Cellular Endosomal Escape via Photothermal Generated Membrane Disruption and Relaxation

Endosomal escape is a well-known hindrance for successful delivery of macromolecular drugs, genes and imaging agents into cell cytoplasm. Plasmonic-based photothermal heat generation and thermal relaxation/disruption of endosomal membranes is an approach to overcome this restriction. In this study, a pulsed laser (800nm) was used to induce heating of the NIR-transparent media with near infrared light-absorbing hollow gold nanoshells (HGNs) in living SK-BR-3 human breast carcinoma cells. In order to confirm endosomal escape SK-BR-3 cells were transfected to express RaB5a-GFP fusion. Upon early endosome formation, a localized protein within the endosome fluoresces identifying the endosome. Fluorophores of a different wavelength (Cy5) were attached to the HGNs surface via oligonucleotides, 24 bases in length, for fluorescent identification and tracking of the particles within the cells. Laser-induced elevated temperatures of the HGNs caused a heat gradient around the HGN clusters within the endosome. It was observed that upon cellular irradiation with a pulsed NIR laser thermal disruption and relaxation of the endosomal membrane occurred and the HGNs were released into the cytoplasm of the cell. Prior to laser irradiation, endosomal encapsulation of the fluorescently tagged HGNs within the GFP expressing endosome was confirmed via colocalization of the fluorescent markers. After irradiation, colocalization was lost and tagged HGNs were observed within the cytoplasm of the cell. The heating effect is localized resulting in no heating of the cellular fluid outside the endosomes. Endosome escape was not observed in cells that were irradiated with the laser where no HGNs were present under equal conditions.