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Hum Gene Ther. 2016 Oct;27(10):860-869. doi: 10.1089/hum.2016.091.

Automated Enrichment, Transduction, and Expansion of Clinical-Scale CD62L+ T Cells for Manufacturing of Gene Therapy Medicinal Products.

Author information

1
1 Cellular Therapy Center, Institute of Cellular Therapeutics , Hannover Medical School, Hannover, Germany.
2
2 GMP Development Unit, Institute of Cellular Therapeutics , Integrated Research and Treatment Center for Transplantation, Hannover Medical School, Hannover, Germany.
3
3 Miltenyi Biotec, Bergisch Gladbach, Germany .
4
4 Institute of Transfusion Medicine , Hannover Medical School, Hannover, Germany.

Abstract

Multiple clinical studies have demonstrated that adaptive immunotherapy using redirected T cells against advanced cancer has led to promising results with improved patient survival. The continuously increasing interest in those advanced gene therapy medicinal products (GTMPs) leads to a manufacturing challenge regarding automation, process robustness, and cell storage. Therefore, this study addresses the proof of principle in clinical-scale selection, stimulation, transduction, and expansion of T cells using the automated closed CliniMACS® Prodigy system. Naïve and central memory T cells from apheresis products were first immunomagnetically enriched using anti-CD62L magnetic beads and further processed freshly (n = 3) or split for cryopreservation and processed after thawing (n = 1). Starting with 0.5 × 108 purified CD3+ T cells, three mock runs and one run including transduction with green fluorescent protein (GFP)-containing vector resulted in a median final cell product of 16 × 108 T cells (32-fold expansion) up to harvesting after 2 weeks. Expression of CD62L was downregulated on T cells after thawing, which led to the decision to purify CD62L+CD3+ T cells freshly with cryopreservation thereafter. Most important in the split product, a very similar expansion curve was reached comparing the overall freshly CD62L selected cells with those after thawing, which could be demonstrated in the T cell subpopulations as well by showing a nearly identical conversion of the CD4/CD8 ratio. In the GFP run, the transduction efficacy was 83%. In-process control also demonstrated sufficient glucose levels during automated feeding and medium removal. The robustness of the process and the constant quality of the final product in a closed and automated system give rise to improve harmonized manufacturing protocols for engineered T cells in future gene therapy studies.

PMID:
27562135
PMCID:
PMC5035932
DOI:
10.1089/hum.2016.091
[Indexed for MEDLINE]
Free PMC Article

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