The monoculture of cord-blood-derived CD34 ⁺ cells by an automated, membrane-based dynamic perfusion system with a novel cytokine cocktail

Summary

Human leukocyte antigen (HLA)-matched cord blood (CB) transplantation is a procedure for the treatment of certain hematological malignancies, hemoglobinopathies, and autoimmune disorders. However, one of the challenges is to provide a sufficient number of T cell-depleted hematopoietic stem and progenitor cells. Currently, only 4%–5% of the CB units stored in CB banks contain enough CD34 ⁺ cells for engrafting 70-kg patients. To support this clinical need, we have developed an automated expansion protocol for CB-derived CD34 ⁺ cells in the Quantum system’s dynamic perfusion bioreactor using a novel cytokine cocktail comprised of stem cell factor (SCF), thrombopoietin (TPO), fms-like tyrosine kinase 3 ligand (Flt-3L), interleukin-3 (IL-3), IL-6, glial cell line-derived neurotrophic factor (GDNF), StemRegenin 1 (SR-1), and a fibronectin-stromal-cell-derived factor-1 (SDF-1)-coated membrane. In an 8-day expansion of a 2 × 10 ⁶ positively selected CD34 ⁺ cell inoculum from 3 donor lineages, the mean cell harvest and cell viability were 1.02 × 10 ⁸ cells and 95.5%, respectively, and the mean frequency of the CD45 ⁺CD34 ⁺ immunophenotype was 54.3%. The mean differentiated cell frequencies were 0.5% for lymphocytes, 15.8% for neutrophils, and 15.4% for platelets. These results demonstrate that the automated monoculture protocol can support the expansion of CD34 ⁺ cells with minimal lymphocyte residual.

Highlights

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  Expansion of CD34 ⁺ cells in a perfusion bioreactor using a novel cytokine cocktail

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  8-day expansion of CD34 ⁺ cells produces a 51-fold increase over 3 donor lineages

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  Differentiated cell types are minimized during expansion

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  Monoculture supports the expansion of CD34 ⁺ cells with minimal lymphocyte residual

Abstract

In this article, Jones and colleagues show the development of an automated expansion protocol for CB-derived CD34 ⁺ cells in the Quantum system’s perfusion bioreactor using a novel cytokine cocktail with GDNF and a fibronectin-SDF-1-coated membrane. In an 8-day expansion of 2 × 10 ⁶ positively selected CD34 ⁺ cells, the mean cell harvest and cell viability were 1.02 × 10 ⁸ cells and 95.5%, respectively, and the mean CD45 ⁺CD34 ⁺ immunophenotype frequency was 54.3% with minimal lymphocyte residual.