CN105330740B - 抗pd-1抗体及其应用 - Google Patents
抗pd-1抗体及其应用 Download PDFInfo
- Publication number
- CN105330740B CN105330740B CN201410369300.7A CN201410369300A CN105330740B CN 105330740 B CN105330740 B CN 105330740B CN 201410369300 A CN201410369300 A CN 201410369300A CN 105330740 B CN105330740 B CN 105330740B
- Authority
- CN
- China
- Prior art keywords
- antibody
- cell
- polynucleotides
- seq
- heavy chain
- 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.)
- Active
Links
- 101710089372 Programmed cell death protein 1 Proteins 0.000 title description 3
- 102100023990 60S ribosomal protein L17 Human genes 0.000 title 1
- 206010028980 Neoplasm Diseases 0.000 claims abstract description 34
- 239000003814 drug Substances 0.000 claims abstract description 14
- 229940079593 drug Drugs 0.000 claims abstract description 9
- 108091033319 polynucleotide Proteins 0.000 claims description 30
- 102000040430 polynucleotide Human genes 0.000 claims description 30
- 239000002157 polynucleotide Substances 0.000 claims description 30
- 230000014509 gene expression Effects 0.000 claims description 27
- 125000003275 alpha amino acid group Chemical group 0.000 claims description 22
- 201000011510 cancer Diseases 0.000 claims description 17
- 239000013604 expression vector Substances 0.000 claims description 15
- 239000008194 pharmaceutical composition Substances 0.000 claims description 14
- 206010058467 Lung neoplasm malignant Diseases 0.000 claims description 12
- 201000001441 melanoma Diseases 0.000 claims description 11
- 201000005202 lung cancer Diseases 0.000 claims description 10
- 208000020816 lung neoplasm Diseases 0.000 claims description 10
- 208000035473 Communicable disease Diseases 0.000 claims description 8
- 239000000203 mixture Substances 0.000 claims description 6
- 108090000765 processed proteins & peptides Proteins 0.000 claims description 5
- 230000001105 regulatory effect Effects 0.000 claims description 5
- 238000000926 separation method Methods 0.000 claims description 5
- 208000031886 HIV Infections Diseases 0.000 claims description 3
- 208000037357 HIV infectious disease Diseases 0.000 claims description 3
- 208000033519 human immunodeficiency virus infectious disease Diseases 0.000 claims description 3
- 102000004196 processed proteins & peptides Human genes 0.000 claims description 3
- 241000700721 Hepatitis B virus Species 0.000 claims description 2
- 229920001184 polypeptide Polymers 0.000 claims description 2
- 239000012634 fragment Substances 0.000 abstract description 29
- 108010074708 B7-H1 Antigen Proteins 0.000 description 63
- 102000008096 B7-H1 Antigen Human genes 0.000 description 63
- 210000004027 cell Anatomy 0.000 description 55
- 210000001744 T-lymphocyte Anatomy 0.000 description 52
- 238000000034 method Methods 0.000 description 27
- 241000699666 Mus <mouse, genus> Species 0.000 description 19
- 230000002401 inhibitory effect Effects 0.000 description 17
- 102000000588 Interleukin-2 Human genes 0.000 description 16
- 108010002350 Interleukin-2 Proteins 0.000 description 16
- 230000000694 effects Effects 0.000 description 12
- 102100024213 Programmed cell death 1 ligand 2 Human genes 0.000 description 11
- 230000028993 immune response Effects 0.000 description 11
- 108020004414 DNA Proteins 0.000 description 10
- 102100037850 Interferon gamma Human genes 0.000 description 10
- 108010074328 Interferon-gamma Proteins 0.000 description 10
- 101100407308 Mus musculus Pdcd1lg2 gene Proteins 0.000 description 10
- 108700030875 Programmed Cell Death 1 Ligand 2 Proteins 0.000 description 10
- 230000002708 enhancing effect Effects 0.000 description 10
- 210000004881 tumor cell Anatomy 0.000 description 10
- 230000008485 antagonism Effects 0.000 description 9
- 239000000427 antigen Substances 0.000 description 9
- 108091007433 antigens Proteins 0.000 description 9
- 102000036639 antigens Human genes 0.000 description 9
- 239000008363 phosphate buffer Substances 0.000 description 9
- 230000004913 activation Effects 0.000 description 8
- 108090000623 proteins and genes Proteins 0.000 description 8
- 102000017420 CD3 protein, epsilon/gamma/delta subunit Human genes 0.000 description 7
- 230000036039 immunity Effects 0.000 description 7
- 210000003819 peripheral blood mononuclear cell Anatomy 0.000 description 7
- 229920001223 polyethylene glycol Polymers 0.000 description 7
- 241001465754 Metazoa Species 0.000 description 6
- 230000006044 T cell activation Effects 0.000 description 6
- 210000003719 b-lymphocyte Anatomy 0.000 description 6
- 230000006870 function Effects 0.000 description 6
- 239000000243 solution Substances 0.000 description 6
- 230000000638 stimulation Effects 0.000 description 6
- 238000002965 ELISA Methods 0.000 description 5
- 101001117317 Homo sapiens Programmed cell death 1 ligand 1 Proteins 0.000 description 5
- 108010050904 Interferons Proteins 0.000 description 5
- 102000014150 Interferons Human genes 0.000 description 5
- OUYCCCASQSFEME-QMMMGPOBSA-N L-tyrosine Chemical compound OC(=O)[C@@H](N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-QMMMGPOBSA-N 0.000 description 5
- 108091028043 Nucleic acid sequence Proteins 0.000 description 5
- 102100024216 Programmed cell death 1 ligand 1 Human genes 0.000 description 5
- 210000004443 dendritic cell Anatomy 0.000 description 5
- 201000010099 disease Diseases 0.000 description 5
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 5
- 238000005516 engineering process Methods 0.000 description 5
- 230000005764 inhibitory process Effects 0.000 description 5
- 230000003993 interaction Effects 0.000 description 5
- 229940079322 interferon Drugs 0.000 description 5
- 210000004698 lymphocyte Anatomy 0.000 description 5
- 230000004048 modification Effects 0.000 description 5
- 238000012986 modification Methods 0.000 description 5
- 108020003175 receptors Proteins 0.000 description 5
- 102000005962 receptors Human genes 0.000 description 5
- 230000006798 recombination Effects 0.000 description 5
- 238000005215 recombination Methods 0.000 description 5
- 210000001519 tissue Anatomy 0.000 description 5
- OUYCCCASQSFEME-UHFFFAOYSA-N tyrosine Natural products OC(=O)C(N)CC1=CC=C(O)C=C1 OUYCCCASQSFEME-UHFFFAOYSA-N 0.000 description 5
- 238000005406 washing Methods 0.000 description 5
- PHEDXBVPIONUQT-UHFFFAOYSA-N Cocarcinogen A1 Natural products CCCCCCCCCCCCCC(=O)OC1C(C)C2(O)C3C=C(C)C(=O)C3(O)CC(CO)=CC2C2C1(OC(C)=O)C2(C)C PHEDXBVPIONUQT-UHFFFAOYSA-N 0.000 description 4
- 230000006907 apoptotic process Effects 0.000 description 4
- 230000000903 blocking effect Effects 0.000 description 4
- 108020001507 fusion proteins Proteins 0.000 description 4
- 102000037865 fusion proteins Human genes 0.000 description 4
- 239000007928 intraperitoneal injection Substances 0.000 description 4
- 239000003446 ligand Substances 0.000 description 4
- 210000002540 macrophage Anatomy 0.000 description 4
- 230000002093 peripheral effect Effects 0.000 description 4
- PHEDXBVPIONUQT-RGYGYFBISA-N phorbol 13-acetate 12-myristate Chemical compound C([C@]1(O)C(=O)C(C)=C[C@H]1[C@@]1(O)[C@H](C)[C@H]2OC(=O)CCCCCCCCCCCCC)C(CO)=C[C@H]1[C@H]1[C@]2(OC(C)=O)C1(C)C PHEDXBVPIONUQT-RGYGYFBISA-N 0.000 description 4
- 238000012216 screening Methods 0.000 description 4
- 239000007790 solid phase Substances 0.000 description 4
- 238000007920 subcutaneous administration Methods 0.000 description 4
- 230000009385 viral infection Effects 0.000 description 4
- UAIUNKRWKOVEES-UHFFFAOYSA-N 3,3',5,5'-tetramethylbenzidine Chemical compound CC1=C(N)C(C)=CC(C=2C=C(C)C(N)=C(C)C=2)=C1 UAIUNKRWKOVEES-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
- 241000283707 Capra Species 0.000 description 3
- 241000699802 Cricetulus griseus Species 0.000 description 3
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 3
- 108010001336 Horseradish Peroxidase Proteins 0.000 description 3
- 102100040678 Programmed cell death protein 1 Human genes 0.000 description 3
- 239000006146 Roswell Park Memorial Institute medium Substances 0.000 description 3
- 241000700605 Viruses Species 0.000 description 3
- 230000003213 activating effect Effects 0.000 description 3
- 125000000539 amino acid group Chemical group 0.000 description 3
