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Novel coronavirus (sars-cov-2) spike protein binding molecule and application thereof

A SARS-COV-2 and spike protein technology, applied in antiviral immunoglobulin, application, antiviral agents, etc., can solve the problems that it is difficult to become a specific drug for SARS-COV-2, and the treatment effect is not good.

Active Publication Date: 2022-03-18
SHENZHEN IMMUNOTHERAPY BIOTECH CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] Aiming at the problem that the existing antiviral drugs have no specificity to the novel coronavirus, the therapeutic effect is not good, and it is difficult to become a specific drug for SARS-COV-2, the present invention provides a novel coronavirus (SARS-COV-2) Spike protein binding molecules and applications thereof

Method used

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  • Novel coronavirus (sars-cov-2) spike protein binding molecule and application thereof
  • Novel coronavirus (sars-cov-2) spike protein binding molecule and application thereof
  • Novel coronavirus (sars-cov-2) spike protein binding molecule and application thereof

Examples

Experimental program
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Effect test

Embodiment 1

[0075] Screening of single domain antibodies against SARS-COV-2-Spike protein

[0076] 1.1 Library Construction

[0077] 1.1.1 Immunity

[0078] The alpaca was immunized with the Spike-RBD protein of the new coronavirus, and immunized at the 1st, 2nd, 4th, and 6th week, respectively, for a total of 4 times, and each immunization dose was 300ug.

[0079] 1.1.2 Extraction of total RNA

[0080] Get 50ml of peripheral blood of alpaca after the 6th week of immunization, separate lymphocytes, use Trizol to extract the total RNA of lymphocytes, and adopt the ultraviolet spectrophotometer to detect the results of the extracted RNA: OD260 / 280=1.97, OD260 / 230= 2.14, indicating that the extracted RNA is not significantly degraded, and the purity is good; the total RNA concentration is 937.5ng / μL. The extracted total RNA was used for agarose gel electrophoresis, and the results were as follows: figure 1 As shown, two bands of 28S and 18S can be seen.

[0081] 1.1.3 RNA reverse transc...

Embodiment 2

[0142] 1.1 Preparation of Fc fusion protein of SARS-COV-2-Spike protein single domain antibody

[0143] According to the amino acid sequence of the constant region of human immunoglobulin (IgG1) on the protein database Uniprot, the amino acid sequence of the human IgG1-Fc region (SEQ ID NO: 85) was obtained. By reverse transcription PCR, the nucleic acid fragment encoding human IgG1-Fc (nucleic acid sequence such as SEQ ID NO: 107) was obtained from human PBMC total RNA, and then the SARS-COV-2-Spike protein single domain antibody and Fc were obtained by overlapping PCR The nucleic acid fragment encoding the fusion protein was recombined into the vector pCDNA4 (Invitrogen, Cat V86220).

[0144] The successfully constructed pCDNA4 plasmid containing the nucleic acid fragment of the fusion protein of SARS-COV-2-Spike protein single domain antibody and Fc was transfected into HEK293 cells for expression. Specifically, the recombinant expression plasmid was diluted with Freestyle...

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Abstract

The invention relates to the technical field of biomedicine, and specifically discloses a novel coronavirus (SARS‑COV‑2) spike protein binding molecule and an application thereof. The binding molecule can specifically bind the spike protein of SARS-COV-2 and comprises at least one immunoglobulin single variable domain. The SARS-COV-2-Spike protein binding molecule provided by the present invention can specifically bind the SARS-COV-2-Spike protein, and effectively block the binding of the SARS-COV-2-Spike protein to the ACE2 receptor of human cells, and then Block the infection process of SARS‑COV‑2 to cells, inhibit the infection and amplification of SARS‑COV‑2, play a long-term inhibitory effect on SARS‑COV‑2 in vivo, and effectively prevent the recurrence of SARS‑COV‑2 in vivo .

Description

technical field [0001] The invention relates to the technical field of biomedicine, in particular to a novel coronavirus (SARS-COV-2) spike protein binding molecule and an application thereof. Background technique [0002] At present, there is no specific drug for COVID-19, and it is urgent to develop effective drugs quickly. Although broad-spectrum small-molecule antiviral drugs such as remdesivir and favipiravir have been discovered to have certain curative effects on COVID-19, they are not specific for SARS-COV-2, and the therapeutic effect is limited, so it is difficult to become a COVID-19 patient. -19 specific medicine. Contents of the invention [0003] Aiming at the problem that the existing antiviral drugs have no specificity to the novel coronavirus, the therapeutic effect is not good, and it is difficult to become a specific drug for SARS-COV-2, the present invention provides a novel coronavirus (SARS-COV-2) Spike protein binding molecules and applications the...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K16/10C12N15/13C12P21/02A61K39/42A61K47/68A61P31/14G01N33/569
CPCC07K16/10A61K47/6801A61P31/14G01N33/56983C07K2317/22C07K2317/569C07K2317/565C07K2317/76C07K2317/92A61K2039/505G01N2333/165Y02A50/30
Inventor 张军方
Owner SHENZHEN IMMUNOTHERAPY BIOTECH CO LTD