, ferroptosis markers (Fe2+, ROS, lipid peroxidation, GPX4), xenograft tumor assay\",\n      \"journal\": \"Journal of biochemical and molecular toxicology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — ChIP + luciferase validation + functional rescue + in vivo model, single lab\",\n      \"pmids\": [\"39692484\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2018,\n      \"finding\": \"Let-7d inhibits colorectal cancer cell proliferation by targeting CST1, which acts through the p65 (NF-κB) pathway; CST1 siRNA knockdown and let-7d mimics both suppress proliferation, and CST1 is a direct target of let-7d (inferred from luciferase assay context in the study).\",\n      \"method\": \"siRNA knockdown, let-7d mimic transfection, Western blot, RT-qPCR, functional proliferation assays\",\n      \"journal\": \"International journal of oncology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 — pathway placement via KD with proliferation readout, limited mechanistic detail on CST1–p65 direct link\",\n      \"pmids\": [\"29845224\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2021,\n      \"finding\": \"ENO1 acts upstream of CST1 in thyroid carcinoma; the mTOR/HIF1α pathway regulates ENO1, which in turn regulates CST1 expression; knockdown of CST1 reverses the enhanced tumorigenicity caused by ENO1 overexpression, placing CST1 downstream of ENO1 in this oncogenic axis.\",\n      \"method\": \"ENO1 knockdown/overexpression, CST1 knockdown rescue experiments, Western blot, proliferation/migration assays\",\n      \"journal\": \"Frontiers in cell and developmental biology\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 — genetic epistasis by rescue experiment, single lab, limited direct mechanistic detail for CST1 itself\",\n      \"pmids\": [\"33968941\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"BRD9 cooperates with the transcription factor FOXP1 (shown by ChIP-qPCR interaction) to regulate CST1 expression in gallbladder cancer; BRD9 inhibitor I-BRD9 suppresses CST1 expression and inhibits the PI3K/AKT pathway, thereby reducing GBC cell proliferation.\",\n      \"method\": \"siRNA knockdown, CHIP-qPCR, mRNA sequencing, Western blot, BRD9 inhibitor (I-BRD9) treatment, in vivo nude mouse GBC model\",\n      \"journal\": \"Gene therapy\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — ChIP-qPCR + transcriptomic + pharmacological inhibition + in vivo model, single lab\",\n      \"pmids\": [\"39306629\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2025,\n      \"finding\": \"UBE2D2 stabilizes CST1 through autophagy-dependent degradation suppression; CST1 in turn stabilizes GPX4 to inhibit ferroptosis in gastric cancer; UBE2D2 knockdown reduces CST1 and GPX4, increases ROS accumulation, and induces ferroptosis via the CST1–GPX4 axis.\",\n      \"method\": \"Proteomic screening, in vitro and in vivo functional experiments (proliferation, invasion, migration, EMT), ROS/ferroptosis assays, Western blot for CST1/GPX4\",\n      \"journal\": \"International journal of biological macromolecules\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — proteomic identification of CST1 as UBE2D2 downstream target + functional rescue, single lab\",\n      \"pmids\": [\"40912429\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2011,\n      \"finding\": \"CST1 protein accumulates intracellularly within vesicular structures during cellular senescence in human fibroblasts; CST1 expression at mRNA and protein levels is strongly and consistently upregulated upon senescence induction regardless of trigger, consistent with its function as a lysosomal cysteine protease inhibitor.\",\n      \"method\": \"Immunoblotting, immunofluorescence cytochemistry, comparison across multiple senescence-inducing conditions and cell types\",\n      \"journal\": \"The journals of gerontology. Series A, Biological sciences and medical sciences\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — direct subcellular localization by immunofluorescence + multiple conditions, single lab\",\n      \"pmids\": [\"21636832\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2026,\n      \"finding\": \"NFATC2 transcriptionally activates CST1 in cholangiocarcinoma; CST1 suppresses cellular senescence by stabilizing SOX4 protein (ectopic SOX4 rescues senescence induced by CST1 depletion); this axis promotes CCA tumor growth and metastasis.