Synthesis of glycosylated polyethylenimine with reduced toxicity and high transfecting efficiency

Bioorganic and Medicinal Chemistry Letters

Volumn 10, Issue 11, 2000, Pages 1233-1235

  Leclercq, Françoise ^a   Dubertret, Catherine ^a   Pitard, Bruno ^a   Scherman, Daniel ^a   Herscovici, Jean ^a  

^a SANOFI   (France)

Author keywords

[No Author keywords available]

Indexed keywords

DNA; POLYETHYLENEIMINE DERIVATIVE; SODIUM BOROHYDRIDE; TITANIUM DERIVATIVE;

ARTICLE; DNA TRANSFECTION; DRUG SYNTHESIS; HUMAN; HUMAN CELL; STRUCTURE ANALYSIS;

CELL SURVIVAL; GLYCOSYLATION; POLYETHYLENEIMINE; TRANSFECTION; TUMOR CELLS, CULTURED;

EID: 0034608330     PISSN: 0960894X     EISSN: None     Source Type: Journal    
DOI: 10.1016/S0960-894X(00)00195-5     Document Type: Article

References (30)

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25. A typical experimental process for PEI glycosylation is illustrated for 3a as follow: PEI 25 KDa from Aldrich (500 mg, 0.02 mmol) was dissolved in 20 mL of anhydrous ethanol. Maltose (252 mg, 0.7 mmol) was added and the resulting solution was stirred under nitogen atmosphere for 15 min. Titanium isopropoxide (0.295 mL; 1 mmol) was slowly added and the resulting mixture was kept stirring overnight. Sodium borohydride (28.5 mg; 0.75 mmol) was then added and stirring was carried on for 8 h. The reaction mixture was filtered and concentrated to 10 mL. The resulting solution was finally dialyzed for 12 h (exclusion size membrane 12000).Yield 80%.
26. DNA (plasmid vector pXL2774) is dissolved in NaCl 150 mM at 80 mg/mL. PEI polymers were diluted in water at double used concentration and mixed volume to volume with DNA solution. Immediately prior to transfection, cells were washed twice with 500 mL of serum free-medium. 50 mL of polymer/DNA complexes solution were added dropwise to the cells (three wells per ratio condition). When the cells were transfected in "absence of FCS", the medium was supplemented by fetal calf serum 2 h after transfection. After transfection, cells were incubated at 37 °C in humidified air with 5% CO2 for an additional 44 h, up to the evaluation of the efficiency of gene transduction.The gene expression was monitored using the Luciferase Assay System (promega) according to the manufactuer protocols; the toxicity of the polymer/DNA mixture was assassed by cell lysat concentrations. Results were reported as enzymatic activity per mg of lysate protein.
27. (a) Pitard, B.; Aguerre, O.; Airiau, M.; Lachages, A. M.; Boukhnikachvili, T.; Byk, G.; Dubertret, C.; Herviou, C.; Mayaux, J. F.; Crouzet, J. Proc. Natl. Acad. Sci. USA 1997, 94, 14412.
28. (b) Pitard, B.; Oudrhiri, N.; Vigneron, J. P.; Hauchecorne, M.; Aguerre, O.; Toury, R.; Airiau, M.; Ramasawmy, R.; Scherman, D.; Crouzet, J.; Lehn, J. M.; Lehn, P. Proc. Natl. Acad. Sci. USA 1999, 96, 2621.
29. Polyplexes of different, N/P ratios (expressed as moles of amines per mol of phosphate) were obtained by mixing equal volumes of various concentrations of PEI with plasmid DNA at the final concentration of 250 mg DNA/mL in 5% glucose and 20 mM NaCl.
30. a,b) using a Coulter N4 Plus particle analyzer (Coulter). The means particle diameter was obtained from the unimodal fit analysis following dynamic light scattering done at a 90° angle at 20 °C. All samples were measured at a DNA concentration of 10 μg/mL in 5% glucose and 20 mM NaCl.