Pharmacological inhibitors of the mitogen-activated protein kinase (MAPK) kinase/MAPK cascade interact synergistically with UCN-01 to induce mitochondrial dysfunction and apoptosis in human leukemia cells

interactions between the checkpoint abrogator UCN-01 and several pharmacolo gical inhibitors of the mitogen-activated protein kinase (MAPK) kinase (MEK )/MAPK pathway have been examined in a variety of human leukemia cell lines . Exposure of U937 monocytic leukemia cells to a marginally toxic concentra tion of UCN-01 (e.g., 150 nM) for 18 h resulted in phosphorylation/activati on of p42/44 MAPK. Coadministration of the MEK inhibitor PD184352 (10 muM) blocked UCN-01-induced MAPK activation and was accompanied by marked mitoch ondrial damage (e.g., cytochrome c release and loss of Delta Psi (m)), casp ase activation, DNA fragmentation, and apoptosis. Similar interactions were noted in the case of of her MEK; inhibitors (e.g., PD98059; U0136) as well as in multiple other leukemia cell types (e.g., HL-60, Jurkat, CCRF-CEM, a nd Raji). Coadministration of PD184352 and UCN-01 resulted in reduced bindi ng of the cdc25C phosphatase to 14-3-3 proteins, enhanced dephosphorylation /activation of p34(cdc2) and diminished phosphorylation of cyclic AMP-respo nsive element binding protein. The ability of UCN-01, when combined with PD 184352, to antagonize cdc25C/14-3-3 protein binding, promote dephosphorylat ion of p34(cdc2), and potentiate apoptosis was mimicked by the ataxia telan gectasia mutation inhibitor caffeine. In contrast, cotreatment of cells wit h UCN-01 and PD184352 did not substantially increase c-Jun-NH2-terminal kin ase activation nor did if alter expression of Bcl-2, Bcl-x(L), Bar, or X-in hibitor of apoptosis. However, coexposure of U937 cells to UCN-01 and PD184 352 induced a marked increase in p38 MAPK activation. Moreover, SB203580, w hich inhibits multiple kinases including p38 MAPK, partially antagonized ce ll death. Lastly, although UCN-01 c PD184352 did not induce p21(CIP1), stab le expression of a p21(CIP1) antisense construct significantly increased su sceptibility to this drug combination. Together, these findings indicate th at exposure of leukemic cells to UCN-01 leads to activation of the MAPK; ca scade and that interruption of this process by R MEK inhibition triggers pe rturbations in several signaling and cell cycle regulatory pathways that cu lminate in mitochondrial injury, caspase activation, and apoptosis. They al so raise the possibility. that disrupting multiple signaling pathways, e.g. , by combining UCN-01 with MEK inhibitors, may represent a novel antileukem ic strategy.