CN102085377B - Immunogenic composition and methods - Google Patents
Immunogenic composition and methods Download PDFInfo
- Publication number
- CN102085377B CN102085377B CN201010587086.4A CN201010587086A CN102085377B CN 102085377 B CN102085377 B CN 102085377B CN 201010587086 A CN201010587086 A CN 201010587086A CN 102085377 B CN102085377 B CN 102085377B
- Authority
- CN
- China
- Prior art keywords
- antigen
- compositions
- vsv
- sequence
- plasmid
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired - Lifetime
Links
- 239000000203 mixture Substances 0.000 title claims abstract description 229
- 238000000034 method Methods 0.000 title claims abstract description 63
- 230000002163 immunogen Effects 0.000 title claims description 76
- 239000013612 plasmid Substances 0.000 claims abstract description 185
- 241000711975 Vesicular stomatitis virus Species 0.000 claims abstract description 141
- 239000000427 antigen Substances 0.000 claims abstract description 141
- 102000036639 antigens Human genes 0.000 claims abstract description 141
- 108091007433 antigens Proteins 0.000 claims abstract description 141
- 108020004414 DNA Proteins 0.000 claims abstract description 136
- 230000001105 regulatory effect Effects 0.000 claims abstract description 38
- 230000014509 gene expression Effects 0.000 claims abstract description 25
- 108091028043 Nucleic acid sequence Proteins 0.000 claims abstract description 24
- 230000001939 inductive effect Effects 0.000 claims abstract description 14
- 150000007523 nucleic acids Chemical group 0.000 claims abstract description 11
- 230000010076 replication Effects 0.000 claims abstract description 8
- 210000004027 cell Anatomy 0.000 claims description 67
- 241000700605 Viruses Species 0.000 claims description 62
- 102000004127 Cytokines Human genes 0.000 claims description 34
- 108090000695 Cytokines Proteins 0.000 claims description 34
- 241000713311 Simian immunodeficiency virus Species 0.000 claims description 32
- 230000001276 controlling effect Effects 0.000 claims description 32
- 230000004044 response Effects 0.000 claims description 29
- 108090000765 processed proteins & peptides Proteins 0.000 claims description 27
- 238000012360 testing method Methods 0.000 claims description 21
- 239000002773 nucleotide Substances 0.000 claims description 18
- 125000003729 nucleotide group Chemical group 0.000 claims description 18
- 102000013462 Interleukin-12 Human genes 0.000 claims description 17
- 108010065805 Interleukin-12 Proteins 0.000 claims description 17
- 230000000844 anti-bacterial effect Effects 0.000 claims description 13
- 239000012634 fragment Substances 0.000 claims description 13
- 238000002360 preparation method Methods 0.000 claims description 12
- 241000282414 Homo sapiens Species 0.000 claims description 10
- 230000004927 fusion Effects 0.000 claims description 10
- 102000039446 nucleic acids Human genes 0.000 claims description 9
- 108020004707 nucleic acids Proteins 0.000 claims description 9
- 239000000546 pharmaceutical excipient Substances 0.000 claims description 8
- 229920001184 polypeptide Polymers 0.000 claims description 8
- 102000004196 processed proteins & peptides Human genes 0.000 claims description 8
- 241000233866 Fungi Species 0.000 claims description 7
- 239000003085 diluting agent Substances 0.000 claims description 7
- 239000003814 drug Substances 0.000 claims description 6
- 244000045947 parasite Species 0.000 claims description 6
- 210000004881 tumor cell Anatomy 0.000 claims description 6
- LEBVLXFERQHONN-UHFFFAOYSA-N 1-butyl-N-(2,6-dimethylphenyl)piperidine-2-carboxamide Chemical compound CCCCN1CCCCC1C(=O)NC1=C(C)C=CC=C1C LEBVLXFERQHONN-UHFFFAOYSA-N 0.000 claims description 5
- 229960003150 bupivacaine Drugs 0.000 claims description 5
- 230000005875 antibody response Effects 0.000 claims description 4
- 108010017213 Granulocyte-Macrophage Colony-Stimulating Factor Proteins 0.000 claims description 2
- 101710149951 Protein Tat Proteins 0.000 claims description 2
- 102000004457 Granulocyte-Macrophage Colony-Stimulating Factor Human genes 0.000 claims 1
- 230000005867 T cell response Effects 0.000 claims 1
- 238000003149 assay kit Methods 0.000 claims 1
- 230000003053 immunization Effects 0.000 abstract description 25
- 238000002649 immunization Methods 0.000 abstract description 25
- 241000251539 Vertebrata <Metazoa> Species 0.000 abstract description 22
- 201000010099 disease Diseases 0.000 abstract description 19
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 abstract description 19
- 230000001225 therapeutic effect Effects 0.000 abstract description 5
- 238000011282 treatment Methods 0.000 abstract description 5
- 230000028993 immune response Effects 0.000 abstract description 4
- 108090000623 proteins and genes Proteins 0.000 description 70
- 230000036039 immunity Effects 0.000 description 66
- 238000011081 inoculation Methods 0.000 description 57
- 241000725303 Human immunodeficiency virus Species 0.000 description 33
- 239000002671 adjuvant Substances 0.000 description 28
- 150000001413 amino acids Chemical class 0.000 description 26
- 229940024606 amino acid Drugs 0.000 description 24
- 235000001014 amino acid Nutrition 0.000 description 24
- 238000002560 therapeutic procedure Methods 0.000 description 23
- 241001465754 Metazoa Species 0.000 description 22
- 206010022000 influenza Diseases 0.000 description 17
- 238000005728 strengthening Methods 0.000 description 17
- 108010074328 Interferon-gamma Proteins 0.000 description 15
- 239000002299 complementary DNA Substances 0.000 description 14
- 230000002787 reinforcement Effects 0.000 description 14
- 230000003612 virological effect Effects 0.000 description 14
- 108091032973 (ribonucleotides)n+m Proteins 0.000 description 13
- 101150082239 G gene Proteins 0.000 description 13
- 241000713772 Human immunodeficiency virus 1 Species 0.000 description 13
- 102100037850 Interferon gamma Human genes 0.000 description 13
- 241000124008 Mammalia Species 0.000 description 13
- -1 cation lipid Chemical class 0.000 description 11
- 238000006243 chemical reaction Methods 0.000 description 11
- 239000003589 local anesthetic agent Substances 0.000 description 11
- 102000004169 proteins and genes Human genes 0.000 description 11
- 210000002966 serum Anatomy 0.000 description 11
- 230000033228 biological regulation Effects 0.000 description 10
- 235000018102 proteins Nutrition 0.000 description 10
- 241000220317 Rosa Species 0.000 description 9
- 210000001744 T-lymphocyte Anatomy 0.000 description 8
- 230000000694 effects Effects 0.000 description 8
- 238000005516 engineering process Methods 0.000 description 8
- 239000003623 enhancer Substances 0.000 description 8
- 108700004026 gag Genes Proteins 0.000 description 8
- 101150098622 gag gene Proteins 0.000 description 8
- 230000008488 polyadenylation Effects 0.000 description 8
- 239000013598 vector Substances 0.000 description 8
- 241000894006 Bacteria Species 0.000 description 7
- 239000000654 additive Substances 0.000 description 7
- 230000000996 additive effect Effects 0.000 description 7
- 239000002585 base Substances 0.000 description 7
- 230000008859 change Effects 0.000 description 7
- 210000000805 cytoplasm Anatomy 0.000 description 7
- 108010030074 endodeoxyribonuclease MluI Proteins 0.000 description 7
- 108700004025 env Genes Proteins 0.000 description 7
- 230000005847 immunogenicity Effects 0.000 description 7
- 230000006698 induction Effects 0.000 description 7
- 239000003351 stiffener Substances 0.000 description 7
- 238000002965 ELISA Methods 0.000 description 6
- 230000000890 antigenic effect Effects 0.000 description 6
- 101150030339 env gene Proteins 0.000 description 6
- 238000011534 incubation Methods 0.000 description 6
- 230000004048 modification Effects 0.000 description 6
- 238000012986 modification Methods 0.000 description 6
- 210000005105 peripheral blood lymphocyte Anatomy 0.000 description 6
- 108091033319 polynucleotide Proteins 0.000 description 6
- 102000040430 polynucleotide Human genes 0.000 description 6
- 239000002157 polynucleotide Substances 0.000 description 6
- 230000008569 process Effects 0.000 description 6
- 239000000047 product Substances 0.000 description 6
- 230000001681 protective effect Effects 0.000 description 6
- 108020003519 protein disulfide isomerase Proteins 0.000 description 6
- 238000001890 transfection Methods 0.000 description 6
- 208000030507 AIDS Diseases 0.000 description 5
- 241000588724 Escherichia coli Species 0.000 description 5
- 102100030698 Interleukin-12 subunit alpha Human genes 0.000 description 5
- 206010028980 Neoplasm Diseases 0.000 description 5
- 102100037837 Nucleoporin Nup37 Human genes 0.000 description 5
- 229920001213 Polysorbate 20 Polymers 0.000 description 5
- 241000607142 Salmonella Species 0.000 description 5
- 241000701033 Simian cytomegalovirus Species 0.000 description 5
- 102100022203 Tumor necrosis factor receptor superfamily member 25 Human genes 0.000 description 5
- 230000015572 biosynthetic process Effects 0.000 description 5
- 239000000839 emulsion Substances 0.000 description 5
- XUWPJKDMEZSVTP-LTYMHZPRSA-N kalafungina Chemical compound O=C1C2=C(O)C=CC=C2C(=O)C2=C1[C@@H](C)O[C@H]1[C@@H]2OC(=O)C1 XUWPJKDMEZSVTP-LTYMHZPRSA-N 0.000 description 5
- 239000007788 liquid Substances 0.000 description 5
- 238000005457 optimization Methods 0.000 description 5
- 239000000256 polyoxyethylene sorbitan monolaurate Substances 0.000 description 5
- 235000010486 polyoxyethylene sorbitan monolaurate Nutrition 0.000 description 5
- 230000008521 reorganization Effects 0.000 description 5
- 108091008146 restriction endonucleases Proteins 0.000 description 5
- 239000004094 surface-active agent Substances 0.000 description 5
- 238000013518 transcription Methods 0.000 description 5
- 230000035897 transcription Effects 0.000 description 5
- 229960005486 vaccine Drugs 0.000 description 5
- 108091026890 Coding region Proteins 0.000 description 4
- 102000004190 Enzymes Human genes 0.000 description 4
- 108090000790 Enzymes Proteins 0.000 description 4
- 102100026878 Interleukin-2 receptor subunit alpha Human genes 0.000 description 4
- 101150062031 L gene Proteins 0.000 description 4
- 241000187479 Mycobacterium tuberculosis Species 0.000 description 4
- 238000012408 PCR amplification Methods 0.000 description 4
- 101710137500 T7 RNA polymerase Proteins 0.000 description 4
- 241000700618 Vaccinia virus Species 0.000 description 4
- 238000004458 analytical method Methods 0.000 description 4
- 230000002421 anti-septic effect Effects 0.000 description 4
- 238000013459 approach Methods 0.000 description 4
- PYMYPHUHKUWMLA-WDCZJNDASA-N arabinose Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)C=O PYMYPHUHKUWMLA-WDCZJNDASA-N 0.000 description 4
- PYMYPHUHKUWMLA-UHFFFAOYSA-N arabinose Natural products OCC(O)C(O)C(O)C=O PYMYPHUHKUWMLA-UHFFFAOYSA-N 0.000 description 4
- 230000001580 bacterial effect Effects 0.000 description 4
- SRBFZHDQGSBBOR-UHFFFAOYSA-N beta-D-Pyranose-Lyxose Natural products OC1COC(O)C(O)C1O SRBFZHDQGSBBOR-UHFFFAOYSA-N 0.000 description 4
- 210000001151 cytotoxic T lymphocyte Anatomy 0.000 description 4
- 238000010586 diagram Methods 0.000 description 4
- 230000002068 genetic effect Effects 0.000 description 4
- 239000001963 growth medium Substances 0.000 description 4
- 230000001900 immune effect Effects 0.000 description 4
- 208000015181 infectious disease Diseases 0.000 description 4
- 230000005764 inhibitory process Effects 0.000 description 4
- 238000010255 intramuscular injection Methods 0.000 description 4
- 239000007927 intramuscular injection Substances 0.000 description 4
- 230000003472 neutralizing effect Effects 0.000 description 4
- 239000013600 plasmid vector Substances 0.000 description 4
- 239000000843 powder Substances 0.000 description 4
- 238000000746 purification Methods 0.000 description 4
- 238000011084 recovery Methods 0.000 description 4
- 238000010839 reverse transcription Methods 0.000 description 4
- 238000005406 washing Methods 0.000 description 4
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 4
- DIGQNXIGRZPYDK-WKSCXVIASA-N (2R)-6-amino-2-[[2-[[(2S)-2-[[2-[[(2R)-2-[[(2S)-2-[[(2R,3S)-2-[[2-[[(2S)-2-[[2-[[(2S)-2-[[(2S)-2-[[(2R)-2-[[(2S,3S)-2-[[(2R)-2-[[(2S)-2-[[(2S)-2-[[(2S)-2-[[2-[[(2S)-2-[[(2R)-2-[[2-[[2-[[2-[(2-amino-1-hydroxyethylidene)amino]-3-carboxy-1-hydroxypropylidene]amino]-1-hydroxy-3-sulfanylpropylidene]amino]-1-hydroxyethylidene]amino]-1-hydroxy-3-sulfanylpropylidene]amino]-1,3-dihydroxypropylidene]amino]-1-hydroxyethylidene]amino]-1-hydroxypropylidene]amino]-1,3-dihydroxypropylidene]amino]-1,3-dihydroxypropylidene]amino]-1-hydroxy-3-sulfanylpropylidene]amino]-1,3-dihydroxybutylidene]amino]-1-hydroxy-3-sulfanylpropylidene]amino]-1-hydroxypropylidene]amino]-1,3-dihydroxypropylidene]amino]-1-hydroxyethylidene]amino]-1,5-dihydroxy-5-iminopentylidene]amino]-1-hydroxy-3-sulfanylpropylidene]amino]-1,3-dihydroxybutylidene]amino]-1-hydroxy-3-sulfanylpropylidene]amino]-1,3-dihydroxypropylidene]amino]-1-hydroxyethylidene]amino]-1-hydroxy-3-sulfanylpropylidene]amino]-1-hydroxyethylidene]amino]hexanoic acid Chemical compound C[C@@H]([C@@H](C(=N[C@@H](CS)C(=N[C@@H](C)C(=N[C@@H](CO)C(=NCC(=N[C@@H](CCC(=N)O)C(=NC(CS)C(=N[C@H]([C@H](C)O)C(=N[C@H](CS)C(=N[C@H](CO)C(=NCC(=N[C@H](CS)C(=NCC(=N[C@H](CCCCN)C(=O)O)O)O)O)O)O)O)O)O)O)O)O)O)O)N=C([C@H](CS)N=C([C@H](CO)N=C([C@H](CO)N=C([C@H](C)N=C(CN=C([C@H](CO)N=C([C@H](CS)N=C(CN=C(C(CS)N=C(C(CC(=O)O)N=C(CN)O)O)O)O)O)O)O)O)O)O)O)O DIGQNXIGRZPYDK-WKSCXVIASA-N 0.000 description 3
- 108020004705 Codon Proteins 0.000 description 3
- 206010011831 Cytomegalovirus infection Diseases 0.000 description 3
- 102000004163 DNA-directed RNA polymerases Human genes 0.000 description 3
- 108090000626 DNA-directed RNA polymerases Proteins 0.000 description 3
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 3
- 101710154606 Hemagglutinin Proteins 0.000 description 3
- 101000679903 Homo sapiens Tumor necrosis factor receptor superfamily member 25 Proteins 0.000 description 3
- 102000003792 Metallothionein Human genes 0.000 description 3
- 108090000157 Metallothionein Proteins 0.000 description 3
- 101710093908 Outer capsid protein VP4 Proteins 0.000 description 3
- 101710135467 Outer capsid protein sigma-1 Proteins 0.000 description 3
- 101710176177 Protein A56 Proteins 0.000 description 3
- 241000714474 Rous sarcoma virus Species 0.000 description 3
- 241000580858 Simian-Human immunodeficiency virus Species 0.000 description 3
- 102100040112 Tumor necrosis factor receptor superfamily member 10B Human genes 0.000 description 3
- 230000003321 amplification Effects 0.000 description 3
- 230000036436 anti-hiv Effects 0.000 description 3
- 239000000969 carrier Substances 0.000 description 3
- 230000001413 cellular effect Effects 0.000 description 3
- 150000001875 compounds Chemical class 0.000 description 3
- 230000008034 disappearance Effects 0.000 description 3
- 238000002474 experimental method Methods 0.000 description 3
- 239000013613 expression plasmid Substances 0.000 description 3
- 239000008187 granular material Substances 0.000 description 3
- 238000002347 injection Methods 0.000 description 3
- 239000007924 injection Substances 0.000 description 3
- 239000002502 liposome Substances 0.000 description 3
- 210000004962 mammalian cell Anatomy 0.000 description 3
- 230000001404 mediated effect Effects 0.000 description 3
- 238000012544 monitoring process Methods 0.000 description 3
- 238000003199 nucleic acid amplification method Methods 0.000 description 3
- 229910052760 oxygen Inorganic materials 0.000 description 3
- 239000001301 oxygen Substances 0.000 description 3
- 238000012856 packing Methods 0.000 description 3
- 229920000642 polymer Polymers 0.000 description 3
- 229920005862 polyol Polymers 0.000 description 3
- 150000003077 polyols Chemical class 0.000 description 3
- 102000005962 receptors Human genes 0.000 description 3
- 108020003175 receptors Proteins 0.000 description 3
- 210000000664 rectum Anatomy 0.000 description 3
- 230000003362 replicative effect Effects 0.000 description 3
- 238000012163 sequencing technique Methods 0.000 description 3
- 239000000126 substance Substances 0.000 description 3
- 210000001519 tissue Anatomy 0.000 description 3
- 230000005026 transcription initiation Effects 0.000 description 3
- 241000606748 Actinobacillus pleuropneumoniae Species 0.000 description 2
- 102000007469 Actins Human genes 0.000 description 2
- 108010085238 Actins Proteins 0.000 description 2
- 102000002659 Amyloid Precursor Protein Secretases Human genes 0.000 description 2
- 108010043324 Amyloid Precursor Protein Secretases Proteins 0.000 description 2
- 241000193830 Bacillus <bacterium> Species 0.000 description 2
- 241000335423 Blastomyces Species 0.000 description 2
- 241000588807 Bordetella Species 0.000 description 2
- 241000588832 Bordetella pertussis Species 0.000 description 2
- 241000589969 Borreliella burgdorferi Species 0.000 description 2
- 241000283690 Bos taurus Species 0.000 description 2
- 241000222120 Candida <Saccharomycetales> Species 0.000 description 2
- 241001647372 Chlamydia pneumoniae Species 0.000 description 2
- 241001647378 Chlamydia psittaci Species 0.000 description 2
- 241000606153 Chlamydia trachomatis Species 0.000 description 2
- 241000193449 Clostridium tetani Species 0.000 description 2
- UDMBCSSLTHHNCD-UHFFFAOYSA-N Coenzym Q(11) Natural products C1=NC=2C(N)=NC=NC=2N1C1OC(COP(O)(O)=O)C(O)C1O UDMBCSSLTHHNCD-UHFFFAOYSA-N 0.000 description 2
- 241000759568 Corixa Species 0.000 description 2
- 241001337994 Cryptococcus <scale insect> Species 0.000 description 2
- 241000223935 Cryptosporidium Species 0.000 description 2
- 241000701022 Cytomegalovirus Species 0.000 description 2
- 241000194032 Enterococcus faecalis Species 0.000 description 2
- 101710177291 Gag polyprotein Proteins 0.000 description 2
- 108700028146 Genetic Enhancer Elements Proteins 0.000 description 2
- 241000224466 Giardia Species 0.000 description 2
- DHMQDGOQFOQNFH-UHFFFAOYSA-N Glycine Chemical compound NCC(O)=O DHMQDGOQFOQNFH-UHFFFAOYSA-N 0.000 description 2
- 102000003886 Glycoproteins Human genes 0.000 description 2
- 108090000288 Glycoproteins Proteins 0.000 description 2
- 241000590002 Helicobacter pylori Species 0.000 description 2
- 241000700721 Hepatitis B virus Species 0.000 description 2
- 241000606831 Histophilus somni Species 0.000 description 2
- 241000228402 Histoplasma Species 0.000 description 2
- 101000610604 Homo sapiens Tumor necrosis factor receptor superfamily member 10B Proteins 0.000 description 2
- 206010061598 Immunodeficiency Diseases 0.000 description 2
- 108060003951 Immunoglobulin Proteins 0.000 description 2
- 102100029604 Interferon alpha-inducible protein 27, mitochondrial Human genes 0.000 description 2
- 102000008070 Interferon-gamma Human genes 0.000 description 2
- 241000222697 Leishmania infantum Species 0.000 description 2
- 241000222732 Leishmania major Species 0.000 description 2
- 241000589929 Leptospira interrogans Species 0.000 description 2
- 241000186781 Listeria Species 0.000 description 2
- 101710125418 Major capsid protein Proteins 0.000 description 2
- 241000712079 Measles morbillivirus Species 0.000 description 2
- 101710151805 Mitochondrial intermediate peptidase 1 Proteins 0.000 description 2
- 241000588655 Moraxella catarrhalis Species 0.000 description 2
- 241000186367 Mycobacterium avium Species 0.000 description 2
- 241000204022 Mycoplasma gallisepticum Species 0.000 description 2
- 108091034117 Oligonucleotide Proteins 0.000 description 2
- 229930182555 Penicillin Natural products 0.000 description 2
- JGSARLDLIJGVTE-MBNYWOFBSA-N Penicillin G Chemical compound N([C@H]1[C@H]2SC([C@@H](N2C1=O)C(O)=O)(C)C)C(=O)CC1=CC=CC=C1 JGSARLDLIJGVTE-MBNYWOFBSA-N 0.000 description 2
- ISWSIDIOOBJBQZ-UHFFFAOYSA-N Phenol Chemical compound OC1=CC=CC=C1 ISWSIDIOOBJBQZ-UHFFFAOYSA-N 0.000 description 2
- 241000223960 Plasmodium falciparum Species 0.000 description 2
- 241000233872 Pneumocystis carinii Species 0.000 description 2
- 239000004721 Polyphenylene oxide Substances 0.000 description 2
- ZTHYODDOHIVTJV-UHFFFAOYSA-N Propyl gallate Chemical compound CCCOC(=O)C1=CC(O)=C(O)C(O)=C1 ZTHYODDOHIVTJV-UHFFFAOYSA-N 0.000 description 2
- 241000588770 Proteus mirabilis Species 0.000 description 2
- 241000588767 Proteus vulgaris Species 0.000 description 2
- 241000293871 Salmonella enterica subsp. enterica serovar Typhi Species 0.000 description 2
- 241000242678 Schistosoma Species 0.000 description 2
- 241000607768 Shigella Species 0.000 description 2
- 241000191967 Staphylococcus aureus Species 0.000 description 2
- 241000191963 Staphylococcus epidermidis Species 0.000 description 2
- 241000193985 Streptococcus agalactiae Species 0.000 description 2
- 241000193998 Streptococcus pneumoniae Species 0.000 description 2
- 241000193996 Streptococcus pyogenes Species 0.000 description 2
- RAHZWNYVWXNFOC-UHFFFAOYSA-N Sulphur dioxide Chemical compound O=S=O RAHZWNYVWXNFOC-UHFFFAOYSA-N 0.000 description 2
- 241000223997 Toxoplasma gondii Species 0.000 description 2
- 102000004357 Transferases Human genes 0.000 description 2
- 108090000992 Transferases Proteins 0.000 description 2
- 241000224526 Trichomonas Species 0.000 description 2
- 241000607626 Vibrio cholerae Species 0.000 description 2
- 108020000999 Viral RNA Proteins 0.000 description 2
- 208000036142 Viral infection Diseases 0.000 description 2
- 230000005856 abnormality Effects 0.000 description 2
- 230000009471 action Effects 0.000 description 2
