CN117430665B - 甲型流感病毒t细胞抗原表位肽及其应用 - Google Patents
甲型流感病毒t细胞抗原表位肽及其应用 Download PDFInfo
- Publication number
- CN117430665B CN117430665B CN202311379347.7A CN202311379347A CN117430665B CN 117430665 B CN117430665 B CN 117430665B CN 202311379347 A CN202311379347 A CN 202311379347A CN 117430665 B CN117430665 B CN 117430665B
- Authority
- CN
- China
- Prior art keywords
- influenza
- virus
- epitope peptide
- cells
- antigen
- Prior art date
- Legal status (The legal status 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 status listed.)
- Active
Links
- 108090000765 processed proteins & peptides Proteins 0.000 title claims abstract description 91
- 210000001744 T-lymphocyte Anatomy 0.000 title claims abstract description 80
- 241000712431 Influenza A virus Species 0.000 title claims abstract description 66
- 239000000427 antigen Substances 0.000 claims abstract description 66
- 108091007433 antigens Proteins 0.000 claims abstract description 56
- 102000036639 antigens Human genes 0.000 claims abstract description 56
- 229960005486 vaccine Drugs 0.000 claims abstract description 24
- 210000000612 antigen-presenting cell Anatomy 0.000 claims abstract description 12
- 239000003814 drug Substances 0.000 claims abstract description 11
- 238000002360 preparation method Methods 0.000 claims abstract description 11
- 229940079593 drug Drugs 0.000 claims abstract description 6
- 125000003275 alpha amino acid group Chemical group 0.000 claims abstract description 3
- 210000004027 cell Anatomy 0.000 claims description 32
- 102000025850 HLA-A2 Antigen Human genes 0.000 claims description 24
- 108010074032 HLA-A2 Antigen Proteins 0.000 claims description 24
- 108020004707 nucleic acids Proteins 0.000 claims description 8
- 102000039446 nucleic acids Human genes 0.000 claims description 8
- 150000007523 nucleic acids Chemical class 0.000 claims description 8
- 238000004153 renaturation Methods 0.000 claims description 5
- 238000002255 vaccination Methods 0.000 claims description 5
- 230000000890 antigenic effect Effects 0.000 claims description 4
- 230000000694 effects Effects 0.000 claims description 4
- 238000012216 screening Methods 0.000 claims description 3
- 102000004169 proteins and genes Human genes 0.000 abstract description 10
- 108090000623 proteins and genes Proteins 0.000 abstract description 10
- 238000012827 research and development Methods 0.000 abstract description 6
- 230000007969 cellular immunity Effects 0.000 abstract description 4
- 230000005847 immunogenicity Effects 0.000 abstract description 4
- 238000009169 immunotherapy Methods 0.000 abstract description 4
- 102000004196 processed proteins & peptides Human genes 0.000 description 23
- 206010022000 influenza Diseases 0.000 description 16
- 210000001266 CD8-positive T-lymphocyte Anatomy 0.000 description 11
- 241000700605 Viruses Species 0.000 description 11
- 230000028993 immune response Effects 0.000 description 9
- 238000001514 detection method Methods 0.000 description 7
- 239000000178 monomer Substances 0.000 description 6
- 230000006907 apoptotic process Effects 0.000 description 5
- 239000000243 solution Substances 0.000 description 5
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 4
- 238000011161 development Methods 0.000 description 4
- 238000000684 flow cytometry Methods 0.000 description 4
- 230000036737 immune function Effects 0.000 description 4
- 238000011534 incubation Methods 0.000 description 4
- 208000037797 influenza A Diseases 0.000 description 4
- 229920001184 polypeptide Polymers 0.000 description 4
- 102000011786 HLA-A Antigens Human genes 0.000 description 3
- 108010075704 HLA-A Antigens Proteins 0.000 description 3
- 108010002350 Interleukin-2 Proteins 0.000 description 3
- 230000006044 T cell activation Effects 0.000 description 3
- 238000002835 absorbance Methods 0.000 description 3
- 230000003213 activating effect Effects 0.000 description 3
- 230000004913 activation Effects 0.000 description 3
- 230000000903 blocking effect Effects 0.000 description 3
- 210000004369 blood Anatomy 0.000 description 3
- 239000008280 blood Substances 0.000 description 3
- 230000017188 evasion or tolerance of host immune response Effects 0.000 description 3
- 238000002474 experimental method Methods 0.000 description 3
