FIG. 9.

Caspase-3 or caspase-9 knockdown modestly but significantly protects cells from sorafenib-mediated lethality. (A) Protein lysates were prepared from two clones of U937 cells transfected with caspase-9 shRNA (casp9-shRNA3 and casp9-shRNA8) and from GFP-shRNA-transfected cells and subjected to Western blot analysis (upper). Alternatively, these cells were treated with 10 μM sorafenib (Sor) or 2.5 μM VP16 for 24 h, after which the extent of apoptosis was determined using an annexin V staining assay (lower). (B) Protein lysates were prepared from two clones of U937 cells transfected with caspase-3 shRNA (casp3-shRNA10 and casp3-shRNA11) and from GFP-shRNA-transfected cells and subjected to Western blot analysis (upper). Alternatively, these cells were exposed to sorafenib (10 μM) and VP16 (2.5 μM) for 24 h and then subjected to an annexin V staining assay (lower). (C and D) Casp4-shRNA3 (C), casp9-shRNA8 (D), and GFP-shRNA cells were treated with sorafenib (10 μM) for the designated intervals or with thapsigargin (Tg; 0.5 μM) for 16 h, after which protein lysates were prepared and subjected to Western blot analysis. *, significantly less than values for controls (P < 0.05). C, control; Tub, antitubulin antibody.