WO2008026225A2 - A vaccine for chikungunya virus infection - Google Patents
A vaccine for chikungunya virus infection Download PDFInfo
- Publication number
- WO2008026225A2 WO2008026225A2 PCT/IN2007/000383 IN2007000383W WO2008026225A2 WO 2008026225 A2 WO2008026225 A2 WO 2008026225A2 IN 2007000383 W IN2007000383 W IN 2007000383W WO 2008026225 A2 WO2008026225 A2 WO 2008026225A2
- Authority
- WO
- WIPO (PCT)
- Prior art keywords
- virus
- seq
- chikungunya
- hrs
- cells
- Prior art date
Links
- 229960005486 vaccine Drugs 0.000 title claims abstract description 51
- 208000004293 Chikungunya Fever Diseases 0.000 title claims abstract description 9
- 206010067256 Chikungunya virus infection Diseases 0.000 title claims abstract description 9
- 241000700605 Viruses Species 0.000 claims abstract description 167
- 241001502567 Chikungunya virus Species 0.000 claims abstract description 71
- 239000000203 mixture Substances 0.000 claims abstract description 49
- 238000000034 method Methods 0.000 claims abstract description 47
- 238000009472 formulation Methods 0.000 claims abstract description 42
- 239000000427 antigen Substances 0.000 claims abstract description 32
- 108091007433 antigens Proteins 0.000 claims abstract description 32
- 102000036639 antigens Human genes 0.000 claims abstract description 32
- 208000015181 infectious disease Diseases 0.000 claims abstract description 24
- 238000000746 purification Methods 0.000 claims abstract description 20
- 230000002163 immunogen Effects 0.000 claims abstract description 19
- 230000028993 immune response Effects 0.000 claims abstract description 9
- 230000001681 protective effect Effects 0.000 claims abstract description 9
- 230000002458 infectious effect Effects 0.000 claims abstract description 8
- 238000001727 in vivo Methods 0.000 claims abstract description 4
- 108090000623 proteins and genes Proteins 0.000 claims description 35
- 102000004169 proteins and genes Human genes 0.000 claims description 27
- WSFSSNUMVMOOMR-UHFFFAOYSA-N Formaldehyde Chemical compound O=C WSFSSNUMVMOOMR-UHFFFAOYSA-N 0.000 claims description 26
- 230000014509 gene expression Effects 0.000 claims description 22
- 201000009182 Chikungunya Diseases 0.000 claims description 20
- 238000002360 preparation method Methods 0.000 claims description 19
- 230000002779 inactivation Effects 0.000 claims description 16
- 238000004113 cell culture Methods 0.000 claims description 15
- 239000002245 particle Substances 0.000 claims description 15
- VEZXCJBBBCKRPI-UHFFFAOYSA-N beta-propiolactone Chemical compound O=C1CCO1 VEZXCJBBBCKRPI-UHFFFAOYSA-N 0.000 claims description 12
- 229960000380 propiolactone Drugs 0.000 claims description 12
- 241000701447 unidentified baculovirus Species 0.000 claims description 12
- 238000000338 in vitro Methods 0.000 claims description 11
- 239000013598 vector Substances 0.000 claims description 11
- 241000588724 Escherichia coli Species 0.000 claims description 10
- 150000003839 salts Chemical class 0.000 claims description 9
- 235000000346 sugar Nutrition 0.000 claims description 9
- 240000004808 Saccharomyces cerevisiae Species 0.000 claims description 8
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 claims description 8
- 230000000890 antigenic effect Effects 0.000 claims description 8
- 102000007056 Recombinant Fusion Proteins Human genes 0.000 claims description 7
- 108010008281 Recombinant Fusion Proteins Proteins 0.000 claims description 7
- 239000000872 buffer Substances 0.000 claims description 7
- 230000002068 genetic effect Effects 0.000 claims description 7
- 239000008363 phosphate buffer Substances 0.000 claims description 7
- 239000000126 substance Substances 0.000 claims description 7
- 150000008163 sugars Chemical class 0.000 claims description 7
- 238000005199 ultracentrifugation Methods 0.000 claims description 7
- CIWBSHSKHKDKBQ-JLAZNSOCSA-N Ascorbic acid Chemical compound OC[C@H](O)[C@H]1OC(=O)C(O)=C1O CIWBSHSKHKDKBQ-JLAZNSOCSA-N 0.000 claims description 6
- 241000238631 Hexapoda Species 0.000 claims description 6
- 241000282412 Homo Species 0.000 claims description 6
- 230000006978 adaptation Effects 0.000 claims description 6
- 239000003795 chemical substances by application Substances 0.000 claims description 6
- 238000010790 dilution Methods 0.000 claims description 6
- 239000012895 dilution Substances 0.000 claims description 6
- 239000007788 liquid Substances 0.000 claims description 6
- 238000004519 manufacturing process Methods 0.000 claims description 6
- 239000013612 plasmid Substances 0.000 claims description 6
- 108091034117 Oligonucleotide Proteins 0.000 claims description 5
- WNROFYMDJYEPJX-UHFFFAOYSA-K aluminium hydroxide Chemical compound [OH-].[OH-].[OH-].[Al+3] WNROFYMDJYEPJX-UHFFFAOYSA-K 0.000 claims description 5
- 229910021502 aluminium hydroxide Inorganic materials 0.000 claims description 5
- 238000003306 harvesting Methods 0.000 claims description 5
- 230000000415 inactivating effect Effects 0.000 claims description 5
- 239000007981 phosphate-citrate buffer Substances 0.000 claims description 5
- 239000003381 stabilizer Substances 0.000 claims description 5
- 108091006905 Human Serum Albumin Proteins 0.000 claims description 4
- 102000008100 Human Serum Albumin Human genes 0.000 claims description 4
- CZMRCDWAGMRECN-UGDNZRGBSA-N Sucrose Chemical compound O[C@H]1[C@H](O)[C@@H](CO)O[C@@]1(CO)O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 CZMRCDWAGMRECN-UGDNZRGBSA-N 0.000 claims description 4
- 229930006000 Sucrose Natural products 0.000 claims description 4
- JLCPHMBAVCMARE-UHFFFAOYSA-N [3-[[3-[[3-[[3-[[3-[[3-[[3-[[3-[[3-[[3-[[3-[[5-(2-amino-6-oxo-1H-purin-9-yl)-3-[[3-[[3-[[3-[[3-[[3-[[5-(2-amino-6-oxo-1H-purin-9-yl)-3-[[5-(2-amino-6-oxo-1H-purin-9-yl)-3-hydroxyoxolan-2-yl]methoxy-hydroxyphosphoryl]oxyoxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(5-methyl-2,4-dioxopyrimidin-1-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(6-aminopurin-9-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(6-aminopurin-9-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(6-aminopurin-9-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(6-aminopurin-9-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxyoxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(5-methyl-2,4-dioxopyrimidin-1-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(4-amino-2-oxopyrimidin-1-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(5-methyl-2,4-dioxopyrimidin-1-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(5-methyl-2,4-dioxopyrimidin-1-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(6-aminopurin-9-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(6-aminopurin-9-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(4-amino-2-oxopyrimidin-1-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(4-amino-2-oxopyrimidin-1-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(4-amino-2-oxopyrimidin-1-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(6-aminopurin-9-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(4-amino-2-oxopyrimidin-1-yl)oxolan-2-yl]methyl [5-(6-aminopurin-9-yl)-2-(hydroxymethyl)oxolan-3-yl] hydrogen phosphate Polymers Cc1cn(C2CC(OP(O)(=O)OCC3OC(CC3OP(O)(=O)OCC3OC(CC3O)n3cnc4c3nc(N)[nH]c4=O)n3cnc4c3nc(N)[nH]c4=O)C(COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3CO)n3cnc4c(N)ncnc34)n3ccc(N)nc3=O)n3cnc4c(N)ncnc34)n3ccc(N)nc3=O)n3ccc(N)nc3=O)n3ccc(N)nc3=O)n3cnc4c(N)ncnc34)n3cnc4c(N)ncnc34)n3cc(C)c(=O)[nH]c3=O)n3cc(C)c(=O)[nH]c3=O)n3ccc(N)nc3=O)n3cc(C)c(=O)[nH]c3=O)n3cnc4c3nc(N)[nH]c4=O)n3cnc4c(N)ncnc34)n3cnc4c(N)ncnc34)n3cnc4c(N)ncnc34)n3cnc4c(N)ncnc34)O2)c(=O)[nH]c1=O JLCPHMBAVCMARE-UHFFFAOYSA-N 0.000 claims description 4
- 238000004440 column chromatography Methods 0.000 claims description 4
- 238000000432 density-gradient centrifugation Methods 0.000 claims description 4
- 239000003599 detergent Substances 0.000 claims description 4
- 239000013604 expression vector Substances 0.000 claims description 4
- 238000002523 gelfiltration Methods 0.000 claims description 4
- 238000004255 ion exchange chromatography Methods 0.000 claims description 4
- 239000012931 lyophilized formulation Substances 0.000 claims description 4
- 239000012528 membrane Substances 0.000 claims description 4
- 229920001223 polyethylene glycol Polymers 0.000 claims description 4
- 238000001556 precipitation Methods 0.000 claims description 4
- 102000004196 processed proteins & peptides Human genes 0.000 claims description 4
- 108090000765 processed proteins & peptides Proteins 0.000 claims description 4
- 239000011780 sodium chloride Substances 0.000 claims description 4
- 239000005720 sucrose Substances 0.000 claims description 4
- 238000011282 treatment Methods 0.000 claims description 4
- 238000000108 ultra-filtration Methods 0.000 claims description 4
- HDTRYLNUVZCQOY-UHFFFAOYSA-N α-D-glucopyranosyl-α-D-glucopyranoside Natural products OC1C(O)C(O)C(CO)OC1OC1C(O)C(O)C(O)C(CO)O1 HDTRYLNUVZCQOY-UHFFFAOYSA-N 0.000 claims description 3
- OWEGMIWEEQEYGQ-UHFFFAOYSA-N 100676-05-9 Natural products OC1C(O)C(O)C(CO)OC1OCC1C(O)C(O)C(O)C(OC2C(OC(O)C(O)C2O)CO)O1 OWEGMIWEEQEYGQ-UHFFFAOYSA-N 0.000 claims description 3
- GUBGYTABKSRVRQ-XLOQQCSPSA-N Alpha-Lactose Chemical compound O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@H]1O[C@@H]1[C@@H](CO)O[C@H](O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-XLOQQCSPSA-N 0.000 claims description 3
- 108010010803 Gelatin Proteins 0.000 claims description 3
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 claims description 3
- GUBGYTABKSRVRQ-QKKXKWKRSA-N Lactose Natural products OC[C@H]1O[C@@H](O[C@H]2[C@H](O)[C@@H](O)C(O)O[C@@H]2CO)[C@H](O)[C@@H](O)[C@H]1O GUBGYTABKSRVRQ-QKKXKWKRSA-N 0.000 claims description 3
- GUBGYTABKSRVRQ-PICCSMPSSA-N Maltose Natural products O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@@H](CO)OC(O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-PICCSMPSSA-N 0.000 claims description 3
- 241000124008 Mammalia Species 0.000 claims description 3
- 229910019142 PO4 Inorganic materials 0.000 claims description 3
- 239000002202 Polyethylene glycol Substances 0.000 claims description 3
- 101710132906 Structural polyprotein Proteins 0.000 claims description 3
- HDTRYLNUVZCQOY-WSWWMNSNSA-N Trehalose Natural products O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@@H](O)[C@@H](CO)O1 HDTRYLNUVZCQOY-WSWWMNSNSA-N 0.000 claims description 3
- HDTRYLNUVZCQOY-LIZSDCNHSA-N alpha,alpha-trehalose Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CO)O[C@@H]1O[C@@H]1[C@H](O)[C@@H](O)[C@H](O)[C@@H](CO)O1 HDTRYLNUVZCQOY-LIZSDCNHSA-N 0.000 claims description 3
- ILRRQNADMUWWFW-UHFFFAOYSA-K aluminium phosphate Chemical compound O1[Al]2OP1(=O)O2 ILRRQNADMUWWFW-UHFFFAOYSA-K 0.000 claims description 3
- 229940001007 aluminium phosphate Drugs 0.000 claims description 3
- 229910000147 aluminium phosphate Inorganic materials 0.000 claims description 3
- 239000011668 ascorbic acid Substances 0.000 claims description 3
- 229960005070 ascorbic acid Drugs 0.000 claims description 3
- 235000010323 ascorbic acid Nutrition 0.000 claims description 3
- WQZGKKKJIJFFOK-VFUOTHLCSA-N beta-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 claims description 3
- GUBGYTABKSRVRQ-QUYVBRFLSA-N beta-maltose Chemical compound OC[C@H]1O[C@H](O[C@H]2[C@H](O)[C@@H](O)[C@H](O)O[C@@H]2CO)[C@H](O)[C@@H](O)[C@@H]1O GUBGYTABKSRVRQ-QUYVBRFLSA-N 0.000 claims description 3
- 238000012258 culturing Methods 0.000 claims description 3
- 239000008273 gelatin Substances 0.000 claims description 3
- 229920000159 gelatin Polymers 0.000 claims description 3
- 235000019322 gelatine Nutrition 0.000 claims description 3
- 235000011852 gelatine desserts Nutrition 0.000 claims description 3
- 239000008103 glucose Substances 0.000 claims description 3
- 238000011534 incubation Methods 0.000 claims description 3
- 239000008101 lactose Substances 0.000 claims description 3
- NBIIXXVUZAFLBC-UHFFFAOYSA-K phosphate Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 claims description 3
- 238000007920 subcutaneous administration Methods 0.000 claims description 3
- 108091035707 Consensus sequence Proteins 0.000 claims description 2
- 238000001042 affinity chromatography Methods 0.000 claims description 2
- 238000005119 centrifugation Methods 0.000 claims description 2
- 230000002209 hydrophobic effect Effects 0.000 claims description 2
- 238000004191 hydrophobic interaction chromatography Methods 0.000 claims description 2
- 239000002955 immunomodulating agent Substances 0.000 claims description 2
- 238000007918 intramuscular administration Methods 0.000 claims description 2
- 238000001990 intravenous administration Methods 0.000 claims description 2
- 239000011159 matrix material Substances 0.000 claims description 2
- 150000002894 organic compounds Chemical class 0.000 claims description 2
- 239000003960 organic solvent Substances 0.000 claims description 2
- 239000002671 adjuvant Substances 0.000 claims 5
- 125000003275 alpha amino acid group Chemical group 0.000 claims 4
- 108020004511 Recombinant DNA Proteins 0.000 claims 3
- 239000008194 pharmaceutical composition Substances 0.000 claims 3
- 102000004407 Lactalbumin Human genes 0.000 claims 2
- 108090000942 Lactalbumin Proteins 0.000 claims 2
- 239000000654 additive Substances 0.000 claims 2
- GHCZTIFQWKKGSB-UHFFFAOYSA-N 2-hydroxypropane-1,2,3-tricarboxylic acid;phosphoric acid Chemical compound OP(O)(O)=O.OC(=O)CC(O)(C(O)=O)CC(O)=O GHCZTIFQWKKGSB-UHFFFAOYSA-N 0.000 claims 1
- 108091028043 Nucleic acid sequence Proteins 0.000 claims 1
- 150000001450 anions Chemical class 0.000 claims 1
- 229910000389 calcium phosphate Inorganic materials 0.000 claims 1
- 239000001506 calcium phosphate Substances 0.000 claims 1
- 235000011010 calcium phosphates Nutrition 0.000 claims 1
- 238000004587 chromatography analysis Methods 0.000 claims 1
- 238000001962 electrophoresis Methods 0.000 claims 1
- 238000005194 fractionation Methods 0.000 claims 1
- 230000003308 immunostimulating effect Effects 0.000 claims 1
- 150000002484 inorganic compounds Chemical class 0.000 claims 1
- 229910010272 inorganic material Inorganic materials 0.000 claims 1