\",\n      \"method\": \"Gain- and loss-of-function experiments, senescence-associated β-galactosidase activity, SASP factor measurement, multi-omics profiling, ectopic SOX4 rescue experiments, murine orthotopic liver implantation and pulmonary metastasis models\",\n      \"journal\": \"Cell death discovery\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — genetic epistasis rescue + multi-omics + in vivo models, single lab\",\n      \"pmids\": [\"41881961\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2024,\n      \"finding\": \"ENO1 silencing in breast cancer cells promotes autophagy-dependent ferroptosis and inhibits glycolysis; CST1 overexpression reverses these effects of ENO1 silencing, placing CST1 downstream of ENO1 in regulating the mTOR-autophagy-ferroptosis axis.\",\n      \"method\": \"siRNA knockdown, overexpression rescue, ECAR/glycolysis assays, iron/lipid peroxidation assays, autophagy markers, mTOR pathway inhibitor/activator experiments, in vivo xenograft\",\n      \"journal\": \"Drug development research\",\n      \"confidence\": \"Low\",\n      \"confidence_rationale\": \"Tier 3 — rescue epistasis experiment for CST1, mechanism primarily attributed to ENO1/mTOR axis\",\n      \"pmids\": [\"39503165\"],\n      \"is_preprint\": false\n    },\n    {\n      \"year\": 2002,\n      \"finding\": \"CST1 mRNA expression in adult human tissues is tissue-specific, with predominant expression in submandibular and parotid glands, lacrimal gland, gallbladder epithelium, seminal vesicle, and tracheal glands; immunohistochemistry localized expression to serous-type secretory end pieces, consistent with a secretory/protective function of CST1 as a cysteine protease inhibitor.\",\n      \"method\": \"RNase protection assay (gene-specific), immunohistochemistry across 23 adult human tissues\",\n      \"journal\": \"DNA and cell biology\",\n      \"confidence\": \"Medium\",\n      \"confidence_rationale\": \"Tier 2 — direct localization by IHC + quantitative tissue expression profiling with multiple tissues\",\n      \"pmids\": [\"11879580\"],\n      \"is_preprint\": false\n    }\n  ],\n  \"current_model\": \"CST1 (Cystatin SN) is a secreted cysteine protease inhibitor that, in cancer contexts, promotes tumor progression by: (1) interacting with GPX4 and recruiting the deubiquitinase OTUB1 to stabilize GPX4 and thereby suppress ferroptosis; (2) activating Wnt/β-catenin, PI3K/AKT, and MEK/ERK/CREB signaling pathways to drive migration and invasion; (3) being transcriptionally activated by upstream regulators including TFAP2C, TFAP2A, NFATC2, and BRD9/FOXP1 complexes; and (4) acting downstream of ENO1 to modulate glycolysis and autophagy-dependent ferroptosis, while in inflammatory airway disease it amplifies eosinophilic inflammation via AKT-SERPINB2 signaling and cytokine crosstalk with epithelial cells.\"\n}\n```","stage2_raw":"```json\n{\n  \"mechanistic_narrative\": \"CST1 (Cystatin SN) is a secreted cysteine protease inhibitor predominantly expressed in exocrine glands and mucosal epithelia that has been co-opted in multiple cancer types to suppress ferroptosis and promote tumor progression [PMID:11879580, PMID:36369321]. CST1 physically interacts with GPX4 and recruits the deubiquitinase OTUB1 to stabilize GPX4 protein, thereby reducing lipid peroxidation and blocking ferroptotic cell death; this axis is transcriptionally activated by TFAP2C, TFAP2A, NFATC2, and BRD9/FOXP1 complexes in a tissue-dependent manner [PMID:36369321, PMID:38243003, PMID:39692484, PMID:41881961, PMID:39306629]. Beyond ferroptosis suppression, CST1 activates Wnt/β-catenin, PI3K/AKT, and MEK/ERK/CREB signaling to drive migration and invasion, and in airway epithelial cells it participates in a TSLP/IL-33 positive feedback loop that amplifies eosinophilic inflammation via AKT–SERPINB2 signaling [PMID:33658852, PMID:36354198, PMID:36848349, PMID:29845224, PMID:37075800]. CST1 also accumulates in intracellular vesicular structures during cellular senescence and suppresses senescence in cholangiocarcinoma by stabilizing SOX4 [PMID:21636832, PMID:41881961].