The main challenges for the delivery of genetic material (i.e. DNA, RNA, etc.) lie in the poor cellular penetration and targeting delivery. For example, negatively charged DNA biomacromolecule cannot be taken up by cell membranes without the aid of transporters or ‘vectors’ owing to its unfavourable surface charge and hydrodynamic volume. Additionally, the vectors should be equipped with an imaging agent (e.g. fluorescent entity) to improve the therapeutic DNA localization and its diagnostics.
Therefore, in our studies, we entrapped the DNA material (model genetic material from herring) in the newly-designed non-viral vectors, i.e, multifunctional nanocarriers obtained by layer-by-layer adsorption of DNA and oppositely charged polysaccharides (dextran or chitosan, DEX/CHIT) on the nanoemulsion core, loaded by cyanine IR-780 (used as the fluorescent marker) and stabilized by gemini N,N’-di-n-dodecyl-N,N,N’,N’-tetramethyl ethane-1,2-diaminium dibromide. Firstly, we determined ternary-phase diagrams of the surfactant–oil–water systems by titration method. Then, the stability of obtained nanoemulsion (Ne) with IR-780 solubilized in the isopropyl myristate oleic phase was evaluated by turbidimetry and ζ-potential. In the next step, CHIT, DNA and DEX layers were subsequently deposited on the Ne core. Finally, all IR-780-loaded nanocarriers covered by (DEX/CHIT/DNA/CHIT)2 bilayers with the size were subjected to colloidal stability studies and biological analysis on human cancer cell lines (MCF-7/WT, A549, MEWO), i.e., cytotoxic activity evaluated by MTT-based cytotoxity assay and bioimaging analyzed by CLSM and TIRFM.
Our results prove that the fabricated colloidal stable oil-core multilayer nanocapsules can be suitable as promising multifunctional nanocarriers for gene therapy and cancer diagnostics.
Acknowledgement: This work was financed by the National Science Center (Poland) under Grant No. 2012/05/B/ST4/00095.
urszula.bazylinska@pwr.wroc.pl
