TWI421093B - 胜肽-聚合物綴合物 - Google Patents
胜肽-聚合物綴合物 Download PDFInfo
- Publication number
- TWI421093B TWI421093B TW098125858A TW98125858A TWI421093B TW I421093 B TWI421093 B TW I421093B TW 098125858 A TW098125858 A TW 098125858A TW 98125858 A TW98125858 A TW 98125858A TW I421093 B TWI421093 B TW I421093B
- Authority
- TW
- Taiwan
- Prior art keywords
- conjugate
- moiety
- peptide
- inf
- polymer
- Prior art date
Links
- 229920000642 polymer Polymers 0.000 title claims description 49
- 229920001223 polyethylene glycol Polymers 0.000 claims description 27
- OXCMYAYHXIHQOA-UHFFFAOYSA-N potassium;[2-butyl-5-chloro-3-[[4-[2-(1,2,4-triaza-3-azanidacyclopenta-1,4-dien-5-yl)phenyl]phenyl]methyl]imidazol-4-yl]methanol Chemical group [K+].CCCCC1=NC(Cl)=C(CO)N1CC1=CC=C(C=2C(=CC=CC=2)C2=N[N-]N=N2)C=C1 OXCMYAYHXIHQOA-UHFFFAOYSA-N 0.000 claims description 24
- 108010051696 Growth Hormone Chemical group 0.000 claims description 15
- 102000018997 Growth Hormone Human genes 0.000 claims description 15
- 239000000122 growth hormone Chemical group 0.000 claims description 15
- 125000000524 functional group Chemical group 0.000 claims description 12
- 229910052799 carbon Inorganic materials 0.000 claims description 11
- 239000002202 Polyethylene glycol Substances 0.000 claims description 10
- 125000003118 aryl group Chemical group 0.000 claims description 9
- 108090000467 Interferon-beta Proteins 0.000 claims description 8
- 102000003996 Interferon-beta Human genes 0.000 claims description 8
- 125000000304 alkynyl group Chemical group 0.000 claims description 8
- 229960001388 interferon-beta Drugs 0.000 claims description 8
- 125000000539 amino acid group Chemical group 0.000 claims description 7
- 125000001072 heteroaryl group Chemical group 0.000 claims description 7
- 125000000217 alkyl group Chemical group 0.000 claims description 6
- 125000004432 carbon atom Chemical group C* 0.000 claims description 6
- 229920001427 mPEG Polymers 0.000 claims description 6
- 125000006374 C2-C10 alkenyl group Chemical group 0.000 claims description 5
- 125000000592 heterocycloalkyl group Chemical group 0.000 claims description 5
- 125000004433 nitrogen atom Chemical group N* 0.000 claims description 5
- 125000006552 (C3-C8) cycloalkyl group Chemical group 0.000 claims description 4
- 229910052757 nitrogen Inorganic materials 0.000 claims description 4
- 229910052739 hydrogen Inorganic materials 0.000 claims description 3
- 125000003827 glycol group Chemical group 0.000 claims 5
- 239000000562 conjugate Substances 0.000 description 50
- 239000000243 solution Substances 0.000 description 26
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 22
- 108090000623 proteins and genes Proteins 0.000 description 21
- 102000003951 Erythropoietin Human genes 0.000 description 19
- 108090000394 Erythropoietin Proteins 0.000 description 19
- 239000000203 mixture Substances 0.000 description 18
- 235000018102 proteins Nutrition 0.000 description 18
- 102000004169 proteins and genes Human genes 0.000 description 18
- 239000000872 buffer Substances 0.000 description 16
- 229940105423 erythropoietin Drugs 0.000 description 16
- 238000000034 method Methods 0.000 description 15
- 108090000765 processed proteins & peptides Proteins 0.000 description 14
- 108010013127 Met-human growth hormone Proteins 0.000 description 13
- 238000002360 preparation method Methods 0.000 description 13
- -1 tributyl Chemical group 0.000 description 13
- 150000001299 aldehydes Chemical class 0.000 description 12
- 210000004027 cell Anatomy 0.000 description 12
- 241000588724 Escherichia coli Species 0.000 description 11
- 241000700159 Rattus Species 0.000 description 11
- 239000011541 reaction mixture Substances 0.000 description 11
- 239000011780 sodium chloride Substances 0.000 description 11
- 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 10
- 150000001413 amino acids Chemical group 0.000 description 10
- 210000002966 serum Anatomy 0.000 description 10
- 239000000725 suspension Substances 0.000 description 9
- 238000010521 absorption reaction Methods 0.000 description 8
- 208000015181 infectious disease Diseases 0.000 description 8
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 8
- 208000007502 anemia Diseases 0.000 description 7
- 210000004369 blood Anatomy 0.000 description 7
- 239000008280 blood Substances 0.000 description 7
- 150000002430 hydrocarbons Chemical class 0.000 description 7
- 210000003000 inclusion body Anatomy 0.000 description 7
- 239000002243 precursor Substances 0.000 description 7
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 description 6
- 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 6
- 238000002965 ELISA Methods 0.000 description 6
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 6
- 241000711549 Hepacivirus C Species 0.000 description 6
- HEMHJVSKTPXQMS-UHFFFAOYSA-M Sodium hydroxide Chemical compound [OH-].[Na+] HEMHJVSKTPXQMS-UHFFFAOYSA-M 0.000 description 6
- 238000003756 stirring Methods 0.000 description 6
- 239000006228 supernatant Substances 0.000 description 6
- 238000003786 synthesis reaction Methods 0.000 description 6
- 239000004215 Carbon black (E152) Substances 0.000 description 5
- 108090000790 Enzymes Proteins 0.000 description 5
- 102000004190 Enzymes Human genes 0.000 description 5
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 5
- 241000700721 Hepatitis B virus Species 0.000 description 5
- 125000003342 alkenyl group Chemical group 0.000 description 5
- 238000005119 centrifugation Methods 0.000 description 5
- 150000001875 compounds Chemical class 0.000 description 5
- 238000000855 fermentation Methods 0.000 description 5
- 230000004151 fermentation Effects 0.000 description 5
- 239000008103 glucose Substances 0.000 description 5
- 229930195733 hydrocarbon Natural products 0.000 description 5
- 239000002609 medium Substances 0.000 description 5
- 239000008188 pellet Substances 0.000 description 5
- 239000000863 peptide conjugate Substances 0.000 description 5
- 125000006239 protecting group Chemical group 0.000 description 5
- 239000002904 solvent Substances 0.000 description 5
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical compound [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 4
- 108010000521 Human Growth Hormone Proteins 0.000 description 4
- 102000002265 Human Growth Hormone Human genes 0.000 description 4
- 239000000854 Human Growth Hormone Substances 0.000 description 4
- NQRYJNQNLNOLGT-UHFFFAOYSA-N Piperidine Chemical compound C1CCNCC1 NQRYJNQNLNOLGT-UHFFFAOYSA-N 0.000 description 4
- 102000007056 Recombinant Fusion Proteins Human genes 0.000 description 4
- 108010008281 Recombinant Fusion Proteins Proteins 0.000 description 4
- VMHLLURERBWHNL-UHFFFAOYSA-M Sodium acetate Chemical compound [Na+].CC([O-])=O VMHLLURERBWHNL-UHFFFAOYSA-M 0.000 description 4
- XSQUKJJJFZCRTK-UHFFFAOYSA-N Urea Chemical compound NC(N)=O XSQUKJJJFZCRTK-UHFFFAOYSA-N 0.000 description 4
- 239000004480 active ingredient Substances 0.000 description 4
- 125000003277 amino group Chemical group 0.000 description 4
- BFNBIHQBYMNNAN-UHFFFAOYSA-N ammonium sulfate Chemical compound N.N.OS(O)(=O)=O BFNBIHQBYMNNAN-UHFFFAOYSA-N 0.000 description 4
- 229910052921 ammonium sulfate Inorganic materials 0.000 description 4
- 235000011130 ammonium sulphate Nutrition 0.000 description 4
- 230000015572 biosynthetic process Effects 0.000 description 4
- 229940041514 candida albicans extract Drugs 0.000 description 4
- 150000001721 carbon Chemical group 0.000 description 4
- WIIZWVCIJKGZOK-RKDXNWHRSA-N chloramphenicol Chemical compound ClC(Cl)C(=O)N[C@H](CO)[C@H](O)C1=CC=C([N+]([O-])=O)C=C1 WIIZWVCIJKGZOK-RKDXNWHRSA-N 0.000 description 4
- 229960005091 chloramphenicol Drugs 0.000 description 4
- BPHPUYQFMNQIOC-NXRLNHOXSA-N isopropyl beta-D-thiogalactopyranoside Chemical compound CC(C)S[C@@H]1O[C@H](CO)[C@H](O)[C@H](O)[C@H]1O BPHPUYQFMNQIOC-NXRLNHOXSA-N 0.000 description 4
- 229930027917 kanamycin Natural products 0.000 description 4
- 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 4
- 229960000318 kanamycin Drugs 0.000 description 4
- 229930182823 kanamycin A Natural products 0.000 description 4
- 125000005647 linker group Chemical group 0.000 description 4
- HQKMJHAJHXVSDF-UHFFFAOYSA-L magnesium stearate Chemical compound [Mg+2].CCCCCCCCCCCCCCCCCC([O-])=O.CCCCCCCCCCCCCCCCCC([O-])=O HQKMJHAJHXVSDF-UHFFFAOYSA-L 0.000 description 4
- 238000002731 protein assay Methods 0.000 description 4
- 150000003839 salts Chemical class 0.000 description 4
- 239000001632 sodium acetate Substances 0.000 description 4
- 235000017281 sodium acetate Nutrition 0.000 description 4
- 239000012064 sodium phosphate buffer Substances 0.000 description 4
- 230000001225 therapeutic effect Effects 0.000 description 4
- 239000012138 yeast extract Substances 0.000 description 4
- WVDDGKGOMKODPV-UHFFFAOYSA-N Benzyl alcohol Chemical compound OCC1=CC=CC=C1 WVDDGKGOMKODPV-UHFFFAOYSA-N 0.000 description 3
- LYCAIKOWRPUZTN-UHFFFAOYSA-N Ethylene glycol Chemical compound OCCO LYCAIKOWRPUZTN-UHFFFAOYSA-N 0.000 description 3
- 238000005481 NMR spectroscopy Methods 0.000 description 3
- 239000011543 agarose gel Substances 0.000 description 3
- 235000013877 carbamide Nutrition 0.000 description 3
- 238000006243 chemical reaction Methods 0.000 description 3
- 239000003153 chemical reaction reagent Substances 0.000 description 3
- 239000003795 chemical substances by application Substances 0.000 description 3
- KRKNYBCHXYNGOX-UHFFFAOYSA-N citric acid Chemical compound OC(=O)CC(O)(C(O)=O)CC(O)=O KRKNYBCHXYNGOX-UHFFFAOYSA-N 0.000 description 3
- 238000000502 dialysis Methods 0.000 description 3
