Abstract
2‐Methoxyestradiol (2‐ME), an endogenous metabolite of 17β‐estradiol, is present in human blood and urine. Here we show for the first time that 2‐ME significantly inhibited the growth of normal prostate epithelial cells and androgen‐dependent LNCaP and androgen‐independent DU145 prostate cancer cells. This growth inhibition was accompanied by a twofold increase in the G~2~/M population, with a concomitant decrease in the G~1~ population, as shown by cell‐cycle analysis. 2‐ME treatment affected the cell‐cycle progression of prostate cancer cells specifically by blocking cells in the G~2~ phase. Immunoblot analysis of the key cell‐cycle regulatory proteins in the G~2~/M phase showed a 14‐fold increase in the expression of p21 and an eightfold increase in the expression of p34 cell division cycle 2 (cdc2). We also found an accumulation of phosphorylated cdc2 after 2‐ME treatment. Furthermore, Wee 1 kinase was detectable after 2‐ME treatment. 2‐ME treatment also led to an increase in the activity of caspase‐3, followed by apoptosis, as shown by terminal deoxynucleotidyl transferase–mediated deoxyuridine 5‐triphosphate–biotin nick end‐labeling and fluorescein isothiocyanate–poly(ADP‐ribose) polymerase assay. Estrogen receptor levels did not change after treatment with 2‐ME. Examination of the signaling pathways that mediate 2‐ME–induced apoptosis showed reduction in the level of p53 expression and its DNA‐binding activity. Given the fact that p53 mutations are common in patients with metastatic prostate cancer, our finding that 2‐ME–mediated growth inhibition of human prostate cancer cells occurred in a p53‐independent manner has considerable clinical significance. These findings, combined with the limited toxicity of 2‐ME, may have significant implications for alternative treatment of advanced prostate cancer. © 2001 Wiley‐Liss, Inc.