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.
Highlights
IntroductionSIGNIFICANT PROGRESS HAS BEEN MADE in the last decade by using redirected immune response against advanced cancer, especially for leukemia.[1,2] Clinical Phase I/II studies have shown the successful treatment with autologous T cells redirected against CD19 leukemic cells by using recombinant chimeric antigen receptors (CARs), which consist of a single-chain variable fragment (scFv) linked to intracellular signaling domains.Long-term remission in those patients with B cell malignancies has been observed.[2,3] In addition, a broad range of different cancer target antigens under clinical investigation using CAR-expressing T cells redirected against CD20, CD30, CD138, c-Met, EGFRvIII, ErbB2, FAB, GD2, HER2, WT1, PSMA, NY-ESO1, and others have been reviewed.[4,5,6] In a similar way, both autologous and allogeneic natural killer (NK) effector cells,[7] as well as the NK cell lineNK-92, can be redirected against cancer
Clinical Phase I/II studies have shown the successful treatment with autologous T cells redirected against CD19 leukemic cells by using recombinant chimeric antigen receptors (CARs), which consist of a single-chain variable fragment linked to intracellular signaling domains
These experiments led to the decision to purify naıve and central memory T cells from fresh apheresis products and consequent cryopreservation
Summary
SIGNIFICANT PROGRESS HAS BEEN MADE in the last decade by using redirected immune response against advanced cancer, especially for leukemia.[1,2] Clinical Phase I/II studies have shown the successful treatment with autologous T cells redirected against CD19 leukemic cells by using recombinant chimeric antigen receptors (CARs), which consist of a single-chain variable fragment (scFv) linked to intracellular signaling domains.Long-term remission in those patients with B cell malignancies has been observed.[2,3] In addition, a broad range of different cancer target antigens under clinical investigation using CAR-expressing T cells redirected against CD20, CD30, CD138, c-Met, EGFRvIII, ErbB2, FAB, GD2, HER2, WT1, PSMA, NY-ESO1, and others have been reviewed.[4,5,6] In a similar way, both autologous and allogeneic natural killer (NK) effector cells,[7] as well as the NK cell lineNK-92, can be redirected against cancer. SIGNIFICANT PROGRESS HAS BEEN MADE in the last decade by using redirected immune response against advanced cancer, especially for leukemia.[1,2] Clinical Phase I/II studies have shown the successful treatment with autologous T cells redirected against CD19 leukemic cells by using recombinant chimeric antigen receptors (CARs), which consist of a single-chain variable fragment (scFv) linked to intracellular signaling domains. Long-term remission in those patients with B cell malignancies has been observed.[2,3] In addition, a broad range of different cancer target antigens under clinical investigation using CAR-expressing T cells redirected against CD20, CD30, CD138, c-Met, EGFRvIII, ErbB2, FAB, GD2, HER2, WT1, PSMA, NY-ESO1, and others have been reviewed.[4,5,6] In a similar way, both autologous and allogeneic natural killer (NK) effector cells,[7] as well as the NK cell line.
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