CN101094867B - 用于固相肽合成的方法 - Google Patents
用于固相肽合成的方法 Download PDFInfo
- Publication number
- CN101094867B CN101094867B CN2005800435731A CN200580043573A CN101094867B CN 101094867 B CN101094867 B CN 101094867B CN 2005800435731 A CN2005800435731 A CN 2005800435731A CN 200580043573 A CN200580043573 A CN 200580043573A CN 101094867 B CN101094867 B CN 101094867B
- Authority
- CN
- China
- Prior art keywords
- gly
- glu
- resin
- pro
- peptide
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Expired - Fee Related
Links
- 239000007790 solid phase Substances 0.000 title claims description 32
- 238000000034 method Methods 0.000 title abstract description 20
- 238000010647 peptide synthesis reaction Methods 0.000 title description 2
- 239000011347 resin Substances 0.000 claims description 97
- 229920005989 resin Polymers 0.000 claims description 97
- 108090000765 processed proteins & peptides Proteins 0.000 claims description 46
- 125000006239 protecting group Chemical group 0.000 claims description 33
- DTQVDTLACAAQTR-UHFFFAOYSA-N Trifluoroacetic acid Chemical compound OC(=O)C(F)(F)F DTQVDTLACAAQTR-UHFFFAOYSA-N 0.000 claims description 26
- 239000002253 acid Substances 0.000 claims description 13
- 238000003776 cleavage reaction Methods 0.000 claims description 6
- 125000000484 butyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 claims description 5
- 229910052760 oxygen Inorganic materials 0.000 claims description 4
- 230000007017 scission Effects 0.000 claims description 4
- 125000002221 trityl group Chemical group [H]C1=C([H])C([H])=C([H])C([H])=C1C([*])(C1=C(C(=C(C(=C1[H])[H])[H])[H])[H])C1=C([H])C([H])=C([H])C([H])=C1[H] 0.000 claims description 4
- 125000003545 alkoxy group Chemical group 0.000 claims description 3
- DTGKSKDOIYIVQL-WEDXCCLWSA-N (+)-borneol Chemical compound C1C[C@@]2(C)[C@@H](O)C[C@@H]1C2(C)C DTGKSKDOIYIVQL-WEDXCCLWSA-N 0.000 claims 1
- DTGKSKDOIYIVQL-MRTMQBJTSA-N Isoborneol Natural products C1C[C@@]2(C)[C@H](O)C[C@@H]1C2(C)C DTGKSKDOIYIVQL-MRTMQBJTSA-N 0.000 claims 1
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical compound [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 claims 1
- CKDOCTFBFTVPSN-UHFFFAOYSA-N borneol Natural products C1CC2(C)C(C)CC1C2(C)C CKDOCTFBFTVPSN-UHFFFAOYSA-N 0.000 claims 1
- 125000002915 carbonyl group Chemical group [*:2]C([*:1])=O 0.000 claims 1
- DTGKSKDOIYIVQL-UHFFFAOYSA-N dl-isoborneol Natural products C1CC2(C)C(O)CC1C2(C)C DTGKSKDOIYIVQL-UHFFFAOYSA-N 0.000 claims 1
- 239000001301 oxygen Substances 0.000 claims 1
- 108010055460 bivalirudin Proteins 0.000 abstract description 13
- 230000002194 synthesizing effect Effects 0.000 abstract 1
- 238000010511 deprotection reaction Methods 0.000 description 31
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 description 26
- 230000002378 acidificating effect Effects 0.000 description 19
- 238000006243 chemical reaction Methods 0.000 description 17
- -1 aromatic carbenium ions Chemical class 0.000 description 16
- 239000000243 solution Substances 0.000 description 16
- 150000001413 amino acids Chemical class 0.000 description 14
- 238000005804 alkylation reaction Methods 0.000 description 13
- 239000000562 conjugate Substances 0.000 description 13
- 230000029936 alkylation Effects 0.000 description 12
- 125000003088 (fluoren-9-ylmethoxy)carbonyl group Chemical group 0.000 description 11
- OUYCCCASQSFEME-QMMMGPOBSA-N L-tyrosine Chemical compound OC(=O)[C@@H](N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-QMMMGPOBSA-N 0.000 description 11
- 235000001014 amino acid Nutrition 0.000 description 11
- 210000004899 c-terminal region Anatomy 0.000 description 11
- 239000000047 product Substances 0.000 description 11
- OUYCCCASQSFEME-UHFFFAOYSA-N tyrosine Natural products OC(=O)C(N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-UHFFFAOYSA-N 0.000 description 11
- 239000003875 Wang resin Substances 0.000 description 8
- NERFNHBZJXXFGY-UHFFFAOYSA-N [4-[(4-methylphenyl)methoxy]phenyl]methanol Chemical compound C1=CC(C)=CC=C1COC1=CC=C(CO)C=C1 NERFNHBZJXXFGY-UHFFFAOYSA-N 0.000 description 8
- 239000002516 radical scavenger Substances 0.000 description 8
- 238000007086 side reaction Methods 0.000 description 8
- OIRCOABEOLEUMC-GEJPAHFPSA-N bivalirudin Chemical compound C([C@@H](C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N1[C@@H](CCC1)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC=1C=CC(O)=CC=1)C(=O)N[C@@H](CC(C)C)C(O)=O)NC(=O)[C@H](CC(O)=O)NC(=O)CNC(=O)[C@H](CC(N)=O)NC(=O)CNC(=O)CNC(=O)CNC(=O)CNC(=O)[C@H]1N(CCC1)C(=O)[C@H](CCCNC(N)=N)NC(=O)[C@H]1N(CCC1)C(=O)[C@H](N)CC=1C=CC=CC=1)C1=CC=CC=C1 OIRCOABEOLEUMC-GEJPAHFPSA-N 0.000 description 7
- 229960001500 bivalirudin Drugs 0.000 description 7
- 229910052739 hydrogen Inorganic materials 0.000 description 7
- 239000001257 hydrogen Substances 0.000 description 7
- 238000010532 solid phase synthesis reaction Methods 0.000 description 7
- 239000002904 solvent Substances 0.000 description 7
- 125000001493 tyrosinyl group Chemical group [H]OC1=C([H])C([H])=C(C([H])=C1[H])C([H])([H])C([H])(N([H])[H])C(*)=O 0.000 description 7
- UHOVQNZJYSORNB-UHFFFAOYSA-N Benzene Chemical compound C1=CC=CC=C1 UHOVQNZJYSORNB-UHFFFAOYSA-N 0.000 description 6
- 239000004793 Polystyrene Substances 0.000 description 6
- 230000008878 coupling Effects 0.000 description 6
- 238000005859 coupling reaction Methods 0.000 description 6
- 238000005520 cutting process Methods 0.000 description 6
- 230000000694 effects Effects 0.000 description 6
- 239000000203 mixture Substances 0.000 description 6
- 229920000642 polymer Polymers 0.000 description 6
- 229920002223 polystyrene Polymers 0.000 description 6
- DURPTKYDGMDSBL-UHFFFAOYSA-N 1-butoxybutane Chemical compound CCCCOCCCC DURPTKYDGMDSBL-UHFFFAOYSA-N 0.000 description 5
- 125000001711 D-phenylalanine group Chemical group [H]N([H])[C@@]([H])(C(=O)[*])C([H])([H])C1=C([H])C([H])=C([H])C([H])=C1[H] 0.000 description 5
- 238000010168 coupling process Methods 0.000 description 5
- 229940006607 hirudin Drugs 0.000 description 5
- 230000008569 process Effects 0.000 description 5
- 238000010008 shearing Methods 0.000 description 5
- DHMQDGOQFOQNFH-UHFFFAOYSA-N Glycine Chemical compound NCC(O)=O DHMQDGOQFOQNFH-UHFFFAOYSA-N 0.000 description 4
- SECXISVLQFMRJM-UHFFFAOYSA-N N-Methylpyrrolidone Chemical compound CN1CCCC1=O SECXISVLQFMRJM-UHFFFAOYSA-N 0.000 description 4
- ISWSIDIOOBJBQZ-UHFFFAOYSA-N Phenol Chemical compound OC1=CC=CC=C1 ISWSIDIOOBJBQZ-UHFFFAOYSA-N 0.000 description 4
- 239000002585 base Substances 0.000 description 4
- 230000015572 biosynthetic process Effects 0.000 description 4
- 150000002431 hydrogen Chemical class 0.000 description 4
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 description 4
- 230000004048 modification Effects 0.000 description 4
- 238000012986 modification Methods 0.000 description 4
- 238000000926 separation method Methods 0.000 description 4
- 239000000126 substance Substances 0.000 description 4
- 238000010189 synthetic method Methods 0.000 description 4
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 4
- JFLSOKIMYBSASW-UHFFFAOYSA-N 1-chloro-2-[chloro(diphenyl)methyl]benzene Chemical compound ClC1=CC=CC=C1C(Cl)(C=1C=CC=CC=1)C1=CC=CC=C1 JFLSOKIMYBSASW-UHFFFAOYSA-N 0.000 description 3
- 108010016626 Dipeptides Proteins 0.000 description 3
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 3
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 description 3
- 238000002474 experimental method Methods 0.000 description 3
- 238000004128 high performance liquid chromatography Methods 0.000 description 3
- 238000004895 liquid chromatography mass spectrometry Methods 0.000 description 3
- 238000011068 loading method Methods 0.000 description 3
- 239000000463 material Substances 0.000 description 3
- 125000000956 methoxy group Chemical group [H]C([H])([H])O* 0.000 description 3
