Potentiation of 1-beta-D-arabinofuranosylcytosine-mediated mitochondrial damage and apoptosis in human leukemia cells (U937) overexpressing bcl-2 by the kinase inhibitor 7-hydroxystaurosporine (UCN-01).
Antileukemic interactions between the nucleoside analog 1-beta-D-arabinofuranosylcytosine (ara-C) and the kinase inhibitor 7-hydroxystaurosporine (UCN-01) have been examined in relation to Bcl-2 expression/phosphorylation, mitochondrial damage, caspase activation, and loss of clonogenic potential. Subsequent exposure of ara-C-pretreated U937 cells (1 microM; 6 hr) to UCN-01 (300 nM; 24 hr) resulted in marked potentiation of pro-caspase-3 and -9 cleavage/activation, poly(ADP-ribose)polymerase degradation, diminished mitochondrial membrane potential (Deltapsi(m)), enhanced cytochrome c release, reduction in the S-phase fraction, and induction of classic apoptotic morphologic features. Enforced expression of full-length Bcl-2 significantly protected cells (at 24 hr) from ara-C/UCN-01-induced caspase activation and apoptosis, but was ineffective in preventing loss of Deltapsi(m) and cytochrome c release. Ectopic expression of a Bcl-2 N-terminal phosphorylation loop-deleted protein (Bcl-2Delta(32-80)) was more potent than its full-length counterpart in blocking drug-induced loss of Deltapsi(m, ) caspase activation, and apoptotic morphology, but not cytochrome c release. Examination of cells at later intervals revealed that ectopic expression of Bcl-2 or Bcl-2Delta(32-80) could only delay, but not prevent, mitochondrial damage, caspase activation, and cell death induced by ara-C/UCN-01 treatment. Despite their initial ability to inhibit apoptosis, neither full-length nor truncated Bcl-2 protein restored clonogenic potential to drug-treated cells. These findings indicate that subsequent exposure of ara-C-pretreated human leukemia cells to UCN-01 potently triggers mitochondrial damage and apoptosis, and that these events are postponed but not prevented by ectopic expression of Bcl-2 or its phosphorylation loop-deleted counterpart.
Tang
L
L
Department of Microbiology, Medical College of Virginia, Virginia Commonwealth University, Richmond, VA, USA.
Boise
L H
LH
Dent
P
P
Grant
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D000470:Alkaloids / Q000494:pharmacology*; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis; D003561:Cytarabine / Q000494:pharmacology*; D004357:Drug Synergism; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D007938:Leukemia / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D008928:Mitochondria / Q000187:drug effects*; D011493:Protein Kinase C / Q000037:antagonists & inhibitors; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism* / Q000502:physiology; D019311:Staurosporine / Q000031:analogs & derivatives; D020298:U937 Cells
D000470:Alkaloids; D000970:Antineoplastic Agents; D004791:Enzyme Inhibitors; D019253:Proto-Oncogene Proteins c-bcl-2; D003561:Cytarabine; C054852:7-hydroxystaurosporine; D011493:Protein Kinase C; D019311:Staurosporine