Membrane Fusion Mediated Intracellular Delivery of Lipid Bilayer Coated Mesoporous Silica Nanoparticles.

Protein delivery into the cytosol of cells is a challenging topic in the field of nanomedicine, because cellular uptake and endosomal escape are typically inefficient, hampering clinical applications. In this contribution cuboidal mesoporous silica nanoparticles (MSNs) containing disk-shaped cavities with a large pore diameter (10 nm) are studied as a protein delivery vehicle using cytochrome-c (cytC) as a model membrane-impermeable protein. To ensure colloidal stability, the MSNs are coated with a fusogenic lipid bilayer (LB) and cellular uptake is induced by a complementary pair of coiled-coil (CC) lipopeptides. Coiled-coil induced membrane fusion leads to the efficient cytosolic delivery of cytC and triggers apoptosis of cells. Delivery of these LB coated MSNs in the presence of various endocytosis inhibitors strongly suggests that membrane fusion is the dominant mechanism of cellular uptake. This method is potentially a universal way for the efficient delivery of any type of inorganic nanoparticle or protein into cells mediated by CC induced membrane fusion.

Application of Coiled Coil Peptides in Liposomal Anticancer Drug Delivery Using a Zebrafish Xenograft Model.

The complementary coiled coil forming peptides E4 [(EIAALEK)4] and K4 [(KIAALKE)4] are known to trigger liposomal membrane fusion when tethered to lipid vesicles in the form of lipopeptides. In this study, we examined whether these coiled coil forming peptides can be used for drug delivery applications. First, we prepared E4 peptide modified liposomes containing the far-red fluorescent dye TO-PRO-3 iodide (E4-Lipo-TP3) and confirmed that E4-liposomes could deliver TP3 into HeLa cells expressing K4 peptide on the membrane (HeLa-K) under cell culture conditions in a selective manner. Next, we prepared doxorubicin-containing E4-liposomes (E4-Lipo-DOX) and confirmed that E4-liposomes could also deliver DOX into HeLa-K cells. Moreover, E4-Lipo-DOX showed enhanced cytotoxicity toward HeLa-K cells compared to free doxorubicin. To prove the suitability of E4/K4 coiled coil formation for in vivo drug delivery, we injected E4-Lipo-TP3 or E4-Lipo-DOX into zebrafish xenografts of HeLa-K. As a result, E4-liposomes delivered TP3 to the implanted HeLa-K cells, and E4-Lipo-DOX could suppress cancer proliferation in the xenograft when compared to nontargeted conditions (i.e., zebrafish xenograft with free DOX injection). These data demonstrate that coiled coil formation enables drug selectivity and efficacy in vivo. It is envisaged that these findings are a step forward toward biorthogonal targeting systems as a tool for clinical drug delivery.