WO1999020652A1 - Facteur stimulant les colonies de granulocytes de felins - Google Patents
Facteur stimulant les colonies de granulocytes de felins Download PDFInfo
- Publication number
- WO1999020652A1 WO1999020652A1 PCT/JP1998/004809 JP9804809W WO9920652A1 WO 1999020652 A1 WO1999020652 A1 WO 1999020652A1 JP 9804809 W JP9804809 W JP 9804809W WO 9920652 A1 WO9920652 A1 WO 9920652A1
- Authority
- WO
- WIPO (PCT)
- Prior art keywords
- protein
- feline
- stimulating factor
- activity
- amino acid
- Prior art date
Links
- 108010017080 Granulocyte Colony-Stimulating Factor Proteins 0.000 title claims abstract description 63
- 102000004269 Granulocyte Colony-Stimulating Factor Human genes 0.000 title claims abstract description 59
- 241000282324 Felis Species 0.000 title claims abstract description 31
- 108090000623 proteins and genes Proteins 0.000 claims abstract description 51
- 102000004169 proteins and genes Human genes 0.000 claims abstract description 28
- 230000000694 effects Effects 0.000 claims abstract description 21
- 238000004519 manufacturing process Methods 0.000 claims abstract description 9
- 239000004480 active ingredient Substances 0.000 claims abstract description 4
- 241000282326 Felis catus Species 0.000 claims description 45
- 238000000034 method Methods 0.000 claims description 30
- 125000003275 alpha amino acid group Chemical group 0.000 claims description 25
- 150000001413 amino acids Chemical class 0.000 claims description 14
- 210000000440 neutrophil Anatomy 0.000 claims description 9
- 238000012258 culturing Methods 0.000 claims description 5
- 239000003814 drug Substances 0.000 claims description 5
- 229940124597 therapeutic agent Drugs 0.000 claims description 5
- 239000013604 expression vector Substances 0.000 claims description 4
- 239000008194 pharmaceutical composition Substances 0.000 claims description 3
- 239000000427 antigen Substances 0.000 claims 1
- 102000036639 antigens Human genes 0.000 claims 1
- 108091007433 antigens Proteins 0.000 claims 1
- 238000009395 breeding Methods 0.000 claims 1
- 230000001488 breeding effect Effects 0.000 claims 1
- 230000015271 coagulation Effects 0.000 claims 1
- 238000005345 coagulation Methods 0.000 claims 1
- 238000002405 diagnostic procedure Methods 0.000 claims 1
- 210000003714 granulocyte Anatomy 0.000 claims 1
- 238000003259 recombinant expression Methods 0.000 claims 1
- 230000004936 stimulating effect Effects 0.000 claims 1
- 208000004235 neutropenia Diseases 0.000 abstract description 7
- 206010028980 Neoplasm Diseases 0.000 abstract description 4
- 210000000265 leukocyte Anatomy 0.000 abstract description 2
- 230000002411 adverse Effects 0.000 abstract 1
- 125000000539 amino acid group Chemical group 0.000 abstract 1
- 210000004493 neutrocyte Anatomy 0.000 abstract 1
- 230000001737 promoting effect Effects 0.000 abstract 1
- 210000004027 cell Anatomy 0.000 description 49
- 108020004414 DNA Proteins 0.000 description 36
- 239000002773 nucleotide Substances 0.000 description 30
- 125000003729 nucleotide group Chemical group 0.000 description 30
- 239000002609 medium Substances 0.000 description 19
- 239000013615 primer Substances 0.000 description 18
- 239000013598 vector Substances 0.000 description 17
- 239000012634 fragment Substances 0.000 description 16
- 241000701447 unidentified baculovirus Species 0.000 description 16
- 108091008146 restriction endonucleases Proteins 0.000 description 15
- 239000002299 complementary DNA Substances 0.000 description 14
- 239000000047 product Substances 0.000 description 13
- 238000010367 cloning Methods 0.000 description 12
- 108020001019 DNA Primers Proteins 0.000 description 11
- 239000003155 DNA primer Substances 0.000 description 11
- 101000746367 Homo sapiens Granulocyte colony-stimulating factor Proteins 0.000 description 10
- 239000000243 solution Substances 0.000 description 10
- 241000700605 Viruses Species 0.000 description 9
- 230000004071 biological effect Effects 0.000 description 9
- 238000006243 chemical reaction Methods 0.000 description 9
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 8
- 239000012228 culture supernatant Substances 0.000 description 8
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Chemical compound O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 8
- 108091032973 (ribonucleotides)n+m Proteins 0.000 description 7
- 241000588724 Escherichia coli Species 0.000 description 7
- 241000238631 Hexapoda Species 0.000 description 7
- 238000002360 preparation method Methods 0.000 description 7
- 238000012546 transfer Methods 0.000 description 7
- 241000282414 Homo sapiens Species 0.000 description 6
- 229960000723 ampicillin Drugs 0.000 description 6
- AVKUERGKIZMTKX-NJBDSQKTSA-N ampicillin Chemical compound C1([C@@H](N)C(=O)N[C@H]2[C@H]3SC([C@@H](N3C2=O)C(O)=O)(C)C)=CC=CC=C1 AVKUERGKIZMTKX-NJBDSQKTSA-N 0.000 description 6
- 230000003321 amplification Effects 0.000 description 6
- 239000012153 distilled water Substances 0.000 description 6
- 238000003199 nucleic acid amplification method Methods 0.000 description 6
- 239000013612 plasmid Substances 0.000 description 6
- 229920000936 Agarose Polymers 0.000 description 5
- 108091028043 Nucleic acid sequence Proteins 0.000 description 5
- 239000000203 mixture Substances 0.000 description 5
- 239000002953 phosphate buffered saline Substances 0.000 description 5
- 238000010839 reverse transcription Methods 0.000 description 5
- 238000011282 treatment Methods 0.000 description 5
- FWMNVWWHGCHHJJ-SKKKGAJSSA-N 4-amino-1-[(2r)-6-amino-2-[[(2r)-2-[[(2r)-2-[[(2r)-2-amino-3-phenylpropanoyl]amino]-3-phenylpropanoyl]amino]-4-methylpentanoyl]amino]hexanoyl]piperidine-4-carboxylic acid Chemical compound C([C@H](C(=O)N[C@H](CC(C)C)C(=O)N[C@H](CCCCN)C(=O)N1CCC(N)(CC1)C(O)=O)NC(=O)[C@H](N)CC=1C=CC=CC=1)C1=CC=CC=C1 FWMNVWWHGCHHJJ-SKKKGAJSSA-N 0.000 description 4
- KFZMGEQAYNKOFK-UHFFFAOYSA-N Isopropanol Chemical compound CC(C)O KFZMGEQAYNKOFK-UHFFFAOYSA-N 0.000 description 4
