Exploring continuous and integrated strategies for the up- and downstream processing of human mesenchymal stem cells

Cunha, Bárbara; Aguiar, Tiago; Silva, Marta M.; Silva, Ricardo J.S.; Sousa, Marcos F. Q.; Pineda, Earl; Peixoto, Cristina; Carrondo, Manuel J.T.; Serra, Margarida; Alves, Paula M.

doi:10.1016/j.jbiotec.2015.02.023

Cited by 52 publications

(50 citation statements)

References 26 publications

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“…In the following sections, we describe the operation parameters (Table ) and methodology used (i) for A549 Cell Growth and Onco‐Ad5 Production and (ii) for the expansion of hMSC. It is important to highlight that the selection of the operation parameters for ST bioreactors was based in previous data obtained by our group . For the PBS‐VW, the same operation parameters (e.g., pO 2 , pH, cell inoculum concentration, microcarrier type and concentration, temperature) were used and the agitation rate used was estimated experimentally having as criteria the minimum agitation required for full suspension of the microcarriers ( N S1 ).…”

Section: Methodsmentioning

confidence: 99%

Production of oncolytic adenovirus and human mesenchymal stem cells in a single‐use, Vertical‐Wheel bioreactor system: Impact of bioreactor design on performance of microcarrier‐based cell culture processes

Sousa

Silva

Giroux

et al. 2015

Biotechnology Progress

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Anchorage-dependent cell cultures are used for the production of viruses, viral vectors, and vaccines, as well as for various cell therapies and tissue engineering applications. Most of these applications currently rely on planar technologies for the generation of biological products. However, as new cell therapy product candidates move from clinical trials towards potential commercialization, planar platforms have proven to be inadequate to meet large-scale manufacturing demand. Therefore, a new scalable platform for culturing anchorage-dependent cells at high cell volumetric concentrations is urgently needed. One promising solution is to grow cells on microcarriers suspended in single-use bioreactors. Toward this goal, a novel bioreactor system utilizing an innovative Vertical-Wheel™ technology was evaluated for its potential to support scalable cell culture process development. Two anchorage-dependent human cell types were used: human lung carcinoma cells (A549 cell line) and human bone marrow-derived mesenchymal stem cells (hMSC). Key hydrodynamic parameters such as power input, mixing time, Kolmogorov length scale, and shear stress were estimated. The performance of Vertical-Wheel bioreactors (PBS-VW) was then evaluated for A549 cell growth and oncolytic adenovirus type 5 production as well as for hMSC expansion. Regarding the first cell model, higher cell growth and number of infectious viruses per cell were achieved when compared with stirred tank (ST) bioreactors. For the hMSC model, although higher percentages of proliferative cells could be reached in the PBS-VW compared with ST bioreactors, no significant differences in the cell volumetric concentration and expansion factor were observed. Noteworthy, the hMSC population generated in the PBS-VW showed a significantly lower percentage of apoptotic cells as well as reduced levels of HLA-DR positive cells. Overall, these results showed that process transfer from ST bioreactor to PBS-VW, and scale-up was successfully carried out for two different microcarrier-based cell cultures. Ultimately, the data herein generated demonstrate the potential of Vertical-Wheel bioreactors as a new scalable biomanufacturing platform for microcarrier-based cell cultures of complex biopharmaceuticals.

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Section: Methodsmentioning

confidence: 99%

Production of oncolytic adenovirus and human mesenchymal stem cells in a single‐use, Vertical‐Wheel bioreactor system: Impact of bioreactor design on performance of microcarrier‐based cell culture processes

Sousa

Silva

Giroux

et al. 2015

Biotechnology Progress

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“…Downstream operations are becoming of significant importance for stem cell bioprocessing, and initial theoretical discussions on separation techniques possibly useful for the cell therapy field as reviewed by Diogo et al [63] are becoming a reality. There is a growing rate of studies on bioprocesses and methods required for the clarification and volume reduction of MSC suspensions using membranes and tangential [64,65] or dead-end [66] filtration. Established techniques in other biotechnological fields are also successfully adopted such as the use of expanded bed chromatography for the washing of MSC suspension resulted in improved efficiency [67].…”

Section: Downstream Bio-processes and Process Integrationmentioning

confidence: 99%

Bioprocess engineering strategies for autologous human MSC-based therapies: one size does not fit all

Papantoniou¹,

Lambrechts²,

Aerts³

2017

Cell Gene Therapy Insights

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“…The cultivation of UCM-hMSCs was recently demonstrated using gelatin-based microcarriers and XFM in a controlled STR culture with a working volume of 800 mL [41]. To increase process efficiency, an automated continuous process (e.g., without intervention for media replacement) was developed based on continuous perfusion in a STR and the cells retained their differentiation capacity [95]. Starting with 0.25 × 10 5 cells/mL in 400 mL at 40-60 rpm in Mesencult™-XF media, continuous perfusion was carried out including a cell retention device, such as the ATF-System for microcarriers by Repligen Corporation, with dilution rates of 0.2 d .…”

Section: Stirred Tank Reactormentioning

confidence: 99%

“…The best trade-off between cell recovery (89%) and protein clearance (67%) was achieved using an intermediate expansion bed rate (1.4) which also retained the cell characteristics. A further diafiltration step can be introduced using a CPA solution for formulation, fill and finish [95]. This reduces the overall diafiltration volumes and achieves a product purity sufficient for clinical applications.…”

Section: Separation Clarification and Concentrationmentioning

confidence: 99%

The Challenge of Human Mesenchymal Stromal Cell Expansion: Current and Prospective Answers

Elseberg¹,

Leber²,

Weidner³

et al. 2017

New Insights Into Cell Culture Technology

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In the field of cell therapy, allogenic human mesenchymal stromal cells (hMSCs) are often used in clinical trials, creating a demand for cell mass production using efficient dynamic bioreactor systems. As an advanced therapy medicinal product (ATMP), such cells should meet certain special requirements, including product specifications requiring a production process compatible with good manufacturing practice (GMP). The development of processes in which the cells are the product therefore remains a significant challenge. This chapter describes the requirements at different steps in the upstream and downstream phases of such dynamic processes. Potential solutions are presented and future prospects are discussed, including the selection of media and carriers for the strictly adherent growing cells, allowing efficient cell adhesion and detachment. Strategies for dynamic cultivation in bioreactors are described in detail for fixed-bed and stirred-tank reactors based on GMP requirements and the integration of process analytical technology (PAT). Following cell harvest, separation and purification, the formulation and storage of the product are also described. Finally, the chapter covers important cell quality characteristics necessary for the approval of ATMPs.

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Exploring continuous and integrated strategies for the up- and downstream processing of human mesenchymal stem cells

Cited by 52 publications

References 26 publications

Production of oncolytic adenovirus and human mesenchymal stem cells in a single‐use, Vertical‐Wheel bioreactor system: Impact of bioreactor design on performance of microcarrier‐based cell culture processes

Production of oncolytic adenovirus and human mesenchymal stem cells in a single‐use, Vertical‐Wheel bioreactor system: Impact of bioreactor design on performance of microcarrier‐based cell culture processes

Bioprocess engineering strategies for autologous human MSC-based therapies: one size does not fit all

The Challenge of Human Mesenchymal Stromal Cell Expansion: Current and Prospective Answers

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