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WO2025201493A1 - Dll3 binding molecule and use thereof - Google Patents

Dll3 binding molecule and use thereof

Info

Publication number
WO2025201493A1
WO2025201493A1 PCT/CN2025/085579 CN2025085579W WO2025201493A1 WO 2025201493 A1 WO2025201493 A1 WO 2025201493A1 CN 2025085579 W CN2025085579 W CN 2025085579W WO 2025201493 A1 WO2025201493 A1 WO 2025201493A1
Authority
WO
WIPO (PCT)
Prior art keywords
amino acid
seq
dll3
hcdr2
hcdr1
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
Application number
PCT/CN2025/085579
Other languages
French (fr)
Chinese (zh)
Inventor
黄冰
贾鸽子
王健
郑海林
杨媛媛
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
Nona Biosciences Suzhou Co Ltd
Original Assignee
Nona Biosciences Suzhou Co Ltd
Priority date (The priority date 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 date listed.)
Filing date
Publication date
Application filed by Nona Biosciences Suzhou Co Ltd filed Critical Nona Biosciences Suzhou Co Ltd
Publication of WO2025201493A1 publication Critical patent/WO2025201493A1/en
Pending legal-status Critical Current
Anticipated expiration legal-status Critical

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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
    • C07K16/30—Immunoglobulins [IGs], e.g. monoclonal or polyclonal antibodies against material from animals or humans against receptors, cell surface antigens or cell surface determinants from tumour cells

