Metabolic priming of GD2 TRAC-CAR T cells during manufacturing promotes memory phenotypes while enhancing persistence

Dan Cappabianca; Dan Pham; Matthew H. Forsberg; Madison Bugel; Anna Tommasi; Anthony Lauer; Jolanta Vidugiriene; Brookelyn Hrdlicka; Alexandria McHale; Quaovi H. Sodji; Melissa C. Skala; Christian M. Capitini; Krishanu Saha

Molecular Therapy: Methods & Clinical Development (Jun 2024)

Metabolic priming of GD2 TRAC-CAR T cells during manufacturing promotes memory phenotypes while enhancing persistence

Dan Cappabianca,
Dan Pham,
Matthew H. Forsberg,
Madison Bugel,
Anna Tommasi,
Anthony Lauer,
Jolanta Vidugiriene,
Brookelyn Hrdlicka,
Alexandria McHale,
Quaovi H. Sodji,
Melissa C. Skala,
Christian M. Capitini,
Krishanu Saha

Affiliations

Dan Cappabianca: Wisconsin Institute for Discovery, University of Wisconsin-Madison, Madison, WI 53715, USA; Department of Biomedical Engineering, University of Wisconsin-Madison, Madison, WI 53715, USA
Dan Pham: Department of Biomedical Engineering, University of Wisconsin-Madison, Madison, WI 53715, USA; Morgridge Institute for Research, University of Wisconsin-Madison, Madison, WI 53715, USA
Matthew H. Forsberg: Department of Pediatrics, University of Wisconsin School of Medicine and Public Health, Madison, WI 53705, USA
Madison Bugel: Wisconsin Institute for Discovery, University of Wisconsin-Madison, Madison, WI 53715, USA; Department of Biomedical Engineering, University of Wisconsin-Madison, Madison, WI 53715, USA
Anna Tommasi: Wisconsin Institute for Discovery, University of Wisconsin-Madison, Madison, WI 53715, USA; Department of Biomedical Engineering, University of Wisconsin-Madison, Madison, WI 53715, USA
Anthony Lauer: Promega Corporation, Fitchburg, WI 53711, USA
Jolanta Vidugiriene: Promega Corporation, Fitchburg, WI 53711, USA
Brookelyn Hrdlicka: Wisconsin Institute for Discovery, University of Wisconsin-Madison, Madison, WI 53715, USA; Department of Biomedical Engineering, University of Wisconsin-Madison, Madison, WI 53715, USA
Alexandria McHale: Wisconsin Institute for Discovery, University of Wisconsin-Madison, Madison, WI 53715, USA; Department of Biomedical Engineering, University of Wisconsin-Madison, Madison, WI 53715, USA
Quaovi H. Sodji: University of Wisconsin Carbone Cancer Center, University of Wisconsin-Madison, Madison, WI 53705, USA; Department of Human Oncology, University of Wisconsin School of Medicine and Public Health, Madison, WI 53705, USA
Melissa C. Skala: Department of Biomedical Engineering, University of Wisconsin-Madison, Madison, WI 53715, USA; Morgridge Institute for Research, University of Wisconsin-Madison, Madison, WI 53715, USA; University of Wisconsin Carbone Cancer Center, University of Wisconsin-Madison, Madison, WI 53705, USA; Corresponding author: Melissa C. Skala, Department of Biomedical Engineering, University of Wisconsin-Madison, Madison, WI 53715, USA.
Christian M. Capitini: Department of Pediatrics, University of Wisconsin School of Medicine and Public Health, Madison, WI 53705, USA; University of Wisconsin Carbone Cancer Center, University of Wisconsin-Madison, Madison, WI 53705, USA; Corresponding author: Christian M. Capitini, Department of Pediatrics, University of Wisconsin School of Medicine and Public Health, Madison, WI 53715, USA.
Krishanu Saha: Wisconsin Institute for Discovery, University of Wisconsin-Madison, Madison, WI 53715, USA; Department of Biomedical Engineering, University of Wisconsin-Madison, Madison, WI 53715, USA; University of Wisconsin Carbone Cancer Center, University of Wisconsin-Madison, Madison, WI 53705, USA; Corresponding author: Krishanu Saha, Wisconsin Institute for Discovery, University of Wisconsin-Madison, Madison, WI 53715, USA.

Journal volume & issue: Vol. 32, no. 2
p. 101249

Abstract

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Manufacturing chimeric antigen receptor (CAR) T cell therapies is complex, with limited understanding of how medium composition impacts T cell phenotypes. CRISPR-Cas9 ribonucleoproteins can precisely insert a CAR sequence while disrupting the endogenous T cell receptor alpha constant (TRAC) gene resulting in TRAC-CAR T cells with an enriched stem cell memory T cell population, a process that could be further optimized through modifications to the medium composition. In this study we generated anti-GD2 TRAC-CAR T cells using ''metabolic priming'' (MP), where the cells were activated in glucose/glutamine-low medium and then expanded in glucose/glutamine-high medium. T cell products were evaluated using spectral flow cytometry, metabolic assays, cytokine production, cytotoxicity assays in vitro, and potency against human GD2+ xenograft neuroblastoma models in vivo. Compared with standard TRAC-CAR T cells, MP TRAC-CAR T cells showed less glycolysis, higher CCR7/CD62L expression, more bound NAD(P)H activity, and reduced IFN-γ, IL-2, IP-10, IL-1β, IL-17, and TGF-β production at the end of manufacturing ex vivo, with increased central memory CAR T cells and better persistence observed in vivo. MP with medium during CAR T cell biomanufacturing can minimize glycolysis and enrich memory phenotypes ex vivo, which could lead to better responses against solid tumors in vivo.

Published in Molecular Therapy: Methods & Clinical Development

ISSN: 2329-0501 (Online)
Publisher: Elsevier
Country of publisher: United States
LCC subjects: Science: Biology (General): Genetics; Science: Biology (General): Cytology
Website: http://www.cell.com/molecular-therapy-family/methods/latest-content

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