Chitosan Nanoplexes For Target siRNA Delivery

DSpace Repository


Dateien:

URI: http://hdl.handle.net/10900/69518
http://nbn-resolving.de/urn:nbn:de:bsz:21-dspace-695189
http://dx.doi.org/10.15496/publikation-10932
Dokumentart: PhDThesis
Date: 2016
Language: English
Faculty: 7 Mathematisch-Naturwissenschaftliche Fakultät
7 Mathematisch-Naturwissenschaftliche Fakultät
Department: Chemie
Advisor: Laufer, Stefan (Prof. Dr.)
Day of Oral Examination: 2015-12-17
DDC Classifikation: 500 - Natural sciences and mathematics
540 - Chemistry and allied sciences
Keywords: Chitosan , Nanopartikel , Polymere
Other Keywords:
nanoparticles
gene delivery
License: http://tobias-lib.uni-tuebingen.de/doku/lic_mit_pod.php?la=de http://tobias-lib.uni-tuebingen.de/doku/lic_mit_pod.php?la=en
Order a printed copy: Print-on-Demand
Show full item record

Abstract:

In recent years biopharmaceutical companies have started to turn their attention towards small interfering RNA (siRNA) therapeutics, as they offer a strategy to address therapeutically interesting targets that are not “druggable” with classic small molecule inhibitors. siRNAs can be easily adapted to any target of interest; they operate upstream of the protein production by silencing the messenger RNA (mRNA) before it is translated into pathogenic or disease-related proteins. However, the successful application of siRNA based therapy strongly depends on the development of an efficient delivery system to (1) specifically target a particular cell type, (2) protect the siRNA from ribonucleases enzymatic degradation, (3) be taken up by the cells, and (4) release the siRNA in the cytoplasm where the targeted mRNA is located. Therefore, the main focus of this study was to design, synthesize and characterize an efficient and specific delivery system for siRNA. Chitosan is a versatile, biocompatible and biodegradable polymer with high positive charge density. As such, it can be modified and optimized with various domains, thus proving to be very suitable as a gene delivery system. Despite the advantage of being biocompatible and biodegradable, chitosan has also some drawbacks, one of them being low solubility at physiological pH which can influence the formation of nanoplex (NP) aggregation. Upon contact with biological fluids, chitosan:siRNA NP may unspecifically interact with the charged biological components. Therefore, chitosan was first conjugated with increasing grafting ratios of hydrophilic polyethylene glycol (PEG). The systematical analysis of various chitosan_PEGs enabled the identification of a defined PEG ratio with a low impact on the advantageous physicochemical characteristics of chitosan NP, whilst maintaining high gene transfection efficiency. Further work was focused on a specific targeting of the NP. Two different strategies were pursued: targeting with a small molecule inhibitor as well as targeting via a specific antibody fragment. The exposed position of the targeting moiety on the surface of the NP, outside of any shielding effect, is one of the basic requirements for the effective addressing of the target. Therefore the main building block of the linker system had to be based on a spacer system. PEG was the obvious choice as the spacer moiety due to the positive results with 6 chitosan_PEG NP. A heterofunctional linker system was developed, with one end selective to the targeting moiety and other end selective to the modified chitosan. The siRNA delivery system was designed with the aim of incorporating the NP on the surface of implantable medical devices. Therefore, a layer-by-layer coating approach of solid surfaces was tested with chitosan_PEG/siRNA NP with regards to layer deposition and release of the NP over time. Overall, this study provided a delivery system with high transfection efficiency, achieved by an optimized PEG ratio. The developed system was further modified for specific cell targeting with both biological molecules and chemical inhibitors via a highly flexible heterofunctional linker system. Together with the layer by layer coating, the basis for a highly flexible and efficient targeted siRNA delivery system was successfully developed, providing a broad know-how for the further development of various siRNA delivery systems.

This item appears in the following Collection(s)