Abstract
Lentivirus-mediated gene transfer is an efficient method to introduce a variety of transgenes to human T cells. Here we describe a protocol to transduce human CD4+, CD8+, or CD4+ regulatory T cells. To illustrate the method, we use transduction with lentivirus encoding an HLA-A2-specific chimeric antigen receptor (CAR) and a transduction marker as an example. Methods to isolate, transduce, purify, and expand CD4+ and CD8+ T cells as well as regulatory T cells are provided. We also describe how to carry out cytotoxicity or suppression assays to assess the function of the resulting CAR T cell or CAR regulatory T cells, respectively.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
References
Boardman DA, Levings MK (2019) Cancer immunotherapies repurposed for use in autoimmunity. Nat Biomed Eng 3:259–263
June CH, Riddell SR, Schumacher TN (2015) Adoptive cellular therapy: a race to the finish line. Sci Transl Med 7:280ps287
June CH, Sadelain M (2018) Chimeric antigen receptor therapy. N Engl J Med 379:64–73
Maldini CR, Ellis GI, Riley JL (2018) CAR T cells for infection, autoimmunity and allotransplantation. Nat Rev Immunol 18:605–616
Cieri N, Mastaglio S, Oliveira G, Casucci M, Bondanza A, Bonini C (2014) Adoptive immunotherapy with genetically modified lymphocytes in allogeneic stem cell transplantation. Immunol Rev 257:165–180
Roth TL, Puig-Saus C, Yu R, Shifrut E, Carnevale J, Li PJ et al (2018) Reprogramming human T cell function and specificity with non-viral genome targeting. Nature 559:405–409
Ando D, Meyer K (2017) Gene editing: regulatory and translation to clinic. Hematol Oncol Clin North Am 31:797–808
de Waal Malefyt R, Verma S, Bejarano MT, Ranes-Goldberg M, Hill M, Spits H (1993) CD2/LFA-3 or LFA-1/ICAM-1 but not CD28/B7 interactions can augment cytotoxicity by virus-specific CD8+ cytotoxic T lymphocytes. Eur J Immunol 23:418–424
de Waal Malefyt R, Yssel H, de Vries JE (1993) Direct effects of IL-10 on subsets of human CD4+ T cell clones and resting T cells. Specific inhibition of IL-2 production and proliferation. J Immunol 150:4754–4765
Suhoski MM, Golovina TN, Aqui NA, Tai VC, Varela-Rohena A, Milone MC et al (2007) Engineering artificial antigen-presenting cells to express a diverse array of co-stimulatory molecules. Mol Ther 15:981–988
Kutner RH, Zhang XY, Reiser J (2009) Production, concentration and titration of pseudotyped HIV-1-based lentiviral vectors. Nat Protoc 4:495–505
Dawson NA, Lamarche C, Hoeppli RE, Bergqvist P, Fung VC, McIver E et al (2019) Systematic testing and specificity mapping of alloantigen-specific chimeric antigen receptors in regulatory T cells. JCI Insight 4:e123672
MacDonald KG, Hoeppli RE, Huang Q, Gillies J, Luciani DS, Orban PC et al (2016) Alloantigen-specific regulatory T cells generated with a chimeric antigen receptor. J Clin Invest 126:1413–1424
Dawson NAJ, Rosado-Sánchez I, Novakovsky GE, Fung VCW, Huang Q, McIver E et al (2020) Functional effects of chimeric antigen receptor co-receptor signaling domains in human regulatory T cells. Sci Transl Med 12:eaaz3866
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2021 Springer Science+Business Media, LLC, part of Springer Nature
About this protocol
Cite this protocol
Fung, V.C.W., Rosado-Sánchez, I., Levings, M.K. (2021). Transduction of Human T Cell Subsets with Lentivirus. In: Annunziato, F., Maggi, L., Mazzoni, A. (eds) T-Helper Cells. Methods in Molecular Biology, vol 2285. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1311-5_19
Download citation
DOI: https://doi.org/10.1007/978-1-0716-1311-5_19
Published:
Publisher Name: Humana, New York, NY
Print ISBN: 978-1-0716-1310-8
Online ISBN: 978-1-0716-1311-5
eBook Packages: Springer Protocols