Cationic angelica polysaccharide nanoparticle gene delivery system and preparation method thereof

A gene delivery system and cationization technology, applied in the field of cationized angelica polysaccharide nanoparticle gene delivery system, can solve the problems of inducing host immune response, limited loading capacity, and restricting wide application, etc., and achieve strong proliferation, good packaging and release Effect, the effect of simple preparation process

Active Publication Date: 2011-08-17
JIANGSU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0003] Commonly used GDSs include viral vectors and non-viral vectors. Although the former shows high transfection efficiency, its potential tumorigenicity, induction of host immune response, limited loading capacity, and high cost restrict its use in gene expression. Wide range of applications in therapeutic areas

Method used

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  • Cationic angelica polysaccharide nanoparticle gene delivery system and preparation method thereof
  • Cationic angelica polysaccharide nanoparticle gene delivery system and preparation method thereof
  • Cationic angelica polysaccharide nanoparticle gene delivery system and preparation method thereof

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Experimental program
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preparation example Construction

[0043] Preparation of refined angelica polysaccharide:

[0044] Take the dried root of Angelica sinensis, crush it, and extract it with water and alcohol (according to the following process: hot water extraction: material-to-liquid ratio: 1:5~20, extraction temperature: 60~90°C, extraction time 2~5h / time, extraction times: 2 times. After combining the two extracts, concentrate by rotary evaporation to 1 / 5-1 / 10 of the volume of the original extract, and then add 95% ethanol to the concentrate until the final concentration of ethanol is 65%-85%), Freeze-drying, Angelica sinensis crude polysaccharide, trichloroacetic acid method to remove protein, dialysis (molecular weight cut-off>3500Da); successively use DEAE-52 cellulose resin (eluent: double distilled water and 0.05~0.5mol / L NaOH) and SephadexG- 100 dextran gel resin (eluent: 0.1mol / LNaCl) was used to separate and purify it, and the molecular weight distribution of the isolated product was determined by gel chromatography....

Embodiment 1

[0046] Take 0.2g refined angelica polysaccharide, dissolve it in 10ml phosphate buffer (pH=7); dissolve 0.2g carbonylimidazole, the linker for activating hydroxyl groups, in 5ml dichloromethane, and under the protection of nitrogen, first dissolve it in the polysaccharide solution Add 0.05ml of catalyst triethylamine, then slowly add the dichloromethane solution of the hydroxyl linker into the polysaccharide solution, stir at a constant speed, and complete the addition within 60 minutes. After the addition, react at room temperature for 120 minutes to obtain an activated polysaccharide solution; Molecular weight PEI was dissolved in 10ml of phosphate buffer, added catalyst triethylamine, slowly added to the activated polysaccharide solution in the dark, under the protection of nitrogen, at room temperature, the addition was completed within 120min, and reacted for 10h in the dark, at room temperature, The whole reaction was carried out under uniform stirring; after the reaction...

Embodiment 2

[0049] Take 0.8g of refined Angelica polysaccharide, dissolve it in 20ml of phosphate buffer (pH=7); Under the protection of nitrogen, first add 0.8ml of catalyst triethylamine to the polysaccharide solution, then slowly add the dichloromethane solution of the hydroxyl linker into the polysaccharide solution, stir at a constant speed, and complete the addition within 45 minutes. After the addition, react at room temperature for 90 minutes , to obtain an activated polysaccharide solution; dissolve 30 g of small molecular weight PEI in 10 ml of phosphate buffer, add catalyst triethylamine, and slowly add it to the activated polysaccharide solution in the dark, under nitrogen protection, at room temperature, and add within 150 min. After the reaction was completed, the reaction was carried out at room temperature for 10 h in the dark, and the whole reaction was carried out under constant stirring; after the reaction was completed, the solution was dialyzed (cutoff molecular weight...

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Abstract

The invention discloses a cationic angelica polysaccharide nanoparticle gene delivery system. The system is a gene delivery system of angelica polysaccharide combined DNA (Deoxyribonucleic Acid) plasmids modified by amine compounds, wherein molecular weight distribution of angelica polysaccharides is 30 to 50KD and 80 to 100KD; the mass ratio of the cationic angelica polysaccharide to the DNA plasmids is (1-200):1; and the particle diameter of the cationic angelica polysaccharide-DNA plasmid nancomposite is 21 to 77nm. The system has the characteristics that: 1, the angelica polysaccharide has various biological activities such as immune regulation activity, anti-aging activity, anticoagulation activity and the like, is safe and biologically degradable, does not have immunogenicity and isprepared with a simple, economic and convenient process; and 2, all the three cationic angelica polysaccharides have good DNA plasmid combination effect and gene delivery expression effect. Positive charges carried by primary amine, secondary amine and tertiary amine groups combined with saccharide chains can be effectively combined with the DNA plasmids with negative charges through an electrostatic effect, so that the plasmids are protected from being degraded by various enzymes inside and outside cells.

Description

technical field [0001] The invention relates to an angelica polysaccharide and a gene transfer system, in particular to a cationic angelica polysaccharide nanoparticle gene transfer system. Background technique [0002] Gene therapy has opened up its broad application prospects in the field of biomedicine, and can be used to treat genetic and acquired diseases such as hemophilia, cystic fibrosis, gynecological diseases, etc. [See: Jay Lozier. Gene therapy of the hemophilias. Seminars in Hematology, 2004, 41 (4): 287~296. Uta Griesenbach, A. Christopher Boyd. Pre-clinical and clinical endpoint assays for cystic fibrosis gene therapy. Journal of Cystic Fibrosis, 2005, 4 (2): 89~ 100. Memy H, Hassan, Essam E, Othman, Daniela Hornung, Ayman A1-Hendy. Gene therapy of benign gynecological diseases. Advanced Drug Delivery Reviews, 2009, 61(10):822~835.]. The key to gene therapy technology is to be able to effectively deliver foreign genes into the nucleus and make them express eff...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/63B82Y5/00
Inventor 徐希明王淼余江南邓纹纹曹霞
Owner JIANGSU UNIV
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