- 239000002480 mineral oil Substances 0.000 claims 1
- 235000010446 mineral oil Nutrition 0.000 claims 1
- 150000007523 nucleic acids Chemical group 0.000 claims 1
- 239000002773 nucleotide Substances 0.000 claims 1
- 125000003729 nucleotide group Chemical group 0.000 claims 1
- 239000010452 phosphate Substances 0.000 claims 1
- 238000011321 prophylaxis Methods 0.000 claims 1
- QORWJWZARLRLPR-UHFFFAOYSA-H tricalcium bis(phosphate) Chemical compound [Ca+2].[Ca+2].[Ca+2].[O-]P([O-])([O-])=O.[O-]P([O-])([O-])=O QORWJWZARLRLPR-UHFFFAOYSA-H 0.000 claims 1
- 238000002255 vaccination Methods 0.000 claims 1
- 108010067390 Viral Proteins Proteins 0.000 abstract description 5
- 230000003053 immunization Effects 0.000 abstract description 4
- 238000002649 immunization Methods 0.000 abstract description 4
- 229940031626 subunit vaccine Drugs 0.000 abstract description 4
- 210000004027 cell Anatomy 0.000 description 84
- 210000003501 vero cell Anatomy 0.000 description 24
- 230000035931 haemagglutination Effects 0.000 description 21
- 235000018102 proteins Nutrition 0.000 description 20
- 241000699670 Mus sp. Species 0.000 description 18
- 210000002966 serum Anatomy 0.000 description 18
- 241001465754 Metazoa Species 0.000 description 13
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 12
- 241000282414 Homo sapiens Species 0.000 description 11
- 238000002955 isolation Methods 0.000 description 11
- 238000012360 testing method Methods 0.000 description 11
- 238000002965 ELISA Methods 0.000 description 10
- 241000283973 Oryctolagus cuniculus Species 0.000 description 10
- 238000010367 cloning Methods 0.000 description 10
- 201000010099 disease Diseases 0.000 description 10
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 10
- 230000003612 virological effect Effects 0.000 description 10
- 238000006386 neutralization reaction Methods 0.000 description 9
- 230000002441 reversible effect Effects 0.000 description 9
- 210000002845 virion Anatomy 0.000 description 9
- 101710172711 Structural protein Proteins 0.000 description 8
- 239000012634 fragment Substances 0.000 description 8
- 238000002347 injection Methods 0.000 description 8
- 239000007924 injection Substances 0.000 description 8
- 108091032973 (ribonucleotides)n+m Proteins 0.000 description 7
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 7
- 238000003556 assay Methods 0.000 description 7
- 239000012091 fetal bovine serum Substances 0.000 description 7
- 230000006698 induction Effects 0.000 description 7
- 239000002609 medium Substances 0.000 description 7
- 239000000047 product Substances 0.000 description 7
- 238000002415 sodium dodecyl sulfate polyacrylamide gel electrophoresis Methods 0.000 description 7
- 239000006144 Dulbecco’s modified Eagle's medium Substances 0.000 description 6
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 6
- 241000699666 Mus <mouse, genus> Species 0.000 description 6
- LOKCTEFSRHRXRJ-UHFFFAOYSA-I dipotassium trisodium dihydrogen phosphate hydrogen phosphate dichloride Chemical compound P(=O)(O)(O)[O-].[K+].P(=O)(O)([O-])[O-].[Na+].[Na+].[Cl-].[K+].[Cl-].[Na+] LOKCTEFSRHRXRJ-UHFFFAOYSA-I 0.000 description 6
- 230000005764 inhibitory process Effects 0.000 description 6
- 230000003472 neutralizing effect Effects 0.000 description 6
- 239000002953 phosphate buffered saline Substances 0.000 description 6
- 230000000644 propagated effect Effects 0.000 description 6
- 241000255925 Diptera Species 0.000 description 5
- 108700026244 Open Reading Frames Proteins 0.000 description 5
- 206010058874 Viraemia Diseases 0.000 description 5
- 230000008901 benefit Effects 0.000 description 5
- 210000004899 c-terminal region Anatomy 0.000 description 5
- 239000013553 cell monolayer Substances 0.000 description 5
- 238000006243 chemical reaction Methods 0.000 description 5
- 239000000499 gel Substances 0.000 description 5
- 230000001404 mediated effect Effects 0.000 description 5
- 241000710929 Alphavirus Species 0.000 description 4
- 241000271566 Aves Species 0.000 description 4
- 241000235058 Komagataella pastoris Species 0.000 description 4
- 241000700159 Rattus Species 0.000 description 4
- 208000035415 Reinfection Diseases 0.000 description 4
- 210000000234 capsid Anatomy 0.000 description 4
- 230000000295 complement effect Effects 0.000 description 4
- 239000002299 complementary DNA Substances 0.000 description 4
- 230000000120 cytopathologic effect Effects 0.000 description 4
- 238000005516 engineering process Methods 0.000 description 4
- 210000003743 erythrocyte Anatomy 0.000 description 4
- 239000012669 liquid formulation Substances 0.000 description 4
- 230000001105 regulatory effect Effects 0.000 description 4
- 239000012679 serum free medium Substances 0.000 description 4
- 230000017105 transposition Effects 0.000 description 4
- 230000009385 viral infection Effects 0.000 description 4
- 241000272814 Anser sp. Species 0.000 description 3
- 208000002109 Argyria Diseases 0.000 description 3
- 208000006820 Arthralgia Diseases 0.000 description 3
- 241000282693 Cercopithecidae Species 0.000 description 3
- 101900351172 Chikungunya virus Structural polyprotein Proteins 0.000 description 3
- FBPFZTCFMRRESA-FSIIMWSLSA-N D-Glucitol Natural products OC[C@H](O)[C@H](O)[C@@H](O)[C@H](O)CO FBPFZTCFMRRESA-FSIIMWSLSA-N 0.000 description 3
- FBPFZTCFMRRESA-KVTDHHQDSA-N D-Mannitol Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-KVTDHHQDSA-N 0.000 description 3
- FBPFZTCFMRRESA-JGWLITMVSA-N D-glucitol Chemical compound OC[C@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-JGWLITMVSA-N 0.000 description 3
- 108020004414 DNA Proteins 0.000 description 3
- 206010012310 Dengue fever Diseases 0.000 description 3
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 3
- 229930195725 Mannitol Natural products 0.000 description 3
- 241000235648 Pichia Species 0.000 description 3
- 239000004480 active ingredient Substances 0.000 description 3
- 235000001014 amino acid Nutrition 0.000 description 3
- 150000001413 amino acids Chemical class 0.000 description 3
- BFNBIHQBYMNNAN-UHFFFAOYSA-N ammonium sulfate Chemical compound N.N.OS(O)(=O)=O BFNBIHQBYMNNAN-UHFFFAOYSA-N 0.000 description 3
- 229910052921 ammonium sulfate Inorganic materials 0.000 description 3
- 235000011130 ammonium sulphate Nutrition 0.000 description 3
- 239000001166 ammonium sulphate Substances 0.000 description 3
- 238000010171 animal model Methods 0.000 description 3
- 210000004556 brain Anatomy 0.000 description 3
- 230000001413 cellular effect Effects 0.000 description 3
- 208000025729 dengue disease Diseases 0.000 description 3
- 230000000694 effects Effects 0.000 description 3
- RTZKZFJDLAIYFH-UHFFFAOYSA-N ether Substances CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 3
- 230000012010 growth Effects 0.000 description 3
- 239000000594 mannitol Substances 0.000 description 3
- 235000010355 mannitol Nutrition 0.000 description 3
- 239000003550 marker Substances 0.000 description 3
- 229940068196 placebo Drugs 0.000 description 3
- 239000000902 placebo Substances 0.000 description 3
- 229920001184 polypeptide Polymers 0.000 description 3
- 238000013207 serial dilution Methods 0.000 description 3
- 230000000405 serological effect Effects 0.000 description 3
- 239000002356 single layer Substances 0.000 description 3
- 239000000243 solution Substances 0.000 description 3
- 239000000600 sorbitol Substances 0.000 description 3
- 108090000565 Capsid Proteins Proteins 0.000 description 2
- 241000700198 Cavia Species 0.000 description 2
- 102100023321 Ceruloplasmin Human genes 0.000 description 2
- 208000001490 Dengue Diseases 0.000 description 2
- SXRSQZLOMIGNAQ-UHFFFAOYSA-N Glutaraldehyde Chemical compound O=CCCCC=O SXRSQZLOMIGNAQ-UHFFFAOYSA-N 0.000 description 2
- 206010061192 Haemorrhagic fever Diseases 0.000 description 2
- 208000032843 Hemorrhage Diseases 0.000 description 2
- 108091034057 RNA (poly(A)) Proteins 0.000 description 2
- 238000003917 TEM image Methods 0.000 description 2
- 108090000631 Trypsin Proteins 0.000 description 2
- 102000004142 Trypsin Human genes 0.000 description 2
- COQLPRJCUIATTQ-UHFFFAOYSA-N Uranyl acetate Chemical compound O.O.O=[U]=O.CC(O)=O.CC(O)=O COQLPRJCUIATTQ-UHFFFAOYSA-N 0.000 description 2
- 230000010530 Virus Neutralization Effects 0.000 description 2
- 229940037003 alum Drugs 0.000 description 2
- 230000003321 amplification Effects 0.000 description 2
- 230000005875 antibody response Effects 0.000 description 2
- 230000002238 attenuated effect Effects 0.000 description 2
- 210000004369 blood Anatomy 0.000 description 2
- 239000008280 blood Substances 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 230000034994 death Effects 0.000 description 2
- 238000011161 development Methods 0.000 description 2
- 238000001493 electron microscopy Methods 0.000 description 2
- 238000000605 extraction Methods 0.000 description 2
- 239000001963 growth medium Substances 0.000 description 2
- 125000000487 histidyl group Chemical group [H]N([H])C(C(=O)O*)C([H])([H])C1=C([H])N([H])C([H])=N1 0.000 description 2
- 230000001900 immune effect Effects 0.000 description 2
- 238000010166 immunofluorescence Methods 0.000 description 2
- 230000005847 immunogenicity Effects 0.000 description 2
- 238000011081 inoculation Methods 0.000 description 2
- 238000007912 intraperitoneal administration Methods 0.000 description 2
- BPHPUYQFMNQIOC-NXRLNHOXSA-N isopropyl beta-D-thiogalactopyranoside Chemical compound CC(C)S[C@@H]1O[C@H](CO)[C@H](O)[C@H](O)[C@H]1O BPHPUYQFMNQIOC-NXRLNHOXSA-N 0.000 description 2
- 238000000464 low-speed centrifugation Methods 0.000 description 2
- 238000002156 mixing Methods 0.000 description 2
- 238000003199 nucleic acid amplification method Methods 0.000 description 2
- 230000009465 prokaryotic expression Effects 0.000 description 2
- 238000011084 recovery Methods 0.000 description 2
- 230000009467 reduction Effects 0.000 description 2
- 230000035945 sensitivity Effects 0.000 description 2
- 239000013605 shuttle vector Substances 0.000 description 2
- 239000007787 solid Substances 0.000 description 2
- 239000002904 solvent Substances 0.000 description 2
- 241000894007 species Species 0.000 description 2
- 238000001228 spectrum Methods 0.000 description 2
- 238000007447 staining method Methods 0.000 description 2
- 239000006228 supernatant Substances 0.000 description 2
- 208000024891 symptom Diseases 0.000 description 2
- 238000004448 titration Methods 0.000 description 2
- 238000001890 transfection Methods 0.000 description 2
- 239000012588 trypsin Substances 0.000 description 2
- 108010087967 type I signal peptidase Proteins 0.000 description 2
- 238000001262 western blot Methods 0.000 description 2
- LINMATFDVHBYOS-MBJXGIAVSA-N (2s,3r,4s,5r,6r)-2-[(5-bromo-1h-indol-3-yl)oxy]-6-(hydroxymethyl)oxane-3,4,5-triol Chemical compound O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@H]1OC1=CNC2=CC=C(Br)C=C12 LINMATFDVHBYOS-MBJXGIAVSA-N 0.000 description 1
- 102000040650 (ribonucleotides)n+m Human genes 0.000 description 1
- JKMHFZQWWAIEOD-UHFFFAOYSA-N 2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid Chemical compound OCC[NH+]1CCN(CCS([O-])(=O)=O)CC1 JKMHFZQWWAIEOD-UHFFFAOYSA-N 0.000 description 1
- ZOOGRGPOEVQQDX-UUOKFMHZSA-N 3',5'-cyclic GMP Chemical compound C([C@H]1O2)OP(O)(=O)O[C@H]1[C@@H](O)[C@@H]2N1C(N=C(NC2=O)N)=C2N=C1 ZOOGRGPOEVQQDX-UUOKFMHZSA-N 0.000 description 1
- 241000256111 Aedes <genus> Species 0.000 description 1
- 229920001817 Agar Polymers 0.000 description 1
- 241000283690 Bos taurus Species 0.000 description 1
- OYPRJOBELJOOCE-UHFFFAOYSA-N Calcium Chemical compound [Ca] OYPRJOBELJOOCE-UHFFFAOYSA-N 0.000 description 1
- 241000283707 Capra Species 0.000 description 1
- 241000700199 Cavia porcellus Species 0.000 description 1
- 241000288673 Chiroptera Species 0.000 description 1
- 241000282552 Chlorocebus aethiops Species 0.000 description 1
- 241000867607 Chlorocebus sabaeus Species 0.000 description 1
- 241000272201 Columbiformes Species 0.000 description 1
- 208000035473 Communicable disease Diseases 0.000 description 1
- 241000036171 Coquillettidia perturbans Species 0.000 description 1
- 241000699800 Cricetinae Species 0.000 description 1
- 206010011703 Cyanosis Diseases 0.000 description 1
- 102000004163 DNA-directed RNA polymerases Human genes 0.000 description 1
- 108090000626 DNA-directed RNA polymerases Proteins 0.000 description 1
- 206010011906 Death Diseases 0.000 description 1
- 241000725619 Dengue virus Species 0.000 description 1
- 101710201734 E3 protein Proteins 0.000 description 1
- 101710091045 Envelope protein Proteins 0.000 description 1
- 206010066919 Epidemic polyarthritis Diseases 0.000 description 1
- 241000283086 Equidae Species 0.000 description 1
- 208000010201 Exanthema Diseases 0.000 description 1
- 241000282326 Felis catus Species 0.000 description 1
- 241000287828 Gallus gallus Species 0.000 description 1
- 229930182566 Gentamicin Natural products 0.000 description 1
- CEAZRRDELHUEMR-URQXQFDESA-N Gentamicin Chemical compound O1[C@H](C(C)NC)CC[C@@H](N)[C@H]1O[C@H]1[C@H](O)[C@@H](O[C@@H]2[C@@H]([C@@H](NC)[C@@](C)(O)CO2)O)[C@H](N)C[C@@H]1N CEAZRRDELHUEMR-URQXQFDESA-N 0.000 description 1
- 108090000288 Glycoproteins Proteins 0.000 description 1
- 102000003886 Glycoproteins Human genes 0.000 description 1
- 206010053759 Growth retardation Diseases 0.000 description 1
- 206010061218 Inflammation Diseases 0.000 description 1
- 102100034349 Integrase Human genes 0.000 description 1
- 101710125507 Integrase/recombinase Proteins 0.000 description 1
- QNAYBMKLOCPYGJ-REOHCLBHSA-N L-alanine Chemical compound C[C@H](N)C(O)=O QNAYBMKLOCPYGJ-REOHCLBHSA-N 0.000 description 1
- 241000282562 Macaca radiata Species 0.000 description 1
- FYYHWMGAXLPEAU-UHFFFAOYSA-N Magnesium Chemical compound [Mg] FYYHWMGAXLPEAU-UHFFFAOYSA-N 0.000 description 1
- 240000003492 Neolamarckia cadamba Species 0.000 description 1
- 206010067482 No adverse event Diseases 0.000 description 1
- 101800000515 Non-structural protein 3 Proteins 0.000 description 1
- 241000710944 O'nyong-nyong virus Species 0.000 description 1
- 241000282520 Papio Species 0.000 description 1
