The influence of lentivirus-mediated CXCR4 RNA interference on hepatic metastasis of colorectal cancer

Wang,Tian-Bao; Hu,Bao-Guang; Liu,Da-Wei; Shi,Han-Ping; Dong,Wen-Guang

doi:10.3892/ijo.2014.2348

The influence of lentivirus-mediated CXCR4 RNA interference on hepatic metastasis of colorectal cancer

Authors:
- Tian-Bao Wang
- Bao-Guang Hu
- Da-Wei Liu
- Han-Ping Shi
- Wen-Guang Dong
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Affiliations: Department of Surgery, The First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong 510080, P.R. China, Department of Surgery, Prince of Wales Hospital, The Chinese University of Hong Kong, Shatin, Hong Kong, P.R. China, Department of Pathology, The First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong 510080, P.R. China
Published online on: March 19, 2014 https://doi.org/10.3892/ijo.2014.2348
Pages: 1861-1869
Copyright: © Wang et al. This is an open access article distributed under the terms of Creative Commons Attribution License [CC BY_NC 3.0].

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Abstract

The aim of this study was to construct a lentiviral vector of CXCR4-siRNA (Lenti-CXCR4-siRNA) and investigate whether the vector can inhibit the growth, migration, invasion and hepatic metastasis of colorectal cancer (CRC). RT-PCR and western blotting were employed to identify the ideal RNA interference sequence. Lenti-CXCR4-siRNA was constructed and transfected into the SW480 cell line. We used RT-PCR and western blotting to measure the expression of CXCR4 RNA and protein, respectively; the MTS assay to assess the proliferation of SW480 cells; transwell chambers to estimate the inhibitory effect on migration and invasion; and the Balb/c nude mouse model of CRC to examine the inhibition of hepatic metastasis. The relative expression of the CXCR4 gene and protein was 5.4 and 18.95%, respectively, in the siCXCR4 group. The genes in the expression plasmid pLenti-CXCR4-siRNA were in the correct order. In the SW480, nonsense control (NC) and the Lenti-CXCR4-siRNA groups CXCR4 RNA levels were, respectively, 0.54±0.06, 1.00±0.03 and 0.11±0.04 (P=0.0001); CXCR4 protein levels were 0.60±0.03, 0.72±0.03 and 0.18±0.02 (P=0.0001); the OD value was 1.38±0.04 (P=0.0050), 1.28±0.05 (P=0.0256) and 0.92±0.06; SW480 cell number in migration test was 32±6.85, 32.63±1.69 and 0.75±0.71 (P=0.0000); SW480 cell number in the invasion test was 29.13±10.3, 30.38±6.09 and 0.63±0.74 (P=0.0000); hepatic metastasis number was 7.10±3.98 (P=0.034), 7.50±4.09 (P=0.019) and (3.50±2.51); hepatic metastasis mean weight (in g) was 2.25±2.51 (P=0.000), 2.11±2.38 (P=0.000) and 1.45±2.07. Lenti-CXCR4-siRNA constructs were correctly constructed and effectively inhibit the expression of CXCR4 RNA and protein, reducing the proliferation, migration, invasion capacity of SW480 cells and hepatic metastasis of CRC.