- 150000001413 amino acids Chemical class 0.000 description 3
- 230000000259 anti-tumor effect Effects 0.000 description 3
- 238000013459 approach Methods 0.000 description 3
- 230000033228 biological regulation Effects 0.000 description 3
- 239000000969 carrier Substances 0.000 description 3
- 230000008859 change Effects 0.000 description 3
- 238000010367 cloning Methods 0.000 description 3
- 238000013461 design Methods 0.000 description 3
- 238000002474 experimental method Methods 0.000 description 3
- 210000004602 germ cell Anatomy 0.000 description 3
- 230000013595 glycosylation Effects 0.000 description 3
- 238000006206 glycosylation reaction Methods 0.000 description 3
- 230000005934 immune activation Effects 0.000 description 3
- 230000001900 immune effect Effects 0.000 description 3
- 239000004615 ingredient Substances 0.000 description 3
- 238000011081 inoculation Methods 0.000 description 3
- 238000005259 measurement Methods 0.000 description 3
- 230000001404 mediated effect Effects 0.000 description 3
- 210000001616 monocyte Anatomy 0.000 description 3
- 210000001672 ovary Anatomy 0.000 description 3
- 238000006366 phosphorylation reaction Methods 0.000 description 3
- 239000013641 positive control Substances 0.000 description 3
- 230000004044 response Effects 0.000 description 3
- 229940124597 therapeutic agent Drugs 0.000 description 3
- 101710153593 Albumin A Proteins 0.000 description 2
- 208000023275 Autoimmune disease Diseases 0.000 description 2
- 108091008875 B cell receptors Proteins 0.000 description 2
- 241000894006 Bacteria Species 0.000 description 2
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 2
- 201000009030 Carcinoma Diseases 0.000 description 2
- 102000004127 Cytokines Human genes 0.000 description 2
- 108090000695 Cytokines Proteins 0.000 description 2
- 102100039498 Cytotoxic T-lymphocyte protein 4 Human genes 0.000 description 2
- LYCAIKOWRPUZTN-UHFFFAOYSA-N Ethylene glycol Chemical compound OCCO LYCAIKOWRPUZTN-UHFFFAOYSA-N 0.000 description 2
- 229920001917 Ficoll Polymers 0.000 description 2
- 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 2
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 2
- 208000009329 Graft vs Host Disease Diseases 0.000 description 2
- 102000006354 HLA-DR Antigens Human genes 0.000 description 2
- 108010058597 HLA-DR Antigens Proteins 0.000 description 2
- 101000889276 Homo sapiens Cytotoxic T-lymphocyte protein 4 Proteins 0.000 description 2
- 101000611936 Homo sapiens Programmed cell death protein 1 Proteins 0.000 description 2
- 108060003951 Immunoglobulin Proteins 0.000 description 2
- 108700005091 Immunoglobulin Genes Proteins 0.000 description 2
- 102000006496 Immunoglobulin Heavy Chains Human genes 0.000 description 2
- 108010019476 Immunoglobulin Heavy Chains Proteins 0.000 description 2
- 206010061218 Inflammation Diseases 0.000 description 2
- 108090000978 Interleukin-4 Proteins 0.000 description 2
- 238000011789 NOD SCID mouse Methods 0.000 description 2
- 102000004160 Phosphoric Monoester Hydrolases Human genes 0.000 description 2
- 108090000608 Phosphoric Monoester Hydrolases Proteins 0.000 description 2
- 229920001213 Polysorbate 20 Polymers 0.000 description 2
- 108020004511 Recombinant DNA Proteins 0.000 description 2
- 240000004808 Saccharomyces cerevisiae Species 0.000 description 2
- 235000014680 Saccharomyces cerevisiae Nutrition 0.000 description 2
- 210000004241 Th2 cell Anatomy 0.000 description 2
- 108060008682 Tumor Necrosis Factor Proteins 0.000 description 2
- 108060008683 Tumor Necrosis Factor Receptor Proteins 0.000 description 2
- 239000002671 adjuvant Substances 0.000 description 2
- 230000003321 amplification Effects 0.000 description 2
- 239000005557 antagonist Substances 0.000 description 2
- 230000001363 autoimmune Effects 0.000 description 2
- WQZGKKKJIJFFOK-VFUOTHLCSA-N beta-D-glucose Chemical compound OC[C@H]1O[C@@H](O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-VFUOTHLCSA-N 0.000 description 2
- 239000003124 biologic agent Substances 0.000 description 2
- 210000002798 bone marrow cell Anatomy 0.000 description 2
- 238000007385 chemical modification Methods 0.000 description 2
- 239000003795 chemical substances by application Substances 0.000 description 2
- 230000001684 chronic effect Effects 0.000 description 2
- 230000004540 complement-dependent cytotoxicity Effects 0.000 description 2
- 238000005094 computer simulation Methods 0.000 description 2
- 230000004940 costimulation Effects 0.000 description 2
- 239000012228 culture supernatant Substances 0.000 description 2
- 238000011982 device technology Methods 0.000 description 2
- 238000010790 dilution Methods 0.000 description 2
- 239000012895 dilution Substances 0.000 description 2
- 235000019441 ethanol Nutrition 0.000 description 2
- 238000011156 evaluation Methods 0.000 description 2
- 239000012894 fetal calf serum Substances 0.000 description 2
- 208000024908 graft versus host disease Diseases 0.000 description 2
- 210000002216 heart Anatomy 0.000 description 2
- 238000002649 immunization Methods 0.000 description 2
- 230000005847 immunogenicity Effects 0.000 description 2
- 102000018358 immunoglobulin Human genes 0.000 description 2
- 230000001976 improved effect Effects 0.000 description 2
- 238000001727 in vivo Methods 0.000 description 2
- 238000011534 incubation Methods 0.000 description 2
- 230000004054 inflammatory process Effects 0.000 description 2
- 238000007918 intramuscular administration Methods 0.000 description 2
- 238000001990 intravenous administration Methods 0.000 description 2
- 201000005296 lung carcinoma Diseases 0.000 description 2
- 210000005210 lymphoid organ Anatomy 0.000 description 2
- HQKMJHAJHXVSDF-UHFFFAOYSA-L magnesium stearate Chemical compound [Mg+2].CCCCCCCCCCCCCCCCCC([O-])=O.CCCCCCCCCCCCCCCCCC([O-])=O HQKMJHAJHXVSDF-UHFFFAOYSA-L 0.000 description 2
- 210000004962 mammalian cell Anatomy 0.000 description 2
- 239000000463 material Substances 0.000 description 2
- 230000007246 mechanism Effects 0.000 description 2
- 238000003199 nucleic acid amplification method Methods 0.000 description 2
- 102000039446 nucleic acids Human genes 0.000 description 2
- 108020004707 nucleic acids Proteins 0.000 description 2
- 150000007523 nucleic acids Chemical class 0.000 description 2
- 210000000496 pancreas Anatomy 0.000 description 2
- 230000026731 phosphorylation Effects 0.000 description 2
- 239000013612 plasmid Substances 0.000 description 2
- 235000010486 polyoxyethylene sorbitan monolaurate Nutrition 0.000 description 2
- 239000000256 polyoxyethylene sorbitan monolaurate Substances 0.000 description 2
- 238000002360 preparation method Methods 0.000 description 2
- 238000004393 prognosis Methods 0.000 description 2
- 230000001737 promoting effect Effects 0.000 description 2
- 102000004169 proteins and genes Human genes 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
- 238000011160 research Methods 0.000 description 2
- 238000003757 reverse transcription PCR Methods 0.000 description 2
- 230000028327 secretion Effects 0.000 description 2
- 229910052710 silicon Inorganic materials 0.000 description 2
- 239000010703 silicon Substances 0.000 description 2
- 210000003491 skin Anatomy 0.000 description 2
- 239000006228 supernatant Substances 0.000 description 2
- 230000001629 suppression Effects 0.000 description 2
- 239000000725 suspension Substances 0.000 description 2
- 102000003390 tumor necrosis factor Human genes 0.000 description 2
- 102000003298 tumor necrosis factor receptor Human genes 0.000 description 2
- 210000003556 vascular endothelial cell Anatomy 0.000 description 2
- 230000003442 weekly effect Effects 0.000 description 2
- KIUKXJAPPMFGSW-DNGZLQJQSA-N (2S,3S,4S,5R,6R)-6-[(2S,3R,4R,5S,6R)-3-Acetamido-2-[(2S,3S,4R,5R,6R)-6-[(2R,3R,4R,5S,6R)-3-acetamido-2,5-dihydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-2-carboxy-4,5-dihydroxyoxan-3-yl]oxy-5-hydroxy-6-(hydroxymethyl)oxan-4-yl]oxy-3,4,5-trihydroxyoxane-2-carboxylic acid Chemical compound CC(=O)N[C@H]1[C@H](O)O[C@H](CO)[C@@H](O)[C@@H]1O[C@H]1[C@H](O)[C@@H](O)[C@H](O[C@H]2[C@@H]([C@@H](O[C@H]3[C@@H]([C@@H](O)[C@H](O)[C@H](O3)C(O)=O)O)[C@H](O)[C@@H](CO)O2)NC(C)=O)[C@@H](C(O)=O)O1 KIUKXJAPPMFGSW-DNGZLQJQSA-N 0.000 description 1