- 239000003085 diluting agent Substances 0.000 description 3
- 239000002270 dispersing agent Substances 0.000 description 3
- 210000003743 erythrocyte Anatomy 0.000 description 3
- 238000005227 gel permeation chromatography Methods 0.000 description 3
- 239000007788 liquid Substances 0.000 description 3
- 125000002950 monocyclic group Chemical group 0.000 description 3
- 230000004766 neurogenesis Effects 0.000 description 3
- 229910052760 oxygen Inorganic materials 0.000 description 3
- 210000002706 plastid Anatomy 0.000 description 3
- 229920000233 poly(alkylene oxides) Polymers 0.000 description 3
- 229920001184 polypeptide Polymers 0.000 description 3
- 239000002244 precipitate Substances 0.000 description 3
- 238000011552 rat model Methods 0.000 description 3
- 230000002829 reductive effect Effects 0.000 description 3
- 239000007974 sodium acetate buffer Substances 0.000 description 3
- 239000007787 solid Substances 0.000 description 3
- 229910052717 sulfur Inorganic materials 0.000 description 3
- PUPZLCDOIYMWBV-UHFFFAOYSA-N (+/-)-1,3-Butanediol Chemical compound CC(O)CCO PUPZLCDOIYMWBV-UHFFFAOYSA-N 0.000 description 2
- 238000000035 BCA protein assay Methods 0.000 description 2
- KRKNYBCHXYNGOX-UHFFFAOYSA-K Citrate Chemical compound [O-]C(=O)CC(O)(CC([O-])=O)C([O-])=O KRKNYBCHXYNGOX-UHFFFAOYSA-K 0.000 description 2
- RGSFGYAAUTVSQA-UHFFFAOYSA-N Cyclopentane Chemical compound C1CCCC1 RGSFGYAAUTVSQA-UHFFFAOYSA-N 0.000 description 2
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N DMSO Substances CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 2
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 2
- 108010005716 Interferon beta-1a Proteins 0.000 description 2
- 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 2
- JGFZNNIVVJXRND-UHFFFAOYSA-N N,N-Diisopropylethylamine (DIPEA) Chemical compound CCN(C(C)C)C(C)C JGFZNNIVVJXRND-UHFFFAOYSA-N 0.000 description 2
- 108090000854 Oxidoreductases Proteins 0.000 description 2
- 102000004316 Oxidoreductases Human genes 0.000 description 2
- KDLHZDBZIXYQEI-UHFFFAOYSA-N Palladium Chemical compound [Pd] KDLHZDBZIXYQEI-UHFFFAOYSA-N 0.000 description 2
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 description 2
- 239000004698 Polyethylene Substances 0.000 description 2
- 239000007983 Tris buffer Substances 0.000 description 2
- 125000004036 acetal group Chemical group 0.000 description 2
- DHKHKXVYLBGOIT-UHFFFAOYSA-N acetaldehyde Diethyl Acetal Natural products CCOC(C)OCC DHKHKXVYLBGOIT-UHFFFAOYSA-N 0.000 description 2
- 150000001241 acetals Chemical class 0.000 description 2
- 239000002253 acid Substances 0.000 description 2
- 150000001412 amines Chemical class 0.000 description 2
- 235000001014 amino acid Nutrition 0.000 description 2
- 150000001450 anions Chemical class 0.000 description 2
- 125000002178 anthracenyl group Chemical group C1(=CC=CC2=CC3=CC=CC=C3C=C12)* 0.000 description 2
- 239000007900 aqueous suspension Substances 0.000 description 2
- 125000001584 benzyloxycarbonyl group Chemical group C(=O)(OCC1=CC=CC=C1)* 0.000 description 2
- 239000002775 capsule Substances 0.000 description 2
- 239000004202 carbamide Substances 0.000 description 2
- PVAWKBXCZQBVLC-UHFFFAOYSA-N carbonic acid;1-hydroxypyrrolidine-2,5-dione Chemical compound OC(O)=O.ON1C(=O)CCC1=O PVAWKBXCZQBVLC-UHFFFAOYSA-N 0.000 description 2
- 125000002915 carbonyl group Chemical group [*:2]C([*:1])=O 0.000 description 2
- 150000007942 carboxylates Chemical class 0.000 description 2
- 239000000969 carrier Substances 0.000 description 2
- 239000003054 catalyst Substances 0.000 description 2
- 150000001768 cations Chemical class 0.000 description 2
- 238000003776 cleavage reaction Methods 0.000 description 2
- 125000000753 cycloalkyl group Chemical group 0.000 description 2
- 150000004985 diamines Chemical class 0.000 description 2
- 201000010099 disease Diseases 0.000 description 2
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 2
- 238000004090 dissolution Methods 0.000 description 2
- 239000002552 dosage form Substances 0.000 description 2
- 239000003814 drug Substances 0.000 description 2
- 239000003995 emulsifying agent Substances 0.000 description 2
- 239000000839 emulsion Substances 0.000 description 2
- 239000003623 enhancer Substances 0.000 description 2
- 239000001963 growth medium Substances 0.000 description 2
- 125000005842 heteroatom Chemical group 0.000 description 2
- 238000004128 high performance liquid chromatography Methods 0.000 description 2
- 229940088597 hormone Drugs 0.000 description 2
- 239000005556 hormone Substances 0.000 description 2
- 238000005984 hydrogenation reaction Methods 0.000 description 2
- 230000002209 hydrophobic effect Effects 0.000 description 2
- 230000028993 immune response Effects 0.000 description 2
- 229940124622 immune-modulator drug Drugs 0.000 description 2
- 238000001727 in vivo Methods 0.000 description 2
- 230000003993 interaction Effects 0.000 description 2
- 238000007917 intracranial administration Methods 0.000 description 2
- 210000003734 kidney Anatomy 0.000 description 2
- 239000008101 lactose Substances 0.000 description 2
- 235000019359 magnesium stearate Nutrition 0.000 description 2
- 238000004519 manufacturing process Methods 0.000 description 2
- BQJCRHHNABKAKU-KBQPJGBKSA-N morphine Chemical compound O([C@H]1[C@H](C=C[C@H]23)O)C4=C5[C@@]12CCN(C)[C@@H]3CC5=CC=C4O BQJCRHHNABKAKU-KBQPJGBKSA-N 0.000 description 2
- 231100000252 nontoxic Toxicity 0.000 description 2
- 230000003000 nontoxic effect Effects 0.000 description 2
- 239000003921 oil Substances 0.000 description 2
- 235000019198 oils Nutrition 0.000 description 2
- 125000004430 oxygen atom Chemical group O* 0.000 description 2
- 239000008194 pharmaceutical composition Substances 0.000 description 2
- 229920000573 polyethylene Polymers 0.000 description 2
- 239000000843 powder Substances 0.000 description 2
- 239000000047 product Substances 0.000 description 2
- 238000005932 reductive alkylation reaction Methods 0.000 description 2
- 230000007017 scission Effects 0.000 description 2
- 208000037921 secondary disease Diseases 0.000 description 2
- 239000011734 sodium Substances 0.000 description 2
- BEOOHQFXGBMRKU-UHFFFAOYSA-N sodium cyanoborohydride Chemical compound [Na+].[B-]C#N BEOOHQFXGBMRKU-UHFFFAOYSA-N 0.000 description 2
- 239000001488 sodium phosphate Substances 0.000 description 2
- 229910000162 sodium phosphate Inorganic materials 0.000 description 2
- 238000010189 synthetic method Methods 0.000 description 2
- LENZDBCJOHFCAS-UHFFFAOYSA-N tris Chemical compound OCC(N)(CO)CO LENZDBCJOHFCAS-UHFFFAOYSA-N 0.000 description 2
- RYFMWSXOAZQYPI-UHFFFAOYSA-K trisodium phosphate Chemical compound [Na+].[Na+].[Na+].[O-]P([O-])([O-])=O RYFMWSXOAZQYPI-UHFFFAOYSA-K 0.000 description 2
- 238000000108 ultra-filtration Methods 0.000 description 2
- QFLWZFQWSBQYPS-AWRAUJHKSA-N (3S)-3-[[(2S)-2-[[(2S)-2-[5-[(3aS,6aR)-2-oxo-1,3,3a,4,6,6a-hexahydrothieno[3,4-d]imidazol-4-yl]pentanoylamino]-3-methylbutanoyl]amino]-3-(4-hydroxyphenyl)propanoyl]amino]-4-[1-bis(4-chlorophenoxy)phosphorylbutylamino]-4-oxobutanoic acid Chemical compound CCCC(NC(=O)[C@H](CC(O)=O)NC(=O)[C@H](Cc1ccc(O)cc1)NC(=O)[C@@H](NC(=O)CCCCC1SC[C@@H]2NC(=O)N[C@H]12)C(C)C)P(=O)(Oc1ccc(Cl)cc1)Oc1ccc(Cl)cc1 QFLWZFQWSBQYPS-AWRAUJHKSA-N 0.000 description 1
- 125000000027 (C1-C10) alkoxy group Chemical group 0.000 description 1
- 125000000008 (C1-C10) alkyl group Chemical group 0.000 description 1
- 125000000081 (C5-C8) cycloalkenyl group Chemical group 0.000 description 1
- WRIDQFICGBMAFQ-UHFFFAOYSA-N (E)-8-Octadecenoic acid Natural products CCCCCCCCCC=CCCCCCCC(O)=O WRIDQFICGBMAFQ-UHFFFAOYSA-N 0.000 description 1
- DXIYRKVXLUOOON-UHFFFAOYSA-N (hydroxyamino)phosphonic acid Chemical compound ONP(O)(O)=O DXIYRKVXLUOOON-UHFFFAOYSA-N 0.000 description 1
- UNILWMWFPHPYOR-KXEYIPSPSA-M 1-[6-[2-[3-[3-[3-[2-[2-[3-[[2-[2-[[(2r)-1-[[2-[[(2r)-1-[3-[2-[2-[3-[[2-(2-amino-2-oxoethoxy)acetyl]amino]propoxy]ethoxy]ethoxy]propylamino]-3-hydroxy-1-oxopropan-2-yl]amino]-2-oxoethyl]amino]-3-[(2r)-2,3-di(hexadecanoyloxy)propyl]sulfanyl-1-oxopropan-2-yl Chemical compound O=C1C(SCCC(=O)NCCCOCCOCCOCCCNC(=O)COCC(=O)N[C@@H](CSC[C@@H](COC(=O)CCCCCCCCCCCCCCC)OC(=O)CCCCCCCCCCCCCCC)C(=O)NCC(=O)N[C@H](CO)C(=O)NCCCOCCOCCOCCCNC(=O)COCC(N)=O)CC(=O)N1CCNC(=O)CCCCCN\1C2=CC=C(S([O-])(=O)=O)C=C2CC/1=C/C=C/C=C/C1=[N+](CC)C2=CC=C(S([O-])(=O)=O)C=C2C1 UNILWMWFPHPYOR-KXEYIPSPSA-M 0.000 description 1
- LQJBNNIYVWPHFW-UHFFFAOYSA-N 20:1omega9c fatty acid Natural products CCCCCCCCCCC=CCCCCCCCC(O)=O LQJBNNIYVWPHFW-UHFFFAOYSA-N 0.000 description 1
- QSBYPNXLFMSGKH-UHFFFAOYSA-N 9-Heptadecensaeure Natural products CCCCCCCC=CCCCCCCCC(O)=O QSBYPNXLFMSGKH-UHFFFAOYSA-N 0.000 description 1
- 239000004475 Arginine Substances 0.000 description 1
- 238000009010 Bradford assay Methods 0.000 description 1
- 239000012619 Butyl Sepharose® Substances 0.000 description 1
- 125000005865 C2-C10alkynyl group Chemical group 0.000 description 1
- BHPQYMZQTOCNFJ-UHFFFAOYSA-N Calcium cation Chemical compound [Ca+2] BHPQYMZQTOCNFJ-UHFFFAOYSA-N 0.000 description 1
- 229920002134 Carboxymethyl cellulose Polymers 0.000 description 1
- 108091026890 Coding region Proteins 0.000 description 1
- XDTMQSROBMDMFD-UHFFFAOYSA-N Cyclohexane Chemical compound C1CCCCC1 XDTMQSROBMDMFD-UHFFFAOYSA-N 0.000 description 1
- LVZWSLJZHVFIQJ-UHFFFAOYSA-N Cyclopropane Chemical compound C1CC1 LVZWSLJZHVFIQJ-UHFFFAOYSA-N 0.000 description 1
- FBPFZTCFMRRESA-KVTDHHQDSA-N D-Mannitol Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-KVTDHHQDSA-N 0.000 description 1
- 108020004414 DNA Proteins 0.000 description 1
