Inhibition of protein kinase C activator-mediated induction of p21(CIP1) and p27(KIP1) by deoxycytidine analogs in human leukemia cells - Relationship to apoptosis and differentiation

Events accompanying sequential exposure of U937 leukemic cells to the deoxy cytidine (dCyd) analogs 1-[beta-D-arabinofuranosyl]cytosine (ara-C) or 2',2 '-difluorodeoxycytidine (gemcitabine; dFdC) followed by two protein kinase C (PKC) activators [bryostatin 1 (BRY) or phorbol 12'-myristate 13'-acetate (PMA)] exhibiting disparate differentiation-inducing abilities were charac terized. A 24-hr exposure to 10 nM BRY or PMA after a 6-hr incubation with 1 mu M ara-C or 100 nM dFdC resulted in equivalent increases in apoptosis, caspase-3 activation, and polyADP-ribose polymerase degradation, as well as identical DNA cleavage patterns. BRY and PMA did not modify retention of t he lethal ara-C metabolite ara-CTP or alter ara-CTP/dCTP ratios. Unexpected ly, pretreatment of cells with ara C or dFdC opposed BRY- and PMA-related. induction of the cyclin-dependent kinase inhibitors (CDKIs) p21(CIP1) and/o r p27(KIP1). These effects were not mimicked by the DNA polymerase inhibito r aphidicolin or by VP-16, a potent inducer of apoptosis. Inhibition of PKC activator-induced CDKI expression by ara-C and dFdC did not lead to redist ribution of proliferating cell nuclear antigen but was accompanied by sub-a dditive or antagonistic effects on leukemic cell differentiation. Sequentia l exposure of cells to ara-C followed by BRY or PMA led to substantial redu ctions in clonogenicity that could not be attributed solely to apoptosis. F inally, pretreatment of cells with ara-C attenuated PMA- and BRY-mediated a ctivation of mitogen-activated protein kinase, an enzyme implicated in CDKI induction. Collectively, these findings suggest that pretreatment of leuke mic cells with certain dCyd analogs interferes with CDKI induction by the P KC activators PMA and BRY, and that this action may contribute to modulatio n of apoptosis and differentiation in cells exposed sequentially to these a gents. BIOCHEM PHARMACOL 58;1:121-131, 1999. (C) 1999 Elsevier Science Inc.