- 238000004519 manufacturing process Methods 0.000 description 3
- 238000000034 method Methods 0.000 description 3
- 230000035772 mutation Effects 0.000 description 3
- 239000013642 negative control Substances 0.000 description 3
- 210000003819 peripheral blood mononuclear cell Anatomy 0.000 description 3
- 230000002093 peripheral effect Effects 0.000 description 3
- 239000013641 positive control Substances 0.000 description 3
- YBJHBAHKTGYVGT-ZKWXMUAHSA-N (+)-Biotin Chemical compound N1C(=O)N[C@@H]2[C@H](CCCCC(=O)O)SC[C@@H]21 YBJHBAHKTGYVGT-ZKWXMUAHSA-N 0.000 description 2
- BFSVOASYOCHEOV-UHFFFAOYSA-N 2-diethylaminoethanol Chemical compound CCN(CC)CCO BFSVOASYOCHEOV-UHFFFAOYSA-N 0.000 description 2
- 238000002965 ELISA Methods 0.000 description 2
- 102100025137 Early activation antigen CD69 Human genes 0.000 description 2
- 102100030385 Granzyme B Human genes 0.000 description 2
- 101000934374 Homo sapiens Early activation antigen CD69 Proteins 0.000 description 2
- 101001009603 Homo sapiens Granzyme B Proteins 0.000 description 2
- 101000914514 Homo sapiens T-cell-specific surface glycoprotein CD28 Proteins 0.000 description 2
- PXHVJJICTQNCMI-UHFFFAOYSA-N Nickel Chemical compound [Ni] PXHVJJICTQNCMI-UHFFFAOYSA-N 0.000 description 2
- 108010090804 Streptavidin Proteins 0.000 description 2
- 230000024932 T cell mediated immunity Effects 0.000 description 2
- 108091008874 T cell receptors Proteins 0.000 description 2
- 102000016266 T-Cell Antigen Receptors Human genes 0.000 description 2
- 102100027213 T-cell-specific surface glycoprotein CD28 Human genes 0.000 description 2
- 239000007983 Tris buffer Substances 0.000 description 2
- 230000000840 anti-viral effect Effects 0.000 description 2
- 238000006243 chemical reaction Methods 0.000 description 2
- 238000010790 dilution Methods 0.000 description 2
- 239000012895 dilution Substances 0.000 description 2
- 229940031551 inactivated vaccine Drugs 0.000 description 2
- 208000015181 infectious disease Diseases 0.000 description 2
- 108010061181 influenza matrix peptide (58-66) Proteins 0.000 description 2
- 238000005342 ion exchange Methods 0.000 description 2
- 230000001404 mediated effect Effects 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 239000002808 molecular sieve Substances 0.000 description 2
- 238000012544 monitoring process Methods 0.000 description 2
- 210000005259 peripheral blood Anatomy 0.000 description 2
- 239000011886 peripheral blood Substances 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- URGAHOPLAPQHLN-UHFFFAOYSA-N sodium aluminosilicate Chemical compound [Na+].[Al+3].[O-][Si]([O-])=O.[O-][Si]([O-])=O URGAHOPLAPQHLN-UHFFFAOYSA-N 0.000 description 2
- 239000011780 sodium chloride Substances 0.000 description 2
- 230000000638 stimulation Effects 0.000 description 2
- 229940126585 therapeutic drug Drugs 0.000 description 2
- 230000001225 therapeutic effect Effects 0.000 description 2
- 239000013638 trimer Substances 0.000 description 2
- LENZDBCJOHFCAS-UHFFFAOYSA-N tris Chemical compound OCC(N)(CO)CO LENZDBCJOHFCAS-UHFFFAOYSA-N 0.000 description 2
- 239000011534 wash buffer Substances 0.000 description 2
- 238000005406 washing Methods 0.000 description 2
- 102000000412 Annexin Human genes 0.000 description 1
- 108050008874 Annexin Proteins 0.000 description 1
- 108020005544 Antisense RNA Proteins 0.000 description 1
- 239000004475 Arginine Substances 0.000 description 1
- YASYEJJMZJALEJ-UHFFFAOYSA-N Citric acid monohydrate Chemical compound O.OC(=O)CC(O)(C(O)=O)CC(O)=O YASYEJJMZJALEJ-UHFFFAOYSA-N 0.000 description 1
- 208000034657 Convalescence Diseases 0.000 description 1
- 238000011537 Coomassie blue staining Methods 0.000 description 1
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 description 1
- 241001198387 Escherichia coli BL21(DE3) Species 0.000 description 1
- 102000001398 Granzyme Human genes 0.000 description 1
- 108060005986 Granzyme Proteins 0.000 description 1
- 102000008949 Histocompatibility Antigens Class I Human genes 0.000 description 1
- 108010088652 Histocompatibility Antigens Class I Proteins 0.000 description 1
- 241000701044 Human gammaherpesvirus 4 Species 0.000 description 1
- MHAJPDPJQMAIIY-UHFFFAOYSA-N Hydrogen peroxide Chemical compound OO MHAJPDPJQMAIIY-UHFFFAOYSA-N 0.000 description 1
