Gene expression imaging by enzymatic catalysis of a fluorescent probe via membrane-anchored β-glucuronidase

Y. C. Su, K. H. Chuang, Y. M. Wang, C. M. Cheng, S. R. Lin, J. Y. Wang, J. J. Hwang, B. M. Chen, K. C. Chen, S. Roffler, T. L. Cheng

Research output: Contribution to journalArticle

25 Citations (Scopus)

Abstract

Development of nonimmunogenic and specific reporter genes to monitor gene expression in vivo is important for the optimization of gene therapy protocols. We developed a membrane-anchored form of mouse β-glucuronidase (mβG) as a reporter gene to hydrolyze a nonfluorescent glucuronide probe (fluorescein di-β-D-glucuronide, (FDGlcU) to a highly fluorescent reporter to assess the location and persistence of gene expression. A functional β-glucuronidase (βG) was stably expressed on the surface of murine CT26 colon adenocarcinoma cells where it selectively hydrolyzed the cell-impermeable FDGlcU probe. FDGlcU was also preferentially converted to fluorescent probe by (βG) on CT26 tumors. The fluorescent intensity in βG-expressing CT26 tumors was 240 times greater than the intensity in control tumors. Selective imaging of gene expression was also observed after intratumoral injection of adenoviral βG vector into carcinoma xenografts. Importantly, mβG did not induce an antibody response after hydrodynamic plasmid immunization of Balbβc mice, indicating that the reporter gene product displayed low immunogenicity. A membrane-anchored form of human βG also allowed in vivo imaging, demonstrating that human βG can be employed for imaging. This imaging system therefore, displays good selectivity with low immunogenicity and may help assess the location, magnitude and duration of gene expression in living animals and humans.

Original languageEnglish
Pages (from-to)565-574
Number of pages10
JournalGene Therapy
Volume14
Issue number7
DOIs
Publication statusPublished - Apr 2007
Externally publishedYes

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Glucuronidase
Catalysis
Fluorescent Dyes
Glucuronides
Reporter Genes
Gene Expression
Membranes
Fluorescein
Neoplasms
Hydrodynamics
Heterografts
Genetic Therapy
Antibody Formation
Immunization
Colon
Adenocarcinoma
Plasmids
Carcinoma
Injections

ASJC Scopus subject areas

  • Genetics

Cite this

Gene expression imaging by enzymatic catalysis of a fluorescent probe via membrane-anchored β-glucuronidase. / Su, Y. C.; Chuang, K. H.; Wang, Y. M.; Cheng, C. M.; Lin, S. R.; Wang, J. Y.; Hwang, J. J.; Chen, B. M.; Chen, K. C.; Roffler, S.; Cheng, T. L.

In: Gene Therapy, Vol. 14, No. 7, 04.2007, p. 565-574.

Research output: Contribution to journalArticle

Su, YC, Chuang, KH, Wang, YM, Cheng, CM, Lin, SR, Wang, JY, Hwang, JJ, Chen, BM, Chen, KC, Roffler, S & Cheng, TL 2007, 'Gene expression imaging by enzymatic catalysis of a fluorescent probe via membrane-anchored β-glucuronidase', Gene Therapy, vol. 14, no. 7, pp. 565-574. https://doi.org/10.1038/sj.gt.3302896
Su, Y. C. ; Chuang, K. H. ; Wang, Y. M. ; Cheng, C. M. ; Lin, S. R. ; Wang, J. Y. ; Hwang, J. J. ; Chen, B. M. ; Chen, K. C. ; Roffler, S. ; Cheng, T. L. / Gene expression imaging by enzymatic catalysis of a fluorescent probe via membrane-anchored β-glucuronidase. In: Gene Therapy. 2007 ; Vol. 14, No. 7. pp. 565-574.
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