- 241000287127 Passeridae Species 0.000 description 1
- 241001494479 Pecora Species 0.000 description 1
- 241000288906 Primates Species 0.000 description 1
- 101800000980 Protease nsP2 Proteins 0.000 description 1
- 101710188315 Protein X Proteins 0.000 description 1
- 206010037660 Pyrexia Diseases 0.000 description 1
- 101800001758 RNA-directed RNA polymerase nsP4 Proteins 0.000 description 1
- 108091081062 Repeated sequence (DNA) Proteins 0.000 description 1
- 241000283984 Rodentia Species 0.000 description 1
- 241000710942 Ross River virus Species 0.000 description 1
- 241000710961 Semliki Forest virus Species 0.000 description 1
- 229920002684 Sepharose Polymers 0.000 description 1
- 239000005708 Sodium hypochlorite Substances 0.000 description 1
- 101800001473 Spike glycoprotein E2 Proteins 0.000 description 1
- 108091027544 Subgenomic mRNA Proteins 0.000 description 1
- 101000865057 Thermococcus litoralis DNA polymerase Proteins 0.000 description 1
- 241000710924 Togaviridae Species 0.000 description 1
- KZSNJWFQEVHDMF-UHFFFAOYSA-N Valine Natural products CC(C)C(N)C(O)=O KZSNJWFQEVHDMF-UHFFFAOYSA-N 0.000 description 1
- 241000251539 Vertebrata <Metazoa> Species 0.000 description 1
- 108010087302 Viral Structural Proteins Proteins 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 210000000577 adipose tissue Anatomy 0.000 description 1
- 235000004279 alanine Nutrition 0.000 description 1
- 235000011124 aluminium ammonium sulphate Nutrition 0.000 description 1
- LCQXXBOSCBRNNT-UHFFFAOYSA-K ammonium aluminium sulfate Chemical compound [NH4+].[Al+3].[O-]S([O-])(=O)=O.[O-]S([O-])(=O)=O LCQXXBOSCBRNNT-UHFFFAOYSA-K 0.000 description 1
- 229960000723 ampicillin Drugs 0.000 description 1
- AVKUERGKIZMTKX-NJBDSQKTSA-N ampicillin Chemical compound C1([C@@H](N)C(=O)N[C@H]2[C@H]3SC([C@@H](N3C2=O)C(O)=O)(C)C)=CC=CC=C1 AVKUERGKIZMTKX-NJBDSQKTSA-N 0.000 description 1
- 239000003708 ampul Substances 0.000 description 1
- 210000004102 animal cell Anatomy 0.000 description 1
- 238000005349 anion exchange Methods 0.000 description 1
- 206010003246 arthritis Diseases 0.000 description 1
- 230000005540 biological transmission Effects 0.000 description 1
- 229960000074 biopharmaceutical Drugs 0.000 description 1
- 210000000988 bone and bone Anatomy 0.000 description 1
- 244000309464 bull Species 0.000 description 1
- 239000011575 calcium Substances 0.000 description 1
- 229910052791 calcium Inorganic materials 0.000 description 1
- 230000000747 cardiac effect Effects 0.000 description 1
- 230000030833 cell death Effects 0.000 description 1
- 239000006285 cell suspension Substances 0.000 description 1
- 230000008859 change Effects 0.000 description 1
- 239000013043 chemical agent Substances 0.000 description 1
- 210000003837 chick embryo Anatomy 0.000 description 1
- 210000004978 chinese hamster ovary cell Anatomy 0.000 description 1
- 230000000052 comparative effect Effects 0.000 description 1
- 238000012790 confirmation Methods 0.000 description 1
- 210000002808 connective tissue Anatomy 0.000 description 1
- 238000011109 contamination Methods 0.000 description 1
- 239000012228 culture supernatant Substances 0.000 description 1
- 206010061428 decreased appetite Diseases 0.000 description 1
- 238000002405 diagnostic procedure Methods 0.000 description 1
- 239000003085 diluting agent Substances 0.000 description 1
- 210000001840 diploid cell Anatomy 0.000 description 1
- 231100000673 dose–response relationship Toxicity 0.000 description 1
- 239000003937 drug carrier Substances 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 230000004064 dysfunction Effects 0.000 description 1
- 230000002500 effect on skin Effects 0.000 description 1
- 238000010828 elution Methods 0.000 description 1
- ZMMJGEGLRURXTF-UHFFFAOYSA-N ethidium bromide Chemical compound [Br-].C12=CC(N)=CC=C2C2=CC=C(N)C=C2[N+](CC)=C1C1=CC=CC=C1 ZMMJGEGLRURXTF-UHFFFAOYSA-N 0.000 description 1
- 229960005542 ethidium bromide Drugs 0.000 description 1
- 238000011156 evaluation Methods 0.000 description 1
- 201000005884 exanthem Diseases 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 210000002950 fibroblast Anatomy 0.000 description 1
- PGBHMTALBVVCIT-VCIWKGPPSA-N framycetin Chemical compound N[C@@H]1[C@@H](O)[C@H](O)[C@H](CN)O[C@@H]1O[C@H]1[C@@H](O)[C@H](O[C@H]2[C@@H]([C@@H](N)C[C@@H](N)[C@@H]2O)O[C@@H]2[C@@H]([C@@H](O)[C@H](O)[C@@H](CN)O2)N)O[C@@H]1CO PGBHMTALBVVCIT-VCIWKGPPSA-N 0.000 description 1
- 210000001035 gastrointestinal tract Anatomy 0.000 description 1
- 238000001641 gel filtration chromatography Methods 0.000 description 1
- 238000003205 genotyping method Methods 0.000 description 1
- 229960002518 gentamicin Drugs 0.000 description 1
- 235000011187 glycerol Nutrition 0.000 description 1
- 208000037824 growth disorder Diseases 0.000 description 1
- 231100000001 growth retardation Toxicity 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 210000002216 heart Anatomy 0.000 description 1
- 238000010211 hemagglutination inhibition (HI) assay Methods 0.000 description 1
- 108010028403 hemagglutinin esterase Proteins 0.000 description 1
- 208000031169 hemorrhagic disease Diseases 0.000 description 1
- 230000002008 hemorrhagic effect Effects 0.000 description 1
- 230000023597 hemostasis Effects 0.000 description 1
- 208000021760 high fever Diseases 0.000 description 1
- 230000002631 hypothermal effect Effects 0.000 description 1
- 125000001841 imino group Chemical group [H]N=* 0.000 description 1
- 210000004201 immune sera Anatomy 0.000 description 1
- 229940042743 immune sera Drugs 0.000 description 1
- 230000036039 immunity Effects 0.000 description 1
- 230000000951 immunodiffusion Effects 0.000 description 1
- 238000001114 immunoprecipitation Methods 0.000 description 1
- 230000004054 inflammatory process Effects 0.000 description 1
- 239000002054 inoculum Substances 0.000 description 1
- 230000003993 interaction Effects 0.000 description 1
- 230000000968 intestinal effect Effects 0.000 description 1
- 230000003834 intracellular effect Effects 0.000 description 1
- 238000010255 intramuscular injection Methods 0.000 description 1
- 239000007927 intramuscular injection Substances 0.000 description 1
- 238000005342 ion exchange Methods 0.000 description 1
- 150000002500 ions Chemical class 0.000 description 1
- 210000001503 joint Anatomy 0.000 description 1
- 229960000318 kanamycin Drugs 0.000 description 1
- SBUJHOSQTJFQJX-NOAMYHISSA-N kanamycin Chemical compound O[C@@H]1[C@@H](O)[C@H](O)[C@@H](CN)O[C@@H]1O[C@H]1[C@H](O)[C@@H](O[C@@H]2[C@@H]([C@@H](N)[C@H](O)[C@@H](CO)O2)O)[C@H](N)C[C@@H]1N SBUJHOSQTJFQJX-NOAMYHISSA-N 0.000 description 1
- 229930027917 kanamycin Natural products 0.000 description 1
- 229930182823 kanamycin A Natural products 0.000 description 1
- 210000003734 kidney Anatomy 0.000 description 1
- 239000010410 layer Substances 0.000 description 1
- 231100000518 lethal Toxicity 0.000 description 1
- 230000001665 lethal effect Effects 0.000 description 1
- 150000002632 lipids Chemical class 0.000 description 1
- 210000004185 liver Anatomy 0.000 description 1
- 230000005923 long-lasting effect Effects 0.000 description 1
- 210000004072 lung Anatomy 0.000 description 1
- 239000011777 magnesium Substances 0.000 description 1
- 229910052749 magnesium Inorganic materials 0.000 description 1
- 238000012423 maintenance Methods 0.000 description 1
- 206010025482 malaise Diseases 0.000 description 1
- 210000004962 mammalian cell Anatomy 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 230000007246 mechanism Effects 0.000 description 1
- 230000003278 mimic effect Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 230000000877 morphologic effect Effects 0.000 description 1
- 229940031348 multivalent vaccine Drugs 0.000 description 1
- 210000003205 muscle Anatomy 0.000 description 1
- 230000017074 necrotic cell death Effects 0.000 description 1
- 229940053050 neomycin sulfate Drugs 0.000 description 1
- 210000000653 nervous system Anatomy 0.000 description 1
- 230000007935 neutral effect Effects 0.000 description 1
- 238000010899 nucleation Methods 0.000 description 1
- 230000008520 organization Effects 0.000 description 1
- 230000007918 pathogenicity Effects 0.000 description 1
- 230000010412 perfusion Effects 0.000 description 1
- 239000000546 pharmaceutical excipient Substances 0.000 description 1
- 230000000704 physical effect Effects 0.000 description 1
- 238000000053 physical method Methods 0.000 description 1
- 238000011020 pilot scale process Methods 0.000 description 1
- 238000003752 polymerase chain reaction Methods 0.000 description 1
- 229920005862 polyol Polymers 0.000 description 1
- 150000003077 polyols Chemical class 0.000 description 1
- 229920000036 polyvinylpyrrolidone Polymers 0.000 description 1
- 239000001267 polyvinylpyrrolidone Substances 0.000 description 1
- 235000013855 polyvinylpyrrolidone Nutrition 0.000 description 1
- 238000012809 post-inoculation Methods 0.000 description 1
- 239000003755 preservative agent Substances 0.000 description 1
- 230000002335 preservative effect Effects 0.000 description 1
- 230000035755 proliferation Effects 0.000 description 1
- 230000001902 propagating effect Effects 0.000 description 1
- 238000003908 quality control method Methods 0.000 description 1
- 238000003127 radioimmunoassay Methods 0.000 description 1
- 206010037844 rash Diseases 0.000 description 1
- 238000000472 rate-zonal centrifugation Methods 0.000 description 1
- 238000003259 recombinant expression Methods 0.000 description 1
- 230000010076 replication Effects 0.000 description 1
- 230000004044 response Effects 0.000 description 1
- 238000003757 reverse transcription PCR Methods 0.000 description 1
- 238000009938 salting Methods 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- 238000004062 sedimentation Methods 0.000 description 1
- 238000012163 sequencing technique Methods 0.000 description 1
- 210000002027 skeletal muscle Anatomy 0.000 description 1
- 210000003491 skin Anatomy 0.000 description 1
- SUKJFIGYRHOWBL-UHFFFAOYSA-N sodium hypochlorite Chemical compound [Na+].Cl[O-] SUKJFIGYRHOWBL-UHFFFAOYSA-N 0.000 description 1
- 230000003381 solubilizing effect Effects 0.000 description 1
- 238000000527 sonication Methods 0.000 description 1
- 235000010356 sorbitol Nutrition 0.000 description 1
- 238000002336 sorption--desorption measurement Methods 0.000 description 1
- 230000009295 sperm incapacitation Effects 0.000 description 1
- 238000010561 standard procedure Methods 0.000 description 1
- 230000036435 stunted growth Effects 0.000 description 1
- 150000005846 sugar alcohols Chemical class 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- 238000004114 suspension culture Methods 0.000 description 1
- 230000009885 systemic effect Effects 0.000 description 1
- 210000001519 tissue Anatomy 0.000 description 1
- 230000000451 tissue damage Effects 0.000 description 1
- 231100000827 tissue damage Toxicity 0.000 description 1
- 231100000419 toxicity Toxicity 0.000 description 1
- 230000001988 toxicity Effects 0.000 description 1
- 230000009466 transformation Effects 0.000 description 1
- 238000004627 transmission electron microscopy Methods 0.000 description 1
- 230000014599 transmission of virus Effects 0.000 description 1
- 210000003932 urinary bladder Anatomy 0.000 description 1
- 239000004474 valine Substances 0.000 description 1
- 125000002987 valine group Chemical group [H]N([H])C([H])(C(*)=O)C([H])(C([H])([H])[H])C([H])([H])[H] 0.000 description 1
- 238000012795 verification Methods 0.000 description 1
- 239000005723 virus inoculator Substances 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/005—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from 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
- 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
-
- 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
- C12N7/00—Viruses; Bacteriophages; Compositions thereof; Preparation or purification thereof
-
- 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
- A61K2039/525—Virus
- A61K2039/5252—Virus inactivated (killed)
-
- 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
- C12N2710/14011—Baculoviridae
- C12N2710/14111—Nucleopolyhedrovirus, e.g. autographa californica nucleopolyhedrovirus
- C12N2710/14141—Use of virus, viral particle or viral elements as a vector
- C12N2710/14143—Use of virus, viral particle or viral elements as a vector viral genome or elements thereof as genetic vector
-
- 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
- C12N2770/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA ssRNA viruses positive-sense
- C12N2770/00011—Details
- C12N2770/36011—Togaviridae
- C12N2770/36111—Alphavirus, e.g. Sindbis virus, VEE, EEE, WEE, Semliki
- C12N2770/36122—New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
-
- 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
- C12N2770/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA ssRNA viruses positive-sense
- C12N2770/00011—Details
- C12N2770/36011—Togaviridae
- C12N2770/36111—Alphavirus, e.g. Sindbis virus, VEE, EEE, WEE, Semliki
- C12N2770/36134—Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
-
- 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
- C12N2770/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA ssRNA viruses positive-sense
- C12N2770/00011—Details
- C12N2770/36011—Togaviridae
- C12N2770/36111—Alphavirus, e.g. Sindbis virus, VEE, EEE, WEE, Semliki
- C12N2770/36151—Methods of production or purification of viral material
-
- 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
- C12N2770/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA ssRNA viruses positive-sense
- C12N2770/00011—Details
- C12N2770/36011—Togaviridae
- C12N2770/36111—Alphavirus, e.g. Sindbis virus, VEE, EEE, WEE, Semliki
- C12N2770/36161—Methods of inactivation or attenuation
- C12N2770/36163—Methods of inactivation or attenuation by chemical treatment
-
- 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 vaccine formulation capable of eliciting protective immune response against Chikungunya virus (CHIKV) infection in humans and other mammalian hosts.
