Large-scale production of red blood cells from stem cells: What are the technical challenges ahead?
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Transcript of Large-scale production of red blood cells from stem cells: What are the technical challenges ahead?
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Large-scale production of red blood cells from stem cells: What are the technical challeng
es ahead?
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cytokine
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Main advances in RBC culture in view of the large-scale production of clinical grade cells
A biological odyssey
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On the way to clinical applications
(i) Are there any fundamental barriers to large-scalemanufacture of RBC?
(ii) Is it possible to generate clinically relevant numbers of cells?
Three economic questions arise
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The price
① 5 × 107 cells/mL, it would take 40 liters of culture media with a total cost of $8330 to generate enough cells (without consideringthe investment in facilities)
②Zeuner et al. proposed a production price $8000–15 000 per unit of blood.
Timmins, N. E., Nielsen, L. K., Blood cell manufacture: current methods and future challenges. Trends Biotechnol. 2009, 27, 415–422.
Zeuner, A., Martelli, F., Vaglio, S., Federici, G. et al., Concise review: stem cell-derived erythrocytes as upcoming players in blood transfusion. Stem Cells 2012, 30, 1587–1596.
$700–1200 hospital cost
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biotechnological breakthrough
seven 175-cm2 flasks
expansion and dilution
CD34+ RBC
2 tennis courts
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medium price
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control of the pH
pH
lactate
cell metabolism slows down with higher cell density
ammoniametabolic byproducts
pH decreasemodification of osmolarity
direct growth inhibitor
glucose and glutamine metabolisms
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DO level
Dissolved oxygen
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How to improve the culture conditions
• It will be necessary to monitor at the same time the differentiation of the cells, which will imply a compromise between expansion and maturation.
• For RBC production, the complete medium is too expensive, with growth factors and other proteins representing more than 90% of the price. Thus, a new method to eliminate waste products while conserving the expensive molecules is needed.
• All the downstream process will have to be revised, with critical improvements to implement in the separation and purification procedures.
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RBC production in bioreactors
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Day 21 expanded cell populations comprised a pure population of erythroid cells of mixed maturity
EBs at different stages of maturation
21 Day
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Ultra-high-yield manufacture of RBC
CD71
CD235
107 108
500 units of RBC per UCB donation
per UCB donation (5 × 106 CD34+ cells/ donation)
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hollow-fiber bioreactor
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Bioreactor perfusion schematic
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Bioreactor parameters