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1.	Wichmann MW, Beukes E, Esufali ST, Plaumann L and Maddern G: Five-year results of surgical colorectal cancer treatment in rural Australia. ANZ J Surg. 83:112–117. 2013.
2.	Shin HN, Moon HH and Ku JL: Stromal cell-derived factor-1α and macrophage migration-inhibitory factor induce metastatic behavior in CXCR4-expressing colon cancer cells. Int J Mol Med. 30:1537–1543. 2012.
3.	Yasuoka H, Kodama R, Hirokawa M, Takamura Y, Miyauchi A, Sanke T and Nakamura Y: CXCR4 expression in papillary thyroid carcinoma: induction by nitric oxide and correlation with lymph node metastasis. BMC Cancer. 8:2742008. View Article : Google Scholar : PubMed/NCBI
4.	Cavallaro S: CXCR4/CXCL12 in non-small-cell lung cancer metastasis to the brain. Int J Mol Sci. 14:1713–1727. 2013. View Article : Google Scholar : PubMed/NCBI
5.	Sekiya R, Kajiyama H, Sakai K, Umezu T, Mizuno M, Shibata K, Yamamoto E, Fujiwara S, Nagasaka T and Kikkawa F: Expression of CXCR4 indicates poor prognosis in patients with clear cell carcinoma of the ovary. Hum Pathol. 43:904–910. 2012. View Article : Google Scholar : PubMed/NCBI
6.	Jung SJ, Kim CI, Park CH, Chang HS, Kim BH, Choi MS and Jung HR: Correlation between chemokine receptor CXCR4 expression and prognostic factors in patients with prostate cancer. Korean J Urol. 52:607–611. 2011. View Article : Google Scholar : PubMed/NCBI
7.	Wang L, Huang T, Chen W, Gao X, Zhou T, Wu Z and Sun Y: Silencing of CXCR4 by RNA interference inhibits cell growth and metastasis in human renal cancer cells. Oncol Rep. 28:2043–2048. 2012.PubMed/NCBI
8.	Gros SJ, Kurschat N, Drenckhan A, Dohrmann T, Forberich E, Effenberger K, Reichelt U, Hoffman RM, Pantel K, Kaifi JT and Izbicki JR: Involvement of CXCR4 chemokine receptor in metastastic HER2-positive esophagealcancer. PLoS One. 7:e472872012. View Article : Google Scholar : PubMed/NCBI
9.	Zhong W, Chen W, Zhang D, Sun J, Li Y, Zhang J, Gao Y, Zhou W and Li S: CXCL12/CXCR4 axis plays pivotal roles in the organ-specific metastasis of pancreatic adenocarcinoma: a clinical study. Exp Ther Med. 4:363–369. 2012.
10.	Lee HJ and Jo DY: The role of the CXCR4/CXCL12 axis and its clinical implications in gastric cancer. Histol Histopathol. 27:1155–1161. 2012.PubMed/NCBI
11.	Zhang SS, Han ZP, Jing YY, Tao SF, Li TJ, Wang H, Wang Y, Li R, Yang Y, Zhao X, Xu XD, Yu ED, Rui YC, Liu HJ, Zhang L and Wei LX: CD133(+)CXCR4(+) colon cancer cells exhibit metastatic potential and predict poor prognosis of patients. BMC Med. 10:852012.
12.	Wang TB, Chen ZG, Wei XQ, Wei B and Dong WG: Serum vascular endothelial growth factor-C and lymphangiogenesis are associated with the lymph node metastasis and prognosis of patients with colorectal cancer. ANZ J Surg. 81:894–899. 2011.PubMed/NCBI
13.	De Falco V, Guarino V, Avilla E, Castellone MD, Salerno P, Salvatore G, Faviana P, Basolo F, Santoro M and Melillo RM: Biological role and potential therapeutic targeting of the chemokine receptor CXCR4 in undifferentiated thyroid cancer. Cancer Res. 67:11821–11829. 2007.PubMed/NCBI
14.	Li JK, Yu L, Shen Y, Zhou LS, Wang YC and Zhang JH: Inhibition of CXCR4 activity with AMD3100 decreases invasion of human colorectal cancer cells in vitro. World J Gastroenterol. 14:2308–2313. 2008. View Article : Google Scholar : PubMed/NCBI
15.	Kim JM, Lee YH, Ku CR and Lee EJ: The cyclic pentapep-tide d-Arg3FC131, a CXCR4 antagonist, induces apoptosis of somatotrope tumor and inhibits tumor growth in nude mice. Endocrinology. 152:536–544. 2011. View Article : Google Scholar : PubMed/NCBI
