Therapeutic efficacy of an oncolytic adenovirus containing RGD ligand in minor capsid protein IX and Fiber, Δ24DoubleRGD, in an ovarian cancer model

Lena J Gamble, Hideyo Ugai, Minghui Wang, Anton V Borovjagin, Qiana L Matthews


Ovarian cancer is the leading cause of gynecological disease death despite advances in medicine. Therefore, novel strategies are required for ovarian cancer therapy. Conditionally replicative adenoviruses (CRAds), genetically modified as anti-cancer therapeutics, are one of the most attractive candidate agents for cancer therapy. However, a paucity of coxsackie B virus and adenovirus receptor (CAR) expression on the surface of ovarian cancer cells has impeded treatment of ovarian cancer using this approach.
This study sought to engineer a CRAd with enhanced oncolytic ability in ovarian cancer cells, “Δ24DoubleRGD.” Δ24DoubleRGD carries an arginine-glycine-aspartate (RGD) motif incorporated into both fiber and capsid protein IX (pIX) and its oncolytic efficacy was evaluated in ovarian cancer. In vitro analysis of cell viability showed that infection of ovarian cancer cells with Δ24DoubleRGD leads to increased cell killing relative to the control CRAds. Data from this study suggested that not only an increase in number of RGD motifs on the CRAd capsid, but also a change in the repertoir of targeted integrins could lead to enhanced oncolytic potency of Δ24DoubleRGD in ovarian cancer cells in vitro. In an intraperitoneal model of ovarian cancer, mice injected with Δ24DoubleRGD showed, however, a similar survival rate as mice treated with control CRAds.


Conditionally replicative adenoviruses (CRAds); Ovarian Cancer; arginine-glycine-aspartate (RGD) motif; capsid-modification; anti-cancer therapy


Barnes MN, Coolidge CJ, Hemminki A, Alvarez RD & Curiel DT 2002 Conditionally replicative adenoviruses for ovarian cancer therapy. Mol Cancer Ther 1 435-439.

Bauerschmitz GJ, Lam JT, Kanerva A, Suzuki K, Nettelbeck DM, Dmitriev I, Krasnykh V, Mikheeva GV, Barnes MN, Alvarez RD, et al. 2002 Treatment of ovarian cancer with a tropism modified oncolytic adenovirus. Cancer Res 62 1266-1270.

Blackwell JL, Li H, Gomez-Navarro J, Dmitriev I, Krasnykh V, Richter CA, Shaw DR, Alvarez RD, Curiel DT & Strong TV 2000 Using a tropism modified adenoviral vector to circumvent inhibitory factors in ascites fluid. Hum Gene Ther 11 1657-1669.

Borovjagin AV, McNally LR, Wang M, Curiel DT, Mac-Dougall MJ & Zinn KR 2010 Noninvasive monitoring of mRFP1 and mCherry labeled oncolytic adenoviruses in an orthotopic breast cancer model by spectral imaging. Mol Imaging 9 59-75.

Brüning A, Köhler T, Quist S, Wang-Gohrke S, Moebus VJ, Kreienberg R & Runnebaum IB 2001 Adenoviral transduction efficiency of ovarian cancer cells can be limited by loss of integrin beta3 subunit expression and increased by reconstitution of integrin alphavbeta3. Hum Gene There 12 391-399.

Campos SK, Parrott MB & Barry MA 2004 Avidin-based targeting and purification of a protein IX modified, metabolically biotinylated adenoviral vector. Mol Ther 9 942-954.

Cannistra SA, Ottensmeier C, Niloff J, Orta B & DiCarlo J 1995 Expression and function of beta 1 and alpha v beta 3 integrins in ovarian cancer. Gynecol Oncol 58 216-225.

Carmeliet P 2000 Mechanisms of angiogenesis and arterio-genesis. Nat Med 6 389-395.

Carreiras F, Denoux Y, Staedel C, Lehmann M, Sichel F & Gauduchon P 1996 Expression and localization of alpha v integrins and their ligand vitronectin in normal ovarian epithelium and in ovarian carcinoma. Gynecol Oncol 62 260-267.

Chartier C, Degryse E, Gantzer M, Dieterle A, Pavirani A & Mehtali M 1996 Efficient generation of recombinant adenovirus vectors by homologous recombination in Escherichia coli. J Virol 70 4805-4810.

Choi M, Fuller CD, Thomas CR & Wang SJ 2008 Conditional survival in ovarian cancer: results from the SEER dataset 1988-2001. Gynecol Oncol 109 203-209.

Contag CH, Jenkins D, Contag PR & Negrin RS 2000 Use of reporter genes for optical measurements of neoplastic disease in vivo. Neoplasia 2 41-52.

