CN115925937A - Antibody or fragment thereof and pharmaceutical application thereof - Google Patents
Antibody or fragment thereof and pharmaceutical application thereof Download PDFInfo
- Publication number
- CN115925937A CN115925937A CN202210936286.9A CN202210936286A CN115925937A CN 115925937 A CN115925937 A CN 115925937A CN 202210936286 A CN202210936286 A CN 202210936286A CN 115925937 A CN115925937 A CN 115925937A
- Authority
- CN
- China
- Prior art keywords
- antibody
- fragment
- cancer
- seq
- amino acid
- 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
- 239000012634 fragment Substances 0.000 title claims abstract description 49
- 206010028980 Neoplasm Diseases 0.000 claims abstract description 12
- 125000003275 alpha amino acid group Chemical group 0.000 claims description 22
- 239000013598 vector Substances 0.000 claims description 17
- 241001529936 Murinae Species 0.000 claims description 16
- 108020004707 nucleic acids Proteins 0.000 claims description 16
- 102000039446 nucleic acids Human genes 0.000 claims description 16
- 150000007523 nucleic acids Chemical class 0.000 claims description 16
- 239000000427 antigen Substances 0.000 claims description 14
- 102000036639 antigens Human genes 0.000 claims description 14
- 108091007433 antigens Proteins 0.000 claims description 14
- 239000003814 drug Substances 0.000 claims description 9
- 201000011510 cancer Diseases 0.000 claims description 8
- 239000008194 pharmaceutical composition Substances 0.000 claims description 8
- 208000002154 non-small cell lung carcinoma Diseases 0.000 claims description 5
- 208000029729 tumor suppressor gene on chromosome 11 Diseases 0.000 claims description 5
- 206010005003 Bladder cancer Diseases 0.000 claims description 4
- 206010035226 Plasma cell myeloma Diseases 0.000 claims description 4
- 208000007097 Urinary Bladder Neoplasms Diseases 0.000 claims description 4
- 201000001441 melanoma Diseases 0.000 claims description 4
- 201000005112 urinary bladder cancer Diseases 0.000 claims description 4
- 206010006187 Breast cancer Diseases 0.000 claims description 3
- 208000026310 Breast neoplasm Diseases 0.000 claims description 3
- 208000037845 Cutaneous squamous cell carcinoma Diseases 0.000 claims description 3
- 208000000461 Esophageal Neoplasms Diseases 0.000 claims description 3
- 208000022072 Gallbladder Neoplasms Diseases 0.000 claims description 3
- 208000017604 Hodgkin disease Diseases 0.000 claims description 3
- 208000021519 Hodgkin lymphoma Diseases 0.000 claims description 3
- 208000010747 Hodgkins lymphoma Diseases 0.000 claims description 3
- 208000005016 Intestinal Neoplasms Diseases 0.000 claims description 3
- 208000008839 Kidney Neoplasms Diseases 0.000 claims description 3
- 206010023825 Laryngeal cancer Diseases 0.000 claims description 3
- 206010058467 Lung neoplasm malignant Diseases 0.000 claims description 3
- 206010025323 Lymphomas Diseases 0.000 claims description 3
- 208000003445 Mouth Neoplasms Diseases 0.000 claims description 3
- 208000034578 Multiple myelomas Diseases 0.000 claims description 3
- 208000001894 Nasopharyngeal Neoplasms Diseases 0.000 claims description 3
- 206010061306 Nasopharyngeal cancer Diseases 0.000 claims description 3
- 208000015914 Non-Hodgkin lymphomas Diseases 0.000 claims description 3
- 206010030155 Oesophageal carcinoma Diseases 0.000 claims description 3
- 206010061902 Pancreatic neoplasm Diseases 0.000 claims description 3
- 206010060862 Prostate cancer Diseases 0.000 claims description 3
- 208000000236 Prostatic Neoplasms Diseases 0.000 claims description 3
- 206010038389 Renal cancer Diseases 0.000 claims description 3
- 208000005718 Stomach Neoplasms Diseases 0.000 claims description 3
- 208000024770 Thyroid neoplasm Diseases 0.000 claims description 3
- 201000005188 adrenal gland cancer Diseases 0.000 claims description 3
- 208000024447 adrenal gland neoplasm Diseases 0.000 claims description 3
- 201000009036 biliary tract cancer Diseases 0.000 claims description 3
- 208000020790 biliary tract neoplasm Diseases 0.000 claims description 3
- 201000010099 disease Diseases 0.000 claims description 3
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 claims description 3
- 201000004101 esophageal cancer Diseases 0.000 claims description 3
- 208000024519 eye neoplasm Diseases 0.000 claims description 3
- 201000010175 gallbladder cancer Diseases 0.000 claims description 3
- 206010017758 gastric cancer Diseases 0.000 claims description 3
- 201000002313 intestinal cancer Diseases 0.000 claims description 3
- 201000010982 kidney cancer Diseases 0.000 claims description 3
- 206010023841 laryngeal neoplasm Diseases 0.000 claims description 3
- 208000032839 leukemia Diseases 0.000 claims description 3
- 208000012987 lip and oral cavity carcinoma Diseases 0.000 claims description 3
- 201000007270 liver cancer Diseases 0.000 claims description 3
- 208000014018 liver neoplasm Diseases 0.000 claims description 3
- 201000005202 lung cancer Diseases 0.000 claims description 3
- 208000020816 lung neoplasm Diseases 0.000 claims description 3
- 208000015486 malignant pancreatic neoplasm Diseases 0.000 claims description 3
- 206010061311 nervous system neoplasm Diseases 0.000 claims description 3
- 201000002528 pancreatic cancer Diseases 0.000 claims description 3
- 208000008443 pancreatic carcinoma Diseases 0.000 claims description 3
- 208000028466 reproductive system neoplasm Diseases 0.000 claims description 3
- 201000010106 skin squamous cell carcinoma Diseases 0.000 claims description 3
- 201000011549 stomach cancer Diseases 0.000 claims description 3
- 201000002510 thyroid cancer Diseases 0.000 claims description 3
- 108010003723 Single-Domain Antibodies Proteins 0.000 claims description 2
- 238000004519 manufacturing process Methods 0.000 claims description 2
- 239000000546 pharmaceutical excipient Substances 0.000 claims description 2
- 230000002265 prevention Effects 0.000 claims description 2
- 102000008096 B7-H1 Antigen Human genes 0.000 abstract description 31
- 108010074708 B7-H1 Antigen Proteins 0.000 abstract description 31
- 101001117317 Homo sapiens Programmed cell death 1 ligand 1 Proteins 0.000 abstract description 10
- 102000048776 human CD274 Human genes 0.000 abstract description 8
- 238000002474 experimental method Methods 0.000 abstract description 7
- 230000000694 effects Effects 0.000 abstract description 4
- 238000012216 screening Methods 0.000 abstract description 4
- 238000010494 dissociation reaction Methods 0.000 abstract description 3
- 230000005593 dissociations Effects 0.000 abstract description 3
- 230000005847 immunogenicity Effects 0.000 abstract description 3
- 230000009466 transformation Effects 0.000 abstract description 3
- 230000005764 inhibitory process Effects 0.000 abstract description 2
- 230000003285 pharmacodynamic effect Effects 0.000 abstract description 2
- 210000004027 cell Anatomy 0.000 description 53
- 238000001514 detection method Methods 0.000 description 28
- 238000001943 fluorescence-activated cell sorting Methods 0.000 description 28
- 210000004408 hybridoma Anatomy 0.000 description 18
- 239000012228 culture supernatant Substances 0.000 description 16
- 239000006285 cell suspension Substances 0.000 description 14
