Generation of CAR+ T Lymphocytes Using the Sleeping Beauty Transposon System

  • Leonardo Chicaybam
  • Luiza Abdo
  • Martín H. BonaminoEmail author
Part of the Methods in Molecular Biology book series (MIMB, volume 2086)


Adoptive immunotherapy of cancer using T cells expressing chimeric antigen receptors (CARs) is now an approved treatment for non-Hodgkin lymphoma (NHL) and B cell acute lymphoblastic leukemia (B-ALL), inducing high response rates in patients. The infusion products are generated by using retro- or lentiviral transduction to induce CAR expression in T cells followed by an in vitro expansion protocol. However, use of viral vectors is cumbersome and is associated with increased costs due to the required high titers, replication-competent retrovirus (RCR) detection and production/use in a biosafety level 2 culture rooms, and additional quality control tests. Nonviral methods, like the Sleeping Beauty transposon system, can stably integrate in the genome of target cells and can be delivered using straightforward methods like electroporation. This chapter describes a protocol for T cell genetic modification using Sleeping Beauty transposon system and electroporation with the Lonza Nucleofector II device for the stable expression of CAR molecules in T lymphocytes.

Key words

Chimeric antigen receptor Sleeping beauty transposon T lymphocyte CD19 Electroporation 


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Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2020

Authors and Affiliations

  • Leonardo Chicaybam
    • 1
    • 2
  • Luiza Abdo
    • 1
  • Martín H. Bonamino
    • 1
    • 2
    Email author
  1. 1.Programa de Carcinogênese Molecular—Coordenação de PesquisaInstituto Nacional de CâncerRio de JaneiroBrazil
  2. 2.Vice-Presidência de Pesquisa e Coleções Biológicas (VPPCB)Fundação Instituto Oswaldo Cruz (FIOCRUZ)Rio de JaneiroBrazil

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