- BXRNXXXXHLBUKK-UHFFFAOYSA-N piperazine-2,5-dione Chemical compound O=C1CNC(=O)CN1 BXRNXXXXHLBUKK-UHFFFAOYSA-N 0.000 description 3
- 235000018102 proteins Nutrition 0.000 description 3
- 108090000623 proteins and genes Proteins 0.000 description 3
- 102000004169 proteins and genes Human genes 0.000 description 3
- 239000000376 reactant Substances 0.000 description 3
- HNICLNKVURBTKV-NDEPHWFRSA-N (2s)-5-[[amino-[(2,2,4,6,7-pentamethyl-3h-1-benzofuran-5-yl)sulfonylamino]methylidene]amino]-2-(9h-fluoren-9-ylmethoxycarbonylamino)pentanoic acid Chemical compound C12=CC=CC=C2C2=CC=CC=C2C1COC(=O)N[C@H](C(O)=O)CCCN=C(N)NS(=O)(=O)C1=C(C)C(C)=C2OC(C)(C)CC2=C1C HNICLNKVURBTKV-NDEPHWFRSA-N 0.000 description 2
- YDNMHDRXNOHCJH-UHFFFAOYSA-N 3-aminopyrrolidine-2,5-dione Chemical compound NC1CC(=O)NC1=O YDNMHDRXNOHCJH-UHFFFAOYSA-N 0.000 description 2
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 2
- COLNVLDHVKWLRT-MRVPVSSYSA-N D-phenylalanine Chemical compound OC(=O)[C@H](N)CC1=CC=CC=C1 COLNVLDHVKWLRT-MRVPVSSYSA-N 0.000 description 2
- CKLJMWTZIZZHCS-REOHCLBHSA-N L-aspartic acid Chemical compound OC(=O)[C@@H](N)CC(O)=O CKLJMWTZIZZHCS-REOHCLBHSA-N 0.000 description 2
- BZLVMXJERCGZMT-UHFFFAOYSA-N Methyl tert-butyl ether Chemical compound COC(C)(C)C BZLVMXJERCGZMT-UHFFFAOYSA-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
- XUYPXLNMDZIRQH-LURJTMIESA-N N-acetyl-L-methionine Chemical compound CSCC[C@@H](C(O)=O)NC(C)=O XUYPXLNMDZIRQH-LURJTMIESA-N 0.000 description 2
- JUJWROOIHBZHMG-UHFFFAOYSA-N Pyridine Chemical compound C1=CC=NC=C1 JUJWROOIHBZHMG-UHFFFAOYSA-N 0.000 description 2
- CZPWVGJYEJSRLH-UHFFFAOYSA-N Pyrimidine Chemical compound C1=CN=CN=C1 CZPWVGJYEJSRLH-UHFFFAOYSA-N 0.000 description 2
- BLRPTPMANUNPDV-UHFFFAOYSA-N Silane Chemical compound [SiH4] BLRPTPMANUNPDV-UHFFFAOYSA-N 0.000 description 2
- 229940122388 Thrombin inhibitor Drugs 0.000 description 2
- 238000009825 accumulation Methods 0.000 description 2
- 238000005903 acid hydrolysis reaction Methods 0.000 description 2
- 125000003368 amide group Chemical group 0.000 description 2
- 150000001408 amides Chemical class 0.000 description 2
- 238000004458 analytical method Methods 0.000 description 2
- 239000003146 anticoagulant agent Substances 0.000 description 2
- 230000000903 blocking effect Effects 0.000 description 2
- 239000003153 chemical reaction reagent Substances 0.000 description 2
- 239000000460 chlorine Substances 0.000 description 2
- 229910052801 chlorine Inorganic materials 0.000 description 2
- 239000011162 core material Substances 0.000 description 2
- 238000006073 displacement reaction Methods 0.000 description 2
- 238000001035 drying Methods 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 150000002148 esters Chemical class 0.000 description 2
- 238000001914 filtration Methods 0.000 description 2
- 125000000524 functional group Chemical group 0.000 description 2
- 238000007327 hydrogenolysis reaction Methods 0.000 description 2
- 238000010930 lactamization Methods 0.000 description 2
- 239000011159 matrix material Substances 0.000 description 2
- 229930182817 methionine Natural products 0.000 description 2
- 238000002156 mixing Methods 0.000 description 2
- 239000002245 particle Substances 0.000 description 2
- 125000000951 phenoxy group Chemical group [H]C1=C([H])C([H])=C(O*)C([H])=C1[H] 0.000 description 2
- 150000003053 piperidines Chemical class 0.000 description 2
- 239000003880 polar aprotic solvent Substances 0.000 description 2
- 230000036316 preload Effects 0.000 description 2
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 description 2
- 238000002271 resection Methods 0.000 description 2
- 238000003756 stirring Methods 0.000 description 2
- 239000000725 suspension Substances 0.000 description 2
- 238000003786 synthesis reaction Methods 0.000 description 2
- 238000012360 testing method Methods 0.000 description 2
- HNKJADCVZUBCPG-UHFFFAOYSA-N thioanisole Chemical group CSC1=CC=CC=C1 HNKJADCVZUBCPG-UHFFFAOYSA-N 0.000 description 2
- 239000003868 thrombin inhibitor Substances 0.000 description 2
- 238000005406 washing Methods 0.000 description 2
- 125000001834 xanthenyl group Chemical group C1=CC=CC=2OC3=CC=CC=C3C(C12)* 0.000 description 2
- MTCFGRXMJLQNBG-REOHCLBHSA-N (2S)-2-Amino-3-hydroxypropansäure Chemical compound OC[C@H](N)C(O)=O MTCFGRXMJLQNBG-REOHCLBHSA-N 0.000 description 1
- CBPJQFCAFFNICX-IBGZPJMESA-N (2s)-2-(9h-fluoren-9-ylmethoxycarbonylamino)-4-methylpentanoic acid Chemical compound C1=CC=C2C(COC(=O)N[C@@H](CC(C)C)C(O)=O)C3=CC=CC=C3C2=C1 CBPJQFCAFFNICX-IBGZPJMESA-N 0.000 description 1
- MATSKSZAPIUFCB-UHFFFAOYSA-N 2,3-bis(ethenyl)phenol Chemical compound OC1=CC=CC(C=C)=C1C=C MATSKSZAPIUFCB-UHFFFAOYSA-N 0.000 description 1
- 125000002103 4,4'-dimethoxytriphenylmethyl group Chemical group [H]C1=C([H])C([H])=C(C([H])=C1[H])C(*)(C1=C([H])C([H])=C(OC([H])([H])[H])C([H])=C1[H])C1=C([H])C([H])=C(OC([H])([H])[H])C([H])=C1[H] 0.000 description 1
- 239000004475 Arginine Substances 0.000 description 1
- 241000208199 Buxus sempervirens Species 0.000 description 1
- 0 CNC(CCCOc1cc(*)c(C*)cc1)=O Chemical compound CNC(CCCOc1cc(*)c(C*)cc1)=O 0.000 description 1
- 229930182832 D-phenylalanine Natural products 0.000 description 1
- SCCPDJAQCXWPTF-VKHMYHEASA-N Gly-Asp Chemical compound NCC(=O)N[C@H](C(O)=O)CC(O)=O SCCPDJAQCXWPTF-VKHMYHEASA-N 0.000 description 1
- 241000237677 Hirudinaria Species 0.000 description 1
- 241000237902 Hirudo medicinalis Species 0.000 description 1
- AYFVYJQAPQTCCC-GBXIJSLDSA-N L-threonine Chemical compound C[C@@H](O)[C@H](N)C(O)=O AYFVYJQAPQTCCC-GBXIJSLDSA-N 0.000 description 1
- 239000004472 Lysine Substances 0.000 description 1
- 108010033276 Peptide Fragments Proteins 0.000 description 1
- 102000007079 Peptide Fragments Human genes 0.000 description 1
- MTCFGRXMJLQNBG-UHFFFAOYSA-N Serine Natural products OCC(N)C(O)=O MTCFGRXMJLQNBG-UHFFFAOYSA-N 0.000 description 1
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 1
- NINIDFKCEFEMDL-UHFFFAOYSA-N Sulfur Chemical compound [S] NINIDFKCEFEMDL-UHFFFAOYSA-N 0.000 description 1
- 239000005864 Sulphur Substances 0.000 description 1
- AYFVYJQAPQTCCC-UHFFFAOYSA-N Threonine Natural products CC(O)C(N)C(O)=O AYFVYJQAPQTCCC-UHFFFAOYSA-N 0.000 description 1
- 239000004473 Threonine Substances 0.000 description 1
- 108090000190 Thrombin Proteins 0.000 description 1
- 208000007536 Thrombosis Diseases 0.000 description 1
- YXFVVABEGXRONW-UHFFFAOYSA-N Toluene Natural products CC1=CC=CC=C1 YXFVVABEGXRONW-UHFFFAOYSA-N 0.000 description 1
- VORIUEAZEKLUSJ-UHFFFAOYSA-M [(6-chlorobenzotriazol-1-yl)oxy-(dimethylamino)methylidene]-dimethylazanium;trifluoroborane;fluoride Chemical compound [F-].FB(F)F.C1=C(Cl)C=C2N(OC(N(C)C)=[N+](C)C)N=NC2=C1 VORIUEAZEKLUSJ-UHFFFAOYSA-M 0.000 description 1
- 125000002252 acyl group Chemical group 0.000 description 1
- 125000005073 adamantyl group Chemical group C12(CC3CC(CC(C1)C3)C2)* 0.000 description 1
- 230000002411 adverse Effects 0.000 description 1
- 239000003513 alkali Substances 0.000 description 1
- 125000005907 alkyl ester group Chemical group 0.000 description 1
- 125000000217 alkyl group Chemical group 0.000 description 1
- 125000004390 alkyl sulfonyl group Chemical group 0.000 description 1
- 238000005937 allylation reaction Methods 0.000 description 1
- 125000003277 amino group Chemical group 0.000 description 1
- 230000002785 anti-thrombosis Effects 0.000 description 1
- 229940127219 anticoagulant drug Drugs 0.000 description 1
- 239000004019 antithrombin Substances 0.000 description 1
- 239000012736 aqueous medium Substances 0.000 description 1
- ODKSFYDXXFIFQN-UHFFFAOYSA-N arginine Natural products OC(=O)C(N)CCCNC(N)=N ODKSFYDXXFIFQN-UHFFFAOYSA-N 0.000 description 1
- 125000000637 arginyl group Chemical group N[C@@H](CCCNC(N)=N)C(=O)* 0.000 description 1
- 125000004104 aryloxy group Chemical group 0.000 description 1
- 235000003704 aspartic acid Nutrition 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- 125000004429 atom Chemical group 0.000 description 1
