CAR-T-cell therapy has shown considerable advance in recent years, being approved by regulatory agencies in US, Europe, and Japan for the treatment of refractory patients with CD19+ B-cell leukemia or diffuse large B-cell lymphoma. Current methods for CAR-T-cell production use viral vectors for T-cell genetic modification and can take up to 15 days to generate the infusion product. The development of simple and less costly manufacturing protocols is needed in order to meet the increasing demand for this therapy. In this present work, we generated 19BBz CAR-T cells in 8 days using a protocol based on the non-viral transposon-based vector Sleeping Beauty. The expanded cells display mostly a central memory phenotype, expressing higher levels of inhibitory receptors when compared with mock cells. In addition, CAR-T cells were cytotoxic against CD19+ leukemia cells in vitro and improved overall survival rates of mice xenografted with human RS4;11 or Nalm-6 B-cell leukemias. Infused CAR-T cells persisted for up to 28 days, showing that they are capable of long-term persistence and antitumor response. Altogether, these results demonstrate the effectiveness of our protocol and pave the way for a broader application of CAR-T-cell therapy.
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We would like to thank the Brazilian National Cancer Institute (INCA) Blood Bank and Hemotherapy Service for sample preparation and cell irradiation.
Funding for this study was provided by the Brazilian Ministry of Health, Brazilian National Cancer Institute (INCA), CNPq, CAPES, INCT Synthetic Biology (465603/2014-9), Ary Frauzino Cancer Foundation (FAF), Oncobiology Program - UFRJ, INOVA FIOCRUZ program, ICGEB/CNPq (405231/2015-6).
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Chicaybam, L., Abdo, L., Viegas, M. et al. Transposon-mediated generation of CAR-T cells shows efficient anti B-cell leukemia response after ex vivo expansion. Gene Ther (2020) doi:10.1038/s41434-020-0121-4