Objective: In this study, we inhibited the expression of VEGF by short hairpin RNA (shRNA) in SKOV3 ovarian cancer cells in vitro and in vivo to explore the antitumor efficacy of shRNA in ovarian cancer cells. Methods: ShRNA targeting VEGF was cloned into pGenesil-2 plasmid vector and then transfected into SKOV3 ovarian cancer cells using liposome. Silencing of VEGF expression was measured by RT-PCR and ELISA assays. Furthermore, the growth inhibition capacity of shRNA on SKOV3 intraperitoneal ovarian carcinomatosis was tested in nude mice. Tumor weight was measured. Microvessel density, number of apoptotic cells and proliferation index in tumor tissues were assessed by CD31, TUNEL and PCNA immunostaining. Results: shRNA targeting VEGF significantly silenced VEGF expression in SKOV3 ovarian cancer cells, confirmed by RT-PCR and ELISA assays (p < 0.05). In vivo, the shRNA reduced tumor weight by ∼60.3% compared with control groups (p < 0.05), accompanied with angiogenesis inhibition (p < 0.01) and apoptosis induction (p < 0.01). Conclusion: Our data indicated that shRNA-mediated silencing of VEGF might be a promising therapeutic strategy against ovarian cancer by reducing angiogenesis and inducing apoptosis.

1.
du Bois A, Luck HJ, Meier W, Adams HP, Mobus V, Costa S, Bauknecht T, Richter B, Warm M, Schroder W, Olbricht S, Nitz U, Jackisch C, Emons G, Wagner U, Kuhn W, Pfisterer J: A randomized clinical trial of cisplatin/paclitaxel versus carboplatin/paclitaxel as first-line treatment of ovarian cancer. J Natl Cancer Inst 2003;95:1320–1329.
2.
Szotek PP, Pieretti-Vanmarcke R, Masiakos PT, Dinulescu DM, Connolly D, Foster R, Dombkowski D, Preffer F, Maclaughlin DT, Donahoe PK: Ovarian cancer side population defines cells with stem cell-like characteristics and Mullerian inhibiting substance responsiveness. Proc Natl Acad Sci USA 2006;103:11154–11159.
3.
Jemal A, Siegel R, Ward E, Hao Y, Xu J, Murray T, Thun MJ: Cancer statistics, 2008. CA Cancer J Clin 2008;58:71–96.
4.
Folkman J: What is the evidence that tumors are angiogenesis dependent? J Natl Cancer Inst 1990;82:4–6.
5.
Robinson CJ, Stringer SE: The splice variants of vascular endothelial growth factor (VEGF) and their receptors. J Cell Sci 2001;114:853–865.
6.
Bachelder RE, Crago A, Chung J, Wendt MA, Shaw LM, Robinson G, Mercurio AM: Vascular endothelial growth factor is an autocrine survival factor for neuropilin-expressing breast carcinoma cells. Cancer Res 2001;61:5736–5740.
7.
Ohm JE, Carbone DP: VEGF as a mediator of tumor-associated immunodeficiency. Immunol Res 2001;23:263–272.
8.
Ferrara N: Molecular and biological properties of vascular endothelial growth factor. J Mol Med 1999;77:527–543.
9.
Abu-Jawdeh GM, Faix JD, Niloff J, Tognazzi K, Manseau E, Dvorak HF, Brown LF: Strong expression of vascular permeability factor (vascular endothelial growth factor) and its receptors in ovarian borderline and malignant neoplasms. Lab Invest 1996;74:1105–1115.
10.
Cooper BC, Ritchie JM, Broghammer CL, Coffin J, Sorosky JI, Buller RE, Hendrix MJ, Sood AK: Preoperative serum vascular endothelial growth factor levels: significance in ovarian cancer. Clin Cancer Res 2002;8:3193–3197.
11.
Chen CA, Cheng WF, Lee CN, Chen TM, Kung CC, Hsieh FJ, Hsieh CY: Serum vascular endothelial growth factor in epithelial ovarian neoplasms: correlation with patient survival. Gynecol Oncol 1999;74:235–240.
12.
Boocock CA, Charnock-Jones DS, Sharkey AM, McLaren J, Barker PJ, Wright KA, Twentyman PR, Smith SK: Expression of vascular endothelial growth factor and its receptors flt and KDR in ovarian carcinoma. J Natl Cancer Inst 1995;87:506–516.
