Fig. 6.

Effect of GWARJD10 on apoptosis marker (Cleaved PARP) and cell cycle marker (Cyclin D1) in androgen-dependent LNCaP cells and its castrate-resistant variant C4-2 cells. LNCaP or C4-2 cells were treated with (+) or without (−) 2 nmol/l androgen R1881 and zero (C), 1 or 5 μmol/l of lead compound GWARJD10 for 48 and 72 hr. Cells were then harvested and protein levels were determined by Western blot analysis. β-actin was used as loading control and normalization. Percentage of each protein band compared to C (%) at each timepoint was calculated after normalizing each band against β-actin. Representative blots are shown. The level of Cleaved PARP (89 kDa) did not change under treatment conditions. Cyclin D1 (37 kDa) level was markedly reduced in R1881 treated LNCaP and C4-2 cells after 72 hr. The western blots were run three times for cell lysates from three to four different experiments per condition with similar results. Statistical analysis of bands across the experiments showed significantly lower (^*) average cyclin D1 of 65% in LNCaP cells (P = 0.001, N = 3) and 71% in C4-2 cells (P = 0.05, N = 4) compared to control.