Definitions

  • DLL3 Delta-Like Ligand 3
  • SCLC small cell lung cancer
  • LNEC large cell neuroendocrine carcinoma
  • DLL3 is expressed in other tumor types of neuroendocrine origin, such as melanoma, glioblastoma multiforme, small cell bladder cancer, metastatic castration-resistant prostate cancer, and neuroendocrine lung tumors.
  • DLL3 In tumor cells, DLL3 is specifically expressed on the cell surface. In contrast, only a few normal cell types (such as neurons, pancreatic islet cells, and pituitary cells) express DLL3, and in normal cells, DLL3 expression is confined to the cytoplasm.
  • the DLL3 expression profile (high, homogeneous cell surface expression in tumors versus low, cytoplasmic expression in subsets of normal tissues) enables the development of therapies that specifically target tumor cells (e.g., SCLC cells) with DLL3.
  • therapies that specifically target tumor cells (e.g., SCLC cells) with DLL3.
  • tumor cells e.g., SCLC cells
  • DLL3-specific drugs against tumors (e.g., SCLC and other neuroendocrine tumors).
  • DLL3-targeting antibody drugs include Rova-T (Rovalpituzumab tesirine), an antibody-drug conjugate developed by AbbVie.
  • Rova-T TM is an antibody-drug conjugate that targets DLL3 in small cell lung cancer (SCLC) tumors, where it is internalized and releases the toxin pyrrolobenzodiazepine, which induces cell death.
  • TCE molecules such as Amgen's tarlatamab, Harpoon Therapeutics' HPN328, Boehringer Ingelheim's BI 764532, Qilu Pharmaceutical's QLS31904, Roche's RO7616789, and Phanes Therapeutics' PT217 are all in Phase I clinical trials.
  • Tarlatamab (AMG 757), developed by Amgen, has been optimized to extend the half-life of the BiTE molecule, activating T cells to target tumors expressing DLL3.
  • a Phase I clinical study of AMG 757 for SCLC patients (NCT03319940) is ongoing.
  • Chimeric antigen receptor T cell therapy genetically modifies the patient's autologous T cells to guide the patient's T cells to express chimeric receptors for tumor antigens, and then re-injects these cells into the patient's body to attack and kill target cells.
  • AMG119 is an adoptive cell therapy that modifies the patient's autologous T cells to express a transmembrane chimeric receptor (CAR) targeting DLL3 and attack DLL3-positive cells.
  • CAR transmembrane chimeric receptor
  • DLL3 As an inhibitory ligand for Notch signaling, DLL3 is closely associated with various tumors. In SCLC, in particular, extensive basic and clinical research evidence indicates that DLL3 is a promising therapeutic target. Therefore, exploring antibody drugs targeting DLL3 for cancer treatment is expected to provide more options for cancer treatment.
  • the present invention provides a DLL3 binding molecule comprising a heavy chain variable region, wherein the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1 comprises the sequence shown in SEQ ID NO: 18 or 32 or an amino acid sequence that differs from the sequence shown in SEQ ID NO: 18 or 32 by no more than 2 amino acids in amino acid addition, deletion or substitution; and/or HCDR2 comprises the sequence shown in SEQ ID NO: 69, 74, 79, 80, 87 or 90 or an amino acid sequence that differs from the sequence shown in SEQ ID NO: 69, 74, 79, 80, 87 or 90 by no more than 2 amino acids in amino acid addition, deletion or substitution.
  • Sequence; and/or HCDR3 comprises the sequence shown in SEQ ID NO: 126, 127, 128, 129, 133, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 146, 147, 148, 149, 150, 151 or 152, or an amino acid sequence that differs from the sequence shown in SEQ ID NO: 126, 127, 128, 129, 133, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 146, 147, 148, 149, 150, 151 or 152 by amino acid addition, deletion or substitution of no more than 2 amino acids.
  • the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 18, 60, 126, respectively; or HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 19, 60, 126, respectively; or HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 18, 61, 126, respectively; or HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 20, 62, 127, respectively; or HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NOs: 18, 63, and 128, respectively; or HCDR1, HCDR2, and HCDR3 comprise the amino acid sequences shown in SEQ ID NOs: 21, 64, and 129, respectively; or HCDR1, HCDR2, and HCDR3
  • R2 and HCDR3 contain the amino acid sequences shown in SEQ ID NO: 18, 64, 131, respectively; or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NO: 19, 68, 126, respectively; or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NO: 18, 68, 126, respectively; or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NO: 20, 69, 132, respectively; or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NO: 24, 66, 126, respectively; or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NO: 24, 66, 126, respectively; or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NO: 24, 66, 126, respectively; or CDR2 and HCDR3 comprise
  • the heavy chain variable region comprises 1) SEQ ID NO: 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 185, 186, 187, 188, 189, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 202, 213,
  • the invention provides a multispecific antigen-binding construct comprising a first antigen-binding moiety that binds DLL3, wherein the first antigen-binding moiety comprises the DLL3-binding molecule of the first aspect of the invention.
  • the multispecific antigen-binding construct comprises a first antigen-binding moiety that binds DLL3 and a second antigen-binding moiety that binds a second antigen, wherein the first antigen-binding moiety comprises the DLL3-binding molecule of the first aspect of the invention.
  • the present invention provides a chimeric antigen receptor comprising the DLL3 binding molecule of the first aspect of the invention.
  • the invention provides an antibody conjugate comprising the DLL3 binding molecule of the first aspect of the invention or the multispecific antigen-binding construct of the second aspect of the invention conjugated to at least one therapeutic agent.
  • the present invention provides use of the DLL3 binding molecule of the first aspect of the invention, the multispecific antigen-binding construct of the second aspect of the invention, the immune effector cell of the fourth aspect of the invention, the antibody conjugate of the fifth aspect of the invention, or the pharmaceutical composition of the sixth aspect of the invention in the preparation of a medicament for treating cancer.
  • the present invention provides a polynucleotide encoding the DLL3 binding molecule of the first aspect of the invention or the multispecific antigen-binding construct of the second aspect of the invention.
  • 1A-1D show the binding activity of candidate antibodies to human DLL3 protein.
  • 2A-2D show the binding activity of candidate antibodies to cynomolgus monkey DLL3 protein.
  • Figures 4A-4D show the binding activity of candidate antibodies to CHO-K1 cells overexpressing human DLL3.
  • 10A-10B show the internalization of candidate antibodies in SHP-77 tumor cells.
  • antibody refers to an immunoglobulin or a fragment thereof that specifically binds to an antigenic epitope through at least one antigen binding site. Antibody encompasses antibody fragments.
  • the term “antibody” includes synthetic antibodies, recombinantly produced antibodies, multispecific antibodies (e.g., bispecific antibodies), human antibodies, non-human antibodies, humanized antibodies, fully humanized antibodies, heavy chain antibodies, nanobodies, chimeric antibodies, intracellular antibodies, and antibody fragments, such as but not limited to Fab fragments, Fab' fragments, F(ab') 2 fragments, Fv fragments, disulfide-linked Fv (dsFv), Fd fragments, Fd' fragments, single-chain Fv (scFv), single-chain Fab (scFab), diabodies, anti-idiotypic (anti-Id) antibodies, or antigen-binding fragments of any of the above antibodies.
  • the antibodies provided herein include members of any immunoglobulin class (e.g., IgG, IgM, IgD, IgE, IgA, and IgY), any class (e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2), or subclass (e.g., IgG2a and IgG2b).
  • immunoglobulin class e.g., IgG, IgM, IgD, IgE, IgA, and IgY
  • any class e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2b
  • subclass e.g., IgG2a and IgG2b.
  • the antibodies of the present invention are heavy chain antibodies, and/or are fully humanized antibodies.
  • an "antibody fragment” or “antigen-binding fragment” of an antibody refers to any portion of a full-length antibody that is less than full-length but contains at least a portion of the variable region (e.g., one or more CDRs and/or one or more antibody binding sites) of the antibody that binds to an antigen and thus retains binding specificity and at least a portion of the specific binding ability of the full-length antibody.
  • an antigen-binding fragment refers to an antibody fragment that contains an antigen-binding portion that binds to the same antigen as the antibody from which the antibody fragment was derived.
  • Antibody fragments include antibody derivatives produced by enzymatic treatment of full-length antibodies, as well as synthetically produced derivatives, such as recombinantly produced derivatives.
  • Antibodies include antibody fragments. Examples of antibody fragments include, but are not limited to, Fab, Fab', F(ab') 2 , single-chain Fv (scFv), Fv, dsFv, diabodies, Fd and Fd' fragments, and other fragments, including modified fragments (see, e.g., Methods in Molecular Biology, Vol 207: Recombinant Antibodies for Cancer Therapy Methods and Protocols (2003); Chapter 1; p 3-25, Kipriyanov).
  • the fragments can include multiple chains linked together, for example, by disulfide bonds and/or by peptide linkers.
  • Antibody fragments generally contain at least or about 50 amino acids, and typically at least or about 200 amino acids.
  • Antigen-binding fragments include any antibody fragment that, when inserted into an antibody framework (e.g., by replacing the corresponding regions), results in an antibody that immunospecifically binds to an antigen.
  • heavy-chain-only antibody and “heavy-chain antibody” are used interchangeably and in their broadest sense to refer to antibodies that lack conventional antibody light chains and contain only a heavy chain variable region and a heavy chain constant region that does not contain CH1 (e.g., Fc fragment).
  • Fc fragment generally refers to a crystallizable fragment of a conventional antibody or heavy chain antibody after papain digestion.
  • the Fc fragment of IgG and heavy chain antibodies can include part of the hinge region, CH2 and CH3.
  • the Fc fragment can include at least part of the hinge region (e.g., all or part of the hinge region), CH2 and CH3.
  • variable region i.e., "binding domain” allows the binding molecule to selectively recognize an epitope on an antigen and specifically bind to the epitope. That is, for example, the light chain variable region (VL) domain and the heavy chain variable region (VH) domain of the binding molecule of an antibody, or these complementary determining regions (CDR) subgroups combine to form a variable region that determines a three-dimensional antigen binding site. More specifically, the antigen binding site is determined by three CDRs on each VH and VL chain.
  • the heavy chain antibody of the present invention has an antigen binding site determined by three CDRs on the VH chain.
  • CDRs complementary determining regions
  • FR framework
  • the binding domain formed by the positioned CDRs determines a surface that is complementary to the epitope on the immunoreactive antigen. This complementary surface promotes the non-covalent binding of the antibody to its complementary epitope.
  • the amino acids that make up the CDRs and framework regions, respectively, in any given heavy or light chain variable region can be identified by conventional methods (see, "Sequences of Proteins of Immunological Interest," Kabat, E.
  • the CDRs (CDRL or LCDR) of the light chain variable region may be referred to as LCDR1, LCDR2, and LCDR3, and the CDRs (CDRH or HCDR) of the heavy chain variable region may be referred to as HCDR1, HCDR2, and HCDR3.
  • the amino acid sequences of CDRs are all shown according to the Chothia definition rules (the sequences in the claims of the present invention are also shown according to the Chothia definition rules).
  • the CDR of an antibody can be defined in the art by a variety of methods, such as Chothia based on the three-dimensional structure of the antibody and the topology of the CDR loop (see, for example, Chothia, C. et al., Nature, 342, 877-883 (1989); and Al-Lazikani, B. et al., J. Mol. Biol., 273, 927-948 (1997)), Kabat based on antibody sequence variability (see, for example, Kabat, E.A.
  • CDR complementary determining region
  • the terms "CDR” and "complementarity determining region" of a given antibody or a region thereof (such as a variable region) should be understood to cover the complementary determining region defined by any of the above-mentioned known schemes described by the present invention.
  • the scope of protection claimed in the claims of the present invention is based on the sequences shown in the Chothia definition rules, the amino acid sequences corresponding to the definition rules of other CDRs should also fall within the scope of protection of the present invention.
  • antibodies defined by specific CDR sequences defined herein the scope of said antibodies also encompasses antibodies whose variable region sequences comprise said specific CDR sequences, but whose declared CDR boundaries differ from the specific CDR boundaries defined herein due to the application of a different scheme (e.g., a different assignment system rule or combination).
  • framework region and “framework region” are used interchangeably.
  • framework region refers to those amino acid residues in the antibody variable region excluding the CDR sequences as defined above.
  • disulfide bond includes a covalent bond formed between two sulfur atoms.
  • the amino acid cysteine contains a sulfhydryl group that can form a disulfide bond or bridge a second sulfhydryl group.
  • percent (%) sequence identity or “sequence identity” of amino acid sequences has the art-recognized definition of the percentage of identity between two polypeptide sequences as determined by sequence alignment (e.g., by manual inspection or a publicly known algorithm). This can be determined using methods known to those skilled in the art, for example, using publicly available computer software such as BLAST, BLAST-2, Clustal Omega, and FASTA software.
  • Affinity or "binding affinity” is a measure of the strength of the non-covalent binding between an antibody and an antigen. Affinity can be determined using conventional techniques known in the art, such as membrane interferometry (e.g., using the Octet Fortebio assay system), radioimmunoassay, surface plasmon resonance, enzyme-linked immunosorbent assay (ELISA), or flow cytometry (FACS).
  • membrane interferometry e.g., using the Octet Fortebio assay system
  • radioimmunoassay e.g., using the Octet Fortebio assay system
  • ELISA enzyme-linked immunosorbent assay
  • FACS flow cytometry
  • binding molecule "A” has a higher specificity for a given epitope than binding molecule "B,” or it can be said that binding molecule "A” specifically binds to epitope "C” with a higher specificity than its specificity for a related epitope "D.”
  • endocytosis refers to the process by which a substance to be internalized is enclosed by the cell membrane and then buds into a vesicle containing the internalized substance, bringing the external substance into the cell. This is one way that substances are transported across the cytoplasmic membrane.
  • antibody internalization refers to the induction of receptor-mediated endocytosis when a cell surface receptor binds to its specific antibody.
  • DLL3 antibodies can mediate the internalization of cell-surface-expressed DLL3 protein by binding to the extracellular portion of DLL3. The endocytic effect of antibodies can be measured using conventional techniques known in the art, such as labeling with internalizing antibodies.
  • polynucleotide and “nucleic acid” are used interchangeably to refer to a polymer of deoxyribonucleotides (deoxyribonucleic acid, DNA) or a polymer of ribonucleotides (ribonucleic acid, RNA).
  • Polynucleotide sequence “nucleic acid sequence,” and “nucleotide sequence” are used interchangeably to refer to the order of nucleotides in a polynucleotide.
  • an isolated nucleic acid molecule is one that is separated from other nucleic acid molecules present in the natural source of the nucleic acid molecule.
  • An "isolated" nucleic acid molecule such as a cDNA molecule, can be substantially free of other cellular material or culture medium when produced by recombinant techniques, or substantially free of chemical precursors or other chemical components when chemically synthesized.
  • Exemplary isolated nucleic acid molecules provided herein include isolated nucleic acid molecules that encode provided DLL3 binding molecules.
  • expression includes transcription and/or translation of a nucleotide sequence.
  • expression can involve the production of transcripts and/or polypeptides.
  • Viral vectors include, but are not limited to, retroviral vectors (including lentiviral vectors), adenoviral vectors, adeno-associated viral vectors, herpes virus vectors, pox virus vectors, and baculovirus vectors, etc.
  • a "host cell” is a cell that is used to receive, maintain, replicate, and amplify a vector.
  • a host cell can also be used to express a polypeptide encoded by the vector. When the host cell divides, the nucleic acid contained in the vector is replicated, thereby amplifying the nucleic acid.
  • the host cell can be a eukaryotic cell or a prokaryotic cell. Suitable host cells include, but are not limited to, CHO cells, various COS cells, HeLa cells, and HEK cells such as HEK 293 cells.
  • a "subject in need thereof” can include a subject already suffering from a disease; a subject susceptible to a disease; and a subject in need of prevention of a disease.
  • a DLL3 binding molecule can include any molecule that specifically binds to DLL3.
  • the DLL3 binding molecule is a polypeptide or protein, such as an antibody.
  • the DLL3 binding molecule is an antibody or antigen-binding fragment thereof directed against DLL3.
  • DLL3-binding molecules include synthetic antibodies, recombinantly produced antibodies, multispecific antibodies (e.g., bispecific antibodies), human antibodies, non-human antibodies, humanized antibodies, fully humanized antibodies, heavy chain antibodies, nanobodies, chimeric antibodies, intrabodies, and antibody fragments, such as, but not limited to, Fab fragments, Fab' fragments, F(ab')2 fragments, Fv fragments, disulfide-linked Fv (dsFv), Fd fragments, Fd' fragments, single-chain Fv (scFv), single-chain Fab (scFab), diabodies, anti-idiotypic (anti-Id) antibodies, or antigen-binding fragments of any of the foregoing.
  • multispecific antibodies e.g., bispecific antibodies
  • human antibodies e.g., non-human antibodies, humanized antibodies, fully humanized antibodies, heavy chain antibodies, nanobodies, chimeric antibodies, intrabodies, and antibody fragments, such as, but not limited to,
  • the DLL3 binding molecule is a heavy chain antibody or antigen-binding fragment thereof comprising a heavy chain variable region.
  • the DLL3 binding molecule is a heavy chain antibody. In another embodiment, the DLL3 binding molecule is a fully humanized antibody. In a preferred embodiment, the DLL3 binding molecule is a heavy chain antibody and is a fully humanized antibody.
  • the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 3 is substituted. In one embodiment, the amino acid at position 3 in SEQ ID NO: 18 is substituted with N (SEQ ID NO: 19; GFNFSDY).
  • the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 4 is substituted. In one embodiment, the amino acid at position 4 in SEQ ID NO: 18 is substituted with L (SEQ ID NO: 20; GFTLSDY).
  • the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 4 is substituted. In one embodiment, the amino acid at position 4 in SEQ ID NO: 18 is substituted with 1 (SEQ ID NO: 21; GFTISDY).
  • the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 3 is substituted. In some embodiments, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 4 is substituted. In some embodiments, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 3 and the amino acid at position 4 are substituted. In one embodiment, the amino acid at position 3 in SEQ ID NO: 18 is substituted with N, and the amino acid at position 4 in SEQ ID NO: 18 is substituted with I (SEQ ID NO: 22; GFNISDY).
  • the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 3 is substituted. In some embodiments, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 4 is substituted. In some embodiments, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 3 and the amino acid at position 4 are substituted. In one embodiment, the amino acid at position 3 in SEQ ID NO: 18 is substituted with N, and the amino acid at position 4 in SEQ ID NO: 18 is substituted with L (SEQ ID NO: 23; GFNLSDY).
  • the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 3 is substituted. In some embodiments, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 5 is substituted. In some embodiments, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 3 and the amino acid at position 5 are substituted. In one embodiment, the amino acid at position 3 in SEQ ID NO: 18 is substituted with N, and the amino acid at position 5 in SEQ ID NO: 18 is substituted with N (SEQ ID NO: 24; GFNFNDY).
  • the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 6 is substituted. In one embodiment, the amino acid at position 6 in SEQ ID NO: 18 is substituted with N (SEQ ID NO: 26; GFTFSNY).
  • the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 7 is substituted. In one embodiment, the amino acid at position 7 in SEQ ID NO: 18 is substituted with H (SEQ ID NO: 27; GFTFSDH).
  • the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 6 is substituted. In one embodiment, the amino acid at position 6 in SEQ ID NO: 18 is substituted with T (SEQ ID NO: 28; GFTFSTY).
  • the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 3 is substituted. In one embodiment, the amino acid at position 3 in SEQ ID NO: 18 is substituted with A (SEQ ID NO: 29; GFAFSDY).
