DC cell targeted nano SARS-CoV2 S protein polypeptide pool vaccine and preparation method thereof

A protein peptide, cell targeting technology, applied in the field of biomedicine, to achieve the effects of safe production technology, enhanced immune effect, and improved uptake and presentation efficiency

Pending Publication Date: 2021-07-30
DALIAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In the face of COVID-19, no effective treatment drugs have been developed yet. The key is to study safe and effective vaccines to prevent the spread of the disease

Method used

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  • DC cell targeted nano SARS-CoV2 S protein polypeptide pool vaccine and preparation method thereof
  • DC cell targeted nano SARS-CoV2 S protein polypeptide pool vaccine and preparation method thereof
  • DC cell targeted nano SARS-CoV2 S protein polypeptide pool vaccine and preparation method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] Example 1 Preparation of gelatin nanoparticles, mannose modified gelatin nanoparticles, nanoparticles encapsulated polypeptide

[0043] Weigh 0.5g gelatin into a 100mL beaker, add 10mL double distilled water, stir to dissolve at 50°C, add 10mL acetone while stirring, let stand at room temperature for 20min, discard the supernatant, then add 10mL double distilled water, 50°C thoroughly Dissolve the bottom precipitate, add 1mol / L HCl solution, adjust the pH to 2.5; add 30mL of acetone dropwise, then add 2mL of glutaraldehyde containing 0.5% diluted with acetone, keep stirring at room temperature 1000rpm for 5h, after volatilization of acetone and glutaraldehyde Dilute 10 times with double distilled water, use 40KD dialysis bags, dialyze in double distilled water for 24 hours to prepare gelatin nanoparticles, and store them statically at -4°C.

[0044] Take 20 mL of gelatin nanoparticles with a concentration of 1 mg / mL in a 50 mL beaker, add 0.68 g of mannose, add 2 mL of ...

Embodiment 2

[0049] Example 2 Prediction, Analysis, Screening and Synthesis of Polypeptide Epitopes

[0050] The key to researching peptide epitope vaccines is to find out the immunogenic epitope from the functional protein sequence of COVID-19 coronavirus, and use bioinformatics technology to predict the epitope of S protein.

[0051] After searching the amino acid sequence of the corresponding virus protein through NCBI, the HLA-A2 restricted CTL cell epitope was predicted, and the IEDB website and NetMHC 4.0Server website were used to predict the class I MHC molecular binding site. Cross-screen the obtained epitopes for epitopes with high affinity. Epitopes with %Rank<0.5 tend to form strong bonds, and epitopes with 0.5<%Rank <4 tend to form weak bonds; peptides with IC50 values<50nM are considered high affinity, <500nM It is an intermediate affinity, <5000 nM low affinity; then the screened epitope is screened for allergens and analyzed for physical and chemical properties, and finally...

Embodiment 3D

[0057] Encapsulation Efficiency and Release Rate Detection of Encapsulation Efficiency of Example 3DC Targeted Nanoparticles

[0058] Mix 1 mg / mL MnGNP with 10 μg / mL fluorescently labeled polypeptide, shake and wrap on a micro shaker at 4°C, centrifuge at 10,000 rpm for 10 min, detect the fluorescence intensity of the supernatant and precipitate, and calculate the encapsulation efficiency of nanoparticles. The result is as Figure 4 .

[0059] Encapsulation efficiency = polypeptide in precipitation / total amount of added polypeptide × 100%. The results showed that the maximum encapsulation efficiency of the polypeptide was 49%.

[0060] Release rate: Put the gelatin nanoparticles wrapped with polypeptides in a constant temperature shaker at 37°C, take them out at 0h, 3h, 6h, 9h, 12h, and 24h in sequence, centrifuge at 10,000rpm for 10min, and detect the fluorescence intensity of the precipitate and supernatant respectively After each test, the precipitate was resuspended wit...

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Abstract

The invention belongs to the field of biological medicines, and particularly relates to a DC cell targeted nano SARS-CoV2S protein polypeptide pool vaccine and a preparation method thereof. Nanoparticles are modified by mannose, so that the vaccine is specifically targeted to DC cells, the acquisition of SARS-CoV2S antigens by the DC cells is accelerated, the maturation of the DC cells is promoted, the antigens are efficiently presented to CTL cells, and the proliferation of the CTL cells is promoted; the killing effect of CTL cells is enhanced, so that the immune efficacy of the polypeptide vaccine is improved. According to the invention, polypeptide prediction analysis is carried out by utilizing a bioinformatics technology, the DC cells are efficiently activated by utilizing a nanoparticle targeting technology, the immune action of the SARS-CoV2S resistant cells is effectively activated, and a safer and more effective DC cell targeting nano SARS-CoV2S protein polypeptide pool vaccine is provided for prevention and control of COVID-19.

Description

technical field [0001] The invention belongs to the field of biomedicine, and in particular relates to a DC cell-targeted nano SARS-CoV2 S protein polypeptide pool vaccine and a preparation method thereof. Background technique [0002] At present, China is promoting the research and development of five technical routes in parallel, including inactivated vaccines, adenovirus vector vaccines, recombinant protein vaccines, nucleic acid vaccines, and attenuated influenza vaccines, and three vaccines have been conditionally used. Inactivated vaccines refer to the selection of highly immunogenic pathogens, which are inactivated by physical or chemical methods after a large number of artificial cultures, destroying the replication ability of the virus, making it lose pathogenicity but retain immunogenicity. Inactivated vaccines have the advantages of mature technical route, fast early research and development, inability to reproduce in vivo, and will not cause corresponding disease...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/215A61K9/51A61K47/42A61K47/54A61K47/69A61P31/14C07K14/165
CPCA61K39/12A61K9/5169A61K47/549A61K47/6931A61P31/14C07K14/005C12N2770/20022C12N2770/20034
Inventor 王钦富王拱辰黄晶曹钧雄王美辰李晨阳
Owner DALIAN UNIV
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