Abstract
BackgroundThe design of efficient drug delivery vectors requires versatile formulations able to simultaneously direct a multitude of molecular targets and to bypass the endosomal recycling pathway of cells. Liposomal-based vectors need the decoration of the lipid surface with specific peptides to fulfill the functional requirements. The unspecific binding of peptides to the lipid surface is often accompanied with uncontrolled formulations and thus preventing the molecular mechanisms of a successful therapy.ResultsWe present a simple synthesis pathway to anchor cysteine-terminal peptides to thiol-reactive lipids for adequate and quantitative liposomal formulations. As a proof of concept, we have synthesized two different lipopeptides based on (a) the truncated Fibroblast Growth Factor (tbFGF) for cell targeting and (b) the pH sensitive and fusogenic GALA peptide for endosomal scape.ConclusionsThe incorporation of these two lipopeptides in the liposomal formulation improves the fibroblast cell targeting and promotes the direct delivery of cargo molecules to the cytoplasm of the cell.
Highlights
The design of efficient drug delivery vectors requires versatile formulations able to simultaneously direct a multitude of molecular targets and to bypass the endosomal recycling pathway of cells
As a proof of concept, our results show that the incorporation of both synthesized bioconjugated lipopeptides in liposomal formulations improved the cell targeting and promoted the direct delivery of cargo molecules in the cytosolic moiety of cultured mouse embryonic fibroblasts (MEFs)
A first disulfide exchange occurs between 2-2-pyridyl disulfide (DPDS) and the thiol group of DPTE in acidic conditions
Summary
Lipid–peptide conjugation through pyridyl disulfide reaction chemistry The cysteine-containing tbFGF and a variant of the GALA peptide (GALA-Cys, carrying the cysteine residue at the carboxy terminal end) were conjugated to the thiol-reactive lipid 1,2-dipalmitoyl-sn-glycero3-phospho-thio-ethanol (DPTE) accomplished with two pyridyl disulfide exchange reactions (Fig. 1). After the optimization of the minimal concentration of ATP required its encapsulation into POPC liposomes carrying 10% mol of DPTE-tbFGF and 10% mol of DPTE-GALA (Additional file 1: Figure S4), MEFs were incubated for 6 h with POPC liposomes containing 10% mol of DPTE-tbFGF and increasing amounts of DPTEGALA (0, 5 and 10% mol) and loaded with 50 mM ATP. The change of the cellular ATP concentrations in MEFs upon exposure to a free ATP or b DPTE-tbFGF- and DPTE-GALA containing POPC liposomes with encapsulated ATP. In this case, MEFs were incubated for 6 h at 37 °C and the cellular ATP levels were measured after 12 h with Luciferin/Luciferase based assay (see main text for details)
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