  • the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 3 is substituted. In some embodiments, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 6 is substituted. In some embodiments, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 3 and the amino acid at position 6 are substituted. In one embodiment, the amino acid at position 3 in SEQ ID NO: 18 is substituted with I, and the amino acid at position 6 in SEQ ID NO: 18 is substituted with N (SEQ ID NO: 30; GFIFSNY).
  • the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 3 is substituted. In some embodiments, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 6 is substituted. In some embodiments, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 3 and the amino acid at position 6 are substituted. In one embodiment, the amino acid at position 3 in SEQ ID NO: 18 is substituted with I, and the amino acid at position 6 in SEQ ID NO: 18 is substituted with S (SEQ ID NO: 31; GFIFSSY).
  • the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 4 is substituted. In some embodiments, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 6 is substituted. In some embodiments, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 4 and the amino acid at position 6 are substituted. In one embodiment, the amino acid at position 4 in SEQ ID NO: 18 is substituted with I, and the amino acid at position 6 in SEQ ID NO: 18 is substituted with N (SEQ ID NO: 33; GFTISNY).
  • the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 2 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 3 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 2 and the amino acid at position 3 are substituted. In one embodiment, the amino acid at position 2 in SEQ ID NO: 69 is substituted with A, and the amino acid at position 3 in SEQ ID NO: 69 is substituted with N (SEQ ID NO: 61; SANGST).
  • the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 1 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 2 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 1 and the amino acid at position 2 are substituted. In one embodiment, the amino acid at position 1 in SEQ ID NO: 69 is substituted with T, and the amino acid at position 2 in SEQ ID NO: 69 is substituted with G (SEQ ID NO: 62; TGSGST).
  • the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 2 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 5 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 2 and the amino acid at position 5 are substituted. In one embodiment, the amino acid at position 2 in SEQ ID NO: 69 is substituted with G, and the amino acid at position 5 in SEQ ID NO: 69 is substituted with R (SEQ ID NO: 65; SGSGRT).
  • the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 2 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 6 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 2 and the amino acid at position 6 are substituted. In one embodiment, the amino acid at position 2 in SEQ ID NO: 69 is substituted with G, and the amino acid at position 6 in SEQ ID NO: 69 is substituted with I (SEQ ID NO: 68; SGSGSI).
  • the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 5 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 6 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 5 and the amino acid at position 6 are substituted. In one embodiment, the amino acid at position 5 in SEQ ID NO: 69 is substituted with D, and the amino acid at position 6 in SEQ ID NO: 69 is substituted with S (SEQ ID NO: 70; SSSGDS).
  • the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 5 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 6 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 5 and the amino acid at position 6 are substituted. In one embodiment, the amino acid at position 5 in SEQ ID NO: 69 is substituted with N, and the amino acid at position 6 in SEQ ID NO: 69 is substituted with S (SEQ ID NO: 73; SSSGNS).
  • the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 3 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 6 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 3 and the amino acid at position 6 are substituted. In one embodiment, the amino acid at position 3 in SEQ ID NO: 69 is substituted with D, and the amino acid at position 6 in SEQ ID NO: 69 is substituted with S (SEQ ID NO: 75; SSDGSS).
  • the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 2 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 6 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 2 and the amino acid at position 6 are substituted. In one embodiment, the amino acid at position 2 in SEQ ID NO: 69 is substituted with G, and the amino acid at position 6 in SEQ ID NO: 69 is substituted with D (SEQ ID NO: 76; SGSGSD).
  • the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 3 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 6 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 3 and the amino acid at position 6 are substituted. In one embodiment, the amino acid at position 3 in SEQ ID NO: 69 is substituted with D, and the amino acid at position 6 in SEQ ID NO: 69 is substituted with N (SEQ ID NO: 83; SSDGSN).
  • the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 2 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 6 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 2 and the amino acid at position 6 are substituted. In one embodiment, the amino acid at position 2 in SEQ ID NO: 69 is substituted with T, and the amino acid at position 6 in SEQ ID NO: 69 is substituted with P (SEQ ID NO: 89; STSGSP).
  • the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 74 (SEQ ID NO: 74; SGSDNN), wherein the amino acid at position 1 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 74 (SEQ ID NO: 74; SGSDNN), wherein the amino acid at position 3 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 74 (SEQ ID NO: 74; SGSDNN), wherein the amino acid at position 1 and the amino acid at position 3 are substituted. In one embodiment, the amino acid at position 1 in SEQ ID NO: 74 is substituted with N, and the amino acid at position 3 in SEQ ID NO: 74 is substituted with N (SEQ ID NO: 72; NGNDNN).
  • the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 79 (SEQ ID NO: 79; NSDGST), wherein the amino acid at position 2 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 79 (SEQ ID NO: 79; NSDGST), wherein the amino acid at position 6 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 79 (SEQ ID NO: 79; NSDGST), wherein the amino acid at position 2 and the amino acid at position 6 are substituted. In one embodiment, the amino acid at position 2 in SEQ ID NO: 79 is substituted with G, and the amino acid at position 6 in SEQ ID NO: 79 is substituted with N (SEQ ID NO: 77; NGDGSN).
  • the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 79 (SEQ ID NO: 79; NSDGST), wherein the amino acid at position 6 is substituted. In one embodiment, the amino acid at position 6 in SEQ ID NO: 79 is substituted with Y (SEQ ID NO: 78; NSDGSY).
  • the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 79 (SEQ ID NO: 79; NSDGST), wherein the amino acid at position 3 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 79 (SEQ ID NO: 79; NSDGST), wherein the amino acid at position 5 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 79 (SEQ ID NO: 79; NSDGST), wherein the amino acid at position 3 and the amino acid at position 5 are substituted. In one embodiment, the amino acid at position 3 in SEQ ID NO: 79 is substituted with N, and the amino acid at position 5 in SEQ ID NO: 79 is substituted with N (SEQ ID NO: 81; NSNGNT).
  • the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 79 (SEQ ID NO: 79; NSDGST), wherein the amino acid at position 6 is substituted. In one embodiment, the amino acid at position 6 in SEQ ID NO: 79 is substituted with D (SEQ ID NO: 84; NSDGSD).
  • the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 79 (SEQ ID NO: 79; NSDGST), wherein the amino acid at position 6 is substituted. In one embodiment, the amino acid at position 6 in SEQ ID NO: 79 is substituted with N (SEQ ID NO: 85; NSDGSN).
  • the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 79 (SEQ ID NO: 79; NSDGST), wherein the amino acid at position 6 is substituted. In one embodiment, the amino acid at position 6 in SEQ ID NO: 79 is substituted with S (SEQ ID NO: 88; NSDGSS).
  • the HCDR3 sequence comprises the amino acid sequence of SEQ ID NO: 126 (SEQ ID NO: 126; ENGNSDY), wherein the amino acid at position 2 is substituted. In one embodiment, the amino acid at position 2 in SEQ ID NO: 126 is substituted with S (SEQ ID NO: 131; ESGNSDY).
  • the HCDR3 sequence comprises the amino acid sequence of SEQ ID NO: 127 (SEQ ID NO: 127; GLGEFSLGI), wherein the amino acid at position 9 is substituted. In one embodiment, the amino acid at position 9 in SEQ ID NO: 127 is substituted with S (SEQ ID NO: 130; GLGEFSLGS).
  • the HCDR3 sequence comprises the amino acid sequence of SEQ ID NO: 127 (SEQ ID NO: 127; GLGEFSLGI), wherein the amino acid at position 5 is substituted. In one embodiment, the amino acid at position 5 in SEQ ID NO: 127 is substituted with L (SEQ ID NO: 132; GLGELSLGI).
  • the HCDR3 sequence comprises the amino acid sequence of SEQ ID NO: 133 (SEQ ID NO: 133; ERGSTGGWFDP), wherein the amino acid at position 2 is substituted.
  • the amino acid at position 2 in SEQ ID NO: 133 is substituted with K (SEQ ID NO: 134; EKGSTGGWFDP).
  • the HCDR3 sequence comprises the amino acid sequence of SEQ ID NO: 139 (SEQ ID NO: 139; GGDTVVVPEFDY), wherein the amino acid at position 2 is substituted. In some embodiments, the HCDR3 sequence comprises the amino acid sequence of SEQ ID NO: 139 (SEQ ID NO: 139; GGDTVVVPEFDY), wherein the amino acid at position 7 is substituted. In some embodiments, the HCDR3 sequence comprises the amino acid sequence of SEQ ID NO: 139 (SEQ ID NO: 139; GGDTVVVPEFDY), wherein the amino acid at position 2 and the amino acid at position 7 are substituted. In one embodiment, the amino acid at position 2 in SEQ ID NO: 139 is substituted with D, and the amino acid at position 7 in SEQ ID NO: 139 is substituted with L (SEQ ID NO: 145; GDDTVVLPEFDY).
  • HCDR1 comprises an amino acid sequence as shown in SEQ ID NO: 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33 or 34.
  • HCDR2 comprises an amino acid sequence as shown in SEQ ID NO: 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89 or 90.
  • HCDR3 comprises an amino acid sequence as shown in SEQ ID NO:126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151 or 152.
  • HCDR1 comprises an amino acid sequence as shown in SEQ ID NO: 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33 or 34
  • HCDR2 comprises an amino acid sequence as shown in SEQ ID NO: 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85 2, 83, 84, 85, 86, 87, 88, 89 or 90
  • HCDR3 comprises the amino acid sequence shown in SEQ ID NO:126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151 or 152.
  • the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:18, 60, 126, respectively.
  • the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:19, 60, 126, respectively.
  • the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:18, 61, 126, respectively.
  • the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 20, 62, 127, respectively.
  • the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:18, 63, 128, respectively.
  • the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 22, 65, and 126, respectively.
  • the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 23, 65, and 126, respectively.
  • the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:18, 66, 126, respectively.
  • the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:18, 67, 130, respectively.
  • the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:18, 64, 131, respectively.
  • the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:19, 68, and 126, respectively.
  • the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:18, 68, 126, respectively.
  • the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 20, 69, 132, respectively.
  • the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 24, 66, 126, respectively.
  • the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:18, 71, 134, respectively.
  • the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 25, 74, 135, respectively.
  • the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 26, 75, 136, respectively.
  • the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 25, 77, 138, respectively.
  • the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 25, 78, 139, respectively.
  • the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 25, 82, 143, respectively.
  • the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 29, 84, 143, respectively.
  • the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:32, 86, and 147, respectively.
  • the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:18, 85, 148, respectively.
  • the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 33, 87, 149, respectively.
  • the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 26, 88, and 150, respectively.
  • the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:18, 89, 151, respectively.
  • the heavy chain variable region comprises 1) SEQ ID NO: 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210 , 211, 212, 213, 214, 215, 216 or 217 having an amino acid sequence that is at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%
  • the heavy chain variable region is fused to another molecule.
  • the other molecule is the Fc domain of an immunoglobulin.
  • the other molecule is the Fc domain of human immunoglobulin G1 (IgG1).
  • the DLL3 binding molecule comprises 1) SEQ ID NO: 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257, 258, 259, 260 , 261, 262, 263, 264 or 265; 2) an amino acid sequence having the same amino acid sequence as SEQ ID NO: 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257, 258 8) an amino acid sequence having at least 80%, at least 85%,
  • the DLL3 binding molecule has at least one of the following characteristics:
  • the DLL3 binding molecules include antibodies to DLL3, such as PR013234, PR013235, PR013236, PR013239, PR013240, PR013241, PR013242, PR013244, PR013246, PR013249, PR013250, PR013252, PR013253, PR013254, PR013256, PR013257, PR013258, PR013259, PR013260, PR013261, PR013262, PR013263, PR013264, PR013363, PR013364, 13240, PR013254, PR013371, PR013377, PR013393, and PR013397.
  • the DLL3 antibody PR013241 comprises a heavy chain variable region (VH),
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 179 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 179.
  • the DLL3 antibody PR013241 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013241 comprises the amino acid sequence shown in SEQ ID NO:227 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:227.
  • the antibody comprises a heavy chain variable region, wherein
  • the heavy chain variable region comprises:
  • HCDR2 comprising the amino acid sequence shown in SEQ ID NO:64, and
  • HCDR3 which comprises the amino acid sequence shown in SEQ ID NO:129.
  • the DLL3 antibody PR013242 comprises a heavy chain variable region (VH),
  • the DLL3 antibody PR013242 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013242 comprises the amino acid sequence shown in SEQ ID NO:228 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:228.
  • the present invention provides an antibody PR013244 or an antigen-binding fragment thereof against DLL3,
  • the antibody comprises a heavy chain variable region, wherein
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 181 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 181.
  • the DLL3 antibody PR013244 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013244 comprises the amino acid sequence shown in SEQ ID NO:229 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:229.
  • the present invention provides an antibody PR013246 or an antigen-binding fragment thereof against DLL3,
  • the DLL3 antibody PR013246 comprises a heavy chain variable region (VH),
  • the DLL3 antibody PR013246 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013246 comprises the amino acid sequence shown in SEQ ID NO:230 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:230.
  • the present invention provides an antibody PR013249 or an antigen-binding fragment thereof against DLL3,
  • the antibody comprises a heavy chain variable region, wherein
  • the heavy chain variable region comprises:
  • HCDR2 comprising the amino acid sequence shown in SEQ ID NO:66, and
  • the DLL3 antibody PR013249 comprises a heavy chain variable region (VH),
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 183 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 183.
  • the DLL3 antibody PR013249 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013249 comprises the amino acid sequence shown in SEQ ID NO:231 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:231.
  • the antibody comprises a heavy chain variable region, wherein
  • the heavy chain variable region comprises:
  • HCDR2 comprising the amino acid sequence shown in SEQ ID NO:67, and
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 184 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 184.
  • the DLL3 antibody PR013250 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013250 comprises the amino acid sequence shown in SEQ ID NO:232 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:232.
  • the antibody comprises a heavy chain variable region, wherein
  • HCDR1 which comprises the amino acid sequence shown in SEQ ID NO:18,
  • HCDR3 which comprises the amino acid sequence shown in SEQ ID NO:131.
  • the DLL3 antibody PR013252 comprises a heavy chain variable region (VH),
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 185 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 185.
  • the DLL3 antibody PR013252 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013252 comprises the amino acid sequence shown in SEQ ID NO:233 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:233.
  • the present invention provides an antibody PR013253 or an antigen-binding fragment thereof against DLL3,
  • the heavy chain variable region comprises:
  • HCDR1 which comprises the amino acid sequence shown in SEQ ID NO:19,
  • the DLL3 antibody PR013253 comprises a heavy chain variable region (VH),
  • the DLL3 antibody PR013253 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013253 comprises the amino acid sequence shown in SEQ ID NO:234 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:234.
  • the present invention provides an antibody PR013254 or an antigen-binding fragment thereof against DLL3,
  • the antibody comprises a heavy chain variable region, wherein
  • the DLL3 antibody PR013254 comprises a heavy chain variable region (VH),
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 187 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 187.
  • the DLL3 antibody PR013254 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013254 comprises the amino acid sequence shown in SEQ ID NO:235 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:235.
  • the present invention provides an antibody PR013256 or an antigen-binding fragment thereof against DLL3,
  • the heavy chain variable region comprises:
  • HCDR1 which comprises the amino acid sequence shown in SEQ ID NO:18,
  • HCDR2 comprising the amino acid sequence shown in SEQ ID NO:68, and
  • HCDR3 which comprises the amino acid sequence shown in SEQ ID NO:126.
  • the DLL3 antibody PR013256 comprises a heavy chain variable region (VH),
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 188 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 188.
  • the DLL3 antibody PR013256 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013256 comprises the amino acid sequence shown in SEQ ID NO:236 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:236.
  • the present invention provides an antibody PR013257 or an antigen-binding fragment thereof against DLL3,
  • the antibody comprises a heavy chain variable region, wherein
  • the heavy chain variable region comprises:
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 189 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 189.
  • the present invention provides an antibody PR013258 or an antigen-binding fragment thereof against DLL3,
  • the antibody comprises a heavy chain variable region, wherein
  • the heavy chain variable region comprises:
  • HCDR1 which comprises the amino acid sequence shown in SEQ ID NO:24,
  • HCDR2 comprising the amino acid sequence shown in SEQ ID NO:66, and
  • HCDR3 which comprises the amino acid sequence shown in SEQ ID NO:126.
  • the DLL3 antibody PR013258 comprises a heavy chain variable region (VH),
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 190 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 190.
  • the DLL3 antibody PR013258 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013258 comprises the amino acid sequence shown in SEQ ID NO:238 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:238.
  • the present invention provides an antibody PR013259 or an antigen-binding fragment thereof against DLL3,
  • the heavy chain variable region comprises:
  • HCDR1 which comprises the amino acid sequence shown in SEQ ID NO:18,
  • HCDR3 which comprises the amino acid sequence shown in SEQ ID NO:133.
  • the DLL3 antibody PR013259 comprises a heavy chain variable region (VH),
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 191 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 191.
  • the present invention provides an antibody PR013260 or an antigen-binding fragment thereof against DLL3,
  • the antibody comprises a heavy chain variable region, wherein
  • the heavy chain variable region comprises:
  • HCDR1 which comprises the amino acid sequence shown in SEQ ID NO:18,
  • HCDR2 comprising the amino acid sequence shown in SEQ ID NO:70, and
  • the DLL3 antibody PR013260 comprises a heavy chain variable region (VH),
  • the DLL3 antibody PR013260 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013260 comprises the amino acid sequence shown in SEQ ID NO: 240 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 240.
  • the present invention provides an antibody PR013261 or an antigen-binding fragment thereof against DLL3,
  • the antibody comprises a heavy chain variable region, wherein
  • the heavy chain variable region comprises:
  • HCDR1 which comprises the amino acid sequence shown in SEQ ID NO:18,
