CN114081943B - 一种水痘-带状疱疹mRNA疫苗组合物及其制备方法和应用 - Google Patents
一种水痘-带状疱疹mRNA疫苗组合物及其制备方法和应用 Download PDFInfo
- Publication number
- CN114081943B CN114081943B CN202111373707.3A CN202111373707A CN114081943B CN 114081943 B CN114081943 B CN 114081943B CN 202111373707 A CN202111373707 A CN 202111373707A CN 114081943 B CN114081943 B CN 114081943B
- Authority
- CN
- China
- Prior art keywords
- varicella
- zoster
- mrna
- vaccine composition
- vaccine
- 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
- 208000007514 Herpes zoster Diseases 0.000 title claims abstract description 54
- 239000000203 mixture Substances 0.000 title claims abstract description 50
- 108700021021 mRNA Vaccine Proteins 0.000 title claims abstract description 42
- 229940126582 mRNA vaccine Drugs 0.000 title claims abstract description 42
- 238000002360 preparation method Methods 0.000 title claims abstract description 12
- 229960005486 vaccine Drugs 0.000 claims abstract description 33
- 101900123149 Varicella-zoster virus Envelope glycoprotein E Proteins 0.000 claims abstract description 19
- 150000002632 lipids Chemical class 0.000 claims abstract description 7
- 239000002105 nanoparticle Substances 0.000 claims abstract description 7
- 229920002477 rna polymer Polymers 0.000 claims abstract description 3
- 239000000243 solution Substances 0.000 claims description 21
- 125000003275 alpha amino acid group Chemical group 0.000 claims description 11
- 239000000872 buffer Substances 0.000 claims description 10
- 108090000623 proteins and genes Proteins 0.000 claims description 10
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 claims description 9
- 108091032973 (ribonucleotides)n+m Proteins 0.000 claims description 8
- 238000000034 method Methods 0.000 claims description 6
- 230000035772 mutation Effects 0.000 claims description 6
- 238000000746 purification Methods 0.000 claims description 5
- 239000000758 substrate Substances 0.000 claims description 5
- 238000013518 transcription Methods 0.000 claims description 5
- NRJAVPSFFCBXDT-HUESYALOSA-N 1,2-distearoyl-sn-glycero-3-phosphocholine Chemical compound CCCCCCCCCCCCCCCCCC(=O)OC[C@H](COP([O-])(=O)OCC[N+](C)(C)C)OC(=O)CCCCCCCCCCCCCCCCC NRJAVPSFFCBXDT-HUESYALOSA-N 0.000 claims description 4
- 108091026898 Leader sequence (mRNA) Proteins 0.000 claims description 4
- 108091036066 Three prime untranslated region Proteins 0.000 claims description 4
- 238000007905 drug manufacturing Methods 0.000 claims description 4
- 238000000338 in vitro Methods 0.000 claims description 4
- 238000002347 injection Methods 0.000 claims description 4
- 239000007924 injection Substances 0.000 claims description 4
- 230000003834 intracellular effect Effects 0.000 claims description 4
- 238000010255 intramuscular injection Methods 0.000 claims description 4
- 239000007927 intramuscular injection Substances 0.000 claims description 4
- 238000004519 manufacturing process Methods 0.000 claims description 4
- 239000012528 membrane Substances 0.000 claims description 4
- 230000008488 polyadenylation Effects 0.000 claims description 4
- 239000000126 substance Substances 0.000 claims description 4
- 230000035897 transcription Effects 0.000 claims description 4
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 claims description 3
- 239000007853 buffer solution Substances 0.000 claims description 3
- 125000003178 carboxy group Chemical group [H]OC(*)=O 0.000 claims description 3
- 238000005457 optimization Methods 0.000 claims description 3
- 230000001954 sterilising effect Effects 0.000 claims description 3
- 238000010254 subcutaneous injection Methods 0.000 claims description 3
- 239000007929 subcutaneous injection Substances 0.000 claims description 3
- 206010036376 Postherpetic Neuralgia Diseases 0.000 claims description 2
- 230000015572 biosynthetic process Effects 0.000 claims description 2
- 238000003776 cleavage reaction Methods 0.000 claims description 2
- 239000003814 drug Substances 0.000 claims description 2
- 239000013612 plasmid Substances 0.000 claims description 2
- 230000007017 scission Effects 0.000 claims description 2
- 238000003786 synthesis reaction Methods 0.000 claims description 2
- 238000005303 weighing Methods 0.000 claims description 2
- 108020004705 Codon Proteins 0.000 claims 1
- 230000002265 prevention Effects 0.000 claims 1
- 230000024932 T cell mediated immunity Effects 0.000 abstract description 6
- 239000002245 particle Substances 0.000 abstract description 6
- 229940021648 varicella vaccine Drugs 0.000 abstract description 5
- 101000807236 Human cytomegalovirus (strain AD169) Membrane glycoprotein US3 Proteins 0.000 abstract description 3
- 208000032420 Latent Infection Diseases 0.000 abstract description 2
- 208000037771 disease arising from reactivation of latent virus Diseases 0.000 abstract description 2
- 230000028996 humoral immune response Effects 0.000 abstract description 2
- 230000000052 comparative effect Effects 0.000 description 23
- 210000004027 cell Anatomy 0.000 description 18
- 210000004989 spleen cell Anatomy 0.000 description 17
- 108091007433 antigens Proteins 0.000 description 14
- 102000036639 antigens Human genes 0.000 description 14
- 239000000427 antigen Substances 0.000 description 11
- 230000004048 modification Effects 0.000 description 9
- 238000012986 modification Methods 0.000 description 9
- 238000005406 washing Methods 0.000 description 8
- 230000000694 effects Effects 0.000 description 7
- 150000007523 nucleic acids Chemical group 0.000 description 7
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 6
- HVYWMOMLDIMFJA-DPAQBDIFSA-N cholesterol Chemical compound C1C=C2C[C@@H](O)CC[C@]2(C)[C@@H]2[C@@H]1[C@@H]1CC[C@H]([C@H](C)CCCC(C)C)[C@@]1(C)CC2 HVYWMOMLDIMFJA-DPAQBDIFSA-N 0.000 description 6
