Knockdown of glucose‑regulated protein 78/binding immunoglobulin heavy chain protein expression by asymmetric small interfering RNA induces apoptosis in prostate cancer cells and attenuates migratory capability

Lu,Tong; Yang,Weimin; Wang,Zhihua; Hu,Zhiquan; Zeng,Xing; Yang,Chunguang; Wang,Ye; Zhang,Yong; Li,Fan; Liu,Zhuo; Wang,Dongbiao; Ye,Zhangqun

doi:10.3892/mmr.2014.2737

Knockdown of glucose‑regulated protein 78/binding immunoglobulin heavy chain protein expression by asymmetric small interfering RNA induces apoptosis in prostate cancer cells and attenuates migratory capability

Authors:
- Tong Lu
- Weimin Yang
- Zhihua Wang
- Zhiquan Hu
- Xing Zeng
- Chunguang Yang
- Ye Wang
- Yong Zhang
- Fan Li
- Zhuo Liu
- Dongbiao Wang
- Zhangqun Ye
View Affiliations

Affiliations: Department of Urology, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan, Hubei 430030, P.R. China
Published online on: October 22, 2014 https://doi.org/10.3892/mmr.2014.2737
Pages: 249-256

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

Glucose‑regulated protein 78 [GRP78, also termed binding immunoglobulin heavy chain protein (Bip)] may be involved in cancer progression and metastasis. However, to date there has been minimal investigation into its potential role in human prostate cancer cells. Recent studies have demonstrated that asymmetric small interfering RNA (asiRNA)‑mediated gene silencing is more effective and longer‑lasting than conventional symmetric siRNA. Thus, the current study aimed to investigate the effects of GRP78‑specific asiRNA on human prostate cancer cells. A series of asiRNAs was synthesized and their efficiency in silencing GRP78 expression in PC‑3 human prostate cancer cells was evaluated. The effects of knockdown using asiRNAs were compared to those of knockdown using symmetric siRNAs. The effect of GRP78 silencing on PC‑3 cell apoptosis and migration, and the possible mechanisms governing these biological processes were examined. Compared with the symmetric siRNA, transfection with the 15 base pair asiRNA (asiGRP78‑3) resulted in greater downregulation of GRP78 expression. GRP78 depletion in PC‑3 cells resulted in increased apoptosis and decreased migration of these cells. Experiments investigating the underlying mechanisms of these effects revealed that knockdown of GRP78 attenuated protein kinase B activation and decreased the expression of pro‑caspase 9, pro‑caspase 3 and vimentin. In conclusion, knockdown of GRP78/Bip expression with asymmetric siRNA led to increased prostate cancer cell apoptosis and reduced cellular migration.

