title: Role of TLR3 in the immunogenicity of replicon plasmid-based vaccines.
abstract: Replicon plasmids encoding an alphavirus RNA replicase constitute an alternative to conventional DNA plasmids with promise for DNA vaccination in humans. Replicase activity amplifies the levels of transgene mRNA through a copying process involving double-stranded (ds) RNA intermediates, which contribute to vaccine immunogenicity by activating innate antiviral responses. Toll-like receptor 3 (TLR3) is a dsRNA innate immune receptor expressed by antigen-presenting dendritic cells (DCs). Here, we test the hypothesis that TLR3 is necessary for the immunogenicity of replicon plasmid-based DNA vaccines. We show that mouse CD8 alpha(+) DC phagocytose dying replicon plasmid-transfected cells in vitro and are activated in a TLR3-dependent manner by dsRNA present within those cells. However, we find that cytotoxic T-cell responses to a replicon plasmid intramuscular vaccine are not diminished in the absence of TLR3 in vivo. Our results underscore the potential role of TLR3 in mediating immune activation by dsRNA-bearing replicon plasmid-transfected cells and indicate that other innate sensing pathways can compensate for TLR3 absence in vivo.
authors.affiliation: Immunobiology Laboratory, Cancer Research UK, London Research Institute, London, UK. sandra.diebold@kcl.ac.uk
authors.forename: S S
authors.initials: SS
authors.lastname: Diebold
authors.forename: O
authors.initials: O
authors.lastname: Schulz
authors.forename: L
authors.initials: L
authors.lastname: Alexopoulou
authors.forename: W W
authors.initials: WW
authors.lastname: Leitner
authors.forename: R A
authors.initials: RA
authors.lastname: Flavell
authors.forename: C
authors.initials: C
authors.lastname: Reis e Sousa
mesh_terms: D000818:Animals; D017209:Apoptosis / Q000276:immunology; D016827:CD8 Antigens / Q000032:analysis; D018414:CD8-Positive T-Lymphocytes / Q000276:immunology; D002522:Chlorocebus aethiops; D018920:Coculture Techniques; D003602:Cytotoxicity, Immunologic / Q000276:immunology; D003713:Dendritic Cells / Q000276:immunology; D015870:Gene Expression / Q000276:immunology; D041722:Genes, Transgenic, Suicide; D005822:Genetic Vectors / Q000276:immunology; D051379:Mice; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D010957:Plasmids / Q000276:immunology; D012330:RNA, Double-Stranded / Q000096:biosynthesis; D012093:Replicon / Q000276:immunology*; D013154:Spleen / Q000276:immunology; D051196:Toll-Like Receptor 3 / Q000276:immunology*; D014162:Transfection; D014611:Vaccination / Q000379:methods; D019444:Vaccines, DNA / Q000276:immunology*; D014709:Vero Cells
chemical_list: D016827:CD8 Antigens; C409469:CD8 antigen, alpha chain; D012330:RNA, Double-Stranded; C519703:TLR3 protein, mouse; D051196:Toll-Like Receptor 3; D019444:Vaccines, DNA