- 229960003852 atezolizumab Drugs 0.000 description 9
- 230000000903 blocking effect Effects 0.000 description 8
- 150000001413 amino acids Chemical class 0.000 description 7
- 238000000034 method Methods 0.000 description 7
- 229940079593 drug Drugs 0.000 description 6
- 230000014509 gene expression Effects 0.000 description 6
- 239000006228 supernatant Substances 0.000 description 6
- 238000012360 testing method Methods 0.000 description 6
- 241000699670 Mus sp. Species 0.000 description 5
- 238000012163 sequencing technique Methods 0.000 description 5
- 238000004088 simulation Methods 0.000 description 5
- 238000003556 assay Methods 0.000 description 4
- 239000001963 growth medium Substances 0.000 description 4
- 238000005259 measurement Methods 0.000 description 4
- 108090000623 proteins and genes Proteins 0.000 description 4
- 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 4
- 238000005406 washing Methods 0.000 description 4
- 101000998947 Homo sapiens Immunoglobulin heavy variable 1-46 Proteins 0.000 description 3
- 101001138133 Homo sapiens Immunoglobulin kappa variable 1-5 Proteins 0.000 description 3
- 102100036888 Immunoglobulin heavy variable 1-46 Human genes 0.000 description 3
- 102100020769 Immunoglobulin kappa variable 1-5 Human genes 0.000 description 3
- 208000002030 Merkel cell carcinoma Diseases 0.000 description 3
- 206010029266 Neuroendocrine carcinoma of the skin Diseases 0.000 description 3
- 102100040678 Programmed cell death protein 1 Human genes 0.000 description 3
- 238000004458 analytical method Methods 0.000 description 3
- 238000004364 calculation method Methods 0.000 description 3
- 208000017763 cutaneous neuroendocrine carcinoma Diseases 0.000 description 3
- 239000013604 expression vector Substances 0.000 description 3
- 229940125645 monoclonal antibody drug Drugs 0.000 description 3
- 238000007747 plating Methods 0.000 description 3
- 239000013641 positive control Substances 0.000 description 3
- 210000004881 tumor cell Anatomy 0.000 description 3
- 241000282693 Cercopithecidae Species 0.000 description 2
- 238000002965 ELISA Methods 0.000 description 2
- 101000611936 Homo sapiens Programmed cell death protein 1 Proteins 0.000 description 2
- 102100024216 Programmed cell death 1 ligand 1 Human genes 0.000 description 2
- 101710089372 Programmed cell death protein 1 Proteins 0.000 description 2
- 210000001744 T-lymphocyte Anatomy 0.000 description 2
- 239000002671 adjuvant Substances 0.000 description 2
- 238000004113 cell culture Methods 0.000 description 2
- 239000002299 complementary DNA Substances 0.000 description 2
- 238000012258 culturing Methods 0.000 description 2
- 238000010790 dilution Methods 0.000 description 2
- 239000012895 dilution Substances 0.000 description 2
- 229950009791 durvalumab Drugs 0.000 description 2
- 238000000684 flow cytometry Methods 0.000 description 2
- 230000003993 interaction Effects 0.000 description 2
- 238000012933 kinetic analysis Methods 0.000 description 2
- 239000003446 ligand Substances 0.000 description 2
- 230000001394 metastastic effect Effects 0.000 description 2
- 206010061289 metastatic neoplasm Diseases 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 239000013612 plasmid Substances 0.000 description 2
- BASFCYQUMIYNBI-UHFFFAOYSA-N platinum Chemical compound [Pt] BASFCYQUMIYNBI-UHFFFAOYSA-N 0.000 description 2
- 229950010131 puromycin Drugs 0.000 description 2
- 230000000717 retained effect Effects 0.000 description 2
- 239000000243 solution Substances 0.000 description 2
- 210000004988 splenocyte Anatomy 0.000 description 2
- 238000006467 substitution reaction Methods 0.000 description 2
- 241000894006 Bacteria Species 0.000 description 1
- 239000006144 Dulbecco’s modified Eagle's medium Substances 0.000 description 1
- 208000009129 Ear Neoplasms Diseases 0.000 description 1
- 241000238631 Hexapoda Species 0.000 description 1
- 108060003951 Immunoglobulin Proteins 0.000 description 1
- 206010061218 Inflammation Diseases 0.000 description 1
- 241000124008 Mammalia Species 0.000 description 1
- 241000699666 Mus <mouse, genus> Species 0.000 description 1
- 206010029519 Non-small cell lung cancer stage III Diseases 0.000 description 1
- 108091028043 Nucleic acid sequence Proteins 0.000 description 1
- 229920001213 Polysorbate 20 Polymers 0.000 description 1
- 230000006044 T cell activation Effects 0.000 description 1
- 101710120037 Toxin CcdB Proteins 0.000 description 1
- 238000001042 affinity chromatography Methods 0.000 description 1
- 230000003321 amplification Effects 0.000 description 1
- 210000004102 animal cell Anatomy 0.000 description 1
- 229940125644 antibody drug Drugs 0.000 description 1
- 210000004507 artificial chromosome Anatomy 0.000 description 1
- 230000005784 autoimmunity Effects 0.000 description 1
- 229950002916 avelumab Drugs 0.000 description 1
- 210000003719 b-lymphocyte Anatomy 0.000 description 1
- 230000001580 bacterial effect Effects 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 229960000074 biopharmaceutical Drugs 0.000 description 1
- 230000007910 cell fusion Effects 0.000 description 1
- 238000003163 cell fusion method Methods 0.000 description 1
- 230000010261 cell growth Effects 0.000 description 1
- 239000003153 chemical reaction reagent Substances 0.000 description 1
- 238000002512 chemotherapy Methods 0.000 description 1
- UHZZMRAGKVHANO-UHFFFAOYSA-M chlormequat chloride Chemical compound [Cl-].C[N+](C)(C)CCCl UHZZMRAGKVHANO-UHFFFAOYSA-M 0.000 description 1
- 230000016396 cytokine production Effects 0.000 description 1
- 238000003113 dilution method Methods 0.000 description 1
- 238000001962 electrophoresis Methods 0.000 description 1
- 238000005516 engineering process Methods 0.000 description 1
- 238000001976 enzyme digestion Methods 0.000 description 1
- 210000003527 eukaryotic cell Anatomy 0.000 description 1
- 230000017188 evasion or tolerance of host immune response Effects 0.000 description 1
- 239000013613 expression plasmid Substances 0.000 description 1
- 238000001914 filtration Methods 0.000 description 1
- 230000002538 fungal effect Effects 0.000 description 1
- 108020001507 fusion proteins Proteins 0.000 description 1
- 102000037865 fusion proteins Human genes 0.000 description 1
- 230000003394 haemopoietic effect Effects 0.000 description 1
- 210000003958 hematopoietic stem cell Anatomy 0.000 description 1
- 230000013632 homeostatic process Effects 0.000 description 1
- 102000048362 human PDCD1 Human genes 0.000 description 1
- 230000028993 immune response Effects 0.000 description 1
- 230000006058 immune tolerance Effects 0.000 description 1
- 230000003053 immunization Effects 0.000 description 1
- 238000002649 immunization Methods 0.000 description 1
- 230000002163 immunogen Effects 0.000 description 1
- 102000018358 immunoglobulin Human genes 0.000 description 1
- 238000009169 immunotherapy Methods 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
- 230000004054 inflammatory process Effects 0.000 description 1
- 238000012423 maintenance Methods 0.000 description 1
- 229940126619 mouse monoclonal antibody Drugs 0.000 description 1
- 201000000050 myeloid neoplasm Diseases 0.000 description 1
- 239000013642 negative control Substances 0.000 description 1
- 239000002547 new drug Substances 0.000 description 1