- 229920005601 base polymer Polymers 0.000 description 1
- 125000000051 benzyloxy group Chemical group [H]C1=C([H])C([H])=C(C([H])=C1[H])C([H])([H])O* 0.000 description 1
- 125000001584 benzyloxycarbonyl group Chemical group C(=O)(OCC1=CC=CC=C1)* 0.000 description 1
- OQFSQFPPLPISGP-UHFFFAOYSA-N beta-carboxyaspartic acid Natural products OC(=O)C(N)C(C(O)=O)C(O)=O OQFSQFPPLPISGP-UHFFFAOYSA-N 0.000 description 1
- 230000002051 biphasic effect Effects 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 244000309464 bull Species 0.000 description 1
- 229910052799 carbon Inorganic materials 0.000 description 1
- 125000003178 carboxy group Chemical group [H]OC(*)=O 0.000 description 1
- 238000010531 catalytic reduction reaction Methods 0.000 description 1
- 238000001311 chemical methods and process Methods 0.000 description 1
- 239000003795 chemical substances by application Substances 0.000 description 1
- 238000005660 chlorination reaction Methods 0.000 description 1
- 238000005345 coagulation Methods 0.000 description 1
- 230000015271 coagulation Effects 0.000 description 1
- 239000000084 colloidal system Substances 0.000 description 1
- 239000012043 crude product Substances 0.000 description 1
- 230000007547 defect Effects 0.000 description 1
- 238000000151 deposition Methods 0.000 description 1
- 239000003085 diluting agent Substances 0.000 description 1
- 238000010790 dilution Methods 0.000 description 1
- 239000012895 dilution Substances 0.000 description 1
- 239000012153 distilled water Substances 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 239000003814 drug Substances 0.000 description 1
- 239000012065 filter cake Substances 0.000 description 1
- 239000000706 filtrate Substances 0.000 description 1
- 239000012530 fluid Substances 0.000 description 1
- 229920002313 fluoropolymer Polymers 0.000 description 1
- 239000004811 fluoropolymer Substances 0.000 description 1
- 239000012634 fragment Substances 0.000 description 1
- 108010001064 glycyl-glycyl-glycyl-glycine Proteins 0.000 description 1
- 125000001475 halogen functional group Chemical group 0.000 description 1
- HNDVDQJCIGZPNO-UHFFFAOYSA-N histidine Natural products OC(=O)C(N)CC1=CN=CN1 HNDVDQJCIGZPNO-UHFFFAOYSA-N 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- 230000008676 import Effects 0.000 description 1
- 239000003112 inhibitor Substances 0.000 description 1
- 230000001788 irregular Effects 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 230000007246 mechanism Effects 0.000 description 1
- GRVDJDISBSALJP-UHFFFAOYSA-N methyloxidanyl Chemical group [O]C GRVDJDISBSALJP-UHFFFAOYSA-N 0.000 description 1
- 238000001471 micro-filtration Methods 0.000 description 1
- 229910052757 nitrogen Inorganic materials 0.000 description 1
- 238000010534 nucleophilic substitution reaction Methods 0.000 description 1
- SBOJXQVPLKSXOG-UHFFFAOYSA-N o-amino-hydroxylamine Chemical group NON SBOJXQVPLKSXOG-UHFFFAOYSA-N 0.000 description 1
- 239000000863 peptide conjugate Substances 0.000 description 1
- 125000001151 peptidyl group Chemical group 0.000 description 1
- 239000012071 phase Substances 0.000 description 1
- 238000005191 phase separation Methods 0.000 description 1
- 150000002989 phenols Chemical class 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
- 229920005990 polystyrene resin Polymers 0.000 description 1
- 230000003389 potentiating effect Effects 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 102000004196 processed proteins & peptides Human genes 0.000 description 1
- 238000012545 processing Methods 0.000 description 1
- 230000017854 proteolysis Effects 0.000 description 1
- UMJSCPRVCHMLSP-UHFFFAOYSA-N pyridine Natural products COC1=CC=CN=C1 UMJSCPRVCHMLSP-UHFFFAOYSA-N 0.000 description 1
- 230000006340 racemization Effects 0.000 description 1
- 230000035484 reaction time Effects 0.000 description 1
- 230000009257 reactivity Effects 0.000 description 1
- 238000004064 recycling Methods 0.000 description 1
- 238000011160 research Methods 0.000 description 1
- 230000004044 response Effects 0.000 description 1
- 150000003839 salts Chemical class 0.000 description 1
- 229910000077 silane Inorganic materials 0.000 description 1
- 239000000741 silica gel Substances 0.000 description 1
- 229910002027 silica gel Inorganic materials 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 239000011877 solvent mixture Substances 0.000 description 1
- 125000006850 spacer group Chemical group 0.000 description 1
- 230000006641 stabilisation Effects 0.000 description 1
- 238000011105 stabilization Methods 0.000 description 1
- 230000003068 static effect Effects 0.000 description 1
- 239000000758 substrate Substances 0.000 description 1
- 235000012976 tarts Nutrition 0.000 description 1
- UGNWTBMOAKPKBL-UHFFFAOYSA-N tetrachloro-1,4-benzoquinone Chemical compound ClC1=C(Cl)C(=O)C(Cl)=C(Cl)C1=O UGNWTBMOAKPKBL-UHFFFAOYSA-N 0.000 description 1
- WROMPOXWARCANT-UHFFFAOYSA-N tfa trifluoroacetic acid Chemical compound OC(=O)C(F)(F)F.OC(=O)C(F)(F)F WROMPOXWARCANT-UHFFFAOYSA-N 0.000 description 1
- 230000001225 therapeutic effect Effects 0.000 description 1
- 229960004072 thrombin Drugs 0.000 description 1
- AAAQKTZKLRYKHR-UHFFFAOYSA-N triphenylmethane Chemical compound C1=CC=CC=C1C(C=1C=CC=CC=1)C1=CC=CC=C1 AAAQKTZKLRYKHR-UHFFFAOYSA-N 0.000 description 1
- LZTRCELOJRDYMQ-UHFFFAOYSA-N triphenylmethanol Chemical group C=1C=CC=CC=1C(C=1C=CC=CC=1)(O)C1=CC=CC=C1 LZTRCELOJRDYMQ-UHFFFAOYSA-N 0.000 description 1
- 238000000108 ultra-filtration Methods 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
- C07K14/81—Protease inhibitors
- C07K14/815—Protease inhibitors from leeches, e.g. hirudin, eglin
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K1/00—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length
- C07K1/04—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length on carriers
- C07K1/042—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length on carriers characterised by the nature of the carrier
Landscapes
- Chemical & Material Sciences (AREA)
- Health & Medical Sciences (AREA)
- Organic Chemistry (AREA)
- Medicinal Chemistry (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Biochemistry (AREA)
- Biophysics (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Life Sciences & Earth Sciences (AREA)
- Molecular Biology (AREA)
- Gastroenterology & Hepatology (AREA)
- Tropical Medicine & Parasitology (AREA)
- Analytical Chemistry (AREA)
- Peptides Or Proteins (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
- Organic Low-Molecular-Weight Compounds And Preparation Thereof (AREA)
Abstract
本发明提供了一种合成二价水蛭肽的新方法。
Description
本发明涉及和改进了抗凝血肽比伐卢定(bivalirudin)，即所谓的“二价水蛭素(hirulog)”的固相肽合成方法。本发明还涉及相应的肽-固相接合产物，该产物含有结合于树脂的仍受保护的肽。
凝血酶抑制剂被认为是很有前途的抗血栓药：通过凝血酶进行的蛋白水解过程是血液凝结控制的关键所在。水蛭素是一种获自吸血水蛭——医用水蛭Hirudo medicinalis的有效治疗性凝血酶肽抑制剂，其包含65个氨基酸。
业已证明了水蛭素第45-65位氨基酸的肽片段短肽类似物(即所谓的二价水蛭素)在治疗血栓症(一种危及生命的病症)中的有效性。
Okayama等(1996，Chem.Pharm.Bull.44：1344-1350)和Steinmetzer等(1999，Eur.J.Biochem.265：598-605)发明了在Wang树脂上固相合成不同二价水蛭素的方法，即将Fmoc氨基酸的C末端通过形成酯结合到被酯化为对-苄氧-苄醇基团的树脂上。使用Wang树脂需要用浓三氟乙酸将肽从树脂上切割下，其中树脂切割等于相伴进行的肽的完全脱保护(global deprotection)。
从Wang树脂上通过酸解切割(Acidolytic cleavage)是在强酸性条件下进行了，且已知如果在酸水解过程中不使用清除剂，则该方法会不可避免地引起作为副反应的Trp残基不良烷基化(Giraud等，1999，J.Peptide Science 5：457-461)。C末端Trp尤其容易发生该副反应(Atherton等，1988，Tetrahedron 44：843-857)。烷基化是由Wang树脂接头苯氧基部分产生的芳香性碳鎓离子所造成的。-当二价水蛭素不含Trp残基时，它们在邻近C末端的位置含有Tyr残基。我们发现并在此首次报告在从Wang树脂切割下时，该Tyr残基同样倾向于对烷基化不稳定，并对产物纯度产生不良的影响。
本发明的目的是消除现有技术中的缺点，提供不同的或改进的二价水蛭肽(Hirulog peptide)合成方法。
通过肽-树脂偶联物和本发明的相应合成方法来实现该目的。
在本发明中提供了一种用于对肽-固相偶联物进行分离和脱保护，最终获得肽的方法，所述肽优选具有式D-Phe-Pro-Arg-Pro-Gly-Gly-Gly-Gly-Asn-Gly-Asp-Phe-Glu-Glu-Tyr-Leu。所述肽-固相偶联物包括式I中的2-氯-三苯甲基手臂(handle)：
