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Good Manufacturing Practice-Compliant Production and Lot-Release of Ex Vivo Expanded Regulatory T Cells As Basis for Treatment of Patients with Autoimmune and Inflammatory Disorders.

ABSTRACT: In recent years, the exploration of regulatory T cell (Treg)-based cellular therapy has become an attractive strategy to ameliorate inflammation and autoimmunity in various clinical settings. The main obstacle to the clinical application of Treg in human is their low number circulating in peripheral blood. Therefore, ex vivo expansion is inevitable. Moreover, isolation of Treg bears the risk of concurrent isolation of unwanted effector cells, which may trigger or deteriorate inflammation upon adoptive Treg transfer. Here, we present a protocol for the GMP-compliant production, lot-release and validation of ex vivo expanded Tregs for treatment of patients with autoimmune and inflammatory disorders. In the presented production protocol, large numbers of Treg, previously enriched from a leukapheresis product by using the CliniMACS^® system, are ex vivo expanded in the presence of anti-CD3/anti-CD28 expander beads, exogenous IL-2 and rapamycin during 21?days. The expanded Treg drug product passed predefined lot-release criteria. These criteria include (i) sterility testing, (ii) assessment of Treg phenotype, (iii) assessment of non-Treg cellular impurities, (iv) confirmation of successful anti-CD3/anti-CD28 expander bead removal after expansion, and (v) confirmation of the biological function of the Treg product. Furthermore, the Treg drug product was shown to retain its stability and suppressive function for at least 1?year after freezing and thawing. Also, dilution of the Treg drug product in 0.9% physiological saline did not affect Treg phenotype and Treg function for up to 90?min. These data indicate that these cells are ready to use in a clinical setting in which a cell infusion time of up to 90?min can be expected. The presented production process has recently undergone on site GMP-conform evaluation and received GMP certification from the Bavarian authorities in Germany. This protocol can now be used for Treg-based therapy of various inflammatory and autoimmune disorders.

SUBMITTER: Wiesinger M

PROVIDER: S-EPMC5662555 | biostudies-literature | 2017

REPOSITORIES: biostudies-literature

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Good Manufacturing Practice-Compliant Production and Lot-Release of <i>Ex Vivo</i> Expanded Regulatory T Cells As Basis for Treatment of Patients with Autoimmune and Inflammatory Disorders.

Wiesinger Manuel M Stoica Diane D Roessner Susanne S Lorenz Carmen C Fischer Anika A Atreya Raja R Neufert Clemens F CF Atreya Imke I Scheffold Alexander A Schuler-Thurner Beatrice B Neurath Markus F MF Schuler Gerold G Voskens Caroline J CJ

Frontiers in immunology 20171026

In recent years, the exploration of regulatory T cell (Treg)-based cellular therapy has become an attractive strategy to ameliorate inflammation and autoimmunity in various clinical settings. The main obstacle to the clinical application of Treg in human is their low number circulating in peripheral blood. Therefore, <i>ex vivo</i> expansion is inevitable. Moreover, isolation of Treg bears the risk of concurrent isolation of unwanted effector cells, which may trigger or deteriorate inflammation ...[more]

PMID: 29123521

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Project description:BackgroundClinical translation of mesenchymal stromal cells (MSCs) necessitates basic characterization of the cell product since variability in biological source and processing of MSCs may impact therapeutic outcomes. Although expression of classical cell surface markers (e.g., CD90, CD73, CD105, and CD44) is used to define MSCs, identification of functionally relevant cell surface markers would provide more robust release criteria and options for quality control. In addition, cell surface expression may distinguish between MSCs from different sources, including bone marrow-derived MSCs and clinical-grade adipose-derived MSCs (AMSCs) grown in human platelet lysate (hPL).MethodsIn this work we utilized quantitative PCR, flow cytometry, and RNA-sequencing to characterize AMSCs grown in hPL and validated non-classical markers in 15 clinical-grade donors.ResultsWe characterized the surface marker transcriptome of AMSCs, validated the expression of classical markers, and identified nine non-classical markers (i.e., CD36, CD163, CD271, CD200, CD273, CD274, CD146, CD248, and CD140B) that may potentially discriminate AMSCs from other cell types. More importantly, these markers exhibit variability in cell surface expression among different cell isolates from a diverse cohort of donors, including freshly prepared, previously frozen, or proliferative state AMSCs and may be informative when manufacturing cells.ConclusionsOur study establishes that clinical-grade AMSCs expanded in hPL represent a homogeneous cell culture population according to classical markers,. Additionally, we validated new biomarkers for further AMSC characterization that may provide novel information guiding the development of new release criteria.Clinical trialsUse of Autologous Bone Marrow Aspirate Concentrate in Painful Knee Osteoarthritis (BMAC): Clinicaltrials.gov NCT01931007 . Registered August 26, 2013. MSC for Occlusive Disease of the Kidney: Clinicaltrials.gov NCT01840540 . Registered April 23, 2013. Mesenchymal Stem Cell Therapy in Multiple System Atrophy: Clinicaltrials.gov NCT02315027 . Registered October 31, 2014. Efficacy and Safety of Adult Human Mesenchymal Stem Cells to Treat Steroid Refractory Acute Graft Versus Host Disease. Clinicaltrials.gov NCT00366145 . Registered August 17, 2006. A Dose-escalation Safety Trial for Intrathecal Autologous Mesenchymal Stem Cell Therapy in Amyotrophic Lateral Sclerosis. Clinicaltrials.gov NCT01609283 . Registered May 18, 2012.

Dataset Information

Good Manufacturing Practice-Compliant Production and Lot-Release of Ex Vivo Expanded Regulatory T Cells As Basis for Treatment of Patients with Autoimmune and Inflammatory Disorders.

Publications

Good Manufacturing Practice-Compliant Production and Lot-Release of <i>Ex Vivo</i> Expanded Regulatory T Cells As Basis for Treatment of Patients with Autoimmune and Inflammatory Disorders.

Similar Datasets

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