- 210000004967 non-hematopoietic stem cell Anatomy 0.000 description 1
- 238000003199 nucleic acid amplification method Methods 0.000 description 1
- 230000003287 optical effect Effects 0.000 description 1
- 239000000825 pharmaceutical preparation Substances 0.000 description 1
- 229910052697 platinum Inorganic materials 0.000 description 1
- 235000010486 polyoxyethylene sorbitan monolaurate Nutrition 0.000 description 1
- 239000000256 polyoxyethylene sorbitan monolaurate Substances 0.000 description 1
- 238000002360 preparation method Methods 0.000 description 1
- 238000004321 preservation Methods 0.000 description 1
- 239000000047 product Substances 0.000 description 1
- 210000001236 prokaryotic cell Anatomy 0.000 description 1
- 230000009465 prokaryotic expression Effects 0.000 description 1
- 238000002331 protein detection Methods 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 239000002994 raw material Substances 0.000 description 1
- 238000010839 reverse transcription Methods 0.000 description 1
- 238000012552 review Methods 0.000 description 1
- 210000002966 serum Anatomy 0.000 description 1
- 230000035939 shock Effects 0.000 description 1
- 238000002415 sodium dodecyl sulfate polyacrylamide gel electrophoresis Methods 0.000 description 1
- 241000894007 species Species 0.000 description 1
- 210000000952 spleen Anatomy 0.000 description 1
- 230000002459 sustained effect Effects 0.000 description 1
- 230000001225 therapeutic effect Effects 0.000 description 1
- 210000001519 tissue Anatomy 0.000 description 1
- 231100000331 toxic Toxicity 0.000 description 1
- 230000002588 toxic effect Effects 0.000 description 1
- 238000001890 transfection Methods 0.000 description 1
- 206010044412 transitional cell carcinoma Diseases 0.000 description 1
- 102000035160 transmembrane proteins Human genes 0.000 description 1
- 108091005703 transmembrane proteins Proteins 0.000 description 1
- 238000012795 verification Methods 0.000 description 1
Images
Landscapes
- Peptides Or Proteins (AREA)
- Preparation Of Compounds By Using Micro-Organisms (AREA)
Abstract
The present invention relates to an antibody or fragment thereof comprising: the antibody or fragment thereof comprises a heavy chain variable region (VH) comprising the H-CDR1, H-CDR2, H-CDR3 shown in sequence in SEQ ID NO 35, SEQ ID NO 36, SEQ ID NO 37 and a light chain variable region (VL) comprising the L-CDR1, L-CDR2, L-CDR3 shown in sequence in SEQ ID NO 38, SEQ ID NO 39, SEQ ID NO 40. The invention provides an antibody which is combined with PD-L1, especially human PD-L1, with high affinity, and the antibodies screened by the invention are improved in the aspects of affinity and dissociation rate to PD-L1; meanwhile, based on the mouse antibody obtained by initial screening, the immunogenicity of the antibody is reduced by performing humanization transformation to the maximum extent; in addition, pharmacodynamic experiments prove that the antibody provided by the invention has better tumor inhibition effect.
Description
Technical Field
The invention relates to the field of biomedicine, in particular to an antibody or a fragment thereof, and also relates to application of the antibody or the fragment thereof.
Background
PD-L1, the gene encoding it, also known as CD274, is a ligand molecule for the programmed death factor PD1, often expressed on hematopoietic and non-hematopoietic cells such as T-cells and B-cells, as well as on various types of tumor cells. PD-L1 is assigned to type 1 transmembrane proteins, which have immunoglobulin V-like and C-like domains. When PD-L1 interacts as a ligand with its receptor molecule PD-1, it inhibits T cell activation and cytokine production. This interaction is important to prevent autoimmunity by maintaining the homeostasis of the immune response during infection or inflammation of normal tissues. At the same time, this interaction plays a crucial role in the induction and maintenance of immune tolerance to itself, which in the tumor microenvironment usually leads to the inactivation of toxic T cells, thus providing immune escape for the tumor cells. The expression of the gene in tumor cells is considered to be one of the criteria for whether many types of human malignant tumors can use PD-1 or PD-L1 antibody drugs, and the malignant tumors comprise melanoma, non-small cell lung cancer and the like.
Several PD-L1 monoclonal antibody drugs are currently on the market, such as the first PD-L1 monoclonal antibody drug Atezolizumab (trade name Tecntriq) developed and produced by GENECECH under Roche, and used for treating advanced bladder cancer with failed or advanced platinum chemotherapy. In 2017, the U.S. Food and Drug Administration (FDA) accelerated the approval of a new drug Bavencio for the treatment of Merkel Cell Carcinoma (MCC) in adults and 12 year old children. The medicine mainly contains Aveluab, and has sustained remission effect on metastatic Merkel cell carcinoma (mMCC). In europe, the approval application for PD-L1 antibody biologics for the treatment of metastatic urothelial cancer is also under FDA priority review. Astrikon, 3 months 2018, also published that its immunotherapy drug, imfinzi (Durvalumab), has formally received us Food and Drug Administration (FDA) approval for the treatment of non-small cell lung cancer stage III (NSCLC) patients who cannot be surgically resected.
At present, the monoclonal antibody drug aiming at PDL1 has a space for improving affinity and dissociation speed, and clinical test data shows that the immunogenicity of the Atezolizumab drug is higher, so that more anti-antibody is generated, and the treatment effect of the drug is greatly influenced.
Disclosure of Invention
Therefore, the invention provides an antibody or a fragment thereof, and the antibody or the fragment thereof can be used for obtaining the antibody which specifically binds to the human PD-L1, particularly has high affinity to the human PD-L1 through hybridoma screening and humanization technology. In view of the above technical problems, it is an object of the present invention to provide an anti-PD-L1 antibody or a fragment thereof, and uses thereof based on the antibody or the fragment thereof. "fragments" of the antibody molecules of the invention encompass various functional fragments of an antibody, for example, antigen-binding portions thereof, such as Fab, F (ab') 2 or scFv fragments.
To achieve the above object, the present invention provides an antibody or a fragment thereof, comprising:
the antibody or fragment thereof comprises a heavy chain variable region (VH) comprising the H-CDR1, H-CDR2, H-CDR3 shown in sequence in SEQ ID NO 35, SEQ ID NO 36, SEQ ID NO 37 and a light chain variable region (VL) comprising the L-CDR1, L-CDR2, L-CDR3 shown in sequence in SEQ ID NO 38, SEQ ID NO 39, SEQ ID NO 40.
Further, the heavy chain variable region comprises a sequence selected from: an amino acid sequence shown in SEQ ID NO. 9 or an amino acid sequence having at least 75% identity to the amino acid sequence shown in SEQ ID NO. 9; and/or, the light chain variable region comprises a sequence selected from: 10 or an amino acid sequence having at least 75% identity to the amino acid sequence shown in SEQ ID No. 10.