16.	Murakami T, Kawada K, Iwamoto M, Akagami M, Hida K, Nakanishi Y, Kanda K, Kawada M, Seno H, Taketo MM and Sakai Y: The role of CXCR3 and CXCR4 in colorectal cancer metastasis. Int J Cancer. 132:276–287. 2013. View Article : Google Scholar : PubMed/NCBI
17.	Heckmann D, Laufs S, Maier P, Zucknick M, Giordano FA, Veldwijk MR, Eckstein V, Wenz F, Zeller WJ, Fruehauf S and Allgayer H: A lentiviral CXCR4 overexpression and knockdown model in colorectal cancer cell lines reveals plerixafor-dependent suppression of SDF-1α-induced migration and invasion. Onkologie. 34:502–508. 2011.PubMed/NCBI
18.	Liang Z, Yoon Y, Votaw J, Goodman MM, Williams L and Shim H: Silencing of CXCR4 blocks breast cancer metastasis. Cancer Res. 65:967–971. 2005.PubMed/NCBI
19.	Smith MC, Luker KE, Garbow JR, Prior JL, Jackson E, Piwnica-Worms D and Luker GD: CXCR4 regulates growth of both primary and metastatic breast cancer. Cancer Res. 64:8604–8612. 2004. View Article : Google Scholar : PubMed/NCBI
20.	Liang Z, Wu H, Reddy S, Zhu A, Wang S, Blevins D, Yoon Y, Zhang Y and Shim H: Blockade of invasion and metastasis of breast cancer cells via targeting CXCR4 with an artificial microRNA. Biochem Biophys Res Commun. 363:542–546. 2007. View Article : Google Scholar : PubMed/NCBI
21.	Beverly AT and Simon PF: CXCL12 (SDF-1)/CXCR4 pathway in cancer. Clin Cancer Res. 16:2927–2931. 2010. View Article : Google Scholar : PubMed/NCBI
22.	Meads MB, Hazlehurst LA and Dalton WS: The bone marrow microenvironment as a tumor sanctuary and contributor to drug resistance. Clin Cancer Res. 14:2519–2526. 2008. View Article : Google Scholar : PubMed/NCBI
23.	Balkwill F: Cancer and the chemokine network. Nat Rev Cancer. 4:540–550. 2004. View Article : Google Scholar
24.	Petit I, Jin D and Rafii S: The SDF-1-CXCR4 signaling pathway: a molecular hub modulating neo-angiogenesis. Trends Immunol. 28:299–307. 2007. View Article : Google Scholar : PubMed/NCBI
25.	Rubie C, Frick VO, Ghadjar P, Wagner M, Justinger C, Faust SK, Vicinus B, Gräber S, Kollmar O and Schilling MK: CXC receptor-4 mRNA silencing abrogates CXCL12-induced migration of colorectal cancer cells. J Transl Med. 9:222011. View Article : Google Scholar : PubMed/NCBI
26.	Wang SC, Lin JK, Wang HS, Yang SH, Li AF and Chang SC: Nuclear expression of CXCR4 is associated with advanced colorectal cancer. Int J Colorectal Dis. 25:1185–1191. 2010. View Article : Google Scholar : PubMed/NCBI
27.	Fukunaga S, Maeda K, Noda E, Inoue T, Wada K and Hirakawa K: Association between expression of vascular endothelial growth factor C, chemokine receptor CXCR4 and lymph node metastasis in colorectal cancer. Oncology. 71:204–211. 2006. View Article : Google Scholar : PubMed/NCBI
28.	Matsusue R, Kubo H, Hisamori S, Okoshi K, Takagi H, Hida K, Nakano K, Itami A, Kawada K, Nagayama S and Sakai Y: Hepatic stellate cells promote liver metastasis of colon cancer cells by the action of SDF-1/CXCR4 axis. Ann Surg Oncol. 16:2645–2653. 2009. View Article : Google Scholar : PubMed/NCBI
29.	Mongan JP, Fadul CE, Cole BF, Zaki BI, Suriawinata AA, Ripple GH, Tosteson TD and Pipas JM: Brain metastases from colorectal cancer: risk factors, incidence, and the possible role of chemokines. Clin Colorectal Cancer. 8:100–105. 2009. View Article : Google Scholar
30.	Speetjens FM, Liefers GJ, Korbee CJ, Mesker WE, van de Velde CJ, van Vlierberghe RL, Morreau H, Tollenaar RA and Kuppen PJ: Nuclear localization of CXCR4 determines prognosis for colorectal cancer patients. Cancer Microenviron. 2:1–7. 2009. View Article : Google Scholar : PubMed/NCBI
