Transfection efficacy and drug release depends upon the PEG derivative in cationic lipoplexes: Evaluation in 3D in vitro model and in vivo.

The goal of the study was to estimate transfection efficacy and drug release in function of the PEG derivative in cationic liposomes and lipoplexes in both 2D and 3D in vitro models as well as in a mouse model (in vivo). For this purpose, cationic PEGylated nanocarriers based on OrnOrnGlu(C16 H33 )2 lipopeptides were fabricated and characterized. The nanocarriers were loaded with DNA plasmid pGL3 or with siRNA targeting 5'-UTR region of Hepatitis C virus, and their transfection efficacies were studied by luciferase test or by PCR technique, respectively. The pGL3-lipoplexes containing PEG derivative b (6 mol % PEG) were selected as the most promising nanocarriers for further in vivo study. In vitro cytotoxicity assay of the pGL3-lipoplexes with the PEG derivative b showed 2- and 1.5-fold enhancements of IC50 levels for HEK293T and HepG2 cells, respectively. Accumulation of the liposomes in the cells was studied by confocal microscopy using both 2D (monolayer culture) and 3D (multicellular spheroids) in vitro models. The PEGylated liposomes were found to penetrate cells more slowly than unmodified ones (without PEG). Thus, maximum liposomes in the HEK293T cells was observed after 1 and 3 h in the case of 2D and 3D in vitro models, respectively. Biodistribution study in mice showed that the PEGylated lipoplexes containing the PEG derivative b were eliminated from the bloodstream more slowly, namely with the doubled half-life time, than unmodified ones. Thus, the enhanced transfection efficacy and prolonged drug release of the PEGylated lipoplexes containing the optimal PEG derivative was demonstrated. This approach could be promising for development of novel siRNA-based drugs.

The mixture of siRNAs targeted to IL-4 and IL-13 genes effectively reduces the airway hyperreactivity and allergic inflammation in a mouse model of asthma.

Bronchial asthma (BA) is one of the most common chronic inflammatory disease of airways. There are huge experimental data indicating that Th2-cytokines IL-4 and IL-13 play a key role in BA pathogenesis. They are implicated in the IgE synthesis, eosinophil infiltration to the lungs and in the development of airway hyperreactivity (AHR), that makes these cytokines the promising targets. Neutralization of IL-4 and IL-13 or its common receptor chain (IL-4Rα) by monoclonal antibodies substantially reduce asthma symptoms. RNA interference provides a novel method for regulation of gene expression by siRNA molecules. In this study we evaluated whether the siRNA targeted to IL-4 and IL-13 reduce BA symptoms in mice model. Experimental BA was induced in BALB/c mice by sensitization to ovalbumin allergen followed by intranasal challenge. The intranasal delivery of siRNAs targeted to IL-4 and IL-13 inhibited the lung expression of these cytokines by more than 50% that led to the attenuation of AHR and pulmonary inflammation; the quantity of eosinophils in lungs which are one of the major inflammatory cells involved in allergic asthma pathogenesis decreased by more than 50% after siRNA treatment. These data support the possibility of a dual IL-4 and IL-13 inhibition by locally delivered siRNAs which in turn leads to the suppression of allergen-induced pulmonary inflammation and AHR.