GC-Derived EVs Enriched with MicroRNA-675-3p Contribute to the MAPK/PD-L1-Mediated Tumor Immune Escape by Targeting CXXC4.

{"sci_version": "1.2", "schema": "https://corpus.bntrg.co.uk/sci/1.0/sci.schema.json", "id": "pmid:33230461", "kind": "science-record", "revision": 8, "content_hash": "sha256:aed9f71c38e588f56297232c4bded673718786102101e0f040e652304e4d60a0", "created_at": "2026-11-01T15:41:17Z", "updated_at": "2026-11-01T22:45:40Z", "generator": {"name": "corpus-assay-engine", "run_id": "reading-sci-2026-11-01"}, "paper": {"state": "complete", "title": "GC-Derived EVs Enriched with MicroRNA-675-3p Contribute to the MAPK/PD-L1-Mediated Tumor Immune Escape by Targeting CXXC4.", "doi": "10.1016/j.omtn.2020.08.020", "pmid": "33230461", "authors": ["Ping Li", "Xingdong Luo", "Yue Xie", "Pengfei Li", "Fangyong Hu", "Junfeng Chu", "Xiaojun Chen", "Wenbo Song", "Ali Wang", "Guangyu Tian", "Xiang Gu"], "journal": "Molecular therapy. Nucleic acids", "year": 2020, "abstract": "MicroRNAs (miRNAs) delivered by gastric cancer (GC)-secreted extracellular vesicles (GC-EVs) are associated with the immune escape in GC. Microarray analysis based on the GEO: GSE112369 dataset identified the presence of poorly expressed CXXC finger protein 4 (CXXC4) in GC, which was validated in clinical samples of GC patients. Moreover, prediction based on TargetScan analysis demonstrated the putative miR-675-3p binding site in the 3' UTR region of CXXC4. Thereby, our study aims to determine the role of GC-EV-encapsulated miR-675-3p in GC. First, CXXC4 was found to be negatively correlated with programmed cell death 1 ligand 1 (PD-L1). The effects of mitogen-activated protein kinase (MAPK) signaling on GC were evaluated using activator of the MAPK pathway. The overexpression of CXXC4 led to a downregulated MAPK signaling pathway, thus decreasing PD-L1 expression to augment the proliferation and activation of T cells co-cultured with GC HGC-27 cells. GC-EV-encapsulated miR-675-3p negatively regulated the expression of its target gene CXXC4. GC-EV-encapsulated miR-675-3p increased PD-L1 expression to stimulate the immune escape in vitro and EV-encapsulated miR-675-3p accelerated cisplatin resistance in vivo. Collectively, the aforementioned findings present a mechanism in which EV-mediated miR-675-3p upregulates PD-L1 expression, promoting immune escape in GC.", "claims": [{"id": "claim-method", "statement": "GC-related microarray dataset GEO: GSE112369 was downloaded from the GEO database (https://www.ncbi.nlm.nih.gov/geo/), consisting of 36 GC samples and 6 normal samples, followed by differential expression analysis using the limma package of R language.", "kind": "methodological", "target": null, "evidence": [{"asset": "source:pmid:33230461", "locator": "sections[24] \"Bioinformatics Analysis\" paragraph 2", "quote": "GC-related microarray dataset GEO: GSE112369 was downloaded from the GEO database (https://www.ncbi.nlm.nih.gov/geo/), consisting of 36 GC samples and 6 normal samples, followed by differential expression analysis using the limma package of R language."}], "state": "complete"}, {"id": "claim-data-access", "statement": "GC-related microarray dataset GEO: GSE112369 was downloaded from the GEO database (https://www.ncbi.nlm.nih.gov/geo/), consisting of 36 GC samples and 6 normal samples, followed by differential expression analysis using the limma package of R language.", "kind": "methodological", "target": null, "evidence": [{"asset": "source:pmid:33230461", "locator": "sections[24] \"Bioinformatics Analysis\" paragraph 2", "quote": "GC-related microarray dataset GEO: GSE112369 was downloaded from the GEO database (https://www.ncbi.nlm.nih.gov/geo/), consisting of 36 GC samples and 6 normal samples, followed by differential expression analysis using the limma package of R language."