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Recombinant SARS-CoV-2 vaccine using human replication-defective adenovirus as vector

A technology of coronavirus and recombinant adenovirus, which is applied in the direction of virus/bacteriophage, virus, viral peptide, etc., to achieve the effect of fast and simple preparation method, good immune protection effect, and good immunogenicity

Active Publication Date: 2020-06-02
ACADEMY OF MILITARY MEDICAL SCI +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

The types of new coronavirus vaccines under research include adenovirus vector vaccines, mRNA vaccines, DNA vaccines, recombinant protein vaccines and inactivated vaccines, and the results of the research have not yet been reported

Method used

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  • Recombinant SARS-CoV-2 vaccine using human replication-defective adenovirus as vector
  • Recombinant SARS-CoV-2 vaccine using human replication-defective adenovirus as vector
  • Recombinant SARS-CoV-2 vaccine using human replication-defective adenovirus as vector

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] Example 1. Preparation of recombinant novel coronavirus vaccine with human replication defective adenovirus as carrier

[0044] 1. S protein gene optimization and synthesis

[0045] The target antigen of the recombinant novel coronavirus vaccine is the S protein of the novel coronavirus strain (Genebank number: NC_045512.2). By optimizing the S protein gene, the expression level of the S protein is increased, thereby improving the immunogenicity of the vaccine.

[0046] First, use the Upgene software (Gao, W.Rzewski, A.Sun, H.Robbins, P.D. & Gambotto, A. UpGene: Application of a web-based DNA codon optimization algorithm. Biotechnol Prog, 2004.20 (2): p.443-8 .) Perform codon optimization to change most of the rare codons in the S protein gene to high frequency codons. Secondly, considering that software optimization may mechanically change codons to the most frequently used codons, protein translation efficiency may not be significantly improved due to the influence ...

Embodiment 2

[0109] Example 2. Immunological evaluation of different constructed recombinant adenoviruses on mouse models

[0110] 1. Vaccine humoral immune response detection

[0111] 100 SPF grade female BALB / c mice (6-8 weeks old) were randomly divided into 10 groups, 10 mice in each group. The mice were immunized with Ad5-nCoV according to the grouping conditions shown in Table 1. The way of intramuscular injection is to inject 100 μL into the inner side of the hind thigh, and the way of nasal drop immunization is to anesthetize the mice with isoflurane and instill 20 μL through the nasal cavity. The grouping situation is shown in Table 1.

[0112] Table 1. Grouping of mice for vaccine humoral immune response detection

[0113]

[0114]

[0115] Blood was collected from the mice at a specific time point after immunization, the serum was separated, and the IgG antibody titer against the new coronavirus S protein in the serum was detected by ELISA. Test results such as Figure...

Embodiment 3

[0129] Example 3. Immunological evaluation of Ad5-nCoV on guinea pig model

[0130] Fifty-six guinea pigs of SPF grade, weighing 200 to 250 grams, were randomly divided into 4 groups, 14 in each group, half male and half male. Guinea pigs were immunized with Ad5-nCoV according to the grouping conditions shown in Table 3. The way of immunization was intramuscular injection of 200 μL in the rear thigh.

[0131] Table 3. Grouping of Ad5-nCoV guinea pig immunogenicity detection

[0132]

[0133] Blood was collected from the guinea pigs at a specific time point after immunization, the serum was separated, and the IgG antibody titer against the novel coronavirus S protein in the serum was detected by ELISA. Test results such as Figure 14 Shown (ns, P≥0.05; *, P<0.05; **, P<0.01; ***, P<0.001; ****, P<0.0001). The results showed that 14 days after Ad5-nCoV immunization in guinea pigs, high levels of serum IgG antibody titers were detected. There was no significant difference...

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Abstract

The invention provides a SARS-CoV-2 vaccine using human type-5 replication-defective adenovirus as a vector. The vaccine uses E1 and E3 to be combined with replication-defective human type-5 adenovirus as the vector and HEK293 cells integrating adenovirus E1 gene as a packaging cell line, and a protective antigen gene carried is the 2019 SARS-CoV-2 S protein gene (Ad5-nCoV) which is subjected to optimization design. After the S protein gene is optimized, the expression level in transfected cells is increased significantly. The vaccine has good immunogenicity in mouse and guinea pig models, andcan induce a body to produce a strong cellular and humoral immune response in a short time. Studies on the protective effect of hACE2 transgenic mice show that after 14 days of single immunization ofAd5-nCoV, the viral load in lung tissue can be significantly reduced, and it is indicated that the vaccine has a good immunoprotective effect on the 2019 SARS-CoV-2. In addition, the vaccine is quick, simple and convenient to prepare, and can be mass-produced in a short period of time to respond to sudden outbreaks.

Description

technical field [0001] The invention relates to a recombinant novel coronavirus vaccine, aimed at preventing the epidemic of novel coronavirus. The invention belongs to the technical field of bioengineering. Background technique [0002] The 2019 novel coronavirus, SARS‑CoV‑2 (also known as 2019‑nCoV), was named by the World Health Organization on January 12, 2020. It is a new strain of coronavirus that has never been found in humans before. This virus is the seventh coronavirus (CoV) that can infect humans. The incubation period of human infection with 2019-nCoV is generally 1 to 14 days. Common signs after infection with 2019-nCoV include respiratory symptoms, fever, cough, shortness of breath, and dyspnea. In more severe cases, infection can lead to pneumonia, severe acute respiratory syndrome, kidney failure, and even death. As of March 17, 2020, my country has reported 81,116 confirmed cases and 3,231 deaths; other countries have reported 98,486 cases and a total of...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/50C07K14/165C12N15/861A61K39/215A61P31/14
CPCC07K14/005C12N15/86A61K39/12A61P31/14C12N2770/20022C12N2710/10043C12N2770/20034A61K2039/575A61K2039/543A61K2039/545A61K2039/57A61K2039/572A61K39/215
Inventor 陈薇吴诗坡侯利华张哲王步森郭强张金龙宋小红付玲张军陈旖赵拯浩朱涛李荩莘春林
Owner ACADEMY OF MILITARY MEDICAL SCI
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