Nonviral vector for efficient gene transfer to human ovarian adenocarcinoma cells.
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.


METHODS
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.


CONCLUSIONS
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.
Kim
C K
CK
Physical Pharmacy Lab, College of Pharmacy, Seoul National University, San 56-1, Shinlim-Dong, Kwanak-Ku, Seoul, 151-742, Korea. ckkim@plaza.snu.ac.kr
Haider
K H
KH
Choi
S H
SH
Choi
E J
EJ
Ahn
W S
WS
Kim
Y B
YB
D000230:Adenocarcinoma / Q000235:genetics* / Q000378:metabolism / Q000628:therapy; D002412:Cations; D002784:Cholesterol / Q000737:chemistry / Q000493:pharmacokinetics; D004247:DNA / Q000008:administration & dosage* / Q000235:genetics; D005229:Fatty Acids, Monounsaturated / Q000737:chemistry / Q000493:pharmacokinetics; D005260:Female; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors / Q000737:chemistry / Q000493:pharmacokinetics*; D020404:Glycerophospholipids / Q000737:chemistry / Q000493:pharmacokinetics; D049452:Green Fluorescent Proteins; D006801:Humans; D008055:Lipids / Q000737:chemistry / Q000493:pharmacokinetics*; D008081:Liposomes; D008164:Luminescent Proteins / Q000096:biosynthesis / Q000235:genetics; D010051:Ovarian Neoplasms / Q000235:genetics* / Q000378:metabolism / Q000628:therapy; D010714:Phosphatidylethanolamines; D000644:Quaternary Ammonium Compounds / Q000737:chemistry / Q000493:pharmacokinetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection; D014407:Tumor Cells, Cultured
C410262:1,2-dioleoyl-glycero-3-phosphatidyl ethanolamine; D002412:Cations; D005229:Fatty Acids, Monounsaturated; D020404:Glycerophospholipids; D008055:Lipids; D008081:Liposomes; D008164:Luminescent Proteins; D010714:Phosphatidylethanolamines; D000644:Quaternary Ammonium Compounds; D049452:Green Fluorescent Proteins; D004247:DNA; D002784:Cholesterol; C070046:1,2-dioleoyloxy-3-(trimethylammonium)propane