Preclinical Efficacy and Safety of CD19CAR Cytokine-Induced Killer Cells Transfected with Sleeping Beauty Transposon for the Treatment of Acute Lymphoblastic Leukemia

Hum Gene Ther. 2018 May;29(5):602-613. doi: 10.1089/hum.2017.207. Epub 2018 Apr 16.


Infusion of patient-derived CD19-specific chimeric antigen receptor (CAR) T cells engineered by viral vectors achieved complete remission and durable response in relapsed and refractory (r/r) B-lineage neoplasms. Here, we expand on those findings by providing a preclinical evaluation of allogeneic non-viral cytokine-induced killer (CIK) cells transfected with the Sleeping Beauty (SB) transposon CD19CAR (CARCIK-CD19). Specifically, thanks to a large-scale 18-day manufacturing process, it was possible to achieve stable CD19CAR expression (62.425 ± 6.399%) and efficient T-cell expansion (23.36 ± 3.00-fold). Frozen/thawed CARCIK-CD19 remained fully functional both in vitro and in an established patient-derived xenograft (PDX) of MLL-ENL rearranged acute lymphoblastic leukemia (ALL). CARCIK-CD19 showed a dose-dependent antitumor response and prolonged persistence in a PDX, bearing the feature of a Philadelphia-like ALL with PAX5/AUTS2 translocation, and in a survival model of lymphoma, achieving complete eradication of disseminated tumors. Finally, the infusion of CARCIK-CD19 proved to be safe and well tolerated in a biodistribution and toxicity model. The infused cells persisted in the hematopoietic and post-injection perfused organs until the end of the study and consisted of CD8+, CD56+, and CAR+ T cells. Overall, these findings provide important implications for non-viral technology and the proof-of-concept that donor-derived CARCIK-CD19 are indeed effective against relapsed ALL, a possibility that will be tested in Phase I/II clinical trials after allogeneic hematopoietic stem-cell transplantation.

Keywords: Sleeping Beauty transposon; acute lymphoblastic leukemia; chimeric antigen receptor; cytokine-induced killer cells.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Antigens, CD19 / genetics
  • Antigens, CD19 / immunology
  • Antigens, CD19 / therapeutic use
  • Cytokine-Induced Killer Cells / immunology*
  • Gene Expression Regulation, Neoplastic / genetics
  • Genetic Vectors / genetics
  • Genetic Vectors / therapeutic use
  • Humans
  • Immunotherapy, Adoptive*
  • Mice
  • Precursor Cell Lymphoblastic Leukemia-Lymphoma / genetics
  • Precursor Cell Lymphoblastic Leukemia-Lymphoma / immunology
  • Precursor Cell Lymphoblastic Leukemia-Lymphoma / pathology
  • Precursor Cell Lymphoblastic Leukemia-Lymphoma / therapy*
  • Receptors, Antigen, T-Cell / genetics
  • Receptors, Antigen, T-Cell / immunology
  • Receptors, Antigen, T-Cell / therapeutic use*
  • T-Lymphocytes / immunology
  • Transfection
  • Xenograft Model Antitumor Assays


  • Antigens, CD19
  • CD19-specific chimeric antigen receptor
  • Receptors, Antigen, T-Cell