{"doi":"10.1063/5.0207941","title":"CAR T cell infiltration and cytotoxic killing within the core of 3D breast cancer spheroids under the control of antigen sensing in microwell arrays","abstract":"The success of chimeric antigen receptor (CAR) T cells in blood cancers has intensified efforts to develop CAR T therapies for solid cancers. In the solid tumor microenvironment, CAR T cell trafficking and suppression of cytotoxic killing represent limiting factors for therapeutic efficacy. Here, we present a microwell platform to study CAR T cell interactions with 3D breast tumor spheroids and determine predictors of anti-tumor CAR T cell function. To precisely control antigen sensing, we utilized a switchable adaptor CAR system that covalently attaches to co-administered antibody adaptors and mediates antigen recognition. Following the addition of an anti-HER2 adaptor antibody, primary human CAR T cells exhibited higher infiltration, clustering, and secretion of effector cytokines. By tracking CAR T cell killing in individual spheroids, we showed the suppressive effects of spheroid size and identified the initial CAR T cell to spheroid area ratio as a predictor of cytotoxicity. We demonstrate that larger spheroids exhibit higher hypoxia levels and are infiltrated by CAR T cells with a suppressed activation state, characterized by reduced expression of IFN-γ, TNF-α, and granzyme B. Spatiotemporal analysis revealed lower CAR T cell numbers and cytotoxicity in the spheroid core compared to the periphery. Finally, increasing CAR T cell seeding density resulted in higher CAR T cell infiltration and cancer cell elimination in the spheroid core. Our findings provide new quantitative insight into CAR T cell function within 3D cancer spheroids. Given its miniaturized nature and live imaging capabilities, our microfabricated system holds promise for screening cellular immunotherapies.","journal":"APL Bioengineering","year":2024,"id":432378,"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":15,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9537,"is_data_producer":false,"deposit_databanks":null,"is_oa":true,"file_count":0,"downloads":0,"has_version_chain":false,"published_date":"2024-01-01","fair_score":null,"fair_percentile":null,"algorithm_id":"datarank_citation_only_1hop_v6","ranking_scope":"data_only","authors":[{"id":1237397,"name":"Matthew S. Laird","orcid":null,"position":1,"is_corresponding":false},{"id":1236953,"name":"Teddi Bishop","orcid":"0009-0003-7733-7402","position":2,"is_corresponding":false},{"id":1236954,"name":"Ruxuan Li","orcid":"0000-0002-3619-368X","position":3,"is_corresponding":false},{"id":1178767,"name":"Dorota E. Jazwinska","orcid":"0000-0002-1950-7747","position":4,"is_corresponding":false},{"id":1025387,"name":"Elisa Ruffo","orcid":"0000-0003-1990-7377","position":5,"is_corresponding":false},{"id":559131,"name":"Jason Lohmueller","orcid":"0000-0001-6089-9992","position":6,"is_corresponding":false},{"id":499879,"name":"Ioannis K. Zervantonakis","orcid":"0000-0003-2386-9553","position":7,"is_corresponding":false},{"id":1236952,"name":"Youngbin Cho","orcid":"0009-0006-6218-4214","position":0,"is_corresponding":true}],"reference_count":77,"raw_metadata":null,"created_at":"2026-07-19T01:59:35.267142Z","pmid":"39049849","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":[]}