- MHZGKXUYDGKKIU-UHFFFAOYSA-N Decylamine Chemical compound CCCCCCCCCCN MHZGKXUYDGKKIU-UHFFFAOYSA-N 0.000 description 1
- 108020005199 Dehydrogenases Proteins 0.000 description 1
- 229920002307 Dextran Polymers 0.000 description 1
- 108090000371 Esterases Proteins 0.000 description 1
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 1
- 108010053070 Glutathione Disulfide Proteins 0.000 description 1
- 102000003886 Glycoproteins Human genes 0.000 description 1
- 108090000288 Glycoproteins Proteins 0.000 description 1
- 101000987586 Homo sapiens Eosinophil peroxidase Proteins 0.000 description 1
- 101001075374 Homo sapiens Gamma-glutamyl hydrolase Proteins 0.000 description 1
- 101000664737 Homo sapiens Somatotropin Proteins 0.000 description 1
- AVXURJPOCDRRFD-UHFFFAOYSA-N Hydroxylamine Chemical compound ON AVXURJPOCDRRFD-UHFFFAOYSA-N 0.000 description 1
- DGAQECJNVWCQMB-PUAWFVPOSA-M Ilexoside XXIX Chemical compound C[C@@H]1CC[C@@]2(CC[C@@]3(C(=CC[C@H]4[C@]3(CC[C@@H]5[C@@]4(CC[C@@H](C5(C)C)OS(=O)(=O)[O-])C)C)[C@@H]2[C@]1(C)O)C)C(=O)O[C@H]6[C@@H]([C@H]([C@@H]([C@H](O6)CO)O)O)O.[Na+] DGAQECJNVWCQMB-PUAWFVPOSA-M 0.000 description 1
- 108010005714 Interferon beta-1b Proteins 0.000 description 1
- 101001110310 Lentilactobacillus kefiri NADP-dependent (R)-specific alcohol dehydrogenase Proteins 0.000 description 1
- 239000004472 Lysine Substances 0.000 description 1
- JLVVSXFLKOJNIY-UHFFFAOYSA-N Magnesium ion Chemical compound [Mg+2] JLVVSXFLKOJNIY-UHFFFAOYSA-N 0.000 description 1
- 229930195725 Mannitol Natural products 0.000 description 1
- 241001465754 Metazoa Species 0.000 description 1
- BZLVMXJERCGZMT-UHFFFAOYSA-N Methyl tert-butyl ether Chemical compound COC(C)(C)C BZLVMXJERCGZMT-UHFFFAOYSA-N 0.000 description 1
- 239000005642 Oleic acid Substances 0.000 description 1
- ZQPPMHVWECSIRJ-UHFFFAOYSA-N Oleic acid Natural products CCCCCCCCC=CCCCCCCCC(O)=O ZQPPMHVWECSIRJ-UHFFFAOYSA-N 0.000 description 1
- 229910019142 PO4 Inorganic materials 0.000 description 1
- 108091005804 Peptidases Proteins 0.000 description 1
- 102000035195 Peptidases Human genes 0.000 description 1
- 108700020962 Peroxidase Proteins 0.000 description 1
- 102000003992 Peroxidases Human genes 0.000 description 1
- 108090000608 Phosphoric Monoester Hydrolases Proteins 0.000 description 1
- 102000004160 Phosphoric Monoester Hydrolases Human genes 0.000 description 1
- 229920003171 Poly (ethylene oxide) Polymers 0.000 description 1
- 239000004743 Polypropylene Substances 0.000 description 1
- 239000004365 Protease Substances 0.000 description 1
- 239000012614 Q-Sepharose Substances 0.000 description 1
- 229920002684 Sepharose Polymers 0.000 description 1
- MTCFGRXMJLQNBG-UHFFFAOYSA-N Serine Natural products OCC(N)C(O)=O MTCFGRXMJLQNBG-UHFFFAOYSA-N 0.000 description 1
- DBMJMQXJHONAFJ-UHFFFAOYSA-M Sodium laurylsulphate Chemical compound [Na+].CCCCCCCCCCCCOS([O-])(=O)=O DBMJMQXJHONAFJ-UHFFFAOYSA-M 0.000 description 1
- 108010073771 Soybean Proteins Proteins 0.000 description 1
- 102000005262 Sulfatase Human genes 0.000 description 1
- NINIDFKCEFEMDL-UHFFFAOYSA-N Sulfur Chemical compound [S] NINIDFKCEFEMDL-UHFFFAOYSA-N 0.000 description 1
- 239000012505 Superdex™ Substances 0.000 description 1
- DTQVDTLACAAQTR-UHFFFAOYSA-N Trifluoroacetic acid Chemical class OC(=O)C(F)(F)F DTQVDTLACAAQTR-UHFFFAOYSA-N 0.000 description 1
- 239000007984 Tris EDTA buffer Substances 0.000 description 1
- 240000008042 Zea mays Species 0.000 description 1
- 235000016383 Zea mays subsp huehuetenangensis Nutrition 0.000 description 1
- 235000002017 Zea mays subsp mays Nutrition 0.000 description 1
- 150000001242 acetic acid derivatives Chemical class 0.000 description 1
- 150000007513 acids Chemical class 0.000 description 1
- 230000006978 adaptation Effects 0.000 description 1
- 239000000443 aerosol Substances 0.000 description 1
- 238000013019 agitation Methods 0.000 description 1
- 125000003172 aldehyde group Chemical group 0.000 description 1
- 125000000033 alkoxyamino group Chemical group 0.000 description 1
- 125000003282 alkyl amino group Chemical group 0.000 description 1
- 125000004422 alkyl sulphonamide group Chemical group 0.000 description 1
- 125000004414 alkyl thio group Chemical group 0.000 description 1
- VREFGVBLTWBCJP-UHFFFAOYSA-N alprazolam Chemical compound C12=CC(Cl)=CC=C2N2C(C)=NN=C2CN=C1C1=CC=CC=C1 VREFGVBLTWBCJP-UHFFFAOYSA-N 0.000 description 1
- 229910000147 aluminium phosphate Inorganic materials 0.000 description 1
- 239000001166 ammonium sulphate Substances 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 239000002518 antifoaming agent Substances 0.000 description 1
- 230000000890 antigenic effect Effects 0.000 description 1
- ODKSFYDXXFIFQN-UHFFFAOYSA-N arginine Natural products OC(=O)C(N)CCCNC(N)=N ODKSFYDXXFIFQN-UHFFFAOYSA-N 0.000 description 1
- 125000001769 aryl amino group Chemical group 0.000 description 1
- 125000004421 aryl sulphonamide group Chemical group 0.000 description 1
- 125000005110 aryl thio group Chemical group 0.000 description 1
- 125000004104 aryloxy group Chemical group 0.000 description 1
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 1
- 229940003504 avonex Drugs 0.000 description 1
- 125000003943 azolyl group Chemical group 0.000 description 1
- 235000019445 benzyl alcohol Nutrition 0.000 description 1
- 150000001576 beta-amino acids Chemical class 0.000 description 1
- 229940021459 betaseron Drugs 0.000 description 1
- 125000002619 bicyclic group Chemical group 0.000 description 1
- 230000004071 biological effect Effects 0.000 description 1
- 210000001185 bone marrow Anatomy 0.000 description 1
- 150000001649 bromium compounds Chemical class 0.000 description 1
- 229910001424 calcium ion Inorganic materials 0.000 description 1
- 150000001720 carbohydrates Chemical class 0.000 description 1
- 150000004649 carbonic acid derivatives Chemical class 0.000 description 1
- 239000001768 carboxy methyl cellulose Substances 0.000 description 1
- 235000010948 carboxy methyl cellulose Nutrition 0.000 description 1
- 150000001733 carboxylic acid esters Chemical class 0.000 description 1
- 239000008112 carboxymethyl-cellulose Substances 0.000 description 1
- 210000000085 cashmere Anatomy 0.000 description 1
- 239000004359 castor oil Substances 0.000 description 1
- 235000019438 castor oil Nutrition 0.000 description 1
- 230000015556 catabolic process Effects 0.000 description 1
- 230000004663 cell proliferation Effects 0.000 description 1
- 239000001913 cellulose Substances 0.000 description 1
- 229920002678 cellulose Polymers 0.000 description 1
- 235000010980 cellulose Nutrition 0.000 description 1
- 229910000420 cerium oxide Inorganic materials 0.000 description 1
- PBAYDYUZOSNJGU-UHFFFAOYSA-N chelidonic acid Natural products OC(=O)C1=CC(=O)C=C(C(O)=O)O1 PBAYDYUZOSNJGU-UHFFFAOYSA-N 0.000 description 1
- 150000001805 chlorine compounds Chemical class 0.000 description 1
- 238000004587 chromatography analysis Methods 0.000 description 1
- 208000020832 chronic kidney disease Diseases 0.000 description 1
- 239000007979 citrate buffer Substances 0.000 description 1
- 150000001860 citric acid derivatives Chemical class 0.000 description 1
- 230000000112 colonic effect Effects 0.000 description 1
- 239000003086 colorant Substances 0.000 description 1
- 229940125904 compound 1 Drugs 0.000 description 1
- 229940125773 compound 10 Drugs 0.000 description 1
- 229940125782 compound 2 Drugs 0.000 description 1
- 229940126214 compound 3 Drugs 0.000 description 1
- 229920001577 copolymer Polymers 0.000 description 1
- 239000012043 crude product Substances 0.000 description 1
- 125000004122 cyclic group Chemical group 0.000 description 1
- XUJNEKJLAYXESH-UHFFFAOYSA-N cysteine Natural products SCC(N)C(O)=O XUJNEKJLAYXESH-UHFFFAOYSA-N 0.000 description 1
- 235000018417 cysteine Nutrition 0.000 description 1
- 125000000151 cysteine group Chemical group N[C@@H](CS)C(=O)* 0.000 description 1
- 238000006731 degradation reaction Methods 0.000 description 1
- KXGVEGMKQFWNSR-LLQZFEROSA-M deoxycholate Chemical compound C([C@H]1CC2)[C@H](O)CC[C@]1(C)[C@@H]1[C@@H]2[C@@H]2CC[C@H]([C@@H](CCC([O-])=O)C)[C@@]2(C)[C@@H](O)C1 KXGVEGMKQFWNSR-LLQZFEROSA-M 0.000 description 1
- 229940009976 deoxycholate Drugs 0.000 description 1
- 238000010511 deprotection reaction Methods 0.000 description 1
- 239000003599 detergent Substances 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 238000002405 diagnostic procedure Methods 0.000 description 1
- 125000004663 dialkyl amino group Chemical group 0.000 description 1
- 125000005266 diarylamine group Chemical group 0.000 description 1
- 235000014113 dietary fatty acids Nutrition 0.000 description 1
- 239000006185 dispersion Substances 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 239000003937 drug carrier Substances 0.000 description 1
- 238000001962 electrophoresis Methods 0.000 description 1
- 230000001804 emulsifying effect Effects 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 125000001495 ethyl group Chemical group [H]C([H])([H])C([H])([H])* 0.000 description 1
- 239000013604 expression vector Substances 0.000 description 1
- 239000000194 fatty acid Substances 0.000 description 1
- 229930195729 fatty acid Natural products 0.000 description 1
- 150000004665 fatty acids Chemical class 0.000 description 1
- 239000000796 flavoring agent Substances 0.000 description 1
- 125000005567 fluorenylene group Chemical group 0.000 description 1
- 238000009472 formulation Methods 0.000 description 1
- 238000005194 fractionation Methods 0.000 description 1
- 239000012634 fragment Substances 0.000 description 1
