Bioprocess integration for human mesenchymal stem cells

From up to downstream processing scale-up to cell proteome characterization

Bárbara Cunha, Tiago Aguiar, Sofia B. Carvalho, Marta M. Silva, Ricardo A. Gomes, Manuel J.T. Carrondo, Patrícia Gomes-Alves, Cristina Peixoto, Margarida Serra, Paula M. Alves

Research output: Contribution to journalArticle

10 Citations (Scopus)

Abstract

To deliver the required cell numbers and doses to therapy, scaling-up production and purification processes (at least to the liter-scale) while maintaining cells’ characteristics is compulsory. Therefore, the aim of this work was to prove scalability of an integrated streamlined bioprocess compatible with current good manufacturing practices (cGMP) comprised by cell expansion, harvesting and volume reduction unit operations using human mesenchymal stem cells (hMSC) isolated from bone marrow (BM-MSC) and adipose tissue (AT-MSC). BM-MSC and AT-MSC expansion and harvesting steps were scaled-up from spinner flasks to 2 L scale stirred tank single-use bioreactor using synthetic microcarriers and xeno-free medium, ensuring high cellular volumetric productivities (50 × 106 cell L−1 day−1), expansion factors (14–16 fold) and cell recovery yields (80%). For the concentration step, flat sheet cassettes (FSC) and hollow fiber cartridges (HF) were compared showing a fairly linear scale-up, with a need to slightly decrease the permeate flux (30–50 LMH, respectively) to maximize cell recovery yield. Nonetheless, FSC allowed to recover 18% more cells after a volume reduction factor of 50. Overall, at the end of the entire bioprocess more than 65% of viable (>95%) hMSC could be recovered without compromising cell's critical quality attributes (CQA) of viability, identity and differentiation potential. Alongside the standard quality assays, a proteomics workflow based on mass spectrometry tools was established to characterize the impact of processing on hMSC's CQA; These analytical tools constitute a powerful tool to be used in process design and development.

Original languageEnglish
Pages (from-to)87-98
Number of pages12
JournalJournal of Biotechnology
Volume248
DOIs
Publication statusPublished - 20 Apr 2017

Fingerprint

Proteome
Stem cells
Mesenchymal Stromal Cells
Proteins
Processing
Recovery
Bioreactors
Purification
Mass spectrometry
Scalability
Assays
Process design
Bone
Productivity
Tissue
Fluxes
Workflow
Fibers
Proteomics
Adipose Tissue

Keywords

  • Cell therapy
  • Human mesenchymal stem cells
  • Mass spectrometry
  • Process development
  • Product characterization
  • Scale-up

Cite this

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title = "Bioprocess integration for human mesenchymal stem cells: From up to downstream processing scale-up to cell proteome characterization",
abstract = "To deliver the required cell numbers and doses to therapy, scaling-up production and purification processes (at least to the liter-scale) while maintaining cells’ characteristics is compulsory. Therefore, the aim of this work was to prove scalability of an integrated streamlined bioprocess compatible with current good manufacturing practices (cGMP) comprised by cell expansion, harvesting and volume reduction unit operations using human mesenchymal stem cells (hMSC) isolated from bone marrow (BM-MSC) and adipose tissue (AT-MSC). BM-MSC and AT-MSC expansion and harvesting steps were scaled-up from spinner flasks to 2 L scale stirred tank single-use bioreactor using synthetic microcarriers and xeno-free medium, ensuring high cellular volumetric productivities (50 × 106 cell L−1 day−1), expansion factors (14–16 fold) and cell recovery yields (80{\%}). For the concentration step, flat sheet cassettes (FSC) and hollow fiber cartridges (HF) were compared showing a fairly linear scale-up, with a need to slightly decrease the permeate flux (30–50 LMH, respectively) to maximize cell recovery yield. Nonetheless, FSC allowed to recover 18{\%} more cells after a volume reduction factor of 50. Overall, at the end of the entire bioprocess more than 65{\%} of viable (>95{\%}) hMSC could be recovered without compromising cell's critical quality attributes (CQA) of viability, identity and differentiation potential. Alongside the standard quality assays, a proteomics workflow based on mass spectrometry tools was established to characterize the impact of processing on hMSC's CQA; These analytical tools constitute a powerful tool to be used in process design and development.",
keywords = "Cell therapy, Human mesenchymal stem cells, Mass spectrometry, Process development, Product characterization, Scale-up",
author = "B{\'a}rbara Cunha and Tiago Aguiar and Carvalho, {Sofia B.} and Silva, {Marta M.} and Gomes, {Ricardo A.} and Carrondo, {Manuel J.T.} and Patr{\'i}cia Gomes-Alves and Cristina Peixoto and Margarida Serra and Alves, {Paula M.}",
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TY - JOUR

T1 - Bioprocess integration for human mesenchymal stem cells

T2 - From up to downstream processing scale-up to cell proteome characterization

AU - Cunha, Bárbara

AU - Aguiar, Tiago

AU - Carvalho, Sofia B.

