Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Original Article
  • Published:

Additive effect of adenovirus-mediated E2F-1 gene transfer and topoisomerase II inhibitors on apoptosis in human osteosarcoma cells

Cancer Gene Therapy volume 8, pages 241–251 (2001)Cite this article

Abstract

Recently, it has been demonstrated that Etoposide, a topoisomerase II inhibitor, can induce apoptosis in MDM2-overexpressing tumor cells by inhibition of MDM2 synthesis. We have previously shown that E2F-1 overexpression induces apoptosis of MDM2-overexpressing sarcoma cells, which is related to the inhibition of MDM2 expression. Therefore, the present study was designed to investigate the in vitro and in vivo effect of combined treatment of adenovirus-mediated E2F-1 and topoisomerase II inhibitors on the growth inhibition and apoptosis in human sarcoma cells. Two human sarcoma cell lines, OsACL and U2OS, were treated with topoisomerase II inhibitors (Etoposide and Adriamycin), alone or in combination with adenoviral vectors expressing β-galactosidase (Ad-LacZ) or E2F-1 (Ad-E2F-1). E2F-1 expression was confirmed by Western blot analysis. Ad-E2F-1 gene transfer at a low dose (multiplicity of infection, 2) markedly increased the sensitivity of human sarcoma cells to topoisomerase II inhibitor treatment. This cooperative effect of E2F-1 and topoisomerase II inhibitors was less marked in SAOS-2 cells (p53 and pRb null). Topoisomerase II inhibitors also cooperated with E2F-1 overexpression to enhance tumor cell killing in an in vivo model using xenografts in nude mice. When combined with Adriamycin or Etoposide, E2F-1 adenovirus therapy resulted in approximately 95% and 85% decrease in tumor size, respectively, compared to controls (P<.05). These results suggest a new chemosensitization strategy that is effective in MDM2-overexpressing tumors and may have clinical utility. Cancer Gene Therapy (2001) 8, 241–251

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Similar content being viewed by others

References

  1. Mertens WC, Bramwell VC . Osteosarcoma and other tumors of bone Curr Opin Oncol 1997; 9:: 360–365

    Google Scholar 

  2. Bruland OS, Pihl A . On the current management of osteosarcoma. A critical evaluation and a proposal for a modified treatment strategy Eur J Cancer 1997; 11:: 1725–1731

    Google Scholar 

  3. de Saint Aubain Somerhausen N, Fletcher CD . Soft-tissue sarcomas: an update Eur J Surg Oncol 1999; 25:: 215–220

    Google Scholar 

  4. Antman KH, Elias AD . Chemotherapy of advanced soft-tissue sarcomas Semin Surg Oncol 1988; 4:: 53–58

    Google Scholar 

  5. Mulligan LM, Matlashewski GJ, Scrable HJ, et al . Mechanisms of p53 loss in human sarcomas Proc Natl Acad Sci USA 1990; 87:: 5863–5867

    Google Scholar 

  6. Miller CW, Aslo A, Tsay C, et al . Frequency and structure of p53 rearrangements in human osteosarcoma Cancer Res 1990; 50:: 7950–7954

    Google Scholar 

  7. Hung J, Anderson R . p53: functions, mutations and sarcomas Acta Orthop Scand Suppl 1997; 273:: 68–73

    Google Scholar 

  8. Yamaguchi T, Toguchida J, Yamamuro T, et al . Allelotype analysis in osteosarcomas: frequent allele loss on 3q, 13q, 17p, and 18q Cancer Res 1992; 52:: 2419–2423

    Google Scholar 

  9. Malkin D, Jolly KW, Barbier N, et al . Germline mutations of the p53 tumor-suppressor gene in children and young adults with second malignant neoplasms N Engl J Med 1992; 326:: 1309–1315

    Google Scholar 

  10. Srivastava S, Zou ZQ, Pirollo K, et al . Germ-line transmission of a mutated p53 gene in a cancer-prone family with Li-Fraumeni syndrome Nature 1990; 348:: 747–749

    Google Scholar 

  11. Cordon-Cardo C, Latres E, Drobnjak M, et al . Molecular abnormalities of mdm2 and p53 genes in adult soft tissue sarcomas Cancer Res 1994; 54:: 794–799

    Google Scholar 

  12. Momand J, Zambetti GP, Olson DC, et al . The mdm-2 oncogene product forms a complex with the p53 protein and inhibits p53-mediated transactivation Cell 1992; 69:: 1237–1245