- 125000002541 furyl group Chemical group 0.000 description 1
- YPZRWBKMTBYPTK-BJDJZHNGSA-N glutathione disulfide Chemical compound OC(=O)[C@@H](N)CCC(=O)N[C@H](C(=O)NCC(O)=O)CSSC[C@@H](C(=O)NCC(O)=O)NC(=O)CC[C@H](N)C(O)=O YPZRWBKMTBYPTK-BJDJZHNGSA-N 0.000 description 1
- 125000005456 glyceride group Chemical group 0.000 description 1
- ZEMPKEQAKRGZGQ-XOQCFJPHSA-N glycerol triricinoleate Natural products CCCCCC[C@@H](O)CC=CCCCCCCCC(=O)OC[C@@H](COC(=O)CCCCCCCC=CC[C@@H](O)CCCCCC)OC(=O)CCCCCCCC=CC[C@H](O)CCCCCC ZEMPKEQAKRGZGQ-XOQCFJPHSA-N 0.000 description 1
- 210000003714 granulocyte Anatomy 0.000 description 1
- 229910052736 halogen Inorganic materials 0.000 description 1
- 150000002367 halogens Chemical class 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- DMEGYFMYUHOHGS-UHFFFAOYSA-N heptamethylene Natural products C1CCCCCC1 DMEGYFMYUHOHGS-UHFFFAOYSA-N 0.000 description 1
- 125000005553 heteroaryloxy group Chemical group 0.000 description 1
- XTBMQKZEIICCCS-UHFFFAOYSA-N hexane-1,5-diamine Chemical compound CC(N)CCCCN XTBMQKZEIICCCS-UHFFFAOYSA-N 0.000 description 1
- 102000044890 human EPO Human genes 0.000 description 1
- 230000007062 hydrolysis Effects 0.000 description 1
- 238000006460 hydrolysis reaction Methods 0.000 description 1
- 230000003301 hydrolyzing effect Effects 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- 125000002883 imidazolyl group Chemical group 0.000 description 1
- 230000005847 immunogenicity Effects 0.000 description 1
- 238000011534 incubation Methods 0.000 description 1
- 230000006698 induction Effects 0.000 description 1
- 238000001802 infusion Methods 0.000 description 1
- 230000000977 initiatory effect Effects 0.000 description 1
- 239000007972 injectable composition Substances 0.000 description 1
- 238000002347 injection Methods 0.000 description 1
- 239000007924 injection Substances 0.000 description 1
- 238000002697 interventional radiology Methods 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
- 238000007919 intrasynovial administration Methods 0.000 description 1
- 238000007913 intrathecal administration Methods 0.000 description 1
- 238000001990 intravenous administration Methods 0.000 description 1
- 150000004694 iodide salts Chemical class 0.000 description 1
- 238000005342 ion exchange Methods 0.000 description 1
- 238000004255 ion exchange chromatography Methods 0.000 description 1
- QXJSBBXBKPUZAA-UHFFFAOYSA-N isooleic acid Natural products CCCCCCCC=CCCCCCCCCC(O)=O QXJSBBXBKPUZAA-UHFFFAOYSA-N 0.000 description 1
- 125000001449 isopropyl group Chemical group [H]C([H])([H])C([H])(*)C([H])([H])[H] 0.000 description 1
- 239000010977 jade Substances 0.000 description 1
- ZLVXBBHTMQJRSX-VMGNSXQWSA-N jdtic Chemical compound C1([C@]2(C)CCN(C[C@@H]2C)C[C@H](C(C)C)NC(=O)[C@@H]2NCC3=CC(O)=CC=C3C2)=CC=CC(O)=C1 ZLVXBBHTMQJRSX-VMGNSXQWSA-N 0.000 description 1
- IZWSFJTYBVKZNK-UHFFFAOYSA-N lauryl sulfobetaine Chemical compound CCCCCCCCCCCC[N+](C)(C)CCCS([O-])(=O)=O IZWSFJTYBVKZNK-UHFFFAOYSA-N 0.000 description 1
- 210000004185 liver Anatomy 0.000 description 1
- 239000007937 lozenge Substances 0.000 description 1
- 239000000314 lubricant Substances 0.000 description 1
- 229910001425 magnesium ion Inorganic materials 0.000 description 1
- 235000009973 maize Nutrition 0.000 description 1
- 239000000594 mannitol Substances 0.000 description 1
- 235000010355 mannitol Nutrition 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 230000002503 metabolic effect Effects 0.000 description 1
- AFVFQIVMOAPDHO-UHFFFAOYSA-M methanesulfonate group Chemical class CS(=O)(=O)[O-] AFVFQIVMOAPDHO-UHFFFAOYSA-M 0.000 description 1
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 description 1
- 239000011785 micronutrient Substances 0.000 description 1
- 235000013369 micronutrients Nutrition 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 108091005601 modified peptides Proteins 0.000 description 1
- 229960005181 morphine Drugs 0.000 description 1
- 125000000740 n-pentyl group Chemical group [H]C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])* 0.000 description 1
- 125000004123 n-propyl group Chemical group [H]C([H])([H])C([H])([H])C([H])([H])* 0.000 description 1
- 125000001624 naphthyl group Chemical group 0.000 description 1
- 230000003472 neutralizing effect Effects 0.000 description 1
- 150000002823 nitrates Chemical class 0.000 description 1
- 239000000346 nonvolatile oil Substances 0.000 description 1
- 238000010899 nucleation Methods 0.000 description 1
- ZQPPMHVWECSIRJ-KTKRTIGZSA-N oleic acid Chemical compound CCCCCCCC\C=C/CCCCCCCC(O)=O ZQPPMHVWECSIRJ-KTKRTIGZSA-N 0.000 description 1
- 239000004006 olive oil Substances 0.000 description 1
- 235000008390 olive oil Nutrition 0.000 description 1
- 239000012044 organic layer Substances 0.000 description 1
- BMMGVYCKOGBVEV-UHFFFAOYSA-N oxo(oxoceriooxy)cerium Chemical compound [Ce]=O.O=[Ce]=O BMMGVYCKOGBVEV-UHFFFAOYSA-N 0.000 description 1
- 239000001301 oxygen Substances 0.000 description 1
- 238000010979 pH adjustment Methods 0.000 description 1
- 229910052763 palladium Inorganic materials 0.000 description 1
- 239000000546 pharmaceutical excipient Substances 0.000 description 1
- 229940124531 pharmaceutical excipient Drugs 0.000 description 1
- 125000001997 phenyl group Chemical group [H]C1=C([H])C([H])=C(*)C([H])=C1[H] 0.000 description 1
- UYWQUFXKFGHYNT-UHFFFAOYSA-N phenylmethyl ester of formic acid Natural products O=COCC1=CC=CC=C1 UYWQUFXKFGHYNT-UHFFFAOYSA-N 0.000 description 1
- 235000021317 phosphate Nutrition 0.000 description 1
- 239000008363 phosphate buffer Substances 0.000 description 1
- 125000002467 phosphate group Chemical group [H]OP(=O)(O[H])O[*] 0.000 description 1
- 150000003013 phosphoric acid derivatives Chemical class 0.000 description 1
- 230000001766 physiological effect Effects 0.000 description 1
- 229920001155 polypropylene Polymers 0.000 description 1
- 229920000136 polysorbate Polymers 0.000 description 1
- 229920002451 polyvinyl alcohol Polymers 0.000 description 1
- 235000019422 polyvinyl alcohol Nutrition 0.000 description 1
- 229910001414 potassium ion Inorganic materials 0.000 description 1
- 239000003755 preservative agent Substances 0.000 description 1
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 description 1
- 102000004196 processed proteins & peptides Human genes 0.000 description 1
- OGHBATFHNDZKSO-UHFFFAOYSA-N propan-2-olate Chemical compound CC(C)[O-] OGHBATFHNDZKSO-UHFFFAOYSA-N 0.000 description 1
- 238000009163 protein therapy Methods 0.000 description 1
- 125000004076 pyridyl group Chemical group 0.000 description 1
- 125000000714 pyrimidinyl group Chemical group 0.000 description 1
- 125000000168 pyrrolyl group Chemical group 0.000 description 1
- 125000002294 quinazolinyl group Chemical group N1=C(N=CC2=CC=CC=C12)* 0.000 description 1
- 230000009257 reactivity Effects 0.000 description 1
- 229940038850 rebif Drugs 0.000 description 1
- 238000011084 recovery Methods 0.000 description 1
- 230000009467 reduction Effects 0.000 description 1
- 238000006722 reduction reaction Methods 0.000 description 1
- 125000006413 ring segment Chemical group 0.000 description 1
- 208000011571 secondary malignant neoplasm Diseases 0.000 description 1
- 230000001953 sensory effect Effects 0.000 description 1
- 229910052708 sodium Inorganic materials 0.000 description 1
- 229910001415 sodium ion Inorganic materials 0.000 description 1
- 235000019333 sodium laurylsulphate Nutrition 0.000 description 1
- 229940001941 soy protein Drugs 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 230000004936 stimulating effect Effects 0.000 description 1
- 238000007920 subcutaneous administration Methods 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 125000001424 substituent group Chemical group 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 108060007951 sulfatase Proteins 0.000 description 1
- 229940124530 sulfonamide Drugs 0.000 description 1
- 150000003456 sulfonamides Chemical class 0.000 description 1
- 239000011593 sulfur Substances 0.000 description 1
- 150000003467 sulfuric acid derivatives Chemical class 0.000 description 1
- 239000000829 suppository Substances 0.000 description 1
- 239000004094 surface-active agent Substances 0.000 description 1
- 239000000375 suspending agent Substances 0.000 description 1
- 239000003765 sweetening agent Substances 0.000 description 1
- 208000024891 symptom Diseases 0.000 description 1
- 238000012360 testing method Methods 0.000 description 1
- 125000000335 thiazolyl group Chemical group 0.000 description 1
- 125000001544 thienyl group Chemical group 0.000 description 1
- 125000003396 thiol group Chemical group [H]S* 0.000 description 1
- BRNULMACUQOKMR-UHFFFAOYSA-N thiomorpholine Chemical compound C1CSCCN1 BRNULMACUQOKMR-UHFFFAOYSA-N 0.000 description 1
- 231100000331 toxic Toxicity 0.000 description 1
- 230000002588 toxic effect Effects 0.000 description 1
- 235000013619 trace mineral Nutrition 0.000 description 1
- 239000011573 trace mineral Substances 0.000 description 1
- 230000009466 transformation Effects 0.000 description 1
- 238000000844 transformation Methods 0.000 description 1
- 238000005199 ultracentrifugation Methods 0.000 description 1
- 150000003672 ureas Chemical class 0.000 description 1
- 150000003673 urethanes Chemical class 0.000 description 1
- 230000002792 vascular Effects 0.000 description 1
- 239000003981 vehicle Substances 0.000 description 1
- 230000029812 viral genome replication Effects 0.000 description 1
- HBOMLICNUCNMMY-XLPZGREQSA-N zidovudine Chemical compound O=C1NC(=O)C(C)=CN1[C@@H]1O[C@H](CO)[C@@H](N=[N+]=[N-])C1 HBOMLICNUCNMMY-XLPZGREQSA-N 0.000 description 1