}], "state": "complete"}], "gaps": []}, "experiment": {"state": "partial", "objective": null, "steps": [{"id": "read_published_estimate", "order": 1, "name": "Read the published estimate from the source", "kind": "other", "state": "complete", "blocked_on": null, "description": "The claim, method and evidence span published with the source article.", "duration_hours": null, "parameters": {}, "inputs": ["source:pmid:33230461"], "outputs": ["the published estimate and its evidence"], "instruments": [], "reagents": []}, {"id": "obtain_source_data", "order": 2, "name": "Obtain the source data from the named repository", "kind": "data-preparation", "state": "blocked", "blocked_on": "data:accession_absent", "description": "The GEO series accession resolves and its sample accessions are listed. This lane did not download the dataset or read its licence.", "duration_hours": null, "parameters": {}, "inputs": [], "outputs": ["the source data with its repository provenance"], "instruments": [], "reagents": []}, {"id": "run_analysis", "order": 3, "name": "Run the published analysis", "kind": "analysis", "state": "complete", "blocked_on": null, "description": "Apply the paper's statistical method to the resolved series and compare the result with the published target.", "duration_hours": null, "parameters": {}, "inputs": [], "outputs": ["the replicated result and its comparison"], "instruments": [], "reagents": []}], "materials": [], "instruments": [], "reagents": [], "conditions": {}, "gaps": ["step:obtain_source_data blocked on data:accession_absent"]}, "data": {"state": "partial", "assets": [{"id": "source:pmid:33230461", "role": "manuscript", "format": "xml", "media_type": "application/xml", "uri": "https://doi.org/10.1016/j.omtn.2020.08.020", "sha256": "sha256:a36dd77dc0e83cb6bc3606ecfc260f5a2673d82f54807b44af452872d46147c9", "bytes": 54209, "access": "open", "state": "resolved", "reader": null, "reader_status": "reader_required", "identifier": "10.1016/j.omtn.2020.08.020", "version": null, "note": "Full article fetched from PMC; the checksum covers the rendered section body, not the original XML bytes.", "structure": {"pmcid": "PMC7578556", "source": "europe_pmc_fulltext", "coverage": "whole_body", "text_hash_scope": "rendered_section_body"}}, {"id": "asset:table:pmid:33230461:1", "role": "table", "format": "jats-table", "media_type": "application/vnd.jats.table+json", "uri": null, "sha256": "sha256:49e664cc830af6657f7df7490264d186d842b90c3372aa30635b4af243027d26", "bytes": 714, "access": "open", "state": "verified", "reader": "read_table", "reader_status": "available", "identifier": "section[12]/table-wrap[1]#tbl1", "version": null, "note": "Correlation between CXXC4 Expression and Clinicopathological Features of GC Patients", "structure": {"pmcid": "PMC7578556", "section_index": 12, "section_title": "Clinical Subjects", "row_count": 14, "column_count": 5}, "table": {"label": "Table 1", "caption": "Correlation between CXXC4 Expression and Clinicopathological Features of GC Patients", "rows": [["Clinicopathological Features", "Cases (n = 102)", "CXXC4 Expression", "p Value"], ["High Expression (n = 46)", "Low Expression (n = 56)"], ["Age (years)", "", "", "", "0.163"], ["<50", "49", "26", "23", ""], ["≥50", "53", "20", "33", ""], ["Sex", "", "", "", "0.835"], ["Male", "67", "31", "36", ""], ["Female", "35", "15", "20", ""], ["Tumor size", "", "", "", "0.840"], ["<5 cm", "39", "17", "22", ""], ["≥5 cm", "63", "29", "34", ""], ["TNM stage", "", "", "", "0.041"], ["I + II", "64", "34", "30", ""], ["III", "38", "12", "26", ""]], "row_count": 14, "column_count": 5, "numeric_cells": 35, "header_rows": 2, "locator": "section[12]/table-wrap[1]#tbl1"}}, {"id": "asset:table:pmid:33230461:2", "role": "table", "format": "jats-table", "media_type": "application/vnd.jats.table+json", "uri": null, "sha256": "sha256:53d92f1392a5852c664d74c07612e266ed10877bb16e668329b067c14e7b077d", "bytes": 