- 239000002773 nucleotide Substances 0.000 description 6
- 125000003729 nucleotide group Chemical group 0.000 description 6
- 102000004169 proteins and genes Human genes 0.000 description 6
- 102000005962 receptors Human genes 0.000 description 6
- 108020003175 receptors Proteins 0.000 description 6
- 230000003248 secreting effect Effects 0.000 description 6
- 238000002255 vaccination Methods 0.000 description 6
- 108010002350 Interleukin-2 Proteins 0.000 description 5
- AFCARXCZXQIEQB-UHFFFAOYSA-N N-[3-oxo-3-(2,4,6,7-tetrahydrotriazolo[4,5-c]pyridin-5-yl)propyl]-2-[[3-(trifluoromethoxy)phenyl]methylamino]pyrimidine-5-carboxamide Chemical compound O=C(CCNC(=O)C=1C=NC(=NC=1)NCC1=CC(=CC=C1)OC(F)(F)F)N1CC2=C(CC1)NN=N2 AFCARXCZXQIEQB-UHFFFAOYSA-N 0.000 description 5
- 238000001514 detection method Methods 0.000 description 5
- 238000010790 dilution Methods 0.000 description 5
- 239000012895 dilution Substances 0.000 description 5
- 239000007788 liquid Substances 0.000 description 5
- 102100032912 CD44 antigen Human genes 0.000 description 4
- 101000868273 Homo sapiens CD44 antigen Proteins 0.000 description 4
- 101001018097 Homo sapiens L-selectin Proteins 0.000 description 4
- 241000701085 Human alphaherpesvirus 3 Species 0.000 description 4
- 102100037850 Interferon gamma Human genes 0.000 description 4
- 108010074328 Interferon-gamma Proteins 0.000 description 4
- 102100033467 L-selectin Human genes 0.000 description 4
- 241001465754 Metazoa Species 0.000 description 4
- 241000699670 Mus sp. Species 0.000 description 4
- DRTQHJPVMGBUCF-XVFCMESISA-N Uridine Chemical compound O[C@@H]1[C@H](O)[C@@H](CO)O[C@H]1N1C(=O)NC(=O)C=C1 DRTQHJPVMGBUCF-XVFCMESISA-N 0.000 description 4
- 230000002238 attenuated effect Effects 0.000 description 4
- 230000000903 blocking effect Effects 0.000 description 4
- 230000003053 immunization Effects 0.000 description 4
- 238000002649 immunization Methods 0.000 description 4
- 108020004707 nucleic acids Proteins 0.000 description 4
- 102000039446 nucleic acids Human genes 0.000 description 4
- 244000052769 pathogen Species 0.000 description 4
- 102000004190 Enzymes Human genes 0.000 description 3
- 108090000790 Enzymes Proteins 0.000 description 3
- 108091028043 Nucleic acid sequence Proteins 0.000 description 3
- 239000006180 TBST buffer Substances 0.000 description 3
- 241000700605 Viruses Species 0.000 description 3
- 239000002671 adjuvant Substances 0.000 description 3
- 238000003556 assay Methods 0.000 description 3
- 238000005119 centrifugation Methods 0.000 description 3
- 235000012000 cholesterol Nutrition 0.000 description 3
- 239000011248 coating agent Substances 0.000 description 3
- 238000000576 coating method Methods 0.000 description 3
- 238000005538 encapsulation Methods 0.000 description 3
- 238000003114 enzyme-linked immunosorbent spot assay Methods 0.000 description 3
- 239000002502 liposome Substances 0.000 description 3
- 230000008823 permeabilization Effects 0.000 description 3
- 238000002791 soaking Methods 0.000 description 3
- 210000000952 spleen Anatomy 0.000 description 3
- 238000010186 staining Methods 0.000 description 3
- 239000006228 supernatant Substances 0.000 description 3
- 239000000725 suspension Substances 0.000 description 3
- 239000011534 wash buffer Substances 0.000 description 3
- 102000040650 (ribonucleotides)n+m Human genes 0.000 description 2
- 102100037435 Antiviral innate immune response receptor RIG-I Human genes 0.000 description 2
- 108091008875 B cell receptors Proteins 0.000 description 2
- 238000002965 ELISA Methods 0.000 description 2
- LYCAIKOWRPUZTN-UHFFFAOYSA-N Ethylene glycol Chemical compound OCCO LYCAIKOWRPUZTN-UHFFFAOYSA-N 0.000 description 2
- 101000669402 Homo sapiens Toll-like receptor 7 Proteins 0.000 description 2
- 206010061598 Immunodeficiency Diseases 0.000 description 2
- 208000029462 Immunodeficiency disease Diseases 0.000 description 2
- 108010014726 Interferon Type I Proteins 0.000 description 2
- 102000002227 Interferon Type I Human genes 0.000 description 2
- 102000004889 Interleukin-6 Human genes 0.000 description 2
- 108090001005 Interleukin-6 Proteins 0.000 description 2
- 102000003960 Ligases Human genes 0.000 description 2
- 108090000364 Ligases Proteins 0.000 description 2
- XUMBMVFBXHLACL-UHFFFAOYSA-N Melanin Chemical compound O=C1C(=O)C(C2=CNC3=C(C(C(=O)C4=C32)=O)C)=C2C4=CNC2=C1C XUMBMVFBXHLACL-UHFFFAOYSA-N 0.000 description 2
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 description 2
- DBMJMQXJHONAFJ-UHFFFAOYSA-M Sodium laurylsulphate Chemical compound [Na+].CCCCCCCCCCCCOS([O-])(=O)=O DBMJMQXJHONAFJ-UHFFFAOYSA-M 0.000 description 2
- 102000002689 Toll-like receptor Human genes 0.000 description 2
- 108020000411 Toll-like receptor Proteins 0.000 description 2
- 102100039390 Toll-like receptor 7 Human genes 0.000 description 2
- 108060008682 Tumor Necrosis Factor Proteins 0.000 description 2
- 102000000852 Tumor Necrosis Factor-alpha Human genes 0.000 description 2
- 108091023045 Untranslated Region Proteins 0.000 description 2
- SIIZPVYVXNXXQG-KGXOGWRBSA-N [(2r,3r,4r,5r)-5-(6-aminopurin-9-yl)-4-[[(3s,4r)-5-(6-aminopurin-9-yl)-3,4-dihydroxyoxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-3-hydroxyoxolan-2-yl]methyl [(2r,4r,5r)-2-(6-aminopurin-9-yl)-4-hydroxy-5-(phosphonooxymethyl)oxolan-3-yl] hydrogen phosphate Polymers C1=NC2=C(N)N=CN=C2N1[C@@H]1O[C@H](COP(O)(=O)OC2[C@@H](O[C@H](COP(O)(O)=O)[C@H]2O)N2C3=NC=NC(N)=C3N=C2)[C@@H](O)[C@H]1OP(O)(=O)OCC([C@@H](O)[C@H]1O)OC1N1C(N=CN=C2N)=C2N=C1 SIIZPVYVXNXXQG-KGXOGWRBSA-N 0.000 description 2
