{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,7,24]],"date-time":"2026-07-24T15:05:22Z","timestamp":1784905522935,"version":"3.55.0"},"reference-count":28,"publisher":"MDPI AG","issue":"8","license":[{"start":{"date-parts":[[2026,7,24]],"date-time":"2026-07-24T00:00:00Z","timestamp":1784851200000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"name":"Ministry of Healthcare of the Republic of Kazakhstan","award":["BR25293293"],"award-info":[{"award-number":["BR25293293"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Bioengineering"],"abstract":"<jats:p>Lentiviral vector (LV) production for CAR-T therapy remains challenging due to the limited scalability of adherent HEK293 cell cultures. To address this, we adapted HEK293FT cells directly to serum-free FreeStyle 293 Expression Medium, generating a novel suspension cell line, HEK293FT-DS. We characterized growth kinetics, stability over 35 passages, and transient transfection parameters (PEI:DNA ratio, cell density). The suspension-adapted cells grew with &gt;93% viability and a specific growth rate of 0.54 day\u22121, and maintained stable viable cell density (1.51 \u00b1 0.03 \u00d7 106 cells\/mL) over 35 passages. The optimal PEI:DNA ratio was 2.5:1, and increasing the transfection cell density to 16 \u00d7 106 cells\/mL boosted functional titers to approximately 9 \u00d7 106 TU\/mL in clarified supernatant. Scaled production (1 L culture supernatant per batch) yielded up to 5 \u00d7 109 TU. The resulting vectors efficiently transduced primary human T cells (57% CAR-positive cells) in a CliniMACS Prodigy-based process. The platform based on suspension-adapted HEK293FT-DS cells enables high-density transfection and provides a cost-effective, scalable alternative to commercial LV production systems, particularly suited for academic CAR-T cell manufacturing.<\/jats:p>","DOI":"10.3390\/bioengineering13080856","type":"journal-article","created":{"date-parts":[[2026,7,24]],"date-time":"2026-07-24T13:53:26Z","timestamp":1784901206000},"page":"856","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Suspension-Adapted HEK293FT Cells Enable High-Density Transfection for Efficient Lentiviral Vector Production in CAR-T Therapy"],"prefix":"10.3390","volume":"13","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-0307-1053","authenticated-orcid":false,"given":"Alexandr","family":"Shevtsov","sequence":"first","affiliation":[{"name":"National Center for Biotechnology, Korgalzhin hwy 13\/5, 010000 Astana, Kazakhstan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0003-2199-5756","authenticated-orcid":false,"given":"Aitolkyn","family":"Kydyrbayeva","sequence":"additional","affiliation":[{"name":"National Center for Biotechnology, Korgalzhin hwy 13\/5, 010000 Astana, Kazakhstan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0009-0001-7868-3396","authenticated-orcid":false,"given":"Gaziza","family":"Nigmatulla","sequence":"additional","affiliation":[{"name":"National Center for Biotechnology, Korgalzhin hwy 13\/5, 010000 Astana, Kazakhstan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-8885-2387","authenticated-orcid":false,"given":"Viktoriya","family":"Keyer","sequence":"additional","affiliation":[{"name":"National Center for Biotechnology, Korgalzhin hwy 13\/5, 010000 Astana, Kazakhstan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Tolganay","family":"Kulatay","sequence":"additional","affiliation":[{"name":"National Center for Biotechnology, Korgalzhin hwy 13\/5, 010000 Astana, Kazakhstan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-1514-9302","authenticated-orcid":false,"given":"Gulzat","family":"Zauatbayeva","sequence":"additional","affiliation":[{"name":"National Center for Biotechnology, Korgalzhin hwy 13\/5, 010000 Astana, Kazakhstan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Bakytkali","family":"Ingirbay","sequence":"additional","affiliation":[{"name":"National Center for Biotechnology, Korgalzhin hwy 13\/5, 010000 Astana, Kazakhstan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Maral","family":"Zhumabekova","sequence":"additional","affiliation":[{"name":"National Center for Biotechnology, Korgalzhin hwy 13\/5, 010000 Astana, Kazakhstan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Dinara","family":"Zharlyganova","sequence":"additional","affiliation":[{"name":"National Center for Biotechnology, Korgalzhin hwy 13\/5, 010000 Astana, Kazakhstan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9880-9382","authenticated-orcid":false,"given":"Alexandr V.","family":"Shustov","sequence":"additional","affiliation":[{"name":"National Center for Biotechnology, Korgalzhin hwy 13\/5, 010000 Astana, Kazakhstan"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2026,7,24]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"376","DOI":"10.1007\/s12032-025-02923-x","article-title":"CAR T-cell therapy in hematologic and solid malignancies: Mechanisms, clinical applications, and future directions","volume":"42","author":"Kuttiappan","year":"2025","journal-title":"Med. 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