Correlating transfection barriers and biophysical properties of cationic polymethacrylates.
Transfection efficiencies of several polymeric gene carriers were compared and correlated quantitatively to the amounts of cellular accumulation of plasmid DNA and to the expression of mRNA by quantitative real-time polymerase chain reaction (real-time PCR). Three polycations polymers with similar chemical structure were used in this study: poly(dimethylamino)ethyl methacrylate (PDMA) homopolymer, PEO-b-PDMA copolymer, and PEO-b-poly(diethylamino)ethyl methacrylate (PEO-b-PDEA) copolymer. Despite their similar chemical structures, the transfection efficiencies were significantly different. PEO-b-PDEA copolymer was significantly less efficient as gene carrier as compared to both PDMA and PEO-b-PDMA. Correlations between cytotoxicity, cellular uptake of plasmid DNA, expression levels of transgene and protein, and the physical properties of the polymers were observed. With the PEO-b-PDEA studies, cytotoxicity was due primarily to the excess of polymers that did not participate in the DNA binding. In addition, the inability of the polymer/DNA polyplexes to interact with cell effectively was identified as a critical barrier for high efficiency of transfection. This study demonstrated that the use of quantitative real-time PCR in combination with physical characterization techniques could provide useful insights into the transfection barrier at different cellular levels.
Tan
J F
JF
Singapore-MIT Alliance, and Department of Chemical Engineering, Massachusetts Institute of Technology, Cambridge, Massachusetts 02139, USA.
Hatton
T A
TA
Tam
K C
KC
Too
H P
HP
D000178:Acrylamides; D002412:Cations; D002460:Cell Line; D002470:Cell Survival; D006801:Humans; D010957:Plasmids / Q000032:analysis / Q000493:pharmacokinetics*; D011092:Polyethylene Glycols; D016133:Polymerase Chain Reaction; D011109:Polymethacrylic Acids; D012333:RNA, Messenger / Q000032:analysis* / Q000096:biosynthesis; D014162:Transfection / Q000592:standards*
D000178:Acrylamides; D002412:Cations; D011109:Polymethacrylic Acids; D012333:RNA, Messenger; C429790:poly(N,N-dimethylacrylamide); D011092:Polyethylene Glycols