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Method for preparing non-viral gene vector of amino acid material

An amino acid, non-viral technology, applied in the field of non-viral gene therapy vectors, can solve the problems of no transfection efficiency and low toxicity, and achieve the effects of high gene transfection efficiency and high recovery rate.

Inactive Publication Date: 2008-07-23
ZHEJIANG UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Non-viral gene carriers have problems such as toxicity, biocompatibility, and degradability. At present, there is a synthetic material polyethyleneimine (PEI). Experiments have found that polyethyleneimine with a molecular weight of 22000-25000 has a very high transfection efficiency. However, there are problems of toxicity and degradability. Polyethyleneimine with small molecular weight (600, 1200, 2000) has low toxicity and is easy to metabolize in vivo, but has almost no transfection efficiency

Method used

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  • Method for preparing non-viral gene vector of amino acid material

Examples

Experimental program
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Effect test

Embodiment 1

[0029] Example 1: Take polyaspartic acid as an example.

[0030] 1) Preparation of polyaspartic acid-iminopropanol polymer

[0031] ● 10g of aspartic acid is catalyzed by 85% phosphoric acid, and dehydrated for 2h at 160-180℃ and reduced to 3mmHg under vacuum. Remove the vacuum pump and add N,N-dimethylformamide (DMF) while it is hot to fully dissolve the solids. Add the reaction solution dropwise to 500 ml of distilled water to get a white flake precipitate, filter, and wash with distilled water To neutrality, dry to obtain 6.8 g of polyamino acid.

[0032]●Take 3 g of polyaspartic acid, dissolve it in 20 ml of DMF, cool, add 7 ml of 3-aminopropanol dropwise under an ice bath, stir for 4 hours at room temperature, and add the reaction solution dropwise to 100 ml of n-propanol During the precipitation, the precipitate was washed with acetone and filtered with suction to obtain a solid. The solid was placed in a round-bottomed flask, pumped with a water pump for 3 hours, and then p...

Embodiment 2

[0038] Take glutamic acid as an example to synthesize polyglutamic acid, and the synthesis process is the same as in Example 1.

[0039] Take 3 g of polyglutamic acid, dissolve it in 18 ml of DMF, cool, add 5 ml of 3-aminoethanol dropwise under an ice bath, stir for 4 hours at room temperature, add the reaction solution dropwise to 100 ml of n-propanol, and precipitate Precipitate, wash with acetone, and filter with suction to obtain a solid. The solid was placed in a round-bottomed flask, pumped with a water pump for 3 hours, and then pumped with an oil pump to completely dry, to obtain a polyglutamic acid-iminoethanol polymer.

[0040] Take 0.5 g of polyglutamic acid-iminoethanol polymer and dissolve it in 6 ml of dimethyl sulfoxide (DMSO). Take 0.8 g of benzotriazole carbonate, dissolve it in 4 ml of DMSO, add it to the polymer solution under nitrogen protection and light-proof conditions, stir while adding, and react for 3 hours to obtain an activated polymer.

[0041] Take 1....

Embodiment 3

[0044] Take lysine as an example to synthesize polylysine, and the synthesis process is the same as in Example 1.

[0045] Take 3 grams of polylysine, dissolve it in 18 ml of DMF, cool, add 8 ml of 3-aminopentanol dropwise under an ice bath, stir for 4 hours at room temperature, and add the reaction solution dropwise to 100 ml of n-propanol. The precipitate was separated out, washed with acetone, and filtered with suction to obtain a solid. The solid was placed in a round bottom flask, pumped with a water pump for 3 hours, and then pumped with an oil pump to completely dry, to obtain a polylysine-iminopentanol polymer.

[0046] Take 0.5 g of polylysine-iminopentanol polymer and dissolve it in 6 ml of dimethyl sulfoxide (DMSO). Take 1.0 g of N-hydroxysuccinimide chloroformate, dissolve it in 5 ml of DMSO, add it to the polymer solution under nitrogen protection and dark conditions, stir while adding, and react for 3 hours to obtain an activated polymerization Things.

[0047] Take ...

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Abstract

The invention provides a modification method of polymer amino acid, in particular to a macromolecular compound of non-viral gene vector material, which is characterized in that the polymer amino acid is generated by polycondensation of amino acid monomer, the ring-opening of the polymer amino acid side group is processed by amino alcohol reagent, the structure modification is processed by the connection reagent, the polyethyleneimine (PEI) is joined by the activation of active hydroxy on the amino alcohol, and that the functional composite material of polymer amino acid-amino alcohol-polyethyleneimine is generated by dialysis and lyophilization. The modification method of polymer amino acid has the advantages of low mammalian toxicity, high gene transfection efficiency and performance of biodegradation.

Description

Technical field [0001] The invention relates to a class of non-viral gene therapy vectors, in particular to a class of modified amino acid materials used as non-viral gene therapy vectors. Background technique [0002] Polyamino acid materials have been widely studied as non-viral gene carrier materials due to their biodegradability and low toxicity. Among them are polylysine, polyaspartic acid, polyornithine, polyglutamic acid and so on. The polyamino acid material has a relatively clear molecular structure because it is obtained by the formation of peptide bonds from amino acid monomers through intermolecular dehydration. Polylysine has a positive charge under physiological conditions and can bind DNA through electrostatic attraction. Polyaspartic acid has an imide ring structure and can be ring-opened by alkaline reagents. Therefore, it can join various functional groups through ring-opening reaction, which has strong remodeling and adds new functions. [0003] Gene drug thera...

Claims

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Application Information

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IPC IPC(8): C12N15/64C08G73/10C08G81/00
Inventor 汤谷平陈丹周峻王青青余海
Owner ZHEJIANG UNIV
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