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Tumor-targeted multifunctional non-viral gene vector and preparation method and application thereof

A gene carrier and tumor targeting technology, applied in the field of medicine, can solve the problems of not fully meeting the clinical needs of gene therapy, unsatisfactory transfection effect, and low transfection efficiency

Active Publication Date: 2021-08-31
WEST CHINA HOSPITAL SICHUAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the usual transfection efficiency of the modified PEI-anionic polymer complex is low, about 40%, and its transfection effect is always unsatisfactory, which cannot fully meet the clinical needs of gene therapy

Method used

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  • Tumor-targeted multifunctional non-viral gene vector and preparation method and application thereof
  • Tumor-targeted multifunctional non-viral gene vector and preparation method and application thereof
  • Tumor-targeted multifunctional non-viral gene vector and preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0086] Embodiment 1, the cationic core of the non-viral gene carrier (HPTHPU nanoparticle) of the present invention: preparation of heparin-PEI-TAT (HPT) nanoparticle

[0087] (1) The synthesis method of PEI-TAT is as follows: first, PEI (500 mg, 0.275 mol, 1.0 eq) with a molecular weight of 1800 was dissolved in 10 mL of chloroform, and then 3-maleimidopropionic acid hydroxysuccinimide The ester (88.5mg, 0.333mmol, 1.2eq) was added to the above solution, and after reacting at room temperature for 3 hours, the reaction solvent, chloroform, was removed by rotary evaporation. The remaining residue was fully dissolved with a mixed solution of 10 mL tetrahydrofuran (THF) and water (the volume ratio of THF and water was 1:1), and then the penetrating peptide Cys-TAT (457 mg, 0.275 mmol, 1.0 eq) was added to the above solution The reaction was continued at room temperature for 6 hours. After the reaction was finished, the reaction solution was dialyzed in pure water for 3 days with...

Embodiment 2

[0092] Embodiment 2, the anionic shell of the non-viral gene carrier (HPTHPU nanoparticle) of the present invention: the preparation of HA-PEG-uPA (HPU)

[0093] Hyaluronic acid (HA, 100mg), EDCI (14.2mg) and NHS (8.5mg) were dissolved together in 30mL MES buffer (pH 5.5, 0.05M), and reacted at room temperature for 2h to fully activate the carboxyl groups on HA. Then Mal-PEG-NH 2 (weight-average molecular weight: 2000, 124 mg) was added to the above reaction solution and continued to react for 24 hours at room temperature. Subsequently, the reaction solution was transferred to a dialysis bag with a molecular weight cut-off of 3500, and then dialyzed in pure water for 3 days to remove unreacted impurities. After dialysis, the liquid in the dialysis bag was reacted with cys-uPA (7.44mg, 0.0124069mmol) in 10mL of THF and water mixed solution (the volume ratio of THF and water was 1:1) for 48h. Finally, the above reaction solution was dialyzed in pure water for 3 days with a dia...

Embodiment 3

[0095] Embodiment 3, the preparation method (not carrying plasmid DNA) when the non-viral gene carrier (HPTHPU nanoparticle) of the present invention is used

[0096] The HPU powder prepared in Example 2 was dissolved in water to obtain an HPU aqueous solution. Then, the solution containing HPT nanoparticles prepared in Example 1 and the HPU aqueous solution were fully mixed according to the mass ratio of HPU powder and HPT nanoparticles at 3:1, and then incubated at room temperature for 25 minutes to obtain HPTHPU nanoparticles.

[0097] According to the preparation method described in Example 3, only by changing the mass ratio of HPT nanoparticles and HPU powder, HPTHPU nanoparticles with different mass ratios of HPT nanoparticles and HPU powder can be obtained. The mass ratio of HPU powder and HPT nanoparticles can also be 0.5:1, 1:1, 1.5:1, 2:1, 2.5:1, 3.5:1, 4:1.

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Abstract

The invention provides a tumor-targeted multifunctional non-viral gene vector and a preparation method and application thereof, and belongs to the field of medicine. The gene vector is prepared from a cationic core and an anionic shell through electrostatic interaction; the cationic core is prepared from heparin sodium, polyethyleneimine and polyethyleneimine-cell-penetrating peptide; the anionic shell is prepared from hyaluronic acid, maleimide polyethylene glycol amino and cysteine-labeled urokinase type plasmin activator; and the polyethyleneimine-cell-penetrating peptide is prepared from polyethyleneimine, 3-maleimide hydroxy succinimide propionate and cell-penetrating peptide. The gene vector is good in safety, capable of efficiently compounding genes, high in cell uptake rate, good in lysosome escape performance, capable of entering the deep part of tumor tissue for treatment and high in transfection efficiency and has good application prospects in the field of gene therapy.

Description

technical field [0001] The invention belongs to the field of medicine, and in particular relates to a tumor-targeting multifunctional non-viral gene carrier and its preparation method and application. Background technique [0002] Gene therapy has attracted more and more attention. It can deliver gene drugs to target cells or tissues, thus providing a highly effective and potential therapy for many diseases such as AIDS and cancer, which are poorly effective or even untreatable. treatment method. It is well known that naked DNA is not suitable for direct use in in vivo treatment because naked DNA is easily degraded and cleared by nucleases in blood or body fluids after entering the body. Therefore, there is an urgent need for a gene carrier that can not only protect the gene drug from degradation, but also efficiently deliver the gene drug to the target cells or tissues. [0003] Gene vectors widely used at present are mainly divided into viral vectors and non-viral vector...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K48/00A61K9/51A61K47/36A61K47/34A61K47/42A61K47/18A61P35/00
CPCA61K48/0041A61K9/5161A61K9/5146A61K9/5169A61P35/00
Inventor 巩长旸宋林江吴秦洁魏于全
Owner WEST CHINA HOSPITAL SICHUAN UNIV
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