Effects of Combined SiRNA-TR and -TERT on Telomerase Activity and Growth of Bladder Transitional Cell Cancer BIU-87 Cells

Overview

Journal J Huazhong Univ Sci Technolog Med Sci

Specialty General Medicine

Date 2010 Jun 18

PMID 20556588

Citations 3

Authors

Wen Cheng

Zhifeng Wei

Jianping Gao

Zhengyu Zhang

Jingping Ge

Kangzhen Jing

Feng Xu

Peng Xie

Affiliations

Soon will be listed here.

Abstract

The effects of combined RNA interference (RNAi) of human telomerase RNA (hTR) and human telomerase reverse transcriptase (hTERT) genes on telomerase activity in a bladder cancer cell line (BIU-87 cells) were investigated by using gene chip technology in vitro with an attempt to evaluate the role of RNAi in the gene therapy of bladder transitional cell cancer (BTCC). Three TR-specific double-stranded small interfering RNAs (siRNAs) and three TERT-specific double-stranded siRNAs were designed to target different regions of TR and TERT mRNA. The phTR-siRNA, phTERT-siRNA, and the combination of both plasmids phTR+phTERT-siRNA were transfected into BIU-87 cells. The expression of hTR and hTERT mRNA was detected by quantitative fluorescent reverse transcription-polymerase chain reaction, and a telomeric repeat amplification protocol was applied to detect telomerase activity. Growth inhibition of BIU-87 cells was measured by MTT assay. Gene chip analysis was performed to evaluate the effects of the combined RNAi of hTR+hTERT genes on telomerase activity and growth of BIU-87 cells in vitro. The results showed that the expression of hTERT and hTR mRNA was inhibited by pRNAT-hTERT-III, pRNAT-hTR-III, and pRNAT-hTR-III+hTERT-III in BIU-87 cells. The inhibition efficiency of pRNAT-hTERT-III, pRNAT-hTR-III, pRNAT-hTERT-III+pRNAT-hTR-III was 67% for TERT mRNA, 41% for TR mRNA, 57% for TR mRNA and 70% for TERT mRNA in BIU-87 cells respectively. The growth of BIU-87 cells was inhibited and telomerase activity was considerably decreased, especially in the cells treated with combined RNAi-hTR and -hTERT. Gene chip analysis revealed that 21 genes were down-regulated (ATM, BAX, BCL2, BCL2L1, BIRC5, CD44, CTNNB1, E2F1, JUN, MCAM, MTA1, MYC, NFKB1, NFKBIA, NME4, PNN, PNN, SERPINE1, THBS1, TNFRSF1A, and UCC1). The results indicated that hTR-siRNA and hTERT-siRNA, especially their combination, siRNA hTR+hTERT, specifically and effectively suppressed the expression of both hTR and hTERT mRNA and telomerase activity. Molecular biological mechanism by which combined siRNA-TR and -TERT inhibited telomerase activity and growth of BIU-87 cells in vitro may involve the down-regulation of the 21 genes.

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References

Angele S, Falconer A, Foster C, Taniere P, Eeles R, Hall J . ATM protein overexpression in prostate tumors: possible role in telomere maintenance. Am J Clin Pathol. 2004; 121(2):231-6. DOI: 10.1309/JTKG-GGKU-RFX3-XMGT. View

Ito H, Kyo S, Kanaya T, Takakura M, Inoue M, Namiki M . Expression of human telomerase subunits and correlation with telomerase activity in urothelial cancer. Clin Cancer Res. 1998; 4(7):1603-8. View

Malerba I, Gribaldo L, Diodovich C, Carloni M, Meschini R, Bowe G . Induction of apoptosis and inhibition of telomerase activity in human bone marrow and HL-60 p53 null cells treated with anti-cancer drugs. Toxicol In Vitro. 2005; 19(4):523-32. DOI: 10.1016/j.tiv.2004.12.006. View

Zou L, Zhang P, Luo C, Tu Z . shRNA-targeted hTERT suppress cell proliferation of bladder cancer by inhibiting telomerase activity. Cancer Chemother Pharmacol. 2005; 57(3):328-34. DOI: 10.1007/s00280-005-0056-x. View