- NHBKXEKEPDILRR-UHFFFAOYSA-N 2,3-bis(butanoylsulfanyl)propyl butanoate Chemical compound CCCC(=O)OCC(SC(=O)CCC)CSC(=O)CCC NHBKXEKEPDILRR-UHFFFAOYSA-N 0.000 description 1
- QCVGEOXPDFCNHA-UHFFFAOYSA-N 5,5-dimethyl-2,4-dioxo-1,3-oxazolidine-3-carboxamide Chemical compound CC1(C)OC(=O)N(C(N)=O)C1=O QCVGEOXPDFCNHA-UHFFFAOYSA-N 0.000 description 1
- GUBGYTABKSRVRQ-XLOQQCSPSA-N Alpha-Lactose Chemical compound O[C@@H]1[C@@H](O)[C@@H](O)[C@@H](CO)O[C@H]1O[C@@H]1[C@@H](CO)O[C@H](O)[C@H](O)[C@H]1O GUBGYTABKSRVRQ-XLOQQCSPSA-N 0.000 description 1
- 241000272525 Anas platyrhynchos Species 0.000 description 1
- 241000272814 Anser sp. Species 0.000 description 1
- 241000193830 Bacillus <bacterium> Species 0.000 description 1
- 206010006187 Breast cancer Diseases 0.000 description 1
- 208000026310 Breast neoplasm Diseases 0.000 description 1
- 208000017897 Carcinoma of esophagus Diseases 0.000 description 1
- 241000282693 Cercopithecidae Species 0.000 description 1
- 241000272194 Ciconiiformes Species 0.000 description 1
- 206010009944 Colon cancer Diseases 0.000 description 1
- FBPFZTCFMRRESA-KVTDHHQDSA-N D-Mannitol Chemical compound OC[C@@H](O)[C@@H](O)[C@H](O)[C@H](O)CO FBPFZTCFMRRESA-KVTDHHQDSA-N 0.000 description 1
- 102000053602 DNA Human genes 0.000 description 1
- 238000008157 ELISA kit Methods 0.000 description 1
- 102000002322 Egg Proteins Human genes 0.000 description 1
- 108010000912 Egg Proteins Proteins 0.000 description 1
- 241000588724 Escherichia coli Species 0.000 description 1
- 208000000461 Esophageal Neoplasms Diseases 0.000 description 1
- 241000233866 Fungi Species 0.000 description 1
- 230000035519 G0 Phase Effects 0.000 description 1
- 230000010190 G1 phase Effects 0.000 description 1
- 208000032612 Glial tumor Diseases 0.000 description 1
- 206010018338 Glioma Diseases 0.000 description 1
- 102000006395 Globulins Human genes 0.000 description 1
- 108010044091 Globulins Proteins 0.000 description 1
- 108010017213 Granulocyte-Macrophage Colony-Stimulating Factor Proteins 0.000 description 1
- 102100039620 Granulocyte-macrophage colony-stimulating factor Human genes 0.000 description 1
- 241000282412 Homo Species 0.000 description 1
- 101000834898 Homo sapiens Alpha-synuclein Proteins 0.000 description 1
- 101000652359 Homo sapiens Spermatogenesis-associated protein 2 Proteins 0.000 description 1
- 101000914514 Homo sapiens T-cell-specific surface glycoprotein CD28 Proteins 0.000 description 1
- 101000818543 Homo sapiens Tyrosine-protein kinase ZAP-70 Proteins 0.000 description 1
- 101000617285 Homo sapiens Tyrosine-protein phosphatase non-receptor type 6 Proteins 0.000 description 1
- 102000018071 Immunoglobulin Fc Fragments Human genes 0.000 description 1
- 108010091135 Immunoglobulin Fc Fragments Proteins 0.000 description 1
- 102000008394 Immunoglobulin Fragments Human genes 0.000 description 1
- 108010021625 Immunoglobulin Fragments Proteins 0.000 description 1
- 102000013463 Immunoglobulin Light Chains Human genes 0.000 description 1
- 108010065825 Immunoglobulin Light Chains Proteins 0.000 description 1
- 241000235058 Komagataella pastoris Species 0.000 description 1
- 150000008575 L-amino acids Chemical class 0.000 description 1
- SHZGCJCMOBCMKK-DHVFOXMCSA-N L-fucopyranose Chemical group C[C@@H]1OC(O)[C@@H](O)[C@H](O)[C@@H]1O SHZGCJCMOBCMKK-DHVFOXMCSA-N 0.000 description 1
- GUBGYTABKSRVRQ-QKKXKWKRSA-N Lactose Natural products OC[C@H]1O[C@@H](O[C@H]2[C@H](O)[C@@H](O)C(O)O[C@@H]2CO)[C@H](O)[C@@H](O)[C@H]1O GUBGYTABKSRVRQ-QKKXKWKRSA-N 0.000 description 1
- 206010050551 Lupus-like syndrome Diseases 0.000 description 1
- 241000701076 Macacine alphaherpesvirus 1 Species 0.000 description 1
- 241000124008 Mammalia Species 0.000 description 1
- 229930195725 Mannitol Natural products 0.000 description 1
- 241000699670 Mus sp. Species 0.000 description 1
- 241000409991 Mythimna separata Species 0.000 description 1
- 229930193140 Neomycin Natural products 0.000 description 1
- 244000061176 Nicotiana tabacum Species 0.000 description 1
- 235000002637 Nicotiana tabacum Nutrition 0.000 description 1
- 108091034117 Oligonucleotide Proteins 0.000 description 1
- 241000283973 Oryctolagus cuniculus Species 0.000 description 1
- 208000037581 Persistent Infection Diseases 0.000 description 1
- 108091000080 Phosphotransferase Proteins 0.000 description 1
- 206010035226 Plasma cell myeloma Diseases 0.000 description 1
- 239000002202 Polyethylene glycol Substances 0.000 description 1
- 241000282405 Pongo abelii Species 0.000 description 1
- 101710094000 Programmed cell death 1 ligand 1 Proteins 0.000 description 1
- 102000003923 Protein Kinase C Human genes 0.000 description 1
- 241000588770 Proteus mirabilis Species 0.000 description 1
- 241000700159 Rattus Species 0.000 description 1
- 238000011579 SCID mouse model Methods 0.000 description 1
- 241000607142 Salmonella Species 0.000 description 1
- 241000235347 Schizosaccharomyces pombe Species 0.000 description 1
- 241001282153 Scopelogadus mizolepis Species 0.000 description 1
- 238000012300 Sequence Analysis Methods 0.000 description 1
- 229920002472 Starch Polymers 0.000 description 1
- 208000005718 Stomach Neoplasms Diseases 0.000 description 1
- 241001655322 Streptomycetales Species 0.000 description 1
- 230000006052 T cell proliferation Effects 0.000 description 1
- 108091008874 T cell receptors Proteins 0.000 description 1
- 102000016266 T-Cell Antigen Receptors Human genes 0.000 description 1
- 102100027213 T-cell-specific surface glycoprotein CD28 Human genes 0.000 description 1
- 206010052779 Transplant rejections Diseases 0.000 description 1
- 241000223259 Trichoderma Species 0.000 description 1
- 206010067584 Type 1 diabetes mellitus Diseases 0.000 description 1
- 102100021125 Tyrosine-protein kinase ZAP-70 Human genes 0.000 description 1
- 102100033019 Tyrosine-protein phosphatase non-receptor type 11 Human genes 0.000 description 1
- 101710116241 Tyrosine-protein phosphatase non-receptor type 11 Proteins 0.000 description 1
- 102100021657 Tyrosine-protein phosphatase non-receptor type 6 Human genes 0.000 description 1
- 208000036142 Viral infection Diseases 0.000 description 1
- 239000012042 active reagent Substances 0.000 description 1
- 239000000654 additive Substances 0.000 description 1
- 230000000996 additive effect Effects 0.000 description 1
- 239000000853 adhesive Substances 0.000 description 1
- 230000001070 adhesive effect Effects 0.000 description 1
- 239000000556 agonist Substances 0.000 description 1
- 125000001118 alkylidene group Chemical group 0.000 description 1
- 230000000735 allogeneic effect Effects 0.000 description 1
- 238000004458 analytical method Methods 0.000 description 1
- 238000010171 animal model Methods 0.000 description 1
- 230000006023 anti-tumor response Effects 0.000 description 1
- 230000002155 anti-virotic effect Effects 0.000 description 1
- 230000000890 antigenic effect Effects 0.000 description 1
- 238000003782 apoptosis assay Methods 0.000 description 1
- 239000007864 aqueous solution Substances 0.000 description 1
- 206010003246 arthritis Diseases 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- 201000004984 autoimmune cardiomyopathy Diseases 0.000 description 1
- 230000004888 barrier function Effects 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- 230000015572 biosynthetic process Effects 0.000 description 1
- 201000001531 bladder carcinoma Diseases 0.000 description 1
- 210000004369 blood Anatomy 0.000 description 1
- 239000008280 blood Substances 0.000 description 1
- 239000000872 buffer Substances 0.000 description 1
- 238000004113 cell culture Methods 0.000 description 1
- 230000004663 cell proliferation Effects 0.000 description 1
- 230000036755 cellular response Effects 0.000 description 1
- 238000005119 centrifugation Methods 0.000 description 1
- 208000019065 cervical carcinoma Diseases 0.000 description 1
- 238000006243 chemical reaction Methods 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 208000029742 colonic neoplasm Diseases 0.000 description 1