- 241000699666 Mus <mouse, genus> Species 0.000 description 4
- 101000746384 Mus musculus Granulocyte colony-stimulating factor Proteins 0.000 description 4
- 238000002835 absorbance Methods 0.000 description 4
- 238000007792 addition Methods 0.000 description 4
- 239000000872 buffer Substances 0.000 description 4
- 238000005119 centrifugation Methods 0.000 description 4
- 230000014509 gene expression Effects 0.000 description 4
- 230000006798 recombination Effects 0.000 description 4
- 239000006228 supernatant Substances 0.000 description 4
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 3
- 241000282472 Canis lupus familiaris Species 0.000 description 3
- 241000282412 Homo Species 0.000 description 3
- ISWSIDIOOBJBQZ-UHFFFAOYSA-N Phenol Chemical compound OC1=CC=CC=C1 ISWSIDIOOBJBQZ-UHFFFAOYSA-N 0.000 description 3
- 108010076504 Protein Sorting Signals Proteins 0.000 description 3
- 238000012217 deletion Methods 0.000 description 3
- 230000037430 deletion Effects 0.000 description 3
- 239000002158 endotoxin Substances 0.000 description 3
- 238000005516 engineering process Methods 0.000 description 3
- 229920006008 lipopolysaccharide Polymers 0.000 description 3
- 239000002244 precipitate Substances 0.000 description 3
- 238000005215 recombination Methods 0.000 description 3
- 238000011084 recovery Methods 0.000 description 3
- 238000006467 substitution reaction Methods 0.000 description 3
- 238000003786 synthesis reaction Methods 0.000 description 3
- 230000009466 transformation Effects 0.000 description 3
- CPELXLSAUQHCOX-UHFFFAOYSA-M Bromide Chemical compound [Br-] CPELXLSAUQHCOX-UHFFFAOYSA-M 0.000 description 2
- HEDRZPFGACZZDS-UHFFFAOYSA-N Chloroform Chemical compound ClC(Cl)Cl HEDRZPFGACZZDS-UHFFFAOYSA-N 0.000 description 2
- 108020004705 Codon Proteins 0.000 description 2
- 102000004127 Cytokines Human genes 0.000 description 2
- 108090000695 Cytokines Proteins 0.000 description 2
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 description 2
- 108010067770 Endopeptidase K Proteins 0.000 description 2
- ZRALSGWEFCBTJO-UHFFFAOYSA-N Guanidine Chemical compound NC(N)=N ZRALSGWEFCBTJO-UHFFFAOYSA-N 0.000 description 2
- 101000716729 Homo sapiens Kit ligand Proteins 0.000 description 2
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 2
- 102100020880 Kit ligand Human genes 0.000 description 2
- 239000005662 Paraffin oil Substances 0.000 description 2
- VMHLLURERBWHNL-UHFFFAOYSA-M Sodium acetate Chemical compound [Na+].CC([O-])=O VMHLLURERBWHNL-UHFFFAOYSA-M 0.000 description 2
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 2
- 108091081024 Start codon Proteins 0.000 description 2
- 101000865057 Thermococcus litoralis DNA polymerase Proteins 0.000 description 2
- IQFYYKKMVGJFEH-XLPZGREQSA-N Thymidine Chemical compound O=C1NC(=O)C(C)=CN1[C@@H]1O[C@H](CO)[C@@H](O)C1 IQFYYKKMVGJFEH-XLPZGREQSA-N 0.000 description 2
- 239000011543 agarose gel Substances 0.000 description 2
- 238000004166 bioassay Methods 0.000 description 2
- 210000004369 blood Anatomy 0.000 description 2
- 239000008280 blood Substances 0.000 description 2
- 201000011510 cancer Diseases 0.000 description 2
- 230000010261 cell growth Effects 0.000 description 2
- 210000004748 cultured cell Anatomy 0.000 description 2
- OPTASPLRGRRNAP-UHFFFAOYSA-N cytosine Chemical compound NC=1C=CNC(=O)N=1 OPTASPLRGRRNAP-UHFFFAOYSA-N 0.000 description 2
- 239000003085 diluting agent Substances 0.000 description 2
- 238000010790 dilution Methods 0.000 description 2
- 239000012895 dilution Substances 0.000 description 2
- LOKCTEFSRHRXRJ-UHFFFAOYSA-I dipotassium trisodium dihydrogen phosphate hydrogen phosphate dichloride Chemical compound P(=O)(O)(O)[O-].[K+].P(=O)(O)([O-])[O-].[Na+].[Na+].[Cl-].[K+].[Cl-].[Na+] LOKCTEFSRHRXRJ-UHFFFAOYSA-I 0.000 description 2
- 210000003527 eukaryotic cell Anatomy 0.000 description 2
- 239000012091 fetal bovine serum Substances 0.000 description 2
- 238000010353 genetic engineering Methods 0.000 description 2
- 208000015181 infectious disease Diseases 0.000 description 2
- PHTQWCKDNZKARW-UHFFFAOYSA-N isoamylol Chemical compound CC(C)CCO PHTQWCKDNZKARW-UHFFFAOYSA-N 0.000 description 2
- 210000003734 kidney Anatomy 0.000 description 2
- 230000007774 longterm Effects 0.000 description 2
- 238000012423 maintenance Methods 0.000 description 2
- 238000002156 mixing Methods 0.000 description 2
- 230000035772 mutation Effects 0.000 description 2
- 150000007523 nucleic acids Chemical class 0.000 description 2
- 210000005259 peripheral blood Anatomy 0.000 description 2
- 239000011886 peripheral blood Substances 0.000 description 2
- 210000001236 prokaryotic cell Anatomy 0.000 description 2
- 210000002966 serum Anatomy 0.000 description 2
- 239000001632 sodium acetate Substances 0.000 description 2
- 235000017281 sodium acetate Nutrition 0.000 description 2
- 241000894007 species Species 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- QKNYBSVHEMOAJP-UHFFFAOYSA-N 2-amino-2-(hydroxymethyl)propane-1,3-diol;hydron;chloride Chemical compound Cl.OCC(N)(CO)CO QKNYBSVHEMOAJP-UHFFFAOYSA-N 0.000 description 1
- NKDFYOWSKOHCCO-YPVLXUMRSA-N 20-hydroxyecdysone Chemical compound C1[C@@H](O)[C@@H](O)C[C@]2(C)[C@@H](CC[C@@]3([C@@H]([C@@](C)(O)[C@H](O)CCC(C)(O)C)CC[C@]33O)C)C3=CC(=O)[C@@H]21 NKDFYOWSKOHCCO-YPVLXUMRSA-N 0.000 description 1
- -1 3-indolyl- Chemical group 0.000 description 1
- DWRXFEITVBNRMK-UHFFFAOYSA-N Beta-D-1-Arabinofuranosylthymine Natural products O=C1NC(=O)C(C)=CN1C1C(O)C(O)C(CO)O1 DWRXFEITVBNRMK-UHFFFAOYSA-N 0.000 description 1
- 101000746369 Canis lupus familiaris Granulocyte colony-stimulating factor Proteins 0.000 description 1
- 102000012410 DNA Ligases Human genes 0.000 description 1
- 108010061982 DNA Ligases Proteins 0.000 description 1
- 230000004543 DNA replication Effects 0.000 description 1
- 239000006144 Dulbecco’s modified Eagle's medium Substances 0.000 description 1
- 238000002965 ELISA Methods 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 101000993347 Gallus gallus Ciliary neurotrophic factor Proteins 0.000 description 1
