CN1886424A - 抗igf-i受体抗体 - Google Patents
抗igf-i受体抗体 Download PDFInfo
- Publication number
- CN1886424A CN1886424A CNA200480034889XA CN200480034889A CN1886424A CN 1886424 A CN1886424 A CN 1886424A CN A200480034889X A CNA200480034889X A CN A200480034889XA CN 200480034889 A CN200480034889 A CN 200480034889A CN 1886424 A CN1886424 A CN 1886424A
- Authority
- CN
- China
- Prior art keywords
- antibody
- ser
- gly
- leu
- val
- 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.)
- Pending
Links
- 102000005962 receptors Human genes 0.000 title claims description 50
- 108020003175 receptors Proteins 0.000 title claims description 50
- 206010028980 Neoplasm Diseases 0.000 claims abstract description 69
- 230000012010 growth Effects 0.000 claims abstract description 49
- 239000012634 fragment Substances 0.000 claims abstract description 48
- 108010021625 Immunoglobulin Fragments Proteins 0.000 claims abstract description 38
- 102000008394 Immunoglobulin Fragments Human genes 0.000 claims abstract description 38
- 238000011282 treatment Methods 0.000 claims abstract description 37
- 206010006187 Breast cancer Diseases 0.000 claims abstract description 19
- 208000026310 Breast neoplasm Diseases 0.000 claims abstract description 19
- 206010009944 Colon cancer Diseases 0.000 claims abstract description 13
- 206010058467 Lung neoplasm malignant Diseases 0.000 claims abstract description 13
- 201000005202 lung cancer Diseases 0.000 claims abstract description 11
- 208000020816 lung neoplasm Diseases 0.000 claims abstract description 11
- 108090000723 Insulin-Like Growth Factor I Proteins 0.000 claims abstract description 8
- 206010033128 Ovarian cancer Diseases 0.000 claims abstract description 8
- 206010060862 Prostate cancer Diseases 0.000 claims abstract description 8
- 208000000236 Prostatic Neoplasms Diseases 0.000 claims abstract description 8
- 102000004218 Insulin-Like Growth Factor I Human genes 0.000 claims abstract description 7
- 206010042863 synovial sarcoma Diseases 0.000 claims abstract description 6
- 210000004027 cell Anatomy 0.000 claims description 157
- 239000003795 chemical substances by application Substances 0.000 claims description 85
- 230000001225 therapeutic effect Effects 0.000 claims description 74
- 238000000034 method Methods 0.000 claims description 57
- 201000011510 cancer Diseases 0.000 claims description 51
- 125000003275 alpha amino acid group Chemical group 0.000 claims description 49
- 108010047041 Complementarity Determining Regions Proteins 0.000 claims description 39
- 230000027455 binding Effects 0.000 claims description 30
- 229930012538 Paclitaxel Natural products 0.000 claims description 28
- BASFCYQUMIYNBI-UHFFFAOYSA-N platinum Substances [Pt] BASFCYQUMIYNBI-UHFFFAOYSA-N 0.000 claims description 28
- 210000004408 hybridoma Anatomy 0.000 claims description 27
- 239000000203 mixture Substances 0.000 claims description 27
- 229960001592 paclitaxel Drugs 0.000 claims description 27
- RCINICONZNJXQF-MZXODVADSA-N taxol Chemical compound O([C@@H]1[C@@]2(C[C@@H](C(C)=C(C2(C)C)[C@H](C([C@]2(C)[C@@H](O)C[C@H]3OC[C@]3([C@H]21)OC(C)=O)=O)OC(=O)C)OC(=O)[C@H](O)[C@@H](NC(=O)C=1C=CC=CC=1)C=1C=CC=CC=1)O)C(=O)C1=CC=CC=C1 RCINICONZNJXQF-MZXODVADSA-N 0.000 claims description 26
- 238000012360 testing method Methods 0.000 claims description 24
- 241001529936 Murinae Species 0.000 claims description 22
- 229910052697 platinum Inorganic materials 0.000 claims description 22
- SDUQYLNIPVEERB-QPPQHZFASA-N gemcitabine Chemical compound O=C1N=C(N)C=CN1[C@H]1C(F)(F)[C@H](O)[C@@H](CO)O1 SDUQYLNIPVEERB-QPPQHZFASA-N 0.000 claims description 21
- 229960005277 gemcitabine Drugs 0.000 claims description 21
- NKANXQFJJICGDU-QPLCGJKRSA-N Tamoxifen Chemical compound C=1C=CC=CC=1C(/CC)=C(C=1C=CC(OCCN(C)C)=CC=1)/C1=CC=CC=C1 NKANXQFJJICGDU-QPLCGJKRSA-N 0.000 claims description 16
- ZDZOTLJHXYCWBA-VCVYQWHSSA-N N-debenzoyl-N-(tert-butoxycarbonyl)-10-deacetyltaxol Chemical compound O([C@H]1[C@H]2[C@@](C([C@H](O)C3=C(C)[C@@H](OC(=O)[C@H](O)[C@@H](NC(=O)OC(C)(C)C)C=4C=CC=CC=4)C[C@]1(O)C3(C)C)=O)(C)[C@@H](O)C[C@H]1OC[C@]12OC(=O)C)C(=O)C1=CC=CC=C1 ZDZOTLJHXYCWBA-VCVYQWHSSA-N 0.000 claims description 15
- GXJABQQUPOEUTA-RDJZCZTQSA-N bortezomib Chemical compound C([C@@H](C(=O)N[C@@H](CC(C)C)B(O)O)NC(=O)C=1N=CC=NC=1)C1=CC=CC=C1 GXJABQQUPOEUTA-RDJZCZTQSA-N 0.000 claims description 15
- 229960003668 docetaxel Drugs 0.000 claims description 15
- 229960001756 oxaliplatin Drugs 0.000 claims description 15
- DWAFYCQODLXJNR-BNTLRKBRSA-L oxaliplatin Chemical compound O1C(=O)C(=O)O[Pt]11N[C@@H]2CCCC[C@H]2N1 DWAFYCQODLXJNR-BNTLRKBRSA-L 0.000 claims description 15
- 229960005267 tositumomab Drugs 0.000 claims description 15
- GHASVSINZRGABV-UHFFFAOYSA-N Fluorouracil Chemical compound FC1=CNC(=O)NC1=O GHASVSINZRGABV-UHFFFAOYSA-N 0.000 claims description 14
- VSRXQHXAPYXROS-UHFFFAOYSA-N azanide;cyclobutane-1,1-dicarboxylic acid;platinum(2+) Chemical compound [NH2-].[NH2-].[Pt+2].OC(=O)C1(C(O)=O)CCC1 VSRXQHXAPYXROS-UHFFFAOYSA-N 0.000 claims description 14
- 229960004562 carboplatin Drugs 0.000 claims description 14
- 229940099039 velcade Drugs 0.000 claims description 14
- 230000010261 cell growth Effects 0.000 claims description 11
- 208000001333 Colorectal Neoplasms Diseases 0.000 claims description 10
- -1 Rinotecan Chemical compound 0.000 claims description 10
- 201000010989 colorectal carcinoma Diseases 0.000 claims description 10
- VSJKWCGYPAHWDS-FQEVSTJZSA-N camptothecin Chemical compound C1=CC=C2C=C(CN3C4=CC5=C(C3=O)COC(=O)[C@]5(O)CC)C4=NC2=C1 VSJKWCGYPAHWDS-FQEVSTJZSA-N 0.000 claims description 9
- XGALLCVXEZPNRQ-UHFFFAOYSA-N gefitinib Chemical compound C=12C=C(OCCCN3CCOCC3)C(OC)=CC2=NC=NC=1NC1=CC=C(F)C(Cl)=C1 XGALLCVXEZPNRQ-UHFFFAOYSA-N 0.000 claims description 9
- 108010083551 iturelix Proteins 0.000 claims description 9
- QRYFGTULTGLGHU-NBERXCRTSA-N iturelix Chemical compound C([C@H](C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCCCNC(C)C)C(=O)N1[C@@H](CCC1)C(=O)N[C@H](C)C(N)=O)NC(=O)[C@H](CCCCNC(=O)C=1C=NC=CC=1)NC(=O)[C@H](CO)NC(=O)[C@@H](CC=1C=NC=CC=1)NC(=O)[C@@H](CC=1C=CC(Cl)=CC=1)NC(=O)[C@@H](CC=1C=C2C=CC=CC2=CC=1)NC(C)=O)CCCNC(=O)C1=CC=CN=C1 QRYFGTULTGLGHU-NBERXCRTSA-N 0.000 claims description 9
- 229960001924 melphalan Drugs 0.000 claims description 9
- SGDBTWWWUNNDEQ-LBPRGKRZSA-N melphalan Chemical compound OC(=O)[C@@H](N)CC1=CC=C(N(CCCl)CCCl)C=C1 SGDBTWWWUNNDEQ-LBPRGKRZSA-N 0.000 claims description 9
- STQGQHZAVUOBTE-UHFFFAOYSA-N 7-Cyan-hept-2t-en-4,6-diinsaeure Natural products C1=2C(O)=C3C(=O)C=4C(OC)=CC=CC=4C(=O)C3=C(O)C=2CC(O)(C(C)=O)CC1OC1CC(N)C(O)C(C)O1 STQGQHZAVUOBTE-UHFFFAOYSA-N 0.000 claims description 8
- VWUXBMIQPBEWFH-WCCTWKNTSA-N Fulvestrant Chemical compound OC1=CC=C2[C@H]3CC[C@](C)([C@H](CC4)O)[C@@H]4[C@@H]3[C@H](CCCCCCCCCS(=O)CCCC(F)(F)C(F)(F)F)CC2=C1 VWUXBMIQPBEWFH-WCCTWKNTSA-N 0.000 claims description 8
- BLCLNMBMMGCOAS-URPVMXJPSA-N Goserelin Chemical compound C([C@@H](C(=O)N[C@H](COC(C)(C)C)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCCN=C(N)N)C(=O)N1[C@@H](CCC1)C(=O)NNC(N)=O)NC(=O)[C@H](CO)NC(=O)[C@H](CC=1C2=CC=CC=C2NC=1)NC(=O)[C@H](CC=1NC=NC=1)NC(=O)[C@H]1NC(=O)CC1)C1=CC=C(O)C=C1 BLCLNMBMMGCOAS-URPVMXJPSA-N 0.000 claims description 8
- 108010069236 Goserelin Proteins 0.000 claims description 8
- 239000005517 L01XE01 - Imatinib Substances 0.000 claims description 8
- 108010023617 abarelix Proteins 0.000 claims description 8
- AIWRTTMUVOZGPW-HSPKUQOVSA-N abarelix Chemical compound C([C@@H](C(=O)N[C@H](CC(N)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCCCNC(C)C)C(=O)N1[C@@H](CCC1)C(=O)N[C@H](C)C(N)=O)N(C)C(=O)[C@H](CO)NC(=O)[C@@H](CC=1C=NC=CC=1)NC(=O)[C@@H](CC=1C=CC(Cl)=CC=1)NC(=O)[C@@H](CC=1C=C2C=CC=CC2=CC=1)NC(C)=O)C1=CC=C(O)C=C1 AIWRTTMUVOZGPW-HSPKUQOVSA-N 0.000 claims description 8
- 229960002184 abarelix Drugs 0.000 claims description 8
- JCKYGMPEJWAADB-UHFFFAOYSA-N chlorambucil Chemical compound OC(=O)CCCC1=CC=C(N(CCCl)CCCl)C=C1 JCKYGMPEJWAADB-UHFFFAOYSA-N 0.000 claims description 8
- 229960004630 chlorambucil Drugs 0.000 claims description 8
- STQGQHZAVUOBTE-VGBVRHCVSA-N daunorubicin Chemical compound O([C@H]1C[C@@](O)(CC=2C(O)=C3C(=O)C=4C=CC=C(C=4C(=O)C3=C(O)C=21)OC)C(C)=O)[C@H]1C[C@H](N)[C@H](O)[C@H](C)O1 STQGQHZAVUOBTE-VGBVRHCVSA-N 0.000 claims description 8
- 229960002258 fulvestrant Drugs 0.000 claims description 8
- 229960002913 goserelin Drugs 0.000 claims description 8
- KTUFNOKKBVMGRW-UHFFFAOYSA-N imatinib Chemical compound C1CN(C)CCN1CC1=CC=C(C(=O)NC=2C=C(NC=3N=C(C=CN=3)C=3C=NC=CC=3)C(C)=CC=2)C=C1 KTUFNOKKBVMGRW-UHFFFAOYSA-N 0.000 claims description 8
- 229960002411 imatinib Drugs 0.000 claims description 8
- 201000008968 osteosarcoma Diseases 0.000 claims description 8
- 229960004641 rituximab Drugs 0.000 claims description 8
- 229960001603 tamoxifen Drugs 0.000 claims description 8
- 229960000303 topotecan Drugs 0.000 claims description 8
- UCFGDBYHRUNTLO-QHCPKHFHSA-N topotecan Chemical compound C1=C(O)C(CN(C)C)=C2C=C(CN3C4=CC5=C(C3=O)COC(=O)[C@]5(O)CC)C4=NC2=C1 UCFGDBYHRUNTLO-QHCPKHFHSA-N 0.000 claims description 8
- XRASPMIURGNCCH-UHFFFAOYSA-N zoledronic acid Chemical class OP(=O)(O)C(P(O)(O)=O)(O)CN1C=CN=C1 XRASPMIURGNCCH-UHFFFAOYSA-N 0.000 claims description 8
- 229960000641 zorubicin Drugs 0.000 claims description 8
- FBTUMDXHSRTGRV-ALTNURHMSA-N zorubicin Chemical compound O([C@H]1C[C@@](O)(CC=2C(O)=C3C(=O)C=4C=CC=C(C=4C(=O)C3=C(O)C=21)OC)C(\C)=N\NC(=O)C=1C=CC=CC=1)[C@H]1C[C@H](N)[C@H](O)[C@H](C)O1 FBTUMDXHSRTGRV-ALTNURHMSA-N 0.000 claims description 8
- RTQWWZBSTRGEAV-PKHIMPSTSA-N 2-[[(2s)-2-[bis(carboxymethyl)amino]-3-[4-(methylcarbamoylamino)phenyl]propyl]-[2-[bis(carboxymethyl)amino]propyl]amino]acetic acid Chemical compound CNC(=O)NC1=CC=C(C[C@@H](CN(CC(C)N(CC(O)=O)CC(O)=O)CC(O)=O)N(CC(O)=O)CC(O)=O)C=C1 RTQWWZBSTRGEAV-PKHIMPSTSA-N 0.000 claims description 7
- AOJJSUZBOXZQNB-VTZDEGQISA-N 4'-epidoxorubicin Chemical compound O([C@H]1C[C@@](O)(CC=2C(O)=C3C(=O)C=4C=CC=C(C=4C(=O)C3=C(O)C=21)OC)C(=O)CO)[C@H]1C[C@H](N)[C@@H](O)[C@H](C)O1 AOJJSUZBOXZQNB-VTZDEGQISA-N 0.000 claims description 7
- VVIAGPKUTFNRDU-UHFFFAOYSA-N 6S-folinic acid Natural products C1NC=2NC(N)=NC(=O)C=2N(C=O)C1CNC1=CC=C(C(=O)NC(CCC(O)=O)C(O)=O)C=C1 VVIAGPKUTFNRDU-UHFFFAOYSA-N 0.000 claims description 7
- 241000796533 Arna Species 0.000 claims description 7
- BFYIZQONLCFLEV-DAELLWKTSA-N Aromasine Chemical compound O=C1C=C[C@]2(C)[C@H]3CC[C@](C)(C(CC4)=O)[C@@H]4[C@@H]3CC(=C)C2=C1 BFYIZQONLCFLEV-DAELLWKTSA-N 0.000 claims description 7
- 102000015790 Asparaginase Human genes 0.000 claims description 7
- 108010024976 Asparaginase Proteins 0.000 claims description 7
- COVZYZSDYWQREU-UHFFFAOYSA-N Busulfan Chemical compound CS(=O)(=O)OCCCCOS(C)(=O)=O COVZYZSDYWQREU-UHFFFAOYSA-N 0.000 claims description 7
- CMSMOCZEIVJLDB-UHFFFAOYSA-N Cyclophosphamide Chemical compound ClCCN(CCCl)P1(=O)NCCCO1 CMSMOCZEIVJLDB-UHFFFAOYSA-N 0.000 claims description 7
- UHDGCWIWMRVCDJ-CCXZUQQUSA-N Cytarabine Chemical compound O=C1N=C(N)C=CN1[C@H]1[C@@H](O)[C@H](O)[C@@H](CO)O1 UHDGCWIWMRVCDJ-CCXZUQQUSA-N 0.000 claims description 7
- 229930189413 Esperamicin Natural products 0.000 claims description 7
- YCKRFDGAMUMZLT-UHFFFAOYSA-N Fluorine atom Chemical compound [F] YCKRFDGAMUMZLT-UHFFFAOYSA-N 0.000 claims description 7
- MPJKWIXIYCLVCU-UHFFFAOYSA-N Folinic acid Natural products NC1=NC2=C(N(C=O)C(CNc3ccc(cc3)C(=O)NC(CCC(=O)O)CC(=O)O)CN2)C(=O)N1 MPJKWIXIYCLVCU-UHFFFAOYSA-N 0.000 claims description 7
- 102100040018 Interferon alpha-2 Human genes 0.000 claims description 7
- 108010079944 Interferon-alpha2b Proteins 0.000 claims description 7
- 239000005551 L01XE03 - Erlotinib Substances 0.000 claims description 7
- HLFSDGLLUJUHTE-SNVBAGLBSA-N Levamisole Chemical compound C1([C@H]2CN3CCSC3=N2)=CC=CC=C1 HLFSDGLLUJUHTE-SNVBAGLBSA-N 0.000 claims description 7
- 206010061535 Ovarian neoplasm Diseases 0.000 claims description 7
- OIRDTQYFTABQOQ-UHTZMRCNSA-N Vidarabine Chemical compound C1=NC=2C(N)=NC=NC=2N1[C@@H]1O[C@H](CO)[C@@H](O)[C@@H]1O OIRDTQYFTABQOQ-UHTZMRCNSA-N 0.000 claims description 7
- 229960000473 altretamine Drugs 0.000 claims description 7
- 229960003272 asparaginase Drugs 0.000 claims description 7
- DCXYFEDJOCDNAF-UHFFFAOYSA-M asparaginate Chemical compound [O-]C(=O)C(N)CC(N)=O DCXYFEDJOCDNAF-UHFFFAOYSA-M 0.000 claims description 7
- 229940120638 avastin Drugs 0.000 claims description 7
- 150000003851 azoles Chemical class 0.000 claims description 7
- 229960002092 busulfan Drugs 0.000 claims description 7
- 229960005395 cetuximab Drugs 0.000 claims description 7
- 229960000975 daunorubicin Drugs 0.000 claims description 7
- UREBDLICKHMUKA-CXSFZGCWSA-N dexamethasone Chemical compound C1CC2=CC(=O)C=C[C@]2(C)[C@]2(F)[C@@H]1[C@@H]1C[C@@H](C)[C@@](C(=O)CO)(O)[C@@]1(C)C[C@@H]2O UREBDLICKHMUKA-CXSFZGCWSA-N 0.000 claims description 7
- 229960003957 dexamethasone Drugs 0.000 claims description 7
- AAKJLRGGTJKAMG-UHFFFAOYSA-N erlotinib Chemical compound C=12C=C(OCCOC)C(OCCOC)=CC2=NC=NC=1NC1=CC=CC(C#C)=C1 AAKJLRGGTJKAMG-UHFFFAOYSA-N 0.000 claims description 7
- LJQQFQHBKUKHIS-WJHRIEJJSA-N esperamicin Chemical compound O1CC(NC(C)C)C(OC)CC1OC1C(O)C(NOC2OC(C)C(SC)C(O)C2)C(C)OC1OC1C(\C2=C/CSSSC)=C(NC(=O)OC)C(=O)C(OC3OC(C)C(O)C(OC(=O)C=4C(=CC(OC)=C(OC)C=4)NC(=O)C(=C)OC)C3)C2(O)C#C\C=C/C#C1 LJQQFQHBKUKHIS-WJHRIEJJSA-N 0.000 claims description 7
- FRPJXPJMRWBBIH-RBRWEJTLSA-N estramustine Chemical compound ClCCN(CCCl)C(=O)OC1=CC=C2[C@H]3CC[C@](C)([C@H](CC4)O)[C@@H]4[C@@H]3CCC2=C1 FRPJXPJMRWBBIH-RBRWEJTLSA-N 0.000 claims description 7
- 229960001842 estramustine Drugs 0.000 claims description 7
- VJJPUSNTGOMMGY-MRVIYFEKSA-N etoposide Chemical compound COC1=C(O)C(OC)=CC([C@@H]2C3=CC=4OCOC=4C=C3[C@@H](O[C@H]3[C@@H]([C@@H](O)[C@@H]4O[C@H](C)OC[C@H]4O3)O)[C@@H]3[C@@H]2C(OC3)=O)=C1 VJJPUSNTGOMMGY-MRVIYFEKSA-N 0.000 claims description 7
- 229960005420 etoposide Drugs 0.000 claims description 7
- 229960000255 exemestane Drugs 0.000 claims description 7
- 229910052731 fluorine Inorganic materials 0.000 claims description 7
- 239000011737 fluorine Substances 0.000 claims description 7
- 229960002949 fluorouracil Drugs 0.000 claims description 7
- VVIAGPKUTFNRDU-ABLWVSNPSA-N folinic acid Chemical compound C1NC=2NC(N)=NC(=O)C=2N(C=O)C1CNC1=CC=C(C(=O)N[C@@H](CCC(O)=O)C(O)=O)C=C1 VVIAGPKUTFNRDU-ABLWVSNPSA-N 0.000 claims description 7
- 235000008191 folinic acid Nutrition 0.000 claims description 7
- 239000011672 folinic acid Substances 0.000 claims description 7
- 229940022353 herceptin Drugs 0.000 claims description 7
- UUVWYPNAQBNQJQ-UHFFFAOYSA-N hexamethylmelamine Chemical compound CN(C)C1=NC(N(C)C)=NC(N(C)C)=N1 UUVWYPNAQBNQJQ-UHFFFAOYSA-N 0.000 claims description 7
- 229960001001 ibritumomab tiuxetan Drugs 0.000 claims description 7
- 229960003881 letrozole Drugs 0.000 claims description 7
- HPJKCIUCZWXJDR-UHFFFAOYSA-N letrozole Chemical compound C1=CC(C#N)=CC=C1C(N1N=CN=C1)C1=CC=C(C#N)C=C1 HPJKCIUCZWXJDR-UHFFFAOYSA-N 0.000 claims description 7
- 229960001691 leucovorin Drugs 0.000 claims description 7
- 229960001614 levamisole Drugs 0.000 claims description 7
- KKZJGLLVHKMTCM-UHFFFAOYSA-N mitoxantrone Chemical compound O=C1C2=C(O)C=CC(O)=C2C(=O)C2=C1C(NCCNCCO)=CC=C2NCCNCCO KKZJGLLVHKMTCM-UHFFFAOYSA-N 0.000 claims description 7
- 229960001156 mitoxantrone Drugs 0.000 claims description 7
- 201000009410 rhabdomyosarcoma Diseases 0.000 claims description 7
- 229940120982 tarceva Drugs 0.000 claims description 7
- 229960005026 toremifene Drugs 0.000 claims description 7
- XFCLJVABOIYOMF-QPLCGJKRSA-N toremifene Chemical compound C1=CC(OCCN(C)C)=CC=C1C(\C=1C=CC=CC=1)=C(\CCCl)C1=CC=CC=C1 XFCLJVABOIYOMF-QPLCGJKRSA-N 0.000 claims description 7
- 229960003636 vidarabine Drugs 0.000 claims description 7
- GBABOYUKABKIAF-GHYRFKGUSA-N vinorelbine Chemical compound C1N(CC=2C3=CC=CC=C3NC=22)CC(CC)=C[C@H]1C[C@]2(C(=O)OC)C1=CC([C@]23[C@H]([C@]([C@H](OC(C)=O)[C@]4(CC)C=CCN([C@H]34)CC2)(O)C(=O)OC)N2C)=C2C=C1OC GBABOYUKABKIAF-GHYRFKGUSA-N 0.000 claims description 7
- 229960002066 vinorelbine Drugs 0.000 claims description 7
- 206010008342 Cervix carcinoma Diseases 0.000 claims description 6
- 206010029260 Neuroblastoma Diseases 0.000 claims description 6
- 208000006105 Uterine Cervical Neoplasms Diseases 0.000 claims description 6
- 201000010881 cervical cancer Diseases 0.000 claims description 6
- 208000021045 exocrine pancreatic carcinoma Diseases 0.000 claims description 6
- 201000001441 melanoma Diseases 0.000 claims description 6
- 238000002360 preparation method Methods 0.000 claims description 6
- 206010035226 Plasma cell myeloma Diseases 0.000 claims description 5
- 208000034578 Multiple myelomas Diseases 0.000 claims description 4
- 230000001093 anti-cancer Effects 0.000 claims description 4
- 239000003937 drug carrier Substances 0.000 claims description 3
- 230000035772 mutation Effects 0.000 claims description 3
- 238000012217 deletion Methods 0.000 claims description 2
- 230000037430 deletion Effects 0.000 claims description 2
- 238000003780 insertion Methods 0.000 claims description 2
- 230000037431 insertion Effects 0.000 claims description 2
- 239000008194 pharmaceutical composition Substances 0.000 claims 2
- BTCSSZJGUNDROE-UHFFFAOYSA-N gamma-aminobutyric acid Chemical compound NCCCC(O)=O BTCSSZJGUNDROE-UHFFFAOYSA-N 0.000 claims 1
- 210000000496 pancreas Anatomy 0.000 claims 1
- 108010031794 IGF Type 1 Receptor Proteins 0.000 abstract description 35
- 210000002966 serum Anatomy 0.000 abstract description 33
- 230000004083 survival effect Effects 0.000 abstract description 27
- 230000000694 effects Effects 0.000 abstract description 24
- 210000004881 tumor cell Anatomy 0.000 abstract description 23
- 102000048143 Insulin-Like Growth Factor II Human genes 0.000 abstract description 21
- 108090001117 Insulin-Like Growth Factor II Proteins 0.000 abstract description 21
- 239000003814 drug Substances 0.000 abstract description 14
- 239000000556 agonist Substances 0.000 abstract description 11
- 229940127089 cytotoxic agent Drugs 0.000 abstract description 11
- 239000002254 cytotoxic agent Substances 0.000 abstract description 11
- 231100000599 cytotoxic agent Toxicity 0.000 abstract description 6
- 229940124597 therapeutic agent Drugs 0.000 abstract description 5
- 238000003384 imaging method Methods 0.000 abstract description 3
- 206010061902 Pancreatic neoplasm Diseases 0.000 abstract description 2
- 208000029742 colonic neoplasm Diseases 0.000 abstract description 2
- 238000003745 diagnosis Methods 0.000 abstract description 2
- 208000015486 malignant pancreatic neoplasm Diseases 0.000 abstract description 2
- 208000008443 pancreatic carcinoma Diseases 0.000 abstract description 2
- 102000038455 IGF Type 1 Receptor Human genes 0.000 abstract 3
- 201000002528 pancreatic cancer Diseases 0.000 abstract 1
- 239000000370 acceptor Substances 0.000 description 122
- 241000699666 Mus <mouse, genus> Species 0.000 description 72
- 108010051242 phenylalanylserine Proteins 0.000 description 60
- 241000282414 Homo sapiens Species 0.000 description 45
- 108010050848 glycylleucine Proteins 0.000 description 43
- 108010089804 glycyl-threonine Proteins 0.000 description 34
- 102100039688 Insulin-like growth factor 1 receptor Human genes 0.000 description 33
- 239000003153 chemical reaction reagent Substances 0.000 description 33
- UIPXCLNLUUAMJU-JBDRJPRFSA-N Ser-Ile-Ser Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CO)C(O)=O UIPXCLNLUUAMJU-JBDRJPRFSA-N 0.000 description 30
- MRQQMVZUHXUPEV-IHRRRGAJSA-N Asp-Arg-Phe Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC1=CC=CC=C1)C(O)=O MRQQMVZUHXUPEV-IHRRRGAJSA-N 0.000 description 29
- NRFGTHFONZYFNY-MGHWNKPDSA-N Leu-Ile-Tyr Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@H](C(O)=O)CC1=CC=C(O)C=C1 NRFGTHFONZYFNY-MGHWNKPDSA-N 0.000 description 29
- SPSSJSICDYYTQN-HJGDQZAQSA-N Met-Thr-Gln Chemical compound CSCC[C@H](N)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@H](C(O)=O)CCC(N)=O SPSSJSICDYYTQN-HJGDQZAQSA-N 0.000 description 29
- 241000699660 Mus musculus Species 0.000 description 29
- JMVQDLDPDBXAAX-YUMQZZPRSA-N Pro-Gly-Gln Chemical compound NC(=O)CC[C@@H](C(O)=O)NC(=O)CNC(=O)[C@@H]1CCCN1 JMVQDLDPDBXAAX-YUMQZZPRSA-N 0.000 description 29
- UIGMAMGZOJVTDN-WHFBIAKZSA-N Ser-Gly-Ser Chemical compound OC[C@H](N)C(=O)NCC(=O)N[C@@H](CO)C(O)=O UIGMAMGZOJVTDN-WHFBIAKZSA-N 0.000 description 29
- KRPKYGOFYUNIGM-XVSYOHENSA-N Thr-Asp-Phe Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CC(=O)O)C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)O)N)O KRPKYGOFYUNIGM-XVSYOHENSA-N 0.000 description 29
- 108010081404 acein-2 Proteins 0.000 description 29
- HUZGPXBILPMCHM-IHRRRGAJSA-N Asn-Arg-Phe Chemical compound [H]N[C@@H](CC(N)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC1=CC=CC=C1)C(O)=O HUZGPXBILPMCHM-IHRRRGAJSA-N 0.000 description 28
- KCGIREHVWRXNDH-GARJFASQSA-N Ser-Leu-Pro Chemical compound CC(C)C[C@@H](C(=O)N1CCC[C@@H]1C(=O)O)NC(=O)[C@H](CO)N KCGIREHVWRXNDH-GARJFASQSA-N 0.000 description 28
- 235000001014 amino acid Nutrition 0.000 description 28
- 150000001413 amino acids Chemical class 0.000 description 28
- 108010057083 glutamyl-aspartyl-leucine Proteins 0.000 description 28
- AYFVRYXNDHBECD-YUMQZZPRSA-N Asp-Leu-Gly Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](CC(C)C)C(=O)NCC(O)=O AYFVRYXNDHBECD-YUMQZZPRSA-N 0.000 description 27
- GEEXORWTBTUOHC-FXQIFTODSA-N Cys-Arg-Ser Chemical compound C(C[C@@H](C(=O)N[C@@H](CO)C(=O)O)NC(=O)[C@H](CS)N)CN=C(N)N GEEXORWTBTUOHC-FXQIFTODSA-N 0.000 description 27
- BBQIWFFTTQTNOC-AVGNSLFASA-N Cys-Phe-Gln Chemical compound C1=CC=C(C=C1)C[C@@H](C(=O)N[C@@H](CCC(=O)N)C(=O)O)NC(=O)[C@H](CS)N BBQIWFFTTQTNOC-AVGNSLFASA-N 0.000 description 27
- CGWHAXBNGYQBBK-JBACZVJFSA-N Glu-Trp-Tyr Chemical compound C([C@H](NC(=O)[C@H](CC=1C2=CC=CC=C2NC=1)NC(=O)[C@H](CCC(O)=O)N)C(O)=O)C1=CC=C(O)C=C1 CGWHAXBNGYQBBK-JBACZVJFSA-N 0.000 description 27
- XHVONGZZVUUORG-WEDXCCLWSA-N Gly-Thr-Lys Chemical compound NCC(=O)N[C@@H]([C@H](O)C)C(=O)N[C@H](C(O)=O)CCCCN XHVONGZZVUUORG-WEDXCCLWSA-N 0.000 description 27
- 108010065920 Insulin Lispro Proteins 0.000 description 27
- FQZPTCNSNPWHLJ-AVGNSLFASA-N Leu-Gln-Lys Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCCCN)C(O)=O FQZPTCNSNPWHLJ-AVGNSLFASA-N 0.000 description 27
- GILLQRYAWOMHED-DCAQKATOSA-N Lys-Val-Ser Chemical compound OC[C@@H](C(O)=O)NC(=O)[C@H](C(C)C)NC(=O)[C@@H](N)CCCCN GILLQRYAWOMHED-DCAQKATOSA-N 0.000 description 27
- HBOABDXGTMMDSE-GUBZILKMSA-N Ser-Arg-Val Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](C(C)C)C(O)=O HBOABDXGTMMDSE-GUBZILKMSA-N 0.000 description 27
- XXXAXOWMBOKTRN-XPUUQOCRSA-N Ser-Gly-Val Chemical compound [H]N[C@@H](CO)C(=O)NCC(=O)N[C@@H](C(C)C)C(O)=O XXXAXOWMBOKTRN-XPUUQOCRSA-N 0.000 description 27
- ZUDXUJSYCCNZQJ-DCAQKATOSA-N Ser-His-Val Chemical compound CC(C)[C@@H](C(=O)O)NC(=O)[C@H](CC1=CN=CN1)NC(=O)[C@H](CO)N ZUDXUJSYCCNZQJ-DCAQKATOSA-N 0.000 description 27
- DEGCBBCMYWNJNA-RHYQMDGZSA-N Thr-Pro-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@@H]1CCCN1C(=O)[C@@H](N)[C@@H](C)O DEGCBBCMYWNJNA-RHYQMDGZSA-N 0.000 description 27
- JAWUQFCGNVEDRN-MEYUZBJRSA-N Thr-Tyr-Leu Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CC1=CC=C(C=C1)O)C(=O)N[C@@H](CC(C)C)C(=O)O)N)O JAWUQFCGNVEDRN-MEYUZBJRSA-N 0.000 description 27
- VHIZXDZMTDVFGX-DCAQKATOSA-N Val-Ser-Leu Chemical compound CC(C)C[C@@H](C(=O)O)NC(=O)[C@H](CO)NC(=O)[C@H](C(C)C)N VHIZXDZMTDVFGX-DCAQKATOSA-N 0.000 description 27
- OWFGFHQMSBTKLX-UFYCRDLUSA-N Val-Tyr-Tyr Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CC1=CC=C(C=C1)O)C(=O)N[C@@H](CC2=CC=C(C=C2)O)C(=O)O)N OWFGFHQMSBTKLX-UFYCRDLUSA-N 0.000 description 27
- 108010078144 glutaminyl-glycine Proteins 0.000 description 27
- LKDIBBOKUAASNP-FXQIFTODSA-N Glu-Ala-Glu Chemical compound OC(=O)CC[C@H](N)C(=O)N[C@@H](C)C(=O)N[C@@H](CCC(O)=O)C(O)=O LKDIBBOKUAASNP-FXQIFTODSA-N 0.000 description 26
- 230000002018 overexpression Effects 0.000 description 26
- JCGMFFQQHJQASB-PYJNHQTQSA-N Ile-Val-His Chemical compound N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CC1=CNC=N1)C(=O)O JCGMFFQQHJQASB-PYJNHQTQSA-N 0.000 description 25
- HPBCTWSUJOGJSH-MNXVOIDGSA-N Leu-Glu-Ile Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O HPBCTWSUJOGJSH-MNXVOIDGSA-N 0.000 description 25
- XKUKSGPZAADMRA-UHFFFAOYSA-N glycyl-glycyl-glycine Natural products NCC(=O)NCC(=O)NCC(O)=O XKUKSGPZAADMRA-UHFFFAOYSA-N 0.000 description 25
- NAXPHWZXEXNDIW-JTQLQIEISA-N Phe-Gly-Gly Chemical compound OC(=O)CNC(=O)CNC(=O)[C@@H](N)CC1=CC=CC=C1 NAXPHWZXEXNDIW-JTQLQIEISA-N 0.000 description 24
- 238000013016 damping Methods 0.000 description 24
- 239000012530 fluid Substances 0.000 description 24
- 108010070409 phenylalanyl-glycyl-glycine Proteins 0.000 description 24
- FMDHKPRACUXATF-ACZMJKKPSA-N Ser-Gln-Ser Chemical compound OC[C@H](N)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CO)C(O)=O FMDHKPRACUXATF-ACZMJKKPSA-N 0.000 description 23
- 108010018006 histidylserine Proteins 0.000 description 23
- GIKOVDMXBAFXDF-NHCYSSNCSA-N Asp-Val-Leu Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CC(C)C)C(O)=O GIKOVDMXBAFXDF-NHCYSSNCSA-N 0.000 description 22
- VPZXBVLAVMBEQI-UHFFFAOYSA-N glycyl-DL-alpha-alanine Natural products OC(=O)C(C)NC(=O)CN VPZXBVLAVMBEQI-UHFFFAOYSA-N 0.000 description 22
- 230000005764 inhibitory process Effects 0.000 description 22
- 108090000623 proteins and genes Proteins 0.000 description 22
- RMODQFBNDDENCP-IHRRRGAJSA-N Pro-Lys-Leu Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC(C)C)C(O)=O RMODQFBNDDENCP-IHRRRGAJSA-N 0.000 description 21
- XHWCDRUPDNSDAZ-XKBZYTNZSA-N Thr-Ser-Glu Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CO)C(=O)N[C@@H](CCC(=O)O)C(=O)O)N)O XHWCDRUPDNSDAZ-XKBZYTNZSA-N 0.000 description 21
- 230000008859 change Effects 0.000 description 21
- 238000006243 chemical reaction Methods 0.000 description 21
- 108010063718 gamma-glutamylaspartic acid Proteins 0.000 description 21
- RCYUBVHMVUHEBM-RCWTZXSCSA-N Pro-Pro-Thr Chemical compound [H]N1CCC[C@H]1C(=O)N1CCC[C@H]1C(=O)N[C@@H]([C@@H](C)O)C(O)=O RCYUBVHMVUHEBM-RCWTZXSCSA-N 0.000 description 20
- BMKNXTJLHFIAAH-CIUDSAMLSA-N Ser-Ser-Leu Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CO)C(=O)N[C@@H](CC(C)C)C(O)=O BMKNXTJLHFIAAH-CIUDSAMLSA-N 0.000 description 20
- LGIMRDKGABDMBN-DCAQKATOSA-N Ser-Val-Lys Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CCCCN)C(=O)O)NC(=O)[C@H](CO)N LGIMRDKGABDMBN-DCAQKATOSA-N 0.000 description 20
- 108010015385 valyl-prolyl-proline Proteins 0.000 description 20
- NFGXHKASABOEEW-UHFFFAOYSA-N 1-methylethyl 11-methoxy-3,7,11-trimethyl-2,4-dodecadienoate Chemical compound COC(C)(C)CCCC(C)CC=CC(C)=CC(=O)OC(C)C NFGXHKASABOEEW-UHFFFAOYSA-N 0.000 description 19
- WMLFFCRUSPNENW-ZLUOBGJFSA-N Asp-Ser-Ala Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](CO)C(=O)N[C@@H](C)C(O)=O WMLFFCRUSPNENW-ZLUOBGJFSA-N 0.000 description 19
- GBYYQVBXFVDJPJ-WLTAIBSBSA-N Gly-Tyr-Thr Chemical compound C[C@H]([C@@H](C(=O)O)NC(=O)[C@H](CC1=CC=C(C=C1)O)NC(=O)CN)O GBYYQVBXFVDJPJ-WLTAIBSBSA-N 0.000 description 19
- VOOINLQYUZOREH-SRVKXCTJSA-N Met-Gln-Leu Chemical compound CC(C)C[C@@H](C(=O)O)NC(=O)[C@H](CCC(=O)N)NC(=O)[C@H](CCSC)N VOOINLQYUZOREH-SRVKXCTJSA-N 0.000 description 19
- 108010086434 alanyl-seryl-glycine Proteins 0.000 description 19
- 108010092854 aspartyllysine Proteins 0.000 description 19
- IOFVWPYSRSCWHI-JXUBOQSCSA-N Ala-Thr-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@H]([C@@H](C)O)NC(=O)[C@H](C)N IOFVWPYSRSCWHI-JXUBOQSCSA-N 0.000 description 18
- ZFBBMCKQSNJZSN-AUTRQRHGSA-N Gln-Val-Gln Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CCC(N)=O)C(O)=O ZFBBMCKQSNJZSN-AUTRQRHGSA-N 0.000 description 18
- FYBFTPLPAXZBOY-KKHAAJSZSA-N Thr-Val-Asp Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CC(O)=O)C(O)=O FYBFTPLPAXZBOY-KKHAAJSZSA-N 0.000 description 18
- MJIJBEYEHBKTIM-BYULHYEWSA-N Asn-Val-Asn Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CC(=O)N)C(=O)O)NC(=O)[C@H](CC(=O)N)N MJIJBEYEHBKTIM-BYULHYEWSA-N 0.000 description 17
- NYQHSUGFEWDWPD-ACZMJKKPSA-N Asp-Gln-Ala Chemical compound C[C@@H](C(=O)O)NC(=O)[C@H](CCC(=O)N)NC(=O)[C@H](CC(=O)O)N NYQHSUGFEWDWPD-ACZMJKKPSA-N 0.000 description 17
- XOKGKOQWADCLFQ-GARJFASQSA-N Gln-Arg-Pro Chemical compound C1C[C@@H](N(C1)C(=O)[C@H](CCCN=C(N)N)NC(=O)[C@H](CCC(=O)N)N)C(=O)O XOKGKOQWADCLFQ-GARJFASQSA-N 0.000 description 17
- SOEGEPHNZOISMT-BYPYZUCNSA-N Gly-Ser-Gly Chemical compound NCC(=O)N[C@@H](CO)C(=O)NCC(O)=O SOEGEPHNZOISMT-BYPYZUCNSA-N 0.000 description 17
- FEHQLKKBVJHSEC-SZMVWBNQSA-N Leu-Glu-Trp Chemical compound C1=CC=C2C(C[C@H](NC(=O)[C@H](CCC(O)=O)NC(=O)[C@@H](N)CC(C)C)C(O)=O)=CNC2=C1 FEHQLKKBVJHSEC-SZMVWBNQSA-N 0.000 description 17
- UWKNTTJNVSYXPC-CIUDSAMLSA-N Lys-Ala-Ser Chemical compound OC[C@@H](C(O)=O)NC(=O)[C@H](C)NC(=O)[C@@H](N)CCCCN UWKNTTJNVSYXPC-CIUDSAMLSA-N 0.000 description 17
- 108010038320 lysylphenylalanine Proteins 0.000 description 17
- 230000000638 stimulation Effects 0.000 description 17
- 108010073969 valyllysine Proteins 0.000 description 17
- BYYNJRSNDARRBX-YFKPBYRVSA-N Gly-Gln-Gly Chemical compound NCC(=O)N[C@@H](CCC(N)=O)C(=O)NCC(O)=O BYYNJRSNDARRBX-YFKPBYRVSA-N 0.000 description 16
- PDIDTSZKKFEDMB-UWVGGRQHSA-N Lys-Pro-Gly Chemical compound [H]N[C@@H](CCCCN)C(=O)N1CCC[C@H]1C(=O)NCC(O)=O PDIDTSZKKFEDMB-UWVGGRQHSA-N 0.000 description 16
- XJDMUQCLVSCRSJ-VZFHVOOUSA-N Ser-Thr-Ala Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](C)C(O)=O XJDMUQCLVSCRSJ-VZFHVOOUSA-N 0.000 description 16
- UOXPLPBMEPLZBW-WDSOQIARSA-N Trp-Val-Lys Chemical compound C1=CC=C2C(C[C@H](N)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CCCCN)C(O)=O)=CNC2=C1 UOXPLPBMEPLZBW-WDSOQIARSA-N 0.000 description 16
