Quantitative activation suppression assay to evaluate human bone marrow–derived mesenchymal stromal cell potency

• Published in: Cytotherapy (Oxford, England) Vol. 17; no. 12; pp. 1675 - 1686
• Main Authors: Salem, Bahey, Miner, Samantha, Hensel, Nancy F., Battiwalla, Minoo, Keyvanfar, Keyvan, Stroncek, David F., Gee, Adrian P., Hanley, Patrick J., Bollard, Catherine M., Ito, Sawa, Barrett, A. John
• Format: Journal Article
• Language: English
• Published: England Elsevier Inc 01.12.2015
• Subjects: Advanced Basic Science; Antibodies - immunology; Antibodies - pharmacology; Biological Assay; Bone Marrow Cells - cytology; CD28 Antigens - immunology; CD3 Complex - immunology; CD4-Positive T-Lymphocytes - cytology; CD4-Positive T-Lymphocytes - immunology; CD40 Ligand - metabolism; Cell Line; Cell Proliferation; Cell Survival - immunology; Coculture Techniques; Humans; immunosuppression; Immunosuppression Therapy - methods; Interferon-gamma - pharmacology; K299; Lymphocyte Activation - immunology; Mesenchymal Stem Cells - cytology; mesenchymal stromal cells; Other; suppression potency assay; immunosuppression; K299; suppression potency assay; mesenchymal stromal cells
• ISSN: 1465-3249; 1477-2566; 1477-2566
• DOI: 10.1016/j.jcyt.2015.08.008

With the increasing use of cell therapies involving immune modulatory cells, there is a need for a simple standardized method to evaluate and compare the suppressive potency of different cell products. We used the Karpas 299 (K299) cell line as the reference suppressor cell to develop a standardized suppression assay to quantify the immune-modulatory capacity of bone marrow–derived mesenchymal stromal cells (BM-MSCs). Healthy donor CD4 T cells were co-cultured with the K299 cell line or with third-party BM-MSCs. After stimulation with anti-CD3/CD28 beads, CD154 activation and proliferation of CD4 T cells were measured to calculate suppression. The K299 cell line reproducibly suppressed both the activation and proliferation of healthy donor CD4 T cells in a dose-dependent manner. A rapid (16-h) assay that was based on activation-suppression was selected for development. In replicate testing, there was an inherent variability of suppression of 11% coefficient of variation between different responder T cells. Suppression by BM-MSCs on different responders correlated with suppression by K299. We therefore used K299 suppression as the reference to define suppression potency of BM-MSCs in K299 Suppression Units. We found that inter-donor variability, passage number, method of manufacture and exposure of BM-MSCs to steroids or interferon-γ all affected BM-MSC potency of suppression. This method provides a platform for standardizing suppressor function to facilitate comparisons between laboratories and for use as a cell product release assay.