- 102100034343 Integrase Human genes 0.000 description 1
- 102000000589 Interleukin-1 Human genes 0.000 description 1
- 108010002352 Interleukin-1 Proteins 0.000 description 1
- 102100021592 Interleukin-7 Human genes 0.000 description 1
- 108010002586 Interleukin-7 Proteins 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- 101001033276 Mus musculus Interleukin-3 Proteins 0.000 description 1
- 241000699670 Mus sp. Species 0.000 description 1
- BACYUWVYYTXETD-UHFFFAOYSA-N N-Lauroylsarcosine Chemical compound CCCCCCCCCCCC(=O)N(C)CC(O)=O BACYUWVYYTXETD-UHFFFAOYSA-N 0.000 description 1
- CHJJGSNFBQVOTG-UHFFFAOYSA-N N-methyl-guanidine Natural products CNC(N)=N CHJJGSNFBQVOTG-UHFFFAOYSA-N 0.000 description 1
- 108700026244 Open Reading Frames Proteins 0.000 description 1
- 101700056750 PAK1 Proteins 0.000 description 1
- 102000045595 Phosphoprotein Phosphatases Human genes 0.000 description 1
- 108700019535 Phosphoprotein Phosphatases Proteins 0.000 description 1
- 101710182846 Polyhedrin Proteins 0.000 description 1
- 108010092799 RNA-directed DNA polymerase Proteins 0.000 description 1
- 239000012980 RPMI-1640 medium Substances 0.000 description 1
- 101000702488 Rattus norvegicus High affinity cationic amino acid transporter 1 Proteins 0.000 description 1
- 102000007056 Recombinant Fusion Proteins Human genes 0.000 description 1
- 108010008281 Recombinant Fusion Proteins Proteins 0.000 description 1
- 102100027910 Serine/threonine-protein kinase PAK 1 Human genes 0.000 description 1
- 108010006785 Taq Polymerase Proteins 0.000 description 1
- 239000007983 Tris buffer Substances 0.000 description 1
- HMNZFMSWFCAGGW-XPWSMXQVSA-N [3-[hydroxy(2-hydroxyethoxy)phosphoryl]oxy-2-[(e)-octadec-9-enoyl]oxypropyl] (e)-octadec-9-enoate Chemical compound CCCCCCCC\C=C\CCCCCCCC(=O)OCC(COP(O)(=O)OCCO)OC(=O)CCCCCCC\C=C\CCCCCCCC HMNZFMSWFCAGGW-XPWSMXQVSA-N 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 238000001042 affinity chromatography Methods 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- IQFYYKKMVGJFEH-UHFFFAOYSA-N beta-L-thymidine Natural products O=C1NC(=O)C(C)=CN1C1OC(CO)C(O)C1 IQFYYKKMVGJFEH-UHFFFAOYSA-N 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 210000000988 bone and bone Anatomy 0.000 description 1
- 238000010322 bone marrow transplantation Methods 0.000 description 1
- 230000030833 cell death Effects 0.000 description 1
- 239000013592 cell lysate Substances 0.000 description 1
- 230000004663 cell proliferation Effects 0.000 description 1
- 239000013043 chemical agent Substances 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 238000002512 chemotherapy Methods 0.000 description 1
- 238000004587 chromatography analysis Methods 0.000 description 1
- 208000037516 chromosome inversion disease Diseases 0.000 description 1
- 238000003776 cleavage reaction Methods 0.000 description 1
- 230000000295 complement effect Effects 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 238000010276 construction Methods 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 201000001546 cyclic hematopoiesis Diseases 0.000 description 1
- 229940104302 cytosine Drugs 0.000 description 1
- 230000003247 decreasing effect Effects 0.000 description 1
- 125000000118 dimethyl group Chemical group [H]C([H])([H])* 0.000 description 1
- SWSQBOPZIKWTGO-UHFFFAOYSA-N dimethylaminoamidine Natural products CN(C)C(N)=N SWSQBOPZIKWTGO-UHFFFAOYSA-N 0.000 description 1
- 231100000673 dose–response relationship Toxicity 0.000 description 1
- 210000002919 epithelial cell Anatomy 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 239000012894 fetal calf serum Substances 0.000 description 1
- 238000007667 floating Methods 0.000 description 1
- 239000011521 glass Substances 0.000 description 1
- 210000003958 hematopoietic stem cell Anatomy 0.000 description 1
- 230000011132 hemopoiesis Effects 0.000 description 1
- 230000016784 immunoglobulin production Effects 0.000 description 1
- 238000002347 injection Methods 0.000 description 1
- 239000007924 injection Substances 0.000 description 1
- 229940100994 interleukin-7 Drugs 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
- 210000004185 liver Anatomy 0.000 description 1
- 238000011866 long-term treatment Methods 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 239000013642 negative control Substances 0.000 description 1
- PGSADBUBUOPOJS-UHFFFAOYSA-N neutral red Chemical compound Cl.C1=C(C)C(N)=CC2=NC3=CC(N(C)C)=CC=C3N=C21 PGSADBUBUOPOJS-UHFFFAOYSA-N 0.000 description 1
- 230000003472 neutralizing effect Effects 0.000 description 1
- 231100000957 no side effect Toxicity 0.000 description 1
- 108020004707 nucleic acids Proteins 0.000 description 1
- 102000039446 nucleic acids Human genes 0.000 description 1
- 239000008188 pellet Substances 0.000 description 1
- 230000000865 phosphorylative effect Effects 0.000 description 1
- 230000035755 proliferation Effects 0.000 description 1
- 238000011002 quantification Methods 0.000 description 1
- 239000011541 reaction mixture Substances 0.000 description 1
- 230000007115 recruitment Effects 0.000 description 1
- 239000000523 sample Substances 0.000 description 1
- 108700004121 sarkosyl Proteins 0.000 description 1
- 229920006395 saturated elastomer Polymers 0.000 description 1
- 230000007017 scission Effects 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- 239000011780 sodium chloride Substances 0.000 description 1
- 239000001509 sodium citrate Substances 0.000 description 1
- NLJMYIDDQXHKNR-UHFFFAOYSA-K sodium citrate Chemical compound O.O.[Na+].[Na+].[Na+].[O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O NLJMYIDDQXHKNR-UHFFFAOYSA-K 0.000 description 1
- AEEZIRBYGITPDN-UHFFFAOYSA-M sodium;dodecanoyl sulfate Chemical compound [Na+].CCCCCCCCCCCC(=O)OS([O-])(=O)=O AEEZIRBYGITPDN-UHFFFAOYSA-M 0.000 description 1
- 239000007921 spray Substances 0.000 description 1
- 210000000130 stem cell Anatomy 0.000 description 1
