MicroRNA-23b Functions as a Tumor Suppressor by Regulating Zeb1 in Bladder Cancer

Shahana Majid,Inik Chang,Guoren Deng,Kirsten Greene,Rajvir Dahiya,Soichiro Yamamura,Yuichiro Tanaka,Altaf A Dar,Sharanjot Saini,Karl X Chai

doi:10.1371/journal.pone.0067686

Abstract

MicroRNAs (miRNAs) are small, non-coding RNAs that regulate gene expression by targeted repression of transcription and translation. In this study we show that miRNA-23b (miR-23b) acts as a tumor suppressor in bladder cancer. Quantitative real-time PCR analysis showed that miR-23b is significantly down-regulated in bladder cancer cell lines and tumor tissues compared to non-malignant cells and normal tissue samples. We also demonstrate that miR-23b expression has a potential to be diagnostic and prognostic biomarker in bladder cancer. High miR-23b expression is positively correlated with higher overall survival of bladder cancer patients as revealed by Kaplan-Meier analysis. ROC analysis showed that miR-23b expression can distinguish between normal and bladder cancer tissues. Further we elucidated the biological significance of miR-23b in bladder cancer. Over-expression of miR-23b in bladder cancer cells inhibited cell proliferation and impaired colony formation. Fluorescence activated cell sorting (FACS) analysis revealed that re-expression of miR-23b in bladder cancer cells induced G0/G1 cell cycle arrest and apoptosis while inhibiting cell migration and invasion. Luciferase reporter assays demonstrated that Zeb1, a crucial regulator of epithelial-to-mesenchymal transition (EMT), is a direct target of miR-23b in bladder cancer. These results show that loss of miR-23b confers a proliferative advantage and promotes bladder cancer cell migration and invasion. Furthermore, re-expression of miR-23b may be a beneficial therapeutic strategy for the treatment of human bladder cancer.

Highlights

Bladder cancer is the fourth most common malignancy in the United States and one of the costliest to clinically manage [1]
Preliminary microRNA microarray data revealed that miR-23b was highly downregulated in bladder cancer cell lines compared to the non-malignant SV-HUC1 cell line
We validated the microarray data by miRNA-quantitative RT-PCR analysis and results confirmed that miR-23b was downregulated in bladder cancer cell lines J82, T24 compared to nonmalignant cell line SV-HUC1 (Figure 1A)

Summary

Introduction

Bladder cancer is the fourth most common malignancy in the United States and one of the costliest to clinically manage [1]. More than 90% of urinary bladder tumors are comprised of transitional cell carcinoma (TCC) that arises from transitional epithelium [2]. Urinary bladder tumors are classified into two distinct categories: non-muscle and muscle invasive bladder cancer [3,4]. Most tumors (75–80%) present as low-grade papillary noninvasive tumors that rarely progress to become lethal but almost always recur. This type of cancer is called ‘‘superficial’’ bladder cancer and requires expensive long-term management. Etiological factors involved in bladder carcinogenesis remain unidentified, and effective molecular markers for the disease are limited

Methods

Results

Conclusion