Effects of the bcr/abl kinase inhibitors AG957 and NSC 680410 on chronic myelogenous leukemia cells in vitro.
The tyrphostin AG957 (NSC 654705) inhibits p210bcr/abl, the transforming kinase responsible for most cases of chronic myelogenous leukemia (CML). The present studies were performed to determine the fate of AG957-treated cells and assess the selectivity of AG957 for CML myeloid progenitors. When K562 cells (derived from a patient with blast crisis CML) were treated with AG957, dose- and time-dependent p210bc/abl down-regulation was followed by mitochondrial release of cytochrome c, activation of caspase-9 and caspase-3, and apoptotic morphological changes. These apoptotic changes were inhibited by transfection with cDNA encoding dominant negative caspase-9 but not dominant-negative FADD or blocking anti-Fas antibodies. In additional experiments, a 24-h AG957 exposure caused dose-dependent inhibition of K562 colony formation in soft agar. To extend these studies to clinical samples of CML, peripheral blood mononuclear cells from 10 chronic phase CML patients and normal controls were assayed for the growth of hematopoietic colonies in vitro in the presence of increasing concentrations of AG957. These assays demonstrated selectivity of AG957 for CML progenitors, with median IC50s (CML versus normal) of 7.3 versus >20 microM AG957 in granulocyte colony-forming cells (P < 0.001), 5.3 versus >20 microM in granulocyte/macrophage colony-forming cells (P < 0.05), and 15.5 versus > 20 microM in erythroid colony-forming cells (P > 0.05). The adamantyl ester of AG957 (NSC 680410) down-regulated p210bcr/abl in K562 cells and inhibited granulocyte colony formation in CML specimens at lower concentrations without enhanced toxicity in normal progenitors. These observations not only demonstrate that AG957-induced p210bcr/abl down-regulation is followed by activation of the cytochrome c/Apaf-1/caspase-9 pathway but also indicate that this class of kinase inhibitor exhibits selectivity worthy of further evaluation.
Svingen
P A
PA
Department of Oncology, Mayo Medical School, Rochester, Minnesota 55905, USA.
Tefferi
A
A
Kottke
T J
TJ
Kaur
G
G
Narayanan
V L
VL
Sausville
E A
EA
Kaufmann
S H
SH
D000218:Adamantane / Q000031:analogs & derivatives* / Q000633:toxicity; D017209:Apoptosis / Q000187:drug effects; D053453:Caspase 9; D020169:Caspases / Q000378:metabolism; D002455:Cell Division / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000633:toxicity*; D016044:Fusion Proteins, bcr-abl / Q000037:antagonists & inhibitors* / Q000235:genetics; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D006801:Humans; D006873:Hydroquinones / Q000633:toxicity*; D020014:K562 Cells; D007700:Kinetics; D015464:Leukemia, Myelogenous, Chronic, BCR-ABL Positive; D011505:Protein-Tyrosine Kinases / Q000037:antagonists & inhibitors; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014162:Transfection; D014410:Tumor Stem Cell Assay; D020032:Tyrphostins / Q000633:toxicity*
D004791:Enzyme Inhibitors; D006873:Hydroquinones; C404397:NSC 680410; D011993:Recombinant Fusion Proteins; D020032:Tyrphostins; C104965:tyrphostin AG957; D011505:Protein-Tyrosine Kinases; D016044:Fusion Proteins, bcr-abl; C506433:CASP9 protein, human; D053453:Caspase 9; D020169:Caspases; D000218:Adamantane