Activation of caspase-3 and regulation of Bcl-2 protein family members in response to ErPC3-treatment and irradiation. PC3 and LNCaP cells were treated with 0-25 µM ErPC3 or irradiated with 2 or 10 Gy. Cells lysates were generated 48 h after treatment, separated by electrophoresis, and protein expression was subsequently analyzed by western blotting. Both cells lines showed a concentration-dependent activation of caspase-3 in response to ErPC3-treatment (A, B). In PC3 cells, cleavage of the caspase-3 substrate PARP could already be detected after treatment with 12.5 µM ErPC3; PARP-cleavage was accompanied by a weak activation of caspase-3 detectable upon treatment with 12.5 µM ErPC3 (A). A weak cleavage of caspase-3 and PARP was also observed when LNCaP cells were treated with 12.5 µM ErPC3, but cleavage was clearly visible after treatment with 25 µM ErPC3 (B). No caspase-3 activation and PARP cleavage was observed in response to ionizing radiation. No change of protein levels of the pro-apoptotic Bak and Bax and the anti-apoptotic Bcl-xL was observed upon irradiation or in response to treatment with ErPC3 (C, D). A slight reduction in the levels of antiapoptotic Bcl-2 was observed upon irradiation in LNCaP and PC3 cells, whereas treatment with ErPC3 reduced the levels of the anti-apoptotic Mcl-1 in LNCaP cells. However, the changes of Mcl-1 expression levels did not correlate with the sensitivity of LNCaP cells to ErPC3.