CAR T cell manufacturing from naive/stem memory T lymphocytes enhances antitumor responses while curtailing cytokine release syndrome
Silvia Arcangeli, … , Attilio Bondanza, Monica Casucci
Silvia Arcangeli, … , Attilio Bondanza, Monica Casucci
Published May 3, 2022
Citation Information: J Clin Invest. 2022;132(12):e150807. https://doi.org/10.1172/JCI150807.
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CAR T cell manufacturing from naive/stem memory T lymphocytes enhances antitumor responses while curtailing cytokine release syndrome

Abstract

Chimeric antigen receptor (CAR) T cell expansion and persistence represent key factors to achieve complete responses and prevent relapses. These features are typical of early memory T cells, which can be highly enriched through optimized manufacturing protocols. Here, we investigated the efficacy and safety profiles of CAR T cell products generated from preselected naive/stem memory T cells (TN/SCM), as compared with unselected T cells (TBULK). Notwithstanding their reduced effector signature in vitro, limiting CAR TN/SCM doses showed superior antitumor activity and the unique ability to counteract leukemia rechallenge in hematopoietic stem/precursor cell–humanized mice, featuring increased expansion rates and persistence together with an ameliorated exhaustion and memory phenotype. Most relevantly, CAR TN/SCM proved to be intrinsically less prone to inducing severe cytokine release syndrome, independently of the costimulatory endodomain employed. This safer profile was associated with milder T cell activation, which translated into reduced monocyte activation and cytokine release. These data suggest that CAR TN/SCM are endowed with a wider therapeutic index compared with CAR TBULK.

Authors

Silvia Arcangeli, Camilla Bove, Claudia Mezzanotte, Barbara Camisa, Laura Falcone, Francesco Manfredi, Eugenia Bezzecchi, Rita El Khoury, Rossana Norata, Francesca Sanvito, Maurilio Ponzoni, Beatrice Greco, Marta Angiola Moresco, Matteo G. Carrabba, Fabio Ciceri, Chiara Bonini, Attilio Bondanza, Monica Casucci

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Figure 1

CAR TN/SCM display an indolent effector signature in vitro.

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CAR TN/SCM display an indolent effector signature in vitro. (A) Schemati...
(A) Schematic representation of CAR T cell manufacturing. Briefly, double-positive CD62L+CD45RA+ TN/SCM cells were isolated by FACS and bulk unselected T cells (TBULK) were employed as control. TN/SCM and TBULK were activated with TransAct (anti-CD3/anti-CD28 [αCD3/28]), transduced with a lentiviral vector (LV) encoding a CD19.28z CAR, and expanded in culture with IL-7 and IL-15. (B) CD19.28z CAR expression (mean fluorescence intensity, MFI; n = 5), (C) TSCM enrichment (n = 16), (D) CD4+/CD8+ ratio (n = 20), and (E) HLA-DR expression (percentage of positive cells, n = 18) at the end of CAR T cell manufacturing. (F) Fold expansion at different days of culture (n = 12). (G) Degranulation assay performed by coculturing CAR TN/SCM, CAR TBULK, and Mock control with CD19+ targets for 24 hours (n = 14 donors challenged against NALM-6, BV173, and ALL-CM CD19+ target cell lines). (H) Killing activity expressed as elimination index (see Methods) and measured by coculturing CAR T cells with CD19+ tumor cells for 4 days at different effector/target (E:T) ratios (n = 15 for CAR TBULK, n = 14 for CAR TN/SCM against NALM-6 cell line; n = 7 for CAR TBULK, n = 6 for CAR TN/SCM against BV173 cell line; n = 8 for CAR TN/SCM, n = 9 for CAR TBULK against ALL-CM cell line). (I) Cytokine (CTK) production after 24-hour coculture of T cells with CD19+ tumor cells at a 1:10 E:T ratio (n = 5 donors challenged against NALM-6, BV173, and ALL-CM cell lines). (J) T cell proliferation after 4-day coculture with CD19+ tumor cells, measured by intracellular staining of Ki-67 (n = 15 donors challenged against NALM-6, BV173, and ALL-CM cell lines). Data are represented as mean ± SEM or mean ± SEM together with overlapping scattered values. *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001 by paired t test (B–E, G, and L) or 2-way ANOVA (F, H, and I).