- the immunogenic formulation comprises purified inactivated Chikungunya virus in a stable formulation.
- the method of adaptation and propagation of the virus in vitro in continuous cell culture in vaccine quality cell lines such as Vero and MRC-5 cells that are approved by FDA/National Regulatory Authority is provided. Purification and methods of inactivation of the virus are discussed.
- the method of preparation and administration of liquid and lyophilized formulations of the virus with added stabilizers is discussed.
- the inactivated virus preparation is non-infectious, immunogenic and elicits protective immune response in mammalian host.
- the immunogenic composition is formulated for in vivo administration to humans.
- the strategy of developing a subunit vaccine using the recombinant virus proteins. as antigens capable of ehciting protective immune response against Chikungunya- virus infection in mammalian hosts.
- the recombinant antigens could potentially find use in diagnosing for the presence of the virus.
- Chikungunya is a physically debilitating disease of humans mainly in Africa and Asia.
- the symptoms include abrupt onset of high fever, rash or hemorrhages, arthralgia and occasional involvement of the nervous system, heart and liver.
- the incapacitation is due to arthralgia, which can persist for years (Sarkar et al., 1965; Rao et al., 1965; Nimmannitya et al., 1969; Schuffenecker et al., 2006).
- the disease is caused by Chikungunya virus (CHIKV), and is spread by Aedes spp.
- the disease can be diagnosed by va ⁇ ous serological t°sts, but definitive ider ⁇ fication requires verification of the genetic material since man , closely related arboviruses cause similar disease. Treatment is only palliative and there is no commercially available vaccine.
- the virus harbors a single-stranded, positive sense RNA genome, and belongs to group
- a arboviruses along with Sindbis and Semliki Forest disease viruses in the alphavirus genus of Togaviridae family (Fauquet et al., 2005).
- the virion is 50-60 nm in size and is inactivated by 70% ethanol, 1% sodium hypochlorite, 2% glutaraldehyde, lipid solvents, moist or dry heat > 58° C, as well as drying.
- Genotyping suggests the existence of three clades: West African, East-Central-South- African and Asian.
- the Asian and African strains form closely-related clades that differ from each other in sequence, antigenicity and virus properties.
- the Asian isolates appear to be more conserved than either of the African clades (Powers et al., 2000; Schuffenecker et al., 2006).
- the recent Indian Ocean outbreak isolates show a characteristic change from Alanine to Valine in position 226 of the envelope glycopriten El from early to later phase of the disease, respectively (Schuffenecker et al., 2006). While the importance of this is not understood, an evolutionary advantage for the virus can be surmised. Not much is known about the viral proteins, their function or pathogenicity.
- the genome consists of -12 kilobases, with a 5' 7mG cap and the 3' poly(A) region, and a base composition of 30% A, 25% C and G, and 20% U.
- the genome has the sequence of 5'-nsPl-nsP2-nsP3-nsP4-(junction region)-C-E3-E2-6K-El-polyA-3'.
- the nonstructural proteins are translated directly from the 5' two-thirds of the genome, and the structural proteins are produced from the 26S subgenomic RNA which is col linear with the 3' one-third of the genome.
- nsPl, nsP2, nsP3, nsP4, C, E3, E2, 6K and El proteins contain 535, 798, 530, 611, 261, 64, 423 and 61 amino acids (Khan et al., 2002; Schuffenecker et al., 2006).
- the envelope proteins can be observed on SDS gels as 62 kD E2/E3.
- Serum with HI titers of >40 generally shows neutralization capacity (Bedekar and Pavri, 1969b). Isolation of virus can be performed in newborn rats or mice, or in animal or insect cell cultures. Vero, African green monkey kidney, BHK21, BSC-I, chick embryo fibroblasts and C6/36 cells have been used for in vitro virus isolation and expansion. The virus replicates fairly rapidly in cell culture. Depending on dose and on the cell line, cytopathic effect can be observed in 12-48 hrs. At multiplicities of 1-5, following a 5-6 hr eclipse period, the intracellular virus titer rises sharply and reaches peak by 12 hrs.
- Extracellular virus can be observed at 8 hrs post-infection and peaks at 12-24 his depending on the cell system and dose of the inoculum (Chain et al., 1966; Higashi et al., 1967; Hahon and Hankins, 1970; Eckels et al., 1970).
- Spread of infection through the monolayer differs in different cell types-, involving both extracellular and cell-to- cell transmission in BHK21 cells, but only the former in L929 and guinea pig lung " cells (Hahon and Zimmerman, 1970). Titers in supernatants can reach as high as that observed with mouse brain preparations (Shah et al., 1964; Paul and Singh, 1968; Umrigar and Kadam, 1974).
- the virus can be concentrated from cell culture supernatant by ultracentrifugation, or precipitation with ammonium sulphate, alum, or polyethylene glycol (Eckels et al., 1970; Klein et al., 1970; Banerjee and Ranadive, 1988; Killington et al., 1996).
- the virus can be further purified by using rate zonal centrifugation, equilibrium density gradient or gel filtration (Eckels et al., 1970; Simizu et al., 1984; Banerjee and Ranadive, 1988). Titration of the virus can be performed by immunofluorescence, ELISA, complement fixation, agar gel immunodiffusion, hemagglutination and inhibition, plaque assays, or neutralization.
- CHIKV a ⁇ phaviruses cause arthritis. Suggested mechanisms include replication leading to cell death and tissue damage, immune- mediated attack on the joints, or lmmune-comptex-mediated inflammation. While Semliki forest virus and Ross River virus have been shown to known to replicate in bone-associated connective tissue in neonates as w .11 as skin and muscle in adult mice (Heise et al., 2000), no such study has been done with CHIKV.
- CHIKV In contrast to Dengue virus, which requires adaptation for infection of animals, CHIKV shows rapid and high fatality on primary inoculation of clinical samples (Myers et al., 1965). However, newborn mice and rats are the only species that show disease (Chakravarthy and Sarkar, 1969). In a dose-dependent manner, rat/mouse pups show illness and high mortality following intracerebral, intraperitoneal or subcutaneous inoculation of patient sera in 3-10 days, yielding 10 5 5 -10 7 0 mouse-LD 5 o of virus per mL of serum (Shah et al., 1964). Animals rapidly manifest sluggishness, severe inappetence, cyanosis, dermal hypothermia, and death.
- mice, rats, guinea pigs, hamsters, hare and rabbits show viremia, but not disease, and develop HI and neutralizing antibodies, whereas adult cats and fowl don't.
- Low titers of HI antibodies without viremia or neutralizing antibodies can be seen in cows, sheep, goats and horses (Mclntosh et al., 1963; Bedekar and Pavri, 1969a; Chakravarthy and Sarkar, 1969).
- the susceptibility of birds is controversial. In one study, white leghorn chicks have been shown to succumb to virus inoculation, and recovered birds develop neutralization antibodies (Bedekar and Pavri, 1969a).
- CHIKV infection (whether clinical or silent) is thought to confer life-long immunity. Because of close antigenic relationship, cross-protection between different strains (Casals, 1957; Porterfield, 1961 ; Shah et al., 1964) as well as reciprocal cross- protection among other alphaviruses (Parks and Price, 1958; Hearn and Rainey, 1963) can be hypothesized, and is demonstrated in animal models. However, there is some evidence that live attenuated alphavirus vaccines may interfere with a subsequent, related vaccine (McClain et al., 1998).
- CHIKV preparations involved either formalin inactivation (Harrison et al., 1967) or tween-ether extraction of virus grown in vitro (Eckels et al., 1970). While formalin kills HA activity, the latter treatment retains the HA activity completely, although both lose infectivity drastically. However, they both elicit similar HA and complement-fixing and neutralization antibodies and also show similar levels of protection in lethal challenge studies (Eckels et al., 1970).
- CHIKV strain 15561 from Thailand (1962 outbreak) was used to develop a small lot of green monkey passaged, formalin- inactivated preparation that was administered to 16 volunteers who showed high immune responses and no adverse effects (Harrison et al., 1971).
- the GMK-passaged virus was further serially passaged by plaqutng 18 times in MRC-5 cells (Levitt et al., 1986), and found to be safe and immunogenic in phase I trial with 15 alphavirus-na ⁇ ve individuals, viremia occurring on day 2-4 post-inoculation (McClain et al., 1988).
- Fig.l A- Control uninfected Vero cells B- CHIK infected Vero cells C- Control MRC-5 cells D- CHIKV infected MRC-5 Cells
- Fig.2 SDS-PAGE of purified CHIKV preparation depicting the El and E2 proteins that appears to be more predominant than the virus antigens.
- the virus sample was run in 12% denaturing SDS-PAGE gel and visualized by silver staining. The two proteins appear to be approximately 46 - 50 kE> in size.
- Fig.3 In vivo toxicity of the purified Chikungunya virus isolate CHK/03/06 after intracerebral injection in mice. 2- day old mice were injected purified Chikungunya virus isolate CHK/03/06 intracerebrally and the control animals were injected with equal volume of PBS.
- Panel B- mice injected with purified virus were found dead on the 6 th day after intracerebral injection of the purified virus preparation.
- Panel C On the 9 th day after intracerebral injection in 2-day old mice, the PBS injected control animal shows normal growth (left) and the mice injected with 1 :64 dilution of the purified virus preparation shows severe growth retardation.
- Fig.4 Transmission Electron Micrograph (TEM) of the CH K/03/06 isolate at 200 K magnification
- Fig.5 Ethidium bromide stained photograph of RT-PCR products of CHK/01/06 and CHK/03/06 virus isolates amplified with gene specific primers for the CHIK virus.
- Fig.6 Haemagglutination (HA) of the CHK/03/06 virus using fresh goose erythrocytes.
- the first two rows is the HA titer of the virus harvest from Vero cells in serial dilution.
- the last two rows is the HA titer of the serially diluted purified CHIK virus preparation showing an increase in the HA titer.
- Fig.7 RT-PCR of the viral structural proteins.
- Panel A Lane 1- Capsid PCR product; Lane 2- 100 bp ladder; Lane 3 - E2 PCR product
- Fig.8 SDS-PAGE of induction of cloned El protein in E.coli expressed cells. Lane 1 - uninduced cells; Lane 2 - protein molecular size marker; Lane 3 - E.coli cells 4 hours after induction with IPTG showing the induction of- 47 kD El protein.
- Fig.9 SDS-PAGE of the induced ⁇ 30 kD Capsid protein in Pichia pastoris; Lane 1 - protein expression at 0 hours after induction with methanol; Lane 2- at 72 hours after induction with methanol; Lane 3 - protein molecular size marker.
- the scope of the present invention includes the procedure for isolation of Chikungunya
- Regulatory Approved cell lines such as Vero and MRC-5 and harvesting the virus from the infected cells in culture.
- the methods described in the present invention are ap p licable to any genotype/strains/genetic variants of Chikungunya virus and is known to hose skilled in the art.
- the scope of the current invention induces methods used for the purification of the virus from the infected cells substantially free of cellular and serum components.
- One of the methods used in the current invention includes the use of proprietary HimaxTM technology for purification of the virus.
- the purified virus is inactivated either by heat or chemically inactivated with one of the inactivating agents that includes but is not limited to the following agents: formalin, beta-propiolactone, glutaraldehye, non-ionic detergents, ascorbic acid etc.
- the vaccine formulation comprises a pharmaceutically accepted buffer such as phosphate or phosphate-citrate buffer in the pH range 6.4 - 7.5 with added stabilizing agents that includes one or more of the following but is not limited to: human serum albumin, gelatin, reducing and non- reducing sugars, amino acids, polyols such as sorbitol and mannitol, glycerol organic and inorganic salts, polyvinyl pyrrolidone etc.
- the stable formulation in a liquid form is suitable for intramuscular/intradermal/subcutaneous/intravenous administration in a human host.
- the stable formulation in a dry lyophilized form can be reconstituted with a suitable solvent before administration.
- the formulations are suitable for oral and intranasal administration in humans.
- the efficacy of the vaccine formulation has been established by various methods such as virus neutralization test and hemagglutination inhibition assay.
- the estimation of the antigenic protein is by using standard protein estimation methods and by ELISA. Potency of the vaccine has been tested in animal models.
- the potency of the vaccine formulation has been tested in doses ranging from 1 ⁇ g upto 200 ⁇ g of the antigen where the inactivated virus is the active ingredient in the formulation.
- a high rate of seroconversion and protective antibodies against the virus has been observed in rabbits that have been administered the formulation.
- the adaptation and growth of the virus in a continuous cell culture in a cell line such as Vero/MRC-5 that is approved by FDA and National Regulatory authorities offers an affordable, reproducible and an easy to be validated method of propagation of Chikungunya virus in continuous cell culture suitable in industrial scale production of a stable formulation of the virus that can be used as a vaccine.
- the stable formulation of the inactivated Chikungunya virus with added stabilizers is immunogenic eliciting protective immune response against Chikungunya virus strains.
- the present invention also describes a strategy for developing a subunit vaccine using recombinant virus proteins as antigens.
- the viral proteins can be expressed as recombinant proteins either in prokaryotic or eukaryotic host cells.
- the methods of cloning and expression described herein are applicable to any genetic variants/mutants of the virus proteins.
- Chikungunya virus particles as an immunogen, adaptation and propagation of the virus in host cell lines to a high titer, determination of the identity of the virus by RT-PCR, electron microscopy; methods of purification and inactivation of the virus, preparation of stable liquid and lyophilized vaccine formulation(s) in a pharmaceutically acceptable carrier suitable for administration in human, viral methods and tests for vaccine potency in animal model have been discussed.
- the active ingredient or immunogen, of the vaccine described in the current invention is inactivated virus particles of Chikungunya virus.
- the methods described in the current invention are applicable to any genotype/strains of Chikungunya virus.
- the virus particles obtained from the clinical isolates from patients' serum have been adapted and propagated in vitro in cell monolayers for several passages. In alternative protocol, the virus particles have been passaged once through 2 day old suckling mice and the virus re-isolated and passaged in cell monolayers in vitro.
- the Chikungunya (CHIK) virus vaccine refers to an active ingredient (immunogen) of an inactivated Chikungunya virus that is produced under cGMP conditions by infecting in a monolayer of Vero cells or MRC-5 cells in continuous cell culture where the cells are used as host cells for culturing the Chikungunya virus.
- the cell line is qualified through various quality control tests and is approved by FDA/National Regulatory Authority as a vaccine quality cell line.
- the virus is propagated in large quantities by growing to a high titer ( ⁇ 10 9 ) in cell culture and purified from infected cells.
- the virus is inactivated either by heat or with an inactivating agent.
- a neutralizing antibody titer of the anti-serum obtained by immunization with the above particles in a stable f »rmulation, as measured by in vitro neutralization assay offers protective immunization in animals.
- the immunogen of a Chikungunya virus vaccine is a representative example of an immunogen of a vaccine against infectious diseases caused by Chikungunya viruses of any strain or genotypic variants of the Chikungunya virus.
- the vaccine of the present invention is provided in a sealed vial or ampoule in a liquid or lyophilized form.