- 229960002555 zidovudine Drugs 0.000 description 1
Classifications
-
- 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
- A61K38/21—Interferons [IFN]
- A61K38/215—IFN-beta
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
-
- 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
- A61K38/18—Growth factors; Growth regulators
- A61K38/1816—Erythropoietin [EPO]
-
- 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
- A61K38/22—Hormones
- A61K38/27—Growth hormone [GH], i.e. somatotropin
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/30—Macromolecular organic or inorganic compounds, e.g. inorganic polyphosphates
- A61K47/34—Macromolecular compounds obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyesters, polyamino acids, polysiloxanes, polyphosphazines, copolymers of polyalkylene glycol or poloxamers
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/30—Macromolecular organic or inorganic compounds, e.g. inorganic polyphosphates
- A61K47/42—Proteins; Polypeptides; Degradation products thereof; Derivatives thereof, e.g. albumin, gelatin or zein
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K47/00—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient
- A61K47/50—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates
- A61K47/51—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent
- A61K47/56—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule
- A61K47/59—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyureas or polyurethanes
- A61K47/60—Medicinal preparations characterised by the non-active ingredients used, e.g. carriers or inert additives; Targeting or modifying agents chemically bound to the active ingredient the non-active ingredient being chemically bound to the active ingredient, e.g. polymer-drug conjugates the non-active ingredient being a modifying agent the modifying agent being an organic macromolecular compound, e.g. an oligomeric, polymeric or dendrimeric molecule obtained otherwise than by reactions only involving carbon-to-carbon unsaturated bonds, e.g. polyureas or polyurethanes the organic macromolecular compound being a polyoxyalkylene oligomer, polymer or dendrimer, e.g. PEG, PPG, PEO or polyglycerol
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P25/00—Drugs for disorders of the nervous system
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
- A61P31/12—Antivirals
- A61P31/14—Antivirals for RNA viruses
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
- A61P31/12—Antivirals
- A61P31/20—Antivirals for DNA viruses
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P37/00—Drugs for immunological or allergic disorders
- A61P37/02—Immunomodulators
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P7/00—Drugs for disorders of the blood or the extracellular fluid
- A61P7/06—Antianaemics
Landscapes
- Health & Medical Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Animal Behavior & Ethology (AREA)
- Medicinal Chemistry (AREA)
- Pharmacology & Pharmacy (AREA)
- General Health & Medical Sciences (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- Engineering & Computer Science (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Epidemiology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Immunology (AREA)
- Gastroenterology & Hepatology (AREA)
- Zoology (AREA)
- Organic Chemistry (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Chemical Kinetics & Catalysis (AREA)
- General Chemical & Material Sciences (AREA)
- Virology (AREA)
- Endocrinology (AREA)
- Communicable Diseases (AREA)
- Oncology (AREA)
- Molecular Biology (AREA)
- Inorganic Chemistry (AREA)
- Diabetes (AREA)
- Biotechnology (AREA)
- Neurosurgery (AREA)
- Neurology (AREA)
- Biomedical Technology (AREA)
- Hematology (AREA)
- Peptides Or Proteins (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
- Medicinal Preparation (AREA)
- Polyethers (AREA)
Description
本發明係關於一種胜肽-聚合物綴合物。
在細胞生物學及重組蛋白質技術上的發展已導致蛋白質治療之發展。
然而,主要困難仍然存在。大部分蛋白質易受蛋白代謝性降解影響,因此具有短的循環時間。其它缺點包括低水溶解度及引發中和性抗體。
聚合物(例如,聚乙二醇(PEG))接附至蛋白質會阻礙蛋白代謝酵素接近蛋白質骨架,此導致蛋白質穩定性提高。此外,亦可改良水溶解度及減少免疫抗原性。已需要有效將聚合物接附至蛋白質的方法。
本發明之觀點係關於式I的聚合物-多胜肽綴合物:
其中R1
、R2
、R3
、R4
及R5
各者各自獨立地為H、C1-10
烷基、C2-10
烯基、C2-10
炔基、芳基、雜芳基、C3-8
環烷基或C3-8
雜環烷基;A1
及A2
各者各自獨立地為聚合物部分(例如,聚環氧化烷部分);G1
、G2
及G3
各者各自獨立地為一鍵結或一連結官能基;P為干擾素-β(INF-β)部分、紅血球生成素(EPO)部分或生長激素(GH)部分;m為0或整數1-10;及n為整數1-10。在這些綴合物中,該INF-β部分、EPO部分或GH部分的N-終端鍵結至G3
。
參照上式,該聚合物-多胜肽綴合物具有一或多個下列特徵:A1
及A2
為具有分子量2-100kD(10-30kD較佳,例如,20kD)的聚環氧化烷部分;G1
及G2
各者為(其中O原子鍵結至A1
或A2
及N原子鍵結至碳原子,如顯示在式I中);G1
及G2
各者為尿素、磺醯胺或醯胺(其中N原子鍵結至碳原子,如顯示在式I中);m為4;n為2;及R1
、R2
、R3
、R4
及R5
各者為H。在這些綴合物的某些中,P為rINF-β Ser17
或在INF-β的N-終端處包含1-4個其它胺基酸殘基之經改質的INF-β部分。
本發明的另一個觀點係關於式II之聚合物-胜肽綴合物:
A-G1
-L-G2
-P式II
其中A為聚合物部分(例如,聚環氧化烷部分);G1
及G2
各者各自獨立地為一鍵結或一連結官能基;L為C2-10
伸烯基或C2-10
伸炔基;及P為INF-β部分、EPO部分或GH部分。在這些綴合物中,INF-β部分、EPO部分或GH部分的N-終端接附至G2
。
參照式II,該聚合物-胜肽綴合物具有一或多個下列特徵:A1
及A2
為具有分子量2-100kD(10-30kD較佳,例如,20kD)的聚環氧化烷部分,G1
及G2
各者為一鍵結,C6
為伸烯基,及R1
、R2
、R3
、R4
與R5
各者皆為H。
本發明的另一個觀點係關於式III之聚合物-胜肽綴合物:
其中R1
、R2
、R3
及R4
各者各自獨立地為H、C1-10
烷基、C2-10
烯基、C2-10
炔基、芳基、雜芳基、C3-8
環烷基或C3-8
雜環烷基;n為整數2-10;A為聚合物部分;G為連結官能基;及P為胜肽部分,該胜肽部分的N-終端之氮原子鍵結至在上述式中所顯示出的部分之碳原子。
參照式III,該聚合物-胜肽綴合物具有一或多個下列特徵:n為1;A為具有分子量10-40kD或20-30kD的聚環氧化烷部分;G為:
其中O原子鍵結至A及N原子鍵結至碳原子;及P為INF部分、EPO部分、GH部分。
於本文中所使用的名稱“C1-10
烷基”指為包含1至10個碳原子之直鏈或分枝的烴單價基團。該烷基的實施例包括甲基、乙基、正丙基、異丙基、三級丁基及正戊基。類似地,名稱“C2-10
烯基”指為包含2至10個碳原子及一或多個雙鍵之直鏈或分枝的烴單價基團。名稱“C2-10
炔基”指為包含2至10個碳原子及一或多個三鍵之直鏈或分枝的烴單價基團。名稱“C2-10
伸烯基”指為包含2至10個碳原子及一或多個雙鍵之直鏈或分枝的烴二價基團。名稱“C2-10
伸炔基”指為包含2至10個碳原子及一或多個三鍵之直鏈或分枝的烴二價基團。
於本文中所使用的名稱“芳基”指為具有至少一個芳香環之烴環系統(單環或雙環)。該芳基部分的實施例包括(但不限於)苯基、萘基及芘基。
於本文中所使用的名稱“雜芳基”指為具有至少一個芳香環(其包含至少一個雜原子(諸如O、N或S)作為該環系統的部分及剩餘為碳)之烴環系統(單環或雙環)。該雜芳基部分的實施例包括(但不限於)呋喃基、吡咯基、噻吩基、唑基、咪唑基、噻唑基、吡啶基、嘧啶基、喹唑啉基及吲哚基。
於本文中所使用的名稱“環烷基”指為僅具有碳環原子之部分或完全飽和的單環或雙環環系統。實施例包括(但不限於)環丙烷、環戊烷及環己烷。
於本文中所使用的名稱“雜環烷基”指為除了碳外,具有一或多個雜原子(例如,O、N或S)作為環原子之部分或完全飽和的單環或雙環環系統。實施例包括(但不限於)哌啶、哌、嗎福啉、硫嗎福啉及1,4-氧氮。
於本文所提到的烷基、烯基、炔基、伸烯基、伸炔基、芳基、雜芳基、環烷基及雜環烷基包括經取代及未經取代的部分二者。該取代基之實施例包括C1
-C10
烷基、C2
-C10
烯基、C2
-C10
炔基、C3
-C8
環烷基、C5
-C8
環烯基、C1
-C10
烷氧基、芳基、芳氧基、雜芳基、雜芳氧基、胺基、C1
-C10
烷基胺基、C1
-C20
二烷基胺基、芳基胺基、二芳基胺基、羥基胺基、烷氧基胺基、C1
-C10
烷基磺醯胺、芳基磺醯胺、羥基、鹵素、硫、C1
-C10
烷硫基、芳硫基、氰基、硝基、醯基、醯氧基、羧基及羧酸酯。
名稱“聚合物部分”指為衍生自線性、分枝或星形聚合物之單價基團。該聚合物部分的分子量可為2-100kD。該聚合物部分之實施例包括(但不限於)聚環氧乙烷、聚乙二醇、聚環氧異丙烷、聚環氧丁烯、聚乙二醇及其共聚物。亦可使用其它聚合物,諸如葡萄聚糖、聚乙烯醇類、聚丙烯醯胺類或以碳水化合物為基礎的聚合物,只要它們不具抗原性、毒性或引起免疫反應。
名稱“多胜肽部分”指為來自天然發生的多胜肽或經改質的多胜肽之單價基團。該天然發生的胜肽可為INF-α2b
、INF-β、GH、EPO及粒性細胞結腸刺激因子或抗體。該經改質的胜肽可例如為在INF-α2b
的N-終端處包含INF及1-4個其它胺基酸殘基之胜肽。此經改質的INF之實施例有,IFN代表INF-α2b
部分,該胺基在其N-終端處鍵結至羰基。
名稱“干擾素-β”指為高度同源可抑制病毒複製及細胞增生且可調節免疫反應的蛋白質家族。參見戴潤克(Derynck)等人,(1980),自然 285
(5766):542-7;及谷口(Taniguchi)等人,(1980),基因 10
(1):11-5。其包含天然發生的INF-β及其官能性同等物(即,具有至少80%(例如,85%、90%、95%或99%)與其野生型對應體相同之多胜肽)二者。INF-β的實施例包括在可商業購得的藥物中之活性成份,諸如雅芳內斯(Avonex)、貝它誰隆(Betaseron)及利比扶(Rebif)。參見例如艾替馬帝發(Etemadifar)M.等人,Acta Neurol. Scand
.,2006,113(5):283-7。
下列列出者為典型人類INF-β蛋白質的胺基酸序列(呈前驅物形式或成熟形式):mtnkcllqia lllcfsttal
smsynllgfl qrssnfqcqk llwqlngrle yclkdrmnfd ipeeikqlqq fqkedaalti yemlqnifai frqdssstgw netivenlla nvyhqinhlk tvleekleke dftrgklmss lhlkryygri lhylkakeys hcawtivrve ilrnfyfinr ltgylrn
(參見基因銀行(Genbank)登錄編號:M28622,1993年4月23日版本;斜體表示部分指為訊息胜肽)mnsfstsafg pvafslglll vlpaafpapv ppgedskdva aphrqpltss eridkqiryi ldgisalrke tcnksnmces skealaennl nlpkmaekdg cfqsgfneet clvkiitgll efevyleylq nrfesseeqa ravqmstkvl iqflqkkakn ldaittpdpt tnaslltklq aqnqwlqdmt thlilrsfke flqsslralr qm
(參見基因銀行登錄編號:CAA00839,1993年12月3日版本)
在一個實施例中,該INF-β為突變型rINF-β Ser17
(重組的INF-β,其中絲胺酸為在天然成熟的INF-β序列中之位置17處之半胱胺酸的取代)。此突變型的胺基酸顯示在下列:synllgflqr ssnfqsqkll wqlngrleyc lkdrmnfdip eeikqlqqfq kedaaltiye mlqnifaifr qdssstgwne tivenllanv yhqinhlktv leeklekedf trgklmsslh lkryygrilh ylkakeyshc awtivrveil rnfyfinrlt gylrn
在另一個實施例中,該INF-β為經改質的天然INF-β,其中1-4個其它胺基酸殘基接附至天然的INF-β之N-終端。
EPO(由肝或腎臟製造)為一種控制紅血球生成或紅血球製造的糖蛋白激素。其包括天然發生的EPO及其官能性同等物二者。參見美國專利5,621,080及美國專利申請案公告20050176627。人類EPO的胺基酸序列(呈前驅物及成熟形式)顯示在下列:mgvhecpawl wlllsllslp lglpvlgapp rlicdsrvle rylleakeae nittgcaehc slnenitvpd tkvnfyawkr mevgqqavev wqglallsea vlrgqallvn ssqpweplql hvdkavsglr slttllralg aqkeaisppd aasaaplrti tadtfrklfr vysnflrgkl klytgeacrt gdr(前驅物)
apprlicdsr vlerylleak eaenittgca ehcslnenit vpdtkvnfya wkrmevgqqa vevwqglall seavlrgqal lvnssqpwep lqlhvdkavs glrslttllr algaqkeais ppdaasaapl rtitadtfrk lfrvysnflr gklklytgea crtgdr(成熟形式)
使用來製得本發明之綴合物的EPO蛋白質可為藉由合適的物種(例如,人類、鼠、豬或牛)所製造之EPO蛋白質(呈前驅物或成熟形式)。在一個實施例中,該EPO蛋白質具有與上述顯示出的胺基酸序列之一有至少80%(例如,85%、90%、95%或99%)相同之胺基酸序列。在另一個實施例中,該EPO為經改質的天然EPO,其中1-4個其它胺基酸殘基接附至天然EPO的N-終端。
名稱“生長激素”指為天然發生的人類生長激素(呈前驅物或成熟形式)及其官能性變異體,即,具有與天然發生的人類生長激素至少80%(例如,85%、90%、95%或99%)相同且擁有與人類生長激素相同的生理學活性之胺基酸序列。在一個實施例中,該生長激素為經改質的天然生長激素,其中1-4個其它胺基酸殘基接附至天然的生長激素之N-終端。天然發生的人類生長激素之胺基酸序列(呈前驅物及成熟形式)顯示在下列:matgsrtsll lafgllclpw lqegsafpti plsrlfdnam lrahrlhqla fdtyqefeea yipkeqkysf lqnpqtslcf sesiptpsnr eetqqksnle llrisllliq swlepvqflr svfanslvyg asdsnvydll kdleegiqtl mgrledgspr tgqifkqtys kfdtnshndd allknyglly cfrkdmdkve tflrivqcrs vegscgf(前驅物)
fptiplsrlf dnamlrahrl hqlafdtyqe feeayipkeq kysflqnpqt slcfsesipt psnreetqqk snlellrisl lliqswlepv qflrsvfans lvygasdsnv ydllkdleeg iqtlmgrled gsprtgqifk qtyskfdtns hnddallkny gllycfrkdm dkvetflriv qcrsvegscg f(成熟形式)
mfptiplsrl fdnamlrahr lhqlafdtyq efeeayipke qkysflqnpq tslcfsesip tpsnreetqq ksnlellris llliqswlep vqflrsvfan slvygasdsn vydllkdlee giqtlmgrle dgsprtgqif kqtyskfdtn shnddallkn ygllycfrkd mdkvetflri vqcrsvegsc gf(改質形式)
使用卡琳(Karlin)及歐次諸(Altschul)之演算法來測量二種胺基酸序列的“同一性百分比”,Proc. Natl. Acad. Sci.
USA 87:2264-68,1990;如在卡琳及歐次諸的Proc. Natl. Acad. Sci.
USA 90:5873-77,1993中修改。此演算法併入歐次諸等人之NBLAST及XBLAST程式(2.0版)中,J. Mol. Biol.
215:403-10,1990。可使用XBLAST程式,刻記(score)=50,字長(wordlength)=3來進行BLAST蛋白質研究,以獲得與使用在本發明中的蛋白質分子同源之胺基酸序列。若於二個序列間存在有缺口時,可使用Gapped BLAST,如描述在歐次諸等人,Nucleic Acids Res.
