Cancer-specific targeting of an adenovirus-delivered herpes simplex virus thymidine kinase suicide gene using translational control

J. Michael Mathis, B. Jill Williams, Don A. Sibley, Jennifer L. Carroll, Jie Li, Yoshinobu Odaka, Shayne Barlow, Cherie Ann O. Nathan, Benjamin D.L. Li, Arrigo DeBenedetti

Research output: Contribution to journalArticle

27 Scopus citations

Abstract

Two technical hurdles, gene delivery and target specificity, have hindered the development of effective cancer gene therapies. In order to circumvent the problem of tumor specificity, the suicide gene, HSV-1 thymidine kinase (HSV-Tk), was modified with a complex 5′ upstream-untranslated region (5′-UTR) that limits efficient translation to cells expressing high levels of the translation initiation factor, eIF4E. Since previous studies have shown that most tumor cells express elevated levels of eIF4E, tumor-specific gene delivery was optimized by incorporation of the 5′-UTR-modified suicide gene (HSV-UTk) into an adenovirus vector (Ad-CMV-UTk). The efficacy of this novel approach of targeting suicide gene expression and limiting cytotoxicity by means of translational restriction was tested in vitro with the use of the human breast cancer cell lines (MCF-7, MDA-MB435, and ZR-75-1). As controls, normal MCF10A, HMEC, and HMSC cell lines that express relatively low levels of eIF4E were used. Real-time reverse-transcription polymerase chain reaction (RT-PCR) was used to quantify HSV-Tk mRNA for cells infected with Ad-CMV-UTk as well as with Ad-CMV-Tk (a control adenovirus in which HSV-Tk is not regulated at the level of translation). Translation of HSV-Tk in the Ad-infected cells was measured by Western blot analysis. In addition, cytotoxicity was determined following treatment with the pro-drug ganciclovir (GCV) using an MTT viability assay. Finally, microPET imaging was used to assess cancer cell-specific expression of HSV-Tk and expression in normal tissues in vivo after intraperitoneal injection of Ad-CMV-Tk or Ad-CMV-UTk. These data collectively showed enhanced cancer cell-specific gene expression and reduced normal tissue gene expression for the Ad-HSV-UTk compared to the Ad-CMV-Tk, leading to increased cancer cell-enhanced GCV cytotoxicity. These results indicate that translational targeting of suicide gene expression in tumor cells in vitro and in vivo is effective and may provide a platform for enhanced cancer gene therapy specificity.

Original languageEnglish
Pages (from-to)1105-1120
Number of pages16
JournalJournal of Gene Medicine
Volume8
Issue number9
DOIs
StatePublished - 1 Sep 2006

Keywords

  • Adenovirus
  • Breast cancer
  • Gene therapy
  • MicroPET
  • Suicide gene
  • Thymidine kinase
  • eIF4E
  • mRNA translation

Fingerprint Dive into the research topics of 'Cancer-specific targeting of an adenovirus-delivered herpes simplex virus thymidine kinase suicide gene using translational control'. Together they form a unique fingerprint.

  • Cite this

    Mathis, J. M., Williams, B. J., Sibley, D. A., Carroll, J. L., Li, J., Odaka, Y., Barlow, S., Nathan, C. A. O., Li, B. D. L., & DeBenedetti, A. (2006). Cancer-specific targeting of an adenovirus-delivered herpes simplex virus thymidine kinase suicide gene using translational control. Journal of Gene Medicine, 8(9), 1105-1120. https://doi.org/10.1002/jgm.935