Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Application of HOIP inhibitor in preparation of medicine for treating type II hereditary hemorrhagic telangiectasia

A technology for telangiectasia and drug treatment, which is applied in the field of biomedicine and can solve problems such as studies that have not yet been reported.

Active Publication Date: 2021-09-10
ACADEMY OF MILITARY MEDICAL SCI
View PDF1 Cites 2 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although the family screening found that the loss of linear ubiquitination is associated with multi-organ autoimmune deficiency and chronic autoinflammation and other diseases, there is no report on the regulation of linear ubiquitination of HHT2 and the application of HOIP inhibitors to treat HHT2

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Application of HOIP inhibitor in preparation of medicine for treating type II hereditary hemorrhagic telangiectasia
  • Application of HOIP inhibitor in preparation of medicine for treating type II hereditary hemorrhagic telangiectasia
  • Application of HOIP inhibitor in preparation of medicine for treating type II hereditary hemorrhagic telangiectasia

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] Example 1 demonstrates that ALK1 270-320aa (amino acids) domain specifically binds to HOIP

[0045] In order to confirm the specific domain of ALK1 binding to HOIP, HEK293T purchased from ATCC was preferred to overexpress the Flag-ALK1 series truncated body ( figure 2 ) and Myc-HOIP plasmid, and the interaction between the two was detected by immunoprecipitation technique.

[0046] The protein co-immunoprecipitation steps are:

[0047] 1. Collect the cells that have been transfected with the plasmid for 48 hours (you can choose culture flask or 6-well plate for transfection);

[0048] 2. Wash 3 times with pre-cooled PBS, centrifuge at 2000rpm for 3min;

[0049] 3. Add 500 μl of lysate (choose HEPES lysate or RIPA lysate according to specific experimental requirements), then choose to lyse on ice for 30 minutes or directly sonicate the cells, and then centrifuge at 12,000 rpm for 10 minutes to obtain the supernatant for later use;

[0050] 4. Take 50 μl of the supern...

Embodiment 2

[0068] Example 2 demonstrates the relationship between HHT2-related mutations and linear ubiquitination

[0069] To further explore the physiological significance of HOIP binding to ALK1, we combined the existing UniProt (https: / / www.uniprot.org / uniprot / P37023) database to screen 8 HHT2-related mutations (T277K, H280R, L285F , L289P, L294R, A306P, L313V, H314Y), using the same method as in Example 1 to overexpress the Flag-ALK1 series mutants and Myc-HOIP plasmids in HEK293T to detect the linear ubiquitination level ( image 3 ), six of the eight mutations (T277K, H280R, L285F, L289P, A306P, H314Y) were found to have significantly enhanced linear ubiquitination levels relative to wild-type ALK1 ( image 3 and Figure 4 ).

[0070] At the same time, it was found that the binding ability of these six mutants to HOIP was significantly enhanced by immunoprecipitation technique ( Figure 5 ). The ubiquitination experiment operation is as follows:

[0071] 1. Collect the cells ...

Embodiment 3

[0077] Example 3 Linear ubiquitination modification inhibits the kinase activity of ALK1

[0078] In order to verify the effect of these mutations on enhancing the ability to bind HOIP and increasing the ubiquitination level, we need to first figure out how the ubiquitination modification affects ALK1 itself. ALK1 is a classic serine-threonine kinase, and its kinase activity is the key to its function. The common detection of ALK1 kinase activity is to detect by constructing and purifying the constitutive activator of ALK1. Therefore, we found that the constitutive activator of ALK1 can phosphorylate the substrate Smad1 through in vitro phosphorylation experiments, but the constitutive activator modified by linear ubiquitination lost its activity. Therefore, the activity of ALK1 kinase after enhanced ubiquitination of mutants was proved by in vitro phosphorylation experiments. The in vitro phosphorylation experiment steps are as follows:

[0079] 1. HEK293T cells were transf...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention discloses application of an HOIP inhibitor in preparation of a medicine for treating type II hereditary hemorrhagic telangiectasia (HHT2). Firstly, a binding domain and a binding mechanism of ALK1 kinase and HOIP are disclosed, and it is found that binding enhancement of an ALK1 mutant and HOIP promotes obvious enhancement of the ubiquitination levels. By inhibiting the activity of HOIP ubiquitin ligase, the ubiquitination level of the ALK1 mutant is reduced, so that an ALK1-Smad1 / 5 signaling pathway is recovered, and symptoms of HHT2 are relieved. The invention proves that the pathogenesis of the ALK1 mutation-induced HHT2 is the reduction of the enzyme activity of ALK1 kinase, and a specific mutant of ALK1 can be inhibited through the HOIP inhibitor, so that the HOIP inhibitor can be used for treating HHT2, and the HOIP inhibitor becomes a potential medicine for treating the specific ALK1 mutation-induced HHT2.

Description

technical field [0001] The invention relates to the field of biomedicine, in particular to the application of a HOIP inhibitor in the preparation of drugs for treating type II human telangiectasia (HHT2). Background technique [0002] Hereditary hemorrhagic telangiectasia (HHT) is an autosomal dominant genetic disease with an incidence rate as high as 1 / 5000. Activin receptor-like kinase 1 (ALK1 Activin receptor-like kinase 1) is the most important pathogenic gene in type II human telangiectasia (HHT2). The ALK1 mutation carried by HHT2 patients often leads to the weakening of the ALK1-Smad1 / 5 signaling pathway in endothelial cells, which can easily induce arteriovenous malformations (AVMs). Capillaries are often found at the junction of arteries and veins, allowing the surrounding tissues surrounded by capillaries to obtain nutrients and oxygen. High-grade arteriovenous malformations result in a direct connection of arteries and veins, and the impact of blood flow at the ...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
IPC IPC(8): A61K45/06A61K31/277A61K31/4704A61K31/4184A61K31/548A61P9/14C12Q1/6883
CPCA61K45/06A61K31/277A61K31/4704A61K31/4184A61K31/548A61P9/14C12Q1/6883C12Q2600/156A61K2300/00
Inventor 张令强付业胜刘翠华崔春萍
Owner ACADEMY OF MILITARY MEDICAL SCI
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com