WO2020215351A1 - Immunopotentialisateur, son procédé de préparation, vaccin contre la grippe aviaire et utilisation de celui-ci - Google Patents

Immunopotentialisateur, son procédé de préparation, vaccin contre la grippe aviaire et utilisation de celui-ci Download PDF

Info

Publication number
WO2020215351A1
WO2020215351A1 PCT/CN2019/085076 CN2019085076W WO2020215351A1 WO 2020215351 A1 WO2020215351 A1 WO 2020215351A1 CN 2019085076 W CN2019085076 W CN 2019085076W WO 2020215351 A1 WO2020215351 A1 WO 2020215351A1
Authority
WO
WIPO (PCT)
Prior art keywords
pentapeptide
bursin
avian influenza
immune enhancer
vaccine
Prior art date
Application number
PCT/CN2019/085076
Other languages
English (en)
Chinese (zh)
Inventor
陈瑞爱
李延鹏
叶俊贤
董楠
杨小云
Original Assignee
肇庆大华农生物药品有限公司
华农（肇庆）生物产业技术研究院有限公司
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by 肇庆大华农生物药品有限公司, 华农（肇庆）生物产业技术研究院有限公司 filed Critical 肇庆大华农生物药品有限公司
Publication of WO2020215351A1 publication Critical patent/WO2020215351A1/fr

Links

Images

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K39/12—Viral antigens
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K39/39—Medicinal preparations containing antigens or antibodies characterised by the immunostimulating additives, e.g. chemical adjuvants
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • A61P31/12—Antivirals
    • A61P31/14—Antivirals for RNA viruses
    • A61P31/16—Antivirals for RNA viruses for influenza or rhinoviruses
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • A61P37/02—Immunomodulators
    • A61P37/04—Immunostimulants
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/555—Medicinal preparations containing antigens or antibodies characterised by a specific combination antigen/adjuvant
    • A61K2039/55511—Organic adjuvants
    • A61K2039/55516—Proteins; Peptides
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/555—Medicinal preparations containing antigens or antibodies characterised by a specific combination antigen/adjuvant
    • A61K2039/55511—Organic adjuvants
    • A61K2039/55583—Polysaccharides
    • C—CHEMISTRY; METALLURGY
    • C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
    • C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
    • C12N2760/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA ssRNA viruses negative-sense
    • C12N2760/00011—Details
    • C12N2760/16011—Orthomyxoviridae
    • C12N2760/16111—Influenzavirus A, i.e. influenza A virus
    • C12N2760/16134—Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
    • Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
    • Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Definitions

