title: Small molecule Toll-like receptor 7 agonists localize to the MHC class II loading compartment of human plasmacytoid dendritic cells.
abstract: TLR7 and TLR8 are intracellular sensors activated by single-stranded RNA species generated during viral infections. Various synthetic small molecules can also activate TLR7 or TLR8 or both through an unknown mechanism. Notably, direct interaction between small molecules and TLR7 or TLR8 has never been shown. To shed light on how small molecule agonists target TLRs, we labeled 2 imidazoquinolines, resiquimod and imiquimod, and one adenine-based compound, SM360320, with 2 different fluorophores [5(6) carboxytetramethylrhodamine and Alexa Fluor 488] and monitored their intracellular localization in human plasmacytoid dendritic cells (pDCs). All fluorescent compounds induced the production of IFN-α, TNF-α, and IL-6 and the up-regulation of CD80 and CD86 by pDCs showing they retained TLR7-stimulating activity. Confocal imaging of pDCs showed that, similar to CpG-B, all compounds concentrated in the MHC class II loading compartment (MIIC), identified as lysosome-associated membrane protein 1(+), CD63, and HLA-DR(+) endosomes. Treatment of pDCs with bafilomycin A, an antagonist of the vacuolar-type proton ATPase controlling endosomal acidification, prevented the accumulation of small molecule TLR7 agonists, but not of CpG-B, in the MIIC. These results indicate that a pH-driven concentration of small molecule TLR7 agonists in the MIIC is required for pDC activation.
authors.affiliation: Novartis Vaccines & Diagnostics, Siena, Italy.
authors.forename: Carla
authors.initials: C
authors.lastname: Russo
authors.forename: Ivan
authors.initials: I
authors.lastname: Cornella-Taracido
authors.forename: Luisa
authors.initials: L
authors.lastname: Galli-Stampino
authors.forename: Rishi
authors.initials: R
authors.lastname: Jain
authors.forename: Edmund
authors.initials: E
authors.lastname: Harrington
authors.forename: Yuko
authors.initials: Y
authors.lastname: Isome
authors.forename: Simona
authors.initials: S
authors.lastname: Tavarini
authors.forename: Chiara
authors.initials: C
authors.lastname: Sammicheli
authors.forename: Sandra
authors.initials: S
authors.lastname: Nuti
authors.forename: M Lamine
authors.initials: ML
authors.lastname: Mbow
authors.forename: Nicholas M
authors.initials: NM
authors.lastname: Valiante
authors.forename: John
authors.initials: J
authors.lastname: Tallarico
authors.forename: Ennio
authors.initials: E
authors.lastname: De Gregorio
authors.forename: Elisabetta
authors.initials: E
authors.lastname: Soldaini
mesh_terms: D000225:Adenine / Q000031:analogs & derivatives* / Q000493:pharmacokinetics; D000634:Aminoquinolines / Q000493:pharmacokinetics*; D000970:Antineoplastic Agents / Q000493:pharmacokinetics; D002478:Cells, Cultured; D003713:Dendritic Cells / Q000378:metabolism*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005455:Fluorescent Antibody Technique; D005456:Fluorescent Dyes; D005802:Genes, MHC Class II / Q000502:physiology*; D006801:Humans; D007093:Imidazoles / Q000493:pharmacokinetics*; D000077271:Imiquimod; D018942:Macrolides / Q000494:pharmacology; D006180:Proton-Translocating ATPases / Q000037:antagonists & inhibitors; D011804:Quinolines / Q000737:chemistry / Q000493:pharmacokinetics; D051199:Toll-Like Receptor 7 / Q000819:agonists* / Q000378:metabolism
chemical_list: D000634:Aminoquinolines; D000970:Antineoplastic Agents; D004791:Enzyme Inhibitors; D005456:Fluorescent Dyes; D007093:Imidazoles; D018942:Macrolides; D011804:Quinolines; C508876:SM360320; C495348:TLR7 protein, human; D051199:Toll-Like Receptor 7; C057620:bafilomycin A; D006180:Proton-Translocating ATPases; D000225:Adenine; D000077271:Imiquimod; C402365:resiquimod