Polyion complex micelles from plasmid DNA and poly(ethylene glycol)-poly(L-lysine) block copolymer as serum-tolerable polyplex system: physicochemical properties of micelles relevant to gene transfection efficiency.
Polyion complex (PIC) micelles composed of the poly(ethylene glycol)-poly(L-lysine) (PEG-PLL) block copolymer and plasmid DNA (pDNA) were investigated in this study from a physicochemical viewpoint to get insight into the structural feature of the PIC micellar vector system to show practical gene transfection efficacy particularly under serum-containing medium. The residual ratio (r) of the lysine units in PEG-PLL to the phosphate units of pDNA in the system significantly affects the size of the PIC micelles evaluated from dynamic light scattering, being decreased from approximately 120 to 80 nm with an increase in the r value for the region with r > or = 1.0. The zeta potential of the complexes slightly increased with r in the same region, yet maintained a very small absolute value and leveled off to a few mV at r approximately 2.0. These results suggest that the micelles are most likely to take the core-shell structure with dense PEG palisades surrounding the PIC core to compartmentalize the condensed pDNA. Furthermore, an increasing r value in the region of r > or = 1 induces a rearrangement of the stoichiometric complex formed at r=1.0 to the non-stoichiometric complex composed of the excess block copolymer. The association number of pDNA and the block copolymer in the micelle was estimated from the apparent micellar molecular weight determined by static light scattering measurements, indicating that a single pDNA molecule was incorporated in each of the micelles prepared from the PEG (Mw=12,000 g/mol)-PLL (polymerization degree of PLL segment: 48) (12-48) block copolymer at r=2.0. These 12-48/pDNA micelles showed a gene expression comparable to the lipofection toward cultured 293 cells, though 100 microM chloroquine was required in the transfection medium. Notably, even in the presence of serum, the PIC micelles achieved appreciable cellular association to attain a high gene expression, which is in sharp contrast with the drastic decrease in the gene expression for lipoplex system in the presence of serum. A virus-comparable size (approximately 100 nm) with a serum-tolerable property of the PIC micelles indeed suggests their promising feasibility as non-viral gene-vector systems used for clinical gene therapy.
Itaka
Keiji
K
Department of Materials Science and Engineering, Graduate School of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8656, Japan.
Yamauchi
Kyosuke
K
Harada
Atsushi
A
Nakamura
Kozo
K
Kawaguchi
Hiroshi
H
Kataoka
Kazunori
K
D001672:Biocompatible Materials / Q000737:chemistry; D001692:Biological Transport; D002460:Cell Line; D004247:DNA / Q000737:chemistry / Q000235:genetics; D004996:Ethidium; D031541:Fluorescence Resonance Energy Transfer; D006801:Humans; D007700:Kinetics; D008027:Light; D008823:Micelles; D008970:Molecular Weight; D010957:Plasmids / Q000737:chemistry* / Q000235:genetics; D011092:Polyethylene Glycols / Q000737:chemistry*; D011107:Polylysine / Q000737:chemistry*; D012542:Scattering, Radiation; D013050:Spectrometry, Fluorescence; D014162:Transfection / Q000379:methods
C099578:(O-methylpoly(ethylene glycol)-O'-succinyl)-N-epsilon-poly(L-lysine); D001672:Biocompatible Materials; D008823:Micelles; D011107:Polylysine; D011092:Polyethylene Glycols; D004247:DNA; D004996:Ethidium