  • the present invention relates to the technical field of veterinary biological products, in particular to an immune enhancer, a preparation method, avian influenza vaccine and application.
  • Avian influenza is an important infectious disease caused by the avian influenza virus, which seriously harms the development of the chicken industry. Vaccine immunization is one of the most effective means to control the epidemic. However, with the promotion and use of vaccines, viruses continue to mutate under the pressure of vaccine selection, and their virulence has increased, which has brought challenges to the prevention and control of epidemics. To solve the above dilemma, clinically, the vaccine strains are constantly replaced. For the prevention and control of avian influenza, the replacement of vaccine strains can indeed have an immediate effect in a short period of time, but in the long run, this will also accelerate the rate of mutation of the avian influenza virus, and the prevention and treatment will be trapped in a cycle.
  • Immune enhancers can improve the function of the body's immune system and enhance the body's non-specific immunity to diseases.
  • the use of appropriate immune enhancers in combination with vaccines is one of the effective ways to enhance the immunity of existing vaccines. Improving vaccine immunity means that fewer vaccines can have better effects. Clinical use can reduce the number of immunizations and reduce stress, which not only saves costs, but also effectively controls diseases.
  • Bursalin is a small molecule peptide that exists in the bursa of poultry and is closely related to immune function.
  • Bursin pentapeptide BP5
  • BP5 Bursin pentapeptide
  • It can induce the differentiation and proliferation of poultry B lymphocyte precursors, increase antibody levels, and enhance humoral immunity; promote the transformation activity of T lymphocytes, thereby enhancing cellular immunity .
  • Two representative polypeptides in the bursin family are BP-14 and BP-5.
  • the immune enhancer BP14 is added to the livestock and poultry vaccine in different doses according to different target animals for joint vaccination; the antibody level of the immunized animals is higher than that of the vaccine group; the test animals produce higher levels of cytokines .
  • the dosage for poultry is 0.1 ⁇ g/feather, and for pigs is 5 ⁇ g/head; referring to Figure 1, BP-14+ vaccine (H5N1 subtype, Re-6 strain) is used, and the titer is 5-8log2.
  • BP-5 the right holder Li Deyuan applied for Chinese patent CN200810243575.0 in 2008 to disclose a bursal pentapeptide whose amino acid sequence is: CYS-LYS-ASP-VAL-TYR.
  • the bursa pentapeptide (BP5) of the invention has simple structure, small molecular weight, no chemical toxicity, no immunogenicity, and simple preparation. It can be extracted from the bursa of chicken or other poultry, or can be chemically synthesized. The cost is very low and can be prepared in large quantities.
  • Bursa pentapeptide can promote the proliferation of T lymphocytes and B lymphocytes, improve the level of humoral and cellular immunity of the body, and can also improve the body's ability to resist peroxidative stress. It is a kind of broad application prospect Drugs or preparations for immunomodulation, immunotherapy and anti-peroxidation. It can be used in the fields of basic research, clinical treatment, nutrition and health care, and cosmetics.
  • the patent records the potential use of bursa pentapeptide in immunotherapy and immune regulation, but does not mention its application in avian influenza vaccine.
  • the recombinant fusion peptide is a fusion of thymosin ⁇ 1 and bursa pentapeptide BP5 through a flexible Linker.
  • the present invention inserts the recombinant T ⁇ 1-BP5 fusion peptide gene into an expression vector, transforms Escherichia coli, and obtains a genetically engineered bacteria that efficiently expresses the recombinant T ⁇ 1-BP5 fusion peptide.
  • the recombinant T ⁇ 1-BP5 fusion peptide is prepared through liquid culture and purification.
  • the fusion peptide The thioredoxin was removed by enterokinase with a His tag at the N-terminal, and then purified by affinity chromatography to obtain a single recombinant T ⁇ 1-BP5 fusion peptide.
  • the recombinant T ⁇ 1-BP5 fusion peptide of the invention can be used as a novel polypeptide immune adjuvant used with vaccines, can effectively enhance the body's cellular immunity and humoral immunity, and has broad application prospects.
  • the invention is based on the bursa pentapeptide BP5, and the gene recombination expression with thymosin ⁇ 1 is carried out to achieve the purpose of immune enhancement.
  • the purpose of the present invention is to conduct a more in-depth study on BP5 and propose a new application formula for BP5 to improve the immune effect of the vaccine.
  • the purpose of the present invention is to provide an immune enhancer.
  • the present invention also discloses the preparation method of the immune enhancer, the avian influenza vaccine using the immune enhancer and the application of the avian influenza vaccine.
  • the immune enhancer can effectively increase the antibody titer expression level of vaccine immunity.
  • the immune enhancer is simple to prepare and has good industrial application prospects.
