Scalable Production of Human Mesenchymal STEM/Stromal CELLS in Microcarrier-Based Culture Systems

SCALABLE
PRODUCTION OF HUMAN MESENCHYMAL STEM/STROMAL CELLS IN MICROCARRIER-BASED
CULTURE SYSTEMS

Joana G.
Carmelo, Department of Bioengineering and IBB - Institute for Biotechnology and
Bioengineering, Instituto Superior Técnico, Universidade de Lisboa, Lisboa,
Portugal

Ana
Fernandes-Platzgummer, Department of Bioengineering and IBB - Institute for Biotechnology
and Bioengineering, Instituto Superior Técnico, Universidade de Lisboa, Lisboa,
Portugal

Jennifer
L. Weber, Corning Life Sciences, Corning Incorporated, Corning, NY 14831, USA

Marie-Maud
Bear, Corning SAS, CETC, Avon 77210, France

Martial
Hervy, Corning SAS, CETC, Avon 77210, France

Maria
Margarida Diogo, Department of Bioengineering and IBB - Institute for Biotechnology
and Bioengineering, Instituto Superior Técnico, Universidade de Lisboa, Lisboa,
Portugal

Cláudia
Lobato da Silva, Department of Bioengineering and IBB - Institute for Biotechnology
and Bioengineering, Instituto Superior Técnico, Universidade de Lisboa,
Portugal

Joaquim
M.S. Cabral, Department of Bioengineering and IBB - Institute for Biotechnology
and Bioengineering, Instituto Superior Técnico, Universidade de Lisboa, Lisboa,
Portugal

T: (+351)
218 419 063, joaquim.cabral@ist.utl.pt

The
clinical application of human mesenchymal stem/stromal cells (MSC) for a wide
range of diseases is at the front line of stem cell-based cellular therapies. Nevertheless,
successful implementation of these cell-based therapies requires the ability to
generate large numbers of cells with well-defined characteristics in a
cost-effective way. To this end, significant research has been done on the
development of microcarrier-based cultures in scalable stirred bioreactors
alongside with the development and evaluation of well-defined serum-free and
xeno-free media formulations which represent important milestones towards the
production of MSC for cellular therapies.

In this work, a
microcarrier-based suspension culture was explored for the scale-up of MSCexpansion
in xeno-free medium using synthetic peptide acrylate surface beads. Cells were
maintained on Corning® Synthemax® II polystyrene and CELLstart?-coated Solohill
plastic microcarriers [1] for 14 days in xeno-free medium. Bone marrow (BM)
derived- and adipose tissue (AT) derived-MSC were seeded at 50,000 cells/mL
with 4.5 cm²/mL of microcarriers in spinner flasks (80 mL volume). 
To maximize cell seeding, the adhesion step was performed in 50% final volume
during the first 24 hours. The efficiency of initial cell adhesion to microcarriers
was similar for both cell types; slightly better cell adhesion was observed on
Synthemax II compared to CELLstart-coated microcarriers: 48% and 43% for AT MSC
and 42% to 37% for BM MSC. The homogenous cell distribution on the first days
of culture resulted in a higher expansion rate with exponential growth from day
4 to day 6 for AT MSC and day 4 to day 9 for BM MSC.  The longer growth phase
observed for BM MSC resulted in higher cell densities of 350,000 cells/mL compared
to 250,000 cell/mL for AT MSC cultures. Expanded cells maintained their
characteristic immunophenotype and multilineage differentiation potential. Ongoing
work includes the translation of this culture system to a fully controlled
stirred-tank bioreactor (1 L) and the study of the impact of different culture
parameters namely feeding regime and dissolved oxygen concentration on cell
productivity. This scalable xeno-free, microcarrier-based culture platform is
anticipated to enable cost effective and robust production of MSC meeting the
needs of the allogeneic ?off-the-shelf? MSC therapy sector.

[1] Dos Santos F, Andrade
PZ, Abecasis MM, et al. Toward a clinical-grade expansion of mesenchymal stem cells from
human sources: a microcarrier-based culture system under xeno-free conditions.
Tissue engineering. Part C, Methods. 17(12), 1201?10 (2011).