- 206010061598 Immunodeficiency Diseases 0.000 description 1
- 102100037850 Interferon gamma Human genes 0.000 description 1
- 108010074328 Interferon-gamma Proteins 0.000 description 1
- 102000043129 MHC class I family Human genes 0.000 description 1
- 108091054437 MHC class I family Proteins 0.000 description 1
- 101710199771 Matrix protein 1 Proteins 0.000 description 1
- 229930192392 Mitomycin Natural products 0.000 description 1
- NWIBSHFKIJFRCO-WUDYKRTCSA-N Mytomycin Chemical compound C1N2C(C(C(C)=C(N)C3=O)=O)=C3[C@@H](COC(N)=O)[C@@]2(OC)[C@@H]2[C@H]1N2 NWIBSHFKIJFRCO-WUDYKRTCSA-N 0.000 description 1
- 108091028043 Nucleic acid sequence Proteins 0.000 description 1
- 229920001213 Polysorbate 20 Polymers 0.000 description 1
- 239000012506 Sephacryl® Substances 0.000 description 1
- 239000012505 Superdex™ Substances 0.000 description 1
- XSQUKJJJFZCRTK-UHFFFAOYSA-N Urea Chemical compound NC(N)=O XSQUKJJJFZCRTK-UHFFFAOYSA-N 0.000 description 1
- 208000027418 Wounds and injury Diseases 0.000 description 1
- ZTOJFFHGPLIVKC-CLFAGFIQSA-N abts Chemical compound S/1C2=CC(S(O)(=O)=O)=CC=C2N(CC)C\1=N\N=C1/SC2=CC(S(O)(=O)=O)=CC=C2N1CC ZTOJFFHGPLIVKC-CLFAGFIQSA-N 0.000 description 1
- 230000008485 antagonism Effects 0.000 description 1
- 239000003443 antiviral agent Substances 0.000 description 1
- ODKSFYDXXFIFQN-UHFFFAOYSA-N arginine Natural products OC(=O)C(N)CCCNC(N)=N ODKSFYDXXFIFQN-UHFFFAOYSA-N 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- 230000004888 barrier function Effects 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 229960002685 biotin Drugs 0.000 description 1
- 235000020958 biotin Nutrition 0.000 description 1
- 239000011616 biotin Substances 0.000 description 1
- 239000004202 carbamide Substances 0.000 description 1
- 230000001413 cellular effect Effects 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 229960002303 citric acid monohydrate Drugs 0.000 description 1
- 239000003184 complementary RNA Substances 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- OOTFVKOQINZBBF-UHFFFAOYSA-N cystamine Chemical compound CCSSCCN OOTFVKOQINZBBF-UHFFFAOYSA-N 0.000 description 1
- 229940099500 cystamine Drugs 0.000 description 1
- UFULAYFCSOUIOV-UHFFFAOYSA-N cysteamine Chemical compound NCCS UFULAYFCSOUIOV-UHFFFAOYSA-N 0.000 description 1
- 230000006378 damage Effects 0.000 description 1
- 230000005860 defense response to virus Effects 0.000 description 1
- 239000008367 deionised water Substances 0.000 description 1
- 229910021641 deionized water Inorganic materials 0.000 description 1
- 239000013024 dilution buffer Substances 0.000 description 1
- 201000010099 disease Diseases 0.000 description 1
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 1
- 238000006073 displacement reaction Methods 0.000 description 1
- 241001493065 dsRNA viruses Species 0.000 description 1
- 238000011156 evaluation Methods 0.000 description 1
- 230000001747 exhibiting effect Effects 0.000 description 1
- 239000013604 expression vector Substances 0.000 description 1
- MHMNJMPURVTYEJ-UHFFFAOYSA-N fluorescein-5-isothiocyanate Chemical compound O1C(=O)C2=CC(N=C=S)=CC=C2C21C1=CC=C(O)C=C1OC1=CC(O)=CC=C21 MHMNJMPURVTYEJ-UHFFFAOYSA-N 0.000 description 1
- 238000010353 genetic engineering Methods 0.000 description 1
- PCHJSUWPFVWCPO-UHFFFAOYSA-N gold Chemical compound [Au] PCHJSUWPFVWCPO-UHFFFAOYSA-N 0.000 description 1
- 239000010931 gold Substances 0.000 description 1
- 229910052737 gold Inorganic materials 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 230000005965 immune activity Effects 0.000 description 1
- 230000007124 immune defense Effects 0.000 description 1
- 230000036039 immunity Effects 0.000 description 1
- 230000001506 immunosuppresive effect Effects 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
- 238000000338 in vitro Methods 0.000 description 1
- 238000001727 in vivo Methods 0.000 description 1
- 230000001939 inductive effect Effects 0.000 description 1
- 208000014674 injury Diseases 0.000 description 1
- 238000002372 labelling Methods 0.000 description 1
- 210000004698 lymphocyte Anatomy 0.000 description 1
- 239000003550 marker Substances 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 230000007246 mechanism Effects 0.000 description 1
