the production of cell therapeutics, as, among other things, cell material (in an
autologous approach) may vary between batches. Process conditions must, therefore, be adapted to the biological starting material, increasing the complexity of the
production process. Here application of a “Digital Twin,” which combines biochemical engineering data of the cultivation system with a mathematical model of the cell
growth, is beneficial, as it tests different process conditions in silico and subsequently proposes optimal parameter combinations for the hMSC production process.
2 In Vitro Expansion Approaches: Current Situation
For the clinical application of hMSCs, the in vitro expansion of the cells represents
an important step. Although recent studies have shown the difference in cell yield
depending on the hMSC source (e.g., bone marrow vs. adipose tissue), the required
therapeutic dose (1–5 million hMSCs/kg body weight) makes in vitro expansion
mandatory independent on the hMSC-type. Therefore, different systems and cultivation strategies have been developed over the years for the expansion of hMSCs,
which will be presented and discussed in the following sections.
2.1 Planar Approach (2D Cultures)
hMSCs are typically isolated by their capacity to adhere to plastic surfaces. Therefore, the simplest way to expand hMSCs is the usage of plastic vessels, such as
T-flask or stacked plate systems, which allow for the expansion of the cells at
laboratory and pilot plant production scale for early-phase clinical trials [15]. Planar
expansion approaches in normal cell culture flasks (e.g., T-flasks) represent a costefficient and easy-to-operate solution. Maximum cell densities for hMSCs from the
human bone marrow, the adipose tissue, and the umbilical cord have been reported
in the literature in the range of 0.05 to 1.0 Â 10
5 cells/cm
2 (PDL 2.8–7.4) for T-flask
cultures performed with serum-containing and serum-free cell culture medium (see
Table 2). Maximum cell densities for CellSTACK cultures were even reported in the
range of 2.5 to 4.2 Â 10
5 cells/cm
2 (¼1.59-2.67 Â 10
9 cells) using hMSCs from the
bone marrow.
However, scale-up of such an hMSC expansion process would require a large
number of cell culture flasks, which is by any means neither economic nor ecologic.
Moreover, handling of multiple flasks in parallel is very labor and cost intensive
(increased facility footprint) and may result in high flask-to-flask variabilities. In
addition, the risk of contamination (e.g., bacteria, mycoplasms) is increased due to
the large number of open manipulations. Alternatives to the normal cell culture
flasks are stacked-plate or multi-tray culture systems, such as cell factories, which
significantly increase the efficiency of the cultivation step by using several layers per
cultivation system (up to 40-layer systems available). Thus, the absolute cell number
Numerical Methods for the Design and Description of In Vitro Expansion. . .
191
autologous approach) may vary between batches. Process conditions must, therefore, be adapted to the biological starting material, increasing the complexity of the
production process. Here application of a “Digital Twin,” which combines biochemical engineering data of the cultivation system with a mathematical model of the cell
growth, is beneficial, as it tests different process conditions in silico and subsequently proposes optimal parameter combinations for the hMSC production process.
2 In Vitro Expansion Approaches: Current Situation
For the clinical application of hMSCs, the in vitro expansion of the cells represents
an important step. Although recent studies have shown the difference in cell yield
depending on the hMSC source (e.g., bone marrow vs. adipose tissue), the required
therapeutic dose (1–5 million hMSCs/kg body weight) makes in vitro expansion
mandatory independent on the hMSC-type. Therefore, different systems and cultivation strategies have been developed over the years for the expansion of hMSCs,
which will be presented and discussed in the following sections.
2.1 Planar Approach (2D Cultures)
hMSCs are typically isolated by their capacity to adhere to plastic surfaces. Therefore, the simplest way to expand hMSCs is the usage of plastic vessels, such as
T-flask or stacked plate systems, which allow for the expansion of the cells at
laboratory and pilot plant production scale for early-phase clinical trials [15]. Planar
expansion approaches in normal cell culture flasks (e.g., T-flasks) represent a costefficient and easy-to-operate solution. Maximum cell densities for hMSCs from the
human bone marrow, the adipose tissue, and the umbilical cord have been reported
in the literature in the range of 0.05 to 1.0 Â 10
5 cells/cm
2 (PDL 2.8–7.4) for T-flask
cultures performed with serum-containing and serum-free cell culture medium (see
Table 2). Maximum cell densities for CellSTACK cultures were even reported in the
range of 2.5 to 4.2 Â 10
5 cells/cm
2 (¼1.59-2.67 Â 10
9 cells) using hMSCs from the
bone marrow.
However, scale-up of such an hMSC expansion process would require a large
number of cell culture flasks, which is by any means neither economic nor ecologic.
Moreover, handling of multiple flasks in parallel is very labor and cost intensive
(increased facility footprint) and may result in high flask-to-flask variabilities. In
addition, the risk of contamination (e.g., bacteria, mycoplasms) is increased due to
the large number of open manipulations. Alternatives to the normal cell culture
flasks are stacked-plate or multi-tray culture systems, such as cell factories, which
significantly increase the efficiency of the cultivation step by using several layers per
cultivation system (up to 40-layer systems available). Thus, the absolute cell number
Numerical Methods for the Design and Description of In Vitro Expansion. . .
191