R-568 reduces cell viability in prostate cancer cells. A&B Cells were seeded in 96-well plates overnight and then treated with R-568 or S-568 at the indicated doses. Control cells received no treatment. After 48 h, viable cells were determined using a MTT assay kit (Sigma, St Louise, MO). The average values of optical densities from each group were presented. Data represents three separate experiments. The red dotted line indicates the IC50 value. C Cells were plated in 12-well plates and treated with R-568 at the indicated doses for 48 h. The control cells received no treatment. Cells were harvested at the end of experiment and stained in 0.4% trypan blue solution. The dead (blue) cells were counted and the average of death rate in each well was presented. D PC-3 cells were plated in 6-well plates and then transfected with negative control siRNA or CaSR siRNA at 100 μM final concentration in the culture media. Two days later, cells were treated with the solvent or R-568 (50 μM) for 48 hours. Cell death rate was assessed using trypan blue exclusion assay as described earlier. INSERT: Two days after the siRNA transfection, PC-3 cells were treated with or without R-568 for 48 h. Cell lysates were subjected to Western blot for assessing CaSR protein levels. Actin blot served as protein loading control. Data represents three different experiments. The asterisk indicates a significant difference (P < 0.05, Student t-test) between R-568 treatment and the control.