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Phenylboronic acid-containing modified polymer material and application thereof in intracellular delivery of proteins and polypeptides

A polymer material, a technology containing phenylboronic acid, applied in the intracellular delivery of proteins and small peptides, in the field of polymer materials, can solve the problem that polymer carriers cannot bind proteins and peptides well

Active Publication Date: 2020-10-13
EAST CHINA NORMAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] In order to solve the problem that polymer carriers in the prior art cannot bind proteins and polypeptides well, the present invention innovatively provides a class of polymer materials modified with phenylboronic acid as carriers for intracellular delivery of proteins, polypeptides and other molecules

Method used

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  • Phenylboronic acid-containing modified polymer material and application thereof in intracellular delivery of proteins and polypeptides
  • Phenylboronic acid-containing modified polymer material and application thereof in intracellular delivery of proteins and polypeptides
  • Phenylboronic acid-containing modified polymer material and application thereof in intracellular delivery of proteins and polypeptides

Examples

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Effect test

Embodiment 1

[0053] Example 1: Synthesis of protein and polypeptide intracellular delivery carriers P1-P6 based on polyamide-amine dendrimers and functional groups containing phenylboronic acid.

[0054] In a specific embodiment, the preparation method of the polymer material modified with phenylboronic acid is as follows: 4-bromomethylphenylboronic acid and the fifth generation polyamidoamine dendrimer are mixed according to different molar ratios (26: 1, 38:1, 96:1, 128:1) were added into the methanol solution, heated and reacted at 70°C for 24 hours, transferred to the dialysis bag, fully dialyzed in methanol and deionized water respectively, separated and purified to obtain the product, respectively Four kinds of phenylboronic acid-modified dendrimers (as shown in formula (9)) with different grafting ratios. according to 1 H NMR nuclear magnetic spectrum characterizes the connection efficiency of phenylboronic acid on the surface of polyamide-amine dendrimer, and calculates the averag...

Embodiment 2

[0062] Example 2: Comparison of protein intracellular delivery efficiency of dendrimers P1-P4 modified with phenylboronic acid

[0063] Using green fluorescein FITC-labeled bovine serum albumin (BSA-FITC) as a model protein, the delivery efficiency of phenylboronic acid-modified dendrimer was evaluated on HeLa cells by detecting intracellular fluorescence, and the optimal material was screened.

[0064] The specific method is as follows: inoculate HeLa cells into 48-well plates overnight, and start the protein delivery experiment when the density of HeLa cells reaches above 80%. After BSA-FITC was thoroughly mixed with P1, P2, P3 and P4 prepared in Example 1 of the present invention, 100 μL of serum-free medium was added, and vortexed to mix. After incubating at room temperature for 30 minutes, 150 μL of serum-free medium was added to the mixed solution. Remove the cell medium, wash twice with PBS, add the medium solution containing the protein complex, and incubate for 4 hou...

Embodiment 3

[0066] Example 3: Comparison of protein intracellular delivery efficiency of dendrimer materials P4-P6 modified with phenylboronic acid

[0067] Using green fluorescein FITC-labeled bovine serum albumin (BSA-FITC) as a model protein, the delivery efficiency of boronic acid compound-modified dendrimer was evaluated on HeLa cells by detecting intracellular fluorescence, and the optimal material was screened.

[0068] The specific method is as follows: inoculate HeLa cells into 48-well plates overnight, and start the protein delivery experiment when the density of HeLa cells reaches above 80%. After fully mixing BSA-FITC with P4, P5, and P6 respectively, add 100 μL of serum-free medium, shake and mix well. After incubating at room temperature for 30 minutes, 150 μL of serum-free medium was added to the mixed solution. Remove the cell medium, wash twice with PBS, add the medium solution containing the protein complex, and incubate for 4 hours in a 37°C incubator. Afterwards, the...

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Abstract

The invention provides an application of a phenylboronic acid-containing modified high polymer material in intracellular delivery of proteins and polypeptides, the phenylboronic acid-containing modified high polymer material is composed of a cationic polymer and a phenylboronic acid-containing functional group, and the phenylboronic acid-containing functional group is covalently connected to the cationic polymer; the cationic polymer comprises a polyamide-amine dendrimer, polypropyleneimine, polylysine and the like; the phenylboronic acid-containing modified high polymer material can be used as an intracellular delivery carrier for proteins and polypeptides, i.e., protein and polypeptide molecules are delivered from extracellular to intracellular carriers. The method for using the phenylboronic acid-modified polymer material as a protein and polypeptide intracellular delivery carrier can achieve high efficiency, is effective to proteins with different molecular weights and isoelectricpoints, can maintain the biological activity of the proteins, has low toxicity to cells, and has good biocompatibility.

Description

technical field [0001] The invention relates to the fields of biotechnology, macromolecule chemistry, cell biology and the like, in particular to a polymer material modified with phenylboronic acid and its application in intracellular delivery of proteins and small peptides. Background technique [0002] Intracellular protein delivery refers to the delivery of biomolecules such as exogenous proteins or polypeptides into the cytoplasm, so as to realize the regulation of cell biological functions and the treatment of diseases. Compared with gene transfection, the functional protein delivered into the cell can respond quickly without changing the genome of the cell, reducing the risk of off-target, and is easily degraded after the effect is completed, thereby avoiding long-term toxic side effects on the cell. However, because proteins or polypeptides are hydrophilic molecules with relatively large molecular weight and few surface charges, they cannot enter cells through the cel...

Claims

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Application Information

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IPC IPC(8): C08G73/02C08G69/48C12Q1/02A61K47/34
CPCC08G73/028C08G69/48G01N33/5008A61K47/34C12N2503/00
Inventor 程义云刘崇懿王辉
Owner EAST CHINA NORMAL UNIVERSITY
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