- 238000002648 combination therapy Methods 0.000 description 1
- 238000004590 computer program Methods 0.000 description 1
- 238000010276 construction Methods 0.000 description 1
- 230000001276 controlling effect Effects 0.000 description 1
- 210000001151 cytotoxic T lymphocyte Anatomy 0.000 description 1
- 230000003013 cytotoxicity Effects 0.000 description 1
- 231100000135 cytotoxicity Toxicity 0.000 description 1
- 230000034994 death Effects 0.000 description 1
- 238000012217 deletion Methods 0.000 description 1
- 230000037430 deletion Effects 0.000 description 1
- 230000001419 dependent effect Effects 0.000 description 1
- 230000030609 dephosphorylation Effects 0.000 description 1
- 238000006209 dephosphorylation reaction Methods 0.000 description 1
- 238000000502 dialysis Methods 0.000 description 1
- 235000014103 egg white Nutrition 0.000 description 1
- 210000000969 egg white Anatomy 0.000 description 1
- 239000003995 emulsifying agent Substances 0.000 description 1
- 201000002491 encephalomyelitis Diseases 0.000 description 1
- 210000002919 epithelial cell Anatomy 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 239000000835 fiber Substances 0.000 description 1
- 238000013467 fragmentation Methods 0.000 description 1
- 238000006062 fragmentation reaction Methods 0.000 description 1
- 206010017758 gastric cancer Diseases 0.000 description 1
- 239000008103 glucose Substances 0.000 description 1
- ZDXPYRJPNDTMRX-UHFFFAOYSA-N glutamine Natural products OC(=O)C(N)CCC(N)=O ZDXPYRJPNDTMRX-UHFFFAOYSA-N 0.000 description 1
- 235000011187 glycerol Nutrition 0.000 description 1
- 230000003394 haemopoietic effect Effects 0.000 description 1
- 201000010536 head and neck cancer Diseases 0.000 description 1
- 208000014829 head and neck neoplasm Diseases 0.000 description 1
- 208000002672 hepatitis B Diseases 0.000 description 1
- 102000048776 human CD274 Human genes 0.000 description 1
- 102000048362 human PDCD1 Human genes 0.000 description 1
- 229920002674 hyaluronan Polymers 0.000 description 1
- 229960003160 hyaluronic acid Drugs 0.000 description 1
- 210000004408 hybridoma Anatomy 0.000 description 1
- WGCNASOHLSPBMP-UHFFFAOYSA-N hydroxyacetaldehyde Natural products OCC=O WGCNASOHLSPBMP-UHFFFAOYSA-N 0.000 description 1
- 238000011502 immune monitoring Methods 0.000 description 1
- 230000006058 immune tolerance Effects 0.000 description 1
- 230000003053 immunization Effects 0.000 description 1
- 230000002766 immunoenhancing effect Effects 0.000 description 1
- 230000002779 inactivation Effects 0.000 description 1
- 230000006698 induction Effects 0.000 description 1
- 208000015181 infectious disease Diseases 0.000 description 1
- 239000003112 inhibitor Substances 0.000 description 1
- 230000015788 innate immune response Effects 0.000 description 1
- 230000003834 intracellular effect Effects 0.000 description 1
- 230000000366 juvenile effect Effects 0.000 description 1
- 210000002510 keratinocyte Anatomy 0.000 description 1
- 210000003734 kidney Anatomy 0.000 description 1
- 239000008101 lactose Substances 0.000 description 1
- 230000002045 lasting effect Effects 0.000 description 1
- 239000002502 liposome Substances 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 210000004185 liver Anatomy 0.000 description 1
- 201000007270 liver cancer Diseases 0.000 description 1
- 208000014018 liver neoplasm Diseases 0.000 description 1
- 210000004072 lung Anatomy 0.000 description 1
- 206010025135 lupus erythematosus Diseases 0.000 description 1
- 210000003563 lymphoid tissue Anatomy 0.000 description 1
- ZLNQQNXFFQJAID-UHFFFAOYSA-L magnesium carbonate Chemical compound [Mg+2].[O-]C([O-])=O ZLNQQNXFFQJAID-UHFFFAOYSA-L 0.000 description 1
- 239000001095 magnesium carbonate Substances 0.000 description 1
- 229910000021 magnesium carbonate Inorganic materials 0.000 description 1
- 235000019359 magnesium stearate Nutrition 0.000 description 1
- 230000036210 malignancy Effects 0.000 description 1
- 239000000594 mannitol Substances 0.000 description 1
- 235000010355 mannitol Nutrition 0.000 description 1
- 238000004519 manufacturing process Methods 0.000 description 1
- 230000035800 maturation Effects 0.000 description 1
- 239000002609 medium Substances 0.000 description 1
- 238000005374 membrane filtration Methods 0.000 description 1
- 238000007799 mixed lymphocyte reaction assay Methods 0.000 description 1
- 238000002156 mixing Methods 0.000 description 1
- 238000000302 molecular modelling Methods 0.000 description 1
- 210000003205 muscle Anatomy 0.000 description 1
- 201000000050 myeloid neoplasm Diseases 0.000 description 1
- 239000013642 negative control Substances 0.000 description 1
- 230000007896 negative regulation of T cell activation Effects 0.000 description 1
- 229960004927 neomycin Drugs 0.000 description 1
- 238000005457 optimization Methods 0.000 description 1
- 210000000056 organ Anatomy 0.000 description 1
- 230000006320 pegylation Effects 0.000 description 1
- 210000005259 peripheral blood Anatomy 0.000 description 1
- 239000011886 peripheral blood Substances 0.000 description 1
- 102000020233 phosphotransferase Human genes 0.000 description 1
- 210000002826 placenta Anatomy 0.000 description 1
- 239000003726 plant lectin Substances 0.000 description 1
- 229920001515 polyalkylene glycol Polymers 0.000 description 1
- 229920001451 polypropylene glycol Polymers 0.000 description 1
- 230000008569 process Effects 0.000 description 1
- 239000000047 product Substances 0.000 description 1
- 230000005522 programmed cell death Effects 0.000 description 1
- 230000035755 proliferation Effects 0.000 description 1
- 238000002818 protein evolution Methods 0.000 description 1
- 229950010131 puromycin Drugs 0.000 description 1
- 230000002441 reversible effect Effects 0.000 description 1
- 206010039073 rheumatoid arthritis Diseases 0.000 description 1
- 238000009938 salting Methods 0.000 description 1
- 238000012772 sequence design Methods 0.000 description 1
- 238000012163 sequencing technique Methods 0.000 description 1
- 210000002966 serum Anatomy 0.000 description 1
- 230000019491 signal transduction Effects 0.000 description 1
- 230000011664 signaling Effects 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 206010041823 squamous cell carcinoma Diseases 0.000 description 1
- 208000017572 squamous cell neoplasm Diseases 0.000 description 1
- 235000019698 starch Nutrition 0.000 description 1
- 239000008107 starch Substances 0.000 description 1
- 210000000130 stem cell Anatomy 0.000 description 1
- 230000004936 stimulating effect Effects 0.000 description 1
- 201000011549 stomach cancer Diseases 0.000 description 1
- 239000000126 substance Substances 0.000 description 1
- 230000008093 supporting effect Effects 0.000 description 1
- 208000024891 symptom Diseases 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 201000000596 systemic lupus erythematosus Diseases 0.000 description 1
- 238000002560 therapeutic procedure Methods 0.000 description 1
- 238000013518 transcription Methods 0.000 description 1
- 230000035897 transcription Effects 0.000 description 1
- 238000001890 transfection Methods 0.000 description 1
- 238000003151 transfection method Methods 0.000 description 1
- 239000012096 transfection reagent Substances 0.000 description 1
- 150000003626 triacylglycerols Chemical class 0.000 description 1
- 210000003171 tumor-infiltrating lymphocyte Anatomy 0.000 description 1
- 241001515965 unidentified phage Species 0.000 description 1
- 238000011144 upstream manufacturing Methods 0.000 description 1
- 208000010570 urinary bladder carcinoma Diseases 0.000 description 1
- 239000013603 viral vector Substances 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
- 239000000080 wetting agent Substances 0.000 description 1
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
- C07K16/2803—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants against the immunoglobulin superfamily
- C07K16/2818—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants against the immunoglobulin superfamily against CD28 or CD152
-
- 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
- A61K39/0005—Vertebrate antigens
- A61K39/0011—Cancer antigens
