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Non-MHC-Dependent Redirected T Cells Against Tumor Cells

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RNA Therapeutics

Part of the book series: Methods in Molecular Biology ((MIMB,volume 629))

Abstract

Adoptive transfer of T cells with restricted tumor specificity provides a promising approach to immunotherapy of cancers. However, the isolation of autologous cytotoxic T cells that recognize tumor-associated antigens is time consuming and fails in many instances. Alternatively, gene modification with tumor antigen-specific T-cell receptors (TCR) or chimeric antigen receptors (CARs) can be used to redirect the specificity of large numbers of immune cells toward the malignant cells. Chimeric antigen receptors are composed of the single-chain variable fragment (scFv) of a tumor-recognizing antibody cloned in frame with human T-cell signaling domains (e.g., CD3ζ, CD28, OX40, 4-1BB), thus combining the specificity of antibodies with the effector functions of cytotoxic T cells. Upon antigen binding, the intracellular signaling domains of the CAR initiate cellular activation mechanisms including cytokine secretion and cytolysis of the antigen-positive target cell.

In this chapter, we provide detailed protocols for large-scale ex vivo expansion of T cells and manufacturing of medium-scale batches of CAR-expressing T cells for translational research by mRNA electroporation. An anti-CD19 chimeric receptor for the targeting of leukemias and lymphomas was used as a model system. We are currently scaling up the protocols to adapt them to cGMP production of a large number of redirected T cells for clinical applications

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Acknowledgments

This work was supported in part by CHILDHOPE (EU contract #037381) and the authors would like to thank Dr. Martin Pulè at UCL Cancer Institute, University College London, UK, for providing the vector for the chimeric antigen receptor. The authors would also thank Dr. Stein Sæebøe-Larssen at our laboratory for providing the pCIpA102 expression vector and Dr. Rainer Loew at EUFETS, Idar-Oberstein, Germany, for the IRES-EGFP construct.

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Authors and Affiliations

  1. Department of Immunology, Radiumhospitalet-Rikshospitalet, University Hospital, Oslo, Norway

    Hilde Almåsbak & Anne-Marie Rasmussen

  2. Department of Cellular Therapy, Radiumhospitalet-Rikshospitalet, University Hospital, Oslo, Norway

    Marianne Lundby

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  1. Hilde Almåsbak
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  2. Marianne Lundby
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  3. Anne-Marie Rasmussen
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Editors and Affiliations

  1. Institute for Cancer Research, Dept. Immunology, Norwegian Radium Hospital, Montebello, Oslo, 0310, Norway

    Mouldy Sioud

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Almåsbak, H., Lundby, M., Rasmussen, AM. (2010). Non-MHC-Dependent Redirected T Cells Against Tumor Cells. In: Sioud, M. (eds) RNA Therapeutics. Methods in Molecular Biology, vol 629. Humana Press. https://doi.org/10.1007/978-1-60761-657-3_28

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  • DOI: https://doi.org/10.1007/978-1-60761-657-3_28

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