title: Synthetic miRNA-mowers targeting miR-183-96-182 cluster or miR-210 inhibit growth and migration and induce apoptosis in bladder cancer cells.
abstract: BACKGROUND
  MicroRNAs (miRNAs) function as endogenous regulators of biological behaviors of human cancers. Several natural non-coding RNAs are reported to inhibit miRNAs by base-pairing interactions. These phenomena raise questions about the ability of artificial device to regulate miRNAs. The purpose of this study is to create synthetic devices that target a single miRNA or a miRNA cluster and to ascertain their therapeutic effects on the phenotypes of bladder cancer cells.
  
  
  RESULTS
  Tandem bulged miRNA binding sites were inserted into the 3' untranslated region (UTR) of the SV-40 promoter-driven Renilla luciferase gene to construct two "miRNA-mowers" for suppression of miR-183-96-182 cluster or miR-210. A third device with tandem repeat sequences not complementary to any known miRNA was generated as an untargeted-control. In functional analyses, bladder cancer T24 and UM-UC-3 cells were transfected with each of the three devices, followed by assays for detection of their impacts. Luciferase assays indicated that the activities of the luciferase reporters in the miRNA-mowers were decreased to 30-50% of the untargeted-control. Using Real-Time qPCR, the expression levels of the target miRNAs were shown to be reduced 2-3-fold by the corresponding miRNA-mower. Cell growth, apoptosis, and migration were tested by MTT assay, flow cytometry assay, and in vitro scratch assay, respectively. Cell growth inhibition, increased apoptosis, and decreased motility were observed in miRNA-mowers-transfected bladder cancer cells.
  
  
  CONCLUSIONS
  Not only a single target miRNA but also the whole members of a target miRNA cluster can be blocked using this modular design strategy. Anti-cancer effects are induced by the synthetic miRNA-mowers in the bladder cancer cell lines. miR-183/96/182 cluster and miR-210 are shown to play oncogenic roles in bladder cancer. A potentially useful synthetic biology platform for miRNA loss-of-function study and cancer treatment has been established in this work.
authors.affiliation: Guangdong and Shenzhen Key Laboratory of Male Reproductive Medicine and Genetics, Institute of Urology, Peking University Shenzhen Hospital, Shenzhen PKU-HKUST Medical Center, Shenzhen, People's Republic of China.
authors.forename: Yuchen
authors.initials: Y
authors.lastname: Liu
authors.forename: Yonghua
authors.initials: Y
authors.lastname: Han
authors.forename: Hu
authors.initials: H
authors.lastname: Zhang
authors.forename: Liping
authors.initials: L
authors.lastname: Nie
authors.forename: Zhimao
authors.initials: Z
authors.lastname: Jiang
authors.forename: Pingping
authors.initials: P
authors.lastname: Fa
authors.forename: Yaoting
authors.initials: Y
authors.lastname: Gui
authors.forename: Zhiming
authors.initials: Z
authors.lastname: Cai
mesh_terms: D017209:Apoptosis / Q000235:genetics / Q000502:physiology; D045744:Cell Line, Tumor; D002465:Cell Movement / Q000235:genetics / Q000502:physiology; D006801:Humans; D035683:MicroRNAs / Q000235:genetics*; D060888:Real-Time Polymerase Chain Reaction; D001749:Urinary Bladder Neoplasms / Q000235:genetics*
chemical_list: C533449:MIRN183 microRNA, human; C543800:MIRN210 microRNA, human; C541492:MIRN96 microRNA, human; D035683:MicroRNAs