- A61K39/001148—Regulators of development
- A61K39/00115—Apoptosis related proteins, e.g. survivin or livin
-
- 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
- A61K39/39558—Antibodies; Immunoglobulins; Immune serum, e.g. antilymphocytic serum against materials from animals against tumor tissues, cells, antigens
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K39/00—Medicinal preparations containing antigens or antibodies
- A61K39/46—Cellular immunotherapy
- A61K39/461—Cellular immunotherapy characterised by the cell type used
- A61K39/4611—T-cells, e.g. tumor infiltrating lymphocytes [TIL], lymphokine-activated killer cells [LAK] or regulatory T cells [Treg]
-
- 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
- A61K39/46—Cellular immunotherapy
- A61K39/464—Cellular immunotherapy characterised by the antigen targeted or presented
- A61K39/4643—Vertebrate antigens
- A61K39/4644—Cancer antigens
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
- A61P31/12—Antivirals
- A61P31/14—Antivirals for RNA viruses
- A61P31/18—Antivirals for RNA viruses for HIV
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
- A61P31/12—Antivirals
- A61P31/20—Antivirals for DNA viruses
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
-
- 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
- C07K16/2803—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants against the immunoglobulin superfamily
- C07K16/2809—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants against the immunoglobulin superfamily against the T-cell receptor (TcR)-CD3 complex
-
- 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
-
- 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
- A61K2039/507—Comprising a combination of two or more separate antibodies
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2239/00—Indexing codes associated with cellular immunotherapy of group A61K39/46
- A61K2239/31—Indexing codes associated with cellular immunotherapy of group A61K39/46 characterized by the route of administration
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2239/00—Indexing codes associated with cellular immunotherapy of group A61K39/46
- A61K2239/38—Indexing codes associated with cellular immunotherapy of group A61K39/46 characterised by the dose, timing or administration schedule
-
- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
- A61K2239/00—Indexing codes associated with cellular immunotherapy of group A61K39/46
- A61K2239/46—Indexing codes associated with cellular immunotherapy of group A61K39/46 characterised by the cancer treated
- A61K2239/57—Skin; melanoma
-
- 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
- A61K39/0005—Vertebrate antigens
- A61K39/0011—Cancer antigens
-
- 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
- C07K2317/56—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
- C07K2317/73—Inducing cell death, e.g. apoptosis, necrosis or inhibition of cell proliferation
-
- 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
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07K—PEPTIDES
- C07K2317/00—Immunoglobulins specific features
- C07K2317/90—Immunoglobulins specific features characterized by (pharmaco)kinetic aspects or by stability of the immunoglobulin
- C07K2317/92—Affinity (KD), association rate (Ka), dissociation rate (Kd) or EC50 value
Abstract
本发明提供了一种新的PD‑1抗体或其功能性片段。特别地,本发明还涉及所述抗体在制备用于治疗肿瘤的药物中的用途。
Description
技术领域
本发明提供了一种新的PD-1抗体或其功能性片段。特别地,本发明还涉及所述抗体在制备用于治疗肿瘤的药物中的用途。
背景技术
T细胞共受体信号是紧密调节免疫反应的重要机制。共信号的细胞表面分子(包括共刺激和共抑制)可分成两类主要的家族:免疫球蛋白(Ig)超家族与肿瘤坏死因子(TNF)-肿瘤坏死因子受体(TNFR)超家族。通常T细胞的激活依赖于HLA-I类或II类分子呈递抗原肽。共受体信号则增加或防止这种激活。例如利用激动剂活化CD28或4-1BB,通过独特的共刺激途径,可以提高周围淋巴器官中淋巴细胞的启动和成熟,或增强其在机体的应答效应。免疫激活也可以通过利用拮抗体阻断共抑制信号通路实现,如程序性细胞死亡蛋白1(PD-1)、B7同系物1(B7-H1,也称为PDL1)通路、细胞毒性T淋巴细胞抗原4(CTLA4)、B/T淋巴细胞衰减子(BTLA)等通路。这些共抑制信号通路在调控免疫耐受中发挥重要作用,提供了限制、终止和/或减弱免疫反应的负信号。共刺激受体介导的免疫激活是通过刺激近膜激酶活化并产生磷酸化级联放大,而共抑制受体如CTLA4,PD1和B/T淋巴细胞衰减因子(BTLA)则招募磷酸酶扭转免疫活化诱导的磷酸化反应。免疫调节生物制剂可广泛地用于治疗免疫相关疾病,抑制诸如移植排斥、自身免疫疾病或炎症性疾病等引起的免疫亢进,或刺激癌症、慢性细菌、病毒感染等免疫机能减退的免疫应答。不同于主流的单克隆抗体和重组融合蛋白为基础的疗法-既通过中和或消耗靶抗原或靶标阳性细胞,免疫调节生物制剂主要通过结合并调控宿主免疫细胞表面的信号分子,来调节抗原特异性T细胞受体(TCR)和B细胞受体(BCR)的信号,从而控制淋巴细胞应答的方向和强度。
PD-1基因在T细胞杂交瘤凋亡时发生上调,被命名为程序细胞死亡蛋白1。PD-1(CD279)表达于活化的T细胞、B细胞,以及活化的骨髓细胞(Ishida Y,Agata Y,ShibaharaK,Honjo T.EMBO J.1992,1:3887-3895),也表达在活化的巨噬细胞、DC和单核细胞,但不存在于他们的幼稚细胞(AgataY,et al.Int.Immunol.1996,8:765-772;Said EA,etal.Nature Med.2010,16:452-459)。这些细胞表面PD-1的表达上调可抑制机体获得性免疫和先天免疫反应。PD-1胞内结构域包含两个酪氨酸位点,一个为免疫受体酪氨酸抑制性受体(ITIM),另一个为免疫受体酪氨酸的开关结构单元(ITSM)。ITSM上酪氨酸的磷酸化招募酪氨酸磷酸酶SHP2和/或SHP1。这些磷酸酶可对ZAP70、CD3和PKC去磷酸化,从而削弱T细胞的信号。PD-1主要通过G0/G1期阻滞抑制T细胞和B细胞增殖细胞,并抑制T细胞产生细胞因子。PD-1表达缺失的动物产生各种自身免疫表型,包括自身免疫性心肌病和狼疮样综合征关节炎和肾炎(Nishimura et al.Immunity.1999,H:141-51;Nishimura etal.Science.2001,291:319-22)。此外,PD-1也在自身免疫性脑脊髓炎、全身性红斑狼疮、移植物抗宿主病(GVHD)、I型糖尿病和类风湿性关节炎等中发挥作用(Salama et al.J ExpMed.2003,198:71-78;Prokunina and Alarcon-Riquelme,Hum MoI Genet.1992,13:R143;Nielsen et al.Lupus.2004,11:510)。
目前报道PD-1配体有两个,PD-L1/B7H1/CD274和PD-L2/B7-DC/CD273(FreemanGJ,et al.J.Exp.Med.2000,192:1027-1034;Latchman Y,et al.Nature Immunol.2001,2:261-268)。PD-L1在免疫细胞如B细胞、树突状细胞、巨噬细胞和T细胞上低水平表达,并在细胞活化时表达上调。PD-L1也表达于如血管内皮细胞、心脏、肺、胰脏、肌肉、角质形成细胞和胎盘等非淋巴器官。PD-L1在非淋巴样组织中的表达揭示,PD-L1可能调节自身反应性T细胞、B细胞以及外周组织中骨髓细胞的功能,也可能参与靶器官的炎症反应。PD-L1的表达主要是通过1型和2型干扰素调节,这也是血管内皮细胞和上皮细胞PD-L1水平的主要调节剂。PD-L1也表达于肿瘤细胞中,并与预后不良密切相关。各种病毒感染可诱导宿主组织大量表达PD-L1。尽管PD-L2转录产物在非造血组织如心脏、肝脏和胰脏被发现,但PD-L2/B7-DC在细胞表面的表达仅限于巨噬细胞和树突状细胞,并取决于IFNγ和Th2细胞因子的产生。PD-L1和PD-L2的表达也受到不同刺激的影响。巨噬细胞上的PD-L1由INFγ诱导,而PD-L2则受IL-4调控。类似现象也出现在树突状细胞上。研究揭示PD-L1可能优先调节Th1应答,而PD-L2则调节Th2细胞应答。PD-L1和PD-L2都可抑制T细胞的增殖、细胞因子的产生以及β1/β2整合素介导的粘附作用。PD-L2也可触发树突状细胞反向信号、导致IL-12产生和T细胞活化。
PD-L1-PD-1调节轴在人体T细胞活化控制与机体免疫耐受维持中发挥关键作用,也被肿瘤细胞以及病毒在慢性病毒感染所利用(Yao S,Chen L.Trends Mol.Med.2006,12:244-246;Zou W,Chen L.Nature Rev.Immunol.2008,8:467-477)。PD-L1在多种人体癌症组织中高度表达(Dong et al,Nat.Med.2002,8:787-9)。PD-L1的表达与某些类型恶性肿瘤的进展和预后不良具有相关性(Thompson RH,et al.Cancer Res.2006,66:3381-3385)。PD-L1-PD1通路也已被证实促进T细胞耗竭(Zajac AJ,et al..J.Exp.Med.1998,188:2205-2213)。肿瘤或病毒引起的PD-L1-PD1通路可通过多种机制来实现宿主免疫监视的逃逸,包括促进T细胞失活、疲劳、反应迟钝和细胞凋亡,诱导Treg细胞扩增,以及增强肿瘤内在性抵抗杀伤和凋亡的能力。通过癌细胞介导的PD-1与PD-L1相互作用导致肿瘤浸润淋巴细胞减少,T细胞受体介导的T细胞增殖受到抑制,并增加免疫逃逸(Dong et al.J.Mol.Med.2003,81:281-7;Blank et al.Cancer Immunol.Immunother.2005,54:307-314;Konishi etal.Clin.Cancer Res.2004,10:5094-100)。
迄今为止,还没有令人满意的方法可诱导针对癌症患者有效的免疫应答,特异性阻断PD-L1-PD-1调节轴,提供了在激活抗肿瘤及抗病毒免疫应答的方法,因此,急需开发设计特异性阻断PD-L1-PD-1调节轴的治疗方法,来克服癌症患者或慢性感染的免疫抑制。
发明内容
一般地,本发明提供了一种新的PD-1抗体或其功能性片段。特别地,本发明的抗体是人源化抗体。
在一个方面,本发明提供了能够特异性结合PD-1的抗体或其功能性片段,其中所述抗体包含重链和轻链,其中(i)所述重链包含分别具有如SEQ ID NO:7、8和11或者9、10和11所示的氨基酸序列的H-CDR1、H-CDR2和H-CDR3;和(ii)所述轻链包含分别具有如SEQ IDNO:12、13和14所示的氨基酸序列的L-CDR1、L-CDR2和L-CDR3。
在一个方面,本发明提供了能够特异性结合PD-1的抗体或其功能性片段,其中(i)所述重链包含具有如SEQ ID NO:1、2、4或5所示氨基酸序列的重链可变区;并且(ii)所述轻链包含具有如SEQ ID NO:3或6所示氨基酸序列的轻链可变区。
在另一个方面,本发明提供了能够特异性结合PD-1的抗体或其功能性片段,其中(i)所述重链包含具有如SEQ ID NO:4或5所示氨基酸序列的重链可变区;并且(ii)所述轻链包含具有如SEQ ID NO:6所示氨基酸序列的轻链可变区。
优选地,本发明的抗PD-1抗体选自10F8、15H6、BA08-1和BA08-2。
在又一个方面,本发明提供了分离的多核苷酸,其编码本发明的抗PD-1抗体或其功能性片段。
在又一个方面,本发明提供了分离的多核苷酸的组合,其包括:编码本发明抗PD-1抗体或其功能性片段之轻链的多核苷酸和编码本发明抗PD-1抗体或其功能性片段之重链的多核苷酸。
在又一个方面,本发明提供了表达载体,其包含本发明的多核苷酸或者本发明的多核苷酸的组合,所述多核苷酸与允许其所编码的多肽在宿主细胞或无细胞表达系统中表达的调节序列有效连接。优选地,所述表达载体是病毒载体或非病毒载体。
在又一个方面,本发明提供了药物组合物,其包含本发明的抗PD-1抗体或其功能性片段,以及可药用载体。
在又一个方面,本发明提供了用于在有需要的对象中治疗或预防癌症或感染性疾病的方法,其包括将本发明的抗PD-1抗体或其功能性片段、多核苷酸、多核苷酸组合、表达载体和/或药物组合物施用给所述对象。在一些特定实施方案中,所述对象已接受或待接受抗CD3抗体治疗。
在又一个方面,本发明提供了用于在有需要的对象中增强T细胞免疫应答的方法,其包括将本发明的抗PD-1抗体或其功能性片段、多核苷酸、多核苷酸组合、表达载体和/或药物组合物施用给所述对象。在一些实施方案中,所述增强T细胞免疫应答包括增强T细胞的细胞因子产生,优选地所述细胞因子包括IL-2和/或IFN-γ。在一些优选的实施方案中,所述增强T细胞的细胞因子产生包括抗CD3抗体刺激的T细胞的细胞因子产生。在另一些优选的实施方案中,所述对象是癌症患者,例如PD-L1阳性癌症患者,优选地肺癌和黑素瘤患者。
在又一个方面,本发明提供了用于在有需要的对象中促进T细胞活化的方法,其包括将本发明的抗PD-1抗体或其功能性片段、多核苷酸、多核苷酸组合、表达载体和/或药物组合物施用给所述对象。优选地,所述方法还包括将抗CD3抗体施用给所述对象。
在又一个方面,本发明提供了用于在有需要的对象中消除PD-L1对T细胞活化之抑制的方法,其包括将本发明的抗PD-1抗体或其功能性片段、多核苷酸、多核苷酸组合、表达载体和/或药物组合物施用给所述对象。优选地,所述方法还包括将抗CD3抗体施用给所述对象。
在又一个方面,本发明提供了促进T细胞活化的方法(优选体外的),其包括使本发明的抗PD-1抗体或其功能性片段、多核苷酸、多核苷酸组合、表达载体和/或药物组合物与T细胞相接触。优选地,所述方法还包括使抗CD3抗体与T细胞相接触。
在又一个方面,本发明提供了消除PD-L1对T细胞活化之抑制的方法(优选体外的),其包括使本发明的抗PD-1抗体或其功能性片段、多核苷酸、多核苷酸组合、表达载体和/或药物组合物与T细胞相接触。优选地,所述方法还包括使抗CD3抗体与T细胞相接触。
在又一个方面,本发明的方法还提供了联合治疗,其包括向有需要的对象施用本发明的抗PD-1抗体和抗CD3抗体
在又一个方面,本发明提供了根据本发明的抗PD-1抗体或其功能性片段在制备用于治疗或预防癌症或感染性疾病的药物中的用途。