View Figures

View References

1	Semenas J, Allegrucci C, Boorjian SA, Mongan NP and Persson JL: Overcoming drug resistance and treating advanced prostate cancer. Curr Drug Targets. 13:1308–1323. 2012. View Article : Google Scholar : PubMed/NCBI
2	Feldman BJ and Feldman D: The development of androgen-independent prostate cancer. Nat Rev Cancer. 1:34–45. 2001. View Article : Google Scholar : PubMed/NCBI
3	Oh WK and Kantoff PW: Management of hormone refractory prostate cancer: current standards and future prospects. J Urol. 160:1220–1229. 1998. View Article : Google Scholar : PubMed/NCBI
4	Misra UK, Mowery Y, Kaczowka S and Pizzo SV: Ligation of cancer cell surface GRP78 with antibodies directed against its COOH-terminal domain up-regulates p53 activity and promotes apoptosis. Mol Cancer Ther. 8:1350–1362. 2009. View Article : Google Scholar : PubMed/NCBI
5	Misra UK, Wang F and Pizzo SV: Transcription factor TFII-I causes transcriptional upregulation of GRP78 synthesis in prostate cancer cells. J Cell Biochem. 106:381–389. 2009. View Article : Google Scholar : PubMed/NCBI
6	Huang LW, Lin CY, Lee CC, Liu TZ and Jeng CJ: Overexpression of GRP78 is associated with malignant transformation in epithelial ovarian tumors. Appl Immunohistochem Mol Morphol. 20:381–385. 2012. View Article : Google Scholar : PubMed/NCBI
7	Lee E, Nichols P, Spicer D, Groshen S, Yu MC and Lee AS: GRP78 as a novel predictor of responsiveness to chemotherapy in breast cancer. Cancer Res. 66:7849–7853. 2006. View Article : Google Scholar : PubMed/NCBI
8	Sun Q, Hua J, Wang Q, et al: Expressions of GRP78 and Bax associate with differentiation, metastasis, and apoptosis in non-small cell lung cancer. Mol Biol Rep. 39:6753–6761. 2012. View Article : Google Scholar : PubMed/NCBI
9	Li H, Song H, Luo J, Liang J, Zhao S and Su R: Knockdown of glucose-regulated protein 78 decreases the invasion, metalloproteinase expression and ECM degradation in hepatocellular carcinoma cells. J Exp Clin Cancer Res. 31:392012. View Article : Google Scholar
10	Wang Q, Shu R, He H, et al: Co-silencing of Birc5 (survivin) and Hspa5 (Grp78) induces apoptosis in hepatoma cells more efficiently than single gene interference. Int J Oncol. 41:652–660. 2012.PubMed/NCBI
11	Prados J, Melguizo C, Roldan H, et al: RNA Interference in the treatment of colon cancer. BioDrugs. 27:317–327. 2013. View Article : Google Scholar : PubMed/NCBI
12	Shi H, Deng JH, Wang Z, Cao KY, Zhou L and Wan H: Knockdown of clusterin inhibits the growth and migration of renal carcinoma cells and leads to differential gene expression. Mol Med Rep. 8:35–40. 2013.PubMed/NCBI
13	Zhao Y, Jian W, Gao W, et al: RNAi silencing of c-Myc inhibits cell migration, invasion, and proliferation in HepG2 human hepatocellular carcinoma cell line: c-Myc silencing in hepatocellular carcinoma cell. Cancer Cell Int. 13:232013. View Article : Google Scholar : PubMed/NCBI
14	Zhu Z, Zhu Z, Pang Z, Xing Y, Wan F, Lan D and Wang H: Short hairpin RNA targeting FOXQ1 inhibits invasion and metastasis via the reversal of epithelial-mesenchymal transition in bladder cancer. Int J Oncol. 42:1271–1278. 2013.PubMed/NCBI
15	Clark PR, Pober JS and Kluger MS: Knockdown of TNFR1 by the sense strand of an ICAM-1 siRNA: dissection of an off-target effect. Nucleic Acids Res. 36:1081–1097. 2008. View Article : Google Scholar : PubMed/NCBI
16	Jackson AL and Linsley PS: Noise amidst the silence: off-target effects of siRNAs? Trends Genet. 20:521–524. 2004. View Article : Google Scholar : PubMed/NCBI
17	Chang CI, Yoo JW, Hong SW, et al: Asymmetric shorter-duplex siRNA structures trigger efficient gene silencing with reduced nonspecific effects. Mol Ther. 17:725–732. 2009. View Article : Google Scholar : PubMed/NCBI
18	Jo SG, Hong SW, Yoo JW, Lee CH, Kim S and Lee DK: Selection and optimization of asymmetric siRNA targeting the human c-MET gene. Mol Cells. 32:543–548. 2011. View Article : Google Scholar : PubMed/NCBI
19	Sun X, Rogoff HA and Li CJ: Asymmetric RNA duplexes mediate RNA interference in mammalian cells. Nat Biotechnol. 26:1379–1382. 2008. View Article : Google Scholar : PubMed/NCBI
