{"doi":"10.1101/2025.04.26.650803","title":"Optimizing canine T cell activation, expansion, and transduction","abstract":"Abstract Dogs are becoming an important model for human cancers, and successfully troubleshooting issues with genetically modified T cell immunotherapy for round cell and solid neoplasms in dogs provides a unique opportunity to improve efficacy, safety, and affordability for humans as well. Unfortunately, T cell activation in dogs for optimal viral transduction has not been determined, restricting advancements in canine T cell immunotherapy. Two αCD3 and two αCD28 antibody clones for canine T cell stimulation have been described in the literature, but no studies have been undertaken to evaluate which αCD3/αCD28 combination is most effective, nor has anyone directly compared the efficacy of the two most popular antibody presentation strategies: antibody-coated plates and antibody-conjugated beads. In evaluating the effects of plate- or bead-bound αCD3 stimulation alone versus αCD3/αCD28 in combination, we tested 12 possible antibody stimulation strategies in addition to evaluating two largely unexplored mitogens in canine T cell transduction, phorbol myristate acetate (PMA) with ionomycin and concanavalin A (ConA). We investigated the impact of these stimulation strategies on canine T cell activation, expansion, and transduction. For stimulation strategies producing the best results, we also examined how each strategy affected the proportions of CD4/CD8 T cell subsets and regulatory T cell (T reg ) prevalence. We determined that, in general, plate-bound antibodies were far superior to bead-bound antibodies for canine T cell stimulation, and that plate-bound αCD3 clone CA17.6F9 in combination with αCD28 clone 5B8 or the mitogen PMA with ionomycin produced better activation and expansion profiles, better transduction, and more desirable T cell subsets that are more likely to improve patient outcomes in dogs suffering from round cell and solid tumors.","journal":"bioRxiv (Cold Spring Harbor Laboratory)","year":2025,"id":565181,"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":0,"citer_count":0,"citers_with_citation_signal":0,"citers_with_endowment":0,"datacite_reuse_total":0,"is_dataset":false,"is_dataset_confidence":0.9593,"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":1439291,"name":"Jennifer C. Holmes","orcid":null,"position":1,"is_corresponding":false},{"id":1439292,"name":"Arianna Z He","orcid":null,"position":2,"is_corresponding":false},{"id":598504,"name":"Paul R. Hess","orcid":"0000-0002-8146-547X","position":3,"is_corresponding":false},{"id":378412,"name":"Christopher L. Mariani","orcid":"0000-0003-3228-6536","position":4,"is_corresponding":false},{"id":341734,"name":"Yevgeny Brudno","orcid":"0000-0003-2963-3293","position":5,"is_corresponding":false},{"id":181268,"name":"Tom Davis","orcid":null,"position":0,"is_corresponding":true}],"reference_count":90,"raw_metadata":{"citation_network_status":"fetched"},"created_at":"2026-07-19T02:56:28.709936Z","pmid":null,"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":[]}