- 238000003756 stirring Methods 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 230000003319 supportive effect Effects 0.000 description 1
- 229940126585 therapeutic drug Drugs 0.000 description 1
- 229940104230 thymidine Drugs 0.000 description 1
- 230000036962 time dependent Effects 0.000 description 1
- 238000013518 transcription Methods 0.000 description 1
- 230000035897 transcription Effects 0.000 description 1
- 238000001890 transfection Methods 0.000 description 1
- LENZDBCJOHFCAS-UHFFFAOYSA-N tris Chemical compound OCC(N)(CO)CO LENZDBCJOHFCAS-UHFFFAOYSA-N 0.000 description 1
- 238000011144 upstream manufacturing Methods 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
- DGVVWUTYPXICAM-UHFFFAOYSA-N β‐Mercaptoethanol Chemical compound OCCS DGVVWUTYPXICAM-UHFFFAOYSA-N 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
- C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
- C07K14/52—Cytokines; Lymphokines; Interferons
- C07K14/53—Colony-stimulating factor [CSF]
- C07K14/535—Granulocyte CSF; Granulocyte-macrophage CSF
-
- 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
- A61K38/193—Colony stimulating factors [CSF]
Definitions
- the present invention relates to a feline granulocyte colony-stimulating factor and a gene encoding the same, and more particularly, to a feline granulocyte colony-stimulating factor useful as a substance mainly acting on neutrophil production and function maintenance in cats. Factors and methods for their manufacture. Background art
- G-CSF granulocyte colony stimulating factor
- Human G-CSF is a secreted protein having a molecular weight of about 19 kDa, and includes a protein consisting of 204 amino acids including a signal sequence consisting of 30 amino acids and a protein consisting of 207 amino acids.
- Va l-Ser-Glu is inserted between Le 35 and Cys 36 of the former.
- the latter G—CSF biological activity is invitro, which is about 1Z10 of the former.
- the former that is, G-CSF, consisting of 174 highly active amino acids, is considered to be mainly present in the body and play an important role.
- a recombinant human G-CSF preparation When a recombinant human G-CSF preparation is administered to a living body, it stimulates neutrophil production at a very high rate and effectively in a dose-dependent and time-dependent manner, increasing the number of peripheral blood neutrophils. And enhances the function of increased mature neutrophils and causes the recruitment of hematopoietic stem cells to the peripheral blood. These effects quickly disappear when administration is stopped. In addition, there are almost no side effects even at very large doses, and long-term administration does not cause stem cell death or antibody production. Thus, it can be applied to supportive care after cancer chemotherapy, treatment of severe infections during cancer treatment, recovery of neutropenia after bone marrow transplantation, and early recovery of hematopoiesis.
- the present inventors cloned a gene encoding cat G-CSF in order to supply a protein having the biological activity of cat G-CSF at low cost, and further possessed the gene.
- An expression vector was prepared, and the present invention was completed.
- the invention of claim 1 of the present application is characterized in that (a) a force having the amino acid sequence of SEQ ID NO: 1, or (b) one or several amino acids in the amino acid sequence of SEQ ID NO: 1 are deleted or substituted.
- it is a protein having an added amino acid sequence and having the activity of feline granulocyte colony stimulating factor (ie, the biological activity of feline G-CSF) It is characterized by the following.
- the above amino acid of SEQ ID NO: 1 has the following characteristics.
- Organism name Cat (feiridaeefelis scatu s)
- the protein having the biological activity of cat G-CSF having the above amino acid sequence (a) was transcribed and translated from a gene fragment having the nucleotide sequence of SEQ ID NO: 2 or a genomic gene encoding cat G-CSF And may be produced by a chemical synthesis method or the like. Further, the base sequence of SEQ ID NO: 2 may be obtained by any method such as a chemical synthesis method, a DNA replication method, a reverse transcription method, and a transcription method.
- the protein having the biological activity of feline G-CSF of the above amino acid sequence (b) in which one or several amino acids of the amino acid sequence of SEQ ID NO: 1 has been deleted, substituted or added is represented by SEQ ID NO:
- the nucleotide corresponding to the amino acid deletion, substitution, or addition is encoded by a gene having a nucleotide sequence in which the nucleotide corresponding to the deletion, substitution, or addition of the amino acid is deleted, substituted, or added. May be used, or may be produced by a chemical synthesis method or the like.
- cat G-CSF protein a protein having the biological activity of cat G-CSF (hereinafter, simply referred to as “cat G-CSF protein”), As a treatment for neutropenia Used effectively.
- This cat G-CSF protein a specific antibody can be obtained by a known method.
- This antibody can be used to immunologically test for the activity of granulocyte colony-stimulating factor in cats. Therefore, it can be used as a sensor for detecting and diagnosing G-CSF levels in cats. it can. Further, it can be used for affinity mouth chromatography for separating and purifying the cat G-CSF protein from the protein produced by the following transformant.
- the above transformant can be produced, for example, as a cat G—CSF producing cell.
- a DNA fragment containing a cat G-CSF gene fragment having the amino acid sequence shown in SEQ ID NO: 1 is ligated to a promoter such as a virus, and the eukaryotic cell is transformed. Can be converted to produce cat G—CSF producing cells.
- a recombinant virus vector such as a baculovirus is used, for example, the method of Smith et al. (Smitheta 1., Mo 1. Cell. Biol., 3, p2156-21665) ), A recombinant virus vector can be created.