\",\n  \"teleology\": [\n    {\n      \"year\": 2002,\n      \"claim\": \"Mapping CST1 tissue expression established that it is a gland-restricted secretory cysteine protease inhibitor rather than a ubiquitously expressed housekeeping gene, framing its later study in mucosal biology and cancer.\",\n      \"evidence\": \"RNase protection assay and immunohistochemistry across 23 human tissues\",\n      \"pmids\": [\"11879580\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No direct demonstration of protease-inhibitory activity in the studied tissues\", \"Protein-level quantitation limited to IHC\"]\n    },\n    {\n      \"year\": 2011,\n      \"claim\": \"Discovery that CST1 accumulates in intracellular vesicular compartments during senescence broadened its biology beyond secretory protease inhibition and suggested an intracellular role linked to lysosomal proteostasis.\",\n      \"evidence\": \"Immunofluorescence and immunoblotting in human fibroblasts across multiple senescence triggers\",\n      \"pmids\": [\"21636832\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No functional test of whether vesicular CST1 is required for senescence phenotype\", \"Identity of the vesicular compartment not resolved (lysosomal vs. other)\"]\n    },\n    {\n      \"year\": 2018,\n      \"claim\": \"Identification of a CST1–TSLP/IL-33 positive feedback loop in nasal epithelial cells and fibroblasts revealed how CST1 amplifies type-2 airway inflammation, establishing its first defined signaling circuit outside of cancer.\",\n      \"evidence\": \"Cytokine stimulation of primary nasal epithelial cells and fibroblasts with RT-qPCR and protein measurement\",\n      \"pmids\": [\"29845224\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism by which CST1 induces TSLP not defined at the receptor level\", \"In vivo airway model not included in this study\"]\n    },\n    {\n      \"year\": 2021,\n      \"claim\": \"Demonstration that CST1 activates Wnt/β-catenin signaling to promote gastric cancer migration and invasion provided the first defined oncogenic pathway downstream of CST1, while ENO1 was placed upstream of CST1 in thyroid carcinoma, indicating CST1 acts as a pathway relay node.\",\n      \"evidence\": \"Luciferase reporter, β-catenin nuclear translocation, Wnt inhibitor rescue (gastric cancer); ENO1 OE/KD with CST1 rescue epistasis (thyroid carcinoma)\",\n      \"pmids\": [\"33658852\", \"33968941\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"No direct binding partner identified for Wnt pathway activation by CST1\", \"ENO1–CST1 regulatory mechanism not resolved at the promoter/protein level\", \"Single-lab findings in each cancer type\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"The critical mechanistic advance was the identification of a CST1–GPX4–OTUB1 ternary complex: CST1 physically binds GPX4 and recruits OTUB1 to deubiquitinate and stabilize GPX4, directly suppressing ferroptosis and promoting metastasis — the first molecular mechanism explaining how a secreted cystatin inhibits cell death.\",\n      \"evidence\": \"Reciprocal co-IP with mass spectrometry, ubiquitination assays, overexpression/knockdown in vitro and nude mouse metastasis model\",\n      \"pmids\": [\"36369321\"],\n      \"confidence\": \"High\",\n      \"gaps\": [\"Structural basis of CST1–GPX4 interaction unknown\", \"Whether catalytic cystatin activity is required for GPX4 stabilization not tested\"]\n    },\n    {\n      \"year\": 2022,\n      \"claim\": \"CST1 was shown to activate PI3K/AKT in airway smooth muscle cells to promote proliferation/migration, paralleling its oncogenic PI3K/AKT activation and extending its signaling repertoire to non-malignant disease.\",\n      \"evidence\": \"siRNA/overexpression with PI3K/AKT inhibitor rescue, EdU/CCK-8, Transwell assays in PDGF-BB-treated ASMCs\",\n      \"pmids\": [\"36354198\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Receptor or cell-surface partner through which secreted CST1 engages PI3K/AKT not identified\"]\n    },\n    {\n      \"year\": 2023,\n      \"claim\": \"Identification of MEK/ERK/CREB activation downstream of CST1 in ESCC and AKT–SERPINB2 axis in asthmatic epithelial cells expanded the signaling portfolio of CST1 and placed it upstream of multiple MAPK and AKT effector branches.