OVCAR-3 cells internalize TAT-peptide modified liposomes by endocytosis

Abstract

For cytosolic delivery of liposomes containing macromolecular drugs, such as proteins or nucleic acids, it would be beneficial to bypass endocytosis to prevent degradation in the lysosomes. Recent reports pointed to the possibility that coupling of TAT-peptides to the outer surface of liposome particles would enable translocation over the cellular plasma membrane. Here, we demonstrate that cellular uptake of TAT-liposomes occurs via endocytosis rather than plasma membrane translocation. The coupling of HIV-1 derived TAT-peptide to liposomes enhances their binding to ovarian carcinoma cells. The binding was inhibited by the presence of heparin or dextran sulfate, indicating that cell surface proteoglycans are involved in the binding interaction. Furthermore, living confocal microscopy studies revealed that binding of the TAT-liposomes to the plasma membrane is followed by intracellular uptake in vesicular structures. Staining the endosomes and lysosomes demonstrated that fluorescent liposomal labels are present within the endosomal and lysosomal compartments. Furthermore, incubation at low temperature or addition of a metabolic or an endocytosis inhibitor blocked cellular uptake. In conclusion, coupling TAT-peptide to the outer surface of liposomes leads to enhanced endocytosis of the liposomes by ovarian carcinoma cells, rather than direct cytosolic delivery by plasma membrane translocation.

Effects of cell‐penetrating peptides and pegylation on transfection efficiency of polyethylenimine in mouse lungs

“…Similar results have been obtained for PEI [36,37]. Thirdly, data collected by Fretz et al [38] strongly suggest that TAT coupled to liposomes does not lead to an advantageous route of internalization compared to the unmodified liposomes. Similar to PEI, recent studies suggest uptake of CPPs is via clathrin-and caveolaemediated endocytosis, and macropinocytosis [1].…”

Cytosolic Delivery of Liposomally Targeted Proteins Induced by Photochemical Internalization

“…PEGliposomes (PEG-L) without an antibody attached were used as a control. Due to the PEG-layer surrounding these liposomes, the cellular association and uptake by OVCAR-3 cells is low as shown previously (9,19).…”

Arginine‐rich cell‐penetrating peptides

“…FACS analysis following trypsin treatment indicated a relatively slow rate of uptake, comparable to that of classical markers of endocytosis. Since then, many other studies have also observed inhibition of cellular uptake at 4°C and with chemical means that induce energy depletion, indicating an energy-dependent process as the major route for the internalization of cell-penetrating peptides [27][28][29][30][31][32][33].…”