POTENTIATION OF ARA-C-INDUCED APOPTOSIS BY THE PROTEIN-KINASE-C ACTIVATOR BRYOSTATIN-1 IN HUMAN LEUKEMIA-CELLS (HL-60) INVOLVES A PROCESS DEPENDENT UPON C-MYC

The role of the nuclear phosphoprotein c-Myc has been examined with re spect to the regulation of 1-beta-D-arabinofuranosylcytosine (ara-C)-i nduced apoptosis in human leukemia cells exposed to bryostatin 1 and o ther pharmacologic protein kinase C (PKC) activators. Pretreatment of HL-60 cells fur 24 hr with 10 nM bryostatin 1 significantly potentiate d the ability of ara-C (10 mu M; 6 hr) to induce apoptosis without red ucing the expression of c-Myc protein. In contrast, equivalent exposur e to the stage 2 tumor-promoting PKC activator mezerein (10 nM) in con junction with ara-C reduced c-Myc levels by 87% and failed to potentia te apoptosis. Co-administration of bryostatin 1 with mezerein before a ra-C prevented down-regulation of c-Myc and augmented cell death, wher eas co-treatment with the calcium ionophore A23187 (250 nM) and bryost atin 1 reduced c-Myc levels by 80% and abrogated the increase in ara-C -induced apoptosis. When cells were exposed for 24 hr to a c-myc antis ense oligonucleotide (AS-ODN; 10 mu M) but not to a scrambled sequence ODN (SS-ODN) prior to ara-C, c-Myc expression was reduced by 81%, and apoptosis and cell viability were unperturbed. However, AS-ODN (but n ot SS-ODN) reduced c-Myc protein in cells pre-exposed to bryostatin 1 by 74% and abrogated potentiation of ara-C-induced apoptosis. The acti ons of c-myc AS-ODN did not stem from proximal G(1) arrest/diffrrentia tion or biochemical events, since they were not associated with a redu ction in the S-phase cell fraction, p21(WAF1/CIP1) induction, pRb hypo phosphorylation, or alterations in ara-C metabolism. Together, these f indings indicate that HL-60 cell apoptosis proceeds by both c-Myc-depe ndent and -independent pathways, and that only the former are involved in the potentiation of ara-C-mediated cell death by bryostatin 1. (C) 1997 Elsevier Science Inc.