M Khodaei1, K Rostamizadeh2, A H Taromchi1, H Monirinasab3, M Fathi4,5. 1. Department of Biotechnology, School of Medicine, Zanjan University of Medical Sciences, Zanjan, Iran. 2. Department of Medicinal Chemistry, School of Pharmacy, Zanjan University of Medical Sciences, Zanjan, Iran. 3. Department of Clinical Biochemistry, School of Medicine, Zanjan University of Medical Sciences, Zanjan, Iran. 4. Department of Clinical Biochemistry, School of Medicine, Zanjan University of Medical Sciences, Zanjan, Iran. m_fathi@zums.ac.ir. 5. Cancer Gene Therapy Research Center, Zanjan University of Medical Sciences, Zanjan, Iran. m_fathi@zums.ac.ir.
Abstract
BACKGROUND AND OBJECTIVE: To use siRNA molecule as a therapeutic agent in gene silencing, an efficient delivery system is necessary. Stability and clearance by reticuloendothelial of siRNA still remains the major challenges for clinical application. Herein, we could develop new lipid-polymer hybrid nanoparticles (LPHNP) as a siRNA carrier to silence insulin-like growth factor type I (IGF-1R) gene overexpression in MCF-7 human breast cancer cell line. METHODS: Dimethyldioctadecylammonium bromide-methoxy poly(ethylene glycol)-poly (ε-caprolactone) (DDAB-mPEG-PCL) LPHNPs were synthesized using a single step nanoprecipitation method and characterized by dynamic light scattering (DLS) and atomic force microscopy (AFM) microscope. Cytotoxicity of the nanoparticles was assessed in the MCF7 cell line using 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. Desired LPHNP-siRNA complex was determined using different Nitrogen:Phosphate ratio (N/P) ratios and gel retardation. To determine the encapsulation efficiency of siRNA (%) in LPHNP, its absorbance was measured. The effect of the siRNA-LPHNP complex on IGF-1R silencing was assessed by reverse transcription-polymerase chain reaction (RT-PCR) RESULTS: LPHNP was synthesized using a single-step sonication method with a size below 100 nM. The viability of cells treated with hybrid nanoparticles was significantly greater than the corresponding cationic lipid (P < 0.01). As demonstrated by gel retardation assay, efficient siRNA binding to LPHNP occurred at N/P equal to 40 and siRNA encapsulation efficiency was found to be 95% ± 4 at this ratio. LPHNP-IGF-1R siRNA complex could be able to down-regulate the target more efficiently when it compared with the corresponded controls (P < 0.001). CONCLUSION: In conclusion, our results suggest that DDAB cationic lipid and mPEG-PCL copolymer hybrid nanoparticle may be a good candidate for efficient siRNA delivery.
BACKGROUND AND OBJECTIVE: To use siRNA molecule as a therapeutic agent in gene silencing, an efficient delivery system is necessary. Stability and clearance by reticuloendothelial of siRNA still remains the major challenges for clinical application. Herein, we could develop new lipid-polymer hybrid nanoparticles (LPHNP) as a siRNA carrier to silence insulin-like growth factor type I (IGF-1R) gene overexpression in MCF-7 human breast cancer cell line. METHODS: Dimethyldioctadecylammonium bromide-methoxy poly(ethylene glycol)-poly (ε-caprolactone) (DDAB-mPEG-PCL) LPHNPs were synthesized using a single step nanoprecipitation method and characterized by dynamic light scattering (DLS) and atomic force microscopy (AFM) microscope. Cytotoxicity of the nanoparticles was assessed in the MCF7 cell line using 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. Desired LPHNP-siRNA complex was determined using different Nitrogen:Phosphate ratio (N/P) ratios and gel retardation. To determine the encapsulation efficiency of siRNA (%) in LPHNP, its absorbance was measured. The effect of the siRNA-LPHNP complex on IGF-1R silencing was assessed by reverse transcription-polymerase chain reaction (RT-PCR) RESULTS: LPHNP was synthesized using a single-step sonication method with a size below 100 nM. The viability of cells treated with hybrid nanoparticles was significantly greater than the corresponding cationic lipid (P < 0.01). As demonstrated by gel retardation assay, efficient siRNA binding to LPHNP occurred at N/P equal to 40 and siRNA encapsulation efficiency was found to be 95% ± 4 at this ratio. LPHNP-IGF-1R siRNA complex could be able to down-regulate the target more efficiently when it compared with the corresponded controls (P < 0.001). CONCLUSION: In conclusion, our results suggest that DDAB cationic lipid and mPEG-PCL copolymer hybrid nanoparticle may be a good candidate for efficient siRNA delivery.
Authors: Femke M van de Water; Otto C Boerman; Alfons C Wouterse; Janny G P Peters; Frans G M Russel; Rosalinde Masereeuw Journal: Drug Metab Dispos Date: 2006-05-19 Impact factor: 3.922