其中：A＝Boc-D-Phe-Pro-Arg(R2)-Pro-Gly-Gly-Gly-Gly-Asn(R3)-Gly-Asp(R4)-Phe-Glu(R5)-Glu(R6)-Ile-Pro-Glu(R7)-Glu(R8)-Tyr(R9)-Leu-O-，或A＝Fmoc-D-Phe-Pro-Arg(R2)-Pro-Gly-Gly-Gly-Gly-Asn(R3)-Gly-Asp(R4)-Phe-Glu(R5)-Glu(R6)-Ile-Pro-Glu(R7)-Glu(R8)-Tyr(R9)-Leu-O-，或A＝NH2-D-Phe-Pro-Arg(R2)-Pro-Gly-Gly-Gly-Gly-Asn(R3)-Gly-Asp(R4)-Phe-Glu(R5)-Glu(R6)-Ile-Pro-Glu(R7)-Glu(R8)-Tyr(R9)-Leu-O-，且其中R2、R3、R4、R5、R7、R8、R9是氨基侧链保护基团，R1是不溶性固相。
上述肽序列是二价水蛭素-8(Hirulog-8)的肽序列(描述于EP-489070)。它是水蛭素(65聚体，天然存在的强效凝血酶抑制剂)的20聚体二价衍生物。它是由水蛭素上功能上重要的、相连的结构基序组成的：水蛭素上的活性位点结合基序D-Phe-Pro-Arg-Pro和羧基末端序列Asn9到Leu20，通过四甘氨酸间隔物桥接。在此，‘-D-Phe-’是指D-苯丙氨酸，不同于给定氨基酸(在此为Phe)的天然L-对映体。
任选地，在本发明的另一目的中，式I中的基团A可为以下基团中的任意基团：
1.A＝P-X1-Tyr(R9)-X2-，其中X1是肽基部分，任选地包括位于各单独氨基酸侧链上的保护基，其具有0-200个，优选1-100个，最优选2-50个氨基酸，其中X2是与固相通过-O-或-NH-连接的任选地在侧链或末端被保护的单个氨基酸残基，X2最好不是Trp、Cys或Arg，且其中P是H(即得到NH2)或保护基，该保护基优选正交保护基(orthogonal protection group)或如下所定义的在强酸性条件下可去除的保护基，更优选该保护基选自：Boc、Fmoc、Dde、Nps、Alloc、Z。
应理解的是，根据上文所述，当其X2残基的侧链中包含氨基官能团(例如赖氨酸、高赖氨酸或诸如3-赖氨酸的结构性异构体)时，X2残基可以与树脂仅通过氨基醚官能团结合。反之，-O-连接可以是通过侧链或Cα羧基或侧链羟基官能团与手臂或接头相连的酯键或醚键。
2.A＝P-X1-Tyr(R9)-Leu-O或P-X1-Tyr(R9)-X2
3.A＝P-X1-Asp(R4)-Phe-Glu(R5)-Glu(R6)-Ile-Pro-Glu(R7)-Glu(R8)-Tyr(R9)-Leu-O或P-X1-Asp(R4)-Phe-Glu(R5)-Glu(R6)-Ile-Pro-Glu(R7)-Glu(R8)-Tyr(R9)-X2
4.A＝P-X1-[Gly]0-3-Asn(R3)-Gly-Asp(R4)-Phe-Glu(R5)-Glu(R6)-Ile-Pro-Glu(R7)-Glu(R8)-Tyr(R9)-Leu-O或P-X1--[Gly]0-3-Asn(R3)-Gly-Asp(R4)-Phe-Glu(R5)-Glu(R6)-Ile-Pro-Glu(R7)-Glu(R8)-Tyr(R9)-X2
5.A＝P-X1-Arg(R2)-Pro-Gly-Gly-Gly-Asn(R3)-Gly-Asp(R4)-Phe-Glu(R5)-Glu(R6)-Ile-Pro-Glu(R7)-Glu(R8)-Tyr(R9)-Leu-O-或P-X1-Arg(R2)-Pro-Gly-Gly-Gly-Asn(R3)-Gly-Asp(R4)-Phe-Glu(R5)-Glu(R6)-Ile-Pro-Glu(R7)-Glu(R8)-Tyr(R9)-X2
P、X1、X2的定义通用于各种A和所得的肽-固相偶联物的可能实施方式中。
我们发现并在此首次报道：从Wang树脂上切下时，所述Tyr残基也同样倾向于发生不规则的烷基化，这会对产品的纯度产生不良影响。对于二价水蛭素而言，这种修饰似乎受到邻位效应的促进，这与Atherton等对Trp的研究结果类似；然而，Tyr的烷基化(例如在精氨酸脱保护中)从未作为普遍现象被报道过，这与Trp的情形明显不同(Atherton等，1989，Solid phase synthesis：A practicalapproach，《固相合成：实践进展》IRL press，Oxford)。此外，Atherton的研究是仅限于C末端Trp，而以C末端到N末端的方向合成的二价水蛭肽中的Tyr残基，仅是邻末端的(juxtaproximal)，即生长中肽链C末端的倒数第二个残基。事后发现，不限于任何理论，这可能是因为苯氧基部分在亲电子取代中比一般芳基化合物反应性更强。事实上，酚类是从树脂上进行酸解剪切时的清除剂(scavenging agent)(D.S.King等，1990，Int.J.Peptid Protein Res.，36，255)。至此，技术人员仍不曾描述过或提及所述副反应(side-reaction)，至今仍相信仅末端Trp容易被无规烷基化。因此，直至今日，Wang树脂仍被广泛用于二价水蛭素的合成。
可通过本领域熟知的常规固相方法合成本发明的肽-固相偶联物，Bodanszky等的Principles of Peptide Synthesis，(《肽合成原理》第二版，SpringerVerlag Berlin Heidelberg 1989)对此类方法有详细描述，可作为参考。由于固相连接的酸不稳定性，这样的合成策略中必须利用Fmoc化学方法进行固相合成期间的偶联反应。只有最后的封端D-Phe残基可以是Boc-或Fmoc保护的。可在肽仍连于树脂上时消除该Fmoc保护，可采用例如20％哌啶或其它Fmoc脱保护碱试剂的标准处理进行，并由此获得了本发明的肽-树脂偶联物(conjugate)，但其带有游离的N-末端氨基。然而，这样的早期Fmoc脱保护，早早地暴露出N-末端，会在通过酸解从树脂上分离，尤其是在强酸性条件下分离的同时进行全面脱保护时，使得或可能使得所述游离N-末端D-Phe残基更容易发生外消旋。因此，更为优选的是，对封端D-Phe残基进行Boc-保护，或用在强酸条件下易于去除的其它保护基保护，从而无需单独的Fmoc脱保护步骤。为了清楚起见，这包括例如Z-(苯甲氧基-羰基-)保护基，该保护基可在本发明所述的(特别是)强酸性条件被切除，尽管已知氢解或HF促进的切割更为有效。再次指出，封端D-Phe上的单独的Fmoc脱保护步骤早早地暴露出的N-末端(以游离α-氨基封端，在式I中表示为H-D-Phe-或NH2-D-Phe，两者同义)，会在通过酸解从树脂上分离同时进行全面脱保护时使得脱保护后的游离N-末端D-Phe容易发生外消旋化，虽然这不是一种优选，但仍是本发明可行的实施方式之一。所述的一步分离或剪切的同时全面脱保护可在溶剂混合物(例如TFA和DCM水性混合物)中进行。
总体而言，根据本发明，既可以在氨基酸侧链(优选的N-末端的保护基)脱保护或完全脱保护的同时也可以在此前的起始步骤中将式I中的受保护的肽从树脂上切下。在后一种实施方式中，依次是先在弱酸条件下从树脂上进行切割，然后是在强酸条件下切除所有残留的保护基(全面脱保护)。
无论是在两种条件中的任何一种之下，尤其是2-氯-三苯甲基-树脂(简称CTC树脂)以及例如市售的十分相似的4-甲氧基-或4-甲基-三苯甲基-树脂，或者，在本发明包括的它树脂也一样或在相当程度上十分适用于在剪切和/或脱保护时避免近端酪氨酸被不当修饰。当近端酪氨酸(即位于C末端的倒数第二个酪氨酸)在肽从树脂上切下时的同时脱保护时，这防止了该酪氨酸不为所需的烷基化。由于卤代，CTC树脂可以使得受保护的肽和酪氨酸在非常温和的酸解反应条件(例如在0.5％三氟乙酸(TFA)的二氯甲烷(DMC)溶液中)下从树脂上被有效地切下，在该条件下大部分侧链和N-末端保护基通常不会受到影响，由此通过及时分离不同的脱保护事件而防止了烷基化。下文中，以CTC树脂(作为固相或树脂的最优选实施方式)进行的具体实施例默认(tacidly)也可用本发明说明书和权利要求中所描述的其它树脂进行。
根据本发明中的定义，相对于弱酸性条件的强酸性条件是指采用浓度至少50％(v/v)的三氟乙酸(TFA)溶液。并且，反过来，需要用强酸条件来去除的保护基是至少可以80％TFA去除的保护基。因此，在本发明全文中上述的定义不包括需要更强酸(例如HF)的保护基。将弱酸性条件定义为具有0.01％(v/v)到＜50％的TFA，优选0.1％到30％的TFA。
在任一种模式中，值得注意的是，本发明的肽基部分表现为出乎意料地没有不期望的近端酪氨酸的烷基化，且完全没有二酮哌嗪副反应(在肽从树脂上切下时可能会发生的另一副反应，且已知其对于C末端最后两个氨基酸的性质尤为敏感)。不希望受限于理论，据推测，肽链上近CTC树脂手臂第2位上的酪氨酸恰处于最佳距离和间隔，从而显现出一定程度的芳性苯基部分的稳定化作用和疏水性堆积作用，避免了例如环化现象，这一环化现象正是二酮哌嗪形成的前奏。
CTC树脂的加载(load)通常是通过联苯-2’-氯苯基-氯甲烷衍生物(因此将其称为CTC，即氯化氯-三苯甲基的简称)的亲核取代进行的，已知其是有效的。作为一种选择，也有市售的经预载的Fmoc-氨基-酸-CTC树脂。
保护基和它们的化学性质是本领域熟知的，且有广泛的记载(参见Bodanszky，同上)。毋庸置疑，不同的保护基R2-R9分别适于保护独特的各氨基酸侧链，因为不同的化学部分需要使用不同保护基。例如，通常可用三苯甲基或Boc来保护组氨酸，用Boc或烯丙氧羰基来保护赖氨酸，用叔丁醚或烯丙酯来保护天冬氨酸。通常将苏氨酸、丝氨酸和酪氨酸也保护为叔丁醚。在下文中还将讨论对精氨酸的保护。可施用不同的脱保护模式，例如通过Pd催化的还原酰基转移反应费力地(laborously)去除烯丙基化的保护基。Z(苯甲氧基羰基)基团使用起来较不方便，这是因为其需要氢解才能有效去除。优选地，保护基R2-R9是酸不稳定性的，‘不稳定’是指在弱酸性或强酸性条件下，在DCM溶液中孵育5小时，所述各保护基的切除率至少为20％。根据本发明所述，更优选的是，在上文所述的强酸性条件下并仅在该条件下(即通过强酸性条件下的酸解方式)去除所述保护基R2-R9。
R1是不溶性的固相，通常为聚合物固相，例如交联的聚苯乙烯/1％二乙烯基苯酚共聚物。通常(但不是严格要求)，本发明操作中该固相R1当然还可以呈现出多个以肽基A官能化的2-氯-三苯甲基-手臂部分，而非仅限于式I中明示的那一个。更重要的是，为了用于由Merrifiled首次提出的固相合成中，聚合物固相的最小粒径需能够形成具有足够大小的易过滤或易沉淀颗粒的的真悬浮液而不是胶体特性。除了直接用CTC手臂或接头(例如Bayer的4-羧基三苯甲基衬里(liner)，Bayer等，13th American Peptide Symposium，Hodges等编，ESCOM，Leiden，1994，第156页)衍生的聚苯乙烯主体聚合物(base polymer)所述主体聚合物中的个别苯部分已经被衍生化从而参与形成2-氯-三苯甲基官能)以外，还有其它主体聚合物，例如纯PEG或混合PEG树脂(例如Tentagel)或任选的混合或接枝树脂(hybrid or grafted resins)，例如通过PEG间隔基将2-CTC接头(如Bayer接头)接枝到聚苯乙烯基聚合物上而不是直接使该接头与聚苯乙烯基聚合物反应。将PEG加入树脂提供了更具两亲性的树脂，并由此获得更佳的操作性，例如在DCM/TFA混合物中进行一步分离和脱保护时的操作性，虽然负载容量可能会成问题。应注意的是，当然存在严格意义上的不溶性PEG树脂。但在Bayer等的一种技术(Nature 1972，第237卷，第512f页)记载了一种模仿固相分离原理但严格在溶液中工作的基于PEG聚合物的技术，其中的肽-树脂偶联物仍然是可溶的，并提供了均一的单相系统。根据本发明上下文的优选含义中，这样的树脂特性包括在本发明的“不溶性”定义中，因为这样的树脂特性基本上允许通过微滤或超滤技术在微观水平上快速简便地基于大小来进行分离。在更优选的含义中，“不溶性”是指在用于肽合成的给定溶剂系统中，两相系统中的一相为真正的固体悬浮相。
优选地，所述固相的粒度小于700目(粒度如US标准局所定义，可在例如Chemie-Lexikon，7.Auflage，1973，Franck’sche Verlagshandlung，W.Keller&Co.Stuttgart/Germany检索到)。
优选地，本发明的2-氯-三苯甲基-官能化固相的粒度为50-600目(如US标准局所定义)，更优选60-400目，最优选100-300目。
可用不同的保护基来保护本发明的酪氨酸，例如叔丁醚，或者Z-，或更优选2-溴-Z酯。同样，也可采用三苯甲基醇保护基，例如2-氯-三苯甲基或4-甲氧基或4，4’甲氧基-三苯甲基。优选地，R9是三苯甲基或叔丁基保护基。更优选地，R9是叔丁基(tBu)保护基，这是指酪氨酰侧链被修饰为叔丁醚。tBu基团仅可在强酸性条件下被有效去除。
优选地，单独和尤其是结合其它优选的实施方式，精氨酸保护基R2选自：五甲基二氢苯并呋喃基(Pbf)、金刚烷氧基-羰基和异冰片基-氧-羰基、五甲基色原磺酰基(pentamethylenchromanesulfonyl，Pmc)、4-甲氧基-2，3，6-三甲基苯磺酰基(Mtr)及其4-叔丁基-2，3，5，6-四甲基同系物(Tart)或Boc，它们仅在上文所述的强酸性条件下被切除。R2更优选为Pbf、Pmc、Mtr，最优选为Pbf；在强酸性条件下且通常在水性介质中进行的侧链完全脱保护后，采用Pmc、Mtr和尤其是Pbf保护未观察到脱保护酪氨酸的旁观烷基化(bystander-alkylation)。Pbf的切除率最高。
众所周知羧基-保护基可用于Glu、Asp，例如可使用Mpe、O-1-金刚烷基、O-苄基，甚至于更简单的烷基酯，虽然不是很常用。为了简便起见，通常并优选独立地将叔丁基用作保护基R4、R5、R6、R7、R8。
保护基R3至关重要，因为其存在于二价水蛭素-8中的上述序列Gly-Asp中，该二肽序列尤其倾向于发生副反应而形成天冬酰亚胺。天冬酰亚胺的形成会在线性合成过程中接着的每一轮偶联反应中于受保护的肽中发生，反应程度轻微(0.1-0.5％)，但在最后可表现出聚积效应。同样，虽然优选用三苯甲基保护基或其2-氯和4-甲基或4-甲氧基衍生物，但也可使用金刚烷基保护基。最优选使用三苯甲基保护基。
还应注意到的是，在线性合成中可使用Nα-烷基保护二肽模块来代替侧链和Nα保护氨基酸偶联；该二肽具有干扰二级结构的作用，使得合成产率和纯度更佳。例如Fmoc-Gly-(N-Hmb)Gly-OH和Fmoc-Gly-(N-Dmb)Gly-OH可从EMD Biosciences(Novabiochem)市售购得。应理解的是，该N-烷基基团在本发明中不被认为是保护基，因此，它们的使用或存在是任选的且并不排除在式I结构之外。
在基本上如各权利要求中所述的式I所示肽-偶联物的分离和脱保护的优选方法中，使用了二步骤连续方案，首先在弱酸性条件下进行酸解以从CTC树脂上切下受保护的肽，然后在强酸性条件下去除残留的保护基。
采用这一方案的原因在于式I肽-固相偶联物的一步全面脱保护中完全脱保护产物和疏水性接合离析物对溶剂要求相互抵触，为此进行折中处理对产物的纯度和产率都有负面影响。该连续多步骤方法消除了上述内在缺陷，从而能够更好地控制不同的反应，并由此优化产率。根据本发明，其令人惊讶的效果还包括完全抑制了作为副反应的二酮哌嗪形成。
因此，设计了一种对如前所述式I所示肽-固相偶联物进行分离和脱保护以获得式D-Phe-Pro-Arg-Pro-Gly-Gly-Gly-Gly-Asn-Gly-Asp-Phe-Glu-Glu-Ile-Pro-Glu-Glu-Tyr-Leu所示的肽的方法，该方法的特征在于：在第一步中，通过弱酸性条件下的处理，从2-氯三苯甲基手臂上切下受保护的肽，优选使用0.1-10％的TFA在极性疏质子溶剂中的溶液，在第二步中，如上所述地在强酸性条件下去除保护基。
优选地，所述第一步在极性疏质子溶剂中进行，所述溶剂为二氯甲烷。与其它溶剂(例如NMP，N-甲基吡咯烷酮)相比，这是进行该反应的最佳溶剂。还可以(而非必须)在溶剂中包含清除剂，尤其是用于第二脱保护步骤中的溶剂系统，清除剂其存在的量为反应液(reaction broth)的0.1-10％(w/w)，用于防止酪氨酸的芳核再次发生不为所需的烷基化。清除剂阻断了去除保护基时产生的反应性烷基-碳鎓离子中间体(可能在第一步中的剪切反应期间已轻微发生)。
清除剂的例子为苯甲硫醚(其还具有第二性的酸解促进效果-该第二作用和苯甲硫的替代物在Bodanszky M等，Int.J.Peptide Protein Res.23：287中有所讨论)。不具有该酸解效果的其它清除剂的例子是苯酚和/或三烷基硅烷(Stierandova等，Int.J.Peptide Protein Res.43，1994，31-38)。
优选在第一步从树脂上切下或分离后，通过与吡啶混合直接淬灭反应，然后通过与水混合来回收步骤1的产物。通过该方法可最简单有效地回收产物。
在本发明的另一实施方式中，涉及的肽与式I的肽-固相偶联物基本上被要求，唯一的不同是凝血酶切割位点-Arg(R2)-Pro-不是标准的肽键而是化学修饰的拟易裂键(pseudoscissile)或‘psi’键(酰胺键的置换用增加的括号中带‘psi’前缀的原子表示，参见Rudinger等，Drug Desgin Vol.II，编辑Ariens，E.，AcademicPress，New York，p.319(1971)。更优选，该psi置换是-Arg[psiCH2NH]Pro-(Kline，T等，1991，Hirolog peptides with scissile bond replacements resistant to thrombincleavage，带有易裂键置换的抗凝血酶切割的二价水蛭肽，Biochem.Biophys.Res.Commun.177，1049-1055)。最简单的是，例如，该psi键在固相合成中立即用预制的Fmoc保护的psi二肽与生长中的接合状态的肽正常偶联来导入。
本发明的另一目的是将上述实施方式和方法扩展到包含上述CTC树脂以外的树脂部分的肽-固相偶联物，这样扩展后，所述方法仍可在如前所述的弱酸性或温和酸性条件下从树脂上切下肽部分。
作为另一目的，设计了式A-W所示的肽-树脂偶联物，其中A可为任何前文所述A的实施方式，任选地包含各氨基酸侧链保护基团，其中所存在的R2-R9如上所述，其中W是一优选为不溶性的固相或固相复合物，其在肽部分可在弱酸性条件下得以切除并具有下所示的树脂手臂或接头
a.式II
前提是包含残基X2的A总是通过-O-与所示手臂或接头连接，
