Nanoboyutlu Polimerik Miseller Sistemler İle Gen Tedavisine Yönelik Yaklaşımlar

Abstract

Gene therapy is an effective treatment method that has received considerable attention recently in the treatment of many genetic diseases. The success of gene therapy depends on the development of an efficient carrier for the most safe and successful delivery of the therapeutic gene to the target tissue or cell. Recently, non-viral carrier systems, particularly polymeric systems based on cationic polymers, have received considerable attention in biomedical applications. In this thesis we aimed to prepare an efficient nanocarrier system with a novel biocompatible and biodegradable block copolymer synthesis for the successful transfer of the therapeutic BikDD gene used in the apoptotic gene treatment of breast cancer to the breast cancer cells. Also it is aimed to transfer this prepared carrier system to the MCF-7 and MDA-MB-468 cancer cells with high efficiency and to evaluate the in vitro cellular activity. In our study, cationic polymeric micelles which are among the promising gene delivery agents, have been used because of the low size, biocompatibility, high structural stability, long blood circulation time and adjustable amphiphilic properties. The production and characterization of suitable micelle formulations have been made by using PEtOx based block copolymers. In our thesis study, the biocompatible and biodegradable PEtOx polymer was used to form the hydrophilic block of micelle formulations and was proposed as a new alternative agent to the poly (ethylene glycol) (PEG) polymer with stealth feature.The physicochemical properties of the micelles which prepared by PEtOx-co-PEI-b-PEI cationic copolymer that contain a positive charge source linear Poli (ethyleneimine) (PEI) obtained by two different ratio (30% and 60%) with the partial hydrolysis of poly (2-ethyl-2-oxazoline) (PEtOx) were compared. In our study we aimed to reduce the various disadvantages of linear PEI, such as toxicity, depending on the production method of the block copolymer and the intense complex formation with DNA by utilizing the positive charge of PEI.Micelle/gene complexes were prepared by condansing the therapeutic BikDD gene with the micelles that prepared by block copolymers which contain the two different hydrolysis degrees of PEI and these complexes were characterized by comparing their characterizations, stability studies, in vitro cell viability assays, gene transfection efficiencies, gene expression analyzes and BikDD gene-induced apoptotic death efficiency. Among the micelles in these two different hydrolysis degrees, % 60 hydrolised micelles caused highest apoptozis mediated cellular death as 40% in MCF-7 cells and low cyototoxicity. In addition, when compared to PPP30 miceller systems, transfection efficiency was found to be approximately 6-fold higher in MCF-7 cells. The results showed that the efficiency of the micellar systems increased due to the increase in hydrolysis degree. All the obtained results within this thesis study showed that the new micellar systems prepared with PEtOx based block copolymers are effective carrier systems with successful results of apoptotic gene therapy in breast cancer. In particular, the PPP60 micelle / BikDD gene formulation was identified as the most effective formulation with low size, narrow polydispersity range, high positive charge, effective DNAse-I and serum stability, high gene condensation capacity, bioavailability in living cells, high transfection efficiency, intracellular expression and apoptosis inducing properties In conclusion, it was the first study to prepare a novel non-viral gene carrier micellar system from block copolymer containing PEI which synthesized by partial hydrolysis of PEtOx and to investigate the therapeutic gene transfer for apoptotic gene therapy of cancer. The obtained results have contributed to the literature in this respect.