- ZMJPCIAEJKVKMQ-UHFFFAOYSA-M [4-[[4-[benzyl(methyl)amino]phenyl]-[4-(dimethylamino)phenyl]methylidene]cyclohexa-2,5-dien-1-ylidene]-dimethylazanium;chloride Chemical compound [Cl-].C1=CC(N(C)C)=CC=C1C(C=1C=CC(=CC=1)N(C)CC=1C=CC=CC=1)=C1C=CC(=[N+](C)C)C=C1 ZMJPCIAEJKVKMQ-UHFFFAOYSA-M 0.000 description 2
- SHGAZHPCJJPHSC-YCNIQYBTSA-N all-trans-retinoic acid Chemical compound OC(=O)\C=C(/C)\C=C\C=C(/C)\C=C\C1=C(C)CCCC1(C)C SHGAZHPCJJPHSC-YCNIQYBTSA-N 0.000 description 2
- DRTQHJPVMGBUCF-PSQAKQOGSA-N beta-L-uridine Natural products O[C@H]1[C@@H](O)[C@H](CO)O[C@@H]1N1C(=O)NC(=O)C=C1 DRTQHJPVMGBUCF-PSQAKQOGSA-N 0.000 description 2
- -1 but not limited to Substances 0.000 description 2
- 238000006243 chemical reaction Methods 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 210000000805 cytoplasm Anatomy 0.000 description 2
- 210000001151 cytotoxic T lymphocyte Anatomy 0.000 description 2
- 239000008367 deionised water Substances 0.000 description 2
- 229910021641 deionized water Inorganic materials 0.000 description 2
- 230000004069 differentiation Effects 0.000 description 2
- 239000003085 diluting agent Substances 0.000 description 2
- 230000004727 humoral immunity Effects 0.000 description 2
- 230000001900 immune effect Effects 0.000 description 2
- 230000007813 immunodeficiency Effects 0.000 description 2
- 230000001939 inductive effect Effects 0.000 description 2
- 229940100601 interleukin-6 Drugs 0.000 description 2
- 210000004698 lymphocyte Anatomy 0.000 description 2
- 235000013336 milk Nutrition 0.000 description 2
- 239000008267 milk Substances 0.000 description 2
- 210000004080 milk Anatomy 0.000 description 2
- 230000001717 pathogenic effect Effects 0.000 description 2
- 230000037361 pathway Effects 0.000 description 2
- 229920001223 polyethylene glycol Polymers 0.000 description 2
- 239000000047 product Substances 0.000 description 2
- 229930002330 retinoic acid Natural products 0.000 description 2
- 210000002966 serum Anatomy 0.000 description 2
- 230000011664 signaling Effects 0.000 description 2
- 238000007619 statistical method Methods 0.000 description 2
- 229940031626 subunit vaccine Drugs 0.000 description 2
- 238000013519 translation Methods 0.000 description 2
- 229960001727 tretinoin Drugs 0.000 description 2
- DRTQHJPVMGBUCF-UHFFFAOYSA-N uracil arabinoside Natural products OC1C(O)C(CO)OC1N1C(=O)NC(=O)C=C1 DRTQHJPVMGBUCF-UHFFFAOYSA-N 0.000 description 2
- 229940045145 uridine Drugs 0.000 description 2
- 230000009385 viral infection Effects 0.000 description 2
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 2
- 125000000954 2-hydroxyethyl group Chemical group [H]C([*])([H])C([H])([H])O[H] 0.000 description 1
- SXIFAEWFOJETOA-UHFFFAOYSA-N 4-hydroxy-butyl Chemical group [CH2]CCCO SXIFAEWFOJETOA-UHFFFAOYSA-N 0.000 description 1
- GOZMBJCYMQQACI-UHFFFAOYSA-N 6,7-dimethyl-3-[[methyl-[2-[methyl-[[1-[3-(trifluoromethyl)phenyl]indol-3-yl]methyl]amino]ethyl]amino]methyl]chromen-4-one;dihydrochloride Chemical compound Cl.Cl.C=1OC2=CC(C)=C(C)C=C2C(=O)C=1CN(C)CCN(C)CC(C1=CC=CC=C11)=CN1C1=CC=CC(C(F)(F)F)=C1 GOZMBJCYMQQACI-UHFFFAOYSA-N 0.000 description 1
- 101710127675 Antiviral innate immune response receptor RIG-I Proteins 0.000 description 1
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 1
- HSBMBJDVXFRTTD-UHFFFAOYSA-N C(CCCCC)C(C(=O)OCCCCCCOC(C(CCCCCCCC)CCCCCC)=O)CCCCCCCC Chemical compound C(CCCCC)C(C(=O)OCCCCCCOC(C(CCCCCCCC)CCCCCC)=O)CCCCCCCC HSBMBJDVXFRTTD-UHFFFAOYSA-N 0.000 description 1
- 238000011740 C57BL/6 mouse Methods 0.000 description 1
- 241000283707 Capra Species 0.000 description 1
- 201000006082 Chickenpox Diseases 0.000 description 1
- 108700010070 Codon Usage Proteins 0.000 description 1
- 241000699802 Cricetulus griseus Species 0.000 description 1
- 102000004127 Cytokines Human genes 0.000 description 1
- 108090000695 Cytokines Proteins 0.000 description 1
- 108010041986 DNA Vaccines Proteins 0.000 description 1
- 229940021995 DNA vaccine Drugs 0.000 description 1
- 102000010834 Extracellular Matrix Proteins Human genes 0.000 description 1
- 108010037362 Extracellular Matrix Proteins Proteins 0.000 description 1
- 108010087819 Fc receptors Proteins 0.000 description 1
- 102000009109 Fc receptors Human genes 0.000 description 1
- 208000003098 Ganglion Cysts Diseases 0.000 description 1
- 101800001586 Ghrelin Proteins 0.000 description 1
- 102400000442 Ghrelin-28 Human genes 0.000 description 1
- 108090000288 Glycoproteins Proteins 0.000 description 1
- 102000003886 Glycoproteins Human genes 0.000 description 1
- 208000031957 HIV carrier Diseases 0.000 description 1
- 101000952099 Homo sapiens Antiviral innate immune response receptor RIG-I Proteins 0.000 description 1
- 101000917858 Homo sapiens Low affinity immunoglobulin gamma Fc region receptor III-A Proteins 0.000 description 1
- 101000917839 Homo sapiens Low affinity immunoglobulin gamma Fc region receptor III-B Proteins 0.000 description 1
- 101000904196 Homo sapiens Pancreatic secretory granule membrane major glycoprotein GP2 Proteins 0.000 description 1
- 102000008986 Janus Human genes 0.000 description 1
- 108050000950 Janus Proteins 0.000 description 1
- 102000015617 Janus Kinases Human genes 0.000 description 1
- 108010024121 Janus Kinases Proteins 0.000 description 1
- 102100029185 Low affinity immunoglobulin gamma Fc region receptor III-B Human genes 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
- 101150053046 MYD88 gene Proteins 0.000 description 1
- 108700018351 Major Histocompatibility Complex Proteins 0.000 description 1
- 241001092142 Molina Species 0.000 description 1
- 102100024019 Pancreatic secretory granule membrane major glycoprotein GP2 Human genes 0.000 description 1
- 239000002202 Polyethylene glycol Substances 0.000 description 1