其中R″′是固相，R″1、R″2、R″3相同或不同，各自独立地为氢、4-或4′-(C1-C4烷基)或4-或4′-(C1-C4烷氧基)，但是R″1和R″2不可同时是氢，当R″1是H时，R″任选地为2-Cl，且其中更优选或最优选的是，式II的手臂或接头选自2-氯-三苯甲基、4-甲氧基-三苯甲基、4-甲基三苯甲基，
b.或式III
(其可通过用Bayer的4-羧基三苯甲基接头的酰化氨基-或羟基官能化树脂衍生而来)前提是同样包含残基X2的A通过-O-连接到所述手臂或接头，R″′如上所定义，
c.或式IV
其中R″′是固相或聚合物树脂，而R″1、R″2、R″3相同或不同，独立地为氢、C1-C4烷基或C1-C4烷氧基，但R″1和R″2不可同时为氢，其中L是A(L＝A)或其中L是式V
在另一优选的实施方式中，所述树脂手臂具有式VI，R″′、R″1和R″2的定义同上，
如前所述，更为优选的是树脂和树脂手臂具有式VII，R″′、R″1和R″2的定义同上，
在另一更为优选的实施方式中，任选包括残基X2的A通过-O-与式VII的手臂或接头连接，R″1和R″2不同时为氢时，R″1和R″2独立地为氢、甲基或甲氧基；当包含残基X2的A通过-N-与式VII的手臂或接头连接时，R″1和R″2独立地为甲基或甲氧基，优选甲氧基。更为优选的是，同样包含X2的A通过-O-官能团结合于手臂，R″1是氢，R″2是甲基或甲氧基，优选A是树脂或树脂手臂。最优选R″2为甲基。
树脂或树脂手臂组合部分原则上可为用于合成的任何树脂，例如Merrifield用来与羟苄基-苯基接头部分(作为树脂的一部分)一起使用或Wang用来与羟基-苄基-对苄氧基部分一起使用的聚苯乙烯-二苯乙烯树脂，所述一起使用的部分包括例如可进一步接枝更为酸不稳定性的接头，或者可使得后一种接头整合或直接连接到树脂上的那些。原则上，用于合成的固相树脂必须包含至少一个作为其一部分的接头或手臂，所述接头或手臂是固相核心材料的一部分；可将该接头或手臂认为是固定化的保护基(Guillier等，Chem.Rev.100，2091-2157，2000)。例子为例如：Sieber树脂、相关的呫吨基(related xanthenyl)型PAL-手臂树脂、Rink酰胺树脂、Rink酸性树脂、更复杂的PEG接枝的聚苯乙烯树脂(例如基于tentagel的Novasyn TG(Novabiochem，Merck Biosciences，德国)，现成的此类树脂具有不同接枝手臂(例如2′-氯-三苯甲基))，或通过将功能性手臂接枝到基质材料(例如硅胶)上构建而成的树脂。优选的是，如果所述树脂是三苯甲基树脂或树脂手臂，那么它是选用4-甲氧基或4，4′-二甲氧基-三苯甲基树脂。用于本发明中的树脂具有标准的粒径，约为50-500目，更优选100-400目。如式IV中所示的树脂或固相R″′应理解为含有交联的聚合物基质材料，该材料可通过任何种类的化学惰性烷基、烷氧基、芳氧基或烷基酯间隔基或接头(其被认为是R″′的构成部分)结合于式IV-VII所示的手臂部分。然而，应注意的是，除了对从树脂上剪切的条件有影响以外，树脂材料的性质，尤其是手臂基团的化学性质，将极大地影响偶联和内酰胺化反应(lactamisation)的效力，但影响的机制尚不甚明了。树脂上阶段(on-resin stage)中成熟肽的产率根据所用树脂或树脂手臂的类型不同而不同。出于这一原因，在本发明的优选实施方式中，树脂或树脂手臂如式IV所示(如权利要求所述)，更优选具有式VI(如权利要求所述)，最优选的是式VII(如权利要求所述)。此类树脂或树脂手臂的例子是(4-甲氧基苯基)-甲基-和(4-甲基苯基)-甲基-聚苯乙烯(Atkinson等，2000，J.Org.Chem.65，5048)，以O-或N-连接键连接到肽部分的树脂，和它们的PEG-树脂衍生物。其它的例子为例如：酸不稳定性HMPB-MBHA或HMPB-BHA树脂(Sieber等，1987，Tetrahedron Lett.28，6147)、酸不稳定性Rink酰胺树脂或Rink酸性树脂(Rink等，1987，Tetrahedron Lett.28，3787)。术语“酸不稳定性”是指在环境温度下在2-10％TFA的二氯甲烷溶液中至少1小时，发生实质上可定量的切割。令人惊讶的是，采用具有联苯-甲基结构核心基序的此类优选树脂提高了线性合成过程中偶联反应和内酰胺化反应的效率；值得注意的是该树脂可在更低的反应温度例如15-25℃，而在例如三苯甲烷树脂上实现有效偶联所需的标准温度则是40℃。
实验
1.Boc-D-Phe-Pro-Arg(Pbf)-Pro-Gly-Gly-Gly-Gly-Asn(Trt)-Gly-Asp(tBu)-Phe-Glu(tBu)Glu(tBu)-Ile-Pro-Glu(tBu)-Glu(tBu)-Tyr(tBu)-Leu-O-2-CTC(受保护的二价水蛭素-8，详见EP-489070，通过羧基末端以酯键连接到2-CTC树脂)的合成
所有试剂均获自EMD Biosciences(Madison，WI/U.S.A.；Novabiochem-brand)。以Fmoc-Leu-OH预载的基于聚苯乙烯的2-ClTrt(CTC)树脂(Cbl Patras，Greece)以其基体聚合物计为100-200目，以预装载后的最终CTC树脂产物计为60-200目。加载密度约为0.60mmol/g。各氨基酸以Fmoc氨基酸形式提供，或当其为D-Phe时，作为已受Boc-保护的Boc-D-Phe形式提供。在二氯甲烷/N-甲基吡咯烷酮(NMP)溶液中，在Hünig-Base(二异丙基-乙基-胺，DIEA)的存在下，用TCTU进行偶联。通常，采用1.5当量Fmoc氨基酸或Boc保护的氨基酸，但Fmoc-Arg(Pbf)的偶联反应使用2.5当量。类似地，Fmoc-Arg(Pbf)的偶联反应时间由标准的60分钟(30℃)延长到90分钟。过程中，采用Kaiser测试或Chloranil测试方法进行偶联效力的控制。
在30℃用3-4个循环的20％哌啶的NMP溶液(期间采用NMP适当清洗)来进行Fmoc的脱保护。
2.Boc-D-Phe-Pro-Arg(Pbf)-Pro-Gly-Gly-Gly-Gly-Asn(Trt)-Gly-Asp(tBu)-Phe-Glu(tBu)Glu(tBu)-Ile-Pro-Glu(tBu)-Glu(tBu)-Tyr(tBu)-Leu-OH的合成
经3轮15分钟、15℃、2％(w/w)TFA、l％(w/w)三乙基硅烷(TES)、在二氯甲烷溶液中的反应完成从48.3g树脂(如实施例1所述制得)上的剪切。通过氮气鼓泡对反应物进行搅拌；反应物的颜色随反应轮数从黄色/橙色变为褐色。每轮反应后，通过将反应液全部注入稀嘧啶(嘧啶/乙醇1∶9(v/v))直接淬灭剪切反应。然后用玻璃料(frit)过滤分离出树脂用于下一轮反应。汇集所有的滤液，真空(RotaVap)下浓缩至橙色半流体状，用DCM洗涤，重悬于400mk双蒸水中，在室温下搅拌，过滤，以水洗涤，干燥。所得为分析级(～90％纯度)的28,8g的浅黄色粉末。用HPLC和LC-MS分析产物。
3.全面脱保护，NH2-D-Phe-Pro-Arg-Pro-Gly-Gly-Gly-Gly-Asn-Gly-Asp-Phe-Glu-Glu-Ile-Pro-Glu-Glu-Tyr-Leu-OH的合成
在剪切混合物(cleavage cocktail，‘CC’)的DCM稀释液(DCM∶‘CC’＝1∶10(v/v))中进行全面脱保护。‘CC’由混合比为(％w/w)89∶2.5∶2.5∶5.0∶1的TFA/苯甲硫醚/苯酚/水/TES组成。将0.1g的实验2干燥产物溶于10ml如上所述以‘CC’稀释的DCM，在室温下搅拌5小时。然后通过加入50ml甲基-叔丁醚(MTBE，Fluka Chemie，Buchs/Switzerland)回收产物，搅拌下水浴30分钟将反应物冷却到0℃，滤除期间形成的盐沉淀物。滤饼用MTBE洗涤数次，并在室温下进行干燥，得到0.8g的粗产物，以HPLC测定的纯度约为55％。步骤2和3的联合总产率约为55％。
4.Wang树脂或CTC树脂上二价水蛭素-8或其C末端四肽片段合成的比较剪切实验和LC-MS分析
采用HPLC LC-MS分析方法，结果显示，从树脂上剪切下来并在强酸性条件下全面脱保护后，使用Wang树脂的肽产品有1-10％的被烷基化，从CTC树脂上切下后则没有观察到这样的修饰。经MS分析定位发生在酪氨酰残基上的修饰。合成步骤如上所述。
Claims (4)
1.式I所示的肽-树脂偶联物，
其中R1是不溶性固相，所述固相是聚合的且按照US标准局粒度为100-400目其中
A＝Boc-D-Phe-Pro-Arg(R2)-Pro-Gly-Gly-Gly-Gly-Asn(R3)-Gly-Asp(R4)-Phe-Glu(R5)-Glu(R6)-Ile-Pro-Glu(R7)-Glu(R8)-Tyr(R9)-Leu-O-或A＝Fmoc-D-Phe-Pro-Arg(R2)-Pro-Gly-Gly-Gly-Gly-Asn(R3)-Gly-Asp(R4)-Phe-Glu(R5)-Glu(R6)-Ile-Pro-Glu(R7)-Glu(R8)-Tyr(R9)-Leu-O-或A＝NH2-D-Phe-Pro-Arg(R2)-Pro-Gly-Gly-Gly-Gly-Asn(R3)-Gly-Asp(R4)-Phe-Glu(R5)-Glu(R6)-Ile-Pro-Glu(R7)-Glu(R8)-Tyr(R9)-Leu-O-，且其中R2是五甲基二氢苯并呋喃基、金刚烷氧基-羰基或异冰片氧基羰基，R9是叔丁基，R3-R8是酸不稳定性保护基。
2.如权利要求1所述的肽-树脂偶联物，其特征在于，R2是Pbf，R3-R9是需要至少50％三氟乙酸来去除的酸不稳定性保护基。
3.如权利要求2所述的肽-树脂偶联物，其特征在于，R3是三苯甲基，R4、R5、R6、R7和R8是叔丁基。
4.如权利要求1所述的肽-树脂偶联物，其特征在于，凝血酶切割位点-Arg(R2)-Pro-是-Arg[psiCH2NH]Pro-。
Applications Claiming Priority (3)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| EP04024812.2 | 2004-10-19 | ||
| EP04024812 | 2004-10-19 | ||
| PCT/EP2005/011226 WO2006045503A1 (en) | 2004-10-19 | 2005-10-19 | Method for solid phase peptide synthesis |
Related Child Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN2011101120914A Division CN102225966B (zh) | 2004-10-19 | 2005-10-19 | 用于固相肽合成的方法 |
Publications (2)
| Publication Number | Publication Date |
|---|---|
| CN101094867A CN101094867A (zh) | 2007-12-26 |
| CN101094867B true CN101094867B (zh) | 2011-08-24 |
Family
ID=35427609
Family Applications (2)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN2011101120914A Expired - Fee Related CN102225966B (zh) | 2004-10-19 | 2005-10-19 | 用于固相肽合成的方法 |
| CN2005800435731A Expired - Fee Related CN101094867B (zh) | 2004-10-19 | 2005-10-19 | 用于固相肽合成的方法 |
Family Applications Before (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN2011101120914A Expired - Fee Related CN102225966B (zh) | 2004-10-19 | 2005-10-19 | 用于固相肽合成的方法 |
Country Status (11)
| Country | Link |
|---|---|
| US (2) | USRE46830E1 (zh) |
| EP (1) | EP1737889B1 (zh) |
| JP (1) | JP4903709B2 (zh) |
| CN (2) | CN102225966B (zh) |
| AT (1) | ATE480561T1 (zh) |
| DE (1) | DE602005023429D1 (zh) |
| DK (1) | DK1737889T3 (zh) |
| ES (1) | ES2352204T3 (zh) |
| IL (1) | IL182698A (zh) |
| PT (1) | PT1737889E (zh) |
| WO (1) | WO2006045503A1 (zh) |
Families Citing this family (17)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CA2618494A1 (en) * | 2005-09-14 | 2007-03-22 | Novetide, Ltd. | Process for production of bivalirudin |
| WO2008109079A2 (en) * | 2007-03-01 | 2008-09-12 | Novetide, Ltd. | High purity peptides |
| US7598343B1 (en) | 2008-07-27 | 2009-10-06 | The Medicines Company | Pharmaceutical formulations of bivalirudin and processes of making the same |
| US7582727B1 (en) | 2008-07-27 | 2009-09-01 | The Medicinces Company | Pharmaceutical formulations of bivalirudin and processes of making the same |
| JP2012502045A (ja) | 2008-09-03 | 2012-01-26 | サイノファーム タイワン リミテッド | プラムリンチドの製造方法 |
| WO2010075983A1 (en) * | 2008-12-29 | 2010-07-08 | Lonza Braine Sa | Process for the production of bivalirudin |
| CN101475631B (zh) * | 2009-01-08 | 2011-08-17 | 苏州中科天马肽工程中心有限公司 | 比伐卢定的液相合成方法 |
| WO2010117725A2 (en) | 2009-04-06 | 2010-10-14 | Novetide, Ltd. | Production of peptides containing poly-gly sequences using fmoc chemistry |
| CN101555274B (zh) * | 2009-05-15 | 2013-08-21 | 海南双成药业股份有限公司 | 一种多肽固相合成比法卢定粗品的制备方法 |
| WO2013042129A1 (en) | 2011-09-23 | 2013-03-28 | Natco Pharma Limited | Improved process for preparation of bivalirudin |
| US9388212B2 (en) | 2013-02-21 | 2016-07-12 | Chemical & Biopharmaceutical Laboratories Of Patras S.A. | Solid phase peptide synthesis via side chain attachment |
| JP6382488B2 (ja) * | 2013-02-21 | 2018-08-29 | ケミカル アンド バイオファーマシューティカル ラボラトリーズ オブ パトラ エス．エー． | 側鎖結合を介する固相ペプチド合成 |
| GB201310921D0 (en) | 2013-06-19 | 2013-07-31 | Chemical & Biopharmaceutical Lab Of Patras S A | Peptide-resin conjugate and use thereof |
| WO2016047794A1 (ja) * | 2014-09-26 | 2016-03-31 | 株式会社カネカ | 疎水性ペプチドの製造法 |
| CN108383905A (zh) * | 2016-12-30 | 2018-08-10 | 江苏金斯瑞生物科技有限公司 | 一种比伐卢定的制备方法 |
| CN112111001B (zh) * | 2019-06-19 | 2021-10-29 | 翰宇药业（武汉）有限公司 | 胸腺肽Tα-1的合成方法 |
| JP2020138975A (ja) * | 2020-05-18 | 2020-09-03 | ケミカル アンド バイオファーマシューティカル ラボラトリーズ オブ パトラ エス．エー． | 側鎖結合を介する固相ペプチド合成 |
Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1088836A (zh) * | 1992-12-30 | 1994-07-06 | 中国科学院生物物理研究所 | 重组水蛭素及其复合物用于制备预防和治疗血栓病药物 |
| CN1036934C (zh) * | 1991-12-07 | 1998-01-07 | 赫彻斯特股份公司 | 具有改善稳定性的异水蛭素的制备方法 |