  • HCDR2 comprising the amino acid sequence shown in SEQ ID NO:71, and
  • HCDR3 which comprises the amino acid sequence shown in SEQ ID NO:134.
  • the DLL3 antibody PR013261 comprises a heavy chain variable region (VH),
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 193 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 193.
  • the DLL3 antibody PR013261 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013261 comprises the amino acid sequence shown in SEQ ID NO:241 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:241.
  • the antibody comprises a heavy chain variable region, wherein
  • HCDR1 which comprises the amino acid sequence shown in SEQ ID NO:25,
  • HCDR2 comprising the amino acid sequence shown in SEQ ID NO:72, and
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 194 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 194.
  • the present invention provides an antibody PR013263 or an antigen-binding fragment thereof against DLL3,
  • the antibody comprises a heavy chain variable region, wherein
  • the heavy chain variable region comprises:
  • HCDR1 which comprises the amino acid sequence shown in SEQ ID NO:18,
  • HCDR2 comprising the amino acid sequence shown in SEQ ID NO:73, and
  • HCDR3 which comprises the amino acid sequence shown in SEQ ID NO:134.
  • the DLL3 antibody PR013263 comprises a heavy chain variable region (VH),
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 195 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 195.
  • the DLL3 antibody PR013263 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013263 comprises the amino acid sequence shown in SEQ ID NO:243 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:243.
  • the present invention provides an antibody PR013264 or an antigen-binding fragment thereof against DLL3,
  • the antibody comprises a heavy chain variable region, wherein
  • HCDR2 comprising the amino acid sequence shown in SEQ ID NO:74, and
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 196 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 196.
  • the DLL3 antibody PR013264 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013264 comprises the amino acid sequence shown in SEQ ID NO:244 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:244.
  • the antibody comprises a heavy chain variable region, wherein
  • the heavy chain variable region comprises:
  • HCDR1 which comprises the amino acid sequence shown in SEQ ID NO:26,
  • HCDR2 comprising the amino acid sequence shown in SEQ ID NO:75, and
  • HCDR3 which comprises the amino acid sequence shown in SEQ ID NO:136.
  • the DLL3 antibody PR013363 comprises a heavy chain variable region (VH),
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 197 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 197.
  • the DLL3 antibody PR013363 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013363 comprises the amino acid sequence shown in SEQ ID NO:245 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:245.
  • the present invention provides an antibody PR013364 or an antigen-binding fragment thereof against DLL3,
  • the heavy chain variable region comprises:
  • HCDR1 which comprises the amino acid sequence shown in SEQ ID NO:27,
  • HCDR2 comprising the amino acid sequence shown in SEQ ID NO:76, and
  • the DLL3 antibody PR013364 comprises a heavy chain variable region (VH),
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 198 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 198.
  • the antibody comprises a heavy chain variable region, wherein
  • the heavy chain variable region comprises:
  • HCDR2 comprising the amino acid sequence shown in SEQ ID NO:77, and
  • HCDR3 which comprises the amino acid sequence shown in SEQ ID NO:138.
  • the DLL3 antibody PR013367 comprises a heavy chain variable region (VH),
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 199 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 199.
  • the DLL3 antibody PR013367 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013367 comprises the amino acid sequence shown in SEQ ID NO:247 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:247.
  • the present invention provides an antibody PR013368 or an antigen-binding fragment thereof against DLL3,
  • the heavy chain variable region comprises:
  • HCDR3 which comprises the amino acid sequence shown in SEQ ID NO:139.
  • the present invention provides an antibody PR013370 or an antigen-binding fragment thereof against DLL3,
  • HCDR1 which comprises the amino acid sequence shown in SEQ ID NO:18,
  • the DLL3 antibody PR013370 comprises a heavy chain variable region (VH),
  • the DLL3 antibody PR013370 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013370 comprises the amino acid sequence shown in SEQ ID NO:249 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:249.
  • the present invention provides an antibody PR013371 or an antigen-binding fragment thereof against DLL3.
  • the antibody comprises a heavy chain variable region, wherein
  • HCDR1 which comprises the amino acid sequence shown in SEQ ID NO:26,
  • HCDR2 comprising the amino acid sequence shown in SEQ ID NO:80, and
  • the DLL3 antibody PR013371 comprises a heavy chain variable region (VH),
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 202 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 202.
  • the DLL3 antibody PR013371 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013371 comprises the amino acid sequence shown in SEQ ID NO: 250 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 250.
  • the antibody comprises a heavy chain variable region, wherein
  • the heavy chain variable region comprises:
  • HCDR1 which comprises the amino acid sequence shown in SEQ ID NO:26,
  • HCDR2 comprising the amino acid sequence shown in SEQ ID NO:81, and
  • HCDR3 which comprises the amino acid sequence shown in SEQ ID NO:142.
  • the DLL3 antibody PR013372 comprises a heavy chain variable region (VH),
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 203 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 203.
  • the DLL3 antibody PR013372 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013372 comprises the amino acid sequence shown in SEQ ID NO:251 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:251.
  • the present invention provides an antibody PR013376 or an antigen-binding fragment thereof against DLL3,
  • the antibody comprises a heavy chain variable region, wherein
  • the heavy chain variable region comprises:
  • HCDR2 comprising the amino acid sequence shown in SEQ ID NO:82, and
  • HCDR3 which comprises the amino acid sequence shown in SEQ ID NO:143.
  • the DLL3 antibody PR013376 comprises a heavy chain variable region (VH),
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 204 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 204.
  • the DLL3 antibody PR013376 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013376 comprises the amino acid sequence shown in SEQ ID NO:252 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:252.
  • the present invention provides an antibody PR013377 or an antigen-binding fragment thereof against DLL3,
  • the antibody comprises a heavy chain variable region, wherein
  • the heavy chain variable region comprises:
  • HCDR1 which comprises the amino acid sequence shown in SEQ ID NO:18,
  • HCDR3 which comprises the amino acid sequence shown in SEQ ID NO:126.
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 205 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 205.
  • the DLL3 antibody PR013377 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013377 comprises the amino acid sequence shown in SEQ ID NO:253 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:253.
  • the present invention provides an antibody PR013378 or an antigen-binding fragment thereof against DLL3,
  • the antibody comprises a heavy chain variable region, wherein
  • HCDR1 which comprises the amino acid sequence shown in SEQ ID NO:28,
  • HCDR2 comprising the amino acid sequence shown in SEQ ID NO:83, and
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 206 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 206.
  • the DLL3 antibody PR013378 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013378 comprises the amino acid sequence shown in SEQ ID NO:254 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:254.
  • the present invention provides an antibody PR013381 or an antigen-binding fragment thereof against DLL3,
  • the antibody comprises a heavy chain variable region, wherein
  • the heavy chain variable region comprises:
  • HCDR1 which comprises the amino acid sequence shown in SEQ ID NO:29,
  • HCDR2 comprising the amino acid sequence shown in SEQ ID NO:84, and
  • HCDR3 which comprises the amino acid sequence shown in SEQ ID NO:143.
  • the DLL3 antibody PR013381 comprises a heavy chain variable region (VH),
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 207 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 207.
  • the DLL3 antibody PR013381 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013381 comprises the amino acid sequence shown in SEQ ID NO:255 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:255.
  • HCDR1 which comprises the amino acid sequence shown in SEQ ID NO:26,
  • HCDR2 comprising the amino acid sequence shown in SEQ ID NO:85, and
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 208 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 208.
  • the present invention provides an antibody PR013384 or an antigen-binding fragment thereof against DLL3,
  • the antibody comprises a heavy chain variable region, wherein
  • the heavy chain variable region comprises:
  • HCDR1 which comprises the amino acid sequence shown in SEQ ID NO:30,
  • HCDR3 which comprises the amino acid sequence shown in SEQ ID NO:136.
  • the DLL3 antibody PR013384 comprises a heavy chain variable region (VH),
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 209 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 209.
  • the DLL3 antibody PR013384 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013384 comprises the amino acid sequence shown in SEQ ID NO:257 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:257.
  • the present invention provides an antibody PR013387 or an antigen-binding fragment thereof against DLL3,
  • HCDR3 which comprises the amino acid sequence shown in SEQ ID NO:146.
  • the DLL3 antibody PR013387 comprises a heavy chain variable region (VH),
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 210 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 210.
  • the present invention provides an antibody PR013392 or an antigen-binding fragment thereof against DLL3,
  • HCDR1 which comprises the amino acid sequence shown in SEQ ID NO:18,
  • the DLL3 antibody PR013392 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013392 comprises the amino acid sequence shown in SEQ ID NO:259 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:259.
  • the heavy chain variable region comprises:
  • HCDR1 which comprises the amino acid sequence shown in SEQ ID NO:32,
  • HCDR2 comprising the amino acid sequence shown in SEQ ID NO:86, and
  • the DLL3 antibody PR013393 comprises a heavy chain variable region (VH),
  • the DLL3 antibody PR013393 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013393 comprises the amino acid sequence shown in SEQ ID NO:260 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:260.
  • the heavy chain variable region comprises:
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 214 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 214.
  • the DLL3 antibody PR013397 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013397 comprises the amino acid sequence shown in SEQ ID NO:262 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:262.
  • the present invention provides an antibody PR013399 or an antigen-binding fragment thereof against DLL3.
  • HCDR3 which comprises the amino acid sequence shown in SEQ ID NO:150.
  • the DLL3 antibody PR013399 comprises a heavy chain variable region (VH),
  • the DLL3 antibody PR013399 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013399 comprises the amino acid sequence shown in SEQ ID NO:263 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:263.
  • the antibody comprises a heavy chain variable region, wherein
  • the DLL3 antibody PR013402 comprises a heavy chain variable region (VH),
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 216 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 216.
  • the DLL3 antibody PR013402 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013402 comprises the amino acid sequence shown in SEQ ID NO:264 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:264.
  • the antibody comprises a heavy chain variable region, wherein
  • HCDR1 which comprises the amino acid sequence shown in SEQ ID NO:34,
  • HCDR2 comprising the amino acid sequence shown in SEQ ID NO:90, and
  • the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 217 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 217.
  • modification refers to the presence of changes to a natural amino acid, non-natural amino acid, natural amino acid polypeptide, or non-natural amino acid polypeptide. Such changes or modifications can be obtained by post-synthetic modification of a natural amino acid, non-natural amino acid, natural amino acid polypeptide, or non-natural amino acid polypeptide; or by co-translational or post-translational modification of a natural amino acid, non-natural amino acid, natural amino acid polypeptide, or non-natural amino acid polypeptide.
  • the invention provides a multispecific antigen-binding construct comprising a first antigen-binding moiety that binds DLL3, wherein the first antigen-binding moiety comprises the DLL3-binding molecule of the first aspect of the invention.
  • the multispecific antigen-binding construct comprises a first antigen-binding moiety that binds DLL3 and a second antigen-binding moiety that binds a second antigen, wherein the first antigen-binding moiety comprises the DLL3-binding molecule of the first aspect of the invention.
  • first antigen-binding portion and “second antigen-binding portion” refer to amino acid sequences that contain an antigen-binding site and are capable of binding to an antigen epitope, and are defined within the meaning of an antibody or an antigen-binding fragment.
  • the second antigen binding portion can be an antibody or antigen binding fragment that binds to any antigenic epitope of interest.
  • the second antigen is an antigen other than DLL3.
  • Antigens that the second antigen binding portion can specifically bind to may include: tumor antigens (e.g., tumor-associated antigens and tumor-specific antigens), immunomodulatory receptors, and immune checkpoint molecules.
  • tumor antigens e.g., tumor-associated antigens and tumor-specific antigens
  • immunomodulatory receptors e.g., immunomodulatory receptors, and immune checkpoint molecules.
  • tumor antigens e.g., tumor-associated antigens and tumor-specific antigens
  • immunomodulatory receptors e.g., antigen-associated antigens and tumor-specific antigens
  • immune checkpoint molecules e.g., tumor-associated antigens and tumor-specific antigens
  • tumor-associated antigen refers to an antigen that is highly expressed in tumor cells and also present in healthy cells but
  • the second antigen binding portion specifically binds SIRP ⁇ , PD-1, PD-L1, LAG3, TIM-3, CTLA-4, VISTA, GPC3, EGFR, HER-2, CD19, CD20, CD33, CD40, CD73, OX40, CD3, TIP-1, folate receptor alpha (FOLR1) and/or other antigens.
  • the first antigen binding portion and the second antigen binding portion can optionally be connected by a linker. In some embodiments, the first antigen binding portion and the second antigen binding portion are not connected by a linker. In other embodiments, the first antigen binding portion and the second antigen binding portion are connected by a linker, such as a peptide linker or a chemical bond. Preferably, the first antigen binding portion and the second antigen binding portion are connected by a peptide linker.
  • Exemplary peptide linkers can include but are not limited to polyglycine (G), polyalanine (A), polyserine (S) or a combination thereof, such as GGAS, GGGS, GGGSG or (G 4 S) n , wherein n is an integer of 1-20.
  • the present invention also provides a chimeric antigen receptor (CAR) comprising a DLL3 binding molecule of the present invention (e.g., an anti-DLL3 antibody of the present invention or an antigen-binding fragment thereof).
  • CAR chimeric antigen receptor
  • the term "chimeric antigen receptor (CAR)" refers to a polypeptide comprising at least an extracellular domain, a transmembrane domain, and an intracellular T cell receptor activation signaling domain, wherein the extracellular domain binds to or specifically binds to a target on a monospecific or multispecific antibody through a monospecific or multispecific antibody.
  • CAR is a recombinant receptor that simultaneously provides antigen binding and activates T cell function.
  • CAR structure and engineering are described in, for example, Dotti G. et al., (2014) Immunol Rev. 257(1):107-126, which is incorporated herein by reference.
  • the extracellular region comprises a DLL3 binding molecule of the invention (e.g., an anti-DLL3 antibody of the invention or an antigen-binding fragment thereof), such that the CAR of the invention binds to cells expressing DLL3 (e.g., cancer cells).
  • a DLL3 binding molecule of the invention e.g., an anti-DLL3 antibody of the invention or an antigen-binding fragment thereof
  • the CAR of the invention binds to cells expressing DLL3 (e.g., cancer cells).
  • the intracellular signaling region further comprises at least one co-stimulatory domain.
  • the co-stimulatory domain can promote the activation of CAR-expressing immune effector cells after binding to the antigen targeted by the extracellular region.
  • the present invention also relates to a polynucleotide encoding the CAR of the present invention and an expression vector comprising the polynucleotide.
  • the present invention also provides an immune effector cell that expresses the CAR of the present invention on the cell surface.
  • the immune effector cell is selected from T lymphocytes (e.g., cytotoxic T cells (CTL)), macrophages, natural killer cells (NK) and natural killer T cells (NKT).
  • TTL cytotoxic T cells
  • NK natural killer cells
  • NKT natural killer T cells
  • Immune effector cells can target DLL3-positive diseased cells (e.g., DLL3-positive cancer cells) and be activated to start effector functions, for example, causing the death of DLL3-positive cancer cells.
  • the present invention further provides an antibody conjugate comprising a DLL3-binding molecule of the invention (e.g., an anti-DLL3 antibody or antigen-binding fragment thereof) or a multispecific antigen-binding construct of the invention conjugated to at least one therapeutic agent.
  • An antibody-drug conjugate is a typical antibody conjugate, wherein the therapeutic agent can be, for example, a cytotoxic agent.
  • the therapeutic agent can be selected from cytotoxic agents, therapeutic antibodies (e.g., antibodies or antigen-binding fragments thereof that specifically bind to another antigen), radioactive isotopes, oligonucleotides and their analogs (e.g., interfering RNA), biologically active peptides, protein toxins (e.g., diphtheria toxin, ricin toxin), and enzymes (e.g., urease).
  • therapeutic antibodies e.g., antibodies or antigen-binding fragments thereof that specifically bind to another antigen
  • radioactive isotopes e.g., oligonucleotides and their analogs (e.g., interfering RNA), biologically active peptides, protein toxins (e.g., diphtheria toxin, ricin toxin), and enzymes (e.g., urease).
  • therapeutic antibodies e.g., antibodies or antigen-binding fragments thereof that specifically bind to another
  • Cytotoxic agents refer to substances that inhibit or reduce the activity, function and/or kill cells.
  • examples of cytotoxic agents may include, but are not limited to, maytansinoids (e.g., maytansine), auristatins (e.g., MMAF, MMAE, MMAD), duostatins, cryptophycins, vinca alkaloids (e.g., vinblastine, vincristine), colchicines, dolastatins, taxanes, paclitaxel, docetaxel, cabazitaxel, enediyne antibiotics, cytochalasins, camptothecins, anthracyclines (e.g., daunorubicin, dihydroxyanthracindione, doxorubicin), cytotoxic antibiotics (e.g., mitomycin, actinomycin), dactinomycin, dactin, dactycin, dactin ... [0013] Examples of the
  • the therapeutic agent is selected from cytotoxic agents, chemotherapeutic agents, radioisotopes, immune checkpoint inhibitors, antibodies targeting tumor-specific antigens, and other anti-tumor drugs.
  • the therapeutic agent is a cytotoxic agent.
  • the therapeutic agent is a radioisotope.
  • the therapeutic agent is an immune checkpoint inhibitor.
  • the therapeutic agent can be conjugated to the DLL3-binding molecule of the invention (e.g., an anti-DLL3 antibody or antigen-binding fragment thereof) or the multispecific antigen-binding construct of the invention via a linker using any technique known in the art.
  • the linker can comprise a reactive group for covalent conjugation, such as an amine, hydroxylamine, maleimido, carboxyl, phenyl, thiol, sulfhydryl, or hydroxyl group.
  • the present invention also provides a pharmaceutical composition
  • a DLL3 binding molecule of the present invention e.g., an anti-DLL3 antibody or antigen-binding fragment thereof of the present invention
  • a multispecific antigen-binding construct e.g., an immune effector cell or an antibody conjugate, and a pharmaceutically acceptable carrier.
  • suitable carriers can be selected from a buffer (e.g., citrate buffer, acetate buffer, phosphate buffer, histidine buffer, histidine salt buffer), an isotonic agent (e.g., trehalose, sucrose, mannitol, sorbitol, lactose, glucose), a non-ionic surfactant (e.g., polysorbate 80, polysorbate 20, poloxamer), or a combination thereof.
  • a buffer e.g., citrate buffer, acetate buffer, phosphate buffer, histidine buffer, histidine salt buffer
  • an isotonic agent e.g., trehalose, sucrose, mannitol, sorbitol, lactose, glucose
  • a non-ionic surfactant e.g., polysorbate 80, polysorbate 20, poloxamer
  • compositions provided herein may be in a variety of dosage forms, including but not limited to solid, semisolid, liquid, powder, or lyophilized forms.
