Application of intervening BOK in preparation of medicine for treating COVID-19

A drug and pneumonia technology, which is applied in the application field of intervening BOK in the preparation of drugs for the treatment of new coronary pneumonia, can solve the problems that there are no specific drugs, etc., and achieve the effect of reducing the infiltration of alveolar cavity fluid and reducing lung damage

Inactive Publication Date: 2021-10-22
THE FIFTH AFFILIATED HOSPITAL SUN YAT SEN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

No specific drug exists for COVID-19, although several antiviral drugs are being actively tested, including the nucleotide analog remdesivir

Method used

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  • Application of intervening BOK in preparation of medicine for treating COVID-19
  • Application of intervening BOK in preparation of medicine for treating COVID-19
  • Application of intervening BOK in preparation of medicine for treating COVID-19

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] Example 1 Detection of Apoptosis After NCl-H292 and Ea.hy926 Transient Plasmids Expressing SARS-CoV-2 Membrane Protein

[0029] 1. Detection object:

[0030] NCl-H292 and Ea.hy926: Transient empty plasmid and plasmid expressing membrane protein, respectively.

[0031] 2. The detection method specifically includes the following steps:

[0032] (1) 1×10^5 cells per well were plated overnight, and the next day, using the transfection reagent lipo2000, 1.5 μg plasmid (Vector represents empty expression plasmid, M-Flag represents SARS-CoV-2 expression with Flag tag) Membrane protein) was transferred into the cells, and the medium was changed after 6 hours.

[0033] (2) Trypsinize for 48 hours, centrifuge at 1000 rpm for 5 minutes, and wash the cells once with 1×PBS.

[0034] (3) Cells were resuspended in apoptosis staining buffer, stained with Annexin-V / PI (Annexin-V-FITC, PI-PE, stained at room temperature for 25 minutes, tested on the machine, apoptosis was measured by ...

Embodiment 2

[0037] Example 2 Knockout and recovery of BOK affect the ability of SARS-CoV-2 membrane protein to induce apoptosis

[0038] 1. Detection of cell objects:

[0039] BOK knockout H292 cell lines (KO1 and KO2), on the basis of the BOK knockout cell line H292, use lentivirus to restore BOK expression cell lines (KO1+BOK+GFP and KO2+BOK+GFP), KO+ Vector+GFP cell line represents a cell line stably transfected with lentivirus empty load.

[0040] 2. The detection method specifically includes the following steps:

[0041] (1) 1×10^5 cells per well were plated overnight, and the next day, 1.5 μg of the corresponding plasmid (Vector represents empty expression plasmid, M-Flag represents SARS-CoV- 2 membrane protein) into the cells, and change the medium after 6 hours.

[0042] (2) Trypsinization for 48 hours, centrifugation at 1000rpm for 5 minutes, lysing the cells with protein lysate, and detecting the expression of apoptosis-related proteins by Western blot. .

[0043] 3. Experime...

Embodiment 3

[0045] Example 3 Effect of SARS-CoV-2 Membrane Protein Expression on Mouse Lung Tissue

[0046] 1. Detection object:

[0047] Transduction of mouse lung tissue with lentivirus expressing SARS-CoV-2 membrane protein.

[0048] 2. The detection method specifically includes the following steps:

[0049] (1) Preparation of lentivirus expressing SARS-CoV-2 membrane protein: connect the membrane protein sequence of SARS-CoV-2 to the pLVX destination vector, sequence and verify that the connection is successful, and transfer competent amplification of the destination plasmid and helper plasmid (pMD2G and psPAX2). According to the target plasmid: pMD2G: psPAX2 = 10:7:5, use PEI transfection reagent to transfect HEK293T cell packaging virus, change the medium for 8 hours, collect the supernatant of virus particles for 48 hours, collect the virus particles by ultra-high speed centrifugation, resuspend in PBS and prepare for tracheal injection contents;

[0050] (2) The mice were rand...

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Abstract

The invention belongs to the technical field of biological medicines, and discloses application of intervening BOK in preparation of a medicine for treating COVID-19. In order to search a new immunotherapy scheme for comprehensive treatment of COVID-19, the lung tissue of patients with COVID-19 is researched, and a research result prove that the expression quantity of BOK in the lung tissue cells of the patients with COVID-19 is increased, and BOK gene knockout has an effect on inhibiting new coronal virus membrane protein from inducing apoptosis of human lung cells and vascular endothelial cells; and the effect of inducing lung tissue apoptosis by the new crown virus membrane protein can be inhibited by injecting lentivirus into the trachea of a mouse to express shRNA aiming at BOK, and the knock-down of BOK indicates that BOK is expected to be applied to the comprehensive treatment of COVID-19. The BOK is made into a medicine for treating COVID-19 so as to relieve lung injury and alveolar cavity fluid infiltration of patients with COVID-19, and the medicine is suitable for protecting lung tissues and organs in the development process of COVID-19.

Description

technical field [0001] The present invention relates to the technical field of biomedicine, and more specifically, relates to the application of intervening BOK in the preparation of drugs for the treatment of new coronary pneumonia. Background technique [0002] Coronavirus disease 2019 (COVID-19) is a respiratory disease caused by acute respiratory syndrome coronavirus 2 (SARS-CoV-2), most patients with COVID-19 show mild to moderate symptoms, but about Fifteen percent progress to severe pneumonia, and about 5% eventually develop acute respiratory distress syndrome (ARDS), septic shock and / or multiorgan failure. Clinical treatment mainly includes symptomatic management and oxygen therapy, and provides mechanical ventilation for patients with respiratory failure. No specific drug exists for the treatment of COVID-19, although several antiviral drugs, including the nucleotide analog remdesivir, are being actively tested. In addition to vaccine development and approaches th...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K45/00A61K31/7088A61P11/00A61P31/14
CPCA61K45/00A61K31/7088A61P11/00A61P31/14
Inventor 黄曦杨扬王之影
Owner THE FIFTH AFFILIATED HOSPITAL SUN YAT SEN UNIV
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