Family Cites Families (152)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| US4093610A (en) | 1977-01-31 | 1978-06-06 | American Home Products Corporation | Process for producing triglycyl-lysine vasopressin and intermediates therefor |
| US4108846A (en) | 1977-02-01 | 1978-08-22 | Hoffmann-La Roche Inc. | Solid phase synthesis with base N alpha-protecting group cleavage |
| US4169141A (en) | 1978-01-30 | 1979-09-25 | Shering Corporation | 1-Peptidyl derivatives of di-O-aminoglycosyl-1,3-diaminocyclitol antibacterial agents |
| US5789540A (en) | 1987-01-23 | 1998-08-04 | Merrell Pharmaceuticals Inc. | Anticoagulant peptides |
| US6005071A (en) | 1987-01-23 | 1999-12-21 | Merrell Pharmaceuticals Inc. | Anticoagulant peptides |
| US6239101B1 (en) | 1989-07-05 | 2001-05-29 | Oklahoma Medical Research Foundation | Thrombin binding polypeptides |
| US5240913A (en) | 1989-08-18 | 1993-08-31 | Biogen, Inc. | Inhibitors of thrombin |
| US5196404B1 (en) * | 1989-08-18 | 1996-09-10 | Biogen Inc | Inhibitors of thrombin |
| US5663141A (en) | 1989-12-01 | 1997-09-02 | Basf Aktiengesellschaft | Hirudin/polyalkylene glycol conjugates and hirudin muteins |
| US5624822A (en) | 1989-12-22 | 1997-04-29 | Basf Aktiengesellschaft | Hirudin fusion proteins and preparation of hirudin |
| AU641215B2 (en) | 1990-02-13 | 1993-09-16 | Merrell Dow Pharmaceuticals Inc. | Stabilized sulfonate, sulfate, phosphonate and phosphate derivatives of hirudin |
| DE4016596A1 (de) | 1990-05-23 | 1991-11-28 | Hoechst Ag | Ein neues kupplungsreagenz fuer die peptidsynthese |
| US6060451A (en) | 1990-06-15 | 2000-05-09 | The National Research Council Of Canada | Thrombin inhibitors based on the amino acid sequence of hirudin |
| US5574012A (en) | 1990-07-24 | 1996-11-12 | Merrell Pharmaceuticals Inc. | Analogs of hirudin having anti-platelet activity |
| ES2129749T3 (es) | 1990-11-08 | 1999-06-16 | Japan Energy Corp | Vector de secrecion, microorganismos transformados que contienen el indicado vector y produccion de un producto a partir de dicho microorganismo. |
| US5242810A (en) | 1990-12-07 | 1993-09-07 | Biogen, Inc. | Bifunctional inhibitors of thrombin and platelet activation |
| AU656947B2 (en) | 1991-02-04 | 1995-02-23 | Akzo N.V. | Factor IIa inhibitors |
| DE4103649A1 (de) | 1991-02-07 | 1992-08-13 | Basf Ag | Neue antikoagulatorisch wirksame peptide |
| CA2101942C (en) * | 1991-02-08 | 2001-01-30 | Richard T. Dean | Technetium-99m labeled polypeptides for imaging |
| US5767235A (en) | 1991-03-05 | 1998-06-16 | Nippon Mining Company Limited | Anticoagulant hirudin variants and methods for their production |
| US6514730B1 (en) | 1991-03-21 | 2003-02-04 | Consortium für elektrochemische Industrie GmbH | Secretion of hirudin derivatives |
| GB9112825D0 (en) | 1991-06-14 | 1991-07-31 | Ici Plc | Process for making peptides |
| US5837808A (en) | 1991-08-20 | 1998-11-17 | Baxter International Inc. | Analogs of hirudin |
| GB9118669D0 (en) | 1991-08-30 | 1991-10-16 | Univ Southampton | Preparation of peptides by a soliphase synthesis and intermediates therefor |
| DE4209110A1 (de) | 1991-11-26 | 1993-05-27 | Basf Ag | Neue thrombininhibitorische proteine aus landblutegeln |
| US5371184A (en) | 1992-02-05 | 1994-12-06 | Mallinckrodt Medical, Inc. | Radiolabelled peptide compounds |
| FR2687681B1 (fr) | 1992-02-20 | 1995-10-13 | Transgene Sa | Conjugues polyethyleneglycol-hirudine, leur procede de preparation et leur emploi pour le traitement des thromboses. |
| GB9209032D0 (en) | 1992-04-25 | 1992-06-10 | Ciba Geigy Ag | New peptide derivatives |
| US5968476A (en) | 1992-05-21 | 1999-10-19 | Diatide, Inc. | Technetium-99m labeled peptides for thrombus imaging |
| WO1994017817A1 (en) | 1993-02-12 | 1994-08-18 | Corvas International, Inc. | Inhibitors of thrombosis |
| US5656600A (en) | 1993-03-25 | 1997-08-12 | Corvas International, Inc. | α-ketoamide derivatives as inhibitors of thrombosis |
| DE4310632A1 (de) | 1993-04-01 | 1994-10-06 | Merck Patent Gmbh | Lineare Adhäsionsinhibitoren |
| IT1265023B1 (it) | 1993-04-16 | 1996-10-28 | Dev Biotechnological Processes | Inibitori della trombina, loro preparazione ed uso per applicazioni terapeutiche, profilattiche e diagnostiche |
| US5443827A (en) | 1993-05-03 | 1995-08-22 | President And Fellows Of Harvard College | Fibrin-targeted inhibitors of thrombin |
| CN1124964A (zh) | 1993-06-11 | 1996-06-19 | 默里尔药物公司 | 三功能抗凝血酶和抗血小板的肽 |
| US5457177A (en) | 1993-06-16 | 1995-10-10 | Merck & Co., Inc. | Thrombin receptor radioligands |
| DE4323754C1 (de) | 1993-07-15 | 1994-12-01 | Gruenenthal Gmbh | Bifunktionelle Urokinasevarianten mit verbesserten fibrinolytischen Eigenschaften und thrombinhemmender Wirkung |
| JPH09500140A (ja) | 1993-07-19 | 1997-01-07 | レゾリューション・ファーマスーティカルズ・インコーポレーテッド | Ｎ▲下３▼ｓ立体配置を有するヒドラジノ型放射性核種キレート形成剤 |
| US5449761A (en) | 1993-09-28 | 1995-09-12 | Cytogen Corporation | Metal-binding targeted polypeptide constructs |
| US5972648A (en) | 1993-09-28 | 1999-10-26 | Japan Energy Corporation | Hirudin analogs, methods of manufacture thereof and anticoagulant compositions having these as active ingredients |
| FR2710917B1 (fr) | 1993-10-07 | 1995-11-24 | Adir | Nouveaux dérivés de peptides thérapeutiquement actifs dans la cascade de coagulation sanguine, leur procédé de préparation et les compositions pharmaceutiques qui les contiennent. |
| DE69426569T2 (de) | 1993-10-25 | 2001-05-10 | National Research Council Of Canada, Ottawa | Bivalente thrombininhibitore |
| US7060484B1 (en) | 1993-11-12 | 2006-06-13 | Gilead Sciences, Inc. | Polypeptides and coagulation therapy |
| US6156540A (en) | 1993-12-22 | 2000-12-05 | Human Genome Sciences, Inc. | Thrombin inhibitor |
| DE4404168A1 (de) | 1994-02-10 | 1995-08-17 | Hoechst Ag | Hirudinderivate und Verfahren zu deren Herstellung |
| US6335160B1 (en) | 1995-02-17 | 2002-01-01 | Maxygen, Inc. | Methods and compositions for polypeptide engineering |
| US5662885A (en) | 1994-07-22 | 1997-09-02 | Resolution Pharmaceuticals Inc. | Peptide derived radionuclide chelators |
| US5759542A (en) | 1994-08-05 | 1998-06-02 | New England Deaconess Hospital Corporation | Compositions and methods for the delivery of drugs by platelets for the treatment of cardiovascular and other diseases |
| KR0141651B1 (ko) | 1994-09-07 | 1998-06-15 | 강재헌 | 금속이온 친화성 크로마토그라피를 이용한 히루딘의 정제방법 |
| DE4440892A1 (de) | 1994-11-17 | 1996-05-23 | Gruenenthal Gmbh | Proteine mit fibrinolytischen und gerinnungshemmenden Eigenschaften |
| DE4442665A1 (de) | 1994-11-30 | 1996-06-05 | Gruenenthal Gmbh | Chimäre Proteine mit fibrinolytischen und thrombinhemmenden Eigenschaften |
| US5747453A (en) | 1995-06-06 | 1998-05-05 | Alza Corporation | Method for increasing the electrotransport flux of polypeptides |
| US5817758A (en) | 1995-06-07 | 1998-10-06 | Terrapin Technologies, Inc. | P-nitrobenzyl side-chain protection for solid-phase synthesis |
| US5767078A (en) | 1995-06-07 | 1998-06-16 | Johnson; Dana L. | Agonist peptide dimers |
| DE69627191T2 (de) | 1995-06-09 | 2004-01-08 | The Regents Of The University Of Michigan, Ann Arbor | Bradikininanaloge als selektive thrombininhibitoren |
| IL119466A (en) | 1995-11-03 | 2001-08-26 | Akzo Nobel Nv | Thrombin inhibitors, their preparation and pharmaceutical compositions containing them |
| US5910481A (en) | 1995-11-13 | 1999-06-08 | Immuno Ag | Hybrid proteins with modified activity |
| US6333189B1 (en) | 1996-06-06 | 2001-12-25 | Alza Corporation | Method of making an electrotransport device |
| US7176282B1 (en) | 1996-09-09 | 2007-02-13 | Zealand Pharma A/S | Solid-phase peptide synthesis and agent for use in such synthesis |
| CZ295838B6 (cs) | 1996-09-09 | 2005-11-16 | Zealand Pharma A/S | Způsob výroby peptidů |
| US6265204B1 (en) | 1997-01-17 | 2001-07-24 | Genencor International, Inc. | DNA sequences, vectors, and fusion polypeptides for secretion of polypeptides in filamentous fungi |