    Google Scholar 

  13. Oliner JD, Pietenpol JA, Thiagalingam S, et al . Oncoprotein MDM2 conceals the activation domain of tumour suppressor p53 Nature 1993; 362:: 857–860

    Google Scholar 

  14. Chen J, Marechal V, Levine AJ . Mapping of the p53 and mdm2 interaction domains Mol Cell Biol 1993; 13:: 4107–4114

    PubMed Central  Google Scholar 

  15. Finlay CA . The mdm-2 oncogene can overcome wild-type p53 suppression of transformed cell growth Mol Cell Biol 1993; 13:: 301–306

    PubMed Central  Google Scholar 

  16. Haupt Y, Maya R, Kazaz A, et al . Mdm2 promotes the rapid degradation of p53 Nature 1997; 387:: 296–299

    Google Scholar 

  17. Haupt Y, Barak Y, Oren M . Cell type–specific inhibition of p53-mediated apoptosis by mdm2 EMBO J 1996; 15:: 1596–1606

    PubMed Central  Google Scholar 

  18. Montes de Oca Luna R, Wagner DS, Lozano G . Rescue of early embryonic lethality in mdm2-deficient mice by deletion of p53 Nature 1995; 378:: 203–206

    Google Scholar 

  19. Jones SN, Roe AE, Donehower LA, et al . Rescue of embryonic lethality in Mdm2-deficient mice by absence of p53 Nature 1995; 378:: 206–208

    Google Scholar 

  20. Xiao ZX, Chen J, Levine AJ, et al . Interaction between the retinoblastoma protein and the oncoprotein MDM2 Nature 1995; 375:: 694–698

    Google Scholar 

  21. Martin K, Trouche D, Hagemeier C, et al . Stimulation of E2F1/DP1 transcriptional activity by MDM2 oncoprotein Nature 1995; 375:: 691–694

    Google Scholar 

  22. Dyson N . The regulation of E2F by pRB-family proteins Genes Dev 1998; 12:: 2245–2262

    Google Scholar 

  23. Singh P, Wong SH, Hong W . Overexpression of E2F-1 in rat embryo fibroblasts leads to neoplastic transformation EMBO J 1994; 13:: 3329–3338

    Google Scholar 

  24. Johnson DG, Cress WD, Jakoi L, et al . Oncogenic capacity of the E2F1 gene Proc Natl Acad Sci USA 1994; 91:: 12823–12827

    Google Scholar 

  25. Xu G, Livingston DM, Krek W . Multiple members of the E2F transcription factor family are the products of oncogenes Proc Natl Acad Sci USA 1995; 92:: 1357–13561

    Google Scholar 

  26. Helin K . Regulation of cell proliferation by the E2F transcription factors Curr Opin Genet Dev 1998; 8:: 28–35

    Google Scholar 

  27. Pierce AM, Fisher SM, Conti CJ, et al . Deregulated expression of E2F1 induces hyperplasia and cooperates with ras in skin tumor development Oncogene 1998; 16:: 1267–1276

    Google Scholar 

  28. Yamasaki L, Jacks T, Bronson R, et al . Tumor induction and tissue atrophy in mice lacking E2F-1 Cell 1996; 85:: 537–548

    Google Scholar 

  29. Field SJ, Tsai FY, Kuo F, et al . E2F-1 functions in mice to promote apoptosis and suppress proliferation Cell 1996; 85:: 549–561

    Google Scholar 

  30. Pan H, Yin C, Dyson NJ, et al . Key roles for E2F1 in signaling p53-dependent apoptosis and in cell division within developing tumors Mol Cell 1998; 2:: 283–292

    Google Scholar 

  31. Yamasaki L, Bronson R, Williams BO, et al . Loss of E2F-1 reduces tumorigenesis and extends the lifespan of Rb1(±)mice Nat Genet 1998; 18:: 360–364

    Google Scholar 

  32. DeGregori J, Kowalik T, Nevins JR . Cellular targets for activation by the E2F1 transcription factor include DNA synthesis- and G1/S-regulatory genes Mol Cell Biol 1995; 15:: 4215–4224

    Google Scholar 

  33. Wu XW, Levine AJ . P53 and E2F-1 cooperate to mediate apoptosis Proc Natl Acad Sci USA 1994; 91:: 3602–3606

    Google Scholar 

  34. Shan B, Lee WH . Deregulated expression of E2F-1 induces S-phase entry and leads to apoptosis Mol Cell Biol 1994; 14:: 8166–8173

    Google Scholar 

  35. Qin XQ, Livingston DM, Ewen M . The transcription factor E2F-1 is a downstream target of RB action Mol Cell Biol 1995; 15:: 742–755

