Abstract
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Purpose: In prostate carcinomas, gene amplifications (e.g. c-MYC, HER2/NEU, androgen receptor gene) have been found and being related to progressive tumor growth. Using comparative genomic hybridization (CGH), we recently detected DNA copy number gains at chromosome 3q25-q27 in 35% of analyzed tumor samples. In the present study, we set out to further characterize this amplicon by cytogenetic and molecular genetic techniques and to identify a probable target gene. Material and Methods: Prostate carcinoma cell lines (PC3, DU145, DU145MN1) and 22 tumor samples from radical prostatectomy specimens were screened for chromosomal aberrations with CGH. To narrow down the amplification unit, 26 overlapping BAC clones covering the 3q amplicon were hybridized on the prostate carcinoma cell lines. DNA copy number changes of 6 candidate genes (hTERC, SAMD7, TLOC1/SEC62, SKIL, SLC2A2, PIK3CA) located within the BAC clones with the highest copy-number status were analyzed using a modified quantification assay based on the LightCycler™ system. Additionally, mRNA expression levels of these genes were measured using relative real time PCR quantification. Results: The 3q amplicon could be narrowed down to 700 kb at 3q26.2. Positional and functional candidates revealed the following amplification frequencies in the primary tumor samples: hTERC 9%, SAMD7 32%, TLOC1/SEC62 59%, SKIL 0%, SLC2A2 27%, PIK3CA 9%. Interestingly, TLOC1/SEC62 with the highest amplification frequency showed a 3-14 fold overexpression in the 13 prostate carcinomas analyzed compared to normal prostate fibroblasts. TLOC1/Sec62-p was also demonstrated to be overexpressed by western blot analysis. Conclusions: Using molecular cytogenetic techniques, we identified TLOC1/SEC62 as the best candidate within the 3q amplification unit in prostate cancer. Intriguingly, the TLOC1/SEC62 gene copy number was increased in prostate tumors from patients, who had a lower risk of and a longer time to progression following radical prostatectomy. TLOC1/Sec62 protein is a component of the endoplasmic reticulum and is supposed to be active in a dual manner as part of the protein translocase complex and as a calcium ion channel. Supported by Deutsche Krebshilfe: 70-2936 Wu I
[Proc Amer Assoc Cancer Res, Volume 47, 2006]