Indirect imaging of cardiac-specific transgene expression using a bidirectional two-step transcriptional amplification strategy

I. Y. Chen, O. Gheysens, S. Ray, Q. Wang, P. Padmanabhan, R. Paulmurugan, A. M. Loening, Martin G Rodriguez-Porcel, J. K. Willmann, A. Y. Sheikh, C. H. Nielsen, G. Hoyt, C. H. Contag, R. C. Robbins, S. Biswal, J. C. Wu, S. S. Gambhir

Research output: Contribution to journalArticle

22 Scopus citations

Abstract

Transcriptional targeting for cardiac gene therapy is limited by the relatively weak activity of most cardiac-specific promoters. We have developed a bidirectional plasmid vector, which uses a two-step transcriptional amplification (TSTA) strategy to enhance the expression of two optical reporter genes, firefly luciferase (fluc) and Renilla luciferase (hrluc), driven by the cardiac troponin T (cTnT) promoter. The vector was characterized in vitro and in living mice using luminometry and bioluminescence imaging to assess its ability to mediate strong, correlated reporter gene expression in a cardiac cell line and the myocardium, while minimizing expression in non-cardiac cell lines and the liver. In vitro, the TSTA system significantly enhanced cTnT-mediated reporter gene expression with moderate preservation of cardiac specificity. After intramyocardial and hydrodynamic tail vein delivery of an hrluc-enhanced variant of the vector, long-term fluc expression was observed in the heart, but not in the liver. In both the cardiac cell line and the myocardium, fluc expression correlated well with hrluc expression. These results show the vector's ability to effectively amplify and couple transgene expression in a cardiac-specific manner. Further replacement of either reporter gene with a therapeutic gene should allow non-invasive imaging of targeted gene therapy in living subjects.

Original languageEnglish (US)
Pages (from-to)827-838
Number of pages12
JournalGene Therapy
Volume17
Issue number7
DOIs
StatePublished - 2010

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Keywords

  • bidirectional construct
  • bioluminescence imaging
  • cardiac specificity
  • plasmid vector
  • reporter gene expression
  • two-step transcriptional amplification

ASJC Scopus subject areas

  • Molecular Medicine
  • Molecular Biology
  • Genetics
  • Medicine(all)

Cite this

Chen, I. Y., Gheysens, O., Ray, S., Wang, Q., Padmanabhan, P., Paulmurugan, R., Loening, A. M., Rodriguez-Porcel, M. G., Willmann, J. K., Sheikh, A. Y., Nielsen, C. H., Hoyt, G., Contag, C. H., Robbins, R. C., Biswal, S., Wu, J. C., & Gambhir, S. S. (2010). Indirect imaging of cardiac-specific transgene expression using a bidirectional two-step transcriptional amplification strategy. Gene Therapy, 17(7), 827-838. https://doi.org/10.1038/gt.2010.30