T cells for suicide gene therapy: Activation, functionality and clinical relevance

Hollatz, Gabriele; Grez, Manuel; Mastaglio, Sara; Quaritsch, R; Huenecke, Sabine; Ciceri, Fabio; Bonini, Chiara; Esser, Ruth; Klingebiel, Thomas; Kreuter, J.; Koehl, Ulrike

doi:10.1016/j.jim.2007.11.007

Cited by 8 publications

(5 citation statements)

References 43 publications

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“…All clinical scale runs resulted in a nearly identical conversion of the CD4/CD8 ratio. This is in line with the results of Hollatz 20 using bead-coupled or soluble antibodies + IL-2. In addition, the viability and expansion rate of the overall CD3 + T cells at harvest was equal between cryopreserved and fresh CD62L-selected T cells.…”

Section: Discussionsupporting

confidence: 91%

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

et al. 2016

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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.

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Section: Discussionsupporting

confidence: 91%

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

et al. 2016

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“…These findings suggest that incorporation of the CD137 intracellular domain mediates antigen-independent activity that is similar to that provided by the natural 4-1BB receptor in T cells following ligation [23]. As a result of the enhanced proliferation observed following the initial activation of T cells via αCD3/αCD28-coated beads used to enhance T-cell transduction [24, 25] (Fig 4b), CAR+ T cells expressing the αCD19-BB-ζ receptor had relatively low CAR-driven proliferation (Fig. 4a[iii]).…”

Section: Resultsmentioning

confidence: 99%

Chimeric Receptors Containing CD137 Signal Transduction Domains Mediate Enhanced Survival of T Cells and Increased Antileukemic Efficacy In Vivo

et al. 2009

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Persistence of T cells engineered with chimeric antigen receptors (CARs) has been a major barrier to use of these cells for molecularly targeted adoptive immunotherapy. To address this issue, we created a series of CARs that contain the TCR-ζ signal transduction domain with the CD28 and/or CD137 (4-1BB) intracellular domains in tandem. After short-term expansion, primary human T cells were subjected to lentiviral gene transfer, resulting in large numbers of cells with >85% CAR expression. In an immunodeficient mouse xenograft model of primary human pre-B-cell acute lymphoblastic leukemia, human T cells expressing anti-CD19 CARs containing CD137 exhibited the greatest anti-leukemic efficacy and prolonged (>6 months) survival in vivo, and were significantly more effective than cells expressing CARs containing TCR-ζ alone or CD28-ζ signaling receptors. We uncovered a previously unrecognized, antigen-independent effect of CARs expressing the CD137 cytoplasmic domain that likely contributes to the enhanced antileukemic efficacy and survival in tumor bearing mice. Furthermore, our studies revealed significant discrepancies between in vitro and in vivo surrogate measures of CAR efficacy. Together these results suggest that incorporation of the CD137 signaling domain in CARs should improve the persistence of CARs in the tumor microenvironment and hence maximize their antitumor activity.

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“…To have adequate numbers of cells for the experiments, T cells were expanded for 7 to 9 days with CD3/CD28 T-cell expander beads (Invitrogen, Carlsbad, CA) in X-vivo 15 medium without phenol red (Lonza, Walkersville, MD) supplemented with 5% FBS (HyClone, Logan, UT), 25 μg/ml gentamicin (Invitrogen) and 100 U/ml IL-2 (PeproTech, Rocky Hill, NJ) as described previously (37). After expansion, the T cells expressed a memory phenotype as a result of stimulation with CD3/CD28 beads and IL-2 (39–42). …”

Section: Methodsmentioning

confidence: 99%

Identification of Radiation-Induced Expression Changes in Nonimmortalized Human T Cells

et al. 2010

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In the event of a radiation accident or attack, it will be imperative to quickly assess the amount of radiation exposure to accurately triage victims for appropriate care. RNA-based radiation dosimetry assays offer the potential to rapidly screen thousands of individuals in an efficient and cost-effective manner. However, prior to the development of these assays, it will be critical to identify those genes that will be most useful to delineate different radiation doses. Using global expression profiling, we examined expression changes in nonimmortalized T cells across a wide range of doses (0.15–12 Gy). Because many radiation responses are highly dependent on time, expression changes were examined at three different times (3, 8, and 24 h). Analyses identified 61, 512 and 1310 genes with significant linear dose-dependent expression changes at 3, 8 and 24 h, respectively. Using a stepwise regression procedure, a model was developed to estimate in vitro radiation exposures using the expression of three genes (CDKN1A, PSRC1 and TNFSF4) and validated in an independent test set with 86% accuracy. These findings suggest that RNA-based expression assays for a small subset of genes can be employed to develop clinical biodosimetry assays to be used in assessments of radiation exposure and toxicity.

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T cells for suicide gene therapy: Activation, functionality and clinical relevance

Cited by 8 publications

References 43 publications

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

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

Chimeric Receptors Containing CD137 Signal Transduction Domains Mediate Enhanced Survival of T Cells and Increased Antileukemic Efficacy In Vivo

Identification of Radiation-Induced Expression Changes in Nonimmortalized Human T Cells

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T cells for suicide gene therapy: Activation, functionality and clinical relevance

Cited by 8 publications

References 43 publications

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

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

Chimeric Receptors Containing CD137 Signal Transduction Domains Mediate Enhanced Survival of T Cells and Increased Antileukemic Efficacy In Vivo

Identification of Radiation-Induced Expression Changes in Nonimmortalized Human T Cells

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

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