- 230000006870 function Effects 0.000 description 16
- NXRNRBOKDBIVKQ-CXTHYWKRSA-N Ile-Tyr-Tyr Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CC1=CC=C(C=C1)O)C(=O)N[C@@H](CC2=CC=C(C=C2)O)C(=O)O)N NXRNRBOKDBIVKQ-CXTHYWKRSA-N 0.000 description 15
- KAFOIVJDVSZUMD-UHFFFAOYSA-N Leu-Gln-Gln Natural products CC(C)CC(N)C(=O)NC(CCC(N)=O)C(=O)NC(CCC(N)=O)C(O)=O KAFOIVJDVSZUMD-UHFFFAOYSA-N 0.000 description 15
- KIZIOFNVSOSKJI-CIUDSAMLSA-N Leu-Ser-Cys Chemical compound CC(C)C[C@@H](C(=O)N[C@@H](CO)C(=O)N[C@@H](CS)C(=O)O)N KIZIOFNVSOSKJI-CIUDSAMLSA-N 0.000 description 15
- AMXMBCAXAZUCFA-RHYQMDGZSA-N Thr-Leu-Arg Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O AMXMBCAXAZUCFA-RHYQMDGZSA-N 0.000 description 15
- 238000011160 research Methods 0.000 description 15
- GJBUAAAIZSRCDC-GVXVVHGQSA-N Glu-Leu-Val Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](C(C)C)C(O)=O GJBUAAAIZSRCDC-GVXVVHGQSA-N 0.000 description 14
- STVHDEHTKFXBJQ-LAEOZQHASA-N Gly-Glu-Ile Chemical compound [H]NCC(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O STVHDEHTKFXBJQ-LAEOZQHASA-N 0.000 description 14
- KAFOIVJDVSZUMD-DCAQKATOSA-N Leu-Gln-Gln Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCC(N)=O)C(O)=O KAFOIVJDVSZUMD-DCAQKATOSA-N 0.000 description 14
- DIBZLYZXTSVGLN-CIUDSAMLSA-N Lys-Ser-Ser Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H](CO)C(=O)N[C@@H](CO)C(O)=O DIBZLYZXTSVGLN-CIUDSAMLSA-N 0.000 description 14
- UQFYNFTYDHUIMI-WHFBIAKZSA-N Ser-Gly-Ala Chemical compound OC(=O)[C@H](C)NC(=O)CNC(=O)[C@@H](N)CO UQFYNFTYDHUIMI-WHFBIAKZSA-N 0.000 description 14
- 108010068265 aspartyltyrosine Proteins 0.000 description 14
- 230000000452 restraining effect Effects 0.000 description 14
- 238000005406 washing Methods 0.000 description 14
- 238000002965 ELISA Methods 0.000 description 13
- GNRMAQSIROFNMI-IXOXFDKPSA-N Phe-Thr-Ser Chemical compound [H]N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CO)C(O)=O GNRMAQSIROFNMI-IXOXFDKPSA-N 0.000 description 13
- KCZGSXPFPNKGLE-WDSOQIARSA-N Trp-Met-His Chemical compound CSCC[C@@H](C(=O)N[C@@H](CC1=CN=CN1)C(=O)O)NC(=O)[C@H](CC2=CNC3=CC=CC=C32)N KCZGSXPFPNKGLE-WDSOQIARSA-N 0.000 description 13
- 238000007413 biotinylation Methods 0.000 description 13
- 230000006287 biotinylation Effects 0.000 description 13
- 230000014509 gene expression Effects 0.000 description 13
- JBGSZRYCXBPWGX-BQBZGAKWSA-N Ala-Arg-Gly Chemical compound OC(=O)CNC(=O)[C@@H](NC(=O)[C@@H](N)C)CCCN=C(N)N JBGSZRYCXBPWGX-BQBZGAKWSA-N 0.000 description 12
- OLGCWMNDJTWQAG-GUBZILKMSA-N Asn-Glu-Lys Chemical compound NCCCC[C@@H](C(O)=O)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@@H](N)CC(N)=O OLGCWMNDJTWQAG-GUBZILKMSA-N 0.000 description 12
- KPSHWSWFPUDEGF-FXQIFTODSA-N Asp-Pro-Ala Chemical compound OC(=O)[C@H](C)NC(=O)[C@@H]1CCCN1C(=O)[C@@H](N)CC(O)=O KPSHWSWFPUDEGF-FXQIFTODSA-N 0.000 description 12
- UGVQELHRNUDMAA-BYPYZUCNSA-N Gly-Ala-Gly Chemical compound [NH3+]CC(=O)N[C@@H](C)C(=O)NCC([O-])=O UGVQELHRNUDMAA-BYPYZUCNSA-N 0.000 description 12
- KKBWDNZXYLGJEY-UHFFFAOYSA-N Gly-Arg-Pro Natural products NCC(=O)NC(CCNC(=N)N)C(=O)N1CCCC1C(=O)O KKBWDNZXYLGJEY-UHFFFAOYSA-N 0.000 description 12
- 108010047562 NGR peptide Proteins 0.000 description 12
- MECLEFZMPPOEAC-VOAKCMCISA-N Thr-Leu-Lys Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCCCN)C(=O)O)N)O MECLEFZMPPOEAC-VOAKCMCISA-N 0.000 description 12
- 231100000433 cytotoxic Toxicity 0.000 description 12
- 230000001472 cytotoxic effect Effects 0.000 description 12
- 239000013613 expression plasmid Substances 0.000 description 12
- JVTHIXKSVYEWNI-JRQIVUDYSA-N Thr-Asn-Tyr Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(O)=O JVTHIXKSVYEWNI-JRQIVUDYSA-N 0.000 description 11
- 238000004458 analytical method Methods 0.000 description 11
- 108010037850 glycylvaline Proteins 0.000 description 11
- 230000026731 phosphorylation Effects 0.000 description 11
- 238000006366 phosphorylation reaction Methods 0.000 description 11
- 238000007789 sealing Methods 0.000 description 11
- 239000006228 supernatant Substances 0.000 description 11
- DDPXDCKYWDGZAL-BQBZGAKWSA-N Asn-Gly-Arg Chemical compound NC(=O)C[C@H](N)C(=O)NCC(=O)N[C@H](C(O)=O)CCCN=C(N)N DDPXDCKYWDGZAL-BQBZGAKWSA-N 0.000 description 10
- GMUOCGCDOYYWPD-FXQIFTODSA-N Asn-Pro-Ser Chemical compound [H]N[C@@H](CC(N)=O)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CO)C(O)=O GMUOCGCDOYYWPD-FXQIFTODSA-N 0.000 description 10
- 101000599951 Homo sapiens Insulin-like growth factor I Proteins 0.000 description 10
- KZTLZZQTJMCGIP-ZJDVBMNYSA-N Thr-Val-Thr Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H]([C@@H](C)O)C(O)=O KZTLZZQTJMCGIP-ZJDVBMNYSA-N 0.000 description 10
- 238000010276 construction Methods 0.000 description 10
- 230000008569 process Effects 0.000 description 10
- 238000012545 processing Methods 0.000 description 10
- 239000012679 serum free medium Substances 0.000 description 10
- OVQJAKFLFTZDNC-GUBZILKMSA-N Arg-Pro-Asp Chemical compound [H]N[C@@H](CCCNC(N)=N)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CC(O)=O)C(O)=O OVQJAKFLFTZDNC-GUBZILKMSA-N 0.000 description 9
- RVKIPWVMZANZLI-UHFFFAOYSA-N H-Lys-Trp-OH Natural products C1=CC=C2C(CC(NC(=O)C(N)CCCCN)C(O)=O)=CNC2=C1 RVKIPWVMZANZLI-UHFFFAOYSA-N 0.000 description 9
- 102000003746 Insulin Receptor Human genes 0.000 description 9
- 108010001127 Insulin Receptor Proteins 0.000 description 9
- 108010087066 N2-tryptophyllysine Proteins 0.000 description 9
- OZPDGESCTGGNAD-CIUDSAMLSA-N Ser-Ser-Lys Chemical compound NCCCC[C@@H](C(O)=O)NC(=O)[C@H](CO)NC(=O)[C@@H](N)CO OZPDGESCTGGNAD-CIUDSAMLSA-N 0.000 description 9
- WYOBRXPIZVKNMF-IRXDYDNUSA-N Tyr-Tyr-Gly Chemical compound C([C@H](N)C(=O)N[C@@H](CC=1C=CC(O)=CC=1)C(=O)NCC(O)=O)C1=CC=C(O)C=C1 WYOBRXPIZVKNMF-IRXDYDNUSA-N 0.000 description 9
- QHSSPPHOHJSTML-HOCLYGCPSA-N Val-Trp-Gly Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CC1=CNC2=CC=CC=C21)C(=O)NCC(=O)O)N QHSSPPHOHJSTML-HOCLYGCPSA-N 0.000 description 9
- 238000004140 cleaning Methods 0.000 description 9
- 238000001943 fluorescence-activated cell sorting Methods 0.000 description 9
- 230000002401 inhibitory effect Effects 0.000 description 9
- 102000013415 peroxidase activity proteins Human genes 0.000 description 9
- 108040007629 peroxidase activity proteins Proteins 0.000 description 9
- 239000000523 sample Substances 0.000 description 9
- 239000000243 solution Substances 0.000 description 9
- 239000000758 substrate Substances 0.000 description 9
- QAPSNMNOIOSXSQ-YNEHKIRRSA-N 1-[(2r,4s,5r)-4-[tert-butyl(dimethyl)silyl]oxy-5-(hydroxymethyl)oxolan-2-yl]-5-methylpyrimidine-2,4-dione Chemical compound O=C1NC(=O)C(C)=CN1[C@@H]1O[C@H](CO)[C@@H](O[Si](C)(C)C(C)(C)C)C1 QAPSNMNOIOSXSQ-YNEHKIRRSA-N 0.000 description 8
- DXVMJJNAOVECBA-WHFBIAKZSA-N Asn-Gly-Asn Chemical compound NC(=O)C[C@H](N)C(=O)NCC(=O)N[C@@H](CC(N)=O)C(O)=O DXVMJJNAOVECBA-WHFBIAKZSA-N 0.000 description 8
- NKKFVJRLCCUJNA-QWRGUYRKSA-N Lys-Gly-Lys Chemical compound NCCCC[C@H](N)C(=O)NCC(=O)N[C@H](C(O)=O)CCCCN NKKFVJRLCCUJNA-QWRGUYRKSA-N 0.000 description 8
- 108010079364 N-glycylalanine Proteins 0.000 description 8
- 239000000427 antigen Substances 0.000 description 8
- 102000036639 antigens Human genes 0.000 description 8
- 108091007433 antigens Proteins 0.000 description 8
- OHDRQQURAXLVGJ-HLVWOLMTSA-N azane;(2e)-3-ethyl-2-[(e)-(3-ethyl-6-sulfo-1,3-benzothiazol-2-ylidene)hydrazinylidene]-1,3-benzothiazole-6-sulfonic acid Chemical compound [NH4+].[NH4+].S/1C2=CC(S([O-])(=O)=O)=CC=C2N(CC)C\1=N/N=C1/SC2=CC(S([O-])(=O)=O)=CC=C2N1CC OHDRQQURAXLVGJ-HLVWOLMTSA-N 0.000 description 8
- 239000007853 buffer solution Substances 0.000 description 8
- 230000000295 complement effect Effects 0.000 description 8
- 239000002299 complementary DNA Substances 0.000 description 8
- 235000018102 proteins Nutrition 0.000 description 8
- 102000004169 proteins and genes Human genes 0.000 description 8
- UOWVMDUEMSNCAV-WYENRQIDSA-N rachelmycin Chemical compound C1([C@]23C[C@@H]2CN1C(=O)C=1NC=2C(OC)=C(O)C4=C(C=2C=1)CCN4C(=O)C1=CC=2C=4CCN(C=4C(O)=C(C=2N1)OC)C(N)=O)=CC(=O)C1=C3C(C)=CN1 UOWVMDUEMSNCAV-WYENRQIDSA-N 0.000 description 8
- CLNJSLSHKJECME-BQBZGAKWSA-N Pro-Gly-Ala Chemical compound OC(=O)[C@H](C)NC(=O)CNC(=O)[C@@H]1CCCN1 CLNJSLSHKJECME-BQBZGAKWSA-N 0.000 description 7
- XQJCEKXQUJQNNK-ZLUOBGJFSA-N Ser-Ser-Ser Chemical compound OC[C@H](N)C(=O)N[C@@H](CO)C(=O)N[C@@H](CO)C(O)=O XQJCEKXQUJQNNK-ZLUOBGJFSA-N 0.000 description 7
- UIRVSEPRMWDVEW-RNXOBYDBSA-N Trp-Tyr-Phe Chemical compound C1=CC=C(C=C1)C[C@@H](C(=O)O)NC(=O)[C@H](CC2=CC=C(C=C2)O)NC(=O)[C@H](CC3=CNC4=CC=CC=C43)N UIRVSEPRMWDVEW-RNXOBYDBSA-N 0.000 description 7
- PZTZYZUTCPZWJH-FXQIFTODSA-N Val-Ser-Ser Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CO)C(=O)N[C@@H](CO)C(=O)O)N PZTZYZUTCPZWJH-FXQIFTODSA-N 0.000 description 7
- YAJCHEVQCOHZDC-QMMNLEPNSA-N actrapid Chemical compound C([C@@H](C(=O)N[C@@H](CC(C)C)C(=O)N[C@H]1CSSC[C@H]2C(=O)N[C@H](C(=O)N[C@@H](CO)C(=O)N[C@H](C(=O)N[C@@H](C(N[C@@H](CO)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC=3C=CC(O)=CC=3)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CC=3C=CC(O)=CC=3)C(=O)N[C@@H](CSSC[C@H](NC(=O)[C@H](C(C)C)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CC=3C=CC(O)=CC=3)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](C)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](C(C)C)NC(=O)[C@H](CC(C)C)NC(=O)[C@H](CC=3N=CNC=3)NC(=O)[C@H](CO)NC(=O)CNC1=O)C(=O)NCC(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)NCC(=O)N[C@@H](CC=1C=CC=CC=1)C(=O)N[C@@H](CC=1C=CC=CC=1)C(=O)N[C@@H](CC=1C=CC(O)=CC=1)C(=O)N[C@@H]([C@H](C)O)C(=O)N1[C@@H](CCC1)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H]([C@H](C)O)C(O)=O)C(=O)N[C@@H](CC(N)=O)C(O)=O)=O)CSSC[C@@H](C(N2)=O)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@H](C(C)C)NC(=O)[C@@H](NC(=O)CN)[C@H](C)CC)[C@H](C)CC)[C@H](C)O)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@@H](NC(=O)[C@@H](NC(=O)[C@@H](N)CC=1C=CC=CC=1)C(C)C)C(N)=O)C1=CNC=N1 YAJCHEVQCOHZDC-QMMNLEPNSA-N 0.000 description 7
- 201000010099 disease Diseases 0.000 description 7
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 7
- 238000011156 evaluation Methods 0.000 description 7
- 238000011534 incubation Methods 0.000 description 7
- 229950004152 insulin human Drugs 0.000 description 7
- 239000000047 product Substances 0.000 description 7
- 108010029020 prolylglycine Proteins 0.000 description 7
- ZUNMTUPRQMWMHX-LSJOCFKGSA-N Asp-Val-Val Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](C(C)C)C(O)=O ZUNMTUPRQMWMHX-LSJOCFKGSA-N 0.000 description 6
- 241000282326 Felis catus Species 0.000 description 6
- SJNZALDHDUYDBU-IHRRRGAJSA-N Lys-Arg-Lys Chemical compound NCCCC[C@H](N)C(=O)N[C@@H](CCCN=C(N)N)C(=O)N[C@@H](CCCCN)C(O)=O SJNZALDHDUYDBU-IHRRRGAJSA-N 0.000 description 6
- 241001494479 Pecora Species 0.000 description 6
- 102000003993 Phosphatidylinositol 3-kinases Human genes 0.000 description 6
- 108090000430 Phosphatidylinositol 3-kinases Proteins 0.000 description 6
- VCYJKOLZYPYGJV-AVGNSLFASA-N Pro-Arg-Leu Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC(C)C)C(O)=O VCYJKOLZYPYGJV-AVGNSLFASA-N 0.000 description 6
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 6
- XSQUKJJJFZCRTK-UHFFFAOYSA-N Urea Chemical compound NC(N)=O XSQUKJJJFZCRTK-UHFFFAOYSA-N 0.000 description 6
- HTONZBWRYUKUKC-RCWTZXSCSA-N Val-Thr-Val Chemical compound CC(C)[C@H](N)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](C(C)C)C(O)=O HTONZBWRYUKUKC-RCWTZXSCSA-N 0.000 description 6
- 230000008485 antagonism Effects 0.000 description 6
- 230000006907 apoptotic process Effects 0.000 description 6
- 230000035578 autophosphorylation Effects 0.000 description 6
- 230000012447 hatching Effects 0.000 description 6
- 238000002347 injection Methods 0.000 description 6
- 239000007924 injection Substances 0.000 description 6
- 239000006166 lysate Substances 0.000 description 6
- 239000011782 vitamin Substances 0.000 description 6
- 229940088594 vitamin Drugs 0.000 description 6
- 235000013343 vitamin Nutrition 0.000 description 6
- 229930003231 vitamin Natural products 0.000 description 6
- 150000003722 vitamin derivatives Chemical class 0.000 description 6
- OBVSBEYOMDWLRJ-BFHQHQDPSA-N Ala-Gly-Thr Chemical compound C[C@@H](O)[C@@H](C(O)=O)NC(=O)CNC(=O)[C@H](C)N OBVSBEYOMDWLRJ-BFHQHQDPSA-N 0.000 description 5
- ALMIMUZAWTUNIO-BZSNNMDCSA-N Asp-Tyr-Tyr Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(O)=O ALMIMUZAWTUNIO-BZSNNMDCSA-N 0.000 description 5
- XSBGUANSZDGULP-IUCAKERBSA-N Gln-Gly-Lys Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)NCC(=O)N[C@@H](CCCCN)C(O)=O XSBGUANSZDGULP-IUCAKERBSA-N 0.000 description 5
- JXFLPKSDLDEOQK-JHEQGTHGSA-N Gln-Gly-Thr Chemical compound C[C@@H](O)[C@@H](C(O)=O)NC(=O)CNC(=O)[C@@H](N)CCC(N)=O JXFLPKSDLDEOQK-JHEQGTHGSA-N 0.000 description 5
- WCORRBXVISTKQL-WHFBIAKZSA-N Gly-Ser-Ser Chemical compound NCC(=O)N[C@@H](CO)C(=O)N[C@@H](CO)C(O)=O WCORRBXVISTKQL-WHFBIAKZSA-N 0.000 description 5
- FFALDIDGPLUDKV-ZDLURKLDSA-N Gly-Thr-Ser Chemical compound [H]NCC(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CO)C(O)=O FFALDIDGPLUDKV-ZDLURKLDSA-N 0.000 description 5
- 102100037852 Insulin-like growth factor I Human genes 0.000 description 5
- 241000880493 Leptailurus serval Species 0.000 description 5
- 241001465754 Metazoa Species 0.000 description 5
- LMKKMCGTDANZTR-BZSNNMDCSA-N Tyr-Phe-Asp Chemical compound C([C@H](N)C(=O)N[C@@H](CC=1C=CC=CC=1)C(=O)N[C@@H](CC(O)=O)C(O)=O)C1=CC=C(O)C=C1 LMKKMCGTDANZTR-BZSNNMDCSA-N 0.000 description 5
- NUQZCPSZHGIYTA-HKUYNNGSSA-N Tyr-Trp-Gly Chemical compound C1=CC=C2C(=C1)C(=CN2)C[C@@H](C(=O)NCC(=O)O)NC(=O)[C@H](CC3=CC=C(C=C3)O)N NUQZCPSZHGIYTA-HKUYNNGSSA-N 0.000 description 5
- HQYVQDRYODWONX-DCAQKATOSA-N Val-His-Ser Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CC1=CN=CN1)C(=O)N[C@@H](CO)C(=O)O)N HQYVQDRYODWONX-DCAQKATOSA-N 0.000 description 5
- 239000002253 acid Substances 0.000 description 5
- 230000006909 anti-apoptosis Effects 0.000 description 5
- 230000000259 anti-tumor effect Effects 0.000 description 5
- 238000002648 combination therapy Methods 0.000 description 5
- 238000011284 combination treatment Methods 0.000 description 5
- 238000005516 engineering process Methods 0.000 description 5
- 102000044162 human IGF1 Human genes 0.000 description 5
- 230000036039 immunity Effects 0.000 description 5
- 239000011049 pearl Substances 0.000 description 5
- 239000013612 plasmid Substances 0.000 description 5
- 108090000765 processed proteins & peptides Proteins 0.000 description 5
- 230000019491 signal transduction Effects 0.000 description 5
- 239000000126 substance Substances 0.000 description 5
- WSTYNZDAOAEEKG-GWJSGULQSA-N tingenone Chemical class CC1=C(O)C(=O)C=C2[C@@](CC[C@]3([C@@H]4C[C@H](C(C[C@@]4(CC[C@@]33C)C)=O)C)C)(C)C3=CC=C21 WSTYNZDAOAEEKG-GWJSGULQSA-N 0.000 description 5
- 108010077037 tyrosyl-tyrosyl-phenylalanine Proteins 0.000 description 5
- 108091032973 (ribonucleotides)n+m Proteins 0.000 description 4
- QNJIRRVTOXNGMH-GUBZILKMSA-N Asn-Gln-Lys Chemical compound NCCCC[C@@H](C(O)=O)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@@H](N)CC(N)=O QNJIRRVTOXNGMH-GUBZILKMSA-N 0.000 description 4
- 108010076667 Caspases Proteins 0.000 description 4
- 102000011727 Caspases Human genes 0.000 description 4
- 241000699802 Cricetulus griseus Species 0.000 description 4
- AOJJSUZBOXZQNB-TZSSRYMLSA-N Doxorubicin Chemical compound O([C@H]1C[C@@](O)(CC=2C(O)=C3C(=O)C=4C=CC=C(C=4C(=O)C3=C(O)C=21)OC)C(=O)CO)[C@H]1C[C@H](N)[C@H](O)[C@H](C)O1 AOJJSUZBOXZQNB-TZSSRYMLSA-N 0.000 description 4
- 102000004190 Enzymes Human genes 0.000 description 4
- 108090000790 Enzymes Proteins 0.000 description 4
- MFNUFCFRAZPJFW-JYJNAYRXSA-N Glu-Lys-Phe Chemical compound OC(=O)CC[C@H](N)C(=O)N[C@@H](CCCCN)C(=O)N[C@H](C(O)=O)CC1=CC=CC=C1 MFNUFCFRAZPJFW-JYJNAYRXSA-N 0.000 description 4
- SOYWRINXUSUWEQ-DLOVCJGASA-N Glu-Val-Val Chemical compound CC(C)[C@@H](C(O)=O)NC(=O)[C@H](C(C)C)NC(=O)[C@@H](N)CCC(O)=O SOYWRINXUSUWEQ-DLOVCJGASA-N 0.000 description 4
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 4
- FBGXMKUWQFPHFB-JBDRJPRFSA-N Ile-Ser-Cys Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CO)C(=O)N[C@@H](CS)C(=O)O)N FBGXMKUWQFPHFB-JBDRJPRFSA-N 0.000 description 4
- MVJRBCJCRYGCKV-GVXVVHGQSA-N Leu-Val-Gln Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CCC(N)=O)C(O)=O MVJRBCJCRYGCKV-GVXVVHGQSA-N 0.000 description 4
- NPBGTPKLVJEOBE-IUCAKERBSA-N Lys-Arg Chemical compound NCCCC[C@H](N)C(=O)N[C@H](C(O)=O)CCCNC(N)=N NPBGTPKLVJEOBE-IUCAKERBSA-N 0.000 description 4
- ZUGVARDEGWMMLK-SRVKXCTJSA-N Lys-Ser-Lys Chemical compound NCCCC[C@H](N)C(=O)N[C@@H](CO)C(=O)N[C@H](C(O)=O)CCCCN ZUGVARDEGWMMLK-SRVKXCTJSA-N 0.000 description 4
- BDFHWFUAQLIMJO-KXNHARMFSA-N Lys-Thr-Pro Chemical compound C[C@H]([C@@H](C(=O)N1CCC[C@@H]1C(=O)O)NC(=O)[C@H](CCCCN)N)O BDFHWFUAQLIMJO-KXNHARMFSA-N 0.000 description 4
- 241000124008 Mammalia Species 0.000 description 4
- UIMCLYYSUCIUJM-UWVGGRQHSA-N Pro-Gly-Lys Chemical compound NCCCC[C@@H](C(O)=O)NC(=O)CNC(=O)[C@@H]1CCCN1 UIMCLYYSUCIUJM-UWVGGRQHSA-N 0.000 description 4
- 229920002684 Sepharose Polymers 0.000 description 4
- BPMRXBZYPGYPJN-WHFBIAKZSA-N Ser-Gly-Asn Chemical compound [H]N[C@@H](CO)C(=O)NCC(=O)N[C@@H](CC(N)=O)C(O)=O BPMRXBZYPGYPJN-WHFBIAKZSA-N 0.000 description 4
- YMTLKLXDFCSCNX-BYPYZUCNSA-N Ser-Gly-Gly Chemical compound OC[C@H](N)C(=O)NCC(=O)NCC(O)=O YMTLKLXDFCSCNX-BYPYZUCNSA-N 0.000 description 4
- 108010090804 Streptavidin Proteins 0.000 description 4
- 229940123237 Taxane Drugs 0.000 description 4
- DGDCHPCRMWEOJR-FQPOAREZSA-N Thr-Ala-Tyr Chemical compound C[C@@H](O)[C@H](N)C(=O)N[C@@H](C)C(=O)N[C@H](C(O)=O)CC1=CC=C(O)C=C1 DGDCHPCRMWEOJR-FQPOAREZSA-N 0.000 description 4
- KKPOGALELPLJTL-MEYUZBJRSA-N Thr-Lys-Tyr Chemical compound C[C@@H](O)[C@H](N)C(=O)N[C@@H](CCCCN)C(=O)N[C@H](C(O)=O)CC1=CC=C(O)C=C1 KKPOGALELPLJTL-MEYUZBJRSA-N 0.000 description 4
- HUPLKEHTTQBXSC-YJRXYDGGSA-N Thr-Ser-Tyr Chemical compound C[C@@H](O)[C@H](N)C(=O)N[C@@H](CO)C(=O)N[C@H](C(O)=O)CC1=CC=C(O)C=C1 HUPLKEHTTQBXSC-YJRXYDGGSA-N 0.000 description 4
- IQFYYKKMVGJFEH-XLPZGREQSA-N Thymidine Chemical compound O=C1NC(=O)C(C)=CN1[C@@H]1O[C@H](CO)[C@@H](O)C1 IQFYYKKMVGJFEH-XLPZGREQSA-N 0.000 description 4
- 230000004913 activation Effects 0.000 description 4
- 238000001261 affinity purification Methods 0.000 description 4
- 108010050025 alpha-glutamyltryptophan Proteins 0.000 description 4
- 125000003277 amino group Chemical group 0.000 description 4
- 239000005557 antagonist Substances 0.000 description 4
- 238000003556 assay Methods 0.000 description 4
- 230000003305 autocrine Effects 0.000 description 4
- 230000004069 differentiation Effects 0.000 description 4
- 230000029087 digestion Effects 0.000 description 4
- 125000002228 disulfide group Chemical group 0.000 description 4
- 229940088598 enzyme Drugs 0.000 description 4
- 108010010147 glycylglutamine Proteins 0.000 description 4
- 230000006872 improvement Effects 0.000 description 4
- 238000001727 in vivo Methods 0.000 description 4
- 239000003112 inhibitor Substances 0.000 description 4
- 125000005647 linker group Chemical group 0.000 description 4
- 230000001404 mediated effect Effects 0.000 description 4
- 231100000350 mutagenesis Toxicity 0.000 description 4
- 238000011275 oncology therapy Methods 0.000 description 4
- 230000003076 paracrine Effects 0.000 description 4
- 238000001556 precipitation Methods 0.000 description 4
- 230000002829 reductive effect Effects 0.000 description 4
- 238000012216 screening Methods 0.000 description 4
- 238000002415 sodium dodecyl sulfate polyacrylamide gel electrophoresis Methods 0.000 description 4
- 239000002904 solvent Substances 0.000 description 4
- 238000001890 transfection Methods 0.000 description 4
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 4
- JKMHFZQWWAIEOD-UHFFFAOYSA-N 2-[4-(2-hydroxyethyl)piperazin-1-yl]ethanesulfonic acid Chemical compound OCC[NH+]1CCN(CCS([O-])(=O)=O)CC1 JKMHFZQWWAIEOD-UHFFFAOYSA-N 0.000 description 3
- 101100230376 Acetivibrio thermocellus (strain ATCC 27405 / DSM 1237 / JCM 9322 / NBRC 103400 / NCIMB 10682 / NRRL B-4536 / VPI 7372) celI gene Proteins 0.000 description 3
- WYPUMLRSQMKIJU-BPNCWPANSA-N Ala-Arg-Tyr Chemical compound [H]N[C@@H](C)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(O)=O WYPUMLRSQMKIJU-BPNCWPANSA-N 0.000 description 3
- WBDWQKRLTVCDSY-WHFBIAKZSA-N Asp-Gly-Asp Chemical compound OC(=O)C[C@H](N)C(=O)NCC(=O)N[C@@H](CC(O)=O)C(O)=O WBDWQKRLTVCDSY-WHFBIAKZSA-N 0.000 description 3
- 102000000844 Cell Surface Receptors Human genes 0.000 description 3
- 108010001857 Cell Surface Receptors Proteins 0.000 description 3
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 3
- WSFSSNUMVMOOMR-UHFFFAOYSA-N Formaldehyde Chemical compound O=C WSFSSNUMVMOOMR-UHFFFAOYSA-N 0.000 description 3
- 230000035519 G0 Phase Effects 0.000 description 3
- 230000010190 G1 phase Effects 0.000 description 3
- XZLLTYBONVKGLO-SDDRHHMPSA-N Gln-Lys-Pro Chemical compound C1C[C@@H](N(C1)C(=O)[C@H](CCCCN)NC(=O)[C@H](CCC(=O)N)N)C(=O)O XZLLTYBONVKGLO-SDDRHHMPSA-N 0.000 description 3
- WPLGNDORMXTMQS-FXQIFTODSA-N Glu-Gln-Ser Chemical compound OC(=O)CC[C@H](N)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CO)C(O)=O WPLGNDORMXTMQS-FXQIFTODSA-N 0.000 description 3
- JVZLZVJTIXVIHK-SXNHZJKMSA-N Glu-Trp-Ile Chemical compound CC[C@H](C)[C@@H](C(=O)O)NC(=O)[C@H](CC1=CNC2=CC=CC=C21)NC(=O)[C@H](CCC(=O)O)N JVZLZVJTIXVIHK-SXNHZJKMSA-N 0.000 description 3
- RJIVPOXLQFJRTG-LURJTMIESA-N Gly-Arg-Gly Chemical compound OC(=O)CNC(=O)[C@@H](NC(=O)CN)CCCN=C(N)N RJIVPOXLQFJRTG-LURJTMIESA-N 0.000 description 3
- OCQUNKSFDYDXBG-QXEWZRGKSA-N Gly-Arg-Ile Chemical compound CC[C@H](C)[C@@H](C(O)=O)NC(=O)[C@@H](NC(=O)CN)CCCN=C(N)N OCQUNKSFDYDXBG-QXEWZRGKSA-N 0.000 description 3
- VXKCPBPQEKKERH-IUCAKERBSA-N Gly-Arg-Pro Chemical compound NC(N)=NCCC[C@H](NC(=O)CN)C(=O)N1CCC[C@H]1C(O)=O VXKCPBPQEKKERH-IUCAKERBSA-N 0.000 description 3
- GNPVTZJUUBPZKW-WDSKDSINSA-N Gly-Gln-Ser Chemical compound [H]NCC(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CO)C(O)=O GNPVTZJUUBPZKW-WDSKDSINSA-N 0.000 description 3
- TVTZEOHWHUVYCG-KYNKHSRBSA-N Gly-Thr-Thr Chemical compound [H]NCC(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H]([C@@H](C)O)C(O)=O TVTZEOHWHUVYCG-KYNKHSRBSA-N 0.000 description 3
- YGHSQRJSHKYUJY-SCZZXKLOSA-N Gly-Val-Pro Chemical compound CC(C)[C@@H](C(=O)N1CCC[C@@H]1C(=O)O)NC(=O)CN YGHSQRJSHKYUJY-SCZZXKLOSA-N 0.000 description 3
- 239000007995 HEPES buffer Substances 0.000 description 3
- 108010054477 Immunoglobulin Fab Fragments Proteins 0.000 description 3
- 102000001706 Immunoglobulin Fab Fragments Human genes 0.000 description 3
- 239000005411 L01XE02 - Gefitinib Substances 0.000 description 3
- YTJFXEDRUOQGSP-DCAQKATOSA-N Lys-Pro-Ser Chemical compound [H]N[C@@H](CCCCN)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CO)C(O)=O YTJFXEDRUOQGSP-DCAQKATOSA-N 0.000 description 3
- 101150018665 MAPK3 gene Proteins 0.000 description 3
- 241000283973 Oryctolagus cuniculus Species 0.000 description 3
- LLGTYVHITPVGKR-RYUDHWBXSA-N Phe-Gln-Gly Chemical compound [H]N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CCC(N)=O)C(=O)NCC(O)=O LLGTYVHITPVGKR-RYUDHWBXSA-N 0.000 description 3
- BIYWZVCPZIFGPY-QWRGUYRKSA-N Phe-Gly-Ser Chemical compound [H]N[C@@H](CC1=CC=CC=C1)C(=O)NCC(=O)N[C@@H](CO)C(O)=O BIYWZVCPZIFGPY-QWRGUYRKSA-N 0.000 description 3
- SPLBRAKYXGOFSO-UNQGMJICSA-N Pro-Phe-Thr Chemical compound C[C@H]([C@@H](C(=O)O)NC(=O)[C@H](CC1=CC=CC=C1)NC(=O)[C@@H]2CCCN2)O SPLBRAKYXGOFSO-UNQGMJICSA-N 0.000 description 3
- QKWYXRPICJEQAJ-KJEVXHAQSA-N Pro-Tyr-Thr Chemical compound C[C@H]([C@@H](C(=O)O)NC(=O)[C@H](CC1=CC=C(C=C1)O)NC(=O)[C@@H]2CCCN2)O QKWYXRPICJEQAJ-KJEVXHAQSA-N 0.000 description 3
- 239000006146 Roswell Park Memorial Institute medium Substances 0.000 description 3
- WPSKTVVMQCXPRO-BWBBJGPYSA-N Thr-Ser-Ser Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CO)C(=O)N[C@@H](CO)C(O)=O WPSKTVVMQCXPRO-BWBBJGPYSA-N 0.000 description 3
- LDKDSFQSEUOCOO-RPTUDFQQSA-N Tyr-Thr-Phe Chemical compound [H]N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC1=CC=CC=C1)C(O)=O LDKDSFQSEUOCOO-RPTUDFQQSA-N 0.000 description 3
- ANHVRCNNGJMJNG-BZSNNMDCSA-N Tyr-Tyr-Cys Chemical compound C1=CC(=CC=C1C[C@@H](C(=O)N[C@@H](CC2=CC=C(C=C2)O)C(=O)N[C@@H](CS)C(=O)O)N)O ANHVRCNNGJMJNG-BZSNNMDCSA-N 0.000 description 3
- 230000003187 abdominal effect Effects 0.000 description 3
- 238000002835 absorbance Methods 0.000 description 3
- 230000009824 affinity maturation Effects 0.000 description 3
- 108010005233 alanylglutamic acid Proteins 0.000 description 3
- 125000000539 amino acid group Chemical group 0.000 description 3
- 230000000692 anti-sense effect Effects 0.000 description 3
- 239000004202 carbamide Substances 0.000 description 3
- 238000012219 cassette mutagenesis Methods 0.000 description 3
- 230000022131 cell cycle Effects 0.000 description 3
- 230000003915 cell function Effects 0.000 description 3
- 239000006285 cell suspension Substances 0.000 description 3
- 238000010367 cloning Methods 0.000 description 3
- 230000008878 coupling Effects 0.000 description 3
- 238000010168 coupling process Methods 0.000 description 3
- 238000005859 coupling reaction Methods 0.000 description 3
- 239000012228 culture supernatant Substances 0.000 description 3
- 229940079593 drug Drugs 0.000 description 3
- 210000004602 germ cell Anatomy 0.000 description 3
- XBGGUPMXALFZOT-UHFFFAOYSA-N glycyl-L-tyrosine hemihydrate Natural products NCC(=O)NC(C(O)=O)CC1=CC=C(O)C=C1 XBGGUPMXALFZOT-UHFFFAOYSA-N 0.000 description 3
- 108010033719 glycyl-histidyl-glycine Proteins 0.000 description 3
- 239000001963 growth medium Substances 0.000 description 3
- 230000002163 immunogen Effects 0.000 description 3
- 238000001114 immunoprecipitation Methods 0.000 description 3
- 238000010253 intravenous injection Methods 0.000 description 3
- 108010044374 isoleucyl-tyrosine Proteins 0.000 description 3
- 108010057821 leucylproline Proteins 0.000 description 3
- 108010057952 lysyl-phenylalanyl-lysine Proteins 0.000 description 3
- 210000001161 mammalian embryo Anatomy 0.000 description 3
- 238000004519 manufacturing process Methods 0.000 description 3
- 108010022588 methionyl-lysyl-proline Proteins 0.000 description 3
- 230000004048 modification Effects 0.000 description 3
- 238000012986 modification Methods 0.000 description 3
- 208000002154 non-small cell lung carcinoma Diseases 0.000 description 3
- 108020004707 nucleic acids Proteins 0.000 description 3
- 102000039446 nucleic acids Human genes 0.000 description 3
- 150000007523 nucleic acids Chemical class 0.000 description 3
- 239000002773 nucleotide Substances 0.000 description 3
- 125000003729 nucleotide group Chemical group 0.000 description 3
- 210000001672 ovary Anatomy 0.000 description 3
- 238000002823 phage display Methods 0.000 description 3
- 230000035479 physiological effects, processes and functions Effects 0.000 description 3
- 102000004196 processed proteins & peptides Human genes 0.000 description 3
- 239000011780 sodium chloride Substances 0.000 description 3
- 210000004988 splenocyte Anatomy 0.000 description 3
- DKPFODGZWDEEBT-QFIAKTPHSA-N taxane Chemical class C([C@]1(C)CCC[C@@H](C)[C@H]1C1)C[C@H]2[C@H](C)CC[C@@H]1C2(C)C DKPFODGZWDEEBT-QFIAKTPHSA-N 0.000 description 3
- 108010038745 tryptophylglycine Proteins 0.000 description 3
- 208000029729 tumor suppressor gene on chromosome 11 Diseases 0.000 description 3
- 239000011735 vitamin B7 Substances 0.000 description 3
- QWPXBEHQFHACTK-KZVYIGENSA-N (10e,12e)-86-chloro-12,14,4-trihydroxy-85,14-dimethoxy-33,2,7,10-tetramethyl-15,16-dihydro-14h-7-aza-1(6,4)-oxazina-3(2,3)-oxirana-8(1,3)-benzenacyclotetradecaphane-10,12-dien-6-one Chemical compound CN1C(=O)CC(O)C2(C)OC2C(C)C(OC(=O)N2)CC2(O)C(OC)\C=C\C=C(C)\CC2=CC(OC)=C(Cl)C1=C2 QWPXBEHQFHACTK-KZVYIGENSA-N 0.000 description 2
- WXPZDDCNKXMOMC-AVGNSLFASA-N (2s)-1-[(2s)-2-[[(2s)-1-(2-aminoacetyl)pyrrolidine-2-carbonyl]amino]-5-(diaminomethylideneamino)pentanoyl]pyrrolidine-2-carboxylic acid Chemical compound NCC(=O)N1CCC[C@H]1C(=O)N[C@@H](CCCNC(N)=N)C(=O)N1[C@H](C(O)=O)CCC1 WXPZDDCNKXMOMC-AVGNSLFASA-N 0.000 description 2
- 206010069754 Acquired gene mutation Diseases 0.000 description 2
- KXEVYGKATAMXJJ-ACZMJKKPSA-N Ala-Glu-Asp Chemical compound C[C@H](N)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(O)=O)C(O)=O KXEVYGKATAMXJJ-ACZMJKKPSA-N 0.000 description 2
- REAQAWSENITKJL-DDWPSWQVSA-N Ala-Met-Asp-Tyr Chemical compound [H]N[C@@H](C)C(=O)N[C@@H](CCSC)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(O)=O REAQAWSENITKJL-DDWPSWQVSA-N 0.000 description 2
- RTZCUEHYUQZIDE-WHFBIAKZSA-N Ala-Ser-Gly Chemical compound C[C@H](N)C(=O)N[C@@H](CO)C(=O)NCC(O)=O RTZCUEHYUQZIDE-WHFBIAKZSA-N 0.000 description 2
- LSMDIAAALJJLRO-XQXXSGGOSA-N Ala-Thr-Glu Chemical compound [H]N[C@@H](C)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CCC(O)=O)C(O)=O LSMDIAAALJJLRO-XQXXSGGOSA-N 0.000 description 2
- JJHBEVZAZXZREW-LFSVMHDDSA-N Ala-Thr-Phe Chemical compound C[C@@H](O)[C@H](NC(=O)[C@H](C)N)C(=O)N[C@@H](Cc1ccccc1)C(O)=O JJHBEVZAZXZREW-LFSVMHDDSA-N 0.000 description 2
- XCIGOVDXZULBBV-DCAQKATOSA-N Ala-Val-Lys Chemical compound CC(C)[C@H](NC(=O)[C@H](C)N)C(=O)N[C@@H](CCCCN)C(O)=O XCIGOVDXZULBBV-DCAQKATOSA-N 0.000 description 2
- 108010088751 Albumins Proteins 0.000 description 2
- NLXLAEXVIDQMFP-UHFFFAOYSA-N Ammonia chloride Chemical compound [NH4+].[Cl-] NLXLAEXVIDQMFP-UHFFFAOYSA-N 0.000 description 2
- GXXWTNKNFFKTJB-NAKRPEOUSA-N Arg-Ile-Ser Chemical compound [H]N[C@@H](CCCNC(N)=N)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CO)C(O)=O GXXWTNKNFFKTJB-NAKRPEOUSA-N 0.000 description 2
