title: Synthetic tetracycline-controllable shRNA targeting long non-coding RNA HOXD-AS1 inhibits the progression of bladder cancer.
abstract: Long non-coding RNAs (lncRNAs) have been proved to act as key molecules in cancer development and progression. Dysregulation of lncRNAs is discovered in various tumor tissues and cancer cells where they can serve as oncogenes or tumor suppressors. Long non-coding RNA HOXD-AS (HOXD cluster antisense RNA 1) has recently been identified to be involved in the development of several cancers including neuroblastoma, adenocarcinomas and breast cancer. However, the role of HOXD-AS1 in bladder cancer remains unknown.
  
  
  
  The synthetic tetracycline-controllable shRNA was used to modulate the level of HOXD-AS1 by adding different concentrations of doxycycline (dox). RT-qPCR was used to detect the expression level of HOXD-AS1. Cell proliferation was determined by CCK-8 assay and EdU incorporation experiment when HOXD-AS1 was knocked down. We used wound-healing assay for detecting the effect of HOXD-AS1 on cell migration. Eventually, cell apoptosis was determined by caspase 3 ELISA assay and flow cytometry assay.
  
  
  
  In this study, we found that the expression level of HOXD-AS1 was significantly increased in bladder cancer tissues and cells. Furthermore, high expression of HOXD-AS1 was significantly related to tumor size, histological grade and TNM stage. In vitro assays confirmed that knockdown of HOXD-AS1 suppressed cell proliferation/migration and increased the rate of apoptotic cell in bladder cancer cells. At last, we used the important element of synthetic biology, tetracycline(tet)-controllable switch, to construct tet-controllable shRNA vectors which can modulate the expression of HOXD-AS1 in a dosage-dependent manner.
  
  
  
