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PLoS One. 2013 May 31;8(5):e64138. doi: 10.1371/journal.pone.0064138. Print 2013.

Manufacture of clinical-grade CD19-specific T cells stably expressing chimeric antigen receptor using Sleeping Beauty system and artificial antigen presenting cells.

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  • 1Division of Pediatrics, Children's Cancer Hospital, The University of Texas MD Anderson Cancer Center, Houston, Texas, United States of America.

Abstract

Adoptive transfer of T cells expressing a CD19-specific chimeric antigen receptor (CAR) is being evaluated in multiple clinical trials. Our current approach to adoptive immunotherapy is based on a second generation CAR (designated CD19RCD28) that signals through a CD28 and CD3-ζ endodomain. T cells are electroporated with DNA plasmids from the Sleeping Beauty (SB) transposon/transposase system to express this CAR. Stable integrants of genetically modified T cells can then be retrieved when co-cultured with designer artificial antigen presenting cells (aAPC) in the presence of interleukin (IL)-2 and 21. Here, we reveal how the platform technologies of SB-mediated transposition and CAR-dependent propagation on aAPC were adapted for human application. Indeed, we have initiated clinical trials in patients with high-risk B-lineage malignancies undergoing autologous and allogeneic hematopoietic stem-cell transplantation (HSCT). We describe the process to manufacture clinical grade CD19-specific T cells derived from healthy donors. Three validation runs were completed in compliance with current good manufacturing practice for Phase I/II trials demonstrating that by 28 days of co-culture on γ-irradiated aAPC ∼10(10) T cells were produced of which >95% expressed CAR. These genetically modified and propagated T cells met all quality control testing and release criteria in support of infusion.

PMID:
23741305
[PubMed - indexed for MEDLINE]
PMCID:
PMC3669363
Free PMC Article

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