- liquid formulation it can either be subcutaneously / intramuscularly / intradermally/intravenously injected or orally / intranasally administered to a subject to be vaccinated in an amount of about 0.05ml to 5 ml per person.
- a dry lyophilized formulation it is injected after being re-solubilized with a suitable solubilizing solution.
- the method that is applicable to strains of Chikungunya virus being used in the current invention is applicable to any Chikungunya virus strain with a broad antigenic spectrum.
- the broad spectrum antigenic response would offer a satisfactory immune protection against plural strains of CHIK virus in addition to the virus strain used in production of the vaccine.
- a divalent or polyvalent vaccine may be prepared by mixing vaccines produced from two or more CHIK virus strains that have been genetically confirmed as CHIK viruses and is mixed in a suitable ratio based on the antigenic protein content. Such mixing would provide a vaccine preparation having a broader antigenic spectrum for protection against the infection.
- Vero cclls/MRC-5 cells with the virus, and maintaining the infected cells in continuous
- a culturing method involves infecting the host cell monolayer with the virus and harvesting the virus in sufficient quantities from the infected host ceils.
- the virus grown in cell layers could include a population of the extracellular vims that is obtained in the supernatant of the infected cell culture that can be harvested by centrifugation, and harvesting the virus that is cell associated by sonication and centrifuga ⁇ on.
- a cell line that can be propagated in vitro in culture can be used as a host for virv" culture.
- diploid cell lines such as MRC-5 and WI-38 and serially passaged cell lines such as Vero, BHK-21 , CHO cells etc. can be used.
- permissive cells are selected which allow the virus to grow well.
- Vero ATCC No. CCL-81
- BHK-21 ATCC No. CCL-IO
- C6/C3 ATCC No. CRL- 1660
- One such cell line used in the current invention is Vero cells which have been validated for use as a host cell for vaccine production.
- the validated Vero cell lines conforms to the Requirements for Biological Substances No.50 regarding requirements for use of cells for the production of biologicals recommended by the World Health Organization (WHO) thereby confirming these cell lines as qualified for producing a vaccine (WHO Technical report Series, No. 878, pp 19-52, 1998).
- An inactivating agent such as formalin, beta-propiolactone, and glutaraldehyde is added to a virus suspension to inactivate the virus.
- the amount to be added is about 0.001% to 0.4% (v/v)
- the inactivation temperature is about 2-8 DEG C to about 40 DEG C and the inactivation duration mainly depends upon the inactivation temperature. For e.g. it could range between 2 hours to72 hours at 37 DEG C and about 12 hours to 250 hours at 2 - 8 DEG C.
- Inactivation is also effective at intermediate temperatures of around 22 DEG C for a period of 2-120 hours.
- Inactivation of the virus can also be carried out by non-ionic detergents and ascorbic acid.
- Heat inactivation is effective for Chikungunya virus at temperatures at around 56-58 DEG C for one hour.
- Purification of the virus is conducted by physical means or chemical means and preferably b; ' a combination of both.
- Physical methods utilize the physical properties of the virus such as density, size, mass, sedimentation coefficient etc. and includes any of the following techniques but is not limited to: zonal ultra-centrifugation, density gradient centrifugation, ultrafiltration with membranes with size cut offs ranging from 50 - 1000 kDa to remove serum and cellular components.
- Purification through chemical means employs methods such as adsorption/desorption through chemical or physioch ⁇ mical reactions and includes for example purification by ultracentrifugation, density gradient centrifugation, ion exchange chromatography, affinity chromatography, hydrophobic interaction chromatography, gel filtration chromatography, salting with inorganic salts one such example being ammonium sulphate, and by the use of proprietary HimaxTM technology, organic salts, organic solvents, aluminium phosphate, aluminium hydroxide and organic compounds such as .polyethylene glycol. Purification of the virus is achieved by either one or a combination of two or more of the above mentioned methods.
- Concentration of the virus is achieved by low - speed centrifugation with ultrafiltration membrane, precipitation with salts or purification with Himax rM Technology Inactivation of the virus particles can be achieved either before or after purification of the virus.
- the virus can be visualized by a negative staining method using 2% (w/v) uranyl acetate can be observed under an electron microscope at a magnification of about 20,000 to about 200,000.
- the genetic identity of the CHIK virus was confirmed by sequencing the cDNA complementary to the genomic RNA of the strains. More specifically, genomic RNA was extracted from virus infected cells or from the purified CHIKV after pelleting by ultracentrifugation or isolation on a density gradient. Thereafter, a region encoding an envelope proicin or non-structural protein was further amplified by reverse ⁇ transcriptase polymerase chain reaction (RT-PCR) using a pair of primers and the base sequence of the resultant cDNA is determined by dideoxy chain termination method. The imino acid sequence encoded by the base sequence was decoded using universal codes and the genetic identity of the virus was confirmed as Chikungunya virus.
- RT-PCR reverse ⁇ transcriptase polymerase chain reaction
- An inactivated virus particle of the present invention is diluted with any suitable diluent that is pharmaceutically acceptable so as to obtain the desired titer.
- the buffer used in the formulation may be phosphate or phosphate-citrate buffer.
- a vaccine may optionally contain preservative(s), stabilizer(s) etc. Reducing and non-reducing sugars, sugar alcohols such sorbitol and mannitol, glycerol, amino acids, human serum albumin is added in the range of 0.01% to 10% for the liquid formulation and upto 60% of the total solids for a lyophilized formulation.
- Such a stable formulation of the immunogen either in a liquid or in a lyophilized form and after reconstitution in a pharmaceutically acceptable buffer or water is suitable for administration intradermally/subcutaneously/intramuscularly/intravenously in human host and is also suitable for oral and intranasal administration.
- the vaccine for Chikungunya virus alternatively could be a subunit vaccine prepared with the structural proteins such as, Capsid, El and E2 and E3 glycoproteins of the Chikungunya virus strains either cloned and expressed individually and purified for constituting a vaccine formulation, or by expressing the entire structural polyprotein consisting of the Capsid, E3, E2, 6K polypeptide and El proteins that is cloned as a single polypeptide with a potential to assemble into a Virus Like Particle (VLP) in eukaryotic host cells such as yeast, in insect cells via baculovirus mediated expression, and in mammalian cells after being processed by host signal peptidases.
- VLP Virus Like Particle
- the VLPs are highly immunogenic as they mimic the native virus particle in structure when processed correctly for assembly by the host signal peptidases. They contain multiple copies of the antigenic proteins in their assembled srructure.
- the viral proteins either expressed in prokaryotic or eukaryotic host cells are purified free of host cell proteins.
- the purified structural proteins can be administered in a formulation and by the methods described above. The addition of Histidine residues to either end of the recombinant proteins while cloning facilitates purification of the recombinant proteins.
- virus particles obtained according to the present invention, as well the recombinant viral proteins can be used as a reagent (as an antigen) in the diagnostic tests e.g. as an antigen in an immunoprecipitation method, a hemagglutination inhibition (HI) test, complement fixation (CF) reaction, ELISA, radioimmunoassay, immunofluorescence, Western Blot and the like. More specifically using the entire or a part of an inactivated virus particle of the present invention, a diagnostic assay with high sensitivity and specificity for detecting infection by different strains of the Chikungunya virus can be provided.
- a diagnostic assay with high sensitivity and specificity for detecting infection by different strains of the Chikungunya virus can be provided.
- a part of an inactivated virus particle refers to a fraction of the virus which retains desired antigenicity and is derived from the virus particles including for example, structural protein solubilized during the purification step described in the given examples or expressed in a recombinant expression system.
- Polyclonal antibodies or monoclonal antibodies specific for the virus can be used in a diagnostic assay for Chikungunya virus infection.
- the vaccine' formulations were tested in Balb/c mice 11 and rabbits.
- the animals in each group were injected intrape ⁇ toneally with about 0.2 to 0.5ml /mouse of serially diluted vaccine preparation ranging in dose from 1 ⁇ g to about 500 ⁇ g for the different test groups.
- a booster dose was given 7 - 14 days after the first administration of the antigen.
- a formulation of the inactivated virus preparation formulated in aluminium hydroxide in combination with oligonucleotides gave a higher immune response.
- the resultant serum was assayed by in vitro neutralization tests and the antibody titer was determined by ELISA. Seroconversion was observed in the animals immunized with the vaccine formulation.
- Vera cell line (ATCC No. CCL-81), BHK- 21 cells and MRC-5 cells were used as candidate cell lines for Chikungunya virus (CHIKV) culture and good viral propagation in each cell line was observed.
- Vero cells and BHK-21 cells were each prepared in a growth medium consisting of DMEM (Dulbecco's Modified Eagle Medium; Sigma-Aldrich Catalog # D5523 and used per the manufacturer's instructions) containing 5% fetal bovine serum (FBS). They were statically incubated at 37° C until reaching 80 - 100 % confluence of the monolayer. Thereafter the number of cells was counted.
- DMEM Dulbecco's Modified Eagle Medium
- FBS fetal bovine serum
- MRC-5 cells were prepared in growth medium consisting of MEM (Minimal Eagle's Medium) buffered to neutral pH with Hepes buffer and consisting of 5% FBS, was statically incubated at 37 0 C for 6 days and thereafter the cells were counted. In an alternate procedure, Vero cells and MRC-5 cells were cultured in serum free medium. For scaling the production of cells for virus infection, one cryo vial containing 5 x 10 6 viable Vero cells from working cell bank were used for seeding one T 175 Cell culture grade flask. DMEM containing 5 % FBS and 50 ⁇ g/ml of neomycin sulfate was used for revival and replenishing the cells.
- MEM Minimum Eagle's Medium
- AfteF 90 % of the confluence of the cell monolayer in T 175 flasks the cells were trypsinized and propagated further in cell factories / cell stacks (CFlO).
- the same DMEM medium is used for propagation ( ⁇ 2.0 L/ CFlO).
- Chikungunya virus was isolated from serum of infected patients who had the classical clinical symptoms of Chikungunya virus infection.
- the Indian isolate(s) of the virus were used in the development of the vaccine formulation.
- two isolates namely, CHK/03/06 and CHK/01/06 were characterized.
- the blood sample drawn from the patient was transported at 4 0 C and the serum was separated.
- the serum was diluted 1 :1 with phosphate buffered saline (PBS) and 0.5 ml of the diluted serum was used for infection of Vero cells (ATCC No. CCL-81) in 25 2 cm flask and incubated at 34 0 C to 37 0 C in serum free medium.
- PBS phosphate buffered saline
- the control fiask of Vero cells was treated with equal volume of Ix PBS.
- the medium for propagation of Vero cells was DMEM containing 1% fetal bovine serum (FBS)
- FBS fetal bovine serum
- the cell and virus culture were carried out in serum free medium.
- the virus was harvested 30 - 48 hours after infection.
- the cytopathic effect (cpe) of Chikungunya viru-' isolate CHK/03/06 in infected Vero cells was as observed in F g. 1 B, and IA is the uninfected control.
- the virus was also cultured under similar conditions in MRC-5 cells and the cytopathic effect of CHK/03/06 observed in MRC-5 cells is depicted in Fig. 1 D.
- FIG. 1C is the uninfected control MRC-5 cells.
- the cpe of the virus in BHK21 cells is observed in Fig.l F and that of the control uninfected BHK-21 cells is depicted in Fig. 1 E.
- Vero and MRC-5 cells were treated with chemical agents such as trypsin at low concentrations ranging from 0.01% to 1% and with or without divalent ions such as calcium and magnesium before infection.
- CHK/03/06 virus isolates were propagated in continuous cell culture in Vero cells (ATCC No. CCL-81) and in BHK-21 cells and MRC-5 cells.
- the medium for the infection was DMEM containing 0% - 1% FBS.
- the cytopathic effect was near total in Vero and BHK21 cells and the virus was present largely in the extracellular medium.
- a TCID 50 of the virus upto 10 9 /ml was obtained.
- the virus isolates were passaged serially 23 times in Vero cells and were found to be stable in continuous cell culture. The virus titer significantly enhanced after passaging through mouse brain. Virus infection and propagation were also alternately tested in a serum free medium.
- EXAMPLE 4 Purification of the virus: The two virus isolates were purified from the infected Vero cell monolayers by methods that include but not is limited to: low-speed centrifugation to remove much of the cellular debris and serum components, ultracentrifugation, sucrose density gradient centrifugation, and ultrafiltration through 100 - 1000 kDa membrane. The virus was further purified by ion exchange column chromatography and by gel filtration in Sepharose CL 4B or a matrix of similar property. The fractions containing the virus were pooled and precipitated with salts that included one of the following: PEG in the presence of 0.020M - 0.20 M NaCl, precipitation using HimaxTM technology, ammonium sulphate, alum etc.
- the infectivity of the virions was checked by re-infection of the Vero, BHK 21 and MRC-5 cells and was found to b? infectious.
- the concentrated virus was resuspended in phosphate buffer, pH 7.2 for checking in 7.5 % - 12% SDS-PAGE and virus proteins were visualized by silver staining.
- High purification of the virus was achieved also by ion exchange chromatography and hydrophobic interaction column chromatography and by the use of MonolithTM columns with salt elution.
- the purity of the virus preparation was checked by silver staining of SDS-PAGE gel and by Western blot using anti-CHIKV antisera raised in rabbit. Purification on the Monolith anion exchange columns gave a higher recovery of infectivity.
- Infectivity of the virus was also monitored during different steps of a pilot scale purification that eliminates serum and host cell contamination of the virus and TCID 50 of 10 5 to 10 7 /ml could be routinely obtained and is being further optimized.
- the purity of the virus preparation was checked by SDS-PAGE was found to be of good purity.
- the El and E2 envelope glycoproteins could be easily detected. See Fig 2.
- the purified virus was had infectivity as determined by determining the infectious count of the virus and by intracerebral injection of the purified virus fraction in 2-day old mice, at various dilutions. Neat purified virus and several serial dilutions resulted in death when injected intracerebrally. The mice showed severe retarded growth when compared to PBS injected control mice as observed in Fig.3
- Electron microscopy of the virus The Transmission Electron microscopy (Hitachi H- 7500) of the CHIK vims after ultracentrifugation was carried out by negative staining method using 2% uranyl acetate is depicted in Fig.4.
- RT-PCR Reverse-Transcription- Polymerase Chain Reaction
- the virus genomic RNA was extracted from the virus culture in Vero cells using standard protocols. The total RNA was reverse transcribed in different reactions using oligodT (I S mer) and by primers specific for Chikungunya virus sequences. Alternatively, the PCR was carried out on the reverse transcribed product using gene ⁇ ecific forward and reverse primers as given below. The PCR was carried out using Vent DNA polymerase (New England Biolabs). A few examples of the RT-PCR of the cDNA of CHIK virus ⁇ isolates are depicted in Fig. 5. The sequence of few of the forward primers and reverse primers used in the some of the PCR reactions are as follows:
- CHK SP FPl 5'CTAAATAGGTACGCACTACAGC 3'
- CHK SP FP2 5' TGGACTCCGCGCCCTACTATC 3'
- CHKSPFP3 5' TACTCAGGAGGCCGGTTCAC 3'
- CHK SP RP4 5' GTGTCCCATTGTTCCAG 3'
- CHK SP RP5 5' GTGAACCGGCCTCCTGAGTA 3'
- the RT-PCR amplified cDNA fragments were sequenced by dideoxy chain termination method.
- the deduced protein sequences were determined using universal genetic codes. Additional regions of the Chikungunya virus genome sequences were amplified with other gene specific piimers, and sequences of the amplified DNA were determined by dideoxy chain te ⁇ nination method.