  • preferred dosage forms include, for example, injection solutions and lyophilized powders.
  • compositions provided herein can be administered to a subject by any method known in the art, for example, by systemic or topical administration.
  • Routes of administration include, but are not limited to, parenteral (e.g., intravenous, intraperitoneal, intradermal, intramuscular, subcutaneous, or intracavitary), topical (e.g., intratumoral), epidural, or mucosal (e.g., intranasal, oral, vaginal, rectal, sublingual, or topical).
  • parenteral e.g., intravenous, intraperitoneal, intradermal, intramuscular, subcutaneous, or intracavitary
  • topical e.g., intratumoral
  • epidural e.g., intranasal, oral, vaginal, rectal, sublingual, or topical
  • the pharmaceutical composition is suitable for intravenous, intramuscular, subcutaneous, parenteral, spinal, or epidermal administration (e.g., by injection or infusion).
  • the exact dosage administered will depend on various factors, such as the metabolic kinetic properties of the pharmaceutical composition, the duration of treatment, the excretion rate of the specific compound, the purpose of the treatment, the route of administration, and the condition of the subject, such as the patient's age, health status, weight, sex, diet, medical history, and other factors well known in the medical field.
  • the DLL3-binding molecules of the invention e.g., anti-DLL3 antibodies or antigen-binding fragments thereof), multispecific antigen-binding constructs, or antibody conjugates can be administered at a dosage ranging from about 0.0001 to 100 mg/kg, more typically 0.01 to 20 mg/kg of the subject's body weight.
  • the dosage can be 0.3 mg/kg, 1 mg/kg, 3 mg/kg, 5 mg/kg, 10 mg/kg, or 20 mg/kg, or within the range of 1-20 mg/kg.
  • Exemplary treatment regimens entail dosing once a week, once every two weeks, once every three weeks, once every four weeks, once a month, once every three months, once every three to six months, or initially with a shorter dosing interval followed by a longer dosing interval.
  • the present invention relates to use of a DLL3 binding molecule (e.g., an anti-DLL3 antibody or antigen-binding fragment thereof), a multispecific antigen-binding construct, an immune effector cell, an antibody conjugate, or a pharmaceutical composition of the invention in the preparation of a medicament for treating a disease in a subject.
  • a DLL3 binding molecule e.g., an anti-DLL3 antibody or antigen-binding fragment thereof
  • a multispecific antigen-binding construct e.g., an immune effector cell, an antibody conjugate, or a pharmaceutical composition of the invention in the preparation of a medicament for treating a disease in a subject.
  • the present invention also relates to the DLL3 binding molecules of the present invention (eg, the anti-DLL3 antibodies or antigen-binding fragments thereof), multispecific antigen-binding constructs, immune effector cells, antibody conjugates or pharmaceutical compositions of the present invention for use in treating diseases.
  • DLL3 binding molecules of the present invention eg, the anti-DLL3 antibodies or antigen-binding fragments thereof
  • multispecific antigen-binding constructs eg. the antigen-binding constructs, immune effector cells, antibody conjugates or pharmaceutical compositions of the present invention for use in treating diseases.
  • the present invention also provides a method of treating a disease in a subject, comprising administering to the subject a therapeutically effective amount of a DLL3-binding molecule of the invention (e.g., an anti-DLL3 antibody or antigen-binding fragment thereof), a multispecific antigen-binding construct, an immune effector cell, an antibody conjugate, or a pharmaceutical composition.
  • a DLL3-binding molecule of the invention e.g., an anti-DLL3 antibody or antigen-binding fragment thereof
  • a multispecific antigen-binding construct e.g., an immune effector cell, an antibody conjugate, or a pharmaceutical composition.
  • a "therapeutically effective dose” refers to a dose that results in a decrease in the severity of disease symptoms, an increase in the frequency and duration of asymptomatic periods of the disease, or prevents damage or disability caused by the suffering of the disease.
  • a therapeutically effective dose is used for anti-proliferative effects, preventing further tumor development, reducing tumor size, reducing tumor vascularity, reducing the number of cancer cells, inhibiting, delaying or reducing tumor and/or malignant cell growth and/or metastasis in cancer patients, and/or reducing one or more symptoms associated with the disease that can be observed.
  • the therapeutically effective dose can vary depending on many different factors, including the mode of administration, the target site, the patient's physiological state, whether the patient is human or other animal, other drugs administered, and whether the treatment is prophylactic or therapeutic.
  • the patient is human, but non-human mammals, including transgenic animals, can also be treated.
  • the therapeutic dose can be titrated using conventional methods known to those skilled in the art to optimize safety and efficacy.
  • a "therapeutically effective dose" of a DLL3 binding molecule e.g., an anti-DLL3 antibody or antigen-binding fragment thereof), multispecific antigen-binding construct, immune effector cell, antibody conjugate, or pharmaceutical composition of the invention preferably inhibits cell growth or tumor growth by at least about 10%, preferably at least about 20%, more preferably at least about 30%, more preferably at least about 40%, more preferably at least about 50%, more preferably at least about 60%, more preferably at least about 70%, and more preferably at least about 80%.
  • the ability to inhibit tumor growth can be evaluated in animal model systems predictive of efficacy against human tumors.
  • an effective amount of a DLL3 binding molecule e.g., an anti-DLL3 antibody or antigen-binding fragment thereof
  • multispecific antigen-binding construct e.g., immune effector cell, antibody conjugate, or pharmaceutical composition of the invention is capable of reducing tumor size or otherwise alleviating symptoms in a subject, such as preventing and/or treating metastasis or recurrence.
  • Such an amount can be determined by one skilled in the art based on factors such as the size of the subject, the severity of the subject's symptoms, and the specific composition or route of administration selected.
  • the disease is associated with abnormal DLL3 expression.
  • abnormal expression refers to excessively high or low protein expression levels in a sample compared to a normal sample (or a standard sample, such as a sample from a subject not suffering from a disease associated with abnormal DLL3 expression).
  • the disease is characterized by high DLL3 expression.
  • DLL3 is highly expressed in tissues (e.g., tumor tissue and adjacent tissues) of subjects suffering from or suspected of suffering from a disease (e.g., small cell lung cancer), while DLL3 is low in the corresponding tissues of subjects not suffering from the disease.
  • the disease as described above is cancer.
  • the cancer is a DLL3-positive tumor.
  • the cancer is selected from small cell lung cancer (SCLC); large cell neuroendocrine carcinoma (LCNEC); neuroendocrine tumors of various tissues including the kidney, genitourinary tract (e.g., bladder, prostate, ovary, cervix, and endometrium), gastrointestinal tract (e.g., stomach, colon), thyroid (e.g., medullary thyroid carcinoma), pancreas, and lung; glioma or pseudoneuroendocrine tumor (pNET).
  • SCLC small cell lung cancer
  • LNEC large cell neuroendocrine carcinoma
  • pNET pseudoneuroendocrine tumor
  • the DLL3-binding molecules of the invention e.g., anti-DLL3 antibodies or antigen-binding fragments thereof
  • multispecific antigen-binding constructs e.g., immune effector cells, antibody conjugates, or pharmaceutical compositions of the invention
  • other treatment methods including but not limited to surgery, chemotherapy, radiotherapy, targeted therapy, immunotherapy, hormone therapy, angiogenesis inhibition, and palliative care.
  • the DLL3 binding molecules of the present invention are further used in combination with one or more therapeutic agents selected from the group consisting of chemotherapeutic agents, radioisotopes, immune checkpoint inhibitors, and tumor antigen-targeted drugs.
  • chemotherapeutic agents may include, for example, antimetabolites, alkylating agents, cytotoxic agents, topoisomerase inhibitors, and microtubule inhibitors.
  • Tumor antigen-targeted drugs include, but are not limited to, drugs that target tumor-associated antigens and tumor-specific antigens.
  • therapeutic agents may include, for example, angiogenesis inhibitors, heparin deacetylase (HDAC) inhibitors, hedgehog signaling pathway blockers, mTOR inhibitors, p53/mdm2 inhibitors, PARP inhibitors, proteasome inhibitors (e.g., bortezomib, carfilzomib, ixazomib, marizomib, oprozomib), and tyrosine kinase inhibitors (e.g., BTK inhibitors).
  • HDAC heparin deacetylase
  • the DLL3 binding molecules e.g., anti-DLL3 antibodies or antigen-binding fragments thereof), multispecific antigen-binding constructs, immune effector cells, antibody conjugates, or pharmaceutical compositions of the present invention are used in combination with a radioisotope.
  • the DLL3 binding molecules e.g., anti-DLL3 antibodies or antigen-binding fragments thereof
  • multispecific antigen-binding constructs, immune effector cells, antibody conjugates, or pharmaceutical compositions of the present invention are used in combination with a tumor antigen-targeted drug.
  • the present invention provides a polynucleotide encoding a DLL3 binding molecule of the present invention (eg, an anti-DLL3 antibody or antigen-binding fragment thereof of the present invention) or a multispecific antigen-binding construct.
  • the present invention also provides an expression vector comprising a polynucleotide of the present invention.
  • the expression vector may further comprise additional polynucleotide sequences, such as regulatory sequences and antibiotic resistance genes.
  • the polynucleotide of the present invention may be present in one or more expression vectors.
  • the polynucleotide of the present invention is prepared as a recombinant nucleic acid. Recombinant nucleic acids can be prepared using techniques well known in the art, such as chemical synthesis, DNA recombination techniques (e.g., polymerase chain reaction (PCR) technology), etc.
  • PCR polymerase chain reaction
  • the present invention further provides a host cell comprising a polynucleotide or expression vector of the present invention.
  • the polynucleotide or expression vector of the present invention can be introduced into a suitable host cell using various methods known in the art. Such methods include, but are not limited to, liposome transfection, electroporation, viral transduction, and calcium phosphate transfection.
  • host cells are used to express the DLL3-binding molecules of the invention (e.g., anti-DLL3 antibodies or antigen-binding fragments thereof) or multispecific antigen-binding constructs of the invention.
  • host cells include, but are not limited to, prokaryotic cells (e.g., bacteria, such as E. coli) and eukaryotic cells (e.g., yeast, insect cells, mammalian cells).
  • prokaryotic cells e.g., bacteria, such as E. coli
  • eukaryotic cells e.g., yeast, insect cells, mammalian cells.
  • Mammalian host cells suitable for antibody expression include, but are not limited to, human cervical carcinoma cells (HeLa cells), human embryonic kidney cells (HEK cells, such as HEK 293 cells), Chinese hamster ovary (CHO) cells, and other mammalian cells suitable for antibody expression.
  • HeLa cells human cervical carcinoma cells
  • HEK cells human embryonic kidney cells
  • CHO Chinese ham
  • the present invention further provides a method for producing a DLL3-binding molecule of the present invention (e.g., an anti-DLL3 antibody or antigen-binding fragment thereof of the present invention) or a multispecific antigen-binding construct of the present invention, the method comprising:
  • DLL3-binding molecules of the invention e.g., anti-DLL3 antibodies or antigen-binding fragments thereof
  • multispecific antigen-binding constructs of the invention e.g., multispecific antigen-binding constructs of the invention
  • kits comprising a DLL3-binding molecule of the invention (e.g., an anti-DLL3 antibody or antigen-binding fragment thereof), a multispecific antigen-binding construct, an immune effector cell, an antibody conjugate, or a pharmaceutical composition, and instructions for use.
  • the kit may also comprise a suitable container.
  • the kit further comprises a device for administration.
  • Kits generally include a label indicating the intended use and/or instructions for use of the kit contents.
  • label includes any written or recorded material provided on or with the kit, or otherwise accompanying the kit.
  • Example 1 Obtaining a fully human anti-DLL3 HCAb antibody
  • Antibodies specifically binding to DLL3 can be obtained by immunizing experimental animals (e.g., mice, rats, rabbits, sheep, camels, etc.) with the DLL3 antigen.
  • the resulting antibodies are non-human.
  • these molecules need to be humanized using antibody engineering techniques to reduce immunogenicity and improve drugability.
  • the humanization process is technically complex, and humanized molecules often have reduced affinity for the antigen.
  • advances in transgenic technology have enabled the creation of genetically engineered mice that carry a human immunoglobulin repertoire and lack the endogenous murine immune repertoire.
  • the antibodies produced by these transgenic mice possess fully human sequences, eliminating the need for further humanization and significantly improving the efficiency of therapeutic antibody development.
  • HCAb mice (Harbour Antibodies BV, WO2002/085945A3) are transgenic mice carrying a human immunoglobulin repertoire, capable of producing novel heavy-chain-only antibodies that are half the size of traditional IgG antibodies.
  • the antibodies it produces only have human antibody "heavy chain” variable domains and mouse Fc constant domains. Due to the absence of light chains, the antibodies almost solve the problems of light chain mispairing and heterodimerization, making this technology platform capable of developing products that are difficult to achieve with traditional antibody platforms.
  • mice transgenic mice expressing human DLL3-his tags, using recombinant human DLL3-his tagged protein (Acro, #DL3-H52H4).
  • each mouse was injected with 50 ⁇ g of immunogen protein and 0.2 mL of complete Freund's adjuvant (CFA, Sigma, #F5881) into the abdominal cavity, axillary lymph nodes, and inguinal lymph nodes, respectively.
  • CFA complete Freund's adjuvant
  • mouse serum was collected and diluted 10-fold to obtain six concentrations (1:100, 1:1000, 1:10,000, 1:100,000, and 1:1,000,000).
  • ELISA titers were determined using ELISA plates coated with huDLL3-his protein (Acro, #DL3-H52H4), cynoDLL3-his protein (Acro, #DL3-C52H9), or proteins unrelated to DLL3.
  • Flow cytometry was used to analyze the specific reactivity of two concentrations of mouse serum (1:100 and 1:1000) against CHO-K1/humanDLL3 cells, CHO-K1/cynoDLL3 cells, and CHO-K1 blast cells expressing high DLL3 expression.
  • a blank control (PB) consisted of serum from pre-immunized mice.
  • CHO-K1/human DLL3, CHO-K1/cyno DLL3, or CHO-K1 cells were incubated with serially diluted mouse serum for 1 hour at 4°C. After washing twice, the cells were incubated with a secondary antibody, Alexa FluorTM 488 goat anti-mouse IgG Fc ⁇ specific (Jackson, #115-545-071), for 1 hour at 4°C. After washing twice, the cells were resuspended and analyzed by flow cytometry (BD Canto II). CHO-K1 cells served as a background control.
  • the plasmid encoding the HCAb antibody is transfected into mammalian host cells (such as human embryonic kidney cells HEK293), and the purified anti-DLL3 recombinant heavy chain antibody can be obtained using conventional recombinant protein expression and purification techniques.
  • the plasmid was purified using a large extraction kit (Macherey-Nagel, Plasmids were extracted using Xtra Midi (Extra Midi) to improve their purity and quality, and then filtered through a 0.22 ⁇ m filter (millpore). The purified plasmids were used for transfection.
  • HEK293 cells were cultured in FreeStyle TM F17 Expression Medium (Thermo, A1383504).
  • the target protein is then eluted with 0.1 M glycine (pH 3.5), adjusted to neutrality with Tris-HCl (pH 8.0), and concentrated using an ultrafiltration tube (Millipore, UFC901024) to PBS buffer to obtain a purified anti-DLL3 heavy chain antibody solution.
  • the antibody concentration was determined by NanoDrop assay at 280 nm absorbance, and the antibody purity was determined by SEC-HPLC and SDS-PAGE.
  • the plate was washed three times with PBST buffer (pH 7.4, containing 0.05% tween-20). 250 mL of 2% BSA blocking buffer was added, and the plate was incubated at 37°C for 1 hour. The blocking buffer was discarded and the plate was washed three times with PBST buffer (pH 7.4, containing 0.05% Tween-20). Then, 100 mL of the antibody to be tested was added to each well in a 5-fold serial dilution. The plate was incubated at 37°C for 1 hour. An isotype antibody was used as a control.
  • 18 antibodies such as PR013234, PR013235, PR013236, PR013239, PR013240, PR013241, PR013244, PR013246, PR013254, PR013256, PR013257, PR013258, PR013259, PR013261, PR013364, PR013377, PR013392, and PR013402 have excellent binding activity to human DLL3, monkey DLL3, and mouse DLL3, especially PR013364, PR013377, and PR013402, with maximum values and EC50 values comparable to those of the reference antibody PR012621.
  • 11 antibodies have binding activity to human DLL3 and monkey DLL3, but do not bind to mouse DLL3, such as PR013363, PR013367, PR013368, PR013370, PR013371, PR013376, PR013378, PR013382, PR013384, PR013387, and PR013399.
  • PR013372 and PR013394 only bind to human DLL3.
  • Example 3 FACS detection of the binding ability of anti-DLL3 HCAb monoclonal antibody at the cellular level
  • CHO-K1 cell line hamster ovary cell subline
  • human DLL3 CHO-K1/humanDLL3, also referred to as CHO-K1-humanDLL3, Nona Bio
  • CHO-K1 cell line overexpressing cynomolgus monkey DLL3 CHO-K1/cynoDLL3, also referred to as CHO-K1-cynoDLL3, Nona Bio
  • CHO-K1 mother cell line CCL-61) or the cell line SHP-77 (human small cell lung cancer cells) that highly expresses human DLL3 ( CRL-2195), DMS79 (human small cell lung cancer cells) CRL-2049) was used for antibody binding experiments at the cellular level.
  • CHO-K1/humanDLL3 cells, CHO-K1/cynoDLL3 cells, CHO-K1 cells, SHP-77 cells or DMS79 cells were digested and collected, and resuspended in PBS containing 2% FBS. The cell density was adjusted to 1 ⁇ 10 6 cells/mL. 100 ⁇ L cells/well were seeded in a 96-well V-bottom plate (Corning, #3894), followed by the addition of 100 ⁇ L/well of a 5-fold concentration gradient dilution of the antibody to be tested at 2 times the final concentration. The cells were placed at 4°C and incubated in the dark for 2 hours.
  • the PCR plate was sealed and placed in a real-time fluorescence quantitative PCR instrument (Bio-Rad CFX96 PCR System), first incubated at 25°C for 5 minutes, then gradually heated from 25°C to 95°C with a gradient of 0.2°C/0.2 minutes, and the temperature was lowered to 25°C at the end of the test.
  • the FRET scanning mode was used and Bio-Rad CFX Maestro software was used for data analysis and calculation of the sample Tm.
  • the extracellular domain of DLL3 consists of several distinct subdomains:
  • Human DLL3EGF2 aa 274-310
  • Detailed information about the recombinant plasmids overexpressing the DLL3 subdomain is shown in Table 14 and Figure 11A. These plasmids were transiently transfected into HEK239T cells. Cells were harvested 48 hours later, and the transfection efficiency of the recombinant cell lines was determined using a FITC-labeled anti-FLAG antibody. Untransfected HEK93T cells served as a negative control for transfection efficiency. Primary antibody staining was performed using 1 mg/mL DLL3 antibody. hIgG1 was used as an isotype negative reference antibody.
  • the binding of the anti-DLL3 antibody to the transiently transfected cells containing the subdomain was measured by flow cytometry using a fluorescent secondary antibody (Alexa Fluor 647-labeled affinity-purified goat anti-human IgG, specifically reactive with Fc ⁇ fragments, Jackson, #109-605-098, 1:1000 dilution) as a secondary antibody.
  • a fluorescent secondary antibody Alexa Fluor 647-labeled affinity-purified goat anti-human IgG, specifically reactive with Fc ⁇ fragments, Jackson, #109-605-098, 1:1000 dilution
  • FIG 11B shows that recombinant plasmids overexpressing DLL3 subdomains were transiently transfected into HEK293T cells.
  • Figures 12A-12J show that DLL3 antibodies bind to different regions of the extracellular end of human DLL3, and the results are shown in Table 15.
  • PR013234 anti-DLL3 N1003P003A12 HCAb hIgG1(C220S)
  • PR013235 anti-DLL3 N1003P003B12 HCAb hIgG1(C220S)
  • PR013236 anti-DLL3 N1003P003D01 HCAb hIgG1(C220S)
  • PR013240 anti-DLL3 N1003P005G03 HCAb hIgG1(C220S)
  • PR013242 anti-DLL3 N1003P008A11 HCAb hIgG1(C220S)
  • PR013249 anti-DLL3 N1003P013C05 HCAb hIgG1(C220S)
  • PR013253 anti-DLL3 N1003P018C04 HCAb hIgG1(C220S)
  • PR013260 anti-DLL3 N1003P024E01 HCAb hIgG1(C220S)
  • PR013261 anti-DLL3 N1003P024G01 HCAb hIgG1(C220S)
  • PR013263 anti-DLL3 N1003P028F03 HCAb hIgG1(C220S)
  • PR013363 anti-DLL3 N1003P041D05 HCAb hIgG1(C220S)
  • PR013364 anti-DLL3 N1003P041G02 HCAb hIgG1(C220S)
  • PR013368 anti-DLL3 N1003P044G07 HCAb hIgG1(C220S)
  • PR013370 anti-DLL3 N1003P046B02 HCAb hIgG1(C220S)
  • PR013372 anti-DLL3 N1003P046G02 HCAb hIgG1(C220S)
  • PR013376 anti-DLL3 N1003P047C08 HCAb hIgG1(C220S)
  • PR013377 anti-DLL3 N1003P047D04 HCAb hIgG1(C220S)
  • PR013378 anti-DLL3 N1003P047D06 HCAb hIgG1(C220S)
  • PR013381 anti-DLL3 N1003P048B11 HCAb hIgG1(C220S)
  • PR013382 anti-DLL3 N1003P049A11 HCAb hIgG1(C220S)
  • PR013384 anti-DLL3 N1003P051B09 HCAb hIgG1(C220S)
  • PR013387 anti-DLL3 N1003P052G11 HCAb hIgG1(C220S)
  • PR013393 anti-DLL3 N1003P054C07 HCAb hIgG1(C220S)
  • PR013397 anti-DLL3 N1003P054E04 HCAb hIgG1(C220S)
  • PR013406 anti-DLL3 N1003P058H08 HCAb hIgG1(C220S)

