Ex vivo cytokine expansion of peripheral blood 5-fluorouracil-treated CD34-positive chronic myeloid leukaemia cells increases the selection of Ph-negative cells.
It has been shown that normal early progenitor or stem cells persist in the blood and bone marrow of patients with Philadelphia chromosome (Ph)-positive chronic myeloid leukaemia (CML); it is also known that normal haemopoietic progenitors can be expanded ex vivo in the presence of various cytokine combinations. The selection of normal (Ph-negative) progenitor cells from CML patients would potentially be of considerable clinical value for ex vivo purging and autologous transplantation. To obtain these cells. CD34-positive (progenitor) cells from the peripheral blood (PB) of CML patients were pretreated with 5-fluorouracil (5FU) (5 microg/ml) to suppress Ph-positive cells and then grown in suspension culture for 7 d with a combination of cytokines. We compared two combinations of cytokines: interleukin-1alpha (IL1alpha) and interleukin-3 (IL3) with either Flt3-ligand (FLT3L) or stem cell factor (SCF). Using these two combinations, we obtained the same degree of day 14 CFU-GM expansion (3.1 +/- 0.5 and 3.4 +/- 0.7 fold expansion). FISH analysis showed that 5FU pretreatment significantly favoured a higher frequency of Ph-negative cells after expansion in liquid culture. Moreover, after 5FU pretreatment, the mean (+/-SEM) percentage of Ph negativity was significantly greater for IL1alpha-IL3-FLT3L compared to IL1alpha-IL3-SCF (19.1 +/- 2.5% v 14.8 +/- 2.3%, P=0.009, n = 7). The output long-term culture initiating cells (LTC-IC) which could only be detected after 5FU pretreatment and the combination, IL1alpha-IL3-FLT3L were all Ph negative by FISH analysis. Thus, a subset of Ph-negative cells was selected from CML PB by 5FU and expanded using the combination of cytokines IL1alpha-IL3-FLT3L and IL1alpha-IL3-SCF. Ex vivo expansion of putatively normal haemopoietic progenitor cells is feasible in CML.
Mahon
F X
FX
Laboratoire de Greffe de Moelle, UMR CNRS 5540, Université Victor Segalen Bordeaux 2, France.
Pigeonnier-Lagarde
V
V
Chahine
H
H
Barbot
C
C
Jazwiec
B
B
Ripoche
J
J
Reiffers
J
J
D018952:Antigens, CD34 / Q000097:blood*; D002455:Cell Division; D016207:Cytokines / Q000494:pharmacology*; D005472:Fluorouracil / Q000494:pharmacology*; D016044:Fusion Proteins, bcr-abl / Q000378:metabolism; D006801:Humans; D017404:In Situ Hybridization, Fluorescence; D007375:Interleukin-1 / Q000494:pharmacology; D007377:Interleukin-3 / Q000494:pharmacology; D054438:Leukemia, Myeloid, Chronic, Atypical, BCR-ABL Negative / Q000473:pathology*; D008565:Membrane Proteins / Q000494:pharmacology; D019089:Stem Cell Factor / Q000494:pharmacology; D013234:Stem Cells / Q000473:pathology; D014407:Tumor Cells, Cultured / Q000473:pathology
D018952:Antigens, CD34; D016207:Cytokines; D007375:Interleukin-1; D007377:Interleukin-3; D008565:Membrane Proteins; D019089:Stem Cell Factor; C084881:flt3 ligand protein; D016044:Fusion Proteins, bcr-abl; D005472:Fluorouracil