A tetracycline-inducible CRISPR/Cas9 system, targeting two long non-coding RNAs, suppresses the malignant behavior of bladder cancer cells

Abstract

Clustered regularly interspaced short palindromic repeats (CRISPR) associated protein 9 (Cas9) technology has been applied in varied biological studies, including cancer studies. However, stable mRNA expression of Cas9 has potential risks in future gene therapy. Therefore, in the present study, a tetracycline-inducible switch was used to control the mRNA expression of Cas9. Long non-coding RNAs (lncRNAs) may be important functional regulators in tumor development, including in bladder cancer. RNA was designed to simultaneously target two lncRNAs, PVT1 and ANRIL, which are considered to be bladder cancer oncogenes. The mRNA expression of Cas9 was controlled by doxycycline. Reverse transcription-quantitative polymerase chain reaction revealed that the expression of PVT1 and ANRIL was significantly inhibited by the tetracycline-inducible CRISPR/Cas9 system. Functional assays demonstrated that this system could inhibit proliferation, induce apoptosis and suppress cell migration. Therefore, the tetracycline-inducible CRISPR/Cas9 system was demonstrated to repress the malignant behavior of bladder cancer cells by controlling the expression of Cas9 and simultaneously targeting two oncogenic lncRNAs.