- HGKHPCFTRQDHCU-IUCAKERBSA-N Arg-Pro-Gly Chemical compound NC(N)=NCCC[C@H](N)C(=O)N1CCC[C@H]1C(=O)NCC(O)=O HGKHPCFTRQDHCU-IUCAKERBSA-N 0.000 description 2
- ULBHWNVWSCJLCO-NHCYSSNCSA-N Arg-Val-Glu Chemical compound OC(=O)CC[C@@H](C(O)=O)NC(=O)[C@H](C(C)C)NC(=O)[C@@H](N)CCCN=C(N)N ULBHWNVWSCJLCO-NHCYSSNCSA-N 0.000 description 2
- WLVLIYYBPPONRJ-GCJQMDKQSA-N Asn-Thr-Ala Chemical compound [H]N[C@@H](CC(N)=O)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](C)C(O)=O WLVLIYYBPPONRJ-GCJQMDKQSA-N 0.000 description 2
- BWJZSLQJNBSUPM-FXQIFTODSA-N Asp-Pro-Asn Chemical compound OC(=O)C[C@H](N)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CC(N)=O)C(O)=O BWJZSLQJNBSUPM-FXQIFTODSA-N 0.000 description 2
- ZBYLEBZCVKLPCY-FXQIFTODSA-N Asp-Ser-Arg Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](CO)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O ZBYLEBZCVKLPCY-FXQIFTODSA-N 0.000 description 2
- HTSSXFASOUSJQG-IHPCNDPISA-N Asp-Tyr-Trp Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CC1=CNC2=C1C=CC=C2)C(O)=O HTSSXFASOUSJQG-IHPCNDPISA-N 0.000 description 2
- 238000012935 Averaging Methods 0.000 description 2
- DWRXFEITVBNRMK-UHFFFAOYSA-N Beta-D-1-Arabinofuranosylthymine Natural products O=C1NC(=O)C(C)=CN1C1C(O)C(O)C(CO)O1 DWRXFEITVBNRMK-UHFFFAOYSA-N 0.000 description 2
- 101100218515 Candida albicans (strain SC5314 / ATCC MYA-2876) BBP gene Proteins 0.000 description 2
- BVKZGUZCCUSVTD-UHFFFAOYSA-L Carbonate Chemical compound [O-]C([O-])=O BVKZGUZCCUSVTD-UHFFFAOYSA-L 0.000 description 2
- IGXWBGJHJZYPQS-SSDOTTSWSA-N D-Luciferin Chemical compound OC(=O)[C@H]1CSC(C=2SC3=CC=C(O)C=C3N=2)=N1 IGXWBGJHJZYPQS-SSDOTTSWSA-N 0.000 description 2
- CYCGRDQQIOGCKX-UHFFFAOYSA-N Dehydro-luciferin Natural products OC(=O)C1=CSC(C=2SC3=CC(O)=CC=C3N=2)=N1 CYCGRDQQIOGCKX-UHFFFAOYSA-N 0.000 description 2
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 description 2
- BJGNCJDXODQBOB-UHFFFAOYSA-N Fivefly Luciferin Natural products OC(=O)C1CSC(C=2SC3=CC(O)=CC=C3N=2)=N1 BJGNCJDXODQBOB-UHFFFAOYSA-N 0.000 description 2
- PKVWNYGXMNWJSI-CIUDSAMLSA-N Gln-Gln-Gln Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCC(N)=O)C(O)=O PKVWNYGXMNWJSI-CIUDSAMLSA-N 0.000 description 2
- MADFVRSKEIEZHZ-DCAQKATOSA-N Gln-Gln-Lys Chemical compound C(CCN)C[C@@H](C(=O)O)NC(=O)[C@H](CCC(=O)N)NC(=O)[C@H](CCC(=O)N)N MADFVRSKEIEZHZ-DCAQKATOSA-N 0.000 description 2
- UFNSPPFJOHNXRE-AUTRQRHGSA-N Gln-Gln-Val Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](C(C)C)C(O)=O UFNSPPFJOHNXRE-AUTRQRHGSA-N 0.000 description 2
- GQZDDFRXSDGUNG-YVNDNENWSA-N Gln-Ile-Gln Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CCC(N)=O)C(O)=O GQZDDFRXSDGUNG-YVNDNENWSA-N 0.000 description 2
- ZZLDMBMFKZFQMU-NRPADANISA-N Gln-Val-Ala Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](C)C(O)=O ZZLDMBMFKZFQMU-NRPADANISA-N 0.000 description 2
- CXRWMMRLEMVSEH-PEFMBERDSA-N Glu-Ile-Asn Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CC(N)=O)C(O)=O CXRWMMRLEMVSEH-PEFMBERDSA-N 0.000 description 2
- XTZDZAXYPDISRR-MNXVOIDGSA-N Glu-Ile-Lys Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CCCCN)C(=O)O)NC(=O)[C@H](CCC(=O)O)N XTZDZAXYPDISRR-MNXVOIDGSA-N 0.000 description 2
- DWBBKNPKDHXIAC-SRVKXCTJSA-N Glu-Leu-Met Chemical compound CSCC[C@@H](C(O)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@@H](N)CCC(O)=O DWBBKNPKDHXIAC-SRVKXCTJSA-N 0.000 description 2
- QXUPRMQJDWJDFR-NRPADANISA-N Glu-Val-Ser Chemical compound CC(C)[C@H](NC(=O)[C@@H](N)CCC(O)=O)C(=O)N[C@@H](CO)C(O)=O QXUPRMQJDWJDFR-NRPADANISA-N 0.000 description 2
- DTPOVRRYXPJJAZ-FJXKBIBVSA-N Gly-Arg-Thr Chemical compound C[C@@H](O)[C@@H](C(O)=O)NC(=O)[C@@H](NC(=O)CN)CCCN=C(N)N DTPOVRRYXPJJAZ-FJXKBIBVSA-N 0.000 description 2
- DTRUBYPMMVPQPD-YUMQZZPRSA-N Gly-Gln-Arg Chemical compound [H]NCC(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O DTRUBYPMMVPQPD-YUMQZZPRSA-N 0.000 description 2
- XLFHCWHXKSFVIB-BQBZGAKWSA-N Gly-Gln-Gln Chemical compound NCC(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCC(N)=O)C(O)=O XLFHCWHXKSFVIB-BQBZGAKWSA-N 0.000 description 2
- ADZGCWWDPFDHCY-ZETCQYMHSA-N Gly-His-Gly Chemical compound OC(=O)CNC(=O)[C@@H](NC(=O)CN)CC1=CN=CN1 ADZGCWWDPFDHCY-ZETCQYMHSA-N 0.000 description 2
- VEPBEGNDJYANCF-QWRGUYRKSA-N Gly-Lys-Lys Chemical compound NCCCC[C@@H](C(O)=O)NC(=O)[C@@H](NC(=O)CN)CCCCN VEPBEGNDJYANCF-QWRGUYRKSA-N 0.000 description 2
- WTUSRDZLLWGYAT-KCTSRDHCSA-N Gly-Trp-Ile Chemical compound CC[C@H](C)[C@@H](C(=O)O)NC(=O)[C@H](CC1=CNC2=CC=CC=C21)NC(=O)CN WTUSRDZLLWGYAT-KCTSRDHCSA-N 0.000 description 2
- FFKJUTZARGRVTH-KKUMJFAQSA-N His-Ser-Tyr Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)N[C@@H](CO)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(O)=O FFKJUTZARGRVTH-KKUMJFAQSA-N 0.000 description 2
- IOVUXUSIGXCREV-DKIMLUQUSA-N Ile-Leu-Phe Chemical compound CC[C@H](C)[C@H](N)C(=O)N[C@@H](CC(C)C)C(=O)N[C@H](C(O)=O)CC1=CC=CC=C1 IOVUXUSIGXCREV-DKIMLUQUSA-N 0.000 description 2
- MLSUZXHSNRBDCI-CYDGBPFRSA-N Ile-Pro-Val Chemical compound CC[C@H](C)[C@@H](C(=O)N1CCC[C@H]1C(=O)N[C@@H](C(C)C)C(=O)O)N MLSUZXHSNRBDCI-CYDGBPFRSA-N 0.000 description 2
- 102000004877 Insulin Human genes 0.000 description 2
- 108090001061 Insulin Proteins 0.000 description 2
- XEEYBQQBJWHFJM-UHFFFAOYSA-N Iron Chemical compound [Fe] XEEYBQQBJWHFJM-UHFFFAOYSA-N 0.000 description 2
- XUJNEKJLAYXESH-REOHCLBHSA-N L-Cysteine Chemical compound SC[C@H](N)C(O)=O XUJNEKJLAYXESH-REOHCLBHSA-N 0.000 description 2
- HGCNKOLVKRAVHD-UHFFFAOYSA-N L-Met-L-Phe Natural products CSCCC(N)C(=O)NC(C(O)=O)CC1=CC=CC=C1 HGCNKOLVKRAVHD-UHFFFAOYSA-N 0.000 description 2
- FBOZXECLQNJBKD-ZDUSSCGKSA-N L-methotrexate Chemical compound C=1N=C2N=C(N)N=C(N)C2=NC=1CN(C)C1=CC=C(C(=O)N[C@@H](CCC(O)=O)C(O)=O)C=C1 FBOZXECLQNJBKD-ZDUSSCGKSA-N 0.000 description 2
- CCQLQKZTXZBXTN-NHCYSSNCSA-N Leu-Gly-Ile Chemical compound [H]N[C@@H](CC(C)C)C(=O)NCC(=O)N[C@@H]([C@@H](C)CC)C(O)=O CCQLQKZTXZBXTN-NHCYSSNCSA-N 0.000 description 2
- UCBPDSYUVAAHCD-UWVGGRQHSA-N Leu-Pro-Gly Chemical compound CC(C)C[C@H](N)C(=O)N1CCC[C@H]1C(=O)NCC(O)=O UCBPDSYUVAAHCD-UWVGGRQHSA-N 0.000 description 2
- AIQWYVFNBNNOLU-RHYQMDGZSA-N Leu-Thr-Val Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](C(C)C)C(O)=O AIQWYVFNBNNOLU-RHYQMDGZSA-N 0.000 description 2
- 108010000817 Leuprolide Proteins 0.000 description 2
- DDWFXDSYGUXRAY-UHFFFAOYSA-N Luciferin Natural products CCc1c(C)c(CC2NC(=O)C(=C2C=C)C)[nH]c1Cc3[nH]c4C(=C5/NC(CC(=O)O)C(C)C5CC(=O)O)CC(=O)c4c3C DDWFXDSYGUXRAY-UHFFFAOYSA-N 0.000 description 2
- KNKHAVVBVXKOGX-JXUBOQSCSA-N Lys-Ala-Thr Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H](C)C(=O)N[C@@H]([C@@H](C)O)C(O)=O KNKHAVVBVXKOGX-JXUBOQSCSA-N 0.000 description 2
- GPJGFSFYBJGYRX-YUMQZZPRSA-N Lys-Gly-Asp Chemical compound NCCCC[C@H](N)C(=O)NCC(=O)N[C@H](C(O)=O)CC(O)=O GPJGFSFYBJGYRX-YUMQZZPRSA-N 0.000 description 2
- NROQVSYLPRLJIP-PMVMPFDFSA-N Lys-Trp-Tyr Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H](CC1=CNC2=C1C=CC=C2)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(O)=O NROQVSYLPRLJIP-PMVMPFDFSA-N 0.000 description 2
- 108091054455 MAP kinase family Proteins 0.000 description 2
- 102000043136 MAP kinase family Human genes 0.000 description 2
- 101150116844 MSL5 gene Proteins 0.000 description 2
- MHQXIBRPDKXDGZ-ZFWWWQNUSA-N Met-Gly-Trp Chemical compound C1=CC=C2C(C[C@H](NC(=O)CNC(=O)[C@@H](N)CCSC)C(O)=O)=CNC2=C1 MHQXIBRPDKXDGZ-ZFWWWQNUSA-N 0.000 description 2
- HSJIGJRZYUADSS-IHRRRGAJSA-N Met-Lys-Leu Chemical compound [H]N[C@@H](CCSC)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC(C)C)C(O)=O HSJIGJRZYUADSS-IHRRRGAJSA-N 0.000 description 2
- NWIBSHFKIJFRCO-WUDYKRTCSA-N Mytomycin Chemical compound C1N2C(C(C(C)=C(N)C3=O)=O)=C3[C@@H](COC(N)=O)[C@@]2(OC)[C@@H]2[C@H]1N2 NWIBSHFKIJFRCO-WUDYKRTCSA-N 0.000 description 2
- YBAFDPFAUTYYRW-UHFFFAOYSA-N N-L-alpha-glutamyl-L-leucine Natural products CC(C)CC(C(O)=O)NC(=O)C(N)CCC(O)=O YBAFDPFAUTYYRW-UHFFFAOYSA-N 0.000 description 2
- 108091007491 NSP3 Papain-like protease domains Proteins 0.000 description 2
- 108091028043 Nucleic acid sequence Proteins 0.000 description 2
- 108091034117 Oligonucleotide Proteins 0.000 description 2
- 108091007960 PI3Ks Proteins 0.000 description 2
- 229930182555 Penicillin Natural products 0.000 description 2
- JGSARLDLIJGVTE-MBNYWOFBSA-N Penicillin G Chemical compound N([C@H]1[C@H]2SC([C@@H](N2C1=O)C(O)=O)(C)C)C(=O)CC1=CC=CC=C1 JGSARLDLIJGVTE-MBNYWOFBSA-N 0.000 description 2
- 102000007079 Peptide Fragments Human genes 0.000 description 2
- 108010033276 Peptide Fragments Proteins 0.000 description 2
- FRPVPGRXUKFEQE-YDHLFZDLSA-N Phe-Asp-Val Chemical compound [H]N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](C(C)C)C(O)=O FRPVPGRXUKFEQE-YDHLFZDLSA-N 0.000 description 2
- MCIXMYKSPQUMJG-SRVKXCTJSA-N Phe-Ser-Ser Chemical compound [H]N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CO)C(=O)N[C@@H](CO)C(O)=O MCIXMYKSPQUMJG-SRVKXCTJSA-N 0.000 description 2
- RAGOJJCBGXARPO-XVSYOHENSA-N Phe-Thr-Asp Chemical compound OC(=O)C[C@@H](C(O)=O)NC(=O)[C@H]([C@H](O)C)NC(=O)[C@@H](N)CC1=CC=CC=C1 RAGOJJCBGXARPO-XVSYOHENSA-N 0.000 description 2
- YCEWAVIRWNGGSS-NQCBNZPSSA-N Phe-Trp-Ile Chemical compound C([C@H](N)C(=O)N[C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O)C1=CC=CC=C1 YCEWAVIRWNGGSS-NQCBNZPSSA-N 0.000 description 2
- ZSKJPKFTPQCPIH-RCWTZXSCSA-N Pro-Arg-Thr Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H]([C@@H](C)O)C(O)=O ZSKJPKFTPQCPIH-RCWTZXSCSA-N 0.000 description 2
- HJSCRFZVGXAGNG-SRVKXCTJSA-N Pro-Gln-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@H](CCC(N)=O)NC(=O)[C@@H]1CCCN1 HJSCRFZVGXAGNG-SRVKXCTJSA-N 0.000 description 2
- DXTOOBDIIAJZBJ-BQBZGAKWSA-N Pro-Gly-Ser Chemical compound [H]N1CCC[C@H]1C(=O)NCC(=O)N[C@@H](CO)C(O)=O DXTOOBDIIAJZBJ-BQBZGAKWSA-N 0.000 description 2
- OWQXAJQZLWHPBH-FXQIFTODSA-N Pro-Ser-Asn Chemical compound [H]N1CCC[C@H]1C(=O)N[C@@H](CO)C(=O)N[C@@H](CC(N)=O)C(O)=O OWQXAJQZLWHPBH-FXQIFTODSA-N 0.000 description 2
- WWXNZNWZNZPDIF-SRVKXCTJSA-N Pro-Val-Arg Chemical compound NC(N)=NCCC[C@@H](C(O)=O)NC(=O)[C@H](C(C)C)NC(=O)[C@@H]1CCCN1 WWXNZNWZNZPDIF-SRVKXCTJSA-N 0.000 description 2
- 102000004022 Protein-Tyrosine Kinases Human genes 0.000 description 2
- 108090000412 Protein-Tyrosine Kinases Proteins 0.000 description 2
- 239000012980 RPMI-1640 medium Substances 0.000 description 2
- 230000018199 S phase Effects 0.000 description 2
- KMWFXJCGRXBQAC-CIUDSAMLSA-N Ser-Cys-Lys Chemical compound C(CCN)C[C@@H](C(=O)O)NC(=O)[C@H](CS)NC(=O)[C@H](CO)N KMWFXJCGRXBQAC-CIUDSAMLSA-N 0.000 description 2
- YPUSXTWURJANKF-KBIXCLLPSA-N Ser-Gln-Ile Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O YPUSXTWURJANKF-KBIXCLLPSA-N 0.000 description 2
- KJMOINFQVCCSDX-XKBZYTNZSA-N Ser-Gln-Thr Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H]([C@@H](C)O)C(O)=O KJMOINFQVCCSDX-XKBZYTNZSA-N 0.000 description 2
- KDGARKCAKHBEDB-NKWVEPMBSA-N Ser-Gly-Pro Chemical compound C1C[C@@H](N(C1)C(=O)CNC(=O)[C@H](CO)N)C(=O)O KDGARKCAKHBEDB-NKWVEPMBSA-N 0.000 description 2
- XNCUYZKGQOCOQH-YUMQZZPRSA-N Ser-Leu-Gly Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CC(C)C)C(=O)NCC(O)=O XNCUYZKGQOCOQH-YUMQZZPRSA-N 0.000 description 2
- GYDFRTRSSXOZCR-ACZMJKKPSA-N Ser-Ser-Glu Chemical compound OC[C@H](N)C(=O)N[C@@H](CO)C(=O)N[C@H](C(O)=O)CCC(O)=O GYDFRTRSSXOZCR-ACZMJKKPSA-N 0.000 description 2
- UBTNVMGPMYDYIU-HJPIBITLSA-N Ser-Tyr-Ile Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H]([C@@H](C)CC)C(O)=O UBTNVMGPMYDYIU-HJPIBITLSA-N 0.000 description 2
- CDBYLPFSWZWCQE-UHFFFAOYSA-L Sodium Carbonate Chemical compound [Na+].[Na+].[O-]C([O-])=O CDBYLPFSWZWCQE-UHFFFAOYSA-L 0.000 description 2
- UIIMBOGNXHQVGW-UHFFFAOYSA-M Sodium bicarbonate Chemical compound [Na+].OC([O-])=O UIIMBOGNXHQVGW-UHFFFAOYSA-M 0.000 description 2
- 239000004098 Tetracycline Substances 0.000 description 2
- NJEMRSFGDNECGF-GCJQMDKQSA-N Thr-Ala-Asp Chemical compound C[C@@H](O)[C@H](N)C(=O)N[C@@H](C)C(=O)N[C@H](C(O)=O)CC(O)=O NJEMRSFGDNECGF-GCJQMDKQSA-N 0.000 description 2
- YDWLCDQXLCILCZ-BWAGICSOSA-N Thr-His-Tyr Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC1=CNC=N1)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(O)=O YDWLCDQXLCILCZ-BWAGICSOSA-N 0.000 description 2
- MXDOAJQRJBMGMO-FJXKBIBVSA-N Thr-Pro-Gly Chemical compound C[C@@H](O)[C@H](N)C(=O)N1CCC[C@H]1C(=O)NCC(O)=O MXDOAJQRJBMGMO-FJXKBIBVSA-N 0.000 description 2
- 239000007983 Tris buffer Substances 0.000 description 2
- SVGAWGVHFIYAEE-JSGCOSHPSA-N Trp-Gly-Gln Chemical compound C1=CC=C2C(C[C@H](N)C(=O)NCC(=O)N[C@@H](CCC(N)=O)C(O)=O)=CNC2=C1 SVGAWGVHFIYAEE-JSGCOSHPSA-N 0.000 description 2
- YTCNLMSUXPCFBW-SXNHZJKMSA-N Trp-Ile-Glu Chemical compound [H]N[C@@H](CC1=CNC2=C1C=CC=C2)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CCC(O)=O)C(O)=O YTCNLMSUXPCFBW-SXNHZJKMSA-N 0.000 description 2
- PHKQVWWHRYUCJL-HJOGWXRNSA-N Tyr-Phe-Tyr Chemical compound [H]N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(O)=O PHKQVWWHRYUCJL-HJOGWXRNSA-N 0.000 description 2
- TYGHOWWWMTWVKM-HJOGWXRNSA-N Tyr-Tyr-Phe Chemical compound C([C@H](N)C(=O)N[C@@H](CC=1C=CC(O)=CC=1)C(=O)N[C@@H](CC=1C=CC=CC=1)C(O)=O)C1=CC=C(O)C=C1 TYGHOWWWMTWVKM-HJOGWXRNSA-N 0.000 description 2
- RMRFSFXLFWWAJZ-HJOGWXRNSA-N Tyr-Tyr-Tyr Chemical compound C([C@H](N)C(=O)N[C@@H](CC=1C=CC(O)=CC=1)C(=O)N[C@@H](CC=1C=CC(O)=CC=1)C(O)=O)C1=CC=C(O)C=C1 RMRFSFXLFWWAJZ-HJOGWXRNSA-N 0.000 description 2
- 108010064997 VPY tripeptide Proteins 0.000 description 2
- SLLKXDSRVAOREO-KZVJFYERSA-N Val-Ala-Thr Chemical compound C[C@H]([C@@H](C(=O)O)NC(=O)[C@H](C)NC(=O)[C@H](C(C)C)N)O SLLKXDSRVAOREO-KZVJFYERSA-N 0.000 description 2
- CVUDMNSZAIZFAE-UHFFFAOYSA-N Val-Arg-Pro Natural products NC(N)=NCCCC(NC(=O)C(N)C(C)C)C(=O)N1CCCC1C(O)=O CVUDMNSZAIZFAE-UHFFFAOYSA-N 0.000 description 2
- BZOSBRIDWSSTFN-AVGNSLFASA-N Val-Leu-Met Chemical compound CC(C)C[C@@H](C(=O)N[C@@H](CCSC)C(=O)O)NC(=O)[C@H](C(C)C)N BZOSBRIDWSSTFN-AVGNSLFASA-N 0.000 description 2
- CXWJFWAZIVWBOS-XQQFMLRXSA-N Val-Lys-Pro Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CCCCN)C(=O)N1CCC[C@@H]1C(=O)O)N CXWJFWAZIVWBOS-XQQFMLRXSA-N 0.000 description 2
- PGBMPFKFKXYROZ-UFYCRDLUSA-N Val-Tyr-Phe Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CC1=CC=C(C=C1)O)C(=O)N[C@@H](CC2=CC=CC=C2)C(=O)O)N PGBMPFKFKXYROZ-UFYCRDLUSA-N 0.000 description 2
- 229940009456 adriamycin Drugs 0.000 description 2
- 238000011091 antibody purification Methods 0.000 description 2
- 239000002246 antineoplastic agent Substances 0.000 description 2
- 108010013835 arginine glutamate Proteins 0.000 description 2
- 108010089975 arginyl-glycyl-aspartyl-serine Proteins 0.000 description 2
- 108010084758 arginyl-tyrosyl-aspartic acid Proteins 0.000 description 2
- 108010060035 arginylproline Proteins 0.000 description 2
- 108010077245 asparaginyl-proline Proteins 0.000 description 2
- 210000003719 b-lymphocyte Anatomy 0.000 description 2
- IQFYYKKMVGJFEH-UHFFFAOYSA-N beta-L-thymidine Natural products O=C1NC(=O)C(C)=CN1C1OC(CO)C(O)C1 IQFYYKKMVGJFEH-UHFFFAOYSA-N 0.000 description 2
- 230000004663 cell proliferation Effects 0.000 description 2
- 239000013599 cloning vector Substances 0.000 description 2
- 239000012141 concentrate Substances 0.000 description 2
- 108091036078 conserved sequence Proteins 0.000 description 2
- NKLPQNGYXWVELD-UHFFFAOYSA-M coomassie brilliant blue Chemical compound [Na+].C1=CC(OCC)=CC=C1NC1=CC=C(C(=C2C=CC(C=C2)=[N+](CC)CC=2C=C(C=CC=2)S([O-])(=O)=O)C=2C=CC(=CC=2)N(CC)CC=2C=C(C=CC=2)S([O-])(=O)=O)C=C1 NKLPQNGYXWVELD-UHFFFAOYSA-M 0.000 description 2
- 238000005336 cracking Methods 0.000 description 2
- 238000005520 cutting process Methods 0.000 description 2
- 230000003013 cytotoxicity Effects 0.000 description 2
- 231100000135 cytotoxicity Toxicity 0.000 description 2
- 230000007547 defect Effects 0.000 description 2
- 230000001419 dependent effect Effects 0.000 description 2
- 238000013461 design Methods 0.000 description 2
- 238000001514 detection method Methods 0.000 description 2
- 239000003085 diluting agent Substances 0.000 description 2
- 238000010494 dissociation reaction Methods 0.000 description 2
- 230000005593 dissociations Effects 0.000 description 2
- 238000009826 distribution Methods 0.000 description 2
- 239000012636 effector Substances 0.000 description 2
- 230000002124 endocrine Effects 0.000 description 2
- 239000002532 enzyme inhibitor Substances 0.000 description 2
- 238000002474 experimental method Methods 0.000 description 2
- 239000013604 expression vector Substances 0.000 description 2
- 229960002584 gefitinib Drugs 0.000 description 2
- 108010049041 glutamylalanine Proteins 0.000 description 2
- 108010028188 glycyl-histidyl-serine Proteins 0.000 description 2
- 108010017446 glycyl-prolyl-arginyl-proline Proteins 0.000 description 2
- 108010079413 glycyl-prolyl-glutamic acid Proteins 0.000 description 2
- 239000003102 growth factor Substances 0.000 description 2
- 231100000652 hormesis Toxicity 0.000 description 2
- 229940088597 hormone Drugs 0.000 description 2
- 239000005556 hormone Substances 0.000 description 2
- 230000001900 immune effect Effects 0.000 description 2
- 230000003053 immunization Effects 0.000 description 2
- 238000002649 immunization Methods 0.000 description 2
- 230000008676 import Effects 0.000 description 2
- 238000001802 infusion Methods 0.000 description 2
- NOESYZHRGYRDHS-UHFFFAOYSA-N insulin Chemical compound N1C(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(NC(=O)CN)C(C)CC)CSSCC(C(NC(CO)C(=O)NC(CC(C)C)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CCC(N)=O)C(=O)NC(CC(C)C)C(=O)NC(CCC(O)=O)C(=O)NC(CC(N)=O)C(=O)NC(CC=2C=CC(O)=CC=2)C(=O)NC(CSSCC(NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2C=CC(O)=CC=2)NC(=O)C(CC(C)C)NC(=O)C(C)NC(=O)C(CCC(O)=O)NC(=O)C(C(C)C)NC(=O)C(CC(C)C)NC(=O)C(CC=2NC=NC=2)NC(=O)C(CO)NC(=O)CNC2=O)C(=O)NCC(=O)NC(CCC(O)=O)C(=O)NC(CCCNC(N)=N)C(=O)NCC(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC=CC=3)C(=O)NC(CC=3C=CC(O)=CC=3)C(=O)NC(C(C)O)C(=O)N3C(CCC3)C(=O)NC(CCCCN)C(=O)NC(C)C(O)=O)C(=O)NC(CC(N)=O)C(O)=O)=O)NC(=O)C(C(C)CC)NC(=O)C(CO)NC(=O)C(C(C)O)NC(=O)C1CSSCC2NC(=O)C(CC(C)C)NC(=O)C(NC(=O)C(CCC(N)=O)NC(=O)C(CC(N)=O)NC(=O)C(NC(=O)C(N)CC=1C=CC=CC=1)C(C)C)CC1=CN=CN1 NOESYZHRGYRDHS-UHFFFAOYSA-N 0.000 description 2
- 230000003834 intracellular effect Effects 0.000 description 2
- 238000007912 intraperitoneal administration Methods 0.000 description 2
- 230000007794 irritation Effects 0.000 description 2
- 210000003734 kidney Anatomy 0.000 description 2
- 208000032839 leukemia Diseases 0.000 description 2
- GFIJNRVAKGFPGQ-LIJARHBVSA-N leuprolide Chemical compound CCNC(=O)[C@@H]1CCCN1C(=O)[C@H](CCCNC(N)=N)NC(=O)[C@H](CC(C)C)NC(=O)[C@@H](CC(C)C)NC(=O)[C@@H](NC(=O)[C@H](CO)NC(=O)[C@H](CC=1C2=CC=CC=C2NC=1)NC(=O)[C@H](CC=1N=CNC=1)NC(=O)[C@H]1NC(=O)CC1)CC1=CC=C(O)C=C1 GFIJNRVAKGFPGQ-LIJARHBVSA-N 0.000 description 2
- 229960004338 leuprorelin Drugs 0.000 description 2
- 238000012417 linear regression Methods 0.000 description 2
- 239000007788 liquid Substances 0.000 description 2
- 201000005296 lung carcinoma Diseases 0.000 description 2
- 108010054155 lysyllysine Proteins 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 239000012528 membrane Substances 0.000 description 2
- 230000004060 metabolic process Effects 0.000 description 2
- 229960000485 methotrexate Drugs 0.000 description 2
- 229940126619 mouse monoclonal antibody Drugs 0.000 description 2
- 238000002703 mutagenesis Methods 0.000 description 2
- 239000003471 mutagenic agent Substances 0.000 description 2
- 125000002347 octyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])C([H])([H])[H] 0.000 description 2
- 229940049954 penicillin Drugs 0.000 description 2
- 239000000546 pharmaceutical excipient Substances 0.000 description 2
- YBYRMVIVWMBXKQ-UHFFFAOYSA-N phenylmethanesulfonyl fluoride Chemical compound FS(=O)(=O)CC1=CC=CC=C1 YBYRMVIVWMBXKQ-UHFFFAOYSA-N 0.000 description 2
- 210000002826 placenta Anatomy 0.000 description 2
- 108091033319 polynucleotide Proteins 0.000 description 2
- 102000040430 polynucleotide Human genes 0.000 description 2
- 239000002157 polynucleotide Substances 0.000 description 2
- 229920001184 polypeptide Polymers 0.000 description 2
- 229920002981 polyvinylidene fluoride Polymers 0.000 description 2
- 238000004321 preservation Methods 0.000 description 2
- 238000000734 protein sequencing Methods 0.000 description 2
- 238000000746 purification Methods 0.000 description 2
- RXWNCPJZOCPEPQ-NVWDDTSBSA-N puromycin Chemical compound C1=CC(OC)=CC=C1C[C@H](N)C(=O)N[C@H]1[C@@H](O)[C@H](N2C3=NC=NC(=C3N=C2)N(C)C)O[C@@H]1CO RXWNCPJZOCPEPQ-NVWDDTSBSA-N 0.000 description 2
- 238000012372 quality testing Methods 0.000 description 2
- 239000000376 reactant Substances 0.000 description 2
- 230000004044 response Effects 0.000 description 2
- 108091008146 restriction endonucleases Proteins 0.000 description 2
- 238000012163 sequencing technique Methods 0.000 description 2
- 108010026333 seryl-proline Proteins 0.000 description 2
- 238000002741 site-directed mutagenesis Methods 0.000 description 2
- PUZPDOWCWNUUKD-UHFFFAOYSA-M sodium fluoride Chemical compound [F-].[Na+] PUZPDOWCWNUUKD-UHFFFAOYSA-M 0.000 description 2
- 230000037439 somatic mutation Effects 0.000 description 2
- 210000000952 spleen Anatomy 0.000 description 2
- 238000010186 staining Methods 0.000 description 2
- QTENRWWVYAAPBI-YCRXJPFRSA-N streptomycin sulfate Chemical compound OS(O)(=O)=O.OS(O)(=O)=O.OS(O)(=O)=O.CN[C@H]1[C@H](O)[C@@H](O)[C@H](CO)O[C@H]1O[C@@H]1[C@](C=O)(O)[C@H](C)O[C@H]1O[C@@H]1[C@@H](N=C(N)N)[C@H](O)[C@@H](N=C(N)N)[C@H](O)[C@H]1O.CN[C@H]1[C@H](O)[C@@H](O)[C@H](CO)O[C@H]1O[C@@H]1[C@](C=O)(O)[C@H](C)O[C@H]1O[C@@H]1[C@@H](N=C(N)N)[C@H](O)[C@@H](N=C(N)N)[C@H](O)[C@H]1O QTENRWWVYAAPBI-YCRXJPFRSA-N 0.000 description 2
- 238000010254 subcutaneous injection Methods 0.000 description 2
- 239000007929 subcutaneous injection Substances 0.000 description 2
- 229930101283 tetracycline Natural products 0.000 description 2
- 229960002180 tetracycline Drugs 0.000 description 2
- 235000019364 tetracycline Nutrition 0.000 description 2
- 150000003522 tetracyclines Chemical class 0.000 description 2
- 238000002560 therapeutic procedure Methods 0.000 description 2
- 125000000101 thioether group Chemical group 0.000 description 2
- 108010033670 threonyl-aspartyl-tyrosine Proteins 0.000 description 2
- 108010061238 threonyl-glycine Proteins 0.000 description 2
- 229940104230 thymidine Drugs 0.000 description 2
- 210000001519 tissue Anatomy 0.000 description 2
- 238000004448 titration Methods 0.000 description 2
- 238000000954 titration curve Methods 0.000 description 2
- 238000003146 transient transfection Methods 0.000 description 2
- LENZDBCJOHFCAS-UHFFFAOYSA-N tris Chemical compound OCC(N)(CO)CO LENZDBCJOHFCAS-UHFFFAOYSA-N 0.000 description 2
- 230000004614 tumor growth Effects 0.000 description 2
- 210000003462 vein Anatomy 0.000 description 2
- OGWKCGZFUXNPDA-UHFFFAOYSA-N vincristine Natural products C1C(CC)(O)CC(CC2(C(=O)OC)C=3C(=CC4=C(C56C(C(C(OC(C)=O)C7(CC)C=CCN(C67)CC5)(O)C(=O)OC)N4C=O)C=3)OC)CN1CCC1=C2NC2=CC=CC=C12 OGWKCGZFUXNPDA-UHFFFAOYSA-N 0.000 description 2
- OGWKCGZFUXNPDA-XQKSVPLYSA-N vincristine Chemical compound C([N@]1C[C@@H](C[C@]2(C(=O)OC)C=3C(=CC4=C([C@]56[C@H]([C@@]([C@H](OC(C)=O)[C@]7(CC)C=CCN([C@H]67)CC5)(O)C(=O)OC)N4C=O)C=3)OC)C[C@@](C1)(O)CC)CC1=C2NC2=CC=CC=C12 OGWKCGZFUXNPDA-XQKSVPLYSA-N 0.000 description 2
- 229960004528 vincristine Drugs 0.000 description 2
- 230000003442 weekly effect Effects 0.000 description 2
- 108010027345 wheylin-1 peptide Proteins 0.000 description 2
- XVZCXCTYGHPNEM-IHRRRGAJSA-N (2s)-1-[(2s)-2-[[(2s)-2-amino-4-methylpentanoyl]amino]-4-methylpentanoyl]pyrrolidine-2-carboxylic acid Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](CC(C)C)C(=O)N1CCC[C@H]1C(O)=O XVZCXCTYGHPNEM-IHRRRGAJSA-N 0.000 description 1
- VEEGZPWAAPPXRB-BJMVGYQFSA-N (3e)-3-(1h-imidazol-5-ylmethylidene)-1h-indol-2-one Chemical compound O=C1NC2=CC=CC=C2\C1=C/C1=CN=CN1 VEEGZPWAAPPXRB-BJMVGYQFSA-N 0.000 description 1
- HZKLCOYAVAAQRD-VGMNWLOBSA-N (3s)-3-[[2-[[(2s)-2-amino-5-(diaminomethylideneamino)pentanoyl]amino]acetyl]amino]-4-[[(1r)-1-carboxyethyl]amino]-4-oxobutanoic acid Chemical compound OC(=O)[C@@H](C)NC(=O)[C@H](CC(O)=O)NC(=O)CNC(=O)[C@@H](N)CCCN=C(N)N HZKLCOYAVAAQRD-VGMNWLOBSA-N 0.000 description 1
- BMGMINKVTPDDRZ-UHFFFAOYSA-N 2-acetamido-n-[1-[[5-(diaminomethylideneamino)-1-oxopentan-2-yl]amino]-4-methyl-1-oxopentan-2-yl]-4-methylpentanamide;n-[1-[[5-(diaminomethylideneamino)-1-oxopentan-2-yl]amino]-4-methyl-1-oxopentan-2-yl]-4-methyl-2-(propanoylamino)pentanamide Chemical compound CC(C)CC(NC(C)=O)C(=O)NC(CC(C)C)C(=O)NC(C=O)CCCN=C(N)N.CCC(=O)NC(CC(C)C)C(=O)NC(CC(C)C)C(=O)NC(C=O)CCCN=C(N)N BMGMINKVTPDDRZ-UHFFFAOYSA-N 0.000 description 1
- TVZGACDUOSZQKY-LBPRGKRZSA-N 4-aminofolic acid Chemical compound C1=NC2=NC(N)=NC(N)=C2N=C1CNC1=CC=C(C(=O)N[C@@H](CCC(O)=O)C(O)=O)C=C1 TVZGACDUOSZQKY-LBPRGKRZSA-N 0.000 description 1
- SVBXIUDNTRTKHE-CIUDSAMLSA-N Ala-Arg-Glu Chemical compound [H]N[C@@H](C)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCC(O)=O)C(O)=O SVBXIUDNTRTKHE-CIUDSAMLSA-N 0.000 description 1
- DWINFPQUSSHSFS-UVBJJODRSA-N Ala-Arg-Trp Chemical compound N[C@@H](C)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC1=CNC2=CC=CC=C12)C(=O)O DWINFPQUSSHSFS-UVBJJODRSA-N 0.000 description 1
- BTYTYHBSJKQBQA-GCJQMDKQSA-N Ala-Asp-Thr Chemical compound C[C@H]([C@@H](C(=O)O)NC(=O)[C@H](CC(=O)O)NC(=O)[C@H](C)N)O BTYTYHBSJKQBQA-GCJQMDKQSA-N 0.000 description 1
- AWZKCUCQJNTBAD-SRVKXCTJSA-N Ala-Leu-Lys Chemical compound C[C@H](N)C(=O)N[C@@H](CC(C)C)C(=O)N[C@H](C(O)=O)CCCCN AWZKCUCQJNTBAD-SRVKXCTJSA-N 0.000 description 1
- HOVPGJUNRLMIOZ-CIUDSAMLSA-N Ala-Ser-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@H](CO)NC(=O)[C@H](C)N HOVPGJUNRLMIOZ-CIUDSAMLSA-N 0.000 description 1
- KLKARCOHVHLAJP-UWJYBYFXSA-N Ala-Tyr-Cys Chemical compound C[C@H](N)C(=O)N[C@@H](Cc1ccc(O)cc1)C(=O)N[C@@H](CS)C(O)=O KLKARCOHVHLAJP-UWJYBYFXSA-N 0.000 description 1
- 102100027211 Albumin Human genes 0.000 description 1
- 102000009027 Albumins Human genes 0.000 description 1
- 108020004774 Alkaline Phosphatase Proteins 0.000 description 1
- 102000002260 Alkaline Phosphatase Human genes 0.000 description 1
- 102000010565 Apoptosis Regulatory Proteins Human genes 0.000 description 1
- 108010063104 Apoptosis Regulatory Proteins Proteins 0.000 description 1
- OGUPCHKBOKJFMA-SRVKXCTJSA-N Arg-Glu-Lys Chemical compound NCCCC[C@@H](C(O)=O)NC(=O)[C@H](CCC(O)=O)NC(=O)[C@@H](N)CCCN=C(N)N OGUPCHKBOKJFMA-SRVKXCTJSA-N 0.000 description 1
- AOHKLEBWKMKITA-IHRRRGAJSA-N Arg-Phe-Ser Chemical compound C1=CC=C(C=C1)C[C@@H](C(=O)N[C@@H](CO)C(=O)O)NC(=O)[C@H](CCCN=C(N)N)N AOHKLEBWKMKITA-IHRRRGAJSA-N 0.000 description 1
- JJIBHAOBNIFUEL-SRVKXCTJSA-N Arg-Pro-Met Chemical compound CSCC[C@@H](C(=O)O)NC(=O)[C@@H]1CCCN1C(=O)[C@H](CCCN=C(N)N)N JJIBHAOBNIFUEL-SRVKXCTJSA-N 0.000 description 1
- FRBAHXABMQXSJQ-FXQIFTODSA-N Arg-Ser-Ser Chemical compound [H]N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CO)C(=O)N[C@@H](CO)C(O)=O FRBAHXABMQXSJQ-FXQIFTODSA-N 0.000 description 1
- VLIJAPRTSXSGFY-STQMWFEESA-N Arg-Tyr-Gly Chemical compound NC(N)=NCCC[C@H](N)C(=O)N[C@H](C(=O)NCC(O)=O)CC1=CC=C(O)C=C1 VLIJAPRTSXSGFY-STQMWFEESA-N 0.000 description 1
- OKZOABJQOMAYEC-NUMRIWBASA-N Asn-Gln-Thr Chemical compound [H]N[C@@H](CC(N)=O)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H]([C@@H](C)O)C(O)=O OKZOABJQOMAYEC-NUMRIWBASA-N 0.000 description 1
- RTFXPCYMDYBZNQ-SRVKXCTJSA-N Asn-Tyr-Asn Chemical compound [H]N[C@@H](CC(N)=O)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CC(N)=O)C(O)=O RTFXPCYMDYBZNQ-SRVKXCTJSA-N 0.000 description 1
- KTDWFWNZLLFEFU-KKUMJFAQSA-N Asn-Tyr-His Chemical compound C1=CC(=CC=C1C[C@@H](C(=O)N[C@@H](CC2=CN=CN2)C(=O)O)NC(=O)[C@H](CC(=O)N)N)O KTDWFWNZLLFEFU-KKUMJFAQSA-N 0.000 description 1
- WQAOZCVOOYUWKG-LSJOCFKGSA-N Asn-Val-Val Chemical compound CC(C)[C@@H](C(=O)N[C@@H](C(C)C)C(=O)O)NC(=O)[C@H](CC(=O)N)N WQAOZCVOOYUWKG-LSJOCFKGSA-N 0.000 description 1
- DXQOQMCLWWADMU-ACZMJKKPSA-N Asp-Gln-Ser Chemical compound OC(=O)C[C@H](N)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CO)C(O)=O DXQOQMCLWWADMU-ACZMJKKPSA-N 0.000 description 1
- NRIFEOUAFLTMFJ-AAEUAGOBSA-N Asp-Gly-Trp Chemical compound [H]N[C@@H](CC(O)=O)C(=O)NCC(=O)N[C@@H](CC1=CNC2=C1C=CC=C2)C(O)=O NRIFEOUAFLTMFJ-AAEUAGOBSA-N 0.000 description 1
- KQBVNNAPIURMPD-PEFMBERDSA-N Asp-Ile-Glu Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CCC(O)=O)C(O)=O KQBVNNAPIURMPD-PEFMBERDSA-N 0.000 description 1
- ORRJQLIATJDMQM-HJGDQZAQSA-N Asp-Leu-Thr Chemical compound C[C@@H](O)[C@@H](C(O)=O)NC(=O)[C@H](CC(C)C)NC(=O)[C@@H](N)CC(O)=O ORRJQLIATJDMQM-HJGDQZAQSA-N 0.000 description 1
- IOXWDLNHXZOXQP-FXQIFTODSA-N Asp-Met-Ser Chemical compound CSCC[C@@H](C(=O)N[C@@H](CO)C(=O)O)NC(=O)[C@H](CC(=O)O)N IOXWDLNHXZOXQP-FXQIFTODSA-N 0.000 description 1
- HICVMZCGVFKTPM-BQBZGAKWSA-N Asp-Pro-Gly Chemical compound OC(=O)C[C@H](N)C(=O)N1CCC[C@H]1C(=O)NCC(O)=O HICVMZCGVFKTPM-BQBZGAKWSA-N 0.000 description 1
- FAUPLTGRUBTXNU-FXQIFTODSA-N Asp-Pro-Ser Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CO)C(O)=O FAUPLTGRUBTXNU-FXQIFTODSA-N 0.000 description 1
- XYPJXLLXNSAWHZ-SRVKXCTJSA-N Asp-Ser-Tyr Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H](CO)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(O)=O XYPJXLLXNSAWHZ-SRVKXCTJSA-N 0.000 description 1
- MNQMTYSEKZHIDF-GCJQMDKQSA-N Asp-Thr-Ala Chemical compound [H]N[C@@H](CC(O)=O)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](C)C(O)=O MNQMTYSEKZHIDF-GCJQMDKQSA-N 0.000 description 1
