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Method for diagnosing and treating arteriosclerosis using blood flow change site-targeted nanovesicles

A technology of nanovesicles and arteriosclerosis, which is applied in the direction of peptides containing positioning/targeting motifs, nanomedicine, nanotechnology, etc., and can solve problems such as difficulties

Pending Publication Date: 2022-04-01
努迈斯有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Despite attempts to develop nanovesicle-based therapeutics for a variety of fatal diseases and injuries, including cardiovascular disease, kidney injury, liver disease, and neurological disease, the size was maintained at high yields during a long and difficult manufacturing process Uniformity and consistency of active ingredients remain a challenge

Method used

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  • Method for diagnosing and treating arteriosclerosis using blood flow change site-targeted nanovesicles
  • Method for diagnosing and treating arteriosclerosis using blood flow change site-targeted nanovesicles
  • Method for diagnosing and treating arteriosclerosis using blood flow change site-targeted nanovesicles

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preparation example Construction

[0069]Moreover, the present invention provides a method for preparing the above-mentioned nanovesicles, which comprises obtaining from stem cells transfected with a vector transfected with a coding sequence of a signal peptide-blood flow disturbance site-targeting peptide-transmembrane protein-green fluorescent protein Steps for displaying on the surface of nanovesicles targeting peptides at sites of blood flow disturbances.

[0070] In the present specification, "blood flow disorder" refers to abnormal and irregular blood flow due to the structural characteristics of blood vessels, and is an event of early arteriosclerosis leading to dysfunction of vascular endothelial cells.

[0071] In this specification, "nanovesicles (NV)" refers to artificially pulverizing cells by passing them through a filter to prepare by self-assembly, which should be understood as different from exosomes extracted from cells .

[0072] The present invention provides stem cell-derived nanovesicles f...

Embodiment 1

[0101] Example 1: Preparation and Characterization of PREY Peptide-Nanovesicles

[0102] Design and cloning of plasmids for PREY expression

[0103] The plasmid that expresses and localizes PREY on the outside of the cell membrane consists of the N-terminal on the outer side-promoter-signal peptide-PREY-V5-transmembrane protein-green fluorescent protein-C-terminus on the inner side. As the signal peptide, signal peptide F (BKU002587, Korean Human Gene Bank, Korea) or signal peptide R (BKU008396, Korean Human Gene Bank, Korea) was used. In the plasmid sequence, i) a signal peptide induces the localization of the PREY peptide to the outside of the cell membrane; and ii) the V5 tag and green fluorescent protein monitor the position and expression level of PREY. Expression vectors were cloned by using the NEB Gibson Assembly kit (New England Biolabs, MA) according to the manufacturer's instructions. Each type of signal peptide and transmembrane protein was separately amplified b...

experiment example 1

[0137] Experimental Example 1: Displaying RPEY on the outside of the membrane

[0138] In order to target nanovesicles to sites of blood flow disturbance, PREY as a targeting peptide is externally displayed by expressing a specially designed plasmid across the membrane of mesenchymal stem cells (PMSC: PREY-expressing human mesenchymal stem cells) . The plasmid was designed to express the signal peptide and the PREY ectodomain at the N-terminal, whereby the signal peptide induces localization from the mesenchymal stem cell membrane to the outer side of the PREY ectodomain and inactivates the induction signal by cleavage.

[0139] Also, the ability of 3 transmembrane proteins for improving the expression of PREY, ie, exosomal marker (CD86) and mesenchymal stem cell markers (CD105 and CD271), was tested. In this way, the ability of PREY to search and target sites of blood flow disturbance is maximized. After PREY is connected to the V5 tag, it is connected to the green fluoresc...

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Abstract

The present invention relates to a method for diagnosing and treating arteriosclerosis using a nanovesicle targeted at a blood flow change site, and more particularly, to: a stem cell-derived nanovesicle as an anti-arteriosclerosis diagnosis and treatment platform using a peptide capable of targeting at a blood flow disorder site; the nanovesicles provide strong anti-inflammatory and endothelial repair promoting effects similar to those of mesenchymal stem cells, and thus can be used as a novel diagnostic and therapeutic agent capable of preventing the onset of arteriosclerosis.

Description

technical field [0001] The invention relates to a method for diagnosing and treating arteriosclerosis by targeting nanovesicles at blood flow change sites. Background technique [0002] Atherosclerosis is the leading cause of death, but current diagnostic methods are unable to detect its early etiological signals associated with irreversible cascades. Existing prevention and treatment options aimed at lowering cholesterol levels, blood pressure, or plaque formation are commonly used, although substantial risk of disease progression remains. The occurrence of disturbed blood flow as an early arteriosclerotic event of stenosis at bifurcations, tortuous regions, or the periphery leads to dysfunction of vascular endothelial cells (EC). In normal blood flow, endothelial cells line up along the direction of blood flow and maintain anti-inflammatory and anti-thrombotic functions. On the contrary, atherosclerosis and blood flow disorder stimulate endothelial cell dysfunction while...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K38/17A61K35/28C07K7/08C12N15/62C12N15/85G01N33/68G01N33/554A61P9/10B82Y5/00
CPCA61K35/28A61P9/10C07K7/08G01N33/6893G01N2800/323C07K7/06C07K2319/03C12N5/0663C12N2501/65C12N2503/00C12N5/0607A61K38/00G01N33/554A61K38/177C07K2319/02
Inventor 成学俊尹正基
Owner 努迈斯有限公司
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