Phospho-Akt pathway activation and inhibition depends on N-cadherin or phospho-EGFR expression in invasive human bladder cancer cell lines☆

Abstract 

Objectives

A particular interest in epithelial-mesenchymal transition (EMT), which takes place during embryonic development, provided potential mechanisms involved in the progression of many epithelial tumors, including bladder cancer (BC). The phospho-Akt signaling pathway is supposed to be involved in invasion and progression of human tumors, including BC. Moreover, it has been demonstrated in bladder cancer cell lines that N-cadherin or phospho-epithelial growth factor receptor (EGFR) expression are correlated to tumor progression. Our objectives were to evaluate the potential phospho-Akt pathway involvement in N-cadherin and/or phospho-EGFR positive BC cell lines and to evaluate the prognostic value of E- and N-cadherin expression in patients undergoing cystectomy for invasive BC.

Materials and methods

We screened a panel of invasive and noninvasive BC cell lines for E- and N-cadherin, phospho-EGFR, and phospho-Akt expression using the Western blot technique (WB). The potential role of N-cadherin in invasion was assessed by Matrigel assays with and without the N-cadherin blocking monoclonal antibody GC-4. Then we used the Affymetrix microarray technique to evaluate the prognostic value of E- and N-cadherin expression in 30 patients undergoing a cystectomy for invasive BC.

Results

N-cadherin and phospho-EGFR expression are associated with Akt activation and with invasive behavior modulation. Even if Akt activation is sufficient in promoting invasion, its inactivation by LY294002 (PI-3 kinase inhibitor) is less efficient on invasion than inhibition of N-cadherin and phospho-EGFR by GC-4 (monoclonal antibody) and gefitinib (anti-tyrosine kinase), respectively. N-cadherin and phospho-EGFR inhibition decreased phospho-Akt activation but also caused restoration and reinforcing of E-cadherin expression, respectively, while phospho-Akt inhibition did not have any impact on E-cadherin expression. In a group of high-risk bladder tumors (T₁G₃), N- and E-cadherin expression could be considered as a prognostic marker. In a group of patients with invasive BC (pT₂-T₄) undergoing cystectomy, we showed a shorter overall survival when BC expressed N-cadherin (P = 0.0064) and when E-cadherin expression was down-regulated (P = 0.00165). The N _positive /E _negative profile has the worst prognosis (P = 0.00153).

Conclusions

We confirmed the partial responsibility of p-Akt activation in invasion of some BC cell lines expressing N-cadherin or p-EGFR and also the potential role of N-cadherin and p-EGFR as target in cancer therapy. N/E- cadherin expression profile has a significant prognostic value in invasive BC.

Keywords: Urothelial bladder neoplasms, Invasion, Epithelial-mesenchymal transition, N-cadherin, E-cadherin, Phospho-EGFR, Phospho-Akt, Targeted therapy