- UDMBCSSLTHHNCD-KQYNXXCUSA-N adenosine 5'-monophosphate Chemical compound C1=NC=2C(N)=NC=NC=2N1[C@@H]1O[C@H](COP(O)(O)=O)[C@@H](O)[C@H]1O UDMBCSSLTHHNCD-KQYNXXCUSA-N 0.000 description 2
- 229950006790 adenosine phosphate Drugs 0.000 description 2
- 239000000443 aerosol Substances 0.000 description 2
- ILRRQNADMUWWFW-UHFFFAOYSA-K aluminium phosphate Chemical compound O1[Al]2OP1(=O)O2 ILRRQNADMUWWFW-UHFFFAOYSA-K 0.000 description 2
- VSCWAEJMTAWNJL-UHFFFAOYSA-K aluminium trichloride Chemical compound Cl[Al](Cl)Cl VSCWAEJMTAWNJL-UHFFFAOYSA-K 0.000 description 2
- DZHSAHHDTRWUTF-SIQRNXPUSA-N amyloid-beta polypeptide 42 Chemical compound C([C@@H](C(=O)N[C@@H](C)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@H](C(=O)NCC(=O)N[C@@H](CO)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CCCCN)C(=O)NCC(=O)N[C@@H](C)C(=O)N[C@H](C(=O)N[C@@H]([C@@H](C)CC)C(=O)NCC(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCSC)C(=O)N[C@@H](C(C)C)C(=O)NCC(=O)NCC(=O)N[C@@H](C(C)C)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](C)C(O)=O)[C@@H](C)CC)C(C)C)NC(=O)[C@H](CC=1C=CC=CC=1)NC(=O)[C@@H](NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CCCCN)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@H](CC=1N=CNC=1)NC(=O)[C@H](CC=1N=CNC=1)NC(=O)[C@@H](NC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](CC=1C=CC(O)=CC=1)NC(=O)CNC(=O)[C@H](CO)NC(=O)[C@H](CC(O)=O)NC(=O)[C@H](CC=1N=CNC=1)NC(=O)[C@H](CCCNC(N)=N)NC(=O)[C@H](CC=1C=CC=CC=1)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](C)NC(=O)[C@@H](N)CC(O)=O)C(C)C)C(C)C)C1=CC=CC=C1 DZHSAHHDTRWUTF-SIQRNXPUSA-N 0.000 description 2
- 239000003242 anti bacterial agent Substances 0.000 description 2
- 230000000843 anti-fungal effect Effects 0.000 description 2
- 229940121375 antifungal agent Drugs 0.000 description 2
- WGQKYBSKWIADBV-UHFFFAOYSA-O benzylaminium Chemical compound [NH3+]CC1=CC=CC=C1 WGQKYBSKWIADBV-UHFFFAOYSA-O 0.000 description 2
- 230000037396 body weight Effects 0.000 description 2
- 210000000988 bone and bone Anatomy 0.000 description 2
- 244000309466 calf Species 0.000 description 2
- 201000011510 cancer Diseases 0.000 description 2
- 239000002775 capsule Substances 0.000 description 2
- 230000006727 cell loss Effects 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 229940038705 chlamydia trachomatis Drugs 0.000 description 2
- OSASVXMJTNOKOY-UHFFFAOYSA-N chlorobutanol Chemical compound CC(C)(O)C(Cl)(Cl)Cl OSASVXMJTNOKOY-UHFFFAOYSA-N 0.000 description 2
- 230000000295 complement effect Effects 0.000 description 2
- 238000010276 construction Methods 0.000 description 2
- 230000001086 cytosolic effect Effects 0.000 description 2
- 230000001419 dependent effect Effects 0.000 description 2
- 238000013461 design Methods 0.000 description 2
- 238000001514 detection method Methods 0.000 description 2
- UREBDLICKHMUKA-CXSFZGCWSA-N dexamethasone Chemical compound C1CC2=CC(=O)C=C[C@]2(C)[C@]2(F)[C@@H]1[C@@H]1C[C@@H](C)[C@@](C(=O)CO)(O)[C@@]1(C)C[C@@H]2O UREBDLICKHMUKA-CXSFZGCWSA-N 0.000 description 2
- 229940079593 drug Drugs 0.000 description 2
- 239000003937 drug carrier Substances 0.000 description 2
- CBOQJANXLMLOSS-UHFFFAOYSA-N ethyl vanillin Chemical compound CCOC1=CC(C=O)=CC=C1O CBOQJANXLMLOSS-UHFFFAOYSA-N 0.000 description 2
- 239000013604 expression vector Substances 0.000 description 2
- 229940037467 helicobacter pylori Drugs 0.000 description 2
- 230000008348 humoral response Effects 0.000 description 2
- 210000000987 immune system Anatomy 0.000 description 2
- 102000018358 immunoglobulin Human genes 0.000 description 2
- 230000003308 immunostimulating effect Effects 0.000 description 2
- 230000001976 improved effect Effects 0.000 description 2
- 239000004615 ingredient Substances 0.000 description 2
- 239000002917 insecticide Substances 0.000 description 2
- 238000003780 insertion Methods 0.000 description 2
- 230000037431 insertion Effects 0.000 description 2
- NOESYZHRGYRDHS-UHFFFAOYSA-N insulin Chemical compound N1C(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(NC(=O)CN)C(C)CC)CSSCC(C(NC(CO)C(=O)NC(CC(C)C)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CCC(N)=O)C(=O)NC(CC(C)C)C(=O)NC(CCC(O)=O)C(=O)NC(CC(N)=O)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CSSCC(NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2C=CC(O)=CC=2)NC(=O)C(CC(C)C)NC(=O)C(C)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2NC=NC=2)NC(=O)C(CO)NC(=O)CNC2=O)C(=O)NCC(=O)NC(CCC(O)=O)C(=O)NC(CCCNC(N)=N)C(=O)NCC(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC(O)=CC=3)C(=O)NC(C(C)O)C(=O)N3C(CCC3)C(=O)NC(CCCCN)C(=O)NC(C)C(O)=O)C(=O)NC(CC(N)=O)C(O)=O)=O)NC(=O)C(C(C)CC)NC(=O)C(CO)NC(=O)C(C(C)O)NC(=O)C1CSSCC2NC(=O)C(CC(C)C)NC(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CC(N)=O)NC(=O)C(NC(=O)C(N)CC=1C=CC=CC=1)C(C)C)CC1=CN=CN1 NOESYZHRGYRDHS-UHFFFAOYSA-N 0.000 description 2
- 229940117681 interleukin-12 Drugs 0.000 description 2
- 210000000265 leukocyte Anatomy 0.000 description 2
- 238000004519 manufacturing process Methods 0.000 description 2
- 239000003550 marker Substances 0.000 description 2
- 239000002609 medium Substances 0.000 description 2
- 239000012528 membrane Substances 0.000 description 2
- 125000001419 myristoyl group Chemical group O=C([*])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 description 2
- 229910052757 nitrogen Inorganic materials 0.000 description 2
- 230000007170 pathology Effects 0.000 description 2
- 229940049954 penicillin Drugs 0.000 description 2
- 229910052698 phosphorus Inorganic materials 0.000 description 2
- 210000002381 plasma Anatomy 0.000 description 2
- 229920000570 polyether Polymers 0.000 description 2
- 230000002265 prevention Effects 0.000 description 2
- 229940007042 proteus vulgaris Drugs 0.000 description 2
- 230000008707 rearrangement Effects 0.000 description 2
- 230000002829 reductive effect Effects 0.000 description 2
- 230000003252 repetitive effect Effects 0.000 description 2
- 230000002441 reversible effect Effects 0.000 description 2
- 210000003296 saliva Anatomy 0.000 description 2
- 230000028327 secretion Effects 0.000 description 2
- 238000007493 shaping process Methods 0.000 description 2
- DAEPDZWVDSPTHF-UHFFFAOYSA-M sodium pyruvate Chemical compound [Na+].CC(=O)C([O-])=O DAEPDZWVDSPTHF-UHFFFAOYSA-M 0.000 description 2
- 239000003381 stabilizer Substances 0.000 description 2
- 229940031000 streptococcus pneumoniae Drugs 0.000 description 2
- 239000000725 suspension Substances 0.000 description 2
- 238000013268 sustained release Methods 0.000 description 2
- 239000012730 sustained-release form Substances 0.000 description 2
- 208000024891 symptom Diseases 0.000 description 2
- 239000003053 toxin Substances 0.000 description 2
- 231100000765 toxin Toxicity 0.000 description 2
- 230000001131 transforming effect Effects 0.000 description 2
- 238000013519 translation Methods 0.000 description 2
- 241000701161 unidentified adenovirus Species 0.000 description 2
- 241001430294 unidentified retrovirus Species 0.000 description 2
- 210000001215 vagina Anatomy 0.000 description 2
- 210000003501 vero cell Anatomy 0.000 description 2
- 230000029812 viral genome replication Effects 0.000 description 2
- 230000009385 viral infection Effects 0.000 description 2
- 210000000605 viral structure Anatomy 0.000 description 2
- 230000000007 visual effect Effects 0.000 description 2
- DRHZYJAUECRAJM-DWSYSWFDSA-N (2s,3s,4s,5r,6r)-6-[[(3s,4s,4ar,6ar,6bs,8r,8ar,12as,14ar,14br)-8a-[(2s,3r,4s,5r,6r)-3-[(2s,3r,4s,5r,6s)-5-[(2s,3r,4s,5r)-4-[(2s,3r,4r)-3,4-dihydroxy-4-(hydroxymethyl)oxolan-2-yl]oxy-3,5-dihydroxyoxan-2-yl]oxy-3,4-dihydroxy-6-methyloxan-2-yl]oxy-5-[(3s,5s, Chemical compound O([C@H]1[C@H](O)[C@H](O[C@H]([C@@H]1O[C@H]1[C@@H]([C@@H](O)[C@@H](O)[C@@H](CO)O1)O)O[C@H]1CC[C@]2(C)[C@H]3CC=C4[C@@H]5CC(C)(C)CC[C@@]5([C@@H](C[C@@]4(C)[C@]3(C)CC[C@H]2[C@@]1(C=O)C)O)C(=O)O[C@@H]1O[C@H](C)[C@@H]([C@@H]([C@H]1O[C@H]1[C@@H]([C@H](O)[C@@H](O[C@H]2[C@@H]([C@@H](O[C@H]3[C@@H]([C@@](O)(CO)CO3)O)[C@H](O)CO2)O)[C@H](C)O1)O)O)OC(=O)C[C@@H](O)C[C@H](OC(=O)C[C@@H](O)C[C@@H]([C@@H](C)CC)O[C@H]1[C@@H]([C@@H](O)[C@H](CO)O1)O)[C@@H](C)CC)C(O)=O)[C@@H]1OC[C@@H](O)[C@H](O)[C@H]1O DRHZYJAUECRAJM-DWSYSWFDSA-N 0.000 description 1
- ASWBNKHCZGQVJV-UHFFFAOYSA-N (3-hexadecanoyloxy-2-hydroxypropyl) 2-(trimethylazaniumyl)ethyl phosphate Chemical compound CCCCCCCCCCCCCCCC(=O)OCC(O)COP([O-])(=O)OCC[N+](C)(C)C ASWBNKHCZGQVJV-UHFFFAOYSA-N 0.000 description 1
- YYGNTYWPHWGJRM-UHFFFAOYSA-N (6E,10E,14E,18E)-2,6,10,15,19,23-hexamethyltetracosa-2,6,10,14,18,22-hexaene Chemical compound CC(C)=CCCC(C)=CCCC(C)=CCCC=C(C)CCC=C(C)CCC=C(C)C YYGNTYWPHWGJRM-UHFFFAOYSA-N 0.000 description 1
- ALSTYHKOOCGGFT-KTKRTIGZSA-N (9Z)-octadecen-1-ol Chemical class CCCCCCCC\C=C/CCCCCCCCO ALSTYHKOOCGGFT-KTKRTIGZSA-N 0.000 description 1
- 102000040650 (ribonucleotides)n+m Human genes 0.000 description 1
- NWUYHJFMYQTDRP-UHFFFAOYSA-N 1,2-bis(ethenyl)benzene;1-ethenyl-2-ethylbenzene;styrene Chemical compound C=CC1=CC=CC=C1.CCC1=CC=CC=C1C=C.C=CC1=CC=CC=C1C=C NWUYHJFMYQTDRP-UHFFFAOYSA-N 0.000 description 1
- FJLUATLTXUNBOT-UHFFFAOYSA-N 1-Hexadecylamine Chemical compound CCCCCCCCCCCCCCCCN FJLUATLTXUNBOT-UHFFFAOYSA-N 0.000 description 1
- CHHHXKFHOYLYRE-UHFFFAOYSA-M 2,4-Hexadienoic acid, potassium salt (1:1), (2E,4E)- Chemical compound [K+].CC=CC=CC([O-])=O CHHHXKFHOYLYRE-UHFFFAOYSA-M 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
- OYJPMMSSLQTURE-UHFFFAOYSA-N 2-methylheptatriacontane Chemical compound CCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCCC(C)C OYJPMMSSLQTURE-UHFFFAOYSA-N 0.000 description 1
- WXNZTHHGJRFXKQ-UHFFFAOYSA-N 4-chlorophenol Chemical compound OC1=CC=C(Cl)C=C1 WXNZTHHGJRFXKQ-UHFFFAOYSA-N 0.000 description 1
- QRXMUCSWCMTJGU-UHFFFAOYSA-N 5-bromo-4-chloro-3-indolyl phosphate Chemical compound C1=C(Br)C(Cl)=C2C(OP(O)(=O)O)=CNC2=C1 QRXMUCSWCMTJGU-UHFFFAOYSA-N 0.000 description 1
- 108010042708 Acetylmuramyl-Alanyl-Isoglutamine Proteins 0.000 description 1
- 208000003200 Adenoma Diseases 0.000 description 1
- 206010001233 Adenoma benign Diseases 0.000 description 1
- 102000009027 Albumins Human genes 0.000 description 1
- 108010088751 Albumins Proteins 0.000 description 1
- 241000186033 Alloiococcus Species 0.000 description 1
- 208000031295 Animal disease Diseases 0.000 description 1
- 239000004475 Arginine Substances 0.000 description 1
- 241000244186 Ascaris Species 0.000 description 1
- 102100022717 Atypical chemokine receptor 1 Human genes 0.000 description 1
- 241000711404 Avian avulavirus 1 Species 0.000 description 1
- 241001519465 Avian metapneumovirus Species 0.000 description 1
- 102100031746 Bone sialoprotein 2 Human genes 0.000 description 1
- 102100021943 C-C motif chemokine 2 Human genes 0.000 description 1
- 101710155857 C-C motif chemokine 2 Proteins 0.000 description 1
- 102100032367 C-C motif chemokine 5 Human genes 0.000 description 1
- 108010029697 CD40 Ligand Proteins 0.000 description 1
- 101150013553 CD40 gene Proteins 0.000 description 1
- 102100032937 CD40 ligand Human genes 0.000 description 1
- 108010084313 CD58 Antigens Proteins 0.000 description 1
- 101100289995 Caenorhabditis elegans mac-1 gene Proteins 0.000 description 1
- 241000712083 Canine morbillivirus Species 0.000 description 1
- 108010022366 Carcinoembryonic Antigen Proteins 0.000 description 1
- 102000012406 Carcinoembryonic Antigen Human genes 0.000 description 1
- 102000011727 Caspases Human genes 0.000 description 1
- 108010076667 Caspases Proteins 0.000 description 1
- 102000053642 Catalytic RNA Human genes 0.000 description 1
- 108090000994 Catalytic RNA Proteins 0.000 description 1
- 206010007882 Cellulitis Diseases 0.000 description 1
- 241000282693 Cercopithecidae Species 0.000 description 1
- 241000711969 Chandipura virus Species 0.000 description 1
- 108010055166 Chemokine CCL5 Proteins 0.000 description 1
- 241001432959 Chernes Species 0.000 description 1
- 206010008631 Cholera Diseases 0.000 description 1
- 108010049048 Cholera Toxin Proteins 0.000 description 1
- 102000009016 Cholera Toxin Human genes 0.000 description 1
- 241000223203 Coccidioides Species 0.000 description 1
- 241000711573 Coronaviridae Species 0.000 description 1
- 241000186216 Corynebacterium Species 0.000 description 1
- 241000186227 Corynebacterium diphtheriae Species 0.000 description 1
- 241000699800 Cricetinae Species 0.000 description 1
- KDXKERNSBIXSRK-RXMQYKEDSA-N D-lysine Chemical compound NCCCC[C@@H](N)C(O)=O KDXKERNSBIXSRK-RXMQYKEDSA-N 0.000 description 1
- 102000016928 DNA-directed DNA polymerase Human genes 0.000 description 1
- 108010014303 DNA-directed DNA polymerase Proteins 0.000 description 1
- 101100450350 Danio rerio helz gene Proteins 0.000 description 1
- 102100037840 Dehydrogenase/reductase SDR family member 2, mitochondrial Human genes 0.000 description 1
- 101710088341 Dermatopontin Proteins 0.000 description 1
- 108010016626 Dipeptides Proteins 0.000 description 1
- 206010059866 Drug resistance Diseases 0.000 description 1
- 239000006144 Dulbecco’s modified Eagle's medium Substances 0.000 description 1
- 108010069091 Dystrophin Proteins 0.000 description 1
- 102000001039 Dystrophin Human genes 0.000 description 1
- 101150029707 ERBB2 gene Proteins 0.000 description 1
- 102100027723 Endogenous retrovirus group K member 6 Rec protein Human genes 0.000 description 1
- 241000991587 Enterovirus C Species 0.000 description 1
- 101710091045 Envelope protein Proteins 0.000 description 1
- 241000283073 Equus caballus Species 0.000 description 1
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 1
- 241001136487 Eurotium Species 0.000 description 1
- 241000711475 Feline infectious peritonitis virus Species 0.000 description 1
- 241000282324 Felis Species 0.000 description 1
- 102000004641 Fetal Proteins Human genes 0.000 description 1
- 108010003471 Fetal Proteins Proteins 0.000 description 1
- 102000018233 Fibroblast Growth Factor Human genes 0.000 description 1
- 108050007372 Fibroblast Growth Factor Proteins 0.000 description 1
- 108010022355 Fibroins Proteins 0.000 description 1
- 108010010803 Gelatin Proteins 0.000 description 1
- 230000010558 Gene Alterations Effects 0.000 description 1
- 108700007698 Genetic Terminator Regions Proteins 0.000 description 1
- JZNWSCPGTDBMEW-UHFFFAOYSA-N Glycerophosphorylethanolamin Natural products NCCOP(O)(=O)OCC(O)CO JZNWSCPGTDBMEW-UHFFFAOYSA-N 0.000 description 1
- 239000004471 Glycine Substances 0.000 description 1
- 206010018612 Gonorrhoea Diseases 0.000 description 1
- 108010017080 Granulocyte Colony-Stimulating Factor Proteins 0.000 description 1
- 102000004269 Granulocyte Colony-Stimulating Factor Human genes 0.000 description 1
- 102100039620 Granulocyte-macrophage colony-stimulating factor Human genes 0.000 description 1
- 239000007995 HEPES buffer Substances 0.000 description 1
- 241000606768 Haemophilus influenzae Species 0.000 description 1
- 102100031573 Hematopoietic progenitor cell antigen CD34 Human genes 0.000 description 1
- 208000032843 Hemorrhage Diseases 0.000 description 1
- 241000893570 Hendra henipavirus Species 0.000 description 1
- 241000711549 Hepacivirus C Species 0.000 description 1
- 241000709721 Hepatovirus A Species 0.000 description 1
- 101000678879 Homo sapiens Atypical chemokine receptor 1 Proteins 0.000 description 1
- 101000911390 Homo sapiens Coagulation factor VIII Proteins 0.000 description 1
- 101000777663 Homo sapiens Hematopoietic progenitor cell antigen CD34 Proteins 0.000 description 1
- 101001018097 Homo sapiens L-selectin Proteins 0.000 description 1
- 101000623901 Homo sapiens Mucin-16 Proteins 0.000 description 1
- 101000979333 Homo sapiens Neurofilament light polypeptide Proteins 0.000 description 1
- 101000622137 Homo sapiens P-selectin Proteins 0.000 description 1
- 101000612089 Homo sapiens Pancreas/duodenum homeobox protein 1 Proteins 0.000 description 1
- 101000934346 Homo sapiens T-cell surface antigen CD2 Proteins 0.000 description 1
- 101100100117 Homo sapiens TNFRSF10B gene Proteins 0.000 description 1
- 101000679921 Homo sapiens Tumor necrosis factor receptor superfamily member 21 Proteins 0.000 description 1
- 241000701074 Human alphaherpesvirus 2 Species 0.000 description 1
- 241000342334 Human metapneumovirus Species 0.000 description 1
- 241000701806 Human papillomavirus Species 0.000 description 1
- 208000029462 Immunodeficiency disease Diseases 0.000 description 1
- 102000004877 Insulin Human genes 0.000 description 1
- 108090001061 Insulin Proteins 0.000 description 1
- 108090000723 Insulin-Like Growth Factor I Proteins 0.000 description 1
- 102100025323 Integrin alpha-1 Human genes 0.000 description 1
- 102100022339 Integrin alpha-L Human genes 0.000 description 1
- 108010041341 Integrin alpha1 Proteins 0.000 description 1
- 108010055795 Integrin alpha1beta1 Proteins 0.000 description 1
- 108010028750 Integrin-Binding Sialoprotein Proteins 0.000 description 1
- 108010064593 Intercellular Adhesion Molecule-1 Proteins 0.000 description 1
- 108010064600 Intercellular Adhesion Molecule-3 Proteins 0.000 description 1
- 102100037877 Intercellular adhesion molecule 1 Human genes 0.000 description 1
- 102100037872 Intercellular adhesion molecule 2 Human genes 0.000 description 1
- 101710148794 Intercellular adhesion molecule 2 Proteins 0.000 description 1
- 102100037871 Intercellular adhesion molecule 3 Human genes 0.000 description 1
- 102000006992 Interferon-alpha Human genes 0.000 description 1
- 108010047761 Interferon-alpha Proteins 0.000 description 1
- 102000003996 Interferon-beta Human genes 0.000 description 1
- 108090000467 Interferon-beta Proteins 0.000 description 1
- 101710194995 Interleukin-12 subunit alpha Proteins 0.000 description 1
- 102100036701 Interleukin-12 subunit beta Human genes 0.000 description 1
- 101710187487 Interleukin-12 subunit beta Proteins 0.000 description 1
- CKLJMWTZIZZHCS-REOHCLBHSA-N L-aspartic acid Chemical compound OC(=O)[C@@H](N)CC(O)=O CKLJMWTZIZZHCS-REOHCLBHSA-N 0.000 description 1
- ZDXPYRJPNDTMRX-VKHMYHEASA-N L-glutamine Chemical compound OC(=O)[C@@H](N)CCC(N)=O ZDXPYRJPNDTMRX-VKHMYHEASA-N 0.000 description 1
- AGPKZVBTJJNPAG-WHFBIAKZSA-N L-isoleucine Chemical compound CC[C@H](C)[C@H](N)C(O)=O AGPKZVBTJJNPAG-WHFBIAKZSA-N 0.000 description 1
- 102100033467 L-selectin Human genes 0.000 description 1
- KZSNJWFQEVHDMF-BYPYZUCNSA-N L-valine Chemical compound CC(C)[C@H](N)C(O)=O KZSNJWFQEVHDMF-BYPYZUCNSA-N 0.000 description 1
- 102000004407 Lactalbumin Human genes 0.000 description 1
- 108090000942 Lactalbumin Proteins 0.000 description 1
- 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 description 1
- 108010064548 Lymphocyte Function-Associated Antigen-1 Proteins 0.000 description 1
- 108090000542 Lymphotoxin-alpha Proteins 0.000 description 1
- 102000004083 Lymphotoxin-alpha Human genes 0.000 description 1
- 108010059343 MM Form Creatine Kinase Proteins 0.000 description 1
- 241000282553 Macaca Species 0.000 description 1
- 241000701076 Macacine alphaherpesvirus 1 Species 0.000 description 1
- 102000007651 Macrophage Colony-Stimulating Factor Human genes 0.000 description 1
- 108010046938 Macrophage Colony-Stimulating Factor Proteins 0.000 description 1
- 241001293418 Mannheimia haemolytica Species 0.000 description 1
- 108010060408 Member 25 Tumor Necrosis Factor Receptors Proteins 0.000 description 1
- 201000009906 Meningitis Diseases 0.000 description 1
- 241000711513 Mononegavirales Species 0.000 description 1
- 241000713333 Mouse mammary tumor virus Species 0.000 description 1
- 102100034256 Mucin-1 Human genes 0.000 description 1
- 108010008707 Mucin-1 Proteins 0.000 description 1
- 102100023123 Mucin-16 Human genes 0.000 description 1
- 108010063954 Mucins Proteins 0.000 description 1
- 241000711386 Mumps virus Species 0.000 description 1
- 241000711408 Murine respirovirus Species 0.000 description 1
- 101100450352 Mus musculus Helz gene Proteins 0.000 description 1
- 241000186364 Mycobacterium intracellulare Species 0.000 description 1
- 102100026057 Myosin regulatory light chain 2, atrial isoform Human genes 0.000 description 1
- 101710098224 Myosin regulatory light chain 2, atrial isoform Proteins 0.000 description 1
- FFDGPVCHZBVARC-UHFFFAOYSA-N N,N-dimethylglycine Chemical compound CN(C)CC(O)=O FFDGPVCHZBVARC-UHFFFAOYSA-N 0.000 description 1
- 101150064037 NGF gene Proteins 0.000 description 1
- 241000588652 Neisseria gonorrhoeae Species 0.000 description 1
- 241000588650 Neisseria meningitidis Species 0.000 description 1
- 108010025020 Nerve Growth Factor Proteins 0.000 description 1
- 102000015336 Nerve Growth Factor Human genes 0.000 description 1
- 241000526636 Nipah henipavirus Species 0.000 description 1
- 108090001074 Nucleocapsid Proteins Proteins 0.000 description 1
- 241000283973 Oryctolagus cuniculus Species 0.000 description 1
- 102000004067 Osteocalcin Human genes 0.000 description 1
- 108090000573 Osteocalcin Proteins 0.000 description 1
- 101150084044 P gene Proteins 0.000 description 1
- 102100023472 P-selectin Human genes 0.000 description 1
- 101150044441 PECAM1 gene Proteins 0.000 description 1
- 229910019142 PO4 Inorganic materials 0.000 description 1
- 102100041030 Pancreas/duodenum homeobox protein 1 Human genes 0.000 description 1
- 208000002606 Paramyxoviridae Infections Diseases 0.000 description 1
- 241000606856 Pasteurella multocida Species 0.000 description 1
- 241001494479 Pecora Species 0.000 description 1
- 108010081690 Pertussis Toxin Proteins 0.000 description 1
- 102100029251 Phagocytosis-stimulating peptide Human genes 0.000 description 1
- BELBBZDIHDAJOR-UHFFFAOYSA-N Phenolsulfonephthalein Chemical compound C1=CC(O)=CC=C1C1(C=2C=CC(O)=CC=2)C2=CC=CC=C2S(=O)(=O)O1 BELBBZDIHDAJOR-UHFFFAOYSA-N 0.000 description 1
- 102000007982 Phosphoproteins Human genes 0.000 description 1