Oeggerli M, Tomovska S, Schraml P, Calvano-Forte D, Schafroth S, Simon R . E2F3 amplification and overexpression is associated with invasive tumor growth and rapid tumor cell proliferation in urinary bladder cancer. Oncogene. 2004; 23(33):5616-23. DOI: 10.1038/sj.onc.1207749. View

Wu K, Grandori C, Amacker M, POLACK A, Lingner J, Dalla-Favera R . Direct activation of TERT transcription by c-MYC. Nat Genet. 1999; 21(2):220-4. DOI: 10.1038/6010. View

Toshikuni N, Nouso K, Higashi T, Nakatsukasa H, Onishi T, Kaneyoshi T . Expression of telomerase-associated protein 1 and telomerase reverse transcriptase in hepatocellular carcinoma. Br J Cancer. 2000; 82(4):833-7. PMC: 2374388. DOI: 10.1054/bjoc.1999.1008. View

Lord R, Salonga D, Danenberg K, Peters J, DeMeester T, Park J . Telomerase reverse transcriptase expression is increased early in the Barrett's metaplasia, dysplasia, adenocarcinoma sequence. J Gastrointest Surg. 2000; 4(2):135-42. DOI: 10.1016/s1091-255x(00)80049-9. View

Pirocanac E, Nassirpour R, Yang M, Wang J, Nardin S, Gu J . Bax-induction gene therapy of pancreatic cancer. J Surg Res. 2002; 106(2):346-51. DOI: 10.1006/jsre.2002.6473. View

10.

Metcalfe J, Parkhill J, Campbell L, Stacey M, Biggs P, Byrd P . Accelerated telomere shortening in ataxia telangiectasia. Nat Genet. 1996; 13(3):350-3. DOI: 10.1038/ng0796-350. View

11.

Rhyu M . Telomeres, telomerase, and immortality. J Natl Cancer Inst. 1995; 87(12):884-94. DOI: 10.1093/jnci/87.12.884. View

12.

DuBois M, Haimberger Z, McIntosh M, Gottschling D . A quantitative assay for telomere protection in Saccharomyces cerevisiae. Genetics. 2002; 161(3):995-1013. PMC: 1462171. DOI: 10.1093/genetics/161.3.995. View

13.

Suenaga M, Yamaguchi A, Soda H, Orihara K, Tokito Y, Sakaki Y . Antiproliferative effects of gefitinib are associated with suppression of E2F-1 expression and telomerase activity. Anticancer Res. 2006; 26(5A):3387-91. View

14.

Elbashir S, Lendeckel W, Tuschl T . RNA interference is mediated by 21- and 22-nucleotide RNAs. Genes Dev. 2001; 15(2):188-200. PMC: 312613. DOI: 10.1101/gad.862301. View

15.

Lin Y, Uemura H, Fujinami K, Hosaka M, Harada M, Kubota Y . Telomerase activity in primary prostate cancer. J Urol. 1997; 157(3):1161-5. View

16.

Wu X, Deng Y, Wang G, Tao K . Combining siRNAs at two different sites in the EGFR to suppress its expression, induce apoptosis, and enhance 5-fluorouracil sensitivity of colon cancer cells. J Surg Res. 2006; 138(1):56-63. DOI: 10.1016/j.jss.2006.06.027. View

17.

Watanabe N . [Telomerase, cell immortality and cancer]. Hokkaido Igaku Zasshi. 2001; 76(3):127-32. View

18.

Lancelin F, Anidjar M, Villette J, Soliman A, Teillac P, Le Duc A . Telomerase activity as a potential marker in preneoplastic bladder lesions. BJU Int. 2000; 85(4):526-31. DOI: 10.1046/j.1464-410x.2000.00466.x. View

19.

Suzuki T, Suzuki Y, Fujioka T . Expression of the catalytic subunit associated with telomerase gene in human urinary bladder cancer. J Urol. 1999; 162(6):2217-20. DOI: 10.1016/S0022-5347(05)68162-1. View

20.

Qi L, Strong M, Karim B, Armanios M, Huso D, Greider C . Short telomeres and ataxia-telangiectasia mutated deficiency cooperatively increase telomere dysfunction and suppress tumorigenesis. Cancer Res. 2003; 63(23):8188-96. View