在又一个方面,本发明提供了根据本发明的抗PD-1抗体或其功能性片段在制备用于增强T细胞免疫应答的药物中的用途。在一些实施方案中,所述增强T细胞免疫应答包括增强T细胞的细胞因子产生,优选地所述细胞因子包括IL-2和/或IFN-γ。在一些优选的实施方案中,所述增强T细胞的细胞因子产生包括抗CD3抗体刺激的T细胞的细胞因子产生。
在一些实施方案中,本发明的抗PD-1抗体或其功能性片段可用于治疗PD-L1阳性癌症和PD-1阴性癌症。在一些特定实施方案中,所述癌症为肺癌或黑素瘤(例如PD-L1阳性肺癌或黑素瘤和/或PD-L1阴性肺癌或黑素瘤),所述感染性疾病为HIV感染或乙型肝炎病毒感染。
在一些特定实施方案中,根据本发明的抗PD-1抗体或其功能性片段阻断PD-1与PD-L1的相互作用和/或PD-1与PD-L2的相互作用。
在一些优选的实施方案中,本发明的抗PD-1抗体或其功能性片段还包含人IgG4或IgG1重链恒定区和人κ轻链恒定区。
本发明还涉及一种筛选和制备如上所述人源化抗体的方法:用人PD-1蛋白免疫BLAC/C小鼠,经流式细胞分离仪筛选高滴度小鼠的抗原特异性B细胞,用RT-PCR方法克隆出抗体重链和轻链可变区基因,再用293细胞表达重组抗体。用蛋白A纯化后,经过亲和力及阻断与PD-L1结合活性的筛选,最终选出具有出人意料的高PD-1亲和力及T细胞活化能力的抗体10F8和15H6。抗体10F8和15H6的重链可变区的氨基酸序列分别如SEQ ID NO:1和2所示;两个抗体含有相同轻链可变区的氨基酸序列,如SEQ ID NO:3所示。抗体10F8的重链CDR(H-CDR1、H-CDR2和H-CDR3)的氨基酸序列分别如SEQ ID NO:7、8和11所示,15H6的重链CDR(H-CDR1、H-CDR2和H-CDR3)的氨基酸序列分别如SEQ ID NO:9、10和11所示。抗体10F8和15H6的轻链CDR(L-CDR1、L-CDR2和L-CDR3)的氨基酸序列分别如SEQ ID NO:12、13和14所示。根据其重链可变区FR1、FR2、FR3和FR4序列对人抗体基因序列库进行比较,找出与重链可变区FR1、FR2、FR3和FR4序列相似的人胚系(germline)抗体可变区相对应序列候选系列。然后经过计算机模拟(in silicon)方法,分析候选系列序列与HLA-DR分子结合的亲和性,选出亲和性最低的框架序列,从而最终确立重链可变区的FR1、FR2、FR3和FR4的人源化序列。在此框架结构的基础上,应用计算机分子模型分析分析出支持CDR构型所需保留的鼠抗体对应框架氨基酸残基。对应10F8的人源化抗体BA08-1和对应15H6的人源化抗体BA08-2的重链可变区氨基酸序列分别如SEQ ID NO:4和5所示。同样分析其鼠抗轻链可变区的序列,根据其轻链可变区FR1、FR2、FR3和FR4序列对人抗体基因序列库(NCBI Ig BLAST)进行比较,找出轻链可变区FR1、FR2、FR3和FR4序列相似的人胚系(germline)抗体可变区相对应候选序列,然后经过计算机(in silicon)分析所述序列与HLA-DR分子结合的亲和性,选出亲和性最低的框架序列,从而最终确立轻链可变区的FR1、FR2、FR3和FR4的人源化序列。在此框架结构的基础上,应用计算机分子摸型分析鼠抗体的可变区立体空间结构,分析出支持CDR构型所需保留的鼠抗体轻链对应框架氨基酸残基。对应10F8和15H6的人源化抗体BA08-1和人源化抗体BA08-2的轻链可变区如SEQ ID NO:6所示。
附图说明:
图1显示抗体BA08-1和BA-08-2以高亲和力结合PD-1蛋白。
图2显示抗体BA08-1和BA-08-2阻断PD-L1与PD-1的结合。
图3显示抗体BA08-1和BA-08-2显著增强了T细胞的IL-2产生,柱从左到右分别显示无抗体阴性对照、阳性对照(单独用抗CD3抗体)、PD-L1对由抗CD3抗体所刺激的IL-2表达的抑制(添加PD-L1和抗CD3抗体)、本发明抗体BA08-1对PD-L1抑制作用的拮抗(添加BA08-1、PD-L1和抗CD3抗体)、本发明抗体BA08-2对PD-L1抑制作用的拮抗(添加BA08-2、PD-L1和抗CD3抗体)、以及MK3475对PD-L1抑制作用的拮抗(添加MK3475、PD-L1和抗CD3抗体),具体如图下的图例所示。
图4显示抗体BA08-1和BA-08-2显著增强了T细胞的IFN-γ产生,柱从左到右分别显示无CD3抗体对照、阳性对照(单独用抗CD3抗体)、PD-L1对由抗CD3抗体所刺激的IFN-γ表达的抑制(添加PD-L1和抗CD3抗体)、本发明抗体BA08-1对PD-L1抑制作用的拮抗(添加BA08-1、PD-L1和抗CD3抗体)、本发明抗体BA08-2对PD-L1抑制作用的拮抗(添加BA08-2、PD-L1和抗CD3抗体)、以及MK3475对PD-L1抑制作用的拮抗(添加MK3475、PD-L1和抗CD3抗体),具体如图下的图例所示。
图5显示抗体BA08-1和BA-08-2对淋巴细胞产生细胞因子的影响。
图6显示抗体BA08-1和BA-08-2对肿瘤细胞(包括黑素瘤细胞和肺癌细胞)抑制活化T细胞产生IL-2的影响,其中PD-L1为固定化PD-L1。
图7显示采用人肺癌的Hu-PBL SCID小鼠评价人源化抗PD-1抗体的体内抗肿瘤效应。
图8显示评价人源化抗PD-1抗体针对黑素瘤的体内抗肿瘤效应。
具体实施方式:
除非另有定义,本文使用的所有科技术语具有本领域普通技术人员所理解的相同含义。关于本领域的定义及术语,专业人员具体可参考Current Protocols in MolecularBiology(Ausubel)。氨基酸残基的缩写是本领域中所用的指代20个常用L-氨基酸之一的标准3字母和/或1字母代码。
本发明提供了能够结合程序性死亡因子1(PD-1)的抗PD-1抗体及其功能性片段。本发明的抗体或其功能性片段具有以下特性的至少一种:能够以高亲和力阻断PD-1和PD-L1的相互作用;能够与PD-1以高特异性结合;激活肿瘤特异性T细胞,从而杀伤肿瘤细胞;显著增强T细胞免疫应答,例如增强T细胞的细胞因子(包括IFNγ和IL-2)产生;等免疫效应物的水平大大增加。
本发明还提供了人源化抗PD-1抗体及其功能性片段。例如,所述人源化抗体由免疫小鼠产生的鼠源抗体经由计算机模拟设计并结合酵母展示技术而得到。
在不实质性影响抗体活性的前提下,本领域技术人员可以对本发明的序列替换、添加和/或缺失一个或更多个(例如1、2、3、4、5、6、7、8、9或10个或更多个)氨基酸,以获得所述抗体或其功能性片段之序列的变体。它们都被视为包括在本发明保护的范围内。如在可变区将具有类似性质的氨基酸进行替换。本发明所述变体的序列可以与其来源序列有至少有95%、96%、97%、98%或99%的一致性。本发明所述的序列一致性可以使用序列分析软件测量。例如使用缺省参数的计算机程序BLAST,尤其是BLASTP或TBLASTN。
本文所使用的术语“抗体”涵盖全长抗体(例如,IgG1或IgG4抗体)、其各种功能性片段(例如可仅包含抗原结合部分,如Fab、F(ab’)2或scFv片段)以及经过修饰的抗体(例如人源化、糖基化等等)。本发明包括具有修饰的糖基化模式的抗PD-1抗体。在一些应用中,进行修饰以除去不期望的糖基化位点可以是有用的,或在寡糖链上不存在岩藻糖部分以例如增强抗体依赖性细胞毒性(ADCC)功能的抗体。在另一些应用中,可进行半乳糖基化修饰以改变补体依赖性细胞毒性(CDC)。
本文所使用的术语“功能性片段”旨在表示保留全长抗体功能的片段,例如抗原结合片段,尤其是指下述抗体片段:如Fv、scFv(sc指单链)、Fab、F(ab’)2、Fab’、scFv-Fc片段或者双抗体(diabody)、或者通过化学修饰或通过掺入脂质体中应能够增加半寿期的任何片段,所述化学修饰例如添加聚(亚烷基)二醇如聚乙二醇(“聚乙二醇化,PEG化”)(被称为Fv-PEG、scFv-PEG、Fab-PEG、F(ab′)2-PEG或Fab′-PEG的聚乙二醇化片段)(“PEG”为聚乙二醇)。
本领域技术人员可以将编码本发明所述抗PD-1抗体的DNA分子克隆到载体中,进而转化宿主细胞。因此,本发明还提供了一种重组DNA载体,其含有编码本发明所述抗PD-1抗体的DNA分子。
优选地,所述重组DNA载体是一种表达载体,本领域技术人员将所述抗体的DNA分子克隆到表达载体中,转化宿主细胞,通过诱导表达获得抗体。本发明的表达载体含有编码的抗PD-1抗体的重链可变区、轻链可变区和/或恒定区的DNA序列。然而,也可有分别构建两种表达载体,一种含有重链可变区和恒定区,另一种含有轻链可变区和恒定区,一同转染哺乳动物。在一个优选的实施方案中,所述表达载体进一步含有启动子和编码分泌信号肽的DNA序列,以及至少一种用于筛选的抗药基因。
本发明所述宿主细胞可以为其为原核宿主细胞、真核宿主细胞或噬菌体。,所述原核宿主细胞可以为大肠杆菌、枯草杆菌、链霉菌或奇异变形菌等。所述真核宿主细胞,可以为如巴斯德毕赤酵母、酿酒酵母、裂殖酵母、木霉等真菌,如草地粘虫等昆虫细胞,如烟草等植物细胞,如BHK细胞、CHO细胞、COS细胞、骨髓瘤细胞等哺乳动物细胞。在一些实施方案中,本发明所述宿主细胞优选为哺乳动物细胞,更优选BHK细胞、CHO细胞、NSO细胞或COS细胞。
本文使用的术语“药物组合物”表示组合在一起以实现某种特定目的的至少一种药物以及任选地可药用载体或辅料的组合。在某些实施方案中,所述药物组合物包括在时间和/或空间上分开的组合,只要其能够共同作用以实现本发明的目的。例如,所述药物组合物中所含的成分(例如根据本发明的抗体、核酸分子、核酸分子组合和/或缀合物)可以以整体施用于对象,或者分开施用于对象。当所述药物组合物中所含的成分分开地施用于对象时,所述成分可以同时或依次施用于对象。优选地,所述可药用载体是水、缓冲水溶液、等渗盐溶液如PBS(磷酸盐缓冲液)、葡萄糖、甘露醇、右旋葡萄糖、乳糖、淀粉、硬脂酸镁、纤维素、碳酸镁、0.3%甘油、透明质酸、乙醇或聚亚烷基二醇如聚丙二醇、甘油三酯等。所用可药用载体的类型尤其依赖于根据本发明的组合物是否配制为用于口服、鼻、皮内、皮下、肌内或静脉施用。根据本发明的组合物可包含润湿剂、乳化剂或缓冲液物质作为添加剂。
根据本发明的药物组合物可通过任何适宜的途径施用,例如可口服、鼻、皮内、皮下、肌内或静脉内施用。
在一个相关方面,本发明提供了为抗PD-1抗体与第二治疗剂的组合的药物组合物。在一个实施方案中,所述第二治疗剂是有利地与抗PD-1抗体组合的任意试剂。可有利地与抗PD-1抗体组合的示例性试剂包括但不限于抑制PD-1活性的其他试剂(包括其他抗体或其抗原结合片段、肽抑制剂、小分子拮抗剂等)和/或干扰PD-1上游或下游信号转导的试剂。优选地,所述第二治疗剂是抗CD3抗体。
本文使用的术语“PD-L1阳性癌症或感染性疾病”旨在表示由PD-1表达所导致或者以PD-1表达为症状/特征的癌症或感染性疾病。所述癌症包括但不限于肺癌,肝癌,卵巢癌,宫颈癌,皮肤癌,膀胱癌,结肠癌,乳腺癌,神经胶质瘤,肾癌,胃癌,食道癌,口腔鳞状细胞癌,头颈癌。所述感染性疾病包括但不限于HIV病毒感染和乙型肝炎病毒感染。
本文使用的“治疗有效量”是指足以显示其对于所施用对象益处的剂量。施用的实际量,以及施用的速率和时间过程会取决于所治疗者的自身情况和严重程度。治疗的处方(例如对剂量的决定等)最终是全科医生及其它医生的责任并依赖其做决定,通常考虑所治疗的疾病、患者个体的情况、递送部位、施用方法以及对于医生来说已知的其它因素。
本文所使用的术语“对象”是指哺乳动物,如人类,但也可以是其它动物,如野生动物(如苍鹭、鹳、鹤等),家畜(如鸭、鹅等)或实验动物(如猩猩、猴子、大鼠、小鼠、兔子、豚鼠等)。
提供了以下实施例以证明并进一步解释本发明的一些优选的实施方式和方面,不应被解释为限制其范围。
实施例
实施例1.小鼠抗PD-1单克隆抗体的产生
用包含PD-1胞外部分的重组融合蛋白PD-1-mFc抗原(25μg)(Sino BiologicalInc,Cat lot:10377-H08H)皮下(SC)免疫6-10周龄的BALB/C小鼠。小鼠首次免疫采用抗原混合弗氏完全佐剂(F5881,Sigma)接种,随后采用抗原混合弗氏不完全佐剂(F5506,Sigma)皮下免疫(共计6次免疫接种,分别于第1、7、14、28、60和64天)。免疫反应通过眼眶静脉丛采血监测。ELISA筛选抗血清,用PBS将纯化的重组PD-1的融合蛋白(Sino Biological Inc,Cat lot:10377-H08H)稀释成1μg/ml,包被微孔板,100μl/孔,4℃孵育过夜。然后用200μl/孔含5%胎牛血清、0.05%吐温20的PBS溶液进行封闭。PD-1免疫小鼠血清经梯度稀释后加入各孔,室温下孵育1小时。用PBS/Tween20溶液洗板后,加入辣根过氧化物酶偶联的山羊抗小鼠IgG多克隆抗体(Jackson Immunoresearch Labs,Cat#:115-035-044)室温孵育1小时。洗板后,用TMB底物(Pierce,Cat#34021,)显色,OD 450检测。根据滴度比较,将抗PD-1免疫球蛋白高滴度的小鼠用于PD-1特异性B细胞分离。通过荧光活化细胞分选(FACS)根据与生物素标记的PD-1的结合来分离各个小鼠B谱系细胞,通过RT-PCR来扩增相应的全长Ig重链(H)可变区和Ig轻链(L)可变区基因转录本。根据制造商的方案,将扩增产物克隆到293表达系统(Life Technology)中。使用蛋白A柱来纯化所产生的单克隆抗体,通过与PD-1的结合以及阻断PD-1和PD-L1相互作用的能力来进一步分析所得的单克隆抗体。根据抗体阻断PD-L1与PD-1结合的能力,10F8和15H6被选作候选克隆进一步用于人源化抗体的开发研究。
实施例2.人源化单克隆抗PD-1受体抗体重链可变区序列的设计
编码10F8、15H6单克隆抗体的重链可变区的氨基酸序列分别如SEQ ID NO:1和SEQID NO:2所示。通过与已知的人胚系免疫球蛋白重链序列进行比较,选出低免疫原性人胚系框架序列,最终确定人源化10F8和15H6重链可利用来自人胚系VH 3-66区段、未确定D区段和人胚系JH4区段。人源化10F8(即BA08-1)和人源化15H6(即BA08-2)重链可变区的氨基酸序列分别为SEQ ID NO:4及SEQ ID NO:5。
表1.本发明抗体的重链CDR(H-CDR)
10F8 | 15H6 | |
H-CDR1 | SEQ ID No:7 | SEQ ID No:9 |
H-CDR2 | SEQ ID No:8 | SEQ ID No:10 |
H-CDR3 | SEQ ID No:11 | SEQ ID No:11 |
实施例3.人源化单克隆抗PD-1受体抗体轻链可变区序列的设计
10F8及15H6轻链可变区的氨基酸序列一致(SEQ ID NO:3)。通过与已知的人胚系免疫球蛋白重链序列进行比较,选出低免疫原性人胚系框架序列,最终确定人源化10F8和15H6轻链可利用来自人胚系VK 3-11区段和JK4区段。人源化10F8(BA08-1)和人源化15H6(BA08-2)共有的轻链可变区的氨基酸序列分别为SEQ ID NO:6。
表2.本发明抗体的轻链CDR(L-CDR)
10F8 | 15H6 | |
L-CDR1 | SEQ ID No:12 | SEQ ID No:12 |
L-CDR2 | SEQ ID No:13 | SEQ ID No:13 |
L-CDR3 | SEQ ID No:14 | SEQ ID No:14 |
实施例4.人源化抗PD-1受体单克隆抗体表达及制备
(人IgG4重链恒定区IgG4 Fc片段序列(J Ellison,J Buxbaum,L Hood-DNA,1981)和轻链恒定区κ片段序列(Hieter,P.A.,Max,E.E.,Seidman,J.G.,Maizel,J.V.Jr.,andLeder,P.Cell.1980;22:197-207)分别由IDT公司(Integrated DNA Technologies,Coralville,Iowa)合成。以pcDNA3为骨架来构建本发明的载体pBA-H4和pBA-Ck,pBA-H4(包含人IgG4重链恒定区IgG4 Fc)和pBA-Ck(包含人轻链恒定区κ片段)载体由Bioabs公司构建,在pBA-H4中VH和CH使用CMV启动子,嘌呤霉素抗性基因使用PGK启动子,在pBA-Ck中VL使用CMV启动子,新霉素抗性基因使用SV40启动子。根据抗体重链可变区序列和轻链可变区蛋白质序列,设计编码重链和轻链可变区的DNA序列,并根据在CHO细胞的优化表达,进一步优化编码重链和轻链可变区的DNA序列,其中编码本发明人源化抗PD-1抗体重链可变区的DNA序列分别如SEQ ID NO:15(BA08-1)和16(BA08-2)所示,编码本发明人源化抗PD-1抗体轻链可变区的DNA序列如SEQ ID NO:17所示。。编码优化的重链和轻链可变区的DNA由IDT公司(Integrated DNA Technologies,Coralville,Iowa)合成。经用HD克隆试剂盒(Clontech Cat#:638910),将所合成的含有BA08-1和BA08-2重链DNA片段直接克隆到NheI酶-线性化的pBA-H4载体上,将所合成的含有BA08-1和BA08-2轻链DNA片段直接克隆到BsiWI酶-线性化的pBA-Ck载体上。转化DH5α细菌,提取质粒并进行测序,测序结果与所设计人源化抗体的DNA编码序列一致。CHO-S细胞(购自Invitrogen)培养条件为1×CD-CHO(购自GIBCO),1×HT(购自GIBCO),8mM谷氨酰胺(购自GIBCO),培养于37℃,8%CO2温箱。抗体BA08-1和BA08-2的重链和轻链质粒共同转染CHO-S细胞,转染方法按照DMRIE-C转染试剂盒(购自Invitrogen)说明书操作。转染后的第三天,细胞培养于上述的培养液并加有500μg/ml G418(购自GIBCO)和12.5μg/ml嘌呤霉素(购自Sigma)进行加压筛选。加压约14天后,挑选出的阳性克隆培养于六孔板中,并用直接ELISA的方法检测抗体表达量。挑选出表达率最高的阳性克隆,大批量培养10天后,离心收集培养上清,采用中蛋白A的亲和层析柱(购自GE)纯化后,透析至PBS,并经过0.22um膜过滤后,用于各项研究。
实施例5.抗体与PD-1结合特异性和相对亲合力
采用基于蛋白质的ELISA法测定本发明抗体与人PD-1的相对结合情况。简言之,用PBS将纯化的重组PD-1融合蛋白(Sino Biological Inc,Cat#:10377-H08H)稀释成1μg/ml,包被微孔板,100μl/孔,4℃孵育过夜。然后用200μl/孔含5%胎牛血清、0.05%吐温20的PBS溶液进行封闭。本发明的抗PD-1抗体、MK3475(本发明人使用本发明的表达载体根据已公开的MK3475序列构建而成的类MK-3475对照,以下简称为MK3475,具体构建方案与本发明抗体类似)和IgG对照经梯度稀释后加入各孔,室温下孵育1小时。用PBS/Tween20溶液洗板后,加入辣根过氧化物酶偶联的山羊抗人IgG多克隆抗体(Jackson Immunoresearch Labs,Cat#:109-035-088)室温孵育1小时。洗板后,用TMB底物(Cat.34021,Pierce)显色,OD 450检测。ELISA结果如图1所示。采用Blitz仪器(Pall life Science)测定的亲和力见表3。