- 108091036414 Polyinosinic:polycytidylic acid Proteins 0.000 description 1
- 229920001213 Polysorbate 20 Polymers 0.000 description 1
- 229930185560 Pseudouridine Natural products 0.000 description 1
- PTJWIQPHWPFNBW-UHFFFAOYSA-N Pseudouridine C Natural products OC1C(O)C(CO)OC1C1=CNC(=O)NC1=O PTJWIQPHWPFNBW-UHFFFAOYSA-N 0.000 description 1
- 241001454523 Quillaja saponaria Species 0.000 description 1
- 235000009001 Quillaja saponaria Nutrition 0.000 description 1
- 208000005400 Synovial Cyst Diseases 0.000 description 1
- 108091008874 T cell receptors Proteins 0.000 description 1
- 102000016266 T-Cell Antigen Receptors Human genes 0.000 description 1
- 102000008230 Toll-like receptor 3 Human genes 0.000 description 1
- 108010060885 Toll-like receptor 3 Proteins 0.000 description 1
- 206010046980 Varicella Diseases 0.000 description 1
- 229940124925 Zostavax Drugs 0.000 description 1
- 238000002835 absorbance Methods 0.000 description 1
- 230000003213 activating effect Effects 0.000 description 1
- 239000012190 activator Substances 0.000 description 1
- 230000033289 adaptive immune response Effects 0.000 description 1
- 239000011543 agarose gel Substances 0.000 description 1
- 238000007605 air drying Methods 0.000 description 1
- XAGFODPZIPBFFR-UHFFFAOYSA-N aluminium Chemical compound [Al] XAGFODPZIPBFFR-UHFFFAOYSA-N 0.000 description 1
- 229910052782 aluminium Inorganic materials 0.000 description 1
- 229910000147 aluminium phosphate Inorganic materials 0.000 description 1
- 150000001413 amino acids Chemical class 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 238000010171 animal model Methods 0.000 description 1
- 230000000840 anti-viral effect Effects 0.000 description 1
- 230000003305 autocrine Effects 0.000 description 1
- 210000003719 b-lymphocyte Anatomy 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- WGDUUQDYDIIBKT-UHFFFAOYSA-N beta-Pseudouridine Natural products OC1OC(CN2C=CC(=O)NC2=O)C(O)C1O WGDUUQDYDIIBKT-UHFFFAOYSA-N 0.000 description 1
- 230000037396 body weight Effects 0.000 description 1
- KQNZDYYTLMIZCT-KQPMLPITSA-N brefeldin A Chemical compound O[C@@H]1\C=C\C(=O)O[C@@H](C)CCC\C=C\[C@@H]2C[C@H](O)C[C@H]21 KQNZDYYTLMIZCT-KQPMLPITSA-N 0.000 description 1
- JUMGSHROWPPKFX-UHFFFAOYSA-N brefeldin-A Natural products CC1CCCC=CC2(C)CC(O)CC2(C)C(O)C=CC(=O)O1 JUMGSHROWPPKFX-UHFFFAOYSA-N 0.000 description 1
- 230000015556 catabolic process Effects 0.000 description 1
- 125000002091 cationic group Chemical group 0.000 description 1
- 238000004113 cell culture Methods 0.000 description 1
- 210000003855 cell nucleus Anatomy 0.000 description 1
- 230000001413 cellular effect Effects 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 210000004978 chinese hamster ovary cell Anatomy 0.000 description 1
- 238000006731 degradation reaction Methods 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 238000001962 electrophoresis Methods 0.000 description 1
- 230000012202 endocytosis Effects 0.000 description 1
- 230000002708 enhancing effect Effects 0.000 description 1
- 238000006911 enzymatic reaction Methods 0.000 description 1
- 210000003743 erythrocyte Anatomy 0.000 description 1
- 210000002744 extracellular matrix Anatomy 0.000 description 1
- 239000012091 fetal bovine serum Substances 0.000 description 1
- 238000000684 flow cytometry Methods 0.000 description 1
- 239000012634 fragment Substances 0.000 description 1
- GNKDKYIHGQKHHM-RJKLHVOGSA-N ghrelin Chemical compound C([C@H](NC(=O)[C@@H](NC(=O)[C@H](CO)NC(=O)CN)COC(=O)CCCCCCC)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CO)C(=O)N1[C@@H](CCC1)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC=1N=CNC=1)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CO)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CCCCN)C(=O)N1[C@@H](CCC1)C(=O)N1[C@@H](CCC1)C(=O)N[C@@H](C)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(N)=O)C(=O)N1[C@@H](CCC1)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O)C1=CC=CC=C1 GNKDKYIHGQKHHM-RJKLHVOGSA-N 0.000 description 1
- 150000002334 glycols Chemical class 0.000 description 1
- 230000013595 glycosylation Effects 0.000 description 1
- 238000006206 glycosylation reaction Methods 0.000 description 1
- 210000002443 helper t lymphocyte Anatomy 0.000 description 1
- 230000002949 hemolytic effect Effects 0.000 description 1
- 238000004128 high performance liquid chromatography Methods 0.000 description 1
- WGCNASOHLSPBMP-UHFFFAOYSA-N hydroxyacetaldehyde Natural products OCC=O WGCNASOHLSPBMP-UHFFFAOYSA-N 0.000 description 1
- 210000001822 immobilized cell Anatomy 0.000 description 1
- 230000036737 immune function Effects 0.000 description 1
- 230000028993 immune response Effects 0.000 description 1
- 230000036039 immunity Effects 0.000 description 1
- 239000003018 immunosuppressive agent Substances 0.000 description 1
- 229940124589 immunosuppressive drug Drugs 0.000 description 1
- 230000001771 impaired effect Effects 0.000 description 1
- 238000001727 in vivo Methods 0.000 description 1
- 238000011534 incubation Methods 0.000 description 1
- 230000006698 induction Effects 0.000 description 1
- 208000015181 infectious disease Diseases 0.000 description 1
- 239000003112 inhibitor Substances 0.000 description 1
- 230000015788 innate immune response Effects 0.000 description 1
- 239000002054 inoculum Substances 0.000 description 1
- 230000010354 integration Effects 0.000 description 1
- 230000003993 interaction Effects 0.000 description 1
- 229960001201 live attenuated varicella Drugs 0.000 description 1
- 238000011068 loading method Methods 0.000 description 1
- 239000006166 lysate Substances 0.000 description 1
- 201000001441 melanoma Diseases 0.000 description 1
- 230000002503 metabolic effect Effects 0.000 description 1
