Treatment of phenylketonuria using minicircle-based naked-DNA gene transfer to murine liver

Authors

  • Hiu Man Viecelli,

    1. Division of Metabolism, Department of Pediatrics, University of Zurich, Zurich, Switzerland; and affiliated with the Children's Research Center Zurich
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  • Richard P. Harbottle,

    1. Section of Molecular Medicine, National Heart and Lung Institute, Imperial College London, London, United Kingdom
    Current affiliation:
    1. DNA Vector Research Group, Deutsches Krebsforschungszentrum (DKFZ), Heidelberg, Germany
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  • Suet Ping Wong,

    1. Section of Molecular Medicine, National Heart and Lung Institute, Imperial College London, London, United Kingdom
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  • Andrea Schlegel,

    1. Swiss HPB and Transplant Center, Department of Surgery, University Hospital Zurich, Zurich, Switzerland
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  • Marinee K. Chuah,

    1. Department of Gene Therapy & Regenerative Medicine, Free University of Brussels, Brussels, Belgium
    2. Department of Cardiovascular Sciences, Center for Molecular and Vascular Biology, University of Leuven, Leuven, Belgium
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  • Thierry VandenDriessche,

    1. Department of Gene Therapy & Regenerative Medicine, Free University of Brussels, Brussels, Belgium
    2. Department of Cardiovascular Sciences, Center for Molecular and Vascular Biology, University of Leuven, Leuven, Belgium
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  • Cary O. Harding,

    1. Departments of Molecular and Medical Genetics and Pediatrics, Oregon Health & Science University, Portland, OR, USA
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  • Beat Thöny

    Corresponding author
    1. Division of Metabolism, Department of Pediatrics, University of Zurich, Zurich, Switzerland; and affiliated with the Children's Research Center Zurich
    • Address reprint requests to: Prof. Beat Thöny, University of Zurich, Division of Metabolism, Steinwiesstrasse 75, CH-8032, Zurich, Switzerland. E-mail: beat.thony@kispi.uzh.ch; fax: +41 44 266 7169.

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  • Potential conflict of interest: Nothing to report.

  • Supported by grants from the Children's Research Center Zurich (to H.M.V.), the Swiss National Science Foundation (no. 310030-122045 to B.T.), the National Institute of Health (research grant no. 1R01HD057033 to C.O.H. and B.T.), and the Stiftung für wissenschaftliche Forschung der Universitüt Zürich (to B.T.).

Abstract

Host immune response to viral vectors, persistence of nonintegrating vectors, and sustained transgene expression are among the major challenges in gene therapy. To overcome these hurdles, we successfully used minicircle (MC) naked-DNA vectors devoid of any viral or bacterial sequences for the long-term treatment of murine phenylketonuria, a model for a genetic liver defect. MC-DNA vectors expressed the murine phenylalanine hydroxylase (Pah) complementary DNA (cDNA) from a liver-specific promoter coupled to a de novo designed hepatocyte-specific regulatory element, designated P3, which is a cluster of evolutionary conserved transcription factor binding sites. MC-DNA vectors were subsequently delivered to the liver by a single hydrodynamic tail vein (HTV) injection. The MC-DNA vector normalized blood phenylalanine concomitant with reversion of hypopigmentation in a dose-dependent manner for more than 1 year, whereas the corresponding parental plasmid did not result in any phenylalanine clearance. MC vectors persisted in an episomal state in the liver consistent with sustained transgene expression and hepatic PAH enzyme activity without any apparent adverse effects. Moreover, 14-20% of all hepatocytes expressed transgenic PAH, and the expression was observed exclusively in the liver and predominately around pericentral areas of the hepatic lobule, while there was no transgene expression in periportal areas. Conclusion: This study demonstrates that MC technology offers an improved safety profile and has the potential for the genetic treatment of liver diseases. (Hepatology 2014;60:1035–1043)

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