APOPTOSIS IN 7-HYDROXYSTAUROSPURINE-TREATED T-LYMPHOBLASTS CORRELATESWITH ACTIVATION OF CYCLIN-DEPENDENT KINASE-1 AND KINASE-2

7-Hydroxystaurosporine (UCN-01) is a potent inhibitor of protein kinas e C (PKC) isozymes alpha, beta, and gamma [Seynaeve et al., Mel. Pharm acol, 45: 1207-1214, 1994] that also has antitumor effects in vivo. To determine whether inhibition of PKC can be related to inhibition of c ell growth with induction of apoptosis, we compared the effects of UCN -01 to those of the highly selective bisindolylmaleimide PKC antagonis t CF 109203X in leukemic T-cell lines. Both compounds potently inhibit ed PKC activity when added to T-cell membrane preparations and reverse d phorbol ester-induced c-fos gene expression in intact cells. However , whereas UCN-01 potently inhibited growth of Jurkat, Molt-3, Molt-4, and Hut-78 cells (IC50 = 20-65 nM, irreversible after 24 h of exposure ), GF 109203X had IC(50)s for cell growth of 3.6-5.0 mu M. Less than 3 h after addition, UCN-01 but not CF 109203X-treated cells displayed l oss of cells with G(2)-M DNA content, appearance of a hypodiploid DNA fraction, and evidence of internucleosomal DNA fragmentation. Six h af ter treatment, cells appeared to accumulate with S-phase DNA content. These effects correlated with selective UCN-01 but not CF 109203X-indu ced decrease in total and tyrosine phosphorylation of cyclin-dependent kinases (cdks) 1 and 2, and with increases in the histone H1 kinase a ctivities of cdk1 and cdk2. UCN-01 was relatively less potent in inhib ition of properly activated cdk1 and cdk2 when added in vitro to H1 ki nase assays (IC50 = 1000 and 600 nM, respectively). We conclude that i nhibition of PKC alone is not sufficient to account for the actions of UCN-01 and are led to the hypothesis that inappropriate cdk activatio n either correlates with or actually mediates cell growth inhibition w ith apoptosis in T lymphoblasts exposed to UCN-01.