Skip to main content

Thank you for visiting 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:

Optimization of radiation controlled gene expression by adenoviral vectors in vitro


The radiation-inducible EGR-1-promoter has been used in different gene therapy approaches in order to enhance and locally restrict therapeutic efficacy. The aim of this study was to reduce nonspecific gene expression in the absence of irradiation (IR) in an adenoviral vector. Rat rhabdomyosarcoma R1H tumor cells were infected with adenoviral vectors expressing either EGFP or HSV-TK under control of the murine EGR-1 promoter/enhancer. Cells were irradiated at 0–6 Gy. Gene expression was determined by FACS-analysis (EGFP), or crystal violet staining (HSV-TK). The bovine growth hormone polyadenylation signal (BGH pA) was used as insulating sequence and was introduced upstream or upstream and downstream of the expression cassette. Infected R1H cells displayed IR dose-dependent EGFP expression. Cells treated with IR, AdEGR.TK and ganciclovir displayed a survival of 17.3% (6 Gy). However, significant gene expression was observed in the absence of IR with EGR.TK and EGR.EGFP constructs. Introduction of BGHpA upstream or upstream and downstream of expression cassette resulted in decreased nonspecific cytotoxicity by a factor of 1.6–2.3 with minor influence on the induced level of cytotoxicity. Introduction of insulating sequences in adenoviral vectors might allow tighter temporospatial control of gene expression by the radiation-inducible EGR-1 promoter.

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

Access options

Buy this article

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

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5

Similar content being viewed by others



multiplicity of infection

CArG Box:

CC(A/T)6GG like sequence






early growth response gene


bovine growth hormone


  1. Gómez-Navarro J, Curiel DT, Douglas JT . Gene therapy for cancer. Eur J Cancer. 1999;35:867–885.

    Article  PubMed  Google Scholar 

  2. Hallahan DE, Mauceri HJ, Seung LP, et al. Spatial and temporal control of gene therapy using ionizing radiation. Nat Med. 1995;1:786–791.

    Article  CAS  PubMed  Google Scholar 

  3. Hallahan DE, Sukhatme VP, Virudachalam S, Kufe DW, Weichselbaum RR . Protein kinase C mediates X-ray inducibility of nuclear signal transducers, Egr-1, and c-jun. Proc Natl Acad Sci USA. 1991;88:2152–2160.

    Google Scholar 

  4. Datta R, Rubin E, Sukhatme VP, et al. Ionizing radiation activates transcription of the EGR1 gene via CArG elements. Proc Natl Acad Sci USA. 1992;89:10149–10153.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Datta R, Taneja N, Sukhatme VP, Qureshi SA, Weichselbaum R, Kufe DW . Reactive oxygen intermediates target CC(A/T)6GG sequences to mediate activation of the early growth response 1 transcription factor gene by ionizing radiation. Proc Natl Acad Sci USA. 1993;90:2419–2422.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  6. Joki T, Nakamura M, Ohno T . Activation of the radiosensitive EGR-1 promoter induces expression of the herpes simplex virus thymidine kinase gene and sensitivity of human glioma cells to gancyclovir. Hum Gene Ther. 1995;6:1507–1513.

    Article  CAS  PubMed  Google Scholar 

  7. Kawashita Y, Ohtsuru A, Kaneda Y, et al. Regression of hepatocellular carcinoma in vitro and in vivo by radiosensitizing suicide gene therapy under the inducible and spatial control of radiation. Hum Gene Ther. 1999;10:1509–1519.

    Article  CAS  PubMed  Google Scholar 

  8. Manome Y, Kunieda T, Wen PY, Koga T, Kufe DW, Ohno T . Transgene expression in malignant glioma using a replication-defective adenoviral vector containing the EGR-1 promoter: activation by ionizing radiation or uptake by radioactive iododeoxyuridine. Hum Gene Ther. 1998;9:1409–1417.

    Article  CAS  PubMed  Google Scholar 

  9. Marples B, Scott SD, Hendry JH, Embleton MJ, Lashford LS, Margison GP . Development of synthetic promoters for radiation-mediated gene therapy. Gene Therapy. 2000;7:511–517.

    Article  CAS  PubMed  Google Scholar 

  10. Mauceri HJ, Hanna NN, Wayne JD, Hallahan DE, Hellman S, Weichselbaum RR . Tumor necrosis factor a (TNF-α) gene therapy targeted by ionizing radiation selectively damages tumor vasculature. Cancer Res. 1996;56:4311–4314.

    CAS  PubMed  Google Scholar 

  11. Meyer RG, Kupper JH, Kandolf R, Rodemann HP . Early growth response-1 gene (Egr-1) promoter induction by ionizing radiation in U87 malignant glioma cells in vitro. Eur J Biochem. 2002;269:337–346.