- RKXVTTIQNKPCHU-KKHAAJSZSA-N Asp-Val-Thr Chemical compound C[C@@H](O)[C@@H](C(O)=O)NC(=O)[C@H](C(C)C)NC(=O)[C@@H](N)CC(O)=O RKXVTTIQNKPCHU-KKHAAJSZSA-N 0.000 description 1
- 241000894006 Bacteria Species 0.000 description 1
- 102100032985 CCR4-NOT transcription complex subunit 7 Human genes 0.000 description 1
- 108050006912 CCR4-NOT transcription complex subunit 7 Proteins 0.000 description 1
- 239000008000 CHES buffer Substances 0.000 description 1
- GAGWJHPBXLXJQN-UORFTKCHSA-N Capecitabine Chemical compound C1=C(F)C(NC(=O)OCCCCC)=NC(=O)N1[C@H]1[C@H](O)[C@H](O)[C@@H](C)O1 GAGWJHPBXLXJQN-UORFTKCHSA-N 0.000 description 1
- GAGWJHPBXLXJQN-UHFFFAOYSA-N Capecitabine Natural products C1=C(F)C(NC(=O)OCCCCC)=NC(=O)N1C1C(O)C(O)C(C)O1 GAGWJHPBXLXJQN-UHFFFAOYSA-N 0.000 description 1
- 241000283707 Capra Species 0.000 description 1
- 241000282552 Chlorocebus aethiops Species 0.000 description 1
- JTNKVWLMDHIUOG-IHRRRGAJSA-N Cys-Arg-Phe Chemical compound [H]N[C@@H](CS)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC1=CC=CC=C1)C(O)=O JTNKVWLMDHIUOG-IHRRRGAJSA-N 0.000 description 1
- NIXHTNJAGGFBAW-CIUDSAMLSA-N Cys-Lys-Ser Chemical compound C(CCN)C[C@@H](C(=O)N[C@@H](CO)C(=O)O)NC(=O)[C@H](CS)N NIXHTNJAGGFBAW-CIUDSAMLSA-N 0.000 description 1
- WEAHRLBPCANXCN-UHFFFAOYSA-N Daunomycin Natural products CCC1(O)CC(OC2CC(N)C(O)C(C)O2)c3cc4C(=O)c5c(OC)cccc5C(=O)c4c(O)c3C1 WEAHRLBPCANXCN-UHFFFAOYSA-N 0.000 description 1
- BWGNESOTFCXPMA-UHFFFAOYSA-N Dihydrogen disulfide Chemical compound SS BWGNESOTFCXPMA-UHFFFAOYSA-N 0.000 description 1
- 208000030453 Drug-Related Side Effects and Adverse reaction Diseases 0.000 description 1
- 108090000371 Esterases Proteins 0.000 description 1
- 208000006168 Ewing Sarcoma Diseases 0.000 description 1
- 102000007317 Farnesyltranstransferase Human genes 0.000 description 1
- 108010007508 Farnesyltranstransferase Proteins 0.000 description 1
- 238000012413 Fluorescence activated cell sorting analysis Methods 0.000 description 1
- RZSLYUUFFVHFRQ-FXQIFTODSA-N Gln-Ala-Glu Chemical compound [H]N[C@@H](CCC(N)=O)C(=O)N[C@@H](C)C(=O)N[C@@H](CCC(O)=O)C(O)=O RZSLYUUFFVHFRQ-FXQIFTODSA-N 0.000 description 1
- RBWKVOSARCFSQQ-FXQIFTODSA-N Gln-Gln-Ser Chemical compound NC(=O)CC[C@H](N)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CO)C(O)=O RBWKVOSARCFSQQ-FXQIFTODSA-N 0.000 description 1
- NSORZJXKUQFEKL-JGVFFNPUSA-N Gln-Gly-Pro Chemical compound C1C[C@@H](N(C1)C(=O)CNC(=O)[C@H](CCC(=O)N)N)C(=O)O NSORZJXKUQFEKL-JGVFFNPUSA-N 0.000 description 1
- VZRAXPGTUNDIDK-GUBZILKMSA-N Gln-Leu-Asn Chemical compound CC(C)C[C@@H](C(=O)N[C@@H](CC(=O)N)C(=O)O)NC(=O)[C@H](CCC(=O)N)N VZRAXPGTUNDIDK-GUBZILKMSA-N 0.000 description 1
- SXFPZRRVWSUYII-KBIXCLLPSA-N Gln-Ser-Ile Chemical compound CC[C@H](C)[C@@H](C(=O)O)NC(=O)[C@H](CO)NC(=O)[C@H](CCC(=O)N)N SXFPZRRVWSUYII-KBIXCLLPSA-N 0.000 description 1
- MXOODARRORARSU-ACZMJKKPSA-N Glu-Ala-Ser Chemical compound C[C@@H](C(=O)N[C@@H](CO)C(=O)O)NC(=O)[C@H](CCC(=O)O)N MXOODARRORARSU-ACZMJKKPSA-N 0.000 description 1
- LTUVYLVIZHJCOQ-KKUMJFAQSA-N Glu-Arg-Phe Chemical compound [H]N[C@@H](CCC(O)=O)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC1=CC=CC=C1)C(O)=O LTUVYLVIZHJCOQ-KKUMJFAQSA-N 0.000 description 1
- QXDXIXFSFHUYAX-MNXVOIDGSA-N Glu-Ile-Leu Chemical compound CC(C)C[C@@H](C(O)=O)NC(=O)[C@H]([C@@H](C)CC)NC(=O)[C@@H](N)CCC(O)=O QXDXIXFSFHUYAX-MNXVOIDGSA-N 0.000 description 1
- 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 1
- MFVQGXGQRIXBPK-WDSKDSINSA-N Gly-Ala-Glu Chemical compound NCC(=O)N[C@@H](C)C(=O)N[C@@H](CCC(O)=O)C(O)=O MFVQGXGQRIXBPK-WDSKDSINSA-N 0.000 description 1
- PMNHJLASAAWELO-FOHZUACHSA-N Gly-Asp-Thr Chemical compound [H]NCC(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H]([C@@H](C)O)C(O)=O PMNHJLASAAWELO-FOHZUACHSA-N 0.000 description 1
- KTSZUNRRYXPZTK-BQBZGAKWSA-N Gly-Gln-Glu Chemical compound NCC(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCC(O)=O)C(O)=O KTSZUNRRYXPZTK-BQBZGAKWSA-N 0.000 description 1
- XTQFHTHIAKKCTM-YFKPBYRVSA-N Gly-Glu-Gly Chemical compound NCC(=O)N[C@@H](CCC(O)=O)C(=O)NCC(O)=O XTQFHTHIAKKCTM-YFKPBYRVSA-N 0.000 description 1
- QSVCIFZPGLOZGH-WDSKDSINSA-N Gly-Glu-Ser Chemical compound NCC(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CO)C(O)=O QSVCIFZPGLOZGH-WDSKDSINSA-N 0.000 description 1
- UQJNXZSSGQIPIQ-FBCQKBJTSA-N Gly-Gly-Thr Chemical compound C[C@@H](O)[C@@H](C(O)=O)NC(=O)CNC(=O)CN UQJNXZSSGQIPIQ-FBCQKBJTSA-N 0.000 description 1
- ULZCYBYDTUMHNF-IUCAKERBSA-N Gly-Leu-Glu Chemical compound NCC(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(O)=O)C(O)=O ULZCYBYDTUMHNF-IUCAKERBSA-N 0.000 description 1
- VBOBNHSVQKKTOT-YUMQZZPRSA-N Gly-Lys-Ala Chemical compound [H]NCC(=O)N[C@@H](CCCCN)C(=O)N[C@@H](C)C(O)=O VBOBNHSVQKKTOT-YUMQZZPRSA-N 0.000 description 1
- LOEANKRDMMVOGZ-YUMQZZPRSA-N Gly-Lys-Asp Chemical compound NCCCC[C@H](NC(=O)CN)C(=O)N[C@@H](CC(O)=O)C(O)=O LOEANKRDMMVOGZ-YUMQZZPRSA-N 0.000 description 1
- MHZXESQPPXOING-KBPBESRZSA-N Gly-Lys-Phe Chemical compound [H]NCC(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC1=CC=CC=C1)C(O)=O MHZXESQPPXOING-KBPBESRZSA-N 0.000 description 1
- NTBOEZICHOSJEE-YUMQZZPRSA-N Gly-Lys-Ser Chemical compound [H]NCC(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CO)C(O)=O NTBOEZICHOSJEE-YUMQZZPRSA-N 0.000 description 1
- LCRDMSSAKLTKBU-ZDLURKLDSA-N Gly-Ser-Thr Chemical compound C[C@@H](O)[C@@H](C(O)=O)NC(=O)[C@H](CO)NC(=O)CN LCRDMSSAKLTKBU-ZDLURKLDSA-N 0.000 description 1
- QYZRTBKYBJRGJB-PCMHIUKPSA-N Granisetron hydrochloride Chemical compound Cl.C1=CC=C2C(C(=O)NC3C[C@H]4CCC[C@@H](C3)N4C)=NN(C)C2=C1 QYZRTBKYBJRGJB-PCMHIUKPSA-N 0.000 description 1
- VBOFRJNDIOPNDO-YUMQZZPRSA-N His-Gly-Asn Chemical compound C1=C(NC=N1)C[C@@H](C(=O)NCC(=O)N[C@@H](CC(=O)N)C(=O)O)N VBOFRJNDIOPNDO-YUMQZZPRSA-N 0.000 description 1
- RGPWUJOMKFYFSR-QWRGUYRKSA-N His-Gly-Leu Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)NCC(=O)N[C@@H](CC(C)C)C(O)=O RGPWUJOMKFYFSR-QWRGUYRKSA-N 0.000 description 1
- GNBHSMFBUNEWCJ-DCAQKATOSA-N His-Pro-Asn Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)N1CCC[C@H]1C(=O)N[C@@H](CC(N)=O)C(O)=O GNBHSMFBUNEWCJ-DCAQKATOSA-N 0.000 description 1
- ZNTSGDNUITWTRA-WDSOQIARSA-N His-Trp-Val Chemical compound [H]N[C@@H](CC1=CNC=N1)C(=O)N[C@@H](CC1=CNC2=C1C=CC=C2)C(=O)N[C@@H](C(C)C)C(O)=O ZNTSGDNUITWTRA-WDSOQIARSA-N 0.000 description 1
- 101000911390 Homo sapiens Coagulation factor VIII Proteins 0.000 description 1
- 101001034652 Homo sapiens Insulin-like growth factor 1 receptor Proteins 0.000 description 1
- 101000904173 Homo sapiens Progonadoliberin-1 Proteins 0.000 description 1
- 101001059454 Homo sapiens Serine/threonine-protein kinase MARK2 Proteins 0.000 description 1
- 108010001336 Horseradish Peroxidase Proteins 0.000 description 1
- LEHPJMKVGFPSSP-ZQINRCPSSA-N Ile-Glu-Trp Chemical compound C1=CC=C2C(C[C@H](NC(=O)[C@H](CCC(O)=O)NC(=O)[C@@H](N)[C@@H](C)CC)C(O)=O)=CNC2=C1 LEHPJMKVGFPSSP-ZQINRCPSSA-N 0.000 description 1
- HPCFRQWLTRDGHT-AJNGGQMLSA-N Ile-Leu-Leu Chemical compound CC[C@H](C)[C@H](N)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC(C)C)C(O)=O HPCFRQWLTRDGHT-AJNGGQMLSA-N 0.000 description 1
- 108060003951 Immunoglobulin Proteins 0.000 description 1
- 102100025092 Insulin receptor substrate 2 Human genes 0.000 description 1
- 101710201820 Insulin receptor substrate 2 Proteins 0.000 description 1
- 102000004374 Insulin-like growth factor binding protein 3 Human genes 0.000 description 1
- 108090000965 Insulin-like growth factor binding protein 3 Proteins 0.000 description 1
- 102100034343 Integrase Human genes 0.000 description 1
- 102000011782 Keratins Human genes 0.000 description 1
- 108010076876 Keratins Proteins 0.000 description 1
- FADYJNXDPBKVCA-UHFFFAOYSA-N L-Phenylalanyl-L-lysin Natural products NCCCCC(C(O)=O)NC(=O)C(N)CC1=CC=CC=C1 FADYJNXDPBKVCA-UHFFFAOYSA-N 0.000 description 1
- LHSGPCFBGJHPCY-UHFFFAOYSA-N L-leucine-L-tyrosine Natural products CC(C)CC(N)C(=O)NC(C(O)=O)CC1=CC=C(O)C=C1 LHSGPCFBGJHPCY-UHFFFAOYSA-N 0.000 description 1
- OUYCCCASQSFEME-QMMMGPOBSA-N L-tyrosine Chemical compound OC(=O)[C@@H](N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-QMMMGPOBSA-N 0.000 description 1
- QLQHWWCSCLZUMA-KKUMJFAQSA-N Leu-Asp-Tyr Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@H](C(O)=O)CC1=CC=C(O)C=C1 QLQHWWCSCLZUMA-KKUMJFAQSA-N 0.000 description 1
- CQGSYZCULZMEDE-UHFFFAOYSA-N Leu-Gln-Pro Natural products CC(C)CC(N)C(=O)NC(CCC(N)=O)C(=O)N1CCCC1C(O)=O CQGSYZCULZMEDE-UHFFFAOYSA-N 0.000 description 1
- OMHLATXVNQSALM-FQUUOJAGSA-N Leu-Ile-Pro Chemical compound CC[C@H](C)[C@@H](C(=O)N1CCC[C@@H]1C(=O)O)NC(=O)[C@H](CC(C)C)N OMHLATXVNQSALM-FQUUOJAGSA-N 0.000 description 1
- JNDYEOUZBLOVOF-AVGNSLFASA-N Leu-Leu-Gln Chemical compound [H]N[C@@H](CC(C)C)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(N)=O)C(O)=O JNDYEOUZBLOVOF-AVGNSLFASA-N 0.000 description 1
- QNBVTHNJGCOVFA-AVGNSLFASA-N Leu-Leu-Glu Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](CC(C)C)C(=O)N[C@H](C(O)=O)CCC(O)=O QNBVTHNJGCOVFA-AVGNSLFASA-N 0.000 description 1
- XVZCXCTYGHPNEM-UHFFFAOYSA-N Leu-Leu-Pro Natural products CC(C)CC(N)C(=O)NC(CC(C)C)C(=O)N1CCCC1C(O)=O XVZCXCTYGHPNEM-UHFFFAOYSA-N 0.000 description 1
- POMXSEDNUXYPGK-IHRRRGAJSA-N Leu-Met-His Chemical compound CC(C)C[C@@H](C(=O)N[C@@H](CCSC)C(=O)N[C@@H](CC1=CN=CN1)C(=O)O)N POMXSEDNUXYPGK-IHRRRGAJSA-N 0.000 description 1
- IBSGMIPRBMPMHE-IHRRRGAJSA-N Leu-Met-Lys Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](CCSC)C(=O)N[C@@H](CCCCN)C(O)=O IBSGMIPRBMPMHE-IHRRRGAJSA-N 0.000 description 1
- LCNASHSOFMRYFO-WDCWCFNPSA-N Leu-Thr-Gln Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@H](C(O)=O)CCC(N)=O LCNASHSOFMRYFO-WDCWCFNPSA-N 0.000 description 1
- FBNPMTNBFFAMMH-AVGNSLFASA-N Leu-Val-Arg Chemical compound CC(C)C[C@H](N)C(=O)N[C@@H](C(C)C)C(=O)N[C@H](C(O)=O)CCCN=C(N)N FBNPMTNBFFAMMH-AVGNSLFASA-N 0.000 description 1
- FBNPMTNBFFAMMH-UHFFFAOYSA-N Leu-Val-Arg Natural products CC(C)CC(N)C(=O)NC(C(C)C)C(=O)NC(C(O)=O)CCCN=C(N)N FBNPMTNBFFAMMH-UHFFFAOYSA-N 0.000 description 1
- 108010011078 Leupeptins Proteins 0.000 description 1
- 229940124041 Luteinizing hormone releasing hormone (LHRH) antagonist Drugs 0.000 description 1
- WTZUSCUIVPVCRH-SRVKXCTJSA-N Lys-Gln-Arg Chemical compound NCCCC[C@H](N)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@H](C(O)=O)CCCN=C(N)N WTZUSCUIVPVCRH-SRVKXCTJSA-N 0.000 description 1
- FHIAJWBDZVHLAH-YUMQZZPRSA-N Lys-Gly-Ser Chemical compound NCCCC[C@H](N)C(=O)NCC(=O)N[C@@H](CO)C(O)=O FHIAJWBDZVHLAH-YUMQZZPRSA-N 0.000 description 1
- CANPXOLVTMKURR-WEDXCCLWSA-N Lys-Gly-Thr Chemical compound C[C@@H](O)[C@@H](C(O)=O)NC(=O)CNC(=O)[C@@H](N)CCCCN CANPXOLVTMKURR-WEDXCCLWSA-N 0.000 description 1
- PRSBSVAVOQOAMI-BJDJZHNGSA-N Lys-Ile-Ser Chemical compound OC[C@@H](C(O)=O)NC(=O)[C@H]([C@@H](C)CC)NC(=O)[C@@H](N)CCCCN PRSBSVAVOQOAMI-BJDJZHNGSA-N 0.000 description 1
- SKRGVGLIRUGANF-AVGNSLFASA-N Lys-Leu-Glu Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(O)=O)C(O)=O SKRGVGLIRUGANF-AVGNSLFASA-N 0.000 description 1
- CAVRAQIDHUPECU-UVOCVTCTSA-N Lys-Thr-Thr Chemical compound [H]N[C@@H](CCCCN)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H]([C@@H](C)O)C(O)=O CAVRAQIDHUPECU-UVOCVTCTSA-N 0.000 description 1
- RIPJMCFGQHGHNP-RHYQMDGZSA-N Lys-Val-Thr Chemical compound C[C@H]([C@@H](C(=O)O)NC(=O)[C@H](C(C)C)NC(=O)[C@H](CCCCN)N)O RIPJMCFGQHGHNP-RHYQMDGZSA-N 0.000 description 1
- QWPXBEHQFHACTK-UHFFFAOYSA-N Maytansinol Natural products CN1C(=O)CC(O)C2(C)OC2C(C)C(OC(=O)N2)CC2(O)C(OC)C=CC=C(C)CC2=CC(OC)=C(Cl)C1=C2 QWPXBEHQFHACTK-UHFFFAOYSA-N 0.000 description 1
- SJDQOYTYNGZZJX-SRVKXCTJSA-N Met-Glu-Leu Chemical compound CSCC[C@H](N)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(C)C)C(O)=O SJDQOYTYNGZZJX-SRVKXCTJSA-N 0.000 description 1
- 241000699670 Mus sp. Species 0.000 description 1
- MKWKNSIESPFAQN-UHFFFAOYSA-N N-cyclohexyl-2-aminoethanesulfonic acid Chemical compound OS(=O)(=O)CCNC1CCCCC1 MKWKNSIESPFAQN-UHFFFAOYSA-N 0.000 description 1
- 238000005481 NMR spectroscopy Methods 0.000 description 1
- 108090000526 Papain Proteins 0.000 description 1
- 108010036222 Pepstatins Proteins 0.000 description 1
- 108091005804 Peptidases Proteins 0.000 description 1
- 102000035195 Peptidases Human genes 0.000 description 1
- IILUKIJNFMUBNF-IHRRRGAJSA-N Phe-Gln-Gln Chemical compound [H]N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CCC(N)=O)C(=O)N[C@@H](CCC(N)=O)C(O)=O IILUKIJNFMUBNF-IHRRRGAJSA-N 0.000 description 1
- BELBBZDIHDAJOR-UHFFFAOYSA-N Phenolsulfonephthalein Chemical compound C1=CC(O)=CC=C1C1(C=2C=CC(O)=CC=2)C2=CC=CC=C2S(=O)(=O)O1 BELBBZDIHDAJOR-UHFFFAOYSA-N 0.000 description 1
- 108090000608 Phosphoric Monoester Hydrolases Proteins 0.000 description 1
- 102000004160 Phosphoric Monoester Hydrolases Human genes 0.000 description 1
- 108091000080 Phosphotransferase Proteins 0.000 description 1
- 229920003171 Poly (ethylene oxide) Polymers 0.000 description 1
- 229920001213 Polysorbate 20 Polymers 0.000 description 1
- MLQVJYMFASXBGZ-IHRRRGAJSA-N Pro-Asn-Tyr Chemical compound C1C[C@H](NC1)C(=O)N[C@@H](CC(=O)N)C(=O)N[C@@H](CC2=CC=C(C=C2)O)C(=O)O MLQVJYMFASXBGZ-IHRRRGAJSA-N 0.000 description 1
- MRYUJHGPZQNOAD-IHRRRGAJSA-N Pro-Leu-Lys Chemical compound CC(C)C[C@@H](C(=O)N[C@@H](CCCCN)C(=O)O)NC(=O)[C@@H]1CCCN1 MRYUJHGPZQNOAD-IHRRRGAJSA-N 0.000 description 1
- 102100024028 Progonadoliberin-1 Human genes 0.000 description 1
- 239000004365 Protease Substances 0.000 description 1
- 108010076504 Protein Sorting Signals Proteins 0.000 description 1
- 108010092799 RNA-directed DNA polymerase Proteins 0.000 description 1
- 229920005654 Sephadex Polymers 0.000 description 1
- 239000012507 Sephadex™ Substances 0.000 description 1
- QEDMOZUJTGEIBF-FXQIFTODSA-N Ser-Arg-Asp Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC(O)=O)C(O)=O QEDMOZUJTGEIBF-FXQIFTODSA-N 0.000 description 1
- YUSRGTQIPCJNHQ-CIUDSAMLSA-N Ser-Arg-Glu Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CCC(O)=O)C(O)=O YUSRGTQIPCJNHQ-CIUDSAMLSA-N 0.000 description 1
- QGMLKFGTGXWAHF-IHRRRGAJSA-N Ser-Arg-Phe Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CCCNC(N)=N)C(=O)N[C@@H](CC1=CC=CC=C1)C(O)=O QGMLKFGTGXWAHF-IHRRRGAJSA-N 0.000 description 1
- OOKCGAYXSNJBGQ-ZLUOBGJFSA-N Ser-Asn-Asn Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CC(N)=O)C(O)=O OOKCGAYXSNJBGQ-ZLUOBGJFSA-N 0.000 description 1
- HEQPKICPPDOSIN-SRVKXCTJSA-N Ser-Asp-Tyr Chemical compound OC[C@H](N)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@H](C(O)=O)CC1=CC=C(O)C=C1 HEQPKICPPDOSIN-SRVKXCTJSA-N 0.000 description 1
- HJEBZBMOTCQYDN-ACZMJKKPSA-N Ser-Glu-Asp Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CCC(O)=O)C(=O)N[C@@H](CC(O)=O)C(O)=O HJEBZBMOTCQYDN-ACZMJKKPSA-N 0.000 description 1
- WOUIMBGNEUWXQG-VKHMYHEASA-N Ser-Gly Chemical compound OC[C@H](N)C(=O)NCC(O)=O WOUIMBGNEUWXQG-VKHMYHEASA-N 0.000 description 1
- SNVIOQXAHVORQM-WDSKDSINSA-N Ser-Gly-Gln Chemical compound [H]N[C@@H](CO)C(=O)NCC(=O)N[C@@H](CCC(N)=O)C(O)=O SNVIOQXAHVORQM-WDSKDSINSA-N 0.000 description 1
- FYUIFUJFNCLUIX-XVYDVKMFSA-N Ser-His-Ala Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CC1=CNC=N1)C(=O)N[C@@H](C)C(O)=O FYUIFUJFNCLUIX-XVYDVKMFSA-N 0.000 description 1
- UGHCUDLCCVVIJR-VGDYDELISA-N Ser-His-Ile Chemical compound CC[C@H](C)[C@@H](C(=O)O)NC(=O)[C@H](CC1=CN=CN1)NC(=O)[C@H](CO)N UGHCUDLCCVVIJR-VGDYDELISA-N 0.000 description 1
- YUJLIIRMIAGMCQ-CIUDSAMLSA-N Ser-Leu-Ser Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CO)C(O)=O YUJLIIRMIAGMCQ-CIUDSAMLSA-N 0.000 description 1
- MUJQWSAWLLRJCE-KATARQTJSA-N Ser-Leu-Thr Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H]([C@@H](C)O)C(O)=O MUJQWSAWLLRJCE-KATARQTJSA-N 0.000 description 1
- XGQKSRGHEZNWIS-IHRRRGAJSA-N Ser-Pro-Tyr Chemical compound N[C@@H](CO)C(=O)N1CCC[C@H]1C(=O)N[C@@H](Cc1ccc(O)cc1)C(O)=O XGQKSRGHEZNWIS-IHRRRGAJSA-N 0.000 description 1
- PPCZVWHJWJFTFN-ZLUOBGJFSA-N Ser-Ser-Asp Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CO)C(=O)N[C@@H](CC(O)=O)C(O)=O PPCZVWHJWJFTFN-ZLUOBGJFSA-N 0.000 description 1
- CUXJENOFJXOSOZ-BIIVOSGPSA-N Ser-Ser-Pro Chemical compound C1C[C@@H](N(C1)C(=O)[C@H](CO)NC(=O)[C@H](CO)N)C(=O)O CUXJENOFJXOSOZ-BIIVOSGPSA-N 0.000 description 1
- BIWBTRRBHIEVAH-IHPCNDPISA-N Ser-Tyr-Trp Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CC1=CNC2=C1C=CC=C2)C(O)=O BIWBTRRBHIEVAH-IHPCNDPISA-N 0.000 description 1
- IAOHCSQDQDWRQU-GUBZILKMSA-N Ser-Val-Arg Chemical compound [H]N[C@@H](CO)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CCCNC(N)=N)C(O)=O IAOHCSQDQDWRQU-GUBZILKMSA-N 0.000 description 1
- 102100028904 Serine/threonine-protein kinase MARK2 Human genes 0.000 description 1
- 102000013275 Somatomedins Human genes 0.000 description 1
- 241000133426 Streptomyces zelensis Species 0.000 description 1
- 241001655322 Streptomycetales Species 0.000 description 1
- QAOWNCQODCNURD-UHFFFAOYSA-L Sulfate Chemical compound [O-]S([O-])(=O)=O QAOWNCQODCNURD-UHFFFAOYSA-L 0.000 description 1
- 101000996723 Sus scrofa Gonadotropin-releasing hormone receptor Proteins 0.000 description 1
- XFTYVCHLARBHBQ-FOHZUACHSA-N Thr-Gly-Asn Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)NCC(=O)N[C@@H](CC(N)=O)C(O)=O XFTYVCHLARBHBQ-FOHZUACHSA-N 0.000 description 1
- SIMKLINEDYOTKL-MBLNEYKQSA-N Thr-His-Ala Chemical compound C[C@H]([C@@H](C(=O)N[C@@H](CC1=CN=CN1)C(=O)N[C@@H](C)C(=O)O)N)O SIMKLINEDYOTKL-MBLNEYKQSA-N 0.000 description 1
- QNCFWHZVRNXAKW-OEAJRASXSA-N Thr-Lys-Phe Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CC1=CC=CC=C1)C(O)=O QNCFWHZVRNXAKW-OEAJRASXSA-N 0.000 description 1
- BIBYEFRASCNLAA-CDMKHQONSA-N Thr-Phe-Gly Chemical compound C[C@@H](O)[C@H](N)C(=O)N[C@H](C(=O)NCC(O)=O)CC1=CC=CC=C1 BIBYEFRASCNLAA-CDMKHQONSA-N 0.000 description 1
- MXNAOGFNFNKUPD-JHYOHUSXSA-N Thr-Phe-Thr Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H]([C@@H](C)O)C(O)=O MXNAOGFNFNKUPD-JHYOHUSXSA-N 0.000 description 1
- YGCDFAJJCRVQKU-RCWTZXSCSA-N Thr-Pro-Val Chemical compound CC(C)[C@@H](C(O)=O)NC(=O)[C@@H]1CCCN1C(=O)[C@@H](N)[C@@H](C)O YGCDFAJJCRVQKU-RCWTZXSCSA-N 0.000 description 1
- IEZVHOULSUULHD-XGEHTFHBSA-N Thr-Ser-Val Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CO)C(=O)N[C@@H](C(C)C)C(O)=O IEZVHOULSUULHD-XGEHTFHBSA-N 0.000 description 1
- BBPCSGKKPJUYRB-UVOCVTCTSA-N Thr-Thr-Leu Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC(C)C)C(O)=O BBPCSGKKPJUYRB-UVOCVTCTSA-N 0.000 description 1
- XVHAUVJXBFGUPC-RPTUDFQQSA-N Thr-Tyr-Phe Chemical compound [H]N[C@@H]([C@@H](C)O)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CC1=CC=CC=C1)C(O)=O XVHAUVJXBFGUPC-RPTUDFQQSA-N 0.000 description 1
- MNYNCKZAEIAONY-XGEHTFHBSA-N Thr-Val-Ser Chemical compound C[C@@H](O)[C@H](N)C(=O)N[C@@H](C(C)C)C(=O)N[C@@H](CO)C(O)=O MNYNCKZAEIAONY-XGEHTFHBSA-N 0.000 description 1
- 101710183280 Topoisomerase Proteins 0.000 description 1
- 206010070863 Toxicity to various agents Diseases 0.000 description 1
- 229920004890 Triton X-100 Polymers 0.000 description 1
- 239000013504 Triton X-100 Substances 0.000 description 1
- HYVLNORXQGKONN-NUTKFTJISA-N Trp-Ala-Lys Chemical compound C1=CC=C2C(C[C@H](N)C(=O)N[C@@H](C)C(=O)N[C@@H](CCCCN)C(O)=O)=CNC2=C1 HYVLNORXQGKONN-NUTKFTJISA-N 0.000 description 1
- MKDXQPMIQPTTAW-SIXJUCDHSA-N Trp-Ile-His Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CC1=CN=CN1)C(=O)O)NC(=O)[C@H](CC2=CNC3=CC=CC=C32)N MKDXQPMIQPTTAW-SIXJUCDHSA-N 0.000 description 1
- ILDJYIDXESUBOE-HSCHXYMDSA-N Trp-Ile-Lys Chemical compound CC[C@H](C)[C@@H](C(=O)N[C@@H](CCCCN)C(=O)O)NC(=O)[C@H](CC1=CNC2=CC=CC=C21)N ILDJYIDXESUBOE-HSCHXYMDSA-N 0.000 description 1
- FBGDDUKYOBNZJL-WDSOQIARSA-N Trp-Met-Lys Chemical compound CSCC[C@@H](C(=O)N[C@@H](CCCCN)C(=O)O)NC(=O)[C@H](CC1=CNC2=CC=CC=C21)N FBGDDUKYOBNZJL-WDSOQIARSA-N 0.000 description 1
- PWPJLBWYRTVYQS-PMVMPFDFSA-N Trp-Phe-Leu Chemical compound [H]N[C@@H](CC1=CNC2=C1C=CC=C2)C(=O)N[C@@H](CC1=CC=CC=C1)C(=O)N[C@@H](CC(C)C)C(O)=O PWPJLBWYRTVYQS-PMVMPFDFSA-N 0.000 description 1
- DVLHKUWLNKDINO-PMVMPFDFSA-N Trp-Tyr-Leu Chemical compound [H]N[C@@H](CC1=CNC2=C1C=CC=C2)C(=O)N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CC(C)C)C(O)=O DVLHKUWLNKDINO-PMVMPFDFSA-N 0.000 description 1
- 101710162629 Trypsin inhibitor Proteins 0.000 description 1
- 229940122618 Trypsin inhibitor Drugs 0.000 description 1
- PEVVXUGSAKEPEN-AVGNSLFASA-N Tyr-Asn-Glu Chemical compound [H]N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CC(N)=O)C(=O)N[C@@H](CCC(O)=O)C(O)=O PEVVXUGSAKEPEN-AVGNSLFASA-N 0.000 description 1
- WPVGRKLNHJJCEN-BZSNNMDCSA-N Tyr-Asp-Phe Chemical compound C([C@H](N)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CC=1C=CC=CC=1)C(O)=O)C1=CC=C(O)C=C1 WPVGRKLNHJJCEN-BZSNNMDCSA-N 0.000 description 1
- CTDPLKMBVALCGN-JSGCOSHPSA-N Tyr-Gly-Val Chemical compound [H]N[C@@H](CC1=CC=C(O)C=C1)C(=O)NCC(=O)N[C@@H](C(C)C)C(O)=O CTDPLKMBVALCGN-JSGCOSHPSA-N 0.000 description 1
- NXRGXTBPMOGFID-CFMVVWHZSA-N Tyr-Ile-Asn Chemical compound [H]N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H]([C@@H](C)CC)C(=O)N[C@@H](CC(N)=O)C(O)=O NXRGXTBPMOGFID-CFMVVWHZSA-N 0.000 description 1
- DZKFGCNKEVMXFA-JUKXBJQTSA-N Tyr-Ile-His Chemical compound CC[C@H](C)[C@H](NC(=O)[C@@H](N)Cc1ccc(O)cc1)C(=O)N[C@@H](Cc1cnc[nH]1)C(O)=O DZKFGCNKEVMXFA-JUKXBJQTSA-N 0.000 description 1
- BSCBBPKDVOZICB-KKUMJFAQSA-N Tyr-Leu-Asp Chemical compound [H]N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CC(O)=O)C(O)=O BSCBBPKDVOZICB-KKUMJFAQSA-N 0.000 description 1
- KSCVLGXNQXKUAR-JYJNAYRXSA-N Tyr-Leu-Glu Chemical compound [H]N[C@@H](CC1=CC=C(O)C=C1)C(=O)N[C@@H](CC(C)C)C(=O)N[C@@H](CCC(O)=O)C(O)=O KSCVLGXNQXKUAR-JYJNAYRXSA-N 0.000 description 1
- SZEIFUXUTBBQFQ-STQMWFEESA-N Tyr-Pro-Gly Chemical compound [H]N[C@@H](CC1=CC=C(O)C=C1)C(=O)N1CCC[C@H]1C(=O)NCC(O)=O SZEIFUXUTBBQFQ-STQMWFEESA-N 0.000 description 1
- YOTRXXBHTZHKLU-BVSLBCMMSA-N Tyr-Trp-Met Chemical compound C([C@H](N)C(=O)N[C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H](CCSC)C(O)=O)C1=CC=C(O)C=C1 YOTRXXBHTZHKLU-BVSLBCMMSA-N 0.000 description 1
- JAQGKXUEKGKTKX-HOTGVXAUSA-N Tyr-Tyr Chemical compound C([C@H](N)C(=O)N[C@@H](CC=1C=CC(O)=CC=1)C(O)=O)C1=CC=C(O)C=C1 JAQGKXUEKGKTKX-HOTGVXAUSA-N 0.000 description 1
- PVPAOIGJYHVWBT-KKHAAJSZSA-N Val-Asn-Thr Chemical compound C[C@H]([C@@H](C(=O)O)NC(=O)[C@H](CC(=O)N)NC(=O)[C@H](C(C)C)N)O PVPAOIGJYHVWBT-KKHAAJSZSA-N 0.000 description 1
- TZVUSFMQWPWHON-NHCYSSNCSA-N Val-Asp-Leu Chemical compound CC(C)C[C@@H](C(=O)O)NC(=O)[C@H](CC(=O)O)NC(=O)[C@H](C(C)C)N TZVUSFMQWPWHON-NHCYSSNCSA-N 0.000 description 1
- VFOHXOLPLACADK-GVXVVHGQSA-N Val-Gln-Leu Chemical compound CC(C)C[C@@H](C(=O)O)NC(=O)[C@H](CCC(=O)N)NC(=O)[C@H](C(C)C)N VFOHXOLPLACADK-GVXVVHGQSA-N 0.000 description 1
- WBAJDGWKRIHOAC-GVXVVHGQSA-N Val-Lys-Gln Chemical compound [H]N[C@@H](C(C)C)C(=O)N[C@@H](CCCCN)C(=O)N[C@@H](CCC(N)=O)C(O)=O WBAJDGWKRIHOAC-GVXVVHGQSA-N 0.000 description 1
- QRVPEKJBBRYISE-XUXIUFHCSA-N Val-Lys-Ile Chemical compound CC[C@H](C)[C@@H](C(=O)O)NC(=O)[C@H](CCCCN)NC(=O)[C@H](C(C)C)N QRVPEKJBBRYISE-XUXIUFHCSA-N 0.000 description 1
- CFIBZQOLUDURST-IHRRRGAJSA-N Val-Tyr-Cys Chemical compound CC(C)[C@@H](C(=O)N[C@@H](CC1=CC=C(C=C1)O)C(=O)N[C@@H](CS)C(=O)O)N CFIBZQOLUDURST-IHRRRGAJSA-N 0.000 description 1
- 102000005789 Vascular Endothelial Growth Factors Human genes 0.000 description 1
- 108010019530 Vascular Endothelial Growth Factors Proteins 0.000 description 1
- 208000036142 Viral infection Diseases 0.000 description 1
- PVNFMCBFDPTNQI-UIBOPQHZSA-N [(1S,2R,5S,6S,16E,18E,20R,21S)-11-chloro-21-hydroxy-12,20-dimethoxy-2,5,9,16-tetramethyl-8,23-dioxo-4,24-dioxa-9,22-diazatetracyclo[19.3.1.110,14.03,5]hexacosa-10,12,14(26),16,18-pentaen-6-yl] acetate [(1S,2R,5S,6S,16E,18E,20R,21S)-11-chloro-21-hydroxy-12,20-dimethoxy-2,5,9,16-tetramethyl-8,23-dioxo-4,24-dioxa-9,22-diazatetracyclo[19.3.1.110,14.03,5]hexacosa-10,12,14(26),16,18-pentaen-6-yl] 3-methylbutanoate [(1S,2R,5S,6S,16E,18E,20R,21S)-11-chloro-21-hydroxy-12,20-dimethoxy-2,5,9,16-tetramethyl-8,23-dioxo-4,24-dioxa-9,22-diazatetracyclo[19.3.1.110,14.03,5]hexacosa-10,12,14(26),16,18-pentaen-6-yl] 2-methylpropanoate [(1S,2R,5S,6S,16E,18E,20R,21S)-11-chloro-21-hydroxy-12,20-dimethoxy-2,5,9,16-tetramethyl-8,23-dioxo-4,24-dioxa-9,22-diazatetracyclo[19.3.1.110,14.03,5]hexacosa-10,12,14(26),16,18-pentaen-6-yl] propanoate Chemical class CO[C@@H]1\C=C\C=C(C)\Cc2cc(OC)c(Cl)c(c2)N(C)C(=O)C[C@H](OC(C)=O)[C@]2(C)OC2[C@H](C)[C@@H]2C[C@@]1(O)NC(=O)O2.CCC(=O)O[C@H]1CC(=O)N(C)c2cc(C\C(C)=C\C=C\[C@@H](OC)[C@@]3(O)C[C@H](OC(=O)N3)[C@@H](C)C3O[C@@]13C)cc(OC)c2Cl.CO[C@@H]1\C=C\C=C(C)\Cc2cc(OC)c(Cl)c(c2)N(C)C(=O)C[C@H](OC(=O)C(C)C)[C@]2(C)OC2[C@H](C)[C@@H]2C[C@@]1(O)NC(=O)O2.CO[C@@H]1\C=C\C=C(C)\Cc2cc(OC)c(Cl)c(c2)N(C)C(=O)C[C@H](OC(=O)CC(C)C)[C@]2(C)OC2[C@H](C)[C@@H]2C[C@@]1(O)NC(=O)O2 PVNFMCBFDPTNQI-UIBOPQHZSA-N 0.000 description 1
- OUUYBRCCFUEMLH-YDALLXLXSA-N [(1s)-2-[4-[bis(2-chloroethyl)amino]phenyl]-1-carboxyethyl]azanium;chloride Chemical compound Cl.OC(=O)[C@@H](N)CC1=CC=C(N(CCCl)CCCl)C=C1 OUUYBRCCFUEMLH-YDALLXLXSA-N 0.000 description 1
- ZVIAAUGUGGKIDY-UHFFFAOYSA-N [Br].N1N=NN=C1.C1(=CC=CC=C1)C=1C=CC=CC1 Chemical compound [Br].N1N=NN=C1.C1(=CC=CC=C1)C=1C=CC=CC1 ZVIAAUGUGGKIDY-UHFFFAOYSA-N 0.000 description 1
- YRKCREAYFQTBPV-UHFFFAOYSA-N acetylacetone Chemical compound CC(=O)CC(C)=O YRKCREAYFQTBPV-UHFFFAOYSA-N 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 239000000654 additive Substances 0.000 description 1
- 230000000996 additive effect Effects 0.000 description 1
- 230000001464 adherent effect Effects 0.000 description 1
- 238000001042 affinity chromatography Methods 0.000 description 1
- 230000001270 agonistic effect Effects 0.000 description 1
- 108010008685 alanyl-glutamyl-aspartic acid Proteins 0.000 description 1
- 108010087924 alanylproline Proteins 0.000 description 1
- 229960005310 aldesleukin Drugs 0.000 description 1
- 108700025316 aldesleukin Proteins 0.000 description 1
- 229940100198 alkylating agent Drugs 0.000 description 1
- 239000002168 alkylating agent Substances 0.000 description 1
- 230000003281 allosteric effect Effects 0.000 description 1
- 229960003896 aminopterin Drugs 0.000 description 1
- 235000019270 ammonium chloride Nutrition 0.000 description 1
- 239000004037 angiogenesis inhibitor Substances 0.000 description 1
- 229940121369 angiogenesis inhibitor Drugs 0.000 description 1
- 230000001772 anti-angiogenic effect Effects 0.000 description 1
- 229940046836 anti-estrogen Drugs 0.000 description 1
- 230000001833 anti-estrogenic effect Effects 0.000 description 1
- 230000000340 anti-metabolite Effects 0.000 description 1
- 229940044684 anti-microtubule agent Drugs 0.000 description 1
- 229940125644 antibody drug Drugs 0.000 description 1
- 229940100197 antimetabolite Drugs 0.000 description 1
- 239000002256 antimetabolite Substances 0.000 description 1
- 108010086780 arginyl-glycyl-aspartyl-alanine Proteins 0.000 description 1
- 108010059459 arginyl-threonyl-phenylalanine Proteins 0.000 description 1
- 108010094001 arginyl-tryptophyl-arginine Proteins 0.000 description 1
- 108010093581 aspartyl-proline Proteins 0.000 description 1
- 238000010009 beating Methods 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- 102000005936 beta-Galactosidase Human genes 0.000 description 1
- 108010005774 beta-Galactosidase Proteins 0.000 description 1
- 229960000397 bevacizumab Drugs 0.000 description 1
- 230000002146 bilateral effect Effects 0.000 description 1
- 230000003115 biocidal effect Effects 0.000 description 1
- 230000008033 biological extinction Effects 0.000 description 1
- 229960002685 biotin Drugs 0.000 description 1
- 239000011616 biotin Substances 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 230000017531 blood circulation Effects 0.000 description 1
- 230000037058 blood plasma level Effects 0.000 description 1
- 238000007664 blowing Methods 0.000 description 1
- 230000037396 body weight Effects 0.000 description 1
- 210000001185 bone marrow Anatomy 0.000 description 1
- 229960001467 bortezomib Drugs 0.000 description 1
- 210000000481 breast Anatomy 0.000 description 1
- 238000004364 calculation method Methods 0.000 description 1
- 244000309466 calf Species 0.000 description 1
- 229930195731 calicheamicin Natural products 0.000 description 1
- HXCHCVDVKSCDHU-LULTVBGHSA-N calicheamicin Chemical compound C1[C@H](OC)[C@@H](NCC)CO[C@H]1O[C@H]1[C@H](O[C@@H]2C\3=C(NC(=O)OC)C(=O)C[C@](C/3=C/CSSSC)(O)C#C\C=C/C#C2)O[C@H](C)[C@@H](NO[C@@H]2O[C@H](C)[C@@H](SC(=O)C=3C(=C(OC)C(O[C@H]4[C@@H]([C@H](OC)[C@@H](O)[C@H](C)O4)O)=C(I)C=3C)OC)[C@@H](O)C2)[C@@H]1O HXCHCVDVKSCDHU-LULTVBGHSA-N 0.000 description 1
- 239000012830 cancer therapeutic Substances 0.000 description 1
- 229960004117 capecitabine Drugs 0.000 description 1
- 230000004856 capillary permeability Effects 0.000 description 1
- 125000003178 carboxy group Chemical group [H]OC(*)=O 0.000 description 1
- 239000013592 cell lysate Substances 0.000 description 1
- 230000001413 cellular effect Effects 0.000 description 1
- 230000005754 cellular signaling Effects 0.000 description 1
- 230000007541 cellular toxicity Effects 0.000 description 1
- 231100000045 chemical toxicity Toxicity 0.000 description 1
- 230000004087 circulation Effects 0.000 description 1
- 239000007979 citrate buffer Substances 0.000 description 1
- 238000012875 competitive assay Methods 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 239000013068 control sample Substances 0.000 description 1
- 238000007796 conventional method Methods 0.000 description 1
- 238000012258 culturing Methods 0.000 description 1
- 230000009089 cytolysis Effects 0.000 description 1
- 239000000824 cytostatic agent Substances 0.000 description 1
- 230000001085 cytostatic effect Effects 0.000 description 1