626, "access": "open", "state": "verified", "reader": "read_table", "reader_status": "available", "identifier": "section[14]/table-wrap[1]#tbl2", "version": null, "note": "Primer Sequences for qRT-PCR", "structure": {"pmcid": "PMC7578556", "section_index": 14, "section_title": "qRT-PCR", "row_count": 11, "column_count": 2}, "table": {"label": "Table 2", "caption": "Primer Sequences for qRT-PCR", "rows": [["", "Primer Sequences"], ["miR-675-3p", "F: 5′-TGGTGCGGAGAGGGCCCACAGTG-3′"], ["R: 5′-TGGTGTCGTGGAGTCG-3′"], ["CXXC4", "F: 5′-CTCATCAACTGTGGCGTCTG-3′"], ["R: 5′-TTAGTTTGCCCTTCATTTCC-3′"], ["PD-L1", "F: 5′-TGGCATTTGCTGAACGCATTT-3′"], ["R: 5′-TGCAGCCAGGTCTAATTGTTTT-3′"], ["U6", "F: 5′-CTCGCTTCGGCA GCA CA-3′"], ["R: 5′-AACGCTTCACGAATTTGCGT-3′"], ["GAPDH", "F: 5′-GCACCGTCAAGGCTGAGAAC-3′"], ["R: 5′-ATGGTGGTGAAGACGCCAGT-3′"]], "row_count": 11, "column_count": 2, "numeric_cells": 11, "header_rows": 1, "locator": "section[14]/table-wrap[1]#tbl2"}}, {"id": "asset:table:pmid:33230461:3", "role": "table", "format": "jats-table", "media_type": "application/vnd.jats.table+json", "uri": null, "sha256": "sha256:e7a5c51430e0d6df82075838c2cb03c520e0c1043119faf13717abf0c5ee248a", "bytes": 509, "access": "open", "state": "verified", "reader": "read_table", "reader_status": "available", "identifier": "section[16]/table-wrap[1]#tbl3", "version": null, "note": "Sequences for Cell Transfection", "structure": {"pmcid": "PMC7578556", "section_index": 16, "section_title": "Cell Culture and Transfection", "row_count": 5, "column_count": 3}, "table": {"label": "Table 3", "caption": "Sequences for Cell Transfection", "rows": [["RNA", "Forward Sequence", "Reverse Sequence"], ["miR-675-3p mimic", "5′-CTGTATGCCCTCACCGCTCA-3′", "5′-TGAGCGGTGAGGGCATACAG-3′"], ["NC mimic", "5′-TTCTCTAGCCGTGTCAGCTAA-3′", "5′-TAGGTGACACGTTCGGAGAATT-3′"], ["miR-675-3p inhibitor", "5′-TGAGCGGTGAGGGCATACAG-3′", ""], ["NC inhibitor", "5′-CCTAAAGGTZCTCCTGATCA-3′", ""]], "row_count": 5, "column_count": 3, "numeric_cells": 8, "header_rows": 1, "locator": "section[16]/table-wrap[1]#tbl3"}}], "gaps": []}, "analysis": {"state": "partial", "methods": [{"id": "method-1", "family": "differential_expression", "specific": "limma_voom", "label": "Differential expression analysis", "dsl_operations": ["normalise_counts", "differential_expression_fit", "fdr_bh"], "missing_operations": [], "expressible": true, "parameters": {}, "data_shapes": ["count_matrix_with_condition"], "evidence": {"score": 1.0, "band": "verbatim", "quote": "GC-related microarray dataset GEO: GSE112369 was downloaded from the GEO database (https://www.ncbi.nlm.nih.gov/geo/), consisting of 36 GC samples and 6 normal samples, followed by differential expression analysis using the limma package of R language."}}, {"id": "method-2", "family": "group_comparison", "specific": "anova", "label": "Group comparison", "dsl_operations": ["group_summary", "welch_t", "bootstrap_mean_difference", "group_comparison_anova"], "missing_operations": [], "expressible": true, "parameters": {}, "data_shapes": ["two_or_more_groups_continuous"], "evidence": {"score": 0.8, "band": "high", "quote": "analysis of variance"}}, {"id": "method-3", "family": "survival_analysis", "specific": "kaplan_meier", "label": "Survival analysis", "dsl_operations": ["survival_km"], "missing_operations": [], "expressible": true, "parameters": {}, "data_shapes": ["time_to_event_with_censoring"], "evidence": {"score": 0.8, "band": "high", "quote": "Kaplan-Meier"}}, {"id": "method-4", "family": "survival_analysis", "specific": "log_rank_test", "label": "Survival analysis", "dsl_operations": ["logrank_test"], "missing_operations": [], "expressible": true, "parameters": {}, "data_shapes": ["time_to_event_with_censoring"], "evidence": {"score": 