Further, the antibody or fragment thereof is an anti-PD-L1 antibody or fragment thereof.
Further, the antibody includes any form of monoclonal antibody, single chain antibody, diabody, single domain antibody, nanobody, fully or partially humanized antibody, and chimeric antibody, or the antigen binding fragment is a half-antibody or an antigen binding fragment of an antibody or a half-antibody, and the antibody of the antigen binding fragment includes scFv, bsFv, dsFv, (dsFv) 2 、Fab、Fab'、F(ab') 2 Fv, and IgG.
Further, the antibody or fragment thereof further comprises a constant region of human or murine origin comprising a heavy chain constant region (CH) of human or murine origin and/or a light chain constant region (CL), wherein the heavy chain constant region comprises an antibody or fragment thereof of IgG, igA, igM, igD or IgE and the light chain constant region comprises a kappa or lambda type.
Further, the antibody comprises a monoclonal antibody which is a monoclonal antibody, a mouse monoclonal antibody, a chimeric monoclonal antibody or a humanized monoclonal antibody, wherein the heavy chain constant region of the monoclonal antibody is of an IgG1 or IgG4 subtype, and the light chain constant region of the monoclonal antibody is of a kappa type;
the heavy chain constant region of the monoclonal antibody comprises the amino acid sequence shown in SEQ ID NO. 1 or an amino acid sequence with at least 75% identity to the amino acid sequence shown in SEQ ID NO. 1;
the light chain constant region of the monoclonal antibody comprises the amino acid sequence set forth in SEQ ID NO. 2 or an amino acid sequence having at least 75% identity to the amino acid sequence set forth in SEQ ID NO. 2.
The nucleic acid molecules of the invention may be cloned into vectors for transformation or transfection of host cells. Thus in a further aspect, the invention also provides a vector comprising a nucleic acid molecule of the invention. The vector can be a eukaryotic expression vector, a prokaryotic expression vector, an artificial chromosome, a phage vector and the like. The vectors or nucleic acid molecules of the invention may be used to transform or transfect host cells for purposes such as preservation or antibody expression. Thus, in a further aspect, the present invention provides a host cell comprising or transformed or transfected with a nucleic acid molecule and/or vector of the invention. The host cell may be any prokaryotic or eukaryotic cell, such as a bacterial or insect, fungal, plant or animal cell.
The antibodies or fragments thereof provided by the present invention can be obtained using any method known in the art. For example, the heavy chain variable region and/or the light chain variable region of the antibody may be obtained from the nucleic acid molecule provided by the present invention, or the heavy chain and/or the light chain of the antibody may be obtained and then assembled into an antibody with optional other domains of the antibody; alternatively, the host cell provided by the present invention is cultured under conditions that allow the host cell to express the heavy chain variable region and/or the light chain variable region of the antibody or the heavy chain and/or the light chain of the antibody to assemble into the antibody. Optionally, the method further comprises the step of recovering the produced antibody.
The antibodies or fragments thereof, nucleic acid molecules, vectors and/or host cells provided by the present invention may be comprised in a pharmaceutical composition, more particularly in a pharmaceutical preparation, for use for various purposes according to the actual needs. Thus, in a further aspect, the invention also provides a pharmaceutical composition comprising an antibody or fragment thereof, a nucleic acid molecule, a vector and/or a host cell according to the invention, and optionally a pharmaceutically acceptable excipient.
In yet another aspect, the present invention also provides the use of the antibody or fragment thereof, nucleic acid molecule, vector, host cell and/or pharmaceutical composition for the manufacture of a medicament for the prevention or treatment of a disease including a tumor or cancer, such as non-small cell lung cancer, melanoma, bladder cancer, merkel lymphoma, cutaneous squamous cell carcinoma, lung cancer, hodgkin lymphoma, kidney cancer, liver cancer, esophageal cancer, non-hodgkin lymphoma, breast cancer, thyroid cancer, stomach cancer, intestinal cancer, nasopharyngeal cancer, pancreatic cancer, prostate cancer, leukemia, laryngeal cancer, oral cancer, ear-eye tumor, biliary tract cancer, gall bladder cancer, adrenal cancer, reproductive system tumor, multiple myeloma, nervous system tumor, urothelial cell cancer.
Accordingly, the present invention also provides a method of preventing or treating a disease comprising a tumor or cancer, such as non-small cell lung cancer, melanoma, bladder cancer, mercker's lymphoma, cutaneous squamous cell carcinoma, lung cancer, hodgkin's lymphoma, kidney cancer, liver cancer, esophageal cancer, non-hodgkin's lymphoma, breast cancer, thyroid cancer, stomach cancer, intestinal cancer, nasopharyngeal cancer, pancreatic cancer, prostate cancer, leukemia, laryngeal cancer, oral cancer, ear and eye tumor, biliary tract cancer, gallbladder cancer, adrenal cancer, reproductive system tumor, multiple myeloma, nervous system tumor, urothelial cell cancer, comprising administering the antibody or fragment thereof, nucleic acid molecule, vector, host cell and/or pharmaceutical composition of the invention to a subject in need thereof. Preferably, the subject is a mammal; more preferably, the subject is a human.
In yet another aspect, the invention provides a kit comprising an antibody or fragment thereof, a nucleic acid molecule, a vector, a host cell and/or a pharmaceutical composition of the invention. The kit may be for therapeutic or diagnostic use.
Compared with the prior art, the invention has the beneficial effects that the invention provides a novel antibody capable of binding PD-L1, especially human PD-L1 with high affinity. Compared with the existing anti-PD-L1 antibody, the antibody has the following characteristics: compared with the existing antibody (taking Atezolizumab (Tecnriq) of Genetech as an example), the antibody screened by the method is improved in the aspects of affinity and dissociation rate to PD-L1; meanwhile, based on the mouse antibody obtained by initial screening, the immunogenicity of the antibody is reduced by performing humanization transformation to the maximum extent; in addition, pharmacodynamic experiments prove that the antibody provided by the invention has better tumor inhibition effect.
Drawings
FIG. 1 shows the results of FACS detection of the binding of the culture supernatant of hybridoma cell lines of the present invention to antigen human PD-L1;
FIG. 2 shows the FACS detection result of the binding of the culture supernatant of the hybridoma cell line of the present invention to the antigen monkey PD-L1;
FIG. 3 shows the result of FACS detection of blocking the binding of PD-L1 to cells by culture supernatant of hybridoma cells according to an embodiment of the present invention;
FIG. 4 shows the results of FACS detection of the binding of the humanized antibody to the antigen human PD-L1 according to the embodiment of the present invention;
FIG. 5 shows the results of FACS detection of the binding of the humanized antibody of the present invention to the antigen monkey PD-L1;
FIG. 6 shows the results of FACS detection of the humanized antibody of the present invention blocking the binding of PD-L1 to cells;
wherein, the abscissa in fig. 1-6 is the parameter after the logarithm of the concentration is taken.
Detailed Description
In order that the objects and advantages of the invention will be more clearly understood, the invention is further described below with reference to examples; it should be understood that the specific embodiments described herein are merely illustrative of the invention and are not intended to limit the invention.