- 229960003151 mercaptamine Drugs 0.000 description 1
- 229960004857 mitomycin Drugs 0.000 description 1
- 230000003472 neutralizing effect Effects 0.000 description 1
- 229910052759 nickel Inorganic materials 0.000 description 1
- GEVPUGOOGXGPIO-UHFFFAOYSA-N oxalic acid;dihydrate Chemical compound O.O.OC(=O)C(O)=O GEVPUGOOGXGPIO-UHFFFAOYSA-N 0.000 description 1
- 244000052769 pathogen Species 0.000 description 1
- 230000001575 pathological effect Effects 0.000 description 1
- 230000002085 persistent effect Effects 0.000 description 1
- 239000000256 polyoxyethylene sorbitan monolaurate Substances 0.000 description 1
- 235000010486 polyoxyethylene sorbitan monolaurate Nutrition 0.000 description 1
- 238000010837 poor prognosis Methods 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 238000004393 prognosis Methods 0.000 description 1
- 230000000069 prophylactic effect Effects 0.000 description 1
- 230000001681 protective effect Effects 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 108020003175 receptors Proteins 0.000 description 1
- 238000009877 rendering Methods 0.000 description 1
- NLJMYIDDQXHKNR-UHFFFAOYSA-K sodium citrate Chemical compound O.O.[Na+].[Na+].[Na+].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O NLJMYIDDQXHKNR-UHFFFAOYSA-K 0.000 description 1
- 238000002415 sodium dodecyl sulfate polyacrylamide gel electrophoresis Methods 0.000 description 1
- 230000004936 stimulating effect Effects 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
- 230000001629 suppression Effects 0.000 description 1
- 230000004083 survival effect Effects 0.000 description 1
- 230000008685 targeting Effects 0.000 description 1
- 238000010998 test method Methods 0.000 description 1
- 241000712461 unidentified influenza virus Species 0.000 description 1
- 230000009385 viral infection Effects 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
- C07K7/04—Linear peptides containing only normal peptide links
- C07K7/06—Linear peptides containing only normal peptide links having 5 to 11 amino acids
-
- 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
- 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
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/705—Receptors; Cell surface antigens; Cell surface determinants
- C07K14/70503—Immunoglobulin superfamily
- C07K14/70539—MHC-molecules, e.g. HLA-molecules
-
- 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
- C12N5/00—Undifferentiated human, animal or plant cells, e.g. cell lines; Tissues; Cultivation or maintenance thereof; Culture media therefor
- C12N5/06—Animal cells or tissues; Human cells or tissues
- C12N5/0602—Vertebrate cells
- C12N5/0634—Cells from the blood or the immune system
- C12N5/0636—T lymphocytes
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N21/00—Investigating or analysing materials by the use of optical means, i.e. using sub-millimetre waves, infrared, visible or ultraviolet light
- G01N21/62—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light
- G01N21/63—Systems in which the material investigated is excited whereby it emits light or causes a change in wavelength of the incident light optically excited
- G01N21/64—Fluorescence; Phosphorescence
- G01N21/6486—Measuring fluorescence of biological material, e.g. DNA, RNA, cells
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/53—Immunoassay; Biospecific binding assay; Materials therefor
- G01N33/569—Immunoassay; Biospecific binding assay; Materials therefor for microorganisms, e.g. protozoa, bacteria, viruses
- G01N33/56966—Animal cells
- G01N33/56977—HLA or MHC typing
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N33/00—Investigating or analysing materials by specific methods not covered by groups G01N1/00 - G01N31/00
- G01N33/48—Biological material, e.g. blood, urine; Haemocytometers
- G01N33/50—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing
- G01N33/94—Chemical analysis of biological material, e.g. blood, urine; Testing involving biospecific ligand binding methods; Immunological testing involving narcotics or drugs or pharmaceuticals, neurotransmitters or associated receptors
- G01N33/9446—Antibacterials
-
- 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
- A61K2039/51—Medicinal preparations containing antigens or antibodies comprising whole cells, viruses or DNA/RNA