  • LB liquid medium Luria-Bertani liquid medium
  • IPTG isopropyl thiogalactoside, Isopropyl ⁇ -D-Thiogalactoside;
  • SDS-PAGE electrophoresis polyacrylamide gel electrophoresis
  • Ni column nickel column
  • Binding buffer nickel column purification protein buffer
  • Elution buffer elution buffer
  • pET-32a expression vector, fusion protein type prokaryotic high-efficiency expression vector, commercially available;
  • TE buffer (10mM Tris-HCl; 1mM EDTA): TE buffer, Tris-HCl concentration is 10mM, EDTA concentration is 1mM;
  • HindIII Restriction endonuclease, commercially available
  • T4 DNA Ligase T4 DNA ligase, commercially available
  • E.coli DH5 ⁇ Escherichia coli DH5 ⁇ ;
  • RosettaTM (DE3) Rosetta (DE3) E. coli expression strain
  • PBS Phosphate buffered saline solution.
  • an immune enhancer comprising bursin pentapeptide and astragalus polysaccharide; the weight ratio of the bursin pentapeptide and astragalus polysaccharide is 1:5 to 1:20.
  • the amino acid sequence of the bursin pentapeptide described herein is one or more tandem repeats of the amino acid sequence Cys-Lys-Arg-Val-Tyr, preferably 1-6 tandem repeats; In specific embodiments, five tandem repeats of the amino acid sequence Cys-Lys-Arg-Val-Tyr are used as the bursin pentapeptide used in the experiment.
  • the weight ratio of the bursin pentapeptide and astragalus polysaccharide is 1:15.
  • the present invention also discloses an avian influenza vaccine, which contains the above-mentioned immune enhancer; the dosage of the immune enhancer in the avian influenza vaccine is 5-15 ⁇ l/feather.
  • the antigen in the avian influenza vaccine is the H9N2 subtype avian influenza virus inactivated antigen or the H5N1 subtype avian influenza virus inactivated antigen.
  • the present invention also discloses a method for preparing the immune enhancer as described above, which is mixed and dissolved in water with cystin pentapeptide and astragalus polysaccharide.
  • Step 1 Construct the gene of bursin pentapeptide
  • Step 1 is specifically: taking 5'-TGCAAACGCGTGTAC-3' as the BP5 gene, adding EcoRI and HindIII restriction sites and protective bases before and after the fragment to obtain the fragment sequence F and the reverse complementary sequence R of the fragment sequence F.
  • the reverse complement sequence R and the fragment sequence F constitute the nucleic acid fragment of the gene of bursin pentapeptide, the fragment length of the fragment sequence F is 93 bp, and the nucleic acid fragment of the gene of bursin pentapeptide is synthesized by Shenggong Bioengineering (Shanghai) Co., Ltd.
  • Fragment sequence F (refer to SEQ ID NO: 1 in the sequence list for details):
  • the reverse complementary sequence R of the fragment sequence F (for details, please refer to the sequence list SEQ ID NO: 2):
  • the sequence of the EcoR I restriction site is as follows: GAATTC;
  • HindIII restriction site is shown as underlined: AAGCTT;
  • Step 2 Link the gene of bursin pentapeptide to the expression vector pET-32a to obtain the recombinant plasmid pET-32a-(BP5) 5 ;
  • Step 2 specifically: Take 1 ⁇ g of nucleic acid fragment and dissolve it in 50 ⁇ L TE buffer solution, the buffer solution contains 10mM Tris-HCl and 1mM EDTA, and use EcoRI and HindIII endonuclease to double digestion of gene fragments; meanwhile, use EcoRI and HindIII endonuclease for double digestion of pET-32a vector;
  • the ligation product was transformed into the standard E. coli strain E. coli DH5 ⁇ , the positive colonies were verified with vector sequencing primers, and the positive clones were sequenced; the recombinant plasmid pET-32a-(BP5) 5 with the correct sequence was stored.
  • Step 3 Introduce the recombinant plasmid pET-32a-(BP5) 5 into E. coli and induce expression to obtain an expression product;
  • Plasmid introduction Take 5 ⁇ L of recombinant plasmid pET-32a-(BP5) 5 and add it to 100 ⁇ L of Escherichia coli engineered bacteria RosettaTM (DE3), mix well on ice for 30 minutes, and put the mixture in a water bath at 42°C for 90 s. After taking it out, quickly stand still on ice Leave for 3 minutes to complete the plasmid transformation to obtain the seed bacterial solution;
  • Step 4 Purify and dry the expression product to obtain bursin pentapeptide
  • Product purification Pack the Ni column, add slowly to the medium supernatant after the Binding buffer is equilibrated; after all samples flow through the Ni column, rinse with an appropriate amount of Binding buffer to remove impurities, and then slowly add the Elution buffer, and collect at the peak Eluent; the eluate is renatured by concentration gradient dialysis to obtain soluble (BP5) 5 -thioredoxin with a size of about 22KDa; enterokinase is added to the purified product to remove thioredoxin to obtain (BP5) 5 fusion Peptide; Ni column chromatography and dialysis refolding of the above-mentioned product (BP5) 5 fusion peptide are performed again to obtain a high purity (BP5) 5 fusion peptide; the product freeze-drying process is the bursin pentapeptide used in the present invention.
  • Step 5 Mix the bursin pentapeptide and astragalus polysaccharide and dissolve it in water for injection.
  • amino acid sequence of the (BP5) 5 fusion peptide prepared by the above method is (for details, please refer to the sequence list SEQ ID NO: 3):
  • the repeating sequence is C-K-R-V-Y.
  • the present invention also discloses an application of the immune enhancer as described above, which is used as an immune enhancing adjuvant for avian influenza vaccine.
  • the invention adopts the combined use of cystatin pentapeptide and astragalus polysaccharide, which has a synergistic effect on the effect, and the compatibility with the avian influenza vaccine can obtain an ideal immune effect.
  • the avian influenza vaccine using the immune enhancer of the present invention has a 3 titer (log 2) higher than the avian influenza vaccine without the immune enhancer during the entire immune cycle. .
  • Figure 1 is a table of antibody levels of the H9N2 subtype avian influenza vaccine immunity test of the present invention
  • Figure 2 is a table of antibody levels of the H5N1 subtype avian influenza vaccine immunity test of the present invention
  • Figure 3 is a table of antibody levels of the H9N2 subtype avian influenza vaccine of the present invention with the addition of bursin pentapeptide immunoassay;
  • Figure 4 is a table of antibody levels of the H9N2 subtype avian influenza vaccine of the present invention added with Astragalus polysaccharides in the immune test;
  • Figure 5 is an SDS-PAGE electrophoresis diagram of the expression of recombinant (BP5) 5 fusion peptide in E. coli.
  • the components of 500 vaccines include:
  • the oil phase contains 84.6ml white oil (EXonMobil), 5.4ml Spencer-80 (Zhaoqing Chaoneng Industrial Co., Ltd.);
  • the water phase contains 52.6ml H9N2 embryo fluid inactivated antigen (10 7 EID 50 /0.1mL, Zhaoqing Dahuanong Biopharmaceutical Co., Ltd.), 2.4ml Tween-80 (Zhaoqing Super Energy Industrial Co., Ltd.), and 5ml immune enhancer.
  • the water phase contains 10 ⁇ l/feather of the immune enhancer, and its components are: bursin pentapeptide 1wt%; astragalus polysaccharide 15wt%; and the balance water.
  • the preparation method of the immune enhancer is:
  • Step 1 Construct the gene of bursin pentapeptide
  • the fragment length is 93 bp, which is synthesized by Bioengineering (Shanghai) Co., Ltd.
  • Step 2 Link the gene of bursin pentapeptide to the expression vector pET-32a to obtain the recombinant plasmid pET-32a-(BP5) 5 ;
  • Step 3 Introduce the recombinant plasmid pET-32a-(BP5) 5 into E. coli and induce expression to obtain an expression product;
  • Plasmid introduction Take 5 ⁇ L of recombinant plasmid pET-32a-(BP5) 5 and add it to 100 ⁇ L of Escherichia coli engineered bacteria RosettaTM (DE3), mix well on ice for 30 minutes, and put the mixture in a water bath at 42°C for 90 s. After taking it out, quickly stand still on ice Leave for 3 min to complete plasmid transformation.
  • DE3 Escherichia coli engineered bacteria RosettaTM
  • Step 4 Purify and dry the expression product to obtain bursin pentapeptide
  • Product purification Pack the Ni column, add the culture supernatant after the Binding buffer is equilibrated; after all the samples flow through the Ni column, rinse with an appropriate amount of Binding buffer to remove impurities, and then slowly add the Elution buffer, collect and elute at the peak
  • the eluate is renatured by concentration gradient dialysis to obtain soluble BP5-thioredoxin with a size of about 22KDa; enterokinase is added to the purified product to remove thioredoxin to obtain (BP5) 5 fusion peptide;
  • the product is subjected to Ni column chromatography and dialysis refolding to obtain a high-purity (BP5) 5 fusion peptide; the product freeze-drying process is the bursin pentapeptide used in the present invention.
  • Step 5 Mix the bursin pentapeptide and astragalus polysaccharide and dissolve it in water for injection.
  • the BP5 used in the following examples, comparative examples, and immunoassays are all the bursin pentapeptide prepared in step 4 of this example.
  • the H9N2 subtype avian influenza vaccine is substantially the same as in Example 1.
  • the immune enhancer components are: bursin pentapeptide 1 wt%; astragalus polysaccharide 5 wt%; the balance water.
  • the immune enhancer in the vaccine is 15 ⁇ l/feather.
  • the H9N2 subtype avian influenza vaccine is roughly the same as in Example 1.
  • the immune enhancer components are: cystin pentapeptide 1wt%; astragalus polysaccharide 20wt%; the balance water.
  • the immune booster in the vaccine is 5 ⁇ l/feather.
  • the components of 500 vaccines include:
  • the oil phase contains 84.6ml white oil (EXonMobil), 5.4ml Spencer-80 (Zhaoqing Chaoneng Industrial Co., Ltd.);
  • the water phase contains 52.6ml H5N1 embryo fluid inactivated antigen (10 8 EID 50 /0.1mL, Guangdong Wenshi Dahuanong Biotechnology Co., Ltd.), 2.4ml Tween-80 (Zhaoqing Super Energy Industrial Co., Ltd.), 5ml immune enhancer .
  • the water phase contains 10 ⁇ l/feather of the immune enhancer, and its components are: bursin pentapeptide 1wt%; astragalus polysaccharide 15wt%; and the balance water.
  • the preparation method of the immune enhancer is the same as in Example 1.
  • the H9N2 subtype avian influenza vaccine is basically the same as in Example 1.
  • the immune enhancer does not contain astragalus polysaccharides, and its components are: cystin pentapeptide 1 wt%; the balance water.
  • the H9N2 subtype avian influenza vaccine is roughly the same as in Example 1.
  • the immune enhancer does not contain cystin pentapeptide, and its components are: its components are: astragalus polysaccharide 15wt%; the balance is water.
  • A conventional H9N2 subtype inactivated avian influenza vaccine, subcutaneous injection on the back of the neck, 0.3ml/feather
  • B inactivated vaccine + APS-BP5 (Example 1)
  • C APS-BP5 control group (10 ⁇ l/0.3ml/feather, APS-BP5 concentration content is the same as in Example 1);
  • D PBS control group (each immunized 0.3ml/feather).
  • the specific antibody level was detected by the hemagglutination inhibition test (HI test).
  • HI test hemagglutination inhibition test
  • 3 weeks after immunization the avian influenza virus (H9) antibody level of the APS-BP5 vaccine group began to be significantly higher than that of the conventional vaccine group (p ⁇ 0.01), and continued until the end of the test. From 3 weeks after immunization to the end of the test, the antibody level of the APS-BP5 vaccine group was always higher than that of the conventional vaccine group by more than 3 titers. In the fourth week, its antibody level was higher than that of the conventional vaccine group by 7 titers. The degree is greater than 11 (log2).
  • A conventional H9N2 subtype inactivated avian influenza vaccine, subcutaneous injection at the back of the neck, 0.3ml/feather
  • B inactivated vaccine + BP5 (right Proportion 1)
  • C BP5 control group (10 ⁇ l/0.3ml/feather, the concentration of BP5 is the same as that of Comparative Example 1)
  • D PBS control group (each immunized 0.3ml/feather).
  • the level of specific antibodies was detected by the hemagglutination inhibition test (HI test), as shown in Figure 2.
  • HI test hemagglutination inhibition test
  • the level of avian influenza virus (H9) antibodies in the bursin pentapeptide adjuvant vaccine group began to gradually be higher than that of the conventional vaccine Group (p ⁇ 0.01), and continued until the end of the experiment. From 3 weeks after immunization to the end of the experiment, the antibody level of the bursin pentapeptide vaccine group was always higher than the conventional vaccine group by about 2 titers.
  • A conventional H9N2 subtype inactivated avian influenza vaccine, subcutaneous injection at the back of the neck, 0.3ml/feather
  • B inactivated vaccine + APS (right Proportion 2)
  • C APS control group (10 ⁇ l/0.3ml/feather, the APS concentration content is the same as that of Comparative Example 2);
  • D PBS control group (each immunized 0.3ml/feather).
  • the specific antibody level was detected by the hemagglutination inhibition test (HI test), as shown in Figure 3.
  • HI test hemagglutination inhibition test
  • the avian influenza virus (H9) antibody level of the astragalus polysaccharide adjuvant vaccine group was slightly higher than that of the conventional vaccine group, and it could continue To the end of the test.
  • A conventional H5N1 subtype inactivated avian influenza vaccine, subcutaneous injection at the back of the neck, 0.3ml/feather
  • B H5N1 inactivated vaccine + APS- BP5 (Example 4)
  • C APS-BP5 control group (10 ⁇ l/0.3ml/feather, the concentration of APS-BP5 is the same as in Example 4);
  • D PBS control group (each immunized 0.3ml/feather)
  • the specific antibody level was detected by the hemagglutination inhibition test (HI test). As shown in Figure 4, 2 weeks after immunization, the avian influenza virus (H5) antibody level of the APS-BP5 vaccine group began to be significantly higher than that of the conventional vaccine group (p ⁇ 0.01), and continued until the end of the test. From 3 weeks after immunization to the end of the test, the antibody level of the APS-BP5 vaccine group was always higher than the conventional vaccine group by more than 3 titers.
  • H5 hemagglutination inhibition test
  • Bursalin is a small molecule peptide that exists in the bursa of poultry and is closely related to immune function.
  • Bursin pentapeptide BP5
  • BP5 Bursin pentapeptide
  • It can induce the differentiation and proliferation of poultry B lymphocyte precursors, increase antibody levels, and enhance humoral immunity; promote the transformation activity of T lymphocytes, thereby enhancing cellular immunity .
  • Polysaccharide compound is a kind of good immunomodulator, and its role in improving and improving the immune function of animals has attracted more and more attention.
  • Astragalus polysaccharide is the main bioactive component in the root extract of Astragalus. Studies have shown that Astragalus polysaccharide can stimulate the activity of macrophages, promote the proliferation of T cells, promote the expression of surface antigens in lymphocytes, induce strong humoral and cellular immune responses, and play an important role in non-specific and specific immune responses. effect. In general, Astragalus polysaccharide has multiple functions such as immune regulation, improvement of immune response, and alleviation of immune dysfunction caused by environmental stress.
  • the combined use of the two immune enhancers has a synergistic effect on the effect, and the combination with the avian flu vaccine can obtain an ideal immune effect.

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Immunology (AREA)
  • Chemical & Material Sciences (AREA)
  • Virology (AREA)
  • Veterinary Medicine (AREA)
  • Public Health (AREA)
  • Medicinal Chemistry (AREA)
  • General Health & Medical Sciences (AREA)
  • Animal Behavior & Ethology (AREA)
  • Pharmacology & Pharmacy (AREA)
  • General Chemical & Material Sciences (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Epidemiology (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Organic Chemistry (AREA)
  • Microbiology (AREA)
  • Mycology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Pulmonology (AREA)
  • Engineering & Computer Science (AREA)
  • Molecular Biology (AREA)
  • Communicable Diseases (AREA)
  • Oncology (AREA)
  • Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
  • Peptides Or Proteins (AREA)

Abstract

L'invention concerne un immunopotentialisateur comprenant un pentapeptide de bursine et un polysaccharide d'astragale, le rapport pondéral du pentapeptide de bursine au polysaccharide d'astragale étant compris entre 1:5 et 1:20. L'immunopotentialisateur est simple à préparer et peut améliorer le niveau d'expression du titre d'anticorps pour l'immunité vaccinale. L'invention concerne également un procédé de préparation de l'immunopotentialisateur, un vaccin contre la grippe aviaire utilisant cet immunopotentialisateur et l'application de l'immunopotentialisateur dans le vaccin contre la grippe aviaire.
PCT/CN2019/085076 2019-04-23 2019-04-29 Immunopotentialisateur, son procédé de préparation, vaccin contre la grippe aviaire et utilisation de celui-ci WO2020215351A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
CN201910331019.7A CN109966484B (zh) 2019-04-23 2019-04-23 一种免疫增强剂、制备方法、禽流感疫苗和应用
CN201910331019.7 2019-04-23

Publications (1)

Publication Number Publication Date
WO2020215351A1 true WO2020215351A1 (fr) 2020-10-29

Family

ID=67086003

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/CN2019/085076 WO2020215351A1 (fr) 2019-04-23 2019-04-29 Immunopotentialisateur, son procédé de préparation, vaccin contre la grippe aviaire et utilisation de celui-ci

Country Status (2)

Country Link
CN (1) CN109966484B (fr)
WO (1) WO2020215351A1 (fr)

Families Citing this family (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN111544587B (zh) * 2020-06-29 2023-06-23 肇庆大华农生物药品有限公司 一种禽流感疫苗佐剂及其应用
CN111603558A (zh) * 2020-06-29 2020-09-01 肇庆大华农生物药品有限公司 一种免疫增强剂及其在禽流感疫苗中的应用
WO2022000205A1 (fr) * 2020-06-29 2022-01-06 肇庆大华农生物药品有限公司 Adjuvant de vaccin contre la grippe aviaire et son utilisation
CN116813795B (zh) * 2023-05-25 2024-01-30 华中农业大学 重组AaLS-BSP融合肽、制备方法及应用

Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101434650A (zh) * 2008-12-23 2009-05-20 李德元 法氏囊五肽、其衍生肽及其应用
CN102000332A (zh) * 2010-11-16 2011-04-06 南京农业大学 一种用于鸭禽流感口服黏膜免疫的复合佐剂
CN103145854A (zh) * 2013-03-05 2013-06-12 河南科技大学 重组Tβ4-BP5融合肽、基因、工程菌及应用

Family Cites Families (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101255189A (zh) * 2008-04-07 2008-09-03 广东大华农动物保健品有限公司 一种动物疫苗免疫增强剂及其生产方法

Patent Citations (3)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN101434650A (zh) * 2008-12-23 2009-05-20 李德元 法氏囊五肽、其衍生肽及其应用
CN102000332A (zh) * 2010-11-16 2011-04-06 南京农业大学 一种用于鸭禽流感口服黏膜免疫的复合佐剂
CN103145854A (zh) * 2013-03-05 2013-06-12 河南科技大学 重组Tβ4-BP5融合肽、基因、工程菌及应用

Also Published As

Publication number Publication date
CN109966484A (zh) 2019-07-05
CN109966484B (zh) 2023-08-01

Similar Documents

Publication Publication Date Title
WO2020215351A1 (fr) Immunopotentialisateur, son procédé de préparation, vaccin contre la grippe aviaire et utilisation de celui-ci
ES2322213T3 (es) Proteina nap de helicobacter pylori.
RU2677799C2 (ru) Модифицированные суперспиральные белки с улучшенными свойствами
CN110408637B (zh) 一种草鱼出血病酵母口服疫苗及应用
CN115725002B (zh) 一种大肠杆菌特异性抗原融合蛋白及其重组乳酸乳球菌
CN107488234B (zh) 禽腺病毒Hexon与鸡传染性法氏囊病病毒VP2融合抗原、亚单位疫苗及其制备方法
CN111548395A (zh) 一种口蹄疫病毒二价多表位重组病毒样颗粒及其应用
CN103897065A (zh) 牛Asia1/O型口蹄疫双价多表位疫苗及其制备方法和应用
CN110606875A (zh) 一种用于制备口蹄疫疫苗的分子内佐剂及其应用和口蹄疫疫苗
CN102178950B (zh) 一种亚单位疫苗免疫佐剂及应用
WO2013013602A1 (fr) Procédé de préparation et de renaturation d'interféron aviaire γ
CN104119442A (zh) 牛a型口蹄疫多表位疫苗及其制备方法和应用
CN111607605B (zh) 一种多价表位和亚单位疫苗的构建方法
CN101376887B (zh) 猪口蹄疫重组免疫复合肽的制备方法及应用
WO2013102492A1 (fr) Gènes synthétiques codant pour des fragments peptidiques des protéines de la myéline naturelle pour l'induction de la tolérance orale, fragment d'adn contenant ces gènes, moyens d'obtenir ces peptides dans un système microbien (bactérien) et leur application médicale
CN104250304A (zh) 一种融合蛋白及其编码的疫苗组合物与应用
CN112402598A (zh) 针对鸭疫里默氏杆菌感染引起的通用型亚单位疫苗
CN107245105B (zh) HN-VP233-221aa融合蛋白及其制备方法和应用
CN107281487B (zh) 一种兽用疫苗佐剂
CN114262719B (zh) 副鸡嗜血杆菌三价基因工程亚单位疫苗制备方法及运用
CN115819625B (zh) 一种大肠杆菌四价抗原融合多肽
CN114292313B (zh) 猪巴氏杆菌毒素亚单位疫苗
CN110564750B (zh) 一种鲫造血器官坏死症酵母口服疫苗及应用
CN102206258A (zh) 甲型副伤寒沙门氏菌NmpC亚单位疫苗及其制备方法
CN114957489B (zh) 一种猪轮状病毒重组蛋白及其应用

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 19925845

Country of ref document: EP

Kind code of ref document: A1

NENP Non-entry into the national phase

Ref country code: DE

122 Ep: pct application non-entry in european phase

Ref document number: 19925845

Country of ref document: EP

Kind code of ref document: A1