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

Cappabianca, D.; Pham, D.; Forsberg, M. H.; Bugel, M.; Tommasi, A.; Lauer, A.; Vidugiriene, J.; Hrdlicka, B.; McHale, A.; Sodji, Q.; Skala, M. C.; Capitini, C. M.; Saha, K.

doi:10.1101/2024.01.31.575774

bioRxiv · 10.1101/2024.01.31.575774

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

Cappabianca, D.·Pham, D.·Forsberg, M. H.·Bugel, M.·Tommasi, A.·Lauer, A.·Vidugiriene, J.·Hrdlicka, B.·McHale, A.·Sodji, Q.·Skala, M. C.·Capitini, C. M.·Saha, K.

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Abstract

Manufacturing Chimeric Antigen Receptor (CAR) T cell therapies is complex, with limited understanding of how media composition impact 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 media 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 media and then expanded in glucose/glutamine high media. 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 to 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-{gamma}, IL-2, IP-10, IL-1{beta}, IL-17, and TGF{beta} production at the end of manufacturing ex vivo, with increased central memory CAR T cells and better persistence observed in vivo. Metabolic priming with media 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.

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Cappabianca, D., Pham, D., Forsberg, M. H., Bugel, M., Tommasi, A., Lauer, A., Vidugiriene, J., Hrdlicka, B., McHale, A., Sodji, Q., Skala, M. C., Capitini, C. M., Saha, K.. 2024-02-02. Metabolic priming of GD2 TRAC-CAR T cellsduring manufacturing promotes memory phenotypes while enhancing persistence. https://doi.org/10.1101/2024.01.31.575774

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