\",\n      \"evidence\": \"Western blot for phospho-MEK/ERK/CREB, dual-luciferase for miR-942-5p targeting CST1 (ESCC); CST1 OE/KD with RNA-seq and p-AKT/SERPINB2 readout plus OVA mouse asthma model (airway)\",\n      \"pmids\": [\"36848349\", \"37075800\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Direct target/receptor mediating CST1-to-MAPK signal remains unknown\", \"SERPINB2 functional role downstream of AKT not fully dissected\"]\n    },\n    {\n      \"year\": 2024,\n      \"claim\": \"Upstream transcriptional control of CST1 was resolved: TFAP2C in breast cancer and BRD9/FOXP1 in gallbladder cancer each directly activate the CST1 promoter, linking epigenetic/transcription factor programs to the CST1–GPX4 ferroptosis suppression axis.\",\n      \"evidence\": \"ChIP + dual-luciferase reporter + siRNA rescue + xenograft models (TFAP2C); ChIP-qPCR + I-BRD9 inhibitor + mRNA-seq + in vivo GBC model (BRD9/FOXP1)\",\n      \"pmids\": [\"38243003\", \"39306629\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Whether TFAP2C and BRD9/FOXP1 act in the same or distinct cancer contexts is unexplored\", \"Promoter elements mediating each TF's binding not mapped at nucleotide resolution\"]\n    },\n    {\n      \"year\": 2025,\n      \"claim\": \"TFAP2A was validated as a second AP-2 family transcriptional activator of CST1 (in lung adenocarcinoma), and UBE2D2 was found to stabilize CST1 protein via suppression of autophagy-dependent degradation, together revealing both transcriptional and post-translational regulatory layers controlling CST1 abundance.\",\n      \"evidence\": \"ChIP + dual-luciferase + ferroptosis markers + xenograft (TFAP2A); proteomic screening + autophagy/ferroptosis assays + in vivo functional studies (UBE2D2)\",\n      \"pmids\": [\"39692484\", \"40912429\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Autophagy receptor or E3 ligase targeting CST1 for autophagic degradation not identified\", \"Whether UBE2D2 ubiquitinates CST1 directly or acts indirectly is unresolved\"]\n    },\n    {\n      \"year\": 2026,\n      \"claim\": \"NFATC2 was identified as a transcriptional activator of CST1 in cholangiocarcinoma, and CST1 was shown to suppress cellular senescence by stabilizing SOX4, revealing a new anti-senescence function beyond ferroptosis suppression.\",\n      \"evidence\": \"Multi-omics profiling, gain/loss-of-function, SOX4 rescue, SA-β-gal assay, orthotopic liver and pulmonary metastasis models\",\n      \"pmids\": [\"41881961\"],\n      \"confidence\": \"Medium\",\n      \"gaps\": [\"Mechanism by which CST1 stabilizes SOX4 protein not defined (direct binding vs. intermediate)\", \"Relationship between senescence suppression and ferroptosis suppression by CST1 not tested\"]\n    },\n    {\n      \"year\": null,\n      \"claim\": \"The cell-surface receptor or binding partner through which extracellular CST1 activates intracellular signaling cascades (Wnt, PI3K/AKT, MEK/ERK) remains unidentified, and whether its classical cysteine protease inhibitory activity is required for any of its oncogenic or anti-ferroptotic functions has not been tested.\",\n      \"evidence\": \"\",\n      \"pmids\": [],\n      \"confidence\": \"High\",\n      \"gaps\": [\"No receptor identified for CST1 signal transduction\", \"Protease-inhibitory activity not dissected from signaling functions using catalytic-dead mutants\", \"Structural basis of CST1–GPX4 interaction unknown\"]\n    }\n  ],\n  \"mechanism_profile\": {\n    \"molecular_activity\": [\n      {\"term_id\": \"GO:0098772\", \"supporting_discovery_ids\": [0, 6, 7, 11]},\n      {\"term_id\": \"GO:0140096\", \"supporting_discovery_ids\": [0, 12]}\n    ],\n    \"localization\": [\n      {\"term_id\": \"GO:0005576\", \"supporting_discovery_ids\": [15]},\n      {\"term_id\": \"GO:0031410\", \"supporting_discovery_ids\": [12]}\n    ],\n    \"pathway\": [\n      {\"term_id\": \"R-HSA-5357801\", \"supporting_discovery_ids\": [0, 6, 7, 11, 14]},\n      {\"term_id\": \"R-HSA-162582\", \"supporting_discovery_ids\": [1, 2, 3, 4, 10]},\n      {\"term_id\": \"R-HSA-1643685\", \"supporting_discovery_ids\": [0, 1, 2, 6, 7, 10, 13]},\n      {\"term_id\": \"R-HSA-168256\", \"supporting_discovery_ids\": [4, 5]}\n    ],\n    \"complexes\": [],\n    \"partners\": [\"GPX4\", \"OTUB1\", \"TFAP2C\", \"TFAP2A\", \"NFATC2\", \"BRD9\", \"SOX4\", \"UBE2D2\"],\n    \"other_free_text\": []\n  }\n}\n```"}