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Journal of Virology, October 2008, p. 10153-10161, Vol. 82, No. 20
0022-538X/08/$08.00+0     doi:10.1128/JVI.01133-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Construction of a Fully Retargeted Herpes Simplex Virus 1 Recombinant Capable of Entering Cells Solely via Human Epidermal Growth Factor Receptor 2 {triangledown} ,{dagger}

Laura Menotti,1 Arianna Cerretani,1 Hartmut Hengel,2 and Gabriella Campadelli-Fiume1*

Department of Experimental Pathology, Section on Microbiology and Virology, Alma Mater Studiorum-University of Bologna, Via San Giacomo, 12, 40126 Bologna, Italy,1 Institute for Virology, Heinrich-Heine-University, Universitätsstrasse 1, D-40225 Düsseldorf, Germany2

Received 30 May 2008/ Accepted 29 July 2008

A novel frontier in the treatment of tumors that are difficult to treat is oncolytic virotherapy, in which a replication-competent virus selectively infects and destroys tumor cells. Herpes simplex virus (HSV) represents a particularly attractive system. Effective retargeting to tumor-specific receptors has been achieved by insertion in gD of heterologous ligands. Previously, our laboratory generated an HSV retargeted to human epidermal growth factor receptor 2 (HER2), a receptor overexpressed in about one-third of mammary tumors and in some ovarian tumors. HER2 overexpression correlates with increased metastaticity and poor prognosis. Because HER2 has no natural ligand, the inserted ligand was a single-chain antibody to HER2. The objective of this work was to genetically engineer an HSV that selectively targets the HER2-expressing tumor cells and that has lost the ability to enter cells through the natural gD receptors, HVEM and nectin1. Detargeting from nectin1 was attempted by two different strategies, point mutations and insertion of the single-chain antibody at a site in gD different from previously described sites of insertion. We report that point mutations at gD amino acids 34, 215, 222, and 223 failed to generate a nectin1-detargeted HSV. An HSV simultaneously detargeted from nectin1 and HVEM and retargeted to HER2 was successfully engineered by moving the site of single-chain antibody insertion at residue 39, i.e., in front of the nectin1-interacting surface and not lateral to it, and by deleting amino acid residues 6 to 38. The resulting recombinant, R-LM113, entered cells and spread from cell to cell solely via HER2.

* Corresponding author. Mailing address: Department of Experimental Pathology, Section on Microbiology and Virology, Alma Mater Studiorum, University of Bologna, Via San Giacomo, 12, 40126 Bologna, Italy. Phone: 39 051 2094733/34. Fax: 39 051 2094735. E-mail: gabriella.campadelli{at}unibo.it

{triangledown} Published ahead of print on 6 August 2008.

{dagger} Supplemental material for this article may be found at http://jvi.asm.org/.

Journal of Virology, October 2008, p. 10153-10161, Vol. 82, No. 20
0022-538X/08/$08.00+0     doi:10.1128/JVI.01133-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.





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