    Google Scholar 

  36. Nip J, Strom DK, Fee BE, et al . E2F-1 cooperates with topoisomerase II inhibition and DNA damage to selectively augment p53-independent apoptosis Mol Cell Biol 1997; 17:: 1049–1056

    Google Scholar 

  37. Meng RD, Phillips P, El-Deiry WS . p53-independent increase in E2F-1 expression enhances the cytotoxic effects of Etoposide and of Adriamycin Int J Oncol 1999; 14:: 5–14

    Google Scholar 

  38. Yang HL, Dong YB, Elliott MJ, et al . Adenovirus-mediated E2F-1 gene transfer inhibits MDM2 expression and efficiently induces apoptosis in MDM2-overexpressing tumor cells Clin Cancer Res 1999; 5:: 2242–2250

    Google Scholar 

  39. Casper ES, Christman KL, Schwartz GK, et al . Edatrexate in patients with soft tissue sarcoma. Activity in malignant fibrous histiocytoma Cancer 1993; 72:: 766–770

    Google Scholar 

  40. Alvegard TA, Sigurdsson H, Mouridsen H, et al . Adjuvant chemotherapy with doxorubicin in high-grade soft tissue sarcoma: a randomized trial of the Scandinavian Sarcoma Group J Clin Oncol 1989; 7:: 1504–1513

    Google Scholar 

  41. Demetri GD, Elias AD . Results of single-agent and combination chemotherapy for advanced soft tissue sarcomas. Implications for decision making in the clinic Hematol Oncol Clin North Am 1995; 9:: 765–785

    Google Scholar 

  42. Demetri GD, Elias AD . Results of single-agent and combination chemotherapy for advanced soft tissue sarcomas. Implications for decision making in the clinic Hematol Oncol Clin North Am 1995; 9:: 765–785

    Google Scholar 

  43. Mouridsen HT, Bastholt L, Somers R, et al . Adriamycin versus epirubicin in advanced soft tissue sarcomas. A randomized phase II/phase III study of the EORTC Soft Tissue and Bone Sarcoma Group Eur J Cancer Clin Oncol 1987; 23:: 1477–1483

    Google Scholar 

  44. Dombernowsky P, Buesa J, Pinedo HM, et al . VP-16 in advanced soft tissue sarcoma: a phase II study of the EORTC soft tissue and bone sarcoma group Eur J Cancer Clin Oncol 1987; 23:: 579–580

    Google Scholar 

  45. French BA, Mazur W, Ali NM, et al . Percutaneous transluminal in vivo gene transfer by recombinant adenovirus in normal porcine coronary arteries, atherosclerotic arteries, and two models of coronary restenosis Circulation 1994; 90:: 2402–2413

    Google Scholar 

  46. Polyak K, Waldman T, He TC, et al . Genetic determinants of p53-induced apoptosis and growth arrest Genes Dev 1996; 10: 1945–1952

    CAS  PubMed  Google Scholar 

  47. Graham FL, Prevec L . Manipulation of adenovirus vectors In: Murray EJ, ed. Gene Transfer and Expression Protocols. Methods in Molecular Biology Clifton, NJ: The Humana Press; 1991: 109–128

  48. Yang HL, Dong YB, Elliott MJ, et al . Caspase activation and changes in Bcl-2 family member protein expression associated with E2F-1–mediated apoptosis in human esophageal cancer cells Clin Cancer Res 2000; 6:: 1579–1589

    Google Scholar 

  49. Kowalik TF, DeGregori J, Leone G, et al . E2F1-specific induction of apoptosis and p53 accumulation, which is blocked by Mdm2 Cell Growth Differ 1998; 9:: 113–118

    Google Scholar 

  50. Kowalik TF, DeGregori J, Schwarz JK, et al . E2F-1 overexpression in quiescent fibroblasts leads to induction of cellular DNA synthesis and apoptosis J Virol 1995; 69:: 2491–2500

    PubMed  PubMed Central  Google Scholar 

  51. Hsieh JK, Fredersdorf S, Kouzarides T, et al . E2F1-induced apoptosis requires DNA binding but not transactivation and is inhibited by the retinoblastoma protein through direct interaction Genes Dev 1997; 11:: 1840–1852

    Google Scholar 

  52. Dong YB, Yang HL, Elliott MJ, et al . Adenovirus-mediated E2F-1 gene transfer efficiently induces apoptosis in melanoma cells Cancer 1999; 86:: 2021–2033

