An immobilized nanoparticle-based platform for efficient gene knockdown of targeted cells in the circulation

Abstract

It is well established that specific interaction between adhesion molecules of endothelial cells and receptors on leukocytes can separate and recruit leukocytes from the bloodstream to sites of inflammation and coagulation. Previously, we showed that P-selectin can be absorbed onto the surface of a bloodcompatible microrenathane tube, and the P-selectin-coated surface could successfully capture P-selectin receptor-positive stem cells from physiological shear flow in vitro and from the bloodstream in vivo. In this paper, P-selectin was covalently attached to the surface of nanoscale liposomes to create targeting nanoparticles (NPs). Small interfering RNA (siRNA) was encapsulated by these nanoscale liposomes, and the liposomes were stabilized by PEGylation with DSPE-PEG2000. Experiments showed that these P-selectin-, PEGylated-, nanoscale-liposomes (PS-DSPE-PEG NPs) could be absorbed onto the inner surface of microrenathane tubing. The coated surface could specifically capture targeted cells from physiological shear flow, efficiently deliver encapsulated siRNA into adherent cells and dramatically silence the targeted gene neutrophil elastase. With this device, we create a high localized concentration for siRNA delivery in the circulatory system, providing circulating target cells adequate time to interact with therapeutic materials. SiRNA is efficaciously delivered into specific target cells, thereby providing a powerful tool for highly efficient siRNA transfection and other therapeutic materials delivery in circulation. The method should prove especially useful for diseases derived from disorders of blood cells.

E-selectin liposomal and nanotube-targeted delivery of doxorubicin to circulating tumor cells

“…1) were prepared using a thin lipid film hydration method [32,33] with 125 mM ammonium sulfate (Sigma-Aldrich, St. Louis, MO, USA) followed by 10 freeze–thaw cycles and then extrusion as previously described [34] to prepare empty liposomes (EL). To prepare L-DXR, DXR HCL (Sigma-Aldrich) was encapsulated within ELs using the ammonium sulfate remote loading method as described previously [33] at a doxorubicin-to-lipid ratio of 0.2:1 (w/w).…”

“…Recombinant human E-selectin/Fc chimera (rhE/Fc) (R&D Systems, Minneapolis, MN) was conjugated to 1,2-Distearoyl-sn-Glycero-3-Phosphoethanolamine-N-Maleimide 2000 (DSPE-PEG 2000 maleimide) (Avanti Polar Lipids, Alabaster, AL, USA) via thiolation, and conjuguates were covalently attached to diluted EL or L-DXR as previously described [34] to make two forms of targeted liposomes, E-selectin-PEG-conjugated liposomal doxorubicin (ES-PEG L-DXR) and E-selectin-PEG-conjugated empty liposomes (ES-PEG EL). The volume ratio of EL or L-DXR in phosphate-buffered saline (PBS) and rhE/Fc-DSPE-PEG 2000 maleimide to the diluted liposomes was 1:1 and 1:12, respectively.…”

Glycomechanics of the Metastatic Cascade: Tumor Cell–Endothelial Cell Interactions in the Circulation

“…FUT3 siRNA can be delivered with a novel flow-based method using P-selectin-conjugated nanoliposomes. 61 At a physiological range of wall shear stress, the number of cancer cells recruited to the microtube surface (rolling/adherent) decreased dramatically after FUT3 knockdown, indicating the essential roles of FUTs in selectin-mediated cell adhesion. 175 To date, a myriad of ligands for the three selectins both on leukocytes and on cancer cells have been discovered.…”

Nanostructured Surfaces to Target and Kill Circulating Tumor Cells While Repelling Leukocytes

“…Our lab has previously demonstrated that selectin proteins and selectin-coated nanoparticles can be immobilized on the inner surfaces of microtubes for the capture of circulating cells [31], as well as for use as in vitro models of the microvasculature to examine the rolling adhesion of leukocytes [32] and cancer cells [33]. In this study, smooth microtubes were coated with only ESPEG L-DXR.…”