sa-ecf-oct-dna
DNA; Studies from the United States, Japan and Germany add new findings to DNA body of knowledge
New findings from the United States, Japan and Germany describe advances in DNA.
Study 1: Plasmid size up to 20 kbp does not limit effective in vivo lung gene transfer using compacted DNA nanoparticles.
According to recent research from the United States, "Nanoparticles consisting of single molecules of DNA condensed with polyethylene glycol-substituted lysine 30-mers efficiently transfect lung epithelium following intrapulmonary administration. Nanoparticles formulated with lysine polymers having different counter ions at the time of DNA mixing have distinct geometric shapes: trifluoroacetate or acetate counter ions produce ellipsoids or rods, respectively."
"Based on intracytoplasmic micro-injection studies, nanoparticle ellipsoids having a minimum diameter less than the 25-nm nuclear membrane pore efficiently transfect nondividing cells," said Tamara L. Fink and colleagues at Copernicus Therapeutics, Inc. "This 25-nm size restriction corresponds to a 5.8-kbp plasmid when compacted into spheroids, whereas the 8- to 11-nm diameter of rod-like particles is smaller than the nuclear pore diameter."
"In mice, up to 50% of lung cells are transfected after dosing with a rod-like compacted 6.9-kbp lacZ expression plasmid, and correction of the CFTR chloride channel was observed in humans following intranasal administration of a rod-like compacted 8.3-kbp plasmid," reported the scientists. "To further investigate the potential size and shape limitations of DNA nanoparticles for in vivo lung delivery, reporter gene activity of ellipsoidal and rod-like compacted luciferase plasmids ranging in size between 5.3 and 20.2 kbp was investigated."
"Equivalent molar reporter gene activities were observed for each formulation, indicating that micro-injection size limitations do not apply to the in vivo gene transfer setting," the researchers reported.
Fink and associates published their study in Gene Therapy (Plasmid size up to 20 kbp does not limit effective in vivo lung gene transfer using compacted DNA nanoparticles. Gene Ther, 2006;13(13):1048-1051).
For additional information, contact Mark J. Cooper, Copernicus Therapeutics Inc., 11000 Cedar Avenue, Suite 145, Cleveland, OH 44106, USA. mcooper@cgsys.com.
Study 2: According to a study from Japan, cationic lipid and plasmid DNA layered onto gold surface allows stent-assisted gene transfer.
"Intravascular stent-assisted gene transfer is an advanced approach for the therapy of vascular diseases such as atherosclerosis and stenosis. This approach requires a stent that allows local and efficient administration of therapeutic genes to the target cells at the vascular wall.
"To create such a stent, a method was developed for loading plasmid DNA onto the metal surface. The method involves the formation of self-assembled monolayer on the noble metal surface followed by electrostatic layer-by-layer (LBL) assembly of a cationic lipid/plasmid DNA complex and free plasmid DNA," wrote F. Yamauchi and colleagues, Kyoto University.
"In this in vitro feasibility study, the thin plainer film and the wire of gold were used as a substrate. The LBL assembly process was characterized by surface plasmon resonance spectroscopy and static contact angle measurement. Plasmid DNA loaded in the multilayer exhibited improved resistance against nuclease digestion. When cultured directly on the DNA-loaded surface, cells were transfected to express exogenous gene in the DNA loading-dependent manner," reported the authors.
The researchers concluded, "Plasmid DNA could also be transferred to endothelial cells from its apical side by placing the DNA-loaded gold wire onto the cell layer."
Yamauchi and colleagues published their study in Biomaterials (Layer-by-layer assembly of cationic lipid and plasmid DNA onto gold surface for stent-assisted gene transfer. Biomaterials, 2006;27(18):3497-3504).
Additional information can be obtained by contacting H. Iwata, Kyoto University, Institute of Frontier Medical Science, Sakyo Ku, 53 Kawahara Cho, Kyoto 6068507, Japan.
Study 3: A study from Germany has described nano-carriers for DNA delivery to the lung based upon a covalently-coupled TAT-derived peptide.
"Gene therapy aimed at the respiratory epithelium holds therapeutic potential for diseases such as cystic fibrosis and lung cancer. Polyethylenimine (PEI) has been utilized for gene delivery to the airways."
"In this study," wrote E. Kleemann and colleagues, University of Marburg, "we describe a new modification of PEI, in which an oligopeptide related to the protein transduction domain of HIV-1 TAT was covalently coupled to 25 kDa PEI (PEI) through a heterobifunctional polyethylenglycol (PEG) spacer resulting in a TAT-PEG-PEI conjugate."
"Improved DNA reporter gene complexation and protection was observed for small (approximately 90 nm) polyplexes as well as significantly improved stability against polyanions, Alveofact bronchial alveolar lining fluid and DNase."
The researchers explained, "To determine polyplex toxicity in vitro, MTT assays were performed and, for in vivo testing, the mice bronchial alveolar lavage was investigated for total cell counts, quantity of neutrophils, total protein and TNF-alpha concentration. All parameters suggest significantly lower toxicity for TAT-PEG-PEI."
"Transfection efficiencies of both PEI and TAT-PEG-PEI polyplexes with DNA were studied under in vitro conditions (A549) and in mice after intratracheal instillation. While luciferase expression in A549 cells was much lower for TAT-PEG-PEI (0.2 ng/mg protein) than for PEI (2 ng/mg), significantly higher transfection efficiencies for TAT PEG-PEI were detected in mice."
"Reporter gene expression was distributed through bronchial and alveolar tissue. Thus, TAT-PEG-PEI represents a new approach to non-viral gene carriers for lung therapy, comprising protection for plasmid DNA, low toxicity and significantly enhanced transfection efficiency under in vivo conditions," the authors concluded.
Kleemann and colleagues published their study in the Journal of Controlled Release (Nano-carriers for DNA delivery to the lung based upon a TAT-derived peptide covalently coupled to PEG-PEI. J Control Release, 2005;109(1-3 Sp. Is):299-316
Obesity, Fitness & Wellness Week
September 30, 2006
