{"doi":"10.1016/j.iotech.2025.101052","title":"Assessment of CD73 activity in breast cancer-derived small extracellular vesicles: application to monitoring of patients’ responses to immunotherapy","abstract":"Background We previously discovered that small extracellular vesicles (sEV) isolated from melanoma cells produce immunosuppressive adenosine (ADO) via the ATP→ADP→AMP→ADO pathway and that CD73 is the ‘gateway' ecto-nucleotidase used by melanoma sEV to generate ADO. Here we extend these findings to CD39(+)CD73(+) and CD39(+)CD73(−) sEV from breast cancer cells. Materials and methods sEV were isolated from supernatants of a triple-negative breast cancer cell line ± the genetic knockout of CD73. A newly developed high pressure liquid chromatography assay with fluorescence detection was used for assessment of N 6 -etheno-AMP conversion to N 6 -etheno-ADO by sEV. PSB12379 (selective CD73 inhibitor) and anti-CD73 antibodies were used to inhibit/neutralize CD73 activity in sEV. Results Untreated sEV isolated from CD39(+)CD73(+) breast cancer cells readily metabolized N 6 -etheno-AMP to N 6 -etheno-ADO, and this activity was abolished by PSB12379. sEV from CD39(+)CD73(−) breast cancer cells were unable to metabolize N 6 -etheno-AMP to N 6 -etheno-ADO. Effects of three different anti-CD73 antibodies on CD73 activity in sEV were examined. Only one antibody, the direct binding pocket inhibitor of CD73, but not antibodies that allosterically inhibit recombinant CD73, attenuated conversion of N 6 -etheno-AMP to N 6 -etheno-ADO by cancer-derived sEV. Conclusions In breast cancer-derived sEV, as in melanoma-derived sEV, CD73 is the gateway enzyme regulating ADO formation from upstream AMP. The quantitation in sEV of N 6 -etheno-AMP conversion to N 6 -etheno-ADO ± neutralizing anti-CD73 antibodies provides a measure of the ability of these antibodies to suppress ADO production and could potentially serve as a personalized predictor of CD73 activity in patients with cancer.","journal":"Immuno-Oncology Technology","year":2025,"id":552104,"datarank":0.0,"base_score":0.0,"endowment":0.0,"self_citation_contribution":0.0,"citation_network_contribution":0.0,"self_endowment_contribution":0.0,"citer_contribution":0.0,"corpus_percentile":null,"corpus_rank":null,"citation_count":1,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9626,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2025-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":796579,"name":"Sarita Sehra","orcid":null,"position":1,"is_corresponding":false},{"id":1448311,"name":"T Chadderton","orcid":null,"position":2,"is_corresponding":false},{"id":500942,"name":"Manti Guha","orcid":"0000-0003-1576-3580","position":3,"is_corresponding":false},{"id":333581,"name":"Matthew C. Stubbs","orcid":"0000-0003-3771-7083","position":4,"is_corresponding":false},{"id":469161,"name":"Cynthia Timmers","orcid":"0000-0002-0509-0805","position":5,"is_corresponding":false},{"id":264558,"name":"Edwin K. Jackson","orcid":"0000-0002-8101-6009","position":6,"is_corresponding":false},{"id":225451,"name":"Theresa L. Whiteside","orcid":"0000-0001-7316-6181","position":0,"is_corresponding":true}],"reference_count":33,"raw_metadata":null,"created_at":"2026-07-19T02:54:29.318076Z","pmid":"40390761","pmcid":null,"fwci":null,"citation_percentile":null,"influential_citations":0,"oa_status":null,"license":null,"views":0,"total_file_size_bytes":0,"version_count":0,"fair_f":null,"fair_a":null,"fair_i":null,"fair_r":null,"fair_zscore":null,"fair_rationale":null,"fair_model":null,"fair_agent_version":null,"fair_fulltext_source":null,"fair_has_llm":null,"fair_computed_at":null,"clinical_trials":[],"software_tools":[],"db_accessions":[],"linked_datasets":[],"topics":[]}