Corney DC, Flesken-Nikitin A, Choi J & Nikitin AY 2008 Role of p53 and Rb in ovarian cancer. Adv Exp Med Biol 622 99-9117.

Dmitriev I, Krasnykh V, Miller CR, Wang M, Kashentseva E, Mikheeva G, Belousova N & Curiel DT 1998 An adeno-virus vector with genetically modified fibers demonstrates expanded tropism via utilization of a coxsackievirus and adenovirus receptor independent cell entry mechanism. J Virol 72 9706-9713.

Dmitriev IP, Kashentseva EA & Curiel DT 2002 Engineering of adenovirus vectors containing heterologous peptide sequences in the C terminus of capsid protein IX. J Virol 76 6893-6899.

Dyson N & Harlow E 1992 Adenovirus E1A targets key regulators of cell proliferation. Cancer Surv 12 161-195.

Edinger M, Sweeney TJ, Tucker AA, Olomu AB, Negrin RS & Contag CH 1999 Noninvasive assessment of tumor cell proliferation in animal models. Neoplasia 1 303-310.

Elkas J, Baldwin R, Pegram M, Tseng Y & Karlan B 1999 Immunoglobulin in ovarian cancer ascites inhibits viral infection: implications for adenoviral mediated gene therapy. Gynecol Oncol 72 456.

Flint J & Shenk T 1997 Viral transactivating proteins. Annu Rev Genet 31 177-212.

Friedlander M, Brooks PC, Shaffer RW, Kincaid CM, Varner JA & Cheresh DA 1995 Definition of two angiogenic pathways by distinct alpha v integrins. Science 270 1500-1502.

Fueyo J, Gomez-Manzano C, Alemany R, Lee PS, McDon-nell TJ, Mitlianga P, Shi YX, Levin VA, Yung WK & Kyritsis AP 2000 A mutant oncolytic adenovirus targeting the Rb pathway produces anti-glioma effect in vivo. Oncogene 19 2-12.

Fueyo J, Gomez-Manzano C, Bruner JM, Saito Y, Zhang B, Zhang W, Levin VA, Yung WK & Kyritsis AP 1996 Hypermethylation of the CpG island of p16/CDKN2 correlates with gene inactivation in gliomas. Oncogene 13 1615-1619.

Gamble LJ, Borovjagin AB & Matthews QL 2010 Role of RGD-containing ligands in targeting cellular integrins: Applications for ovarian cancer virotherapy (Review). Exp Ther Med 1 233-240.

Goldberg I, Davidson B, Reich R, Gotlieb WH, Ben-Baruch G, Bryne M, Berner A, Nesland JM & Kopolovic J 2001 Alphav integrin expression is a novel marker of poor prognosis in advanced stage ovarian carcinoma. Clin Cancer Res 7 4073-4079.

Goldman MJ & Wilson JM 1995 Expression of alpha v beta 5 integrin is necessary for efficient adenovirus mediated gene transfer in the human airway. J Virol 69 5951-5958.

Hedley SJ, Chen J, Mountz JD, Li J, Curiel DT, Korokhov N & Kovesdi I 2006 Targeted and shielded adenovectors for cancer therapy. Cancer Immunol Immunother 55 1412-1419.

Heise C, Hermiston T, Johnson L, Brooks G, Sampson-Johannes A, Williams A, Hawkins L & Kirn D 2000 An adenovirus E1A mutant that demonstrates potent and selective systemic anti-tumoral efficacy. Nat Med 6 1134-1139.

Heise C & Kirn DH 2000 Replication selective adenovi-ruses as oncolytic agents. J Clin Invest 105 847-851.

Hemmi S, Geertsen R, Mezzacasa A, Peter I & Dummer R 1998 The presence of human coxsackievirus and adenovirus receptor is associated with efficient adenovirus-mediated transgene expression in human melanoma cell cultures. Hum Gene Ther 9 2363-2373.

Hynes RO 1999 Cell adhesion: old and new questions. Trends Cell Biol 9 33-37. Initiative IAV 2010 Understanding Preexisting Immunity. AIDS Vaccine Clinical Trials.

Jemal A, Siegel R, Ward E, Hao Y, Xu J, Murray T & Thun MJ 2008 Cancer statistics, 2008. CA Cancer J Clin 58 71-96.

Kaur S, Kenny HA, Jagadeeswaran S, Zillhardt MR, Montag AG, Kistner E, Yamada SD, Mitra AK & Lengyel E 2009 {beta}3-integrin expression on tumor cells inhibits tumor progression, reduces metastasis, and is associated with a favorable prognosis in patients with ovarian cancer. Am J Pathol 175 2184-2196.

