Protein Corona Attenuates the Targeting of Antitumor Sialyl Lewis X-Decorated Liposomes to Vascular Endothelial Cells under Flow Conditions.

Previously, we showed in the human umbilical vein endothelial cells (HUVECs) model that a liposome formulation of melphalan lipophilic prodrug (MlphDG) decorated with selectin ligand tetrasaccharide Sialyl Lewis X (SiaLeX) undergoes specific uptake by activated cells and in an in vivo tumor model causes a severe antivascular effect. Here, we cultured HUVECs in a microfluidic chip and then applied the liposome formulations to study their interactions with the cells in situ under hydrodynamic conditions close to capillary blood flow using confocal fluorescent microscopy. The incorporation of 5 to 10% SiaLeX conjugate in the bilayer of MlphDG liposomes increased their consumption exclusively by activated endotheliocytes. The increase of serum concentration from 20 to 100% in the flow resulted in lower liposome uptake by the cells. To elucidate the possible roles of plasma proteins in the liposome-cell interactions, liposome protein coronas were isolated and analyzed by shotgun proteomics and immunoblotting of selected proteins. Proteomic analysis showed that a gradual increase in SiaLeX content correlated with the overall enrichment of the liposome-associated proteins with several apolipoproteins, including the most positively charged one, ApoC1, and serum amyloid A4, associated with inflammation, on the one hand, and a decrease in the content of bound immunoglobulins, on the other. The article discusses the potential interference of the proteins in the binding of liposomes to selectins of endothelial cells.

Liposomal Formulation of a PLA2-Sensitive Phospholipid-Allocolchicinoid Conjugate: Stability and Activity Studies In Vitro.

To assess the stability and efficiency of liposomes carrying a phospholipase A2-sensitive phospholipid-allocolchicinoid conjugate (aC-PC) in the bilayer, egg phosphatidylcholine and 1-palmitoyl-2-oleoylphosphatidylglycerol-based formulations were tested in plasma protein binding, tubulin polymerization inhibition, and cytotoxicity assays. Liposomes L-aC-PC10 containing 10 mol. % aC-PC in the bilayer bound less plasma proteins and were more stable in 50% plasma within 4 h incubation, according to calcein release and FRET-based assays. Liposomes with 25 mol. % of the prodrug (L-aC-PC25) were characterized by higher storage stability judged by their hydrodynamic radius evolution yet enhanced deposition of blood plasma opsonins on their surface according to SDS-PAGE and immunoblotting. Notably, inhibition of tubulin polymerization was found to require that the prodrug should be hydrolyzed to the parent allocolchicinoid. The L-aC-PC10 and L-aC-PC25 formulations demonstrated similar tubulin polymerization inhibition and cytotoxic activities. The L-aC-PC10 formulation should be beneficial for applications requiring liposome accumulation at tumor or inflammation sites.