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Synthetic miRNA sponges driven by mutant hTERT promoter selectively inhibit the progression of bladder cancer

  • Research Article
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Tumor Biology

Abstract

The mutant promoter of human telomerase reverse transcriptase (hTERT) shows high transcriptional activity in bladder cancer cells. Some up-regulated microRNAs (miRNAs) are reported as oncogenic factors in bladder cancer. Previous studies report that miRNAs can be inhibited by base-pairing interactions. The purpose of this study is to construct a synthetic device driven by mutant hTERT promoter to suppress four up-regulated miRNAs and to verify its effects on phenotypes of bladder cancer cells and human normal cells. Tandem bulged miRNA binding sites targeting oncogenic miRNAs were inserted into the 3′ untranslated region (3′ UTR) of mutant hTERT promoter-driven Renilla luciferase gene to construct a synthetic tumor-specific device, miRNA sponges. A negative control was generated by using tandem repeated sequences without targeting any known miRNA. Bladder cancer cells (T24, 5637, UM-UC-3) and human fiber cells (HFC) were transfected with devices. Various functional assays were used to detect the effects of this device. The activity of the mutant hTERT promoter detected by luciferase assay was about three times as large as the wild-type hTERT promoter in bladder cancer cells, while it could not be measured in HFC. Other assays indicated that the synthetic device can significantly inhibit cell growth, decrease motility, and induce apoptosis in bladder cancer cells but not in HFC. A synthetic biology platform is employed to construct tumor-specific miRNA sponges that can be used to target oncogenic miRNAs to inhibit the progression of bladder cancer cells without affecting normal cells.

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References

  1. Burger M, Catto JW, Dalbagni G, Grossman HB, Herr H, Karakiewicz P, et al. Epidemiology and risk factors of urothelial bladder cancer. Eur Urol. 2013;63(2):234–41.

    Article  PubMed  Google Scholar 

  2. Falke J, Witjes JA. Contemporary management of low-risk bladder cancer. Nat Rev Urol. 2011;8:42–9.

    Article  PubMed  Google Scholar 

  3. Lei AQ, Cheng L, Pan CX. Current treatment of metastatic bladder cancer and future directions. Expert Rev Anticancer Ther. 2011;11(12):1851–62.

    Article  CAS  PubMed  Google Scholar 

  4. Kim WJ, Bae SC. Molecular biomarkers in urothelial bladder cancer. Cancer Sci. 2008;99(4):646–52.

    Article  CAS  PubMed  Google Scholar 

  5. Marta GN, Hanna SA, Gadia R, Correa SF, Silva JL, Carvalho Hde A. The role of radiotherapy in urinary bladder cancer: current status. Int Braz J Urol. 2012;38(2):144–53.

    Article  PubMed  Google Scholar 

  6. Racioppi M, D’Agostino D, Totaro A, Pinto F, Sacco E, D’Addessi A, et al. Value of current chemotherapy and surgery in advanced and metastatic bladder cancer. Urol Int. 2012;88(3):249–58.

    Article  CAS  PubMed  Google Scholar 

  7. Harley CB. Telomerase and cancer therapeutics. Nat Rev Cancer. 2008;8(3):167–79.

    Article  CAS  PubMed  Google Scholar 

  8. Verdun RE, Karlseder J. Replication and protection of telomeres. Nature. 2007;447(7147):924–31.

    Article  CAS  PubMed  Google Scholar 

  9. Wu S, Huang P, Li C, Huang Y, Li X, Wang Y, et al. Telomerase reverase transcriptase gene promotor mutations help discern the origin of urogenital tumors: a genomic and molecular study. Eur Urol. 2014;65(2):274–7.

    Article  CAS  PubMed  Google Scholar 

  10. Bartel DP. MicroRNAs: genomics, biogenesis, mechanism, and function. Cell. 2004;116(2):281–97.

    Article  CAS  PubMed  Google Scholar 

  11. Pasquinelli AE. MicroRNAs and their targets: recognition, regulation and an emerging reciprocal relationship. Nat Rev Genet. 2011;13(4):271–82.

    Google Scholar 

  12. Yoshino H, Seki N, Itesako T, Chiyomaru T, Nakagawa M, Enokida H. Aberrant expression of microRNAs in bladder cancer. Nat Rev Urol. 2013;10(7):396–404.

    Article  CAS  PubMed  Google Scholar 

  13. Liu Y, Han Y, Zhang H, Nie L, Jiang Z, Fa P, et al. Synthetic miRNA-mowers targeting miR-183-96-182 cluster or miRNA-210 inhibit growth and migration and induce apoptosis in bladder cancer cells. PLoS One. 2012;7(12):e52280.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  14. He L, Thomson JM, Hemann MT, Hernando-Monge E, Mu D, Goodson S, et al. A microRNA polycistron as a potential human oncogene. Nature. 2005;435(7043):828–33.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Serrano L. Synthetic biology: promises and challenges. Mol Syst Biol. 2007;3:158.