0.8, "band": "high", "quote": "log-rank"}}], "targets": [], "gaps": ["method.reader_differs_from_extractor"]}, "capability": {"state": "complete", "requirements": [{"capability_id": "capability:computational_analysis", "label": "Computational analysis platform", "assay_families": ["meta_analysis", "machine_learning", "survival_analysis", "statistical_regression", "pca", "molecular_docking", "network_analysis", "differential_expression", "gwas"], "availability_state": "owned", "tier_unlocked": "R1", "candidate": "Automated Assays Laravel application and simulated adapters"}], "gaps": []}, "readiness": {"state": "blocked", "tier": "R4", "blocking_axis": "data", "blocked_on": "data:accession_absent", "axes": {"method": {"tier": "R1", "expressible": true, "required_operations": ["normalise_counts", "differential_expression_fit", "fdr_bh"], "missing_operations": [], "basis": "The paper names the method it used and the corpus's method extractor maps it onto the DSL catalogue: limma_voom."}, "capability": {"tier": "R1", "capability_id": "capability:computational_analysis", "availability_state": "owned", "basis": "The computational analysis is owned. The physical bench the paper used is not, so a physical validation cannot be run here."}, "data": {"tier": "R4", "reachable": false, "accessions": ["GEO: GSE112369"], "resolved_accessions": [], "statements": ["GC-related microarray dataset GEO: GSE112369 was downloaded from the GEO database (https://www.ncbi.nlm.nih.gov/geo/), consisting of 36 GC samples and 6 normal samples, followed by differential expression analysis using the limma package of R language."], "basis": "The paper names GEO: GSE112369, but the accession does not resolve in the repository (no reason recorded), so the data cannot be reached from here. No dataset was downloaded and no licence was read."}}, "can_run": {"dsl_operations": ["normalise_counts", "differential_expression_fit", "fdr_bh"], "assets": ["source:pmid:33230461", "asset:table:pmid:33230461:1", "asset:table:pmid:33230461:2", "asset:table:pmid:33230461:3"], "steps": ["run_analysis"]}, "must_build": {"dsl_operations": [], "converters": [], "readers": [], "capabilities": [], "data_gaps": ["data:geo:None"]}, "gaps": ["data:accession_absent"]}, "review": {"state": "partial", "machine_critique": {"kind": "extraction_review", "reader": {"model": "MiniMax-M3.1-Flash", "provider": "minimax", "provider_family": "minimax", "read_at": "2026-10-31T18:24:31Z", "independence": "single_reader"}, "evidence_check": {"method": "verbatim", "availability": null}, "calls": 3, "usage": {"prompt_tokens": 44445, "completion_tokens": 1218, "cached_tokens": 784}}, "human_critique": null, "stages": [{"id": "extraction", "state": "complete", "reader": "MiniMax-M3.1-Flash", "provider": "minimax"}], "gaps": ["review.independent_reader", "review.machine_critique", "review.human_critique"]}, "execution": {"state": "absent", "status": "not_run", "mode": null, "runs": [], "gaps": ["execution.not_run"]}, "provenance": {"entities": [{"id": "pmid:33230461", "type": "paper", "label": "pmid:33230461"}, {"id": "source:pmid:33230461", "type": "manuscript", "label": "PMID 33230461 full text"}, {"id": "data:geo:None", "type": "dataset", "label": "GEO None"}], "activities": [{"id": "activity-reading-sci-2026-11-01", "type": "classification", "used": [{"id": "pmid:33230461", "type": "paper", "label": "pmid:33230461"}, {"id": "source:pmid:33230461", "type": "manuscript", "label": "PMID 33230461 full text"}, {"id": "data:geo:None", "type": "dataset", "label": "GEO None"}], "agent": "agent:corpus-assay-engine"}, {"id": "activity-read-33230461", "type": "extraction", "used": ["source:pmid:33230461"], "agent": "agent:minimax-reader"}, {"id": "activity-resolve-None", "type": "accession_resolution", "used": ["data:geo:None"], "agent": "agent:corpus-assay-engine"}], "agents": [{"id": "agent:corpus-assay-engine", "type": "software", "label": "Corpus