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Co-expression of p21Waf1/Cip1 in adenovirus vectors improves expression of a second transgene

Gene Therapy volume 16, pages 574–578 (2009)Cite this article

Abstract

First-generation adenoviral (Ad) vectors are frequently used vectors for experimental and clinical gene transfer. Earlier it has been shown that parallel overexpression of the cell cycle regulator p21Waf1/Cip1 (p21) or antiapoptotic bcl-2 from a second vector reduces cytotoxicity and improves transgene expression. Here, we investigate whether the co-expression of p21 and α1-antitrypsin from a single vector improves vector safety and α1-antitrypsin expression. Cell lines (A549 and HeLa) and primary cells (small airway epithelial cells and hepatocytes) were infected with adenovirus vectors transducing α1-antitrypsin with (AdCMV.p21-RSV.hAAT) or without (AdRSV.hAAT) p21. α1-Antitrypsin expression and cytotoxicity were analyzed using western blot/ELISA and LDH/ALT/AST assays, respectively. Cell cycle profiles were determined by flow cytometry. Co-expression of p21 strongly increased the α1-antitrypsin expression in all cell types and at all doses tested. No changes in ALT/AST from hepatocytes and only minor increases in the LDH release in A549 and HeLa were observed with either vector. Cell cycle profiles were also not affected adversely. Incorporation of p21 in Ad vectors together with a gene of interest improves the vector performance; such vectors will allow the application of lower doses and thereby reduce immunological side effects.

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Acknowledgements

We gratefully acknowledge the technical assistance of Margitta Schüman. This work was funded by a Grant (10134771) from the ProFIT program of the Investitionsbank Berlin.

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Authors and Affiliations

  1. Division of Pneumology, Department of Infectious Diseases and Pneumology, Charité–Universitätsmedizin Berlin, Charitéplatz 1, Berlin, Germany

    A Schumacher, S Horvat, C Woischwill, G Wolff & C Witt

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  1. A Schumacher
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  2. S Horvat
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  3. C Woischwill
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  4. G Wolff
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  5. C Witt
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Correspondence to A Schumacher.

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Supplementary Information accompanies the paper on Gene Therapy website (http://www.nature.com/gt)

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Schumacher, A., Horvat, S., Woischwill, C. et al. Co-expression of p21Waf1/Cip1 in adenovirus vectors improves expression of a second transgene. Gene Ther 16, 574–578 (2009). https://doi.org/10.1038/gt.2009.2

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