13.
Paley PJ, Staskus KA, Gebhard K, Mohanraj D, Twiggs LB, Carson LF, Ramakrishnan S: Vascular endothelial growth factor expression in early stage ovarian carcinoma. Cancer 1997;80:98–106.
14.
Oehler MK, Caffier H: Prognostic relevance of serum vascular endothelial growth factor in ovarian cancer. Anticancer Res 2000;20:5109–5112.
15.
Heidersbach A, Gaspar-Maia A, McManus MT, Ramalho-Santos M: RNA interference in embryonic stem cells and the prospects for future therapies. Gene Ther 2006;13:478–486.
16.
Pai SI, Lin YY, Macaes B, Meneshian A, Hung CF, Wu TC: Prospects of RNA interference therapy for cancer. Gene Ther 2006;13:464–477.
17.
Hannon GJ, Rossi JJ: Unlocking the potential of the human genome with RNA interference. Nature 2004;431:371–378.
18.
Reich SJ, Fosnot J, Kuroki A, Tang W, Yang X, Maguire AM, Bennett J, Tolentino MJ: Small interfering RNA (siRNA) targeting VEGF effectively inhibits ocular neovascularization in a mouse model. Mol Vis 2003;9:210–216.
19.
Lin X, Chen X, Wei Y, Zhao J, Fan L, Wen Y, Wu H, Zhao X: Efficient inhibition of intraperitoneal human ovarian cancer growth and prolonged survival by gene transfer of vesicular stomatitis virus matrix protein in nude mice. Gynecol Oncol 2007;104:540–546.
20.
Oka N, Soeda A, Inagaki A, Onodera M, Maruyama H, Hara A, Kunisada T, Mori H, Iwama T: VEGF promotes tumorigenesis and angiogenesis of human glioblastoma stem cells. Biochem Biophys Res Commun 2007;360:553–559.
21.
Lin XJ, Chen XC, Wang L, Wei YQ, Kan B, Wen YJ, He X, Zhao X: Dynamic progression of an intraperitoneal xenograft model of human ovarian cancer and its potential for preclinical trials. J Exp Clin Cancer Res 2007;26:467–474.
22.
Markman M, Walker JL: Intraperitoneal chemotherapy of ovarian cancer: a review, with a focus on practical aspects of treatment. J Clin Oncol 2006;24:988–994.
23.
Jackson MW, Roberts JS, Heckford SE, Ricciardelli C, Stahl J, Choong C, Horsfall DJ, Tilley WD: A potential autocrine role for vascular endothelial growth factor in prostate cancer. Cancer Res 2002;62:854–859.
24.
Tian X, Song S, Wu J, Meng L, Dong Z, Shou C: Vascular endothelial growth factor: Acting as an autocrine growth factor for human gastric adenocarcinoma cell MGC803. Biochem Biophys Res Commun 2001;286:505–512.
25.
Petersen I: Antiangiogenesis, anti-VEGF(R) and outlook. Recent Results Cancer Res 2007;176:189–199.
26.
Ellis LM, Hicklin DJ: VEGF-targeted therapy: mechanisms of anti-tumour activity. Nat Rev Cancer 2008;8:579–591.
27.
Drewe E, Powell RJ: Clinically useful monoclonal antibodies in treatment. J Clin Pathol 2002;55:81–85.
28.
Kong HL, Crystal RG: Gene therapy strategies for tumor antiangiogenesis. J Natl Cancer Inst 1998;90:273–286.
29.
Nabel GJ, Nabel EG, Yang ZY, Fox BA, Plautz GE, Gao X, Huang L, Shu S, Gordon D, Chang AE: Direct gene transfer with DNA-liposome complexes in melanoma: expression, biologic activity, and lack of toxicity in humans. Proc Natl Acad Sci USA 1993;90:11307–11311.
30.
Spankuch B, Matthess Y, Knecht R, Zimmer B, Kaufmann M, Strebhardt K: Cancer inhibition in nude mice after systemic application of U6 promoter-driven short hairpin RNAs against PLK1. J Natl Cancer Inst 2004;96:862–872.
31.
Wyatt CA, Geoghegan JC, Brinckerhoff CE: Short hairpin RNA-mediated inhibition of matrix metalloproteinase-1 in MDA-231 cells: effects on matrix destruction and tumor growth. Cancer Res 2005;65:11101–11108.
