Expression of vhs and VP16 during HSV-1 helper virus-free amplicon packaging enhances titers

Gene Therapy
W J BowersH J Federoff

Abstract

Recently developed helper virus-free methods of herpes simplex virus (HSV) amplicon vector packaging provide stocks that are virtually devoid of the cytotoxic component normally associated with traditional helper virus-based packaging methods. These approaches involve cotransfection of amplicon plasmid DNA with either a five-cosmid set or a bacterial artificial chromosome (BAC) that contains the HSV genome without its cognate pac signals. Helper virus-free amplicon packaging produces low-titer stocks (<10(5) expressing particles/ml) that exhibit a high frequency of pseudotransduction. In an effort to enhance amplicon titers, we introduced in trans a genomic copy of the virion host shutoff (vhs) protein-encoding gene UL41 into both cosmid- and BAC-based packaging strategies. Cotransfection of this plasmid with the amplicon and packaging reagents results in a 10-fold higher amplicon titer, and stocks that do not exhibit the pseudotransduction phenomenon. To further enhance packaging efficiency, the HSV transcriptional activator VP16 was introduced into packaging cells 1 day before the packaging components. Pre-loading of packaging cells with VP16 led to an additional enhancement of amplicon titers, an effect that did not occur in...Continue Reading

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Citations

Oct 24, 2002·Neurobiology of Aging·Kathleen A Maguire-ZeissHoward J Federoff
Jan 1, 2003·Methods : a Companion to Methods in Enzymology·Steven W WangWilliam H Klein
Apr 5, 2008·Molecular Therapy : the Journal of the American Society of Gene Therapy·Maria E FrazerWilliam J Bowers
Jul 27, 2007·Molecular Therapy : the Journal of the American Society of Gene Therapy·Elise B PetersonWilliam J Bowers
Dec 29, 2010·Proceedings of the National Academy of Sciences of the United States of America·Polyxeni PhilippidouSimon Halegoua
Feb 10, 2009·Human Gene Therapy·Yahui Grace ChiuHoward J Federoff
Aug 19, 2007·International Immunology·Mitra AzadnivIan N Crispe
Oct 28, 2008·Blood·Sandrine DaubeufKhaled Tolba
Dec 14, 2011·The Journal of Neuroscience : the Official Journal of the Society for Neuroscience·Guillermo García-AlíasVictor L Arvanian
Mar 25, 2009·Cellular Immunology·Simona BancosIan Nicholas Crispe
Aug 4, 2007·Biochemical and Biophysical Research Communications·Kuei-Cheng LimHoward J Federoff
Jan 18, 2005·Neurobiology of Aging·William J BowersHoward J Federoff
Jun 16, 2001·Molecular Therapy : the Journal of the American Society of Gene Therapy·X ChenH J Federoff
May 25, 2002·Molecular Therapy : the Journal of the American Society of Gene Therapy·Eric R DetraitEric K Richfield
Jul 4, 2002·Molecular Therapy : the Journal of the American Society of Gene Therapy·William J BowersRobert D Frisina
Feb 25, 2003·Molecular Therapy : the Journal of the American Society of Gene Therapy·John A OlschowkaHoward J Federoff
Sep 29, 2004·Molecular Therapy : the Journal of the American Society of Gene Therapy·Angelika OehmigXandra O Breakefield
Jan 18, 2006·Molecular Therapy : the Journal of the American Society of Gene Therapy·William J BowersHoward J Federoff
Dec 4, 2009·Viruses·Suresh de Silva, William J Bowers
Oct 7, 2005·BioTechniques·Rachael L NeveWilliam A Carlezon
Sep 22, 2006·The Journal of Gene Medicine·M W HaltermanH J Federoff
Apr 8, 2020·Molecular Therapy. Methods & Clinical Development·Iván Fernández-FríasJavier Díaz-Nido

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