Preferred embodiments of the present invention are described below with reference to the accompanying drawings. It should be understood by those skilled in the art that these embodiments are only for explaining the technical principle of the present invention, and do not limit the scope of the present invention.
It should be noted that in the description of the present invention, the terms of direction or positional relationship indicated by the terms "upper", "lower", "left", "right", "inner", "outer", etc. are based on the directions or positional relationships shown in the drawings, which are only for convenience of description, and do not indicate or imply that the device or element must have a specific orientation, be constructed in a specific orientation, and be operated, and thus, should not be construed as limiting the present invention.
Furthermore, it should be noted that, in the description of the present invention, unless otherwise explicitly specified or limited, the terms "mounted," "connected," and "connected" are to be construed broadly, and may be, for example, fixedly connected, detachably connected, or integrally connected; can be mechanically or electrically connected; they may be connected directly or indirectly through intervening media, or they may be interconnected between two elements. The specific meanings of the above terms in the present invention can be understood by those skilled in the art according to specific situations.
The experimental procedures in the following examples are all conventional ones unless otherwise specified. The raw materials and reagents used in the following examples are all commercially available products unless otherwise specified.
Atezolizumab (Reference): manufactured by Genetech under the trade name Tecntriq, disclosed in US2016/0319022A1, and see SEQ ID NO 6 and SEQ ID NO 7 for the light and heavy chains, respectively.
EXAMPLE 1 preparation of hybridoma cells according to the examples of the invention
The PD-L1 protein (KP 1008 PDL1-mFc kyinno) is used as immunogen to immunize mice, 5 mice are selected to immunize subcutaneously, 5 mice are immunized intramuscularly, the adjuvant adopts quinkanody 5W water-soluble adjuvant, the titer is measured 2 weeks after the boosting immunization, two mice with high titer are selected to carry out immune shock, and cell fusion is carried out after 3 days.
The cell fusion method specifically comprises the steps of taking serum of 2 mice to be fused, dissecting and taking spleens, separating splenocytes, fusing the splenocytes with cultured myeloma cells, laying the fused cells on a 96-well plate, adding a selective culture medium for screening, changing the culture medium after 7 days, performing ELISA detection after 10 days, and selecting cells with OD (optical density) values more than 10 times of that of a negative control for flow cytometry detection.
Selecting double positive cells, performing subclone plating by a cell limiting dilution method, and selecting monoclonal cells. And (3) taking the selected monoclonal cells, taking culture supernatant, performing ELISA detection and flow cytometry detection, and selecting the cells of double positive for amplification culture.
EXAMPLE 2 FACS detection of binding of PD-L1 to culture supernatants of hybridoma cells of examples of the present invention
293T-PDL1 (KC-0205 293T-PDL1 kyinno) cells were prepared into a cell suspension at a cell concentration of 1X 107 cells/ml in PBS containing 2% FBS.
50 μ l of cell suspension was added to each flow tube (sample tube), while the antibody to be detected (purified hybridoma antibody) and a blank (PBS) were added to each well, starting at 20ug/mL, diluted 3.16-fold, diluted 8 doses-graded, and incubated at 4 ℃ for 60 minutes. To each flow tube, 1ml of flow buffer was added, centrifuged at 1200rpm for 5 minutes, the supernatant was discarded, and the washing was repeated three times. Control tubes 1 (without addition of culture supernatant and secondary antibody below, only cell suspension) and 2 (without addition of culture supernatant, only cell suspension and secondary antibody below) were set simultaneously.
Then 100. Mu.l of flow buffer was added to each flow tube for resuspension, 5. Mu.l of PE-labeled anti-mouse Fc-labeled secondary antibody (Biolegend, cat. No. 409304) was added according to the experimental requirements, incubated at 4 ℃ for 30 minutes in the dark, then 1ml of flow buffer was added, centrifuged at 1200rpm for 5 minutes at room temperature, the supernatant was discarded, and the washing was repeated three times.
Add 250. Mu.l of flow buffer to each flow tube again, resuspend and mix well, and test on the machine.
The results of the FACS binding measurements are shown in table 1 and figure 1.
TABLE 1 FACS detection of the binding of culture supernatants of hybridoma cells to PD-L1
| Numbering | Name (R) | EC50(μg/mL) | Fluorescence intensity max |
| PL-29 | 26G9-5 | 0.2189 | 12113 |
| PL-73 | 54F9-4 | 0.3098 | 10474 |
| PL-80 | 60A12-1 | 0.3784 | 12215 |
| PL-82 | 48B10-6 | 0.4460 | 10728 |
EXAMPLE 3 FACS detection of binding of culture supernatants of hybridoma cells of the examples of the present invention to cynoPD-L1
293T-cynoPDL1 (KC-1001, kyinno) cells were prepared into a cell suspension at a cell concentration of 1X 107 cells/ml in PBS containing 2% FBS.
To each flow tube (sample tube) was placed 50 μ l of cell suspension, while to each well was added the antibody to be detected (purified hybridoma antibody) and a blank control (PBS), starting at 20ug/mL, diluted 3.16 fold, diluted 8doses gradient, and incubated at 4 ℃ for 60 minutes. To each flow tube, 1ml of flow buffer was added, centrifuged at 1200rpm for 5 minutes, the supernatant was discarded, and the washing was repeated three times. Control tubes 1 (without addition of culture supernatant and secondary antibody below, only cell suspension) and 2 (without addition of culture supernatant, only cell suspension and secondary antibody below) were set simultaneously.
Then 100. Mu.l of flow buffer was added to each flow tube for resuspension, 5. Mu.l of PE-labeled anti-mouse Fc-labeled secondary antibody (Biolegend, cat. No. 409304) was added according to the experimental requirements, incubated at 4 ℃ for 30 minutes in the dark, then 1ml of flow buffer was added, centrifuged at 1200rpm for 5 minutes at room temperature, the supernatant was discarded, and the washing was repeated three times.
Add 250. Mu.l of flow buffer to each flow tube again, resuspend and mix well, and test on the machine.
The results of the FACS binding measurements are shown in table 2 and figure 2.
TABLE 2 FACS detection of hybridoma cell culture supernatants associated with cynoPD-L1
| Numbering | Name (R) | EC50(μg/mL) | Fluorescence intensity max |
| PL-29 | 26G9-5 | 0.3073 | 15356 |
| PL-73 | 54F9-4 | 0.3636 | 12176 |
| PL-80 | 60A12-1 | 0.3710 | 12581 |
| PL-82 | 48B10-6 | 0.2974 | 5278 |
EXAMPLE 4 FACS detection of blocking binding of PD-L1 to cells by culture supernatants of hybridoma cells according to the examples of the invention
An extracellular region (from position 24 to position 170 of SEQ ID NO: 8) gene fragment of human PD-1 protein (SEQ ID NO: 8) was synthesized, constructed into PLVX-IRES-PURO (Clontech), transformed into DH 5. Alpha. Competent cells, plated, and cultured overnight at 37 ℃. Selecting a monoclonal colony, shaking bacteria, carrying out enzyme digestion identification, extracting plasmids, selecting correct clones to extract plasmids through sequencing verification, and storing for later use.
