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The hyperactive Sleeping Beauty transposase SB100X improves the genetic modification of T cells to express a chimeric antigen receptor

Gene Therapy volume 18, pages 849–856 (2011)Cite this article

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Abstract

Sleeping Beauty (SB3) transposon and transposase constitute a DNA plasmid system used for therapeutic human cell genetic engineering. Here we report a comparison of SB100X, a newly developed hyperactive SB transposase, to a previous generation SB11 transposase to achieve stable expression of a CD19-specific chimeric antigen receptor (CAR3) in primary human T cells. The electro-transfer of SB100X expressed from a DNA plasmid or as an introduced mRNA species had superior transposase activity in T cells based on the measurement of excision circles released after transposition and emergence of CAR expression on T cells selectively propagated upon CD19+ artificial antigen-presenting cells. Given that T cells modified with SB100X and SB11 integrate on average one copy of the CAR transposon in each T-cell genome, the improved transposition mediated by SB100X apparently leads to an augmented founder effect of electroporated T cells with durable integration of CAR. In aggregate, SB100X improves SB transposition in primary human T cells and can be titrated with an SB transposon plasmid to improve the generation of CD19-specific CAR+ T cells.

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Abbreviations

SB:

Sleeping Beauty

CAR:

chimeric antigen receptor

aAPC:

artificial antigen presenting cell

PBMC:

peripheral blood mononuclear cells

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Acknowledgements

We thank Dr Perry Hackett University of Minnesota for Sleeping Beauty system and Dr Carl June from University of Pennsylvania for assistance generating the K562-aAPC. We are grateful to the Flow Cytometry Core Laboratory at MDACC. Cancer Prevention Research Institute of Texas, Department of Defense, PO1 (CA100265), CCSG (CA16672), RO1s (CA124782, CA120956), R33 (CA116127), Alex's Lemonade Stand Foundation, Alliance for Cancer Gene Therapy, Burroughs Wellcome Fund, Gillson Longenbaugh Foundation, Harry T Mangurian, Jr, Foundation, Institute of Personalized Cancer Therapy, Leukemia and Lymphoma Society, Lymphoma Research Foundation, Miller Foundation, National Foundation for Cancer Research, Pediatric Cancer Research Foundation, National Marrow Donor Program, William Lawrence and Blanche Hughes Foundation.

Author information

Authors and Affiliations

  1. Division of Pediatrics, Children's Cancer Hospital, The University of Texas Graduate School of Biomedical Sciences,

    Z Jin, S Maiti, H Huls, H Singh, S Olivares, D A Lee & L J N Cooper

  2. The University of Texas MD Anderson Cancer Center, Houston,, TX, USA

    Z Jin, S Maiti, H Huls, H Singh, S Olivares, D A Lee & L J N Cooper

  3. Max Delbrück Center for Molecular Medicine, Berlin, Germany

    L Mátés, Z Izsvák & Z Ivics

  4. University of Debrecen, Debrecen, Hungary

    Z Izsvák & Z Ivics

  5. Stem Cell Transplantation and Cellular Therapy, University of Texas MD Anderson Cancer Center, Houston, TX, USA

    R E Champlin

Authors
  1. Z Jin
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  2. S Maiti
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  3. H Huls
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  4. H Singh
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  7. Z Izsvák
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  9. D A Lee
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  11. L J N Cooper
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Correspondence to L J N Cooper.

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Supplementary Information accompanies the paper on Gene Therapy website

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Jin, Z., Maiti, S., Huls, H. et al. The hyperactive Sleeping Beauty transposase SB100X improves the genetic modification of T cells to express a chimeric antigen receptor. Gene Ther 18, 849–856 (2011). https://doi.org/10.1038/gt.2011.40

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