Figure 3

GUTK inhibited colony forming and tumorigenic capacity following induction of cell cycle re-entry. Quiescent LNCaP and PC-3 cells (50 cells/well) were induced to re-enter the cell cycle in the presence or absence of GUTK at GI₂₅, GI₅₀ and GI₇₅ for 24, 48 and 72 h. Thereafter, the cells were cultured in the fresh full medium for additional 2 weeks. Emerging cell colonies were fixed, stained with crystal violet and imaged for counting. Representative image and quantification data of mean±S.D. of three independent experiments in LNCaP (a and b) and PC-3 (c and d). *P<0.05, **P<0.01, ***P<0.001 compared with the respective vehicle control. For in vivo analysis, 5-week-old male BALB/c nude mice were randomly divided into two groups (5 mice per group). Quiescent PC-3 cells were induced to re-enter the cell cycle by plating at a low density and treated either with GUTK at GI₇₅ or with DMSO control for 72 h. The pre-treated cells were then subcutaneously injected into the left flank of male nude mice (day 1) and monitored for tumor formation. Tumor volume (e) and animal body weight (f) were measured every second day. At day 31, the tumors were resected, photographed (g) and weighed (h). The resected tumors were stained for H&E (i) and Ki-67 (j). Scale bar=20 μm. All data are expressed as mean±S.D., *P<0.05, **P<0.01 compared with the DMSO pre-treated control