JNK inhibitor SP600125 promotes the formation of polymerized tubulin, leading to G2/M phase arrest, endoreduplication, and delayed apoptosis

ABSTRACT

The JNK inhibitor SP600125 strongly inhibits cell proliferation in many human cancer cells by blocking cell-cycle progression and inducing apoptosis. Despite extensive study, the mechanism by which SP600125 inhibits mitosis-related effects in human leukemia cells remains unclear. We investigated the effects of SP600125 on the inhibition of cell proliferation and the cell cycle, and on microtubule dynamics in vivo and in vitro. Treatment of synchronized leukemia cells with varying concentrations of SP600125 results in significant G2/M cell cycle arrest with elevated p21 levels, phosphorylation of histone H3 within 24 h, and endoreduplication with elevated Cdk2 protein levels after 48 h. SP600125 also induces significant abnormal microtubule dynamics in vivo. High concentrations of SP600125 (200 microMeter) were required to disorganize microtubule polymerization in vitro. Additionally, SP600125-induced delayed apoptosis and cell death was accompanied by significant poly ADP-ribose polymerase (PARP) cleavage and caspase-3 activity in the late phase (at 72 h). Endoreduplication showed a greater increase in ectopic Bcl-2-expressing U937 cells at 72 h than in wild-type U937 cells without delayed apoptosis. These results indicate that Bcl-2 suppresses apoptosis and SP600125-induced G2/M arrest and endoreduplication. Therefore, we suggest that SP600125 induces mitotic arrest by inducing abnormal spindle microtubule dynamics.