- A61K2039/515—Animal cells
- A61K2039/5154—Antigen presenting cells [APCs], e.g. dendritic cells or macrophages
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K2319/00—Fusion polypeptide
-
- 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
- C12N2510/00—Genetically modified cells
-
- 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
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2333/00—Assays involving biological materials from specific organisms or of a specific nature
- G01N2333/435—Assays involving biological materials from specific organisms or of a specific nature from animals; from humans
- G01N2333/705—Assays involving receptors, cell surface antigens or cell surface determinants
- G01N2333/70503—Immunoglobulin superfamily, e.g. VCAMs, PECAM, LFA-3
- G01N2333/70517—CD8
-
- G—PHYSICS
- G01—MEASURING; TESTING
- G01N—INVESTIGATING OR ANALYSING MATERIALS BY DETERMINING THEIR CHEMICAL OR PHYSICAL PROPERTIES
- G01N2500/00—Screening for compounds of potential therapeutic value
- G01N2500/10—Screening for compounds of potential therapeutic value involving cells
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Immunology (AREA)
- Engineering & Computer Science (AREA)
- Molecular Biology (AREA)
- General Health & Medical Sciences (AREA)
- Biomedical Technology (AREA)
- Organic Chemistry (AREA)
- Biochemistry (AREA)
- Medicinal Chemistry (AREA)
- Virology (AREA)
- Cell Biology (AREA)
- Hematology (AREA)
- Biotechnology (AREA)
- Urology & Nephrology (AREA)
- Zoology (AREA)
- Microbiology (AREA)
- Genetics & Genomics (AREA)
- Pathology (AREA)
- Physics & Mathematics (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Pharmacology & Pharmacy (AREA)
- General Physics & Mathematics (AREA)
- Analytical Chemistry (AREA)
- Veterinary Medicine (AREA)
- Animal Behavior & Ethology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Public Health (AREA)
- Wood Science & Technology (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Food Science & Technology (AREA)
- Biophysics (AREA)
- Pulmonology (AREA)
- Mycology (AREA)
- Epidemiology (AREA)
- Toxicology (AREA)
- Gastroenterology & Hepatology (AREA)
- General Engineering & Computer Science (AREA)
- Tropical Medicine & Parasitology (AREA)
Abstract
本发明属于免疫治疗技术领域，具体涉及甲型流感病毒T细胞抗原表位肽及其应用。本发明要解决的技术问题是目前在甲型流感病毒领域尚没有开发一种用于甲型流感病毒通用疫苗的T细胞抗原表位肽。本发明的技术方案甲型流感病毒T细胞抗原表位肽，其氨基酸序列如SEQ ID No.10所示。本发明提供了的抗原表位肽具有很强的免疫原性，能诱导抗原特异性的CD8+T细胞；其可以和HLA‑A2重链和HLA‑A2轻链β2m蛋白组装成pMHC复合物；或直接负荷到抗原递呈细胞，能活化T细胞，有效诱导T细胞免疫，可用于甲型流感病毒通用疫苗研发、制备和药物研发、临床治疗。
Description
技术领域
本发明属于免疫治疗技术领域，具体涉及甲型流感病毒T细胞抗原表位肽及其应用。
背景技术
流感依然是严重威胁人类健康的，特别是儿童、老人、孕妇和免疫功能低下人群的重要疾病，加强对流感相关的研究依然具有重大的科学和社会意义。
由于有效抗病毒药物的缺失，对流感及其并发症的治疗主要是对症治疗。疫苗接种是目前比较有效的预防措施，多以诱导机体主动产生保护性抗体为主。但是由于甲型流感病毒是单链反义RNA病毒，极易发生变异，产生抗原漂移(antigenic drift)和抗原转变(antigenic shift)，使中和性抗体失去效应，从而导致疫苗失效。相对应的，细胞免疫应答识别高度保守的递呈抗原，因此在不同的甲型流感病毒亚型之间呈现高效的交叉保护能力。譬如，甲型流感病毒(IAV)基质蛋白1(Matrix Protein 1，M1)58到66位GILGFVFTL(M158-66 GIL，简称GIL)特异性的CD8+T细胞免疫应答被发现占据了高达80％的抗甲型流感病毒免疫应答。因此，研究抗甲型流感病毒的细胞免疫应答有望更好地理解抗病毒免疫的机制，为研制广谱高效的抗甲型流感病毒疫苗提供理论与实验基础。
鉴于当前甲型流感病毒变异不断出现，为将来开发通用型甲型流感病毒疫苗和广谱治疗药物提供了契机。通用疫苗是一类选择即使病毒抗原发生变异后仍能有效保护免疫人群特性的广谱疫苗。因此，开发出在甲型流感病毒发生各类变异后，依旧能够给疫苗免疫人群提供有效免疫保护的通用疫苗以及广谱治疗性药物迫在眉睫。
研究显示，T细胞免疫应答在病毒感染后机体抗病毒防御以及机体免疫病理损伤过程中发挥了重要的作用，尤其是CD8+T细胞，其抗原特异性免疫活性11年后依然存在，说明了CD8+T细胞免疫应答在抗甲型流感病毒免疫防御中的重要作用及其在疫苗研发中的重要地位。CD8+T细胞免疫应答的第一步就是T细胞通过其表面的抗原识别受体特异性识别被病毒感染的细胞所递呈的抗原表位肽。因此，抗原表位肽是T细胞特异性识别病毒、发挥免疫保护作用的重要关键分子，是免疫检测、免疫治疗和疫苗研发的关键靶向分子。
CD8+T细胞通过T细胞受体(TCR)识别pMHC活化，杀灭病毒感染细胞、清除病毒，从而发挥抗病毒的细胞免疫作用。因此，鉴定可以有效活化T细胞抗原表位肽，在甲型流感病毒病毒抗原发生变异后仍能诱导有效T细胞免疫保护，是开发甲型流感病毒通用型疫苗和广谱治疗性药物的关键之一。
免疫逃逸是免疫抑制病原体通过其结构和非结构产物，拮抗、阻断和抑制机体的免疫应答。甲型流感病毒因为经常地持续性地发生突变，产生了大量的变异株，严重的威胁了疫苗的免疫屏障效果。
发明内容
本发明要解决的技术问题是目前在甲型流感病毒领域尚没有开发一种用于甲型流感病毒通用疫苗的T细胞抗原表位肽。
本发明的技术方案是甲型流感病毒T细胞抗原表位肽，其氨基酸序列如SEQ IDNo.10所示。
进一步的，本发明还提供了编码所述抗原表位肽的核酸分子。
本发明还提供了含有所述抗原表位肽的pMHC复合物。
进一步的，所述pMHC复合物由HLA-A2重链、HLA-A2轻链β2m和所述抗原表位肽复性得到。
其中，HLA-A2重链、HLA-A2轻链β2m和所述抗原表位肽的摩尔比为1︰2︰10。
本发明还进一步提供了一种抗原肽-抗原递呈细胞复合物，为表面负荷所述抗原表位肽的抗原递呈细胞。
其中，所述抗原递呈细胞为CD8+T细胞。
优选的，所述CD8+T细胞为T2-A2细胞。
本发明还提供了上述抗原表位肽、编码所述抗原表位肽的核酸分子、pMHC复合物和/或抗原肽-抗原递呈细胞复合物在制备甲型流感病毒药物中的应用。
本发明还提供了上述抗原表位肽、编码所述抗原表位肽的核酸分子、pMHC复合物和/或抗原肽-抗原递呈细胞复合物在筛选甲型流感病毒药物中的应用。
本发明还进一步提供了上述抗原表位肽、编码所述抗原表位肽的核酸分子、pMHC复合物和/或抗原肽-抗原递呈细胞复合物在制备甲型流感病毒疫苗中的应用。
