Abstract
Chimeric antigen receptor T cell (CAR-T) therapies have now entered mainstream clinical practice with two approved autologous CAR-T products targeting CD19 and numerous other products in early and late phase clinical trials. This has led to a demand for highly sensitive, specific, and easily reproducible methods to monitor CAR-T cells in patients. Here we describe a flow cytometry based protocol for detection of allogeneic CAR-T cells and for monitoring their phenotype and numbers in blood and bone marrow of patients following CAR-T treatment.
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Porter D, Hwang W, Frey N, Lacey S, Shaw P, Loren A, Bagg A, Marcucci K, Shen A, Gonzalez V, Ambrose D, Grupp S, Chew A, Zheng Z, Milone M, Levine B, Melenhorst J, June C (2015) Chimeric antigen receptor T cells persist and induce sustained remissions in relapsed refractory chronic lymphocytic leukemia. Sci Transl Med 7:303–139
Giavridis T, van der Stegen SJC, Eyquem J, Hamieh M, Piersigilli A (2018) Sadelain CAR T cell-induced cytokine release syndrome is mediated by macrophages and abated by IL-1 blockade. Nat Med 24(6):731–738
Graham C, Jozwik A, Pepper A, Benjamin R (2018) Allogeneic CAR-T cells: more than ease of access? Cells 7(10). https://doi.org/10.3390/cells7100155
Torikai H, Reik A, Liu PQ, Zhou Y, Zhang L, Maiti S, Huls H, Miller JC, Kebriaei P, Rabinovich B, Lee DA, Champlin RE, Bonini C, Naldini L, Rebar EJ, Gregory PD, Holmes MC, Cooper LJ (2012) A foundation for universal T-cell based immunotherapy: T cells engineered to express a CD19-specific chimeric-antigen-receptor and eliminate expression of endogenous TCR. Blood 119(24):5697–5705
Ren J, Liu X, Fang C, Jiang S, June CH, Zhao Y (2017) Multiplex genome editing to generate universal CAR T cells resistant to PD1 inhibition. Clin Cancer Res 23(9):2255–2266
Philip B, Kokalaki E, Mekkaoui L, Thomas S, Straathof K, Flutter B, Marin V, Marafioti T, Chakraverty R, Linch D, Quezada SA, Peggs KS, Pule M (2014) A highly compact epitope-based marker/suicide gene for easier and safer T-cell therapy. Blood 124(8):1277–1287
Poirot L, Philip B, Schiffer-Mannioui C, Le Clerre D, Chion-Sotinel I, Derniame S, Potrel P, Bas C, Lemaire L, Galetto R, Lebuhotel C, Eyquem J, Cheung GW, Duclert A, Gouble A, Arnould S, Peggs K, Pule M, Scharenberg AM, Smith J (2015) Multiplex genome-edited T-cell manufacturing platform for “off-the-shelf” adoptive T-cell immunotherapies. Cancer Res 75(18):3853–3864
Wang H, Du X, Chen WH, Lou J, Xiao HL, Pan YM, Chen H, An N, Zhang QX (2018) Establishment of a quantitative polymerase chain reaction assay for monitoring chimeric antigen receptor T cells in peripheral blood. Transplant Proc 50(1):104–109
Kalina T, JG Marvelde TE, van der Velden VHJ, Flores-Montero J, Thűrner D, Böttcher S, Cullen M, Lhermitte L, Sedek L, Mendonça A, Hrusak O, van Dongen JJM, Orfao A (2012) EuroFlow standardization of flow cytometer instrument settings and immunophenotyping protocols. Section 3. Design and evaluation of EuroFlow standard operating procedure for establishing optimal compensation settings. Leukemia 26:1986–2010
Stewart CC, Stewart SJ (1997) Titrating antibodies. In: Robinson JP et al (eds) Current protocols in cytometry. J. Wiley & Sons, New York, pp 4.1–4.4
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Jozwik, A., Dunlop, A., Sanchez, K., Benjamin, R. (2020). Monitoring Allogeneic CAR-T Cells Using Flow Cytometry. In: Katz, S., Rabinovich, P. (eds) Cell Reprogramming for Immunotherapy. Methods in Molecular Biology, vol 2097. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0203-4_19
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DOI: https://doi.org/10.1007/978-1-0716-0203-4_19
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Publisher Name: Humana, New York, NY
Print ISBN: 978-1-0716-0202-7
Online ISBN: 978-1-0716-0203-4
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