本发明的抗体显示出出人意料的与PD-1的高结合亲和力和结合特异性。
表3.抗PD-1抗体结合动力学
抗体 | ka | kd | KD |
BA08-1 | 1.22E+6 | 3.9E-6 | 4.8E-12 |
BA08-2 | 1.15E+6 | 7.91E-6 | 9.1E-12 |
实施例6.抗PD-1抗体阻断PD-L1配体与PD-1受体结合
通过阻断PD-1结合其配体的方式对本发明的人源化抗PD-1抗体进行测试。具体地,1ug/ml未标记的hPD-L1/Fc(R&D Systems,Cat#156-B7-100)包被96孔板16小时。0.5ug/ml PD-1蛋白预先与不同浓度重组抗PD-1抗体37℃孵育30分钟后加入微孔板反应。结合到包被的PD-L1的PD-1蛋白采用小鼠抗人PD-1抗体(eBioscience,Cat#14-9989-8214)杂交,并进一步采用辣根过氧化物酶偶联的山羊抗小鼠方法检测。洗板后,用TMB底物(Pierce,Cat#34021)显色,OD 450检测。如图2所示,抗PD-1 BA08-1和BA08-2抗体特异性地阻断PD-1与其配体PD-L1(图2)的结合,显著优于MK-3475的阻断效果。因此,本发明的抗体实现了出人意料更高的对PD-L1和PD-1之结合的阻断。
实施例7.抗PD-1抗体对PD-L1介导的人T细胞活性抑制的影响
用Ficoll法从健康供体分离人外周血单核细胞(PBMC)。静息的人外周血T细胞经CD3+T细胞富集柱阴性选择(R&DSystems)获得。采用500ng/mL抗CD3抗体和1ug/ml重组人PD-L1/Fc包被96孔板4℃过夜。每孔加入1ug本发明的抗PD-1抗体或MK3475,并在37℃孵育4小时。加入分离的T细胞RMPI悬液(2×104),收集培养2天后的上清液,分别采用IL-2 ELISA试剂盒(eBioscience Cat# 88-7025-88)和IFN-γELISA试剂盒(R&D Systems)按照使用说明对IL-2和IFN-γ表达情况进行检测。分别如图3和图4所示,柱从左到右分别显示无CD3抗体对照、阳性对照(单独用抗CD3抗体)、PD-L1对由抗CD3抗体所刺激的IL-2或IFN-γ表达的抑制(添加PD-L1和抗CD3抗体)、本发明抗体BA08-1对PD-L1抑制作用的拮抗(添加BA08-1、PD-L1和抗CD3抗体)、本发明抗体BA08-2对PD-L1抑制作用的拮抗(添加BA08-2、PD-L1和抗CD3抗体)、以及MK3475对PD-L1抑制作用的拮抗(添加MK3475、PD-L1和抗CD3抗体)。抗PD-1BA08-1和BA08-2抗体都显著消除了PD-L1对CD3刺激的T细胞产生IL-2和IFN-γ的抑制作用(**p<0.01),而且其免疫增强效果显著高于MK-3475analog(*p<0.05)。
实施例8.抗PD-1抗体对混合淋巴细胞反应中细胞因子表达的影响
应用异源混合淋巴细胞(MLR)反应来检测阻断PD-L1/PD-1途径对效应性淋巴细胞的影响。测定抗PD-1人单克隆抗体存在与否对T细胞分泌IFN-γ的情况。采用CD4+阴性选择试剂盒(MiltenyiBiotech)从PBMC纯化获得人CD4+T细胞。树突状细胞来源于经1000U/mlIL-4和500U/ml GM-CSF(R&D Biosystems)七天培养的纯化单核细胞。每个MLR反应中包含105个纯化的T细胞和104个异源性树突状细胞,总体积200μl。人源化抗PD-1单克隆抗体以不同浓度加入到培养板中,细胞在37℃培养5天后,取100μl培养上清测量细胞因子含量。IL-2的测量采用实施例7中同样的方法进行,结果如图5所示,抗PD-1 BA08-1和BA08-2抗体显著促进T细胞活化,并剂量依赖性地增加IL-2表达,而且其效果显著优于MK-3475。
实施例9.抗PD-1抗体对肿瘤细胞抑制活化T细胞产生IL-2的影响
人黑色素瘤细胞系A2058、人肺癌细胞系HCC827细胞购自ATCC(Manassas,VA)。重组人干扰素(IFN)-γ、佛波醇12-肉豆蔻酸酯13-乙酸酯(PMA)和植物凝集素(PHA)购自Sigma-Aldrich公司(St.Louis,MO)。黑色素瘤和肺癌细胞在完全RPMI 1640培养基中长至80%汇合度后,加入500U/mL重组人IFN-γ处理48小时以上调PD-L1表达,并通过流式细胞仪用小鼠抗人PD-L1抗体(BD Pharmingen,Cat#557924)进行确证。静息的人外周血T细胞经CD3+T细胞富集柱阴性选择(R&D Systems)获得T细胞用1μg/mL PHA和50ng/mL PMA处理过夜,随后加入到1ug/ml重组PD-L1-Fc蛋白(Corning,NY)预包被的96孔板(在图6中标记为PD-L1),或者在3ug/ml抗体(IgG对照、本发明的抗PD-1抗体BA08-1或BA08-2、或者MK3475)存在的条件下,按6∶1比例(肿瘤细胞/T细胞)加入IFN-γ处理后的肿瘤细胞孵育48小时(在图6中标记为A2058和HCC827),或者无任何进一步处理(对照,在图6中标记为PMA+PHA)。收集上清,用ELISA检测IL-2表达情况。如图6所示,固定化的PD-L1可抑制激活的T细胞产生IL-2的能力,抗PD-1 BA08-1和BA08-2抗体显著消除了PD-L1的抑制作用。同样,IFN-γ预处理肿瘤细胞也表现出抑制活化T细胞介导的IL-2表达,其抑制作用也可被抗PD-1 BA08-1和BA08-2抗体所阻断(*p<0.05),而且本发明抗体的效果显著优于MK-3475。
实施例10.本发明抗PD-1抗体用于治疗肺癌的体内实验
为了评价人源化抗PD-1抗体的体内效应,6周龄的NOD-SCID小鼠在实验第1天,皮下接种从组织培养获得的1×107人肺癌HCC827细胞。第8天肿瘤平均100mm3(55-150mm3)开始治疗,每组包含6只动物,均腹腔注射来自同一批次的人PBMC。人PBMC制备方法如上文所述。通过Ficoll法纯化获得PBMC 3小时内,采用腹腔注射接种含1×107人PBMC的RPMI悬液到每只小鼠,并于当天开始腹腔注射抗体,给药剂量5mg/kg,2次/一周,共计4次剂量。每周用游标卡尺对小鼠肿瘤进行2次测量,使用下面公式计算肿瘤体积:体积=(长x宽2)/2。如图7所示,人源化抗PD-1抗体均表现出有效而持久的抗肿瘤活性,而且本发明抗体的效果显著优于MK-3475。抗PD-1 BA08-1和BA08-2抗体在第32天,比对照组相比,其抑瘤率分为84.5%和77.8%(p<0.01),而且比MK3475样品66%抑瘤率有明显提高(p<0.05)。
实施例11.本发明抗PD-1抗体用于治疗黑色素瘤的体内实验
采用6周龄的NOD-SCID小鼠,皮下同时接种从组织培养获得的2×106A375人黑色素瘤细胞与1×106同一肿瘤细胞刺激的人T细胞。肿瘤细胞刺激的人T细胞获取方法如下:用RosetteSep试机盒(Stemcell technologies)从健康人外周血单核细胞中阴性选择T细胞与用2微克丝列霉素C16小时处理的A375人黑色素瘤细胞在含有50U/ml重组IL-2的10%FBS-RPMI培养液中共培养7天后收集备用。每组包含6只动物,当天开始治疗,腹腔注射抗PD-1抗体,3mg/kg,2次/一周,共计4次剂量。每周用游标卡尺对小鼠肿瘤进行2次测量,使用下面公式计算肿瘤体积:体积=(长x宽2)/2。如图8所示,抗PD-1 BA08-1和BA08-2抗体在第35天,比对照组相比,其抑瘤率分为82%和72%(p<0.01),而且比MK3475样品58%抑瘤率有明显提高(p<0.05)。
以上描述地仅是优选实施方案,其只作为示例而不限制实施本发明所必需特征的组合。所提供的标题并不意指限制本发明的多种实施方案。术语例如“包含”、“含”和“包括”不意在限制。此外,除非另有说明,没有数词修饰时包括复数形式,以及“或”、“或者”意指“和/或,,。除非本文另有定义,本文使用的所有技术和科学术语的意思与本领域技术人员通常理解的相同。
本申请中提及的所有公开物和专利通过引用方式并入本文。不脱离本发明的范围和精神,本发明的所描述的方法和组合物的多种修饰和变体对于本领域技术人员是显而易见的。虽然通过具体的优选实施方式描述了本发明,但是应该理解所要求保护的本发明不应该被不适当地局限于这些具体实施方式。事实上,那些对于相关领域技术人员而言显而易见的用于实施本发明的所描述的模式的多种变体意在包括在随附的权利要求的范围内。
Claims (10)
1.能够特异性结合PD-1的抗体,其中所述抗体包含重链和轻链,其中
(i)所述重链包含如SEQ ID NO:7、8和11所示的氨基酸序列的H-CDR1、H-CDR2和H-CDR3,或者如SEQ ID NO:9、10和11所示的氨基酸序列的H-CDR1、H-CDR2和H-CDR3;和
(ii)所述轻链包含如SEQ ID NO:12、13和14所示的氨基酸序列的L-CDR1、L-CDR2和L-CDR3。
2.根据权利要求1所述的抗体,其中
(i)所述重链包含具有如SEQ ID NO:1、2、4或5所示氨基酸序列的重链可变区;并且
(ii)所述轻链包含具有如SEQ ID NO:3或6所示氨基酸序列的轻链可变区。
3.根据权利要求1所述的抗体,其中
(i)所述重链包含具有如SEQ ID NO:4或5所示氨基酸序列的重链可变区;并且
(ii)所述轻链包含具有如SEQ ID NO:6所示氨基酸序列的轻链可变区。
4.分离的多核苷酸,其编码根据前述权利要求任一项所述的抗体。
5.分离的多核苷酸的组合物,其包括:编码根据权利要求1-3中任一项所述抗体之轻链的多核苷酸和编码根据权利要求1-3中任一项所述抗体之重链的多核苷酸。
6.表达载体,其包含根据权利要求4的多核苷酸或者根据权利要求5的多核苷酸的组合物,所述多核苷酸与允许其所编码的多肽在宿主细胞或无细胞表达系统中表达的调节序列有效连接。
7.药物组合物,其包含权利要求1至3中任一项的抗体,以及可药用载体。
8.权利要求1至3中任一项的抗体、权利要求4的多核苷酸、权利要求5的多核苷酸组合物、权利要求6的表达载体或权利要求7的药物组合物在制备治疗或预防癌症的药物中的用途。
9.根据权利要求8的用途,其中所述癌症为肺癌或黑素瘤。
10.权利要求1至3中任一项的抗体、权利要求4的多核苷酸、权利要求5的多核苷酸组合物、权利要求6的表达载体或权利要求7的药物组合物在制备治疗或预防感染性疾病的药物中的用途,所述感染性疾病为HIV感染或乙型肝炎病毒感染。
Priority Applications (10)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN201410369300.7A CN105330740B (zh) | 2014-07-30 | 2014-07-30 | 抗pd-1抗体及其应用 |
US15/328,462 US10414821B2 (en) | 2014-07-30 | 2015-08-28 | Anti-PD-1 antibody and use thereof |
PCT/CN2015/088384 WO2016015685A1 (zh) | 2014-07-30 | 2015-08-28 | 抗pd-1抗体及其应用 |
BR112017000664A BR112017000664A2 (pt) | 2014-07-30 | 2015-08-28 | anticorpo anti-pd-1 e seu uso |
AU2015295946A AU2015295946B2 (en) | 2014-07-30 | 2015-08-28 | Anti-PD-1 antibody and use thereof |
CA2955541A CA2955541C (en) | 2014-07-30 | 2015-08-28 | Anti-pd-1 antibody and use thereof |
EP15826387.1A EP3176180B1 (en) | 2014-07-30 | 2015-08-28 | Anti-pd-1 antibody and use thereof |
KR1020177002113A KR102540601B1 (ko) | 2014-07-30 | 2015-08-28 | 항-pd-1 항체 및 이의 용도 |
RU2017102399A RU2661678C1 (ru) | 2014-07-30 | 2015-08-28 | Антитело против белка запрограммированной смерти клетки 1 (pd-1) и его применение |
JP2017505240A JP6654186B2 (ja) | 2014-07-30 | 2015-08-28 | 抗pd−1抗体およびその用途 |
Applications Claiming Priority (1)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
CN201410369300.7A CN105330740B (zh) | 2014-07-30 | 2014-07-30 | 抗pd-1抗体及其应用 |
Publications (2)
Publication Number | Publication Date |
---|---|
CN105330740A CN105330740A (zh) | 2016-02-17 |
CN105330740B true CN105330740B (zh) | 2018-08-17 |
Family
ID=55216788
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
CN201410369300.7A Active CN105330740B (zh) | 2014-07-30 | 2014-07-30 | 抗pd-1抗体及其应用 |
Country Status (10)
Country | Link |
---|---|
US (1) | US10414821B2 (zh) |
EP (1) | EP3176180B1 (zh) |
JP (1) | JP6654186B2 (zh) |
KR (1) | KR102540601B1 (zh) |
CN (1) | CN105330740B (zh) |
AU (1) | AU2015295946B2 (zh) |
BR (1) | BR112017000664A2 (zh) |
CA (1) | CA2955541C (zh) |
RU (1) | RU2661678C1 (zh) |
WO (1) | WO2016015685A1 (zh) |
Families Citing this family (45)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
RS59480B1 (sr) | 2013-12-12 | 2019-12-31 | Shanghai hengrui pharmaceutical co ltd | Pd-1 antitelo, njegov fragment koji se vezuje na antigen, i njegova medicinska primena |
TWI693232B (zh) | 2014-06-26 | 2020-05-11 | 美商宏觀基因股份有限公司 | 與pd-1和lag-3具有免疫反應性的共價結合的雙抗體和其使用方法 |
MX2017015046A (es) | 2015-05-29 | 2018-05-17 | Agenus Inc | Anticuerpos anti-antigeno 4 del linfocito t citotoxico (ctla-4) y metodos para uso de los mismos. |
TWI773646B (zh) | 2015-06-08 | 2022-08-11 | 美商宏觀基因股份有限公司 | 結合lag-3的分子和其使用方法 |
BR112018000768A2 (pt) | 2015-07-13 | 2018-09-25 | Cytomx Therapeutics Inc | anticorpos anti-pd-1, anticorpos anti-pd-1 ativáveis e métodos de uso dos mesmos |
CA2991628C (en) | 2015-07-16 | 2020-04-07 | Bioxcel Therapeutics, Inc. | A novel approach for treatment of cancer using immunomodulation |
EA201890296A1 (ru) | 2015-07-30 | 2018-08-31 | Макродженикс, Инк. | Pd-1-связывающие молекулы и способы их применения |
CN114605548A (zh) | 2015-09-01 | 2022-06-10 | 艾吉纳斯公司 | 抗-pd-1抗体及其使用方法 |
AU2016332725A1 (en) | 2015-09-29 | 2018-03-22 | Celgene Corporation | PD-1 binding proteins and methods of use thereof |
GEP20217328B (en) | 2015-12-14 | 2021-12-10 | Macrogenics Inc | Bispecific molecules having immunoreactivity with pd-1 and ctla-4, and methods of use thereof |
BR112018074463A2 (pt) | 2016-05-27 | 2019-03-06 | Agenus Inc. | anticorpos anti-tim-3 e métodos de uso dos mesmos. |
KR102531889B1 (ko) | 2016-06-20 | 2023-05-17 | 키맵 리미티드 | 항-pd-l1 및 il-2 사이토카인 |
BR112019002258A2 (pt) * | 2016-08-05 | 2019-05-14 | Y-Biologics Inc. | anticorpo que se liga a pd-1 ou fragmento de ligação ao antígeno do anticorpo, método para produzir o mesmo e composição para prevenir ou tratar câncer |