Kelly FJ, Miller CR, Buchsbaum DJ, Gomez-Navarro J, Barnes MN, Alvarez RD & Curiel DT 2000 Selectivity of TAG-72-targeted adenovirus gene transfer to primary ovar-ian carcinoma cells versus autologous mesothelial cells in vitro. Clin Cancer Res 6 4323-4333.

Khuu H, Conner M, Vanderkwaak T, Shultz J, Gomez-Navarro J, Alvarez RD, Curiel DT & Siegal GP 1999 Detec-tion of Coxsackie Adenovirus Receptor (CAR) Immunoreactivity in Ovarian Tumors of Epithelial Derivation. Appl Immun Mol Morph 7 266.

Kim M, Zinn KR, Barnett BG, Sumerel LA, Krasnykh V, Curiel DT & Douglas JT 2002 The therapeutic efficacy of adenoviral vectors for cancer gene therapy is limited by a low level of primary adenovirus receptors on tumour cells. Eur J Cancer 38 1917-1926.

Kimball KJ, Preuss MA, Barnes MN, Wang M, Siegal GP, Wan W, Kuo H, Saddekni S, Stockard CR, Grizzle WE, Harris RD, Aurigemma R, Curiel DT & Alvarez RD 2010 A phase I study of a tropism modified conditionally replicative adenovirus for recurrent malignant gynecologic diseases. Clin Cancer Res 16 5277-5287.

Kimball KJ, Rivera AA, Zinn KR, Icyuz M, Saini V, Li J, Zhu ZB, Siegal GP, Douglas JT, Curiel DT, et al. 2009 Novel infectivity enhanced oncolytic adenovirus with a capsid incorporated dual imaging moiety for monitoring virotherapy in ovarian cancer. Mol Imaging 8 264-277.

Kirn D 2000 Replication-selective oncolytic adenoviruses: virotherapy aimed at genetic targets in cancer. Oncogene 19 6660-6669.

Le LP, Everts M, Dmitriev IP, Davydova JG, Yamamoto M & Curiel DT 2004 Fluorescently labeled adenovirus with pIX-EGFP for vector detection. Mol Imaging 3 105-116.

Li J, Le L, Sibley DA, Mathis JM & Curiel DT 2005 Genetic incorporation of HSV-1 thymidine kinase into the ade-novirus protein IX for functional display on the virion. Virology 338 247-258.

Liapis H, Adler LM, Wick MR & Rader JS 1997 Expression of alpha(v)beta3 integrin is less frequent in ovarian epithelial tumors of low malignant potential in contrast to ovarian carcinomas. Hum Pathol 28 443-449.

Liu Y, Heyman M, Wang Y, Falkmer U, Hising C, Szekely L & Einhorn S 1994 Molecular analysis of the retinoblastoma gene in primary ovarian cancer cells. Int J Cancer 58 663-667.

Maizel JJ, White D & Scharff M 1968 The polypeptides of adenovirus. I. Evidence for multiple protein components in the virion and a comparison of types 2, 7A, and 12. Virology 36 115-125.

Mathias P, Wickham T, Moore M & Nemerow G 1994 Multiple adenovirus serotypes use alpha v integrins for infection. J Virol 68 6811-6814.

Matthews QL, Sibley DA, Wu H, Li J, Stoff-Khalili MA, Waehler R, Mathis JM & Curiel DT 2006 Genetic incorporation of a herpes simplex virus type 1 thymidine kinase and firefly luciferase fusion into the adenovirus protein IX for functional display on the virion. Mol Imaging 5 510-519.

Meulenbroek RA, Sargent KL, Lunde J, Jasmin BJ & Parks RJ 2004 Use of adenovirus protein IX (pIX) to display large polypeptides on the virion-generation of fluorescent virus through the incorporation of pIX-GFP. Mol Ther 9 617-624.

Miller CR, Buchsbaum DJ, Reynolds PN, Douglas JT, Gillespie GY, Mayo MS, Raben D & Curiel DT 1998 Differential susceptibility of primary and established human glioma cells to adenovirus infection: targeting via the epidermal growth factor receptor achieves fiber receptor independent gene transfer. Cancer Res 58 5738-5748.

Nagel H, Maag S, Tassis A, Nestle FO, Greber UF & Hemmi S 2003 The alphavbeta5 integrin of hematopoietic and nonhematopoietic cells is a transduction receptor of RGD-4C fiber-modified adenoviruses. Gene Ther 10 1643-1653. NCI 2009 Ovarian Cancer Homepage. Cancer Topics. Bethesda Maryland.

