Inhibition of long non-coding RNA UCA1 by CRISPR/Cas9 attenuated malignant phenotypes of bladder cancer

// Shuai Zhen 1, 2, * , Ling Hua 3, * , Yun-Hui Liu 4 , Xiao-Min Sun 5 , Meng-Meng Jiang 6 , Wei Chen 5 , Le Zhao 1, 2 , Xu Li 1, 2 1 Center for Translational Medicine, The First Affiliated Hospital, Xi’an Jiaotong University, Xi’an 710061, P.R. China 2 Key Laboratory for Tumor Precision Medicine of Shaanxi Province, The First Affiliated Hospital of Xi’an Jiaotong University, Xi’an 710061, P.R. China 3 Department of Veterinary Medicine, Rongchang Campus of Southwest University, Chongqing 402460, P.R. China 4 Department of Pharmacology and Toxicology, Beijing Institute of Radiation Medicine, Beijing 100850, P.R. China 5 Center for Laboratory Medicine, The First Affiliated Hospital of Xi’an Jiaotong University, Xi’an 710061, P.R. China 6 State Key Laboratory of Cancer Biology, The Fourth Military Medical University, Xi’an 710032, Shaanxi, China * These authors have contributed equally to this work Correspondence to: Xu Li, email: lixu56@mail.xjtu.edu.cn Le Zhao, email: zhaole2@mail.xjtu.edu.cn Keywords: CRISPR/Cas9, long non-coding RNA, UCA1, bladder cancer Received: January 16, 2016      Accepted: December 12, 2016      Published: December 25, 2016 ABSTRACT CRISPR/Cas9 is a novel and effective genome editing technique, but its application is not widely expanded to manipulate long non-coding RNA (lncRNA) expression. The lncRNA urothelial carcinoma-associated 1 (UCA1) is upregulated in bladder cancer and promotes the progression of bladder cancer. Here, we design gRNAs specific to UCA1 and construct CRISPR/Cas9 systems targeting UCA1. Single CRISPR/Cas9-UCA1 can effectively inhibit UCA1 expression when transfected into 5637 and T24 bladder cancer cells, while the combined transfection of the two most effective CRISPR/Cas9-UCA1s can generate more satisfied inhibitory effect. CRISPR/Cas9-UCA1s attenuate UCA1 expression via targeted genome-specific DNA cleavage, resulting in the significant inhibition of cell proliferation, migration and invasion in vitro and in vivo . The mechanisms associated with the inhibitory effect of CRISPR/Cas9-UCA1 on malignant phenotypes of bladder cancer are attributed to the induction of cell cycle arrest at G1 phase, a substantial increase of apoptosis, and an enhanced activity of MMPs. Additionally, urinary UCA1 can be used as a non-invasive diagnostic marker for bladder cancer as revealed by a meta-analysis. Collectively, our data suggest that CRISPR/Cas9 technique can be used to down-modulate lncRNA expression, and urinary UCA1 may be used as a non-invasive marker for diagnosis of bladder cancer.