Application of PRMT5 specific inhibitor in prevention of coronavirus infection

A coronavirus and inhibitor technology, applied in the fields of biotechnology and biomedicine, can solve problems such as the reduction of vaccine defense capabilities

Active Publication Date: 2021-10-26
XUZHOU MEDICAL UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

For example, the Indian variant and has been reported to be able to increase the speed of transmission several times and reduce the defense ability of existing vaccines by many times

Method used

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  • Application of PRMT5 specific inhibitor in prevention of coronavirus infection
  • Application of PRMT5 specific inhibitor in prevention of coronavirus infection
  • Application of PRMT5 specific inhibitor in prevention of coronavirus infection

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0082] Example 1. PRMT5 mediates SDMA modification of ACE2 and promotes ACE2-RBD binding

[0083] HEK-293T cells overexpressing Flag-ACE2 and GFP-RBD were treated with DMSO, PRMT5 inhibitor GSK3326595 (100nM), and PRMT1 inhibitor AMI-1 (8.8μM), respectively. After 48 hours, co-immunoprecipitation experiments (Co- IP), to detect changes in the binding ability of ACE2 and RBD. The experimental results are shown in Figure 1A, only GSK3326595 can significantly inhibit the binding of ACE2 and RBD.

[0084] In HEK-293T cells overexpressing Flag-ACE2, Co-IP assay was used to detect the monomethylation modification (MMA) and symmetrical dimethylation modification (SDMA) of ACE2, as well as the binding of ACE2 and PRMT5. figure 1 B shows that ACE2 can bind to PRMT5, and ACE2 will be modified by MMA and SDMA methylation.

[0085] After HEK-293T cells overexpressing Flag-ACE2 were treated with DMSO or PRMT5 inhibitor GSK3326595 (100nM) for 48 hours, the changes of MMA and SDMA modific...

Embodiment 2

[0087] Example 2. Effects of ACE2 methylation sites and glycosylation sites on binding to RBD

[0088] LC-MS mass spectrometry was used to detect the arginine methylation modification of Flag-ACE2 protein. R644 and R671 are the arginine methylation modification sites identified by mass spectrometry, the results are as follows figure 2 A and 2B are shown.

[0089] The Flag-ACE2-R644K and Flag-ACE2-R671K point mutation plasmids were constructed, and the Flag-ACE2-WT / R644K / R671K plasmids were overexpressed in HEK-293T cells, respectively, and the SDMA of ACE2 and the binding of ACE2-PRMT5 were detected by IP-Flag-ACE2 change of circumstances. The result is as image 3 Shown in A: only the R671 site mutation significantly weakened the SDMA modification, indicating that R671 is the main methylation site for symmetric dimethylation (SDMA) in ACE2.

[0090] The GFP-RBD and Flag-ACE2-WT / R671K plasmids were co-transfected in HEK-293T cells, and the binding of Flag-ACE2 and GFP-R...

Embodiment 3

[0094] Example 3, Effect of GSK3326595 Concentration on the Binding of ACE2-RBD and Cell Proliferation

[0095] In BEAS lung epithelial cells and HK2 kidney normal epithelial cells, respectively, under the treatment of different concentrations of GSK3326595 (100nM, 50nM, 25nM, 10nM), the cell proliferation was detected by CCK-8 assay. The result is as Figure 4 As shown in A and 4B, when the concentration of GSK3326595 is 25 nM or lower than 10 nM, there is basically no effect on cell proliferation.

[0096] In HEK-293T cells overexpressing GFP-RBD and Flag-ACE2, treated with different concentrations of GSK3326595 (100nM, 50nM, 25nM, 10nM) for 48 hours, IP-GFP-RBD was used to detect ACE2-RBD binding. The result is as Figure 4 As shown in C, when the concentration of GSK3326595 is 25nM, it can also significantly inhibit the binding of ACE2-RBD.

[0097] The above experiments show that: when the concentration of GSK3326595 is 25nM, it basically has little effect on cell pr...

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Abstract

The invention relates to the field of biotechnology and biomedicine, in particular to application of a PRMT5 specific inhibitor in prevention of coronavirus infection. More specifically, the invention provides a gene editing reagent of a PRMT5 inhibitor and/or ACE2 protein and application of a pharmaceutical composition composed of the gene editing reagent in preparation of products for preventing coronavirus infection/treating diseases caused by coronavirus infection. Preferably, the coronavirus comprises a novel 2019 coronavirus.

Description

technical field [0001] The present invention relates to the fields of biotechnology and biomedicine, in particular to the application of PRMT5 specific inhibitors in the prevention of coronavirus infection. Background technique [0002] The SARS-CoV-2 virus mainly infects the human body by binding to human ACE2. Many studies have reported that SARS-CoV2 infects the human body by combining the receptor binding domain (RBD) of Spike1 (new crown spike protein) with ACE2. [0003] With the spread of the SARS-CoV2 epidemic, more and more mutant strains of the SARS-CoV2 virus have been reported. Examples include South African variants (Beta variants) and Indian variants (Delta variants). On the one hand, these mutant virus strains make the virus spread faster, and on the other hand, the protective ability of the vaccine is continuously reduced. For example, the Indian variant has been reported to be able to increase the speed of transmission several times and reduce the defense...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K45/00A61K45/06A61K31/506A61P31/14A61P11/00
CPCA61K45/00A61K45/06A61K31/506A61P31/14A61P11/00A61K2300/00
Inventor 李中伟白津郑骏年王雯雯
Owner XUZHOU MEDICAL UNIV
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