Cationic Nanohydrogel Particles as Potential siRNA Carriers for Cellular Delivery

Oligonucleotides such as short, double-stranded RNA (siRNA) or plasmid DNA (pDNA) promise high potential in gene therapy. For pharmaceutical application, however, adequate drug carriers are required. Among various concepts progressing in the market or final development, nanosized hydrogel particles may serve as novel transport media especially for siRNA. In this work, a new concept of synthesizing polymeric cationic nanohydrogels was developed, which offers a promising strategy to complex and transport siRNA into cells. For this purpose, amphiphilic reactive ester block copolymers were synthesized by RAFT polymerization of pentafluorophenyl methacrylate as reactive ester monomer together with tri(ethylene glycol)methyl ether methacrylate. In polar aprotic solvents, a self-assembly of these polymers could be observed leading to the formation of nanometer-sized polymer aggregates. The resulting superstructures were used to convert the reactive precursor block copolymers with amine-containing cross-linker molecules into covalently stabilized hydrogel particles. Detailed dynamic light scattering studies showed that the structure of the self-assembled aggregates can permanently be locked-in by this process. This method offers a new possibility to synthesize precise nanohydrogels of different size starting from various block copolymers. Moreover, <i>via</i> reactive ester approach, further functionalities could be attached to the nanoparticle, such as fluorescent dyes, which allowed distinct tracing of the hydrogels during complexation with siRNA or cell uptake experiments. In this respect, cellular uptake of the particles themselves as well as with its payload could be detected successfully. Looking ahead, these novel cationic nanohydrogel particles may serve as a new platform for proper siRNA delivery systems

Click Modification of Multifunctional Liposomes Bearing Hyperbranched Polyether Chains

Aiming at controlled modification of liposomal surface structures, we describe a postpreparational approach for surface derivatization of a new type of multifunctional, sterically stabilized liposomes. Application of dual centrifugation (DC) resulted in high encapsulation efficiencies above 50% at very small batch sizes with a total volume of 150 μL, which were conductive to fast and efficient optimization of variegated surface modification reactions. Cholesterol-polymer amphiphiles, including complex hyperbranched polyether structures bearing 1–4 terminal alkynes, were used in DC formulations to provide steric stabilization. The alkyne moieties were explored as anchors for the conjugation of small molecules to the liposomal surface via click chemistry, binding 350–450 fluorophores per liposome as examples for surface active molecules. Using Förster resonance energy transfer (FRET) spectroscopy, the conjugation reaction as well as the uptake of FRET-labeled liposomes by RBE4 cells was monitored, and the distribution of the fluorescent lipids among cellular structures and membranes could be studied. Thus, the combination of clickable hyperbranched amphiphiles and dual centrifugation provides access to well-defined liposomal formulations with a variety of surface moieties