Investigation of the role of t(4;6) and 6q15 deletion in prostate cancer development and progression

Abstract

The recent identification of TMPRSS2:ETS gene fusions highlighted the importance of chromosomal rearrangement and fusion gene development in prostate tumourigenesis. We previously reported a recurrent translocation, t(4;6), in prostate cancer (PCa) with the breakpoints identified at 4q22 and 6q15 in the LNCaP cell line. A small deletion adjacent to the 6q15 breakpoint was found, which is consistent with the frequently lost chromosome region previously reported and detected by our micro-array analysis. Here I accessed the prevalence of the t(4;6)(q22;q15) and the 6q15 deletion and also looked for relevant candidate tumour suppressor genes (TSGs) in PCa. Using fluorescence in situ hybridisation (FISH) analysis, t(4;6)(q22;q15) was detected in 78 of 667 clinical, localised PCa samples. Statistical analysis showed it was not independently associated with patient outcome but occurred more frequently in high clinical T stage, high tumour volume specimens and in those with high baseline PSA (prostate-specific antigen) (P=0.001, 0.001 and 0.01 respectively). We hypothesised that both t(4;6)(q22;q15) and the 6q15 deletion contribute to the inactivation of TSGs at 6q15, which are associated with PCa development and progression. Therefore I investigated the TSGs located in this region. Using FISH analysis, this deletion was confirmed in 46% (13 in 28) of the PCa samples. Using Exon array to systematically detect inactivated genes in this region, four candidate genes, CNR1, PNRC1, GJA10 and BACH2 were identified with common down-regulation. The real-time PCR analysis validated these exon array results. However, with additional controls and clinical cancer samples, only two genes (CNR1 and BACH2) still showed significant down-regulation. CNR1 protein expression was absent in 76.8% (43 in 56) of PCa cases whereas its 5 expression was shown in 83.9% (26 in 31) of benign prostatic hyperplasia (BPH) samples as demonstrated by immunohistochemistry (IHC) analysis (p<0.001). In contrast, cytoplasmic expression of BACH2 was slightly less in the examined PCa cases (14.0%, 12 in 86) compared with BPH samples (6.9%, 2 in 29). However, nuclear expression of BACH2 was significantly higher in prostate cancer cases (45.3%) than it was in BPH samples (10.3%) (p=0.001). Five of the 43 PCa samples without CNR1 expression were selected for sequencing and one of the five samples had insertion in the CNR1 genomic DNA. The cellular function study of PCa cell lines indicated that the CNR1 might function as a tumour suppressor involved in cell proliferation and invasion. In conclusion, both t(4;6)(4q22;6q15) and the 6q15 deletion are frequent events and the gene CNR1 is a candidate TSG at this deletion region in PCa.