title: Clinical scale ex vivo manufacture of neutrophils from hematopoietic progenitor cells.
abstract: Dose-intensive chemotherapy results in an obligatory period of severe neutropenia during which patients are at high risk of infection. While patient support with donor neutrophils is possible, this option is restricted due to donor availability and logistic complications. To overcome these problems, we explored the possibility of large scale ex vivo manufacture of neutrophils from hematopoietic progenitor cells (HPC). CD34+ HPC isolated from umbilical cord blood (UCB) and mobilized peripheral blood (mPB) were expanded in serum-free medium supplemented with stem cell factor, granulocyte colony stimulating factor, and a thrombopoietin peptide mimetic. After 15 days of cultivation a 5,800-fold expansion in cell number was achieved for UCB, and up to 4,000-fold for mPB, comprising 40% and 60% mature neutrophils respectively. Ex vivo expanded neutrophils exhibited respiratory burst activity similar to that for donor neutrophils, and were capable of killing Candida albicans in vitro. These yields correspond to a more than 10-fold improvement over current methods, and are sufficient for the production of multiple neutrophil transfusion doses per HPC donation. To enable clinical scale manufacture, we adapted our protocol for use in a wave-type bioreactor at a volume of 10 L. This is the first demonstration of a large scale bioprocess suitable for routine manufacture of a mature blood cell product from HPC, and could enable prophylactic neutrophil support for chemotherapy patients.
authors: Australian Institute for Bioengineering and Nanotechnology, The University of Queensland, Brisbane, QLD 4072, Australia.
authors: Nicholas E
authors: NE
authors: Timmins
authors: Emma
authors: E
authors: Palfreyman
authors: Flavia
authors: F
authors: Marturana
authors: Stefanie
authors: S
authors: Dietmair
authors: Sanna
authors: S
authors: Luikenga
authors: Genghis
authors: G
authors: Lopez
authors: Yoke Lin
authors: YL
authors: Fung
authors: Robyn
authors: R
authors: Minchinton
authors: Lars K
authors: LK
authors: Nielsen
mesh_terms: D000328:Adult; D002176:Candida albicans / Q000276:immunology; D018929:Cell Culture Techniques / Q000379:methods*; D018797:Cell Cycle Proteins / Q000378:metabolism; D049109:Cell Proliferation; D016895:Culture Media, Serum-Free; D005312:Fetal Blood / Q000166:cytology*; D006412:Hematopoietic Stem Cells; D006801:Humans; D050296:Microbial Viability; D009504:Neutrophils / Q000276:immunology / Q000502:physiology*
chemical_list: C515965:CDCA4 protein, human; D018797:Cell Cycle Proteins; D016895:Culture Media, Serum-Free