25(17):3389-3402,1997中。當使用BLAST及Gapped BLAST程式時,可使用各別程式(例如,XBLAST及NBLAST)的預設參數。
名稱“連結官能基”指為二價官能基,其一端連接至聚合物部分及其它端連接至胜肽部分。實施例包括(但不限於)-O-、-S-、羧酸鹽、羰基、碳酸鹽、醯胺、胺基甲酸鹽、尿素、碸基、亞碸基、胺基、亞胺基、羥基胺基、膦酸鹽或磷酸鹽基團。
上述描述的胜肽-聚合物綴合物可呈自由態形式或呈鹽形式(若合適的話)。例如,可在陰離子與於本發明之胜肽-聚合物綴合物上的正電荷基團(例如,胺基)間形成鹽。合適的陰離子包括氯化物、溴化物、碘化物、硫酸鹽、硝酸鹽、磷酸鹽、檸檬酸鹽、甲磺酸鹽、三氟醋酸鹽及醋酸鹽。同樣地,亦可在陽離子與於本發明之多胜肽-聚合物綴合物上的負電荷基團(例如,羧酸鹽)間形成鹽。合適的陽離子包括鈉離子、鉀離子、鎂離子、鈣離子及銨陽離子(諸如四甲基銨離子)。
此外,該胜肽-聚合物綴合物可具有一或多個雙鍵或一或多個不對稱中心。此綴合物可發生如為外消旋鹽、外消旋混合物、單一鏡像物、各別的非鏡像異構物、非鏡像異構混合物及順-或反-或E-或Z-雙鍵異構形式。
本發明之聚合物-胜肽綴合物的實施例顯示在下列:
其中mPEG代表具有分子量20kD之經甲氧基封端的聚乙二醇,及rINF-β Ser17
、EPO及GH的N-終端接附至顯示在上述結構中之最右邊的碳。
某些蛋白質具有治療用途。因此,本發明之綴合物(包括胜肽部分)可使用來治療疾病。例如,INF-β為一種用來治療HCV或HBV感染之免疫調節用藥法。參見例如,血管及介入性放射學期刊(Journal of Vascular and Interventional Radiology)
13(2002):191-196。因此,在本發明的範圍內包括使用上述描述的INF-β-聚合物綴合物來治療肝炎C病毒(HCV)感染或肝炎B病毒(HBV)感染之方法。至於另一個實施例,EPO為一種由腎臟製造以促進在骨髓中形成紅血球之激素。其已經使用作為免疫調節用藥法,以治療產生自慢性腎臟病的貧血、AIDS的齊多夫定(zidovudine)治療之二級貧血症及與癌相關的貧血症。近來的研究已亦發現EPO提高神經生成且在中風後恢復上扮演關鍵性角色。參見例如,P.T.蔡(Tsai),神經科學期刊,2006,26:1269。因此,本發明的另一個觀點係關於一種藉由上述描述的EPO-聚合物綴合物來治療貧血或提高神經生成之方法。
包含上述描述的INF-β-聚合物綴合物以使用來治療HCV感染或HBV感染之組成物;及包含上述描述的EPO-聚合物綴合物以使用來治療貧血或提高神經生成之組成物;和該綴合物之治療用途及使用來製造用來治療HCV感染、HBV感染或貧血、或用來提高神經生成的藥劑亦在本發明之範圍內。
在下列描述中提出本發明之一或多個具體實例的細節。將從該描述及從申請專利範圍中明瞭本發明之其它特徵、目標及優點。
可藉由在化學技藝中熟知的合成方法來製備本發明之胜肽-聚合物綴合物。例如,可結合具有一或多個活性官能基的連結子分子與二個對在該連結子分子上之官能基具有官能基反應性的聚合物分子。隨後,讓包含官能基的胜肽分子與該連結子分子之官能基反應,以形成本發明的胜肽-聚合物綴合物。於本文中提供二個闡明的合成方法。
下列方法1顯示出製備式I之胜肽-聚合物綴合物的實施例。二胺化合物1(其包含乙縮醛基團)與碳酸N-羥基琥珀醯亞胺酯mPEG(即,化合物2)反應,以形成二-聚乙烯二醇化的(pegylated)化合物3,其隨後轉換成醛4。此醛化合物與具有自由態胺基的胜肽H-P經由還原性烷基化反應,以獲得本發明之胜肽-聚合物綴合物。
下列方法2顯示出製備式II之胜肽-聚合物綴合物的實施例。化合物6具有一聚合物部分及一醛官能基。其可與具有自由態胺基官能基之胜肽7反應。隨後還原(例如,藉由氫化或藉由NaBH3
CN)所產生的產物8,以獲得胜肽-聚合物綴合物9。
下列方法3為製備式III之胜肽-聚合物綴合物的實施例。具有乙縮醛基團的化合物10(其可從β-胺基酸製備)與碳酸N-羥基琥珀醯亞胺酯mPEG 2反應,以形成聚乙烯二醇化的化合物11,其隨後轉換成醛12。此醛化合物與具有自由態胺基的胜肽H-P經由還原性烷基化反應,以獲得想要的化合物13。
上述描述的化學反應包括使用溶劑、試劑、觸媒、保護基團與去保護基團試劑及某些反應條件。它們可額外地包括加入或移除合適的保護基團之步驟(在特別描述於本文之步驟前或後),以最後允許用於胜肽-聚合物綴合物之合成。此外,可使用另一種程序或順序來進行多個合成步驟,以獲得想要的多胜肽-聚合物綴合物。在合成合適的胜肽-聚合物綴合物中有用的合成化學轉換及保護基團方法(保護及去保護)在技藝中已知,及包括例如描述在下列中的那些:R.拉若克(Larock),綜合性有機轉換(Comprehensive Organic Transformations)
,VCH出版社(1989);T.W.格林尼(Greene)及P.G.M.瓦刺(Wuts),有機合成之保護基(Protective Groups in Organic Synthesis)
,第2版,約翰威利及宋斯(John Wiley and Sons)(1991);L.費賽爾(Fieser)及M.費賽爾,用於有機合成之費賽爾及費賽爾試劑(Fieser and Fieser’s Reagents for Organic Synthesis)
,約翰威利及宋斯(1994);及L.帕奎特(Paquette)編輯,用於有機合成之試劑百科全書(Encyclopedia of Reagents for Organic Synthesis)
,約翰威利及宋斯(1995)及其後續版本。
從而合成的胜肽-聚合物綴合物可藉由諸如離子交換色層分析法、凝膠層析色層分析法、電泳、透析、超微濾法或超速離心法之方法進一步純化。
本發明之胜肽-聚合物綴合物可呈具醫藥活性的綴合物形式。再者,其可藉由酵素地裂解(例如,透過水解)在胜肽部分與聚合物部分間之連結來活體內釋放醫藥活性胜肽。牽涉活體內裂解連結之酵素的實施例包括氧化酵素(例如,過氧化酶類、胺氧化酶類或脫氫酶類)、還原酵素(例如,酮基還原酶類)及水解酵素(例如蛋白酶、酯酶、硫酸酯酶或磷酸酯酶)。
因此,本發明的一個觀點係關於一種給藥有效量之一或多種上述描述的胜肽-聚合物綴合物來治療病症(例如,HCV或HBV感染或貧血)的方法。特別是,可藉由將有效量的一或多種胜肽-聚合物綴合物給藥至患者來治療之疾病。此患者可由保健專家根據任何合適的診斷方法之結果鑑別。
如於本文中所使用,名稱“治療”定義為將一包含胜肽-聚合物綴合物之組成物施加或給藥至患有病症、該病症之症狀、該病症之二級疾病或病症,或關於該病症之體質的患者(人類或動物),其目的為治癒、緩和、解除、補救或改善該病症、該病症之症狀、該病症之二級疾病或病症、或關於該病症之體質。“有效量”指為在受治療的患者上授予治療效應之胜肽-聚合物綴合物的量。治療效應可為目標(即,可由某些測試或標誌測量)或主觀(即,患者提供跡象或感覺效應)。
為了實施本發明之方法,具有一或多種上述提及的綴合物之組成物可非經腸道、口服、鼻、直腸、局部或頰給藥。如於本文中所使用的名稱“非經腸式”指為皮下、皮內、靜脈內、肌肉內、關節內、動脈內、滑膜內、胸骨內、鞘內、病灶內、腹膜內、氣管內或顱內注射,和任何合適的輸液技術。
無菌可注射組成物可為在無毒非經腸道之可接受的稀釋劑或溶劑中之溶液或懸浮液,諸如在1,3-丁二醇中的溶液。在可使用之可接受的媒劑及溶劑當中,其可為甘露醇、水、侖爵氏(Ringer)溶液及等滲壓的氯化鈉溶液。此外,習知上使用非揮發性油作為溶劑或懸浮媒質(例如,合成的單或雙甘油酯)。脂肪酸(諸如,油酸及其甘油酯衍生物)在注射物質之製備上有用,如天然醫藥可接受的油,諸如橄欖油或蓖麻油,特別是其聚氧乙基化的形式。這些油溶液或懸浮液亦可包含一長鏈醇稀釋劑或分散劑、或羧甲基纖維素或類似的分散劑。為了調配之目的,亦可使用其它通常使用的界面活性劑,諸如屯(Tween)類或史班(Span)類或在醫藥可接受的固體、液體或其它劑型之製造中通常使用的其它類似的乳化劑或生物效性促進劑。
用於口服給藥的組成物可為任何口服可接受的劑形,包括膠囊、錠劑、乳液及水性懸浮液、分散液與溶液。在錠劑的實例中,通常使用之載劑包括乳糖及玉黍蜀粉。典型上亦加入潤滑劑,諸如硬脂酸鎂。對以膠囊形式口服給藥來說,有用的稀釋劑包括乳糖及乾玉黍蜀粉。當口服給藥水性懸浮液或乳液時,該活性成份可懸浮或溶解在油相中與乳化或懸浮劑結合。若必要時,可加入某些變甜、風味或著色劑。
可根據在醫藥調配物的技藝中熟知之技術來製備鼻用氣霧劑或吸入組成物。例如,此組成物可使用苄醇或其它合適的防腐劑、提高生物效性的吸收促進劑、碳氟化合物及/或其它在技藝中已知的溶解或分散劑來製備,如為在鹽液中的溶液。亦可以用於直腸給藥的栓劑形式來給藥具有一或多種上述描述的化合物之組成物。
例行地使用一醫藥可接受的載劑與一或多種上述提及之活性綴合物。在該醫藥組合物中之載劑必需在觀念上“可接受”,其與該組成物之活性成份(及能穩定該活性成份較佳)相容且對欲治療的患者無毒。可使用一或多種增溶劑作為用來傳遞上述提及的化合物之醫藥賦形劑。其它載劑之實施例包括膠態氧化矽、硬脂酸鎂、纖維素、硫酸月桂酯鈉及D&C黃色#10。
下列實施例僅解釋為闡明用且無論如何不以任何方式限制本揭示的剩餘部分。沒有進一步詳細地闡述,咸信熟習該項技術者可根據於本文中之描述將本發明使用至其最大程度。於本文中所引用的全部公告其全文藉此以參考方式併入本文。
根據描述在Bioconjugate Chem.1993,4,568-569中的方法,從購買自美國CA的桑拜歐公司(SunBio Inc.)之20kD mPEGOH來製備20kD PEGO(C=O)OSu。
將6-(1,3-二氧五圜-2-基)己烷-1,5-二胺在二氯甲烷中之溶液(11.97克包含9.03毫克二胺(47.8微莫耳)的溶液)加入至包含20kD PEGO(C=O)OSu(1.72克,86.0微莫耳)的燒瓶。在PEGO(C=O)OSu完全溶解後,加入N,N-二異丙基乙基胺(79微升,478微莫耳)。在室溫下攪拌該反應混合物24小時,然後,隨著攪拌逐滴加入甲基三級丁基醚(200毫升)。收集所產生的沉澱及在真空中乾燥,以獲得二-PEG乙縮醛(1.69克,98%),如為白色固體。
1
H NMR(400MHz,d6
-DMSO)δ7.16(t,J=5.2赫茲,1H),
7.06(d,J=8.8赫茲,1H),4.76(t,J=4.8赫茲,1H),4.10-3.95(m,4H),1.80-1.65(m,1H),1.65-1.50(m,1H),1.48-1.10(m,6H)。
將二-PEG乙縮醛(4.0克,0.2毫莫耳)懸浮在pH 2.0緩衝液(檸檬酸,40毫升)中。在35℃下攪拌該反應混合物24小時,然後以二氯甲烷(3×50毫升)萃取。在硫酸鎂上乾燥結合的有機層,濃縮,然後再溶解於二氯甲烷(20毫升)中。隨著攪拌將該溶液逐滴加入至甲基三級丁基醚(400毫升)。收集所產生的沉澱及在減壓下乾燥,以獲得二-PEG醛(3.8克,95%),如為白色固體。
1
H NMR(400MHz,d6
-DMSO)δ9.60(s,1H),7.24(d,J=8.4赫茲,1H),7.16(t,J=5.2赫茲,1H),4.10-3.95(m,4H),3.95-3.80(m,1H),3.00-2.85(m,2H),2.58-2.36(m,2H),1.46-1.15(m,6H)。
再者,以下列方式製備二-PEG醛:藉由苄氧基羰基保護商業可獲得的同源離胺酸(homo-lysine)(中國的阿斯塔泰克醫藥有限公司(Astatech Pharmaceutical Co.,Ltd.))之二個胺基。酯化及還原該N經保護的同源離胺酸以形成一醛化合物。隨後,以乙二醇保護該醛基團。然後,藉由於鈀觸媒存在下氫化來移除該苄氧基羰基保護基團。在溫和的鹼性條件下,讓該N經去保護的化合物與活化的mPEGOH(南韓的桑拜歐化學有限公司)反
應。在pH 2.0檸檬酸緩衝液(德國的西格瑪亞得富(Sigma-Aldrich))中,於25℃下攪拌所產生的產物72小時,以移除醛保護基團。獲得109克的二-PEG聚合物醛(產率:95%)。純度大於97.7%(藉由HPLC測量)及大於95%(藉由1
H NMR分析測量)。
將編碼出人類INF-β Ser17
的DNA片斷選殖進入表現載體pET24a中,以產生一表現質體rhINF-β Ser17
-pET24a。將此表現質體轉化至大腸桿菌中,且選擇、培養正轉化株(即,攜帶表現質體的選殖物),並將所產生的大腸桿菌培養物貯存在-80℃下。
在37℃及200rpm下,將10微升上述提及之經貯存的大腸桿菌培養物灌輸進入200毫升由泰瑞費克(Terrific)肉湯及甘油所組成之播種媒質中約15小時。將150毫升如此獲得之大腸桿菌培養物轉移至2.5升包含葡萄糖(10克/升)、MgSO4
.7H2
O(0.7克/升)、(NH4
)2
HPO4
(4克/升)、KH2
PO4
(3克/升)、K2
HPO4
(6克/升)、檸檬酸鹽(1.7克/升)、酵母菌萃取物(10克/升)、卡那黴素(50毫克/毫升)、氯黴素(50毫克/毫升)、抗發泡劑及微量元素(包括FeSO4
.7H2
O(10毫克/升)、ZnSO4
.7H2
O(2.25毫克/升)、CuSO4
.5H2
O(1毫克/升)、MnSO4
.H2
O(0.5毫克/升)、H3
BO3
(0.3毫克/升)、CaCl2
.2H2
O(2毫克/升)、(NH4
)6
Mo7
O24
(0.1毫克/升)、EDTA(0.84毫克/升)及Cl(50毫克/升))的培養媒質,並在37℃下培養。當該大腸桿菌培養物的OD600
到達120至140時,將IPTG(1M)
加入至該培養物以引發rhINF-β Ser17
之表現性。在37℃及300rpm下培養所引發的培養物3小時。當需要時,在培養期間,將包含800克/升葡萄糖及20克/升MgSO4
的飼育媒質加入至該大腸桿菌培養物。
讓如上所述所獲得的大腸桿菌培養物接受離心以採集大腸桿菌細胞。將該細胞再懸浮於PBS緩衝液(0.1M Na2
HPO4
,0.15M NaCl)中及在APV均質化機中破裂。在10,000rpm,4℃下離心如此獲得之均質化溶液15分鐘。收集沉澱物(包括包涵體),將其再懸浮於PBS中及在室溫下攪拌20-30分鐘,以形成一懸浮液。將NaOH(6N)加入至該懸浮液以將其pH調整至12,以允許包含在該包涵體中之蛋白質溶解。約2分鐘後,以6N HCl將該懸浮液的pH值調整至7.5。然後,讓該懸浮液接受離心及收集從而形成的上層液,使用分光光度計來測量其蛋白質濃度。讓該上層液與再摺疊緩衝液(TEA,pH 8.3)混合及沒有攪拌在室溫下培養24-48小時。然後,使用由微孔有限公司(Millipore Inc.)所提供的TFF系統及PLCCC匣(PLCCC cassette)濃縮及透析其。讓所產生的溶液接受超微濾、透析及以SPFF瓊脂糖凝膠管柱分餾。使用另一種SPFF瓊脂糖凝膠管柱進一步分餾如此獲得之餾分A9及A10(包括重組蛋白質rhINF-β Ser17
