Amphiphilic Polypeptides for VEGF siRNA Delivery into Retinal Epithelial Cells

Abstract

Polyethyleneimine, poly-L-lysine, chitosan and some others cationic polymers have been thoroughly studied as nucleic acid delivery systems in gene therapy. However, the drug release from these systems proceeds at a very low rate due to extremely high binding between a carrier and gene material. To reduce these interactions and to enhance drug release, we developed a set of amphiphilic polypeptides containing positively and negatively charged amino acids as well as a hydrophobic one. The copolymers obtained were characterized by size-exclusion chromatography, static light scattering, HPLC amino acid analysis and 1HNMR spectroscopy. All copolymers formed particles due to a self-assembly in aqueous media. Depending on polypeptide composition, the formation of particles with hydrodynamic diameters from 180 to 900 nm was observed. Stability of polymer particles, loading and release efficiency were carefully studied. Cellular uptake of the particles was efficient and their cytotoxicity was negligible. The application of polymer carriers, containing siRNA, to vascular endothelial growth factor (VEGF-A165) silencing of ARPE-19 cells was successful. The gene silencing was confirmed by suppression of both messenger RNA and protein expression.

Polypeptide Self-Assembled Nanoparticles as Delivery Systems for Polymyxins B and E

“…The composition of the copolymers was determined by the quantitative HPLC (LC-20 Shimadzu system with refractometric RID-20A detector, Shimadzu, Kyoto, Japan) analysis of free amino acids. For this, the copolymer samples were hydrolyzed to free amino acids and analyzed as described in [ 36 ].…”

Niosome-Based Approach for In Situ Gene Delivery to Retina and Brain Cortex as Immune-Privileged Tissues

“…Most of those cationic polymers include chitosans [55], polyethylenimine [56], or poly (L-lysine) [57]. Moreover, hybrid compounds, made by a combination of both polycationic and polyanionic polymers, and with different organic and inorganic materials such as polymers, magnetite, or lipids can be used also to deliver genetic material for different purposes [58][59][60][61][62].…”

Avoiding the Pitfalls of siRNA Delivery to the Retinal Pigment Epithelium with Physiologically Relevant Cell Models

“…The suitability of a certain carrier varies depending on the tissue, thus careful in vitro testing in physiologically relevant tissue models is required to select the optimal siRNA delivery system for the tissue of interest. In the case of the RPE, various carriers have been evaluated for their siRNA knockdown efficacy in vitro [22][23][24][25][26]; unfortunately, these studies have been performed using dividing cells that do not appropriately represent the terminally differentiated, post-mitotic RPE found in vivo [27]. For example, our recent study [28] has shown that the promising knockdown levels obtained with dividing RPE cells did not translate to adequate efficacy in differentiated cells.…”