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Long Terminal Repeat CRISPR-CAR-Coupled "Universal" T Cells Mediate Potent Anti-leukemic Effects

Georgiadis, C; Preece, R; Nickolay, L; Etuk, A; Petrova, A; Ladon, D; Danyi, A; ... Qasim, W; + view all (2018) Long Terminal Repeat CRISPR-CAR-Coupled "Universal" T Cells Mediate Potent Anti-leukemic Effects. Molecular Therapy , 26 (5) pp. 1215-1227. 10.1016/j.ymthe.2018.02.025. Green open access

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Abstract

Gene editing can be used to overcome allo-recognition, which otherwise limits allogeneic T cell therapies. Initial proof-of-concept applications have included generation of such "universal" T cells expressing chimeric antigen receptors (CARs) against CD19 target antigens combined with transient expression of DNA-targeting nucleases to disrupt the T cell receptor alpha constant chain (TRAC). Although relatively efficient, transgene expression and editing effects were unlinked, yields variable, and resulting T cell populations heterogeneous, complicating dosing strategies. We describe a self-inactivating lentiviral "terminal" vector platform coupling CAR expression with CRISPR/Cas9 effects through incorporation of an sgRNA element into the ΔU3 3' long terminal repeat (LTR). Following reverse transcription and duplication of the hybrid ΔU3-sgRNA, delivery of Cas9 mRNA resulted in targeted TRAC locus cleavage and allowed the enrichment of highly homogeneous (>96%) CAR+ (>99%) TCR- populations by automated magnetic separation. Molecular analyses, including NGS, WGS, and Digenome-seq, verified on-target specificity with no evidence of predicted off-target events. Robust anti-leukemic effects were demonstrated in humanized immunodeficient mice and were sustained longer than by conventional CAR+TCR+ T cells. Terminal-TRAC (TT) CAR T cells offer the possibility of a pre-manufactured, non-HLA-matched CAR cell therapy and will be evaluated in phase 1 trials against B cell malignancies shortly.

Type: Article
Title: Long Terminal Repeat CRISPR-CAR-Coupled "Universal" T Cells Mediate Potent Anti-leukemic Effects
Location: United States
Open access status: An open access version is available from UCL Discovery
DOI: 10.1016/j.ymthe.2018.02.025
Publisher version: https://doi.org/10.1016/j.ymthe.2018.02.025
Language: English
Additional information: This version is the author accepted manuscript. For information on re-use, please refer to the publisher’s terms and conditions.
Keywords: B-ALL, CAR T cells, CRISPR/Cas9 gene editing, lentiviral vector gene therapy
UCL classification: UCL
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Population Health Sciences > UCL GOS Institute of Child Health
UCL > Provost and Vice Provost Offices > School of Life and Medical Sciences > Faculty of Population Health Sciences > UCL GOS Institute of Child Health > Infection, Immunity and Inflammation Dept
URI: https://discovery.ucl.ac.uk/id/eprint/10047336
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