    Article  PubMed  PubMed Central  Google Scholar 

  16. Ebert MS, Neilson JR, Sharp PA. MicroRNA sponges: competitive inhibitors of small RNAs in mammalian cells. Nat Methods. 2007;4(9):721–6.

    Article  CAS  PubMed  Google Scholar 

  17. Liang CC, Park AY, Guan JL. In vitro scratch assay: a convenient and inexpensive method for analysis of cell migration in vitro. Nat Protoc. 2007;2(2):329–33.

    Article  CAS  PubMed  Google Scholar 

  18. Wang T, Yuan J, Feng N, Li Y, Lin Z, Jiang Z, et al. Hsa-miR-1 downregulates long non-coding RNA urothelial cancer associated 1 in bladder cancer. Tumour Biol. 2014;35(10):10075–84.

    Article  CAS  PubMed  Google Scholar 

  19. Png KJ, Halberg N, Yoshida M, Tavazoie SF. A microRNA regulation that mediates endothelial recruitment and metastasis by cancer cells. Nature. 2011;481(7380):190–4.

    Article  PubMed  Google Scholar 

  20. Kandalam MM, Beta M, Maheswari UK, Swaminathan S, Krishnakumar S. Oncogenic microRNA17-92 cluster is regulated by epithelial cell adhesion molecule and could be a potential therapeutic target in retinoblastoma. Mol Vis. 2012;18:2279–87.

    CAS  PubMed  PubMed Central  Google Scholar 

  21. Zhao S, Yao D, Chen J, Ding N. Circulating miRNA-20a and miRNA-203 for screening lymph node metastasis in early stage cervical cancer. Genet Test Mol Biomarkers. 2013;17(8):631–6.

    Article  CAS  PubMed  Google Scholar 

  22. Yan Z, Wang J, Wang C, Jiao Y, Qi W, Che S. miR-96/HBP1/Wnt/β-catenin regulatory circuitry promoters glioma growth. FEBS Lett. 2014;588(17):3038–46.

    Article  CAS  PubMed  Google Scholar 

  23. Zhang QH, Sun HM, Zheng RZ, Li YC, Zhang Q, Cheng P, et al. Meta-analysis of microRNA-183 family expression in human cancer studies comparing cancer tissues with noncancerous tissues. Gene. 2013;527(1):26–32.

    Article  CAS  PubMed  Google Scholar 

  24. Low KC, Tergaonkar V. Telomerase: central regulator of all of the hallmarks of cancer. Trends Biochem Sci. 2013;38(9):426–34.

    Article  CAS  PubMed  Google Scholar 

  25. Shay JW, Zou Y, Hiyama E, Wright WE. Telomerase and cancer. Hum Mol Genet. 2001;10(7):677–85.

    Article  CAS  PubMed  Google Scholar 

  26. Cong YS, Wen J, Bacchetti S. The human telomerase catalytic subunit hTERT: organization of the gene and characterization of the promoter. Hum Mol Genet. 1999;8(1):137–42.

    Article  CAS  PubMed  Google Scholar 

  27. Gladych M, Wojtyla A, Rubis B. Human telomerase expression regulation. Biochem Cell Biol. 2011;89(4):359–76.

    Article  CAS  PubMed  Google Scholar 

  28. Wojtyla A, Gladych M, Rubis B. Human telomerase activity regulation. Mol Biol Rep. 2011;38(5):3339–49.

    Article  CAS  PubMed  Google Scholar 

  29. Horn S, Figl A, Rachakonda PS, Fischer C, Sucker A, Gast A, et al. TERT promoter mutations in familial and sporadic melanoma. Science. 2013;339(6122):959–61.

    Article  CAS  PubMed  Google Scholar 

  30. Huang FW, Hodis E, Xu MJ, Kryukov GV, Chin L, Garraway LA. Highly recurrent TERT promoter mutations in human melanoma. Science. 2013;339(6122):957–9.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  31. Rachakonda PS, Hosen I, de Verdier PJ, Fallah M, Heidenreich B, Ryk C, et al. TERT promoter mutations in bladder cancer affect patient survival and disease recurrence through modification by a common polymorphism. Proc Natl Acad Sci U S A. 2013;110(43):17426–31.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Kyo S, Takakura M, Fujiwara T, Inoue M. Understanding and exploiting hTERT promoter regulation for diagnosis and treatment of human cancers. Cancer Sci. 2008;99(8):1528–38.

    Article  CAS  PubMed  Google Scholar 

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Acknowledgments

This research was supported by the National Key Basic Research Program of China (973 Program) (2014CB745201), National Natural Science Foundation of China [81402103] and Basic Research Program of Shenzhen (JCYJ20120614155650545). The authors thank all the donors whose names were not included in the author list, but who participated in this program.

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Correspondence to Yu-Chen Liu, Wei-Ren Huang or Zhi-Ming Cai.

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Zhuang, CL., Fu, X., Liu, L. et al. Synthetic miRNA sponges driven by mutant hTERT promoter selectively inhibit the progression of bladder cancer. Tumor Biol. 36, 5157–5163 (2015). https://doi.org/10.1007/s13277-015-3169-9

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  • DOI: https://doi.org/10.1007/s13277-015-3169-9

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