    Article  CAS  PubMed  Google Scholar 

  12. Steinwaerder DS, Lieber A . Insulation from viral transcriptional regulatory elements improves inducible transgene expression from adenovirus vectors in vitro and in vivo. Gene Therapy. 2000;7:556–567.

    Article  CAS  PubMed  Google Scholar 

  13. Vassaux G, Hurst HC, Lemoine NR . Insulation of a conditionally expressed transgene in an adenoviral vector. Gene Therapy. 1999;6:1192–1197.

    Article  CAS  PubMed  Google Scholar 

  14. Barendsen GW, Broerse JJ . Experimental radiotherapy of a rat rhabdomyosarcoma with 15 MeV neutrons and 300 kV X-rays I: effects of single exposures. Eur J Cancer. 1969;5:373–391.

    Article  CAS  PubMed  Google Scholar 

  15. Reinhold HS . A cell dispersion technique for use in quantitative transplantation studies with solid tumours. Eur J Cancer. 1965;1:67–71.

    Article  CAS  PubMed  Google Scholar 

  16. Sambrook J, Fritsch EF, Maniatis T . Molecular cloning. In: Nolan C, ed. A Laboratory Manual, 2nd edn. New York: Cold Spring Harbor Laboratory Press; 1989.

    Google Scholar 

  17. Anton M, Graham FL . Site-specific recombination mediated by an adenovirus vector expressing the Cre recombinase protein: a molecular switch for control of gene expression. J Virol. 1995;69:4600–4606.

    CAS  PubMed  PubMed Central  Google Scholar 

  18. Tsai-Morris C-H, Cao X, Sukhatme VP . 5′flanking sequence and genomic structure of Egr-1, a murine mitogen inducible zinc finger encoding gene. Nucleic Acids Res. 1988;16:8835–8846.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Bett AJ, Haddara W, Prevec L, Graham FL . An efficient and flexible system for construction of adenovirus vectors with insertions or deletions in early regions 1 and 3. Proc Natl Acad Sci USA. 1994;91:8802–8806.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  20. Hitt M, Bett AJ, Addison CL, et al. Techniques for human adenovirus vector construction and characterization. In: Adolph KW, ed. Viral Gene Techniques. San Diego, New York, Boston, London, Sydney, Tokyo, Toronto: Academic Press; 1995: pp 13–30.

    Chapter  Google Scholar 

  21. Seung LP, Mauceri HJ, Beckett MA, Hallahan DE, Hellman S, Weichselbaum RR . Genetic radiotherapy overcomes tumor resistance to cytotoxic agents. Cancer Res. 1995;55:5561–5565.

    CAS  PubMed  Google Scholar 

  22. Scott SD, Joiner MC, Marples B . Optimizing radiation-responsive gene promoters for radiogenetic cancer therapy. Gene Therapy. 2002;9:1396–1402.

    Article  CAS  PubMed  Google Scholar 

  23. Scott SD, Marples B, Hendry JH, et al. A radiation-controlled molecular switch for use in gene therapy of cancer. Gene Therapy. 2000;7:1121–1125.

    Article  CAS  PubMed  Google Scholar 

  24. Schmidt M, Heimberger T, Gruensfelder P, Schler G, Hoppe F . Inducible promoters for gene therapy of head and neck cancer: an in vitro study. Eur Arch Otorhinolaryngol. 2004;261:208–215.

    Article  PubMed  Google Scholar 

  25. Santiago FS, Lowe HC, Day FL, Chesterman CN, Khachigian LM . Early groth response factor-1 induction by injury is triggered by release and paracrine activation by fibroblast growth factor-2. Am J Pathol. 1999;154:937–944.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  26. Buvoli M, Langer SJ, Bialik S, Leinwand LA . Potential limitations of transcription terminators used as transgene insulators in adenoviral vectors. Gene Therapy. 2002;9:227–231.

    Article  CAS  PubMed  Google Scholar 

  27. Pederson LC, Buchsbaum DJ, Vickers SM, et al. Molecular chemotherapy combined with radiation therapy enhances killing of cholangiocarcinoma cells in vitro and in vivo. Cancer Res. 1997;57:4325–4332.

    CAS  PubMed  Google Scholar 

Download references


The skillful technical assistance of S Wegerer and B Essien is greatly acknowledged. We thank P Wendt for FACS Analysis. Plasmid pE425 was kindly provided by VP Sukhatme. This work was supported by Deutsche Forschungsgemeinschaft (grant number WU 335/1-1). IEOG was supported by a DAAD PhD scholarship.

Author information

Authors and Affiliations


Corresponding author

Correspondence to Martina Anton.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Anton, M., Gomaa, I., von Lukowicz, T. et al. Optimization of radiation controlled gene expression by adenoviral vectors in vitro. Cancer Gene Ther 12, 640–646 (2005).

Download citation

  • Received:

  • Published:

  • Issue Date:

  • DOI:


This article is cited by


Quick links