本发明还进一步提供了上述抗原表位肽、编码所述抗原表位肽的核酸分子、pMHC复合物和/或抗原肽-抗原递呈细胞复合物在制备评估甲型流感病毒疫苗接种效果的药物的应用。
本发明的有益效果：本发明提供了一种甲型流感病毒T细胞抗原表位肽，其具有很强的免疫原性，能诱导抗原特异性的CD8+T细胞；其可以和HLA-A2重链和HLA-A2轻链β2m蛋白组装成pMHC复合物；或直接负荷到抗原递呈细胞，能活化T细胞，有效诱导T细胞免疫，避免甲型流感病毒突变株的免疫逃逸，并在流感疫苗接种者和康复者中均可检测到特异性的CD8+T细胞，可用于甲型流感病毒通用疫苗研发、制备和药物研发、临床治疗。
本发明还利用该甲型流感病毒CD8+T细胞抗原表位肽制备成带有PE荧光通道的pMHC复合物，可以用于甲型流感病毒疫苗接种者和感染康复者外周血内抗原特异性T细胞的检测，并用于体外的T细胞活化实验，这些甲型流感病毒CD8+T细胞抗原表位肽可以用于制备针对各种甲型流感病毒突变株的通用疫苗、甲型流感病毒相关的免疫检测以及广谱治疗性药物。这些甲型流感病毒CD8+T细胞抗原表位肽制备成带有PE荧光通道的pMHC复合物，或直接负荷到抗原递呈细胞，从而活化T细胞，可用于甲型流感病毒疫苗研发、制备和药物研发、临床治疗，可应用于：
1)甲型流感病毒疫苗的研发和制备：甲型流感病毒发生变异后，该多个T细胞表位能诱导机体产生免疫应答生成抗原特异性T细胞。故该T细胞表位是甲型流感病毒通用疫苗的候选抗原表位肽。
2)检测是否具有抵抗甲型流感病毒感染的细胞免疫功能：待检者体内检测到甲型流感病毒抗原特异性T细胞，代表机体已经产生了T细胞免疫功能，根据该抗原表位肽制备成荧光通道的pMHC复合物标记的抗原特异性CD8+T的比例，可以评价机体T细胞免疫功能强弱及对甲型流感病毒感染可能性。
3)评估疫苗接种后的效果：接种者体内检测到甲型流感病毒抗原特异性T细胞，代表机体已经产生了T细胞免疫功能，根据其比例，可以评估机体再次感染甲型流感病毒可能性。
4)监测病情：可用于对密切接触者、医学观察者、疑似和确诊患者的病情变化监测。
5)预后判断：如果机体不能产生T细胞免疫应答，或抗原特异性T细胞比例持续减少，则预示预后不佳。
附图说明
图1、甲型流行性感冒病毒(IAV)T细胞抗原表位的筛选和鉴定，A和B为T2稳定性实验。A为抗β2m的流式细胞检测，B为A的统计图。C：光敏肽置换实验ELISA的结果。Blank(空白对照):no peptides；Negative ctrl(阴性对照)：EBV virus peptide IVTDFSVIK；Positive ctrl(阳性对照)，influenza A M1 peptide GILGFVFTL(下同)。
图2、HLA-A2的pMHC复合物的制备。A为pMHC复合物分子筛(Sephacryl S-200)纯化结果四份不同样品(图中数字标识为1，2，3，4)，数字1-4代表采集的第一到第四个峰值。B为考马斯蓝染色的15％ SDS-PAGE结果；M：蛋白分子量marker，图中数字1-4对应图A中1-4的峰，其中红色标记的2代表目的峰，为HLA的重链(a)和轻链(β2m)。
图3、甲型流感病毒T细胞抗原表位的免疫原性鉴定。A和B：16小时后CD69(上排)和CD137(下排)分子的检测，B为A的统计图。
图4、抗原表位混合刺激后检测到抗原特异性T细胞的生成；A为抗原表位混合刺激健康人CD8+T细胞0天(上排)和7天(下排)后带有荧光通道的pMHC复合物可以检测到抗原特异性T细胞的生成，B为A的统计图。
图5、表位刺激的T细胞介导的T2细胞凋亡比例。A-B(上排)：刺激培养7天时间T2A2细胞的凋亡百分比；(下排)第7天末，T2A2细胞凋亡百分比。
图6、检测活化CD8+T细胞中IFN-γ(上排)和Granzyme B(下排)的表达情况(A)。B为A的统计图。
图7、评估流感灭活疫苗接种后14天和甲型流感病毒感染康复者7天后体内特异性CD8+T细胞的比例；A-B：流式细胞术检测HLA-A2+流感灭活疫苗接种者上述4种(T细胞活化时，可以产生的抗原特异性CD8+T表位)肽的特异性CD8+T细胞；C-D：流式细胞术检测HLA-A2+甲型流感病毒感染康复者上述4种(T细胞活化时，可以产生的抗原特异性CD8+T表位)肽的特异性CD8+T细胞。
具体实施方式
T2-A2细胞是一种通过重组基因工程技术表达人MHC-I类分子HLA-A2的抗原递呈细胞系。只有有效的抗原表位肽才能被其递呈，从而在其细胞表面形成稳定的pMHC复合物，因此可以作为刺激T细胞的人工抗原递呈细胞。
T细胞抗原表位肽单独无法工作，必须以pMHC复合物或抗原肽-抗原递呈细胞复合物的方式进行T细胞活化。本发明利用MHC的单体和鉴定出的甲型流感病毒T细胞表位进行联合复性，制备了pMHC复合物。将鉴定出的甲型流感病毒T细胞抗原表位肽负荷在抗原递呈细胞表面(T2-A2细胞)，制备了抗原肽-抗原递呈细胞复合物，然后用制备好的pMHC复合物去标记CD8+T细胞，发现其抗原表位肽可以有效活化健康人外周血中T细胞，并具有很强的免疫原性，也可以有效杀伤携带甲型流感病毒抗原的靶细胞。该T细胞抗原表位肽组装成的带有PE荧光通道的pMHC复合物，可以在甲型流感病毒疫苗接种者和康复者中均可检测到。证明新发现的甲型流感病毒CD8+T细胞抗原表位肽能有效诱导T细胞免疫，避免病毒突变的免疫逃逸，可以作为通用型疫苗或应用于免疫治疗等。
下面结合说明书附图及具体实施例对本发明作出进一步地详细阐述，所述实施例只用于解释本发明，并非用于限定本发明的范围。下述实施例中所使用的试验方法如无特殊说明，均为常规方法；所使用的材料、试剂等，如无特殊说明，为可从商业途径得到的试剂和材料。
实施例1甲型流感病毒HLA-A2限制性抗原表位肽的预测
以美国NIH抗原表位数据库(IEDB)提供的MHC-I类分子预测工具(http://tools.iedb.org/mhci/)进行HLA-A2限制性抗原表位预测。招募的志愿者是接种甲型流感病毒裂解疫苗，疫苗采用毒诸位采用WHO和欧盟推荐的A1型、A3型和B型甲型流感病毒(2021/2022)的流行株或相似株(北半球)。疫苗的主要成分是含有以下毒株(2021/2022)的抗原(an A/Victoria/2570/2019(H1N1)pdm09；an A/Cambodia/e0826360/2020(H3N2)；aB/Washington/02/2019(B/Victoria lineage)-like virus)。选用最常见的H1N1和H3N2在Gisaid数据库中找到对应的序列编号(EPI_ISL_417210；EPI_ISL_806547)进行抗原表位蛋白的预测。最后得到11种候选的甲型流感病毒特异性HLA-A2+的CD8+T细胞抗原表位肽，如表1所示。
表1甲型流感病毒病毒T细胞11种抗原表位肽
| 编号 | 开始位置 | 结束位置 | 长度 | 序列 | 序列号 |
| F1 | 413 | 421 | 9 | NMLSTVLGV | SEQ ID No.1 |
| F2 | 46 | 54 | 9 | FMYSDFHFI | SEQ ID No.2 |
| F3 | 18 | 26 | 9 | STICFFMQI | SEQ ID No.3 |
| F4 | 275 | 283 | 9 | CLPACVYGL | SEQ ID No.4 |
| F5 | 373 | 381 | 9 | AMDSNTLEL | SEQ ID No.5 |
| F6 | 24 | 32 | 9 | MQIAILITT | SEQ ID No.6 |
| F7 | 458 | 466 | 9 | FQGRGVFEL | SEQ ID No.7 |
| F8 | 122 | 130 | 9 | AIMDKNIIL | SEQ ID No.8 |
| F9 | 218 | 226 | 9 | CVNGSCFTV | SEQ ID No.9 |
| F10 | 249 | 257 | 9 | KADTKILFI | SEQ ID No.10 |
| F11 | 128 | 136 | 9 | IVLEANFSV | SEQ ID No.11 |
实施例2甲型流感病毒HLA-A2限制性抗原表位肽的鉴定
人工合成实施例1预测得到的候选T细胞抗原表位肽(南京金斯瑞生物科技有限公司)，配置成浓度为20μM。取对数生长状态T2-A2细胞(T2-A2为马萨诸塞大学医学院(University of Massachusetts Medical School)Anna Gil博士的赠与)，种植到96孔板，每孔105个，分布设置空白孔、阴性对照肽(EB病毒，IVTDFSVIK)、阳性对照肽(甲型流感M1肽，SEQ ID No.12：GILGFVFTL)和各合成候选T细胞抗原表位肽，每组3个复孔，终体积200μL。37℃孵育4小时后离心洗涤两次，以FITC anti-human HLA-A2(β2m)抗体标记，4℃避光孵育30分钟后，以流式细胞仪检测。实验共进行3次。
结果如图1的A和B所示，结果显示，11种抗原多肽均可以被抗原递呈细胞有效递呈信息给T细胞，说明这些肽是T细胞抗原表位肽。
实施例3抗原多肽形成pMHC复合物的检测
采用ELISA法检测实施例2中筛选得到的11种甲型流感病毒T细胞抗原表位肽。具体操作如下：