20	Daneshmand S, Quek ML, Lin E, et al: Glucose-regulated protein GRP78 is up-regulated in prostate cancer and correlates with recurrence and survival. Hum Pathol. 38:1547–1552. 2007. View Article : Google Scholar : PubMed/NCBI
21	Lv L, Xiao XY, Gu ZH, Zeng FQ, Huang LQ and Jiang GS: Silencing USP22 by asymmetric structure of interfering RNA inhibits proliferation and induces cell cycle arrest in bladder cancer cells. Mol Cell Biochem. 346:11–21. 2011. View Article : Google Scholar : PubMed/NCBI
22	Yin Y, Chen X, Zhang CD, et al: Asymmetric siRNA targeting the bcl-2 gene inhibits the proliferation of cancer cells in vitro and in vivo. Int J Oncol. 42:253–260. 2013.PubMed/NCBI
23	Yuan Z, Wu X, Liu C, Xu G and Wu Z: Asymmetric siRNA: new strategy to improve specificity and reduce off-target gene expression. Hum Gene Ther. 23:521–532. 2012. View Article : Google Scholar : PubMed/NCBI
24	Zhang LH and Zhang X: Roles of GRP78 in physiology and cancer. J Cell Biochem. 110:1299–1305. 2010. View Article : Google Scholar : PubMed/NCBI
25	Philippova M, Ivanov D, Joshi MB, et al: Identification of proteins associating with glycosylphosphatidylinositol-anchored T-cadherin on the surface of vascular endothelial cells: role for Grp78/BiP in T-cadherin-dependent cell survival. Mol Cell Biol. 28:4004–4017. 2008. View Article : Google Scholar
26	Shani G, Fischer WH, Justice NJ, Kelber JA, Vale W and Gray PC: GRP78 and Cripto form a complex at the cell surface and collaborate to inhibit transforming growth factor beta signaling and enhance cell growth. Mol Cell Biol. 28:666–677. 2008. View Article : Google Scholar : PubMed/NCBI
27	Cohen M and Petignat P: Purified autoantibodies against glucose-regulated protein 78 (GRP78) promote apoptosis and decrease invasiveness of ovarian cancer cells. Cancer Lett. 309:104–109. 2011. View Article : Google Scholar
28	Rasche L, Duell J, Morgner C, et al: The natural human IgM antibody PAT-SM6 induces apoptosis in primary human multiple myeloma cells by targeting heat shock protein GRP78. PLoS One. 8:e634142013. View Article : Google Scholar : PubMed/NCBI
29	Suzuki T, Lu J, Zahed M, Kita K and Suzuki N: Reduction of GRP78 expression with siRNA activates unfolded protein response leading to apoptosis in HeLa cells. Arch Biochem Biophys. 468:1–14. 2007. View Article : Google Scholar : PubMed/NCBI
30	Xing X, Li Y, Liu H, Wang L and Sun L: Glucose regulated protein 78 (GRP78) is overexpressed in colorectal carcinoma and regulates colorectal carcinoma cell growth and apoptosis. Acta Histochem. 113:777–782. 2011. View Article : Google Scholar : PubMed/NCBI
31	Chang YJ, Huang YP, Li ZL and Chen CH: GRP78 knockdown enhances apoptosis via the down-regulation of oxidative stress and Akt pathway after epirubicin treatment in colon cancer DLD-1 cells. PLoS One. 7:e351232012. View Article : Google Scholar : PubMed/NCBI
32	Fu Y, Wey S, Wang M, et al: Pten null prostate tumorigenesis and AKT activation are blocked by targeted knockout of ER chaperone GRP78/BiP in prostate epithelium. Proc Natl Acad Sci USA. 105:19444–19449. 2008. View Article : Google Scholar : PubMed/NCBI
33	Zhang LH, Yang XL, Zhang X, Cheng JX and Zhang W: Association of elevated GRP78 expression with increased astrocytoma malignancy via Akt and ERK pathways. Brain Res. 1371:23–31. 2011. View Article : Google Scholar : PubMed/NCBI
34	Cardone MH, Roy N, Stennicke HR, et al: Regulation of cell death protease caspase-9 by phosphorylation. Science. 282:1318–1321. 1998. View Article : Google Scholar : PubMed/NCBI
35	Li P, Nijhawan D, Budihardjo I, et al: Cytochrome c and dATP-dependent formation of Apaf-1/caspase-9 complex initiates an apoptotic protease cascade. Cell. 91:479–489. 1997. View Article : Google Scholar : PubMed/NCBI
36	Chiu CC, Lin CY, Lee LY, et al: Glucose-regulated protein 78 regulates multiple malignant phenotypes in head and neck cancer and may serve as a molecular target of therapeutic intervention. Mol Cancer Ther. 7:2788–2797. 2008. View Article : Google Scholar : PubMed/NCBI
37	McInroy L and Määttä A: Down-regulation of vimentin expression inhibits carcinoma cell migration and adhesion. Biochem Biophys Res Commun. 360:109–114. 2007. View Article : Google Scholar : PubMed/NCBI