- the sequence of the RT-PCR products confirmed the identity of the Chikungunya virus isolates.
- the purified virions of CHK/03/06 isolates were heat inactivated at temperatures ranging from 50 0 C - 60 0 C for 30 - 60 min.
- the infectivity of the virions was checked by re-infection of the Vero cells. No re-infection was observed.
- the purified virions of CHK/03/06 isolate were inactivated by chemical methods that includes but is not limited to one of the following methods: inactivation at concentrations ranging from 0.01% to 0-.5% formalin (formaldehyde) for 2 hours at 37 0 C followed by incubation at 4 0 C foi a period of 48-96 hours.
- the virions inactivated at the various concentrations of formalin were found to be non-infectious when re- infected in Vero cells after atleast three serial passages.
- the purified virions were inactivated w ith beta-propiolactone (BPL) at various concentrations ranging from 1 : 500 to 1 :3000 of the BPL: virus and for two hours at 37 DEG C, followed by incubation at varying time intervals of 48 - 200 hours at 4 0 C. No re-infection of the virus was observed after three serial passages of the virus at a ratio of BPL: virus at 1 :500 dilution to 1 : 1000 dilution. Inactivation of the virus could also be successfully carried out 22 DEG C for varying number of hours ranging from 48 -96 hrs. No infectivity was observed with the formalin and beta-propiolactone inactivated virions at the range of concentrations and for the various time periods tested.
- BPL beta-propiolactone
- the virus infection titer of CHK/03/06 isolate was counted in PFUs (plaque-forming units/ml) by a plaque-counting method using Vero cells and by determining the TCID 50 (Tissue culture infectious dose) by standard- protocols.
- the plaques could be ready by 36-48 hours and the titers could be determined by 30- 48 hours.
- various passages of the Vero-adapted virus ⁇ yielded titers ranging upto IO 8 3 TCID 50 / mk
- An increase in titer was observed after passaging the virus through 2-day old mouse brain.
- the vims was plaque purified from passage 2 sample in Vero cells.
- Chikungunya virus antigen of the CHK/03/06 isolate was measured by protein estimation by BCA method and by ELISA using rabbit anti-Chikungunya virus antisera. Polyclonal antibodies were raised in rabbit by injecting 100 ⁇ g of the purified Chikungunya virus CHK/03/06 isolate intradermally
- the hemagglutination titer was estimated by standard procedure using goose red blood cell suspension. Haemagglutination inhibition of the immune sera from rabbit and convalescent human sera were determined by standard protocols. Both the rabbit antisera and convalescent human sera showed haemagglutination inhibition.
- the HA titer of the purified virus was higher than the harvested neat virus sample. The HA titer of infected virus sample and the purified virus sample after serial dilutions are depicted in Fig. 6
- mice Fifteen one month old Balb/c mice were used in each group.' The animals in each group were injected intraperitoneally with about 0.2 ml to 0.5ml" /mouse of serially diluted vaccine preparation of the CHK/03/06 isolate ranging in dose from 1 ⁇ g to about 200 ⁇ g for the different test groups. The animals were boosted 14 days after the first immunization.- Blood was collected either at 7 and 14 days after the booster injection. Equal amount of serum was pooled for each group and complement was inactivated at 56 0 C for about 30 min. The resultant serum was used for neutralizing test as an immune serum and for estimation of antibody titer by ELISA. In an alternate experiment, similar amount of the inactivated virus preparation was administered intramuscularly.
- All the formulations contained the viral antigen along with aluminium hydroxide in 40 mM phosphate buffer, pH 6.8 - 7.2 containing 150 mM NaCl, 50 ⁇ g/dose oligonucleotides and varying amounts of excipients as mentioned in the Example below.
- Chikungunya virus antigen CHK/03/06 was prepared in 40 mM phosphate buffer, pH 6.9 - 7.2 with or without 154 mM NaCl.
- the viral preparations were formulated in a liquid formulation with aluminium hydroxide/aluminium phosphate containing eilher one or combination of sugars such as sucrose, maltose, trehalose, lactose, glucose, mannitol or sorbitol.
- sugars such as sucrose, maltose, trehalose, lactose, glucose, mannitol or sorbitol.
- the presence of sugars in the range of 0.5% -10% and preferably in the range of 0.5% to 5% conferred good stability on the formulation as determined by accelerated stability at 37 DEG C for three weeks.
- the CHK/03/06 virus isolate was used as the source for cloning and expression of all viral antigens with the sequences given in SEQ ID NO. i ⁇ to SEQ ID NO. 10.
- the complete open reading frame of the Chikungunya virus Structural polyprotein encoded by the SEQ ID NO.l was amplified by RT-PCR of the viral genomic RNA using the primers CHKCPFP as the forward primer and CHKElRP as the reverse primer to obtain a ⁇ 3776 bp PCR fragment.
- the PCR fragment was digested with Ndel and BamHl and cloned into the Ndel and BamHl sites of the prokaryotic expression vector, pETl IB and the recombinant plasmid containing the insert was transformed in E.coli DH5a.
- the open reading frame encoding the Chikungunya virus structural polyprotein, the SEQ ID NO.2 was cloned and expressed in a similar manner using CHKCKOZ AKFP as the forward primer and CHKE 1RP2 as the reverse primer to obtain a fragment of similar size.
- the primer sequence CHKCKOZAKFP has EcoRl site and CHKE1RP2 has a HindIII and Notl sites to facilitate cloning into baculovirus vector pFastBac (Invitrogen Corporation, Carlsbad, USA) and Pichia vector pPIC3.5K (Invitrogen Corporation, Carlsbad, USA) respectively.
- the SEQ ID NO. 2 was further amplified with a C-terminal primer that introduced 6- Histidine residu .s at the C-terminal end to obtain SEQ ID NO. 3.
- SEQ ID NO.l and SEQ ID NO.2 encode the protein of SEQ ID NO.4.
- SEQ ID NO. 3 encodes the protein of SEQ ID NO.5 that has the 6 His residues at the C-terminal end. The 6-Histidine residues at the C-terminal end of SEQ ID NO. 5, facilitates the purification of the expressed protein on Ni+ affinity column.
- Both SEQ ID No.4 and SEQ ID NO.5 have been expressed for assembly of the virus like particle in yeast and in baculovirus mediated expression in Sf9 cells
- the PCR gene fragments corresponding to SEQ ID NO.2, and SEQ ID NO.3 were digested with EcoRl and Not 1 and gel purified by standard protocols and cloned into EcoRl and Notl sites of the yeast expression vector pPIC3.5K (Invitrogen Corporation, Carlsbad, USA). The positive clones were selected after confirmation by PCR using the same primers.
- the recombinant plasmid encoding the complete structural protein of SEQ ID NO.4 and SEQ ID NO.5 was transformed into Pichia Pastoris GSl 15 as per the manufacturers (Invitrogen) instruction and as outlined in their protocols.
- the PCR amplified fragment has been cloned into the AOXl locus and expressed under the AOXl promoter by methanol induction.
- ORF Open Reading Frame
- the method utilizes a site specific transposition of the expression cassette such as the recombinant pFastBac vector with the cloned inserts as described above into a baculovirus shuttle vector (bacmiri) propagated in E.coli.
- Recombinant pFastBac vector containing one of the inserts SEQ ID NO.4 or SEQ ID NO.5 cloned under the control of ,he polyhedron promoter is transformed into competent cells of E.coli Max Efficiency DHlOBacTM, that contains a baculovirus shuttle vector (bMON 14272) and a helper plasmid (pMON7124) that facilitates transposition to allow efficient re-generation of the recombinant bacmid following the transposition of the pFastBac recombinant constructs containing the SEQ ID NO. 4 or SEQ ID NO. 5.
- E.coli Max Efficiency DHlOBacTM that contains a baculovirus shuttle vector (bMON 14272) and a helper plasmid (pMON7124) that facilitates transposition to allow efficient re-generation of the recombinant bacmid following the transposition of the pFastBac recombinant constructs containing
- the recombinant bacmid were selected on ampicillin, gentamicin and kanamycin containing plates by blue/white selection using bluo-gal and IPTG.
- the recombinant bacmid was isolated by standard protocols similar to that of isolation of plasmid DNA and 1 ⁇ g of the bacmid DNA was used for transfection with the use of CellfectinTM reagent into Sf9 insect cells that were grown in Grace's insect cell medium (Invitrogen Corporation, USA).
- the methods used for transfection, isolation and titration of Pl viral stock are exactly as described in the User's manual of Bac-to-Bac Baculovirus Expression system as given above.
- each of the structural antigens such as the Capsid, E3, E2, 6K polypeptide and El structural proteins corresponding to the following sequence IDs: SEQ ID NO.6, SEQ ID NO.7, SEQ ID NO.8, SEQ ID.N0.9 and SEQ ID NO. 10 respectively encoded in the structural polyprotein sequence were amplified by PCR using gene specific primers as outlined below in the first step of amplification.
- reverse primers that encode for 6 histidine residues were used for PCR (primer sequences not given) of each of the individual genes and cloned into pETl l B for prokaryotic expression in E.coli and in pFastBac vector for baculovirus mediated expression in insect cells.
- CHKE1RP2 5' AACAAGCTTGCGGCCGCTTAGTGCCTGCTGAACGACACG 3'
- CHKCPFP 5' ACAGAATTCATATGGAGTTCATCCCAACCCAAAC y
- CHKE2RP 5' TCCAAGCTTGGATCCTTACGCTTTAGCTGTTCTGATGCAGC 3 '
- the recombinant antigens expressed in either prokaryotic or eukaryotic expression system can be used for diagnostic purpose such as in ELISA.
- ELISA has been established using the inactivated whole virus and using rabbit antisera. Similar technique can be established using the purifed recombinant antigens instead of the whole virus as antigen.
- Polyclonal antisera or monoclonal antibody against the virus antigens particularly the structural antigens can be used as immunotherapeutic agent.
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Virology (AREA)
- Medicinal Chemistry (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Microbiology (AREA)
- Wood Science & Technology (AREA)
- Zoology (AREA)
- Biochemistry (AREA)
- Engineering & Computer Science (AREA)
- Immunology (AREA)
- Veterinary Medicine (AREA)
- Molecular Biology (AREA)
- Public Health (AREA)
- Animal Behavior & Ethology (AREA)
- Pharmacology & Pharmacy (AREA)
- Biomedical Technology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Mycology (AREA)
- Biophysics (AREA)
- Epidemiology (AREA)
- Gastroenterology & Hepatology (AREA)
- General Engineering & Computer Science (AREA)
- Biotechnology (AREA)
- Communicable Diseases (AREA)
- Oncology (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
- Micro-Organisms Or Cultivation Processes Thereof (AREA)
Abstract
Description
Claims
Priority Applications (4)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
BRPI0716145-0A BRPI0716145B1 (en) | 2006-09-01 | 2007-08-31 | VACCINE COMPOSITION AND METHOD FOR PREPARING A VACCINE COMPOSITION FOR CHIKUNGUNYA VIRUS |
CN200780035576.XA CN101516395B (en) | 2006-09-01 | 2007-08-31 | A vaccine for chikungunya virus infection |
EP07827554.2A EP2073839B1 (en) | 2006-09-01 | 2007-08-31 | A vaccine for chikungunya virus infection |
US12/439,509 US8865184B2 (en) | 2006-09-01 | 2007-08-31 | Vaccine for chikungunya virus infection |
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
IN1583/CHE/2006 | 2006-09-01 | ||
IN1583CH2006 | 2006-09-01 |
Publications (2)
Publication Number | Publication Date |
---|---|
WO2008026225A2 true WO2008026225A2 (en) | 2008-03-06 |
WO2008026225A3 WO2008026225A3 (en) | 2008-09-25 |
Family
ID=39136377
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
PCT/IN2007/000383 WO2008026225A2 (en) | 2006-09-01 | 2007-08-31 | A vaccine for chikungunya virus infection |
Country Status (5)
Country | Link |
---|---|
US (1) | US8865184B2 (en) |
EP (1) | EP2073839B1 (en) |
CN (1) | CN101516395B (en) |
BR (1) | BRPI0716145B1 (en) |
WO (1) | WO2008026225A2 (en) |
Cited By (19)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2009031045A2 (en) * | 2007-09-07 | 2009-03-12 | Institut Pasteur | Anti-chikungunya monoclonal antibodies and uses thereof |
WO2009131683A3 (en) * | 2008-04-21 | 2010-04-29 | Gen-Probe Incorporated | Method for detecting chikungunya virus |
WO2010062396A2 (en) | 2008-11-26 | 2010-06-03 | Government Of The United States Of America , As Represented By The Secretary, Department Of Health And Human Services | Virus like particle compositions and methods of use |
EP2274442A2 (en) * | 2008-04-04 | 2011-01-19 | The Trustees Of The University Of Pennsylvania | Consensus sequences of chikungunya viral proteins, nucleic acid molecules encoding the same, and compositions and methods for using the same |
EP2374816A1 (en) * | 2010-04-07 | 2011-10-12 | Humalys | Binding molecules against Chikungunya virus and uses thereof |
WO2011124635A1 (en) * | 2010-04-07 | 2011-10-13 | Humalys | Binding molecules against chikungunya virus and uses thereof |
EP2377927A1 (en) * | 2010-04-14 | 2011-10-19 | Millipore Corporation | Methods of producing high titer, high purity virus stocks and methods of use thereof |
EP2460822A3 (en) * | 2006-03-15 | 2012-09-26 | Institut Pasteur | Novel isolated and purified strains of chikungunya virus and polynucleotides and polypeptides sequences, diagnostic and immunogenical uses thereof |
WO2012073257A3 (en) * | 2010-11-30 | 2012-10-11 | Bharat Biotech International Limited | Vaccine formulation for prophylaxis and treatment of chandipura virus infections in mammals |
WO2012172574A1 (en) | 2011-06-17 | 2012-12-20 | Bharat Biotech International Limited | Vaccine composition comprising an inactivated chikungunya virus strain |
WO2014151855A1 (en) * | 2013-03-14 | 2014-09-25 | Inviragen, Inc. | Compositions and methods for live, attenuated alphavirus formulations |