在室温下，用100μL的0.5μg mL-1链霉亲和素在96个U形板上孵育16～18小时，用洗涤缓冲液(BioLegend,Cat#420201,US)洗涤3次，并在室温下用稀释缓冲液(0.5M Tris pH8.0，1M NaCl，1％ BSA，0.2％吐温20)封闭30分钟。光敏肽(SEQ ID No.13：KILGFVFJV)与MHC((BioLegend,Cat#280003,US))形成的pMHC复合物设为HLA空白对照，甲型流感M1肽(GILGFVFTL)设为阳性对照，EB病毒肽(SEQ ID No.14：IVTDFSVIK)设为阴性对照。然后，分别添加用365nm三用紫外线分析仪(Qilin Bell，Cat#1903274)置换好的20μL实施例1中的11种流感病毒T细胞抗原表位肽的稀释液(400μM)和20μL Flex-TTM单体(200μg/mL)的稀释液100μL到96孔板，并在温度为37℃的细胞培养箱里孵育1小时。使用洗涤缓冲液洗涤3次后，添加100μL稀释HRP结合抗体(BioLegend,Cat#280303,US)，并在37℃下继续孵育1小时，然后洗涤。添加100μL底物溶液(10.34mL去离子水，1.2mL 0.1M柠檬酸一水合物/柠檬酸三钠二水合物，pH 4.0，240μL 40mM ABTS，120μL过氧化氢溶液)，并在室温避光孵育8min。使用50μL终止溶液(2％w/v草酸二水合物)终止反应。在30分钟内，用酶标仪在450nm波长下测定吸光度(OD值)。
结果如图1的C所示，结果显示11种抗原多肽均可以形成pMHC复合物。
实施例4抗原表位肽的pMHC复合物单体的制备
将HLA-A2α链和β2m的核苷酸序列分别构建到pET28a表达载体上，转入大肠杆菌BL21(DE3)进行蛋白的表达，使用镍柱(Biorad，Cat#7324610，US)亲和层析对HLA-A2蛋白(HLA-A2重链a链、HLA-A2轻链β2m)进行纯化，将纯化的蛋白HLA-A2α链蛋白、β2m蛋白和实施例1中表1中11种抗原表位肽分别按照1︰2︰10的摩尔比逐步滴加至40mL复性溶液(5M尿素、0.4M精氨酸、100mM Tris、3.7mM胱胺、6.3mM半胱胺和2mM EDTA)中进行复性，得到抗原表位肽的pMHC复合物单体。
pMHC复合物单体经过DEAE离子交换柱进一步纯化，以0.5M NaCl洗脱，并根据OD280nm紫外吸收峰收集蛋白。随后经DEAE离子交换柱纯化后的蛋白根据分子量大小，通过Superdex 75pg分子筛纯化，以PBS洗脱，并根据OD280 nm紫外吸收峰收集不同分子量蛋白。图2A和2B所示，经过纯化后得到了抗原表位肽的pMHC复合物三聚体(HLA-A2重链a链+HLA-A2轻链β2m+抗原肽的三聚体)。
实施例5甲型流感病毒HLA-A2限制性抗原表位肽活化T细胞
T2细胞通过表达HLA-A2分子(T2-A2，PMID:34414379；PMID:35194575；PMID:35116022；PMID:37117789)来活化T细胞。分离健康志愿者的外周静脉血中的单个核淋巴细胞(PBMCs)，并进一步分离CD8+T细胞。将T2-A2细胞用CFSE标记，用20μg/mL丝裂霉素处理20分钟后，分别用实施例1中11种不同抗原肽孵育。
具体做法为：
将96孔板中每个孔种0.5×106个CD8+T细胞分别与表1中的11条抗原表位肽负荷的0.5×106个T2-A2细胞共培养(共11个孔)，并用1μg/mL抗人CD28抗体和50IU/mL IL-2共刺激。每隔两天补充50IU/mL IL-2和20μM抗原表位肽。
分离健康志愿者外周静脉血中CD8+T细胞，和抗原多肽负荷的T2细胞共同培养，加入1μg/mL抗人CD28抗体和50IU/mL IL-2共刺激，培养16小时后检测T细胞活化分子CD69和CD137的表达。
将实施例4得到的30μL抗原表位肽的pMHC复合物单体与3.3μL PE链霉亲和素(BioLegend Cat#405203，US)在96孔板中混合，并在4℃避光孵育30分钟后，再加入2.4μL封闭溶液(1.6μL 50mM生物素(Thermo Fisher，Cat#B20656，US))和198.4μL PBS以停止反应，并在4～8℃下培养过夜，即可得到带有PE荧光通道的pMHC复合物。
培养7天后计算T2-A2的存活数目百分比、用带有PE荧光通道的pMHC复合物标记特异性CD8+T细胞比例和T2-A2细胞上的凋亡标记物Annexin V-APC百分比，检测特异性CD8+T细胞释放IFN-γ和GZMB的情况。与CD8+T细胞培养7天后IAV介导T2凋亡百分比：CFSE标记的T2A2作为靶细胞，检测残留的靶细胞数目，计算方法为T2细胞的50％减去存活细胞的百分比。
结果如图3A和3B所示，实施例2中筛选得到的11种甲型流感病毒T细胞抗原表位肽均能够活化T细胞。其中，F3对应的T细胞抗原表位肽STICFFMQI、F6对应的T细胞抗原表位肽MQIAILITT、F10对应的T细胞抗原表位肽KADTKILFI和F11对应的T细胞抗原表位肽IVLEANFSV活化CD8+T细胞的能力较强，如图4A和4B所示。同时以上11条肽活化的特异性CD8+T可以杀伤靶细胞，如图5A和5B所示；特异性的CD8+T细胞释放IFN-γ和GZMB，如图6A和6B所示。
实施例6甲型流感病毒HLA-A2限制性抗原表位肽活化T细胞
分离甲型流感病毒接种灭活疫苗第二针后14天和甲型流感病毒感染康复者7天后的志愿者外周静脉血中的PBMCs，并鉴定其HLA亚型，HLA-A2阳性PBMCs样品进一步用实施例5中带有PE荧光通道的pMHC复合物和CD8-APC抗体进行染色，然后进行流式上机检测。
结果如图7所示，结果显示，F3、F6、F10和F114种抗原表位肽的带有荧光通道的pMHC复合物可以识别甲型流感病毒疫苗接种者(图7A和7B)和甲型流感病毒感染康复者(图7C和7D)体内产生的抗原特异性CD8+T细胞。
最后所应当说明的是，以上实施例仅用以说明本发明的技术方案而非对本发明保护范围的限制，对于本领域的普通技术人员来说，在上述说明及思路的基础上还可以做出其它不同形式的变化或变动，这里无需也无法对所有的实施方式予以穷举。凡在本发明的精神和原则之内所作的任何修改、等同替换和改进等，均应包含在本发明权利要求的保护范围之内。
Claims (7)
1.甲型流感病毒T细胞抗原表位肽，其特征在于：其氨基酸序列如SEQ ID No.10所示。
2.编码权利要求1所述抗原表位肽的核酸分子。
3.含有所述权利要求1所述抗原表位肽的pMHC复合物，其特征在于：由HLA-A2重链、HLA-A2轻链β2m和所述抗原表位肽复性得到。
4.根据权利要求3所述pMHC复合物，其特征在于：HLA-A2重链、HLA-A2轻链β2m和所述抗原表位肽的摩尔比为1︰2︰10。
5.一种抗原肽-抗原递呈细胞复合物，其特征在于：为表面负荷权利要求1所述抗原表位肽的抗原递呈细胞；所述抗原递呈细胞为T2-A2细胞。
6.权利要求1所述抗原表位肽、权利要求2所述编码抗原表位肽的核酸分子、权利要求3或4所述pMHC复合物和/或权利要求5所述抗原肽-抗原递呈细胞复合物在制备甲型流感病毒药物或筛选甲型流感病毒药物中的应用。
7.权利要求1所述抗原表位肽、权利要求2所述编码抗原表位肽的核酸分子、权利要求3或4所述pMHC复合物和/或权利要求5所述抗原肽-抗原递呈细胞复合物在制备甲型流感病毒疫苗或制备评估甲型流感病毒疫苗接种效果的药物中的应用。
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202311379347.7A CN117430665B (zh) | 2023-10-24 | 2023-10-24 | 甲型流感病毒t细胞抗原表位肽及其应用 |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202311379347.7A CN117430665B (zh) | 2023-10-24 | 2023-10-24 | 甲型流感病毒t细胞抗原表位肽及其应用 |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN117430665A CN117430665A (zh) | 2024-01-23 |
| CN117430665B true CN117430665B (zh) | 2024-06-04 |
Family
ID=89556323
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202311379347.7A Active CN117430665B (zh) | 2023-10-24 | 2023-10-24 | 甲型流感病毒t细胞抗原表位肽及其应用 |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN117430665B (zh) |
Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| KR20080093382A (ko) * | 2007-04-16 | 2008-10-21 | 주식회사 제넥신 | 독감 예방용 백신 조성물 |
| CN101395181A (zh) * | 2006-09-15 | 2009-03-25 | 美国弗劳恩霍夫股份有限公司 | 流感抗体、组合物和相关方法 |
| WO2011046996A2 (en) * | 2009-10-13 | 2011-04-21 | The Johns Hopkins University | Consensus sequence for influenza a virus |
| CN102741295A (zh) * | 2009-03-10 | 2012-10-17 | 贝勒研究院 | 靶向抗原呈递细胞的抗病毒疫苗 |
| CN110290805A (zh) * | 2017-01-03 | 2019-09-27 | 埃默杰克斯疫苗控股有限公司 | 通用流感疫苗组合物 |
| CN115667302A (zh) * | 2020-02-06 | 2023-01-31 | 亚洲大学校产学协力团 | 将抗原衍生t细胞抗原表位或含所述表位的肽呈递到细胞表面的融合抗体及含所述融合抗体的组合物 |
| CN116396366A (zh) * | 2022-02-24 | 2023-07-07 | 暨南大学 | 一种新型冠状病毒t细胞抗原表位肽及其在制备疫苗中的应用 |
-
2023
- 2023-10-24 CN CN202311379347.7A patent/CN117430665B/zh active Active