- 108010089430 Phosphoproteins Proteins 0.000 description 1
- 102000004160 Phosphoric Monoester Hydrolases Human genes 0.000 description 1
- 108090000608 Phosphoric Monoester Hydrolases Proteins 0.000 description 1
- 241000288906 Primates Species 0.000 description 1
- 102000007066 Prostate-Specific Antigen Human genes 0.000 description 1
- 108010072866 Prostate-Specific Antigen Proteins 0.000 description 1
- 101710188315 Protein X Proteins 0.000 description 1
- 241000125945 Protoparvovirus Species 0.000 description 1
- 102100028688 Putative glycosylation-dependent cell adhesion molecule 1 Human genes 0.000 description 1
- 239000012980 RPMI-1640 medium Substances 0.000 description 1
- 241000711798 Rabies lyssavirus Species 0.000 description 1
- 108020004511 Recombinant DNA Proteins 0.000 description 1
- 241000725643 Respiratory syncytial virus Species 0.000 description 1
- 206010039101 Rhinorrhoea Diseases 0.000 description 1
- 206010051497 Rhinotracheitis Diseases 0.000 description 1
- 241000711897 Rinderpest morbillivirus Species 0.000 description 1
- 241000702670 Rotavirus Species 0.000 description 1
- 241001138501 Salmonella enterica Species 0.000 description 1
- 102000040739 Secretory proteins Human genes 0.000 description 1
- 108091058545 Secretory proteins Proteins 0.000 description 1
- MTCFGRXMJLQNBG-UHFFFAOYSA-N Serine Natural products OCC(N)C(O)=O MTCFGRXMJLQNBG-UHFFFAOYSA-N 0.000 description 1
- 241000607764 Shigella dysenteriae Species 0.000 description 1
- 241000700584 Simplexvirus Species 0.000 description 1
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 1
- 229920002125 Sokalan® Polymers 0.000 description 1
- 102000013275 Somatomedins Human genes 0.000 description 1
- 108091081024 Start codon Proteins 0.000 description 1
- 108010090804 Streptavidin Proteins 0.000 description 1
- 229930006000 Sucrose Natural products 0.000 description 1
- 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 description 1
- 102100025237 T-cell surface antigen CD2 Human genes 0.000 description 1
- 239000004098 Tetracycline Substances 0.000 description 1
- BHEOSNUKNHRBNM-UHFFFAOYSA-N Tetramethylsqualene Natural products CC(=C)C(C)CCC(=C)C(C)CCC(C)=CCCC=C(C)CCC(C)C(=C)CCC(C)C(C)=C BHEOSNUKNHRBNM-UHFFFAOYSA-N 0.000 description 1
- 108010022394 Threonine synthase Proteins 0.000 description 1
- 102100023935 Transmembrane glycoprotein NMB Human genes 0.000 description 1
- 101800001690 Transmembrane protein gp41 Proteins 0.000 description 1
- 241000197881 Trichocephalus Species 0.000 description 1
- 241001489151 Trichuris Species 0.000 description 1
- 108010084754 Tuftsin Proteins 0.000 description 1
- 108060008682 Tumor Necrosis Factor Proteins 0.000 description 1
- 108060008683 Tumor Necrosis Factor Receptor Proteins 0.000 description 1
- 102000000852 Tumor Necrosis Factor-alpha Human genes 0.000 description 1
- 102100022205 Tumor necrosis factor receptor superfamily member 21 Human genes 0.000 description 1
- 102100040245 Tumor necrosis factor receptor superfamily member 5 Human genes 0.000 description 1
- 208000035896 Twin-reversed arterial perfusion sequence Diseases 0.000 description 1
- 206010046865 Vaccinia virus infection Diseases 0.000 description 1
- KZSNJWFQEVHDMF-UHFFFAOYSA-N Valine Natural products CC(C)C(N)C(O)=O KZSNJWFQEVHDMF-UHFFFAOYSA-N 0.000 description 1
- 102000005789 Vascular Endothelial Growth Factors Human genes 0.000 description 1
- 108010019530 Vascular Endothelial Growth Factors Proteins 0.000 description 1
- 108700005077 Viral Genes Proteins 0.000 description 1
- HCHKCACWOHOZIP-UHFFFAOYSA-N Zinc Chemical compound [Zn] HCHKCACWOHOZIP-UHFFFAOYSA-N 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 230000001154 acute effect Effects 0.000 description 1
- 239000011543 agarose gel Substances 0.000 description 1
- 239000003513 alkali Substances 0.000 description 1
- 230000002009 allergenic effect Effects 0.000 description 1
- UPEZCKBFRMILAV-UHFFFAOYSA-N alpha-Ecdysone Natural products C1C(O)C(O)CC2(C)C(CCC3(C(C(C(O)CCC(C)(C)O)C)CCC33O)C)C3=CC(=O)C21 UPEZCKBFRMILAV-UHFFFAOYSA-N 0.000 description 1
- 102000013529 alpha-Fetoproteins Human genes 0.000 description 1
- 108010026331 alpha-Fetoproteins Proteins 0.000 description 1
- RZJRJXONCZWCBN-UHFFFAOYSA-N alpha-octadecene Natural products CCCCCCCCCCCCCCCCCC RZJRJXONCZWCBN-UHFFFAOYSA-N 0.000 description 1
- AZDRQVAHHNSJOQ-UHFFFAOYSA-N alumane Chemical class [AlH3] AZDRQVAHHNSJOQ-UHFFFAOYSA-N 0.000 description 1
- 150000001412 amines Chemical class 0.000 description 1
- 125000004103 aminoalkyl group Chemical group 0.000 description 1
- 229940126575 aminoglycoside Drugs 0.000 description 1
- SWLVFNYSXGMGBS-UHFFFAOYSA-N ammonium bromide Chemical compound [NH4+].[Br-] SWLVFNYSXGMGBS-UHFFFAOYSA-N 0.000 description 1
- 239000002870 angiogenesis inducing agent Substances 0.000 description 1
- 230000008485 antagonism Effects 0.000 description 1
- 230000002141 anti-parasite Effects 0.000 description 1
- 229940088710 antibiotic agent Drugs 0.000 description 1
- 239000003096 antiparasitic agent Substances 0.000 description 1
- 239000007864 aqueous solution Substances 0.000 description 1
- 239000003125 aqueous solvent Substances 0.000 description 1
- 108700003859 araC Genes Proteins 0.000 description 1
- 101150044616 araC gene Proteins 0.000 description 1
- ODKSFYDXXFIFQN-UHFFFAOYSA-N arginine Natural products OC(=O)C(N)CCCNC(N)=N ODKSFYDXXFIFQN-UHFFFAOYSA-N 0.000 description 1
- 238000003491 array Methods 0.000 description 1
- 235000003704 aspartic acid Nutrition 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- 108010088716 attachment protein G Proteins 0.000 description 1
- 230000002238 attenuated effect Effects 0.000 description 1
- 230000001363 autoimmune Effects 0.000 description 1
- WXNRAKRZUCLRBP-UHFFFAOYSA-N avridine Chemical compound CCCCCCCCCCCCCCCCCCN(CCCN(CCO)CCO)CCCCCCCCCCCCCCCCCC WXNRAKRZUCLRBP-UHFFFAOYSA-N 0.000 description 1
- 229950010555 avridine Drugs 0.000 description 1
- 210000003719 b-lymphocyte Anatomy 0.000 description 1
- 229960000190 bacillus calmette–guérin vaccine Drugs 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- OQFSQFPPLPISGP-UHFFFAOYSA-N beta-carboxyaspartic acid Natural products OC(=O)C(N)C(C(O)=O)C(O)=O OQFSQFPPLPISGP-UHFFFAOYSA-N 0.000 description 1
- 229920002988 biodegradable polymer Polymers 0.000 description 1
- 239000004621 biodegradable polymer Substances 0.000 description 1
- 230000006287 biotinylation Effects 0.000 description 1
- 238000007413 biotinylation Methods 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 210000001124 body fluid Anatomy 0.000 description 1
- 239000010839 body fluid Substances 0.000 description 1
- 108010006025 bovine growth hormone Proteins 0.000 description 1
- 210000004899 c-terminal region Anatomy 0.000 description 1
- 239000001506 calcium phosphate Substances 0.000 description 1
- 229910000389 calcium phosphate Inorganic materials 0.000 description 1
- 235000011010 calcium phosphates Nutrition 0.000 description 1
- 150000001720 carbohydrates Chemical class 0.000 description 1
- 125000003178 carboxy group Chemical group [H]OC(*)=O 0.000 description 1
- 125000002091 cationic group Chemical group 0.000 description 1
- 238000004113 cell culture Methods 0.000 description 1
- 230000010261 cell growth Effects 0.000 description 1
- 239000013553 cell monolayer Substances 0.000 description 1
- 230000036755 cellular response Effects 0.000 description 1
- 239000007795 chemical reaction product Substances 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 239000005482 chemotactic factor Substances 0.000 description 1
- 229960004926 chlorobutanol Drugs 0.000 description 1
- 239000003593 chromogenic compound Substances 0.000 description 1
- 238000010367 cloning Methods 0.000 description 1
- 239000000701 coagulant Substances 0.000 description 1
- 239000003636 conditioned culture medium Substances 0.000 description 1
- 230000000139 costimulatory effect Effects 0.000 description 1
- 210000004748 cultured cell Anatomy 0.000 description 1
- 230000001186 cumulative effect Effects 0.000 description 1
- 230000009849 deactivation Effects 0.000 description 1
- 239000008367 deionised water Substances 0.000 description 1
- 229910021641 deionized water Inorganic materials 0.000 description 1
- 239000000412 dendrimer Substances 0.000 description 1
- 229920000736 dendritic polymer Polymers 0.000 description 1
- 238000000432 density-gradient centrifugation Methods 0.000 description 1
- 230000008021 deposition Effects 0.000 description 1
- 239000003599 detergent Substances 0.000 description 1
- 229960003957 dexamethasone Drugs 0.000 description 1
- 229960000633 dextran sulfate Drugs 0.000 description 1
- 206010012601 diabetes mellitus Diseases 0.000 description 1
- 102000004419 dihydrofolate reductase Human genes 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- 108700003601 dimethylglycine Proteins 0.000 description 1
- 206010013023 diphtheria Diseases 0.000 description 1
- 239000002270 dispersing agent Substances 0.000 description 1
- PRAKJMSDJKAYCZ-UHFFFAOYSA-N dodecahydrosqualene Natural products CC(C)CCCC(C)CCCC(C)CCCCC(C)CCCC(C)CCCC(C)C PRAKJMSDJKAYCZ-UHFFFAOYSA-N 0.000 description 1
- 239000002552 dosage form Substances 0.000 description 1
- 238000001962 electrophoresis Methods 0.000 description 1
- 206010014599 encephalitis Diseases 0.000 description 1
- 239000002158 endotoxin Substances 0.000 description 1
- 230000002708 enhancing effect Effects 0.000 description 1
- 238000005837 enolization reaction Methods 0.000 description 1
- 239000002662 enteric coated tablet Substances 0.000 description 1
- 238000001976 enzyme digestion Methods 0.000 description 1
- 210000002919 epithelial cell Anatomy 0.000 description 1
- 239000003797 essential amino acid Substances 0.000 description 1
- 235000020776 essential amino acid Nutrition 0.000 description 1
- BEFDCLMNVWHSGT-UHFFFAOYSA-N ethenylcyclopentane Chemical compound C=CC1CCCC1 BEFDCLMNVWHSGT-UHFFFAOYSA-N 0.000 description 1
- 125000001495 ethyl group Chemical group [H]C([H])([H])C([H])([H])* 0.000 description 1
- 229940073505 ethyl vanillin Drugs 0.000 description 1
- 238000011156 evaluation Methods 0.000 description 1
- 238000000855 fermentation Methods 0.000 description 1
- 230000004151 fermentation Effects 0.000 description 1
- 229940126864 fibroblast growth factor Drugs 0.000 description 1
- 238000012757 fluorescence staining Methods 0.000 description 1
- 230000002538 fungal effect Effects 0.000 description 1
- 229940044627 gamma-interferon Drugs 0.000 description 1
- 208000021302 gastroesophageal reflux disease Diseases 0.000 description 1
- 239000008273 gelatin Substances 0.000 description 1
- 229920000159 gelatin Polymers 0.000 description 1
- 235000019322 gelatine Nutrition 0.000 description 1
- 235000011852 gelatine desserts Nutrition 0.000 description 1
- 238000001476 gene delivery Methods 0.000 description 1
- 229930182478 glucoside Natural products 0.000 description 1
- 150000008131 glucosides Chemical class 0.000 description 1
- 229960005150 glycerol Drugs 0.000 description 1
- 150000002337 glycosamines Chemical class 0.000 description 1
- 208000001786 gonorrhea Diseases 0.000 description 1
- 230000009643 growth defect Effects 0.000 description 1
- 239000003102 growth factor Substances 0.000 description 1
- 230000035931 haemagglutination Effects 0.000 description 1
- 229940047650 haemophilus influenzae Drugs 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 230000003862 health status Effects 0.000 description 1
- 239000000185 hemagglutinin Substances 0.000 description 1
- 230000002949 hemolytic effect Effects 0.000 description 1
- HNDVDQJCIGZPNO-UHFFFAOYSA-N histidine Natural products OC(=O)C(N)CC1=CN=CN1 HNDVDQJCIGZPNO-UHFFFAOYSA-N 0.000 description 1
- 102000057593 human F8 Human genes 0.000 description 1
- 239000000413 hydrolysate Substances 0.000 description 1
- 230000002209 hydrophobic effect Effects 0.000 description 1
- 230000008105 immune reaction Effects 0.000 description 1
- 230000007813 immunodeficiency Effects 0.000 description 1
- 230000004957 immunoregulator effect Effects 0.000 description 1
- 238000002513 implantation Methods 0.000 description 1
- 230000006872 improvement Effects 0.000 description 1
- 238000001727 in vivo Methods 0.000 description 1
- 230000002779 inactivation Effects 0.000 description 1
- 230000002458 infectious effect Effects 0.000 description 1
- 230000001524 infective effect Effects 0.000 description 1
- 230000015788 innate immune response Effects 0.000 description 1
- 229910052500 inorganic mineral Inorganic materials 0.000 description 1
- 229940125396 insulin Drugs 0.000 description 1
- 102000006495 integrins Human genes 0.000 description 1
- 108010044426 integrins Proteins 0.000 description 1
- 230000003993 interaction Effects 0.000 description 1
- 238000001361 intraarterial administration Methods 0.000 description 1
- 238000007918 intramuscular administration Methods 0.000 description 1
- 238000007912 intraperitoneal administration Methods 0.000 description 1
- 238000001990 intravenous administration Methods 0.000 description 1
- 239000003456 ion exchange resin Substances 0.000 description 1
- 229920003303 ion-exchange polymer Polymers 0.000 description 1
- 229960000310 isoleucine Drugs 0.000 description 1
- AGPKZVBTJJNPAG-UHFFFAOYSA-N isoleucine Natural products CCC(C)C(N)C(O)=O AGPKZVBTJJNPAG-UHFFFAOYSA-N 0.000 description 1
- 229930027917 kanamycin Natural products 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
- 229930182823 kanamycin A Natural products 0.000 description 1
- 210000003734 kidney Anatomy 0.000 description 1
- 238000002372 labelling Methods 0.000 description 1
- 239000008101 lactose Substances 0.000 description 1
- 230000002045 lasting effect Effects 0.000 description 1
- 208000032839 leukemia Diseases 0.000 description 1
- GZQKNULLWNGMCW-PWQABINMSA-N lipid A (E. coli) Chemical class O1[C@H](CO)[C@@H](OP(O)(O)=O)[C@H](OC(=O)C[C@@H](CCCCCCCCCCC)OC(=O)CCCCCCCCCCCCC)[C@@H](NC(=O)C[C@@H](CCCCCCCCCCC)OC(=O)CCCCCCCCCCC)[C@@H]1OC[C@@H]1[C@@H](O)[C@H](OC(=O)C[C@H](O)CCCCCCCCCCC)[C@@H](NC(=O)C[C@H](O)CCCCCCCCCCC)[C@@H](OP(O)(O)=O)O1 GZQKNULLWNGMCW-PWQABINMSA-N 0.000 description 1
- 150000002632 lipids Chemical class 0.000 description 1
- 229920006008 lipopolysaccharide Polymers 0.000 description 1
- 210000004185 liver Anatomy 0.000 description 1
- 206010025135 lupus erythematosus Diseases 0.000 description 1
- 210000004698 lymphocyte Anatomy 0.000 description 1
- 201000004792 malaria Diseases 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 238000005259 measurement Methods 0.000 description 1
- 108020004999 messenger RNA Proteins 0.000 description 1
- LXCFILQKKLGQFO-UHFFFAOYSA-N methylparaben Chemical compound COC(=O)C1=CC=C(O)C=C1 LXCFILQKKLGQFO-UHFFFAOYSA-N 0.000 description 1
- 235000013336 milk Nutrition 0.000 description 1
- 239000008267 milk Substances 0.000 description 1
- 210000004080 milk Anatomy 0.000 description 1
- 239000011707 mineral Substances 0.000 description 1
- 239000002480 mineral oil Substances 0.000 description 1
- 235000010446 mineral oil Nutrition 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 238000010369 molecular cloning Methods 0.000 description 1
- 229940035032 monophosphoryl lipid a Drugs 0.000 description 1
- 229910000402 monopotassium phosphate Inorganic materials 0.000 description 1
- 235000019796 monopotassium phosphate Nutrition 0.000 description 1
- 239000010413 mother solution Substances 0.000 description 1
- 210000000214 mouth Anatomy 0.000 description 1
- 210000004877 mucosa Anatomy 0.000 description 1
- 201000006417 multiple sclerosis Diseases 0.000 description 1
- BSOQXXWZTUDTEL-ZUYCGGNHSA-N muramyl dipeptide Chemical compound OC(=O)CC[C@H](C(N)=O)NC(=O)[C@H](C)NC(=O)[C@@H](C)O[C@H]1[C@H](O)[C@@H](CO)O[C@@H](O)[C@@H]1NC(C)=O BSOQXXWZTUDTEL-ZUYCGGNHSA-N 0.000 description 1
- 125000001446 muramyl group Chemical group N[C@@H](C=O)[C@@H](O[C@@H](C(=O)*)C)[C@H](O)[C@H](O)CO 0.000 description 1
- 210000003205 muscle Anatomy 0.000 description 1
- 238000002703 mutagenesis Methods 0.000 description 1
- 231100000350 mutagenesis Toxicity 0.000 description 1
- 230000035772 mutation Effects 0.000 description 1
- 208000010753 nasal discharge Diseases 0.000 description 1
- 108700004028 nef Genes Proteins 0.000 description 1
- 101150023385 nef gene Proteins 0.000 description 1
- 229940052778 neisseria meningitidis Drugs 0.000 description 1
- 229940053128 nerve growth factor Drugs 0.000 description 1
- 230000001537 neural effect Effects 0.000 description 1
- 210000002569 neuron Anatomy 0.000 description 1
- 108091027963 non-coding RNA Proteins 0.000 description 1
- 102000042567 non-coding RNA Human genes 0.000 description 1
- QTNLALDFXILRQO-UHFFFAOYSA-N nonadecane-1,2,3-triol Chemical compound CCCCCCCCCCCCCCCCC(O)C(O)CO QTNLALDFXILRQO-UHFFFAOYSA-N 0.000 description 1
- 230000030648 nucleus localization Effects 0.000 description 1
- 229940038384 octadecane Drugs 0.000 description 1
- 239000003921 oil Substances 0.000 description 1
- 230000008520 organization Effects 0.000 description 1
- 150000004880 oxines Chemical class 0.000 description 1
- 101150000558 p37 gene Proteins 0.000 description 1
- 229940090668 parachlorophenol Drugs 0.000 description 1
- 230000003071 parasitic effect Effects 0.000 description 1
- 238000007911 parenteral administration Methods 0.000 description 1
- 239000002245 particle Substances 0.000 description 1
- 229940051027 pasteurella multocida Drugs 0.000 description 1
- 230000007918 pathogenicity Effects 0.000 description 1
- 230000037361 pathway Effects 0.000 description 1
- 210000005259 peripheral blood Anatomy 0.000 description 1
- 239000011886 peripheral blood Substances 0.000 description 1
- 230000002093 peripheral effect Effects 0.000 description 1
- 230000002085 persistent effect Effects 0.000 description 1
- 239000000825 pharmaceutical preparation Substances 0.000 description 1
- 229960003742 phenol Drugs 0.000 description 1
- 229960003531 phenolsulfonphthalein Drugs 0.000 description 1
- NBIIXXVUZAFLBC-UHFFFAOYSA-K phosphate Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 description 1
- 239000010452 phosphate Substances 0.000 description 1
- 150000008104 phosphatidylethanolamines Chemical class 0.000 description 1
- PJNZPQUBCPKICU-UHFFFAOYSA-N phosphoric acid;potassium Chemical compound [K].OP(O)(O)=O PJNZPQUBCPKICU-UHFFFAOYSA-N 0.000 description 1
- 230000035479 physiological effects, processes and functions Effects 0.000 description 1
- 108010086662 phytohemagglutinin-M Proteins 0.000 description 1
- 108700004029 pol Genes Proteins 0.000 description 1
- 101150088264 pol gene Proteins 0.000 description 1
- 229920001983 poloxamer Polymers 0.000 description 1
- 229920000768 polyamine Polymers 0.000 description 1
- 235000010241 potassium sorbate Nutrition 0.000 description 1
- 239000004302 potassium sorbate Substances 0.000 description 1
- 229940069338 potassium sorbate Drugs 0.000 description 1
- NNGFQKDWQCEMIO-UHFFFAOYSA-M potassium;hydron;phosphonato phosphate Chemical compound [K+].OP(O)(=O)OP(O)([O-])=O NNGFQKDWQCEMIO-UHFFFAOYSA-M 0.000 description 1
- 230000003449 preventive effect Effects 0.000 description 1
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 description 1
- 239000000473 propyl gallate Substances 0.000 description 1
- 229940075579 propyl gallate Drugs 0.000 description 1
- 235000010388 propyl gallate Nutrition 0.000 description 1
- 238000012797 qualification Methods 0.000 description 1
- 150000004053 quinones Chemical class 0.000 description 1
- ZAHRKKWIAAJSAO-UHFFFAOYSA-N rapamycin Natural products COCC(O)C(=C/C(C)C(=O)CC(OC(=O)C1CCCCN1C(=O)C(=O)C2(O)OC(CC(OC)C(=CC=CC=CC(C)CC(C)C(=O)C)C)CCC2C)C(C)CC3CCC(O)C(C3)OC)C ZAHRKKWIAAJSAO-UHFFFAOYSA-N 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 229940047431 recombinate Drugs 0.000 description 1
- 230000006798 recombination Effects 0.000 description 1
- 238000005215 recombination Methods 0.000 description 1
- 239000012925 reference material Substances 0.000 description 1
- 230000022532 regulation of transcription, DNA-dependent Effects 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 201000009410 rhabdomyosarcoma Diseases 0.000 description 1
- 206010039073 rheumatoid arthritis Diseases 0.000 description 1
- 108091092562 ribozyme Proteins 0.000 description 1
- 201000005404 rubella Diseases 0.000 description 1
- 150000003839 salts Chemical class 0.000 description 1
- 229930182490 saponin Natural products 0.000 description 1
- 150000007949 saponins Chemical class 0.000 description 1
- 235000017709 saponins Nutrition 0.000 description 1
- 230000003248 secreting effect Effects 0.000 description 1
- 230000035945 sensitivity Effects 0.000 description 1
- 229940007046 shigella dysenteriae Drugs 0.000 description 1
- 230000035939 shock Effects 0.000 description 1
- 239000002356 single layer Substances 0.000 description 1
- 229960002930 sirolimus Drugs 0.000 description 1
- QFJCIRLUMZQUOT-HPLJOQBZSA-N sirolimus Chemical compound C1C[C@@H](O)[C@H](OC)C[C@@H]1C[C@@H](C)[C@H]1OC(=O)[C@@H]2CCCCN2C(=O)C(=O)[C@](O)(O2)[C@H](C)CC[C@H]2C[C@H](OC)/C(C)=C/C=C/C=C/[C@@H](C)C[C@@H](C)C(=O)[C@H](OC)[C@H](O)/C(C)=C/[C@@H](C)C(=O)C1 QFJCIRLUMZQUOT-HPLJOQBZSA-N 0.000 description 1
- 229940054269 sodium pyruvate Drugs 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 239000000243 solution Substances 0.000 description 1
- 239000002904 solvent Substances 0.000 description 1
- 235000010199 sorbic acid Nutrition 0.000 description 1
- 229940075582 sorbic acid Drugs 0.000 description 1
- 239000004334 sorbic acid Substances 0.000 description 1