- a DNA fragment encoding the feline G-CSF protein was ligated to a baculovirus polynuclear promoter, the transfer vector was introduced into insect cells together with the baculovirus DNA, and the resulting recombinant baculovirus was A recombinant virus clone that produces the activity of cat G-CSF can be selected, and the recombinant clone can be infected into insect cells to produce cells that produce cat G-CSF protein.
- insects that produce feline G-CSF protein can be produced by infecting insects with the recombinant clone.
- the host cell is a prokaryotic cell
- a feline G—CSF protein-encoding DNA fragment is ligated to an expression vector, and the prokaryotic cell is transformed into a feline G— CSF-producing cells can be created.
- Gene manipulation technology can control the expression efficiency by, for example, adding or improving a signal sequence, suitably selecting a host-vector system, or improving a gene expression control site. Alternatively, it may be obtained as a sugar chain linked by selection of a host.
- the desired feline G-CSF protein By culturing the transformed cells and separating and recovering the produced protein, the desired feline G-CSF protein can be obtained.
- cat G—CSF protein can be produced in large quantities and can be used as a therapeutic agent for neutropenia in cats.
- FIG. 1 the amino acid sequences predicted from the nucleotide sequences of the human and mouse G—CSF genes and the cat G—CSF gene are listed together so that the homologous portion is maximized.
- the amino acid sequence is the same as the amino acid sequence of the human G—CSF gene, the deletion is indicated by “—”.
- the feline G—CSF protein having the amino acid sequence of SEQ ID NO: 1 is as follows: Can be obtained.
- genomic DNA extracted from CRFK (ATC C strain number CCL-94), which is a cat kidney-derived cell, was type II, and was prepared based on the gene sequence already reported in human 'mouse' dogs. Amplify the target gene region by PCR (polymerase reaction) using synthetic DNA primers. This is inserted into, for example, a commercially available vector to determine the nucleotide sequence.
- a genomic DNA region adjacent to the cloned region is amplified by the PCR method using a synthetic DNA primer prepared based on the determined nucleotide sequence, and the amplified DNA fragment is inserted into the above vector to determine the nucleotide sequence.
- the nucleotide sequence of the genomic DNA containing the cDNA region encoding the cat G—CSF protein can be determined.
- Cloning of cDNA can be performed as follows. That is, the above CRFK cells are stimulated with LPS (lipopolysaccharide), and the RNA of the cells is extracted. CDNA is prepared for the RNA, and a portion encoding cDNA protein is amplified by PCR using a synthetic DNA primer prepared based on the base sequence of genomic DNA.
- LPS lipopolysaccharide
- insect cells are infected with the recombinant baculovirus incorporated into the amplified cDNA fragment and expressed, and the biological activity of the obtained protein is confirmed to confirm the desired cat G—CSF protein protein. Can be confirmed.
- the cat kidney-derived epithelial cell CRFK (Hacchouji strain number 94) is a DMEM medium (Nissui Pharmaceutical Co., Ltd.) containing lOQ / o fetal calf serum (FCS; Gibeo BRL). was used for the culture.
- Preparation of genomic DNA was performed as follows. Discard the CRFK medium cultured in 26 plates of 9 cm Petri dish, wash with phosphate buffered saline (PB-S), remove cells with a scraper, and divide into 4 50 ml centrifuge tubes. Collected. The cells were suspended by adding 10 ml of PBS to each tube, and then centrifuged at 1,000 rpm for 10 minutes.
- PB-S phosphate buffered saline
- HMW buffer (10 mM Tris-HCl (pH 8.0), 15 OmM sodium chloride, 1 OmM ethylenediaminetetraacetic acid (pH 8.0), 15 ml of 1% sodium lauroyl sulfate) and protease K (100 g / m 1) were added, suspended, and heated at 37 ° C. to inactivate the protein. The mixture was extracted twice with phenol and once with phenol-black mouth. To the collected aqueous layer was added twice the volume of ethanol, mixed well, and cooled at 120 V for 2 hours. The pellet generated by centrifugation at 6000 rpm for 30 minutes at 4 ° C was suspended in 500 z1 of distilled water per tube.
- Hind5 '-1 5' -CGCCAAGCTTCAGAGCTTCCTGCTCAAGT-3 '
- Step 1 Remove paraffin oil from the product.
- the product of Step 1 was electrophoresed on a 1.5% agarose gel, stained with bromide medium, and then exposed to ultraviolet light at about 860 bp under a Gene Clean Kit Ver. (Biol 01, Inc.) according to the attached protocol.
- the excised DNA fragment was digested with restriction enzymes EcoRI and Hindill. (2-2) Cloning of PCR products
- a commercially available pUC18 vector is digested with EcoRI and Hindill, and ligated using the DNA fragment obtained in the above (2-1) and a Ligation kit manufactured by TAKARA. Was. Using this reaction solution, a transformation reaction was performed on Escherichia coli XL I-B 1 ue which had been made into a complex.
- the transformed Escherichia coli was selected on a 9 cm diameter agarose plate prepared in LB medium containing 100 g / m1 of ampicillin by adding 2% of 5-bromo-14-clo-mouth 3-indolyl- / 3-
- the test was performed on a plate coated with 201 each of D-galactoside and 100 mM isopropyl-1- ⁇ -D-thiogalactoside.
- the cells were cultured at 37 ° C, and white colonies were randomly selected from the colonies grown on the plate, and subsequently cultured in 2 ml of an LB medium containing 100 ⁇ g / m1 of ampicillin.
- Plasmid DNA was extracted and purified using QiagenPlamidM Midi Kit (Biol01, Inc.) according to the protocol attached.
- the purified plasmid DNA was digested with restriction enzymes EcoRI and Hindill to obtain a clone incorporating the DNA fragment of the desired size.
- nucleotide sequence was determined using the sequencer A (373) by the dideoxy sequence method using a cycle sequence kit manufactured by ABI.
- kitl 5 one CTCCACCACCCCTCTCCAGC one 3'
- kit2 5 '-CGCGGATCCTGCAGCGCAGTGCCATCAGCCTGG 1 3'
- kit3 5 one GGCCTCCTGCAGGCCCTGGC one 3'
- kit4 5 'one CGCGGATCCGCTGGACATCACCGACTTTGCTATC one 3'
- the modified part of the kit uses restriction enzymes (Sau 3A I from TAKARA, N 1 a III and T sp 509 I from New England Bio 1 ab) not used in the attached protocol as genome.