Patent Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101395181A (zh) * | 2006-09-15 | 2009-03-25 | 美国弗劳恩霍夫股份有限公司 | 流感抗体、组合物和相关方法 |
| KR20080093382A (ko) * | 2007-04-16 | 2008-10-21 | 주식회사 제넥신 | 독감 예방용 백신 조성물 |
| CN102741295A (zh) * | 2009-03-10 | 2012-10-17 | 贝勒研究院 | 靶向抗原呈递细胞的抗病毒疫苗 |
| WO2011046996A2 (en) * | 2009-10-13 | 2011-04-21 | The Johns Hopkins University | Consensus sequence for influenza a virus |
| CN110290805A (zh) * | 2017-01-03 | 2019-09-27 | 埃默杰克斯疫苗控股有限公司 | 通用流感疫苗组合物 |
| CN115667302A (zh) * | 2020-02-06 | 2023-01-31 | 亚洲大学校产学协力团 | 将抗原衍生t细胞抗原表位或含所述表位的肽呈递到细胞表面的融合抗体及含所述融合抗体的组合物 |
| CN116396366A (zh) * | 2022-02-24 | 2023-07-07 | 暨南大学 | 一种新型冠状病毒t细胞抗原表位肽及其在制备疫苗中的应用 |
Also Published As
| Publication number | Publication date |
|---|---|
| CN117430665A (zh) | 2024-01-23 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN114751965B (zh) | 一种新型冠状病毒t细胞抗原表位肽及其在制备疫苗中的应用 | |
| CN111548396B (zh) | 一种新型冠状病毒T细胞抗原表位肽、pMHC及其制备和应用 | |
| CN112876542B (zh) | 一种新型冠状病毒t细胞的抗原表位肽及其应用 | |
| Crowley et al. | Boosting of cross-reactive antibodies to endemic coronaviruses by SARS-CoV-2 infection but not vaccination with stabilized spike | |
| Manigold et al. | Highly differentiated, resting gn-specific memory CD8+ T cells persist years after infection by andes hantavirus | |
| Aspinall et al. | Immunity in the elderly: the role of the thymus | |
| Crough et al. | Contemporaneous fluctuations in T cell responses to persistent herpes virus infections | |
| De Wolf et al. | An Arthritis‐Suppressive and Treg Cell–Inducing CD4+ T Cell Epitope Is Functional in the Context of HLA‐Restricted T Cell Responses | |
| Lohia et al. | Identification of conserved peptides comprising multiple T cell epitopes of matrix 1 protein in H1N1 influenza virus | |
| CN106397574A (zh) | 抗原表位肽及其应用 | |
| Selvavinayagam et al. | Factors associated with the decay of anti-SARS-CoV-2 S1 IgG antibodies among recipients of an adenoviral vector-based AZD1222 and a whole-virion inactivated BBV152 vaccine | |
| Birnbaum et al. | Heat shock proteins and experimental autoimmune encephalomyelitis: II: environmental infection and extra-neuraxial inflammation alter the course of chronic relapsing encephalomyelitis | |
| CN117430665B (zh) | 甲型流感病毒t细胞抗原表位肽及其应用 | |
| CN108440656B (zh) | 结核分枝杆菌pka蛋白及其融合蛋白在辅助诊断肺结核病中的应用 | |
| CN117430664B (zh) | 一种甲型流感病毒t细胞抗原表位肽及其应用 | |
| US20210338806A1 (en) | METHODS OF GENERATING VACCINES AGAINST NOVEL CORONAVIRUS, NAMED SARS-COV-2 COMPRISING VARIABLE EPITOPE LIBRARIES (VELs) AS IMMUNOGENS | |
| CN117402214B (zh) | 甲型流感病毒cd8+t细胞抗原表位肽及其应用 | |
| CN117402213B (zh) | 一种甲型流感病毒cd8+t细胞抗原表位肽及其应用 | |
| CN101370513A (zh) | 由肿瘤抗原衍生的优化隐蔽性肽组成的免疫原性多肽及其用途 | |
| WO2014165866A2 (en) | Methods for immune-based diagnosis, prevention and personalized treatment of narcolepsy | |
| Saruhan-Direskeneli et al. | Human HSP 60 peptide responsive T cell lines are similarly present in both Behcet's disease patients and healthy controls | |
| Hayney | Pharmacogenomics and infectious diseases: impact on drug response and applications to disease management | |
| US20040141995A1 (en) | MHC class I-restricted and MHC class II-restricted EBNA1 peptides | |
| EP4145132A1 (en) | Methods and kit for determining the antibody status and t-cell immunity against sars-cov-2 | |
| Yajima et al. | SARS-CoV-2 spike protein-derived immunogenic peptides that are promiscuously presented by several HLA-class II molecules and their potential for inducing acquired immunity |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| CB03 | Change of inventor or designer information |
Inventor after: Chen Guobing Inventor after: Long Ze Inventor after: Xiao Chanchan Inventor after: Li Shumin Inventor after: Ren Zhiyao Inventor after: Gao Wen Inventor after: Jiang Wei Inventor after: Luo Junhong Inventor after: Wang Pengcheng Inventor after: Gao Lijuan Inventor before: Xiao Chanchan Inventor before: Long Ze Inventor before: Chen Guobing Inventor before: Li Shumin Inventor before: Ren Zhiyao Inventor before: Gao Wen Inventor before: Jiang Wei Inventor before: Luo Junhong Inventor before: Wang Pengcheng Inventor before: Gao Lijuan |
|
| CB03 | Change of inventor or designer information | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant |