Abstract
Mitoxantrone, a new anthraquinone, showed inhibitory an effect on protein kinase C (PKC) activity. Its IC50, value was 4.4μg/ml (8.5μM), which is much lower than those of the well-known anthracyclines daunorubicin and doxorubicin, the IC50 values of which are more than 100μg/ml (>170μM). Kinetic studies demonstrated that mitoxantrone inhibited PKC in a competitive manner with respect to histone H1, and its K1 value was 6.3μM (K1 values of daunorubicin and doxorubicin were 0.89 and 0.15mM, respectively), and in a non-competitive manner with respect to phosphatidylserine and ATP. Inhibition of phosphorylation by mitoxantrone was observed with various substrates including S6 peptide, myelin basic protein and its peptide substrate derived from the amino-terminal region. Their IC50, values were 0.49μg/ml (0.95μM), 1.8μg/ml (3.5μM), and 0.82μg/ml (1.6μM), respectively. Mitoxantrone did not markedly inhibit the activity of cyclic AMP-dependent protein kinase, casein kinase I or casein kinase II, at concentrations of less than 10μg/ml. On the other hand, brief exposure (5min) of HL60 cells to mitoxantrone caused the inhibition of cell growth with an IC50, value of 52ng/ml (0.1μM). In HL60 cells, most of the PKC activity (about 90%) was detected in the cytosolic fraction. When HL60 cells exposed to 10μg/ml mitoxantrone for 5min were observed with fluorescence microscopy, the fluorescence elicited from mitoxantrone was detected in the extranuclear area. These results indicated that mitoxantrone is a potent inhibitor of PKC, and this inhibition may be one of the mechanisms of antitumor activity of mitoxantrone.
