Figure 3

AA005 induces cell death through caspase-3-independent mechanisms. (A) SW620 cells were treated with or without 1 μM AA005 for the indicated times. Treatment with 20 μM camptothecin for 36 h was used as positive control. Proteins PARP-1, and caspase-3 were detected by western blot, standardized to actin. ΔPARP-1, and ΔCaspase-3 indicate cleaved 85-kDa fragment of PARP-1 and active Caspase-3 fragment proteins, respectively. (B) SW620 cells were preincubated with 100 μM Z-VAD.fmk for 1 h and then treated with or without 1 μM AA005 for 48 h, and with 20 μM camptothecin for 36 h. Annexin-V/PI double-stained cells and cell death were measured by flow cytometry. Results show mean ± S.D. *P < 0.05, versus camptothecin-treated group. (C, D) NB4 cells were preincubated with 40 μM Z-VAD.fmk for 1 h and then treated with or without 1 μM camptothecin (C), or 1 μM AA005 (D), for indicated times. Annexin-V/PI double-stained cells and dead cells were measured on flow cytometry. Values show mean ± S.D. of triplicates in an independent experiment.