Application of polymers in nucleic acid delivery

Gene therapy and immunotherapy are powerful techniques in the treatment of many life threatening diseases. The major challenge in these therapies is to seek a safe and efficient delivery carrier for gene and antigen materials. Carriers are designed to protect these molecules from degradation, improve their stability and facilitate the delivery of them to the site of action. This research study aims to develop appropriate carriers for small interferencing RNA (siRNA), DNA, antigen and ajuvant respectively. In the case of siRNA, material encompassing mannose, polyethylene glycol (PEG) and polyethylenimine (PEI) was investigated. Two structures were assembled: in one construct, mannose was conjugated to PEI directly (Mannose-PEI-PEG) whilst in a second construct; the mannose was conjugated to PEI via a PEG spacer (PEI-PEGmannose). Confocal microscopy images suggested a faster escape and release of siRNA into the perinuclear region when siRNA was complexed with mannose-PEG-PEI. Mannosylation and PEGylation generated significant toxicity reduction compared to unmodified PEI alone. Real-time polymerase chain reaction (RT-PCR) results showed a significant decrease on mRNA knockdown when using modified PEIs. It was found that PEI-PEG-mannose was a stronger candidate for siRNA delivery because it displays lower toxicity, higher uptake efficiency and higher relative knockdown efficiencies. In the case of pDNA delivery, dextran was introduced to reduce the toxicity generated by PEI. PEI 2000 was more effective than PEI 800 in condensing DNA and inducing transfection when incorporated with dextran. The toxicity of dextran-PEI was greatly reduced when compared to unmodified PEI. Dextran-PEI was able to generate significantly higher transfection efficiencies than PEI alone in the presence of serum. An