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Nonviral vector for efficient gene transfer to human ovarian adenocarcinoma cells
C K Kim1, K H Haider, S H Choi
1Physical Pharmacy Lab, College of Pharmacy, Seoul National University, San 56-1, Shinlim-Dong, Kwanak-Ku, Seoul, 151-742, Korea. ckkim@plaza.snu.ac.kr
Objective:
Various strategies have been attempted to design efficient protocols for ovarian cancer gene therapy but there has been little progress in their clinical application. In this study, we formulated and evaluated a new cationic liposome prepared with dioleoyltrimethylaminopropane (DOTAP), 1,2-dioleoyl-3-phosphophatidylethanolamine (DOPE), and cholesterol (Chol) (DDC) for plasmid DNA transfer into ovarian cancer cells.
Method:
The DDC liposome was prepared by mixing the DOTAP:DOPE:Cholin a 1:0.7:0.3 molar ratio using the extrusion method. Plasmid DNA (pEGFP-C1) and DDC were complexed at various weight ratios to find the optimum condition and the percentage of transfected cells was determined by selecting a green fluorescence protein (GFP) expressing cells in flow cytometry. The transfection efficiency of the DDC liposome was compared with 3[N-(N,N-dimethylaminoethylene) carbamoyl] cholesterol (DC-Chol)/DOPE liposome and commercially available lifopectin.
Results:
The optimal transfection of plasmid DNA was achieved at a 1:4 (w/w) ratio of DDC to DNA. The DDC/DNA complex exhibited higher transfection efficiency in human ovarian cancer cells (OVCAR-3 and SK-OV-3 cells) compared to that in other types of cell lines (NCI-NIH:522 and HepG2 cells). Flow cytometric analysis revealed that the DDC/DNA complex exhibited an over fourfold increase in GFP expression levels compared with DC-Chol/DOPE or lipofectin in OVCAR-3 cells. This result was further confirmed by confocal microscopy and RT-PCR analysis.
Conclusion:
These results suggest that our newly formulated cationic liposome (DDC) appears to be a promising nonviral vector for treating ovarian adenocarcinoma because of its selective high gene transfer ability in ovarian cancer cells.
Insights
A novel cationic liposome (DDC) effectively delivers plasmid DNA to ovarian cancer cells, showing promise for gene therapy. This DDC liposome demonstrates superior transfection efficiency compared to existing methods.
Area of Science:
- Biotechnology
- Gene Therapy
- Nanomedicine
Background:
- Ovarian cancer gene therapy faces challenges with inefficient delivery systems.
- Developing effective nonviral vectors is crucial for clinical translation.
Purpose of the Study:
- To formulate and evaluate a new cationic liposome (DDC) for enhanced plasmid DNA transfer into ovarian cancer cells.
- To assess the transfection efficiency of DDC liposomes compared to established methods.
Main Methods:
- DDC liposomes were prepared using dioleoyltrimethylaminopropane (DOTAP), 1,2-dioleoyl-3-phosphophatidylethanolamine (DOPE), and cholesterol (Chol) at a 1:0.7:0.3 molar ratio.
- Complexation of DDC with plasmid DNA (pEGFP-C1) was optimized, and transfection efficiency was quantified using flow cytometry for green fluorescence protein (GFP) expression.
- DDC liposome performance was benchmarked against DC-Chol/DOPE liposomes and Lipofectin.
Main Results:
- Optimal plasmid DNA transfection was achieved at a 1:4 (w/w) DDC to DNA ratio.
- DDC/DNA complexes demonstrated significantly higher transfection efficiency in ovarian cancer cell lines (OVCAR-3, SK-OV-3) compared to other cell types.
- GFP expression levels were over fourfold higher with DDC/DNA complexes in OVCAR-3 cells versus DC-Chol/DOPE or Lipofectin, confirmed by confocal microscopy and RT-PCR.
Conclusions:
- The novel DDC cationic liposome is a promising nonviral vector for ovarian cancer gene therapy.
- DDC exhibits selective and high gene transfer capabilities in ovarian cancer cells.
- This formulation represents a potential advancement for treating ovarian adenocarcinoma.