WO2017009873A1 (en) * | 2015-07-16 | 2017-01-19 | Bharat Biotech International Limited | Vaccine compositions |
WO2017172643A1 (en) * | 2016-03-31 | 2017-10-05 | Takeda Vaccines, Inc. | Compositions and methods for stabilizing alphaviruses with improved formulations |
WO2017197034A1 (en) * | 2016-05-10 | 2017-11-16 | Najit Technologies, Inc. | Inorganic polyatomic oxyanions for protecting against antigenic damage during pathogen inactivation for vaccine production |
US10718771B2 (en) | 2018-02-09 | 2020-07-21 | Chung Yuan Christian University | Recombinant baculoviruses and their uses in detecting arthropod-borne virus |
US11478541B2 (en) | 2017-11-03 | 2022-10-25 | Takeda Vaccines, Inc. | Method for inactivating Zika virus and for determining the completeness of inactivation |
EP3393506B1 (en) * | 2015-12-23 | 2024-03-13 | Valneva SE | Virus purification |
US11975062B2 (en) | 2017-11-30 | 2024-05-07 | Takeda Vaccines, Inc. | Zika vaccines and immunogenic compositions, and methods of using the same |
US12011477B2 (en) | 2017-09-21 | 2024-06-18 | Valneva Se | Method of producing pharmaceutical compositions comprising immunogenic chikungunya virus CHIKV-delta5NSP3 |
Families Citing this family (12)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
DK2519266T3 (en) | 2009-12-31 | 2017-11-06 | Medigen Inc | Infectious DNA vaccines against Chikungunya virus |
CN103476788A (en) * | 2010-12-10 | 2013-12-25 | 新加坡科技研究局 | Immunogenic chikungunya virus peptides |
MA41517A (en) * | 2015-04-14 | 2017-12-19 | Univ Vanderbilt | NEUTRALIZATION OF THE ANTIBODY-MEDIATED CHIKUNGUNYA VIRUS |
WO2017123932A1 (en) * | 2016-01-13 | 2017-07-20 | United States, As Represented By The Secretary Of The Army | Inactived vaccine for chikungunya virus |
CN105906694B (en) * | 2016-05-05 | 2019-05-14 | 中国人民解放军军事医学科学院微生物流行病研究所 | Datum hole Kenya virus-specific detects antigen and its application |
WO2017210215A1 (en) | 2016-05-31 | 2017-12-07 | The Government Of The United States Of America As Represented By The Secretary Of The Army | Zika virus vaccine and methods of production |
CN111050795B (en) * | 2017-07-03 | 2023-09-22 | 巴拉特生物技术国际有限公司 | Vaccine compositions based on synthetic polypeptide epitopes |
EP3755709A1 (en) | 2018-02-22 | 2020-12-30 | Euroimmun Medizinische Labordiagnostika AG | Assay for the diagnosis of viral infections |
EP3530668A1 (en) | 2018-02-22 | 2019-08-28 | Euroimmun Medizinische Labordiagnostika AG | A novel assay for the diagnosis of viral infections |
BR102020012760A2 (en) | 2019-07-02 | 2022-03-08 | Euroimmun Medizinische Labordiagnostika Ag | ASSAY FOR THE DIAGNOSIS OF INFECTIONS CAUSED BY NEMATODES |
AU2020370603A1 (en) * | 2019-10-25 | 2022-05-12 | Bavarian Nordic A/S | Chikungunya virus-like particle vaccine and methods of using the same |
CN117771353B (en) * | 2024-02-27 | 2024-05-14 | 中国医学科学院医学生物学研究所 | MRNA vaccine of chikungunya virus with envelope protein as target and preparation method thereof |
Family Cites Families (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US20100233209A1 (en) * | 2005-08-11 | 2010-09-16 | Higgs Stephen T | Chikungunya virus infectious clones and uses therefor |
CA2545597A1 (en) * | 2006-03-15 | 2007-09-15 | Institut Pasteur | Novel isolated and purified strains of chikungunya virus and polynucleotides and polypeptides sequences, diagnostic and immunogenical uses thereof |
-
2007
- 2007-08-31 EP EP07827554.2A patent/EP2073839B1/en active Active
- 2007-08-31 BR BRPI0716145-0A patent/BRPI0716145B1/en active IP Right Grant
- 2007-08-31 US US12/439,509 patent/US8865184B2/en active Active
- 2007-08-31 WO PCT/IN2007/000383 patent/WO2008026225A2/en active Application Filing
- 2007-08-31 CN CN200780035576.XA patent/CN101516395B/en active Active
Non-Patent Citations (44)
Title |
---|
"8th Report of the International Committee on Taxonomy of Viruses", 2005 |
"ATCC Microbes and Cell at Work", 1991, AMERICAN TYPE CULTURE COLLECTION, pages: 144 |
BANERJEE K; RANADIVE SN: "Oligonucleotide fingerprinting of Chikungunya virus strains", IND. J. MED. RES., vol. 87, 1988, pages 531 - 541 |
BEDEKAR SD; PAVRI KM: "Studies with Chikungunya virus. Part I. Susceptibility of birds and small mammals", IND. J. MED. RES., vol. 57, 1969, pages 1181 - 1192 |
BEDEKAR SD; PAVRI KM: "Studies with Chikungunya virus. Part II. Serologiacl survey of humans and animals in India", IND. J. MED. RES., vol. 57, 1969, pages 1193 - 1197 |
CASALS J.: "The arthropod-borne group of animal viruses", TRANS. N. Y. ACAD. SCI., vol. 19, 1957, pages 219 - 235 |
CHAIN MMT; DOANE RW; MCLEAN DM: "Morphological development of Chikungunya virus", CAN. J. MICROBIOL., vol. 12, 1966, pages 895 - 899 |
CHAKRAVARTHY SK; SARKAR JK: "Susceptibility of new born and adult laboratory animals to Chikungunya virus", IND. J. MED. RES., vol. 57, 1969, pages 1157 - 1164 |
CHATURVEDI UC; MEHROTRA NK; MATHUR A; KAPOOR AK; MEHROTRA RM: "Chikungunya virus HI antibodies in the population of Lucknow and Kanpur", IND. JOUR. MED. RES., vol. 58, 1970, pages 297 - 301 |
ECKELS KH; HARRISON VR; HETRICK FM: "Chikungunya virus vaccine prepared by Tween-ether extraction", APPLIED MICROBIOL., vol. 19, 1970, pages 321 - 325, XP002487088 |
EDELMAN R; TACKET CO; WASSERMAN SS; BODISON SA; PERRY JG; MAGNIAFICO JA: "Phase II safety and immunogenicity study of live chikungunya virus vaccine TSI-GSD-218", AM. J. TROP. MED. HYG., vol. 62, 2000, pages 681 - 685, XP002487087 |
GIOVARELLI M; VIANO I; ZUCCA M; VALBONESI R; DIANZANI F: "Effect of anti- D-chain-specific immunosuppression on Chikungunya virus encephalitis of mice", INFECT. IMMUN, vol. 16, 1977, pages 849 - 852, XP002409145 |
HAHON N; HANKINS WA: "Assay for Chikungunya virus in cell monolayers by immunofluorescence", APPLIED MICROBIOL., vol. 19, 1970, pages 224 - 231 |
HAHON N; ZIMMERMAN WD: "Chikungunya virus infection of cell monolayers by cell-to-cell and extracellular transmission", APPLIED MICROBIOL., vol. 19, 1970, pages 389 - 391 |
HANNOUN: "Arbovirus haemgglutinins: differential susceptibility to trypsin", NATURE, vol. 219, 1968, pages 753 - 755 |
HARRISON VR; BINN LN; RANDALL R: "Comparative immunogenicities of chikungunya vaccines prepared in avian and mammalian tissues", AMER. J. TROP. MED. HYG., vol. 16, 1967, pages 786 - 791, XP009102713 |
HARRISON VR; ECKELS KH; BARTELLONI PJ; HAMPTON C: "Production and evaluation of a formalin-killed chikungunya vaccine", J. IMMUNOL., vol. 107, 1971, pages 643 - 647, XP002487089 |
HEARN HJ; RAINEY CT: "Cross-protection in aminals infected with group A arboviruses", J. IMMUNOL., vol. 90, 1963, pages 720 - 724 |
HEISE MT; SIMPSON DA; JOHNSTON JE: "Sindbis-group alphavirus replication in periosteum and endosteum of long bones in adult mice", J. VIROL., vol. 74, 2000, pages 9294 - 9299 |
HIGASHI N; MATSUMOTO A; TABATA K; NAGATOMO Y: "Electron microscope study of development of Chikungunya virus in green monkey kidney stable (Vero) cells", VIROLOGY, vol. 33, 1967, pages 55 - 69, XP023053123, DOI: doi:10.1016/0042-6822(67)90093-1 |
KHAN AH; MORITA K; PARQUET MC; HASEBE F; MATHENGE EGM; IGARASHI A: "Complete nucleotide sequence of chikungunya virus and evidence for an internal polyadenylation site", J. GEN. VIROL., vol. 83, 2002, pages 3075 - 3084, XP002458289 |
KILLINGTON RA; STOKES A; HIERHOLZER JC: "Virus purification. In ''Virology methods manual", 1996, ACADEMIC PRESS, pages: 71 - 89 |
KLIEN F; MAHLANDT BG; COCKEY RR; LINCOLN RE: "Concentration of Rift valley fever and Chikungunya viruses by precipitation", APPLIED MICROBIOL., vol. 20, 1970, pages 346 - 350 |
LANCIOTTI RS; LUDWIG ML; RWAGUMA EB; LUTWAMA JJ; KRAM TM ET AL.: "Emergence of epidemic O'nyong-nyong fever in Uganda after a 35-year absence: genetic characterization of the virus", VIROLOGY, vol. 252, 1998, pages 252 - 268 |
LEVITT NH; RAMSBURG HH; HASTY SE; REPIK PM; COLE FE JR; LUPTON HW: "Development of an attenuated strain of chikungunya virus for use in vaccine production", VACCINE, vol. 4, 1986, pages 157 - 162, XP023710291, DOI: doi:10.1016/0264-410X(86)90003-4 |
MCCLAIN DJ; PITTMAN PR; RAMSBURG HH; NELSON GO; ROSSI CA; MANGIAFICO JA; SCHMALJOHN AL; MALINOSKI FJ.: "Immunologic interference from sequential administration of live attenuated alphavirus vaccines", J. INFECT. DIS., vol. 177, 1998, pages 634 - 641, XP009102703 |
MCLNTOSH BM; PATERSON HE; MCGILLIVRAY G; DESOUSA J.: "Further studies on the Chikungunya outbreak in Southern Rhodesia in 1962. I. Mosquitoes, wild primates and birds in relation to the epidemic", ANN. TROP. MED. PARASITOL., vol. 58, pages 45 - 51 |
MYERS RM; CAREY DE; REUBEN R; JESUDASS ES; DE RANITZ C; JADHAV M: "The 1964 epidemic of Dengue-like fever in South India: isolation of Chikungunya virus from human sera and from mosquitoes", IND. J. MED. RES., vol. 53, 1965, pages 694 - 701 |
NIMMANNITYA S; HALSTEAD SB; COHEN SN; MARGIOTTA MR: "Dengue and · chikungunya virus infection in man in Thailand, 1962-1964. I. Observations on hospitalized patients with hemorrhagic fever", AM. J. TROP. MED. HYG., vol. 18, 1969, pages 954 - 971 |
PARKS JJ; PRICE WJ.: "Studies on immunologic overlap among certain arthropod-borne viruses", AM. J. TROP. MED. HYG., vol. 67, 1958, pages 187 - 206 |
PAUL SD; SINGH KRP: "Experimental infection of Macaca radiata with Chikungunya virus and transmission of virus by mosquitoes", IND. J. MED. RES., vol. 56, 1968, pages 802 - 810 |
PORTERFIELD JS.: "Cross-neutralization studies with group A arthropod-borne viruses", BULL WHO, vol. 24, 1961, pages 735 - 741 |
POWERS AM; BRAULT AC; TESH RB; WEAVER SC: "Re-emergence of chikungunya and o'nyong-nyong viruses: evidence for distinct geographical lineages and distant evolutionary relationships", J. GEN. VIROL., vol. 81, 2000, pages 471 - 479, XP002458290 |
RANADIVE SN; BANERJEE K: "Cloning and expression of Chikungunya virus genes coding structural proteins in Escherichia coli", IND. J. MED. RES., vol. 91, 1990, pages 386 - 392, XP008101388 |
RAO TR; CAREY DE; PAVRI KM: "Preliminary isolation and identification of Chikungunya virus from cases of Dengue-like illness in Madras city", IND. J. MED. RES., vol. 53, 1965, pages 689 - 693 |
RAVI V.: "Re-emergence of chikungunya virus in India", IND. J. MED. MICROBIOL., vol. 24, 2006, pages 83 - 84 |
SARKAR JK; CHATTERJEE JK; CHAKRAVARTI SK; MITRA AC: "Chikungunya virus infection with haemorrhagic manifestations", IND. JOUR. MED. RES., vol. 53, 1965, pages 921 - 925 |
SARKAR JK; CHATTERJEE SN; CHAKRAVARTY SK: "Haemorrhagic fever in Calcutta: some epidemiological observations", IRID: J. MED. RES., vol. 52, 1964, pages 651 - 659 |
SCHUFFENECKER I; ITEMAN I; MICHAULT A; MURRI S; FRANGEUL L; VANEY M-C; LAVENIR R; PARDIGON N; REYNES, J-M; PETTINELLI F: "Genome microevolution of Chikungunya viruses causing the Indian Ocean outbreak", PLOS MED., vol. 3, 2006, pages E263 |
SHAW KV; CLARENCE JG, JR.; BANERJEE G.: "Virological investigation of the epidemic of haemorrhagic fever in Calcutta: isolation of three strains of Chikungunya virus", IND. J. MED. RES., vol. 52, 1964, pages 676 - 682 |
SIMIZU B; YAMAMOTO K; HASHIMOTO K; OGATA T: "Structural proteins of Chikungunya virus", J. VIROL., vol. 51, 1984, pages 254 - 258 |
UMRIGAR MD; KADAM SS: "Comparative sensitivity of suckling mice and Vero cells for primary isolation of Chikungunya virus", IND. J. MED. RES., vol. 62, 1974, pages 1893 - 1895 |
WEISS HJ; HALSTEAD SB; RUSS SB: "Hemorrhagic disease in rodents caused by Chikungunya virus. 1. Studies of hemostasis", PROC. SOC. EXP. BIOL. MED., vol. 119, 1965, pages 427 - 432 |
WHO TECHNICAL REPORT SERIES, NO. 878, 1998, pages 19 - 52 |
Cited By (74)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
EP2460822A3 (en) * | 2006-03-15 | 2012-09-26 | Institut Pasteur | Novel isolated and purified strains of chikungunya virus and polynucleotides and polypeptides sequences, diagnostic and immunogenical uses thereof |
WO2009031045A3 (en) * | 2007-09-07 | 2009-04-23 | Pasteur Institut | Anti-chikungunya monoclonal antibodies and uses thereof |
WO2009031045A2 (en) * | 2007-09-07 | 2009-03-12 | Institut Pasteur | Anti-chikungunya monoclonal antibodies and uses thereof |
AU2008294415B2 (en) * | 2007-09-07 | 2014-09-04 | Institut Pasteur | Anti-chikungunya monoclonal antibodies and uses thereof |
EP2274442A4 (en) * | 2008-04-04 | 2011-10-05 | Univ Pennsylvania | Consensus sequences of chikungunya viral proteins, nucleic acid molecules encoding the same, and compositions and methods for using the same |
US8852609B2 (en) | 2008-04-04 | 2014-10-07 | The Trustees Of The University Of Pennsylvania | Consensus sequences of chikungunya viral proteins, nucleic acid molecules encoding the same, and compositions and methods for using the same |
CN101990580A (en) * | 2008-04-04 | 2011-03-23 | 宾夕法尼亚州立大学托管会 | Consensus sequences of chikungunya viral proteins, nucleic acid molecules encoding the same, and compositions and methods for using the same |
EP2274442A2 (en) * | 2008-04-04 | 2011-01-19 | The Trustees Of The University Of Pennsylvania | Consensus sequences of chikungunya viral proteins, nucleic acid molecules encoding the same, and compositions and methods for using the same |
JP2011517959A (en) * | 2008-04-21 | 2011-06-23 | ジェン−プロウブ インコーポレイテッド | Method for detecting chikungunya virus |
EP3957754A1 (en) * | 2008-04-21 | 2022-02-23 | Gen-Probe Incorporated | Method for detecting chikungunya virus |
US10344341B2 (en) | 2008-04-21 | 2019-07-09 | Gen-Probe Incorporated | Method for detecting chikungunya virus |
US9273365B2 (en) | 2008-04-21 | 2016-03-01 | Gen-Probe Incorporated | Method for detecting Chikungunya virus |
EP3392351A1 (en) * | 2008-04-21 | 2018-10-24 | Gen-Probe Incorporated | Method for detecting chikungunya virus |
EP2811037A1 (en) * | 2008-04-21 | 2014-12-10 | Gen-Probe Incorporated | Method for detecting Chikungunya virus |
WO2009131683A3 (en) * | 2008-04-21 | 2010-04-29 | Gen-Probe Incorporated | Method for detecting chikungunya virus |