  Our research suggested that high expression of HOXD-AS1 may be involved in the bladder cancer carcinogenesis through inhibiting the phenotypes and activating endogenous cancer-related molecular pathways. Therefore, HOXD-AS1 may act as an oncogene and provide a potential attractive therapeutic target for bladder cancer. In addition, the synthetic tetracycline-controllable shRNA may provide a novel method for cancer research in vitro assays.
authors.affiliation: Key Laboratory of Medical Reprogramming Technology, Shenzhen Second People's Hospital, Clinical Institute of Shantou University Medical College, First Affiliated Hospital of Shenzhen University, Shenzhen, 518039, Guangdong Province, People's Republic of China.|Shantou University Medical College, Shantou, 515041, Guangdong Province, People's Republic of China.
authors.forename: Jianfa
authors.initials: J
authors.lastname: Li
authors.affiliation: Key Laboratory of Medical Reprogramming Technology, Shenzhen Second People's Hospital, Clinical Institute of Shantou University Medical College, First Affiliated Hospital of Shenzhen University, Shenzhen, 518039, Guangdong Province, People's Republic of China.|Shantou University Medical College, Shantou, 515041, Guangdong Province, People's Republic of China.|Guangdong and Shenzhen Key Laboratory of Male Reproductive Medicine and Genetics, Institute of Urology, Peking University Shenzhen Hospital, Shenzhen PKU-HKUST Medical Center, Shenzhen, 518036, People's Republic of China.
authors.forename: Chengle
authors.initials: C
authors.lastname: Zhuang
authors.affiliation: Key Laboratory of Medical Reprogramming Technology, Shenzhen Second People's Hospital, Clinical Institute of Shantou University Medical College, First Affiliated Hospital of Shenzhen University, Shenzhen, 518039, Guangdong Province, People's Republic of China.
authors.forename: Yuchen
authors.initials: Y
authors.lastname: Liu
authors.affiliation: Key Laboratory of Medical Reprogramming Technology, Shenzhen Second People's Hospital, Clinical Institute of Shantou University Medical College, First Affiliated Hospital of Shenzhen University, Shenzhen, 518039, Guangdong Province, People's Republic of China.|Anhui Medical University, Hefei, 230000, Anhui Province, People's Republic of China.
authors.forename: Mingwei
authors.initials: M
authors.lastname: Chen
authors.affiliation: Shantou University Medical College, Shantou, 515041, Guangdong Province, People's Republic of China.
authors.forename: Yincong
authors.initials: Y
authors.lastname: Chen
authors.affiliation: Key Laboratory of Medical Reprogramming Technology, Shenzhen Second People's Hospital, Clinical Institute of Shantou University Medical College, First Affiliated Hospital of Shenzhen University, Shenzhen, 518039, Guangdong Province, People's Republic of China.|Shantou University Medical College, Shantou, 515041, Guangdong Province, People's Republic of China.
authors.forename: Zhicong
authors.initials: Z
authors.lastname: Chen
authors.affiliation: Key Laboratory of Medical Reprogramming Technology, Shenzhen Second People's Hospital, Clinical Institute of Shantou University Medical College, First Affiliated Hospital of Shenzhen University, Shenzhen, 518039, Guangdong Province, People's Republic of China.|Anhui Medical University, Hefei, 230000, Anhui Province, People's Republic of China.
authors.forename: Anbang
authors.initials: A
authors.lastname: He
authors.affiliation: Key Laboratory of Medical Reprogramming Technology, Shenzhen Second People's Hospital, Clinical Institute of Shantou University Medical College, First Affiliated Hospital of Shenzhen University, Shenzhen, 518039, Guangdong Province, People's Republic of China.|Shantou University Medical College, Shantou, 515041, Guangdong Province, People's Republic of China.
authors.forename: Junhao
authors.initials: J
authors.lastname: Lin
authors.affiliation: Key Laboratory of Medical Reprogramming Technology, Shenzhen Second People's Hospital, Clinical Institute of Shantou University Medical College, First Affiliated Hospital of Shenzhen University, Shenzhen, 518039, Guangdong Province, People's Republic of China.|Shantou University Medical College, Shantou, 515041, Guangdong Province, People's Republic of China.
authors.forename: Yonghao
authors.initials: Y
authors.lastname: Zhan
authors.affiliation: Key Laboratory of Medical Reprogramming Technology, Shenzhen Second People's Hospital, Clinical Institute of Shantou University Medical College, First Affiliated Hospital of Shenzhen University, Shenzhen, 518039, Guangdong Province, People's Republic of China.|Shantou University Medical College, Shantou, 515041, Guangdong Province, People's Republic of China.
authors.forename: Li
authors.initials: L
authors.lastname: Liu
authors.affiliation: Key Laboratory of Medical Reprogramming Technology, Shenzhen Second People's Hospital, Clinical Institute of Shantou University Medical College, First Affiliated Hospital of Shenzhen University, Shenzhen, 518039, Guangdong Province, People's Republic of China.
authors.forename: Wen
authors.initials: W
authors.lastname: Xu
authors.affiliation: Shanghai-MOST Key Laboratory of Health and Disease Genomics, Chinese National Human Genome Centerat Shanghai, Shanghai, 200000, People's Republic of China.
authors.forename: Guoping
authors.initials: G
authors.lastname: Zhao
authors.affiliation: Department of Urology, Peking University First Hospital, Institute of Urology, Peking University, National Urological Cancer Center, Beijing, 100034, People's Republic of China.
authors.forename: Yinglu
authors.initials: Y
authors.lastname: Guo
authors.affiliation: Key Laboratory of Medical Reprogramming Technology, Shenzhen Second People's Hospital, Clinical Institute of Shantou University Medical College, First Affiliated Hospital of Shenzhen University, Shenzhen, 518039, Guangdong Province, People's Republic of China.
authors.forename: Hanwei
authors.initials: H
authors.lastname: Wu
authors.affiliation: Key Laboratory of Medical Reprogramming Technology, Shenzhen Second People's Hospital, Clinical Institute of Shantou University Medical College, First Affiliated Hospital of Shenzhen University, Shenzhen, 518039, Guangdong Province, People's Republic of China. caizhiming@yahoo.com.cn.|Shantou University Medical College, Shantou, 515041, Guangdong Province, People's Republic of China. caizhiming@yahoo.com.cn.|Anhui Medical University, Hefei, 230000, Anhui Province, People's Republic of China. caizhiming@yahoo.com.cn.|Department of Urology, Peking University First Hospital, Institute of Urology, Peking University, National Urological Cancer Center, Beijing, 100034, People's Republic of China. caizhiming@yahoo.com.cn.
authors.forename: Zhiming
authors.initials: Z
authors.lastname: Cai
authors.affiliation: Key Laboratory of Medical Reprogramming Technology, Shenzhen Second People's Hospital, Clinical Institute of Shantou University Medical College, First Affiliated Hospital of Shenzhen University, Shenzhen, 518039, Guangdong Province, People's Republic of China. pony8980@163.com.|Shantou University Medical College, Shantou, 515041, Guangdong Province, People's Republic of China. pony8980@163.com.|Anhui Medical University, Hefei, 230000, Anhui Province, People's Republic of China. pony8980@163.com.|Department of Urology, Peking University First Hospital, Institute of Urology, Peking University, National Urological Cancer Center, Beijing, 100034, People's Republic of China. pony8980@163.com.
authors.forename: Weiren
authors.initials: W
authors.lastname: Huang
mesh_terms: D053148:Caspase 3 / Q000378:metabolism; D045744:Cell Line, Tumor; D002465:Cell Movement; D049109:Cell Proliferation; D018450:Disease Progression; D005260:Female; D015972:Gene Expression Regulation, Neoplastic; D055785:Gene Knockdown Techniques; D006801:Humans; D008297:Male; D009367:Neoplasm Staging; D062085:RNA, Long Noncoding / Q000235:genetics*; D034741:RNA, Small Interfering / Q000235:genetics*; D015398:Signal Transduction; D058615:Synthetic Biology; D013754:Tetracyclines / Q000494:pharmacology*; D047368:Tumor Burden; D015854:Up-Regulation; D001749:Urinary Bladder Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000473:pathology*
chemical_list: D062085:RNA, Long Noncoding; D034741:RNA, Small Interfering; D013754:Tetracyclines; C000625564:long non-coding RNA HOXD-AS1, human; C505407:CASP3 protein, human; D053148:Caspase 3
keywords: Bladder cancer; HOXD-AS1; Synthetic biology; Tetracycline-controllable shRNA; lncRNA