- HAAZLUGHYHWQIW-KVQBGUIXSA-N dGTP Chemical compound C1=NC=2C(=O)NC(N)=NC=2N1[C@H]1C[C@H](O)[C@@H](COP(O)(=O)OP(O)(=O)OP(O)(O)=O)O1 HAAZLUGHYHWQIW-KVQBGUIXSA-N 0.000 description 1
- 230000034994 death Effects 0.000 description 1
- 230000003111 delayed effect Effects 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 230000018109 developmental process Effects 0.000 description 1
- 238000002405 diagnostic procedure Methods 0.000 description 1
- 238000000502 dialysis Methods 0.000 description 1
- 238000009792 diffusion process Methods 0.000 description 1
- 230000008034 disappearance Effects 0.000 description 1
- 239000002552 dosage form Substances 0.000 description 1
- 231100000673 dose–response relationship Toxicity 0.000 description 1
- 238000004043 dyeing Methods 0.000 description 1
- 238000001976 enzyme digestion Methods 0.000 description 1
- 229940125532 enzyme inhibitor Drugs 0.000 description 1
- 210000002615 epidermis Anatomy 0.000 description 1
- 229930013356 epothilone Natural products 0.000 description 1
- HESCAJZNRMSMJG-KKQRBIROSA-N epothilone A Chemical class C/C([C@@H]1C[C@@H]2O[C@@H]2CCC[C@@H]([C@@H]([C@@H](C)C(=O)C(C)(C)[C@@H](O)CC(=O)O1)O)C)=C\C1=CSC(C)=N1 HESCAJZNRMSMJG-KKQRBIROSA-N 0.000 description 1
- 210000003743 erythrocyte Anatomy 0.000 description 1
- 239000000328 estrogen antagonist Substances 0.000 description 1
- 239000000284 extract Substances 0.000 description 1
- 210000002950 fibroblast Anatomy 0.000 description 1
- 229960000390 fludarabine Drugs 0.000 description 1
- GIUYCYHIANZCFB-FJFJXFQQSA-N fludarabine phosphate Chemical compound C1=NC=2C(N)=NC(F)=NC=2N1[C@@H]1O[C@H](COP(O)(O)=O)[C@@H](O)[C@@H]1O GIUYCYHIANZCFB-FJFJXFQQSA-N 0.000 description 1
- GNBHRKFJIUUOQI-UHFFFAOYSA-N fluorescein Chemical compound O1C(=O)C2=CC=CC=C2C21C1=CC=C(O)C=C1OC1=CC(O)=CC=C21 GNBHRKFJIUUOQI-UHFFFAOYSA-N 0.000 description 1
- 229940060037 fluorine Drugs 0.000 description 1
- 238000011010 flushing procedure Methods 0.000 description 1
- 238000009472 formulation Methods 0.000 description 1
- 238000004108 freeze drying Methods 0.000 description 1
- 239000005338 frosted glass Substances 0.000 description 1
- 210000001035 gastrointestinal tract Anatomy 0.000 description 1
- 238000012637 gene transfection Methods 0.000 description 1
- 208000005017 glioblastoma Diseases 0.000 description 1
- 239000008103 glucose Substances 0.000 description 1
- 108010067216 glycyl-glycyl-glycine Proteins 0.000 description 1
- XLXSAKCOAKORKW-UHFFFAOYSA-N gonadorelin Chemical compound C1CCC(C(=O)NCC(N)=O)N1C(=O)C(CCCN=C(N)N)NC(=O)C(CC(C)C)NC(=O)CNC(=O)C(NC(=O)C(CO)NC(=O)C(CC=1C2=CC=CC=C2NC=1)NC(=O)C(CC=1NC=NC=1)NC(=O)C1NC(=O)CC1)CC1=CC=C(O)C=C1 XLXSAKCOAKORKW-UHFFFAOYSA-N 0.000 description 1
- 239000002474 gonadorelin antagonist Substances 0.000 description 1
- MFWNKCLOYSRHCJ-BTTYYORXSA-N granisetron Chemical compound C1=CC=C2C(C(=O)N[C@H]3C[C@H]4CCC[C@@H](C3)N4C)=NN(C)C2=C1 MFWNKCLOYSRHCJ-BTTYYORXSA-N 0.000 description 1
- 229960003727 granisetron Drugs 0.000 description 1
- 239000003324 growth hormone secretagogue Substances 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 239000003652 hormone inhibitor Substances 0.000 description 1
- 102000057593 human F8 Human genes 0.000 description 1
- 235000003642 hunger Nutrition 0.000 description 1
- 239000012216 imaging agent Substances 0.000 description 1
- 230000028993 immune response Effects 0.000 description 1
- 102000018358 immunoglobulin Human genes 0.000 description 1
- 239000002955 immunomodulating agent Substances 0.000 description 1
- 229940121354 immunomodulator Drugs 0.000 description 1
- 230000002584 immunomodulator Effects 0.000 description 1
- 239000012133 immunoprecipitate Substances 0.000 description 1
- 238000002513 implantation Methods 0.000 description 1
- 238000000338 in vitro Methods 0.000 description 1
- 238000011503 in vivo imaging Methods 0.000 description 1
- 238000011081 inoculation Methods 0.000 description 1
- 229940125396 insulin Drugs 0.000 description 1
- 102000028416 insulin-like growth factor binding Human genes 0.000 description 1
- 108091022911 insulin-like growth factor binding Proteins 0.000 description 1
- 230000003993 interaction Effects 0.000 description 1
- 230000031146 intracellular signal transduction Effects 0.000 description 1
- 229940084651 iressa Drugs 0.000 description 1
- 229910052742 iron Inorganic materials 0.000 description 1
- 108010031424 isoleucyl-prolyl-proline Proteins 0.000 description 1
- 230000009916 joint effect Effects 0.000 description 1
- 108010090333 leucyl-lysyl-proline Proteins 0.000 description 1
- 108010073472 leucyl-prolyl-proline Proteins 0.000 description 1
- 108010012058 leucyltyrosine Proteins 0.000 description 1
- 239000003446 ligand Substances 0.000 description 1
- 239000008176 lyophilized powder Substances 0.000 description 1
- 230000002934 lysing effect Effects 0.000 description 1
- 239000012139 lysis buffer Substances 0.000 description 1
- 239000003550 marker Substances 0.000 description 1
- 230000008774 maternal effect Effects 0.000 description 1
- 230000007246 mechanism Effects 0.000 description 1
- 230000010534 mechanism of action Effects 0.000 description 1
- HAWPXGHAZFHHAD-UHFFFAOYSA-N mechlorethamine Chemical class ClCCN(C)CCCl HAWPXGHAZFHHAD-UHFFFAOYSA-N 0.000 description 1
- 108020004999 messenger RNA Proteins 0.000 description 1
- 230000002503 metabolic effect Effects 0.000 description 1
- 229910021645 metal ion Inorganic materials 0.000 description 1
- MYWUZJCMWCOHBA-VIFPVBQESA-N methamphetamine Chemical compound CN[C@@H](C)CC1=CC=CC=C1 MYWUZJCMWCOHBA-VIFPVBQESA-N 0.000 description 1
- 239000003226 mitogen Substances 0.000 description 1
- 230000002297 mitogenic effect Effects 0.000 description 1
- 229960004857 mitomycin Drugs 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 230000009149 molecular binding Effects 0.000 description 1
- 238000010369 molecular cloning Methods 0.000 description 1
- 238000012544 monitoring process Methods 0.000 description 1
- 239000000178 monomer Substances 0.000 description 1
- 210000003205 muscle Anatomy 0.000 description 1
- 201000000050 myeloid neoplasm Diseases 0.000 description 1
- 230000017095 negative regulation of cell growth Effects 0.000 description 1
- 238000006386 neutralization reaction Methods 0.000 description 1
- 230000003472 neutralizing effect Effects 0.000 description 1
- 230000001254 nonsecretory effect Effects 0.000 description 1
- 238000011580 nude mouse model Methods 0.000 description 1
- 238000006384 oligomerization reaction Methods 0.000 description 1
- 230000003287 optical effect Effects 0.000 description 1
- 229940026778 other chemotherapeutics in atc Drugs 0.000 description 1
- 229940055729 papain Drugs 0.000 description 1
- 235000019834 papain Nutrition 0.000 description 1
- 239000000137 peptide hydrolase inhibitor Substances 0.000 description 1
- 229960003531 phenolsulfonphthalein Drugs 0.000 description 1
- 108010012581 phenylalanylglutamate Proteins 0.000 description 1
- 208000028591 pheochromocytoma Diseases 0.000 description 1
- 239000008363 phosphate buffer Substances 0.000 description 1
- 102000020233 phosphotransferase Human genes 0.000 description 1
- 230000004962 physiological condition Effects 0.000 description 1
- 230000035790 physiological processes and functions Effects 0.000 description 1
- 239000006187 pill Substances 0.000 description 1
- 210000002381 plasma Anatomy 0.000 description 1
- 229920001223 polyethylene glycol Polymers 0.000 description 1
- 229920000151 polyglycol Polymers 0.000 description 1
- 239000010695 polyglycol Substances 0.000 description 1
- 239000000256 polyoxyethylene sorbitan monolaurate Substances 0.000 description 1
- 235000010486 polyoxyethylene sorbitan monolaurate Nutrition 0.000 description 1
- 239000011148 porous material Substances 0.000 description 1
- 239000002244 precipitate Substances 0.000 description 1
- XJMOSONTPMZWPB-UHFFFAOYSA-M propidium iodide Chemical compound [I-].[I-].C12=CC(N)=CC=C2C2=CC=C(N)C=C2[N+](CCC[N+](C)(CC)CC)=C1C1=CC=CC=C1 XJMOSONTPMZWPB-UHFFFAOYSA-M 0.000 description 1
- 235000019633 pungent taste Nutrition 0.000 description 1
- 239000012521 purified sample Substances 0.000 description 1
- 229950010131 puromycin Drugs 0.000 description 1
- 238000012207 quantitative assay Methods 0.000 description 1
- 238000000163 radioactive labelling Methods 0.000 description 1
- 238000001959 radiotherapy Methods 0.000 description 1
- 239000011541 reaction mixture Substances 0.000 description 1
- 229940047431 recombinate Drugs 0.000 description 1
- 230000009467 reduction Effects 0.000 description 1
- 230000008929 regeneration Effects 0.000 description 1
- 238000011069 regeneration method Methods 0.000 description 1
- 230000001105 regulatory effect Effects 0.000 description 1
- 239000003488 releasing hormone Substances 0.000 description 1
- 230000008521 reorganization Effects 0.000 description 1
- 239000011347 resin Substances 0.000 description 1
- 229920005989 resin Polymers 0.000 description 1
- 238000003757 reverse transcription PCR Methods 0.000 description 1
- 238000012552 review Methods 0.000 description 1
- PYWVYCXTNDRMGF-UHFFFAOYSA-N rhodamine B Chemical compound [Cl-].C=12C=CC(=[N+](CC)CC)C=C2OC2=CC(N(CC)CC)=CC=C2C=1C1=CC=CC=C1C(O)=O PYWVYCXTNDRMGF-UHFFFAOYSA-N 0.000 description 1
- 239000012266 salt solution Substances 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 238000002864 sequence alignment Methods 0.000 description 1
- 230000008771 sex reversal Effects 0.000 description 1
- 235000017557 sodium bicarbonate Nutrition 0.000 description 1
- 229910000030 sodium bicarbonate Inorganic materials 0.000 description 1
- 235000017550 sodium carbonate Nutrition 0.000 description 1
- 229910000029 sodium carbonate Inorganic materials 0.000 description 1
- 235000013024 sodium fluoride Nutrition 0.000 description 1
- 239000011775 sodium fluoride Substances 0.000 description 1
- 239000001488 sodium phosphate Substances 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 238000003153 stable transfection Methods 0.000 description 1
- 230000037351 starvation Effects 0.000 description 1
- 238000007619 statistical method Methods 0.000 description 1
- 230000004936 stimulating effect Effects 0.000 description 1
- 238000007920 subcutaneous administration Methods 0.000 description 1
- 230000009182 swimming Effects 0.000 description 1
- 208000024891 symptom Diseases 0.000 description 1
- 230000002195 synergetic effect Effects 0.000 description 1
- 210000001258 synovial membrane Anatomy 0.000 description 1
- 150000004579 taxol derivatives Chemical class 0.000 description 1
- 125000002456 taxol group Chemical group 0.000 description 1
- 125000003396 thiol group Chemical group [H]S* 0.000 description 1
- 231100000331 toxic Toxicity 0.000 description 1
- 230000002588 toxic effect Effects 0.000 description 1
- 230000001988 toxicity Effects 0.000 description 1
- 231100000419 toxicity Toxicity 0.000 description 1
- 238000012546 transfer Methods 0.000 description 1
- 230000001131 transforming effect Effects 0.000 description 1
- 230000001052 transient effect Effects 0.000 description 1
- 229960000575 trastuzumab Drugs 0.000 description 1
- RYFMWSXOAZQYPI-UHFFFAOYSA-K trisodium phosphate Chemical compound [Na+].[Na+].[Na+].[O-]P([O-])([O-])=O RYFMWSXOAZQYPI-UHFFFAOYSA-K 0.000 description 1
- 229910000406 trisodium phosphate Inorganic materials 0.000 description 1
- 235000019801 trisodium phosphate Nutrition 0.000 description 1
- IHIXIJGXTJIKRB-UHFFFAOYSA-N trisodium vanadate Chemical compound [Na+].[Na+].[Na+].[O-][V]([O-])([O-])=O IHIXIJGXTJIKRB-UHFFFAOYSA-N 0.000 description 1
- 239000002753 trypsin inhibitor Substances 0.000 description 1
- OUYCCCASQSFEME-UHFFFAOYSA-N tyrosine Natural products OC(=O)C(N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-UHFFFAOYSA-N 0.000 description 1
- 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 1
- 108010020532 tyrosyl-proline Proteins 0.000 description 1
- 108010003137 tyrosyltyrosine Proteins 0.000 description 1
- 239000003981 vehicle Substances 0.000 description 1
- 229960004276 zoledronic acid Drugs 0.000 description 1
Images
Classifications
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
- C07K16/18—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans
- C07K16/28—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
- C07K16/18—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans
- C07K16/28—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants
- C07K16/2863—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants against receptors for growth factors, growth regulators
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K39/395—Antibodies; Immunoglobulins; Immune serum, e.g. antilymphocytic serum
- A61K39/39533—Antibodies; Immunoglobulins; Immune serum, e.g. antilymphocytic serum against materials from animals
- A61K39/39541—Antibodies; Immunoglobulins; Immune serum, e.g. antilymphocytic serum against materials from animals against normal tissues, cells
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K16/00—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies
- C07K16/18—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K2039/505—Medicinal preparations containing antigens or antibodies comprising antibodies
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K2317/00—Immunoglobulins specific features
- C07K2317/20—Immunoglobulins specific features characterized by taxonomic origin
- C07K2317/24—Immunoglobulins specific features characterized by taxonomic origin containing regions, domains or residues from different species, e.g. chimeric, humanized or veneered
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K2317/00—Immunoglobulins specific features
- C07K2317/50—Immunoglobulins specific features characterized by immunoglobulin fragments
- C07K2317/55—Fab or Fab'
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K2317/00—Immunoglobulins specific features
- C07K2317/50—Immunoglobulins specific features characterized by immunoglobulin fragments
- C07K2317/56—Immunoglobulins specific features characterized by immunoglobulin fragments variable (Fv) region, i.e. VH and/or VL
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K2317/00—Immunoglobulins specific features
- C07K2317/50—Immunoglobulins specific features characterized by immunoglobulin fragments
- C07K2317/56—Immunoglobulins specific features characterized by immunoglobulin fragments variable (Fv) region, i.e. VH and/or VL
- C07K2317/565—Complementarity determining region [CDR]
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K2317/00—Immunoglobulins specific features
- C07K2317/70—Immunoglobulins specific features characterized by effect upon binding to a cell or to an antigen
- C07K2317/76—Antagonist effect on antigen, e.g. neutralization or inhibition of binding
Landscapes
- Health & Medical Sciences (AREA)
- Chemical & Material Sciences (AREA)
- Immunology (AREA)
- Organic Chemistry (AREA)
- Life Sciences & Earth Sciences (AREA)
- Medicinal Chemistry (AREA)
- General Health & Medical Sciences (AREA)
- Genetics & Genomics (AREA)
- Biophysics (AREA)
- Molecular Biology (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Biochemistry (AREA)
- Engineering & Computer Science (AREA)
- Bioinformatics & Cheminformatics (AREA)
- Public Health (AREA)
- Microbiology (AREA)
- Mycology (AREA)
- Pharmacology & Pharmacy (AREA)
- Epidemiology (AREA)
- Animal Behavior & Ethology (AREA)
- Biomedical Technology (AREA)
- Veterinary Medicine (AREA)
- Peptides Or Proteins (AREA)
- Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
- Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
- Acyclic And Carbocyclic Compounds In Medicinal Compositions (AREA)
- Saccharide Compounds (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
可特异结合并抑制胰岛素样生长因子-I受体的抗体、人源化抗体、表面重构抗体、抗体片段、衍生抗体和上述抗体与细胞毒性试剂形成的偶联物,它们可拮抗IGF-I、IGF-II和血清对肿瘤细胞生长和存活的效应,并且它们实质上不存在激动剂活性。任选地与其他治疗剂联合,所述的抗体及其片段可以用于治疗IGF-I受体表达水平升高的肿瘤,如乳癌、结肠癌、肺癌、卵巢癌、滑膜肉瘤、前列腺癌和胰腺癌,所述的衍生抗体可用于IGF-I受体表达水平升高的肿瘤的诊断和成像。
Description
[01]本发明是2002年6月14提交的母申请10/170,390的部分继续申请,该母申请通过参考整体并入本文。
发明领域
[02]本发明涉及与人胰岛素样生长因子-I受体(IGF-I受体)结合的抗体。更具体地说,本发明涉及可抑制IGF-I受体的细胞功能的抗IGF-I受体抗体。更具体地,本发明涉及可拮抗IGF-I、IGF-II和血清对肿瘤细胞生长和存活的作用的抗体,它们基本上缺乏激动剂活性。本发明也涉及所述抗体的片段,所述抗体的人源化(humanized)和表面重构(resurfaced)的形式,所述抗体的偶联物(conjugates),抗体衍生物(derivatives),和它们在诊断、研究和治疗应用中的用途。本发明进一步涉及改良的抗体或其片段,它们可从上述的抗体及其片段制成。在另一个方面,本发明涉及编码所述抗体或其片段的多聚核苷酸,以及含有所述多聚核苷酸的载体。
发明背景
[03]胰岛素样生长因子-I受体(IGF-I受体)是一种跨膜异源四聚体蛋白,其具有两个细胞外α链和两个跨膜β链,通过二硫键连接成β-α-α-β构型。胰岛素样生长因子-I(IGF-I)和胰岛素样生长因子-II(IGF-II)作为配体通过IGF-I受体的细胞外结构域结合到其上,从而刺激其细胞内的酪氨酸激酶结构域,引起该受体的自体磷酸化(autophosphorylation)和底物磷酸化。IGF-I受体与胰岛素受体同源,在β链的酪氨酸激酶结构域中有84%的高度序列相似性,在α链的细胞外富含半胱氨酸的结构域中有48%的较低序列相似性(Ulrich,A.等,1986,EMBO,5,2503-2512;Fujita-Yamaguchi,Y.等,1986,J.Biol.Chem.,261,16727-16731;LeRoith,D.等,1995,Endocrine Reviews,16,143-163)。IGF-I受体及其配体(IGF-I和IGF-II)在大量的生理学过程中扮演者重要角色,包括在胚胎发生过程中的生长和发育、代谢、成体的细胞增殖和细胞分化(LeRoith,D.,2000,Endocrinology,141,1287-1288;LeRoith,D.,1997,New England J.Med.,336,633-640)。[04]IGF-I和IGF-II均可在血液中作为内分泌激素而发挥作用,此时它们主要与IGF结合蛋白以复合物的形式存在,IGF-I和IGF-II还可作为局部产生的旁分泌和自分泌生长因子而发挥作用(Humbel,R.E.,1990,Eur.J.Biochem.,190,445-462;Cohick,W.S.和Clemmons,D.R.,1993,Annu.Rev.Physiol.55,131-153)。
[05]IGF-I受体被认为涉及促进肿瘤细胞的生长、转化和存活(Baserga,R.等,1997,Biochem.Biophys.Acta,1332,F105-F126;Blakesley,V.A.等,1997,Journal of Endocrinology,152,339-344;Kaleko,M.,Rutter,W.J.和Miller,A.D.1990,Mol.Cell.Biol,10,464-473)。因此,已经知道数种类型的肿瘤表达的IGF-I受体水平较正常水平高,包括乳癌、结肠癌、卵巢癌、滑膜肉瘤和胰腺癌(Khandwala,H.M.等,2000,EndocrineReviews,21,215-244;Werner,H.和LeRoith,D.,1996,Adv.Cancer Res.,68,183-223;Happerfield,L.C.等,1997,J.Pathol,183,412-417;Frier,S.等,1999,Gut,44,704-708;van Dam,P.A.等,1994,J.Clin.Pathol.,47,914-919;Xie,Y.等,1999,Cancer Res.,59,3588-3591;Bergmann,U.等,1995,Cancer Res.,55,2007-2011)。在体外,IGF-I和IGF-II显示可以作为数种人类肿瘤细胞系有效的有丝分裂原,如肺癌、乳癌、结肠癌、骨肉瘤和宫颈癌(Ankrapp,D.P.和Bevan,D.R.,1993,Cancer Res.,53,3399-3404;Cullen,K.J.,1990,Cancer Res.,50,48-53;Hermanto,U.等,2000,Cell Growth & Differentiation,11,655-664;Guo,Y.S.等,1995,J.Am.Coll.Surg.,181,145-154;Kappel,C.C.等,1994,Cancer Res.,54,2803-2807;Steller,M.A.等,1996,Cancer Res.,56,1761-1765)。这些肿瘤和肿瘤细胞系中的数种也可表达高水平的IGF-I或IGF-II,它们可以自分泌或旁分泌的方式刺激其生长(Quinn,K.A.等,1996,J.Biol.Chem.,271,11477-11483)。
[06]流行病学研究已经显示,增高的IGF-I血浆水平(和IGF结合蛋白-3的较低水平)与前列腺癌、结肠癌、肺癌和乳癌的危险性增加有关(Chan,J.M.等,1998,Science,279,563-566;Wolk,A.等,1998,J.Natl.Cancer Inst.,90,911-915;Ma,J.等,1999,J.Natl.Cancer Inst.,91,620-625;Yu,H.等,1999,J.Natl.Cancer Inst.,91,151-156;Hankinson,S.E.等,1998,Lancet,351,1393-1396)。已经有建议,可以采用降低血浆中IGF-I水平或抑制IGF-I受体功能的策略来预防癌症(Wu,Y.等,2002,Cancer Res.,62,1030-1035;Grimberg,A和Cohen P,2000,J.Cell.Physiol,183,1-9)。
[07]IGF-I受体可保护肿瘤细胞免受生长因子缺失(growth factordeprivation),贴壁非依赖性(anchorage-independence)或细胞毒性药物处理所引起的凋亡(Navarro,M.和Baserga,R.,2001,Endocrinology,142,1073-1081;Baserga,R.等,1997,Biochem.Biophys.Acta,1332,F105-F126)。对其促有丝分裂,转化和抗凋亡活性至关重要的IGF-I受体结构域已经通过突变分析被鉴定出来。
[08]例如,IGF-I受体的酪氨酸1251残基被鉴定对于抗凋亡和转化活性非常重要,但与其促有丝分裂活性无关(O′Connor,R.等,1997,Mol.Cell.Biol,17,427-435;Miura,M.等,1995,J.Biol.Chem.,270,22639-22644)。通过配体活化的IGF-I受体的细胞内信号通路涉及胰岛素受体底物(IRS-1和IRS-2)的酪氨酸残基的磷酸化,它们可将磷脂酰肌醇-3-激酶(PI-3-激酶)招集(recruit)至膜上。PI-3激酶的膜结合磷脂产物可活化丝氨酸/苏氨酸激酶Akt,其底物包括促凋亡蛋白BAD,它可被磷酸化成为非活化状态(Datta,S.R.,Brunet,A.和Greenberg,M.E.,1999,Genes & Development,13,2905-2927;Kulik,G,Klippel,A.和Weber,M.J.,1997,Mol.Cell.Biol 17,1595-1606)。在MCF-7人乳癌细胞中IGF-I受体的促有丝分裂信号转导需要PI-3-激酶,不依赖有丝分裂原活化的蛋白激酶,而在分化的大鼠嗜铬细胞瘤PC12细胞中的存活信号转导需要PI-3-激酶和有丝分裂原活化的蛋白激酶通路(Dufourny,B.等,1997,J.Biol.Chem.,212,31163-31171;Parrizas,M.,Saltiel,A.R.和LeRoith,D.,1997,J.Biol.Chem.,272,154-161)。
[09]已经显示,通过反义(anti-sense)策略下调IGF-I受体可降低几种肿瘤细胞系在体内和体外的致肿瘤性,如黑色素瘤、肺癌、卵巢癌、成胶质细胞瘤、成神经细胞瘤和横纹肌肉瘤(Resnicoff,M.等,1994,Cancer Res.,54,4848-4850;Lee,C.-T.等,1996,Cancer Res.,56,3038-3041;Muller,M.等,1998,Int.J.Cancer,77,567-571;Trojan,J.等,1993,Science,259,94-97;Liu,X.等,1998,Cancer Res.,58,5432-5438;Shapiro,D.N.等,1994,J.Clin.Invest.,94,1235-1242)。而且,据报道,IGF-I受体的显性负突变体(dominant negative mutant)可降低过度表达IGF-I受体的转化Rat-1细胞在体内的致肿瘤性和体外的生长(Prager,D.等,1994,Proc.Natl.Acad.Sci.USA,91,2181-2185)。
[10]表达IGF-I受体mRNA的反义分子的肿瘤细胞当被注射进入动物中时,在生物扩散腔(biodiffusion chambers)中会发生大量的凋亡。这种观察使得IGF-I受体成为一种有吸引力的治疗靶标,这是基于这样的假设,即通过抑制IGF-I受体可使肿瘤细胞较正常细胞更易凋亡(Resnicoff,M.等,1995,Cancer Res.,55,2463-2469;Baserga,R.,1995,Cancer Res.,55,249-252)。
[11]在肿瘤细胞中抑制IGF-I受体的功能的另一个策略是应用抗IGF-I受体的抗体,所述抗体可与IGF-I受体的细胞外结构域结合,并抑制其活化。据报道,已经进行了几种尝试来开发抗IGF-I受体的鼠单克隆抗体,其中获得了两种抑制性抗体IR3和1H7,它们的应用已经在几个IGF-I受体研究中被报道。
[12]应用部分纯化的胎盘胰岛素受体制备物来免疫小鼠,开发IR3抗体,这产生选择性地结合胰岛素受体的抗体IR1,以及两种抗体IR2和IR3,这两种抗体显示出对IGF-I受体(生长调节素-C受体)的优先沉淀,但也能微弱地沉淀胰岛素受体,由此开发出IR3抗体(Kull,F.C.等,1983,J.Biol.Chem.,258,6561-6566)。
[13]通过用纯化的胎盘IGF-1受体制备物免疫小鼠开发出1H7抗体,在这个过程中除了产生三种刺激性抗体外,还产生抑制性抗体1H7(Li,S.-L.等,1993,Biochem.Biophys.Res.Commun.,196,92-98;Xiong,L.等,1992,Proc.Natl.Acad.Sci,USA,89,5356-5360)。
[14]在另一个报道中,通过用表达高水平的IGF-I受体的转染3T3细胞免疫小鼠,获得一系列特异针对人IGF-I受体的鼠单克隆抗体,通过结合竞争研究及其对IGF-I与转染3T3细胞结合的抑制或刺激可将它们分为7类(Soos,M.A.等,1992,J.Biol.Chem.,267,12955-12963)。
[15]因此,尽管IR3抗体是体外IGF-I受体研究中最常用的抑制性抗体,但它有一些缺点,即它对表达人IGF-I受体的转染3T3和CHO细胞显示有激动活性(agonistic activity)(Kato,H.等,1993,J.Biol.Chem.,268,2655-2661;Steele-Perkins,G.和Roth,R.A.,1990,Biochem.Biophys.Res.Commun.,171,1244-1251)。同样,在由Soos等人开发的一系列抗体中,最具抑制性的抗体24-57和24-60也在转染3T3细胞中显示出激动活性(Soos,M.A.等,1992,J.Biol.Chem.,267,12955-12963)。尽管,据报道IR3抗体可抑制IGF-I(但不是IGF-II)与完整细胞中和溶解后细胞中表达的受体的结合,它显示出在体外抑制IGF-I和IGF-II刺激细胞中DNA合成的能力(Steele-Perkins,G.和Roth,R.A.,1990,Biochem.Biophys.Res.Commun.,171,1244-1251)。IR3抗体的结合表位已经从嵌合的胰岛素-IGF-I受体构建物中推导出来,是IGF-I受体的223-274区域(Gustafson,T.A.和Rutter,W. J.,1990,J.Biol.Chem.,265,18663-18667;Soos,M.A.等,1992,J.Biol.Chem.,267,12955-12963)。
[16]MCF-7人乳癌细胞系通常被用作模型细胞系来证明体外IGF-I和IGF-II的生长响应(Dufoumy,B.等,1997,J.Biol.Chem.,272,31163-31171)。在MCF-7细胞中,IR3抗体可不完全地阻断在无血清的条件下外源性添加的IGF-I和IGF-II的刺激效应的大约80%。而且,IR3抗体在10%的血清中并不很明显地抑制(小于25%)MCF-7细胞的生长(Cullen,K.J.等,1990,Cancer Res.,50,48-53)。在体外,IR3抗体对血清刺激的MCF-7细胞生长的这种微弱抑制作用与体内研究的结果一致,在体内,IR3抗体的处理并不能明显抑制裸鼠中MCF-7异种移植物的生长(Arteaga,C.L.等,1989,J.Clin.Invest.,84,1418-1423)。
[17]由于IR3和其他已报道的抗体的微弱激动活性,以及它们不能明显抑制肿瘤细胞如MCF-7细胞在血清刺激这样更加生理化的条件下生长(这里所述的血清刺激不同于在无血清条件下外源添加的IGF-I或IGF-II的刺激作用),因此需要能够明显抑制肿瘤细胞的血清刺激性生长,但其本身不显示出明显的激动剂活性的新的抗IGF-I受体抗体。
发明概述
[18]因此,本发明的目的是提供可特异结合胰岛素样生长因子-I受体并通过拮抗该受体而抑制该受体的细胞活性的抗体、抗体片段和抗体衍生物,它们实质上也不具有对该受体的激动剂活性。
[19]因此,在第一个实施方案中,提供了鼠抗体EM 164,在此对其进行了完整的表征,即其轻链和重链可变区的氨基酸序列,轻链和重链可变区的基因的cDNA序列,其CDRs(互补决定区)的鉴定,其表面氨基酸的鉴定,和将其以重组形式进行表达的手段。
[20]在第二个实施方案中,提供了重构表面或人源化的抗体EM 164,其中所述抗体或其片段的暴露于表面的残基在轻链和重链中被替换以便更接近地类似于已知的人抗体表面。这种人源化抗体与鼠EM164相比,在作为治疗或诊断试剂方面的实用性要更强。人源化的抗体EM164的特征在此也完全被揭示,包括其轻链和重链可变区各自的氨基酸序列,轻链和重链可变区的基因的DNA序列,CDRs的鉴定,其表面氨基酸的鉴定,和其以重组形式进行表达的方法的公开。
[21]在第三个实施方案中,提供了一种抗体,它能够在生长刺激物存在的情况下抑制癌症细胞的生长超过大约80%,所述刺激物例如血清、胰岛素样生长因子-I和胰岛素-样生长因子-II。
[22]在第四个实施方案中,提供了一种抗体或抗体片段,其具有重链和轻链,所述重链包含分别具有SEQ ID NOS:1-3中所示的氨基酸序列的CDRs:
SYWMH(SEQ ID NO:1),
EINPSNGRTNYNEKFKR(SEQ ID NO:2),
GRPDYYGSSKWYFDV(SEQ ID NO:3);
所述轻链包含具有SEQ ID NOS:4-6中所示的氨基酸序列的CDRs:
RSSQSIVHSNVNTYLE(SEQ ID NO:4);
KVSNRFS(SEQ ID NO:5);
FQGSHVPPT(SEQ ID NO:6)。
[23]在第五个实施方案中,提供了抗体,其重链具有的氨基酸序列与SEQ ID NO:7中所示的氨基酸序列具有至少90%的序列同一性(identity),SEQ ID NO:7为:
QVQLQQSGAELVKPGASVKLSCKASGYTFTSYWMHWVKQRPGQGLEWIGEINPSNGRTNYNEKFKRKATLTVDKSSSTAYMQLSSLTSEDSAVYYFARGRPDY YGSSKWYFDVWGAGTTVTVSS(SEQ IDNO:7)。
[24]类似地,提供了具有轻链的抗体,所述轻链具有与SEQ ID NO:8描述的氨基酸序列具有至少90%的序列同一性的氨基酸序列,SEQ IDNO:8为:
DVLMTQTPLSLPVSLGDQASISCRSSQSIVHSNVNTYLEWYLQKPGQSPKLLIYKVSNRFSGVPDRFSGSGSGTDFTLRISRVEAEDLGIYYCFQGSHVPPTFGGGTKLEIKR(SEQ ID NO:8)。
[25]在第六个实施方案中,提供了具有人源化或重构表面的轻链可变区的抗体,所述轻链可变区具有对应于SEQ ID NOS:-9-12中的其中一个的氨基酸序列:
DVVMTQTPLSLPVSLGDPASISCRSSQSIVHSNVNTYLEWYLQKPGQSPRLLIYKVSNRFSGVPDRFSGSGAGTDFTLRISRVEAEDLGIYYCFQGSHVPPTFGGGTKLEIKR(SEQ ID NO:9);DVLMTQTPLSLPVSLGDPASISCRSSQSIVHSNVNTYLEWYLQKPGQSPKLLIYKVSNRFSGVPDRFSGSGAGTDFTLRISRVEAEDLGIYYCFQGSHVPPTFGGGTKLEIKR(SEQ ED NO:10);
DVLMTQTPLSLPVSLGDPASISCRSSQSIVHSNVNTYLEWYLQKPGQSPRLLIYKVSNRFSGVPDRFSGSGAGTDFTLRISRVEAEDLGIYYCFQGSHVPPTFGGGTKLEIKR(SEQ ID NO:11);或DVVMTQTPLSLPVSLGDPASISCRSSQSIVHSNVNTYLEWYLQKPGQSPKLLIYKVSNRFSGVPDRFSGSGAGTDFTLRISRVEAEDLGIYYCFQGSHVPPTFGGGTKLEIKR(SEQ ID NO:12)。
[26]类似地,提供了抗体,其具有人源化或重构表面的重链可变区,该区域具有与SEQ ID NO:13对应的氨基酸序列:
QVQLVQSGAEVVKPGASVKLSCKASGYTFTSYWMHWVKQRPGQGLEWIGEINPSNGRTNYNQKFQGKATLTVDKSSSTAYMQLSSLTSEDSAVYYFARGRPDYYGSSKWYFDVWGQGTTVTVSS(SEQ IDNO:13)。
[27]在第七个实施方案中,提供了具有改良的特性的本发明抗体或抗体片断。例如,对IGF-I受体具有更强的亲和性的抗体或抗体片段可通过本发明的抗体或片段的亲和力成熟(affinity maturation)来制备。
[28]本发明进一步提供了所述抗体的偶联物,其中细胞毒性试剂直接地或者通过一个可切割的或不可切割的连接物(linker),被共价连接于本发明的抗体或抗体的表位(epitope)结合片段。在优选的实施方案中,所述细胞毒性试剂是紫杉醇、美登木素类生物碱(maytansinoid)、CC-1065或CC-1065类似物。
[29]本发明进一步提供了进一步被标记的可在研究或诊断应用中被使用的抗体或其片段。在优选的实施方案中,所述标记是放射标记、荧光团(fluorophore)、生色团(chromophore)、显影剂(imaging agent)或金属离子。
[30]也提供了一种诊断方法,其中所述的被标记的抗体或片段被施于怀疑患有癌症的对象,并测定或监测对象体内的标记物的分布。
[31]在第八个实施方案中,本发明提供了通过将本发明的抗体,抗体片段或抗体偶联物单独施用或者与其他细胞毒性试剂或治疗试剂联合施用来治疗患有癌症的对象的方法。所述癌症可以是下列的一种或者多种,例如乳癌、结肠癌、卵巢癌、骨肉瘤、宫颈癌、前列腺癌、肺癌、滑膜肉瘤、前列腺癌或其他还待被确定其中IGF-I受体水平升高的癌症。
[32]在第九个实施方案中,本发明提供了通过或者单独或与其他细胞毒性剂或治疗剂组合,给予本发明的抗体、抗体片段或抗体偶联物,来治疗患有癌症的对象的方法。特别地,优选的细胞毒剂和治疗剂包括多烯紫杉醇(docetaxel)、紫杉酚(paclitaxel)、阿霉素、表阿霉素、环磷酰胺、曲妥珠(trastuzumab)(Herceptin)、卡培他宾(capecitabine)、它莫西芬、托瑞米芬、来曲唑、阿纳曲唑、氟维司群(fulvestrant)、依西美坦、戈舍瑞林(goserelin)、奥沙利铂(oxaliplatin)、卡铂、顺铂、地塞米松、抗排卵肽、贝伐单抗(bevacizumab)(Avastin)、5-氟尿嘧啶、亚叶酸、左旋咪唑、依立替康、依托泊甙、拓朴替康(topotecan)、吉西他滨、长春瑞滨、雌莫司汀、米托蒽醌、阿巴瑞克(abarelix)、唑来瞵酸盐(zoledronate)、链硝脲、妥昔(rituximab)(Rituxan)、柔红霉素、白消安、苯丁酸氮芥、氟阿糖腺苷(fludarabine)、伊马替尼(imatinib)、阿糖胞苷、伊莫单抗(ibritumomab)(Zevalin)、托西莫单抗(tositumomab)(Bexxar)、干扰素α-2b、美法仑(melphalam)、硼替佐米(bortezomib)(Velcade)、六甲蜜胺、门冬酰胺酶、吉非替尼(gefitinib)(Iressa)、埃洛尼替(erlonitib)(Tarceva)、抗-EGF受体抗体(Cetuximab、Abx-EGF)和埃坡霉素(epothilones)。更优选地,治疗剂是铂试剂(诸如卡铂、奥沙利铂、顺铂)、紫杉烷(诸如紫杉酚、多烯紫杉醇)、吉西他滨或喜树碱。
[33]癌症可以是下述举例的癌症中的一种或多种:乳癌、结肠癌、卵巢癌、骨肉瘤、宫颈癌、前列腺癌、肺癌、滑膜肉瘤、胰腺癌、黑素瘤、多发性骨髓瘤、神经母细胞瘤和横纹肌肉瘤、或还待被确定的其中IGF-I受体水平被提高的其他癌症。
[34]在第十个实施方案中,本发明提供了试剂盒,该试剂盒包括本文中描述的一种或多种要素,和使用这些要素的说明书(instruction)。在优选的实施方案中,本发明的试剂盒包括本发明的抗体、抗体片段或偶联物,以及治疗剂。该优选实施方案的说明书包括使用本发明的抗体、抗体片段或偶联物,以及治疗剂,来抑制癌症细胞生长的说明书,和/或使用本发明的抗体、抗体片段或偶联物,以及治疗剂治疗患有癌症的患者的方法的说明书。
附图简述
[35]图1显示了纯化的EM164抗体与过量表达人Y125 1F IGF-I受体或人胰岛素受体的细胞的特异结合的荧光激活细胞分选(FACS)分析。
[36]图2示出了EM164抗体结合生物素化的人IGF-I受体的结合滴定曲线。
[37]图3示出了EM164抗体对生物素化的IGF-I受体结合人乳癌MCF-7细胞的抑制。
[38]图4显示了EM164抗体对MCF-7细胞中IGF-I刺激的IGF-I受体自磷酸化的抑制。
[39]图5显示了EM164抗体对MCF-7细胞中IGF-I刺激的IRS-1磷酸化的抑制。
[40]图6显示了EM164抗体对SaOS-2细胞中IGF-I刺激的信号转导的抑制。
[41]图7显示了通过MTT分析检测的EM164抗体对不同的生长条件下MCF-7细胞的生长和存活的影响。
[42]图8显示了在不同血清浓度下,EM164抗体对MCF-7细胞的生长和存活的影响。
[43]图9示出了EM164抗体对NCI-H838细胞的IGF-I-和血清-刺激的生长和存活的抑制。
[44]图10显示了EM164抗体,紫杉醇或EM164抗体和紫杉醇的联合处理对小鼠中Calu-6肺癌异种移植物生长的影响。
[45]图11示出了人源化EM164抗体(v.1.0)和鼠EM164抗体的结合之间的竞争。
[46]图12显示了鼠抗IGF-I受体抗体EM164的轻链引导区和可变区的cDNA(SEQ ID NO:49)和氨基酸序列(SEQ ID NO:50)。箭头标记了构架1的起始。根据Kabat的3个CDR序列用下划线划出。
[47]图13显示了鼠抗IGF-I受体抗体EM164重链引导区和可变区的cDNA(SEQ ID NO:51)和氨基酸序列(SEQ ID NO:52)。箭头标记了构架1的起始点。3个CDR序列根据Kabat被下划线标出。
[48]图14显示了抗体EM164的轻链和重链CDR氨基酸序列,根据Chothia典型分类定义(Chothis canonical class definitions)确定。也显示了AbM建模软件(AbM modeling software)对重链CDR的定义。
轻链:CDR1是SEQ ID NO:4,CDR2是SEQ ID NO:5,CDR3是SEQ IDNO:6。重链:CDR1是SEQ ID NO:1,CDR2是SEQ ID NO:2,和CDR3是SEQ ID NO:3。AbM重链:CDR1是SEQ ID NO:53,CDR2是SEQ EDNO:54,CDR3是SEQ ED NO:55。
[49]图15显示了Crl(SEQ ID NO:56)和J558.c(SEQ ID NO:57)基因的胚系序列与抗IGF-I受体抗体EM164的轻链和重链氨基酸序列的比对。短划线(-)表示序列的同一性。
[50]图16显示了用于构建和表达重组的嵌合的和人源化的EM164抗体的质粒。A)轻链克隆质粒,B)重链克隆质粒,C)哺乳动物抗体表达质粒。
[51]图17显示了从结构文件组中的127个抗体中筛选出的轻链的10个最同源的氨基酸序列,被用来预测EM164的表面残基,em164 LC(SEQ ID NO:58),2jel(SEQ ID NO:59),2pcp(SEQ ID NO:60),1nqb(SEQ ID NO:61),1kel(SEQ ID NO:62),1hyx(SEQ ID NO.63),1igf(SEQ ID NO:64),1tet(SEQ ID NO:65),1clz(SEQ ID NO:66),1bln(SEQ ID NO:67),1cly(SEQ ID NO:68),保守序列(SEQ ID NO:69)。
[52]图18显示了从结构文件组中的127个抗体中筛选出的重链的10个最同源的氨基酸序列,被用来预测EM164的表面残基,em164 HC(SEQ ID NO:70),1nqb(SEQ ID NO:71),1ngp(SEQ ID NO:72),1fbi(SEQ ID NO:73),1afy(SEQ ID NO:74),1yuh(SEQ ID NO:75),1plg(SEQ ID NO:76),1d5b(SEQ ID NO:77),1ae6(SEQ ID NO:78),1axs(SEQ ID NO:79),3hfl(SEQ ID NO:80),保守序列(SEQ ED NO:81)。
[53]图19显示了所述10个最同源结构的每个(A)轻链和(B)重链可变区残基的平均可及性。数字代表在Kabat抗体序列中的位置编号。
[54]图20显示了鼠EM164(muEM164)和人源化EM164(huEM164)的轻链可变区氨基酸序列。muEM164(SEQ ID NO:82),huEM164 V1.0(SEQ ED NO:83),huEM164 V1.1(SEQ ED NO:84),huEM164 V1.2(SEQ ED NO:85),huEM164 V1.3(SEQ ED NO:86)。
[55]图21显示了鼠EM164抗体(muEM164,SEQ ID NO:87)和人源化EM164抗体(huEM164,SEQ ED NO:88)的重链可变区氨基酸序列。
[56]图22显示了huEM164 v1.0轻链可变区DNA和氨基酸序列(DNA,SEQ ID NO:89,氨基酸SEQ ID NO:90)和重链可变区DNA和氨基酸序列(DNA,SEQ ID NO:91,氨基酸SEQ ID NO:92)。
[57]图23显示了人源化EM164 v1.1的轻链可变区DNA和氨基酸序列(DNA,SEQ ID NO:93;氨基酸SEQ ID NO:94),v1.2的轻链可变区DNA和氨基酸序列(DNA,SEQ ID NO:95;氨基酸SEQ ID NO:96)和v1.3的轻链可变区DNA和氨基酸序列(DNA,SEQ ID NO:97;氨基酸SEQ ID NO:98)。
[58]图24显示了人源化EM164 v1.0抗体和鼠EM164抗体对IGF-I刺激的MCF-7细胞生长和存活的抑制。
[59]图25显示EM164可抑制IGF-I刺激的MCF-7细胞周期。
[60]图26显示EM164可抑制IGF-I和血清的抗凋亡效应。用EM164进行的处理可引起凋亡性细胞死亡,这通过被切割的CK18蛋白水平升高证实。
[61]图27显示了使用EM164抗体,吉西他滨(吉西他滨)或EM164和吉西他滨的联合进行的处理对免疫缺陷小鼠中人BxPC-3胰腺癌异种移植物生长的影响。
发明祥述
[62]本发明发现和改良了可与细胞表面上的人胰岛素样生长因子-I受体(IGF-IR)特异结合的新的抗体。所述抗体和片段具有独特的抑制受体的细胞功能的能力,而没有活化受体本身的能力。因此,以前已知的可特异结合和抑制IGF-IR的抗体即使在缺少IGF-IR配体的情况下也可活化受体,而本发明的抗体或片段可拮抗IGF-IR却又实质上缺少激动剂活性。而且,本发明的抗体和抗体片段可抑制人肿瘤细胞如MCF-7细胞在有血清条件下的生长超过80%,其抑制程度高于使用以前已知的抗IGF-IR抗体所获得的程度。
[63]本发明的实施源自一种鼠抗IGF-IR抗体,在此称为EM164,对此进行了完全的表征,包括轻链和重链的氨基酸序列,CDRs的鉴定,表面氨基酸的鉴定,以及其以重组形式进行表达的方法。
[64]在图15中显示了胚系序列(germline sequence),并将其与EM164的序列进行比对。这样的比较鉴定出EM164中可能的体细胞突变(somatic mutation),包括在轻链CDR1中的一个和重链CDR2中的一个。
[65]抗体EM164和其人源化形式的轻链和重链的一级氨基酸和DNA序列在此被公开。但本发明的范围不限于包含这些序列的抗体和片段。相反,与胰岛素样生长因子-I受体特异结合,并拮抗该受体的生物学活性,而又实质上缺少激动剂活性的所有抗体和片段都包括在本发明的范围之内。因此,抗体和抗体片段可能在其骨架(scaffold),CDRs,轻链和重链的氨基酸序列上与抗体EM164或人源化衍生物有区别,并仍然包括在本发明的范围之内。
[66]通过建模(modeling)鉴定抗体EM164的CDRs,并且其分子结构已经被预测。另外,尽管所述CDRs对于抗原结合部位(epitope)的识别很重要,但它们对于本发明的抗体和片段并不是必需的。因此,提供了具有改善的性质的抗体和片段,例如可通过本发明的抗体的亲和力成熟来产生。