- 230000037353 metabolic pathway Effects 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 239000000178 monomer Substances 0.000 description 1
- WWZKQHOCKIZLMA-UHFFFAOYSA-M octanoate Chemical compound CCCCCCCC([O-])=O WWZKQHOCKIZLMA-UHFFFAOYSA-M 0.000 description 1
- 238000001543 one-way ANOVA Methods 0.000 description 1
- 210000001672 ovary Anatomy 0.000 description 1
- 230000003076 paracrine Effects 0.000 description 1
- 108010089193 pattern recognition receptors Proteins 0.000 description 1
- 102000007863 pattern recognition receptors Human genes 0.000 description 1
- 239000000256 polyoxyethylene sorbitan monolaurate Substances 0.000 description 1
- 235000010486 polyoxyethylene sorbitan monolaurate Nutrition 0.000 description 1
- 229920001184 polypeptide Polymers 0.000 description 1
- 239000005017 polysaccharide Substances 0.000 description 1
- 230000029279 positive regulation of transcription, DNA-dependent Effects 0.000 description 1
- 230000004481 post-translational protein modification Effects 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 description 1
- 102000004196 processed proteins & peptides Human genes 0.000 description 1
- 108090000765 processed proteins & peptides Proteins 0.000 description 1
- 230000000770 proinflammatory effect Effects 0.000 description 1
- PTJWIQPHWPFNBW-GBNDHIKLSA-N pseudouridine Chemical compound O[C@@H]1[C@H](O)[C@@H](CO)O[C@H]1C1=CNC(=O)NC1=O PTJWIQPHWPFNBW-GBNDHIKLSA-N 0.000 description 1
- 238000003908 quality control method Methods 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 230000019491 signal transduction Effects 0.000 description 1
- 235000020183 skimmed milk Nutrition 0.000 description 1
- 230000000087 stabilizing effect Effects 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
- 230000032258 transport Effects 0.000 description 1
- 239000012646 vaccine adjuvant Substances 0.000 description 1
- 229940124931 vaccine adjuvant Drugs 0.000 description 1
- 230000003612 virological effect Effects 0.000 description 1
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
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P25/00—Drugs for disorders of the nervous system
-
- 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
- A61P31/20—Antivirals for DNA viruses
- A61P31/22—Antivirals for DNA viruses for herpes viruses
-
- 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
- A61K2039/53—DNA (RNA) vaccination
-
- 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
- A61K2039/54—Medicinal preparations containing antigens or antibodies characterised by the route of administration
-
- 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
- A61K2039/57—Medicinal preparations containing antigens or antibodies characterised by the type of response, e.g. Th1, Th2
-
- 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
- A61K2039/57—Medicinal preparations containing antigens or antibodies characterised by the type of response, e.g. Th1, Th2
- A61K2039/575—Medicinal preparations containing antigens or antibodies characterised by the type of response, e.g. Th1, Th2 humoral response
-
- 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
- C12N2710/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA dsDNA viruses
- C12N2710/00011—Details
- C12N2710/16011—Herpesviridae
- C12N2710/16711—Varicellovirus, e.g. human herpesvirus 3, Varicella Zoster, pseudorabies
- C12N2710/16734—Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Virology (AREA)
- Animal Behavior & Ethology (AREA)
- Chemical & Material Sciences (AREA)
- Medicinal Chemistry (AREA)
- Veterinary Medicine (AREA)
- Public Health (AREA)
- Pharmacology & Pharmacy (AREA)
- General Health & Medical Sciences (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Organic Chemistry (AREA)
- General Chemical & Material Sciences (AREA)
- Engineering & Computer Science (AREA)
- Biotechnology (AREA)
- Mycology (AREA)
- Communicable Diseases (AREA)
- Oncology (AREA)
- Epidemiology (AREA)
- Microbiology (AREA)
- Immunology (AREA)
- Molecular Biology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Biomedical Technology (AREA)
- Neurology (AREA)
- Neurosurgery (AREA)
- Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
- Peptides Or Proteins (AREA)
Abstract
本发明提供了一种水痘‑带状疱疹mRNA疫苗组合物及其制备方法和应用,所述疫苗组合物中包含编码水痘‑带状疱疹病毒糖蛋白E的信使核糖核酸(mRNA)序列及其衍生序列和脂质纳米颗粒(LNP),通过微流控设备制备为直径20‑400纳米的颗粒。所述疫苗组合物能特异性增强针对水痘‑带状疱疹糖蛋白E的体液免疫应答和细胞免疫应答,可用作不会导致疫苗株潜伏感染的水痘疫苗,亦可用作产能不受限制的带状疱疹疫苗。所述疫苗组合物中各成分均可广泛获取,有效降低了疫苗成本、提高疫苗产量。
Description
技术领域
本发明属于疫苗领域,具体涉及一种水痘-带状疱疹mRNA疫苗组合物及其制备方法和应用。
背景技术
几乎所有儿童成年前均感染过水痘-带状疱疹病毒(Varicella-Zoster Virus,VZV)。初次感染可能产生水痘,水痘自愈后病毒会潜伏在神经节内。伴随年龄老化或其他原因导致的细胞免疫应答削弱(如免疫缺陷病毒感染或临床需要使用免疫抑制药物),潜伏的病毒会在体内再活化,进而导致带状疱疹的发生。
由日本人高桥理明(Michiaki Takahashi)开发的Oka株减毒活疫苗1995年被FDA批准用于接种儿童和成人预防水痘(接种量1 000-5 000 PFU <空斑形成单位,plaqueforming unit>),之后广泛应用于全世界。后续的研究发现,Oka株和野生型病毒一样会建立潜伏感染,进而同样可能导致带状疱疹的发生。因此,未来需要一种更安全的水痘疫苗,通过诱导有效的体液免疫应答替代现有的水痘减毒活疫苗。
默克公司(Merk)2005年上市的减毒活疫苗Zostavax通过对既往感染VZV病毒的50岁以上人群一次性皮下注射高剂量Oka株(接种量19400 PFU)预防带状疱疹,单针售价约200美元。该疫苗对50-59,60-69及70岁以上人群的保护率分别为70%,64%和38%左右。另外,由于相对于制备低滴度的水痘疫苗,技术方面制备和储存高病毒滴度的带状疱疹减毒活疫苗技术难度较大,国内疫苗企业尚无相关产品上市。
葛兰素史克(GSK)2017年底上市的带状疱疹亚单位疫苗Shingrix使用中国仓鼠卵巢细胞(Chinese hamster ovary, CHO)表达保守性病毒糖蛋白E(gE)作为抗原,使用佐剂AS01B有效增强针对VZV-gE特异的细胞免疫应答,临床上对60岁以上各年龄段人群的保护率均高于90%,且在包括HIV携带者在内的免疫缺陷人群中表现出了良好的安全性和有效性。AS01B的关键成分QS21是一种仅能从南美温带地区生长的皂角树Quillaja saponaria树皮中经反向高效液相色谱提取的多糖混合物,目前无法人工合成,存在来源受限、制备过程质量控制难度大(纯化组分为成分复杂的非单体、活性成分对温度敏感)、因具有溶血活性需要加入脱毒剂等局限性。另外根据Shingrix的使用说明,其中的佐剂成分AS01B在使用前需同抗原临时混合(Bedside mix),间接提示该脂质体系统应用于疫苗时稳定性有待进一步提高。以上背景导致Shingrix售价昂贵(150-200美元每针剂,全程接种2针剂),但仍供不应求(2020年84.2%的疫苗在美国本土销售,目前美国以外基本有价无市)。
因此,如何进一步降低水痘疫苗接种导致罹患带状疱疹的风险,并增强细胞免疫应答用作带状疱疹疫苗,是亟需解决的问题。
发明内容