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Abstract

Provided are an anti-DLL3 antibody or an antigen-binding fragment, multispecific antigen-binding construct, immune effector cell, antibody conjugate or pharmaceutical composition thereof, and a use thereof. Also provided are a polynucleotide, an expression vector, and a method for producing the anti-DLL3 antibody or the antigen-binding fragment thereof.

Description

DLL3结合分子及其用途DLL3 binding molecules and their uses

相关申请的交叉引用CROSS-REFERENCE TO RELATED APPLICATIONS

本申请要求2024年3月29日提交的中国专利申请号202410381377.X的优先权，其整体援引并入本文。This application claims priority to Chinese patent application No. 202410381377.X filed on March 29, 2024, which is incorporated herein by reference in its entirety.

技术领域Technical Field

本发明属于生物医药领域。具体地，本发明涉及DLL3结合分子及其用途。The present invention belongs to the field of biomedicine. Specifically, the present invention relates to DLL3 binding molecules and uses thereof.

背景技术Background Art

δ样配体3(Delta-Like Ligand 3，DLL3)为一种附着在细胞表面的单次跨膜蛋白，属于Notch配体家族中的一员。与其他Notch配体不同，DLL3通常表达于高尔基体并在过表达时出现在细胞表面。研究发现DLL3在小细胞肺癌(SCLC)和其他神经内分泌肿瘤比如大细胞神经内分泌癌(large cell neuroendocrine carcinoma，LCNEC)中高表达。并且有研究发现，DLL3在黑色素瘤、多形性胶质母细胞瘤、小细胞膀胱癌、转移性去势抵抗性前列腺癌和神经内分泌肺肿瘤等其他神经内分泌来源的肿瘤类型中也有表达。在肿瘤细胞中，DLL3特异性表达于肿瘤细胞表面。相反，只有少数正常细胞类型表达DLL3(例如神经元、胰岛细胞和垂体细胞)，并且在正常细胞中，DLL3的表达仅在细胞质中。Delta-Like Ligand 3 (DLL3) is a single-pass transmembrane protein attached to the cell surface and a member of the Notch ligand family. Unlike other Notch ligands, DLL3 is normally expressed in the Golgi apparatus and appears on the cell surface when overexpressed. Studies have found that DLL3 is highly expressed in small cell lung cancer (SCLC) and other neuroendocrine tumors, such as large cell neuroendocrine carcinoma (LCNEC). Studies have also found that DLL3 is expressed in other tumor types of neuroendocrine origin, such as melanoma, glioblastoma multiforme, small cell bladder cancer, metastatic castration-resistant prostate cancer, and neuroendocrine lung tumors. In tumor cells, DLL3 is specifically expressed on the cell surface. In contrast, only a few normal cell types (such as neurons, pancreatic islet cells, and pituitary cells) express DLL3, and in normal cells, DLL3 expression is confined to the cytoplasm.

DLL3表达谱(肿瘤中高、均质的细胞表面表达，与正常组织亚群中低、细胞质表达)使得能够开发使用DLL3特异性靶向肿瘤细胞(例如SCLC细胞)的疗法。目前有几项正在进行的临床研究在评估这些DLL3特异性药物针对肿瘤(例如SCLC和其他神经内分泌肿瘤)的效力。The DLL3 expression profile (high, homogeneous cell surface expression in tumors versus low, cytoplasmic expression in subsets of normal tissues) enables the development of therapies that specifically target tumor cells (e.g., SCLC cells) with DLL3. Several ongoing clinical studies are evaluating the efficacy of these DLL3-specific drugs against tumors (e.g., SCLC and other neuroendocrine tumors).

临床前和临床上正在探索多种靶向DLL3的方法，包括抗体药物偶联物(ADC)、T细胞接合器(TCE)分子和嵌合抗原受体(CAR)疗法。其中，靶向DLL3的抗体药包括Rova-T(Rovalpituzumab tesirine)，Rova-TTM是艾伯维(AbbVie)公司研发的一种抗体-药物偶联物，靶向小细胞肺癌(SCLC)肿瘤上的DLL3，被内化并释放毒素吡咯并苯并二氮杂卓诱导细胞死亡。TCE分子，如安进公司的tarlatamab、Harpoon Therapeutics公司的HPN328、Boehringer Ingelheim的BI 764532、齐鲁制药的QLS31904、罗氏公司的RO7616789和Phanes Therapeutics公司的PT217均处于临床I期阶段。由安进开发的tarlatamab(AMG 757)经过优化后延长了BiTE分子半衰期，可激活T细胞靶向表达DLL3的肿瘤。针对SCLC患者的AMG 757临床I期研究(NCT03319940)正在进行中。嵌合抗原受体T细胞疗法通过对患者自体T细胞进行基因改造，引导患者的T细胞表达肿瘤抗原的嵌合受体，并将这些细胞重新注入患者体内以攻击和杀死靶细胞。AMG119是一种过继性细胞疗法，通过对患者的自体T细胞进行修饰使其表达靶向DLL3跨膜嵌合受体(CAR)并攻击DLL3阳性细胞。目前，评估AMG 119用于SCLC治疗的安全性、耐受性和有效性的临床I期研究(NCT03392064)正在在进行中。Various approaches targeting DLL3 are being explored in the preclinical and clinical stages, including antibody-drug conjugates (ADCs), T cell engager (TCE) molecules, and chimeric antigen receptor (CAR) therapies. Among these, DLL3-targeting antibody drugs include Rova-T (Rovalpituzumab tesirine), an antibody-drug conjugate developed by AbbVie. Rova-T ™ is an antibody-drug conjugate that targets DLL3 in small cell lung cancer (SCLC) tumors, where it is internalized and releases the toxin pyrrolobenzodiazepine, which induces cell death. TCE molecules such as Amgen's tarlatamab, Harpoon Therapeutics' HPN328, Boehringer Ingelheim's BI 764532, Qilu Pharmaceutical's QLS31904, Roche's RO7616789, and Phanes Therapeutics' PT217 are all in Phase I clinical trials. Tarlatamab (AMG 757), developed by Amgen, has been optimized to extend the half-life of the BiTE molecule, activating T cells to target tumors expressing DLL3. A Phase I clinical study of AMG 757 for SCLC patients (NCT03319940) is ongoing. Chimeric antigen receptor T cell therapy genetically modifies the patient's autologous T cells to guide the patient's T cells to express chimeric receptors for tumor antigens, and then re-injects these cells into the patient's body to attack and kill target cells. AMG119 is an adoptive cell therapy that modifies the patient's autologous T cells to express a transmembrane chimeric receptor (CAR) targeting DLL3 and attack DLL3-positive cells. Currently, a Phase I clinical study (NCT03392064) evaluating the safety, tolerability, and efficacy of AMG 119 for the treatment of SCLC is underway.

作为Notch信号的抑制性配体，DLL3与多种肿瘤有密切联系。尤其是在SCLC中，大量的基础研究和临床研究证据表明DLL3是治疗SCLC的一种极具潜力的干预靶点。因此，探索针对DLL3的抗体药物用于癌症治疗，有望为肿瘤治疗带来更多选择。As an inhibitory ligand for Notch signaling, DLL3 is closely associated with various tumors. In SCLC, in particular, extensive basic and clinical research evidence indicates that DLL3 is a promising therapeutic target. Therefore, exploring antibody drugs targeting DLL3 for cancer treatment is expected to provide more options for cancer treatment.