- 229940031439 squalene Drugs 0.000 description 1
- TUHBEKDERLKLEC-UHFFFAOYSA-N squalene Natural products CC(=CCCC(=CCCC(=CCCC=C(/C)CCC=C(/C)CC=C(C)C)C)C)C TUHBEKDERLKLEC-UHFFFAOYSA-N 0.000 description 1
- 238000010186 staining Methods 0.000 description 1
- 238000010561 standard procedure Methods 0.000 description 1
- 239000008227 sterile water for injection Substances 0.000 description 1
- 230000001954 sterilising effect Effects 0.000 description 1
- 238000004659 sterilization and disinfection Methods 0.000 description 1
- 230000000638 stimulation Effects 0.000 description 1
- UCSJYZPVAKXKNQ-HZYVHMACSA-N streptomycin Natural products CN[C@H]1[C@H](O)[C@@H](O)[C@H](CO)O[C@H]1O[C@@H]1[C@](C=O)(O)[C@H](C)O[C@H]1O[C@@H]1[C@@H](NC(N)=N)[C@H](O)[C@@H](NC(N)=N)[C@H](O)[C@H]1O UCSJYZPVAKXKNQ-HZYVHMACSA-N 0.000 description 1
- 229960005322 streptomycin Drugs 0.000 description 1
- 238000007920 subcutaneous administration Methods 0.000 description 1
- 239000005720 sucrose Substances 0.000 description 1
- 108010012704 sulfated glycoprotein p50 Proteins 0.000 description 1
- 229940044609 sulfur dioxide Drugs 0.000 description 1
- 235000010269 sulphur dioxide Nutrition 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 239000000829 suppository Substances 0.000 description 1
- 239000000375 suspending agent Substances 0.000 description 1
- 208000011580 syndromic disease Diseases 0.000 description 1
- 230000009885 systemic effect Effects 0.000 description 1
- 239000003826 tablet Substances 0.000 description 1
- 108700004027 tat Genes Proteins 0.000 description 1
- 101150098170 tat gene Proteins 0.000 description 1
- GKCBAIGFKIBETG-UHFFFAOYSA-N tetracaine Chemical compound CCCCNC1=CC=C(C(=O)OCCN(C)C)C=C1 GKCBAIGFKIBETG-UHFFFAOYSA-N 0.000 description 1
- 229960002372 tetracaine Drugs 0.000 description 1
- 229960002180 tetracycline Drugs 0.000 description 1
- 229930101283 tetracycline Natural products 0.000 description 1
- 235000019364 tetracycline Nutrition 0.000 description 1
- 150000003522 tetracyclines Chemical class 0.000 description 1
- 238000010257 thawing Methods 0.000 description 1
- 230000005030 transcription termination Effects 0.000 description 1
- 230000002103 transcriptional effect Effects 0.000 description 1
- 238000003151 transfection method Methods 0.000 description 1
- 238000012546 transfer Methods 0.000 description 1
- 238000003146 transient transfection Methods 0.000 description 1
- 108091007466 transmembrane glycoproteins Proteins 0.000 description 1
- 102000035160 transmembrane proteins Human genes 0.000 description 1
- 108091005703 transmembrane proteins Proteins 0.000 description 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 description 1
- 201000008827 tuberculosis Diseases 0.000 description 1
- IESDGNYHXIOKRW-LEOABGAYSA-N tuftsin Chemical compound C[C@@H](O)[C@H](N)C(=O)N[C@@H](CCCCN)C(=O)N1CCC[C@H]1C(=O)N[C@H](CCCNC(N)=N)C(O)=O IESDGNYHXIOKRW-LEOABGAYSA-N 0.000 description 1
- 229940035670 tuftsin Drugs 0.000 description 1
- 102000003298 tumor necrosis factor receptor Human genes 0.000 description 1
- 241000712461 unidentified influenza virus Species 0.000 description 1
- 238000011144 upstream manufacturing Methods 0.000 description 1
- 239000004474 valine Substances 0.000 description 1
- 239000003981 vehicle Substances 0.000 description 1
- 239000002435 venom Substances 0.000 description 1
- 210000001048 venom Anatomy 0.000 description 1
- 231100000611 venom Toxicity 0.000 description 1
- 229940118696 vibrio cholerae Drugs 0.000 description 1
- 108700026220 vif Genes Proteins 0.000 description 1
- 101150059019 vif gene Proteins 0.000 description 1
- 230000007501 viral attachment Effects 0.000 description 1
- 244000052613 viral pathogen Species 0.000 description 1
- 108700026215 vpr Genes Proteins 0.000 description 1
- 101150024249 vpr gene Proteins 0.000 description 1
- 108700026222 vpu Genes Proteins 0.000 description 1
- 101150090490 vpu gene Proteins 0.000 description 1
- 238000001262 western blot Methods 0.000 description 1
- 239000011701 zinc Substances 0.000 description 1
- 229910052725 zinc Inorganic materials 0.000 description 1
Images
Classifications
-
- 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
- C12N15/00—Mutation or genetic engineering; DNA or RNA concerning genetic engineering, vectors, e.g. plasmids, or their isolation, preparation or purification; Use of hosts therefor
- C12N15/09—Recombinant DNA-technology
- C12N15/63—Introduction of foreign genetic material using vectors; Vectors; Use of hosts therefor; Regulation of expression
- C12N15/79—Vectors or expression systems specially adapted for eukaryotic hosts
- C12N15/85—Vectors or expression systems specially adapted for eukaryotic hosts for animal cells
- C12N15/86—Viral vectors
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K48/00—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K38/00—Medicinal preparations containing peptides
- A61K38/16—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- A61K38/17—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- A61K38/19—Cytokines; Lymphokines; Interferons
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K39/12—Viral antigens
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K39/12—Viral antigens
- A61K39/205—Rhabdoviridae, e.g. rabies virus
-
- 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
- A61K39/21—Retroviridae, e.g. equine infectious anemia virus
-
- 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
- A61K2039/5256—Virus expressing foreign proteins
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K2039/51—Medicinal preparations containing antigens or antibodies comprising whole cells, viruses or DNA/RNA
- A61K2039/53—DNA (RNA) vaccination
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K2039/54—Medicinal preparations containing antigens or antibodies characterised by the route of administration
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K2039/54—Medicinal preparations containing antigens or antibodies characterised by the route of administration
- A61K2039/541—Mucosal route
- A61K2039/543—Mucosal route intranasal
-
- 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
- A61K2039/545—Medicinal preparations containing antigens or antibodies characterised by the dose, timing or administration schedule
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K2039/555—Medicinal preparations containing antigens or antibodies characterised by a specific combination antigen/adjuvant
- A61K2039/55511—Organic adjuvants
- A61K2039/55522—Cytokines; Lymphokines; Interferons
- A61K2039/55527—Interleukins
- A61K2039/55538—IL-12
-
- 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
- C12N2740/00—Reverse transcribing RNA viruses
- C12N2740/00011—Details
- C12N2740/10011—Retroviridae
- C12N2740/15011—Lentivirus, not HIV, e.g. FIV, SIV
- C12N2740/15034—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
- C12N2740/00—Reverse transcribing RNA viruses
- C12N2740/00011—Details
- C12N2740/10011—Retroviridae
- C12N2740/16011—Human Immunodeficiency Virus, HIV
- C12N2740/16034—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
- C12N2760/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA ssRNA viruses negative-sense
- C12N2760/00011—Details
- C12N2760/16011—Orthomyxoviridae
- C12N2760/16111—Influenzavirus A, i.e. influenza A virus
- C12N2760/16134—Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12N—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA
- C12N2760/00—MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA ssRNA viruses negative-sense
- C12N2760/00011—Details
- C12N2760/20011—Rhabdoviridae
- C12N2760/20211—Vesiculovirus, e.g. vesicular stomatitis Indiana virus
- C12N2760/20241—Use of virus, viral particle or viral elements as a vector
- C12N2760/20243—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
- C12N2840/00—Vectors comprising a special translation-regulating system
- C12N2840/20—Vectors comprising a special translation-regulating system translation of more than one cistron
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- General Health & Medical Sciences (AREA)
- Virology (AREA)
- Engineering & Computer Science (AREA)
- Medicinal Chemistry (AREA)
- Veterinary Medicine (AREA)
- Pharmacology & Pharmacy (AREA)
- Epidemiology (AREA)
- Animal Behavior & Ethology (AREA)
- Genetics & Genomics (AREA)
- Public Health (AREA)
- Immunology (AREA)
- Microbiology (AREA)
- Mycology (AREA)
- Zoology (AREA)
- Biotechnology (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Biomedical Technology (AREA)
- Organic Chemistry (AREA)
- Molecular Biology (AREA)
- Wood Science & Technology (AREA)
- General Engineering & Computer Science (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Physics & Mathematics (AREA)
- Hematology (AREA)
- Biophysics (AREA)
- Communicable Diseases (AREA)
- Plant Pathology (AREA)
- Gastroenterology & Hepatology (AREA)
- Biochemistry (AREA)
- Tropical Medicine & Parasitology (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
- Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
Abstract
A method of inducing an antigen-specific immune response in a mammalian subject includes the steps of administering to the subject an effective amount of a first composition comprising a DNA plasmid comprising a DNA sequence encoding an antigen under the control of regulatory sequences directing expression thereof in a mammalian or vertebrate cell. The method also includes administering to the subject an effective amount of a second composition comprising a recombinant vesicular stomatitis virus (rVSV) comprising a nucleic acid sequence encoding the antigen under the control of regulatory sequences directing expression thereof in the mammalian or vertebrate cell. The rVSV is in one embodiment replication competent. Kits for use in immunizations and therapeutic treatments of disease include the components and instructions for practice of this method.
Description
The application is to be on March 23rd, 2004 applying date, the dividing an application of the application for a patent for invention that application number is 200480007818.0, denomination of invention is identical with the present invention.The present invention's part is subsidized (NIH, subsidy NIH NO1-AI05397 and NIH NO1-AI 25458) by the fund from U.S. government.Therefore U.S. government enjoys some right to the present invention.
Background of invention
In order to strengthen the effectiveness of immunogenic composition, already reported panimmunity originality compositions and method, used protein compositions, the compositions based on plasmid and recombinant virus construct as immunogenic composition.Research has before shown the immunogenic composition based on plasmid, in whole body application, make systemic immune system to use secondary general immunization such as the conventional antigen of protein or recombinant virus have more susceptibility (referring to as, Xiang etc., 1997Springer Semin.Iiiimunopathol., 19:257-268; Schneider, J. etc., 1998Nature Med., 4:397; And Sedeguh, M. etc., 1998Proc.Natl.Acad.Sci., USA, 95:7648; Rogers, W.O. etc., 2001Infec. & Immun., 69 (9): 5565-72; Eo, S.K., etc., 2001 J.Immunol., 166 (9): 5473-9; Ramshaw I.A. and Ramsay, A.J., 2000 Immunol.Today, 21 (4): 163-5).
A reinforcement therapy for frequent just exempt from/live vector of use DNA, it comprises by vaccinia virus and carrys out booster immunization.Example in document has comprised above-mentioned immunity inoculation (Hanke T. etc., 2002 Vaccine.20 (15): 1995-8 that HIV (human immunodeficiency virus) (HIV) is carried out recently; Amara R.R. etc., 2002 Vaccine.20 (15): 1949-55; Wee E.G. etc., 2002 J.Gen.Virol., 83 (Pt 1): 75-80; Amara R.R. etc., 2001 Science.292 (5514): 69-74).Just exempt from-strengthen (prime-boost) immunity inoculation and use the vaccinia virus vector antigen expressed of DNA and modification, such as herpes simplex virus-2 glycoprotein D, leishmania infantum (Leishmania infantum) P36/LACK antigen, Plasmodium falciparum (Plasmodium falciparum) TRAP antigen, HIV/SIV antigen, Mus tuberculosis antigen and influenza antigen, already reported this immunity inoculation can bring out specific antibody and cytokine response (referring to, such as Meseda C.A. etc., 2002 J.Infect.Dis., 186 (8): 1065-73; Amara R.R. etc., 2002 J.Virol., 76 (15): 7625-31; Gonzalo R.M. etc., 2002 Vaccine, 20 (7-8): 1226-31; Schneider J. etc., 2001 Vaccine, 19 (32): 4595-602; HelZ. etc., 2001 J.Immunol., 167 (12): 7180-91; McShane H. etc., 2001 Infec. & Immun., 69 (2): 1999 Vaccine such as 681-6 and Degano P., 18 (7-8): 623-32 influenza and malaria model).
Recently, already reported genetic immunization therapy that plasmid adenovirus just exempts to strengthen can induce the immunity of α-fetoprotein specific tumour and protect pig not affected pig cholera (referring to, for example, Meng W.S.2001Cancer Res., 61 (24): 8782-6; Hammond, J.M. etc., 2001 Vet.Microbio., 80 (2): 101-19 and U.S. Patent number 6,210,663).
Already reported that other DNA plasmid virus just exempted from the therapy of strengthening.Referring to, for example, Matano T. etc., 2001 J.Virol., 75 (23): 11891-6 (just exempt from/sendai virus vector of DNA booster immunization).Reported with the DNA that recombinant poxvirus just exempts to strengthen can be used for HIV-1 treatment (referring to, for example, Kent, S.J. etc., 1998J.Virol., 72:10180-8; Robinson, H.L. etc., 1999 Nat.Med., 5:526-34; And Tartaglia, J. etc., 1998 AIDS Res.Human Retrovirus., 14:S291-8).
Although having evaluated many DNA just exempts from/virus reinforcement therapy, and described immunity inoculation therapy still has some shortcomings at present.For example, in experimenter, some above-mentioned viruses of mentioning can produce disease symptoms; Separately there are some may cause restructuring in body.Some viruses are difficult to preparation and/or receive the limited in one's ability of exogenous gene.Also have some viral disadvantages to be to cause the carrier immunity being pre-existing in a large number in human body, and other security consideration.
These need the new and useful immunity inoculation therapy in this area, and discussed antigen is produced cell and the humoral immunoresponse(HI) level strengthening and meets safety, easily preparation and overcome the requirement of mammalian hosts to the innate immunity of carrier when booster immunization is inoculated.
Summary of the invention
The invention provides a kind of new method, compositions and test kit, by just exempting from/strengthen therapy for the immunne response at mammalian subject inducing anti-disease Proantigen or other antigen, compare with using separately the result that DNA plasmid component or recombinant virus component obtain, for described antigen, shown that wonderful cellullar immunologic response works in coordination with stimulation.
In one embodiment, the invention provides a kind of in mammalian subject the new method of inducing antigen-specific immunne response.The method comprises the first compositions that is applied to experimenter's effective dose, described the first compositions comprises the DNA plasmid of the DNA sequence that contains coding for antigens, and described coded sequence is instructed its regulating and controlling sequence of expressing in mammalian cell by DNA plasmid to regulate and control.The method also comprises the second compositions that is applied to experimenter's effective dose, described the second compositions comprises the restructuring vesicular stomatitis virus (rVSV) that contains coding for antigens nucleotide sequence, described coded sequence is subject to the regulation and control of following regulating and controlling sequence, and this regulating and controlling sequence regulates and controls above-mentioned coded sequence expresses by rVSV in mammalian cell.In one embodiment, described restructuring VSVA is attenuation, reproducible virus.In another embodiment, described restructuring VSV is the virus not copying.Can use described the first and second compositionss in any order.In addition, the present invention considers after a kind of compositions, repeatedly to use another compositions repeatedly using wherein again.In one embodiment, be preferably total to administer cytokines.
In another embodiment of the invention, provide a kind of for the immunogenic composition at the former generation antigen-specific immune response of mammalian subject induction antagonism.Described immunogenic composition comprises the first compositions of the DNA plasmid of the DNA sequence that contains the described antigen of encoding, and described coded sequence is subject to the regulation and control of following regulating and controlling sequence, and this regulating and controlling sequence regulates and controls above-mentioned coded sequence by DNA plasmid expression.Said composition also comprises at least one reproducible component, restructuring vesicular stomatitis virus (VSV), and the nucleotide sequence that it comprises the same antigen of encoding, described coded sequence is instructed its regulating and controlling sequence of expressing by the VSV that recombinates to regulate and control.
In another embodiment, the invention provides the test kit using in a kind for the treatment of of antigen specific immune reaction of inducing enhanced level in mammalian subject or prevention method.This test kit comprises, wherein, at least one first compositions, it comprises the DNA plasmid of the DNA sequence that contains the described antigen of encoding, the regulation and control of the modulated regulating and controlling sequence that it expresses in mammal of described coded sequence; At least one second compositions, it comprises a kind of reproducible component, the restructuring vesicular stomatitis virus (rVSV) of the nucleotide sequence that it comprises the described antigen of encoding, the regulation and control of the modulated regulating and controlling sequence that it expresses in mammal of described coded sequence; With the description of implementing said method.
In another embodiment, the invention provides above-mentioned immunogenic composition or its component and produce the purposes in immunoreactive medicine preparing the antigen that induced animal uses in to said composition.
Other side of the present invention and embodiment are disclosed in following detailed description.
Accompanying drawing summary
Figure 1A is the exemplary plasmid DNA schematic diagram of coding simian immunodeficiency virus (SIV) gag p37 albumen.The figure illustrates this plasmid and contain human cytomegalic inclusion disease virus (HCMV) promoter/enhancer of handling gag protein expression, bovine growth hormone polyadenylation site (BGH polyA), origin of replication sequence (ori) and kalamycin resistance (kan
r) marker gene.
Figure 1B is the p35 of coding Rhesus Macacus interleukin 12 and the exemplary bicistronic plasmid DNA schematic diagram of p40 subunit.P35 subunit is regulated and controled by HCMV promoter and has SV40polyA site.P40 subunit is subject to simian cytomegalovirus (SCMV) promoter to regulate and control, have BGH polyA site reverse transcription.This plasmid also contains ori sequence and kan
rgene.
Fig. 2 is presented at by the present invention just to exempt from/strengthen the block diagram that in not classification peripheral blood lymphocytes of therapy institute immune animal (PBMC), VSV N specificity IFN-γ (IFN-γ) ELISpot replys.Coding SIV gag protein D NA plasmid immunity inoculation for leftmost secret note representative, and the scheme of strengthening with the VSV carrier of expressing influenza hemagglutinin (flu HA) albumen.The representative of light gray bar relates to the present invention program who strengthens with the VSV that expresses HIV gag and env albumen again with after the initial immunity inoculation of DNA gag plasmid.Informal voucher is used the scheme of the VSV immunity inoculation of expressing HIV gag and env albumen after representing the initial immunity inoculation of DNA plasmid (con DNA) relating to empty or contrast again.The representative of rightmost secret note relates to after the initial immunity inoculation of con DNA plasmid again by the scheme of expressing the VSV immunity inoculation of flu HA albumen.The result of every group of representative is from 5 animals.
Fig. 3 shows by the present invention just to exempt from/strengthen the therapy institute immune animal block diagram that HIV env 6101 specificity IFN-γ (IFN-γ) ELISpot does not reply in classification PBMC.The DNA plasmid immunity inoculation of coding SIV gag albumen the scheme of strengthening with the VSV carrier of expressing fluHA albumen for leftmost light gray bar representative.The representative of speckle bar relates to the present invention program who strengthens with the VSV that expresses HIV gag and env albumen again with after the initial immunity inoculation of DNA gag plasmid.The representative of grid bar relates to using the scheme of the VSV immunity inoculation of expressing HIV gag and env albumen after the empty initial immunity inoculation of contrast DNA plasmid again.The bar representative of getting ready relates to the scheme of inoculating with the VSV booster immunization of expressing flu HA albumen again with after the initial immunity inoculation of conDNA plasmid.The result of every group of representative is from 5 animals.Asterisk represents to produce statistically significant difference, p=0.0001.
