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Production of erythrocytes from directly isolated or Delta1 Notch ligand expanded CD34 hematopoietic progenitor cells: process characterization, monitoring and implications for manufacture

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posted on 2013-09-18, 13:56 authored by Katie GlenKatie Glen, Victoria L. Workman, Forhad Ahmed, Elizabeth RatcliffeElizabeth Ratcliffe, Adrian J. Stacey, Rob ThomasRob Thomas
Background aims: Economic ex vivo manufacture of erythrocytes at 10 cell doses requires an efficiently controlled bio-process capable of extensive proliferation and high terminal density. High-resolution characterization of the process would identify production strategies for increased efficiency, monitoring and control. Methods: CD34 cord blood cells or equivalent cells that had been pre-expanded for 7 days with Delta1 Notch ligand were placed in erythroid expansion and differentiation conditions in a micro-scale ambr suspension bioreactor. Multiple culture parameters were varied, and phenotype markers and metabolites measured to identify conserved trends and robust monitoring markers. Results: The cells exhibited a bi-modal erythroid differentiation pattern with an erythroid marker peak after 2 weeks and 3 weeks of culture; differentiation was comparatively weighted toward the second peak in Delta1 pre-expanded cells. Both differentiation events were strengthened by omission of stem cell factor and dexamethasone. The cumulative cell proliferation and death, or directly measured CD45 expression, enabled monitoring of proliferative rate of the cells. The metabolic activities of the cultures (glucose, glutamine and ammonia consumption or production) were highly variable but exhibited systematic change synchronized with the change in differentiation state. Conclusions: Erythroid differentiation chronology is partly determined by the heterogeneous CD34 progenitor compartment with implications for input control; Delta1 ligand-mediated progenitor culture can alter differentiation profile with control benefits for engineering production strategy. Differentiation correlated changes in cytokine response, markers and metabolic state will enable scientifically designed monitoring and timing of manufacturing process steps. © 2013 International Society for Cellular Therapy.

Funding

This work was supported by the Engineering and Physical Sciences Research Council (EPSRC).

History

School

  • Mechanical, Electrical and Manufacturing Engineering

Citation

GLEN, K.E. ... et al, 2013. Production of erythrocytes from directly isolated or Delta1 Notch ligand expanded CD34 hematopoietic progenitor cells: process characterization, monitoring and implications for manufacture. Cytotherapy, 15 (9), pp. 1106 - 1117

Publisher

Elsevier / © the authors / International Society for Cellular Therapy

Version

  • VoR (Version of Record)

Publisher statement

http://creativecommons.org/licenses/by/3.0/

Publication date

2013

Notes

This is an Open Access Article. It is published by Elsevier under the Creative Commons Attribution 3.0 Unported Licence (CC BY). Full details of this licence are available at: http://creativecommons.org/licenses/by/3.0/

ISSN

1465-3249

eISSN

1477-2566

Language

  • en

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