Figure 1

Caspase8 inhibition compounds cytotoxicity in ovarian cancer cells treated with IKKβ inhibitor. Caspase8 shRNA toxicity in a sensitization library screen is shown as (a) the log2 ratio of untreated versus IKKβ-inhibited, or (b) log 10 P-value, after 10 days. Caspase8 shRNAs are highlighted in black. (c) Cell viability of Ovcar3, Caov3 and PEO1 cells transduced with control shRNA or either of two different Caspase8 shRNAs was measured by XTT after 7 days of exposure to increasing concentrations of IKKβ inhibitor. Data are shown as fold control shRNA, in the absence of IKKβ inhibitor (DMSO), ±S.E.M., n=8. Asterisks indicate significant differences (P<0.01) at the concentration range used in subsequent studies. (d) Ovcar3 cells were transduced with Caspase8 shRNAs #1, #2, #3 or #4 or control shRNA and exposed to 2.5 μM IKKβ inhibitor or vehicle for 7 days. Viability was measured by XTT and is shown as fold control shRNA and drug control (DMSO). Error bars represent S.E.M., n=8; *P<0.001. (e) Ovcar3 cells expressing control or Caspase8 shRNA #2 were treated with 2.5 μM IKKβ inhibitor for 7 days. (f) Three ovarian cell lines sensitive to IKKβ inhibition (Ovcar3, Caov3 and Igrov1) and one insensitive cell line (Ovcar8) were stained by IHC with NF-κB-p65 and Caspase8 antibodies and the presence of nuclear protein analyzed. IHC scores for relative amounts of nuclear NF-κB-p65 and Caspase8 were measured in four to six uniform fields per sample and averaged. Numbers are average scores ±S.E.M.