Canonical Notch signalling is inactive in urothelial carcinoma

Abstract

BackgroundNotch signalling regulates cell fate in most tissues, promoting precursor cell proliferation in some, but differentiation in others. Accordingly, downregulation or overactivity variously contributes to cancer development. So far, little is known about Notch pathway activity and function in the normal urothelium and in urothelial carcinoma (UC). We have therefore investigated expression of Notch pathway components in UC tissues and cell lines and studied the function of one receptor, NOTCH1, in detail.MethodsExpression of canonical Notch pathway components were studied in UC and normal bladder tissues by immunohistochemistry and quantitative RT-PCR and in UC cell lines and normal cultured urothelial cells by qRT-PCR, immunocytochemistry and Western blotting. Pathway activity was measured by reporter gene assays. Its influence on cell proliferation was investigated by γ-secretase inhibition. Effects of NOTCH1 restoration were followed by measuring cell cycle distribution, proliferation, clonogenicity and nuclear morphology.ResultsNOTCH1 and its ligand, DLL1, were expressed at plasma membranes and in the cytoplasm of cells in the upper normal urothelium layer, but became downregulated in UC tissues, especially in high-stage tumours. In addition, the proteins were often delocalized intracellularly. According differences were observed in UC cell lines compared to normal urothelial cells. Canonical Notch pathway activity in reporter assays was repressed in UC cell lines compared to normal cells and a mammary carcinoma cell line, but was induced by transfected NOTCH1. Inhibitors of Notch signalling acting at the γ-secretase step did not affect UC cell proliferation at concentrations efficacious against a cell line with known Notch activity. Surprisingly, overexpression of NOTCH1 into UC cell lines did not significantly affect short-term cell proliferation, but induced nuclear abnormalities and diminished clonogenicity.ConclusionOur data indicate that canonical Notch signalling is suppressed in urothelial carcinoma mainly through downregulation of NOTCH1. These findings can be explained by proposing that canonical Notch signalling may promote differentiation in the urothelium, like in many squamous epithelia, and its suppression may therefore be advantageous for tumour progression. As an important corollary, inhibition of canonical Notch signalling is unlikely to be efficacious and might be counter-productive in the treatment of urothelial carcinoma.Electronic supplementary materialThe online version of this article (doi:10.1186/1471-2407-14-628) contains supplementary material, which is available to authorized users.

Not all NOTCH Is Created Equal: The Oncogenic Role of NOTCH2 in Bladder Cancer and Its Implications for Targeted Therapy

“…Furthermore, we have demonstrated the proof of concept that these findings can be translated into a therapeutic strategy for patients with bladder cancer by studying the effects of a NOTCH2 inactivating antibody, NRR2Mab. Our results are particularly timely as they build on three recent reports suggesting that NOTCH is a tumor suppressor in bladder cancer (12)(13)(14).…”

“…They are further limited by their dependence on a luminal (and not basal) promoter. Nonetheless, this work establishes the critical role of NOTCH signaling in bladder cancer, and especially highlights the tumor suppressor role of NOTCH1 (14).…”

Tocotrienol-rich mixture inhibits cell proliferation and induces apoptosis via down-regulation of the Notch-1/NF-κB pathways in NSCLC cells

“…21,[27][28][29] Aberrant expression of Notch has been reported in many types of cancer, including pancreatic, colon, lung, cervical, breast, and skin cancers. [30][31][32][33][34][35] Variable expression levels of Notch-1 were observed in a clinical study.…”

Consequences of Disrupted Notch Signaling in Bladder Cancer

“…Now, within a few months, three papers have appeared that, despite different emphases and some divergence in molecular details, agree on the most important issue, namely, that Notch signaling is frequently inactivated in bladder cancer [3][4][5]. This means that drugs inhibiting the pathway overall are unlikely to be beneficial in the treatment of bladder cancer.…”

“…This means that drugs inhibiting the pathway overall are unlikely to be beneficial in the treatment of bladder cancer. Indeed, such inhibitors do not diminish proliferation of urothelial carcinoma (UC) cell lines [3]. Instead, they could exert adverse effects on healthy urothelium if applied for the treatment of cancers with Notch overactivity in other organs.…”