    Google Scholar 

  53. Fueyo J, Gomez-Manzano C, Yung WK, et al . Overexpression of E2F-1 in glioma triggers apoptosis and suppresses tumor growth in vitro and in vivo Nat Med 1998; 4:: 685–690

    Google Scholar 

  54. Frank DK, Liu TJ, Frederick MJ, et al . Combination E2F-1 and p53 gene transfer does not enhance growth inhibition in human squamous cell carcinoma of the head and neck Clin Cancer Res 1998; 4:: 2265–2272

    Google Scholar 

  55. Gomez-Manzano C, Fueyo J, Alameda F, et al . Gene therapy for gliomas: p53 and E2F-1 proteins and the target of apoptosis Int J Mol Med 1999; 3:: 81–85

    Google Scholar 

  56. Phillips AC, Bates S, Ryan KM, et al . Induction of DNA synthesis and apoptosis are separable functions of E2F- 1 Genes Dev 1997; 11:: 1853–1863

    Google Scholar 

  57. DeGregori J, Kowalik T, Nevins JR . Cellular targets for activation by the E2F1 transcription factor include DNA synthesis- and G1/S-regulatory genes Mol Cell Biol 1995 15:: 4215–4224

    Google Scholar 

  58. Isaacs RJ, Davies SL, Wells NJ, et al . Topoisomerases II alpha and beta as therapy targets in breast cancer Anticancer Drugs 1995; 6:: 195–211

    Google Scholar 

  59. Colvin OM . Drug resistance in the treatment of sarcomas Semin Oncol 1997; 24:: 580–591

    Google Scholar 

  60. Meng RD, Shih H, Prabhu NS, et al . Bypass of abnormal MDM2 inhibition of p53-dependent growth suppression Clin Cancer Res 1998; 4:: 251–259

    Google Scholar 

  61. Krek W, Xu G, Livingston DM . Cyclin A kinase regulation of E2F-1 DNA binding function underlies suppression of an S-phase checkpoint Cell 1995; 83:: 1149–1158

    Google Scholar 

  62. Erhardt P, Tomaselli KJ, Cooper GM . Identification of the MDM2 oncoprotein as a substrate for CPP32-like apoptotic proteases J Biol Chem 1997; 272:: 15049–15052

    Google Scholar 

  63. Pochampally R, Fodera B, Chen L, et al . A 60-kDa MDM2 isoform is produced by caspase cleavage in nonapoptotic tumor cells Oncogene 1998; 17:: 2629–2636

    Google Scholar 

  64. Chen L, Marechal V, Moreau J, et al . Proteolytic cleavage of the mdm2 oncoprotein during apoptosis J Biol Chem 1997; 272:: 22966–22973

    Google Scholar 

  65. Holmberg C, Helin K, Sehested M, et al . E2F-1–induced p53-independent apoptosis in transgenic mice Oncogene 1998; 17: 143–155

    CAS  PubMed  Google Scholar 

  66. Hunt KK, Deng J, Liu TJ, et al . Adenovirus-mediated overexpression of the transcription factor E2F-1 induces apoptosis in human breast and ovarian carcinoma cell lines and does not require p53 Cancer Res 1997; 57:: 4722–4726

    Google Scholar 

Download references

Acknowledgements

Financial Support: Supported by American Cancer Society Award 96-55, Alliant Community Trust Fund Award 96-46, the Center for Advanced Surgical Technologies (CAST) of Norton Hospital, and The Mary and Mason Rudd Foundation Award. We are grateful to Dr. T. J. Liu for providing the Ad-E2F-1 vector and Dr. Brent French for providing the Ad5CMVLacZ vector. We thank Sherri Matthews for expert assistance with manuscript preparation.

Author information

Author notes

  1. Hai Liang Yang and Yan Bin Dong: Both authors contributed equally to this work.

Authors and Affiliations

  1. Department of Surgery, University of Louisville, James Graham Brown Cancer Center, Louisville, Kentucky

    Hai Liang Yang, Yan Bin Dong, Mary Jane Elliott, Sandra L Wong & Kelly M McMasters

Authors
  1. Hai Liang Yang
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Yan Bin Dong
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Mary Jane Elliott
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Sandra L Wong
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Kelly M McMasters
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Kelly M McMasters.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Yang, H., Dong, Y., Elliott, M. et al. Additive effect of adenovirus-mediated E2F-1 gene transfer and topoisomerase II inhibitors on apoptosis in human osteosarcoma cells. Cancer Gene Ther 8, 241–251 (2001). https://doi.org/10.1038/sj.cgt.7700301

Download citation

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/sj.cgt.7700301

Keywords

This article is cited by

Search

Advanced search

Quick links