EP3613761A1 (en) * | 2008-11-26 | 2020-02-26 | Government of the United States of America, as represented by the Secretary, Department of Health and Human Services | Virus like particle compositions and methods of use |
AU2009320287B2 (en) * | 2008-11-26 | 2015-10-08 | Government Of The United States Of America, As Represented By The Secretary, Department Of Health And Human Services | Virus like particle compositions and methods of use |
US9353353B2 (en) | 2008-11-26 | 2016-05-31 | The United States Of America, As Represented By The Secretary, Department Of Health And Human Services | Virus-like particles (VLPs) prepared from chikungunya virus structural proteins |
US11992523B2 (en) | 2008-11-26 | 2024-05-28 | The United States Of America, As Represented By The Secretary, Department Of Health And Human Services | Method for producing chikungunya virus (CHIKV) virus-like particles comprising the C, E2, and E1 structural proteins |
US11369674B2 (en) | 2008-11-26 | 2022-06-28 | The United States Of America, As Represented By The Secretary, Department Of Health And Human Services | Treatment method utilizing chikungunya virus (CHIKV) virus-like particles (VLPS) comprising the C, E2 and E1 structural proteins |
US10369208B2 (en) | 2008-11-26 | 2019-08-06 | The United States Of America, As Represented By The Secretary, Department Of Health And Human Services | Methods for the induction of immune responses in a subject compromising administering virus-like particles (VLPS) prepared from Chikungunya virus structural proteins |
WO2010062396A2 (en) | 2008-11-26 | 2010-06-03 | Government Of The United States Of America , As Represented By The Secretary, Department Of Health And Human Services | Virus like particle compositions and methods of use |
EP2370455A4 (en) * | 2008-11-26 | 2012-07-04 | Us Gov Health & Human Serv | Virus like particle compositions and methods of use |
EP2370455A2 (en) * | 2008-11-26 | 2011-10-05 | Government Of The United States Of America, As Represented By The Secretary, Department of Health Human Services | Virus like particle compositions and methods of use |
US20120003266A1 (en) * | 2008-11-26 | 2012-01-05 | The United States of America,as represented by The Secretary, National Institues of Health | Virus like particle compositions and methods of use |
EP2374816A1 (en) * | 2010-04-07 | 2011-10-12 | Humalys | Binding molecules against Chikungunya virus and uses thereof |
US9738704B2 (en) | 2010-04-07 | 2017-08-22 | Agency For Science, Technology And Research | Binding molecules against Chikungunya virus and uses thereof |
WO2011124635A1 (en) * | 2010-04-07 | 2011-10-13 | Humalys | Binding molecules against chikungunya virus and uses thereof |
US9441032B2 (en) | 2010-04-07 | 2016-09-13 | Agency For Science, Technology And Research | Binding molecules against Chikungunya virus and uses thereof |
WO2011130119A3 (en) * | 2010-04-14 | 2012-03-29 | Emd Millipore Corporation | Methods of producing high titer, high purity virus stocks and methods of use thereof |
EP2377927B1 (en) | 2010-04-14 | 2019-05-22 | EMD Millipore Corporation | Methods of producing high titer, high purity mouse minute virus (MMV) stocks and methods of use thereof |
US20120088228A1 (en) * | 2010-04-14 | 2012-04-12 | Millipore Corporation | Methods of producing high titer, high purity virus stocks and methods of use thereof |
US9644187B2 (en) | 2010-04-14 | 2017-05-09 | Emd Millipore Corporation | Methods of producing high titer, high purity virus stocks and methods of use thereof |
WO2011130119A2 (en) * | 2010-04-14 | 2011-10-20 | Millipore Corporation | Methods of producing high titer, high purity virus stocks and methods of use thereof |
KR101549296B1 (en) * | 2010-04-14 | 2015-09-01 | 이엠디 밀리포어 코포레이션 | Methods of producing high titer, high purity virus stocks and methods of use thereof |
EP2377927A1 (en) * | 2010-04-14 | 2011-10-19 | Millipore Corporation | Methods of producing high titer, high purity virus stocks and methods of use thereof |
WO2012073257A3 (en) * | 2010-11-30 | 2012-10-11 | Bharat Biotech International Limited | Vaccine formulation for prophylaxis and treatment of chandipura virus infections in mammals |
KR101792684B1 (en) | 2011-06-17 | 2017-11-02 | 브하라트 바이오테크 인터내셔날 리미티드 | Vaccine composition comprising an inactivated chikungunya virus strain |
JP2014520117A (en) * | 2011-06-17 | 2014-08-21 | バハラ バイオテック インターナショナル リミテッド | Vaccine composition comprising inactivated chikungunya virus strain |
US9844588B2 (en) | 2011-06-17 | 2017-12-19 | Bharat Biotech International Limited | Inactivated chikungunya viruses (CHIKV) comprising an E1-K211E mutation |
AU2012269907B2 (en) * | 2011-06-17 | 2017-05-18 | Bharat Biotech International Limited | Vaccine composition comprising an inactivated chikungunya virus strain |
WO2012172574A1 (en) | 2011-06-17 | 2012-12-20 | Bharat Biotech International Limited | Vaccine composition comprising an inactivated chikungunya virus strain |
US10137186B2 (en) | 2013-03-14 | 2018-11-27 | Takeda Vaccines, Inc. | Compositions and methods for live, attenuated alphavirus formulations |
AU2014236804B2 (en) * | 2013-03-14 | 2018-12-13 | Takeda Vaccines, Inc. | Compositions and methods for live, attenuated alphavirus formulations |
CN105377293A (en) * | 2013-03-14 | 2016-03-02 | 武田疫苗股份有限公司 | Compositions and methods for live, attenuated alphavirus formulations |
CN105377293B (en) * | 2013-03-14 | 2020-07-17 | 武田疫苗股份有限公司 | Compositions and methods for live attenuated alphavirus formulations |
WO2014151855A1 (en) * | 2013-03-14 | 2014-09-25 | Inviragen, Inc. | Compositions and methods for live, attenuated alphavirus formulations |
AU2018236897B2 (en) * | 2013-03-14 | 2020-09-10 | Takeda Vaccines, Inc. | Compositions and methods for live, attenuated alphavirus formulations |
CN111729077A (en) * | 2013-03-14 | 2020-10-02 | 武田疫苗股份有限公司 | Compositions and methods for live attenuated alphavirus formulations |
EP3603667A1 (en) * | 2013-03-14 | 2020-02-05 | Takeda Vaccines, Inc. | Compositions and methods for live, attenuated alphavirus formulations |
US10806781B2 (en) | 2013-03-14 | 2020-10-20 | Takeda Vaccines, Inc. | Compositions and methods for live, attenuated alphavirus formulations |
EP3322441A4 (en) * | 2015-07-16 | 2018-12-19 | Bharat Biotech International Limited | Vaccine compositions |
CN108601825B (en) * | 2015-07-16 | 2023-06-20 | 巴拉特生物技术国际有限公司 | Vaccine composition |
CN108601825A (en) * | 2015-07-16 | 2018-09-28 | 巴拉特生物技术国际有限公司 | Vaccine composition |
EA035921B1 (en) * | 2015-07-16 | 2020-08-31 | Бхарат Байотек Интернэшнл Лимитед | Vaccine compositions for prophylaxis of arbovirus infections |
JP2018527317A (en) * | 2015-07-16 | 2018-09-20 | バハラ バイオテック インターナショナル リミテッド | Vaccine composition |
WO2017009873A1 (en) * | 2015-07-16 | 2017-01-19 | Bharat Biotech International Limited | Vaccine compositions |
EP3393506B1 (en) * | 2015-12-23 | 2024-03-13 | Valneva SE | Virus purification |
WO2017172643A1 (en) * | 2016-03-31 | 2017-10-05 | Takeda Vaccines, Inc. | Compositions and methods for stabilizing alphaviruses with improved formulations |
US10632184B2 (en) | 2016-03-31 | 2020-04-28 | Takeda Vaccines, Inc. | Compositions and methods for stabilizing alphaviruses with improved formulations |
JP2019515046A (en) * | 2016-05-10 | 2019-06-06 | ナジット テクノロジーズ, インコーポレイテッド | Inorganic polyatomic oxyanion to protect against antigen damage during pathogen inactivation for vaccine production |
US11141475B2 (en) | 2016-05-10 | 2021-10-12 | Najit Technologies, Inc. | Inactivating pathogens and producing highly immunogenic inactivated vaccines using a dual oxidation process |
US11633470B2 (en) | 2016-05-10 | 2023-04-25 | Najit Technologies, Inc. | Inactivating pathogens and producing highly immunogenic inactivated vaccines using a dual oxidation process |
WO2017197034A1 (en) * | 2016-05-10 | 2017-11-16 | Najit Technologies, Inc. | Inorganic polyatomic oxyanions for protecting against antigenic damage during pathogen inactivation for vaccine production |
US10744198B2 (en) | 2016-05-10 | 2020-08-18 | Najit Technologies, Inc. | Inorganic polyatomic oxyanions for protecting against antigenic damage during pathogen inactivation for vaccine production |
EP3454895B1 (en) * | 2016-05-10 | 2024-03-27 | Najit Technologies, Inc. | Inorganic polyatomic oxyanions for protecting against antigenic damage during pathogen inactivation for vaccine production |
US11844832B2 (en) | 2016-05-10 | 2023-12-19 | Najit Technologies, Inc. | Inorganic polyatomic oxyanions for protecting against antigenic damage during pathogen inactivation for vaccine production |
US12011477B2 (en) | 2017-09-21 | 2024-06-18 | Valneva Se | Method of producing pharmaceutical compositions comprising immunogenic chikungunya virus CHIKV-delta5NSP3 |
US11648304B2 (en) | 2017-11-03 | 2023-05-16 | Takeda Vaccines, Inc. | Zika vaccines and immunogenic compositions, and methods of using the same |
US11730802B2 (en) | 2017-11-03 | 2023-08-22 | Takeda Vaccines, Inc. | Zika vaccines and immunogenic compositions, and methods of using the same |
US11964008B2 (en) | 2017-11-03 | 2024-04-23 | Takeda Vaccines, Inc. | Method for inactivating zika virus and for determining the completeness of inactivation |
US11478541B2 (en) | 2017-11-03 | 2022-10-25 | Takeda Vaccines, Inc. | Method for inactivating Zika virus and for determining the completeness of inactivation |
US11975062B2 (en) | 2017-11-30 | 2024-05-07 | Takeda Vaccines, Inc. | Zika vaccines and immunogenic compositions, and methods of using the same |
US10718771B2 (en) | 2018-02-09 | 2020-07-21 | Chung Yuan Christian University | Recombinant baculoviruses and their uses in detecting arthropod-borne virus |
Also Published As
Publication number | Publication date |
---|---|
BRPI0716145B1 (en) | 2021-06-22 |
BRPI0716145A2 (en) | 2013-09-17 |
EP2073839B1 (en) | 2016-10-19 |
EP2073839A2 (en) | 2009-07-01 |
US20130022631A1 (en) | 2013-01-24 |
US8865184B2 (en) | 2014-10-21 |
WO2008026225A3 (en) | 2008-09-25 |
CN101516395A (en) | 2009-08-26 |
CN101516395B (en) | 2014-03-26 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
EP2073839B1 (en) | A vaccine for chikungunya virus infection | |
CN108601825B (en) | Vaccine composition | |
Schmaljohn et al. | Antigenic subunits of Hantaan virus expressed by baculovirus and vaccinia virus recombinants | |
Guo et al. | Foot-and-mouth disease virus-like particles produced by a SUMO fusion protein system in Escherichia coli induce potent protective immune responses in guinea pigs, swine and cattle | |
Plana-Duran et al. | Oral immunization of rabbits with VP60 particles confers protection against rabbit hemorrhagic disease | |
AU2010249103B2 (en) | New human rotavirus strains and vaccines | |
EP0433371A1 (en) | Fusion protein of paramyxovirus, method of production using recombinant baculovirus expression vector, vaccine comprising such protein and use thereof | |
Lubroth et al. | Absence of protein 2C from clarified foot-and-mouth disease virus vaccines provides the basis for distinguishing convalescent from vaccinated animals | |
Pekosz et al. | Protection from La Crosse virus encephalitis with recombinant glycoproteins: role of neutralizing anti-G1 antibodies | |
CA2592366C (en) | Recombinant foot and mouth disease vaccine | |
KR20230082653A (en) | Fusion proteins useful for vaccination against rotavirus | |
WO2022080413A1 (en) | Beta coronavirus cold acclimatized strain and vaccine | |
Crisci et al. | Chimeric calicivirus-like particles elicit protective anti-viral cytotoxic responses without adjuvant | |
Aaskov et al. | A candidate Ross River virus vaccine: preclinical evaluation | |
WO2012073257A2 (en) | Vaccine formulation for prophylaxis and treatment of chandipura virus infections in mammals | |
Viaplana et al. | Antigenicity of VP60 structural proteinof rabbit haemorrhagic disease virus | |
PT1131414E (en) | Stable, attenuated rabies virus mutants and live vaccines thereof | |
EP4155393A1 (en) | Attenuated variant of the rift valley fever virus, composition comprising same, and uses thereof | |
BRPI0720319A2 (en) | "NUCLEIC ACID ISOLATED, ISOLATED POLYPEPTIDE, EXPRESSION VECTOR, HOST CELL, METHOD FOR THE PRODUCTION OF A FUSION POLYPTIDE, BAMBOO MOSAIC VIRUS, MUSIC VEQUIC MUSIC VEHICLE And method for the induction of an immune response in an individual " | |
KR20240099257A (en) | Beta coronavirus medicinal liquor | |
Masurel et al. | Vaccination and protection experiments in mice with the human A-1957 and A-1968 strains, and the Equi-2 strain of influenza virus | |
Murdin | Studies of Aphthovirus Subunits and Synthetic Peptides Relevant to Their Use as Vaccines Against Foot and Mouth Disease | |
Found | Equine herpes virus-4 thymidine-kinase-vaccine; expression in mammal host transformant; DNA sequence; potential mono-valent, multivalent live recombinant vaccine vector | |
Nisalakl | Serological Responses to Japanese Encephalitis in Thai Swine | |
MXPA99005452A (en) | Recombinant equine herpesvirus type 1 (ehv-1) comprising a dysfunctional gene 71 region and use thereof as a vaccine |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
WWE | Wipo information: entry into national phase |
Ref document number: 200780035576.X Country of ref document: CN |
|
121 | Ep: the epo has been informed by wipo that ep was designated in this application |
Ref document number: 07827554 Country of ref document: EP Kind code of ref document: A2 |
|
REEP | Request for entry into the european phase |
Ref document number: 2007827554 Country of ref document: EP |
|
WWE | Wipo information: entry into national phase |
Ref document number: 2007827554 Country of ref document: EP |
|
WWE | Wipo information: entry into national phase |
Ref document number: 12439509 Country of ref document: US |
|
NENP | Non-entry into the national phase |
Ref country code: DE |
|
ENP | Entry into the national phase |
Ref document number: PI0716145 Country of ref document: BR Kind code of ref document: A2 Effective date: 20090302 |