MP60-16 CRISPR/CAS9-MEDIATED MIR-210-3P DEPLETION PROMOTED TUMORIGENESIS THROUGH REVIVAL OF TWIST1 IN RENAL CELL CARCINOMA

Field: Biochemistry, Genetics and Molecular Biology (Life Sciences)

You have accessJournal of UrologyKidney Cancer: Basic Research & Pathophysiology II1 Apr 2017MP60-16 CRISPR/CAS9-MEDIATED MIR-210-3P DEPLETION PROMOTED TUMORIGENESIS THROUGH REVIVAL OF TWIST1 IN RENAL CELL CARCINOMA Hirofumi Yoshino, Masaya Yonemori, Kazutaka Miyamoto, Satoshi Kofuji, Nijiro Nohata, Hideki Enokida, and Masayuki Nakagawa Hirofumi YoshinoHirofumi Yoshino More articles by this author , Masaya YonemoriMasaya Yonemori More articles by this author , Kazutaka MiyamotoKazutaka Miyamoto More articles by this author , Satoshi KofujiSatoshi Kofuji More articles by this author , Nijiro NohataNijiro Nohata More articles by this author , Hideki EnokidaHideki Enokida More articles by this author , and Masayuki NakagawaMasayuki Nakagawa More articles by this author View All Author Informationhttps://doi.org/10.1016/j.juro.2017.02.1853AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookTwitterLinked InEmail INTRODUCTION AND OBJECTIVES CRISPR/Cas9 technology was introduced as an efficient, powerful and broadly used genome editing tool. The aim of this study was to utilize the CRISPR/Cas9 system to control miRNA expression in cancer research. METHODS In our miRNA expression signatures in clear cell renal cell carcinoma (ccRCC), we focused on miR-210-3p which was one of the most upregulated miRNAs. We used lenti-CRISPR vector to knock out miR-210-3p with two different single guide RNAs against miR-210-3p. We performed cell function studies and xenograft assay with miR-210-3p- depleted cells. Putative target genes of miR-210-3p was examined in miRNA binding assay. In addition, overall survival between high and low expression of miR-210-3p or the target gene were analyzed by the Kaplan-Meier method. RESULTS In cells transfected with sgRNA targeting miR-210-3p itself, more than 98% miR-210-3p knock out efficiency was observed in all three cell lines (786-o, A498 and Caki2). miR-210-3p depletion significantly increased invasive capacity (P < 0.01) in vitro, and dramatically promoted tumorigenesis in xenograft experiments (P = 0.0039), which was unexpected due to the fact that these miRNAs were up-regulated in RCC. We also found that twist family bHLH transcription factor 1 (TWIST1) was identified as a direct target of miR-210-3p based on target analyses (P < 0.05). The Cancer Genome Atlas (TCGA) database of ccRCC showed that there was a negative correlation between miR-210-3p and TWIST1 expression (P < 0.0001). In accordance with the results in vivo and in vitro analyses, TCGA database showed that the low miR-210-3p expression group had poor survival in comparison with the high group, and the high TWIST1 expression group had poor overall (P = 0.00054) and disease-free survival (P = 0.00347) compared to the low expression group significantly. CONCLUSIONS We utilized the CRISPR/Cas9 system to analyze an upregulated miRNA in cancer. CRISPR/Cas9 successfully suppressed miR-210-3p expression in RCC cells. Moreover, by using CRISPR/Cas9 techniques, we found that tumorigenesis was acquired through enhanced expression of oncogenic TWIST1 due to the suppression of miR-210-3p expression via CRISPR/Cas9 techniques. The introduction of the CRISPR/Cas9 methodologies into studies of miRNAs as well as other non-coding RNAs should provide new possibilities in cancer research. © 2017FiguresReferencesRelatedDetails Volume 197Issue 4SApril 2017Page: e796 Advertisement Copyright & Permissions© 2017MetricsAuthor Information Hirofumi Yoshino More articles by this author Masaya Yonemori More articles by this author Kazutaka Miyamoto More articles by this author Satoshi Kofuji More articles by this author Nijiro Nohata More articles by this author Hideki Enokida More articles by this author Masayuki Nakagawa More articles by this author Expand All Advertisement Advertisement PDF downloadLoading ...