Functional G-CSF pathways in t(8;21) leukemic cells allow for differentiation induction and degradation of AML1-ETO.
BACKGROUND
Efficacy of differentiating agents requires that their specific cellular targets are still expressed and functional in the leukemic cells. One hypothesis to target sensitive cells is to select leukemic clones which harbor disrupted transcription factors. CBFalpha and CBFbeta are core-binding proteins which have been identified as transcription regulators of hematopoietic genes and shown to be altered in numerous leukemias. In M2 AML, the t(8;21) translocation, CBFalpha (AML1) is altered and produced as the AML1-ETO fusion protein. The fusion protein blocks transcription and differentiation mediated by G-CSF. Interestingly, AML1-ETO leukemic cell lines are sensitive to numerous cytokines in vitro and can be induced to differentiate in the presence of G-CSF and PMA.


METHODS
As in the APL differentiation model, primary culture provides a useful tool for therapeutic screening of differentiation inducers, we analysed the in vitro sensitivity of 10 fresh M2 AML t(8;21) leukemic samples to G-CSF and the functionality of G-CSF intracellular pathways. In vitro data were compared with in vivo data from four patients treated with rhG-CSF at the dosage of 5 microg/kg/day i.v. for two to three weeks before the initiation of AML induction chemotherapy and immunophenotypic analysis performed weekly to monitor in vivo differentiation.


RESULTS
In vitro, an increase in CD34+ cells expressing differentiation antigens (CD11b, CD13 or CD15) was noted along with a decrease of immature CD34+/differentiation antigen negative cells. After two weeks of a daily rhG-CSF administration in vivo, a significant, albeit transient, decrease of blast count was achieved, concomitant with an increase in differentiated leukemic cells suggesting that in vivo differentiation occurs. Fresh t(8;21) leukemic cells possess functional G-CSF signaling pathways as normal activity and kinetics of STAT1 and STAT3 binding was observed. Furthermore, differentiation induction leads to a subsequent degradation of the AML1-ETO oncoprotein.


CONCLUSIONS
The data presented here supports the claim that G-CSF can induce in vitro and in vivo differentiation of M2 AML t(8;21) cells.
Da Silva
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N
Laboratoire de Biologie Cellulaire Hématopoïétique (LBCH), INSERM E 00-03, et EA 316 Université Paris 7, Hôpital Saint-Louis, 1 Avenue Claude Vellefaux, 75754 Paris Cedex 10, France. lbch@chu.stlouis.fr
Meyer-Monard
S
S
Menot
M L
ML
Parrado
A
A
Lebel
A
A
Balitrand
N
N
Fenaux
P
P
Micléa
J M
JM
Rousselot
P
P
Degos
L
L
Dombret
H
H
Chomienne
C
C
D015703:Antigens, CD / Q000032:analysis; D017209:Apoptosis; D001773:Blood Cells / Q000378:metabolism / Q000473:pathology; D001854:Bone Marrow Cells / Q000378:metabolism / Q000473:pathology; D002454:Cell Differentiation / Q000187:drug effects; D002891:Chromosomes, Human, Pair 21 / Q000235:genetics* / Q000648:ultrastructure; D002898:Chromosomes, Human, Pair 8 / Q000235:genetics* / Q000648:ultrastructure; D050676:Core Binding Factor Alpha 2 Subunit; D004268:DNA-Binding Proteins / Q000378:metabolism; D016179:Granulocyte Colony-Stimulating Factor / Q000494:pharmacology*; D006801:Humans; D015470:Leukemia, Myeloid, Acute / Q000235:genetics*; D009363:Neoplasm Proteins / Q000378:metabolism*; D015514:Oncogene Proteins, Fusion / Q000378:metabolism*; D000075142:RUNX1 Translocation Partner 1 Protein; D016188:Receptors, Granulocyte Colony-Stimulating Factor / Q000187:drug effects* / Q000502:physiology; D011994:Recombinant Proteins / Q000494:pharmacology; D050794:STAT1 Transcription Factor; D050796:STAT3 Transcription Factor; D015398:Signal Transduction / Q000187:drug effects; D015534:Trans-Activators / Q000378:metabolism; D014157:Transcription Factors / Q000378:metabolism*; D014178:Translocation, Genetic; D014212:Tretinoin / Q000494:pharmacology; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000378:metabolism
C107844:AML1-ETO fusion protein, human; D015703:Antigens, CD; D050676:Core Binding Factor Alpha 2 Subunit; D004268:DNA-Binding Proteins; D009363:Neoplasm Proteins; D015514:Oncogene Proteins, Fusion; D000075142:RUNX1 Translocation Partner 1 Protein; D016188:Receptors, Granulocyte Colony-Stimulating Factor; D011994:Recombinant Proteins; D050794:STAT1 Transcription Factor; C494079:STAT1 protein, human; D050796:STAT3 Transcription Factor; C494086:STAT3 protein, human; D015534:Trans-Activators; D014157:Transcription Factors; D016179:Granulocyte Colony-Stimulating Factor; D014212:Tretinoin