),以豐富化該重組蛋白質(在餾分A8-A10中)。藉由凝膠層析進一步純化(超級戴克斯(Superdex)75 HR 10/300)這些含rhINF-β Ser17
的餾分,以獲得具有純度大於90%之rhINF-β Ser17
蛋白質(1毫克/毫升)。該重組蛋白質之生物活性為>2×107
IU/毫
克蛋白質。
將18.9毫克的rhINF-β Ser17
及1.51克的二PEG醛懸浮在26毫升0.1M磷酸鈉緩衝液(pH 5.0)中。在此溶液中加入400當量的NaCNBH3
(比利時(Belgium)的阿可羅斯有機物(Acros Organics))。在室溫下攪拌該反應混合物16小時,然後接受以25mM tris-HCl(pH 7.8)透析。藉由離子交換管柱純化粗產物,以獲得2毫克的INF-β-二-PEG聚合物。
在250毫升補充以50微升/毫升卡那黴素及50微升/毫升氯黴素之SYN媒質(10克/升之選擇的大豆蛋白腖(soytone)、5克/升酵母菌萃取物及10克/升NaCl)中,灌輸經轉化的大腸桿菌BLR(DE3)-RIL細胞(其攜帶已操作連結至大腸桿菌啟動子的INF-β之編碼序列)。然後,在37℃下,於搖動器培養器中,以220rpm培養細胞過夜(即,16小時)。
將250毫升上述提及之過夜的培養物灌輸進入3.0升鹼性媒質(10克/升葡萄糖、0.7克/升MgSO4
.7H2
O、4克/升(NH4
)2
HPO4
、3克/升KH2
PO4
、6克/升K2
HPO4
、2克/升檸檬酸鹽、10克/升酵母菌萃取物及2克/升異白胺酸)中,並補充以10克/升的基本葡萄糖、0.7克/升的飼育MgSO4
、30毫升的飼育微量元素(10克/升FeSO4
.7H2
O、2.25克/升ZnSO4
.7H2
O、1克/升CuSO4
.5H2
O、0.5克/升MnSO4
.H2
O、0.3克/升H3
BO3
、2克/升CaCl2
.H2
O、0.1克/升(NH4
)6
Mo7
O24
、0.84克/升EDTA、50毫升/升HCl)、25微升/毫升的卡那黴素
及25微升/毫升的氯黴素,並在五升發酵槽(拜歐浮羅(Bioflo)3000;NJ愛迪生(Edison)的布朗斯微克賽恩泰動公司(Brunswick Scientation Co.))中培養。在發酵期間,藉由自動化加入37% NH4
OH溶液將媒質之pH控制在pH 7.1。將溶氧(DO)程度維持在30%。使用程式控制幫浦加入飼育溶液(800克/升葡萄糖、20克/升MgSO4
、50微升/毫升卡那黴素及50微升/毫升氯黴素),其經設定以便當程度超過40-60時進料。當在發酵培養物中的細胞密度(OD600
)到達180至200時,將4毫升1M的異丙基-β-D-1-硫代半乳吡喃糖苷(IPTG)與30毫升的飼育微量元素及25克的酵母菌萃取物一起加入至該發酵培養物以引發INF-β表現性。在IPTG誘發後5小時,藉由離心採集細胞。
以比率大約1:3(潮溼重量克/毫升)將細胞丸粒懸浮在PBS緩衝液(0.1M磷酸鈉,0.15M氯化鈉,pH 7.4)中,藉由微射流均質機破裂,然後在4℃下以10,000rpm離心20分鐘。以PBS緩衝液清洗該含包涵體(IB)的丸粒兩次,如上所述般離心,及懸浮在1升PBS溶液(0.1M磷酸鈉,0.15M氯化鈉,pH 7.4,3%次維特君(zwittergent)3-14,5mM DTT)中。在攪拌30分鐘後,讓該懸浮液接受pH調整,使用6.0M NaOH調整至12,同時攪拌以溶解丸粒。然後,以6N HCl將懸浮液的pH調整至pH 7.5。在10,000rpm下離心20分鐘後,收集包含可溶的INF-β之上層液。
然後,讓該可溶的INF-β接受如下的再摺疊。在10升新鮮製備的再摺疊緩衝液(100mM Tris-HCl(pH 7.6),0.5M L-
精胺酸,2mM EDTA)中稀釋上述提及之上層液,用來形成一再摺疊混合物。沒有攪拌,培養該混合物48小時。在培養後,對著20mM Tris(含有100mM NaCl,0.05%次維特君3-14,pH 7.0)緩衝液來透析該包含再摺疊的重組INF-β之混合物。
將經透析的混合物負載到SP-瓊脂糖凝膠管柱(GE阿默珊藥劑(Amersham Pharmacia))上,其已預先平衡及以20mM Tris-HCl,100mM NaCl緩衝液(pH 7.0)清洗。以包含20mM Tris-HCl緩衝液(pH 7.0)及200mM NaCl的溶液沖提INF-β。根據其在280奈米處之吸收來收集包含INF-β的餾分。藉由疏水性交互作用管柱(GE保健(GE healthcare),丁基瓊脂糖凝膠快速流(Butyl Sepharose Fast Flow))(其已預先平衡及以包含1.0M硫酸銨、20mM醋酸鈉及0.05%次維特君(pH 4.5)的溶液清洗),來進一步純化包含在其中的INF-β。使用包含0.5M硫酸銨及20mM醋酸鈉的溶液沖提INF-β。根據其在280奈米處的吸收收集包含蛋白質之餾分。儲備這些餾分及藉由BCA蛋白質分析(BCATM
蛋白質分析,皮爾斯(Pierce))來測量INF-β濃度。
在水(1.46毫升)中之二-PEG醛(296毫克,7.4微莫耳)溶液中,加入2M磷酸鈉緩衝液(pH 4.0,0.37毫升),次維特君3-14(1.48毫升,10%在水中)及INF-β(14.8毫克在3.7毫升pH 4.5包含20mM醋酸鈉、0.7%硫酸銨及0.05%清潔劑之緩衝液中)。在室溫下攪拌該反應混合物10分鐘,接著加入氰
基硼氫化物水溶液(400mM,92.5微升,37微莫耳)。在黑暗中攪拌該反應混合物40小時及藉由SP HP瓊脂糖凝膠色層分析法純化。根據其滯留時間及在280奈米處的吸收來收集包含想要的PEG-INF-β綴合物之餾分。藉由BCA蛋白質分析(BCATM
蛋白質分析,皮爾斯)來測量綴合物濃度。
在大白鼠模型中進行藥物動力學研究,以比較INF-β及PEG-INF-β之血清半生期。對公大白鼠(250-350克)靜脈內給藥劑量600微克/公斤的INF-β(n=3)及PEG-INF-β(n=3)。在給藥前及在給藥後之0.1、1、2、4、6、10、24、48、72及96小時處,從每隻大白鼠收集血液(250微升)。從該血液製備血清樣品,及藉由酶連接的免疫檢定法(ELISA)分析包含在該樣品中的INF-β量。從最後三個時間點之血清濃度計算,INF-β及PEG-INF-β之血清半生期各別為2小時及20小時。
在水(2.67毫升)中之二-PEG醛(267毫克,6.1微莫耳)溶液中,加入2M磷酸鈉緩衝液(pH 4.0,1毫升)及EPO(10毫克在3.03毫升pH 7.3包含20mM磷酸鈉及150mM NaCl之緩衝液中)。在室溫下攪拌該反應混合物10分鐘,接著加入氰硼氫化鈉水溶液(400mM,100微升,40微莫耳)。在黑暗下攪拌該反應混合物17小時及藉由SP東友珍珠(Toyopearl)管柱(東曹(Tosoh))純化。該管柱與20mM醋酸鈉緩衝液,pH 4.5平衡。將該反應混合物稀釋至濃度0.3-0.4毫克/毫升並將其負載到SP東友珍珠管柱上。根據其滯留時間及在280奈米處的吸收來收集包含想要的PEG-EPO綴合物之餾分。藉由280奈米UV吸收來測量綴合物濃度。
在大白鼠模型中進行藥物動力學研究,以比較EPO及PEG-EPO之血清半生期。對公大白鼠(250-350克)靜脈內給藥劑量25微克/公斤之EPO(n=5)及PEG-EPO(n=5)。在給藥前及給藥後0.088、0.75、1.5、3、6、10、24及48小時,從每隻大白鼠抽出血液(250微升)。對經PEG-EPO治療的大白鼠來說,在給藥後72及96小時處進一步收集血液樣品。從該血液製備血清樣品及以酶連接的免疫檢定法(ELISA)分析以測量包含在其中之EPO量。結果顯示出EPO的血清半生期為9小時,同時PEG-EPO明顯增加(即,38小時)。
將0.2毫克的EPO(台灣的開士米科學公司(Cashmere Scientific Company))及4毫克的二-PEG醛(20當量)懸浮在0.1M磷酸鹽緩衝液(pH 5.0)中。在此溶液中,加入400當量的NaCNBH3
。在室溫下攪拌該反應混合物16小時。HPLC確認EPO-二-PEG聚合物形成。
在上述描述的發酵程序後,培養經轉化能表現出Met-hGH的大腸桿菌BLR(DE3)-RIL細胞,用以表現出Met-hGH。
經由離心採集細胞,且以比率大約1:3(潮溼重量克/毫升)將細胞丸粒懸浮在TE緩衝液(50mM Tris-HCl,1mM EDTA,pH 8.0)中。然後,藉由微射流均質機破裂該等細胞,然後在10,000rpm下離心20分鐘。以TED緩衝液(50mM Tris-HCl,1mM EDTA,2%去氧膽酸鹽,pH 8.0)清洗含包涵體(IB)之丸粒兩次,如上所述般離心,及懸浮在米里Q(MilliQ)水中及在20,000rpm下離心15分鐘。將IB懸浮在400毫升50mM TUD溶液(50mM Tris-HCl,4M尿素,2.5mM DTT,pH 10.0)中,及在20,000rpm下離心該懸浮液20分鐘;收集上層液。
在2.0升新鮮製備的再摺疊緩衝液(50mM Tris-HCl,0.5mM EDTA,5%甘油,10mM GSH/1mM GSSG,pH 8.0)中稀釋上層液。沒有攪拌,培養從而形成的混合物36小時,然後對著20mM Tris緩衝液(pH 7.0)透析。
將經透析包含Met-hGH的混合物負載到Q-瓊脂糖凝膠管柱(PA匹茲堡的GE阿默珊藥劑)上,其已預先平衡及以20mM Tris-HCl緩衝液,pH 7.0清洗。以包含20mM Tris-HCl緩衝液,pH 7.0及100mM NaCl的溶液沖提Met-hGH。收集、儲備包含Met-hGH的餾分(藉由其在280奈米處之吸收測量),且將其負載到疏水性交互作用管柱(PA的匹茲堡GE阿默珊藥劑)上,其已預先平衡及以20mM醋酸鈉緩衝液(pH 7.0),在流速5毫升/分鐘下清洗。以包含20mM醋酸鈉緩衝液及150mM硫酸銨的溶液沖提Met-hGH。收集包含Met-hGH之餾分及接受BCA蛋白質分析(BCATM
蛋白質分析,皮爾斯),以測量Met-hGH濃度。
在水(387微升)中的二-PEG醛(74毫克,1.7微莫耳)溶液中,加入2M磷酸鈉緩衝液(pH 4.0,374微升)及人類GH(22.4毫克在6.5毫升pH 4.5包含20mM醋酸鈉及150mM NaCl之緩衝液中)。在室溫下攪拌該反應混合物10分鐘,接著加入氰硼氫化鈉水溶液(400mM,140微升,56微莫耳)。在黑暗下攪拌該反應混合物17小時及藉由SP XL瓊脂糖凝膠色層分析法純化。根據其滯留時間及在280奈米處的吸收來收集包含想要的聚合物-蛋白質綴合物之餾分。使用布萊德福特(Bradford)方法(皮爾斯,洛克福特(Rockford),IL),藉由蛋白質分析工具組來測量綴合物濃度。
在大白鼠模型中進行藥物動力學研究,以比較Met-hGH及PEG-Met-hGH的血清半生期。對公大白鼠(250-350克)靜脈內給藥劑量100微克/公斤之Met-hGH(n=5)或PEG-Met-hGH(n=5)。在給藥前及在給藥後0.083、1、2、4、8、12及24小時,從經Met-hGH-治療的大白鼠收集血液樣品;及在給藥前及在給藥後0.33、1、4、8、12、24、48、72及96小時,從經PEG-Met-hGH-治療的大白鼠收集。從該血液製備血清樣品及以酶連接的免疫檢定法(ELISA)分析,以測量hGH濃度。Met-hGH及PEG-Met-hGH之血清半生期各別為3小時及35小時。
在本專利說明書中所揭示出的全部特徵可以任何組合結合。在本專利說明書中所揭示的每種特徵可由另一種提供相同、同等或類似目的之特徵置換。因此,除非其它方面有明確描述,否則所揭示的每種特徵僅為同等或類似特徵之一般系列的實施例。
熟習該項技術者可從上述描述中容易地查明本發明之基本特徵而沒有離開其精神及範圍,可對本發明製得多種改變及改質,使其適合於不同用途及條件。因此,其它具體實例亦在下列申請專利範圍之範圍內。
Claims (18)
- 一種下式之胜肽-聚合物綴合物:
- 如申請專利範圍第1項之綴合物,其中P為干擾素-β部分。
- 如申請專利範圍第2項之綴合物,其中P為重組的干擾素-β Ser17 (rINF-β Ser17 )。
- 如申請專利範圍第2項之綴合物,其中P為在N-終端處包含1-4個其它胺基酸殘基之經改質的干擾素-β部分。
- 如申請專利範圍第1項之綴合物,其中P為紅血球生成素部分。
- 如申請專利範圍第1項之綴合物,其中A1 及A2 各者為具有分子量2-100kD的聚乙二醇部分。
- 如申請專利範圍第6項之綴合物,其中A1 及A2 各者為具有分子量10-30kD的聚乙二醇部分。
- 如申請專利範圍第7項之綴合物,其中G1 及G2 各者為:
- 如申請專利範圍第8項之綴合物,其中m為4,n為2及R1 、R2 、R3 、R4 及R5 各者皆為H。
- 如申請專利範圍第9項之綴合物,其中P為干擾素-β部分。
- 如申請專利範圍第10項之綴合物,其中P為rINF-β Ser17 。
- 如申請專利範圍第10項之綴合物,其中P為在N-終端處包含1-4個其它胺基酸殘基之經改質的干擾素-β部分。
- 如申請專利範圍第9項之綴合物,其中P為紅血球生成素部分。
- 如申請專利範圍第9項之綴合物,其中P為生長激素部分。
- 如申請專利範圍第1項之綴合物,其中P為生長激素部分。
- 如申請專利範圍第1項之綴合物,其中該綴合物為:
- 如申請專利範圍第1項之綴合物,其中該綴合物為;
- 如申請專利範圍第1項之綴合物,其中該綴合物為:
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
US8507208P | 2008-07-31 | 2008-07-31 |
Publications (2)
Publication Number | Publication Date |
---|---|
TW201008583A TW201008583A (en) | 2010-03-01 |
TWI421093B true TWI421093B (zh) | 2014-01-01 |
Family
ID=41609027
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
TW098125858A TWI421093B (zh) | 2008-07-31 | 2009-07-31 | 胜肽-聚合物綴合物 |
Country Status (16)
Country | Link |
---|---|
US (2) | US8273343B2 (zh) |
EP (1) | EP2313457B1 (zh) |
JP (1) | JP5639585B2 (zh) |
KR (1) | KR101533757B1 (zh) |
CN (1) | CN102131844B (zh) |
AR (1) | AR072850A1 (zh) |
AU (1) | AU2009276458B2 (zh) |
BR (1) | BRPI0911722B1 (zh) |
EA (1) | EA020347B1 (zh) |
