title: Different antiapoptotic pathways between wild-type and mutated FLT3: insights into therapeutic targets in leukemia.
abstract: An internal tandem duplication (ITD) of the juxtamembrane (JM) domain of FLT3 (FLT3/ITD) has been found in 20% of patients with acute myeloid leukemia (AML) and is correlated with leukocytosis and a poor prognosis. Here, we compared the antiapoptotic effects of wild-type FLT3 (WtFLT3) and FLT3/ITD in terms of the regulation of Bcl-2 family members. In a murine myeloid cell line, 32D, interleukin-3 (IL-3) deprivation induced apoptosis following the down-regulation of Bcl-XL and the dephosphorylation of Bad. However, the expression levels of Bcl-2, Bax, Bak, and Mcl-1 were unchanged. In WtFLT3-transfected 32D (WtFLT3-32D) cells, FLT3 ligand (FL) stimulation did not restore the down-regulation of Bcl-XL but maintained the phosphorylation of Bad. Combined treatment with phosphatidylinositol 3-kinase (PI3K) inhibitor, LY294002, and mitogen-activated protein kinase kinase (MEK) inhibitor, PD98059, dephosphorylated Bad and induced apoptosis in WtFLT3-32D cells stimulated with FL. Induction of nonphosphorylated Bad induced remarkable apoptosis. These findings suggest that the FL stimulation is associated with antiapoptosis through Bad phosphorylation. On the other hand, FLT3/ITD-transfected 32D (FLT3/ITD-32D) cells survived in an IL-3-or FL-deprived state. Furthermore, the dephosphorylation of Bad using LY294002 and PD98059 was insufficient for apoptosis, and the down-regulation of Bcl-XL using antisense treatment was needed to induce apoptosis. FLT3 kinase inhibitor, AG1296, alone not only dephosphorylated Bad but also down-regulated Bcl-XL, leading FLT3/ITD-32D cells into apoptosis. These findings suggest that the antiapoptotic pathways from FLT3/ITD are more divergent than those from WtFLT3 and may represent targets for drug discovery with the potential of inducing selective cell death of human leukemia cells.
authors.affiliation: Department of Hematology, Nagoya University Graduate School of Medicine, Tsurumai-cho 65, Showa-ku, Nagoya 466-8550, Japan.
authors.forename: Yosuke
authors.initials: Y
authors.lastname: Minami
authors.forename: Kazuhito
authors.initials: K
authors.lastname: Yamamoto
authors.forename: Hitoshi
authors.initials: H
authors.lastname: Kiyoi
authors.forename: Ryuzo
authors.initials: R
authors.lastname: Ueda
authors.forename: Hidehiko
authors.initials: H
authors.lastname: Saito
authors.forename: Tomoki
authors.initials: T
authors.lastname: Naoe
mesh_terms: D017209:Apoptosis; D002455:Cell Division; D002460:Cell Line; D002867:Chromones / Q000494:pharmacology; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005419:Flavonoids / Q000494:pharmacology; D006801:Humans; D015151:Immunoblotting; D007938:Leukemia / Q000235:genetics* / Q000473:pathology*; D008024:Ligands; D008565:Membrane Proteins / Q000235:genetics*; D009025:Morpholines / Q000494:pharmacology; D009154:Mutation; D016376:Oligonucleotides, Antisense / Q000494:pharmacology; D019869:Phosphatidylinositol 3-Kinases / Q000378:metabolism; D010766:Phosphorylation; D011233:Precipitin Tests; D011379:Prognosis; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D015398:Signal Transduction; D013997:Time Factors; D014162:Transfection; D051020:bcl-X Protein
chemical_list: C494811:BCL2L1 protein, human; D002867:Chromones; D004791:Enzyme Inhibitors; D005419:Flavonoids; D008024:Ligands; D008565:Membrane Proteins; D009025:Morpholines; D016376:Oligonucleotides, Antisense; D019253:Proto-Oncogene Proteins c-bcl-2; D051020:bcl-X Protein; C084881:flt3 ligand protein; C085911:2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran-4-one; D019869:Phosphatidylinositol 3-Kinases; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one