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Proc Natl Acad Sci U S A. 2015 Mar 31;112(13):E1632-41. doi: 10.1073/pnas.1423556112. Epub 2015 Mar 16.

Herpes simplex viral-vector design for efficient transduction of nonneuronal cells without cytotoxicity.

Author information

1
Department of Microbiology and Molecular Genetics, University of Pittsburgh School of Medicine, Pittsburgh, PA 15219;
2
Department of Microbiology and Molecular Genetics, University of Pittsburgh School of Medicine, Pittsburgh, PA 15219; Section of Pharmacology, Department of Medical Sciences, University of Ferrara, 44121 Ferrara, Italy;
3
Thomas E. Starzl Transplantation Institute, Department of Surgery, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261;
4
Thomas E. Starzl Transplantation Institute, Department of Surgery, University of Pittsburgh School of Medicine, Pittsburgh, PA 15261; Liver Cancer Center, University of Pittsburgh School of Medicine, Pittsburgh, PA 15213; and.
5
Department of Biological Sciences, Northern Kentucky University, Highland Heights, KY 41099.
6
Department of Microbiology and Molecular Genetics, University of Pittsburgh School of Medicine, Pittsburgh, PA 15219; glorioso@pitt.edu.

Abstract

The design of highly defective herpes simplex virus (HSV) vectors for transgene expression in nonneuronal cells in the absence of toxic viral-gene activity has been elusive. Here, we report that elements of the latency locus protect a nonviral promoter against silencing in primary human cells in the absence of any viral-gene expression. We identified a CTCF motif cluster 5' to the latency promoter and a known long-term regulatory region as important elements for vigorous transgene expression from a vector that is functionally deleted for all five immediate-early genes and the 15-kb internal repeat region. We inserted a 16.5-kb expression cassette for full-length mouse dystrophin and report robust and durable expression in dystrophin-deficient muscle cells in vitro. Given the broad cell tropism of HSV, our design provides a nontoxic vector that can accommodate large transgene constructs for transduction of a wide variety of cells without vector integration, thereby filling an important void in the current arsenal of gene-therapy vectors.

KEYWORDS:

HSV vector; ICP0; dystrophin; gene therapy; insulator

PMID:
25775541
PMCID:
PMC4386379
DOI:
10.1073/pnas.1423556112
[Indexed for MEDLINE]
Free PMC Article

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