发明内容Summary of the Invention

在第一方面，本发明提供一种DLL3结合分子，其包含重链可变区，其中所述重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1包含SEQ ID NO:18或32所示的序列或者与SEQ ID NO:18或32所示的序列存在不超过2个氨基酸的氨基酸添加、缺失或取代的差异的氨基酸序列；和/或HCDR2包含SEQ ID NO:69、74、79、80、87或90所示的序列或者与SEQ ID NO:69、74、79、80、87或90所示的序列存在不超过2个氨基酸的氨基酸添加、缺失或取代的差异的氨基酸序列；和/或HCDR3包含SEQ ID NO:126、127、128、129、133、135、136、137、138、139、140、141、142、143、144、146、147、148、149、150、151或152所示的序列或者与SEQ ID NO:126、127、128、129、133、135、136、137、138、139、140、141、142、143、144、146、147、148、149、150、151或152所示的序列存在不超过2个氨基酸的氨基酸添加、缺失或取代的差异的氨基酸序列。In a first aspect, the present invention provides a DLL3 binding molecule comprising a heavy chain variable region, wherein the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1 comprises the sequence shown in SEQ ID NO: 18 or 32 or an amino acid sequence that differs from the sequence shown in SEQ ID NO: 18 or 32 by no more than 2 amino acids in amino acid addition, deletion or substitution; and/or HCDR2 comprises the sequence shown in SEQ ID NO: 69, 74, 79, 80, 87 or 90 or an amino acid sequence that differs from the sequence shown in SEQ ID NO: 69, 74, 79, 80, 87 or 90 by no more than 2 amino acids in amino acid addition, deletion or substitution. Sequence; and/or HCDR3 comprises the sequence shown in SEQ ID NO: 126, 127, 128, 129, 133, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 146, 147, 148, 149, 150, 151 or 152, or an amino acid sequence that differs from the sequence shown in SEQ ID NO: 126, 127, 128, 129, 133, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 146, 147, 148, 149, 150, 151 or 152 by amino acid addition, deletion or substitution of no more than 2 amino acids.

在一实施方案中，所述重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、60、126所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:19、60、126所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、61、126所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:20、62、127所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、63、128所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:21、64、129所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:22、65、126所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:23、65、126所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、66、126所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、64、131所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:19、68、126所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、68、126所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:20、69、132所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:24、66、126所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、70、133所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、70、134所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、71、134所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:25、72、135所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、73、134所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:25、74、135所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:26、75、136所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:27、76、137所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:25、77、138所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:25、78、139所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、79、140所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:26、80、141所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:26、81、142所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:25、82、143所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:28、83、144所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:29、84、143所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:26、85、145所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:30、75、136所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:31、85、146所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:32、86、147所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、85、148所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:33、87、149所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:26、88、150所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、89、151所示的氨基酸序列；或者HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:34、90、152所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 18, 60, 126, respectively; or HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 19, 60, 126, respectively; or HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 18, 61, 126, respectively; or HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 20, 62, 127, respectively; or HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NOs: 18, 63, and 128, respectively; or HCDR1, HCDR2, and HCDR3 comprise the amino acid sequences shown in SEQ ID NOs: 21, 64, and 129, respectively; or HCDR1, HCDR2, and HCDR3 comprise the amino acid sequences shown in SEQ ID NOs: 22, 65, and 126, respectively; or HCDR1, HCDR2, and HCDR3 comprise the amino acid sequences shown in SEQ ID NOs: 23, 65, and 126, respectively; or HCDR1, HCDR2, and HCDR3 comprise the amino acid sequences shown in SEQ ID NOs: 18, 66, and 126, respectively; or HCDR1, HCDR2, and HCDR3 comprise the amino acid sequences shown in SEQ ID NOs: 18, 66, and 126, respectively. R2 and HCDR3 contain the amino acid sequences shown in SEQ ID NO: 18, 64, 131, respectively; or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NO: 19, 68, 126, respectively; or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NO: 18, 68, 126, respectively; or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NO: 20, 69, 132, respectively; or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NO: 24, 66, 126, respectively; or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NO: 24, 66, 126, respectively CDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NOs: 18, 70, and 133, respectively; or HCDR1, HCDR2, and HCDR3 comprise the amino acid sequences shown in SEQ ID NOs: 18, 70, and 134, respectively; or HCDR1, HCDR2, and HCDR3 comprise the amino acid sequences shown in SEQ ID NOs: 18, 71, and 134, respectively; or HCDR1, HCDR2, and HCDR3 comprise the amino acid sequences shown in SEQ ID NOs: 25, 72, and 135, respectively; or HCDR1, HCDR2, and HCDR3 comprise the amino acid sequences shown in SEQ ID NOs: 18, 73, and 134, respectively; or HCDR1 , HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NOs: 25, 74, and 135, respectively; or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NOs: 26, 75, and 136, respectively; or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NOs: 27, 76, and 137, respectively; or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NOs: 25, 77, and 138, respectively; or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NOs: 25, 78, and 139, respectively; or HCD R1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NOs: 18, 79 and 140, respectively; or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NOs: 26, 80 and 141, respectively; or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NOs: 26, 81 and 142, respectively; or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NOs: 25, 82 and 143, respectively; or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NOs: 28, 83 and 144, respectively; or H CDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NOs: 29, 84 and 143, respectively; or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NOs: 26, 85 and 145, respectively; or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NOs: 30, 75 and 136, respectively; or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NOs: 31, 85 and 146, respectively; or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NOs: 32, 86 and 147, respectively; or or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NO: 18, 85, 148, respectively; or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NO: 33, 87, 149, respectively; or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NO: 26, 88, 150, respectively; or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NO: 18, 89, 151, respectively; or HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NO: 34, 90, 152, respectively.

在一实施方案中，所述重链可变区包含1)SEQ ID NO:174、175、176、177、178、179、180、181、182、183、185、186、187、188、189、190、191、192、193、194、195、196、197、198、199、200、201、202、203、204、205、206、207、208、209、210、211、212、213、214、215、216或217的氨基酸序列；2)与SEQ ID NO:174、175、176、177、178、179、180、181、182、183、185、186、187、188、189、190、191、192、193、194、195、196、197、198、199、200、201、202、203、204、205、206、207、208、209、210、211、212、213、214、215、216或217的氨基酸序列具有至少80％、至少85％、至少90％、至少91％、至少92％、至少93％、至少94％、至少95％、至少96％、至少97％、至少98％、至少99％序列相同性的氨基酸序列；或者3)与SEQ ID NO:174、175、176、177、178、179、180、181、182、183、185、186、187、188、189、190、191、192、193、194、195、196、197、198、199、200、201、202、203、204、205、206、207、208、209、210、211、212、213、214、215、216或217的氨基酸序列相比具有一个或多个氨基酸的取代、添加和/或缺失的氨基酸序列，优选地，所述添加、缺失和/或取代不发生在CDR区。In one embodiment, the heavy chain variable region comprises 1) SEQ ID NO: 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 185, 186, 187, 188, 189, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216, or 217; or 3) an amino acid sequence that has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity to the amino acid sequence of SEQ ID NO: 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 185, 186, 187, 188, 190, 201, 202, 203, 204, 205, 206, 207 87, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216 or 217 has one or more amino acid substitutions, additions and/or deletions compared to the amino acid sequence of 87, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216 or 217, preferably, the additions, deletions and/or substitutions do not occur in the CDR regions.

在一实施方案中，所述DLL3结合分子为重链抗体和/或为全人源化抗体。In one embodiment, the DLL3 binding molecule is a heavy chain antibody and/or is a fully humanized antibody.

在一实施方案中，所述DLL3结合分子包含1)SEQ ID NO:222、223、224、225、226、227、228、229、230、231、233、234、235、236、237、238、239、240、241、242、243、244、245、246、247、248、249、250、251、252、253、254、255、256、257、258、259、260、261、262、263、264或265的氨基酸序列；2)与SEQ ID NO:222、223、224、225、226、227、228、229、230、231、233、234、235、236、237、238、239、240、241、242、243、244、245、246、247、248、249、250、251、252、253、254、255、256、257、258、259、260、261、262、263、264或265的氨基酸序列具有至少80％、至少85％、至少90％、至少91％、至少92％、至少93％、至少94％、至少95％、至少96％、至少97％、至少98％、至少99％序列相同性的氨基酸序列；或者3)与SEQ ID NO:222、223、224、225、226、227、228、229、230、231、233、234、235、236、237、238、239、240、241、242、243、244、245、246、247、248、249、250、251、252、253、254、255、256、257、258、259、260、261、262、263、264或265的氨基酸序列相比具有一个或多个氨基酸的取代、添加和/或缺失的氨基酸序列，优选地，所述添加、缺失和/或取代不发生在CDR区。In one embodiment, the DLL3 binding molecule comprises 1) SEQ ID NO: 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257, 258, 259, 260 , 261, 262, 263, 264 or 265; 2) an amino acid sequence having the same amino acid sequence as SEQ ID NO: 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257, 258 , 259, 260, 261, 262, 263, 264 or 265 having at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% sequence identity; or 3) an amino acid sequence that has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% sequence identity to the amino acid sequence of SEQ ID NO: 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 233, 234, 2 264, 265, 266, 267, 268, 269, 270, 271, 272, 273, 274, 275, 276, 277, 278, 279, 280, 281, 282, 283, 284, 285, 286, 287, 288, 289, 300, 301, 302, 303, 304, 305, 306, 307, 308, 309, 310, 311, 312, 313, 314, 315, 316, 317, 318, 319, 320, 321, 322, 323, 324, 325

在第二方面，本发明提供一种多特异性抗原结合构建体，其包含结合DLL3的第一抗原结合部分，其中所述第一抗原结合部分包含本发明第一方面的DLL3结合分子。在一优选实施方案中，多特异性抗原结合构建体包含结合DLL3的第一抗原结合部分以及结合第二抗原的第二抗原结合部分，其中所述第一抗原结合部分包含本发明第一方面的DLL3结合分子。In a second aspect, the invention provides a multispecific antigen-binding construct comprising a first antigen-binding moiety that binds DLL3, wherein the first antigen-binding moiety comprises the DLL3-binding molecule of the first aspect of the invention. In a preferred embodiment, the multispecific antigen-binding construct comprises a first antigen-binding moiety that binds DLL3 and a second antigen-binding moiety that binds a second antigen, wherein the first antigen-binding moiety comprises the DLL3-binding molecule of the first aspect of the invention.

在第三方面，本发明提供一种嵌合抗原受体，其包含本发明第一方面的DLL3结合分子。In a third aspect, the present invention provides a chimeric antigen receptor comprising the DLL3 binding molecule of the first aspect of the invention.

在第四方面，本发明提供一种免疫效应细胞，其在表面表达本发明第三方面的嵌合抗原受体。In a fourth aspect, the present invention provides an immune effector cell expressing the chimeric antigen receptor of the third aspect of the present invention on its surface.

在第五方面，本发明提供一种抗体缀合物，其包含与至少一种治疗剂缀合的本发明第一方面的DLL3结合分子或者本发明第二方面的多特异性抗原结合构建体。In a fifth aspect, the invention provides an antibody conjugate comprising the DLL3 binding molecule of the first aspect of the invention or the multispecific antigen-binding construct of the second aspect of the invention conjugated to at least one therapeutic agent.

在第六方面，本发明提供一种药物组合物，其包含本发明第一方面的DLL3结合分子、本发明第二方面的多特异性抗原结合构建体、本发明第四方面的免疫效应细胞或者本发明第五方面的抗体缀合物，以及药学上可接受的载剂。In a sixth aspect, the present invention provides a pharmaceutical composition comprising the DLL3 binding molecule of the first aspect of the invention, the multispecific antigen-binding construct of the second aspect of the invention, the immune effector cell of the fourth aspect of the invention or the antibody conjugate of the fifth aspect of the invention, and a pharmaceutically acceptable carrier.

在第七方面，本发明提供本发明第一方面的DLL3结合分子、本发明第二方面的多特异性抗原结合构建体、本发明第四方面的免疫效应细胞、本发明第五方面的抗体缀合物或者本发明第六方面的药物组合物在制备用于治疗癌症的药物中的用途。In a seventh aspect, the present invention provides use of the DLL3 binding molecule of the first aspect of the invention, the multispecific antigen-binding construct of the second aspect of the invention, the immune effector cell of the fourth aspect of the invention, the antibody conjugate of the fifth aspect of the invention, or the pharmaceutical composition of the sixth aspect of the invention in the preparation of a medicament for treating cancer.

在第八方面，本发明提供一种多核苷酸，其编码本发明第一方面的DLL3结合分子或者本发明第二方面的多特异性抗原结合构建体。In an eighth aspect, the present invention provides a polynucleotide encoding the DLL3 binding molecule of the first aspect of the invention or the multispecific antigen-binding construct of the second aspect of the invention.

在第九方面，本发明提供一种表达载体，其包含本发明第八方面的多核苷酸。In a ninth aspect, the present invention provides an expression vector comprising the polynucleotide of the eighth aspect of the present invention.

在第十方面，本发明提供一种宿主细胞，其包含本发明第八方面的多核苷酸或本发明第九方面的表达载体。In a tenth aspect, the present invention provides a host cell comprising the polynucleotide of the eighth aspect of the present invention or the expression vector of the ninth aspect of the present invention.

在第十一方面，本发明提供一种产生本发明第一方面的DLL3结合分子或者本发明第二方面的多特异性抗原结合构建体的方法，所述方法包括：In an eleventh aspect, the present invention provides a method of producing the DLL3-binding molecule of the first aspect of the invention or the multispecific antigen-binding construct of the second aspect of the invention, the method comprising:

a)在合适条件下培养本发明第十方面的宿主细胞以表达本发明第一方面的DLL3结合分子或者本发明第二方面的多特异性抗原结合构建体；以及a) culturing the host cell of the tenth aspect of the invention under suitable conditions to express the DLL3 binding molecule of the first aspect of the invention or the multispecific antigen-binding construct of the second aspect of the invention; and

b)从宿主细胞或其培养物分离所述DLL3结合分子或者多特异性抗原结合构建体。b) isolating the DLL3 binding molecule or multispecific antigen-binding construct from the host cell or culture thereof.

附图说明BRIEF DESCRIPTION OF THE DRAWINGS

图1A-1D示出候选抗体与人DLL3蛋白的结合活性。1A-1D show the binding activity of candidate antibodies to human DLL3 protein.

图2A-2D示出候选抗体与食蟹猴DLL3蛋白的结合活性。2A-2D show the binding activity of candidate antibodies to cynomolgus monkey DLL3 protein.