Recovering 293T cells, continuously passaging for 2 times, plating according to 2 multiplied by 106 cells/10 ml/dish, after 16-24 h, when the cell density grows to 70-80%, transfecting the cells with the eukaryotic expression plasmid, and culturing in a 5% CO2 incubator at 37 ℃. Culturing for 12-16 h, removing the culture medium, changing the culture medium, and adding a corresponding amount of Puromycin (the final concentration is 10 mug/ml) after 48-56 h. The cells were cultured continuously for 10 days in DMEM + 10. Mu.g/ml Puromycin containing 10% FBS, if the cell growth state was good. And (5) initially judging to take the polyclonal cell and carrying out flow detection.
Taking positive polyclonal cells, plating each well according to 0.5 cell, picking out monoclonal cells after 7 days, and carrying out expanded culture. And performing FACS detection on the selected monoclonal, selecting positive cells, namely 293T cells stably expressing h-PD1, and naming the cells as the 293T-h-PD1 cells.
The cells were washed once with FACS buffer, plated in 96-deep well plates at 2-5 × 105 cells/well, while the antibody to be tested (hybridoma antibody after purification) and a blank control (PBS) were added to each well, and incubated for 2 hours starting at 20ug/mL, 3.16 fold dilution, 8doses gradient dilution, and 500ng of a pre-mix of the fusion protein PD-L1 (hFC tag) of example 2. Then, 400. Mu.l of FACS buffer was added to each well and washed twice, a PE anti-human IgG Fc secondary antibody was added thereto and incubated for 1 hour, and then 400. Mu.l of FACS buffer was added to each well and washed twice. And (6) performing detection on the machine.
The results of the FACS blockade detection are shown in table 3 and fig. 3.
TABLE 3 FACS detection of hybridoma cell culture supernatants for blocking the binding of PD-L1 to the cells
| Numbering | Name (R) | EC50(μg/mL) | Max/% of blocking rate |
| PL-29 | 26G9-5 | 0.3933 | 99.586 |
| PL-73 | 54F9-4 | 0.4892 | 99.205 |
| PL-80 | 60A12-1 | 0.4251 | 99.615 |
| PL-82 | 48B10-6 | 0.2519 | 99.764 |
Example 5 affinity assay for binding of culture supernatants of hybridoma cells of the examples of the invention to PD-L1
The experimental method comprises the following steps:
And 3, fixing the antibody, wherein the antibody concentration of the purified supernatant adopts 10 mu g/ml.
Step 4. Extracellular regions of human PD-L1 protein (position 19 to position 238 of SEQ ID NO: 3) were diluted with 6 gradients of 200nM, 100nM, 50nM, 25nM, 12.5nM, 6.25nM, respectively, and added to the corresponding wells.
And 5, detecting on the computer. The results are shown in Table 4.
TABLE 4 affinity assay of the binding of culture supernatants of hybridoma cells to PD-L1
And selecting corresponding hybridoma cell strains based on the experiment, extracting RNA from the monoclonal cells, performing reverse transcription to form cDNA, and connecting the cDNA into a sequencing vector for sequencing analysis.
The sequences of exemplary murine antibodies are as follows, with the CDRs (obtained according to the AbM definition method) underlined. The nomenclature of the murine antibody depends on the hybridoma cell line from which it is derived.
A1 (murine antibody 48B 10)
>48B10-VH
QVQLKESGGGLVKPGGSLKLSCAASGFTFSSYAMSWIRQTPEKRLEWVATITSDGRYSYYPDSVKGRFTISRDNAKNTLYLQMSSLRSEDTATYYCARRENWFTYWGQGTLVTVSA
>48B10-VL
DIVITQSPALMSASPGEKVTMTCSASSSVTYMYWYQQKPRSSPKPWIYLTSNLASGVPARFSGSGSGTSFSLTISNMEAEDAATYYCQQWSSGPTFGAGTKLELK
Example 6 humanization of antibodies of the present examples
The A1 antibody (murine antibody 48B 10) was selected for humanization of the antibody. Comprehensively considers the sequence similarity, expression quantity and whether the light and heavy chain combination of the antibody and different humanized templates is used by the antibody which is already prepared, and the like. According to the factors that the similarity value of the light chain and the heavy chain is more than 200, the expression amount is more than 50mg/ml, the antibody of the light chain and the heavy chain is used in combination with the drug and the like, finally, the humanized sequences are respectively selected as templates of the heavy chain and the light chain: IGHV1-46 x 01 (SEQ ID NO: 4) and IGKV1-5 x 01 (SEQ ID NO: 5). And carrying out homologous modeling on the A1 murine monoclonal antibody, and carrying out structural simulation of the Fab region. And finally obtaining the predicted Fab structure of the A1 antibody through homologous modeling calculation.
Through the comparison analysis of the predicted Fab structure and heavy chain with IGHV1-46 x 01 sequence, all CDR regions are replaced by human-derived template, and then structure simulation and kinetic calculation are carried out. Through structural simulation and kinetic analysis, several amino acids affecting the CDR structure are retained, and the final heavy chain version except the CDR region and 68A, 70L, etc. are the original murine amino acids and the other murine amino acids are all replaced with corresponding IGHV1-46 x 01 template human amino acids.
Through the comparison analysis of the predicted Fab structure and light chain with IGKV1-5 x 01 sequence, CDR area is replaced with adult template, and then structure simulation and kinetic calculation are carried out. Through structural simulation and kinetic analysis, several amino acids affecting the CDR structure are retained, and the final light chain version except the CDR area retains the original murine amino acids is replaced with the corresponding IGKV1-5 × 05 template human amino acids.
The sequence of the humanized antibody of the A1 antibody is as follows:
VH humanized sequence 48B10-VH-hz
QVQLVESGGGLVKPGGSLRLSCAASGFTFSSYAMSWIRQAPGKGLEWVATITSDGRYSYYPDSVKGRFTISRDNAKNSLYLQMNSLRAEDTAVYYCARRENWFTYWGQGTTVTVSS
' VL humanized sequence 48B10-VL-hz
DIQITQSPSSLSASVGDRVTITCSASSSVTYMYWYQQKPGSSPKPWIYLTSNLASGVPSRFSGSGSGTDFTLTISSLQPEDFATYYCQQWSSGPTFGQGTRLEIK
The sequence shown in SEQ ID NO. 1 is used as a heavy chain constant region, the sequence shown in SEQ ID NO. 2 is used as a light chain constant region, a primer is redesigned for the DNA sequence of the antibody aiming at the PD-L1 target obtained by sequencing, the corresponding genes of the chimeric antibody and the humanized antibody are synthesized and connected into a eukaryotic expression vector, DH5alpha competent cells are transformed, the cells are cultured in a constant temperature incubator at 37 ℃ for overnight, and monoclonal strains are selected for sequencing and identification. The correct strain was selected, shaken, and then transfected into mammalian expression cells 293F, which were then cultured at 37 ℃ in a 5% CO2 incubator for 7 days.
Collecting the expression supernatant, centrifuging, filtering, selecting protein G affinity chromatography column, purifying, detecting the purity of the purified antibody by SDS-PAGE electrophoresis, detecting the antibody concentration by using BCA protein detection kit, subpackaging, and storing in a refrigerator at-80 ℃ for later use. Among them, the chimeric antibody was named "murine antibody abbreviated (chi)", and the humanized antibody was named "murine antibody abbreviated (hz)".
Example 7 FACS detection of the binding of the antibodies of the invention to PD-L1
The experimental procedure was as in example 3, except that after 50. Mu.l of cell suspension was put into each flow tube, the test antibody, the positive control antibody (Reference) or the negative Reference antibody (hIgG 1) was added at the concentration shown in FIG. 3. Control tubes 1 (no antibody and secondary antibody added, only cell suspension added) and 2 (no antibody added, only cell suspension and secondary antibody added) were set simultaneously.
