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Optimisation of a potency assay for the assessment of immunomodulative potential of clinical grade multipotent mesenchymal stromal cells.

ABSTRACT: Clinical use of multipotent Mesenchymal Stromal Cell (MSC)-based medicinal products requires their production in compliance with Good Manufacturing Practices, thus ensuring that the final drug product meets specifications consistently from batch to batch in terms of cell viability, identity, purity and potency. Potency relates to the efficacy of the medicine in its target clinical indication, so adequate release tests need to be defined and validated as quality controls. Herein we report the design and optimisation of parameters affecting the performance of an in vitro cell-based assay for assessing immunomodulatory potential of clinical grade MSC for human use, based on their capacity to inhibit proliferation of T lymphocytes under strong polyclonal stimuli. The resulting method was demonstrated to be reproducible and relatively simple to execute. Two case studies using clinical grade MSC are presented as examples to illustrate the applicability of the methodology described in this work.

SUBMITTER: Oliver-Vila I

PROVIDER: S-EPMC5809679 | biostudies-literature | 2018 Feb

REPOSITORIES: biostudies-literature

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Optimisation of a potency assay for the assessment of immunomodulative potential of clinical grade multipotent mesenchymal stromal cells.

Oliver-Vila Irene I Ramírez-Moncayo Carmen C Grau-Vorster Marta M Marín-Gallén Sílvia S Caminal Marta M Vives Joaquim J

Cytotechnology 20180110 1

Clinical use of multipotent Mesenchymal Stromal Cell (MSC)-based medicinal products requires their production in compliance with Good Manufacturing Practices, thus ensuring that the final drug product meets specifications consistently from batch to batch in terms of cell viability, identity, purity and potency. Potency relates to the efficacy of the medicine in its target clinical indication, so adequate release tests need to be defined and validated as quality controls. Herein we report the des ...[more]

PMID: 29322348

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Project description:We have previously identified and purified multipotent mesenchymal stromal cell (MSC)-like cells in the highly regenerative endometrial lining of the human uterus (eMSC) as CD140b?CD146? cells. Due to ease of accessibility with minimal morbidity via biopsy, we are proposing to use eMSC in cell-based therapies; however, culture conditions compliant with Good Manufacturing Practice have not been established for eMSC. The aim of this study was to optimize serum-free and xeno-free culture conditions for expansion of eMSC for potential clinical use. Real-time cell assessment (Xcelligence) and MTS viability assays were used to measure attachment and proliferation of freshly isolated, flow cytometry-sorted CD140b?CD146? eMSC cultured in several commercially available and in-house serum-free and xeno-free media in combination with five attachment matrices (fibronectin, collagen, gelatin, laminin, and Cell Start-XF®). Comparisons were made with a standard serum-containing medium, DMEM/F-12/10% fetal bovine serum. Under all conditions examined, eMSC attachment and proliferation was greatest using a fibronectin matrix, with Lonza TP-SF® and our in-house DMEM/SF/FGF2/EGF serum-free xeno-product-containing medium similar to serum-containing medium. Hypoxia increased eMSC proliferation in the DMEM/SF/FGF2/EGF serum-free medium. Culture of eMSC for 7 days on a fibronectin matrix in DMEM/SF/FGF2/EGF serum-free media in 5% O? maintained greater numbers of undifferentiated eMSC expressing CD140b, CD146, and W5C5 compared to culture under similar conditions in Lonza TP-SF medium. However, the percentage of cells expressing typical MSC phenotypic markers, CD29, CD44, CD73, and CD105, were similar for both media. EMSC showed greater expansion in 2D compared to 3D culture on fibronectin-coated microbeads using the optimized DMEM/SF/FGF2/EGF medium in 5% O?. In the optimized 2D culture conditions, eMSC retained CFU activity, multipotency, and MSC surface phenotype, representing the first steps in their preparation for potential clinical use.

Dataset Information

Optimisation of a potency assay for the assessment of immunomodulative potential of clinical grade multipotent mesenchymal stromal cells.

Publications

Optimisation of a potency assay for the assessment of immunomodulative potential of clinical grade multipotent mesenchymal stromal cells.

Similar Datasets

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