Nanoparticulate systems for nucleic acid delivery

Development of carrier systems with controllable physicochemical and delivery properties has opened up the possibility of nanomedicines containing nucleic acids. In the last decades, much effort has been dedicated to two exciting approaches in biomedicine, namely gene and RNA interference (RNAi)-based therapeutics. These two approaches have faced many hurdles as far as delivery issues are concerned, as an efficient cytosolic delivery of siRNA and nuclear delivery of plasmid DNA is necessary for RNAi and gene therapy, respectively. In this PhD project, several biodegradable and non-biodegradable polymeric delivery systems such as TMC, TMC-SH, pHPMA-MPPM, QNPHOS, pDMAEMA, PEI and also carbon nanotubes (CNTs) were studied for nucleic acid delivery in vitro and in vivo. All these carrier systems were able to form nanoparticles with siRNA. However, based on characteristics of the carrier systems such as their chemical structure and cationic charges, the size, zeta potential and colloidal stability of the particles were different. It was shown those polymers with two cationic groups per monomer unit and the polymers which formed disulfide bonds (due to presence of thiol groups in the structure) were able to form stronger complexes with nucleic acid. However, based on the data presented in this thesis, it was concluded that formation of strong siRNA complexes is indeed advantageous, since it enhances the stability of the polyplexes in biological fluids, contributing to an improved siRNA delivery resulting in a high gene silencing activity. On the other hand, formation of too stable polyplexes with DNA with a limited release, resulted in lack of gene expression. The results of this PhD project show high gene silencing activity of thiolated TMC (TMC-SH) (up to 70%), with a low cytotoxicity (less than 20%). Formation of disulfide bonds in the polyplexes based on this polymer, improved the colloidal stability of the particles. Also, pHPMA-MPPM and non thiolated TMC showed high gene silencing activity (up to 70%) after application of endosomal escape methods such as photochemical internalization (PCI). Functionalized CNTs showed a low silencing activity (up to 20%) and a high cytotoxicity (more than 50%). The biodegradable polymers studied in this thesis showed a much lower cytotoxicity than non-biodegradable carrier systems.