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Application of intervening BOK in preparation of medicine for treating novel coronavirus pneumonia

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: 2021-06-04
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 novel coronavirus pneumonia
  • Application of intervening BOK in preparation of medicine for treating novel coronavirus pneumonia
  • Application of intervening BOK in preparation of medicine for treating novel coronavirus pneumonia

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, transfection reagent lipo2000 was used to transfect 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 m...

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 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. Experimenta...

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 novel coronavirus pneumonia. In order to find a new immunotherapy scheme for comprehensive treatment of novel coronavirus pneumonia, the research on the lung tissue of novel coronavirus pneumonia patients finds that the expression quantity of BOK in the lung tissue cell of the novel coronavirus pneumonia patients is increased, BOK gene knockout can inhibit the apoptosis of human lung cells and vascular endothelial cells induced by a novel coronavirus membrane protein, and BOK knockdown by injecting lentivirus into the trachea of a mouse to express shRNA (short hairpin Ribonucleic Acid) aiming at BOK can inhibit the apoptosis of the lung tissue induced by the novel coronavirus membrane protein, which indicates that BOK is expected to be applied to the comprehensive treatment of the novel coronavirus pneumonia and is prepared into the form of a medicine for treating the novel coronavirus pneumonia, which can relieve lung injury and alveolar cavity fluid infiltration of the novel coronavirus pneumonia patients, and is suitable for protecting lung tissues and organs in the novel coronavirus pneumonia process.

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
CPCA61K31/7088A61K45/00A61P11/00A61P31/14
Inventor 黄曦杨扬王之影
Owner THE FIFTH AFFILIATED HOSPITAL SUN YAT SEN UNIV
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