Vaccinia as a vector for tumor-directed gene therapy: Biodistribution of a thymidine kinase-deleted mutant

Markus Puhlmann, Charles K. Brown, Michael Gnant, James Huang, Steven K. Libutti, H. Richard Alexander, David L. Bartlett

Research output: Contribution to journalArticle

126 Scopus citations

Abstract

Tumor-directed gene therapy, such as 'suicide gene' therapy, requires high levels of gene expression in a high percentage of tumor cells in vivo to be effective. Current vector strategies have been ineffective in achieving these goals. This report introduces the attenuated (thymidine kinase (TK)- negative) replication-competent vaccinia virus (VV) as a potential vector for tumor-directed gene therapy by studying the biodistribution of VV in animal tumor models. A TK-deleted recombinant VV (Western Reserve strain) expressing luciferase on a synthetic promoter was constructed. Luciferase activity was measured in vitro after transduction of a variety of human and murine tumor cell lines and in vivo after intraperitoneal (i.p.) delivery in C57BL/6 mice with 7-day i.p. tumors (106 MC-38 cells). Three other in vivo tumor models were examined for tumor-specific gene expression after intravenous delivery of VV (human melanoma in nude mice, adenocarcinoma liver metastasis in immunocompetent mice, and subcutaneous sarcoma in the rat). In addition, a replication-incompetent vaccinia (1 μg of psoralen and ultraviolet light, 365 nm, 4 minutes) was tested in vitro and in vivo and compared with active virus. Luciferase activity in i.p. tumors at 4 days after i.p. injection of VV was >7000-fold higher than lung, >3000-fold higher than liver, and >250- fold higher than ovary. In addition, intravenous injection of VV resulted in markedly higher tumor luciferase activity compared with any other organ in every model tested (up to 188,000-fold higher than liver and 77,000-fold higher than lung). Inactivation of the virus resulted in negligible gene expression in vivo. In summary, VV has a high transduction efficiency in tumor cells with high levels of gene expression. The results suggest a selective in vivo replication of TK-deleted VV in tumor cells. Replication competent, TK-deleted VV appears to be an ideal vector for testing the in vivo delivery of toxic genes to tumor cells.

Original languageEnglish (US)
Pages (from-to)66-73
Number of pages8
JournalCancer gene therapy
Volume7
Issue number1
DOIs
StatePublished - Jan 1 2000

Keywords

  • Biodistribution
  • Gene therapy
  • Luciferase
  • Thymidine kinase
  • Vaccinia

ASJC Scopus subject areas

  • Molecular Medicine
  • Molecular Biology
  • Cancer Research

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    Puhlmann, M., Brown, C. K., Gnant, M., Huang, J., Libutti, S. K., Alexander, H. R., & Bartlett, D. L. (2000). Vaccinia as a vector for tumor-directed gene therapy: Biodistribution of a thymidine kinase-deleted mutant. Cancer gene therapy, 7(1), 66-73. https://doi.org/10.1038/sj.cgt.7700075