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Journal of Virology, April 2009, p. 3249-3257, Vol. 83, No. 7
0022-538X/09/$08.00+0     doi:10.1128/JVI.01721-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Robust In Vivo Transduction of a Genetically Stable Epstein-Barr Virus Episome to Hepatocytes in Mice by a Hybrid Viral Vector{triangledown}

Sean D. Gallaher,1 Jose S. Gil,1 Oliver Dorigo,2 and Arnold J. Berk1,3*

Department of Microbiology, Immunology, and Molecular Genetics,1 Department of Obstetrics and Gynecology, David Geffen UCLA School of Medicine,2 Molecular Biology Institute, University of California at Los Angeles, Los Angeles, California 900953

Received 12 August 2008/ Accepted 12 December 2008

To make a safe, long-lasting gene delivery vehicle, we developed a hybrid vector that leverages the relative strengths of adenovirus and Epstein-Barr virus (EBV). A fully gene-deleted helper-dependent adenovirus (HDAd) is used as the delivery vehicle for its scalability and high transduction efficiency. Upon delivery, a portion of the HDAd vector is recombined to form a circular plasmid. This episome includes two elements from EBV: an EBV nuclear antigen 1 (EBNA1) expression cassette and an EBNA1 binding region. Along with a human replication origin, these elements provide considerable genetic stability to the episome in replicating cells while avoiding insertional mutagenesis. Here, we demonstrate that this hybrid approach is highly efficient at delivering EBV episomes to target cells in vivo. We achieved nearly 100% transduction of hepatocytes after a single intravenous injection in mice. This is a substantial improvement over the transduction efficiency of previously available physical and viral methods. Bioluminescent imaging of vector-transduced mice demonstrated that luciferase transgene expression from the hybrid was robust and compared well to a traditional HDAd vector. Quantitative PCR analysis confirmed that the EBV episome was stable at approximately 30 copies per cell for up to 50 weeks and that it remained circular and extrachromosomal. Approaches for adapting the HDAd-EBV hybrid to a variety of disease targets and the potential benefits of this approach are discussed.

* Corresponding author. Mailing address: Molecular Biology Institute, University of California, Los Angeles, 611 Charles E. Young Drive East, Los Angeles, CA 90095-1570. Phone: (310) 206-6298. Fax: (310) 206-7286. E-mail: berk{at}mbi.ucla.edu

{triangledown} Published ahead of print on 21 January 2009.

Journal of Virology, April 2009, p. 3249-3257, Vol. 83, No. 7
0022-538X/09/$08.00+0     doi:10.1128/JVI.01721-08
Copyright © 2009, American Society for Microbiology. All Rights Reserved.





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