{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,9,9]],"date-time":"2026-09-09T10:17:03Z","timestamp":1788949023303,"version":"build-2803163510"},"reference-count":70,"publisher":"MDPI AG","issue":"3","license":[{"start":{"date-parts":[[2025,1,23]],"date-time":"2025-01-23T00:00:00Z","timestamp":1737590400000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"German Federal Ministry of Education and Research","award":["16GW0322"],"award-info":[{"award-number":["16GW0322"]}]},{"name":"Fraunhofer Cluster of Excellence Immune-Mediated Diseases (CIMD) Cell Therapy and RNA Therapy Platforms","award":["16GW0322"],"award-info":[{"award-number":["16GW0322"]}]},{"name":"Fraunhofer Flagship project \u00abRNAuto\u00bb","award":["16GW0322"],"award-info":[{"award-number":["16GW0322"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["IJMS"],"abstract":"<jats:p>Genetically modified T lymphocytes expressing chimeric antigen receptors (CARs) are becoming increasingly important in the treatment of hematologic malignancies and are also intensively being investigated for other diseases such as autoimmune disorders and HIV. Current CAR T cell therapies predominantly use viral transduction methods which, despite their efficacy, raise safety concerns related to genomic integration and potentially associated malignancies as well as labor- and cost-intensive manufacturing. Therefore, non-viral gene transfer methods, especially mRNA-based approaches, have attracted research interest due to their transient modification and enhanced safety profile. In this study, the optimization of CAR-mRNA for T cell applications is investigated, focusing on the impact of mRNA modifications, in vitro transcription protocols, and purification techniques on the translation efficiency and immunogenicity of mRNA. Furthermore, the refined CAR-mRNA was used to generate transient CAR T cells from acute myeloid leukemia patient samples, demonstrating efficacy in vitro and proof-of-concept for clinically relevant settings. These results highlight the potential of optimized mRNA to produce transient and safe CAR T cells.<\/jats:p>","DOI":"10.3390\/ijms26030965","type":"journal-article","created":{"date-parts":[[2025,1,23]],"date-time":"2025-01-23T12:24:09Z","timestamp":1737635049000},"page":"965","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":17,"title":["Optimal Chimeric Antigen Receptor (CAR)-mRNA for Transient CAR T Cell Generation"],"prefix":"10.3390","volume":"26","author":[{"given":"Reni","family":"Kitte","sequence":"first","affiliation":[{"name":"Fraunhofer Institute for Cell Therapy and Immunology (IZI), Perlickstr. 1, 04103 Leipzig, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Robert","family":"Serfling","sequence":"additional","affiliation":[{"name":"Fraunhofer Institute for Cell Therapy and Immunology (IZI), Perlickstr. 1, 04103 Leipzig, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4233-2268","authenticated-orcid":false,"given":"Ulrich","family":"Blache","sequence":"additional","affiliation":[{"name":"Fraunhofer Institute for Cell Therapy and Immunology (IZI), Perlickstr. 1, 04103 Leipzig, Germany"},{"name":"Fraunhofer Cluster of Excellence Immune-Mediated Diseases (CIMD), Perlickstr. 1, 04103 Leipzig, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Claudius","family":"Seitz","sequence":"additional","affiliation":[{"name":"Fraunhofer Institute for Toxicology and Experimental Medicine (ITEM), Inhoffenstra\u00dfe 7, 38124 Braunschweig, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Selina","family":"Schrader","sequence":"additional","affiliation":[{"name":"Fraunhofer Institute for Toxicology and Experimental Medicine (ITEM), Inhoffenstra\u00dfe 7, 38124 Braunschweig, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ulrike","family":"K\u00f6hl","sequence":"additional","affiliation":[{"name":"Fraunhofer Institute for Cell Therapy and Immunology (IZI), Perlickstr. 1, 04103 Leipzig, Germany"},{"name":"Fraunhofer Cluster of Excellence Immune-Mediated Diseases (CIMD), Perlickstr. 1, 04103 Leipzig, Germany"},{"name":"Institute of Clinical Immunology, Medical Faculty, University of Leipzig, Johannisallee 30, 04103 Leipzig, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Stephan","family":"Fricke","sequence":"additional","affiliation":[{"name":"Fraunhofer Institute for Cell Therapy and Immunology (IZI), Perlickstr. 1, 04103 Leipzig, Germany"},{"name":"Fraunhofer Cluster of Excellence Immune-Mediated Diseases (CIMD), Perlickstr. 1, 04103 Leipzig, Germany"},{"name":"Medicine Campus MEDiC, Technical University of Dresden, Klinikum Chemnitz gGmbH, 09116 Chemnitz, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Christian","family":"B\u00e4r","sequence":"additional","affiliation":[{"name":"Fraunhofer Institute for Toxicology and Experimental Medicine (ITEM), Nikolai-Fuchs-Stra\u00dfe 1, 30625 Hannover, Germany"},{"name":"Fraunhofer Cluster of Excellence Immune-Mediated Diseases (CIMD), Nikolai-Fuchs-Stra\u00dfe 1, 30625 Hannover, Germany"},{"name":"Institute of Molecular and Translational Therapeutic Strategies, Hannover Medical School, Carl-Neuberg-Str. 1, 30625 Hannover, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"U. Sandy","family":"Tretbar","sequence":"additional","affiliation":[{"name":"Fraunhofer Institute for Cell Therapy and Immunology (IZI), Perlickstr. 1, 04103 Leipzig, Germany"},{"name":"Fraunhofer Cluster of Excellence Immune-Mediated Diseases (CIMD), Perlickstr. 1, 04103 Leipzig, Germany"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2025,1,23]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"359","DOI":"10.1038\/s41571-023-00754-1","article-title":"Long-term outcomes following CAR T cell therapy: What we know so far","volume":"20","author":"Cappell","year":"2023","journal-title":"Nat. Rev. Clin. Oncol."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"2124","DOI":"10.1038\/s41591-022-02017-5","article-title":"Anti-CD19 CAR T cell therapy for refractory systemic lupus erythematosus","volume":"28","author":"Mackensen","year":"2022","journal-title":"Nat. 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