31.	Brand S, Dambacher J, Beigel F, Olszak T, Diebold J, Otte JM, Göke B and Eichhorst ST: CXCR4 and CXCL12 are inversely expressed in colorectal cancer cells and modulate cancer cell migration, invasion and MMP-9 activation. Exp Cell Res. 310:117–130. 2005. View Article : Google Scholar : PubMed/NCBI
32.	Ottaiano A, Franco R, Aiello Talamanca A, Liguori G, Tatangelo F, Delrio P, Nasti G, Barletta E, Facchini G, Daniele B, Di Blasi A, Napolitano M, Ieranò C, Calemma R, Leonardi E, Albino V, De Angelis V, Falanga M, Boccia V, Capuozzo M, Parisi V, Botti G, Castello G, Vincenzo Iaffaioli R and Scala S: Overexpression of both CXC chemokine receptor 4 and vascular endothelial growth factor proteins predicts early distant relapse in stage II-III colorectal cancer patients. Clin Cancer Res. 12:2795–2803. 2006. View Article : Google Scholar
33.	Zhu Y, Yang P, Zhang X, Zhang L, Cui G, Wang Q, Lv L, Zhang Y, Xin X, Yan T, Zhao M and Zhang N: The effect and mechanism of CXCR4 silencing on metastasis suppression of human glioma u87 cell line. Anat Rec (Hoboken). 296:1857–1864. 2013. View Article : Google Scholar : PubMed/NCBI
34.	Shen B, Zheng MQ, Lu JW, Jiang Q, Wang TH and Huang XE: CXCL12-CXCR4 promotes proliferation and invasion of pancreatic cancer cells. Asian Pac J Cancer Prev. 14:5403–5408. 2013. View Article : Google Scholar : PubMed/NCBI
35.	Sun X, Charbonneau C, Wei L, Yang W, Chen Q and Terek RM: CXCR4-targeted therapy inhibits VEGF expression and chondrosarcoma angiogenesis and metastasis. Mol Cancer Ther. 12:1163–1170. 2013. View Article : Google Scholar : PubMed/NCBI
36.	Wang B, Wang W, Niu W, Liu E, Liu X, Wang J, Peng C, Liu S, Xu L, Wang L and Niu J: SDF-1/CXCR4 axis promotes directional migration of colorectal cancer cells through upregulation of integrin αvβ6. Carcinogenesis. 35:282–291. 2014.PubMed/NCBI
37.	Xue B, Wu W, Huang K, Xie T, Xu X, Zhang H, Qi C, Ge J and Yu Y: Stromal cell-derived factor-1 (SDF-1) enhances cells invasion by αvβ6 integrin-mediated signaling in ovarian cancer. Mol Cell Biochem. 380:177–184. 2013.
38.	Kiss DL, Windus LC and Avery VM: Chemokine receptor expression on integrin-mediated stellate projections of prostate cancer cells in 3D culture. Cytokine. 64:122–130. 2013. View Article : Google Scholar : PubMed/NCBI
39.	Paddison PJ, Caudy AA, Bernstein E, Hannon GJ and Conklin DS: Short hairpin RNAs (shRNAs) induce sequence-specific silencing in mammalian cells. Genes Dev. 16:948–958. 2002. View Article : Google Scholar : PubMed/NCBI
40.	De Maria R, Miretti S, Iussich S, Olivero M, Morello E, Bertotti A, Christensen JG, Biolatti B, Levine RA, Buracco P and Di Renzo MF: Met oncogene activation qualifies spontaneous canine steosarcoma as a suitable pre-clinical model of human steosarcoma. J Pathol. 218:399–408. 2009.PubMed/NCBI
41.	Chen D, Huang H, Pan C, Wang J, Zhang B and Wang A: Antitumor effects of targeting hTERT lentivirus-mediated RNA interference against KB cell lines. Oncol Res. 17:621–630. 2009. View Article : Google Scholar : PubMed/NCBI
42.	Chunkang C, Rui Y, Feng X, Juan G, Xi Z, Lingyun W, Xiao L and Jianmin W: The roles of SDF-1/CXCR4 axis and its relationship with apoptosis in the myelodysplastic syndromes. Med Oncol. 28(Suppl 1): 494–500. 2011. View Article : Google Scholar : PubMed/NCBI
43.	Yu T, Wu Y, Huang Y, Yan C, Liu Y, Wang Z, Wang X, Wen Y, Wang C and Li L: RNAi targeting CXCR4 inhibits tumor growth through inducing cell cycle arrest and apoptosis. Mol Ther. 20:398–407. 2012. View Article : Google Scholar : PubMed/NCBI
44.	Wang Q, Diao X, Sun J and Chen Z: Regulation of VEGF, MMP-9 and metastasis by CXCR4 in a prostate cancer cell line. Cell Biol Int. 35:897–904. 2011. View Article : Google Scholar : PubMed/NCBI