32.
Shi Y, Zou M, Collison K, Baitei EY, Al-Makhalafi Z, Farid NR, Al-Mohanna FA: Ribonucleic acid interference targeting S100A4 (Mts1) suppresses tumor growth and metastasis of anaplastic thyroid carcinoma in a mouse model. J Clin Endocrinol Metab 2006;91:2373–2379.
33.
Zhdanov RI, Podobed OV, Vlassov VV: Cationic lipid-DNA complexes-lipoplexes-for gene transfer and therapy. Bioelectrochemistry 2002;58:53–64.
34.
Young LS, Mautner V: The promise and potential hazards of adenovirus gene therapy. Gut 2001;48:733–736.
35.
Jiao X, Wang RY, Qiu Q, Alter HJ, Shih JW: Enhanced hepatitis C virus NS3 specific Th1 immune responses induced by co-delivery of protein antigen and CpG with cationic liposomes. J Gen Virol 2004;85:1545–1553.
36.
Zelphati O, Szoka FC Jr: Mechanism of oligonucleotide release from cationic liposomes. Proc Natl Acad Sci USA 1996;93:11493–11498.
37.
Sangare L, Morisset R, Omri A, Ravaoarinoro M: Incorporation rates, stabilities, cytotoxicities and release of liposomal tetracycline and doxycycline in human serum. J Antimicrob Chemother 1998;42:831–834.
38.
Webb MS, Harasym TO, Masin D, Bally MB, Mayer LD: Sphingomyelin-cholesterol liposomes significantly enhance the pharmacokinetic and therapeutic properties of vincristine in murine and human tumour models. Br J Cancer 1995;72:896–904.
39.
Yuan F, Dellian M, Fukumura D, Leunig M, Berk DA, Torchilin VP, Jain RK: Vascular permeability in a human tumor xenograft: molecular size dependence and cutoff size. Cancer Res 1995;55:3752–3756.
40.
Guan H, Zhou Z, Wang H, Jia SF, Liu W, Kleinerman ES: A small interfering RNA targeting vascular endothelial growth factor inhibits Ewing’s sarcoma growth in a xenograft mouse model. Clin Cancer Res 2005;11:2662–2669.
41.
Shaheen RM, Davis DW, Liu W, Zebrowski BK, Wilson MR, Bucana CD, McConkey DJ, McMahon G, Ellis LM: Antiangiogenic therapy targeting the tyrosine kinase receptor for vascular endothelial growth factor receptor inhibits the growth of colon cancer liver metastasis and induces tumor and endothelial cell apoptosis. Cancer Res 1999;59:5412–5416.
42.
Wu Y, Li ZY, Zhao X, Kan B, Wei YQ: Inhibition of ovarian tumor growth by gene therapy with recombinant soluble vascular endothelial growth factor receptor 2. Hum Gene Ther 2006;17:941–948.
43.
Mulkeen AL, Silva T, Yoo PS, Schmitz JC, Uchio E, Chu E, Cha C: Short interfering RNA-mediated gene silencing of vascular endothelial growth factor: effects on cellular proliferation in colon cancer cells. Arch Surg 2006;141:367–374.
Copyright / Drug Dosage / Disclaimer
Copyright: All rights reserved. No part of this publication may be translated into other languages, reproduced or utilized in any form or by any means, electronic or mechanical, including photocopying, recording, microcopying, or by any information storage and retrieval system, without permission in writing from the publisher.
Drug Dosage: The authors and the publisher have exerted every effort to ensure that drug selection and dosage set forth in this text are in accord with current recommendations and practice at the time of publication. However, in view of ongoing research, changes in government regulations, and the constant flow of information relating to drug therapy and drug reactions, the reader is urged to check the package insert for each drug for any changes in indications and dosage and for added warnings and precautions. This is particularly important when the recommended agent is a new and/or infrequently employed drug.
Disclaimer: The statements, opinions and data contained in this publication are solely those of the individual authors and contributors and not of the publishers and the editor(s). The appearance of advertisements or/and product references in the publication is not a warranty, endorsement, or approval of the products or services advertised or of their effectiveness, quality or safety. The publisher and the editor(s) disclaim responsibility for any injury to persons or property resulting from any ideas, methods, instructions or products referred to in the content or advertisements.
You do not currently have access to this content.