Enforced expression of an Flt3 internal tandem duplication in human CD34+ cells confers properties of self-renewal and enhanced erythropoiesis.
To investigate the role of constitutively active internal tandem duplication (ITD) mutants of the Fms-like tyrosine kinase 3 (Flt3) receptor in leukemogenesis, we introduced the Flt3-ITD, W51, into human cord blood CD34+ cells and evaluated their phenotype in diverse hematopoietic assays. W51 expression resulted in a strong proliferative advantage and enhanced erythropoiesis as determined by immunophenotyping, colony assays, and molecular analyses. In MS-5 stromal cocultures, numerous early cobblestone areas (CAs) were generated within 10 to 14 days. Such W51-associated early CAs disappeared by 4 weeks, yet retained self-renewal properties as demonstrated by generation of secondary and tertiary CAs upon replating. This phenotype appears related to the expression of W51 since it was abolished by exposure to the FLT3 inhibitor, AG1295, but not to the c-kit inhibitor PD16. Wild-type Flt3-overexpressing CD34+ cells exposed to high levels of its physiologic ligand did not produce early CAs, highlighting differences in intracellular signaling between wild-type Flt3 and W51. W51-associated signal transducer and activator of transcription 5 (Stat5) activation plays a major role in this phenotype, although additional downstream targets of W51 may be relevant. Flt3-ITD+ acute myeloid leukemia (AML) blasts from patients invariably generated early AG1295-sensitive CAs in MS-5 cocultures, further validating the phenotype observed in transduced CD34+ cells.
Chung
Ki Young
KY
Laboratory of Developmental Hematopoiesis, Cell Biology Program, Memorial Sloan-Kettering Cancer Center, New York, NY 10021, USA.
Morrone
Giovanni
G
Schuringa
Jan Jacob
JJ
Wong
Bryan
B
Dorn
David C
DC
Moore
Malcolm A S
MA
D000818:Animals; D018952:Antigens, CD34 / Q000378:metabolism*; D049109:Cell Proliferation; D002478:Cells, Cultured; D004268:DNA-Binding Proteins / Q000378:metabolism; D004920:Erythropoiesis; D005786:Gene Expression Regulation; D006801:Humans; D016130:Immunophenotyping; D015470:Leukemia, Myeloid, Acute / Q000378:metabolism / Q000473:pathology; D051379:Mice; D008894:Milk Proteins / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D019009:Proto-Oncogene Proteins c-kit / Q000378:metabolism; D020794:Receptor Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D050799:STAT5 Transcription Factor; D013234:Stem Cells / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D015534:Trans-Activators / Q000378:metabolism; D014161:Transduction, Genetic; D020032:Tyrphostins / Q000494:pharmacology; D051941:fms-Like Tyrosine Kinase 3
C407689:6,7-dimethoxy-2-phenylquinoxaline; D018952:Antigens, CD34; D004268:DNA-Binding Proteins; D008894:Milk Proteins; D011518:Proto-Oncogene Proteins; D050799:STAT5 Transcription Factor; D015534:Trans-Activators; D020032:Tyrphostins; C497970:FLT3 protein, human; C497971:Flt3 protein, mouse; D019009:Proto-Oncogene Proteins c-kit; D020794:Receptor Protein-Tyrosine Kinases; D051941:fms-Like Tyrosine Kinase 3