title: Syncytial Mutations Do Not Impair the Specificity of Entry and Spread of a Glycoprotein D Receptor-Retargeted Herpes Simplex Virus.
abstract: Membrane fusion, which is the key process for both initial cell entry and subsequent lateral spread of herpes simplex virus (HSV), requires the four envelope glycoproteins gB, gD, gH, and gL. Syncytial mutations, predominantly mapped to the gB and gK genes, confer hyperfusogenicity on HSV and cause multinucleated giant cells, termed syncytia. Here we asked whether interaction of gD with a cognate entry receptor remains indispensable for initiating membrane fusion of syncytial strains. To address this question, we took advantage of mutant viruses whose viral entry into cells relies on the uniquely specific interaction of an engineered gD with epidermal growth factor receptor (EGFR). We introduced selected syncytial mutations into gB and/or gK of the EGFR-retargeted HSV and found that these mutations, especially when combined, enabled formation of extensive syncytia by human cancer cell lines that express the target receptor; these syncytia were substantially larger than the plaques formed by the parental retargeted HSV strain. We assessed the EGFR dependence of entry and spread separately by using direct entry and infectious center assays, respectively, and we found that the syncytial mutations did not override the receptor specificity of the retargeted viruses at either stage. We discuss the implications of these results for the development of more effective targeted oncolytic HSV vectors.
  
  
  OBJECTIVE
  Herpes simplex virus (HSV) is investigated not only as a human pathogen but also as a promising agent for oncolytic virotherapy. We previously showed that both the initial entry and subsequent lateral spread of HSV can be retargeted to cells expressing tumor-associated antigens by single-chain antibodies fused to a receptor-binding-deficient envelope glycoprotein D (gD). Here we introduced syncytial mutations into the gB and/or gK gene of gD-retargeted HSVs to determine whether viral tropism remained dependent on the interaction of gD with the target receptor. Entry and spread profiles of the recombinant viruses indicated that gD retargeting does not abolish the hyperfusogenic activity of syncytial mutations and that these mutations do not eliminate the dependence of HSV entry and spread on a specific gD-receptor interaction. These observations suggest that syncytial mutations may be valuable for increasing the tumor-specific spreading of retargeted oncolytic HSV vectors.
authors.affiliation: Division of Bioengineering, Advanced Clinical Research Center, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan.|School of Life Sciences, Tokyo University of Pharmacy and Life Sciences, Tokyo, Japan.
authors.forename: Yu
authors.initials: Y
authors.lastname: Okubo
authors.affiliation: Division of Bioengineering, Advanced Clinical Research Center, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan hiuchida-tky@umin.net.
authors.forename: Hiroaki
authors.initials: H
authors.lastname: Uchida
authors.affiliation: School of Life Sciences, Tokyo University of Pharmacy and Life Sciences, Tokyo, Japan.
authors.forename: Aika
authors.initials: A
authors.lastname: Wakata
authors.affiliation: Division of Bioengineering, Advanced Clinical Research Center, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan.|School of Life Sciences, Tokyo University of Pharmacy and Life Sciences, Tokyo, Japan.
authors.forename: Takuma
authors.initials: T
authors.lastname: Suzuki
authors.affiliation: Division of Bioengineering, Advanced Clinical Research Center, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan.|School of Life Sciences, Tokyo University of Pharmacy and Life Sciences, Tokyo, Japan.
authors.forename: Tomoko
authors.initials: T
authors.lastname: Shibata
authors.affiliation: Division of Bioengineering, Advanced Clinical Research Center, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan.|School of Life Sciences, Tokyo University of Pharmacy and Life Sciences, Tokyo, Japan.
authors.forename: Hitomi
authors.initials: H
authors.lastname: Ikeda
authors.affiliation: Department of Molecular Medicine, Research Institute for Frontier Medicine, Sapporo Medical University School of Medicine, Hokkaido, Japan.
authors.forename: Miki
authors.initials: M
authors.lastname: Yamaguchi
authors.affiliation: Department of Microbiology and Molecular Genetics, School of Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania, USA.
authors.forename: Justus B
authors.initials: JB
authors.lastname: Cohen
authors.affiliation: Department of Microbiology and Molecular Genetics, School of Medicine, University of Pittsburgh, Pittsburgh, Pennsylvania, USA.
authors.forename: Joseph C
authors.initials: JC
authors.lastname: Glorioso
authors.affiliation: School of Life Sciences, Tokyo University of Pharmacy and Life Sciences, Tokyo, Japan.
authors.forename: Mitsuo
authors.initials: M
authors.lastname: Tagaya
authors.affiliation: School of Life Sciences, Tokyo University of Pharmacy and Life Sciences, Tokyo, Japan.|Department of Molecular Medicine, Research Institute for Frontier Medicine, Sapporo Medical University School of Medicine, Hokkaido, Japan.
authors.forename: Hirofumi
authors.initials: H
authors.lastname: Hamada
authors.affiliation: Division of Bioengineering, Advanced Clinical Research Center, The Institute of Medical Science, The University of Tokyo, Tokyo, Japan.
authors.forename: Hideaki
authors.initials: H
authors.lastname: Tahara
mesh_terms: D000818:Animals; D016466:CHO Cells; D045744:Cell Line, Tumor; D002470:Cell Survival; D002522:Chlorocebus aethiops; D003412:Cricetulus; D066246:ErbB Receptors / Q000235:genetics / Q000378:metabolism*; D015870:Gene Expression; D015726:Giant Cells / Q000378:metabolism / Q000648:ultrastructure / Q000821:virology; D018259:Herpesvirus 1, Human / Q000235:genetics* / Q000378:metabolism; D054884:Host-Pathogen Interactions; D006801:Humans; D008561:Membrane Fusion; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D050130:Oncolytic Virotherapy; D011991:Receptors, Virus / Q000235:genetics / Q000378:metabolism*; D014709:Vero Cells; D014759:Viral Envelope Proteins / Q000235:genetics* / Q000378:metabolism; D053586:Virus Internalization
chemical_list: D011991:Receptors, Virus; D014759:Viral Envelope Proteins; C039447:glycoprotein D, Human herpesvirus 1; C512478:EGFR protein, human; D066246:ErbB Receptors