title: Improved HSV-1 amplicon packaging using virion host shutoff mutants lacking mRNAse activity.
abstract: Given their generous transgene capacity and inherent neurotropism, herpes simplex virus (HSV-1)-based viral vectors are promising tools for gene delivery to the central nervous system. Despite their widespread pre-clinical use, vector toxicity remains a concern with regard to the use of herpes vectors in humans. One potential source of toxicity stems from the tegument-associated virion host shutoff protein (vhs), which induces translational arrest in the host cell through non-specific mRNAse activity. In the current study we utilized a series of HSV-1 viruses containing a deletion in the U(L)41 open reading frame to investigate: (1) the requirement of intact vhs function in amplicon packaging and (2) whether vhs influences the post-transduction survival of dissociated cortical neurons. Our results demonstrate that while amplicon yield was reduced an order of magnitude, U(L)41 deletion was associated with reduced vector toxicity. Furthermore, partial reconstitution of vhs function using mRNAse-inactive point mutants improved amplicon titers without imparting the toxicity observed with wild-type controls. These findings offer a novel approach to improving the titer and toxicity profiles of HSV-based viral vectors.
authors.affiliation: Department of Neurology, University of Rochester School of Medicine and Dentistry, Rochester, NY 14642, USA. Marc_Halterman@urmc.rochester.edu
authors.forename: M W
authors.initials: MW
authors.lastname: Halterman
authors.forename: R E
authors.initials: RE
authors.lastname: Giuliano
authors.forename: W J
authors.initials: WJ
authors.lastname: Bowers
authors.forename: H J
authors.initials: HJ
authors.lastname: Federoff
mesh_terms: D000818:Animals; D002460:Cell Line; D002470:Cell Survival; D002522:Chlorocebus aethiops; D005814:Genes, Viral; D018259:Herpesvirus 1, Human / Q000235:genetics* / Q000502:physiology; D066298:In Vitro Techniques; D051379:Mice; D008810:Mice, Inbred C57BL; D009154:Mutation; D041681:NIH 3T3 Cells; D009474:Neurons / Q000166:cytology / Q000821:virology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D012367:RNA, Viral / Q000235:genetics / Q000378:metabolism; D012260:Ribonucleases / Q000235:genetics / Q000378:metabolism; D017384:Sequence Deletion; D014709:Vero Cells; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism; D019065:Virus Assembly
chemical_list: D012333:RNA, Messenger; D012367:RNA, Viral; C081688:Ul41 protein, Human herpesvirus 1; D014764:Viral Proteins; C070722:virion host shutoff protein, Simplexvirus; D012260:Ribonucleases