The results of the FACS binding measurements are shown in table 5 and figure 4.
TABLE 5 FACS detection of antibody binding to PD-L1
| Name (R) | EC50(μg/mL) | Fluorescence intensity max |
| Atezolizumab | 0.4584 | 1499 |
| 48B10-4(hz) | 0.377 | 3850 |
EXAMPLE 8 FACS detection of binding of the antibodies of the invention to cynoPD-L1
Antibody binding method to cynoPD-L1 Reference example 3, except that after 50. Mu.l of cell suspension was put into each flow tube, the test antibody, positive control antibody (Reference) or negative Reference antibody (hIgG 1) was added. Control tubes 1 (no antibody and secondary antibody added, only cell suspension added) and 2 (no antibody added, only cell suspension and secondary antibody added) were set simultaneously.
The results of the FACS binding measurements are shown in table 6 and fig. 5.
TABLE 6 FACS detection of antibody binding to PD-L1
| Name(s) | EC50(μg/mL) | Fluorescence intensity max |
| Atezolizumab | 0.5913 | 5034 |
| 48B10-4(hz) | 0.3417 | 8668 |
Example 9 FACS detection of blocking of PD-L1 binding to cells by antibodies of the invention
The experimental procedure was as in example 4, except that the test antibody, the positive control antibody Atezolizumab (Reference) or the negative Reference antibody (hIgG 1) was added to each well.
The results of the FACS blockade detection are shown in table 7 and fig. 6.
TABLE 7 FACS detection of antibody blocking PD-L1 binding to cells
| Name (R) | EC50(μg/mL) | Max/% of blocking rate |
| Atezolizumab | 0.2502 | 99.87 |
| 48B10-4(hz) | 0.5245 | 99.83 |
Example 10 affinity assay for binding of antibodies of the examples of the invention to PD-L1
Extracellular domains of human PD-L1 protein (positions 19 to 238 of SEQ ID NO: 3) were prepared in PBS buffer at a maximum concentration of 30nM and 3-fold diluted to 6 concentrations with a 0 concentration control. The murine, chimeric and humanized antibodies of the invention were prepared as 20nM solutions in PBS.
The affinity of the antibody and antigen was determined using ForteBio Octet as an affinity detection instrument. The specific procedure is as in example 5.
The results are shown in Table 8.
TABLE 8 result of affinity assay of antibody binding to antigen PD-L1
| Antibodies | Antigens | KD(M) | kon(1/Ms) | kdis(1/s) |
| Atezolizumab | Human PD-L1-his | 9.86E-10 | 4.77E+05 | 4.71E-04 |
| 48B10-4(hz) | Human PD-L1-his | 5.95E-10 | 4.26E+05 | 2.54E-04 |
So far, the technical solutions of the present invention have been described in connection with the preferred embodiments shown in the drawings, but it is easily understood by those skilled in the art that the scope of the present invention is obviously not limited to these specific embodiments. Equivalent changes or substitutions of related technical features can be made by those skilled in the art without departing from the principle of the invention, and the technical scheme after the changes or substitutions can fall into the protection scope of the invention.
Claims (12)
1. An antibody or fragment thereof, comprising:
the antibody or fragment thereof comprises a heavy chain variable region (VH) comprising the H-CDR1, H-CDR2, H-CDR3 shown in sequence in SEQ ID NO 35, SEQ ID NO 36, SEQ ID NO 37 and a light chain variable region (VL) comprising the L-CDR1, L-CDR2, L-CDR3 shown in sequence in SEQ ID NO 38, SEQ ID NO 39, SEQ ID NO 40.
2. The antibody or fragment thereof of claim 1, wherein the heavy chain variable region comprises a sequence selected from the group consisting of: an amino acid sequence shown in SEQ ID NO. 9 or an amino acid sequence having at least 75% identity to the amino acid sequence shown in SEQ ID NO. 9; and/or, the light chain variable region comprises a sequence selected from: 10 or an amino acid sequence having at least 75% identity to the amino acid sequence shown in SEQ ID No. 10.
3. The antibody or fragment thereof of claim 2, wherein the antibody or fragment thereof is an anti-PD-L1 antibody or fragment thereof.
4. The antibody or fragment thereof of claim 3, wherein the antibody comprises any of a monoclonal antibody, a single chain antibody, a diabody, a single domain antibody, a nanobody, a fully or partially humanized antibody, and a chimeric antibody, or the antigen-binding fragment is a half-antibody or an antigen-binding fragment of an antibody or a half-antibody, and the antibody of the antigen-binding fragment comprises scFv, bsFv, dsFv, (dsFv) 2 、Fab、Fab'、F(ab') 2 Fv, and IgG.
5. The antibody or fragment thereof of claim 4, wherein the antibody or fragment thereof further comprises a constant region of human or murine origin comprising a heavy chain constant region (CH) of human or murine origin and/or a light chain constant region (CL), wherein the heavy chain constant region comprises an antibody or fragment thereof of IgG, igA, igM, igD or IgE and the light chain constant region comprises a kappa or lambda type.
6. The antibody or fragment thereof of claim 5, wherein the antibody comprises a monoclonal antibody that is murine, chimeric or humanized, wherein the heavy chain constant region of the monoclonal antibody is of the IgG1 or IgG4 subtype and the light chain constant region is of the kappa type;
the heavy chain constant region of the monoclonal antibody comprises the amino acid sequence shown in SEQ ID NO. 1 or an amino acid sequence with at least 75% identity to the amino acid sequence shown in SEQ ID NO. 1;
the light chain constant region of the monoclonal antibody comprises the amino acid sequence set forth in SEQ ID NO. 2 or an amino acid sequence having at least 75% identity to the amino acid sequence set forth in SEQ ID NO. 2.
7. A nucleic acid molecule encoding the antibody or fragment thereof of any one of claims 1 to 6, wherein the antibody or fragment thereof encodes or a heavy chain CDR, a light chain variable region, a heavy chain or a light chain comprised in the antibody or fragment thereof.
8. A vector comprising the nucleic acid molecule of claim 7.
9. A host cell comprising or transformed or transfected with the nucleic acid molecule of claim 7 and/or the vector of claim 8.
10. A pharmaceutical composition comprising the antibody or fragment thereof of any one of claims 1 to 6, the nucleic acid molecule of claim 7, the vector of claim 8, or the host cell of claim 9, and any pharmaceutically acceptable excipient.
11. Use of the antibody or fragment thereof of any one of claims 1 to 6, the nucleic acid molecule of claim 7, the vector of claim 8, the host cell of claim 9, or the pharmaceutical composition of claim 10 in the manufacture of a medicament for the prevention or treatment of a disease comprising a tumor or cancer, including non-small cell lung cancer, melanoma, bladder cancer, merkel lymphoma, cutaneous squamous cell carcinoma, lung cancer, hodgkin lymphoma, kidney cancer, liver cancer, esophageal cancer, non-hodgkin lymphoma, breast cancer, thyroid cancer, gastric cancer, intestinal cancer, nasopharyngeal cancer, pancreatic cancer, prostate cancer, leukemia, laryngeal cancer, oral cancer, otic eye tumor, biliary tract cancer, gallbladder cancer, adrenal cancer, reproductive system tumor, multiple myeloma, nervous system tumor, urothelial cell cancer.