Fig. 4 shows the figure that is just exempted from/strengthened the anti-SIV gag of therapy immune serum p27 antibody titer by enzyme-linked immunosorbent assay (ELISA) the present invention.At the 0th day, the 4th week and the 8th week, use plasmid DNA and at the 15th and the 23rd week, use respectively VSV (Indiana G serotype) and VSV (Chandipura G serotype) reinforcement.By the DNA plasmid immunity inoculation of coding SIV gag albumen, also by the scheme that the VSV carrier of expressing flu HA albumen is strengthened, be expressed as (◆).Relate to DNA gag plasmid and just exempt from by the present invention program that the VSV that expresses HIV gag and env albumen strengthens, to be expressed as (■) again after immunity inoculation.Relate to being expressed as (▲) by the scheme of expressing the VSV immunity inoculation of HIV gag and env albumen again after the empty initial immunity inoculation of contrast DNA plasmid.Relate to being expressed as (●) by the scheme of expressing the VSV immunity inoculation of fluHA albumen again after the initial immunity inoculation of contrast DNA plasmid.The result of every group of representative is from 5 animals.Between each group, statistically significant difference is expressed as p=0.0073 (*); P=0.5941 (#) or p=0.0027
Fig. 5 is the figure that shows SIV gag specificity spot formation cell in animal per 1,000,000 cells of just being exempted from/strengthening therapy immunity inoculation by the evaluation of ELISpot algoscopy with the present invention.At the 0th day, the 4th week and the 8th week, use plasmid DNA and at the 15th and the 23rd week, use respectively VSV (Indiana G serotype) and VSV (Chandipura G serotype) reinforcement.By the DNA plasmid immunity inoculation of coding SIV gag albumen, also by the scheme that the VSV carrier of expressing flu HA albumen is strengthened, be expressed as (◆).Relate to being expressed as (■) by the present invention program that the VSV that expresses HIV gag and env albumen strengthens again after the initial immunity inoculation of DNA gag plasmid.Relate to being expressed as (▲) by the scheme of expressing the VSV immunity inoculation of HIV gag and env albumen again after the empty initial immunity inoculation of contrast DNA plasmid.Relate to being expressed as (●) by the scheme of expressing the VSV immunity inoculation of flu HA albumen again after the initial immunity inoculation of contrast DNA plasmid.The result of every group of representative is from 5 animals.Between each group, statistically significant difference is expressed as p=0.0001 and p=0.0002 by square brackets.
Fig. 6 shows by just exempting from/strengthens the figure in conjunction with the immunne response of the reinforcement causing, shown in symbol as Fig. 5, cause the AIDS protective effect that increases, by measuring cd4 t cell loss minimizing in several days after exciting, determine.
Fig. 7 shows by just exempting from/strengthens the figure in conjunction with the immunne response of the reinforcement causing, shown in symbol as Fig. 5, cause the AIDS protective effect that increases, by measuring blood plasma internal recycle virus (viral copy number/ml) minimizing in several days after exciting, determine.
Detailed Description Of The Invention
The invention provides a kind of in mammalian subject or vertebrate subject the new method of inducing antigen-specific immunne response, by being combined with some component of the immunogenic composition described in prior art, and optimize component to produce wondrous and collaborative effect.Generally speaking, the method comprises the compositions that is applied to experimenter's effective dose, described compositions comprises the DNA plasmid of the DNA sequence that contains the described antigen of encoding, and described coded sequence is instructed it in mammal or vertebrate cells, to pass through the regulation and control of the regulating and controlling sequence of DNA plasmid expression.The method also comprises the step of the compositions that is applied to experimenter's effective dose, and described compositions comprises a kind of reproducible component, restructuring vesicular stomatitis virus (rVSV).This rVSV comprises coding for antigens nucleotide sequence, and described coded sequence is instructed the regulation and control of its regulating and controlling sequence of expressing by rVSV in mammal and vertebrate cells.
The compositions of first using in these two kinds of immunogenic compositions is called just exempts from compositions (prime compositon).The immunogenic composition of using is again called strengthens compositions (boosting composition).Can DNA compositions use as just exempting from compositions, VSV carrier compositions is used as strengthening compositions, vice versa.In one embodiment, before VSV carrier is used as reinforcement compositions, be applied to experimenter at least one times or repeatedly just exempt from compositions.Be applied to experimenter at least one times or repeatedly strengthen compositions thereafter.In addition, the present invention considers after a kind of compositions, repeatedly to use another kind of compositions repeatedly using wherein.The method is further considered and is used the cytokine of effective dose as a step of the method.
Already be surprised to find and used separately DNA plasmid or restructuring VSV compares, and having implemented the method and can induce mammalian subject to produce the immunne response strengthening, being included in replying antigen in CD8+T cell.In fact, shown in the following examples, immunne response is also eager to excel compared with these two kinds of immunogenic composition additive combination are desired.Therefore the immunne response that, this new method is induced has surprising and concertedness enhancing in cell and/or the anti-antigen-reactive of antibody.Referring to, as, following table 1 and Figure 4 and 5, wherein to the peak value of the antigenic specificity cell response of HIV-1gag be use separately that DNA plasmid immunogenic composition replys more than 3 times, or be almost to use separately 9 times that rVSV immunogenic composition is replied.
Although described mammalian subject is primates, people preferably, the present invention is not limited in and is accredited as mammiferous experimenter.Details are as follows for the component of the method, about quoted as proof document, at this, is incorporated herein by reference, and for those skilled in the art, understands in detail.
A.DNA plasmid immunogenic composition
Immunogenic composition of the present invention comprises DNA plasmid, and described DNA plasmid comprises coding and needs to be produced for it DNA sequence of the selected antigen of immunne response.In this plasmid, selected antigen is instructed its regulating and controlling sequence of expressing in mammal or vertebrate cells to regulate and control.This plasmid self component is conventional.
The non-viral plasmid vector can be used in the present invention contains DNA sequence separative and purification, the DNA sequence that described DNA sequence contains the described selected immunogenicity antigen of encoding.This DNA molecular can be derived from virus or non-viral, as being already designed for the antibacterial of encoding exogenous or heterologous nucleic acid sequence.Such plasmid or carrier comprise the sequence that derives from virus or phage.Many non-virus carriers are known in the art and can include but are not limited to plasmid, bacteria carrier, phage vector, " naked " DNA and the DNA condensing with cation lipid or polymer.
The example of bacteria carrier comprises, but be not limited to, be derived from bacillus calmette-guerin vaccine (bacille Calrnette Gu é rin) (BCG), the sequence of Salmonella (Salmonella), shigella dysenteriae (Shigella), escherichia coli (E.coli) and Listerella (Listeria) etc.Suitable plasmid vector comprises, for example, pBR322, pBR325, pACYC177, pACYC184, pUC8, pUC9, pUC18, pUC19, pLG339, pR290, pK37, pKC101, pAC105, pVA51, pKH47, pUB110, pMB9, pBR325, Col E1, pSC101, pBR313, pML21, RSF2124, pCRl, RP4, pBAD18 and pBR328.
The example of suitable derivable coli expression carrier comprises pTrc (Amann etc., 1988Gene, 69:301-315), arabinose expression vector (as, pBAD18, Guzman etc., 1995 J.Bacteriol., 177:4121-4130) and pETIId (Studier etc., 1990Methods in Enzymology, 185:60-89).The expression of pTrc carrier target gene depends on the host RNA polymerase starting from the trp-lac promoter, fusion of heterozygosis and transcribes.The expression of pETIId carrier target gene depends on transcribing of starting from T7gn10-lac promoter, fusion, and the viral rna polymerase T7gn1 that it is coexpression is mediated.This varial polymerases provides from Host Strains BL21 (DE3) or HMS I 74 (DE3), derives from the contained T7gn1 gene of resident (resident) prophage under the regulation and control of lacUV5 promoter transcription.PBAD system depends on the derivable Arabinose promoter that regulates and controls for araC gene.This promoter is induced under arabinose existence condition.
Described promoter and other regulating and controlling sequence are handled antigen and are expressed in required mammal or vertebrate host, can be selected from similarly a series of known promoteres that can be used for this object.Many such promoteres are described below.In one embodiment, immunogenicity DNA plasmid compositions is described below, available promoter is that human cytomegalic inclusion disease virus (HCMV) promoter/enhancer (is described in, as U.S. Patent number 5,168,062 and 5,385, in 839, in this, be incorporated herein by reference) and SCMV promoter/enhancer.
Nucleotide sequence of the present invention, the additional regulating and controlling sequence comprising in molecule or carrier comprises, be not limited to, enhancer sequence, polyadenylation sequence, shear donor sequences and shear receptor sequence, lay respectively at the polypeptide that will be translated and start and the transcription initiation of end and termination site, ribosome binding site for transcriptional domain translation, Epitope tag, nuclear localization sequence, IRES element, Goldberg-Hogness " TATA " element, restriction enzyme site, selected marker thing etc.Enhancer sequence comprises, as, the long terminal repetition or LTRs of the 72bp tandem repetitive sequence of SV40 DNA or retrovirus etc., for increasing transcribing efficiency.
Available other element in DNA plasmid, comprise, as origin of replication, polyadenylation sequence (as BGH polyA, SV40polyA), drug resistance labelling (as kalamycin resistance) etc. can also be selected from many known arrays, comprise described in those embodiment and mention especially as follows.
The selection of promoter and other common carrier element is conventional, and it can obtain many such sequences, for designing plasmid of the present invention.Referring to, as Sambrook etc., MolecularCloning A Laboratory Manual, Cold Spring Harbor Laboratory, New York, and the reference material of quoting as proof (1989), for example, 3.18-3.26 page and 16.17-16.27 page and Ausubel etc., Current Protocols in Molecular Biology, John Wiley & Sons, New York (1989).Those skilled in the art can easily select all spendable plasmid components in the present invention from raw material known in the art, and can obtain from pharmaceuticals industry.The selection of plasmid component and regulating and controlling sequence is without considering limitation of the present invention.
In disclosed patent, describe below the example for the suitable DNA plasmid construction body of immunogenic composition in detail, disclosed content is incorporated herein by reference at this, as, International Patent Publication No. WO 98/17799, WO99/43839 and WO98/17799; And U.S. Patent number 5,593,972,5,817,637,5,830,876 and 5,891,505 etc.
Similarly, selected antigen can be the antigen of identifying in following discussion.In an embodiment of this paper immunogenic composition, described selected antigen is HIV-1 antigen, and such as passing through gag, pol, env, nef, vpr, vpu, vif and tat, one of them is expressed.Other component of preferred described antigen sequence and DNA plasmid is optimized, for example, by selection, be appropriate to object host's codon and by removing arbitrary inhibition sequence, the preparation about antigen be also discussed as follows.
Therefore this immunogenic composition comprises that coding is for expressing a kind of plasmid of single selected antigen host.The method according to this invention, described plasmid compositions also comprises a kind of DNA plasmid, described DNA plasmid contains coding more than the DNA sequence of the described identical selected antigen of a copy.Or described compositions can comprise a kind of plasmid of expressing a plurality of selected antigens.Every kind of antigen is regulated and controled by controlling element or component separately.Or every kind of antigen can be regulated and controled by identical controlling element.In another embodiment, described DNA plasmid compositions can contain a plurality of plasmids, wherein the plasmid-encoded identical or different antigen of each DNA.
In another embodiment, described DNA plasmid immunogenic composition can further comprise, as the component of single DNA plasmid or as the part of the DNA plasmid that contains antigen, the nucleotide sequence of encode required cytokine, lymphokine or other gene adjuvant.The host that nucleotide sequence can obtain above-mentioned suitable adjuvant certainly determines as follows.In an embodiment of institute of the present invention illustration, the required cytokine of using together with DNA plasmid compositions of the present invention is IL-12.
Described DNA plasmid compositions is desirably in pharmaceutically acceptable diluent, excipient or carrier to be used, and for example those are as discussed below.Although described compositions can any selectable route of administration be used, the medication of expectation is comprise described plasmid and use altogether as the bupivacaine intramuscular injection of promoter in one embodiment, is described below.
In an embodiment of the inventive method, wherein said DNA compositions is just to exempt from compositions, and the method is used DNA plasmid at least one times before being included in and using rVSV, and described plasmid comprises coding to induce the sequence of the antigen of required immunne response.In another one embodiment, described DNA just exempts from compositions and also contains the second plasmid, described plasmid-encoded selected cytokine.Also in another one embodiment, following institute illustration, described DNA just exempts from compositions and comprises three kinds of plasmids, a kind of plasmid expression the first antigen, the antigen that the second plasmid expression the second is different and the selected Cytokine adjuvant of the third plasmid expression.As what describe in detail in embodiment, the embodiment that DNA just exempts from compositions comprises the plasmid of three kinds of optimizations, the plasmid (seeing Figure 1A) of the SIV gag p37 gene that (1) coding RNA is optimized; (2) the encode plasmid of two kinds of Rhesus Macacus IL-12 subunit p35 and p40 (is shown in Figure 1B) respectively and the plasmid of (3) coding HIV-1 gp160env gene under the regulation and control of two kinds of promoteres.
When just exempting from compositions, described DNA plasmid compositions was used once or preferably before rVSV strengthens compositions, more than once.When being used as while strengthening compositions, this DNA plasmid compositions is used once or preferably after rVSV just exempts from compositions, more than once.
B.rVSV immunogenic composition
Another kind of immunogenic composition for the inventive method and compositions is vesicular stomatitis virus (VSV) delivery vector reproducible, alive, attenuation.This restructuring VSV comprises the nucleotide sequence of the selected antigen of coding, and described coded sequence is instructed its regulating and controlling sequence of expressing in mammal or vertebrate cells to regulate and control.In the method for the invention, for the antigen of DNA plasmid compositions with identical for the antigen of rVSV immunogenic composition.
VSV is a kind of bovine viral, and it is a member that is called Mononegavirales on taxonomy, comprises 11kb Nonsegmented, strand RNA genome, encode four internal structure albumen and an outside transmembrane protein.From 3 '-5 ' sequentially, the albumen of gene code is called nucleocapsid (N), phosphoprotein (P), stromatin (M), transmembrane glycoprotein (G) and polymerase (L).The VSV that uses technology known in the art separable also " rescue " to live.Referring to, as U.S. Patent number 6,044,886,6,168,943 and 5,789,299; International Publication patent No. WO99/02657; Conzelmann, 1998, Ann.Rev.Genet., 32:123-162; Roberts and Rose, 1998, Virol., 247:1-6; Lawson etc., 1995 Proc.Natl.Acad.Sci., USA 92:4477-4481.In addition, for example, the registration number that the genome sequence of VSV (Indiana) is listed in ncbi database is NC001560.Other VSV sequence, comprises VSV (Chandipura) sequence, can in this data base, obtain, for example, referring to registration number Ay382603, Af128868, V01208, V01207, V01206, M16608, M14715, M14720 and J04350 etc.VSV strain, all can obtain from preservation center such as New Jersey and Indiana etc., for example American type culture collection, Rockville, Maryland (referring to, as registration number VR-1238 and VR-1239.In the file of quoting as proof in full in this description, describe or with reference to other sequence.All documents of quoting as proof in this description are incorporated herein by reference at this.)。
Already shown that VSV genome can hold more than one exogenous gene, at least reached 3,000 bases.These viral genomes are highly stable, can not recombinate, almost rare significant sudden change.In addition,, because copying of they is that Cytoplasm copies, so their genome contains RNA, they can not be incorporated into the genome of infected host cell.Equally, these minus-stranded rna virus have relatively simple transcription regulating nucleotide sequence, can easily handle for inserting effective exogenous gene.Finally, can by change exogenous gene with respect to the position of the initial son of virus transcription to regulate the level of exogenous gene expression.3 '-5 ' gene expression gradient has reflected that the probability that can successfully cross each run into gene within gene termination/gene initial signal when transcribing varial polymerases advances along genomic templates has reduced.Therefore, be positioned at approach 3 ' exogenous gene of end transcription initiation promoter can give full expression to, those be inserted in genome more the exogenous gene of distal position express less.
VSV can copy and reach high-titer in much different cellular type, and great expression virus protein.This not only means that VSV can be used as a kind of delivery vector of effective functional exogenous gene, and approval for the production of the cell line of people's kind biological product in the corresponding accessible level of production of rVSV.
RVSV has and outstanding the crucial former exogenous gene of protective immunity of coding is delivered to the ability of many different cellular types from viral pathogen, cause subsequently real shaping immunogenic protein great expression (Haglund, K., etc., 2000 Virol., 268:112-21; Kahn, J.S. etc., 1999 Virol., 254:81-91; Roberts, A. etc., 1999 J.Virol., 73:3723-32; Rose, N.F. etc., 2000 J.Virol., 74:10903-10 and Schlereth, 2000 J.Virol., the 74:4652-7 such as B.).This immunogen of so expressing can cause the reaction of high lasting virucidin and protectiveness cytotoxic T lymphocyte (CTL) (Roberts, A. etc., 1998 J.Virol., 72:4704-11) simultaneously.
Except the effect of rVSV, the viral gene delivery carrier of this work is safe, because the disease symptoms that wild type VSV can not cause substantially in Healthy People or pathology, (Tesh as the same when a large amount of virus replication even, R.B. etc., 1969 Am.J.Epidemiol., 90:255-61).In addition, the mankind infected by it and be therefore pre-existing in the immunity of VSV be rare.If further attenuation, these rVSV compositionss are applicable to immunocompromised host or and the experimenter of subhealth state.The remarkable advantage that uses in the method VSV carrier is to exist much VSV serotype, and this is due to due to the exchange of VSV viral attachment protein G or modifying.Therefore, carry the different serotypes VSV carrier of identical heterologous antigen can repetitive administration to avoid host immune system to produce the interference of any reaction of the neutralizing antibody for VSV G albumen.
Use the technology of describing before this area can design restructuring VSV, it carries selected antigen and is inserted into the regulating and controlling sequence of any position of VSV under virus transcription promoter regulation.In one embodiment, the heterologous gene of the selected antigen of described coding is inserted between the G and L coding region of VSV.In another one embodiment, heterologous gene can merge the protein loci to G.In another embodiment, heterologous gene merges or is abutted to arbitrary other VSV gene loci.
In another embodiment, gene is in genome diverse location ' reorganization '.Particularly, by N gene ' reorganization ' in genome to diverse location.The recombinant cDNA sequence that clone produces described reorganization comprises modification initial VSV plasmid main chain (pVSV-XN1).Three variants of this initial vector comprise the plasmid that departs from normal gene order (3 '-N-P-M-G-L-5 '), as follows: i) 3 '-P-N-M-G-L-5 '; Ii) 3 '-P-M-N-G-L-5 '; And iii) 3 '-P-M-G-N-L-5 '.The method adopting for generation of the Strategies For The Cloning of these plasmids is described in Ball, and 1999 J.Virol. such as L.A., in 73:4705-12.This technology has been utilized this fact, finds that the gene end/gene initial signal between each coded sequence is almost the same, therefore allows gene rearrangement to build and replaces without importing any nucleotide.Or, some key point sudden changes can be imported to non-coding sequence to produce the restriction enzyme site that contributes to genome rearrangement nearby.
Also in another embodiment of these carriers, the amino acid whose carboxyl terminal coded sequence in 29, G gene cell matter district by the aminoacid of disappearance G gene 5 ' C-terminal by truncate.Or, lack G gene completely.In one embodiment, remove the Cytoplasm district of G gene complete.In another embodiment, remove at least 28, Cytoplasm district aminoacid.In another embodiment, approximately 20 aminoacid in disappearance Cytoplasm district.In another embodiment, approximately 10 or the aminoacid still less in disappearance Cytoplasm district.
It is reported, reorganization genome method and the protein modified method of G all can generating portion growth defect (Flanagan, E.B. wait 2001 J.Virol.75:6107-14; Schnell, M.J. etc., 1998 EMBO J., 17:1289-96).Can estimate that the such modification of VSV can produce more attenuation phenotype or even produce non-replicating VSV, can be used in the various embodiments of the present invention.Referring to, as Johnson, J.E. etc., 1998 Virol., 251:244-52.37; Johnson, J.E., etc., 1997 J.Virol., 71:5060-8.
Similarly, selected antigen can be the following determined antigen of discussing.In an embodiment of immunogenic composition as herein described, selected antigen is a kind of HIV-1 gag and/or env (gp160) gene of clade B virus isolates.Also in one embodiment, described antigen is a kind of HIV-1 pol, nef, vpr, vpu, vif or tat gene.Preferably described antigen sequence is optimized, for example, by selection, be appropriate to object host's codon and/or by removing arbitrary inhibition sequence, the preparation about antigen be also discussed as follows.
In order to overcome the problem that any potential carrier duplicating efficiency reduces in continuous administration, designed similar one group of carrier, carry separately the G gene of different VSV serotypes, can carry out successful booster immunization inoculation.One-level aminoacid sequence from the G albumen of VSV Indiana, New Jersey and Chandipura is substantially divergent, makes the immunity that certain is pre-existing in not get rid of the infection of other kind and copy.Therefore the neutralizing antibody reaction, producing by rVSV (Indiana) should be disturbed copying of rVSV (New Jersey) or rVSV (Chandipura).Can be by using the divergent homologue from VSV Chandipura or VSV New Jersey to replace three kinds of distinct immunocompetence carriers of VSV Indiana G gene formation, to prepare the vehicle group that can allow successfully to carry out continuous immunity inoculation.
Therefore, this rVSV immunogenic composition can comprise a kind of rVSV of the single selected antigen of encoding, for expressing host.The method according to this invention, described rVSV immunogenic composition comprises a kind of rVSV, the nucleotide sequence of the identical selected antigen that it contains the more than one copy of encoding.Or said composition can comprise a kind of rVSV that expresses multiple selected antigen.Every kind of antigen can be regulated and controled by controlling element or component separately.Or every kind of antigen can be regulated and controled by identical controlling element.In another embodiment, described rVSV compositions can comprise multiple rVSV, wherein every kind of rVSV identical or different antigen of encoding.
In another embodiment, described rVSV immunogenic composition can further comprise or use with cytokine, lymphokine or gene adjuvant.Cytokine can albumen form be used, or in above-mentioned plasmid, uses or coded by inserting the insert of cytokine coded sequence in restructuring VSV.For example, described cytokine coded sequence can be inserted into any position of VSV genome and start to express from the initial son of virus transcription.The host that can obtain the suitable adjuvant of nucleotide sequence determines as follows.In an embodiment of institute of the present invention illustration, the required cytokine of using together with rVSV compositions of the present invention is IL-12.
Described rVSV compositions is desirably in pharmaceutically acceptable diluent, excipient or carrier to be used, and for example those are as discussed below.Although described compositions can any selectable route of administration be used, the application process of expectation is nasal injection in one embodiment.
In an embodiment of the inventive method, wherein said rVSV compositions is to strengthen compositions, and the method is included in to be used DNA immunization originality and just exempt to use rVSV at least one times after compositions.This rVSV compositions is expressed the antigen identical with just exempting from compositions.In one embodiment, rVSV compositions comprises the recombinant virus of the additional selected cytokine of coding.In another embodiment, rVSV comprises the sequence of the express cell factor, as IL-12 be present in express as described in the identical rVSV of antigen.In another embodiment, multiple rVSV compositions is used as later stage stiffener.In one embodiment, at least two kinds of rVSV compositionss are used after just exempting from compositions.In another embodiment, at least three kinds of rVSV compositionss are used after just exempting from compositions.
Ideally, as discussed above, each follow-up rVSV compositions has different serotype, but has identical antigen encoding sequence.The arbitrary synthetic serotype that described different serotype is selected from known naturally occurring serotype and provides by handling VSV G albumen.In known method, for the technical description that changes rVSV G albumen in international publication number WO99/32648 and Rose, N.F. etc., 2000 J.Virol., in 74:10903-10.
In another embodiment, every kind of rVSV has different antigen encoding sequences, but has identical VSV G albumen.In another embodiment, every kind of rVSV has different antigen encoding sequences and different VSV G albumen.As describing in detail in embodiment, an embodiment of a series of rVSV reinforcement compositionss comprises the rVSV of two kinds of optimizations, and it contains HIV-1 gp160 env gene, and wherein a kind of rVSV is Indiana serotype, and another kind is Chandipura serotype.
The content according to the present invention, this rVSV immunogenic composition is being used after have a same antigen first exempt from DNA immunization originality compositions, can be used as to strengthen compositions and be applied to host.In arbitrary embodiment of the inventive method, after a rVSV compositions is used, second and arbitrary additional rVSV can be used as stiffener and use.In one embodiment, described additional rVSV stiffener is the phase homologous serotype with same antigen.Or described additional rVSV stiffener has different serotype, continuous administration before using the DNA immunization originality compositions of reinforcement.It is useful already having shown to use at least three kinds of stiffeners.When as reinforcement compositions, described rVSV compositions is continuous administration after just exempting from DNA immunization originality compositions.When conduct is just exempted from compositions, described rVSV compositions is used once or preferably and is used repeatedly.In one embodiment, described additional rVSV stiffener is the phase homologous serotype with same antigen.Or described additional rVSV stiffener has different serotype, continuous administration before using the DNA immunization originality compositions of reinforcement.