Nevins JR 1992 E2F: a link between the Rb tumor suppressor protein and viral oncoproteins. Science 258 424-429. Niederacher D, Yan HY, An HX, Bender HG & Beckmann MW 1999 CDKN2A gene inactivation in epithelial sporadic ovarian cancer. Br J Cancer 80 1920-1926.

Page JG, Tian B, Schweikart K, Tomaszewski J, Harris R, Broadt T, Polley-Nelson J, Noker PE, Wang M, Makhija S, Aurigemma R, Curiel DT & Alvarez RD 2007 Identifying the safety profile of a novel infectivity enhanced condition-ally replicative adenovirus, Ad5-delta24-RGD, in anticipation of a phase I trial for recurrent ovarian cancer. Am J Obstet Gynecol 196 389.e1-9; discussion 389.e9-10.

Ruoslahti E 1996 RGD and other recognition sequences for integrins. Annu Rev Cell Dev Biol 12 697-715.

Sherr CJ 1996 Cancer cell cycles. Science 274 1672-1677.

Stewart PL, Chiu CY, Huang S, Muir T, Zhao Y, Chait B, Mathias P & Nemerow GR 1997 Cryo-EM visualization of an exposed RGD epitope on adenovirus that escapes antibody neutralization. EMBO J 16 1189-1198.

Suzuki K, Fueyo J, Krasnykh V, Reynolds PN, Curiel DT & Alemany R 2001 A conditionally replicative adenovirus with enhanced infectivity shows improved oncolytic potency. Clin Cancer Res 7 120-126.

Takayama K, Ueno H, Pei XH, Nakanishi Y, Yatsunami J & Hara N 1998 The levels of integrin alpha v beta 5 may pre-dict the susceptibility to adenovirus mediated gene transfer in human lung cancer cells. Gene Ther 5 361-368.

Tang Y, Le LP, Matthews QL, Han T, Wu H & Curiel DT 2008 Derivation of a triple mosaic adenovirus based on modification of the minor capsid protein IX. Virology 377 391-400.

Tang Y, Wu H, Ugai H, Matthews QL & Curiel DT 2009 Derivation of a triple mosaic adenovirus for cancer gene therapy. PLoS One 4.

Ulasov IV, Rivera AA, Han Y, Curiel DT, Zhu ZB & Lesniak MS 2007 Targeting adenovirus to CD80 and CD86 receptors increases gene transfer efficiency to malignant glioma cells. J Neurosurg 107 617-627.

Vanderkwaak TJ, Wang M, Gomez-Navarro J, Rancourt C, Dmitriev I, Krasnykh V, Barnes M, Siegal GP, Alvarez R & Curiel DT 1999 An advanced generation of adenoviral vec-tors selectively enhances gene transfer for ovarian cancer gene therapy approaches. Gynecol Oncol 74 227-234.

Vellinga J, Rabelink MJWE, Cramer SJ, van den Wollen-berg DJM, Van der Meulen H, Leppard KN, Fallaux FJ & Hoeben RC 2004 Spacers increase the accessibility of peptide ligands linked to the carboxyl terminus of adenovirus minor capsid protein IX. J Virol 78 3470-3479.

Wang M, Hemminki A, Siegal GP, Barnes MN, Dmitriev I, Krasnykh V, Liu B, Curiel DT & Alvarez RD 2005 Adenoviruses with an RGD-4C modification of the fiber knob elicit a neutralizing antibody response but continue to allow enhanced gene delivery. Gynecol Oncol 96 341-348.

Whyte P, Buchkovich KJ, Horowitz JM, Friend SH, Ray-buck M, Weinberg RA & Harlow E 1988 Association be-tween an oncogene and an anti-oncogene: the adenovirus E1A proteins bind to the retinoblastoma gene product. Na-ture 334 124-129.

Whyte P, Williamson NM & Harlow E 1989 Cellular targets for transformation by the adenovirus E1A proteins. Cell 56 67-75.

Wickham TJ, Mathias P, Cheresh DA & Nemerow GR 1993 Integrins alpha v beta 3 and alpha v beta 5 promote adenovirus internalization but not virus attachment. Cell 73 309-319.

Yaginuma Y, Hayashi H, Kawai K, Kurakane T, Saitoh Y, Kitamura S, Sengoku K & Ishikawa M 1997 Analysis of the Rb gene and cyclin-dependent kinase 4 inhibitor genes (p16INK4 and p15INK4B) in human ovarian carcinoma cell lines. Exp Cell Res 233 233-239.

Full Text: PDF


  • There are currently no refbacks.

Copyright © 2018 Lorem Ipsum Press