AU - Silva, Marta M.

AU - Gomes, Ricardo A.

AU - Carrondo, Manuel J.T.

AU - Gomes-Alves, Patrícia

AU - Peixoto, Cristina

AU - Serra, Margarida

AU - Alves, Paula M.

PY - 2017/4/20

Y1 - 2017/4/20

N2 - To deliver the required cell numbers and doses to therapy, scaling-up production and purification processes (at least to the liter-scale) while maintaining cells’ characteristics is compulsory. Therefore, the aim of this work was to prove scalability of an integrated streamlined bioprocess compatible with current good manufacturing practices (cGMP) comprised by cell expansion, harvesting and volume reduction unit operations using human mesenchymal stem cells (hMSC) isolated from bone marrow (BM-MSC) and adipose tissue (AT-MSC). BM-MSC and AT-MSC expansion and harvesting steps were scaled-up from spinner flasks to 2 L scale stirred tank single-use bioreactor using synthetic microcarriers and xeno-free medium, ensuring high cellular volumetric productivities (50 × 106 cell L−1 day−1), expansion factors (14–16 fold) and cell recovery yields (80%). For the concentration step, flat sheet cassettes (FSC) and hollow fiber cartridges (HF) were compared showing a fairly linear scale-up, with a need to slightly decrease the permeate flux (30–50 LMH, respectively) to maximize cell recovery yield. Nonetheless, FSC allowed to recover 18% more cells after a volume reduction factor of 50. Overall, at the end of the entire bioprocess more than 65% of viable (>95%) hMSC could be recovered without compromising cell's critical quality attributes (CQA) of viability, identity and differentiation potential. Alongside the standard quality assays, a proteomics workflow based on mass spectrometry tools was established to characterize the impact of processing on hMSC's CQA; These analytical tools constitute a powerful tool to be used in process design and development.

AB - To deliver the required cell numbers and doses to therapy, scaling-up production and purification processes (at least to the liter-scale) while maintaining cells’ characteristics is compulsory. Therefore, the aim of this work was to prove scalability of an integrated streamlined bioprocess compatible with current good manufacturing practices (cGMP) comprised by cell expansion, harvesting and volume reduction unit operations using human mesenchymal stem cells (hMSC) isolated from bone marrow (BM-MSC) and adipose tissue (AT-MSC). BM-MSC and AT-MSC expansion and harvesting steps were scaled-up from spinner flasks to 2 L scale stirred tank single-use bioreactor using synthetic microcarriers and xeno-free medium, ensuring high cellular volumetric productivities (50 × 106 cell L−1 day−1), expansion factors (14–16 fold) and cell recovery yields (80%). For the concentration step, flat sheet cassettes (FSC) and hollow fiber cartridges (HF) were compared showing a fairly linear scale-up, with a need to slightly decrease the permeate flux (30–50 LMH, respectively) to maximize cell recovery yield. Nonetheless, FSC allowed to recover 18% more cells after a volume reduction factor of 50. Overall, at the end of the entire bioprocess more than 65% of viable (>95%) hMSC could be recovered without compromising cell's critical quality attributes (CQA) of viability, identity and differentiation potential. Alongside the standard quality assays, a proteomics workflow based on mass spectrometry tools was established to characterize the impact of processing on hMSC's CQA; These analytical tools constitute a powerful tool to be used in process design and development.

KW - Cell therapy

KW - Human mesenchymal stem cells

KW - Mass spectrometry

KW - Process development

KW - Product characterization

KW - Scale-up

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U2 - 10.1016/j.jbiotec.2017.01.014

DO - 10.1016/j.jbiotec.2017.01.014

M3 - Article

VL - 248

SP - 87

EP - 98

JO - Journal of Biotechnology

JF - Journal of Biotechnology

SN - 0168-1656

ER -