Genetic incorporation of the protein transduction domain of Tat into Ad5 fiber enhances gene transfer efficacy

Tie Han, Yizhe Tang, Hideyo Ugai, Leslie E. Perry, Gene P. Siegal, Juan L. Contreras, Hongju Wu

Research output: Contribution to journalArticle

12 Citations (Scopus)

Abstract

Background. Human adenovirus serotype 5 (Ad5) has been widely explored as a gene delivery vector for a variety of diseases. Many target cells, however, express low levels of Ad5 native receptor, the Coxsackie-Adenovirus Receptor (CAR), and thus are resistant to Ad5 infection. The Protein Transduction Domain of the HIV Tat protein, namely PTDtat, has been shown to mediate protein transduction in a wide range of cells. We hypothesize that re-targeting Ad5 vector via the PTDtat motif would improve the efficacy of Ad5-mediated gene delivery. Results. In this study, we genetically incorporated the PTDtat motif into the knob domain of Ad5 fiber, and rescued the resultant viral vector, Ad5.PTDtat. Our data showed the modification did not interfere with Ad5 binding to its native receptor CAR, suggesting Ad5 infection via the CAR pathway is retained. In addition, we found that Ad5.PTDtat exhibited enhanced gene transfer efficacy in all of the cell lines that we have tested, which included both low-CAR and high-CAR decorated cells. Competitive inhibition assays suggested the enhanced infectivity of Ad5.PTDtat was mediated by binding of the positively charged PTDtat peptide to the negatively charged epitopes on the cells' surface. Furthermore, we investigated in vivo gene delivery efficacy of Ad5.PTDtat using subcutaneous tumor models established with U118MG glioma cells, and found that Ad5.PTDtat exhibited enhanced gene transfer efficacy compared to unmodified Ad5 vector as analyzed by a non-invasive fluorescence imaging technique. Conclusion. Genetic incorporation of the PTDtat motif into Ad5 fiber allowed Ad5 vectors to infect cells via an alternative PTDtat targeting motif while retaining the native CAR-mediated infection pathway. The enhanced infectivity was demonstrated in both cultured cells and in in vivo tumor models. Taken together, our study identifies a novel tropism expanded Ad5 vector that may be useful for clinical gene therapy applications.

Original languageEnglish (US)
Article number103
JournalVirology Journal
Volume4
DOIs
StatePublished - 2007
Externally publishedYes

Fingerprint

Adenoviridae
Coxsackie and Adenovirus Receptor-Like Membrane Protein
Genes
Serogroup
Protein Domains
Human Immunodeficiency Virus tat Gene Products
Infection
Human Adenoviruses
Tropism
Optical Imaging
Glioma
Genetic Therapy
Epitopes
Cultured Cells
Neoplasms

ASJC Scopus subject areas

  • Virology
  • Genetics(clinical)

Cite this

Genetic incorporation of the protein transduction domain of Tat into Ad5 fiber enhances gene transfer efficacy. / Han, Tie; Tang, Yizhe; Ugai, Hideyo; Perry, Leslie E.; Siegal, Gene P.; Contreras, Juan L.; Wu, Hongju.

In: Virology Journal, Vol. 4, 103, 2007.

Research output: Contribution to journalArticle

Han, Tie ; Tang, Yizhe ; Ugai, Hideyo ; Perry, Leslie E. ; Siegal, Gene P. ; Contreras, Juan L. ; Wu, Hongju. / Genetic incorporation of the protein transduction domain of Tat into Ad5 fiber enhances gene transfer efficacy. In: Virology Journal. 2007 ; Vol. 4.
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abstract = "Background. Human adenovirus serotype 5 (Ad5) has been widely explored as a gene delivery vector for a variety of diseases. Many target cells, however, express low levels of Ad5 native receptor, the Coxsackie-Adenovirus Receptor (CAR), and thus are resistant to Ad5 infection. The Protein Transduction Domain of the HIV Tat protein, namely PTDtat, has been shown to mediate protein transduction in a wide range of cells. We hypothesize that re-targeting Ad5 vector via the PTDtat motif would improve the efficacy of Ad5-mediated gene delivery. Results. In this study, we genetically incorporated the PTDtat motif into the knob domain of Ad5 fiber, and rescued the resultant viral vector, Ad5.PTDtat. Our data showed the modification did not interfere with Ad5 binding to its native receptor CAR, suggesting Ad5 infection via the CAR pathway is retained. In addition, we found that Ad5.PTDtat exhibited enhanced gene transfer efficacy in all of the cell lines that we have tested, which included both low-CAR and high-CAR decorated cells. Competitive inhibition assays suggested the enhanced infectivity of Ad5.PTDtat was mediated by binding of the positively charged PTDtat peptide to the negatively charged epitopes on the cells' surface. Furthermore, we investigated in vivo gene delivery efficacy of Ad5.PTDtat using subcutaneous tumor models established with U118MG glioma cells, and found that Ad5.PTDtat exhibited enhanced gene transfer efficacy compared to unmodified Ad5 vector as analyzed by a non-invasive fluorescence imaging technique. Conclusion. Genetic incorporation of the PTDtat motif into Ad5 fiber allowed Ad5 vectors to infect cells via an alternative PTDtat targeting motif while retaining the native CAR-mediated infection pathway. The enhanced infectivity was demonstrated in both cultured cells and in in vivo tumor models. Taken together, our study identifies a novel tropism expanded Ad5 vector that may be useful for clinical gene therapy applications.",
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