The example that can express in vivo the suitable rVSV construct of HIV-1 albumen is specified in following embodiment and public publication, as, Rose etc., 2000 Virol., 268:112-121; Rose etc., 2000 J.Virol., 74:10903-10910; 2001 Cell such as Rose, 106:539-549; Rose etc., 2002 J.Virol., 76:2730-2738; Rose etc., 2002 J.Virol., 76:7506-7517, is incorporated herein by reference in this.Additional rVSV carrier is attenuation further, can be undertaken by the VSV structural gene of truncate VSV G albumen or reorganization as mentioned above progressively.According to content of the present invention, non-replicating VSV also can be used.RVSVs has shown balance desired between attenuation and immunocompetence, estimates to be used for the present invention.
In following embodiment, illustrated several exemplary rVSV construct examples have recombination group:
(1)3′N-P-M-G-HIV?env-L-5′
(2)3′N-P-M-G-HIV?gag-L-5′
(3)3′N-P-M-G-SIV?gag-L-5′
In one embodiment, method of the present invention and immunogenic composition have adopted a kind of in these rVSV constructs.In another embodiment, method of the present invention and immunogenic composition have adopted rVSV-HIV env and rVSV-HIV gag.The latter's immunogenic composition brings out the effective humoral immunoresponse(HI) to the gp160 of real shaping simultaneously, and can kill the CTL (for Env and Gag) of the cell of HIV-1 viral infection.The conformational change relevant with receptors bind also experienced in HIV-1 gpl60 and the combination of CD4/ coreceptor expressed in which.Correct gp160/ acceptor interaction has exposed in the hiding virus being present on gp160 and determiner; its be induction antibody-mediated exempt from infected protective effect necessary (referring to; as LaCasse, R.A. etc., 1999 Science.283:357-62).
C. the antigen using in immunogenic composition of the present invention
In the inventive method and compositions, antigenicity or immunogenic composition can be used for strengthening the immunne response of vertebrate host to selected antigen.Selected antigen, when expressing by plasmid DNA or VSV, can be to be derived from Causative virus, antibacterial, fungus or parasitic albumen, polypeptide, peptide, its fragment or fusions.Or selected antigen can be albumen, polypeptide, peptide, its fragment or the fusions that is derived from cancerous cell or tumor cell.In another embodiment, selected antigen can be albumen, polypeptide, peptide, its fragment or the fusions that is derived from anaphylactogen, makes to disturb the generation of IgE to regulate, the abnormality of anaphylactogen to be replied.In another embodiment, selected antigen can be to be derived from host's (self molecule) with a kind of unexpected mode, quantity or those molecules that position was produced or albumen, polypeptide, peptide, its fragment or the fusions of its fragment, for example those from amylaceous before albumen, to there is the disease of amylaceous deposition characteristics in prevention or treatment vertebrate host.In one embodiment of the invention, selected antigen can be albumen, polypeptide, peptide or the fragment that is derived from HIV-1.
The present invention also relates to for strengthening the method for immunogenic composition ability, described immunogenic composition comprises selected antigen, it is derived from the Causative virus that (1) brings out vertebrate host immunne response, antibacterial, fungus or parasite, or cancer antigen or the tumor associated antigen of (2) cancerous cell or tumor cell, it can bring out therapeutic or preventative antitumaous effect in vertebrate host, or (3) anaphylactogen, make to disturb the generation of IgE to regulate, the abnormality of anaphylactogen to be replied, or (4) host (self molecule) is in a kind of unexpected mode, those molecules that quantity or position produce or its part, to reduce above-mentioned unexpected effect.
In another embodiment, the required virus immunity originality compositions of using the present invention just to exempt from/strengthen therapy comprises and relates to those that prevent and/or treat the disease that caused by following virus, and described virus is: be not limited to HIV (human immunodeficiency virus), simian immunodeficiency virus, respiratory syncytial virus, 1-3 type parainfluenza virus, influenza virus, herpes simplex virus, human cytomegalic inclusion disease virus, hepatitis A virus, hepatitis B virus, hepatitis C virus, human papillomavirus, poliovirus, rotavirus, calicivirus, Measles virus, mumps virus, german measles virus, adenovirus, rabies virus, canine distemper virus, rinder pest morbillivirus, human metapneumovirus, avian pneumovirus (being called in the past Turkey's rhinotracheitis virus), Hendra virus, Nipah virus, coronavirus, parvovirus, infectious bovine rhinotrachetis virus, feline leukaemia virus, feline infectious peritonitis virus, bird gomvoro disease virus, new castle disease virus, horse Rec (family name) virus, pig breathes and reproduction syndrome virus, epizootic cellulitis virus and various encephalitis.
In another embodiment, the required bacterial immune originality compositions of using the present invention just to exempt from/strengthen therapy comprises and relates to those that prevent and/or treat the disease that caused by following antibacterial, described antibacterial is: be not limited to influenza (bloodthirsty) bacillus (Haemophilus influenzae) (typing and can not typing), Haemophilus somnus (Haemophilus somnus), Moraxella catarrhalis (Moraxella catarrhalis), streptococcus pneumoniae (Streptococcus pneumoniae), streptococcus pyogenes (Streptococcus pyogenes), streptococcus agalactiae (Streptococcus agalactiae), streptococcus faecalis (Streptococcus faecalis), helicobacter pylori (Helicobacter pylori), meningitis Neisser (family name) bacterium (Neisseriameningitidis), gonorrhea Neisser (family name) bacterium (Neisseria gonorrhoeae), chlamydia trachomatis (Chlamydia trachomatis), Chlamydia pneumoniae (Chlamydia pneumoniae), chlamydia psittaci (Chlamydia psittaci), bordetella pertussis (Bordetella pertussis), Alloiococcus otiditis, Salmonella typhi (Salmonella typhi), Salmonella (Salmonella typhiumrium), Salmonella choleraesuls (Salmonella choleraesuis), escherichia coli (Escherichia coli), Shiga bacillus (Shigella), vibrio cholera (Vibrio cholerae), diphtheria corynebacterium (Corynebacterium diphtheriae), Mycobacterium tuberculosis (Mycobacterium tuberculosis), Mycobacterium avium (Mycobacterium avium)-Mycobacterium intracellulare complex (Mycobacterium intracellulare complex), proteus mirabilis (Proteus mirabilis), proteus vulgaris (Proteus vulgaris), staphylococcus aureus (Staphylococcus aureus), staphylococcus epidermidis (Staphylococcus epidermidis), clostridium tetani (Clostridium tetani), leptospira interrogans (Leptospira interrogans), borrelia burgdorferi (Borrelia burgdorferi), hemolytic Pasteur (family name) bacterium (Pasteurella haemolytica), hemorrhage septic Pasteur (family name) bacterium (Pasteurella multocida), Actinobacillus pleuropneumoniae (Actinobacillus pleuropneumoniae) and Mycoplasma gallisepticum (Mycoplasma gallisepticum).
In another embodiment, the required antifungal that uses the present invention just to exempt from/the strengthen therapy former immunogenic composition that causes a disease comprises relating to and prevents and/or treats by cause a disease those of former caused disease of following fungus, described fungus is caused a disease: be not limited to Eurotium (Aspergillis), Blastomyces (Blastomyces), Candida (Candida), Coccidioides (Coccidiodes), Cryptococcus (Cryptococcus) and Histoplasma (Histoplasma).
In another embodiment, the required antiparasitic immunity originality compositions of using the present invention just to exempt from/strengthen therapy comprises and relates to those that prevent and/or treat the disease that caused by following parasite, described parasite is: be not limited to, leishmania major (Leishmania major), ascarid (Ascaris), Trichocephalus (Trichuris), Giardia (Giardia), Schistosoma (Schistosoma), Cryptosporidium (Cryptosporidium), Trichomonas (Trichomonas), toxoplasma gondii (Toxoplasma gondii) and Pneumocystis carinii (Pneumocystis carinii).
In another embodiment, use the present invention just to exempt from/strengthen the required immunogenic composition of therapy and comprise those that use cancer antigen or tumor associated antigen, described compositions is brought out preventative or therapeutic antitumaous effect in vertebrates, described antigen comprises, be not limited to prostate specific antigen, carcinoembryonic antigen, MUC-1, Her2, CA-125 and MAGE-3.
The nucleotide of above-mentioned listed known antigens and protein sequence can be obtained by the data base such as NCBI by the public at an easy rate, also can obtain from other source, as American type culture collection or university.
In vertebrate host, for regulating the required immunogenic composition that anaphylactogen is replied, it has used of the present inventionly just exempts from/strengthens therapy, comprises that those contain anaphylactogen or its fragment.The example of above-mentioned anaphylactogen is described in U.S. Patent number 5,830,877 and International Patent Publication No. WO 99/51259 in, at this, be incorporated herein by reference.Above-mentioned anaphylactogen comprises, is not limited to pollen, insecticide venom, animal hair, fungal spore and medicine (as penicillin).These immunogenic compositions disturb the generation of IGE antibody, are the known reasons of allergenic response.
In vertebrate host, for regulating the required immunogenic composition to self molecular reaction, it has used the therapy of just exempting from/strengthen of the present invention, comprises that those contain self molecule or its fragment.The example of above-mentioned self molecule comprise G17 molecule related in the related beta chain insulin of diabetes, gastroesophageal reflux disease and in disease the lower antigen that regulates autoimmune reaction, described disease is as multiple sclerosis, lupus and rheumatoid arthritis.Also comprise beta amyloid peptide (also referred to as A β peptide), it is 39-43 amino acid fragment in albumen (APP) before amylaceous, processes APP produce by β or gamma secretase.A β 1-42 peptide has following sequence SEQ ID NO:1:Asp Ala Glu Phe Arg His Asp Ser Gly Tyr Glu Val His His Gln Lys Leu Val Phe Phe Ala Glu Asp Val Gly Ser Asn Lys Gly Ala Ile Ile Gly Leu Met Val Gly Gly Val Val Ile Ala.
At choice and operation antigen sequence, be used for designing DNA plasmid of the present invention and rVSV construct, also need to change the codon that position that selected antigen encoding gene sequence and DNA plasmid will insert adopts, and/or remove inhibition sequence wherein, can be by using as detailed below technology except disinthibiting sequence: U.S. Patent number 5,965,726,5,972,596,6,174,666,6,291,664 and 6,414,132 and International Patent Publication No. WO 01/46408, in this, be incorporated herein by reference.Describe simply, this technology is included in the determined inhibition/unstable sequence of suddenling change in selected gene, preferably carries out multiple point sudden change.
In a specific embodiments of following institute illustration, one or more optimizations HIV-1 antigen of encoding for immunogenicity plasmid of the present invention and the expectation of rVSV compositions, as gag,, pol and nef antigen, or its immunogenic fragments or fusions.Chimeric monkey-human immunodeficiency virus (SHIV) gag and env gene (89.6P) can be used for preparing rVSVs, so that the disease of protecting it to avoid viral infection and alleviating viral load and can confirm in macaque disease model.Following example has proved use SHIV analog, i.e. SIV gag and HIV 89.6Penv.
D. the promoter of using in immunogenicity DNA plasmid or rVSV construct
Suitable promoter for any component of the present invention, can easily be selected among formation type promoter, inducible promoter, tissue-specific promoter etc.The example of nucleic acid molecules formation type that adopt and that the have non-specific activity promoter of code book invention antigen comprises, be not limited to, reverse transcription Rous sarcoma virus (RSV) promoter, reverse transcription LTR promoter (optionally with RSV enhancer), cytomegalovirus (CMV) promoter (optionally with cmv enhancer) (referring to, as Boshart etc., 1985 Cell, 41:521-530), SV40 promoter, dihydrofolate reductase promoter, beta-actin promoter, phosphoglycerokinase (PGK) promoter and EF1 α promoter (Invitrogen).The inducible promoter regulating and controlling for exogenous supplementary compound comprises, be not limited to, Arabinose promoter, zinc induction sheep metallothionein (MT) promoter, mouse mammary adenoma virus (MMTV) promoter of dexamethasone (Dex) induction, T7 polymerase promoter system (WO98/10088), ecdyson insecticide promoter (No etc., 1996 Proc.Natl.Acad.Sci.USA, 93:3346-3351), tetracycline repressible system (Gossen etc., 1992 Proc.Natl.Acad.Sci.USA, 89:5547-5551), tetracycline-inducible (Gossen etc., 1995 Science, 268:1766-1769, also referring to Harvey etc., 1998 Curr.Opira.Chern.Biol., 2:512-518), RU486 inducible system (Wang etc., 1997 Nat.Biotech., 15:239-243 and Wang etc., 1997 Gene Ther., 4:432-441) and rapamycin inducible system (Magari etc., 1997 J.Cliva.Invest., 100:2865-2872).
The inducible promoter that can be used for other type is herein that those are that specificity physiological status regulates and controls, as, temperature or acute stage or only in replicating cell.Useful tissue-specific promoter comprises the gene promoter that is derived from coding skeleton β actin, myosin light chain 2A, dystrophin, muscle creatine kinase, and the more highly active synthetic flesh type promoter of more naturally occurring promoter tool (referring to Li etc., 1999 Nat.Biotech., 17:241-245).The example of known tissue-specific promoter has: for liver (albumin, the .1997 J.Virol. such as Miyatake, 71:5124-32; Hepatitis b virus core promoter, Sandig etc., 1996 Gene Ther., 3:1002-9; Alpha fetal protein (AFP), Arbuthnot etc., 1996 Hum.Gene Ther., 7:1503-14); For bone (osteocalcin, Stein etc., 1997 Mol.Biol.Rep., 24:185-96; Bone sialoprotein, Chen etc., 1996 J.Bone Miner.Res., 11:654-64); For lymphocyte (CD2, Hansal etc., 1988 J.Immunol., 161:1063-8; Heavy chain immunoglobulin; φt cell receptor α chain); For neuron (neuron-specificity enolization alcohol (NSE) promoter, Andersen etc., 1993 Cell.Mol.Neurobiol., 13:503-15; Neurofilament light gene, Piccioli etc., 1991 Proc.Natl.Acad.Sci.USA, 88:5611-5; Neuronal specificity ngf gene, Piccioli etc., 1995Neuron, 15:373-84) etc.Referring to, the known promoter of listing in addition as International Patent Publication No. WO 00/55335 can be used for herein.
E. immunogenic composition of the present invention is used carrier, excipient, adjuvant and formula
No matter can be used for the immunogenic composition in the present invention, be DNA plasmid or rVSV compositions, also comprises acceptable diluent or pharmaceutically acceptable carrier in immunology, for example sterilized water or sterile isotonic saline.Described antigen composition also can mix in a usual manner with above-mentioned diluent or carrier.Term used herein " pharmaceutically acceptable carrier " be intended to comprise any and all solvents, disperse medium, encrusting substance, antibacterial agent and antifungal, etc. ooze with absorption delay agent etc., be applicable to the administration that people or other vertebrate host are carried out.Described suitable carrier is apparent to those skilled in the art, depends primarily on route of administration.
The annexing ingredient that still can exist in immunogenic composition of the present invention has: adjuvant, antiseptic, surfactant and chemical stabilizer, suspending agent or dispersant.Typically, stabilizing agent, adjuvant and antiseptic optimization are determined to effective optimum formula in target human or animal.
1. adjuvant
When jointly using with immunogen or antigen, adjuvant is a kind of material that can strengthen immunne response.Many cytokines or lymphokine tool immunoregulatory activity had already been confirmed, therefore can be used as adjuvant, comprise, but be not limited to, interleukin-11-α, 1-β, 2, 4, 5, 6, 7, 8, 10, 12 (referring to, as U.S. Patent number 5, 723, 127), 13, 14, 15, 16, 17 and 18 (and saltant types), interferon-ALPHA, β and γ, granulocyte macrophage colony stimulating factor (referring to, as U.S. Patent number 5, 078, 996 and ATCC registration number 39900), M-CSF, granulocyte colony-stimulating factor, GSF, and tumor necrosis factor α and β.Also can be used for other adjuvant of the present invention and comprise chemotactic factor, include but not limited to, MCP-1, MIP-1 α, MIP-1 β and RANTES.Adhesion molecule, as selected albumen, for example, L-selects albumen, CD62P and CD62L also to can be used as adjuvant.Other also available adjuvant comprise, be not limited to, mucin quasi-molecule, as CD34, GlyCAM-1 and MadCAM-1, integrin family member, LFA-1 for example, VLA-1, Mac-1 and p150.95, immunoglobulin superfamily member, as PECAM, ICAMs, ICAM-1 for example, ICAM-2 and ICAM-3, CD2 and LFA-3, costimulatory molecules, as CD40 and CD40L, somatomedin, comprise angiogenesis factor, nerve growth factor, fibroblast growth factor, epithelial cell growth factor, B7.2, PDGF, BL-1 and VEGF, acceptor molecule, comprise Fas, TNF receptor, Flt, Apo-1, p55, WSL-1, DR3, TRAMP, Apo-3, AIR, LARD, NGRF, DR4, DR5, KILLER, TRAIL-R2, TRICK2 and DR6.Other adjuvant molecules also comprises caspase (ICE).Also referring to International Patent Publication No. WO 98/17799 and WO99/43839, in this, be incorporated herein by reference.
For strengthening the suitable adjuvant of immunne response, comprise, be not limited to MPL
tM(3-O-deacylated tRNA monophosphoryl lipid A; Corixa, Hamilton, MT), it is described in U.S. Patent number 4,912, in 094, in this, is incorporated herein by reference.What be also suitably used as adjuvant is synthetic lipid A analogue or aminoalkyl glycosamine phosphate cpd (AGP), and or derivatives thereof or analog, can obtain (the Hamilton from Corixa, MT), it is described in U.S. Patent number 6,113, in 918, in this, be incorporated herein by reference.Above-mentioned AGP is 2-[(R)-3-is that myristoyl oxygen base is that myristoyl is amino] ethyl 2-deoxidation-4-O-phosphono-3-O-[(R)-3-myristoyl oxygen base myristoyl]-2-[(R)-3-myristoyl oxygen base myristoyl-amino]-b-D-glucopyranoside, also referred to as 529 (being called in the past RC529).This 529 adjuvant is with aqueous solution form or stable Emulsion form preparation.
Other adjuvant also comprises mineral oil and aqueous emulsion, aluminum salt (Alumen), as aluminum chloride, aluminum phosphate etc., Amphigen, Avridine, L121/ Squalene, D-lactide-polyactide/glucosides, polyether polyol (pluronic polyols), muramyldipeptide, the Bordetella of deactivation (Bordetella), saponins, as Stimulon
tMqS-21 (Antigenics, Framingham, MA.), be described in U.S. Patent number 5,057, in 540, in this, be incorporated herein by reference, and wherein produce granule as ISCOMS (immunostimulating complex), Mycobacterium tuberculosis (Mycobacterium tuberculosis), bacteria lipopolysaccharide, synthetic polynucleotide, as the oligonucleotide that contains CpG motif (U.S. Patent number 6,207,646, in this, be incorporated herein by reference), pertussis toxin, PT (PT), or E.coli LT (LT), particularly LT-K63, LT-R72, PT-K9/G129; Referring to, as International Patent Publication No. WO 93/13302 and WO92/19265, in this, be incorporated herein by reference.
Also can be used as adjuvant: cholera toxin and mutant thereof, be included in (wherein the glycine in the 29th, aminoacid is replaced by other aminoacid (except aspartic acid), is preferably histidine) described in International Patent Publication No. WO 00/18434.Similar CT toxin or mutant, be described in that in International Patent Publication No. WO 02/098368, (wherein the isoleucine in the 16th, aminoacid is replaced by other aminoacid, replace separately or at the serine of the 68th, aminoacid, by other aminoacid, replaced simultaneously, and/or wherein at the valine of the 72nd, aminoacid, by other aminoacid, replaced).Other CT toxin is described in that in International Patent Publication No. WO 02/098369, (wherein the arginine in the 25th, aminoacid is replaced by other aminoacid, and/or insert an aminoacid in the 49th, aminoacid, and/or at aminoacid the 35th and two aminoacid of 36 insertions).
In an embodiment of following institute illustration, required adjuvant is IL-12, and it is expressed from plasmid.Referring to, as U.S. Patent number 5,457,038,5,648,467,5,723,127 and 6,168,923, in this, be incorporated herein by reference.This IL-12 expression plasmid is mixed to the first compositions of exempting from of the embodiment that comprises immunogenicity DNA plasmid.Yet, should be noted that and just exempting from and strengthening between compositions, this plasmid can be separately or be applied to mammal or vertebrate host together with rVSV compositions (or expressing by rVSV).In one embodiment, cytokine can be used as protein and uses.In a preferred embodiment, cytokine can be used as the nucleic acid compositions of the DNA sequence that comprises the Codocyte factor and uses, and described coding is instructed its regulating and controlling sequence of expressing in mammalian cell to regulate and control.In a useful embodiment, Cytokine Plasmids is used together with DNA compositions.In another embodiment, the first administer cytokines between thing and reinforcement compositions of exempting from can used.Also, in another step, cytokine is used in reinforcement step.In another embodiment, cytokine can with just exempt from compositions and use together with strengthening compositions.
At the beginning of wherein said, exempting from compositions is DNA plasmid compositions, and as the following example, plasmid compositions can comprise the DNA sequence of the Codocyte factor, the regulation and control of the modulated regulating and controlling sequence that it expresses in mammal of described coded sequence.In certain embodiments, the existing DNA plasmid of cytokine coded sequence is identical with the existing DNA plasmid of antigen encoding sequence.In another embodiment, the existing DNA plasmid of cytokine coded sequence is different from the DNA plasmid of coding for antigens.
2. promoter or adjuvant
Except above-mentioned carrier, immunogenic composition comprises required polynucleotide molecule, it contains optional polynucleotide promoter or adjuvant, as local anesthetic, peptide, lipid, comprises cation lipid, liposome or fat particles, polycation, as poly-D-lysine, the three-dimensional polycation of branch, as dendritic (dendrimer), carbohydrate, cationic amphiphilic thing, detergent, benzyl ammonium surfactant, or other contributes to polynucleotide to be transferred to the compound of cell.Above-mentioned promoter comprises local anesthetic bupivacaine or tetracaine (referring to U.S. Patent number 5,593,972,5,817,637,5,380,876,5,981,505 and 6,383,512 and International Patent Publication No. WO 98/17799, in this, be incorporated herein by reference).Above-mentioned other non-proprietary example for promoter of the present invention or adjuvant is described in U.S. Patent number 5,703,055; 5,739,118; 5,837,533; With International Patent Publication No. WO 96/10038, be disclosed on April 4th, 1996; And International Patent Publication No. WO 94/16727, be disclosed on August 8th, 1994, in this, be incorporated herein by reference.
Most preferably, described local anesthetic exists with the amount that can form one or more complex with described nucleic acid molecules.When described local anesthetic mixes with nucleic acid molecules of the present invention or plasmid, can form little complex or the granule of the multiple DNA of being wrapped in, and be homogeneous.Therefore, in an embodiment of immunogenic composition of the present invention, by least one plasmid of local anesthetic and the present invention is mixed to form described complex.The formed any single complex of this mixing can comprise multiple different plasmids combination.Or in another embodiment of the present composition, local anesthetic can be pre-mixed respectively with every kind of plasmid.If all plasmids will be used with single bolus administering mode, then different mixture and single compositions are combined to guarantee to exist the plasmid of required ratio in single immunogenic composition.Or, described local anesthetic can with every kind of plasmid independently mix and individual application to obtain required ratio.