WO2018026248A1 (ko) * | 2016-08-05 | 2018-02-08 | 주식회사 와이바이오로직스 | 프로그램화된 세포 사멸 단백질(pd-1)에 대한 신규 항체 및 이의 용도 |
EP3515944A4 (en) | 2016-09-19 | 2020-05-06 | Celgene Corporation | METHODS OF TREATING IMMUNE DISORDERS WITH PD-1 BINDING PROTEINS |
JP2019534859A (ja) | 2016-09-19 | 2019-12-05 | セルジーン コーポレイション | Pd−1結合タンパク質を使用して白斑を治療する方法 |
KR102273634B1 (ko) * | 2016-09-21 | 2021-07-07 | 씨스톤 파마슈티컬즈 | 예정 사멸 1(pd-1)에 대한 신규한 단일클론성 항체 |
IL265800B2 (en) | 2016-10-11 | 2023-10-01 | Agenus Inc | Anti-LAG-3 antibodies and methods of using them |
KR102603681B1 (ko) | 2016-12-07 | 2023-11-17 | 아게누스 인코포레이티드 | 항체 및 이의 사용방법 |
LT3551660T (lt) | 2016-12-07 | 2023-12-27 | Agenus Inc. | Antikūnai prieš ctla-4 ir jų naudojimo būdai |
CN107043420B (zh) * | 2016-12-26 | 2018-07-31 | 中国科学院微生物研究所 | 一种抗pd-1抗体及其应用 |
MX2019008346A (es) | 2017-01-13 | 2019-09-09 | Agenus Inc | Receptores de celulas t que se unen a ny-eso-1 y metodos de uso de estos. |
CN108341871A (zh) * | 2017-01-24 | 2018-07-31 | 三生国健药业(上海)股份有限公司 | 抗pd-1单克隆抗体及其制备方法和应用 |
WO2018162944A1 (en) * | 2017-03-04 | 2018-09-13 | Shenzhen Runshin Bioscience | Recombinant antibodies to programmed death 1 (pd-1) and uses therefor |
KR102629972B1 (ko) | 2017-04-13 | 2024-01-29 | 아게누스 인코포레이티드 | 항-cd137 항체 및 이의 사용 방법 |
CA3062061A1 (en) | 2017-05-01 | 2018-11-08 | Agenus Inc. | Anti-tigit antibodies and methods of use thereof |
CN111278854A (zh) | 2017-09-04 | 2020-06-12 | 艾吉纳斯公司 | 与混合谱系白血病(mll)特异性磷酸肽结合的t细胞受体和其使用方法 |
EP3784688A2 (en) | 2018-04-26 | 2021-03-03 | Agenus Inc. | Heat shock protein-binding peptide compositions and methods of use thereof |
EP3790584A4 (en) | 2018-05-07 | 2022-03-30 | Genmab A/S | METHOD OF TREATMENT OF CANCER WITH A COMBINATION OF AN ANTI-PD-1 ANTIBODY AND AN ANTI-TISSUE FACTOR ANTIBODY-DUG CONJUGATE |
EP3808376A4 (en) | 2018-06-17 | 2021-09-01 | L&L Biopharma Co., Ltd. | CLDN18.2 ANTIBODY, BISPECIFIC ANTIBODY, ADC AND CAR, AND APPLICATIONS THEREOF |
EP3819313A4 (en) * | 2018-07-03 | 2021-09-01 | L&L Biopharma Co., Ltd. | BISPECIFIC ANTIBODIES AND ITS USE |
KR102371173B1 (ko) | 2018-12-21 | 2022-03-04 | 오제 이뮈노테라프틱스 | 인간화된 항-인간-pd-1 항체 |
CN111423510B (zh) | 2019-01-10 | 2024-02-06 | 迈威(上海)生物科技股份有限公司 | 重组抗人pd-1抗体及其应用 |
WO2020255011A1 (en) | 2019-06-18 | 2020-12-24 | Janssen Sciences Ireland Unlimited Company | Combination of hepatitis b virus (hbv) vaccines and anti-pd-1 or anti-pd-l1 antibody |
KR20220041079A (ko) | 2019-06-18 | 2022-03-31 | 얀센 사이언시즈 아일랜드 언리미티드 컴퍼니 | B형 간염 바이러스(hbv) 백신 및 항-pd-1 항체의 조합 |
TWI809286B (zh) | 2019-07-05 | 2023-07-21 | 日商小野藥品工業股份有限公司 | 以pd-1/cd3雙特異性蛋白質所進行之血液性癌症治療 |
JP2022547650A (ja) * | 2019-07-09 | 2022-11-15 | カディラ・ヘルスケア・リミテッド | ヒトプログラム細胞死受容体pd-1に対する抗体 |
EP4011918A4 (en) | 2019-08-08 | 2023-08-23 | ONO Pharmaceutical Co., Ltd. | DUAL SPECIFIC PROTEIN |
CR20220076A (es) | 2019-08-30 | 2022-06-24 | Agenus Inc | Anticuerpos anti-cd96 y sus métodos de uso |
CA3200671A1 (en) | 2020-11-17 | 2022-05-27 | Seagen Inc. | Methods of treating cancer with a combination of tucatinib and an anti-pd-1/anti-pd-l1 antibody |
GB202107994D0 (en) | 2021-06-04 | 2021-07-21 | Kymab Ltd | Treatment of cancer |
WO2023285552A1 (en) | 2021-07-13 | 2023-01-19 | BioNTech SE | Multispecific binding agents against cd40 and cd137 in combination therapy for cancer |
WO2023057534A1 (en) | 2021-10-06 | 2023-04-13 | Genmab A/S | Multispecific binding agents against pd-l1 and cd137 in combination |
WO2023083439A1 (en) | 2021-11-09 | 2023-05-19 | BioNTech SE | Tlr7 agonist and combinations for cancer treatment |
WO2023218046A1 (en) | 2022-05-12 | 2023-11-16 | Genmab A/S | Binding agents capable of binding to cd27 in combination therapy |
Family Cites Families (8)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
CN1753912B (zh) * | 2002-12-23 | 2011-11-02 | 惠氏公司 | 抗pd-1抗体及其用途 |
CN105315373B (zh) * | 2005-05-09 | 2018-11-09 | 小野药品工业株式会社 | 程序性死亡-1(pd-1)的人单克隆抗体及使用抗pd-1抗体来治疗癌症的方法 |
KR101562580B1 (ko) * | 2007-06-18 | 2015-10-22 | 머크 샤프 앤 도메 비.브이. | 사람 프로그램된 사멸 수용체 pd-1에 대한 항체 |
EP3192811A1 (en) * | 2009-02-09 | 2017-07-19 | Université d'Aix-Marseille | Pd-1 antibodies and pd-l1 antibodies and uses thereof |
CA2791930A1 (en) * | 2010-03-11 | 2011-09-15 | Kerry Louise Tyson | Pd-1 antibody |
ES2669310T3 (es) * | 2011-04-20 | 2018-05-24 | Medimmune, Llc | Anticuerpos y otras moléculas que se unen con B7-H1 y PD-1 |
CN103242448B (zh) * | 2013-05-27 | 2015-01-14 | 郑州大学 | 一种全人源化抗pd-1单克隆抗体及其制备方法和应用 |
BR112018000768A2 (pt) * | 2015-07-13 | 2018-09-25 | Cytomx Therapeutics Inc | anticorpos anti-pd-1, anticorpos anti-pd-1 ativáveis e métodos de uso dos mesmos |
-
2014
- 2014-07-30 CN CN201410369300.7A patent/CN105330740B/zh active Active
-
2015
- 2015-08-28 US US15/328,462 patent/US10414821B2/en active Active
- 2015-08-28 RU RU2017102399A patent/RU2661678C1/ru active
- 2015-08-28 AU AU2015295946A patent/AU2015295946B2/en active Active
- 2015-08-28 CA CA2955541A patent/CA2955541C/en active Active
- 2015-08-28 WO PCT/CN2015/088384 patent/WO2016015685A1/zh active Application Filing
- 2015-08-28 JP JP2017505240A patent/JP6654186B2/ja active Active
- 2015-08-28 BR BR112017000664A patent/BR112017000664A2/pt active Search and Examination
- 2015-08-28 KR KR1020177002113A patent/KR102540601B1/ko active IP Right Grant
- 2015-08-28 EP EP15826387.1A patent/EP3176180B1/en active Active
Also Published As
Publication number | Publication date |
---|---|
KR20180035728A (ko) | 2018-04-06 |
EP3176180B1 (en) | 2019-10-30 |
KR102540601B1 (ko) | 2023-06-07 |
CA2955541A1 (en) | 2016-02-04 |
US10414821B2 (en) | 2019-09-17 |
EP3176180A4 (en) | 2018-01-03 |
EP3176180A1 (en) | 2017-06-07 |
AU2015295946A1 (en) | 2017-03-16 |
JP6654186B2 (ja) | 2020-02-26 |
US20170210806A1 (en) | 2017-07-27 |
AU2015295946B2 (en) | 2020-06-25 |
CN105330740A (zh) | 2016-02-17 |
JP2018502044A (ja) | 2018-01-25 |
CA2955541C (en) | 2023-06-27 |
WO2016015685A1 (zh) | 2016-02-04 |
BR112017000664A2 (pt) | 2017-11-07 |
RU2661678C1 (ru) | 2018-07-18 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
CN105330740B (zh) | 抗pd-1抗体及其应用 | |
JP7082620B2 (ja) | 抗pd1モノクローナル抗体、その医薬組成物およびその使用 | |
WO2017148424A1 (zh) | 一种pdl-1抗体、其药物组合物及其用途 | |
CN104945508B (zh) | 针对人程序性死亡受体pd-1的抗体 | |
JP2021508676A (ja) | 抗tigit抗体並びに治療剤及び診断剤としてのその使用 | |
KR20090094822A (ko) | 항-notch3 길항제 항체 및 notch3-관련 질환의 예방 및 치료에 있어서 그의 용도 | |
CN110520441A (zh) | 抗TGF-β抗体及其用途 | |
JP7358478B2 (ja) | ヒト化抗pd-1抗体及びこの使用 | |
CN111303285B (zh) | 靶向ox40的抗体及其制备方法和应用 | |
KR20210142638A (ko) | Cd3 항원 결합 단편 및 이의 응용 | |
KR20200092976A (ko) | 항-cd137 항체 및 이의 용도 | |
CN116178545A (zh) | 一种抗人pd-l1人源化抗体或其抗原结合片段及其应用 | |
KR20230169936A (ko) | Garp 단백질 항체 및 이의 적용 | |
KR102275514B1 (ko) | 조절 t 세포 표면 항원의 에피토프 및 이에 특이적으로 결합하는 항체 | |
CN111205371A (zh) | 一种抗淋巴细胞激活基因3的抗体及应用 | |
WO2022258011A1 (zh) | 抗pd-1人源化抗体或其抗原结合片段及其应用 | |
CN115521378B (zh) | Pd-l1抗体及其用途 | |
TWI833242B (zh) | 抗pd-1人源化抗體或其抗原結合片段、編碼其的核酸、包含其的載體、細胞和藥物組合物及其用途 | |
TW202241958A (zh) | 抗pd-l1抗體及其應用 | |
CN116102655A (zh) | 靶向pd-l1/pd-1的抗体及其应用 | |
KR20240038043A (ko) | 약학적 조성물 및 용도 | |
EP4263613A1 (en) | Cd40 binding molecules and uses thereof | |
KR20230126713A (ko) | Cea6 결합 분자 및 이의 사용 | |
CN117257934A (zh) | 药物组合物及其用途 | |
EA042365B1 (ru) | Бифункциональное антитело против ctla4 и против pd-1, его фармацевтическая композиция и их применение |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
C06 | Publication | ||
PB01 | Publication | ||
C10 | Entry into substantive examination | ||
SE01 | Entry into force of request for substantive examination | ||
C41 | Transfer of patent application or patent right or utility model | ||
TA01 | Transfer of patent application right |
Effective date of registration: 20161206 Address after: The town of Red Bay area 519020 in Guangdong province Zhuhai City Shuanglin area north venture Hong Kong Industrial Zone No. 38 C02 on the west side of building 201 room Applicant after: Zhuhai Lizhu Sheet Resistance To Biotechnology Co., Ltd. Address before: 250100 Shandong City, Ji'nan Province, South Hill Road, building two, unit 9, unit 20, 701 Applicant before: Liu Jie |
|
GR01 | Patent grant | ||
GR01 | Patent grant |