title: Smart tools and orthogonal click-like reactions onto small unilamellar vesicles.
abstract: Click-based reactions were conducted at the surface of small unilamellar vesicles (SUVs) to provide onto-vesicle chemistry with efficient innovative ready-for-use tools. For that purpose, four amphiphilic molecules were designed to insert into bilayers while presenting a reactive functional head. In this manner, a dioleylglycero-ethoxy-ethoxy-ethoxy-ethanamine (DOG-PEG4-NH2) was chosen as a common platform while the reactive amine head was converted into several electrophilic functions. Thus, two dioleylglycerol-based cyclooctyne anchors were prepared: cyclooct-1-yn-3-glycolic acid-based anchor (DOG-COA) and 1-fluorocyclooct-2-ynecarboxylic acid-based anchor (DOG-FCOA). The last one differed from the first one in that a fluorine atom reinforces the electrophilic properties of the unsaturated bond. In addition, a third dioleylglycerol-based triphenylphosphine (DOG-PPh3) was synthesized for the first time. These three innovative amphiphilic anchors were designed to react with any azide-based biomolecule following copper-free Huisgen 1,4-cycloaddition and Staudinger ligation, respectively. A fourth anchor bearing a 3,4-dibromomaleimide ring (DOG-DBM) was also unprecedentedly synthesized, to be further substituted by two thiols. Model reactions conducted in solution with either model biotinyl azide or model biotinyl disulfide gave good to total conversions and excellent isolated yields. The four new anchors were inserted into SUVs whose formula is classically used in in vivo biology. Stability and surface overall electrostatic charge were in the expected range and constant over the study. Then, the functionalized liposomes were ligated to biotin-based reagents and the experimental conditions were finely tuned to optimize the conversion. The biotinyl liposomes were demonstrated functional and totally accessible in an affinity test based on biotin scaffold quantification. Finally, DOG-FCOA's reactivity was confronted to that of DOG-DBM in a 'one-pot' orthogonal reaction. (Biotin-S)2 and TAMRA-N3 (tetramethylcarboxyrhodamine azide) were successively conjugated to the liposome suspension in a successful manner. These data implement and reinforce the interest of bioorthogonal click-like reactions onto lipid nanoparticles.
authors.affiliation: Laboratoire de Conception et Application de Molécules Bioactives, Equipe de BioVectorologie, UMR 7199 - CNRS/Université de Strasbourg, Faculté de Pharmacie, 74 route du Rhin, BP 60024, 67401 Illkirch Cedex, France.
authors.forename: Christophe
authors.initials: C
authors.lastname: Salomé
authors.affiliation: Laboratoire de Conception et Application de Molécules Bioactives, Equipe de BioVectorologie, UMR 7199 - CNRS/Université de Strasbourg, Faculté de Pharmacie, 74 route du Rhin, BP 60024, 67401 Illkirch Cedex, France.
authors.forename: Maria Vittoria
authors.initials: MV
authors.lastname: Spanedda
authors.affiliation: Laboratoire de Conception et Application de Molécules Bioactives, Equipe de BioVectorologie, UMR 7199 - CNRS/Université de Strasbourg, Faculté de Pharmacie, 74 route du Rhin, BP 60024, 67401 Illkirch Cedex, France.
authors.forename: Benoit
authors.initials: B
authors.lastname: Hilbold
authors.affiliation: Laboratoire de Conception et Application de Molécules Bioactives, Equipe de BioVectorologie, UMR 7199 - CNRS/Université de Strasbourg, Faculté de Pharmacie, 74 route du Rhin, BP 60024, 67401 Illkirch Cedex, France.
authors.forename: Etienne
authors.initials: E
authors.lastname: Berner
authors.affiliation: Laboratoire de Conception et Application de Molécules Bioactives, Equipe de BioVectorologie, UMR 7199 - CNRS/Université de Strasbourg, Faculté de Pharmacie, 74 route du Rhin, BP 60024, 67401 Illkirch Cedex, France.
authors.forename: Béatrice
authors.initials: B
authors.lastname: Heurtault
authors.affiliation: Laboratoire de Conception et Application de Molécules Bioactives, Equipe de BioVectorologie, UMR 7199 - CNRS/Université de Strasbourg, Faculté de Pharmacie, 74 route du Rhin, BP 60024, 67401 Illkirch Cedex, France.
authors.forename: Sylvie
authors.initials: S
authors.lastname: Fournel
authors.affiliation: Laboratoire de Conception et Application de Molécules Bioactives, Equipe de BioVectorologie, UMR 7199 - CNRS/Université de Strasbourg, Faculté de Pharmacie, 74 route du Rhin, BP 60024, 67401 Illkirch Cedex, France. Electronic address: frisch@unistra.fr.
authors.forename: Benoit
authors.initials: B
authors.lastname: Frisch
authors.affiliation: Laboratoire de Conception et Application de Molécules Bioactives, Equipe de BioVectorologie, UMR 7199 - CNRS/Université de Strasbourg, Faculté de Pharmacie, 74 route du Rhin, BP 60024, 67401 Illkirch Cedex, France. Electronic address: line.bourel@unistra.fr.
authors.forename: Line
authors.initials: L
authors.lastname: Bourel-Bonnet
mesh_terms: D057930:Click Chemistry; D015394:Molecular Structure; D013501:Surface-Active Agents / Q000138:chemical synthesis / Q000737:chemistry*; D053835:Unilamellar Liposomes / Q000138:chemical synthesis / Q000737:chemistry*
chemical_list: D013501:Surface-Active Agents; D053835:Unilamellar Liposomes
keywords: Copper-free click chemistry; Dibromomaleimide; Double ligation; Liposome; Staudinger