[67]多样的抗体和抗体片段,以及抗体模拟物(mimics)可以很容易地通过在位于一套特定的CDRs的侧面的可变区和恒定区序列中进行突变、删除和/或插入而产生。因此,例如,对于一套既定的CDR,可以通过不同重链的取代而得到不同类型的Ab,由此产生例如IgG1-4、IgM、IgA1-2、IgD、IgE型抗体和同型(isotypes)抗体。同样,在本发明范围内的人造抗体可通过将一组既定的CDRs嵌入进整个合成的骨架中而产生。术语“可变的(variable)”在此用来描述可变结构域的某些部分,它们的序列在抗体之间是不同的,可用于每一特定抗体与其抗原的结合以及特异性。然而,可变性通常并不是在抗体的可变结构域中平均分布的。这种可变性典型地集中在轻链和重链可变结构域内的称为互补性决定区(complementarity determining regions,CDRs)或高变区(hypervariable regions)的三个片段内。可变结构域的更加高度保守的部分称为构架(framework,FR)。重链和轻链的可变结构域每个都包含4个构架区,大部分采取β-片层构型,通过三个CDRs连接,形成环状连接,在一些情况下形成β-片层结构的一部分。各条链中的CDR通过FR区紧靠在一起,并和来自另一条链的CDR一起形成抗体的抗原结合位点(E.A.Kabat等,Sequences of Proteins of ImmunologicalInterest,第五版,1991,NIH)。恒定区不直接参与抗体与抗原的结合,但显示有多种效应器功能(effector function),如该抗体参与抗体依赖的细胞毒性。
[68]人源化抗体或不受其他哺乳动物排斥的抗体可以采用几种技术来产生,如表面重构(resurfacing)和CDR移植(CDR grafting)。在表面重构技术中,将分子建模,统计分析和诱变技术结合起来调整可变区的非CDR表面以使其类似于靶宿主的已知抗体的表面。抗体表面重构的策略和方法和降低抗体在不同宿主中的免疫原性的其它方法在美国专利5,639,641中公开,在此引入以供参考。在CDR移植技术中,鼠重链和轻链CDRs被移植进人的整个构架序列中。
[69]本发明也包括在本说明书中描述的抗体的功能等价物(functionalequivalents)。功能等价物具有与所述抗体相当的结合特性,包括例如,嵌合的,人源化的和单链的抗体以及其片段。产生这样的功能等价物的方法在PCT申请WO 93/21319,欧洲专利申请239,400;PCT申请WO89/09622;欧洲专利申请338,745;和欧洲专利申请EP 332,424中公开,分别在此整体引入以供参考。
[70]功能等价物包括有其氨基酸序列实质上与本发明的抗体的可变区或高变区的氨基酸序列相同的氨基酸序列。“实质上相同”在用于描述氨基酸序列时是指与另一条氨基酸序列具有至少大约90%,更优选至少大约95%的序列同一性的序列,这种同一性可以根据Pearson和Lipman的FASTA搜索方法测定,Proc.Natl.Acad.Sci.USA 85,2444-2448(1988)。
[71]嵌合抗体优选具有实质上或全部地源自人抗体恒定区的恒定区以及实质上或全部地源自除人之外的哺乳动物的可变区序列的可变区。抗体的人源化形式的制备例如可通过将鼠抗体的互补性决定区替换进人构架结构域中来完成,例如,见PCT申请公开号W092/22653。人源化抗体优选具有除了实质上或全部地源自相应的人抗体区的互补性决定区(CDR)和实质上或全部地源自非人的哺乳动物的CDRs之外的恒定区和可变区。
[72]功能等价物也包括单链抗体片段,也称为单链抗体(scFvs)。这些片段含有抗体可变重链氨基酸序列(VH)的至少一个片段,该片段与抗体可变轻链序列(VL)的至少一个片段相连,这些片断间可以具有一个或多个将其互相连接的连接物,也可以没有这样的连接物。这样的连接物可以是短的,柔性的肽,它们是经过选择的,以确保一旦(VL)和(VH)结构域被连接起来便能形成适当的三维折叠,从而保留住作为该单链抗体片段的来源的完整抗体的靶分子结合特异性。通常,(VL)或(VH)序列的羧基末端可通过这样的肽连接物共价相连至互补(VL)和(VH)序列的氨基酸末端。单链抗体片段可通过分子克隆,抗体噬菌体表面展示文库或类似的技术产生。这些蛋白可在真核细胞或原核细胞,包括在细菌中产生。
[73]单链抗体片段包含具有本说明书中描述的完整抗体的可变区或互补性决定区(CDRs)中的至少一个的氨基酸序列,但缺少那些抗体的一些或所有的恒定结构域。这些恒定结构域不是抗原结合所必需的,但构成了完整抗体的结构的主要部分。单链抗体片段因此可以克服在应用含有部分或所有恒定结构域的抗体时相关的一些问题。例如,单链抗体片段往往没有生物分子和重链恒定区之间的不期望的相互作用,或其它不想要的生物学活性。另外,单链抗体片段较完整抗体要显著小,因此较完整抗体有更高的毛细血管通透性,可使单链抗体片段更有效地定位和结合于靶抗原结合位点上。而且,抗体片段可以相对大规模地在原核细胞中产生,因此方便了其生产。而且,单链抗体片段相对小的尺寸使它们在接受者中激发起免疫应答的可能性小于完整抗体。
[74]功能等价物进一步包括具有与完整抗体相同或相当的结合特性的抗体片段。这样的片段含有一个或两个Fab片段或F(ab′)2片段。优选地,所述抗体片段含有完整抗体的所有六个互补性决定区,尽管含有更少的这样的区域,如含有三个,四个或五个CDRs的片段也是有功能的。进一步,功能等价物可以是下面的免疫球蛋白类型的任何一类中的成员或者组合:IgG、IgM、IgA、IgD、或IgE、及其亚类。
[75]在此所述的抗IGF-I受体抗体EM164及其人源化变体的氨基酸和核酸序列的知识,可用来开发同样可结合人IGF-I受体,并抑制IGF-I受体的细胞功能的其他抗体。基于初级抗体(primary antibody)序列的知识,数个研究调查了在抗体序列中不同位置导入一个或多个氨基酸变化对其特性如结合和表达水平的影响(Yang,W.P.等,1995,J.MolBiol,254,392-403;Rader,C.等,1998,Proc.Natl.Acad.Sci.USA,95,8910-8915;Vaughan,T.J.等,1998,Nature Biotechnology,16,535-539)。
[76]在这些研究中,初级抗体的变异体已经通过改变CDR1,CDR2,CDR3或构架区中重链和轻链基因的序列而产生,使用的方法如寡核苷酸介导的位点定向诱变,盒式诱变(cassette mutagenesis),易错PCR,DNA重排(shuffling),或者利用大肠杆菌的增变株(mutator-strains)(Vaughan,T.J.等,1998,Nature Biotechnology,16,535-539;Adey,N.B.等,1996,第16章,277-291页,在″Phage Display of Peptides andProteins″,Eds.Kay,B.K.等,Academic Press)。这些改变初级抗体的序列的方法导致二级抗体(secondary antibodies)的亲和性增加(Gram.,H.等,1992,Proc.Natl.Acad.Sci.USA,89,3576-3580;Boder,E.T.等,2000,Proc.Natl.Acad.Sci.USA,97,10701-10705;Davies,J.和Riechmann,L,1996,Immunotechnolgy,2,169-179;Thompson,J.等,1996,J.Mol Biol,256,77-88;Short,M.K.等,2002,J.Biol.Chem.,277,16365-16370;Fui-ukawa,K.等,2001,J.Biol.Chem.,276,27622-27628)。
[77]通过改变抗体的一个或多个氨基酸残基的类似定向策略,在本发明中描述的抗体序列可以被用来开发具有改善的功能的抗IGF-I受体抗体。
[78]本发明的偶联物包括在此公开的与细胞毒性试剂相连接的抗体,片段和其类似物。优选的细胞毒性试剂是美登木素类生物碱,紫杉烷(taxanes)和CC-1065类似物。所述偶联物可通过体外的方法来制备。为了将细胞毒性试剂与抗体连接,使用了连接基团。适合的连接基团在本领域是为人所熟知的,包括二硫基团,硫醚基团,酸性不稳定(acidlabile)基团,光不稳定基团,肽酶不稳定基团和酯酶不稳定基团。优选的连接基团是二硫基和硫醚基。例如,偶联物可采用二硫化物交换反应(disulfide exchange reaction)或在抗体和细胞毒性试剂之间形成硫醚键而构建。
[79]美登木素类生物碱和美登木素类生物碱类似物属于优选的细胞毒性试剂。合适的美登木素类生物碱的例子包括美登醇和美登醇类似物。合适的美登木素类生物碱在美国专利4,424,219;4,256,746;4,294,757;4,307,016;4,313,946;4,315,929;4,331,598;4,361,650;4,362,663;4,364,866;4,450,254;4,322,348;4,371,533;6,333,410;5,475,092;5,585,499和5,846,545中公开。
[80]紫杉烷也是优选的细胞毒剂。适合用于本发明的紫杉烷公开于美国专利6,372,738和6,340,701中。
[81]CC-1065和其类似物也是优选的用于本发明的细胞毒性药物。CC-1065和其类似物在美国专利6,372,738;6,340,701;5,846,545和5,585,499中公开。
[82]制备这样的细胞毒性偶联物的一种吸引人的候选物是CC-1065,CC-1065是从链霉菌(Streptomyces zelensis)的培养肉汤中分离出的一种有效的抗肿瘤抗生素。CC-1065较普遍使用的抗癌药物,如阿霉素,氨甲蝶呤和长春新碱的体外效力高大约1000倍(B.K.Bhuyan等,Cancer Res.,42,3532-3537(1982))。
[83]细胞毒性药物例如氨甲蝶呤,柔毛霉素,阿霉素,长春新碱,长春碱,美法仑,丝裂霉素C,苯丁酸氮芥,卡奇霉素也适合制备本发明的偶联物,所述药物分子也可通过中间载体分子如血清白蛋白而连接到抗体分子上。
[84]为了诊断性应用,本发明的抗体通常会用可检测的部分(moiety)来标记。所述的可检测的部分可以是能够直接或间接产生可检测信号的任何一种物质。例如,可检测部分可以是放射性同位素,如3H,14C,32P,35S,或131I;荧光或化学发光化合物,如异硫氰酸荧光素,若丹明,或虫萤光素(luciferin);或酶,如碱性磷酸酶,β-半乳糖苷酶或辣根过氧化物酶。
[85]可应用本领域中已知的将抗体与可检测部分偶联的任何方法,包括由Hunter等,Nature 144:945(1962);David等,Biochemistry 13:1014(1974);Pain等,J.Immunol.Meth 40:219(1981);和Nygren,J.Histochem.and Cytochem.30:407(1982)描述的那些方法。
[86]本发明的抗体可以在任何已知的分析方法中被应用,如竞争结合分析,直接和间接夹心分析,和免疫沉淀分析(Zola,MonoclonalAntibodies:A Manual of Techniques,147-158页(CRC Press,Inc.,1987))。
[87]本发明的抗体也用于进行体内成像,其中用可检测部分如不透射线的试剂(radio-opaque agent)或放射性同位素标记的抗体被施予对象,优选施于血流中,然后测定宿主中标记抗体的存在和位置。这种成像技术可用于恶性肿瘤的分级(staging)和治疗。抗体可用任何可在宿主中可检测到的部分来标记,不管是通过核磁共振,放射学或本领域中已知的其他检测方法。
[88]本发明的抗体也可用作亲和纯化试剂。在这个过程中,使用本领域中已知的方法将抗体固定在合适的支持物上,如Sephadex树脂或滤纸上。
[89]基于它们对IGF-I受体在细胞中的功能的抑止,本发明的抗体可以用作生物学研究试剂。
[90]为了进行治疗应用,本发明的抗体或偶联物可以以药学上可接受的剂型施于对象。它们可作为丸剂静脉施与或在一段时间内连续输注,通过肌肉内,皮下,关节内,滑膜内,鞘内,口服,局部或吸入途径给药。抗体也可通过肿瘤内,肿瘤旁,病灶内或病灶旁的途径给药,发挥局部以及全身的治疗效应。合适的药学上可接受的载体,稀释剂和赋形剂是为人所熟知的,可由本领域中的技术人员根据临床情形来确定。合适的载体,稀释剂和/或赋形剂的实例包括:(1)Dulbecco′s磷酸缓冲液,pH大约7.4,含有大约1mg/ml至25mg/ml人血清白蛋白,(2)0.9%盐水(0.9%w/v NaCl),和(3)5%(w/v)葡萄糖。本发明的方法可在体外,体内或离体实施。
[91]在其他治疗性处理中,本发明的抗体,抗体片段或偶联物可与一种或多种其他的治疗性试剂共同施用或相继施用。合适的治疗性药剂包括但不限于细胞毒性试剂或细胞生长抑制试剂。紫杉醇是一种优选的治疗性试剂,也是一种细胞毒剂。
[92]癌症治疗试剂是指那些可杀死癌症细胞或限制癌症细胞的生长而对宿主产生最小的损害的试剂。因此,这样的试剂可以利用癌症细胞特性(例如,代谢,血管化作用或细胞表面抗原呈递)与健康宿主细胞的任何差异。肿瘤形态学的差异是进行干预(intervention)的潜在位点:例如,第二种治疗物可以是抗体如抗VEGF抗体,它可用于延迟实体肿瘤内部的血管化作用,由此可减慢其生长速率。其它的治疗试剂包括但不限于辅助物如盐酸格拉司琼(granisetron HCL),雄性激素抑制剂如亮丙瑞林乙酸盐(leuprolide acetate),抗生素如阿霉素,抗雌激素药如它莫西芬,抗代谢药如α干扰素-2a,细胞毒性药物如紫杉醇,酶抑制剂如ras法尼基-转移酶(ras farnesyl-transferase)抑制剂,免疫调节剂如阿地白介素(aldesleu kin),和氮芥衍生物如盐酸美法仑(melphalan HCl),以及类似物。
[93]可与EM164组合以获得改善的抗-肿瘤效率的治疗剂包括用于肿瘤学实践的多种多样的试剂(参考:Cancer,Principles & Practice ofOncology,DeVita,V.T.,Hellman,S.,Rosenberg,S.A.,第六版,Lippincott-Raven,Philadelphia,2001),诸如多烯紫杉醇、紫杉酚、阿霉素、表阿霉素、环磷酰胺、曲妥珠(Herceptin)、卡培他宾、它莫西芬、托瑞米芬、来曲唑、阿纳曲唑、氟维司群、依西美坦、戈舍瑞林、奥沙利铂、卡铂、顺铂、地塞米松、抗排卵肽、贝伐单抗(Avastin)、5-氟尿嘧啶、亚叶酸、左旋咪唑、依立替康、依托泊甙、拓朴替康、吉西他滨、长春瑞滨、雌莫司汀、米托蒽醌、阿巴瑞克、唑来瞵酸盐、链硝脲、妥昔(Rituxan)、柔红霉素、白消安、苯丁酸氮芥、氟阿糖腺苷、伊马替尼、阿糖胞苷、伊莫单抗(Zevalin)、托西莫单抗(Bexxar)、α干扰素-2b、美法仑、硼替佐米(Velcade)、六甲蜜胺、门冬酰胺酶、吉非替尼(Iressa)、埃洛尼替(Tarceva)、抗-EGF受体抗体(Cetuximab、Abx-EGF)、埃坡霉素,以及细胞毒性药物和针对细胞-表面受体的抗体的偶联物。优选的治疗剂是铂试剂(诸如卡铂、奥沙利铂、顺铂)、紫杉烷(诸如紫杉酚、多烯紫杉醇)、吉西他滨和喜树碱。
[94]一种或多种额外的治疗剂可以在本发明的抗体、抗体片段或偶联物之前、同时或之后给予。本领域技术人员将理解,对于每一个治疗剂来说,特定的给药顺序可能具有优点。类似地,本领域技术人员将理解,对于每一治疗剂,本发明的试剂、抗体、抗体片段或偶联物之间的给药时间长度可以变化。
[95]尽管本领域技术人员将理解到,每一治疗剂的剂量将取决于试剂的特性,优选的剂量可以在约10mg/平方米至约2000mg/平方米,更优选地在约50mg/平方米至约1000mg/平方米范围内。对于优选的试剂诸如铂试剂(卡铂、奥沙利铂、顺铂),优选的剂量约10mg/平方米至约400mg/平方米,对于紫杉烷(紫杉酚、多烯紫杉醇),优选的剂量约20mg/平方米至约150mg/平方米,对于吉西他滨,优选的剂量约100mg/平方米至约2000mg/平方米,对于喜树碱,优选的剂量约50mg/平方米至约350mg/平方米。该治疗剂以及其他治疗剂的剂量可以取决于本发明的抗体、抗体片段或偶联物是否与治疗剂同时给药或顺序给药。
[96]无论是同时给药或顺序给药,本发明的抗体、抗体片段或偶联物和一种或多种额外的治疗剂的给药,可以如上面为治疗应用所描述的那样进行。将会被本领域技术人员理解的是,用于同时给药的合适的药学上可接受的载体、稀释剂和赋形剂取决于正被同时给药的特定治疗剂的特性。
[97]当以含水剂型存在而不是被冻干时,所述抗体典型地以大约0.1mg/ml至100mg/ml的浓度进行配制,尽管在这些范围之外的很大变化也是允许的。为了治疗疾病,抗体或偶联物的合适剂量将依赖于如上面所定义的要治疗的疾病类型、疾病的严重性和病程,而不论所述抗体的施用是为了预防还是治疗、以往治疗的过程、患者的临床病史和对抗体的反应、以及主治医生的判断。抗体可以在一次或在多次治疗过程中被适当地给予患者。
[98]取决于疾病的类型和严重性,对于给药于患者,初始的候选剂量优选约1mg/平方米至约2000mg/平方米的抗体,更优选地,约10mg/平方米至约1000mg/平方米的抗体,无论是例如通过一次或多次分开给药还是连续输注来进行。对于在若干天或更长时间期间反复给药,取决于条件,反复进行治疗,直到获得对疾病症状的理想的抑制。然而,其他剂量方案也可以应用,并不排除在外。
[99]本发明也包括试剂盒,该试剂盒包括一种或多种本文中描述的要素,以及使用这些要素的说明书。在优选的实施方案中,本发明的试剂盒包括本发明的抗体、抗体片段或偶联物,以及治疗剂。该优选实施方案的说明书包括使用本发明的抗体、抗体片段或偶联物和治疗剂,来抑制癌症细胞生长的说明书,和/或使用本发明的抗体、抗体片段或偶联物和治疗剂治疗患有癌症的患者的方法的说明书。
[100]优选地,用于本试剂盒的抗体具有与小鼠杂交瘤EM164(ATCC登记号PTA-4457)产生的鼠抗体EM164同样的氨基酸序列,或该抗体是其表位-结合片段,其中抗体以及抗体片段特异性地结合到胰岛素-样生长因子-I受体。用于试剂盒中的抗体或抗体片段也可以是表面重构形式的EM164抗体、人源化形式的EM164抗体、或改变形式的EM164抗体,该抗体具有至少一个核苷酸突变、剔除或插入。这三种形式的抗体中的每一种抗体或抗体片段都保留与EM164抗体相同的结合特异性。
[101]优选地,用于本试剂盒的治疗剂选自多烯紫杉醇、紫杉酚、阿霉素、表阿霉素、环磷酰胺、曲妥珠(Herceptin)、卡培他宾、它莫西芬、托瑞米芬、来曲唑、阿纳曲唑、氟维司群、依西美坦、戈舍瑞林、奥沙利铂、卡铂、顺铂、地塞米松、抗排卵肽、贝伐单抗(Avastin)、5-氟尿嘧啶、亚叶酸、左旋咪唑、依立替康、依托泊甙、拓朴替康、吉西他滨、长春瑞滨、雌莫司汀、米托蒽醌、阿巴瑞克、唑来瞵酸盐、链硝脲、妥昔(Rituxan)、柔红霉素、白消安、苯丁酸氮芥、氟阿糖腺苷、伊马替尼、阿糖胞苷、伊莫单抗(Zevalin)、托西莫单抗(Bexxar)、α干扰素-2b、美法仑、硼替佐米(Velcade)、六甲蜜胺、门冬酰胺酶、吉非替尼(Iressa)、埃洛尼替(Tarceva)、抗-EGF受体抗体(Cetuximab、Abx-EGF)和埃坡霉素。更优选地,治疗剂是铂试剂(诸如卡铂、奥沙利铂、顺铂)、紫杉烷(诸如紫杉酚、多烯紫杉醇)、吉西他滨或喜树碱。
[102]本发明试剂盒的要素是适合于试剂盒的形式,诸如溶液或冻干粉末。将会被本领域技术人员所理解的是,试剂盒的要素的浓度或数量取将根据试剂盒的每一种要素的特性和目的用途而变化。
[103]试剂盒的说明书中指出的癌症和细胞包括乳腺癌、结肠癌、卵巢癌、骨肉瘤、宫颈癌、前列腺癌、肺癌、滑膜肉瘤、胰腺癌、黑素瘤、多发性骨髓瘤、神经母细胞瘤和横纹肌肉瘤。
实施例
[104]现通过参考下面的实施例来描述本发明,其仅仅是说明性的,而并非限制本发明。
实施例1:鼠EM164抗体
[105]在该这第一个实施例中,公开了本发明的鼠源性抗体的完整的一级氨基酸结构和cDNA序列,以及它的结合特性和将其以重组形式表达的方式。因此,充分且完整地公开了本发明的抗体及其制备,这样在免疫学领域中的普通技术人员将能够制备所述的抗体,而不需要过多的实验。
A.抗-IGF-I受体的单克隆抗体杂交瘤的产生
[106]应用表达带有Y1251F突变的人IGF-I受体的细胞系来进行免疫,因为它可表达大量的IGF-I受体(~107/细胞)。在IGF-I受体的胞浆结构域中的Y1251F突变导致转化和抗凋亡信号传导功能的丧失,但不影响IGF-I结合和IGF-I刺激的促有丝分裂信号转导(O′Connor,R.等,1997,Mol Cell.Biol,17,427-435;Miura,M.等,1995,J.Biol Chem.,270,22639-22644)。另外,突变也不影响抗体的产生,因为本实施例的抗体结合于IGF-I受体的细胞外结构域,该结构域在Y1251F突变体和野生型受体中是相同的。
[107]用带有Y1251F突变的人IGF-I受体基因以及嘌呤霉素抗性基因转染IGF-I受体缺陷型小鼠的3T3样细胞,产生表达带有Y1251F突变的人IGF-I受体的细胞系,用嘌呤霉素(2.5微克/mL)并通过FACS分选对IGF-I受体的高表达进行选择(Miura,M.等,1995,J.Biol.Chem.,270,22639-22644)。具有高水平IGF-I受体表达的细胞系进一步用高浓度的嘌呤霉素如25微克/mL进行选择,这个浓度对大多数细胞是有毒性的。存活的集落被拣出,并选择那些显示出高水平IGF-I受体表达的集落。
[108]6月龄的CAF1/J雌性小鼠在第0天用过量表达Y1251F突变型人IGF-I受体的细胞进行腹膜内免疫(5×105个细胞,悬浮于0.2mL PBS中)。所述动物用0.2mL细胞悬液如下进行加强免疫(boosted):第2天,1×106细胞;第5天,2×106细胞;第7、9、12和23天,1×107细胞。在第26天,处死小鼠,移走其脾脏。
[109]脾脏在两块毛玻璃片之间被磨碎以获得单细胞悬液,用含有青霉素和链霉素的无血清RPMI培养基(SFM)进行清洗。脾细胞沉淀重悬于10mL 0.83%(w/v)氯化铵水溶液中,置于冰上10分钟以裂解血红细胞,然后用无血清的培养基(SFM)清洗。脾细胞(1.2×108)与来自非分泌性鼠骨髓瘤细胞系P3×63Ag8.653(ATCC,Rockville,MD;Cat.#CRL1580)的骨髓瘤细胞(4×107)集中在一个试管中,并用无血清RPMI-1640培养基(SFM)清洗。去掉上清液,细胞沉淀重悬于残余的培养基中。该试管置于37℃的一烧杯水中,以0.5mL/分钟的滴入速率缓慢地加入1.5mL聚乙二醇溶液(50%PEG(w/v),平均分子量1500,在75mM HEPES中,pH 8),同时轻轻振荡试管。等待1分钟后,如下加入10mL SFM:第1分钟1mL,第2分钟2mL,第3分钟7mL。然后再将10mL在1分钟内缓缓加入。细胞通过离心被沉淀,用SFM中清洗,重悬于添加了5%胎牛血清(FBS)、次黄嘌呤/氨蝶呤/胸苷(HAT)、青霉素、链霉素和10%杂交瘤克隆添加剂(HCS)的RPMI-1640生长培养基中。细胞接种在96孔平底组织培养板中,每孔有200μL培养物,其中有2×105个脾细胞。5-7天后,从每孔中移去100μL,用添加了次黄嘌呤/胸苷(HT)和5%FBS的生长培养基替换。用于免疫和产生杂交瘤的一般条件的描述可参见J.Langone和H.Vunakis(Eds.,Methods in Enzymology,Vol.121,″ImmunochemicalTechniques,Part I″;1986;Academic Press,Florida)以及E.Harlow和D.Lane(″Antibodies:A Laboratory Manual″;1988;Cold Spring HarborLaboratory Press,NewYork)。也可使用本领域中技术人员所熟知的其它免疫和杂交瘤产生技术。
[110]通过ELISA筛选杂交瘤克隆的培养物上清液与纯化的人IGF-I受体的结合,通过ELISA和FACS筛选杂交瘤克隆的培养物上清液与过量表达人IGF-I受体的细胞的特异性结合和与过量表达人胰岛素受体的细胞的无法结合,如下所述。对过量表达人IGF-I受体的细胞较过量表达人胰岛素受体的细胞显示出更高结合亲和力的克隆被扩增和亚克隆。亚克隆的培养物上清通过上述的结合分析被进一步筛选。通过这个过程,选择出了亚克隆3F1-C8-D7(EM164),其重链和轻链基因被克隆,并被测序,如下所述。
[111]通过如下的方法分离人IGF-I受体,用于针对来自杂交瘤克隆的上清液与IGF-I受体的结合而进行筛选。用生物素化试剂如硫代-NHS-LC-生物素,硫代-NHS-SS-生物素,或NHS-PEO4-生物素修饰重组的IGF-I,制备出生物素化的IGF-I。生物素化的IGF-I在链亲和素-琼脂糖珠上被吸收,并与过量表达人野生型或Y1251F突变型IGFR的细胞的裂解物一起孵育。用含有2至4M尿素和去垢剂如triton X-100或辛基-β-糖苷的缓冲液清洗和洗脱这些珠子。洗脱出的IGF-I受体用PBS透析,并通过SDS-PAGE在还原条件下分析纯度,结果显示了IGF-I受体的α和β链的条带,其分子量分别为大约135kDa和95kDa。
[112]为了检查杂交瘤上清液与纯化的IGF-I受体的结合,用在pH 9.5的50mM CHES缓冲液中稀释的纯化人IGF-I受体样品(通过亲和性纯化样品的尿素/辛基-β-糖苷洗脱物的透析而制备出来)包被Immulon-4HB ELISA板(Dynatech)(100μL;4℃,过夜)。这些孔用200μL的封闭缓冲液(10mg/mL BSA,溶于含有50mM Tris,150mMNaCl,pH 7.5和0.1%tween-20的TBS-T缓冲液中)来封闭,并与来自杂交瘤克隆的上清液(100μL;在封闭缓冲液中稀释)孵育大约1h至12h,用TBS-T缓冲液清洗,与羊抗鼠-IgG-Fc-抗体-辣根过氧化物酶(HRP)偶联物(100μL;在缓冲液中的浓度为0.8μg/mL;JacksonImmunoResearch Laboratories)共同孵育,然后洗涤并用ABTS/H2O2底物(0.5mg/mL ABTS,0.03%H2O2,在0.1M柠檬酸盐缓冲液中,pH4.2)在405nm处进行检测。典型地,来自3F1杂交瘤亚克隆的上清液在3分钟的显色过程中产生了大约1.2个吸光度单位的信号,与此相比,从其他一些杂交瘤克隆的上清液中获得的值为0.0。这种ELISA的一般条件类似于如E.Harlow和D.Lane所述的抗体结合和检测的标准ELISA条件(″Using Antibodies:A Laboratory Manual″;1999,Cold SpringHarbor Laboratory Press,New York),所述的标准条件也可被应用。
[113]用过量表达人Y1251F-IGF-I受体的细胞系和过量表达人胰岛素受体的细胞系对杂交瘤上清液进行ELISA筛选,筛选出与人IGF-I受体而不与人胰岛素受体特异结合的杂交瘤上清。两种细胞系都是由IGF-I受体缺陷型小鼠的3T3样细胞产生的。过量表达IGF-I受体的细胞和过量表达胰岛素受体的细胞分别从组织培养瓶中通过快速的胰蛋白酶/EDTA处理来获取,然后细胞悬浮在含有10%FBS的生长培养基中,通过离心而被沉淀,在用PBS清洗。将被清洗的细胞(100μL,大约1-3×106细胞/mL)加入到用植物血球凝集素(100μL,20μg/mLPHA)包被的Immulon-2HB板的孔中,离心10分钟,使其粘附于PHA-包被的孔中。轻轻拍打有细胞的板以去掉PBS,然后在37℃干燥过夜。用含有5mg/mL BSA的PBS溶液在37℃封闭孔1h,然后轻轻地用PBS清洗。然后将杂交瘤克隆的等分上清液(100μL;在封闭缓冲液中稀释)加入到含有过量表达IGF-I受体的细胞的孔中和含有过量表达胰岛素受体的细胞的孔中,并在室温下孵育1h。所述孔用PBS清洗,用羊抗鼠-IgG-Fc-抗体-辣根过氧化物酶偶联物(100μL;0.8μg/mL,在封闭缓冲液中)孵育1h,然后清洗,使用ABTS/H2O2底物检测结合。在与过量表达IGF-I受体的细胞孵育后,3F1杂交瘤亚克隆的典型上清液在12分钟显色过程中产生了0.88个吸光度单位的信号,与此相比,与过量表达人胰岛素受体的细胞孵育后获得的值为0.22个吸光度单位。
[114]为了提供纯化的EM164抗体,依照生产厂商的说明书,使杂交瘤在Integra CL 350瓶中(Integra Biosciences,Maryland)生长。从Integra瓶的收获上清液中获得大约0.5-1mg/mL的抗体,这是使用抗体标准品通过ELISA和SDS-PAGE/考马斯亮蓝染色来定量的。在蛋白A-琼脂糖珠的色谱柱上通过亲和色谱来纯化抗体,所述纯化是在标准的纯化条件下进行,即在含有3M NaCl的100mM Tris缓冲液,pH8.9中上样和清洗,然后用含有150mM NaCl的100mM醋酸溶液洗脱。含有抗体的洗脱级分用冷的2M K2HPO4溶液中和,并用PBS在4℃进行透析。通过测量280nm的吸光度(消光系数=1.4mg-1mL cm-1)来测定抗体浓度。纯化的抗体样品在还原条件下通过SDS-PAGE和考马斯亮蓝染色进行分析,结果显示只有抗体的重链和轻链条带,分别在大约55kDa和25kDa处。所述纯化抗体的同型体为具有κ轻链的IgG1。
B.EM164抗体的结合特征
[115]纯化的EM164抗体的特异性结合可使用过量表达人IGF-I受体的细胞和过量表达人胰岛素受体的细胞进行荧光激活细胞分选(FACS)来证实(图1)。使用过量表达IGF-I受体的细胞和过量表达胰岛素受体的细胞(2×105个细胞/mL),在圆底的96孔板中孵育在100μL冷FACS缓冲液(1mg/mL BSA,在Dulbecco′s MEM培养基中)中的EM 164抗体(50-100nM)1h。通过离心细胞将细胞沉淀,用冷的FACS缓冲液通过轻轻吹打进行清洗,然后与羊抗鼠-IgG-抗体-FITC偶联物(100μL;10μg/mL,在FACS缓冲液中)在冰上孵育1h。细胞被沉淀,清洗,重悬在120μL的1%甲醛PBS溶液中。使用FACSCalibur读数器(BD Biosciences)分析培养板。
[116]在过量表达IGF-I受体的细胞与EM164抗体孵育后,获得了很强的荧光变化,相比而言,过量表达胰岛素受体的细胞与EM164抗体的孵育所产生的变化是没有显著意义的(图1),这证明了EM164抗体在其与IGF-I受体的结合中是有选择性的,并不与胰岛素受体结合。对照抗体,抗-IGF-I受体抗体1 H7(Santa Cruz Biotechnology)和抗胰岛素受体α抗体(BD Pharmingen Laboratories),在与过量表达IGF-I受体和胰岛素受体的细胞的孵育后分别产生了荧光的变化(图1)。使用EM164抗体和表达IGF-I受体的人乳癌MCF-7细胞(Dufourny,B.等,1997,J.Biol.Chem.,272,31163-31171),通过FACS分析也观察到了很强的荧光变化,这说明EM164抗体结合于人肿瘤细胞表面上的人IGF-I受体。
[117]通过ELISA滴定抗体在数种浓度下与直接包被的IGF-I受体(应用生物素化的IGF-I亲和纯化得到,如上)或间接俘获的生物素化IGF-I受体的结合,测定EM164抗体与人IGF-I受体结合的解离常数(Kd)。生物素化的IGF-I受体是应用PEO-马来酰亚胺-生物素试剂(Pierce,Molecular Biosciences)对过量表达IGF-I受体的细胞的去垢剂溶解裂解物进行生物素化而制备的,它可应用固定在NHS-琼脂糖珠上的抗IGF-I受体β链抗体进行亲和纯化,并用溶于含有NP-40去垢剂的缓冲液中的2-4M尿素洗脱,并用PBS透析。
[118]EM164抗体与生物素化的IGF-I受体结合的Kd测定是通过将Immulon-2HB板用100μL含有1μg/mL链亲和素的碳酸盐缓冲液(150mM碳酸钠,350mM碳酸氢钠)在4℃过夜包被而进行的。链亲和素包被的孔用200μL封闭缓冲液(含有10mg/mL BSA的TBS-T缓冲液)进行封闭,用TBS-T缓冲液清洗,并与生物素化的IGF-I受体(10至100ng)在室温下孵育4h。然后清洗含有间接俘获的生物素化IGF-I受体的孔,与以不同浓度溶于封闭缓冲液中的EM164抗体(5.1×10-13M至200nM)在室温下孵育2h,然后在4℃孵育过夜。接下来用TBS-T缓冲液清洗孔,与羊抗鼠-IgGH+L-抗体-辣根过氧化物酶偶联物(100μL;0.5μg/mL,在封闭缓冲液中)共同孵育,然后洗涤,应用ABTS/H2O2底物在405nm处检测。Kd的值通过单一位点结合(one-site binding)的非线性回归(non-linear regression)来估计。
[119]类似的结合滴定还通过使用EM164抗体的Fab片段来进行,该片段是通过木瓜酶消化抗体而制备的,如E.Harlow和D.Lane(″UsingAntibodies:A Laboratory Manual″;1999,Cold Spring Harbor LaboratoryPress,New York)所描述。
[120]EM164抗体与生物素化人IGF-I受体结合的结合滴定曲线产生的Kd值为0.1nM(图2)。EM164抗体的Fab片段也与人IGF-I受体非常紧密地结合,Kd值为0.3nM,表明EM164抗体与IGF-I受体的单体结合也非常强烈。
[121]EM164抗体与IGF-I受体结合的这种极低的解离常数值部分是由于非常低的Koff速率造成的,这是通过延迟洗涤与固定的IGF-I受体结合的抗体1-2天后所观察到的强结合信号所证实的。
[122]EM164抗体可用来免疫沉淀IGF-I受体,如通过人乳癌MCF-7细胞的去垢剂溶解裂解物与固定在蛋白G-琼脂糖珠(Pierce ChemicalCompany)上的EM164抗体的孵育所证实的。EM164抗体免疫沉淀物的Western印迹采用兔多克隆抗-IGF-I受体β链(C-末端)抗体(SantaCruz Biotechnology)和羊抗兔-IgG-抗体-辣根过氧化物酶偶联物来检测,然后冲洗,进行增强型化学发光(ECL)检测。MCF-7细胞的EM164免疫沉淀物的Western印迹显示了与IGF-I受体的β链对应的条带,在大约95kDa处,以及与前-IGF-I(pro-IGF-I)受体对应的条带,在大约220kDa处。类似的免疫沉淀也对其他细胞类型实施以检查EM164抗体的结合的种属特异性,结果显示该抗体也与来自cos-7细胞(非洲绿猴)的IGF-I受体结合,但不与3T3细胞(小鼠),CHO细胞(中国仓鼠)或羊成纤维细胞(山羊)的IGF-I受体结合。在MCF-7细胞的裂解物的Western印迹中,EM164抗体不能检测SDS变性的人IGF-I受体,表明它是与天然的,未变性的人IGF-I受体的具有一定构象的表位结合。
[123]EM164抗体的结合结构域进一步应用截短的α链构建物来表征,该构建物含有其侧面带有L1和L2结构域的富含半胱氨酸的结构域(残基1-468),以及与之融合的16-mer-C末端片段(残基704-719),它的末端是C-末端抗原决定部位标记。这种较小的IGF-I受体缺少残基469-703,已经有报道其可结合IGF-I,尽管所述结合不如天然的全长IGF-I受体紧密(Molina,L.等,2000,FEES Letters,467,226-230;Kristensen,C.等,1999,J.Biol.Chem.,274,37251-37356)。因此,制备了截短的IGF-I受体α链构建物,其含有残基1-468以及与之融合的C-末端片断,该片断是残基704-719,侧翼是C-末端myc抗原决定部位标记。构建了表达这种构建物并且也可在人胚胎肾293T细胞中暂时表达该构建物的稳定细胞系。观察到EM164抗体与这种截短的IGF-I受体α链构建物的强烈结合。在被检测的两种抗体中,IR3(Calbiochem)也可与这种截短的α链结合,但1H7抗体(Santa Cruz Biotechnology)不发生结合,表明EM164抗体的抗原决定部位与1H7抗体明显不同。
C.EM164抗体抑制IGF-I对MCF-7细胞的结合
[124]IGF-I与人乳癌MCF-7细胞的结合可被EM164抗体抑制(图3)。MCF-7细胞与或者不与5μg/mL EM164抗体在无血清培养基中孵育2h,然后与50ng/mL生物素化IGF-I在37℃孵育20分钟。然后用无血清培养基洗涤细胞两次,去掉未结合的生物素-IGF-I,然后用含有1%NP-40和蛋白酶抑制剂的50mM HEPES,pH 7.4裂解。Immulon-2HBELISA板用鼠单克隆抗-IGF-I受体β链抗体包被,并将其用于从裂解产物中俘获IGF-I受体和结合的生物素-IGF-I。所述包被抗体与IGF-I受体β链的胞浆C-末端结构域的结合不干扰生物素-IGF-I与IGF-I受体的细胞外结构域的结合。洗涤孔,并与链亲和素-辣根过氧化物酶偶联物孵育,再次洗涤,然后使用ABTS/H2O2底物来检测。5μg/mL EM164抗体对IGF-I与MCF-7细胞结合的抑制实质上是可定量的,这几乎等价于使用缺少生物素-IGF-I的对照时获得的ELISA本底。
[125]除了上述的EM164抗体对IGF-I与MCF-7细胞的结合的抑制情况的分析,下面的分析证明了EM164抗体可高效地取代与MCF-7细胞结合的IGF-I,如生理条件下所需要的,即需要拮抗性的抗IGF-I受体抗体取代已结合的内源性生理配体(如IGF-I或IGF-II)。在这个IGF-I取代分析中,在12孔板中生长的MCF-7细胞遭受血清饥饿(serum-starved),随后与生物素化的IGF-I(20-50ng/mL)在无血清培养基中于37℃(或4℃)孵育1至2h。然后,用EM164抗体或对照抗体(10-100μg/mL)在37℃(或在4℃)处理带有结合的生物素化IGF-I的细胞,处理30min至4h。然后用PBS洗涤细胞,在含有1%NP-40的裂解缓冲液中,在4℃裂解细胞。ELISA如上所述进行,用以从所述裂解产物中俘获IGF-I受体,然后采用链亲和素-辣根过氧化物酶偶联物检测与所述受体结合的生物素化IGF-I。该ELISA证明了EM164抗体在37℃能够几乎完全地取代细胞上预结合的生物素化IGF-I(在30min内为90%,在4h内~100%),在4℃在2h中,取代大约50%。在另一个试验中,NCI-H838肺癌细胞与生物素-IGF-I一起孵育,然后洗涤,与EM164抗体在4℃孵育2h,结果使结合的生物素-IGF-I减少80%。因此,EM164抗体可高效地取代癌细胞上预结合的IGF-I,这在治疗上是非常重要的,因为可以通过取代已结合的内源性生理配体而拮抗IGF-I受体。
[126]MCF-7细胞与EM164抗体在4℃孵育2h(或在37℃孵育30min),根据应用了抗IGF-I受体β链抗体(Santa Cruz Biotechnology;sc-713)的Western印迹分析,这种孵育并不导致IGF-I受体的明显下调,尽管与EM164抗体在37℃长时间孵育2h可引起IGF-I受体25%的下调。因此,在这些短期试验中,EM164抗体在4℃和37℃抑制IGF-I的结合和取代已结合的IGF-I均不能通过由于EM164抗体的结合引起受体下调来解释。EM164抗体有效抑制IGF-I与IGF-I受体的结合和取代预结合的IGF-I的机制很可能是通过结合位点的共享或空间阻遏(stericocclusion)或变构效应所引起的结合竞争。
D.EM164抗体抑制IGF-I受体介导的信号传导
[127]用EM164抗体处理乳癌MCF-7细胞和骨肉瘤SaOS-2细胞几乎完全抑制细胞内的IGF-I受体信号传导,这表现为IGF-I受体自体磷酸化的抑制和其下游效应物如胰岛素受体底物-1(IRS-1),Akt和Erk1/2的磷酸化的抑制(图4-6)。
[128]在图4中,MCF-7细胞在12孔板的常规培养基中生长3天,然后用20μg/mL的EM164抗体(或抗-B4对照抗体)在无血清培养基中处理3h,然后用50ng/mL的IGF-I在37℃刺激20min。然后细胞在冰冷的含有蛋白酶和磷酸酶抑制剂的缓冲液(50mM HEPES缓冲液,pH7.4,1%NP-40,1mM原钒酸钠,100mM氟化钠,10mM焦磷酸钠,2.5mM EDTA,10μM亮肽素,5μM胃蛋白酶抑制剂,1mM PMSF,5mM苯甲脒和5μg/mL抑肽酶)中被裂解。ELISA板用抗IGF-I受体β链C-末端的单克隆抗体TC123进行预包被,与裂解物样本在室温下孵育5h以俘获IGF-I受体。然后洗涤含有被俘获的IGF-I受体的孔,并与生物素化的抗磷酸酪氨酸抗体(PY20;0.25μg/mL;BD TransductionLaboratories)孵育30min,然后洗涤,与链亲和素-辣根过氧化物酶偶联物(0.8μg/mL)孵育30min。洗涤孔,用ABTS/H2O2底物检测。抗B4对照抗体的应用没有显示出对IGF-I刺激的IGF-I受体自体磷酸化作用的抑制。与之相比,在用EM164抗体处理后可获得对IGF-I刺激的IGF-I受体自体磷酸化作用的完全抑制(图4)。
[129]为了证明对胰岛素底物-1(IRS-1)的磷酸化作用的抑制,在应用的ELISA中,使用固定化的抗-IRS-1抗体从裂解物中俘获IRS-1,然后测定可与磷酸化的IRS-1结合的磷脂酰肌醇-3-激酶(PI-3-激酶)的相关p85亚基(Jackson,J.G.等,1998,J.Biol.Chem.,273,9994-10003)。在图5中,MCF-7细胞用5μg/mL的抗体(EM164或IR3)在无血清培养基中处理2h,然后用50ng/mL的IGF-I在37℃刺激10min。抗IRS-1抗体(兔多克隆抗体Upstate Biotechnology)通过与ELISA板上包被的羊抗兔-IgG抗体孵育而被间接俘获,然后通过4℃孵育过夜,用抗IRS-1抗体从细胞裂解物样品中俘获IRS-1。然后孔与鼠单克隆抗-p85-PI-3-激酶抗体(Upstate Biotechnology)孵育4h,然后用羊抗鼠-IgG-抗体-HRP偶联物处理30min。然后洗涤孔,使用ABTS/H2O2底物检测(图5)。如在图5中显示,EM164抗体较IR3抗体更为有效地抑制IGF-I-刺激的IRS-1磷酸化作用,当在缺少IGF-I的情况下与细胞孵育时,EM164抗体对IRS-1磷酸化作用不显示任何激动剂活性。
[130]在SaOS-2细胞(图6)和MCF-7细胞中,其它下游效应物如Akt和Erk1/2的活化,也可以剂量依赖的方式被EM164抗体抑制,如采用裂解物和磷酸化特异性抗体(兔多克隆抗-磷酸化-Ser473Akt多克隆和抗-磷酸化-ERK1/2抗体;Cell Signaling Technology)的Western印迹所显示。pan-ERK抗体证实了所有泳道中上样的蛋白量相等(图6)。用EM164抗体处理SaOS-2细胞并不抑制EGF-刺激的Erk1/2磷酸化,因此证实了EM164抗体对IGF-I受体信号传导通路抑制作用的特异性。
E.EM164抗体抑制IGF-I-、IGF-II-和血清-刺激的人肿瘤细胞的生长和存活
[131]在无血清条件下检测了几种人肿瘤细胞系,检测其对IGF-I的生长和存活反应。这些细胞系在IGF-I,IGF-II或血清存在的情况下用EM164抗体处理,采在2-4天后用MTT分析检测其生长和存活反应。大约1500个细胞被铺板在96孔板上,在含有血清的常规培养基中,第二天用无血清的培养基(添加了铁转运蛋白和BSA的无血清RPMI培养基,或者无酚红的培养基,如Dufourny,B.等,1997,J.Biol.Chem.,272,31163-31171所述)换液。在无血清培养基中生长1天后,细胞用大约75μL 10μg/mL的抗体孵育30min-3h,然后添加25μL的IGF-I(或IGF-II或血清)溶液,获得终浓度为10ng/mL的IGF-I,或20ng/mL的IGF-II,或0.04-10%血清。在一些试验中,在加入EM164抗体之前先用IGF-I刺激细胞15min,或一起加入IGF-I和EM164抗体。然后使细胞再生长2-3天。然后加入MTT溶液(3-(4,5)-二甲基噻唑-2-基)-2,5-二苯基四氮唑溴盐;25μL 5mg/mL的PBS溶液),细胞放回孵箱2-3h。然后去掉培养基,用100μL DMSO换液并混合,在545nm测量板的吸光度。几种人肿瘤细胞系显示在加入IGF-I或IGF-II或血清后的生长和存活反应明显被EM164抗体抑制,不管抗体是在IGF-I之前被加入,或IGF-I在抗体之前被加入,还是IGF-I和抗体一起被加入(表1)。
表1.EM164抗体对IGF-I-刺激的肿瘤细胞生长和存活的抑制作用
肿瘤细胞类型 | 对IGF-I作出响应时的生长倍数(MTT分析:在无血清培养基中,IGF-I处理/未处理细胞)a | 在无血清培养基中,EM164抗体对IGF-I刺激的生长的抑制百分率 | 在1.25-10%血清中,EM164抗体对细胞生长/存活的抑制作用b |
MCF-7(乳房) | 1.7-2.8 | 100% | 85% |
HT-3(宫颈) | 2 | 70-90% | ND |
Colo 205(结肠癌) | 2.3 | 50% | 是 |
HT-29 | 1.5 | 60% | 是 |
NCI-H838(肺癌) | 3 | 100% | 85-90% |
Calu-6 | 1.6-1.8 | 85% | 是 |
SK-LU-1 | 1.4 | 100% | 否 |
NCI-H596 | 1.4 | 100% | 微弱 |
A 549 | 1.2 | 80% | ND |
A 375(黑素瘤) | 1.6 | 90% | 否 |
SK-Mel-37 | 1.4 | 85% | ND |
RD(横纹肌肉瘤) | 1.7 | 85-100% | 是 |
SaOS-2(骨肉瘤) | 2.5 | 100% | 是 |
A431(表皮状瘤) | 2.2 | 85% | 是 |
SK-N-SH(成神经细胞瘤) | 2 | 85% | 30-50% |
a在含有5-10μg/mL EM164抗体的无血清培养基中,响应于10ng/mL IGF-I作用3至4天的细胞的生长/存活的MTT分析。
b基于与对照(有血清但无抗体)的比较,通过MTT分析或集落形成分析,在5-10μg/mL EM164抗体存在的情况下1.25-10%血清中细胞生长所受到的抑制;基于对照(无血清但有抗体,以及有血清但无抗体),定量测定MCF-7,NCI-H838和SK-N-SH细胞所受到的抑制的程度,以解释细胞的自分泌/旁分泌IGF刺激作用。
ND表示由于染色的难度而无数据或数据较小。
[132]EM164抗体强烈地抑制IGF-I或血清刺激的乳癌MCF-7细胞的生长和存活(图7和8)。在一个独立的实验中,EM164抗体强烈地抑制IGF-II刺激的MCF-7细胞生长和存活。以前关于应用市售抗体如IR3抗体的报道显示仅可微弱地抑制血清刺激的MCF-7细胞生长和存活,如在图7中IR3和1H7抗体所证实的(Cullen,K.J.等,1990,Cancer Res.,50,48-53)。与之相比,EM164抗体是血清或IGF刺激的MCF-7细胞生长的有效抑制剂。如在图8中显示,EM164抗体在很宽的血清浓度范围内(0.04-10%血清)都能同样有效地抑制MCF-7细胞的生长和存活。
[133]通过细胞计数来测定EM164抗体对MCF-7细胞的生长抑制。因此,在12孔板中,大约7500个细胞被铺板在含有10%FBS的RPMI培养基中,其中存在或者缺少10μg/mL的EM164抗体。生长5天后,未处理的对照样品的细胞计数为20.5×104个细胞,与之相比,EM164抗体处理的样品的细胞计数仅有1.7×104细胞。用EM164抗体进行的处理可在5天内抑制MCF-7细胞的生长大约12倍。EM164抗体的这种抑制作用明显高于已报道的使用IR3抗体在6天的分析中对MCF-7细胞产生的2.5倍的抑制作用(Rohlik,Q.T.等,1987,Biochem.Biophys.Res.Commun.,149,276-281)。
[134]与对照抗B4抗体相比,IGF-I和血清刺激的非小细胞肺癌系NCI-H838的生长和存活也可被EM164抗体强烈地抑制(图9)。对于NCI-H838和MCF-7细胞,在无血清培养基中用EM164抗体处理所产生的信号较未处理的样品小,推测其原因是EM164抗体也可抑制这些细胞的自分泌和旁分泌IGF-I和IGF-II刺激(图7和9)。HT29结肠癌细胞的集落大小也在用EM164抗体处理后大大减少。
[135]因此,EM164抗体在所有已知的抗IGF-I受体抗体中是独特的,它能有效地抑制肿瘤细胞如MCF-7细胞和NCI-H838细胞的血清刺激性生长超过80%。
[136]EM164抗体引起处于细胞周期的G0/G1期的细胞发生生长停滞,消除了IGF-I的促有丝分裂效应。为了进行细胞周期分析,在存在或不存在EM164(20μg/mL)的情况下用IGF-I(20ng/mL)处理MCF-7细胞1天,然后通过碘化丙啶染色和流式细胞仪进行分析。如在图25中所示,在缺少EM164时对IGF-I刺激作出反应的细胞的周期进程(41%细胞处于S期,50%处于G0/G1期)在EM164处理的细胞(仅有9%处于S期,77%细胞处于G0/G1期)中被抑制。
[137]除了对细胞增殖的抑制作用以外,EM164抗体的处理可导致细胞的凋亡。为了测定细胞凋亡,在存在或缺少EM164的情况下,用IGF-I或血清孵育NCI-H838肺癌细胞1天,测定胱天蛋白酶(caspase)对细胞角蛋白CK18蛋白的切割(图26)。在缺少EM164时,相对于无IGF-I的对照,添加IGF-I或血清产生了更低的胱天蛋白酶切割的CK18信号,这表明IGF-I和血清可防止胱天蛋白酶的活化。EM164的处理可抑制IGF-I和血清的抗凋亡效应,这可由在有EM164存在时所获得的被切割的CK18水平较缺少EM164时要高来说明(图26)。
F.EM164抗体与其他细胞毒性剂和细胞抑制剂协同抑制人肿瘤细胞的生长和存活
[138]EM164抗体与紫杉醇的联合施用较紫杉醇单独施用对非小细胞肺癌Calu6细胞明显更有抑制作用。同样,EM164抗体与喜树碱的联合较喜树碱单独施用对结肠癌HT29细胞的生长和存活明显更有抑制作用。因为并不期望单独的EM164抗体像有机化学毒性药物一样对细胞有毒性,所以在临床条件下,EM164抗体显著的细胞生长抑制效应和化学毒性药物的细胞毒性效应的协同在癌症的联合治疗中是非常有效的。
[139]EM164抗体与抗EGF受体抗体(KS77)的联合效应较单独的EM164抗体或KS77抗体对数种肿瘤细胞系如HT-3细胞,RD细胞,MCF-7细胞和A431细胞的生长和存活具有更明显的抑制作用。因此,将针对两种生长因子受体如IGF-I受体和EGF受体的中和抗体联合起来所产生的协同效应在临床的癌症治疗中也是有用的。
[140]根据EM164抗体作为单一试剂在抑制不同人类癌症细胞系中的效力,如表1所示,使用EM164抗体与其他抗-癌症治疗剂的联合,进行了额外的效力研究。在这些对不同癌症细胞系的研究中,比起或者单独使用EM164或者单独使用其他治疗剂,EM164抗体和其他抗-肿瘤治疗剂的联合治疗产生甚至更加大的抗-癌症效力。因此,EM164抗体和其他抗-肿瘤治疗剂的这些联合应用在抑制癌症细胞增殖和存活中高度有效。可以与EM164联合用于改进抗癌症效力的治疗剂包括用于肿瘤学实践的多种药剂(参考:Cancer,Principles & Practice of Oncology,DeVita,V.T.,Hellman,S.,Rosenberg,S.A.,第六版,Lippincott-Raven,Philadelphia,2001),诸如多烯紫杉醇、紫杉酚、阿霉素、表阿霉素、环磷酰胺、曲妥珠(Herceptin)、卡培他宾、它莫西芬、托瑞米芬、来曲唑、阿纳曲唑、氟维司群、依西美坦、戈舍瑞林、奥沙利铂、卡铂、顺铂、地塞米松、抗排卵肽、贝伐单抗(Avastin)、5-氟尿嘧啶、亚叶酸、左旋咪唑、依立替康、依托泊甙、拓朴替康、吉西他滨、长春瑞滨、雌莫司汀、米托蒽醌、阿巴瑞克、唑来瞵酸盐、链硝脲、妥昔(Rituxan)、柔红霉素、白消安、苯丁酸氮芥、氟阿糖腺苷、伊马替尼、阿糖胞苷、伊莫单抗(Zevalin)、托西莫单抗(Bexxar)、α干扰素-2b、美法仑、硼替佐米(Velcade)、六甲蜜胺、门冬酰胺酶、吉非替尼(Iressa)、埃洛尼替(Tarceva)、抗-EGF受体抗体(Cetuximab、Abx-EGF)、埃坡霉素,以及细胞毒性药剂和针对细胞-表面受体的抗体形成的偶联物。
[141]对于这些联合治疗,EMl64与一种或多种具有不同的作用机制的抗-癌症药剂联合,诸如烷化剂、铂试剂、激素治疗、抗代谢物、拓扑异构酶抑制剂、抗微管剂、分化剂(differentiation agent)、抗血管生成或抗血管化作用治疗、放射治疗、促黄体生成激素释放激素(LHRH)或促性腺激素-释放激素(GnRH)的激动剂和拮抗剂、抑制性抗体或针对细胞-表面受体的小分子抑制剂,和其他化学治疗剂(参考:Cancer,Principles & Practice of Oncology,DeVita,V.T.,Hellman,S.,Rosenberg,S.A.,第六版,Lippincott-Raven,Philadelphia,2001)。在一个实施例中,LHRH拮抗剂抗排卵肽(0.1至10微摩尔)和EM164抗体(0.1至10纳摩尔)的联合,比或者单独使用EM164或者单独使用抗排卵肽,更显著地抑制MCF-7乳癌细胞的增殖。在与铂试剂的联合治疗的实施例中,比起或者单独使用EM164或者单独使用顺铂的抑制,EM164抗体(10微克/ml)和顺铂(0.1-60微克/ml)的联合治疗,产生对MCF-7乳癌细胞的增殖和存活的更大的抑制。
[142]EM164抗体与其他治疗剂的这些联合应用能有效地抵抗若干癌症,包括乳癌、肺癌、结肠癌、前列腺癌、胰腺癌、宫颈癌、卵巢癌、黑素瘤、多发性骨髓瘤、神经母细胞瘤、横纹肌肉瘤和骨肉瘤。EM164抗体和治疗剂或者可以同时给予或者可以按照顺序给予,以进行癌症治疗。
[143]EM164抗体与细胞毒性药物的偶联物在将所述细胞毒药物向过量表达IGF-I受体的肿瘤进行靶向输送的过程中也是有价值的。EM164抗体与放射性标记或其它标记的偶联物可用于过量表达IGF-I受体的肿瘤的治疗和成像。
G.EM164作为单一试剂或与抗癌试剂联合进行处理时对免疫缺陷小鼠中人癌症异种移植物的影响
[144]人非小细胞肺癌Calu-6异种移植物是通过在免疫缺陷小鼠中皮下注射1×107个Calu-6细胞而建立的。如在图10中显示,含有已建立的100m3 Calu-6异种移植物的这些小鼠单独用EM164抗体处理(静脉注射6次,每次0.8mg/小鼠,每周两次)或单独用紫杉醇处理(腹腔注射5次紫杉醇;每两天1次;每次15mg/kg),或联合紫杉醇和EM164抗体进行处理,或单独给予PBS(200μL/小鼠,6次注射,每周2次,静脉注射),每个处理组使用5只小鼠。与PBS对照相比,用EM164抗体处理可明显延缓肿瘤的生长。基于小鼠体重的测量值,没有观察到EM164抗体的毒性。尽管单独的紫杉醇的有效性持续到14天,但到那时肿瘤才开始消退。然而,与单独用紫杉醇进行处理的组相比,在紫杉醇和EM164抗体联合处理的组中,肿瘤的生长明显被延缓。
[145]人胰腺癌异种移植物是通过向5周龄的雌性SCID/ICR小鼠(Taconic)皮下注射含有107个BxPC-3细胞的PBS而建立的(0天)。然后带有80mm3的已建立肿瘤的小鼠用EM164单独处理(13次注射,每次0.8mg/小鼠,尾侧静脉注射,在第12、16、19、23、26、29、36、43、50、54、58、61和64天进行注射),用吉西他滨(吉西他滨)单独处理(腹腔注射两次,每次150mg/kg/小鼠,在第12和19天),用吉西他滨和EM164按照上述方案进行联合处理,单独给予PBS,和单独给予对照抗体(使用与EM164相同的方案),这5个处理组中的每一个都使用5只小鼠。如在图27中所示,EM164的单独处理或与吉西他滨的联合处理都在一开始便导致肿瘤异种移植物的总体消退,在EM164处理组的5只动物中有4只,在联合处理组的所有5只动物中出现了这样的消退。仅在EM164组中的第43天和联合处理组中的第68天见到一只以上的动物发生可测量到的肿瘤再生,这导致了在第74天得到的平均肿瘤体积与对照处理组相比要明显减小(P值分别为0.029和0.002;双侧T-检验;图27)。在另一项研究中,EM164抗体的处理(单独使用或与抗EGF受体抗体联合使用;腹腔注射)可抑制小鼠中已建立的BxPC-3异种移植物的生长。
[146]鼠EM164和人源化的EM164抗体对于小鼠中建立的BxPC-3异种移植物显示出等同的抑制作用,因此证明了人源化的EM164的效价在体内与鼠EM164的效价相同。在不同的EM164抗体给药方式比较中,腹膜内和静脉内注射EM164抗体对于小鼠中建立的BxPC-3异种移植物的生长显示出等同的抑制作用。在另一异种移植物研究中,用EM164抗体治疗显示出能显著延缓小鼠中建立的A-673人横纹肌肉瘤/尤文肉瘤异种移植物的生长。
H.EM164抗体的轻链和重链的克隆和测序
[147]从EM164杂交瘤细胞中纯化出总RNA。使用4-5μg总RNA和寡聚dT或随机六聚物引物进行逆转录酶反应。
[148]采用Co等人所述的RACE方法(J.Immunol.,148,1149-1154(1992))和Wang等人所述的简并引物(J.Immunol.Methods,233,167-177(2000))进行PCR反应。RACE PCR方法需要一个中间步骤来将多聚G尾添加至第一条cDNA链的3’末端上。用Qianeasy(Qiagen)柱纯化RT反应,用50μl的1XNEB缓冲液4洗脱。用0.25mM CoCl2,1mM dGTP和5units的末端转移酶(NEB)在1X NEB缓冲液4中对所述洗脱液进行dG加尾反应。混合物在37℃孵育30分钟,然后将1/5的反应物(10μl)直接加入到PCR反应中作为模板DNA。
[149]除了引物和模板的差异以外,RACE和简并PCR反应是完全相同的。对于RACE PCR是直接使用末端转移酶反应物作为模板,而简并PCR反应是直接使用RT反应混合物。
[150]在RACE和简并PCR反应中,使用相同的3′轻链引物:HindKL-tatagagctcaagcttggatggtgggaagatggatacagttggtgc(SEQ ID NO:14)和3′重链引物:
Bgl2IgG1-ggaagatctatagacagatgggggtgtcgttttggc(SEQ ID NO:15)。
[151]在RACE PCR中,1个多聚C 5′引物同时用于重链和轻链:
EcoPolyC-TATATCTAGAATTCCCCCCCCCCCCCCCCC(SEQ ID NO:16),
而简并5′末端PCR引物为:
对于轻链,为Sac1MK-GGGAGCTCGAYATTGTGMTSACMCARWCTMCA(SEQID NO:17);
对于重链,为下列引物的等量混合物:
EcoR1MH1-CTTCCGGAATTCSARGTNMAGCTGSAGSAGTC(SEQID NO:18)和EcoR1MH2-CTTCCGGAATTCSARGTNMAGCTGSAGSAGTCWGG(SEQ ID NO:19)。
[152]在上述的引物序列中,混合碱基如下定义:H=A+T+C,S=g+C,Y=C+T,K=G+T,M=A+C,R=A+g,W=A+T,V=A+C+G。
[153]使用下述程序进行PCR反应:1)94℃,3分钟,2)94℃,15秒,3)45℃,1分钟,4)72℃,2分钟,5)返回步骤#2循环29次,6)在72℃最后延伸10分钟,PCR反应完成。
[154]采用限制性酶,将通过PCR引物产生的PCR产物克隆进pBluescript II SK+(Stratagene)。
[155]通过常规的方法,对若干独立的轻链和重链克隆测序以鉴定和避免聚合酶产生的可能的序列错误(图12和13)。使用Chothia典型分类定义(Chothia canonical classification definitions),鉴定出三个轻链和重链CDRs(图12-14)。
[156]搜索NCBI IgBlast数据库,表明抗-IGF-I受体抗体的轻链可变区可能衍生自小鼠IgVk Crl胚系基因,而重链可变区可能衍生自IgVhJ558.c胚系基因(图15)。
[157]对鼠EM164抗体蛋白进行测序,以证实图12和13中显示的序列。纯化的EM164抗体的重链和轻链条带从凝胶(SDS-PAGE,还原条件)转移至PVDF膜上,从该PVDF膜上切下,通过蛋白测序进行分析。所述轻链的N-末端序列通过Edman测序测定为:DVLMTQTPLS(SEQ ID NO:20),这与从EM164杂交瘤获得的克隆轻链基因的N-端序列相匹配。
[158]发现重链的N-末端对于Edman蛋白测序来说是封闭的。质量为1129.5(M+H+,单同位素)的重链胰蛋白酶消化肽片段通过源后衰减(post-source decay)(PSD)分成片段,其序列测定为GRPDYYGSSK(SEQ ID NO:21)。质量为2664.2(M+H+,单同位素)的另一个重链胰蛋白酶消化肽片段也通过源后衰减(PSD)分成片段,其序列鉴定为:SSSTAYMQLSSLTSEDSAVYYFAR(SEQ ID NO:22)。这两个序列都与从EM164杂交瘤获得的克隆重链基因的CDR3和框架3(FR3)的序列完全匹配。
I.EM164抗体的重组表达
[159]轻链和重链成对序列被克隆进单一的哺乳动物表达载体中(图16)。人可变序列的PCR引物产生了一些限制性位点,它们可使人信号序列在pBluescriptII克隆载体中被连接,所述可变序列被克隆进哺乳动物表达质粒中,轻链或重链的克隆分别使用了EcoRI和BsiWI或HindIII和ApaI位点(图16)。轻链可变序列符合读框地(in-frame)被克隆到人IgK恒定区上,重链可变序列被克隆到人Igγ恒定区序列中。在最终的表达质粒中,人CMV启动子可驱动轻链和重链cDNA序列的表达。根据本领域中熟知的方法进行重组鼠EM164抗体的表达和纯化。
实施例2:人源化版本的EM164抗体
[160]大体上根据美国专利5,639,641和如下公开的原理和方法进行EM164抗体的表面重构,以提供适合作为治疗或诊断试剂的人源化抗体形式。
A.表面预测
[161]为了预测鼠抗IGF-I受体抗体(EM164)可变区的表面残基,应用了一组具有溶解结构的抗体的可变区残基的溶剂可及性(solventaccessibility)。用MC软件包来计算一组127个独特抗体结构文件(表2)的氨基酸的溶剂可溶性(Pedersen等人,1994,J.Mol.Biol.,235,959-973)。通过序列比对,从这127个结构中确定出10条最相似的轻链和重链氨基酸序列。计算每个可变区残基的平均溶剂可及性(averagesolvent accesibility),具有超过30%的平均可及性的位点被认为是表面残基。通过仅对具有两个相同的侧翼残基的那些结构计算各个残基的可及性,进一步检验平均可及性在25%和35%之间的位点。
表2 来自Brookhaven数据库的用来预测抗IGF-I受体抗体(EM164)的表面的127个抗体结构
用于表面预测的127个Brookhaven结构文件 | |||||||||
2rcs2gfb1yuh1sm31nsn1ncd1mcp1jrh1igi1gpo1fpt1fai | 3hfl2h1p2bfv1tet1opg1nfd1mfb1kb51igm1hil1frg1fbi1dbl | 3hfm2hfl2cgr1vfa1osp1ngp1mim1kel1igt1hyx1fvc1fdl1dfb | 1aif1a6t8fabglb21aj71acy15c81ap21ad01a0q1aqk1ad91a3l | 1a3r1axt1ae61a4j1ay11afv1a5f1b2w1baf1bjm1bln1bbd1bfo | 1bbj1bog1bvl1cly1clz1cbv1axs1adq1cfv1clo1d5b1f581eap | 43c92hrp2dbl1vge1plg1nld1mlb1kip1igy1iai1gaf1fgv1dsf | 4fab2jel2f191yec1psk1nma1mpa1kir1ikf1ibg1ggi1fig1dvf | 6fab2mcp2tb41yed1rmf1nmb1nbv1lve1jel1igc1ghf1flr | 7fab2pcp2fbj1yee1sbs1nqb1ncb1mam1jhl1igf1gig1for |
B.分子建模:
[162]鼠EM164的分子模型采用Oxford分子软件包AbM来产生。抗体构架(framework)的建立是来自具有最相似的氨基酸序列的抗体的结构文件,对于轻链,文件是2jel,对于重链则是1nqb。通过搜索含有非冗余溶解结构(non-redundant solved structures)的C-α结构数据库可建立非正式的CDR(non-canonical CDRs)。CDR的5范围内的残基被确定。
C.人Ab选择
[163]将鼠EM164的表面位置与Kabat数据库(Johnson,G.和Wu,T.T.(2001)Nucleic Acids Research,29:205-206)中人抗体序列的相应位置进行比较。应用抗体数据库管理软件SR(Searle 1998)从天然的重链和轻链人抗体对中提取和比对抗体表面残基。特别考虑到进入CDR的5范围内的位点,选择了具有最相同的表面残基的人抗体表面,以替换鼠抗IGF-I受体抗体的表面残基。
D.PCR诱变
[164]在鼠EM164 cDNA克隆(见上)上进行PCR诱变以构建表面重构的人EM164(在此标为huEM164)。引物组被设计,以产生huEM164的所有受测形式所需的8个氨基酸变化,额外的引物被设计,以选择性地产生两种5残基的变化(表3)。PCR反应按下面的程序来执行:
1)94℃1分钟,2)94℃15秒,3)55℃1分钟,4)72℃1分钟,5)返回#2步骤,循环29次,6)最后在72℃进行最后的延伸步骤4分钟,完成反应。PCR产物用其相应的限制性酶消化,如上所述地克隆进pBluescript克隆载体。克隆被测序以证实所需的氨基酸变化。
表3 用来构建4种人源化EM164抗体的PCR引物
引物 | 序列 | SEQ IDNO: |
Em164hcvv | CAGGTGTACACTCCCAGGTCCAACTGGTGCAGTCTGGGGCTGAAGTGGTGAAGCCTG | 23 |
Em164hcqqgo11 | CAATCAGAAGTTCCAGGGGAAGGCCACAC | 24 |
Em164hcqqgo12 | CCTTCCCCTGGAACTTCTGATTGTAGTTAGTACG | 25 |
Em1641cv3 | CAGGTGTACACTCCGATGTTGTGATGACCCAAACTCC | 26 |
Em164lcl3 | CAGGTGTACACTCCGATGTTTTGATGACCCAAACTCC | 27 |
Em1641cp18 | GACTAGATCTGCAAGAGATGGAGGCT | 28 |
GGATCTCCAAGAC | ||
Em164lcbg12 | TTGCAGATCTAGTCAGAGCATAGTACATAGTAATG | 29 |
Em164r45 | GAATGGTACCTGCAGAAACCAGGCCAGTCTCCAAGGCTCCTGATCTAC | 30 |
Em164a67o11 | GTGGCAGTGGAGCAGGGACAGATTTCAC | 31 |
Em164a67o12 | GAAATCTGTCCCTGCTCCACTGCCACTG | 32 |
E.可变区表面残基
[165]由Pedersen等人(J.Mol.Biol.,235,959-973,1994)和Roguska等人(Protein Eng.,9,895-904,1996)描述的抗体表面重构技术是从预测鼠抗体可变序列的表面残基开始的。表面残基被定义为其总表面积的至少30%可与水分子接触(accessible to a water molecule)的氨基酸。
[166]在这127个抗体结构文件组中10个最同源的抗体被鉴定出(图17和18)。将这些被比对的序列中的每个Kabat位置的溶剂可及性进行平均化,各个残基的相对可及性的分布显示在图19中。轻链和重链均有26个残基的平均相对可及性(relative accessibilities)至少为30%(图19):因此这些残基是EM164的预测的表面残基。有数个残基的平均可及性在25%和35%之间,这些残基被进一步检验,这通过仅对所述残基两侧带有两个完全相同的残基的抗体进行平均化来实施(表4和5)。在进行这样的附加分析后,上面最初鉴定出的表面残基组仍然没有改变。
表4 EM164抗体轻链和重链可变序列的表面残基和平均可及性(ave.acc.)
