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Targeting gene carrier as well as preparation method and applications thereof

A gene carrier and gene technology, applied in the field of bone-targeted gene carrier and its preparation, can solve the problems of low transfection efficiency and lack of targeting, and achieve the effect of low cost, prolonged time and good affinity

Active Publication Date: 2016-10-26
SHENZHEN INST OF ADVANCED TECH CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the transfection efficiency of liposome as a non-viral transgene carrier is generally low and lacks targeting. Therefore, how to improve the targeting and transfection efficiency of liposome non-viral transgene carrier has become a research focus

Method used

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  • Targeting gene carrier as well as preparation method and applications thereof
  • Targeting gene carrier as well as preparation method and applications thereof
  • Targeting gene carrier as well as preparation method and applications thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0080] A method for preparing a bone-targeted gene carrier, comprising the following steps:

[0081] (1) Preparation of phospholipid DSPE-PEG2000-Aln modified by sodium alendronate:

[0082] With 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide (EDC) and N-hydroxysuccinimide (NHS) as activators, 0.24 mmol of DSPE-PEG2000-COOH was Dissolve in MES buffer solution (0.05mol / L, pH=5.5) at room temperature, add EDC and NHS to the above solution and activate the carboxyl group for 2 hours under ice-water bath conditions, wherein, the mole of DSPE-PEG2000-COOH:EDC:NHS The ratio is 0.03:1.25:0.8;

[0083] Dissolve 0.5 mmol of sodium alendronate in an appropriate amount of MES buffer solution (0.05 mol / L, pH=5.5) in advance, then add it into the above-mentioned activated reaction solution, and carry out amidation condensation reaction under stirring conditions for 8 hours, and the The obtained reaction solution was dialyzed in deionized water for 48 hours with a dialysis bag with a molec...

Embodiment 2

[0095] A preparation method of a bone-targeted gene carrier, the difference from Example 1 is that the fluorescent marker NBD-F (4-fluoro-7-nitro-2,1,3-benzoxadioxane oxazole)-labeled dioleoylphosphatidylcholine DOPC (abbreviated as NBD-PC) to replace DOPE in Example 1, and other conditions were the same as Example 1.

[0096] In the bone-targeted gene carrier prepared in Example 2 of the present invention, the molar ratio of each material is DSPE-PEG2000-Aln:DOTAP:NBD-PC:Chol=0.02:2:0.5:0.5. The particle size of the bone targeting gene carrier is 50nm.

[0097] At the same time, in order to highlight the beneficial effects of the present invention, liposomes (abbreviated as "no targeting gene carrier") that have not been modified by alendronate will be used as a contrast, that is, phospholipids that have not been modified by sodium alendronate DSPE-PEG2000 replaces DSPE-PEG2000-Aln in Example 2, wherein the molar ratio of each material in the non-targeting gene carrier is DS...

Embodiment 3

[0099] A method for preparing a bone-targeted gene carrier, comprising the following steps:

[0100] (1) Preparation of phospholipid DSPE-PEG2000-Aln modified by sodium alendronate: same as Example 1;

[0101] (2) Preparation of bone targeting gene carrier:

[0102] DOTAP, DOPE, cholesterol Chol and the DSPE-PEG2000-Aln prepared above were dissolved in chloroform, and added to the flask, and mixed uniformly to obtain the first mixed solution with a total volume of 2mL, wherein the molar ratio of each material was DOTAP: DOPE:Chol:DSPE-PEG2000-Aln=0.02:2:1:1; remove the chloroform in the first mixed solution by rotary evaporation under vacuum condition at 40°C to form a uniform film on the inner wall of the flask, and then Place the above-mentioned flask in a vacuum drying oven to evacuate for 6h, and dry overnight under vacuum to obtain a thin film material (liposome thin layer);

[0103] (3) Hydrate the above-mentioned dry film material with a PBS buffer solution with a pH ...

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Abstract

The invention provides a bone targeting gene carrier. The bone targeting gene carrier is a lipidosome modified by alendronate sodium, wherein the lipidosome modified by alendronate sodium comprises cationic lipid, neutral auxiliary lipoid, cholesterol and phospholipid modified by alendronate sodium, the cationic lipid, the neutral auxiliary lipoid and the cholesterol form a phospholipid layer, the phospholipid modified by alendronate sodium is distributed in the phospholipid layer and forms a vesica structure with the phospholipid layer, and alendronate sodium is exposed out of the phospholipid layer. The bone targeting gene carrier has relatively high targeting property and transfection efficiency for the bone tissue, and can be efficiently expressed near the bone tissue by being loaded with a gene substance. The invention further provides a preparation method and applications of the bone targeting gene carrier.

Description

technical field [0001] The invention relates to the field of non-viral gene delivery systems, in particular to a bone-targeted gene carrier and its preparation method and application. Background technique [0002] Gene therapy is a method of transferring target genes into target cells through gene carriers, and achieves the purpose of treating diseases by adding, blocking, and correcting genes. Gene therapy offers a promising treatment for some major diseases. However, the lack of highly efficient and targeted gene carriers restricts the widespread clinical application of gene therapy. Undoubtedly become the key to the success of gene therapy. [0003] At present, gene vectors mainly include viral vectors and non-viral vectors. Non-viral vectors that have no immune response, low cost, and can be produced repeatedly in large quantities are widely used. Non-viral vectors mainly include liposomes, polyethyleneimine (PEI), chitosan, etc. These non-viral vectors are complexed...

Claims

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Application Information

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IPC IPC(8): C12N15/88A61K48/00
CPCA61K48/0041C12N15/88A61K9/127A61K48/00
Inventor 赵晓丽郑楚萍
Owner SHENZHEN INST OF ADVANCED TECH CHINESE ACAD OF SCI
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