Overexpression of the polycomb group protein enhancer of zeste homologue 2 
(EZH2) has been found in several malignancies, including prostate cancer, with 
an aggressive phenotype. Amplification of the gene has previously been 
demonstrated in several malignancies, but not in prostate cancer. Our goal was 
to evaluate the gene copy number and expression alterations of EZH2 in prostate 
cancer. The copy number of EZH2 in cell lines (LNCaP, DU145, PC-3, 22Rv1), 
xenografts (n = 10), and clinical tumors (n = 191) was studied with fluorescence 
in situ hybridization. All cell lines had a gain of EZH2. Eight of the ten 
xenografts showed an increased copy number of the gene, including one case of 
high-level amplification (>or=5 copies of the gene and EZH2/centromere ratio 
>or=2). 34/125 (27%) of untreated prostate carcinomas showed increased copy 
number, but only one case of low-level amplification (>or=5 copies of the gene 
and EZH2/centromere ratio <2), whereas half (25/46) of the hormone-refractory 
carcinomas showed increased copy number, including seven cases of low-level 
amplification and three cases of high-level amplification (P < 0.0001). 
Expression of EZH2 was significantly (P = 0.0009) higher in hormone-refractory 
prostate cancer compared with that in benign prostatic hyperplasia or untreated 
cancer, according to quantitative real-time RT-PCR assay. Also, the expression 
of EZH2 protein was found to be higher in hormone-refractory tumors than in 
hormone-naïve tumors by immunohistochemistry. The EZH2 gene amplification was 
significantly (P < 0.05) associated with increased EZH2 protein expression. The 
data show that amplification of the EZH2 gene is rare in early prostate cancer, 
whereas a fraction of late-stage tumors contains the gene amplification leading 
to the overexpression of the gene, thus indicating the importance of EZH2 in the 
progression of prostate cancer.