- restriction enzymes Sau 3A I from TAKARA, N 1 a III and T sp 509 I from New England Bio 1 ab
- the underline in the cassette indicates the part corresponding to the recognition sequence of each restriction enzyme.
- the underlined (*) of the cassette and the primer is the newly added restriction enzyme.
- the PCR product was excised by the method described in “(2-1)” and digested with restriction enzymes EcoRI and BamHI.
- kit5 5'-TTCCAGCGCCGGGCAGGAGG-3 '
- kit6 5 'one CGCGGATCCCTGCAGAGCTTCCTGGAGG one 3'
- a commercially available pGEM-3Zf (+) vector is digested with EcoRI and BamHI, and the PCR product obtained in the above (3-1) and the T4-DNA ligase are used. And performed a reign.
- the transformation reaction and selection of transformed Escherichia coli were performed by the method described in the above “(2-2)” to obtain a clone into which a DNA fragment of a desired size had been incorporated.
- the nucleotide sequence was determined by the method described in the above “(2-3)”. However, two or three clones were used for nucleotide sequence determination.
- RNA used the same CRFK as the material for the preparation of genomic DNA.
- Culture CRFK in three 25-cm plastic flasks until confluent, and add LPS from E. coli 0111 (manufactured by SI GMA) to a final concentration of 10 ⁇ g / m 1. And the medium was changed, and the cells were cultured at 37 ° C for 24 hours. After discarding the medium and washing with PBS, the D solution (4 M guanidine thiosinate, 25 mM sodium citrate (pH 7.0), 0.5% N-lauroyl sarcosine acid, 0.1 M 2-mercapto (Ethanol) was added at 1 ml per flask to lyse the cells.
- the 5 'START and 3' END primers have an additional restriction enzyme BamHI recognition sequence at the 5 'end, which is underlined.
- the reverse transcription reaction was carried out as follows using 5 g of the RNA prepared in the above (4) as Type I and Super Scriptll RNAaseH- reverse transcription enzyme manufactured by Gibco BRL. Was. That is, 5 ⁇ g of RNA and 10 pmo 1 of the 3 ′ RT primer were mixed, heated to a total volume of 121 with distilled water treated with DEPC, heated at 70 ° C. for 10 minutes, and then rapidly cooled on ice. Add 1 ⁇ l of 10 mM dNTP, 41 and 0.1 M of DTT to the reverse transcriptase buffer. Was added and mixed well, and reacted at 42 ° C for 55 minutes and at 70 ° C for 15 minutes.
- the obtained amplification product of about 470 bp cDNA was cut out by the method described in the above “(2-1)” and digested with the restriction enzyme BamHI.
- Noculovirus transfection vector pBac PAK1 (C1ontech) is digested with BamHI, treated with phosphatase, and then ligated by TAKARA. Using Kit version 2, according to the attached protocol, the cDNA was ligated with the cDNA obtained in the above “(5-1)”. A transformed reaction of E. coli XL I-B1ue transformed with this reaction solution was performed, and the transformed E. coli was selected using a 9 cm diameter agarose spray prepared on an LB medium containing 100 gZm1 of ampicillin. I went there. Clones inserted in the same direction with respect to polytransfer promoter all over the transfer vector were selected by PCR screening using Taq polymerase (Pharmacia).
- PCR is performed under the conditions shown in the above “(5-1)”, the following DNA primer having the corresponding nucleotide sequence in the cDNA, 3′-III, is synthesized, and the primer is oriented in the same direction as the polyhedrin promoter.
- a commercially available DNA primer having a nucleotide sequence corresponding to the vector side Bac-1 (Clontech).
- nucleotide sequence of SEQ ID NO: 2 are as follows.
- Organism name cat (f e r i d a e f e 1 i s c a t u s)
- the amino acid sequence deduced from the nucleotide sequence consists of 195 amino acids and corresponds to the higher type of human G-CSF activity, but the signal sequence was 9 amino acids less than that of the human mouse.
- high homology % was obtained as shown in Table 1 below. Therefore, This cDNA appeared to encode cat G-CSF.
- the upper row shows the nucleic acid sequence homology
- the lower row shows the amino acid sequence homology
- the nucleotide sequence of the feline genomic gene was also determined. That is, the liver of a 10-month-old cat is excised, cut into small pieces with surgical scissors, and contains 0.6% 308 containing 10% (10 mM Tris ⁇ hydrochloric acid (pH 8.0)-1 mM)
- EDTA EDTA
- proteinase K was added to 100 g / ml, and the protein was digested at 37 ° C with stirring. This mixture was subjected to phenol / chloroform extraction, the aqueous layer was taken, 10 mM 3M sodium acetate ( ⁇ 5.2) was added, and then 200 ml of ethanol was added and stirred. Genomic DNA floating as a precipitate was scooped with a glass rod, washed with 70% ethanol, air-dried, and dissolved in 10 ml of sterile distilled water.
- a DNA primer having a base sequence in the region adjacent to the start codon and the stop codon, 5 ′ OUT primer and 3OUTprimer were synthesized.
- the product amplified in the first PCR was used as a template and amplified in three parts (regions (1) to (3)) corresponding to the exon containing the protein-coding region.
- Table 2 shows the combinations of primers used for each region and the cycle conditions.
- Table 2 Primers used for amplification of each region and cycle conditions
- the 5 'OUT primer is the nucleotide sequence of "(5-3)”
- the Kit 2 primer is the nucleotide sequence of "(3-1)”
- the 3' RT primer is "(5-1)”. showed that.
- the nucleotide sequences of the other primers are as follows.
- Each product of the second PCR was electrophoresed on a 1.5% agarose gel, stained with bromide thidium, and the compound was subjected to Gene Clean S in Kit (Bio101 Inc. ) And cut out according to the attached protocol. After phosphorylating the end of the excised DNA fragment, it was inserted into the Sma1 site of a commercially available pGEN-3Z (+) vector.
- This reaction mixture was mixed with the competent E. coli XLI-B1ue and transformed.
- Clones grown on an LB plate containing 100 / gZm1 of ampicillin were cultured in 5 ml of LB medium containing 10 Q ⁇ g / m1 of ampicillin, and then Qiagen Plasmid Midi K Extract and purify the plasmid DNA by operating it according to the attached protocol and digest it with the restriction enzymes EcoR1 and Sa11 to obtain a clone incorporating the DNA fragment of the desired size. Obtained.
- 2 clones incorporating the DNA fragment of the region (1) or (3) and 4 clones incorporating the DNA fragment of the region (2) were selected, and the nucleotides were obtained by the method described in the above “(2-3)”. The sequence was determined.
- amino acid sequence of Sequence 2 is also the amino acid sequence of the cat genome gene of G—CSF.