Wherein, hereinafter, term " complex (complex) " or " one or more complex " or " complex (complexes) ", for defining the embodiment of immunogenic composition, should be understood to this term and comprise one or more complex.Every kind of complex contains plasmid mixture, or the separated compound that forms complex.Every kind of complex can only contain plasmid or the complex of one type, or the mixture of plasmid or complex, and wherein every kind of complex contains polycistron DNA.Preferably, described complex diameter approximately 50 to about 150nm.When local anesthetic is used as promoter, preferred bupivacaine, preferred amount is about approximately 0.1 percentage by weight of polynucleotide composition total weight to approximately 1.0 percentage by weights.Referring to, International Patent Publication No. WO 99/21591, is incorporated herein by reference in this, and its instruction can be mixed benzyl ammonium surfactant as adjuvant, preferably with the amount of about 0.001-0.03 % by weight, use.According to content of the present invention, the amount of local anesthetic exists with the certain ratio to described nucleic acid molecules, is the local anesthetic of the 0.01-2.5%w/v amount of 1-10 μ g/ml nucleic acid.Another above-mentioned scope is the local anesthetic of the 0.05-1.25%w/v amount of 100 μ g/ml-1mg/ml nucleic acid.
3. other additive of immunogenic composition
In immunogenic composition of the present invention, other additive be can comprise, antiseptic, stable elements comprised, surfactant etc.
Applicable antiseptic example comprises chlorobutanol, potassium sorbate, sorbic acid, sulfur dioxide, propyl gallate, metagin, ethyl vanillin, glycerol, phenol and parachlorophenol.
Spendable applicable stable elements comprises, for example, and casamino acid, sucrose, gelatin, phenol red, N-Z amine, potassium dihydrogen phosphate (monopotassium diphosphate), lactose, lactalbumin hydrolysate and milk powder.
Applicable surfactant comprises, is not limited to freund 's incomplete adjuvant, quinone analog, cetylamine, 18-amine., octadecane amino acid esters, LYSOLECITHIN SUNLECITHIN A, DDA, methoxyl group cetyl glycerol and polyether polyol; Polyamines, for example pyrans, dextran sulfate, poly-IC (poly IC), carbopol; Peptide, for example muramyl peptide and dipeptides, dimethylglycine, tuftsin; Oil emulsion; And mineral coagulant, such as aluminum phosphate etc. and immunostimulating complex (ISCOMS).Plasmid and rVSVs also can mix liposome and be used as immunogenic composition.Described immunogenic composition also can comprise the additive that other is applicable to selected compositions administering mode.Compositions of the present invention also can comprise the polynucleotide of lyophilizing, can make powder, liquid or suspension dosage form together with other pharmaceutically acceptable excipient.Referring to, as Remington:The Science and Practice of Pharmacy, the 2nd volume, the 19th edition (1995), as the 95th chapter Aerosols and International Patent Publication No. WO 99/45966, it is instructed in this and is incorporated herein by reference.
These immunogenic compositions can contain the applicable additive of using by any conventional route of administration.In certain preferred aspects, immunogenic composition of the present invention as follows form for the preparation of using people, for example, liquid, powder, aerosol, tablet, capsule, enteric-coated tablet or capsule, or suppository.Therefore, described immunogenic composition also can include, but not limited to suspension, solution, the Emulsion in oiliness or aqueous solvent, paste and implantable sustained release or biodegradable form.An embodiment for parenteral formula, active component provides with solid form (being powder or granule), before parenteral administration is rebuild compositions, for the carrier with suitable (as sterilized water for injection), rebuilds.Other formula that can be used for parenteral comprises that those are with microcrystalline form, the liposome prepared product or comprise active component as biodegradable polymer system components of usining.The hydrophobic substance that can comprise pharmaceutically acceptable polymer for the compositions of sustained release or implantation, example emulsion, ion exchange resin, slightly soluble polymer or slightly soluble salt.
Immunogenic composition of the present invention is not limited in above-mentioned conventional select, and physiology acceptable carrier, adjuvant or other can be used for the composition of pharmaceutical preparations.The prepared product of these pharmaceutically acceptable compositionss, from said components, has suitable pH isotonicity, stability and other normal attribute well known in the art.
F. the dosage of immunogenic composition of the present invention and approach
In general, for immunogenic composition component of the present invention, suitable " effective dose " or dosage select to depend on the characteristic of the antigen that uses in immunogenic composition, and experimenter's physical qualification, particularly comprise by immune experimenter's routine health, age and body weight.Medication and approach and in immunogenic composition the existence of annexing ingredient also affect the amount of dosage and plasmid and rVSV compositions.Above-mentioned selection and to the rise of effective dose or to lower be also general knowledge known in this field.Induce immune response, preferred protective response, or in patient, produce external source effect and can change along with the variation of these factors without the obvious amount of the needed plasmid of side effect and rVSV.Suitable dosage is easy to be determined by those skilled in the art.
Using people or inhuman antigen vertebrate of the present invention or immunogenic composition can use by being permitted number of ways, include but are not limited to, intranasal, mouth, vagina, rectum, parenteral, Intradermal, percutaneous (referring to, as International Patent Publication No. WO 98/20734, in this, be incorporated herein by reference), intramuscular, intraperitoneal, subcutaneous, intravenous and intra-arterial.Used immunogenic composition characteristic is depended in the selection of suitable pathways, and attending doctor is to patient age, body weight, sex and conventional healthy assessment, and existing antigen and similar factor in immunogenic composition.
In the following embodiment providing, described immunogenicity DNA compositions is used by intramuscular injection (i.m.).In one embodiment, what need is intranasal administration rVSV compositions, but not intramuscular injection.Yet the selection of dosage and route of administration is not limited by the present invention.
Similarly, the selection of immunogenic composition order of administration and the interval between each administration can be determined the clear reaction of the method application according to health status and host by attending doctor or those skilled in the art.Above-mentioned optimization is common practise to those skilled in the art.
G. kit components
Also, in another one embodiment, the invention provides a kind of medicinal reagent box, for preparing to use, produce immunity, preventative or therapeutic treatment, described therapy can be used for treating arbitrary above-mentioned disease or disease, required to antigen immune response.This test kit is designed in mammal or vertebrate subject, induce in the method for high-level antigenic specificity reaction.Described test kit comprises at least one immunogenic composition, and described compositions comprises the DNA plasmid of the DNA sequence of coding for antigens, and described coded sequence is instructed its regulating and controlling sequence of expressing in mammal or vertebrate cells to regulate and control.Preferably, in test kit, provide the repeatedly pre-packing dosage for the DNA immunization originality compositions of multiple dosing.This test kit also comprises at least one immunogenic composition, described compositions contains reproducible restructuring vesicular stomatitis virus (rVSV), the nucleotide sequence that it comprises the same antigen of encoding, described coded sequence is instructed its regulating and controlling sequence of expressing in mammal or vertebrate cells to regulate and control.Preferably, in test kit, provide the repeatedly pre-packing dosage for the rVSV immunogenic composition of multiple dosing.
Wherein above-mentioned immunogenic composition does not comprise DNA plasmid and/or the rVSV expressing such as the cytokine of IL-12, and this test kit also optionally comprises independent cell factor composition or for the repeatedly pre-packing dosage of the cell factor composition of multiple dosing.These cell factor compositions are nucleic acid compositions normally, the DNA sequence that it comprises the selected cytokine of encoding, and described coded sequence is instructed its regulating and controlling sequence of expressing in mammal or vertebrate cells to regulate and control.
Described test kit also comprises described herein just exempt from/adds strong method middle finger and lead the description that immunogenic composition is used.This test kit also comprises description for implementing some analysis, variety carrier, excipient, diluent, adjuvant and above-mentioned etc., and for the device of applying said compositions, such as syringe, sprayer unit etc.Other component can comprise description, applicator stick or container and other compositions of disposable glove, sterilization.
In order to understand better the present invention, provide the following example.Described embodiment object is only illustration and does not limit scope of the present invention.All documents, public publication and the patent in the following example, quoted are incorporated herein by reference in this.
As the following example, prove, in immune experimenter, just exempt from/strengthened scheme of the present invention has wonderful synergism to the induction of antigen specific cell and humoral immunoresponse(HI).In fact, when just exempt from by the present invention/strengthened scheme is induced these whens reaction, repeatedly just exempt from compositions or use the result of repeatedly strengthening compositions and compare with only using, the present composition is very obvious to the synergism of replying.Use the combination of using again rVSV Booster after the DNA plasmid of expressing required antigen, in immune experimenter, producing the T cells with antigenic specificity increasing, be equivalent to surpass arbitrary additive reaction.Similarly, in using the immunization scheme of DNA plasmid and rVSV immunogenic composition, the T cell confirming in the humoral response of required antigen increases unexpected height.Referring to, as following table 1 and Figure 4 and 5.
The preparation of embodiment 1:DNA plasmid
This embodiment has described the exemplary plasmid for one embodiment of the invention, as described in embodiment 2 and 3.These plasmids are not limit by the present invention, but have already been optimized in following test.Use the following DNA immunization originality of standard recombinant dna art designs compositions.The DNA backbone carrier of expressing HIV or SIV gag gene has been used HCMV promoter, BGHpoly A terminator sequence, ColE1 antibacterial origin of replication (ori) and the kalamycin resistance gene for selecting.
A.SIV gag p37 DNA plasmid
Plasmid WLV102 is a kind of bacterial plasmid, it is expressed the required selected antigen SIV gag p37. plasmid WLV102 (4383bp) of immunne response and by the RNA that is derived from SIV, is optimized gag gene (the p37) (Qiu etc. of truncate, 1999 J.Virol., 73:9145-9152) inserting institute in DNA plasmid expression vector WLV001 forms.Gag gene suppresses sequence to optimize RNA by inactivation, therefore allows not dependent form expression of the high-caliber Rev of gag gene, has used the technology discussing in detail in following patent: United States Patent (USP) 5,965,726,5,972,596,6,174,666,6,291,664 and 6,414,132 and International Patent Publication No. WO 01/46408, in this, be incorporated herein by reference.
Described WLV102 plasmid main chain is comprised of three hereditary uniies.The first component is eukaryotic gene expression unit, it contains from HCMV immediate early promoter/enhancer (.1985 such as Boshart, above-mentioned) and the genetic elements of BGH polyadenylation signal (Goodwin EC and Rottman FM, 1982 J.Biol.Claefn.267:16330-16334).This gag gene clone is between SalI and EcoRI site.Second component is chimeric kalamycin resistance (km
r) gene, adenosine phosphate 4 '-nucleotidyl transferase 1a type (referring to U.S. Patent number 5,851,804).This gene had been designed to give limited amount aminoglycosides resistance already, until it can select the antibacterial that comprises this plasmid.The 3rd component is ColE1 antibacterial origin of replication, and for plasmid, the propagation when the bacterial fermentation is essential for it.This plasmid as shown in Figure 1A.
B. the Codocyte factor-rIL-12DNA plasmid
Rhesus Macacus IL-12WLV104 plasmid is the sub-construct of a kind of double starting, the Rhesus Macacus IL-12 of its expressing heterologous dimeric forms.Plasmid WLV103 is the sub-expression plasmid of a kind of double starting, by two genomic constitutions of encoding human IL12 albumen p35 and 40.This plasmid has 6259 nucleotide altogether.Each cistron in WLV103 contains in two interleukin 12 subunits, p35 or p40, and it is regulated and controled by independently controlling element.P35 subunit is subject to HCMV promoter/enhancer and SV40 polyadenylation signal to regulate and control (being cloned between SalI and MluI site).P40 subunit is subject to SCMV promoter and BGH polyadenylation signal to regulate and control (being cloned in XhoI site).
This plasmid main chain is comprised of several components.The first component is eukaryotic gene expression unit, and it contains and is derived from HCMV immediate early promoter/enhancer and SV40 polyadenylation signal (Fitzgerald M and Shenk T., 1981 Cell, 24:251-260) genetic elements.Second component is eukaryotic gene expression unit, with SCMV promoter (Jeang etc., 1987 J.Virol., 61:1559-1570) and BGH polyadenylation signal, forms.The 3rd component is chimeric kalamycin resistance (km
r) gene, adenosine phosphate 4 '-nucleotidyl transferase 1a type.The 4th component is ColE1 antibacterial origin of replication, and for plasmid, the propagation in antibacterial is essential for it.
By restriction enzyme digestion, digest and analyze plasmid vector SIV gag/HIV gag and the IL-12 DNA producing.Plasmid transient transfection is entered to human rhabdomyosarcoma cells, in the DMEM culture medium of adding 10% calf serum (FCS) and antibiotics, cultivate.Then use HIV gag monoclonal antibody specific (ABI) and SIV gag Rhesus Macacus polyclonal serum (being derived from the animal that SIV infects) to carry out Western blotting and analyze the virus protein that these cells are suitably expressed.In supernatant, with the ELISA test kit (R & D Systems) that detects p70 albumen, IL-12 detected.
The described plasmid vector that increases in the DH10B cell transforming is cultivated in the LB culture medium of adding kanamycin, according to manufacturer specification, uses Qiagen test kit purification.Then on agarose gel, carry out electrophoresis, according to known standard analysis DNA.
Every kind of plasmid in 0.25% bupivacaine preparation for following test, concentration is 2.5mg/mL, cumulative volume is 4.0cc.
Embodiment 2: the preparation of restructuring VSV carrier
A.VSV Genomic cDNA clone
Genetic background for the operation of VSV genome cDNA is pVSV-XN1 (Schnell, M.J. wait 1996 J.Virol., 70:2318-23).This clone comprises a kind of VSV Indiana strain (VSV
i) the modification type of cDNA sequence.Described modification comprises adds two kinds of unique restriction enzyme enzyme recognition sites (XhoI and NheI), and adds the copy of VSV gene initial sum VSV gene termination signal.When such as HIV-1
89.6pwhen the exogenous gene of env gp160 or SIV gag p55 or HIV-1gag inserts between XhoI and NheI site suitably, its present position is suitable for expressing under VSV transcriptional control signals-modulating.VSV cDNA sequence is by the DNA sequence side joint of cis acting, and it is necessary that described sequence is that live virus copies rescue.Transcribing of the primary transcript of the whole viral cDNA of T7 rna polymerase promoter guidance.After T7 RNA polymerase is ended to transcribe, ribozyme is sheared described primary transcript to form rna gene group end.
At first, by insert gag gene (HIV clade B) between G and L gene, modify rVSV
igenomic clone (pVSV-XN1), has produced plasmid prVSVi-gag.Similarly, the separated rVSV that preparation contains HIV env gene (genomic clone prVSVi-env)
icDNA clone.Coding region sequence by amplification from the plasmid template that contains HIV HXBc2 gag gene or HIV 6101 strain env genes, inserts pVSV-XN1 by the gag of preparation and env cDNA sequence.Primer for pcr amplification contains the restriction enzyme site that is suitable for carrying out next step clone; 5 ' primer contains XhoI site, and 3 ' primer contains NheI site.The coding region sequence increasing is respectively inserted in pVSV-XN1 to produce the clone who contains gag and the clone who contains env.
Env gene in described insertion pVSV-XN1 is the modification type of coding HIV Env/VSV G fusion rotein.If use splicing PCR (overlap PCR) method that Env Cytoplasm afterbody is replaced with shorter VSV G albuminous cell matter afterbody, after infecting with rVSV-HIV Env carrier, shown the Env that cell surface strengthens express (referring to, as Johnson, Deng, 1998 is above-mentioned; Johnson etc., 1997, above-mentioned).According to disclosed technology, by being Chandipura or New Jersey serotype by Indiana G gene alteration, changed the carrier main chain of these two kinds of plasmids.By utilizing in M gene unique MluI site and transforming the exchange that XhoI site in VSVcDNA clone completes G gene.
Pcr amplification, from the VSV G gene coded sequence of Chandipura or New Jersey strain, is used 5 ' primer extend through MluI site in M gene.3 ' primer used contains with the sequence of G gene 3 ' end homology and corresponding to the additional end sequence in gene-termination/gene-initial signal and XhoI site.At it, already used MluI and XhoI from plasmid main chain enzyme action, the G gene coded sequence that these primers increase can be used for replacing initial Indiana G gene.
B. save the rVSV from cDNA clone
After being imported into cell, genome cDNA plasmid or transcribe from the RNA of genome cDNA plasmid and be not sufficient to start virus replication circulation.Virus genome RNA himself is not the effective template that can be used for translation or copy.Therefore, recombinant virus must be saved from various VSV genome cDNA constructs.Rescue method well known in the art is likely for saving the virus from clone's DNA.Successfully virus rescue needs to exist in cell the VSV N albumen of VSV geneome RNA and coated virus genome RNA, and P and L albumen, and it forms virus mRNA and synthesizes and the required viral RNA-dependent form RNA polymerase of genome duplication.Plasmid by cotransfection VSV genome cDNA and coding VSV N,, the expression plasmid of P and L albumen, in cultured cell, produce all these virus components.
All these plasmids are designed to transcribing by phage t7 RNA polymerase; Correspondingly also with the cell of expressing the recombinant vaccinia virus infection transfection of phage polymerase (MVA/T7 or VTF7-3).Standard method for VSV rescue is described in Lawson etc., and 1995, above-mentioned; With Schnell etc., 1996 J.Virol., in 70:2318-23.The method is improved with more effectively for saving highly attenuated virus, also can make the method consistent with the criterion of administrative organization.In brief, the method is as described below.
Utilize calcium phosphate transfection method to use the helper plasmid transfection of rVSV genomic clone and coding VSV N, P and L gene at 12.5cm
2qualified Vero cell in flask.When dying beginning, add the MVA/T7 of q.s calibrating to provide each cell to have the repeatedly infection of 2 plaque forming units.In transfection, start after 3 hours, cell is carried out to 3 hours heat shock steps to improve the rescue efficiency (Parks, C.L. etc., 1999 J.Virol., 73:3560-6) of several minus-stranded rna virus.After transfection 48-72 hour, by cell and media transfer in larger flask, the monolayer Vero cell that this flask contains establiss, incubation one day or multiple days is to allow the virus amplification of rescue subsequently.The virus of gathering in the crops after this amplification step filters to remove the MVA/T7 that great majority pollute, and for the clone and separate of rVSV bacterial strain.
By total order-checking (consensus sequencing) methods analyst rna virus cdna group, to measure the nucleotide sequence of virus genome RNA.In brief, the viral RNA reverse transcription of purification produces cDNA, then uses gene-specific primer to pass through the genomic overlay region of pcr amplification.Use gel-purified PCR product, then use fluorescence staining terminator to carry out cycle sequencing.Purification sequencing reaction product is also analyzed on automatic sequencer.
C. the specific construct using in embodiment
In order to produce the specificity restructuring VSV carrier for following experiment, the restructuring VSVs that expresses HIV-189.6P gp160 is merged to VSV G albumen cross-film district (rVSV HIV-1envG), and mix and be used as experiment immunization originality compositions with SIVmac239 gag p55 (rVSV SIVgag).The rVSV that expresses influenza hemagglutination fibroin (rVSV fluHA) is used as reference composition.As previously mentioned, the gene that is inserted into the required antigen between VSV G and L transcript unit with encoding is prepared restructuring VSV carrier (Rose etc., 2000 J.Virol.74:10903-10).Following structure is referring to Rose etc., the same.
1. plasmid construction body
Amiya doctor Banerjee of Cleveland Clinic provides the plasmid (Masters, P.S., 1989 Virol., 171:285-290) that contains Chandipura glycoprotein [G (Ch)] gene with open arms.In order to build, contain the VSV carrier that G (Ch) gene replaces Indiana glycoprotein [G (I)] gene, first the mutation of using oligonucleotide to instruct is removed XhoI site from G (Ch) gene, has used complementary primer: 5 '-CCCCTAGTGGGATCTCCAGTGATATTTGGAC (SEQ ID NO:2) and 5 '-GTCCAAATATCACTGGAGATCCCACTAGGGG (SEQ ID NO:3) and Stratagene QuikChange mutagenesis kit.CTCGAG (SEQ ID NO:4) sports CTC
caG (SEQ ID NO:5) has removed XhoI site, but does not affect the aminoacid sequence of G (Ch) albumen.Then use Vent archaeal dna polymerase (New England Biolabs) by this gene of pcr amplification.Forward primer is: 5 '-GATCGATCGAATTC
aCGCGTaACATGACTTCTTCAG (SEQ ID NO:6), for G (Ch) albumen, ATG upstream from start codon contains MluI site (underscore).Reverse primer is: 5 '-GAACGGTCGACGCGCCTCGAGCG
tGATATCTGTTaG
tTTTTTT cATAtCATGTTGTTGGGCTTGAAGATC (SEQ ID NO:7), contains SalI and XhoI site (runic), is then VSV transcription initiation and termination signal (underscore), is then the complementary series of G (Ch) 3 ' coded sequence.
PCR product digests with MluI and SalI and is cloned into (Schnell in pVSVXN-1 carrier, Deng 1996 J.Viral.70:2318-2323), this carrier digests to remove VSV G (I) coded sequence (SalI leaves compatible end for being connected with XhoI) with MluI and XhoI in advance.The prepared plasmid of the method is called pVSV (GCh) XN-1 and contains exogenous gene expression site, by unique XhoI and NheI site side joint between G (Ch) gene and L gene.
Use a kind of method identical with said method to produce and comprise the carrier from G (NJ) protein gene of plasmid pNJG (Gallione, C.J., and J.K.Rose.1983 J.Tirol.:162-169).Forward primer is: 5 '-GATCGATCGAATTCACGCGTAATATGTTGTCTTATCTAATCTTTGC (SEQ NO:8), reverse primer is: 5 '-GGAACGGTCGACGCGCCTCGAGCG
tGATATCTGTTaG
tTTT tTTCATAtTAACGGAAATGAGCCATTTCCACG (SEQ NO:9).The site building for above-mentioned Chandipura is expressed as bold-type letter and underscore sequence, and clone's step subsequently also as mentioned above.The carrier finally obtaining is called pVSV (GNJ) XN-1, and it contains exogenous gene expression site, by unique XhoI and NheI site side joint between G (NJ) gene and VSV L gene.
In order to prepare the carrier that contains HIV Env 89.6G gene, (Johnson etc. 1997 with the gene of 89.6 envelope proteins of VSV G Cytoplasm afterbody with XhoI and NheI enzyme action pVSV-89.6 gp160G, to obtain coding, above-mentioned), and be cloned between the XhoI and NheI site of carrier pVSV (GNJ) XN-1 or pVSV (GCh) XN-1.The gp120 that this gp160G gene code is all and gp41 extracellular region and cross-film district, and thering are 4 aminoacid (N-R-V-R) (SEQ ID NO:10) that merge to 89.6 Env cytoplasmic regions of the 26C-end amino acid of VSV G cytoplasmic region, the sequence I-H-L-C (SEQ ID NO:11) of take is starting point.
2. the recovery of recombinant virus
Use method (Lawson etc. 1995 are above-mentioned) the recovery restructuring VSVs having set up.In brief, on 10-cm culture plate, cultivate baby hamster kidney (BHK) cell to approximately 60% fusion.Then use vTF7-3, a kind of vaccinia virus recombinant of expressing T7 RNA polymerase, infection cell repeatedly, infecting (MOI) is 10, (1987 Mol.Cell.Biol.7:2538-2544 such as Fuerst).After 1 hour, with the encode cell of each culture plate of plasmid transfection of one of above-mentioned three kinds of total length recombinants of 3 μ g pBS-N, 5 μ g pBS-P, 1 μ g pBS-L and 10 μ g.The lipofectamine reagent (Rose etc., 1991 Biotechniques 10:520-525) that contains dimethyl hexatriacontane base ammonium bromide and PHOSPHATIDYL ETHANOLAMINE two oleyl alcohol esters has been used in transfection.Cell was 37 ℃ of incubations 48 hours.Cell conditioned medium liquid is removed to most vaccinia virus by 0.2 μ m filter membrane, be then applied to fresh bhk cell, incubation 48 hours again at 37 ℃.For some recovery, include additional plasmid (pBS-G) and N, P and the L helper plasmid of the coding VSV G of 4 microgram/flat boards.By scanning bhk cell monolayer, to obtain VSV cell pathology result, confirm to infect viral recovery.Then isolated viral plaque on bhk cell, by the virus from single plaque is added on the bhk cell culture plate of 10cm diameter, carries out the cultivation from the virus stocks of single plaque.Then these mother solutions are kept to 80 ℃.After freeze thawing, obtain tiring all 10 from VSV (GI)-89.6G, VSV (GCh)-89.6G and VSV (GNJ)-89.6G
7-10
8within the scope of PFU/ml, this process has reduced by the VSV of about 3 times and has tired.
In order to use in following experiment, it is 0.8cc that each rVSV construct all uses aseptic DME to be diluted to final volume.
Embodiment 3: just exempt from/booster immunization inoculation therapy
A. immunization scheme
By dicistronic dna plasmid, the 5mg of intramuscular injection 5mg coding Rhesus Macacus IL-12 p35 and IL-12 p40, express the next immune Rhesus Macacus (5 every group) of DNA plasmid (3 and 4 groups) of DNA plasmid (1 and 2 group) or the 10mg sky of SIV gag p37 polypeptide.All these DNA plasmids and formula thereof are specified in embodiment 1.DNA immunization inoculation time table is defined in the 0th day carries out initial immunity inoculation, carries out inoculating with secondary booster immunization for the first time at the 4th week and the 8th week.Use syringe needle and syringe to inject at triangular muscle and quricipital 4 positions, each position 1cc.
Then at the 15th week, strengthen Rhesus Macacus, by nasal injection (0.4cc/ nostril, use portable pipette) the restructuring vesicular stomatitis virus (rVSV) of Indiana (I) serotype, its carrier (5x10 based on containing HIV-1gp160 env gene in embodiment 2
6and the second rVSV (the I) (5x10 that contains SIV gag p55 gene pfu),
6pfu) (2 and 3 groups), or the rVSV that contains influenza hemagglutinin gene (I) (1x10
7pfu) (1 and 4 group).At the 23rd week, again strengthen Rhesus Macacus, by nasal injection (portable pipette is used in 0.4cc/ nostril) rVSV (Chandipuri serotype, Ch), its carrier (5x10 based on containing HIV-1gp160 env gene in embodiment 2
6and the second rVSV (the Ch) (5x10 that contains SIV gag p55 gene pfu),
6pfu) (2 and 3 groups), or the rVSV that contains influenza hemagglutinin gene (Ch) (1x10
7pfu) (1 and 4 group).
The closely induction of cell and humoral immunoresponse(HI) in monitoring Rhesus Macacus peripheral blood, and at the antibody response of various mucomembranous surfaces.This monitoring comprises to be carried out enzyme linked immunological point mensuration (ELISpot), IFN-γ and HIV-1 env 6101 peptide storehouses is carried out ELISpot mensuration and IFN-γ and VSV N peptide storehouse are carried out to ELISpot mensuration IFN-γ and SIV gag p55 peptide storehouse.This ELISpot measures the cellullar immunologic response detecting in PBL.
Can assess serum (Fig. 4), nose washing liquid, rectum secretions and the anti-SIV gag of saliva (data do not show) p27 IgG tires and anti-HIV-1 gp160 env tires (data do not show) to check humoral immunoresponse(HI) by ELISA.For serum, the antibody using in ELISA is the natural antibody of embedded virus SHIV89.6 and SHIV89.
B.ELISpot algoscopy detects Mus and people's cell of secrete cytokines
Filter membrane immunity Plaque assay is measured (ELISpot) also referred to as enzyme linked immunological point, and initial exploitation is for detection of the B cell with quantitative single secretory antibody.At first, this technology is to provide a kind of quick and general compensation process for conventional plaque forming cells's algoscopy.Recently, the improvement of ELISpot algoscopy has been improved to its sensitivity, made to 100 specific protein molecules of cell generation per second, can be detected less.These algoscopys have been utilized the known albumen (as cytokine) of relative high concentration in secretory protein cell peripheral environment.Use high-affinity antibody can catch and detect these cellular products.
ELISpot algoscopy has been utilized two kinds of high-affinity cytokine specific antibodies of different epi-positions on anti-same cytokine molecule; Or the combination of two kinds of monoclonal antibodies or a kind of monoclonal antibody and a kind of polyvalent antiserum.On chrominance response basis, ELISpot produces speckle, detects the cytokine of individual cells secretion.This speckle represents initial cytokine institute celliferous " footprint ".Speckle (being spot formation cell or SFC) is permanent, can pass through visual, microscope or quantitative with electronic machine.
ELISpot algoscopy is carried out as follows: by mouse-anti human gamma-interferon (hIFN-γ) monoclonal antibody of 1 μ g/ml concentration, (clone 27, BD-Pharmingen, San Diego CA) coated 96 hole flat ELISpot plate (Millipore, Bedford, MA) spend the night, with the 1x PBS that is added with 0.25%Tween-20, wash 10 times, and seal 2 hours with the PBS containing 5% calf serum (FBS).By Ficoll-Hypaque density gradient centrifugation, separated Rhesus Macacus peripheral blood lymphocyte (PBLs) from freshly extd heparinization whole blood, and be resuspended in containing 50 μ g/ml PHA-M (Sigma) or crossed over open overlapping complete medium (RPMI 1640 culture medium in 11 amino acid whose fifteen amino acid peptide storehouses (final peptide concentration is 1 μ M) of readding this frame of whole SIV gag, be added with 5% FCS, 2mM L-glutaminate, 100 units/mL penicillin, 100 μ g/ml streptomycin sulfates, 1mM Sodium Pyruvate, 1mM HEPES, 100 μ M non essential amino acid) in, or only resuspended in culture medium.
The number of putting into cell in 100 μ L/ holes is 2x10
5pBLs also analyzes in two holes.In 37 ℃ of incubation cells 16 hours, then by first using deionized water wash, then 10 times cell is washed down from culture plate with the 1x PBS washing of adding 0.25%Tween-20., with rabbit multi-resistance anti-hIFN-γ biotinylation detect antibody (0.2 μ g/ hole, Biosource, Camarillo, CA) process culture plate, the 1x PBS dilution containing 1% BSA incubation 2 hours at room temperature for this antibody thereafter.Then with the 1x PBS that adds 0.25% Tween-20, wash culture plate 10 times, and with streptavidin alkali phosphatase conjugate (the Southern Biotech in 100 μ l/ holes, Birmingham AL) process, this conjugate is diluted to 1: 500 with the 1x PBS containing 5% FBS and 0.005% Tween-20 and incubation 2.5 hours more at room temperature.By use, add the 1x PBS washing culture plate of 0.25% Tween-20 and remove unconjugated conjugate 10 times.Then, add chromogenic substrate (100 μ L/ holes, 1-step NBT/BCIP, Pierce, Rockford, IL), after 3 to 5 minutes, water rinses, and air-dry culture plate, with being inverted anatomic microscope to the visual counting of speckle producing.
What ELISpot of the present invention measured is that just exempt from/booster immunization inoculation method is replied middle CD8+T cell (CTLs) and the CD4+T cell number producing of inducing in order to be determined at the present invention, and it is to measure by the generation of IFN-γ.This assessment is until the 25th week (after 3 DNA just exempt to strengthen with 2 VSV) after above-mentioned Rhesus Macacus is processed.
C. result
After the initial immunity inoculation of DNA, at Rhesus Macacus (average 254 SFC/10 of 10 SIV gag/rIL-12 DNA immunizations
6cell) in, have 8 SIVgag-specificity IFN-γ ELISpot easily to be detected only and reply.After SIV gag/IL-12DNA immunity for the second time, 10 immune Rhesus Macacus (average 1133 SFC/10
6cell) all shown that high-caliber ELISpot replys, and with the gag/IL-12 DNA of 1/3 dosage, strengthened the gag-specificity ELISpot in most animals and replied (average 1506 SFC/10
6cell).After rVSV HIVenv/rVSVSIVgag for the first time strengthens, and only accept single rVSV HIVenv/rVSV SIVgag immunity inoculation and compare (average 386 SFC/10
6cell), average SIV gag ELISpot replys obvious raising (3772 SFC/10
6cell).These results have supported cytokine to can be used for strengthening the immunogenicity that DNA just exempts to increase rVSV immunity inoculation.
These outcome record are in following table 1 and Fig. 2,3,4 and 5.Fig. 2 has shown that rVSV N-specificity IFN-γ ELISpot replys in the not classification peripheral blood lymphocytes (PBMC) from these immunity animals after 25 weeks.Fig. 3 has shown that in same sample, HIV env 6101 specificity IFN-γ ELISpot reply.Asterisk represents to produce statistically significant difference, p=0.0001.Fig. 4 has shown tiring of the serum antibody of measuring by ELISA anti-SIV gag p27IgG and anti-HIV gp160 env.By the DNA plasmid immunity inoculation of coding SIV gag albumen, also by the scheme that the VSV carrier of expressing flu HA albumen is strengthened, be expressed as (◆).Relate to DNA gag plasmid and just exempt from by the present invention program that the VSV that expresses HIV gag and env albumen strengthens, to be expressed as (■) again after immunity inoculation.Relate to being expressed as (▲) by the scheme of expressing the VSV immunity inoculation of HIV gag and env albumen again after the empty initial immunity inoculation of contrast DNA plasmid.Relate to being expressed as (●) by the scheme of expressing the VSV immunity inoculation of flu HA albumen again after the initial immunity inoculation of contrast DNA plasmid.The result of every group of representative is from 5 animals.Between each group, statistically significant difference is expressed as p=0.0073 (*); P=0.5941 (#) or p=0.0027
.
Fig. 5 has shown the average that the SIV gag specificity IFN-γ ELISpot in same sample replys.By the DNA plasmid immunity inoculation of coding SIV gag albumen, also by the scheme that the VSV carrier of expressing flu HA albumen is strengthened, be expressed as (◆).Relate to being expressed as (■) by the present invention program that the VSV that expresses HIV gag and env albumen strengthens again after the initial immunity inoculation of DNA gag plasmid.Relate to being expressed as (▲) by the scheme of expressing the VSV immunity inoculation of HIV gag and env albumen again after the empty initial immunity inoculation of contrast DNA plasmid.Relate to being expressed as (●) by the scheme of expressing the VSV immunity inoculation of flu HA albumen again after the initial immunity inoculation of contrast DNA plasmid.The result of every group of representative is from 5 animals.Between each group, statistically significant difference is expressed as p=0.0001 and p=0.0002 by square brackets.
Table 1 has recorded identical result with tabular form.
Table 1:SIV gag/ Rhesus Macacus IL-12 DNA just exempts to reply average with the postvaccinal SIVgag-specificity of rVSV-SIV gag/rVSV-HIVenv booster immunization IFN-γ ELISpot
asIVgag-specificity ELISpot at initial dna immunity inoculation record after 25 weeks replys average.
These measurement results have proved with only using repeatedly the result of just exempting from compositions and repeatedly strengthening compositions and have compared, and of the present inventionly just exempt from/strengthen therapy and have wondrous synergism.Use the combination of using again rVSV Booster after the DNA plasmid of expressing required antigen, in immune experimenter, producing the T cells with antigenic specificity increasing, be equivalent to surpass arbitrary additive reaction.Similarly, in using the immunization scheme of DNA plasmid and rVSV immunogenic composition, the T cell confirming in the humoral response of required antigen increases unexpected height.
Embodiment 4: for the Rhesus Macacus model of anti-HIV immunity inoculation
With first the exempting from embodiment 3/strengthen strategy and according to the described plasmid of same approach and VSV carrier, carry out immune Rhesus Macacus (Rh).Use first just exempt from compositions after approximately 32 weeks, with 330 50% monkey infective dose (MID
50) pathogenicity SIV/HIV recombinant virus SHIV89.6P excite Rhesus Macacus (Reimann etc., 1996 J.Virol., 70:3198-3206; 1996 J.Virol., the 70:6922-6928 such as Reimann).
Excite rear about 50-70 days, detect Animal diseases.Before each immunity inoculation, after one week and two weeks, monitor immediately induction and antibody response that zooblast mediates.Before exciting and the serum of collecting for after exciting 2,4,6,8 and 12 weeks for test to HIV-189.6,89.6P, 6101 and other the neutralizing antibody of clade B initially-separate thing react.In the immunity inoculation stage of test, before exciting and excite latter every two weeks and monitor immediately CD4/CD8 counting.Excite front and excite rear each week, by branched DNA, analyzing and measure immediately viral load in serum.Also check cell and the humoral immunoresponse(HI) of other body fluid (vagina, rectum, nasal discharge and saliva) of animal.
Use the cell-mediated immunne response of several most suitable algoscopy analyses based on cell to required antigen, comprise
51cr-discharges CTL algoscopy, soluble MHC I type tetramer staining, ELISpot algoscopy and cell within a cell factorial analysis.By elisa technique, assess mucoantibody and react, optimize for mucosa sample.Elisa technique by standard also can be assessed serum antibody response.In addition, measure the serum from all immune animals, to determine the neutralizing antibody reaction to HIV Env6101 and other initial clade B separator.Fig. 6 has illustrated by the present invention and has just exempted from/strengthened the immunne response of having strengthened in conjunction with having caused, and causes the AIDS protective effect that increases, by measuring cd4 t cell loss at least 250 days after exciting, reduces and determines.
Except the immunne response that monitoring immunogenic composition causes, also by detecting its emission levels and persistent period, assess the propagation potential of live vector.After each immunity inoculation, starting frequently to obtain at regular intervals nose washing liquid in three weeks exists to measure live virus.Also detect plasmid viral load for determining the live virus copy existing.Fig. 7 shown by just exempting from/strengthen the immunne response in conjunction with the reinforcement causing, and by measuring the minimizing of blood plasma internal recycle virus at least 250 days after exciting, determines.
Compare with control animal, the result of above-mentioned detection probably shows to have very high antigenic specificity CD8+ and CD+T cell in the animal of the immunity according to the present invention.Should expect according to the present invention that the animal of just exempting from/strengthening therapy immunity still keeps fit after HIV exposes, non-immune animal suffers from AIDS.
Compare with other known result of just exempting from/strengthen therapy scheme, just exempt from compositions immunity inoculation with one or many DNA only or result that only one or many rVSV carrier immunity inoculation provides is compared, can estimate that the inventive method has immune animal safety and can print and distribute anti-HIV CTLs and the antibody of higher level.May tool synergism according to first the exempting from of repetition of the present invention/strengthen, and therefore to pre-exposure experimenter tool preventive effect, the experimenter of infected by HIV is had to therapeutical effect.
Citing document in all above-mentioned description is incorporated herein by reference in this.The various variants of the inventive method and component and less variation are apparent to those skilled in the art.
Claims (23)
1. the application of following the first and second compositionss in the medicine for the preparation of inducing antigen-specific immunne response in mammalian subject, wherein
(a) the first described compositions comprises
(i) the DNA plasmid of the DNA sequence that contains coding for antigens, modulated its regulating and controlling sequence by described DNA plasmid expression of described coded sequence regulates and controls; With
(ii) the DNA plasmid of the DNA sequence that comprises the Codocyte factor; And
(b) the second described compositions comprises the restructuring vesicular stomatitis virus VSV (vesicular stomatitis virus) of the nucleotide sequence that contains the described antigen of encoding, the modulated regulating and controlling sequence that it is expressed by restructuring VSV of described coded sequence regulates and controls
Described antigen is albumen, peptide or its fragment or fusant, described antigen is from the member who is selected from antibacterial, virus, fungus, parasite, tumor cell, anaphylactogen and self molecule, described cytokine is selected from IL-12, IL-15, GM-CSF and combination thereof.
2. according to the application of claim 1, wherein said VSV has replication capacity or does not copy.
3. according to the application of claim 1, wherein before using described the second compositions, to experimenter, at least use once described the first compositions.
4. according to the application of claim 3, wherein said the second compositions is used at least one times to experimenter.
5. according to the application of claim 1, wherein before using described the first compositions, to experimenter, at least use once described the second compositions.
6. according to the application of claim 5, wherein said the first compositions is used at least one times to experimenter.
7. according to the application of claim 1, wherein said cytokine coded sequence is present on same DNA plasmid from described antigen encoding sequence or is present on the DNA plasmid different with the DNA plasmid of the described antigen of coding.
8. according to the application of claim 1, wherein said the first compositions further comprises and is selected from people or simian immunodeficiency virus pol, env, nef, vpr, vpu, vif and tat antigen, and the additional antigen of immunogenic fragments or fusions or its combination.
9. according to the application of claim 1, wherein said the first compositions is selected from:
(i) the first compositions that contains described DNA sequence more than one copy, that encode described antigen, wherein, described copy is on identical plasmid or different plasmid; With
(ii) further comprise the first compositions of following DNA plasmid; wherein; the DNA sequence that described DNA plasmid comprises the additional antigen of coding, described additional antigen is different from described the first antigen, and the described DNA sequence of the described antigen of wherein encoding is on identical plasmid or different plasmid.
10. according to the application of claim 1; wherein said immunne response comprises that (i) is better than the situation of only using described DNA plasmid or restructuring VSV to the CD8+T cell response of described antigen; (ii) with only use described first or the situation of the second compositions compare, in to the antibody response of described antigen, there is chemiluminescence.
11. according to the application of claim 1, and wherein said the second compositions also comprises the restructuring VSV that at least one is additional, and this additional restructuring VSV is different from the first restructuring VSV.
12. according to the application of claim 11, and wherein the restructuring of the difference described in each VSV comprises:
(i) different VSV G albumen and different VSV serotype, but there is identical antigen encoding sequence,
(ii) different antigen encoding sequence, but identical VSV G albumen, or
(iii) different antigen encoding sequences and different VSV G albumen.
13. according to the application of claim 1, wherein, the second described compositions comprises at least one additional rVSV, encode identical antigen or different antigen of each rVSV wherein, and each rVSV wherein has identical or different VSV serotype.
14. according to the application of claim 1, wherein, in mammalian subject, inducing antigen-specific immunne response comprises at least two kinds of rVSV compositionss of continuous administration, wherein every kind of rVSV compositions has different VSV serotype and identical antigen encoding sequence, or it has different VSV serotype and different antigen encoding sequences, wherein said not synantigen is albumen, peptide or its fragment or fusant, and described antigen is from the member who is selected from antibacterial, virus, fungus, parasite, tumor cell, anaphylactogen and self molecule.
The application of 15. claim 1, wherein, described mammalian subject is people or non-human primates.
16. according to the application of claim 1, and wherein said the first compositions further comprises pharmaceutically acceptable diluent, excipient or carrier.
The application of 17. claim 16, wherein, described excipient comprises bupivacaine.
18. according to the application of claim 1, and wherein said the second compositions further comprises pharmaceutically acceptable diluent, excipient or carrier.
19. 1 kinds of test kits, comprising:
The first compositions of at least one dosage, comprises the DNA plasmid of the DNA sequence that contains coding for antigens, and modulated its regulating and controlling sequence by described DNA plasmid expression of described coded sequence regulates and controls;
The second compositions of at least one dosage, comprises the restructuring vesicular stomatitis virus VSV of the nucleotide sequence that contains the described antigen of encoding, and the modulated regulating and controlling sequence that it is expressed by described restructuring VSV of described coded sequence regulates and controls; With
For implement the description of the method for inducing antigen-specific immunne response in mammalian subject, wherein said antigen is albumen, peptide or its fragment or fusant, and described antigen is from the member who is selected from antibacterial, virus, fungus, parasite, tumor cell, anaphylactogen and self molecule.
20. according to the test kit of claim 19, and wherein said VSV has replication capacity or do not copy.
21. according to the test kit of claim 19, also comprises cell factor composition.
22. according to the test kit of claim 21, wherein said cell factor composition comprises the nucleic acid compositions that contains DNA plasmid, described DNA plasmid comprises the DNA sequence of the described cytokine of encoding, and modulated its regulating and controlling sequence by described DNA plasmid expression of described coded sequence regulates and controls.
23. according to the application of claim 1 or 14 or according to the test kit of claim 19, and wherein said peptide is polypeptide, and described tumor cell is cancerous cell.
Applications Claiming Priority (6)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| US45787603P | 2003-03-26 | 2003-03-26 | |
| US60/457876 | 2003-03-26 | ||
| US60/457,876 | 2003-03-26 | ||
| US54673304P | 2004-02-23 | 2004-02-23 | |
| US60/546733 | 2004-02-23 | ||
| US60/546,733 | 2004-02-23 |
Related Parent Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN2004800078180A Division CN1764474B (en) | 2003-03-26 | 2004-03-23 | Immunogenic composition and methods |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN102085377A CN102085377A (en) | 2011-06-08 |
| CN102085377B true CN102085377B (en) | 2014-03-12 |
Family
ID=33313369
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201010587086.4A Expired - Lifetime CN102085377B (en) | 2003-03-26 | 2004-03-23 | Immunogenic composition and methods |
Country Status (7)
| Country | Link |
|---|---|
| JP (1) | JP5635950B2 (en) |
| KR (1) | KR101059721B1 (en) |
| CN (1) | CN102085377B (en) |
| CY (1) | CY1110205T1 (en) |
| DE (1) | DE602004027359D1 (en) |
| ES (1) | ES2345198T3 (en) |
| PT (1) | PT1605971E (en) |
Families Citing this family (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN111662885A (en) * | 2020-06-22 | 2020-09-15 | 扬州大学 | Construction, rescue and application of infectious clone of virulent and attenuated strains of two highly homologous genome porcine reproductive and respiratory syndrome viruses |
Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2002097091A1 (en) * | 2001-05-29 | 2002-12-05 | University Of Miami | Generation of hcv-like particles and chimeric hcv virus |
Family Cites Families (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US7153510B1 (en) * | 1995-05-04 | 2006-12-26 | Yale University | Recombinant vesiculoviruses and their uses |
| NZ542652A (en) * | 2003-03-26 | 2008-04-30 | Wyeth Corp | Immunogenic composition and methods |
-
2004
- 2004-03-23 ES ES04722764T patent/ES2345198T3/en not_active Expired - Lifetime
- 2004-03-23 CN CN201010587086.4A patent/CN102085377B/en not_active Expired - Lifetime
- 2004-03-23 KR KR1020057018099A patent/KR101059721B1/en active IP Right Grant
- 2004-03-23 DE DE602004027359T patent/DE602004027359D1/en not_active Expired - Lifetime
- 2004-03-23 PT PT04722764T patent/PT1605971E/en unknown
-
2010
- 2010-07-15 CY CY20101100663T patent/CY1110205T1/en unknown
-
2011
- 2011-07-13 JP JP2011154426A patent/JP5635950B2/en not_active Expired - Lifetime
Patent Citations (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| WO2002097091A1 (en) * | 2001-05-29 | 2002-12-05 | University Of Miami | Generation of hcv-like particles and chimeric hcv virus |
Non-Patent Citations (2)
| Title |
|---|
| Gerd Sutter et al..Novel vaccine delivery systems:solutions to HIV vaccine dilemmas?.《AIDS》.2001,第15卷S142页左栏最后1段-右栏第2段. * |
| Nina F.Rose et al..An Effective AIDS Vaccine Based on Live Attenuated Vesicular Stomatitis Virus Recombinants.《Cell》.2001,第106卷摘要. * |
Also Published As
| Publication number | Publication date |
|---|---|
| KR101059721B1 (en) | 2011-08-29 |
| KR20060002872A (en) | 2006-01-09 |
| JP5635950B2 (en) | 2014-12-03 |
| JP2011251979A (en) | 2011-12-15 |
| CY1110205T1 (en) | 2015-01-14 |
| CN102085377A (en) | 2011-06-08 |
| PT1605971E (en) | 2010-07-29 |
| ES2345198T3 (en) | 2010-09-17 |
| DE602004027359D1 (en) | 2010-07-08 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN1764474B (en) | Immunogenic composition and methods | |
| JP5264840B2 (en) | A plasmid having three complete transcription units and an immunogenic composition for eliciting an immune response against HIV | |
| CN102782136A (en) | Vectors expressing HIV antigens and GM-CSF and related methods for generating an immune response | |
| Billaut-Mulot et al. | Interleukin-18 modulates immune responses induced by HIV-1 Nef DNA prime/protein boost vaccine | |
| IL168614A (en) | Genetic constructs comprising rre and cte, compositions comprising them and method for their production | |
| CN102085377B (en) | Immunogenic composition and methods | |
| CN101001953B (en) | Plasmid having three complete transcriptional units and immunogenic compositions for inducing an immune response to HIV | |
| ZA200507722B (en) | Immunogenic composition and methods |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| GR01 | Patent grant | ||
| GR01 | Patent grant | ||
| CX01 | Expiry of patent term | ||
| CX01 | Expiry of patent term |
Granted publication date: 20140312 |