Development of a process control strategy for the serum-free microcarrier expansion of human mesenchymal stem cells towards cost-effective and commercially viable manufacturing

Abstract

Highlights Superior control in the bioreactor led to improved hMSC quality, consistency and yield  At 25% dissolved oxygen in serum-free media, there was a 500 % increase in the hMSC yield  Process control significantly reduced the variation in hMSC yield to < 15% AbstractHuman Mesenchymal Stem Cells (hMSCs) are advancing through clinical development with the first allogeneic adult hMSC therapy receiving approval in Europe. To enable successful large-scale manufacture of hMSC therapies, increased product consistency and yield, and a reduced batch-tobatch variation must be achieved. This paper addresses ways to reduce variation by controlling the processing conditions, in particular the dissolved oxygen concentration (dO2), and the culture medium.Bone marrow derived hMSCs were cultured in DASGIP DASbox bioreactors on Plastic P-102L microcarriers in FBS-containing and serum free (SFM) media at various dO2 values from 100% to 10%, experiencing the same dO2 value throughout the culture process. The superior control of pH and dO2 in the bioreactor led to improved performances compared to poorly controlled spinner flasks, particularly at reduced dO2 concentrations. At 25% dO2, there was a 300 % increase in the BM-hMSC yield in the bioreactor across the two donor BM-hMSCs in SFM compared to FBS-containing medium.Overall, the process yield increased by an average of around 500% for both donors under controlled conditions in SFM at 25% dO2 in the bioreactor compared to the poorly controlled expansion at atmospheric conditions in FBS-containing medium in spinner flasks. Process control significantly reduced the BM-hMSC variation in yield from 79.1% in FBS-containing medium in spinner flasks to < 15% in controlled SFM bioreactor culture.3 KeywordsProcess control, serum-free, human mesenchymal stem cell, microcarrier expansion, harvest, dissolved oxygen

An Approach towards a GMP Compliant In-Vitro Expansion of Human Adipose Stem Cells for Autologous Therapies

“…The hASCs from the two different donors grew at comparable specific growth rates of between 0.42 and 0.44 d −1 (td = 37.8-39.6 h). The calculated specific growth rates were in a comparable range to literature data for MC-based expansions of hBM-MSCs in serum-free cell culture media [20][21][22]39]. In both cases, glucose was metabolized less efficiently by the hASCs, although -qGlc values (1.34-1.96 pmol/cell/d) were comparable to those in the 2D cultures.…”

Bioprocessing of Human Mesenchymal Stem Cells: From Planar Culture to Microcarrier-Based Bioreactors

“…There are two ways to regulate the temperature in bioreactor culture. One is to maintain the temperature of the water jacket of the bioreactor [40,71] or the metal plate attached to the bioreactor bag [44,74] while the other is directly to control the temperature of the bulk culture by measuring the temperature with a thermocouple or a temperature sensor and then to warm the bioreactor with a heating blanket or pad [36,75]. The former may have some deviations due to the indirect measurement while the latter requires more attention to understand the relative locations of the temperature sensor and the heating source.…”

Expansion of human mesenchymal stem/stromal cells (hMSCs) in bioreactors using microcarriers: lessons learnt and what the future holds

“…Several groups have investigated the use of STRs for microcarrier-based hMSC expansion examples, with different bioreactors using different mechanisms to maintain the culture temperature. The Biostat B Plus® (5L) from Sartorius (Rafiq et al, 2013) and Celligen® from New Brunswick (Mizukami et al, 2016) uses a water jacket, the DASbox® uses aIndividual temperature control with liquid-free heating and cooling (Peltier), whilst in the UniVessel ® (Sartorius) (Schirmaier et al, 2014) and Mobius® 3L, 50L (Applikon) (Lawson et al, 2017) heating blankets are used to keep the temperature at 37 °C (Heathman et al, 2019).…”