- Sf21AE cells which are insect cells, were used as host cells for baculovirus.
- Sf21E cells were cultured in Grace's medium (Gibco BRL) containing 10% fetal bovine serum (FBS), 1% bactotryptosebroth (DIFC0), and 50 g / m1 kanamycin. did.
- FBS fetal bovine serum
- DIFC0 1% bactotryptosebroth
- 50 g / m1 kanamycin 50 g / m1 kanamycin.
- the recombinant virus was cloned by the black method from the virus that appeared in the culture supernatant.
- Cells infected with wild-type baculovirus that have not undergone recombination can be distinguished by the formation of nucleopolynucleotides. That is, after three days from the transflector We click Chillon, culture supernatants were diluted 10000-fold with the medium, the virus diluted into S f 21 AE cells that had been seeded as in 6-well play Bok a lxl 0 6 cell sZwe ll Infection was carried out for 1 hour after adding 200 1 of diluent.
- a medium containing 1% Seaplaque agarose (manufactured by FMC Bio Products) kept at 37 ° C was added, and the mixture was allowed to stand at room temperature for 1 hour to solidify. From above, 1 ml of medium was added and cultured at 27 ° C for 4 days. After confirming that black was formed, PBS containing 0.01% neutral red was added dropwise. The cells were cultured overnight at 27 ° C, and the black portions of the colorless areas were marked after confirming that no nucleus polynuclear was formed. The marked black was removed with a Pasteur pipet, suspended in 5001 medium, and further sonicated to destroy agarose. Agarose was precipitated by centrifugation, and the supernatant was diluted and used for the next black cloning. The same process is repeated two more times, for a total of three times To obtain a recombinant baculovirus.
- Seaplaque agarose manufactured by FMC Bio Products
- the recombinant baculovirus was infected to Sf21AE cells at m.o.i. (multipilicitoyofinfection): about 10 and the culture supernatant was collected 4 days later.
- a culture supernatant was prepared in the same manner as described above, which was infected with Autogr ap a cab l a l i f o r n i c a u c l e a r P o l yh e d o r o s i s vi ir u s (hereinafter abbreviated as AcNPV) as a negative control.
- AcNPV Autogr ap a cab l a l i f o r n i c a u c l e a r P o l yh e d o r o s i s vi ir u
- feline G—CSF The biological activity of feline G—CSF was examined by bioassay using mouse bone marrow-like cells NFS-60, whose proliferation is promoted by mouse G—CSF.
- This quantification system is not a system using cat cells, but if the species specificity of the target cytokine, such as the lymphokine, is low, the biological activity can be measured using the Atssey system used in different species. Is common. For example, Interleukin-1 (Howa rdeta 1., 1 982, J. Exp. Med., 155, 914) and Interleukin-7 (Conloneta 1., 1 989, Blood, 74) , 1 368), and the same NFS-60 cells are used to measure the activity of human G-CSF.
- NF S-60 cells are RPMI 1640 medium (Nissui Pharmaceutical Co., Ltd.) containing 10% FCS and recombinant mouse interleukin 3 (hereinafter abbreviated as ml L-3; manufactured by Pepro Tech EC LTD.). (Manufactured by the company).
- the bioassay was carried out as follows using a cell titer 96-akua skit manufactured by Prmea. That is, the NF S- 60 cells were harvested by centrifugation, L such include a m IL- 3, washed twice with medium and prepared so that the cells number 2 X 1 0 5 Zm 1. This was spread on a 100-well 96-well plate, and the recombinant human G-CSF (manufactured by Gibco BRL) or the culture supernatant sample obtained in the above (6) was serially diluted. Each of the cells was added at a time, and cultured for 24 hours. The assay was performed in duplicate for dilution.
- a protein having the activity of G-CSF can be obtained, and the pharmaceutical composition can be used as a therapeutic agent for neutropenia in cats according to claims 5 and 6.
- the treatment can be provided.
- a gene encoding the G-CSF protein of the cat is obtained, and the gene is expressed by the genetic engineering method of claims 3, 4, and 7 using this gene.
- the gene is expressed by the genetic engineering method of claims 3, 4, and 7 using this gene.
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Medicinal Chemistry (AREA)
- Immunology (AREA)
- Organic Chemistry (AREA)
- Zoology (AREA)
- Gastroenterology & Hepatology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- General Health & Medical Sciences (AREA)
- Molecular Biology (AREA)
- Pharmacology & Pharmacy (AREA)
- Biophysics (AREA)
- Biochemistry (AREA)
- Toxicology (AREA)
- Engineering & Computer Science (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Genetics & Genomics (AREA)
- Epidemiology (AREA)
- Animal Behavior & Ethology (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Peptides Or Proteins (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
Description
Claims
Priority Applications (4)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
EP98950363A EP1033373B1 (en) | 1997-10-23 | 1998-10-23 | Feline granulocyte colony-stimulating factor |
DE69834938T DE69834938T2 (de) | 1997-10-23 | 1998-10-23 | Granulozyten-kolonie-stimulierender faktor aus der katze |
AU96471/98A AU9647198A (en) | 1997-10-23 | 1998-10-23 | Feline granulocyte colony-stimulating factor |
US09/530,095 US6610515B1 (en) | 1997-10-23 | 1998-10-23 | Feline granulocyte colony-stimulating factor |
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
JP29105597 | 1997-10-23 | ||
JP9/291055 | 1997-10-23 |
Publications (1)
Publication Number | Publication Date |
---|---|
WO1999020652A1 true WO1999020652A1 (fr) | 1999-04-29 |
Family
ID=17763854
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
PCT/JP1998/004809 WO1999020652A1 (fr) | 1997-10-23 | 1998-10-23 | Facteur stimulant les colonies de granulocytes de felins |
Country Status (6)
Country | Link |
---|---|
US (1) | US6610515B1 (ja) |
EP (1) | EP1033373B1 (ja) |
AT (1) | ATE329928T1 (ja) |
