butyrolactone-i and Prostatic-Neoplasms

Cardiac digitalis has been considered to be a treatment for breast cancer. Our previous study indicates that digoxin, one member in digitalis, decreases the proliferation of prostate cancer cells, but the mechanisms remain unclear. In the present study, Ca(2+) proved to be an important factor in digoxin-triggered prostate cancer cell death. Because cyclin-dependent kinase (Cdk)5 and p35 cleavage (p25 formation) have been reported to be targets of intracellular Ca(2+), and subsequently correlated to apoptosis, we not only demonstrated first that Cdk5, p35, and p25 proteins were all expressed in prostate cancer cells (including lymph node carcinoma of the prostate (LNCaP) and DU-145 cells), but also showed where p25 formation and Cdk5 kinase activity were affected by treatment with digoxin. The inhibitor of p35 cleavage (calpeptin) was used to reduce p25 formation, and the result suggested that p25 accumulation might be the major cause of digoxin-triggered LNCaP cell death. Butyrolactone-I and roscovitine, two Cdk5 kinase inhibitors, were also found to prevent digoxin-triggered LNCaP cell death. In addition, treatment of siRNA-Cdk5 diminished digoxin-triggered cell death, as compared with the treatments of siRNA-Cdk1 or siRNA-Cdk2, which implies the specific involvement of Cdk5 in digoxin-triggered cell death. Caspase inhibitor set and terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling assay were used to demonstrate that digoxin-triggered LNCaP cell apoptosis through Cdk5 activation. These results suggest that Cdk5/p35 and p25 are novel players in digoxin-triggered prostate cancer cell apoptosis and, therefore, become potential therapeutic targets.

Topics: 4-Butyrolactone; Apoptosis; Calcium; Caspase Inhibitors; Caspases; Cell Line, Tumor; Cyclin-Dependent Kinase 5; Cyclin-Dependent Kinases; Digoxin; Dipeptides; Enzyme Activation; Enzyme Inhibitors; Humans; Male; Nerve Tissue Proteins; Prostatic Neoplasms; Purines; RNA, Small Interfering; Roscovitine

Several naturally occurring cyclin-dependent kinase (CDK) inhibitors have been isolated from different lower organisms. In this report, we examined the effect of one of the CDK inhibitors, butyrolactone I (BL), on the expression of cyclins D2, A and B1 in three human prostatic cancer cell lines (DU145, PC-3, LNCaP) using two colored flow cytometric analysis. The percentage of DU145 cells in the 4C phase of the cell cycle were increased significantly at both 70 microM and 100 microM BL. Furthermore, an additional 8C peak was observed which had double the DNA content of the 4C phase at these concentrations of BL. The appearance of the 8C peak increased gradually and was more evident in DU145 and PC-3 than LNCaP. Cells in the 8C peak had either two nuclei or abnormal nuclei as observed by Papanicolaou stain. BL also increased the amount of cyclin B1 positive cells in the 4C phase. This increase was apparent on day 1 and returned to normal by day 3. Since BL selectively inhibits cyclin-dependent kinase, cyclin B1 might accumulate without being degraded. Other cyclins were not significantly changed by BL. The data demonstrate that BL inhibited Cdc2 of unsynchronized cultured prostate cancer cells, and interrupted the cell cycle progression toward cell division. The BL inhibition of Cdc2 led to the accumulation of cells in the 4C phase without mitosis resulting in an accumulation of cyclin B1. The appearance of cells in the 8C phase may be due to the progression of cells in the 4C phase through the cell cycle skipping mitosis. Cyclin B1 decreased in correlation with the progression through a new cell cycle. These results suggest that BL does not cause a complete arrest of the cell cycle in G2/M but that BL occasionally allows for the skipping of mitosis and subsequent progression through the cell cycle to occur.

Topics: 4-Butyrolactone; Cell Survival; Cyclin B; Cyclin B1; Cyclin-Dependent Kinases; DNA, Neoplasm; Enzyme Inhibitors; G2 Phase; Humans; Male; Mitosis; Prostatic Neoplasms; Tumor Cells, Cultured