Target exosome and preparation method and application thereof, drug delivery system and drug

A delivery system and exosome technology, applied in drug combinations, antineoplastic drugs, pharmaceutical formulations, etc., can solve non-biogenic biocompatibility and safety defects, low transfection efficiency, and difficulty in large-scale preparation And other issues

Pending Publication Date: 2019-09-13
TSINGHUA BERKELEY SHENZHEN INST
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

These nano-drug delivery systems have alleviated the lack of anti-tumor metastasis preparations to a certain extent, but there are still some shortcomings: existing nano-preparations often use a variety of polymer materials, although these polymer materials are good drugs to some extent carrier, but its non-biological origin and defects in biocompatibility and safety have also become important factors that limit its entry into the clinic
Using this method to prepare targeting peptides has the problem of low transfection efficiency, making it difficult to achieve large-scale production

Method used

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  • Target exosome and preparation method and application thereof, drug delivery system and drug

Examples

Experimental program
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Effect test

Embodiment 1

[0040] A method for preparing targeted exosomes, comprising the following steps:

[0041] (1) React 0.5 mg of CD44 antibody with 10 μL of 100 nMTraut’s reagent and 10 μL of 5 mg / mL Cy5-NHS at room temperature for 1 hour, then pass through PD-10 column for purification;

[0042] (2) 0.2 mg of exosomes were reacted with 10 μL of 20 mM NHS-PEG-MAL reagent at room temperature for 1 h, and then purified by PD-10 column;

[0043] (3) Add 20 μL of the product of step (1) to the product of step (2), and react at room temperature for 1 h to obtain targeted exosomes.

Embodiment 2

[0045] Verification of targeted exosome preparation results

[0046] Drop the targeted exosome suspension prepared in Example 1 into a confocal culture dish (the exosomes are labeled with the green fluorescent dye GFP), and the targeting antibody in Example 1 has been coupled during preparation There is a red fluorescent dye Cy5. The fluorescence distribution of exosomes was observed by confocal microscopy. see results figure 1 . figure 1 It is a confocal microscope result map of targeted exosomes according to an embodiment of the present invention. Such as figure 1 as shown, figure 1 The upper middle row shows the fluorescence distribution of exosome aggregates in the exosome suspension, and the lower row shows the fluorescence distribution of small exosome aggregates and single exosomes in the exosome suspension; The first column shows the green fluorescence of exosomes, the second column shows the red fluorescence of the targeting antibody on the targeted exosomes, th...

Embodiment 3

[0049] A method for preparing targeted exosomes, comprising the following steps:

[0050] (1) React 0.5 mg of Her2 antibody with 10 μL of 100 nM Traut's reagent at room temperature for 1 hour and then purify it through a PD-10 column;

[0051] (2) 0.2 mg of exosomes were reacted with 10 μL of 20 mM NHS-PEG-MAL reagent at room temperature for 1 hour, and then purified by NAP-10 column;

[0052] (3) Add 10 μL of the product of step (1) to the product of step (2), and react at room temperature for 1 h to obtain targeted exosomes.

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Abstract

The invention provides a preparation method and application of a target exosome which can effectively improve the modifying efficiency of an exosome so as to realize large-scale preparation, a drug delivery system and a drug. The preparation method of the target exosome comprises the following steps S1, performing sulfhydrylation treatment on a target antibody so as to obtain a sulfhydrylation target antibody; and S2, enabling the sulfhydrylation target antibody to be connected with the exosome through a protein cross-linking agent. The target antibody is subjected to sulfhydrylation modification, so that an amino on the antibody is converted into a sulfydryl; and then the exosome and the antibody are in cross-linking combination so that the target exosome is obtained. Compared with a method of obtaining the target exosome through modification in a plasmid transfection manner in the prior art, the preparation method provided by the invention has the advantages that the problems that the transfection efficiency is low in the plasmid process and large-scale preparation and secretion of mother cells through genetic engineering plasmid transfection are difficult to perform can be solved, industrialization preparation of the target exosome can be realized, and the preparation method has important significance in research and application of the exosome as a delivery carrier and a drug carrier.

Description

technical field [0001] The present invention relates to the field of biotechnology, in particular to a targeted exosome, its preparation method, application, drug delivery system and drug. Background technique [0002] Tumor metastasis is an important cause of tumor death. Tumor metastasis refers to the infiltration of tumor cells from the primary site to surrounding tissues, and the spread to other tissues and organs of the body through the lymphatic system and blood system. This spread increases the mortality of patients. One of the reasons why tumor metastasis is clinically difficult to cure is that it is difficult for drugs to reach the metastatic site. According to the tumor-specific microenvironment and the difference between tumor tissue and normal tissue, researchers have designed a variety of nano-drug delivery systems to inhibit tumor metastasis. For example, loading P-3FaxNeu5Ac into PLGA nanoparticles, targeting sialooligosaccharide nano-drug delivery system, ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K47/46A61K47/42A61K45/00A61P35/04A61P35/00
CPCA61K47/46A61K47/42A61K45/00A61P35/04A61P35/00
Inventor 吴耀炯陈海燕
Owner TSINGHUA BERKELEY SHENZHEN INST
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