针对现有技术中存在的问题,本发明的目的在于提供一种水痘-带状疱疹mRNA疫苗组合物及其制备方法和应用,所述疫苗组合物用作水痘疫苗时不存在因疫苗接种导致潜在罹患带状疱疹的风险;同时可有效增强针对VZV-gE特异的细胞免疫应答,在用作带状疱疹mRNA疫苗时可有效降低疫苗成本、提高疫苗产量。
本发明的目的是通过以下技术方案实现的:
一种水痘-带状疱疹mRNA疫苗组合物,所述疫苗组合物包括编码水痘-带状疱疹病毒糖蛋白E的信使核糖核酸(mRNA)序列及其衍生序列、脂质纳米颗粒(LNP)。
进一步的,所述水痘-带状疱疹病毒糖蛋白E的mRNA序列包括编码如SEQ ID No.1所示的水痘-带状疱疹病毒糖蛋白E的全长氨基酸序列、基于其衍生出的氨基酸序列的核酸序列,编码如SEQ ID No.2所示的水痘-带状疱疹病毒糖蛋白E突变衍生的氨基酸突变序列的核酸序列,编码如SEQ ID No.3所示的水痘-带状疱疹病毒糖蛋白E截取的氨基酸序列的核酸序列、及核苷酸序列优化。
SEQ ID No.1:水痘-带状疱疹病毒糖蛋白E的全长氨基酸序列(gE全长)
MGTVNKPVVG VLMGFGIITG TLRITNPVRA SVLRYDDFHI DEDKLDTNSV YEPYYHSDHA
ESSWVNRGES SRKAYDHNSP YIWPRNDYDG FLENAHEHHG VYNQGRGIDS GERLMQPTQM
SAQEDLGDDT GIHVIPTLNG DDRHKIVNVD QRQYGDVFKG DLNPKPQGQR LIEVSVEENH
PFTLRAPIQR IYGVRYTETW SFLPSLTCTG DAAPAIQHIC LKHTTCFQDV VVDVDCAENT
KEDQLAEISY RFQGKKEADQ PWIVVNTSTL FDELELDPPE IEPGVLKVLR TEKQYLGVYI
WNMRGSDGTS TYATFLVTWK GDEKTRNPTP AVTPQPRGAE FHMWNYHSHV FSVGDTFSLA
MHLQYKIHEA PFDLLLEWLY VPIDPTCQPM RLYSTCLYHP NAPQCLSHMN SGCTFTSPHL
AQRVASTVYQ NCEHADNYTA YCLGISHMEP SFGLILHDGG TTLKFVDTPE SLSGLYVFVV
YFNGHVEAVA YTVVSTVDHF VNAIEERGFP PTAGQPPATT KPKEITPVNP GTSPLLRYAA
WTGGLAAVVL LCLVIFLICT AKRMRVKAYR VDKSPYNQSM YYAGLPVDDF EDSESTDTEE
EFGNAIGGSH GGSSYTVYID KTR;
SEQ ID No.2:水痘-带状疱疹病毒糖蛋白E突变衍生的氨基酸序列(gE突变)
MGTVNKPVVG VLMGFGIITG TLRITNPVRA SVLRYDDFHI DEDKLDTNSV YEPYYHSDHA
ESSWVNRGES SRKAYDHNSP YIWPRNDYDG FLENAHEHHG VYNQGRGIDS GERLMQPTQM
SAQEDLGDDT GIHVIPTLNG DDRHKIVNVD QRQYGDVFKG DLNPKPQGQR LIEVSVEENH
PFTLRAPIQR IYGVRYTETW SFLPSLTCTG DAAPAIQHIC LKHTTCFQDV VVDVDCAENT
KEDQLAEISY RFQGKKEADQ PWIVVNTSTL FDELELDPPE IEPGVLKVLR TEKQYLGVYI
WNMRGSDGTS TYATFLVTWK GDEKTRNPTP AVTPQPRGAE FHMWNYHSHV FSVGDTFSLA
MHLQYKIHEA PFDLLLEWLY VPIDPTCQPM RLYSTCLYHP NAPQCLSHMN SGCTFTSPHL
AQRVASTVYQ NCEHADNYTA YCLGISHMEP SFGLILHDGG TTLKFVDTPE SLSGLYVFVV
YFNGHVEAVA YTVVSTVDHF VNAIEERGFP PTAGQPPATT KPKEITPVNP GTSPLLRYAA
WTGGLAAVVL LCLVIFLICT AKRMRVKAAR VDKSPYNQSM YYAGLPVDDF EDAEAADAEE
EFGNAIGGSH GGSSYTVYID KTR;
SEQ ID No.3:水痘-带状疱疹病毒糖蛋白E截取的氨基酸序列(gE胞外)
MGTVNKPVVG VLMGFGIITG TLRITNPVRA SVLRYDDFHI DEDKLDTNSV YEPYYHSDHA
ESSWVNRGES SRKAYDHNSP YIWPRNDYDG FLENAHEHHG VYNQGRGIDS GERLMQPTQM
SAQEDLGDDT GIHVIPTLNG DDRHKIVNVD QRQYGDVFKG DLNPKPQGQR LIEVSVEENH
PFTLRAPIQR IYGVRYTETW SFLPSLTCTG DAAPAIQHIC LKHTTCFQDV VVDVDCAENT
KEDQLAEISY RFQGKKEADQ PWIVVNTSTL FDELELDPPE IEPGVLKVLR TEKQYLGVYI
WNMRGSDGTS TYATFLVTWK GDEKTRNPTP AVTPQPRGAE FHMWNYHSHV FSVGDTFSLA
MHLQYKIHEA PFDLLLEWLY VPIDPTCQPM RLYSTCLYHP NAPQCLSHMN SGCTFTSPHL
AQRVASTVYQ NCEHADNYTA YCLGISHMEP SFGLILHDGG TTLKFVDTPE SLSGLYVFVV
YFNGHVEAVA YTVVSTVDHF VNAIEERGFP PTAGQPPATT KPKEITPVNP GTSPLLRY。
进一步的,所述水痘-带状疱疹病毒糖蛋白E的mRNA序列包括为实现mRNA疫苗最佳免疫效果进行的非翻译区修饰和核苷酸修饰,包括酶法和化学底物法进行的5’加帽修饰、5’及3’端非翻译区序列引入、3’端多聚腺苷酸序列引入、假尿苷替换尿苷在内的核苷酸修饰等。
进一步的,所述水痘-带状疱疹病毒糖蛋白E的mRNA用量为2.5 μg-150 μg每针剂。
进一步的,所述LNP是由包含且不限于4-(N,N-二甲基氨基)丁酸(二亚油基)甲酯(DLin-MC3-DMA)、((4-羟基丁基)氮杂二基)双(己烷-6,1-二基)双(2-己基癸酸酯)(ALC-0315)、十七烷-9-基-8-((2-羟乙基)(6-氧代-6-((癸氧基)己基)氨基)辛酸酯)(SM-102)在内的阳离子脂质体,二硬脂酰基磷脂酰胆碱(DSPC)、胆固醇(cholesterol)和1,2-二肉豆蔻酰-rac-甘油-3-甲氧基聚乙二醇2000(DMG-PEG2000)等聚乙二醇衍生物组成的混合物。
进一步的,所述疫苗组合物为直径介于20-400纳米的颗粒组合物。
进一步的,所述疫苗组合物的施用方式包括皮下或肌肉注射。
本发明的另一方面:
所述水痘-带状疱疹mRNA疫苗组合物是采用微流控设备制备得到的。
进一步的,所述制备方法包括以下步骤:将脂质纳米颗粒溶解于无水乙醇,形成溶液A;将所述编码水痘-带状疱疹病毒糖蛋白E的信使核糖核酸mRNA溶于柠檬酸缓冲液,形成溶液B;使用微流体纳米药物制造系统将所述溶液A、溶液B混合后,使用PBS缓冲液透析,过滤膜除菌后即得水痘-带状疱疹mRNA疫苗组合物。
所述水痘-带状疱疹mRNA疫苗组合物用于制备预防或改善水痘和/或带状疱疹和/或带状疱疹后神经痛的药物。
本发明相比现有技术的有益效果为:
1、本发明所述水痘-带状疱疹mRNA疫苗组合物相对于其他疫苗生产方式,mRNA可进行体外转录制备,有效降低了疫苗生产成本;其本身理化性质较为均一、生产和纯化过程非容易实现标准化操作,节省了最终抗原特异性的纯化等探索性环节,进一步节省了时间和经济成本;
2、本发明所述水痘-带状疱疹mRNA疫苗组合物内源表达的蛋白抗原相较于异源制备的抗原,进入接种者体内的mRNA翻译成的蛋白具有空间结构、翻译后修饰如糖基化的高保真度,可引起有效针对相应病原体的免疫应答、提升疫苗保护效果;
3、本发明所述水痘-带状疱疹mRNA疫苗组合物与DNA疫苗相比,mRNA疫苗翻译蛋白抗原不需要进入细胞核,在增强了抗原产量的同时避免了基因组整合的风险。另外作为一种体内常见的一过性组分,mRNA比较容易被机体的生理代谢途径清除,不会额外增加接种者机体代谢负担;
4、本发明所述水痘-带状疱疹mRNA疫苗组合物的mRNA可激活多种胞内病原相关模式受体(pathogen related receptors,PRR),具有自佐剂效应,无须额外加入疫苗佐剂。体外转录制备过程中引入的双链RNA可被胞浆中的维甲酸诱导I型基因样受体(retinoicacid-inducible gene-I-like receptors,RLR),包括视黄酸诱导基因I(retinoic-acidinducible gene I,RIG-1)和黑色素分化相关蛋白5(melanoma differentiation-associated 5,MDA5)识别。进入内吞体的RNA则可分别被Toll样受体3(Toll-likereceptor 3,TLR3,识别双链RNA),和TLR7/8(识别单链RNA)识别。激活的受体通过下游特定的接头分子信号传递(即Myd88对TLR7/8,TRIF对TLR3),最终产生I型干扰素(type Iinterferon,IFN I,包括IFN-α和IFN-β)和其他促炎细胞因子(如白细胞介素6(interleukin-6,IL-6)和肿瘤坏死因子α(tumor necrosis factor α,TNF-α))。IFN I进而通过结合自分泌或旁分泌受体,激活Janus激酶信号传导转录活化途径(Janus kinase-signal transducer activator of transcription pathway,JAK-STAT)调节数百种抗病毒免疫的蛋白质基因表达。这些信号通路激活和促进不同的固有免疫反应,进而诱导针对病原体的获得性免疫应答;
5、本发明所述的水痘-带状疱疹mRNA疫苗组合物为实现mRNA疫苗最佳免疫效果进行的非翻译区修饰和核苷酸修饰,包括酶法和化学底物法进行的5’加帽修饰、5’及3’端非翻译区序列引入、3’端多聚腺苷酸序列引入、假尿苷替换尿苷在内的核苷酸修饰等,可进一步提升mRNA稳定性,有效避免机体对外源性mRNA的识别,降低IFN I诱导的2’-5’-寡腺苷酸合成酶(2’-5’-oligoadenylate synthetase,OAS)活性进而通过调节RNaseI活性降低mRNA降解速度、提高蛋白翻译效率、增强疫苗免疫效果;
6、本发明所述水痘-带状疱疹mRNA疫苗组合物分泌表达的蛋白经B细胞受体(Bcell receptor)识别,内吞后经MHC II提呈,进而被CD4+的辅助性T细胞(helper T cell,Th)受体识别。B细胞和Th的相互作用可产生多功能、高亲和力的抗体,进而实现体液免疫,用作水痘疫苗不存在因减毒活疫苗接种导致潜在罹患带状疱疹的风险;
7、本发明所述水痘-带状疱疹mRNA疫苗组合物与外源蛋白类抗原通过效率有限的交叉提呈途径不同,mRNA在胞浆中翻译的蛋白可同病毒感染产生的异源抗原一样被充分处理为多肽并提呈给I型主要组织相容性复合体(major histocompatibility complexclass I,MHC I),进而被CD8+的T细胞受体识别。活化的CD8+细胞毒性T细胞(cytotoxic Tlymphocytes,CTL)能够选择性清除表达外来抗原的细胞如病毒感染的宿主细胞,行使细胞免疫功能用作带状疱疹疫苗;
8、本发明所述水痘-带状疱疹mRNA疫苗组合物的mRNA序列对应的氨基酸序列,通过羧基端胞内区引入突变,可进一步增强VZV-gE特异的体液免疫和细胞免疫应答效果。
附图说明
下面结合附图和实施例对本发明做进一步说明:
图1 显示经实施例1-2制备的水痘-带状疱疹mRNA疫苗组合物粒径(A),多分散性指数(B),mRNA包封率(C)和mRNA完整性(D);
图2 显示经实施例1-2制备的水痘-带状疱疹mRNA疫苗组合物和对比例1-2间隔3周2次肌肉注射实验小鼠终免2周后,诱导的抗原特异性IgG抗体滴度;
图3 显示经实施例1-2制备的水痘-带状疱疹mRNA疫苗组合物和对比例1-2间隔3周2次肌肉注射实验小鼠终免2周后,诱导产生的分泌IFN-γ(A)和IL-2的脾脏细胞数量;
图4 显示经实施例1-2制备的水痘-带状疱疹mRNA疫苗组合物和对比例1-2间隔3周2次肌肉注射实验小鼠终免2周后,诱导产生的分泌IFN-γ的CD4阳性细胞数量在所有脾细胞中所占比例(A)和诱导产生的分泌IFN-γ的CD8阳性细胞数量在所有脾细胞中所占比例(B);
图5 显示经实施例1-2制备的水痘-带状疱疹mRNA疫苗组合物和对比例1-2间隔3周2次肌肉注射实验小鼠终免2周后,诱导产生的同时表达CD44和CD62L的CD4阳性细胞数量在所有脾细胞中所占比例(A)和诱导产生的同时表达CD44和CD62L的CD8阳性细胞数量在所有脾细胞中所占比例(B)。
具体实施方式
实施例1——mRNA制备:
编码水痘-带状疱疹病毒Oka株糖蛋白E全长氨基酸序列(UniProtKB/Swiss-Prot:Q9J3M8.1,见SEQ ID No.1)、羧基端胞内区引入突变的氨基酸序列(Y569A,S593A,S595A,T596A,T598A,见SEQ ID No.2)、仅含有胞外区的氨基酸序列(见SEQ ID No.3)的基因按照哺乳动物密码子偏好优化后,连同5’非翻译区序列、3’非翻译区序列和3’端多聚腺苷酸序列交上海生工合成,构建于质粒pBlueScript II SK(+)上XhoI和BamHI酶切位点间,经线性化后,使用mRNA合成试剂盒(购自上海蓝鹊生物)通过共转录化学底物法加帽方式进行体外转录,并进一步使用RNA纯化试剂盒(购自NEB)纯化。纯化后mRNA浓度使用核酸检测试剂盒Quant-iT RiboGreenR RNA Reagent Kit(购自Thermo Fisher)定量。
实施例2——水痘-带状疱疹mRNA疫苗组合物制备
按照MC3: DSPC: cholesterol: DMG-PEG2000摩尔比为50:10:37.5:2.5称取脂质(购自上海艾伟拓)后溶解于无水乙醇,形成溶液A;将0.1毫克实施例1制备的mRNA溶于100mM,pH 4.0的柠檬酸缓冲液,形成溶液B;使用微流体纳米药物制造系统(加拿大PrecisionNanosystems公司NanoAssemblr Ignite)按照溶液A:溶液B体积比1:3混合后,使用40倍体积PBS缓冲液透析,过0.22微米滤膜除菌后即得水痘-带状疱疹mRNA疫苗组合物。
针对以上实施例2制备得到的各mRNA疫苗组合物,进行如下测定:
1、粒径和多分散性指数
使用纳米粒径检测仪Zetasizer Nao ZS paricle size analyzer(英国马尔文)检测LNP的粒径和多分散性指数。
2、mRNA浓度及包封率
LNP纳米颗粒疫苗于0.1M氢氧化钠和0.1%(w/v)的十二烷基硫酸钠缓冲液室温裂解过夜。使用核酸检测试剂盒Quant-iT RiboGreenR RNA Reagent Kit (购自ThermoFisher)检测核酸浓度并计算核酸荷载效率。
3、mRNA完整性
LNP纳米颗粒疫苗于0.1M氢氧化钠和0.1%(w/v)的十二烷基硫酸钠缓冲液室温裂解过夜。使用1% 变性琼脂糖凝胶进行mRNA完整性检测。
结果如图1所示,实施例1-2使用微流体纳米药物制造系统制备的水痘-带状疱疹mRNA疫苗组合物粒径(图1A)分别为gE全长96.30纳米,gE突变97.92纳米,gE胞外95.52纳米。均一性较好,体现为多分散性指数(图1B)均小于0.2,包括gE全长0.166,gE突变0.160,gE胞外0.132。对于所投0.1毫克mRNA,理论包封率(图1C)分别为gE全长102.05%,gE突变94.91%,gE胞外95.45%。电泳结果(图1D)显示,实施例1制备的mRNA经实施例2处理后,完整性仍十分良好。
对比例1——带状疱疹亚单位疫苗Shingrix(购自葛兰素史克)。
对比例2——CHO表达的gE胞外区糖蛋白(购自普健生物科技有限公司)10微克,双链聚胞嘧啶核苷酸片段(Poly I:C,购自InvivoGen)15微克溶解于PBS,并同等体积铝佐剂(购自Thermo Fisher)混匀为50微升终体积。
针对以上实施例1-2以及比对例1-2制备得到的各疫苗,进一步进行如下测定:
1、统计分析
使用GraphPad Prism 8.0软件,采用one-way ANOVA方法对以上实施例及对比例所得数据进行统计学分析,其中p≥0.05为无显著差异,标注为ns;显著差异中p<0.05标示为*,p<0.01标示为**。
2、动物免疫
间隔3周,肌肉注射实施例2(含7.5微克mRNA)、对比例1(1/10人用剂量)、对比例2制备的疫苗50 µL免疫C57BL/6小鼠2次(6只/组,雌性,初免年龄6周,体重15-18 g,购自北京维通利华实验动物技术有限公司),终免2周后摘取脾脏,心脏取血4 ℃放置过夜后3 500rpm离心20分钟取血清,准备进行后续免疫学分析。
3、抗体滴度检测
溶解于PBS的捕获抗原gE胞外区糖蛋白2 µg/mL每孔100 µL加入96孔酶标板(购自康宁),4 ℃过夜包被后TBST(0.05% Tween20(Sigma) in PBS)洗板1次,每孔200 µL加入5%(w/v)溶于PBS的脱脂奶粉37 ℃封闭1 h,弃掉牛奶后TBST洗4次,每孔100 µL加入1%牛奶梯度稀释的实施例10制备的抗血清37 ℃孵育1 h,TBST洗5次后加入1%牛奶稀释的二抗(1:5000,Goat anti-mouse IgG:HRP购自BioRad)37 ℃孵育1 h,TBST洗5次后每孔加入100 µL按照1:1比例配好的显色液(购自BD),室温下避光放置10分钟后,加入100 µL 1M磷酸终止反应,并于450 nm处检测光吸收值。取OD450>0.1临界的血清稀释浓度作为抗体滴度,最大稀释倍数OD450≤0.1定义为100。
如图2所示,比对例1葛兰素史克的产品Shingrix诱导的抗原特异性抗体滴度(213333)约为对比例2所诱导抗体滴度(101333)的2倍,并略高于实施例中gE胞外区水痘-带状疱疹mRNA疫苗组合物所诱导的抗体滴度(107667),但实施例中gE突变体诱导的抗体滴度(597333)最高,约为gE全长所诱导抗体滴度(333333)的2倍,约为对比例1的Shingrix所诱导抗体滴度的3倍。
4、脾脏淋巴细胞分离
上述1动物免疫部分制备的脾脏置细胞过滤网(无锡耐思生命科技股份有限公司),加入ACK红细胞裂解液室温放置5分钟,1800转/分钟离心后细胞计数,并使用含有10%胎牛血清(购自BI)和双抗的1640培养基(购自Thermo Fisher)重悬为3×106细胞/毫升。使用时100 μL/孔细胞加入96细胞培养板孔板(购自康宁)至最终细胞量3×105细胞/孔。
5、酶联免疫斑点实验 (enzyme linked immunospot assay,ELISPOT)
IL-2及IFN-γ检测试剂盒均购自BD,并按照说明书进行操作,具体步骤如下:包被液稀释捕获抗体后100 μL/孔加入ELISPOT平板,4 ℃包被过夜后弃包被液,200 μL/孔封闭液洗板1次,每孔200 μL加入封闭液室温封闭2h,弃封闭液后加入含有20 μg/mL终浓度gE的1640完全培养基100 μL并加入上述脾脏淋巴细胞分离中得到的等体积稀释好的脾脏细胞(3×106细胞/毫升)37 ℃细胞培养箱过夜。800 g离心5分钟后弃上清,200 μL/孔去离子水清洗2次(每次浸泡5分钟),200 μL/孔清洗液1清洗3次,100 μL/孔加入经稀释液稀释的检测抗体室温孵育2小时后,200 μL/孔清洗液1清洗3次(每次浸泡2分钟),100 μL/孔加入经稀释液稀释的酶偶合物Streptavidin-HRP室温孵育1小时,200 μL/孔清洗液1清洗4次(每次浸泡2分钟),200 μL/孔清洗液2清洗2次后加入100 μL底物溶液反应至合适时间,去离子水清洗终止反应。晾干后使用ELISPOT读板仪器(德国AID Diagnostika GmbH)斑点计数。
如图3所示,实施例中gE突变体水痘-带状疱疹mRNA疫苗组合物所诱导分泌IFN-γ的脾脏细胞数量(图3A,每30万个脾脏细胞中为129个)在所有实施例(包括gE全长诱导产生的102.7个,以及gE胞外区诱导产生的116.5个)最多,并显著高于对比例1(47.3个,p<0.01)和对比例2(P<0.01)。另外,实施例中gE突变体水痘-带状疱疹mRNA疫苗组合物所诱导分泌IL-2的脾脏细胞数量(图3B,每30万个脾脏细胞中为111.7个)仍高于gE全长诱导产生的诱导分泌IL-2的脾脏细胞数量(100.2个)但略低于具有明显离群值的实施例gE胞外区组(118.3个)。相对于对比例2,gE突变体水痘-带状疱疹mRNA疫苗组合物所诱导分泌IL-2的脾脏细胞数量显著升高(p<0.01)。
6、流式细胞分析
上述分离的脾脏淋巴细胞每孔加入终浓度为10μg/mL的gE,连同5 μl/孔的FastImmune(购自BD)37 ℃细胞培养箱孵育2 h,加入蛋白运输抑制剂BrefeldinA(购自Biolegend) 37 ℃细胞培养箱过夜。800 g离心5分钟后弃上清,250 μL staining buffer(购自Biologend)清洗一次,加入50 μL含有5 μg/mL CD16/32抗体(购自Biolegend)的staining buffer 重悬后4 ℃孵育10分钟封闭Fc受体。加入50 μL staining buffer稀释好的抗鼠抗体PerCP-CD4,FITC-CD8a,Brilliant Violet 421-CD62L,Brilliant Violet510-CD44(均购自Biolegend),4 ℃避光染色30分钟,使用250 μL staining buffer清洗1遍,加入200 μL fixation buffer(购自Biolegend)悬浮固定细胞膜室温避光孵育20分钟。离心去上清,250 μL permeabilization wash buffer(购自Biolegend)清洗2次,加入用50μL permeabilization wash buffer稀释好的抗鼠抗体APC-IL-2及PE-IFN-γ(均购自Biolegend)悬浮细胞,室温避光30分钟,250 μl permeabilization wash buffer清洗一次,250 μl PBS清洗一次,100 μl PBS悬浮后使用流式细胞分析仪(Beckman CytoFLEXflow cytometer)检测并使用相关软件FlowJo_V10进行数据分析。
如图4所示,实施例中gE突变体水痘-带状疱疹mRNA疫苗组合物所诱导分泌IFN-γ的CD4阳性细胞数量在所有脾细胞中所占比例(图4A,0.153%)仅略低于具有明显离群值的实施例gE全长组(0.194%),高于实施例gE胞外区组(0.111%)和对比例1(0.054%),并显著高于对比例2(0.025%)。另外,实施例中gE突变体水痘-带状疱疹mRNA疫苗组合物所诱导分泌IFN-γ的CD8阳性细胞数量在所有脾细胞中所占比例(图4B,0.110%)仅略低于具有明显离群值的实施例gE全长组(0.117%),高于实施例gE胞外区组(0.054%)、对比例1(0.108%)和对比例2组(0.046%)。
如图5所示,实施例中gE突变体水痘-带状疱疹mRNA疫苗组合物所诱导的同时表达CD44和CD62L的CD4阳性细胞数量在所有脾细胞中所占比例(图5A,1.448%)最高,并显著高于(p<0.05)对比例2(1.101%)。另外,实施例中gE突变体水痘-带状疱疹mRNA疫苗组合物所诱导的同时表达CD44和CD62L的CD8阳性细胞数量在所有脾细胞中所占比例(图5B,6.717%)仅略低于实施例gE全长组(7.338%),高于实施例gE胞外区组(5.23%)、对比例1(5.757%)和对比例2组(5.788%)。
最后应说明的是,以上仅用以说明本发明的技术方案而非限制,尽管参照较佳布置方案对本发明进行了详细说明,本领域的普通技术人员应当理解,可以对本发明的技术方案进行修改或者等同替换,而不脱离本发明技术方案的精神和范围。
Claims (5)
1.一种水痘-带状疱疹mRNA疫苗组合物,其特征在于,所述疫苗组合物包括编码水痘-带状疱疹病毒糖蛋白E的信使核糖核酸(mRNA)序列、脂质纳米颗粒(LNP);水痘-带状疱疹病毒糖蛋白E的序列为SEQ ID No.2所示;
所述mRNA的制备方法为:编码SEQ ID No.2所述的羧基端胞内区引入突变的氨基酸序列的基因按照哺乳动物密码子偏好优化后,连同5’非翻译区序列、3’非翻译区序列和3’端多聚腺苷酸序列进行合成,构建于质粒pBlueScript II SK(+)上XhoI和BamHI酶切位点间,经线性化后,使用mRNA合成试剂盒通过共转录化学底物法加帽方式进行体外转录,并进一步使用RNA纯化试剂盒纯化;
按照MC3: DSPC: cholesterol: DMG-PEG2000摩尔比为50:10:37.5:2.5称取脂质后溶解于无水乙醇,形成溶液A;将0.1毫克上述步骤制备得到的mRNA溶于100 mM,pH 4.0的柠檬酸缓冲液,形成溶液B;使用微流体纳米药物制造系统按照溶液A:溶液B体积比1:3混合后,使用40倍体积PBS缓冲液透析,过0.22微米滤膜除菌后,即得水痘-带状疱疹mRNA疫苗组合物。
2.如权利要求1所述的水痘-带状疱疹mRNA疫苗组合物,其特征在于,所述水痘-带状疱疹病毒糖蛋白E的mRNA用量为2.5 μg-150 μg每针剂。
3.如权利要求1所述的水痘-带状疱疹mRNA疫苗组合物,其特征在于,所述疫苗组合物为直径介于20-400纳米的颗粒组合物。
4.如权利要求1-3任一项所述的水痘-带状疱疹mRNA疫苗组合物,其特征在于,所述疫苗组合物的施用方式包括皮下或肌肉注射。
5.一种如权利要求1-3任一项所述水痘-带状疱疹mRNA疫苗组合物的应用,其特征在于,所述疫苗组合物用于制备预防或改善水痘和/或带状疱疹和/或带状疱疹后神经痛的药物。
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN202111311113 | 2021-11-08 | ||
CN202111311113X | 2021-11-08 |
Publications (2)