AU (1) | AU9647198A (ja) |
DE (1) | DE69834938T2 (ja) |
WO (1) | WO1999020652A1 (ja) |
Cited By (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2021205401A1 (en) * | 2020-04-09 | 2021-10-14 | Aprilbio Co., Ltd. | Recombinant proteins comprising feline granulocyte colony-stimulating factor and antigen binding fragment for serum albumin, and uses thereof |
Families Citing this family (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US7138691B2 (en) * | 2004-01-22 | 2006-11-21 | International Business Machines Corporation | Selective nitridation of gate oxides |
Citations (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JPS62269693A (ja) * | 1986-05-19 | 1987-11-24 | Chugai Pharmaceut Co Ltd | 新規dna |
WO1991005798A1 (en) * | 1989-10-10 | 1991-05-02 | Amgen Inc. | Compositions and methods for treating or preventing infections in canine and feline animals |
Family Cites Families (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JP3004513B2 (ja) | 1993-09-30 | 2000-01-31 | ゼオン化成株式会社 | 粉体成形用組成物 |
PE20000183A1 (es) * | 1997-07-25 | 2000-03-11 | Schering Corp | Citoquinas de mamiferos y reactivos relacionados |
KR20010085850A (ko) * | 1998-09-29 | 2001-09-07 | 추후제출 | 코돈 변형 유전자의 재편성 |
-
1998
- 1998-10-23 AT AT98950363T patent/ATE329928T1/de not_active IP Right Cessation
- 1998-10-23 US US09/530,095 patent/US6610515B1/en not_active Expired - Fee Related
- 1998-10-23 DE DE69834938T patent/DE69834938T2/de not_active Expired - Lifetime
- 1998-10-23 AU AU96471/98A patent/AU9647198A/en not_active Abandoned
- 1998-10-23 WO PCT/JP1998/004809 patent/WO1999020652A1/ja active IP Right Grant
- 1998-10-23 EP EP98950363A patent/EP1033373B1/en not_active Expired - Lifetime
Patent Citations (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JPS62269693A (ja) * | 1986-05-19 | 1987-11-24 | Chugai Pharmaceut Co Ltd | 新規dna |
WO1991005798A1 (en) * | 1989-10-10 | 1991-05-02 | Amgen Inc. | Compositions and methods for treating or preventing infections in canine and feline animals |
Non-Patent Citations (1)
Title |
---|
NAGATA S., ET AL.: "THE CHROMOSOMAL GENE STRUCTURE AND TWO MRNAS FOR HUMAN GRANYLOCYTE COLONY-STIMULATING FACTOR.", EMBO JOURNAL., OXFORD UNIVERSITY PRESS, SURREY., GB, vol. 05., no. 03., 1 January 1986 (1986-01-01), GB, pages 575 - 581., XP002918598, ISSN: 0261-4189 * |
Cited By (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
WO2021205401A1 (en) * | 2020-04-09 | 2021-10-14 | Aprilbio Co., Ltd. | Recombinant proteins comprising feline granulocyte colony-stimulating factor and antigen binding fragment for serum albumin, and uses thereof |
Also Published As
Publication number | Publication date |
---|---|
AU9647198A (en) | 1999-05-10 |
ATE329928T1 (de) | 2006-07-15 |
DE69834938T2 (de) | 2007-02-01 |
EP1033373B1 (en) | 2006-06-14 |
US6610515B1 (en) | 2003-08-26 |
DE69834938D1 (de) | 2006-07-27 |
EP1033373A4 (en) | 2002-09-25 |
EP1033373A1 (en) | 2000-09-06 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
KR920010225B1 (ko) | 인간암괴사인자 및 그를 코우드하는 dna의 제법 | |
JP4308916B2 (ja) | 異種タンパク質の発現 | |
US20030125538A1 (en) | Amino acid transporters and uses | |
JP4191255B2 (ja) | ケモカイン類に結合する可溶性ワクシニアウイルスタンパク質 | |
EP0784689A1 (en) | Microorganisms as therapeutic delivery systems | |
CA2312000A1 (en) | Mammalian cytokine-like polypeptide-10 | |
JPH10508742A (ja) | ヒトケモカインポリペプチド | |
Cochet et al. | Novel interferon delta genes in mammals: cloning of one gene from the sheep, two genes expressed by the horse conceptus and discovery of related sequences in several taxa by genomic database screening | |
JPS6124599A (ja) | 新規ヒトインタ−フエロン−rポリペプチド誘導体 | |
JPH0641198A (ja) | 抗血液凝固タンパク質とその製造法 | |
US7618635B2 (en) | Super-antigen fusion proteins and the use thereof | |
KR20230172527A (ko) | 범용 클로닝 어댑터를 함유하는 알파바이러스 벡터 | |
JPS6270398A (ja) | ウシインタ−ロイキン−2遺伝子のクロ−ニングおよび同定 | |
HU205617B (en) | Process for producing neturophyl-activating factor, gene coding this and pharmaceutical composition | |
US20080045699A1 (en) | Interleukin-1 Related Gene and Protein | |
WO1999020652A1 (fr) | Facteur stimulant les colonies de granulocytes de felins | |
EP0349569A1 (en) | BEEF INTERLEUKIN 1 g (b). | |
EP1470154B1 (en) | Peptides and nucleic acids of the cathelicidin family, derived from fish, and uses thereof | |
JP2007228814A (ja) | 外来タンパク質の発現用遺伝子カセットおよび外来タンパク質の製造方法 | |
CA2295957C (en) | Canine interleukin 18, canine interleukin 1.beta. converting enzyme, dna sequences encoding these, interleukin 18 production method and canine immune disease remedy | |
WO2002081519A9 (fr) | Nouvelle proteine hybride ifn-thy, preparation et utilisation | |
CN108794645A (zh) | 一种由牛白蛋白、牛干扰素γ和牛干扰素α组成的融合蛋白及其制备方法 | |
JPH11196881A (ja) | 猫顆粒球コロニー刺激因子 | |
Hook et al. | Cloning and expression of the cervine interleukin 4 gene | |
KR100802140B1 (ko) | 돼지 유래 신규 adam3 유전자 및 이를 이용한 돼지정자의 수정능 판별방법 |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
AK | Designated states |
Kind code of ref document: A1 Designated state(s): AU CA KR US |
|
AL | Designated countries for regional patents |
Kind code of ref document: A1 Designated state(s): AT BE CH CY DE DK ES FI FR GB GR IE IT LU MC NL PT SE |
|
121 | Ep: the epo has been informed by wipo that ep was designated in this application | ||
DFPE | Request for preliminary examination filed prior to expiration of 19th month from priority date (pct application filed before 20040101) | ||
WWE | Wipo information: entry into national phase |
Ref document number: 1998950363 Country of ref document: EP |
|
NENP | Non-entry into the national phase |
Ref country code: KR |
|
WWE | Wipo information: entry into national phase |
Ref document number: 09530095 Country of ref document: US |
|
WWP | Wipo information: published in national office |
Ref document number: 1998950363 Country of ref document: EP |
|
NENP | Non-entry into the national phase |
Ref country code: CA |
|
WWG | Wipo information: grant in national office |
Ref document number: 1998950363 Country of ref document: EP |