| US5986065A (en) | 1997-03-10 | 1999-11-16 | Sunol Molecular Corporation | Antibodies for inhibiting blood coagulation and methods of use thereof |
| US6077825A (en) | 1997-03-13 | 2000-06-20 | Auburn University | Antithrombin protein and DNA sequences from black fly |
| GB9708918D0 (en) | 1997-05-01 | 1997-06-25 | Ppl Therapeutics Scotland Ltd | Methods |
| CA2321026A1 (en) | 1998-03-09 | 1999-09-16 | Zealand Pharmaceuticals A/S | Pharmacologically active peptide conjugates having a reduced tendency towards enzymatic hydrolysis |
| US6281331B1 (en) | 1998-03-23 | 2001-08-28 | Trimeris, Inc. | Methods and compositions for peptide synthesis |
| US6451992B1 (en) | 1998-08-20 | 2002-09-17 | Auburn University | Antithrobin nucleotides and proteins from horn fly |
| WO2000061609A2 (de) | 1999-04-09 | 2000-10-19 | Basf Aktiengesellschaft | Prodrugs von thrombininhibitoren |
| ES2154590B1 (es) | 1999-05-20 | 2001-11-01 | Lipotec Sa | Procedimiento de sintesis de peptidos en fase solida |
| US6544750B1 (en) | 1999-08-17 | 2003-04-08 | Thromgen, Inc. | Peptide analogs as selective inhibitors of thrombin activation of protease activated receptor 1 |
| EP1138694A1 (en) | 2000-03-30 | 2001-10-04 | University Of Manitoba | Hirulog-like peptide |
| US6506761B1 (en) | 2000-04-14 | 2003-01-14 | Corvas International, Inc. | Substituted hydrazinyl heteroaromatic inhibitors of thrombin |
| JP2002112782A (ja) | 2000-10-04 | 2002-04-16 | Ajinomoto Co Inc | 抗血栓活性を有する蛋白質及びその製造法 |
| DE10102878A1 (de) | 2001-01-23 | 2002-08-01 | Haemosys Gmbh | Oligo- oder Polyalkylengekoppelte Thrombininhibitoren |
| WO2002079278A1 (en) | 2001-03-30 | 2002-10-10 | Mayo Foundation For Medical Education And Research | Efficient methods for solid phase synthesis using trityl chloride resins |
| WO2002100337A2 (en) | 2001-06-08 | 2002-12-19 | Emory University | Antithrombotic thrombin variants |
| US6703364B2 (en) | 2001-07-23 | 2004-03-09 | Cleveland State University | Thrombin generation inhibitors |
| WO2003022873A1 (de) | 2001-09-10 | 2003-03-20 | Novel Science International Gmbh | Organische verbindungen mit biologischer wirkung als thrombinhemmer und ihre verwendung |
| GB0123262D0 (en) | 2001-09-27 | 2001-11-21 | Adprotech Ltd | Polymeric compounds |
| KR100718431B1 (ko) * | 2001-11-27 | 2007-05-14 | 주식회사 유앤비케미칼 | 트리틸기가 도입된 폴리스티렌 수지의 제조 방법 |
| US7794693B2 (en) | 2002-03-01 | 2010-09-14 | Bracco International B.V. | Targeting vector-phospholipid conjugates |
| WO2003084983A1 (fr) | 2002-04-11 | 2003-10-16 | Daiichi Suntory Pharma Co., Ltd. | Procede de production d'un peptide modifie |
| US7691968B2 (en) | 2002-05-03 | 2010-04-06 | Avecia Biologics Limited | Process for the synthesis of peptides amides by side-chain attachment to a solid phase |
| US6875893B2 (en) | 2002-05-23 | 2005-04-05 | Cephalon, Inc. | Preparations of a sulfinyl acetamide |
| EP1546186A4 (en) | 2002-08-26 | 2006-02-08 | A & Pep Inc | PROCESS FOR SYNTHESIS OF PEPTIDES |
| ATE328909T1 (de) | 2002-12-20 | 2006-06-15 | Hoffmann La Roche | Verfahren zur wiedergewinnung von festphasen harzgebundenem chlortritylchlorid |
| DE10301255A1 (de) | 2003-01-15 | 2004-07-29 | Cpi Creative Pharma International Gmbh | Organische Verbindungen mit biologischer Wirkung als Thrombinhemmer und ihre Verwendung |
| WO2004076484A1 (en) | 2003-02-27 | 2004-09-10 | National Research Council Of Canada | Peptide inhibitors of thrombin as potent anticoagulants |
| US7074765B2 (en) | 2003-05-01 | 2006-07-11 | The Regents Of The University Of Michigan | Synthetic peptide analogs of Arg-Pro-Pro-Gly-Phe as selective inhibitors of thrombin and thrombin activation of protease activated receptors 1 and 4 |
| US7393920B2 (en) | 2003-06-23 | 2008-07-01 | Cem Corporation | Microwave-assisted peptide synthesis |
| WO2004113386A2 (en) | 2003-06-26 | 2004-12-29 | Merck Patent Gmbh | Modified hirudin proteins and t-cell epitopes in hirudin |
| US20070055048A1 (en) | 2003-07-04 | 2007-03-08 | Vincent Cool | Process for supported phase synthesis |
| US7135534B2 (en) | 2003-07-24 | 2006-11-14 | Rajiv Gandhi Centre For Biotechnology | Polymer support for solid phase peptide synthesis and process for preparation thereof |
| GB0318205D0 (en) | 2003-08-02 | 2003-09-03 | Albachem Ltd | Synthetic method |
| ES2295961T3 (es) | 2003-12-31 | 2008-04-16 | F. Hoffmann-La Roche Ag | Proceso para la sintesis peptidica utilizacndo una cantidad reducida de agente de desproteccion. |
| US20080025966A1 (en) | 2004-01-30 | 2008-01-31 | Currie Mark G | Methods And Compositions For The Treatment Of Gastrointestinal disorders |
| US7795205B2 (en) | 2004-04-12 | 2010-09-14 | Canyon Pharmaceuticals, Inc. | Methods for effecting regression of tumor mass and size in a metastasized pancreatic tumor |
| EP1600455A1 (en) | 2004-05-28 | 2005-11-30 | Deutsches Krebsforschungszentrum Stiftung des öffentlichen Rechts | Preparation of solid phase bound peptides or PNAs |
| EP1766011B1 (en) | 2004-06-23 | 2011-05-18 | National Research Council Of Canada | Methods and uses of polypeptide ligands containing linkers |
| US20060172319A1 (en) | 2004-07-12 | 2006-08-03 | Applera Corporation | Mass tags for quantitative analyses |
| US20090054319A1 (en) | 2005-03-17 | 2009-02-26 | Microbia, Inc. | Methods and Compositions for the Treatment of Hypertension and Gastrointestinal Disorders |
| ATE452145T1 (de) | 2005-05-03 | 2010-01-15 | Novetide Ltd | Verfahren zur herstellung von peptidderivaten |
| US7879792B2 (en) | 2005-06-02 | 2011-02-01 | The Regents Of The University Of Michigan | Synthetic peptide inhibitors of thrombin and thrombin activation of protease activated receptors 1 and 4 |
| CA2618494A1 (en) | 2005-09-14 | 2007-03-22 | Novetide, Ltd. | Process for production of bivalirudin |
| ES2428964T3 (es) | 2005-12-09 | 2013-11-12 | Bracco Suisse Sa | Conjugados de vectores específicos de una diana-fosfolípidos |
| US8314208B2 (en) | 2006-02-10 | 2012-11-20 | Cem Corporation | Microwave enhanced N-fmoc deprotection in peptide synthesis |
| CN101033249B (zh) | 2006-03-10 | 2011-05-11 | 周逸明 | 固相多肽合成比筏芦定的制备方法 |
| US20080096819A1 (en) | 2006-05-02 | 2008-04-24 | Allozyne, Inc. | Amino acid substituted molecules |
| WO2007130275A2 (en) | 2006-05-03 | 2007-11-15 | Mallinckrodt Inc. | Composition and method for the release of protected peptides from a resin |
| EP2049561A1 (en) | 2006-07-21 | 2009-04-22 | Solvay (Societe Anonyme) | Process for the manufacture of peptides |
| US20100075375A1 (en) | 2006-10-03 | 2010-03-25 | Novo Nordisk A/S | Methods for the purification of polypeptide conjugates |
| US20100168443A1 (en) | 2006-11-02 | 2010-07-01 | University Of Virginia Patent Foundation | Ethoid-Containing Compounds, Methods for Preparing Ethoid-Containing Compounds, and Methods of Use |
| IE20060841A1 (en) | 2006-11-21 | 2008-05-28 | Ipsen Mfg Ireland Ltd | Boc and fmoc solid phase peptide synthesis |
| CA2913963A1 (en) | 2006-12-08 | 2008-06-19 | Millenium Pharmaceuticals, Inc. | Unit dose formulations and methods of treating thrombosis with an oral factor xa inhibitor |
| CN101668778B (zh) | 2006-12-15 | 2012-10-03 | 中国人民解放军军事医学科学院放射与辐射医学研究所 | 低出血抗凝血融合蛋白的制备及其应用 |
| CN101245110B (zh) | 2007-02-16 | 2010-09-15 | 鲁南制药集团股份有限公司 | 重组中性粒细胞抑制因子和水蛭原嵌合蛋白及其药物组合物 |
| WO2008109079A2 (en) | 2007-03-01 | 2008-09-12 | Novetide, Ltd. | High purity peptides |
| GB0711779D0 (en) | 2007-06-18 | 2007-07-25 | Univ Singapore | Thrombin inhibitor |
| US8206967B2 (en) | 2007-07-06 | 2012-06-26 | Medimmune Limited | Method for production of recombinant human thrombin |
| WO2009014177A1 (ja) | 2007-07-25 | 2009-01-29 | Ajinomoto Co., Inc. | ジベンゾフルベン誘導体の淘汰方法 |
| CN101372512B (zh) | 2007-08-23 | 2011-03-23 | 中国人民解放军军事医学科学院生物工程研究所 | 一类抗凝血多肽及其用途 |
| US20090062511A1 (en) | 2007-09-05 | 2009-03-05 | Raghavendracharyulu Venkata Palle | Process for the preparation of bivalirudin and its pharmaceutical compositions |
| JP2011520844A (ja) | 2008-05-15 | 2011-07-21 | ノボ・ノルデイスク・エー／エス | 固相合成法により調製されたペプチドの精製 |
