Vol. 63, Issue 1, 232-242, January 2003

Bryostatin 1 and UCN-01 Potentiate 1--D-Arabinofuranosylcytosine-Induced Apoptosis in Human Myeloid Leukemia Cells through Disparate Mechanisms

Shujie Wang, Zhiliang Wang, and Steven Grant

Division of Hematology/Oncology, Department of Medicine (S.W., Z.W., S.G.) and the Departments of Pharmacology (S.G.) and Biochemistry (S.W., S.G.), Medical College of Virginia, Virginia Commonwealth University, Richmond, Virginia

The effects of the PKC activator and down-regulator bryostatin 1 and the PKC and Chk1 inhibitor 7-hydroxystaurosporine (UCN-01) were compared with respect to potentiation of 1--D-arabinofuranosylcytosine (ara-C)-induced apoptosis in human myelomonocytic leukemia cells (U937). Whereas bryostatin 1 and UCN-01 both markedly enhanced ara-C-induced mitochondrial injury (e.g., cytochrome c and Smac/DIABLO release, loss of mitochondrial membrane potential), caspase activation, and apoptosis, ectopic expression of an N-terminal loop-deleted Bcl-2 mutant protein protected cells from ara-C/UCN-01- but not ara-C/bryostatin 1-mediated lethality. Conversely, ectopic expression of CrmA or dominant-negative caspase-8 abrogated potentiation of ara-C-mediated apoptosis by bryostatin 1 but not by UCN-01. Exposure of cells to ara-C and bryostatin 1 (but not UCN-01) resulted in sustained release of tumor necrosis factor (TNF) ; moreover, potentiation of ara-C lethality by bryostatin 1 (but not by UCN-01) was reversed by coadministration of TNF soluble receptors or the selective PKC inhibitor bisindolylmaleimide (1 µM). Finally, similar events were observed in the human promyelocytic leukemia cell line HL-60. Together, these findings suggest that potentiation of ara-C lethality in human myeloid leukemia cells by bryostatin 1 but not UCN-01 involves activation of the extrinsic, receptor-mediated apoptotic pathway, and represents a consequence of bryostatin 1-mediated release of TNF-. They also argue that the mechanism by which bryostatin 1 promotes ara-C-induced mitochondrial injury, caspase activation, and apoptosis involves factors other than or in addition to PKC down-regulation or modulation of Bcl-2 phosphorylation status.