RNA interference is a technique to induce specific gene silencing, mediated by siRNA molecules, double stranded RNA molecules (19–27 bp) that, binding to mRNA in the cytoplasm of cells via complementary base pairing, induce mRNA degradation. Compared to the conventional therapies, RNAi represents an emerging paradigm for treatment of many diseases. However, owing to its hydrophilicity and negative charges, siRNA molecules are not readily taken up by cells and due to their chemical nature are susceptible to nuclease action that limits the possibility to effectively administer siRNA through most of the routes. For these reasons much effort have been realized in the development of safe and efficient siRNA delivery systems, including cationic copolymers able to form interpolyelectrolitic complexes (polyplexes) with siRNA. In this study, because of the considerable interest in the chemistry and use of renewable resources, a new cationic copolymer of Inulin (Inu) [1] was synthesized by the coupling of diethylenetriamine (DETA) molecules to Inulin side chains, obtaining Inu-DETA copolymer. The used synthetic procedure involved the Enhanced Microwave Synthesis (EMS) technique [2], in order to combine the employment of this abundant in nature and biocompatible polysaccharide with the possibility to obtain simple, cost-effective and safety siRNA delivery systems. Inu-DETA copolymer, with a derivatization degree of 28% mol/mol, was tested to prepare copolymer/siRNA polyplexes by interpolyelectrolytic interactions. Agarose gel retardation assay showed that this new copolymer was able to stop the electrophoretic run of siRNA starting from copolymer/siRNA weight ratio of 7. Moreover, DLS studies evidenced the copolymer was able to form with siRNA polyplexes in the nanoscaled range and bearing a positive surface charge. Studies to assess complex stability in the presence of albumin showed the ability of this system to resist to the anionic exchange. In addition, biocompatibility of polyplexes was demonstrated by citotocompatibility assays and transfection experiments showed the efficiency of polyplexes to transfect a model cell line, such as JHH6. This results encourage us to continue in this study to obtain efficient inulin based siRNA delivery systems. [1] Stevens C.V. et al. Chemical modification of Inulin: a valuable renewable resource and its industrial applications, Biomacromolecules, 2, 1-16 (2011). [2] Hayes B.L. Recent advances in microwave-assisted synthesis, Aldrichimica Acta, 37(2), 66-77 (2004).
SIMPLE, BIOCOMPATIBLE AND COST-EFFECTIVE INULIN BASED SIRNA DELIVERY SYSTEMS
Sardo C;Cavallaro G;
2014-01-01
Abstract
RNA interference is a technique to induce specific gene silencing, mediated by siRNA molecules, double stranded RNA molecules (19–27 bp) that, binding to mRNA in the cytoplasm of cells via complementary base pairing, induce mRNA degradation. Compared to the conventional therapies, RNAi represents an emerging paradigm for treatment of many diseases. However, owing to its hydrophilicity and negative charges, siRNA molecules are not readily taken up by cells and due to their chemical nature are susceptible to nuclease action that limits the possibility to effectively administer siRNA through most of the routes. For these reasons much effort have been realized in the development of safe and efficient siRNA delivery systems, including cationic copolymers able to form interpolyelectrolitic complexes (polyplexes) with siRNA. In this study, because of the considerable interest in the chemistry and use of renewable resources, a new cationic copolymer of Inulin (Inu) [1] was synthesized by the coupling of diethylenetriamine (DETA) molecules to Inulin side chains, obtaining Inu-DETA copolymer. The used synthetic procedure involved the Enhanced Microwave Synthesis (EMS) technique [2], in order to combine the employment of this abundant in nature and biocompatible polysaccharide with the possibility to obtain simple, cost-effective and safety siRNA delivery systems. Inu-DETA copolymer, with a derivatization degree of 28% mol/mol, was tested to prepare copolymer/siRNA polyplexes by interpolyelectrolytic interactions. Agarose gel retardation assay showed that this new copolymer was able to stop the electrophoretic run of siRNA starting from copolymer/siRNA weight ratio of 7. Moreover, DLS studies evidenced the copolymer was able to form with siRNA polyplexes in the nanoscaled range and bearing a positive surface charge. Studies to assess complex stability in the presence of albumin showed the ability of this system to resist to the anionic exchange. In addition, biocompatibility of polyplexes was demonstrated by citotocompatibility assays and transfection experiments showed the efficiency of polyplexes to transfect a model cell line, such as JHH6. This results encourage us to continue in this study to obtain efficient inulin based siRNA delivery systems. [1] Stevens C.V. et al. Chemical modification of Inulin: a valuable renewable resource and its industrial applications, Biomacromolecules, 2, 1-16 (2011). [2] Hayes B.L. Recent advances in microwave-assisted synthesis, Aldrichimica Acta, 37(2), 66-77 (2004).I documenti in IRIS sono protetti da copyright e tutti i diritti sono riservati, salvo diversa indicazione.
