E2F3 is the main target gene of the 6p22 amplicon with high specificity for human bladder cancer

Abstract

Amplification of 6p22 occurs in about 10-20% of bladder cancers and is associated with enhanced tumour cell proliferation. Candidate target genes for the 6p22 amplicon include E2F3 and the adjacent gene NM_017774. To clarify which gene is representing the main target, we compared the prevalence of the amplification and the functional role of both genes. Amplification of E2F3 and NM_017774 was analysed by fluorescence in situ hybridization on a bladder cancer tissue microarray composed of 2317 cancer samples. Both genes showed amplification in 104 of 893 (11.6%) interpretable tumours and were exclusively found co-amplified. Additional gene expression analysis by real-time polymerase chain reaction in 12 tumour-derived cell lines revealed that amplification of 6p22 was always associated with co-overexpression of E2F3 and NM_017774. Furthermore, RNA interference was used to study the influence of reduced gene expression on cell growth. In tumour cells with and without the 6p22 amplicon, knockdown of E2F3 always lead to unequivocal reduction of proliferation, whereas knockdown of NM_017774 was only capable to slow down cell proliferation in non-amplified cells. Our findings point out that E2F3 but not NM_017774 is driving enhanced proliferation of 6p22 amplified tumour cells. We conclude that E2F3 must be responsible for the growth advantage of 6p22 amplified bladder cancer cells.

Evidence that Igf2 down-regulation in postnatal tissues and up-regulation in malignancies is driven by transcription factor E2f3

“…To test this prediction, we analyzed publicly available expression microarray data from three human malignancies that overexpress E2F3: metastatic prostate cancer (30,31), urinary bladder cancer (32)(33)(34), and Wilms tumor (35). All three types of cancer showed increased E2F3 expression, which is consistent with prior publications (30)(31)(32)(33)(34)(35), and also showed elevated IGF2 levels (Fig. 5).…”

“…Interestingly, E2F3 is the only E2F member for which amplification and overexpression in various types of cancers has been demonstrated (30)(31)(32)(33)(34)(35). More commonly, increased E2F activity results because the malignant cells undergo loss of retinoblastoma protein (RB), which binds to and suppresses E2F activity, or because of mutations in other genes that modulate the phosphorylation and degradation of RB (29).…”

Transcriptomic characterization of fibrolamellar hepatocellular carcinoma

“…The activity of AURKA is significantly enhanced by the PKA-dependent phosphorylation on threonine 288 (81). Other oncogenes that affect the cell cycle that are increased in FLHCC include CCNE1, whose amplification is considered a driver of ovarian cancers (82); CDK6, which has been implicated in tumorigenesis both through phosphorylation of Rb (83) and a phosphorylation-independent effect on proliferation (84); E2F3, which is required for normal cell proliferation (85); and overexpression of E2F3, which has been associated with breast (86), prostate (87), and bladder cancer (88). Our results are consistent with some previous reports on gene expression associated with FLHCC (SI Appendix, Table S14).…”

The origins of bladder cancer

“…56 E2F3a is the predominant binding partner for RB in G0. Overexpression of E2F3 and a second gene from the 6p22 region, CKDAL1, has been documented in bladder tumor tissue microarrays 57 and in bladder cancer cell lines. 58 Hurst 59 has recently demonstrated that there is a functional link between E2F3 overexpression and proliferative advantage in bladder tumor cells with 6p22 amplification; knockdown of CDKAL1 had no effect on cell proliferation.…”