| US7582727B1 (en) | 2008-07-27 | 2009-09-01 | The Medicinces Company | Pharmaceutical formulations of bivalirudin and processes of making the same |
| US7598343B1 (en) | 2008-07-27 | 2009-10-06 | The Medicines Company | Pharmaceutical formulations of bivalirudin and processes of making the same |
| WO2010016551A1 (ja) | 2008-08-06 | 2010-02-11 | 味の素株式会社 | ジベンゾフルベンの除去方法 |
| JP2012502045A (ja) | 2008-09-03 | 2012-01-26 | サイノファーム タイワン リミテッド | プラムリンチドの製造方法 |
| US20110288235A1 (en) | 2008-09-03 | 2011-11-24 | Scinopharm Taiwan Ltd. | Process for the Preparation of Pramlintide |
| WO2010054503A1 (zh) | 2008-11-17 | 2010-05-20 | 中国人民解放军军事医学科学院生物工程研究所 | 抗凝血多肽及其应用 |
| WO2010075983A1 (en) | 2008-12-29 | 2010-07-08 | Lonza Braine Sa | Process for the production of bivalirudin |
| CN101475631B (zh) | 2009-01-08 | 2011-08-17 | 苏州中科天马肽工程中心有限公司 | 比伐卢定的液相合成方法 |
| WO2010117725A2 (en) | 2009-04-06 | 2010-10-14 | Novetide, Ltd. | Production of peptides containing poly-gly sequences using fmoc chemistry |
| GB0907698D0 (en) | 2009-05-05 | 2009-06-10 | Univ Singapore | Method of modifying serine protease inhibitors |
| CN101555274B (zh) | 2009-05-15 | 2013-08-21 | 海南双成药业股份有限公司 | 一种多肽固相合成比法卢定粗品的制备方法 |
| US7803762B1 (en) | 2009-08-20 | 2010-09-28 | The Medicines Company | Ready-to-use bivalirudin compositions |
| GB0915519D0 (en) | 2009-09-04 | 2009-10-07 | King S College London | Ntithrombotic compounds |
| WO2011071799A2 (en) | 2009-12-11 | 2011-06-16 | Dr. Reddy's Laboratories Ltd. | Purification of bivalirudin |
| US7985733B1 (en) | 2010-01-06 | 2011-07-26 | The Medicines Company | Buffer-based method for preparing bivalirudin drug product |
| CN101906150B (zh) | 2010-06-28 | 2013-01-09 | 上海昂博生物技术有限公司 | 一种比法卢定的制备方法 |
| CA2808270A1 (en) | 2010-08-16 | 2012-02-23 | Cem Corporation | Water soluble solid phase peptide synthesis |
| KR20120069288A (ko) | 2010-12-20 | 2012-06-28 | 주식회사 씨트리 | 수용성 당 알코올을 이용한 비바리루딘의 정제방법 |
| CN102260323A (zh) | 2011-05-30 | 2011-11-30 | 杭州诺泰制药技术有限公司 | 固相液相结合制备比伐卢定的方法和检测方法 |
| WO2012165546A1 (ja) | 2011-05-31 | 2012-12-06 | 味の素株式会社 | ペプチドの製造方法 |
| CN102286076B (zh) | 2011-06-23 | 2014-03-12 | 成都圣诺科技发展有限公司 | 比伐卢定的制备方法 |
| CN102336813B (zh) | 2011-07-01 | 2016-01-06 | 上海苏豪逸明制药有限公司 | 一种固相多肽合成蛋白溶菌素的制备方法 |
| US8906681B2 (en) | 2011-08-02 | 2014-12-09 | The Scripps Research Institute | Reliable stabilization of N-linked polypeptide native states with enhanced aromatic sequons located in polypeptide tight turns |
| WO2013042129A1 (en) | 2011-09-23 | 2013-03-28 | Natco Pharma Limited | Improved process for preparation of bivalirudin |
| CN102532274B (zh) | 2012-02-13 | 2014-04-23 | 成都圣诺生物制药有限公司 | 一种比伐卢定的制备方法 |
| CN102641506B (zh) | 2012-04-11 | 2013-10-23 | 承德医学院中药研究所 | 比伐卢定-聚乙二醇化复合物 |
| CN102731624B (zh) | 2012-06-14 | 2015-09-23 | 无锡市凯利药业有限公司 | 一种固相片段法合成比伐卢定的方法 |
| CN102702325B (zh) | 2012-06-19 | 2015-09-23 | 深圳翰宇药业股份有限公司 | 一种抗凝血多肽的制备方法 |
| CN102924575B (zh) | 2012-10-31 | 2014-12-31 | 深圳翰宇药业股份有限公司 | 一种比伐卢定的制备方法 |
-
2005
- 2005-10-19 AT AT05796944T patent/ATE480561T1/de not_active IP Right Cessation
- 2005-10-19 CN CN2011101120914A patent/CN102225966B/zh not_active Expired - Fee Related
- 2005-10-19 ES ES05796944T patent/ES2352204T3/es active Active
- 2005-10-19 US US14/659,770 patent/USRE46830E1/en active Active
- 2005-10-19 DE DE602005023429T patent/DE602005023429D1/de active Active
- 2005-10-19 JP JP2007537195A patent/JP4903709B2/ja not_active Expired - Fee Related
- 2005-10-19 EP EP05796944A patent/EP1737889B1/en active Active
- 2005-10-19 DK DK05796944.6T patent/DK1737889T3/da active
- 2005-10-19 PT PT05796944T patent/PT1737889E/pt unknown
- 2005-10-19 US US11/665,541 patent/US7939629B2/en not_active Ceased
- 2005-10-19 WO PCT/EP2005/011226 patent/WO2006045503A1/en active Application Filing
- 2005-10-19 CN CN2005800435731A patent/CN101094867B/zh not_active Expired - Fee Related
-
2007
- 2007-04-19 IL IL182698A patent/IL182698A/en active IP Right Grant
Patent Citations (2)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1036934C (zh) * | 1991-12-07 | 1998-01-07 | 赫彻斯特股份公司 | 具有改善稳定性的异水蛭素的制备方法 |
| CN1088836A (zh) * | 1992-12-30 | 1994-07-06 | 中国科学院生物物理研究所 | 重组水蛭素及其复合物用于制备预防和治疗血栓病药物 |
Non-Patent Citations (1)
| Title |
|---|
| Ernst Freund和John A. Robinson.Solid-phase synthesis of a putativeheptapeptide intermediatein vancomycin biosynthesis.Chem.Commun..1999,2509-2510. * |
Also Published As
| Publication number | Publication date |
|---|---|
| CN102225966A (zh) | 2011-10-26 |
| US20080287648A1 (en) | 2008-11-20 |
| IL182698A0 (en) | 2007-09-20 |
| DE602005023429D1 (de) | 2010-10-21 |
| ATE480561T1 (de) | 2010-09-15 |
| JP4903709B2 (ja) | 2012-03-28 |
| WO2006045503A1 (en) | 2006-05-04 |
| DK1737889T3 (da) | 2011-01-03 |
| IL182698A (en) | 2013-11-28 |
| CN101094867A (zh) | 2007-12-26 |
| EP1737889B1 (en) | 2010-09-08 |
| US7939629B2 (en) | 2011-05-10 |
| ES2352204T3 (es) | 2011-02-16 |
| USRE46830E1 (en) | 2018-05-08 |
| JP2008517018A (ja) | 2008-05-22 |
| EP1737889A1 (en) | 2007-01-03 |
| CN102225966B (zh) | 2012-12-26 |
| PT1737889E (pt) | 2010-12-13 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN101094867B (zh) | 用于固相肽合成的方法 | |
| DK175042B1 (da) | Kunstharpiks, fremgangsmåde til fremstilling deraf samt fremgangsmåde til fremstilling af peptider og peptidamider under anvendelse af kunstharpiksen | |
| FI70234C (fi) | Foerfarande foer framstaellning av amino-funktionaliserade akrylkopolymerer | |
| TWI453216B (zh) | 比伐盧定（Ｂｉｖａｌｉｒｕｄｉｎ）的製造方法 | |
| US20110160431A1 (en) | Production of peptides containing poly-gly sequences using fmoc chemistry | |
| Brown et al. | Further studies on the protection of histidine side chains in peptide synthesis: the use of the π-benzyloxymethyl group | |
| KR20010071853A (ko) | 비스-트리클로로메틸 카르보네이트를 이용하여 아미노산을결합하는 공정 | |
| CN110204611A (zh) | 一种固相片段法合成比伐卢定 | |
| US4831084A (en) | Resin-linker combination for the solid-phase synthesis of peptides and intermediates | |
| CA2458084A1 (en) | Novel process for octreotide synthesis | |
| CN107778351A (zh) | 一种全固相合成奥曲肽的方法 | |
| Thurieau et al. | New N. alpha.-Guanidinobenzoyl Derivatives of Hirudin-54-65 Containing Stabilized Carboxyl or Phosphoryl Groups on the Side Chain of Phenylalanine-63 | |
| CN115181158A (zh) | Fmoc基团裂解方法 | |
| CN100357313C (zh) | 非靶位点胺基被保护的肽,其制备方法以及使用该肽制备特异性地结合的peg肽的方法 | |
| Pawlak et al. | Synthesis of a novel side‐chain to side‐chain cyclized enkephalin analogue containing a carbonyl bridge | |
| CN109912709A (zh) | 一种酸敏感离子通道抑制剂的制备方法 | |
| CN112679603B (zh) | 固液相结合制备特立帕肽的方法 | |
| AU729393B2 (en) | Serine protease inhibitors | |
| JPH1067796A (ja) | 環状ペプチドの合成法 | |
| Bell et al. | SEQUENTIAL POLYPEPTIDES. Part X. The synthesis of some sequential polypeptide collagen models with functional side‐chains | |
| KR20220008674A (ko) | 링커-레진 화합물, 그 제조방법, 및 그 화합물을 이용한 펩타이드 제조방법 | |
| FEHRENTZ et al. | Solution synthesis of antigenic and inhibiting peptides of the human renin prosegment‐1: [Tyr 47, Nle 53] preprorenin‐(47–60) peptide methyl ester | |
| CN118754966A (zh) | 一种瑞他鲁肽的制备方法 | |
| CN116925188A (zh) | 一种预防或治疗冠状病毒的多肽及其衍生物的合成方法 | |
| CN112390856A (zh) | 一种具有生物活性的多肽eep2和二倍体的固相合成方法 |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| REG | Reference to a national code |
Ref country code: HK Ref legal event code: DE Ref document number: 1113577 Country of ref document: HK |
|
| C14 | Grant of patent or utility model | ||
| GR01 | Patent grant | ||
| REG | Reference to a national code |
Ref country code: HK Ref legal event code: WD Ref document number: 1113577 Country of ref document: HK |
|
| TR01 | Transfer of patent right |
Effective date of registration: 20170527 Address after: Swedish limhamn Patentee after: Polypeptide Laboratory (PPL) holding Co. Address before: Basel, Switzerland Patentee before: LONZA AG |
|
| CF01 | Termination of patent right due to non-payment of annual fee |
Granted publication date: 20110824 |