HK (1) | HK1158236A1 (zh) |
MX (1) | MX2011001167A (zh) |
MY (1) | MY156568A (zh) |
NZ (1) | NZ591167A (zh) |
TW (1) | TWI421093B (zh) |
UA (1) | UA104146C2 (zh) |
WO (1) | WO2010014874A2 (zh) |
Families Citing this family (1)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
HRP20231732T1 (hr) | 2014-11-06 | 2024-03-15 | Pharmaessentia Corporation | Režim doziranja pegiliranog interferona |
Citations (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US20040136952A1 (en) * | 2002-12-26 | 2004-07-15 | Mountain View Pharmaceuticals, Inc. | Polymer conjugates of cytokines, chemokines, growth factors, polypeptide hormones and antagonists thereof with preserved receptor-binding activity |
US20050107277A1 (en) * | 2002-01-18 | 2005-05-19 | Lin Kochung | Polyalkylene polymer compounds and uses thereof |
Family Cites Families (20)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US3643575A (en) * | 1967-10-31 | 1972-02-22 | Nippon Kogaku Kk | Exposure-measuring device in a single-lens reflex camera |
US5919455A (en) * | 1993-10-27 | 1999-07-06 | Enzon, Inc. | Non-antigenic branched polymer conjugates |
US5643575A (en) | 1993-10-27 | 1997-07-01 | Enzon, Inc. | Non-antigenic branched polymer conjugates |
US5951974A (en) * | 1993-11-10 | 1999-09-14 | Enzon, Inc. | Interferon polymer conjugates |
US5824784A (en) * | 1994-10-12 | 1998-10-20 | Amgen Inc. | N-terminally chemically modified protein compositions and methods |
US5932462A (en) * | 1995-01-10 | 1999-08-03 | Shearwater Polymers, Inc. | Multiarmed, monofunctional, polymer for coupling to molecules and surfaces |
EP1191189A1 (de) * | 2000-09-26 | 2002-03-27 | Siemens Aktiengesellschaft | Gasturbinenschaufel |
US7257546B2 (en) * | 2001-09-04 | 2007-08-14 | Yahoo! Inc. | System and method for correlating user data from a content provider and user data from an advertising provider that is stored on autonomous systems |
JP2005525302A (ja) * | 2001-11-20 | 2005-08-25 | ファルマシア・コーポレーション | 化学的に修飾されたヒト成長ホルモンコンジュゲート |
WO2004022630A2 (en) | 2002-09-09 | 2004-03-18 | Nektar Therapeutics Al, Corporation | Water-soluble polymer alkanals |
US20040142870A1 (en) * | 2002-11-20 | 2004-07-22 | Finn Rory F. | N-terminally monopegylated human growth hormone conjugates, process for their preparation, and methods of use thereof |
EP2644206B1 (en) * | 2003-05-23 | 2019-04-03 | Nektar Therapeutics | PEG derivatives containing two PEG chains |
AU2004293103C1 (en) * | 2003-11-24 | 2010-12-02 | Ratiopharm Gmbh | Glycopegylated erythropoietin |
US20060024953A1 (en) * | 2004-07-29 | 2006-02-02 | Papa Rao Satyavolu S | Dual damascene diffusion barrier/liner process with selective via-to-trench-bottom recess |
MX2007002441A (es) | 2004-08-31 | 2007-05-04 | Pharmacia & Upjohn Co Llc | Conjugados de hormona del crecimiento humana y polietilenglicol ramificado con glicerol, proceso para su preparacion y metodos de uso de los mismos. |
US7524318B2 (en) * | 2004-10-28 | 2009-04-28 | Boston Scientific Scimed, Inc. | Ablation probe with flared electrodes |
EP1869079A2 (en) * | 2005-03-11 | 2007-12-26 | Siegfried Ltd. | Di-polymer protein conjugates and processes for their preparation |
WO2006094530A1 (en) * | 2005-03-11 | 2006-09-14 | Siegfried Ltd. | Di-polymer protein conjugates and processes for their preparation |
EP1834963A1 (en) * | 2006-03-13 | 2007-09-19 | Siegfried Ltd. | Di-polymer protein conjugates and processes for their preparation |
CL2008002399A1 (es) * | 2007-08-16 | 2009-01-02 | Pharmaessentia Corp | Conjugado sustancialmente puro que posee una porcion polimerica, una porcion proteica (interferon alfa 2b) y un ligante alifatico de 1 a 10 atomos de carbono, util en el tratamiento de las hepatitis b o c. |
-
2009
- 2009-07-31 BR BRPI0911722-9A patent/BRPI0911722B1/pt active IP Right Grant
- 2009-07-31 JP JP2011521347A patent/JP5639585B2/ja active Active
- 2009-07-31 US US12/533,147 patent/US8273343B2/en active Active
- 2009-07-31 WO PCT/US2009/052347 patent/WO2010014874A2/en active Application Filing
- 2009-07-31 UA UAA201102277A patent/UA104146C2/ru unknown
- 2009-07-31 NZ NZ591167A patent/NZ591167A/en unknown
- 2009-07-31 CN CN200980130470.7A patent/CN102131844B/zh active Active
- 2009-07-31 TW TW098125858A patent/TWI421093B/zh active
- 2009-07-31 EA EA201170278A patent/EA020347B1/ru unknown
- 2009-07-31 AR ARP090102949A patent/AR072850A1/es active IP Right Grant
- 2009-07-31 KR KR1020117004412A patent/KR101533757B1/ko active IP Right Grant
- 2009-07-31 MX MX2011001167A patent/MX2011001167A/es active IP Right Grant
- 2009-07-31 AU AU2009276458A patent/AU2009276458B2/en active Active
- 2009-07-31 MY MYPI2011000446A patent/MY156568A/en unknown
- 2009-07-31 EP EP09803633.8A patent/EP2313457B1/en active Active
-
2010
- 2010-12-29 US US12/981,183 patent/US20110098451A1/en not_active Abandoned
-
2011
- 2011-11-25 HK HK11112846.8A patent/HK1158236A1/zh unknown
Patent Citations (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US20050107277A1 (en) * | 2002-01-18 | 2005-05-19 | Lin Kochung | Polyalkylene polymer compounds and uses thereof |
US20040136952A1 (en) * | 2002-12-26 | 2004-07-15 | Mountain View Pharmaceuticals, Inc. | Polymer conjugates of cytokines, chemokines, growth factors, polypeptide hormones and antagonists thereof with preserved receptor-binding activity |
Also Published As
Publication number | Publication date |
---|---|
NZ591167A (en) | 2012-06-29 |
BRPI0911722B1 (pt) | 2022-09-13 |
US8273343B2 (en) | 2012-09-25 |
US20100029907A1 (en) | 2010-02-04 |
MY156568A (en) | 2016-03-15 |
US20110098451A1 (en) | 2011-04-28 |
KR101533757B1 (ko) | 2015-07-03 |
CN102131844B (zh) | 2016-03-02 |
UA104146C2 (ru) | 2014-01-10 |
HK1158236A1 (zh) | 2012-07-13 |
AU2009276458B2 (en) | 2014-06-19 |
AU2009276458A1 (en) | 2010-02-04 |
TW201008583A (en) | 2010-03-01 |
AR072850A1 (es) | 2010-09-22 |
JP5639585B2 (ja) | 2014-12-10 |
EP2313457B1 (en) | 2020-01-15 |
JP2011529910A (ja) | 2011-12-15 |
MX2011001167A (es) | 2011-04-12 |
BRPI0911722A2 (pt) | 2016-09-13 |
EP2313457A2 (en) | 2011-04-27 |
EA020347B1 (ru) | 2014-10-30 |
EP2313457A4 (en) | 2014-10-01 |
CN102131844A (zh) | 2011-07-20 |
WO2010014874A2 (en) | 2010-02-04 |
EA201170278A1 (ru) | 2011-08-30 |
KR20110059600A (ko) | 2011-06-02 |
WO2010014874A3 (en) | 2010-05-06 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
EP2205281B1 (en) | Protein-polymer conjugates | |
TWI421093B (zh) | 胜肽-聚合物綴合物 | |
EP2509593B1 (en) | Protein-polymer conjugates |