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Multifunctional targeting gene vector as well as preparation method and application

A gene carrier and targeting technology, applied in the field of multifunctional targeted gene carrier and preparation, to achieve the effect of improving gene delivery effect, promoting cell migration and proliferation, and promoting nuclear internalization

Inactive Publication Date: 2018-08-10
TIANJIN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] At present, there is no report on the modification of poly(lactide-co-glycolide)-polyethyleneimine and its use by using multifunctional targeting polypeptide (REDV-G-TAT-G-NLS)

Method used

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  • Multifunctional targeting gene vector as well as preparation method and application
  • Multifunctional targeting gene vector as well as preparation method and application
  • Multifunctional targeting gene vector as well as preparation method and application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0059] A method for preparing a multifunctional targeting gene carrier, comprising the steps of:

[0060] (1) Put 1,8-octanediol, LA and GA into a dry polymerization tube, add stannous octoate as a catalyst, and seal; vacuumize the polymerization tube, fill it with nitrogen, repeat 8 times, and heat up to 140°C for reaction After 28 hours, the crude product of P(LA-co-GA) was obtained; the crude product of P(LA-co-GA) was dissolved in chloroform to obtain a solution, and the solution was added to n-hexane under stirring, a precipitate was formed, and the solid-liquid separation , repeated 2 times, the volume ratio of chloroform and n-hexane was 1:8; the obtained precipitate was dried to constant weight in a vacuum oven to obtain refined P(LA-co-GA); the 1,8 - The molar ratio of octanediol, LA, GA and stannous octoate is 5:140:43:1;

[0061] (2) Dissolve P(LA-co-GA), succinic anhydride, dimethylaminopyridine, and triethylamine in dry 1,4-dioxane at a molar ratio of 1:10:10 in...

Embodiment 2

[0066] A method for preparing a multifunctional targeting gene carrier, comprising the steps of:

[0067] (1) Put 1,8-octanediol, LA and GA into a dry polymerization tube, add stannous octoate as a catalyst, and seal; vacuumize the polymerization tube, fill it with nitrogen, repeat 12 times, and heat up to 150°C Reacted for 24 hours to obtain the crude product of P(LA-co-GA); the crude product of P(LA-co-GA) was dissolved in chloroform to obtain a solution, and the solution was added to n-hexane under stirring, and a precipitate was formed, solid-liquid Separation, repeated 4 times, the volume ratio of chloroform and n-hexane was 1:12; the obtained precipitate was dried to constant weight in a vacuum oven to obtain refined P(LA-co-GA);

[0068] The molar ratio of the 1,8-octanediol, LA, GA and stannous octoate is 5:560:132:2;

[0069] (2) Dissolve P(LA-co-GA), succinic anhydride, dimethylaminopyridine, and triethylamine in dry 1,4-dioxane at a molar ratio of 1:20:20 , reacte...

Embodiment 3

[0074] Preparation method of multifunctional targeting gene carrier

[0075] (1) with embodiment 1 step (1);

[0076] (2) with embodiment 1 step (2);

[0077] (3) According to the molar ratio of 1:15:15, dissolve P(LA-co-GA), EDC and NHS with carboxyl groups in DMSO, stir at room temperature for 1 hour, and add In the DMSO solution of PEI, the molar ratio of P(LA-co-GA) with carboxyl group to PEI was 1:5, stirred at room temperature for 20 hours, placed in a dialysis bag with a molecular weight cut-off of 14,000, and ultra-pure Water dialysis for 2 days, and the dialysate was freeze-dried to obtain P(LA-co-GA)-PEI 10 ;

[0078] (4) Under nitrogen protection, the P(LA-co-GA)-PEI 10 Dissolve in DMSO to obtain solution 2, add OPSS-PEG-NHS DMSO solution with a weight average molecular weight of 2000 dropwise into solution 2, react for 2 hours at room temperature and in the dark, add REDV-G-TAT- The DMSO solution of G-NLS polypeptide was reacted for 4 hours, and the product wa...

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Abstract

The invention discloses a multifunctional targeting gene vector as well as a preparation method and application. The multifunctional targeting gene vector is a polymer represented by P(LA-co-GA)-PEI-PEG-NLS-G-TAT-G-REDV; the multifunctional targeting gene vector disclosed by the invention has a membrane penetration function, a nuclear location function, and a targeting function on endothelial cells; cell extraction of a gene compound prepared from a multifunctional targeting polypeptide modified gene vector is improved through specific recognition of REDV peptide and an integrin receptor on the surfaces of the endothelial cells and the membrane penetration effect of TAT. The nuclear internalization of the gene is promoted through a mutual effect of a nuclear localization signal NLS and a nuclear membrane, so that a gene delivery effect is improved and the aims of improving the transfection efficiency and promoting cell migration and proliferation are realized. The problem of non-viralvectors at present is solved.

Description

technical field [0001] The invention belongs to the technical field of biological materials, and specifically relates to a multifunctional targeting gene carrier, a preparation method and an application. Background technique [0002] Gene therapy is a promising approach for the treatment of congenital and acquired cardiovascular diseases. It is to effectively deliver the therapeutic gene into the target cells through the carrier, and regulate the behavior of the cells through the expression of the genes, so as to achieve the purpose of treatment. The core issue of gene therapy for the treatment of cardiovascular diseases is how to improve the expression efficiency of therapeutic genes in endothelial cells, and gene carriers play an important role in the gene delivery system. In order to promote the transmission of gene complexes in endothelial cells and enhance the expression of target genes, studies have shown that Arg-Glu-Asp-Val (REDV) is a tetrapeptide derived from fibr...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K48/00A61K47/69A61K47/64
CPCA61K47/645A61K47/6907A61K48/0041
Inventor 冯亚凯李茜郝雪芳任相魁郭锦棠
Owner TIANJIN UNIV
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