Systematic microcarrier screening and agitated culture conditions improves human mesenchymal stem cell yield in bioreactors

Abstract

Production of human mesenchymal stem cells for allogeneic cell therapies requires scalable, cost‐effective manufacturing processes. Microcarriers enable the culture of anchorage‐dependent cells in stirred‐tank bioreactors. However, no robust, transferable methodology for microcarrier selection exists, with studies providing little or no reason explaining why a microcarrier was employed. We systematically evaluated 13 microcarriers for human bone marrow‐derived MSC (hBM‐MSCs) expansion from three donors to establish a reproducible and transferable methodology for microcarrier selection. Monolayer studies demonstrated input cell line variability with respect to growth kinetics and metabolite flux. HBM‐MSC1 underwent more cumulative population doublings over three passages in comparison to hBM‐MSC2 and hBM‐MSC3. In 100 mL spinner flasks, agitated conditions were significantly better than static conditions, irrespective of donor, and relative microcarrier performance was identical where the same microcarriers outperformed others with respect to growth kinetics and metabolite flux. Relative growth kinetics between donor cells on the microcarriers were the same as the monolayer study. Plastic microcarriers were selected as the optimal microcarrier for hBM‐MSC expansion. HBM‐MSCs were successfully harvested and characterised, demonstrating hBM‐MSC immunophenotype and differentiation capacity. This approach provides a systematic method for microcarrier selection, and the findings identify potentially significant bioprocessing implications for microcarrier‐based allogeneic cell therapy manufacture.

Process development of human multipotent stromal cell microcarrier culture using an automated high‐throughput microbioreactor

“…A 100 mL BellCo spinner flasks (Bellco Glass Inc., Vineland, NJ), were used for all spinner flask experiments, with a 100 mL working volume and a vessel diameter (T) of 60 mM. For all microcarrier cultures, Plastic P102‐L microcarriers (Pall Life Sciences, Portsmouth, UK) were employed (Rafiq et al, ). The same inoculation concentrations and culture methods were employed as described in Rafiq et al () except where improvements to the process were made (discussed in the Results section).…”

Microfluidic‐Printed Microcarrier for In Vitro Expansion of Adherent Stem Cells in 3D Culture Platform

“…A recent analysis of the experimental processes, cost efficiency, and robustness of adherent cell culture technologies suggested that multi‐layered 2D culture vessels and stirred tank bioreactors with microcarriers could potentially replace conventional 2D culture platforms . Several thorough comparative investigations also indicated that microcarrier‐based 3D cultures provide the most efficient method for the in vitro expansion culturing of adherent cells, not only due to the volume economy arising from the high surface/volume ratio, but also due to the effectiveness of handling large numbers of cultured cells . These considerations elevated plastic microcarriers as the optimal choice for the scale‐up culture of postnatal stem cells, such as hMSCs.…”

Cell and gene therapy manufacturing: the necessity for a cost-based development approach

“…However for many clinical indications, achieving lot sizes of several hundred billion to trillions of cells will be imperative for commercial success; utilizing high-density bioreactor manufacturing platforms, such as stirred-tank bioreactors in conjunction with microcarriers, is likely to be the only way to accomplish this [21][22][23][24][25].…”