EM164表面残基 | |||||
轻链 | 重链 | ||||
EM164 | Kabat# | Ave.Acc. | EM164 | Kabat# | Ave.Acc. |
D | 1 | 45.89 | Q | 1 | 58.19 |
L | 3 | 41.53 | Q | 3 | 34.08 |
T | 7 | 31.40 | Q | 5 | 34.36 |
L | 9 | 50.08 | A | 9 | 38.01 |
L | 15 | 35.45 | L | 11 | 47.72 |
Q | 18 | 39.79 | K | 13 | 46.51 |
R | 24 | 34.36 | P | 14 | 31.49 |
S | 26 | 32.63 | G | 15 | 3 1.42 |
Q | 27 | 34.35 | K | 19 | 34.41 |
N | 28 | 36.38 | K | 23 | 31.23 |
P | 40 | 43.05 | T | 28 | 36.24 |
G | 41 | 46.56 | P | 41 | 44.01 |
Q | 42 | 34.92 | G | 42 | 42.62 |
K | 45 | 30.58 | Q | 43 | 46.85 |
S | 52 | 30.40 | E | 61 | 46.68 |
S | 56 | 41.46 | K | 62 | 44.87 |
G | 57 | 42.41 | K | 64 | 38.92 |
D | 60 | 45.96 | R | 65 | 40.06 |
S | 67 | 38.20 | K | 73 | 35.92 |
R | 77 | 42.61 | S | 74 | 48.91 |
E | 81 | 38.46 | S | 82B | 32.72 |
V | 95E | 34.83 | S | 84 | 35.21 |
K | 103 | 31.10 | E | 85 | 39.62 |
K | 107 | 36.94 | D | 98 | 36.00 |
R | 108 | 60.13 | A | 106 | 37.65 |
A | 109 | 53.65 | S | 113 | 43.42 |
表5
边界表面残基 | |||||
轻链 | 重链 | ||||
EM164 | Kabat# | Ave.Acc. | EM164 | Kabat# | Ave.Acc. |
T | 5 | 28.68 | Q | 3 | 31.62 |
T | 7 | 30.24 | Q | 5 | 36.07 |
P | 12 | 26.59 | P | 14 | 29.88 |
G | 16 | 25.20 | G | 15 | 30.87 |
D | 17 | 25.73 | S | 17 | 25.64 |
S | 20 | 25.37 | K | 19 | 35.06 |
R | 24 | 36.73 | K | 23 | 31.48 |
S | 26 | 31.00 | G | 26 | 30.53 |
Q | 27 | 32.29 | S | 31 | 27.12 |
S | 27A | 29.78 | R | 56 | NA |
V | 27C | 29.05 | T | 68 | 27.71 |
V | 29 | NA | T | 70 | 24.65 |
Q | 42 | 34.92 | S | 75 | 18.80 |
K | 45 | 32.24 | S | 82B | 32.87 |
S | 52 | 30.02 | P | 97 | NA |
R | 54 | 29.50 | Y | 99 | NA |
D | 70 | 26.03 | V | 103 | NA |
R | 74 | NA | T | 111 | 25.95 |
E | 79 | 26.64 | |||
A | 80 | 29.61 | |||
V | 95E | 42.12 | |||
G | 100 | 29.82 | |||
K | 103 | 31.10 | |||
E | 105 | 25.78 |
平均可及性在25%和35%之间的残基被进一步分析,通过对在所述疑问残基两侧有两个完全相同的残基的抗体子集进行平均来实施。这些临界表面位置及其新的平均可及性值被给出。NA是指在这10个最相似的抗体中没有相同侧翼残基的残基。
F.用分子建模来测定哪个残基落入CDR的5内
[167]对上述用AbM软件包产生的分子模型进行分析,以确定在CDR的5范围内的EM164表面残基。为了重构鼠EM164抗体,在CDR之外的所有表面残基都应该变为在人类中的对应物(counterparts),但在CDR的5范围之内的残基要被特别谨慎地对待,因为它们对抗原特异性也可能有贡献。因此,后者的这些残基必须被鉴定出,并在整个人源化过程中都要对它们进行谨慎考虑。在表面重构中被使用的CDR定义结合了重链CDR2的AbM定义和剩余的5个CDRs的Kabat定义(图14)。表6显示了在EM164模型的轻链或重链序列内,处于任何CDR的残基的5范围内的残基。
表6在CDR的5内的EM164抗体框架表面残基CDR的5内的EM164表面残基
轻链 | 重链 |
D1 | T28 |
L3 | K73 |
T7 | S74 |
P40 | |
Q42 | |
K45 | |
G57 | |
D60 | |
E81 |
G.最同源的人表面的鉴定
[168]用于重构EM164的候选人抗体表面在Kabat抗体序列数据库中用SR软件鉴定出来,该软件仅基于抗体数据库来搜索特定的残基位点。为了保留天然的配对,轻链和重链的表面残基一起进行比较。来自Kabat数据库的最同源的人表面按照序列同一性的等级次序进行排列。最顶端的5个表面在表7中给出。然后比较这些表面以鉴定出它们中的哪一个在CDR的5之内需要的变化最小。白血病B细胞抗体,CLL 1.69,要求的表面残基变化数目最小(总共10个),而在这些残基中仅有2个位于CDR的5内。
[169]EM164的全长可变区序列也可与Kabat人抗体数据库进行比对,CLL 1.69再次被鉴定为最相似的人可变区序列。共同地,这些序列比较将人白血病B细胞抗体CLL 1.69鉴定为EM164的人表面的优选选择。
表7从Kabat数据库中提取出的最同源的5个人类序列
5个最同源的人抗体表面 |
抗体 | 轻链 | SEQ IDNO: |
MuEM164 | DLTLLQ PG OKGDSR EKKRA | 33 |
CLL1.69 | DVTLLPPGQRGDAREKKR- | 34 |
MSL5 | DQSLIPPGQKGDSRDKKRA | 35 |
CDP571 | DMSSVRPGQKGSSSDKKR- | 36 |
LC3aPB | EVSGPRPGQRGDSREKKR- | 37 |
SSbPB | EVSGPRPGQRGDSREKKR- | 38 |
抗体 | 重链 | SEQ IDNO: |
MuEM164 | QQQALKPGKK TPGQEKKR KSSSEAS | 39 |
CLL1.69 | QQVAVKPGKKTPGQQKQGKSSSEQS | 40 |
MSL5 | QQQPLKPGKKTPGKDDKGTSNNEQS | 41 |
CDP571 | QQVAVKPGKKTPGQQKKGKSSSEQS | 42 |
LC3aPB | -QVAVKPGKKTPGQQKQGKSSSEQS | 43 |
SSbPB | -QVAVKPGKKTPGQQKQGESSSEQS | 44 |
通过SR产生比对(Pedersen 1993)。位于CDR 5内的EM164表面残基被下划线标记。
H.构建人源化EM164基因
[170]使用如上所述的PCR诱变技术对EM164的10个表面残基(表7)进行变化。因为CLL 1.69的表面残基中的8个不在CDR的5之内,因此这些残基在所有形式的人源化EM164中均从鼠源性转变为人源性(表8和9)。在CDR的5内的两个轻链表面残基(Kabat位点3和45)被改变为人源性或保留为鼠源性。这些选择共同产生出4种构建的EM164人源化形式(图22和23)。
[171]在这4种人源化形式中,1.0型(1.0 version)具有所有的10个人源性表面残基。就CDR附近的改变而言,最保守的形式是1.1型,它保留了在CDR的5之内的所有鼠源性表面残基。所有4种人源化EM164抗体基因都被克隆进抗体表达质粒中(图16)以便用于暂时的和稳定的转染。
表8制备人源化EM164抗体1.0-1.3型时的残基变化
所有形式中均有的变化 | |||||
轻链:muQ18全huP18;muS67全huA67重链:muQ5至huV5;muL11至huV11;muE61至huQ61;muK64至huQ64;muR65至huG65;muA106至huQ106 | |||||
HuEM164变化 | |||||
轻链aa3 | 轻链aa45 | 总数,在5内 | |||
Mu | Hu | mu | hu | 鼠源性残基 | |
V1.0 | V | R | 0 | ||
V1.1 | L | K | 2 | ||
V1.2 | L | R | 1 | ||
V1.3 | V | K | 1 |
I.比较人源化EM164抗体与鼠源性EM164抗体对全长IGF-I受体和截短的IGF-I受体α链结合的亲和性
[172]人源化EM164抗体1.0-1.3的亲和性与鼠源性EM164抗体的亲和性是通过结合竞争分析(binding competition assays)来比较的,其中使用了生物素化全长人IGF-I受体或myc-表位标记的截短的IGF-I受体α链,如上所述。通过在人胚胎肾293T细胞中暂时转染合适的表达载体来获得人源化EM164抗体样本,抗体浓度通过ELISA利用纯化的人源化抗体标准品进行测定。为了进行ELISA结合竞争测定,将人源化抗体样本和多种浓度的鼠EM164抗体的混合物与间接俘获的生物素化全长IGF-I受体或myc-表位标记的截短的IGF-I受体α链一起孵育。平衡后,使用羊抗人Fab’2-抗体-辣根过氧化物酶偶联物检测结合的人源化抗体。就([结合的鼠Ab]/[结合的人源化Ab])对([鼠Ab]/[人源化Ab])进行作图,它理论上产生一条直线,斜率=(Kd人源化Ab/Kd鼠 Ab),由此可以确定人源化抗体和鼠抗体的相对亲和性。
[173]竞争分析的一个实例在图11中显示。Immulon-2HB ELISA板按每孔100μL 5μg/mL的链亲和素在室温用碳酸盐缓冲液包被7h。链亲和素包被的孔用200μL封闭缓冲液(10mg/mL BSA,在TBS-T缓冲液中)封闭1h,用TBS-T缓冲液清洗,与生物素化IGF-I受体(每孔5ng)4℃孵育过夜。然后含有间接俘获的生物素化IGF-I受体的孔被洗涤,与人源化EM164抗体(15.5ng)和鼠抗体(0ng,或16.35ng,或32.7ng,或65.4ng,或163.5ng)的混合物在100μL的封闭缓冲液中室温孵育2h,然后在4℃孵育过夜。然后用TBS-T缓冲液清洗各孔,并与羊抗人-Fab’2-抗体-辣根过氧化物酶偶联物(100μL;1μg/mL,在封闭缓冲液中)孵育1h,然后多次洗涤,使用ABTS/H2O2底物在405nm处检测。
[174]([结合的鼠Ab]/[结合的人源化Ab])对([鼠Ab]/[人源化Ab])的曲线产生一条直线(r2=0.996),斜率(=Kd人源化Ab/Kd鼠Ab)为0.52。因此人源化抗体1.0型较鼠EM164抗体与IGF-I受体的结合更紧密。对于人源化EM164抗体1.0,1.1,1.2和1.3型与鼠EM164抗体对全长IGF-I受体或截短的IGF-I受体α链的结合竞争,可获得类似的梯度值,范围为大约0.5至0.8,这表明所有的EM164抗体人源化形式都具有相似的亲和性,它们都高于作为母本的鼠EM164抗体的亲和性。在重链中带有92F→C突变的EM164抗体的嵌合形式在与鼠EM164抗体的一个类似的结合竞争中显示出大约为3的斜率,表明EM164的92F→C突变体结合IGF-I受体的亲和性较鼠EM164抗体低3倍。人源化EM1641.0型抗体显示出对IGF-I刺激的MCF-7细胞生长和存活的抑制作用,这与鼠EM164抗体相似(图24)。人源化EM164 1.0型抗体对血清刺激的MCF-7细胞生长和存活的抑制作用与鼠EM164抗体的抑制作用相似。
表9
片段 | 轻链 | 重链 |
FR1 | 1-23(在0处偶尔有一个残基,在Vλ链中的10处有一个删除) | 1-30(在0处偶尔有一个残基) |
CDR1 | 24-34(含有可能的插入物,编号为27A,B,C,D,E,F) | 31-35(含有可能的插入物,编号为35A,B) |
FR2 | 35-49 | 36-49 |
CDR2 | 50-56 | 50-65(含有可能的插入物,编号为52A,B,C) |
FR3 | 57-88 | 66-94(含有可能的插入物,编号为82A,B,C) |
CDR3 | 89-97(含有可能的插入物,编号为95A,B,C,D,E,F) | 95-102(含有可能的插入物,编号为100A,B,C,D,E,F,G,H,I,J,K) |
FR4 | 98-107(含有可能的插入物,编号为106 A) | 103-113 |
将Kabat编号系统用于不同形式的EM164 Ab的轻链和重链可变区多肽。根据在多肽链中的位置,将氨基酸残基分组为构架(FR)和互补性决定区(CDR)。取自Kabat等人,Sequences of Proteins of Immunological Interest,第五版,1991,NTH Publication No.91-3242
J.以在此所述的鼠源性和人源化抗体序列作为开始来提供改良的抗IGF-I受体抗体的过程
[175]抗IGF-I受体抗体EM164及其人源化变体的氨基酸和核酸序列被用来开发其他的具有改良特性的抗体,它们也在本发明的范围内。所述改良的特性包括与IGF-I受体的亲和性增加。基于初级抗体(primary antibody)序列的知识,几个研究调查了在抗体序列的不同位点导入一个或多个氨基酸变化对其特性如结合作用和表达水平的影响(Yang,W.P.等,1995,J.Mol Biol,254,392-403;Rader,C.等,1998,Proc.Natl.Acad.Sci.USA,95,8910-8915;Vaughan,T.J.等,1998,NatureBiotechnology,16,535-539)。
[176]在这些研究中,初级抗体的变体已经通过在CDR1,CDR2,CDR3或构架区中改变重链和轻链基因的序列而产生,采用的方法如寡核苷酸介导的位点定向诱变,盒式诱变,易错PCR,DNA重排,或利用大肠杆菌的增变株(Vaughan,T. J.等,1998,Nature Biotechnology,16,535-539;Adey,N.B.等,1996,16章,277-291页,在″Phage Display ofPeptides and Proteins″,Kay,B.K.等人主编,Academic Press)。通过应用标准的筛选技术,改变初级抗体的序列的这些方法已经获得了亲和性增加的次级抗体(secondary antibodies)(Gram,H.等,1992,Proc.Nad.Acad.Sci.USA,89,3576-3580;Boder,E.T.等,2000,Proc.Natl.Acad.Sci.USA,97,10701-10705;Davies,J.和Riechmann,L.,1996,Immunotechnolgy,2,169-179;Thompson,J.等,1996,J.Mol.BioL,256,77-88;Short,M.K.等,2002,J.BioL Chem.,277,16365-16370;Furukawa,K.等,2001,J.Biol.Chem.,276,27622-27628)。
[177]通过改变抗体的一个或多个氨基酸残基的一种相似的定向策略,在本发明中所述的抗体序列可以被用来开发具有改良功能的抗IGF-I受体抗体,如具有合适基团的抗体,所述基团例如在方便进行共价修饰的连接点处的游离氨基基团或硫醇基团,这可以在例如治疗试剂的连接中被应用。
K.鼠的,嵌合的和其它的抗IGF-I受体抗体的可供选择的表达系统
[178]也可利用那些与用来表达人源化抗体的表达质粒(上述)相类似的哺乳动物表达质粒来表达鼠抗IGF-I受体抗体。具有鼠恒定区,包括轻链κ和重链γ-1序列的表达质粒是已知的(McLean等,2000,MolImmunol.,37,837-845)。这些质粒被设计用来接受任何抗体的可变区,如鼠抗-IGF-I受体抗体,这通过简单的限制性消化和克隆来实施。为了建立与表达质粒相容的限制性,通常需要对抗IGF-I受体抗体进行额外的PCR方法。
[179]表达完整鼠抗IGF-I受体抗体的一种可选择的方法是取代嵌合的抗IGF-I受体抗体表达质粒中的人恒定区。利用可变区的序列盒以及轻链和重链恒定区的序列盒来构建嵌合表达质粒(图16)。正如抗体可变区序列可通过限制性消化被克隆进这种表达质粒中一样,单独的限制性消化被用于任何恒定区序列的克隆。κ轻链和γ-1重链cDNA被克隆,所述克隆例如源自鼠杂交瘤RNA,如所述的用于克隆抗IGF-I抗体可变区的RNA。同样,合适的引物可以利用Kabat数据库中获得的序列进行设计(见表10)。例如,应用RT-PCR来克隆恒定区序列,并创造出将这些片段克隆进嵌合抗IGF-I受体抗体表达质粒中所需的限制性位点。然后这种质粒被用于在标准的哺乳动物表达系统如CHO细胞系中表达完整的鼠抗-IGF-I受体抗体。
表10被设计用来分别克隆鼠γ-1恒定区和鼠κ恒定区的引物
鼠恒定区引物 | ||
引物名称 | 引物序列 | SEQ ID NO: |
MuIgG1 C3endX | TTTTTGAGCTCTTATTTACCAGGAGAGTGGGAGAGGCTCTT | 45 |
MuIgG1 C5endH | TTTTAAGCTTGCCAAAACGACACCCCCATCTGTCTAT | 46 |
MuIgKap C3endB | TITTGGATCCTAACACTCATTCCTGTTGAAGC | 47 |
MuIgKao C5endE | TTTTGAATTCGGGCTGATGCTGCACCAACTG | 48 |
引物根据Kabat数据库中获得的序列进行设计(Johnson,G和Wu,T.T.(2001)Nucleic Acids Research,29:205-206)
保藏声明
[180]根据布达佩斯条约的条款,能产生鼠EM164抗体的杂交瘤于2002年6月14日保藏于美国典型培养物保藏中心(American TypeCulture Collection),PO Box 1549,Manassas,VA 20108,保藏登记号(Accession Number)为PTA-4457。
[181]在本公开中引用到了某些专利和已印制的出版物,其教导的内容在此分别以其整体引入作为参考。
[182]尽管本发明已经参照其具体的实施方案作了详细描述,但对于本领域的专业技术人员,明显的是,对其所作的各种变化和修饰不背离其精神和范围。
序列表
<110>伊缪诺金公司(Immunogen,Inc.)
<120>抗IGF-I受体抗体
<130>F190322
<140>10/729,441
<141>2003-12-08
<150>10/170,390
<151>2002-06-14
<160>96
<170>PatentIn version 3.3
<210>1
<211>5
<212>PRT
<213>人工序列
<220>
<223>抗体重链互补决定区
<400>1
Ser Tyr Trp Met His
1 5
<210>2
<211>17
<212>PRT
<213>人工序列
<220>
<223>抗体重链互补决定区
<400>2
Glu Ile Asn Pro Ser Asn Gly Arg Thr Asn Tyr Asn Glu Lys Phe Lys
1 5 10 15
Arg
<210>3
<211>15
<212>PRT
<213>人工序列
<220>
<223>抗体重链互补决定区
<400>3
Gly Arg Pro Asp Tyr Tyr Gly Ser Ser Lys Trp Tyr Phe Asp Val
1 5 10 15
<210>4
<211>16
<212>PRT
<213>人工序列
<220>
<223>抗体轻链互补决定区
<400>4
Arg Ser Ser Gln Ser Ile Val His Ser Asn Val Asn Thr Tyr Leu Glu
1 5 10 15
<210>5
<211>7
<212>PRT
<213>人工序列
<220>
<223>抗体轻链互补决定区
<400>5
Lys Val Ser Asn Arg Phe Ser
1 5
<210>6
<211>9
<212>PRT
<213>人工序列
<220>
<223>抗体轻链互补决定区
<400>6
Phe Gln Gly Ser His Val Pro Pro Thr
1 5
<210>7
<211>124
<212>PRT
<213>人工序列
<220>
<223>抗体重链
<400>7
Gln Val Gln Leu Gln Gln Ser Gly Ala Glu Leu Val Lys Pro Gly Ala
1 5 10 15
Ser Val Lys Leu Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser Tyr
20 25 30
Trp Met His Trp Val Lys Gln Arg Pro Gly Gln Gly Leu Glu Trp Ile
35 40 45
Gly Glu Ile Asn Pro Ser Asn Gly Arg Thr Asn Tyr Asn Glu Lys Phe
50 55 60
Lys Arg Lys Ala Thr Leu Thr Val Asp Lys Ser Ser Ser Thr Ala Tyr
65 70 75 80
Met Gln Leu Ser Ser Leu Thr Ser Glu Asp Ser Ala Val Tyr Tyr Phe
85 90 95
Ala Arg Gly Arg Pro Asp Tyr Tyr Gly Ser Ser Lys Trp Tyr Phe Asp
100 105 110
Val Trp Gly Ala Gly Thr Thr Val Thr Val Ser Ser
115 120
<210>8
<211>113
<212>PRT
<213>人工序列
<223>抗体轻链
<400>8
Asp Val Leu Met Thr Gln Thr Pro Leu Ser Leu Pro Val Ser Leu Gly
1 5 10 15
Asp Gln Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Ile Val His Ser
20 25 30
Asn Val Asn Thr Tyr Leu Glu Trp Tyr Leu Gln Lys Pro Gly Gln Ser
35 40 45
Pro Lys Leu Leu Ile Tyr Lys Val Ser Asn Arg Phe Ser Gly Val Pro
50 55 60
Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Arg Ile
65 70 75 80
Ser Arg Val Glu Ala Glu Asp Leu Gly Ile Tyr Tyr Cys Phe Gln Gly
85 90 95
Ser His Val Pro Pro Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys
100 105 110
Arg
<210>9
<211>113
<212>PRT
<213>人工序列
<220>
<223>人源化轻链可变区
<400>9
Asp Val Val Met Thr Gln Thr Pro Leu Ser Leu Pro Val Ser Leu Gly
1 5 10 15
Asp Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Ile Val His Ser
20 25 30
Asn Val Asn Thr Tyr Leu Glu Trp Tyr Leu Gln Lys Pro Gly Gln Ser
35 40 45
Pro Arg Leu Leu Ile Tyr Lys Val Ser Asn Arg Phe Ser Gly Val Pro
50 55 60
Asp Arg Phe Ser Gly Ser Gly Ala Gly Thr Asp Phe Thr Leu Arg Ile
65 70 75 80
Ser Arg Val Glu Ala Glu Asp Leu Gly Ile Tyr Tyr Cys Phe Gln Gly
85 90 95
Ser His Val Pro Pro Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys
100 105 110
Arg
<210>10
<211>113
<212>PRT
<213>人工序列
<220>
<223>人源化轻链可变区
<400>10
Asp Val Leu Met Thr Gln Thr Pro Leu Ser Leu Pro Val Ser Leu Gly
1 5 10 15
Asp Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Ile Val His Ser
20 25 30
Asn Val Asn Thr Tyr Leu Glu Trp Tyr Leu Gln Lys Pro Gly Gln Ser
35 40 45
Pro Lys Leu Leu Ile Tyr Lys Val Ser Asn Arg Phe Ser Gly Val Pro
50 55 60
Asp Arg Phe Ser Gly Ser Gly Ala Gly Thr Asp Phe Thr Leu Arg Ile
65 70 75 80
Ser Arg Val Glu Ala Glu Asp Leu Gly Ile Tyr Tyr Cys Phe Gln Gly
85 90 95
Ser His Val Pro Pro Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys
100 105 110
Arg
<210>11
<211>113
<212>PRT
<213>人工序列
<220>
<223>人源化轻链可变区
<400>11
Asp Val Leu Met Thr Gln Thr Pro Leu Ser Leu Pro Val Ser Leu Gly
1 5 10 15
Asp Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Ile Val His Ser
20 25 30
Asn Val Asn Thr Tyr Leu Glu Trp Tyr Leu Gln Lys Pro Gly Gln Ser
35 40 45
Pro Arg Leu Leu Ile Tyr Lys Val Ser Asn Arg Phe Ser Gly Val Pro
50 55 60
Asp Arg Phe Ser Gly Ser Gly Ala Gly Thr Asp Phe Thr Leu Arg Ile
65 70 75 80
Ser Arg Val Glu Ala Glu Asp Leu Gly Ile Tyr Tyr Cys Phe Gln Gly
85 90 95
Ser His Val Pro Pro Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys
100 105 110
Arg
<210>12
<211>113
<212>PRT
<213>人工序列
<220>
<223>人源化轻链可变区
<400>12
Asp Val Val Met Thr Gln Thr Pro Leu Ser Leu Pro Val Ser Leu Gly
1 5 10 15
Asp Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Ile Val His Ser
20 25 30
Asn Val Asn Thr Tyr Leu Glu Trp Tyr Leu Gln Lys Pro Gly Gln Ser
35 40 45
Pro Lys Leu Leu Ile Tyr Lys Val Ser Asn Arg Phe Ser Gly Val Pro
50 55 60
Asp Arg Phe Ser Gly Ser Gly Ala Gly Thr Asp Phe Thr Leu Arg Ile
65 70 75 80
Ser Arg Val Glu Ala Glu Asp Leu Gly Ile Tyr Tyr Cys Phe Gln Gly
85 90 95
Ser His Val Pro Pro Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys
100 105 110
Arg
<210>13
<211>124
<212>PRT
<213>人工序列
<220>
<223>人源化重链可变区
<400>13
Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Val Lys Pro Gly Ala
1 5 10 15
Ser Val Lys Leu Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser Tyr
20 25 30
Trp Met His Trp Val Lys Gln Arg Pro Gly Gln Gly Leu Glu Trp Ile
35 40 45
Gly Glu Ile Asn Pro Ser Asn Gly Arg Thr Asn Tyr Asn Gln Lys Phe
50 55 60
Gln Gly Lys Ala Thr Leu Thr Val Asp Lys Ser Ser Ser Thr Ala Tyr
65 70 75 80
Met Gln Leu Ser Ser Leu Thr Ser Glu Asp Ser Ala Val Tyr Tyr Phe
85 90 95
Ala Arg Gly Arg Pro Asp Tyr Tyr Gly Ser Ser Lys Trp Tyr Phe Asp
100 105 110
Val Trp Gly Gln Gly Thr Thr Val Thr Val Ser Ser
115 120
<210>14
<21L>46
<212>DNA
<213>人工序列
<220>
<223>简并3’轻链PCR引物-HindKL
<400>14
tatagagctc aagcttggat ggtgggaaga tggatacagt tggtgc 46
<210>15
<211>36
<212>DNA
<213>人工序列
<220>
<223>简并3’重链PCR引物-Bgl2IgG1
<400>15
ggaagatcta tagacagatg ggggtgtcgt tttggc 36
<210>16
<211>30
<212>DNA
<213>人工序列
<220>
<223>多聚C 5’PCR引物-EcoPolyC
<400>16
tatatctaga attccccccc cccccccccc 30
<210>17
<211>32
<212>DNA
<213>人工序列
<220>
<223>简并5’轻链PCR引物-Sac1MK
<400>17
gggagctcga yattgtgmts acmcarwctm ca 32
<210>18
<211>32
<212>DNA
<213>人工序列
<220>
<223>简并5’重链PCR引物-EcoR1MH1
<220>
<221>misc feature
<222>(18)..(18)
<223>″n″可以是任何核酸
<400>18
cttccggaat tcsargtnma gctgsagsag tc 32
<210>19
<211>35
<212>DNA
<213>人工序列
<220>
<223>简并5’重链PCR引物-EcoR1MH2
<220>
<221>misc feature
<222>(18)..(18)
<223>″n″可以是任何核苷酸
<400>19
cttccggaat tcsargtnma gctgsagsag tcwgg 35
<210>20
<211>10
<212>PRT
<213>小家鼠(Mus musculus)
<400>20
Asp Val Leu Met Thr Gln Thr Pro Leu Ser
1 5 10
<210>21
<211>10
<212>PRT
<213>小家鼠(Mus musculus)
<400>21
Gly Arg Pro Asp Tyr Tyr Gly Ser Ser Lys
1 5 10
<210>22
<211>24
<212>PRT
<213>小家鼠(Mus musculus)
<400>22
Ser Ser Ser Thr Ala Tyr Met Gln Leu Ser Ser Leu Thr Ser Glu Asp
1 5 10 15
Ser Ala Val Tyr Tyr Phe Ala Arg
20
<210>23
<211>57
<212>DNA
<213>人工序列
<220>
<223>PCR引物
<400>23
caggtgtaca ctcccaggtc caactggtgc agtctggggc tgaagtggtg aagcctg 57
<210>24
<211>29
<212>DNA
<213>人工序列
<220>
<223>PCR引物
<400>24
caatcagaag ttccagggga aggccacac 29
<210>25
<211>34
<212>DNA
<213>人工序列
<220>
<223>PCR引物
<400>25
ccttcccctg gaacttctga ttgtagttag tacg 34
<210>26
<211>37
<212>DNA
<213>人工序列
<220>
<223>PCR引物
<400>26
caggtgtaca ctccgatgtt gtgatgaccc aaactcc 37
<210>27
<211>37
<212>DNA
<213>人工序列
<220>
<223>PCR引物
<400>27
caggtgtaca ctccgatgtt ttgatgaccc aaactcc 37
<210>28
<211>39
<212>DNA
<213>人工序列
<220>
<223>PCR引物
<400>28
gactagatct gcaagagatg gaggctggat ctccaagac 39
<210>29
<211>35
<212>DNA
<213>人工序列
<220>
<223>PCR引物
<400>29
ttgcagatct agtcagagca tagtacatag taatg 35
<210>30
<211>48
<212>DNA
<213>人工序列
<220>
<223>PCR引物
<400>30
gaatggtacc tgcagaaacc aggccagtct ccaaggctcc tgatctac 48
<210>31
<211>28
<212>DNA
<213>人工序列
<220>
<223>PCR引物
<400>31
gtggcagtgg agcagggaca gatttcac 28
<210>32
<211>28
<212>DNA
<213>人工序列
<220>
<223>PCR引物
<400>32
gaaatctgtc cctgctccac tgccactg 28
<210>33
<211>19
<212>PRT
<213>人类(Homo sapiens)
<400>33
Asp Leu Thr Leu Leu Gln Pro Gly Gln Lys Gly Asp Ser Arg Glu Lys
1 5 10 15
Lys Arg Ala
<210>34
<211>18
<212>PRT
<213>人类(Homo sapiens)
<400>34
Asp Val Thr Leu Leu Pro Pro Gly Gln Arg Gly Asp Ala Arg Glu Lys
1 5 10 15
Lys Arg
<210>35
<211>19
<212>PRT
<213>人类(Homo sapiens)
<400>35
Asp Gln Ser Leu Ile Pro Pro Gly Gln Lys Gly Asp Ser Arg Asp Lys
1 5 10 15
Lys Arg Ala
<210>36
<211>18
<212>PRT
<213>人类(Homo sapiens)
<400>36
Asp Met Ser Ser Val Arg Pro Gly Gln Lys Gly Ser Ser Ser Asp Lys
1 5 10 15
Lys Arg
<210>37
<211>18
<212>PRT
<213>人类(Homo sapiens)
<400>37
Glu Val Ser Gly Pro Arg Pro Gly Gln Arg Gly Asp Ser Arg Glu Lys
1 5 10 15
Lys Arg
<210>38
<211>18
<212>PRT
<213>人类(Homo sapiens)
<400>38
Glu Val Ser Gly Pro Arg Pro Gly Gln Arg Gly Asp Ser Arg Glu Lys
1 5 10 15
Lys Arg
<210>39
<211>25
<212>PRT
<213>人类(Homo sapiens)
<400>39
Gln Gln Gln Ala Leu Lys Pro Gly Lys Lys Thr Pro Gly Gln Glu Lys
1 5 10 15
Lys Arg Lys Ser Ser Ser Glu Ala Ser
20 25
<210>40
<211>25
<212>PRT
<213>人类(Homo sapiens)
<400>40
Gln Gln Val Ala Val Lys Pro Gly Lys Lys Thr Pro Gly Gln Gln Lys
1 5 10 15
Gln Gly Lys Ser Ser Ser Glu Gln Ser
20 25
<210>41
<211>25
<212>PRT
<213>人类(Homo sapiens)
<400>41
Gln Gln Gln Pro Leu Lys Pro Gly Lys Lys Thr Pro Gly Lys Asp Asp
1 5 10 15
Lys Gly Thr Ser Asn Asn Glu Gln Ser
20 25
<210>42
<211>25
<212>PRT
<213>人类(Homo sapiens)
<400>42
Gln Gln Val Ala Val Lys Pro Gly Lys Lys Thr Pro Gly Gln Gln Lys
1 5 10 15
Lys Gly Lys Ser Ser Ser Glu Gln Ser
20 25
<210>43
<211>24
<212>PRT
<213>人类(Homo sapiens)
<400>43
Gln Val Ala Val Lys Pro Gly Lys Lys Thr Pro Gly Gln Gln Lys Gln
1 5 10 15
Gly Lys Ser Ser Ser Glu Gln Ser
20
<210>44
<211>24
<212>PRT
<213>人类(Homo sapiens)
<400>44
Gln Val Ala Val Lys Pro Gly Lys Lys Thr Pro Gly Gln Gln Lys Gln
1 5 10 15
Gly Glu Ser Ser Ser Glu Gln Ser
20
<210>45
<211>40
<212>DNA
<213>人工序列
<220>
<223>PCR引物
<400>45
ttttgagctc ttatttacca ggagagtggg agaggctctt 40
<210>46
<211>37
<212>DNA
<213>人工序列
<220>
<223>PCR引物
<400>46
ttttaagctt gccaaaacga cacccccatc tgtctat 37
<210>47
<211>32
<212>DNA
<213>人工序列
<220>
<223>PCR引物
<400>47
ttttggatcc taacactcat tcctgttgaa gc 32
<210>48
<211>31
<212>DNA
<213>人工序列
<220>
<223>PCR引物
<400>48
ttttgaattc gggctgatgc tgcaccaact g 31
<210>49
<211>396
<212>DNA
<213>小家鼠(Mus musculus)
<220>
<221>CDS
<222>(1)..(396)
<400>49
atg aag ttg cct gtt agg ctg ttg gtg ctg atg ttc tgg att cct gct 48
Met Lys Leu Pro Val Arg Leu Leu Val Leu Met Phe Trp Ile Pro Ala
l 5 10 15
tcc agt agt gat gtt ttg atg acc caa act cca ctc tcc ctg cct gtc 96
Ser Ser Ser Asp Val Leu Met Thr Gln Thr Pro Leu Ser Leu Pro Val
20 25 30
agt ctt gga gat caa gcc tcc atc tct tgc aga tct agt cag agc att 144
Ser Leu Gly Asp Gln Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Ile
35 40 45
gta cat agt aat gta aac acc tat tta gaa tgg tac ctg cag aaa cca 192
Val His Ser Ash Val Asn Thr Tyr Leu Glu Trp Tyr Leu Gln Lys Pro
50 55 60
ggc cag tct cca aag ctc ctg atc tac aaa gtt tcc aac cga ttt tct 240
Gly Gln Ser Pro Lys Leu Leu Ile Tyr Lys Val Ser Asn Arg Phe Ser
65 70 75 80
ggg gtc cca gac agg ttc agt ggc agt gga tca ggg aca gat ttc aca 288
Gly Val Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr
85 90 95
ctc agg atc agc aga gtg gag gct gag gat ctg gga att tat tac tgc 336
Leu Arg Ile Ser Arg Val Glu Ala Glu Asp Leu Gly Ile Tyr Tyr Cys
100 105 110
ttt caa ggt tca cat gtt cct ccg acg ttc ggt gga ggc acc aag ctg 384
Phe Gln Gly Ser His Val Pro Pro Thr Phe Gly Gly Gly Thr Lys Leu
115 120 125
gaa atc aaa cgg 396
Glu Ile Lys Arg
130
<210>50
<211>132
<212>PRT
<213>小家鼠(Mus musculus)
<400>50
Met Lys Leu Pro Val Arg Leu Leu Val Leu Met Phe Trp Ile Pro Ala
1 5 10 15
Ser Ser Ser Asp Val Leu Met Thr Gln Thr Pro Leu Ser Leu Pro Val
20 25 30
Ser Leu Gly Asp Gln Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Ile
35 40 45
Val His Ser Asn Val Asn Thr Tyr Leu Glu Trp Tyr Leu Gln Lys Pro
50 55 60
Gly Gln Ser Pro Lys Leu Leu Ile Tyr Lys Val Ser Asn Arg Phe Ser
65 70 75 80
Gly Val Pro Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr
85 90 95
Leu Arg Ile Ser Arg Val Glu Ala Glu Asp Leu Gly Ile Tyr Tyr Cys
100 105 110
Phe Gln Gly Ser His Val Pro Pro Thr Phe Gly Gly Gly Thr Lys Leu
115 120 125
Glu Ile Lys Arg
130
<210>51
<211>429
<212>DNA
<213>小家鼠(Mus musculus)
<220>
<221>CDS
<222>(1)..(429)
<400>51
atg gga tgg agc tat atc atc ctc ttt ttg gta gca aca gct aca gaa 48
Met Gly Trp Ser Tyr Ile Ile Leu Phe Leu Val Ala Thr Ala Thr Glu
1 5 10 15
gtc cac tcc cag gtc caa ctg cag cag tct ggg gct gaa ctg gtg aag 96
Val His Ser Gln Val Gln Leu Gln Gln Ser Gly Ala Glu Leu Val Lys
20 25 30
cct ggg gct tca gtg aag ctg tcc tgt aag gct tct ggc tac acc ttc 144
Pro Gly Ala Ser Val Lys Leu Ser Cys Lys Ala Ser Gly Tyr Thr Phe
35 40 45
acc agc tac tgg atg cac tgg gtg aag cag agg cct gga caa ggc ctt 192
Thr Ser Tyr Trp Met His Trp Val Lys Gln Arg Pro Gly Gln Gly Leu
50 55 60
gag tgg att gga gag att aat cct agc aac ggt cgt act aac tac aat 240
Glu Trp Ile Gly Glu Ile Asn Pro Ser Asn Gly Arg Thr Asn Tyr Asn
65 70 75 80
gag aag ttc aag agg aag gcc aca ctg act gta gac aaa tcc tcc agc 288
Glu Lys Phe Lys Arg Lys Ala Thr Leu Thr Val Asp Lys Ser Ser Ser
85 90 95
aca gcc tac atg caa ctc agc agc ctg aca tct gag gac tct gcg gtc 336
Thr Ala Tyr Met Gln Leu Ser Ser Leu Thr Ser Glu Asp Ser Ala Val
100 105 110
tat tac ttt gca aga gga aga cca gat tac tac ggt agt agc aag tgg 384
Tyr Tyr Phe Ala Arg Gly Arg Pro Asp Tyr Tyr Gly Ser Ser Lys Trp
115 120 125
tac ttc gat gtc tgg ggc gca ggg acc acg gtc acc gtc tcc tca 429
Tyr Phe Asp Val Trp Gly Ala Gly Thr Thr Val Thr Val Ser Ser
130 135 140
<210>52
<211>143
<212>PRT
<213>小家鼠(Mus musculus)<400>52
Met Gly Trp Ser Tyr Ile Ile Leu Phe Leu Val Ala Thr Ala Thr Glu
1 5 10 15
Val His Ser Gln Val Gln Leu Gln Gln Ser Gly Ala Glu Leu Val Lys
20 25 30
Pro Gly Ala Ser Val Lys Leu Ser Cys Lys Ala Ser Gly Tyr Thr Phe
35 40 45
Thr Ser Tyr Trp Met His Trp Val Lys Gln Arg Pro Gly Gln Gly Leu
50 55 60
Glu Trp Ile Gly Glu Ile Asn Pro Ser Asn Gly Arg Thr Asn Tyr Asn
65 70 75 80
Glu Lys Phe Lys Arg Lys Ala Thr Leu Thr Val Asp Lys Ser Ser Ser
85 90 95
Thr Ala Tyr Met Gln Leu Ser Ser Leu Thr Ser Glu Asp Ser Ala Val
100 105 110
Tyr Tyr Phe Ala Arg Gly Arg Pro Asp Tyr Tyr Gly Ser Ser Lys Trp
115 120 125
Tyr Phe Asp Val Trp Gly Ala Gly Thr Thr Val Thr Val Ser Ser
130 135 140
<210>53
<211>10
<212>PRT
<213>小家鼠(Mus musculus)
<400>53
Gly Tyr Thr Phe Thr Ser Tyr Trp Met His
1 5 10
<210>54
<211>10
<212>PRT
<213>小家鼠(Mus musculus)
<400>54
Glu Ile Asn Pro Ser Asn Gly Arg Thr Asn
1 5 10
<210>55
<211>15
<212>PRT
<213>小家鼠(Mus musculus)
<400>55
Gly Arg Pro Asp Tyr Tyr Gly Ser Ser Lys Trp Tyr Phe Asp Val
1 5 10 15
<210>56
<211>100
<212>PRT
<213>小家鼠(Mus musculus)
<400>56
Asp Val Leu Met Thr Gln Thr Pro Leu Ser Leu Pro Val Ser Leu Gly
1 5 10 15
Asp Gln Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Ile Val His Ser
20 25 30
Asn Gly Asn Thr Tyr Leu Glu Trp Tyr Leu Gln Lys Pro Gly Gln Ser
35 40 45
Pro Lys Leu Leu Ile Tyr Lys Val Ser Asn Arg Phe Ser Gly Val Pro
50 55 60
Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys Ile
65 70 75 80
Ser Arg Val Glu Ala Glu Asp Leu Gly Val Tyr Tyr Cys Phe Gln Gly
85 90 95
Ser His Val Pro
100
<210>57
<211>98
<212>PRT
<213>小家鼠(Mus musculus)
<400>57
Gln Val Gln Leu Gln Gln Pro Gly Ala Glu Leu Val Lys Pro Gly Ala
1 5 10 15
Ser Val Lys Leu Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser Tyr
20 25 30
Trp Met His Trp Val Lys Gln Arg Pro Gly Gln Gly Leu Glu Trp Ile
35 40 45
Gly Glu Ile Asn Pro Ser Asn Gly Arg Thr Asn Tyr Ash Glu Lys Phe
50 55 60
Lys Ser Lys Ala Thr Leu Thr Val Asp Lys Ser Ser Ser Thr Ala Tyr
65 70 75 80
Met Gln Leu Ser Ser Pro Thr Ser Glu Asp Ser Ala Val Tyr Tyr Cys
85 90 95
Ala Arg
<210>58
<211>113
<212>PRT
<213>人工序列
<220>
<223>合成抗体结构
<400>58
Asp Val Leu Met Thr Gln Thr Pro Leu Ser Leu Pro Val Ser Leu Gly
1 5 10 15
Asp Gln Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Ile Val His Ser
20 25 30
Asn Val Asn Thr Tyr Leu Glu Trp Tyr Leu Gln Lys Pro Gly Gln Ser
35 40 45
Pro Lys Leu Leu Ile Tyr Lys Val Ser Asn Arg Phe Ser Gly Val Pro
50 55 60
Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Arg Ile
65 70 75 80
Ser Arg Val Glu Ala Glu Asp Leu Gly Ile Tyr Tyr Cys Phe Gln Gly
85 90 95
Ser His Val Pro Pro Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys
100 105 110
Arg
<210>59
<211>113
<212>PRT
<213>人工序列
<220>
<223>合成抗体结构
<400>59
Asp Val Leu Met Thr Gln Thr Pro Leu Ser Leu Pro Val Ser Leu Gly
1 5 10 15
Asp Gln Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Ile Val His Ser
20 25 30
Asn Gly Asn Thr Tyr Leu Glu Trp Tyr Leu Gln Lys Pro Gly Gln Ser
35 40 45
Pro Lys Leu Leu Ile Tyr Ser Ile Ser Ser Arg Phe Ser Gly Val Pro
50 55 60
Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys Ile
65 70 75 80
Ser Arg Val Gln Ala Glu Asp Leu Gly Val Tyr Tyr Cys Phe Gln Gly
85 90 95
Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys
100 105 110
Arg
<210>60
<211>113
<212>PRT
<213>人工序列
<220>
<223>合成抗体结构
<400>60
Asp Val Leu Met Thr Gln Thr Pro Leu Ser Leu Pro Val Ser Leu Gly
1 5 10 15
Asp Gln Ala Ser Ile Ser Cys Arg Ser Ser Gln Thr Ile Val Hi s Ser
20 25 30
Asn Gly Asn Thr Tyr Leu Glu Trp Tyr Leu Gln Lys Pro Gly Gln Ser
35 40 45
Pro Lys Leu Leu Ile Tyr Lys Val Thr Asn Arg Phe Ser Gly Val Pro
50 55 60
Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys Ile
65 70 75 80
Ser Arg Val Glu Ala Glu Asp Leu Gly Val Tyr Tyr Cys Phe Gln Gly
85 90 95
Thr His Ala Pro Tyr Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys
100 105 110
Arg
<210>61
<211>113
<212>PRT
<213>人工序列
<220>
<223>合成抗体结构
<400>61
Asp Ile Glu Leu Thr Gln Thr Pro Leu Ser Leu Pro Val Ser Leu Gly
1 5 10 15
Asp Gln Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Ile Val His Ser
20 25 30
Asn Gly Asn Thr Tyr Leu Glu Trp Tyr Leu Gln Lys Pro Gly Gln Ser
35 40 45
Pro Lys Leu Leu Ile Tyr Lys Val Ser Asn Arg Phe Ser Gly Val Pro
50 55 60
Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys Ile
65 70 75 80
Ser Arg Val Glu Ala Glu Asp Leu Gly Val Tyr Tyr Cys Phe Gln Gly
85 90 95
Ser His Val Pro Tyr Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys
100 105 110
Arg
<210>62
<211>113
<212>PRT
<213>人工序列
<220>
<223>合成抗体结构
<400>62
Asp Val Leu Met Thr Gln Thr Pro Leu Ser Leu Pro Val Ser Leu Gly
1 5 10 15
Asp Gln Ala Ser Ile Ser Cys Arg Phe Ser Gln Ser Ile Val His Ser
20 25 30
Asn Gly Asn Thr Tyr Leu Glu Trp Tyr Leu Gln Lys Ser Gly Gln Ser
35 40 45
Pro Lys Leu Leu Ile Tyr Lys Val Ser Asn Arg Phe Ser Gly Val Pro
50 55 60
Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys Ile
65 70 75 80
Ser Arg Val Glu Ala Glu Asp Leu Gly Val Tyr Tyr Cys Phe Gln Gly
85 90 95
Ser His Val Pro Arg Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys
100 105 110
Arg
<210>63
<211>113
<212>PRT
<213>人工序列
<220>
<223>合成抗体结构
<400>63
Glu Leu Val Met Thr Gln Thr Pro Leu Ser Leu Pro Val Ser Leu Gly
1 5 10 15
Asp Gln Ala Ser Ile Ser Cys Arg Ser Ser Gln Thr Ile Val His Ser
20 25 30
Ash Gly Asp Thr Tyr Leu Asp Trp Phe Leu Gln Lys Pro Gly Gln Ser
35 40 45
Pro Lys Leu Leu Ile Tyr Lys Val Ser Asn Arg Phe Ser Gly Val Pro
50 55 60
Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys Ile
65 70 75 80
Ser Arg Val Glu Ala Glu Asp Leu Gly Val Tyr Tyr Cys Phe Gln Gly
85 90 95
Ser His Val Pro Pro Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys
100 105 110
Arg
<210>64
<211>113
<212>PRT
<213>人工序列
<220>
<223>合成抗体结构
<400>64
Asp Val Leu Met Thr Gln Thr Pro Leu Ser Leu Pro Val Ser Leu Gly
1 5 10 15
Asp Gln Ala Ser Ile Ser Cys Arg Ser Asn Gln Thr Ile Leu Leu Ser
20 25 30
Asp Gly Asp Thr Tyr Leu Glu Trp Tyr Leu Gln Lys Pro Gly Gln Ser
35 40 45
Pro Lys Leu Leu Ile Tyr Lys Val Ser Asn Arg Phe Ser Gly Val Pro
50 55 60
Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys Ile
65 70 75 80
Ser Arg Val Glu Ala Glu Asp Leu Gly Val Tyr Tyr Cys Phe Gln Gly
85 90 95
Ser His Val Pro Pro Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys
100 105 110
Arg
<210>65
<211>113
<212>PRT
<213>人工序列
<220>
<223>合成抗体结构
<400>65
Asp Val Leu Met Thr Gln Thr Pro Leu Ser Leu Pro Val Ser Leu Gly
1 5 10 15
Asp Gln Ala Ser Ile Ser Cys Lys Ser Ser Gln Ser Ile Val His Ser
20 25 30
Ser Gly Asn Thr Tyr Phe Glu Trp Tyr Leu Gln Lys Pro Gly Gln Ser
35 40 45
Pro Lys Leu Leu Ile Tyr Lys Val Ser Asn Arg Phe Ser Gly Val Pro
50 55 60
Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys Ile
65 70 75 80
Ser Arg Val Glu Ala Glu Asp Leu Gly Val Tyr Tyr Cys Phe Gln Gly
85 90 95
Ser His Ile Pro Phe Thr Phe Gly Ser Gly Thr Lys Leu Glu Ile Lys
100 105 110
Arg
<210>66
<211>113
<212>PRT
<213>人工序列
<220>
<223>合成抗体结构
<400>66
Asp Val Leu Met Thr Gln Ile Pro Val Ser Leu Pro Val Ser Leu Gly
1 5 10 15
Asp Gln Ala Ser Ile Ser Cys Arg Ser Ser Gln Ile Ile Val His Asn
20 25 30
Asn Gly Asn Thr Tyr Leu Glu Trp Tyr Leu Gln Lys Pro Gly Gln Ser
35 40 45
Pro Gln Leu Leu Ile Tyr Lys Val Ser Asn Arg Phe Ser Gly Val Pro
50 55 60
Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys Ile
65 70 75 80
Ser Arg Val Glu Ala Glu Asp Leu Gly Val Tyr Tyr Cys Phe Gln Gly
85 90 95
Ser His Val Pro Phe Thr Phe Gly Ser Gly Thr Lys Leu Glu Ile Lys
100 105 110
Arg
<210>67
<211>113
<212>PRT
<213>人工序列
<220>
<223>合成抗体结构
<400>67
Asp Val Leu Met Thr Gln Thr Pro Val Ser Leu Ser Val Ser Leu Gly
1 5 10 15
Asp Gln Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Ile Val His Ser
20 25 30
Thr Gly Asn Thr Tyr Leu Glu Trp Tyr Leu Gln Lys Pro Gly Gln Ser
35 40 45
Pro Lys Leu Leu Ile Tyr Lys Ile Ser Asn Arg Phe Ser Gly Val Pro
50 55 60
Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys Ile
65 70 75 80
Ser Arg Val Glu Ala Glu Asp Leu Gly Val Tyr Tyr Cys Phe Gln Ala
85 90 95Ser His Ala Pro Arg Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys
100 105 110
Arg
<210>68
<211>113
<212>PRT
<213>人工序列
<220>
<223>合成抗体结构
<400>68
Asp Val Leu Met Thr Gln Ile Pro Val Ser Leu Pro Val Ser Leu Gly
1 5 10 15
Asp Gln Ala Ser Ile Ser Cys Arg Ser Ser Gln Ile Ile Val His Asn
20 25 30
Asn Gly Asn Thr Tyr Leu Glu Trp Tyr Leu Gln Lys Pro Gly Gln Ser
35 40 45
Pro Gln Leu Leu Ile Tyr Lys Val Ser Asn Arg Phe Ser Gly Val Pro
50 55 60
Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys Ile
65 70 75 80
Ser Arg Val Glu Ala Glu Asp Leu Gly Val Tyr Tyr Cys Phe Gln Gly
85 90 95
Ser His Val Pro Phe Thr Phe Gly Ser Gly Thr Lys Leu Glu Ile Lys
100 105 110
Arg
<210>69
<211>113
<212>PRT
<213>人工序列
<220>
<223>合成抗体结构
<220>
<221>MISC_FEATURE
<222>(28)..(28)
<223>″X″可以是任何氨基酸
<220>
<221>MISC_FEATURE