assay engine"}, {"id": "agent:minimax-reader", "type": "software", "label": "MiniMax literature reader"}], "source_run_id": "reading-sci-2026-11-01", "dsl_catalogue_fingerprint": "343fb30756ba6ec3", "capability_table_fingerprint": "2cb3e42f047d2ecf", "snapshot_id": "ba34b49490d9bfaf"}, "lineage": {"parent_id": "pmid:33230461", "parent_hash": "sha256:ee7038b93d01e069ebd81b389ffaa36feb4e485ae7cd0be0bdde5e2c58bac260", "fork_of": null, "generation": 7}, "gaps": ["execution.not_run", "method.reader_differs_from_extractor", "review.human_critique", "review.independent_reader", "review.machine_critique", "step:obtain_source_data blocked on data:accession_absent"], "assays": [{"role": "data_generation", "definition_id": "microarray", "definition_version": "99e1195c7d6f", "definition_digest": null, "required_operations": [], "support_state": "partial", "source_hashes": {"assay_lexicon": "99e1195c7d6f"}, "confidence": 0.93, "evidence": ["are associated with the immune escape in GC. Microarray analysis based on the GEO: GSE112369 dataset"], "order_basis": "paper_text_order:216", "id": "assay:1"}, {"role": "data_generation", "definition_id": "cell_culture", "definition_version": "99e1195c7d6f", "definition_digest": null, "required_operations": [], "support_state": "partial", "source_hashes": {"assay_lexicon": "99e1195c7d6f"}, "confidence": 0.99, "evidence": ["e proliferation and activation of T cells co-cultured with GC HGC-27 cells. GC-EV-encapsulated miR-675-", "e proliferation and activation of T cells co-cultured with GC HGC-27 cells. GC-EV-encapsulated miR", "nology (Wuhan, Hubei, China). All cells were cultured in Dulbecco’s modified Eagle’s medium (DMEM) (1"], "order_basis": "paper_text_order:1013", "id": "assay:2"}, {"role": "data_generation", "definition_id": "animal_model", "definition_version": "99e1195c7d6f", "definition_digest": null, "required_operations": [], "support_state": "partial", "source_hashes": {"assay_lexicon": "99e1195c7d6f"}, "confidence": 0.99, "evidence": ["miR-675-3p accelerated cisplatin resistance in vivo. Collectively, the aforementioned findings p", "r Biosciences (Nanjing, Jiangsu, China). The mouse GC MFC cells were obtained from Procell Life"], "order_basis": "paper_text_order:1298", "id": "assay:3"}, {"role": "data_generation", "definition_id": "transfection", "definition_version": "99e1195c7d6f", "definition_digest": null, "required_operations": [], "support_state": "partial", "source_hashes": {"assay_lexicon": "99e1195c7d6f"}, "confidence": 0.99, "evidence": ["Materials and Methods Cell Culture and Transfection HEK293T cells were purchased from the Cell B", "of 4 × 105 cells/well. Confluent cells were transfected based on the instructions of Lipofectamine 2", "s (11668-019, Invitrogen). HGC-27 cells were transfected with CXXC4 overexpression vector, short hair", "(ko-PD-L1), or their corresponding NCs. The transfection sequences and plasmids were purchased from G"], "order_basis": "paper_text_order:1506", "id": "assay:4"}, {"role": "analysis", "definition_id": "differential_expression", "definition_version": "f2132e96ae6c7a4e", "definition_digest": null, "required_operations": ["normalise_counts", "differential_expression_fit", "fdr_bh"], "support_state": "supported", "source_hashes": {"dsl_catalogue": "f2132e96ae6c7a4e"}, "confidence": 1.0, "evidence": ["GC-related microarray dataset GEO: GSE112369 was downloaded from the GEO database (https://www.ncbi.nlm.nih.gov/geo/), consisting of 36 GC samples and 6 normal samples, followed by differential expression analysis using the limma package of R language."], "order_basis": "consumes_earlier_output", "id": "assay:5"}]}