Fig. S2.

YK-4-279 induces mitotic arrest and apoptosis. (A) Quantification of percentage of cells at various phases of cell cycle after treatment with DMSO or YK-4-279 for 24 h. Error bars represent ±SD of three independent replicates. **P < 0.01, ***P < 0.001. Unpaired Student t-test was used to assess the significance of difference between the control and treated samples. (B) SK-N-BE(2)-C and CHP-212 were treated with DMSO (D) or two doses of YK-4-279 for 24 h and cell lysates were harvested in RIPA buffer. Blots were probed for several cell cycle and cell death markers. All neuroblastoma cell lines showed increase of Histone H3 (Ser10), cleaved PARP and active caspase-3. Actin was used as a loading control. (C) Immunoblots demonstrating that YK-4-279-induced cell death of SK-N-AS and GIMEN cells is rescued by QVD treatment. Note decreased cleaved caspase-3 and cleaved PARP after QVD treatment. (D) Immunoblotting for acetyl-α-tubulin (Lys40) (indicator of tubulin microtubule stabilization) and phospho-histone H3(ser10) (pHH3). α-tubulin and actin were used as loading controls. (E) Knockdown of wildtype p53 in GIMEN and assessment of YK-4-279 activity. Cells were reverse transfected with either control siRNA or p53 siRNA. After 72 h of incubation whole cell lysates were harvested for western analysis and probed for mitotic and apoptotic markers.