<222>(101)..(101)
<223>″X″可以是任何氨基酸
<400>69
Asp Val Leu Met Thr Gln Thr Pro Leu Ser Leu Pro Val Ser Leu Gly
1 5 10 15
Asp Gln Ala Ser Ile Ser Cys Arg Ser Ser Gln Xaa Ile Val His Ser
20 25 30
Asn Gly Asn Thr Tyr Leu Glu Trp Tyr Leu Gln Lys Pro Gly Gln Ser
35 40 45
Pro Lys Leu Leu Ile Tyr Lys Val Ser Asn Arg Phe Ser Gly Val Pro
50 55 60
Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Lys Ile
65 70 75 80
Ser Arg Val Glu Ala Glu Asp Leu Gly Val Tyr Tyr Cys Phe Gln Gly
85 90 95
Ser His Val Pro Xaa Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys
100 105 110
Arg
<210>70
<211>124
<212>PRT
<213>人工序列
<220>
<223>合成抗体结构
<400>70
Gln Val Gln Leu Gln Gln Ser Gly Ala Glu Leu Val Lys Pro Gly Ala
1 5 10 15
Ser Val Lys Leu Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser Tyr
20 25 30
Trp Met His Trp Val Lys Gln Arg Pro Gly Gln Gly Leu Glu Trp Ile
35 40 45
Gly Glu Ile Asn Pro Ser Asn Gly Arg Thr Asn Tyr Asn Glu Lys Phe
50 55 60
Lys Arg Lys Ala Thr Leu Thr Val Asp Lys Ser Ser Ser Thr Ala Tyr
65 70 75 80
Met Gln Leu Ser Ser Leu Thr Ser Glu Asp Ser Ala Val Tyr Tyr Phe
85 90 95
Ala Arg Gly Arg Pro Asp Tyr Tyr Gly Ser Ser Lys Trp Tyr Phe Asp
100 105 110
Val Trp Gly Ala Gly Thr Thr Val Thr Val Ser Ser
115 120
<210>71
<211>120
<212>PRT
<213>人工序列
<220>
<223>合成抗体结构
<400>71
Gln Val Gln Leu Gln Gln Ser Gly Ala Glu Leu Val Lys Pro Gly Ala
1 5 10 15
Ser Val Lys Leu Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser Tyr
20 25 30
Trp Met His Trp Val Lys Gln Arg Pro Gly Arg Gly Leu Glu Trp Ile
35 40 45
Gly Arg Ile Asp Pro Asn Ser Gly Gly Thr Lys Tyr Asn Glu Lys Phe
50 55 60
Lys Ser Lys Ala Thr Leu Thr Val Asp Lys Pro Ser Ser Thr Ala Tyr
65 70 75 80
Met Gln Leu Ser Ser Leu Thr Ser Glu Asp Ser Ala Val Tyr Tyr Cys
85 90 95
Ala Arg Tyr Asp Tyr Tyr Gly Ser Ser Tyr Phe Asp Tyr Trp Gly Gln
100 105 110
Gly Thr Thr Val Thr Val Ser Ser
115 120
<210>72
<211>120
<212>PRT
<213>人工序列
<220>
<223>合成抗体结构
<400>72
Gln Val Gln Leu Gln Gln Pro Gly Ala Glu Leu Val Lys Pro Gly Ala
1 5 10 15
Ser Val Lys Leu Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser Tyr
20 25 30
Trp Met His Trp Val Lys Gln Arg Pro Gly Arg Gly Leu Glu Trp Ile
35 40 45
Gly Arg Ile Asp Pro Ash Ser Gly Gly Thr Lys Tyr Asn Glu Lys Phe
50 55 60
Lys Ser Lys Ala Thr Leu Thr Val Asp Lys Pro Ser Ser Thr Ala Tyr
65 70 75 80
Met Gln Leu Ser Ser Leu Thr Ser Glu Asp Ser Ala Val Tyr Tyr Cys
85 90 95
Ala Arg Tyr Asp Tyr Tyr Gly Ser Ser Tyr Phe Asp Tyr Trp Gly Gln
100 105 110
Gly Thr Thr Leu Thr Val Ser Ser
115 120
<210>73
<211>122
<212>PRT
<213>人工序列
<220>
<223>合成抗体结构
<400>73
Gln Val Gln Leu Gln Gln Pro Gly Ala Glu Leu Val Lys Pro Gly Ala
1 5 10 15
Ser Val Lys Leu Ser Cys Lys Ala Ser Gly Tyr ThrPhe Thr Ser Tyr
20 25 30
Trp Met His Trp Val Lys Gln Gly Pro Gly Gln Gly Leu Glu Trp Ile
35 40 45
Gly Glu Ile Asp Pro Ser Asp Ser Tyr Pro Asn Tyr Asn Glu Lys Phe
50 55 60
Lys Gly Lys Ala Thr Leu Thr Val Asp Lys Ser Ser Ser Thr Ala Tyr
65 70 75 80
Met Gln Leu Ser Ser Leu Thr Ser Glu Asp Ser Ala Val Tyr Tyr Cys
85 90 95
Ala Ser Leu Tyr Tyr Tyr Gly Thr Ser Tyr Gly Val Leu Asp Tyr Trp
100 105 110
Gly Gln Gly Thr Ser Val Thr Val Ser Ser
115 120
<210>74
<211>120
<212>PRT
<213>人工序列
<220>
<223>合成抗体结构
<400>74
Gln Val Gln Leu Gln Gln Pro Gly Ser Val Leu Val Arg Pro Gly Ala
1 5 10 15
Ser Val Lys Leu Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser Ser
20 25 30
Trp Ile His Trp Ala Lys Gln Arg Pro Gly Gln Gly Leu Glu Trp Ile
35 40 45
Gly Glu Ile His Pro Asn Ser Gly Asn Thr Asn Tyr Asn Glu Lys Phe
50 55 60
Lys Gly Lys Ala Thr Leu Thr Val Asp Thr Ser Ser Ser Thr Ala Tyr
65 70 75 80
Val Asp Leu Ser Ser Leu Thr Ser Glu Asp Ser Ala Val Tyr Tyr Cys
85 90 95
Ala Arg Trp Arg Tyr Gly Ser Pro Tyr Tyr Phe Asp Tyr Trp Gly Gln
100 105 110
Gly Thr Thr Leu Thr Val Ser Ser
115 120
<210>75
<211>118
<212>PRT
<213>人工序列
<220>
<223>合成抗体结构
<400>75
Gln Val Gln Phe Gln Gln Ser Gly Ala Glu Leu Val Lys Pro Gly Ala
1 5 10 15
Ser Val Lys Leu Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser Tyr
20 25 30
Leu Met His Trp Ile Lys Gln Arg Pro Gly Arg Gly Leu Glu Trp Ile
35 40 45
Gly Arg Ile Asp Pro Asn Asn Val Val Thr Lys Phe Asn Glu Lys Phe
50 55 60
Lys Ser Lys Ala Thr Leu Thr Val Asp Lys Pro Ser Ser Thr Ala Tyr
65 70 75 80
Met Glu Leu Ser Ser Leu Thr Ser Glu Asp Ser Ala Val Tyr Tyr Cys
85 90 95
Ala Arg Tyr Ala Tyr Cys Arg Pro Met Asp Tyr Trp Gly Gln Gly Thr
100 105 110
Thr Val Thr Val Ser Ser
115
<210>76
<211>117
<212>PRT
<213>人工序列
<220>
<223>合成抗体结构
<400>76
Gln Ile Gln Leu Gln Gln Ser Gly Pro Glu Leu Val Arg Pro Gly Ala
1 5 10 15
Ser Val Lys Ile Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Asp Tyr
20 25 30
Tyr Ile His Trp Val Lys Gln Arg Pro Gly Glu Gly Leu Glu Trp Ile
35 40 45
Gly Trp Ile Tyr Pro Gly Ser Gly Asn Thr Lys Tyr Asn Glu Lys Phe
50 55 60
Lys Gly Lys Ala Thr Leu Thr Val Asp Thr Ser Ser Ser Thr Ala Tyr
65 70 75 80
Met Gln Leu Ser Ser Leu Thr Ser Glu Asp Ser Ala Val Tyr Phe Cys
85 90 95
Ala Arg Gly Gly Lys Phe Ala Met Asp Tyr Trp Gly Gln Gly Thr Ser
100 105 110
Val Thr Val Ser Ser
115
<210>77
<211>120
<212>PRT
<213>人工序列
<220>
<223>合成抗体结构
<400>77
Gln Val Gln Leu Gln Gln Ser Gly Ala Glu Leu Met Lys Pro Gly Ala
1 5 10 15
Ser Val Lys Ile Ser Cys Lys Ala Thr Gly Tyr Thr Phe Ser Ser Phe
20 25 30
Trp Ile Glu Trp Val Lys Gln Arg Pro Gly His Gly Leu Glu Trp Ile
35 40 45
Gly Glu Ile Leu Pro Gly Ser Gly Gly Thr His Tyr Asn Glu Lys Phe
50 55 60
Lys Gly Lys Ala Thr Phe Thr Ala Asp Lys Ser Ser Asn Thr Ala Tyr
65 70 75 80
Met Gln Leu Ser Ser Leu Thr Ser Glu Asp Ser Ala Val Tyr Tyr Cys
85 90 95
Ala Arg Gly His Ser Tyr Tyr Phe Tyr Asp Gly Asp Tyr Trp Gly Gln
100 105 110
Gly Thr Ser Val Thr Val Ser Ser
115 120
<210>78
<211>120
<212>PRT
<213>人工序列
<220>
<223>合成抗体结构
<400>78
Gln Ile Gln Leu Gln Gln Ser Gly Pro Glu Leu Val Lys Pro Gly Ala
1 5 10 15
Ser Val Lys Ile Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Asp Tyr
20 25 30
Tyr Ile Asn Trp Met Lys Gln Lys Pro Gly Gln Gly Leu Glu Trp Ile
35 40 45
Gly Trp Ile Asp Pro Gly Ser Gly Asn Thr Lys Tyr Asn Glu Lys Phe
50 55 60
Lys Gly Lys Ala Thr Leu Thr Val Asp Thr Ser Ser Ser Thr Ala Tyr
65 70 75 80
Met Gln Leu Ser Ser Leu Thr Ser Glu Asp Thr Ala Val Tyr Phe Cys
85 90 95
Ala Arg Glu Lys Thr Thr Tyr Tyr Tyr Ala Met Asp Tyr Trp Gly Gln
100 105 110
Gly Thr Ser Val Thr Val Ser Ala
115 120
<210>79
<211>120
<212>PRT
<213>人工序列
<220>
<223>合成抗体结构
<400>79
Gln Val Gln Leu Leu Glu Ser Gly Ala Glu Leu Met Lys Pro Gly Ala
1 5 10 15
Ser Val Lys Ile Ser Cys Lys Ala Thr Gly Tyr Thr Phe Ser Ser Phe
20 25 30
Trp Ile Glu Trp Val Lys Gln Arg Pro Gly His Gly Leu Glu Trp Ile
35 40 45
Gly Glu Ile Leu Pro Gly Ser Gly Gly Thr His Tyr Asn Glu Lys Phe
50 55 60
Lys Gly Lys Ala Thr Phe Thr Ala Asp Lys Ser Ser Asn Thr Ala Tyr
65 70 75 80
MetGln Leu Ser Ser Leu Thr Ser Glu Asp Ser Ala Val Tyr Tyr Cys
85 90 95
Ala Arg Gly His Ser Tyr Tyr Phe Tyr Asp Gly Asp Tyr Trp Gly Gln
100 105 110
Gly Thr Ser Val Thr Val Ser Ser
115 120
<210>80
<211>115
<212>PRT
<213>人工序列
<220>
<223>合成抗体结构
<400>80
Val Gln Leu Gln Gln Ser Gly Ala Glu Leu Met Lys Pro Gly Ala Ser
1 5 10 15
Val Lys Ile Ser Cys Lys Ala Ser Gly Tyr Thr Phe Ser Asp Tyr Trp
20 25 30
Ile Glu Trp Val Lys Gln Arg Pro Gly His Gly Leu Glu Trp Ile Gly
35 40 45
Glu Ile Leu Pro Gly Ser Gly Ser Thr Asn Tyr His Glu Arg Phe Lys
50 55 60
Gly Lys Ala Thr Phe Thr Ala Asp Thr Ser Ser Ser Thr Ala Tyr Met
65 70 75 80
Gln Leu Asn Ser Leu Thr Ser Glu Asp Ser Gly Val Tyr Tyr Cys Leu
85 90 95
His Gly Asn Tyr Asp Phe Asp Gly Trp Gly Gln Gly Thr Thr Leu Thr
100 105 110
Val Ser Ser
115
<210>81
<211>121
<212>PRT
<213>人工序列
<220>
<223>合成抗体结构
<220>
<221>MISC_FEATURE
<222>(20)..(20)
<223>″X″可以是任何氨基酸
<220>
<221>MISC_FEATURE
<222>(34)..(34)
<223>″X″可以是任何氨基酸
<220>
<221>MISC_FEATURE
<222>(43)..(43)
<223>″X″可以是任何氨基酸
<220>
<221>MISC_FEATURE
<222>(50)..(50)
<223>″X″可以是任何氨基酸
<220>
<221>MISC_FEATURE
<222>(52)..(52)
<223>″X″可以是任何氨基酸
<220>
<221>MISC_FEATURE
<222>(54)..(54)
<223>″X″可以是任何氨基酸
<220>
<221>MISC_FEATURE
<222>(57)..(57)
<223>″X″可以是任何氨基酸
<220>
<221>MISC_FEATURE
<222>(59)..(59)
<223>″X″可以是任何氨基酸
<220>
<221>MISC_FEATURE
<222>(99)..(99)
<223>″X″可以是任何氨基酸
<220>
<221>MISC_FEATURE
<222>(100)..(100)
<223>″X″可以是任何氨基酸
<220>
<221>MISC_FEATURE
<222>(103)..(108)
<223>″X″可以是任何氨基酸
<220>
<221>MISC_FEATURE
<222>(116)..(116)
<223>″X″可以是任何氨基酸
<400>81
Gln Val Gln Leu Gln Gln Ser Gly Ala Glu Leu Val Lys Pro Gly Ala
1 5 10 15
Ser Val Lys Xaa Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser Tyr
20 25 30
Trp Xaa His Trp Val Lys Gln Arg Pro Gly Xaa Gly Leu Glu Trp Ile
35 40 45
Gly Xaa Ile Xaa Pro Xaa Ser Gly Xaa Thr Xaa Tyr Asn Glu Lys Phe
50 55 60
Lys Gly Lys Ala Thr Leu Thr Val Asp Lys Ser Ser Ser Thr Ala Tyr
65 70 75 80
Met Gln Leu Ser Ser Leu Thr Ser Glu Asp Ser Ala Val Val Tyr Cys
85 90 95
Ala Arg Xaa Xaa Tyr Tyr Xaa Xaa Xaa Xaa Xaa Xaa Asp Tyr Trp Gly
100 105 110
Gln Gly Thr Xaa Val Thr Val Ser Ser
115 120
<210>82
<211>113
<212>PRT
<213>小家鼠(Mus musculus)
<400>82
Asp Val Leu Met Thr Gln Thr Pro Leu Ser Leu Pro Val Ser Leu Gly
1 5 10 15
Asp Gln Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Ile Val His Ser
20 25 30
Asn Val Asn Thr Tyr Leu Glu Trp Tyr Leu Gln Lys Pro Gly Gln Ser
35 40 45
Pro Lys Leu Leu Ile Tyr Lys Val Ser Asn Arg Phe Ser Gly Val Pro
50 55 60
Asp Arg Phe Ser Gly Ser Gly Ser Gly Thr Asp Phe Thr Leu Arg Ile
65 70 75 80
Ser Arg Val Glu Ala Glu Asp Leu Gly Ile Tyr Tyr Cys Phe Gln Gly
85 90 95
Ser His Val Pro Pro Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys
100 105 110
Arg
<210>83
<211>113
<212>PRT
<213>人工序列
<220>
<223>人源化的EM164抗体
<400>83
Asp Val Val Met Thr Gln Thr Pro Leu Ser Leu Pro Val Ser Leu Gly
1 5 10 15
Asp Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Ile Val His Ser
20 25 30
Asn Val Asn Thr Tyr Leu Glu Trp Tyr Leu Gln Lys Pro Gly Gln Ser
35 40 45
Pro Arg Leu Leu Ile Tyr Lys Val Ser Asn Arg Phe Ser Gly Val Pro
50 55 60
Asp Arg Phe Ser Gly Ser Gly Ala Gly Thr Asp Phe Thr Leu Arg Ile
65 70 75 80
Ser Arg Val Glu Ala Glu Asp Leu Gly Ile Tyr Tyr Cys Phe Gln Gly
85 90 95
Ser His Val Pro Pro Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys
100 105 110
Arg
<210>84
<211>113
<212>PRT
<213>人工序列
<220>
<223>人源化的EM164抗体
<400>84
Asp Val Leu Met Thr Gln Thr Pro Leu Ser Leu Pro Val Ser Leu Gly
1 5 10 15
Asp Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Ile Val His Ser
20 25 30
Asn Val Asn Thr Tyr Leu Glu Trp Tyr Leu Gln Lys Pro Gly Gln Ser
35 40 45
Pro Lys Leu Leu Ile Tyr Lys Val Ser Asn Arg Phe Ser Gly Val Pro
50 55 60
Asp Arg Phe Ser Gly Ser Gly Ala Gly Thr Asp Phe Thr Leu Arg Ile
65 70 75 80
Ser Arg Val Glu Ala Glu Asp Leu Gly Ile Tyr Tyr Cys Phe Gln Gly
85 90 95
Ser His Val Pro Pro Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys
100 105 110
Arg
<210>85
<211>113
<212>PRT
<213>人工序列
<220>
<223>人源化的EM164抗体
<400>85
Asp Val Leu Met Thr Gln Thr Pro Leu Ser Leu Pro Val Ser Leu Gly
1 5 10 15
Asp Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Ile Val His Ser
20 25 30
Asn Val Asn Thr Tyr Leu Glu Trp Tyr Leu Gln Lys Pro Gly Gln Ser
35 40 45
Pro Arg Leu Leu Ile Tyr Lys Val Ser Asn Arg Phe Ser Gly Val Pro
50 55 60
Asp Arg Phe Ser Gly Ser Gly Ala Gly Thr Asp Phe Thr Leu Arg Ile
65 70 75 80
Ser Arg Val Glu Ala Glu Asp Leu Gly Ile Tyr Tyr Cys Phe Gln Gly
85 90 95
Ser His Val Pro Pro Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys
100 105 110
Arg
<210>86
<211>113
<212>PRT
<213>人工序列
<220>
<223>人源化的EM164抗体
<400>86
Asp Val Val Met Thr Gln Thr Pro Leu Ser Leu Pro Val Ser Leu Gly
1 5 10 15
Asp Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Ile Val His Ser
20 25 30
Asn Val Asn Thr Tyr Leu Glu Trp Tyr Leu Gln Lys Pro Gly Gln Ser
35 40 45
Pro Lys Leu Leu Ile Tyr Lys Val Ser Ash Arg Phe Ser Gly Val Pro
50 55 60
Asp Arg Phe Ser Gly Ser Gly Ala Gly Thr Asp Phe Thr Leu Arg Ile
65 70 75 80
Ser Arg Val Glu Ala Glu Asp Leu Gly Ile Tyr Tyr Cys Phe Gln Gly
85 90 95
Ser His Val Pro Pro Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys
100 105 110
Arg
<210>87
<211>123
<212>PRT
<213>小家鼠(Mus musculus)
<400>87
Gln Val Gln Leu Gln Gln Ser Gly Ala Glu Leu Val Lys Pro Gly Ala
1 5 10 15
Ser Val Lys Leu Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser Tyr
20 25 30
Trp Met His Trp Val Lys Gln Arg Pro Gly Gln Gly Leu Glu Trp Ile
35 40 45
Gly Glu Ile Asn Pro Ser Asn Gly Arg Thr Asn Tyr Asn Glu Lys Phe
50 55 60
Lys Arg Lys Ala Thr Leu Thr Val Asp Lys Ser Ser Ser Thr Ala Tyr
65 70 75 80
Met Gln Leu Ser Ser Leu Thr Ser Glu Asp Ser Ala Val Tyr Tyr Phe
85 90 95
Ala Arg Gly Arg Pro Asp Tyr Tyr Gly Ser Ser Lys Trp Tyr Phe Asp
100 105 110
Val Trp Gly Ala Gly Thr Thr Val Thr Val Ser
115 120
<210>88
<211>123
<212>PRT
<213>人工序列
<220>
<223>人源化的EM164抗体
<400>88
Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Val Lys Pro Gly Ala
1 5 10 15
Ser Val Lys Leu Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser Tyr
20 25 30
Trp Met His Trp Val Lys Gln Arg Pro Gly Gln Gly Leu Glu Trp Ile
35 40 45
Gly Glu Ile Asn Pro Ser Asn Gly Arg Thr Asn Tyr Asn Gln Lys Phe
50 55 60
Gln Gly Lys Ala Thr Leu Thr Val Asp Lys Ser Ser Ser Thr Ala Tyr
65 70 75 80
Met Gln Leu Ser Ser Leu Thr Ser Glu Asp Ser Ala Val Tyr Tyr Phe
85 90 95
Ala Arg Gly Arg Pro Asp Tyr Tyr Gly Ser Ser Lys Trp Tyr Phe Asp
100 105 110
Val Trp Gly Gln Gly Thr Thr Val Thr Val Ser
115 120
<210>89
<211>339
<212>DNA
<213>人工序列
<220>
<223>人源化EM164抗体的可变区-轻链
<220>
<221>CDS
<222>(1)..(339)
<400>89
gat gtt gtg atg acc caa act cca ctc tcc ctg cct gtc agt ctt gga 48
Asp Val Val Met Thr Gln Thr Pro Leu Ser Leu Pro Val Ser Leu Gly
1 5 10 15
gat cca gcc tcc atc tct tgc aga tct agt cag agc ata gta cat agt 96
Asp Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Ile Val His Ser
20 25 30
aat gta aac acc tat tta gaa tgg tac ctg cag aaa cca ggc cag tct 144
Asn Val Asn Thr Tyr Leu Glu Trp Tyr Leu Gln Lys Pro Gly Gln Ser
35 40 45
cca agg ctc ctg atc tac aaa gtt tcc aac cga ttt tct ggg gtc cca 192
Pro Arg Leu Leu Ile Tyr Lys Val Ser Asn Arg Phe Ser Gly Val Pro
50 55 60
gac agg ttc agt ggc agt gga gca ggg aca gat ttc aca ctc agg atc 240
Asp Arg Phe Ser Gly Ser Gly Ala Gly Thr Asp Phe Thr Leu Arg Ile
65 70 75 80
agc aga gtg gag gct gag gat ctg gga att tat tac tgc ttt caa ggt 288
Ser Arg Val Glu Ala Glu Asp Leu Gly Ile Tyr Tyr Cys Phe Gln Gly
85 90 95
tca cat gtt cct ccg acg ttc ggt gga ggc acc aaa ctg gaa atc aaa 336
Ser His Val Pro Pro Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys
100 105 110
cgt 339
Arg
<210>90
<211>113
<212>PRT
<213>人工序列
<220>
<223>合成的构建物
<400>90
Asp Val Val Met Thr Gln Thr Pro Leu Ser Leu Pro Val Ser Leu Gly
1 5 10 15
Asp Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Ile Val His Ser
20 25 30
Asn Val Asn Thr Tyr Leu Glu Trp Tyr Leu Gln Lys Pro Gly Gln Ser
35 40 45
Pro Arg Leu Leu Ile Tyr Lys Val Ser Asn Arg Phe Ser Gly Val Pro
50 55 60
Asp Arg Phe Ser Gly Ser Gly Ala Gly Thr Asp Phe Thr Leu Arg Ile
65 70 75 80
Ser Arg Val Glu Ala Glu Asp Leu Gly Ile Tyr Tyr Cys Phe Gln Gly
85 90 95
Ser His Val Pro Pro Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys
100 105 110
Arg
<210>91
<211>369
<212>DNA
<213>人工序列
<220>
<223>人源化EM164抗体的可变区-重链
<220>
<221>CDS
<222>(1)..(369)
<400>91
cag gtc caa ctg gtg cag tct ggg gct gaa gtg gtg aag cct ggg gct 48
Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Val Lys Pro Gly Ala
1 5 10 15
tca gtg aag ctg tcc tgt aag get tct ggc tac acc ttc acc agc tac 96
Ser Val Lys Leu Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser Tyr
20 25 30
tgg atg cac tgg gtg aag cag agg cct gga caa ggc ctt gag tgg att 144
Trp Met His Trp Val Lys Gln Arg Pro Gly Gln Gly Leu Glu Trp Ile
35 40 45
gga gag att aat cct agc aac ggt cgt act aac tac aat cag aag ttc 192
Gly Glu Ile Asn Pro Ser Asn Gly Arg Thr Asn Tyr Asn Gln Lys Phe
50 55 60
cag ggg aag gcc aca ctg act gta gac aaa tcc tcc agc aca gcc tac 240
Gln Gly Lys Ala Thr Leu Thr Val Asp Lys Ser Ser Ser Thr Ala Tyr
65 70 75 80
atg caa ctc agc agc ctg aca tct gag gac tct gcg gtc tat tac ttt 288
Met Gln Leu Ser Ser Leu Thr Ser Glu Asp Ser Ala Val Tyr Tyr Phe
85 90 95
gca aga gga aga cca gat tac tac ggt agt agc aag tgg tac ttc gat 336
Ala Arg Gly Arg Pro Asp Tyr Tyr Gly Ser Ser Lys Trp Tyr Phe Asp
100 105 110
gtc tgg ggc caa ggg acc acg gtc acc gtc tcc 369
Val Trp Gly Gln Gly Thr Thr Val Thr Val Ser
115 120
<210>92
<211>123
<212>PRT
<213>人工序列
<220>
<223>合成的构建物
<400>92
Gln Val Gln Leu Val Gln Ser Gly Ala Glu Val Val Lys Pro Gly Ala
1 5 10 15
Ser Val Lys Leu Ser Cys Lys Ala Ser Gly Tyr Thr Phe Thr Ser Tyr
20 25 30
Trp Met His Trp Val Lys Gln Arg Pro Gly Gln Gly Leu Glu Trp Ile
35 40 45
Gly Glu Ile Asn Pro Ser Asn Gly Arg Thr Asn Tyr Asn Gln Lys Phe
50 55 60
Gln Gly Lys Ala Thr Leu Thr Val Asp Lys Ser Ser Ser Thr Ala Tyr
65 70 75 80
Met Gln Leu Ser Ser Leu Thr Ser Glu Asp Ser Ala Val Tyr Tyr Phe
85 90 95
Ala Arg Gly Arg Pro Asp Tyr Tyr Gly Ser Ser Lys Trp Tyr Phe Asp
100 105 110
Val Trp Gly Gln Gly Thr Thr Val Thr Val Ser
115 120
<210>93
<211>339
<212>DNA
<213>人工序列
<220>
<223>人源化EM164 v1.1抗体的轻链可变区
<220>
<221>CDS
<222>(1)..(339)<400>93
gat gtt ttg atg acc caa act cca ctc tcc ctg cct gtc agt ctt gga 48
Asp Val Leu Met Thr Gln Thr Pro Leu Ser Leu Pro Val Ser Leu Gly
1 5 10 15
gat cca gcc tcc atc tct tgc aga tct agt cag agc ata gta cat agt 96
Asp Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Ile Val His Ser
20 25 30
aat gta aac acc tat tta gaa tgg tac ctg cag aaa cca ggc cag tct 144
Asn Val Asn Thr Tyr Leu Glu Trp Tyr Leu Gln Lys Pro Gly Gln Ser
35 40 45
cca aag ctc ctg atc tac aaa gtt tcc aac cga ttt tct ggg gtc cca 192
Pro Lys Leu Leu Ile Tyr Lys Val Ser Asn Arg Phe Ser Gly Val Pro
50 55 60
gac agg ttc agt ggc agt gga gca ggg aca gat ttc aca ctc agg atc 240
Asp Arg Phe Ser Gly Ser Gly Ala Gly Thr Asp Phe Thr Leu Arg Ile
65 70 75 80
agc aga gtg gag gct gag gat ctg gga att tat tac tgc ttt caa ggt 288
Ser Arg Val Glu Ala Glu Asp Leu Gly Ile Tyr Tyr Cys Phe Gln Gly
85 90 95
tca cat gtt cct ccg acg ttc ggt gga ggc acc aaa ctg gaa atc aaa 336
Ser His Val Pro Pro Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys
100 105 110
cgt 339
Arg
<210>94
<211>113
<212>PRT
<213>人工序列
<220>
<223>合成的构建物
<400>94
Asp Val Leu Met Thr Gln Thr Pro Leu Ser Leu Pro Val Ser Leu Gly
1 5 10 15
Asp Pro Ala Ser Ile Ser Cys Arg Ser Ser Gln Ser Ile Val His Ser
20 25 30
Asn Val Asn Thr Tyr Leu Glu Trp Tyr Leu Gln Lys Pro Gly Gln Ser
35 40 45
Pro Lys Leu Leu Ile Tyr Lys Val Ser Asn Arg Phe Ser Gly Val Pro
50 55 60
Asp Arg Phe Ser Gly Ser Gly Ala Gly Thr Asp Phe Thr Leu Arg Ile
65 70 75 80
Ser Arg Val Glu Ala Glu Asp Leu Gly Ile Tyr Tyr Cys Phe Gln Gly
85 90 95
Ser His Val Pro Pro Thr Phe Gly Gly Gly Thr Lys Leu Glu Ile Lys
100 105 110
Arg
<210>95
<211>339
<212>DNA
<213>人工序列
<220>
<223>人源化EM164 v1.2抗体的轻链可变区
<400>95
gatgttttga tgacccaaac tccactctcc ctgcctgtca gtcttggaga tccagcctcc 60
atctcttgca gatctagtca gagcatagta catagtaatg taaacaccta tttagaatgg 120
tacctgcaga aaccaggcca gtctccaagg ctcctgatct acaaagtttc caaccgattt 180
tctggggtcc cagacaggtt cagtggcagt ggagcaggga cagatttcac actcaggatc 240
agcagagtgg aggctgagga tctgggaatt tattactgct ttcaaggttc acatgttcct 300
ccgacgttcg gtggaggcac caaactggaa atcaaacgt 339
<210>96
<211>339
<212>DNA
<213>人工序列
<220>
<223>人源化EM164 v1.3抗体的轻链可变区
<400>96
gatgttgtga tgacccaaac tccactctcc ctgcctgtca gtcttggaga tccagcctcc 60
atctcttgca gatctagtca gagcatagta catagtaatg taaacaccta tttagaatgg 120
tacctgcaga aaccaggcca gtctccaaag ctcctgatct acaaagtttc caaccgattt 180
tctggggtcc cagacaggtt cagtggcagt ggagcaggga cagatttcac actcaggatc 240
agcagagtgg aggctgagga tctgggaatt tattactgct ttcaaggttc acatgttcct 300
ccgacgttcg gtggaggcac caaactggaa atcaaacgt 339
Claims (31)
1.一种组合物,包括:
(a)第一治疗剂,其中所述第一治疗剂是抗体或其表位-结合片段,以及其中所述抗体或所述片段特异性地结合到胰岛素-样生长因子-I受体,所述第一治疗剂选自:
(i)抗体,或其表位-结合片段,具有与鼠抗体EM164相同的氨基酸序列,所述鼠抗体EM164由小鼠杂交瘤EM164(ATCC登记号PTA-4457)产生,
(ii)表面重构抗体,或其表位-结合片段,具有与鼠抗体EM164相同的结合特异性,
(iii)人抗体或人源化的抗体,或其表位-结合片段,具有与鼠抗体EM164相同的结合特异性,
(iv)抗体的功能等价物,或其表位-结合片段,具有与鼠抗体EM164相同的结合特异性,
(v)鼠抗体EM164的变体,或其表位-结合片段,相比起鼠抗体EM164,具有至少一个核苷酸突变、删除或插入,并有与鼠抗体EM164相同的结合特异性,和
(vi)由鼠杂交瘤EM164(ATCC登记号PTA-4457)产生的鼠抗体EM164,或其表位-结合片段,和
(b)第二治疗剂。
2.根据权利要求1所述的组合物,其中所述第二治疗剂选自多烯紫杉醇、紫杉酚、阿霉素、表阿霉素、环磷酰胺、曲妥珠(Herceptin)、卡培他宾、它莫西芬、托瑞米芬、来曲唑、阿纳曲唑、氟维司群、依西美坦、戈舍瑞林、奥沙利铂、卡铂、顺铂、地塞米松、抗排卵肽、贝伐单抗(Avastin)、5-氟尿嘧啶、亚叶酸、左旋咪唑、依立替康、依托泊甙、拓朴替康、吉西他滨、长春瑞滨、雌莫司汀、米托蒽醌、阿巴瑞克、唑来瞵酸盐、链硝脲、妥昔单抗(Rituxan)、柔红霉素、白消安、苯丁酸氮芥、氟阿糖腺苷、伊马替尼、阿糖胞苷、伊莫单抗(Zevalin)、托西莫单抗(Bexxar)、α干扰素-2b、美法仑、硼替佐米(Velcade)、六甲蜜胺、门冬酰胺酶、吉非替尼(Iressa)、埃洛尼替(Tarceva)、抗-EGF受体抗体(Cetuximab、Abx-EGF)和埃坡霉素。
3.根据权利要求1所述的组合物,其中所述第二治疗剂选自卡铂、奥沙利铂、顺铂、紫杉酚、多烯紫杉醇、吉西他滨和喜树碱。
4.根据权利要求1所述的组合物,其中所述第一治疗剂以约1mg/平方米至约2000mg/平方米的剂量给予患者,其中所述第二治疗剂以约10mg/平方米至约2000mg/平方米的剂量给予所述患者。
5.根据权利要求1所述的组合物,其中所述第一治疗剂以约10mg/平方米至约1000mg/平方米的剂量给予患者,其中所述第二治疗剂以约50mg/平方米至约1000mg/平方米的剂量给予所述患者。
6.一种药物组合物,包括根据权利要求1所述的组合物,和药学上可接受的载体或稀释剂。
7.一种组合物,包括
(a)第一治疗剂,其中所述第一治疗剂是包含至少一个互补性决定区的抗体或抗体片段,所述互补性决定区具有选自如下的氨基酸序列
SYWMH (SEQ ID NO:1),
EINPSNGRTNYNEKFKR (SEQ ID NO:2),
GRPDYYGSSKWYFDV (SEQ ID NO:3),
RSSQSIVHSNVNTYLE (SEQ ID NO:4),
KVSNRFS (SEQ ID NO:5),和
FQGSHVPPT (SEQ ID NO:6),和
(b)第二治疗剂。
8.一种组合物,包括
(a)第一治疗剂,其中所述第一治疗剂是包含至少一个重链可变区和至少一个轻链可变区的抗体或抗体片段,其中所述重链可变区包括三个连续的互补-决定区,所述互补-决定区分别具有SEQ ID NOS:1-3描述的氨基酸序列:
SYWMH (SEQ ID NO:1),
EINPSNGRTNYNEKFKR (SEQ ID NO:2),
GRPDYYGSSKWYFDV (SEQ ID NO:3);
以及其中所述轻链可变区包括三个连续的互补-决定区,所述互补-决定区分别具有SEQ ID NOS:4-6描述的氨基酸序列:
RSSQSIVHSNVNTYLE (SEQ ID NO:4),
KVSNRFS (SEQ ID NO:5),
FQGSHVPPT (SEQ ID NO:6),和
b)第二治疗剂。
9.一种组合物,包括:
(a)第一治疗剂,其中所述第一治疗剂是抗体或其片段,其中所述的抗体包含与SEQ ID NO:7描述的氨基酸序列具有至少90%的序列同一性的重链可变区:
QVQLQQSGAELVKPGASVKLSCKASGYTFTSYWMHWVKQRPGQGLEWIGEINPSNGRTNYNEKFKRKATLTVDKSSSTAYMQLSSLTSEDSAVYYFARGRPDYYGSSKWYFDVWGAGTTVTVSS(SEQ IDNO:7)和
(b)第二治疗剂。
10.权利要求9的组合物,其中所述重链可变区与SEQ ID NO:7描述的氨基酸序列具有至少95%的序列同一性。
11.权利要求9的组合物,其中所述重链可变区具有SEQ ID NO:7描述的氨基酸序列。
12.一种组合物,包括:
(a)第一治疗剂,其中所述第一治疗剂是抗体或其片段,其中所述抗体包括轻链可变区,该轻链可变区与SEQ ID NO:8描述的氨基酸序列具有至少90%的序列同一性:
DVLMTQTPLSLPVSLGDQASISCRSSQSIVHSNVNTYLEWYLQKPGQSPKLLIYKVSNRFSGVPDRFSGSGSGTDFTLRISRVEAEDLGIYYCFQGSHVPPTFGGGTKLEIKR(SEQ ID NO:8),和
(b)第二治疗剂。
13.权利要求12的组合物,其中所述轻链可变区与SEQ ID NO:8表示的氨基酸序列具有至少95%的序列同一性。
14.权利要求12的组合物,其中所述轻链可变区具有SEQ ID NO:8表示的氨基酸序列。
15.一种组合物,包括:
(a)第一治疗剂,其中所述第一治疗剂是抗体或其片段,其含有具有选自下列序列的轻链可变区:
DVVMTQTPLSLPVSLGDPASISCRSSQSIVHSNVNTYLEWYLQKPGQSPRLLIYKVSNRFSGVPDRFSGSGAGTDFTLRISRVEAEDLGIYYCFQGSHVPPTFGGGTKLEIKR(SEQ ID NO:9);
DVLMTQTPLSLPVSLGDPASISCRSSQSIVHSNVNTYLEWYLQKPGQSPKLLIYKVSNRFSGVPDRFSGSGAGTDFTLRISRVEAEDLGIYYCFQGSHVPPTFGGGTKLEIKR(SEQ ID NO:10);
DVLMTQTPLSLPVSLGDPASISCRSSQSIVHSNVNTYLEWYLQKPGQSPRLLIYKVSNRFSGVPDRFSGSGAGTDFTLRISRVEAEDLGIYYCFQGSHVPPTFGGGTKLEIKR(SEQ ID NO:11);
DVVMTQTPLSLPVSLGDPASISCRSSQSIVHSNVNTYLEWYLQKPGQSPKLLIYKVSNRFSGVPDRFSGSGAGTDFTLRISRVEAEDLGIYYCFQGSHVPPTFGGGTKLEIKR(SEQ ID NO:12)和
(b)第二治疗剂。
16.一种组合物,包括:
(a)第一治疗剂,其中所述第一治疗剂是抗体或其片段,其含有具有SEQ ID NO:13所示的序列的重链可变区:
QVQLVQSGAEVVKPGASVKLSCKASGYTFTSYWMHWVKQRPGQGLEWIGEINPSNGRTNYNQKFQGKATLTVDKSSSTAYMQLSSLTSEDSAVYYFARGRPDYYGSSKWYFDVWGQGTTVTVSS(SEQ ID NO:13),和
(b)第二治疗剂。
17.权利要求7-16中的任一项所述的组合物,其中所述第二治疗剂选自多烯紫杉醇、紫杉酚、阿霉素、表阿霉素、环磷酰胺、曲妥珠(Herceptin)、卡培他宾、它莫西芬、托瑞米芬、来曲唑、阿纳曲唑、氟维司群、依西美坦、戈舍瑞林、奥沙利铂、卡铂、顺铂、地塞米松、抗排卵肽、贝伐单抗(Avastin)、5-氟尿嘧啶、亚叶酸、左旋咪唑、依立替康、依托泊甙、拓朴替康、吉西他滨、长春瑞滨、雌莫司汀、米托蒽醌、阿巴瑞克、唑来瞵酸盐、链硝脲、妥昔(Rituxan)、柔红霉素、白消安、苯丁酸氮芥、氟阿糖腺苷、伊马替尼、阿糖胞苷、伊莫单抗(Zevalin)、托西莫单抗(Bexxar)、α干扰素-2b、美法仑、硼替佐米(Velcade)、六甲蜜胺、门冬酰胺酶、吉非替尼(Iressa)、埃洛尼替(Tarceva)、抗-EGF受体抗体(Cetuximab、Abx-EGF)和埃坡霉素。
18.权利要求7-16中的任一项所述的组合物,其中所述第二治疗剂选自卡铂、奥沙利铂、顺铂、紫杉酚、多烯紫杉醇、吉西他滨和喜树碱。
19.一种抑制癌细胞生长的方法,包括将所述细胞与权利要求1的组合物接触。
20.一种治疗患有癌症的患者的方法,包括给予所述患者有效剂量的权利要求1的组合物。
21.一种治疗患有癌症的患者的方法,包括给予所述患者有效剂量的权利要求6的药物组合物。
22.权利要求19-21中的任一项的治疗方法,其中所述癌症是选自乳癌、结肠癌、卵巢癌、骨肉瘤、宫颈癌、前列腺癌、肺癌、滑膜肉瘤、胰腺癌、黑素瘤、多发性骨髓瘤、神经母细胞瘤和横纹肌肉瘤的癌症。
23.一种试剂盒,包括
(a)第一治疗剂,所述第一治疗剂是具有与由鼠杂交瘤EM164(ATCC登记号PTA-4457)产生的鼠抗体EM164相同的氨基酸序列的抗体,或其表位-结合片段,其中所述抗体或所述片段特异性地结合到胰岛素-样生长因子-I受体,
(b)第二治疗剂,和
(c)使用说明书。
24.一种抑制癌症细胞生长的方法,包括用下列制剂接触所述细胞:
(a)第一治疗剂,所述第一治疗剂是具有与由鼠杂交瘤EM164(ATCC登记号PTA-4457)产生的鼠抗体EM164相同的氨基酸序列的抗体,或其表位-结合片段,其中所述抗体或所述片段特异性地结合到胰岛素-样生长因子-I受体,和
(b)第二治疗剂。
25.一种治疗患有癌症的患者的方法,包括给予所述患者有效剂量的:
(a)第一治疗剂,所述第一治疗剂是具有与由鼠杂交瘤EM164(ATCC登记号PTA-4457)产生的鼠抗体EM164相同的氨基酸序列的抗体,或其表位-结合片段,其中所述抗体或所述片段特异性地结合到胰岛-样生长因子-I受体,和
(b)第二治疗剂。
26.权利要求24的方法,其中所述细胞同时与所述第一治疗剂和所述第二治疗剂接触。
27.权利要求24的方法,其中所述细胞依次与所述第一治疗剂和所述第二治疗剂接触,并且用两种次序中的任意一种次序。
28.权利要求25的方法,其中所述第一治疗剂和所述第二治疗剂同时给药。
29.权利要求25的方法,其中所述第一治疗剂和所述第二治疗剂依次给药,并且用两种次序中的任意一种次序。
30.权利要求24或25所述的方法,其中所述第二治疗剂选自多烯紫杉醇、紫杉酚、阿霉素、表阿霉素、环磷酰胺、曲妥珠(Herceptin)、卡培他宾、它莫西芬、托瑞米芬、来曲唑、阿纳曲唑、氟维司群、依西美坦、戈舍瑞林、奥沙利铂、卡铂、顺铂、地塞米松、抗排卵肽、贝伐单抗(Avastin)、5-氟尿嘧啶、亚叶酸、左旋咪唑、依立替康、依托泊甙、拓朴替康、吉西他滨、长春瑞滨、雌莫司汀、米托蒽醌、阿巴瑞克、唑来瞵酸盐、链硝脲、妥昔单抗(Rituxan)、柔红霉素、白消安、苯丁酸氮芥、氟阿糖腺苷、伊马替尼、阿糖胞苷、伊莫单抗(Zevalin)、托西莫单抗(Bexxar)、α干扰素-2b、美法仑、硼替佐米(Velcade)、六甲蜜胺、门冬酰胺酶、吉非替尼(Iressa)、埃洛尼替(Tarceva)、抗-EGF受体抗体(Cetuximab、Abx-EGF)和埃坡霉素。
31.权利要求24或25的方法,其中所述第二治疗剂选自卡铂、奥沙利铂、顺铂、紫杉酚、多烯紫杉醇、吉西他滨和喜树碱。
Applications Claiming Priority (2)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
US10/729,441 US8034904B2 (en) | 2002-06-14 | 2003-12-08 | Anti-IGF-I receptor antibody |
US10/729,441 | 2003-12-08 |
Publications (1)
Publication Number | Publication Date |
---|---|
CN1886424A true CN1886424A (zh) | 2006-12-27 |
Family
ID=36616549
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CNA200480034889XA Pending CN1886424A (zh) | 2003-12-08 | 2004-12-07 | 抗igf-i受体抗体 |
Country Status (13)
Country | Link |
---|---|
EP (1) | EP1692176A4 (zh) |
JP (1) | JP2008502589A (zh) |
KR (1) | KR20070001883A (zh) |
CN (1) | CN1886424A (zh) |
AU (1) | AU2004303792A1 (zh) |
BR (1) | BRPI0417406A (zh) |
CA (1) | CA2548065A1 (zh) |
CR (1) | CR8426A (zh) |
EA (1) | EA009807B1 (zh) |
EC (1) | ECSP066595A (zh) |
IL (1) | IL174770A0 (zh) |
MX (1) | MXPA06005540A (zh) |
NO (1) | NO20063155L (zh) |
Cited By (3)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN102905723A (zh) * | 2010-05-11 | 2013-01-30 | Aveo制药公司 | 抗fgfr2抗体 |
CN103509117A (zh) * | 2013-05-06 | 2014-01-15 | 江苏匡亚生物医药科技有限公司 | 抗人her2和人igf-ir的双特异性抗体及其制备方法和用途 |
CN110691794A (zh) * | 2017-05-30 | 2020-01-14 | 帝人制药株式会社 | 抗igf-i受体抗体 |
Families Citing this family (2)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
KR20190067275A (ko) | 2009-12-21 | 2019-06-14 | 제넨테크, 인크. | 항체 제제 |
US9051561B2 (en) | 2011-10-10 | 2015-06-09 | Children's Hospital Los Angeles | Asparaginase and treating diseases associated with asparagine dependence |
Family Cites Families (4)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
JP2002532685A (ja) * | 1998-12-04 | 2002-10-02 | ノバルティス アクチエンゲゼルシャフト | 活性化ビトロネクチンレセプターαVβ3をターゲティングするために有用な方法および組成物 |
EP1399483B1 (en) * | 2001-01-05 | 2010-04-14 | Pfizer Inc. | Antibodies to insulin-like growth factor i receptor |
US7538195B2 (en) * | 2002-06-14 | 2009-05-26 | Immunogen Inc. | Anti-IGF-I receptor antibody |
KR20050109489A (ko) * | 2003-02-13 | 2005-11-21 | 화이자 프로덕츠 인크. | 항-인슐린양 성장인자 i 수용체 항체의 용도 |
-
2004
- 2004-12-07 KR KR1020067010010A patent/KR20070001883A/ko not_active Application Discontinuation
- 2004-12-07 CN CNA200480034889XA patent/CN1886424A/zh active Pending
- 2004-12-07 EA EA200600931A patent/EA009807B1/ru not_active IP Right Cessation
- 2004-12-07 MX MXPA06005540A patent/MXPA06005540A/es active IP Right Grant
- 2004-12-07 EP EP04811082A patent/EP1692176A4/en not_active Ceased
- 2004-12-07 BR BRPI0417406-2A patent/BRPI0417406A/pt not_active IP Right Cessation
- 2004-12-07 AU AU2004303792A patent/AU2004303792A1/en not_active Abandoned
- 2004-12-07 CA CA002548065A patent/CA2548065A1/en not_active Abandoned
- 2004-12-07 JP JP2006543832A patent/JP2008502589A/ja active Pending
-
2006
- 2006-04-04 IL IL174770A patent/IL174770A0/en unknown
- 2006-05-31 EC EC2006006595A patent/ECSP066595A/es unknown
- 2006-06-01 CR CR8426A patent/CR8426A/es not_active Application Discontinuation
- 2006-07-07 NO NO20063155A patent/NO20063155L/no not_active Application Discontinuation
Cited By (5)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN102905723A (zh) * | 2010-05-11 | 2013-01-30 | Aveo制药公司 | 抗fgfr2抗体 |
CN103509117A (zh) * | 2013-05-06 | 2014-01-15 | 江苏匡亚生物医药科技有限公司 | 抗人her2和人igf-ir的双特异性抗体及其制备方法和用途 |
CN103509117B (zh) * | 2013-05-06 | 2016-03-09 | 江苏匡亚生物医药科技有限公司 | 抗人her2和人igf-ir的双特异性抗体及其制备方法和用途 |
CN110691794A (zh) * | 2017-05-30 | 2020-01-14 | 帝人制药株式会社 | 抗igf-i受体抗体 |
CN110691794B (zh) * | 2017-05-30 | 2023-10-27 | 帝人制药株式会社 | 抗igf-i受体抗体 |
Also Published As
Publication number | Publication date |
---|---|
AU2004303792A1 (en) | 2005-07-07 |
CA2548065A1 (en) | 2005-07-07 |
KR20070001883A (ko) | 2007-01-04 |
ECSP066595A (es) | 2006-10-17 |
NO20063155L (no) | 2006-08-11 |
EP1692176A1 (en) | 2006-08-23 |
IL174770A0 (en) | 2006-08-20 |
EP1692176A4 (en) | 2008-11-12 |
EA009807B1 (ru) | 2008-04-28 |
MXPA06005540A (es) | 2006-08-17 |
EA200600931A1 (ru) | 2006-10-27 |
JP2008502589A (ja) | 2008-01-31 |
CR8426A (es) | 2007-12-04 |
BRPI0417406A (pt) | 2007-04-03 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
CN1678633A (zh) | 抗igf-i受体抗体 | |
JP5863926B2 (ja) | ヒト血清アルブミンリンカーおよびそのコンジュゲート | |
CN107849142B (zh) | 拮抗性抗肿瘤坏死因子受体超家族抗体 | |
JP5677972B2 (ja) | ヒト血清アルブミンリンカーおよびそのコンジュゲート | |
US10407511B2 (en) | Covalently linked helicar-anti-helicar antibody conjugates and uses thereof | |
CN1901937A (zh) | 使用拮抗性抗-cd40单克隆抗体治疗多发性骨髓瘤 | |
CN100335132C (zh) | 含有受体酪氨酸激酶抑制剂和血管生成抑制剂的药物组合物及试剂盒 | |
CN1922199A (zh) | Ca6抗原特异性细胞毒性偶联物及其应用方法 | |
CN1620468A (zh) | 新的抗igf-ir抗体及其应用 | |
CN1906214A (zh) | 抗-igfr1抗体治疗性组合 | |
CN101687039B (zh) | 工程化抗αν-整联蛋白杂合抗体 | |
CN1795009A (zh) | 抗-cd33抗体和使用其治疗急性髓性白血病的方法 | |
CN1671837A (zh) | 抗胰岛素样生长因子受体(igfr)的人源中和抗体 | |
CN1662558A (zh) | 人cd22特异性抗体及其治疗和诊断应用 | |
CN101076542A (zh) | 特异性针对肝细胞癌和其他癌的抗体及其用途 | |
CN1929862A (zh) | 拮抗性抗-cd40单克隆抗体治疗慢性淋巴细胞性白血病的应用 | |
CN101035564A (zh) | 人源化的抗5t4抗体和抗5t4抗体/加利车霉素缀合物 | |
CN1898264A (zh) | 用新的抗il13单克隆抗体治疗癌症 | |
CN101074261A (zh) | Trail受体1和/或trail受体2特异性抗体及其应用 | |
CN1905897A (zh) | 拮抗性抗-cd40单克隆抗体和它们的使用方法 | |
CN1961003A (zh) | 人源化抗TGF-β抗体 | |
CN1441677A (zh) | 抗细胞因子抗体或拮抗剂与抗-cd20在b细胞淋巴瘤治疗中的联合应用 | |
CN1646161A (zh) | 自身免疫疾病的药剂和治疗方法 | |
CN1703243A (zh) | 针对erb-b1受体的药物组合物 | |
CN1878790A (zh) | 可中和的hgf表位和与其结合的中和抗体 |
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: 1096693 Country of ref document: HK |
|
REG | Reference to a national code |
Ref country code: HK Ref legal event code: WD Ref document number: 1096693 Country of ref document: HK |
|
C12 | Rejection of a patent application after its publication | ||
RJ01 | Rejection of invention patent application after publication |
Application publication date: 20061227 |