Development of nanoparticular nucleic acid delivery system in breast cancer gene therapy

DEVELOPMENT OF NANOPARTICULAR NUCLEIC ACID DELIVERY SYSTEM IN BREAST CANCER GENE THERAPY ABSTRACT Objective: In this thesis, it is aimed to develop a novel non-viral gene therapy agent which can be used in the treatment of triple negative breast cancer with natural and synthetic polymers. Method: Nucleic acid delivery systems have been developed with natural and synthetic polymers for efficient transfection of plasmid DNA which encodes shRNA for silencing of STAT-3 gene expression in 4T1 triple negative mouse breast cancer cells. Chitosan nanoparticles with low, medium and high molecular weight natural chitosan polymers and polyplex delivery systems with low (linear) and high (branched) molecular weight synthetic polyethylenimine polymers were prepared. Physicochemical characterization was done by measuring of particle size and zeta potantial of the developed nucleic acid delivery systems. The effect of nucleic acid delivery systems on serum stability, cell proliferation, transfection efficiency and cell migration of breast cancer was evaluated. Results: Transfection to 4T1 metastatic breast cancer cells was achieved with linear and branched polyplexes prepared at different polyethyleneimine ratios. It was found that the B2S coded polyplex, which has 237 nm particle size and +21.9 mv zeta potential value and was prepared with the branched PEI at a ratio of 2:1 (w:w) polymer: DNA, was the most effective nucleic acid delivery system. It was observed that the B2S coded polyplex formulation prevented the movement of fibroblast cells in triple negative breast cancer cells which have heterogeneous cell structure (round and spindle), while it provided aggregation by increasing cohesion in the rounded cancer cells. Conclusion: The branched and linear polyethylenimine polyplexes can be used for transfection of nucleic acids. In triple negative breast cancer, suppression of the expression of the STAT-3 gene decelerates the migration rate of cells. Key words: Triple negative breast cancer, STAT-3, 4T1 cell, polyethyleneimine polyplex, chitosan nanoparticle, shRNA, pSIH1-puro-STAT3 shRNA pDNA