Clinical-scale production of granulocyte progenitor and post-progenitor cells using daniplestim, leridistim, Progenipoietin, Promegapoietin and autologous plasma.
BACKGROUND
Supplementation of PBPC autografts with ex vivo expanded PBMC may significantly reduce or eliminate the period of neutropenia associated with high-dose chemotherapy.


METHODS
Unmanipulated growth-factor mobilized PBMC were expanded in media containing daniplestim, leridistim, Promegapoietin, and Progenipoietin (DLPP) and 2% autologous plasma at 4 x 10(5) PBMC/mL, first in 25 cm(2) T-flasks, with sampling on Days 7, 10, 13 and 15, and then in 1264 cm(2) Nunclon Cell Factories, with sampling on Days 7 and 13.


RESULTS
In T25-flasks, maximal CFU-GM expansion ([38.2 +/- 9.5]-fold) occurred on Day 10, whereas maximal total cell expansion ([6.7 +/- 1.1]-fold) occurred on Day 15. Production of CD15(+)CD11b(-) and CD15(+)CD11b(+) granulocytic post-progenitors (3.0 +/- 0.4 x 10(6) and 3.7 +/- 0.9 x 10(6), respectively) was also maximal at Day 15. Compared with the previously studied combination of Flt3L, PIXY321, G-CSF, GM-CSF and Epo, the DLPP cocktail performed similarly, with the exception of yielding larger GM colonies at Day 10 and fewer granulocyte post-progenitors on Day 15. In Cell Factories, CFU-GM were expanded (31.6 +/- 14.5)-fold, while total nonadherent cells were expanded (2.6 +/- 0.5)-fold. The two stack Cell Factory cultures seeded with 1.0 x 10(8) unselected PBMC produced approximately 3.3 x 10(6) CFU-GM and 1.3 x 10(8) myeloid post-progenitors.


CONCLUSIONS
Whereas expansion of cell numbers, CFU-GM and granulocytic post-progenitors in Cell Factories mirrored that achieved in T25-flasks, future preclinical studies with the DLPP cytokine combination may be performed in small volumes, with subsequent translation to the larger volume Cell Factories. Sufficient expansion can be achieved using the DLPP cytokine combination in the Cell Factories to provide the numbers of progenitors required for clinical trials.
Patel
S D
SD
Department of Chemical Engineering, Northwestern University, Evanston and Chicago, IL, USA.
Guo
R
R
Miller
W M
WM
Papoutsakis
E T
ET
Minster
N I
NI
Baum
C M
CM
Winter
J N
JN
D018929:Cell Culture Techniques / Q000379:methods*; D002454:Cell Differentiation / Q000187:drug effects; D002455:Cell Division / Q000187:drug effects; D019070:Cell Lineage / Q000187:drug effects; D016207:Cytokines / Q000494:pharmacology*; D006023:Glycoproteins / Q000494:pharmacology; D016179:Granulocyte Colony-Stimulating Factor / Q000494:pharmacology; D006098:Granulocytes / Q000166:cytology* / Q000187:drug effects*; D006133:Growth Substances / Q000494:pharmacology; D006801:Humans; D016130:Immunophenotyping; D007377:Interleukin-3 / Q000494:pharmacology / Q000031:analogs & derivatives; D007958:Leukocyte Count; D010446:Peptide Fragments; D010455:Peptides / Q000494:pharmacology; D016185:Receptors, Interleukin-3 / Q000378:metabolism; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D013234:Stem Cells / Q000166:cytology* / Q000187:drug effects*; D013926:Thrombopoietin
D016207:Cytokines; D006023:Glycoproteins; D006133:Growth Substances; D007377:Interleukin-3; D010446:Peptide Fragments; D010455:Peptides; D016185:Receptors, Interleukin-3; D011993:Recombinant Fusion Proteins; D011994:Recombinant Proteins; D016179:Granulocyte Colony-Stimulating Factor; C090367:megapoietin protein, Ovis aries; C400711:leridistim; C093340:daniplestim; D013926:Thrombopoietin