Publication Number | Publication Date |
---|---|
CN114081943A CN114081943A (zh) | 2022-02-25 |
CN114081943B true CN114081943B (zh) | 2024-04-02 |
Family
ID=80302430
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CN202111373707.3A Active CN114081943B (zh) | 2021-11-08 | 2021-11-19 | 一种水痘-带状疱疹mRNA疫苗组合物及其制备方法和应用 |
Country Status (1)
Country | Link |
---|---|
CN (1) | CN114081943B (zh) |
Families Citing this family (8)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2023143601A1 (zh) * | 2022-01-30 | 2023-08-03 | 康希诺生物股份公司 | 一种用于核酸递送的新型可电离脂质及其lnp组合物和疫苗 |
CN114404388B (zh) * | 2022-03-31 | 2022-07-26 | 天津外泌体科技有限公司 | 一种细胞外囊泡体外装载mRNA的方法 |
CN114854772A (zh) * | 2022-04-19 | 2022-08-05 | 浙江君怡生物科技有限公司 | 水痘-带状疱疹病毒mRNA疫苗及其制备方法和应用 |
CN115154439B (zh) * | 2022-09-08 | 2022-12-09 | 南京澄实生物科技有限公司 | 一种mRNA脂质纳米颗粒递送系统及其制备方法和应用 |
CN117462666A (zh) * | 2022-09-30 | 2024-01-30 | 烟台派诺生物技术有限公司 | 一种预防或治疗水痘-带状疱疹病毒相关疾病的免疫组合物产品及其制备方法 |
CN116355917B (zh) * | 2023-02-15 | 2023-12-01 | 上海臻上医药科技有限公司 | 一种水痘-带状疱疹病毒mRNA疫苗及其应用 |
CN116606867B (zh) * | 2023-04-28 | 2024-01-26 | 北京启辰生生物科技有限公司 | 一种提高新冠全长S蛋白mRNA完整性的制备方法及用途 |
CN117003894B (zh) * | 2023-08-09 | 2024-04-16 | 成都新诺明生物科技有限公司 | 一种含有IL-2和Fc的gE融合蛋白及其制备方法和应用 |
Citations (8)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US4686101A (en) * | 1985-08-02 | 1987-08-11 | Merck & Co., Inc. | Vaccine against varicella-zoster virus |
US4952674A (en) * | 1986-05-02 | 1990-08-28 | Merck & Co., Inc. | Vaccine against varicella-zoster virus |
CN105669838A (zh) * | 2014-12-04 | 2016-06-15 | 厦门大学 | 来自水痘-带状疱疹病毒gE蛋白的中和表位及针对其的抗体 |
CN108472309A (zh) * | 2015-10-22 | 2018-08-31 | 摩登纳特斯有限公司 | 用于水痘带状疱疹病毒(vzv)的核酸疫苗 |
CN109602901A (zh) * | 2019-01-08 | 2019-04-12 | 成都迈科康生物科技有限公司 | 一种带状疱疹病毒疫苗及其制备方法和应用 |
CN111558036A (zh) * | 2020-05-14 | 2020-08-21 | 中国医学科学院医学生物学研究所 | 一种带状疱疹疫苗组合物及其制备方法和应用 |
CN113144186A (zh) * | 2021-02-09 | 2021-07-23 | 中国医学科学院医学生物学研究所 | 一种水痘-带状疱疹疫苗组合物及其制备方法和应用 |
CN113226364A (zh) * | 2018-11-06 | 2021-08-06 | 牛津大学科技创新有限公司 | 组合物和方法 |
Family Cites Families (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US11045540B2 (en) * | 2017-03-15 | 2021-06-29 | Modernatx, Inc. | Varicella zoster virus (VZV) vaccine |
AU2018352221B2 (en) * | 2017-10-16 | 2024-04-18 | Aadigen, Llc | Peptides and nanoparticles for intracellular delivery of mRNA |
-
2021
- 2021-11-19 CN CN202111373707.3A patent/CN114081943B/zh active Active
Patent Citations (8)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US4686101A (en) * | 1985-08-02 | 1987-08-11 | Merck & Co., Inc. | Vaccine against varicella-zoster virus |
US4952674A (en) * | 1986-05-02 | 1990-08-28 | Merck & Co., Inc. | Vaccine against varicella-zoster virus |
CN105669838A (zh) * | 2014-12-04 | 2016-06-15 | 厦门大学 | 来自水痘-带状疱疹病毒gE蛋白的中和表位及针对其的抗体 |
CN108472309A (zh) * | 2015-10-22 | 2018-08-31 | 摩登纳特斯有限公司 | 用于水痘带状疱疹病毒(vzv)的核酸疫苗 |
CN113226364A (zh) * | 2018-11-06 | 2021-08-06 | 牛津大学科技创新有限公司 | 组合物和方法 |
CN109602901A (zh) * | 2019-01-08 | 2019-04-12 | 成都迈科康生物科技有限公司 | 一种带状疱疹病毒疫苗及其制备方法和应用 |
CN111558036A (zh) * | 2020-05-14 | 2020-08-21 | 中国医学科学院医学生物学研究所 | 一种带状疱疹疫苗组合物及其制备方法和应用 |
CN113144186A (zh) * | 2021-02-09 | 2021-07-23 | 中国医学科学院医学生物学研究所 | 一种水痘-带状疱疹疫苗组合物及其制备方法和应用 |
Non-Patent Citations (2)
Title |
---|
Functions of the ORF9-to-ORF12 Gene Cluster in Varicella-Zoster Virus Replication and in the Pathogenesis of Skin Infection;Xibing Che等;JOURNAL OF VIROLOGY;20080630;第82卷(第12期);第5825-5834页 * |
水痘-带状疱疹病毒糖蛋白E在昆虫细胞中的表达、鉴定及免疫原性分析;齐家龙等;中国生物工程杂志;20190815;第39卷(第8期);第17-24页 * |
Also Published As
Publication number | Publication date |
---|---|
CN114081943A (zh) | 2022-02-25 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
CN114081943B (zh) | 一种水痘-带状疱疹mRNA疫苗组合物及其制备方法和应用 | |
Skountzou et al. | Incorporation of Glycosylphosphatidylinositol-Anchored Granulocyte-MacrophageColony-Stimulating Factor or CD40 Ligand Enhances Immunogenicity of Chimeric Simian Immunodeficiency Virus-Like Particles | |
US20210290759A1 (en) | Immune composition, preparation method therefor, and application thereof | |
CN104271745A (zh) | 用于治疗巨细胞病毒的组合物和方法 | |
Maamary et al. | Newcastle disease virus expressing a dendritic cell-targeted HIV gag protein induces a potent gag-specific immune response in mice | |
Qi et al. | Construction and immunogenic studies of a mFc fusion receptor binding domain (RBD) of spike protein as a subunit vaccine against SARS-CoV-2 infection | |
TW202307212A (zh) | 對抗廣譜冠狀病毒變種的信使rna疫苗 | |
WO2023051701A1 (zh) | 抗SARS-CoV-2感染的mRNA、蛋白以及抗SARS-CoV-2感染的疫苗 | |
WO2021262672A1 (en) | Sars-cov-2 rbd constructs | |
Li et al. | Enhancing the immunogenicity of lipid-nanoparticle mRNA vaccines by adjuvanting the ionizable lipid and the mRNA | |
US11160857B2 (en) | Multivalent enterovirus vaccine compositions and uses related thereto | |
Vijayamahantesh et al. | Modified E2 glycoprotein of hepatitis C virus enhances proinflammatory cytokines and protective immune response | |
CN113144186B (zh) | 一种水痘-带状疱疹疫苗组合物及其制备方法和应用 | |
CN116747298B (zh) | 一种水痘-带状疱疹病毒疫苗及其制备方法和应用 | |
WO2021214703A1 (en) | A vaccine against sars-cov-2 and preparation thereof | |
WO2023160654A1 (zh) | 一种可诱导广谱中和活性重组多组分新冠病毒三聚体蛋白疫苗的制备及应用 | |
CN116410992A (zh) | 预防和/或治疗新型冠状病毒的mRNA、疫苗及其制备方法和应用 | |
CN116024237A (zh) | 一种用于预防裂谷热的mRNA疫苗及其制备方法 | |
CN115820696A (zh) | 治疗性多价HPV mRNA疫苗及其制备方法 | |
CN113832168B (zh) | mRNA疫苗及其制备方法和应用 | |
WO2024055273A1 (zh) | 一种狂犬病mRNA疫苗、其制备及应用 | |
CN117205309B (zh) | 一种流感免疫原组合物和制备方法及其用途 | |
Neeli et al. | Comparison of DNA vaccines with AS03 as an adjuvant and an mRNA vaccine against SARS-CoV-2 | |
CN117003894B (zh) | 一种含有IL-2和Fc的gE融合蛋白及其制备方法和应用 | |
WO2023202711A1 (zh) | 一种基于新型冠状病毒的mRNA疫苗 |
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 | ||
GR01 | Patent grant | ||
GR01 | Patent grant |