Manufacturing Mesenchymal Stromal Cells for the Treatment of Graft-versus-Host Disease: A Survey among Centers Affiliated with the European Society for Blood and Marrow Transplantation

Abstract

The immunosuppressive properties of mesenchymal stromal cells (MSC) have been successfully tested to control clinical severe graft-versus host disease and improve survival. However, clinical studies have not yet provided conclusive evidence of their efficacy largely because of lack of patients' stratification criteria. The heterogeneity of MSC preparations is also a major contributing factor, as manufacturing of therapeutic MSC is performed according to different protocols among different centers. Understanding the variability of the manufacturing protocol would allow a better comparison of the results obtained in the clinical setting among different centers. In order to acquire information on MSC manufacturing we sent a questionnaire to the European Society for Blood and Marrow Transplantation centers registered as producing MSC. Data from 17 centers were obtained and analyzed by means of a 2-phase questionnaire specifically focused on product manufacturing. Gathered information included MSC tissue sources, MSC donor matching, medium additives for ex vivo expansion, and data on MSC product specification for clinical release. The majority of centers manufactured MSC from bone marrow (88%), whilst only 2 centers produced MSC from umbilical cord blood or cord tissue. One of the major changes in the manufacturing process has been the replacement of fetal bovine serum with human platelet lysate as medium supplement. 59% of centers used only third-party MSC, whilst only 1 center manufactured exclusively autologous MSC. The large majority of these facilities (71%) administered MSC exclusively from frozen batches. Aside from variations in the culture method, we found large heterogeneity also regarding product specification, particularly in the markers used for phenotypical characterization and their threshold of expression, use of potency assays to test MSC functionality, and karyotyping. The initial data collected from this survey highlight the variability in MSC manufacturing as clinical products and the need for harmonization. Until more informative potency assays become available, a more homogeneous approach to cell production may at least reduce variability in clinical trials and improve interpretation of results.

Keywords: Cellular therapy; Graft-versus-host disease; Manufacturing; Mesenchymal stromal cells; Product specification; Release criteria.

Figure 1

Clinical MSC production in 17 Good Manufacturing Practice facilities in EBMT centers. (A) Percentage of centers producing MSC from umbilical cord blood or cord tissue, or from BM, either using Ficoll method or adherence from whole BM. (B) Percentage of centers using FBS or hPL from commercially available manufacturers or from pooled expired platelets obtained from blood banks. (C) Distribution of centers using allogeneic or autologous MSC. (D) Percentage of centers delivering fresh or frozen MSC.

Figure 2

Distribution charts on product specification for phenotype on clinical MSC. (A) Distribution chart on the percentage of centers using different acceptance criteria for CD45 expression. (B) Distribution chart on the percentage of centers using different acceptance criteria for expression of markers CD73, CD90, and CD105. (C) Distribution graph on the percentage of centers using other markers.

Figure 3

Distribution chart on release criteria for clinical MSC. (A). Percentage of centers using sterility and absence of mycoplasma species or sterility, absence of mycoplasma species, and endotoxin levels as release criteria for clinical MSCs. (B) Percentage of centers performing analysis of karyotype. (C) Percentage of centers analyzing differentiation ability of MSC: tri-lineage differentiation into adipocytes, osteoblasts, and chondrocytes; two-lineage differentiation into adipocytes and osteoblasts; no differentiation assay carried out. (D) Percentage of centers determining immunosuppressive activity of MSC (potency assay).