Modular Construction of Fluorescent Lipophosphoramidates by Click Chemistry

Abstract

International audienceThe synthesis of fluorescent lipophosphoramidates is reported. Their modular construction allows several variations of the molecular structure including the separation of the fluorescent probe from the lipophosphoramidate moiety by a short methylene or by a tetraethylene glycol spacer. The last step of the convergent synthesis is a Huisgen click reaction, which assembles the fluorescent probe with the lipophosphoramidate part. Five fluorescent probes have been considered in this study including coumarin, NBD, fluorescein, naphthalimide and cyanine

Evaluation of a 7‐Methoxycoumarin‐3‐carboxylic Acid Ester Derivative as a Fluorescent, Cell‐Cleavable, Phosphonate Protecting Group

“…These results show that bulky fluorescent systems can be tolerated by cellular esterases without loss of activity or generation of toxicity relative to the commonly-used POM protection. Fluorescent prodrugs [20] are useful to detect the speed of cellular uptake in a way that is much faster and requires significantly smaller volumes of cells and materials relative to radiolabeling [21] or HPLC [22] approaches, making them amenable to high-throughput applications. While our current results are focused on release of the mono-acid form, we predict a parallel strategy could be used to modify phosphonates to release the di-acid form of compounds that require full deprotection for biological activity.…”

Evaluation of New Fluorescent Lipophosphoramidates for Gene Transfer and Biodistribution Studies after Systemic Administration

“…In addition, an excitation source is needed, and can sometimes cause autofluorescence, phototoxicity or photobleaching phenomena [ 4 ]. Fluorescence technology also requires the use of fluorochromes such as cyanine, naphthalimide, NBD (7-Nitrobenz-2-oxa-1,3-diazol-4-yl), fluorescein and many others, that can be used alone or coupled to different compounds like synthetic carriers [ 5 , 6 , 7 , 8 , 9 ] or peptides [ 10 ]. Among non-viral vectors, cationic lipids are the most commonly used, and some are currently employed in gene therapy clinical trials [ 11 , 12 , 13 , 14 ].…”

“…Thus, different cationic lipids such as lipophosphonates [ 15 , 16 , 17 ] and lipophosphoramidates [ 12 , 13 , 18 , 19 , 20 , 21 ] have been produced, and their high ability to transfect cells in culture and lungs mice were demonstrated [ 13 , 22 , 23 , 24 , 25 , 26 , 27 ]. However, the plasmid type [ 27 , 28 , 29 ], the vector [ 18 , 26 , 29 , 30 , 31 , 32 ] as well as the nucleic acids (NA) or synthetic carrier’s labeling techniques [ 8 , 30 , 33 , 34 , 35 ], impact greatly the distribution in tissues as well as the transgene expression. Labeling liposomes with a fluorescent probe appears as a relevant procedure to establish biodistribution profiles of lipoplexes and their pharmacokinetics, irrespective of administration route [ 26 , 27 , 31 , 32 , 33 , 34 , 36 , 37 , 38 ].…”

“…Labeling liposomes with a fluorescent probe appears as a relevant procedure to establish biodistribution profiles of lipoplexes and their pharmacokinetics, irrespective of administration route [ 26 , 27 , 31 , 32 , 33 , 34 , 36 , 37 , 38 ]. In that sense, fluorescent lipophosphoramidates bearing a single fluorophore localized in the polar head domain were synthesized [ 8 ]. Indeed, the fluorophore (NBD, cyanine or naphtalimide) was coupled with a hydrophobic part that can be associated with other lipidic compounds.…”

“…Indeed, the fluorophore (NBD, cyanine or naphtalimide) was coupled with a hydrophobic part that can be associated with other lipidic compounds. These neutral fluorescent compounds were thus included at a 5% molar ratio with a lipophosphoramidate (KLN47) to produce a fluorescent liposomal solution [ 8 ]. They were incorporated in small proportions to prevent perturbations of the supramolecular assembly as well as potential modifications of the transfection properties [ 8 ].…”