Jiří Pánek

Efficient Condensation of DNA into Environmentally Responsive Polyplexes Produced from Block Catiomers Carrying Amine or Diamine Groups

The intracellular delivery of nucleic acids requires a vector system as they cannot diffuse across lipid membranes. Although polymeric transfecting agents have been extensively investigated, none of the proposed gene delivery vehicles fulfill all of the requirements needed for an effective therapy, namely, the ability to bind and compact DNA into polyplexes, stability in the serum environment, endosome-disrupting capacity, efficient intracellular DNA release, and low toxicity. The challenges are mainly attributed to conflicting properties such as stability vs efficient DNA release and toxicity vs efficient endosome-disrupting capacity. Accordingly, investigations aimed at safe and efficient therapies are still essential to achieving gene therapy clinical success. Taking into account the mentioned issues, herein we have evaluated the DNA condensation ability of poly(ethylene oxide)113-b-poly[2-(diisopropylamino)ethyl methacrylate]50 (PEO113-b-PDPA50), poly(ethylene oxide)113-b-poly[2-(diethylamino)ethyl methacrylate]50 (PEO113-b-PDEA50), poly[oligo(ethylene glycol)methyl ether methacrylate]70-b-poly[oligo(ethylene glycol)methyl ether methacrylate10-co-2-(diethylamino)ethyl methacrylate47-co-2-(diisopropylamino)ethyl methacrylate47] (POEGMA70-b-P(OEGMA10-co-DEA47-co-DPA47), and poly[oligo(ethylene glycol)methyl ether methacrylate]70-b-poly{oligo(ethylene glycol)methyl ether methacrylate10-co-2-methylacrylic acid 2-[(2-(dimethylamino)ethyl)methylamino]ethyl ester44} (POEGMA70-b-P(OEGMA10-co-DAMA44). Block copolymers PEO113-b-PDEA50 and POEGMA70-b-P(OEGMA10-co-DEA47-co-DPA47) were evidenced to properly condense DNA into particles with a desirable size for cellular uptake via endocytic pathways (R(H) ≈ 65-85 nm). The structure of the polyplexes was characterized in detail by scattering techniques and atomic force microscopy. The isothermal titration calorimetric data revealed that the polymer/DNA binding is endothermic; therefore, the process in entropically driven. The combination of results supports that POEGMA70-b-P(OEGMA10-co-DEA47-co-DPA47) condenses DNA more efficiently and with higher thermodynamic outputs than does PEO113-b-PDEA50. Finally, circular dichroism spectroscopy indicated that the conformation of DNA remained the same after complexation and that the polyplexes are very stable in the serum environment.

Tungsten (VI) based “molecular puzzle” photoluminescent nanoparticles easily covered with biocompatible natural polysaccharides via direct chelation

Multimodal probes, which can be simultaneously visualized by multiple imaging modalities, enable the cellular uptake, intracellular fate, biodistribution and elimination to be tracked in organisms. In this study, we report the synthesis of crystalline WO3 and CaWO4 doped with Eu3+ or Tb3+ nanoparticles (size range of 10-160u202fnm) coated with polysaccharides, and these nanoparticles constitute a versatile easy-to-construct modular toolbox for multimodal imaging. The particles adsorb significant amounts of polysaccharides from the solution, providing biocompatibility and may serve as a platform for labeling. For WO3, the sorption is reversible. However, on CaWO4, stable coating is formed. CaWO4/Tb3+ coated with chemisorbed dextrin, mannan, guar gum and sodium alginate successfully underwent endocytosis with HepG2 cells and was visualized using confocal microscopy.

Photoluminescent polysaccharide-coated germanium(IV) oxide nanoparticles

In current biomedically oriented research, the development of a biomimetic nanoparticle platform is of interest to provide a molecular toolbox (i.e., allowing easy modular exchange of its parts depending on actual needs while being nontoxic and allowing real-time recognition and tracking using various methods, such as fluorescence). We report the development of germanium(IV) oxide-polysaccharide composite particles possessing these properties. The nanoparticles are based on a crystalline germanium oxide core with a size range of 20–30 and 300–900xa0nm. Two new simple coating techniques were compared for the preparation of the photoluminescent polysaccharide-coated germanium(IV) oxide nanoparticles. The germanium(IV)-based core allows for in situ polysaccharide attachment via direct chelation. In addition, the nanoparticles were coated with thin layer of silicon oxide. After coating, 3-(triethoxysilyl)propyl isocyanate was grafted onto the surface, and the polysaccharides were immobilized on the particle surface via a covalent urethane linkage, which allows for an even more stable polysaccharide coating than that obtained via chelation. This approach provides access to a new material platform for biological track and image applications.

Polymeric Nanoparticles Stabilized by Surfactants: Kinetic Studies

A kinetic study was carried out to monitor the time-dependent formation of surfactant-stabilized polymeric nanoparticles prepared by controlled phase separation. The nanoparticles were made of poly(methylmethacrylate) or polystyrene and several ionic and nonionic surfactants. The dispersion was prepared by fast mixing a solution of the polymers in organic solvents with a solution of a surfactant in water. We observed the formation of well-defined nanoparticles measured by time-resolved small-angle x-ray scattering (TR-SAXS). Our results suggest that the kinetics for the formation of nanoparticles comprising ionic surfactants is much faster (in the range of milliseconds) than that for nanoparticles comprising nonionic surfactants (on the time scales of several seconds). We were able to observe the transformation of particle surface from transient structure to hard sphere one in real time. Some systems manifest a plethora of structural changes that demands further experimental research.