12. A kit comprising the antibody or fragment thereof of any one of claims 1 to 6, the nucleic acid molecule of claim 7, the vector of claim 8, the host cell of claim 9, or the pharmaceutical composition of claim 10.
Priority Applications (2)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202210936286.9A CN115925937A (en) | 2022-08-05 | 2022-08-05 | Antibody or fragment thereof and pharmaceutical application thereof |
| CN202310978868.8A CN116789833B (en) | 2022-08-05 | 2023-08-04 | Antibody or fragment thereof and pharmaceutical application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202210936286.9A CN115925937A (en) | 2022-08-05 | 2022-08-05 | Antibody or fragment thereof and pharmaceutical application thereof |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN115925937A true CN115925937A (en) | 2023-04-07 |
Family
ID=86551095
Family Applications (2)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202210936286.9A Pending CN115925937A (en) | 2022-08-05 | 2022-08-05 | Antibody or fragment thereof and pharmaceutical application thereof |
| CN202310978868.8A Active CN116789833B (en) | 2022-08-05 | 2023-08-04 | Antibody or fragment thereof and pharmaceutical application thereof |
Family Applications After (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202310978868.8A Active CN116789833B (en) | 2022-08-05 | 2023-08-04 | Antibody or fragment thereof and pharmaceutical application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (2) | CN115925937A (en) |
Citations (6)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106699891A (en) * | 2017-01-25 | 2017-05-24 | 北京天广实生物技术股份有限公司 | Anti-PD-L1 antibody as well as pharmaceutical composition and application of anti-PD-L1 antibody |
| CN107151269A (en) * | 2016-03-04 | 2017-09-12 | 四川科伦博泰生物医药股份有限公司 | A kind of antibody of PDL 1, its medical composition and its use |
| CN108640992A (en) * | 2018-05-08 | 2018-10-12 | 湖南燕山源创生物科技有限公司 | Anti-human PD1 antibody and its application |
| WO2019137397A1 (en) * | 2018-01-10 | 2019-07-18 | 江苏恒瑞医药股份有限公司 | Pd-l1 antibody, antigen-binding fragment thereof, and pharmaceutical use thereof |
| WO2021077250A1 (en) * | 2019-10-21 | 2021-04-29 | 康源博创生物科技(北京)有限公司 | Anti-pd-l1 antibody and pharmaceutical use thereof |
| CN112759647A (en) * | 2019-10-21 | 2021-05-07 | 康源博创生物科技(北京)有限公司 | anti-PD-L1 antibody and pharmaceutical application thereof |
Family Cites Families (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN106478819B (en) * | 2016-10-27 | 2018-12-07 | 常州费洛斯药业科技有限公司 | A kind of monoclonal antibody or antibody fragment for PD-L1 |
| CN112521503A (en) * | 2019-09-19 | 2021-03-19 | 北京慧能安生物科技有限公司 | Monoclonal antibody or antigen binding fragment thereof of anti-human PD-L1, and coding gene and application thereof |
| CN110964111B (en) * | 2019-12-25 | 2020-07-24 | 北京东方百泰生物科技有限公司 | anti-PD-L1 monoclonal antibody, antigen binding fragment thereof and application thereof |
-
2022
- 2022-08-05 CN CN202210936286.9A patent/CN115925937A/en active Pending
-
2023
- 2023-08-04 CN CN202310978868.8A patent/CN116789833B/en active Active
Patent Citations (7)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN107151269A (en) * | 2016-03-04 | 2017-09-12 | 四川科伦博泰生物医药股份有限公司 | A kind of antibody of PDL 1, its medical composition and its use |
| CN106699891A (en) * | 2017-01-25 | 2017-05-24 | 北京天广实生物技术股份有限公司 | Anti-PD-L1 antibody as well as pharmaceutical composition and application of anti-PD-L1 antibody |
| WO2019137397A1 (en) * | 2018-01-10 | 2019-07-18 | 江苏恒瑞医药股份有限公司 | Pd-l1 antibody, antigen-binding fragment thereof, and pharmaceutical use thereof |
| CN111278861A (en) * | 2018-01-10 | 2020-06-12 | 江苏恒瑞医药股份有限公司 | PD-L1 antibody, antigen binding fragment thereof and medical application |
| CN108640992A (en) * | 2018-05-08 | 2018-10-12 | 湖南燕山源创生物科技有限公司 | Anti-human PD1 antibody and its application |
| WO2021077250A1 (en) * | 2019-10-21 | 2021-04-29 | 康源博创生物科技(北京)有限公司 | Anti-pd-l1 antibody and pharmaceutical use thereof |
| CN112759647A (en) * | 2019-10-21 | 2021-05-07 | 康源博创生物科技(北京)有限公司 | anti-PD-L1 antibody and pharmaceutical application thereof |
Also Published As
| Publication number | Publication date |
|---|---|
| CN116789833B (en) | 2023-12-12 |
| CN116789833A (en) | 2023-09-22 |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| TWI830774B (en) | Anti-CD47 antibodies and their applications | |
| US11365260B2 (en) | Agonistic 4-1BB monoclonal antibody | |
| CN110066336B (en) | anti-CD 47 monoclonal antibody, fragment and medical application thereof | |
| WO2017197667A1 (en) | Anti-human pd-l1 humanized monoclonal antibody and application thereof | |
| CN111848800B (en) | PD-L1 single-domain antibody and application thereof | |
| CN111393529B (en) | anti-OX 40 antibodies that non-competitively bind to OX40L | |
| TWI776364B (en) | A kind of BCMA binding protein and its preparation method and application | |
| EP4458851A1 (en) | Anti-gprc5d antibody and use thereof | |
| CN113508139A (en) | Antibodies that bind human LAG-3, methods of making, and uses thereof | |
| CN115386006A (en) | anti-GPRC 5D antibody, preparation method and application thereof | |
| CN112745391A (en) | PD-L1 binding molecules | |
| JP2024028850A (en) | Anti-pd-l1 antibody and pharmaceutical use thereof | |
| CN107108734B (en) | Monoclonal anti-GPC-1 antibodies and uses thereof | |
| CN113817057B (en) | Anti-siglec 15 antibody and application thereof | |
| TWI788788B (en) | Target EpCAM antibody and its preparation and application | |
| CN113527484B (en) | Anti-CD47 monoclonal antibody | |
| CN112759647A (en) | anti-PD-L1 antibody and pharmaceutical application thereof | |
| CN116789833B (en) | Antibody or fragment thereof and pharmaceutical application thereof | |
| CN117769568A (en) | anti-CD 24 antibody and application thereof | |
| WO2018235964A1 (en) | Anti-tgf-beta1 antibodies | |
| TWI804099B (en) | Antibody specifically binding to glycosylated CEACAM5 and preparation method thereof | |
| US20250092124A1 (en) | Antibody molecule against growth and differentiation factor 15 and use thereof | |
| CN114075284B (en) | CD47 binding molecules and uses thereof | |
| US20230002494A1 (en) | Pd-l1 binding molecule | |
| CN119176874A (en) | An antibody or antigen binding fragment thereof capable of binding to LILRB2 |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| WD01 | Invention patent application deemed withdrawn after publication | ||
| WD01 | Invention patent application deemed withdrawn after publication |
Application publication date: 20230407 |