Fusion protein of SARS-CoV-2, and vaccine composition of fusion protein

A vaccine composition, fusion protein technology, applied in the directions of fusion polypeptides, microorganisms, hybrid peptides, etc., can solve the problems of short duration of immune response, multiple vaccinations, poor immunogenicity, etc. The effect of suppressing replication

Inactive Publication Date: 2020-09-15
GUANGZHOU UNIVERSITY OF CHINESE MEDICINE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the small molecular weight SARS-CoV-2RBD (~21KDa) has the disadvantages of poor immunogenicity, inability to induce cytotoxic T lymphocyte response, short duration of the induced immune response, and the need for multiple vaccinations, so it is not suitable to be prepared as a vaccine alone

Method used

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  • Fusion protein of SARS-CoV-2, and vaccine composition of fusion protein
  • Fusion protein of SARS-CoV-2, and vaccine composition of fusion protein
  • Fusion protein of SARS-CoV-2, and vaccine composition of fusion protein

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0032] Example 1: Preparation, identification and content determination of fusion protein RBD-mFc

[0033] According to NCBI (https: / / www.ncbi.nlm.nih.gov) the gene sequence (amino acid sequence such as SEQ ID No. 1), and the gene sequence of the Fc protein in mouse immunoglobulin IgG (the amino acid sequence is shown in SEQ ID No.2), and the fusion protein was prepared by genetic engineering.

[0034] (1) Cloning construction and identification of COVID-19 antigenic protein RBD and mFc fusion protein

[0035]Fully synthesize the nucleotide sequence of RBD-mFc (the nucleotide sequence is shown in SEQ ID No.4, the amino acid sequence is shown in SEQ ID No.3), and introduce specific BamHI and XhoI restriction enzyme cutting sites, the mammalian cell expression vector PCDNA3.1 and RBD-mFc gene, double digestion with BamHI and XhoI at the same time, after digestion, perform 10g / L agarose gel electrophoresis, and pass through the gel Recovery and purification kit to purify the di...

Embodiment 2

[0042] Embodiment 2: the preparation of subunit vaccine

[0043] (1) Dilute the RBD-mFc fusion protein prepared in Example 1 with PBS buffered saline solution, and mix thoroughly to obtain the vaccine RBD-mFc with a recombinant fusion protein content of 10 μg / mL.

[0044] (2) Dilute the RBD-mFc fusion protein prepared in Example 1 with PBS buffered saline solution, and add aluminum adjuvant to mix thoroughly to obtain a vaccine composition RBD-mFc / Al;

[0045] (3) Dilute the RBD-mFc fusion protein prepared in Example 1 with PBS buffered saline solution, add complete Freund's adjuvant, stir to make it a white emulsion, and obtain a recombinant fusion protein with a content of 10 μg / mL. Vaccine composition complete Freund's adjuvant RBD-mFc / FA (RBD-mFc / CFA); use PBS buffered saline solution to dilute the RBD-mFc fusion protein prepared in Example 1, add incomplete Freund's adjuvant, stir Make it into a white emulsion, that is, obtain a vaccine composition incomplete Freund's ...

Embodiment 3

[0046] Example 3: ELISA immunoassay

[0047] (1) Immunization of mice

[0048] Twenty C57BL / 6 mice aged 6-8 weeks were randomly divided into 4 groups, 5 mice in each group, and the prepared vaccine was injected on the 0th and 28th day with the first initial immunization and the second booster immunization scheme, respectively. Only 0.1mL per injection.

[0049] RBD-mFc group: intramuscular injection of the vaccine RBD-mFc prepared in Example 2 on days 0 and 28;

[0050] RBD-mFc / Al group: intramuscular injection of the vaccine composition RBD-mFc / Al prepared in Example 2 on days 0 and 28;

[0051] RBD-mFc / FA group: on the 0th day, the vaccine composition prepared in Example 2 was injected intramuscularly with complete Freund's adjuvant RBD-mFc / FA (RBD-mFc / CFA); on the 28th day, the vaccine composition prepared in Example 2 was injected intramuscularly Vaccine composition Incomplete Freund's adjuvant RBD-mFc / FA (RBD-mFc / IFA).

[0052] Control group: a blank control group, in...

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Abstract

The invention belongs to the technical field of biology, and particularly relates to a fusion protein of SARS-CoV-2, and a vaccine composition of the fusion protein. The fusion protein and the prepared vaccine composition containing the fusion protein overcome the defects of poor immunogenicity and the like of subunit vaccines, can induce the specific immune response aiming at SARS-CoV-2, and achieve the purposes of inhibiting the replication of SARS-CoV-2, inhibiting the spread of SARS-CoV-2 or preventing the settlement of SARS-CoV-2 common strains and variant strains in SARS-CoV-2 host bodies. The fusion protein and the vaccine composition containing the fusion protein can effectively prevent and/or treat novel coronavirus pneumonia (Corona Virus Disease 2019, COVID-19). The fusion protein can be recombined and expressed in large quantities by the aid of the genetic engineering technology, the consumed time is short, and the large-scale production can be facilitated.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a fusion protein of SARS-CoV-2 and a vaccine composition thereof. Background technique [0002] Novel coronavirus disease 2019 (COVID-19) is a viral infectious disease caused by severe acute respiratory syndrome coronavirus 2 (Severe Acute Respiratory Syndrome Coronavirus 2, SARS-CoV-2) in humans. Common signs after a person is infected with coronavirus include respiratory symptoms, fever, cough, shortness of breath, and dyspnea; severe cases can lead to pneumonia, severe acute respiratory syndrome, kidney failure, and even death. This emerging infectious disease is spreading rapidly around the world. As of April 2020, with more than 3 million cases and more than 200,000 deaths worldwide, COVID-19 has become a serious public health problem. Although SARS-CoV-2 seriously endangers human health, there is currently no specific drug for the treatment and prevention of COVID-...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K19/00A61K39/215A61K39/385A61P31/14
CPCC07K14/005A61K39/12A61K39/385A61P31/14C07K2319/30C12N2770/20022C12N2770/20034A61K2039/6056
Inventor 刘中秋廖国超柯昌文戚笑笑柯碧霞杨得盈吴鹏卢琳琳冯潜王彩艳李自波
Owner GUANGZHOU UNIVERSITY OF CHINESE MEDICINE
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