图3A-3D示出候选抗体与小鼠DLL3蛋白的结合活性。3A-3D show the binding activity of candidate antibodies to mouse DLL3 protein.

图4A-4D示出候选抗体与过表达人DLL3的CHO-K1细胞的结合活性。Figures 4A-4D show the binding activity of candidate antibodies to CHO-K1 cells overexpressing human DLL3.

图5A-5D示出候选抗体与过表达食蟹猴DLL3的CHO-K1细胞的结合活性。Figures 5A-5D show the binding activity of candidate antibodies to CHO-K1 cells overexpressing cynomolgus monkey DLL3.

图6A-6D示出候选抗体与CHO-K1母细胞的结合活性。Figures 6A-6D show the binding activity of candidate antibodies to CHO-K1 parent cells.

图7A-7D示出候选抗体与肿瘤细胞SHP-77的结合活性。7A-7D show the binding activity of candidate antibodies to tumor cells SHP-77.

图8A-8D示出候选抗体与肿瘤细胞DMS79的结合活性。8A-8D show the binding activity of candidate antibodies to tumor cells DMS79.

图9A-9B示出候选抗体在过表达人DLL3的CHO-K1细胞上的内吞情况。9A-9B show the internalization of candidate antibodies in CHO-K1 cells overexpressing human DLL3.

图10A-10B示出候选抗体在肿瘤细胞SHP-77上的内吞情况。10A-10B show the internalization of candidate antibodies in SHP-77 tumor cells.

图11A示出人DLL3蛋白胞外段结构域或子结构域的质粒构建信息。图11B示出瞬时转染HEK293T的转染效率。Figure 11A shows the plasmid construction information of the extracellular domain or subdomain of human DLL3 protein. Figure 11B shows the transfection efficiency of transiently transfected HEK293T cells.

图12A-12J示出候选抗体和参照抗体结合人DLL3蛋白的表位区域。12A-12J show the epitope regions to which candidate and reference antibodies bind to human DLL3 protein.

具体实施方式DETAILED DESCRIPTION

定义definition

在本发明中，除非另有说明，否则本文中使用的科学和技术名词具有本领域技术人员所通常理解的含义。并且，本文中所用的蛋白质和核酸化学、分子生物学、细胞和组织培养、微生物学、免疫学相关术语和实验室操作步骤均为相应领域内广泛使用的术语和常规步骤。同时，为了更好地理解本发明，下面提供相关术语的定义和解释。Unless otherwise indicated, scientific and technical terms used herein have the meanings commonly understood by those skilled in the art. Furthermore, terms and laboratory procedures related to protein and nucleic acid chemistry, molecular biology, cell and tissue culture, microbiology, and immunology used herein are those widely used in the respective fields and are standard procedures. To facilitate a better understanding of the present invention, definitions and explanations of relevant terms are provided below.

如本文所用，“至少一个(种)”或“一个(种)或多个(种)”可以表示1、2、3、4、5、6、7、8个(种)或更多个(种)。As used herein, "at least one" or "one or more" may mean 1, 2, 3, 4, 5, 6, 7, 8 or more.

如本文所用，表述“包括”、“包含”、“含有”和“具有”是开放式的，表示包括所列举的元素、步骤或组分但不排除其他未列举的元素、步骤或组分。表述“由……组成”不包括未指定的任何元素、步骤或组分。表述“基本上由……组成”是指范围限于指定的元素、步骤或组分，加上不显著影响要求保护的主题的基本和新颖性质的任选存在的元素、步骤或组分。应当理解，表述“基本上由……组成”和“由……组成”涵盖在表述“包括”的含义之内。As used herein, the expressions "comprises," "comprising," "containing," and "having" are open ended and mean the inclusion of the listed elements, steps, or components but not the exclusion of other unlisted elements, steps, or components. The expression "consisting of excludes any element, step, or component not specified. The expression "consisting essentially of means that the scope is limited to the specified elements, steps, or components, plus optional elements, steps, or components that do not significantly affect the basic and novel properties of the claimed subject matter. It should be understood that the expressions "consisting essentially of" and "consisting of are encompassed within the meaning of the expression "comprising."

如本文所用，多个述及的元素之间的连接术语“和/或”应理解为包括单独的和组合的选项。换言之，“和/或”包括“和”以及“或”。例如，A和/或B包括A、B以及A+B。A、B和/或C包括A、B、C及其任意组合，例如A+B、A+C、B+C以及A+B+C。以“和/或”限定的更多元素以类似方式理解，并且包括其中的任一种及其任意组合。As used herein, the term "and/or" connecting multiple elements should be understood to include both individual and combined options. In other words, "and/or" includes "and" and "or." For example, A and/or B includes A, B, and A+B. A, B, and/or C includes A, B, C, and any combination thereof, such as A+B, A+C, B+C, and A+B+C. More elements qualified with "and/or" are understood in a similar manner and include any one thereof and any combination thereof.

除非另有说明，否则任何数值或数值范围，例如浓度或浓度范围，在任何情况下均应理解为由术语“约”修饰。因此，数值通常包括所述值的±10％。例如，1mg/mL的浓度包括0.9mg/mL至1.1mg/mL。同样的，1％至10％(w/v)的浓度范围包括0.9％(w/v)至11％(w/v)。如本文所用，数值范围的使用明确地包括所有可能的子范围，该范围内的所有单个数值，包括该范围内的整数和分数，除非上下文另外明确指出。Unless otherwise indicated, any numerical value or numerical range, such as concentration or concentration range, is understood to be modified by the term "about" in any case. Thus, numerical values generally include ±10% of the stated value. For example, a concentration of 1 mg/mL includes 0.9 mg/mL to 1.1 mg/mL. Similarly, a concentration range of 1% to 10% (w/v) includes 0.9% (w/v) to 11% (w/v). As used herein, the use of numerical ranges explicitly includes all possible subranges, all individual numerical values within the range, including integers and fractions within the range, unless the context clearly indicates otherwise.

如本文所用，术语“δ样配体3(Delta-Like Ligand 3，DLL3)”是指一种附着在细胞表面的单次跨膜蛋白，属于Notch配体家族中的一员。人DLL3蛋白由619个氨基酸组成，胞外结构域(aa 27-492)含有466个氨基酸，其中包含一个Notch结合DSL结构域(aa176-215)，后跟六个表皮生长因子样结构域(EGF样重复序列)(aa 216-465)。人DLL3的胞外结构域与小鼠DLL3的胞外段(aa 27-488)具有84％的氨基酸相同性。As used herein, the term "Delta-Like Ligand 3 (DLL3)" refers to a single-pass transmembrane protein attached to the cell surface and a member of the Notch ligand family. Human DLL3 protein consists of 619 amino acids, with an extracellular domain (aa 27-492) containing 466 amino acids, including a Notch-binding DSL domain (aa 176-215) followed by six epidermal growth factor-like domains (EGF-like repeats) (aa 216-465). The extracellular domain of human DLL3 shares 84% amino acid identity with the extracellular segment of mouse DLL3 (aa 27-488).

如本文所用，“抗体”指免疫球蛋白或其片段，其通过至少一个抗原结合位点特异性结合抗原表位。抗体涵盖抗体片段。如本文所用，术语“抗体”包括合成抗体、重组产生的抗体、多特异性抗体(例如双特异性抗体)、人抗体、非人抗体、人源化抗体、全人源化抗体、重链抗体、纳米抗体、嵌合抗体、胞内抗体、以及抗体片段，例如但不限于Fab片段、Fab'片段、F(ab’)2片段、Fv片段、二硫键连接的Fv(dsFv)、Fd片段、Fd’片段、单链Fv(scFv)、单链Fab(scFab)、双抗体、抗独特型(抗Id)抗体、或者上述任何抗体的抗原结合片段。本文所提供的抗体包括任何免疫球蛋白类型(例如，IgG、IgM、IgD、IgE、IgA和IgY)、任何类别(例如IgG1、IgG2、IgG3、IgG4、IgA1和IgA2)或亚类(例如，IgG2a和IgG2b)的成员。在优选的实施方案中，本发明的抗体是重链抗体，和/或为全人源化抗体。As used herein, "antibody" refers to an immunoglobulin or a fragment thereof that specifically binds to an antigenic epitope through at least one antigen binding site. Antibody encompasses antibody fragments. As used herein, the term "antibody" includes synthetic antibodies, recombinantly produced antibodies, multispecific antibodies (e.g., bispecific antibodies), human antibodies, non-human antibodies, humanized antibodies, fully humanized antibodies, heavy chain antibodies, nanobodies, chimeric antibodies, intracellular antibodies, and antibody fragments, such as but not limited to Fab fragments, Fab' fragments, F(ab') 2 fragments, Fv fragments, disulfide-linked Fv (dsFv), Fd fragments, Fd' fragments, single-chain Fv (scFv), single-chain Fab (scFab), diabodies, anti-idiotypic (anti-Id) antibodies, or antigen-binding fragments of any of the above antibodies. The antibodies provided herein include members of any immunoglobulin class (e.g., IgG, IgM, IgD, IgE, IgA, and IgY), any class (e.g., IgG1, IgG2, IgG3, IgG4, IgA1, and IgA2), or subclass (e.g., IgG2a and IgG2b). In preferred embodiments, the antibodies of the present invention are heavy chain antibodies, and/or are fully humanized antibodies.

如本文所用，抗体的“抗体片段”或“抗原结合片段”指全长抗体的任何部分，其少于全长，但是至少包含结合抗原的所述抗体的部分可变区(例如一个或多个CDR和/或一个或多个抗体结合位点)，并且因此保留结合特异性以及所述全长抗体的至少部分特异性结合能力。因此，抗原结合片段指包含与衍生抗体片段的抗体结合相同抗原的抗原结合部分的抗体片段。抗体片段包括通过酶促处理全长抗体所产生的抗体衍生物，以及合成产生的衍生物，例如重组产生的衍生物。抗体包括抗体片段。抗体片段的实例包括但不限于Fab、Fab'、F(ab’)2、单链Fv(scFv)、Fv、dsFv、双抗体、Fd和Fd’片段以及其他片段，包括修饰的片段(参见，例如Methods in Molecular Biology,Vol 207:Recombinant Antibodies for Cancer Therapy Methods and Protocols(2003)；Chapter 1；p 3-25,Kipriyanov)。所述片段可以包括连接在一起的多条链，例如通过二硫键和/或通过肽接头。抗体片段一般包含至少或约50个氨基酸，并且典型至少或约200个氨基酸。抗原结合片段包括任何抗体片段，其在被插入抗体框架(例如通过置换相应区域)时获得免疫特异性地结合抗原的抗体。As used herein, an "antibody fragment" or "antigen-binding fragment" of an antibody refers to any portion of a full-length antibody that is less than full-length but contains at least a portion of the variable region (e.g., one or more CDRs and/or one or more antibody binding sites) of the antibody that binds to an antigen and thus retains binding specificity and at least a portion of the specific binding ability of the full-length antibody. Thus, an antigen-binding fragment refers to an antibody fragment that contains an antigen-binding portion that binds to the same antigen as the antibody from which the antibody fragment was derived. Antibody fragments include antibody derivatives produced by enzymatic treatment of full-length antibodies, as well as synthetically produced derivatives, such as recombinantly produced derivatives. Antibodies include antibody fragments. Examples of antibody fragments include, but are not limited to, Fab, Fab', F(ab') 2 , single-chain Fv (scFv), Fv, dsFv, diabodies, Fd and Fd' fragments, and other fragments, including modified fragments (see, e.g., Methods in Molecular Biology, Vol 207: Recombinant Antibodies for Cancer Therapy Methods and Protocols (2003); Chapter 1; p 3-25, Kipriyanov). The fragments can include multiple chains linked together, for example, by disulfide bonds and/or by peptide linkers. Antibody fragments generally contain at least or about 50 amino acids, and typically at least or about 200 amino acids. Antigen-binding fragments include any antibody fragment that, when inserted into an antibody framework (e.g., by replacing the corresponding regions), results in an antibody that immunospecifically binds to an antigen.

如本文所用，术语“仅有重链的抗体(Heavy-chain-only antibody)”和“重链抗体(Heavy-chain antibody)”可互换使用，并且以其最广泛的意义存在，是指缺乏常规抗体轻链的抗体，其仅包含一个重链可变区以及不包含CH1的重链恒定区(例如Fc片段)。As used herein, the terms "heavy-chain-only antibody" and "heavy-chain antibody" are used interchangeably and in their broadest sense to refer to antibodies that lack conventional antibody light chains and contain only a heavy chain variable region and a heavy chain constant region that does not contain CH1 (e.g., Fc fragment).

“Fc片段”一般指常规抗体或重链抗体经木瓜蛋白酶消化的可结晶片段。一般而言，IgG和重链抗体的Fc片段可以包含部分铰链区、CH2和CH3。在本文中，Fc片段可以包含至少部分铰链区(例如铰链区的全部或部分)、CH2和CH3。"Fc fragment" generally refers to a crystallizable fragment of a conventional antibody or heavy chain antibody after papain digestion. Generally speaking, the Fc fragment of IgG and heavy chain antibodies can include part of the hinge region, CH2 and CH3. Herein, the Fc fragment can include at least part of the hinge region (e.g., all or part of the hinge region), CH2 and CH3.

可变区(即“结合结构域”)允许结合分子选择性地识别抗原上的表位并特异性地结合该表位。也就是说，例如抗体的结合分子的轻链可变区(VL)结构域和重链可变区(VH)结构域、或这些互补决定区(CDR)亚组组合形成确定三维抗原结合位点的可变区。更具体地，抗原结合位点由各VH和VL链上的三个CDR确定，例如，本发明的重链抗体，其抗原结合位点由VH链上的三个CDR确定。这些“互补决定区”或“CDR”是氨基酸的非连续短序列，其特异性地定位，以随着抗体在水性环境中采用其三维构型而形成结合结构域。结合结构域中的其余氨基酸称为“构架(FR)”区，显示较小的分子间差异。由定位的CDR形成的结合结构域确定了与免疫反应性抗原上的表位互补的表面。这种互补表面促进了抗体与其互补表位的非共价结合。任何给定的重或轻链可变区中分别组成的CDR和构架区域的氨基酸均可由常规方法鉴定(参见，“免疫学热门蛋白质序列(Sequences of Proteins of Immunological Interest)”，Kabat,E.et al.，美国卫生及公共服务部(U.S.Department of Health and Human Services)，(1983)；和Chothia和Lesk，J.Mol.Biol.,196:901-917(1987)，其通过引用其全文纳入本文)。在本文中，轻链可变区的CDR(CDRL或LCDR)可以称为LCDR1、LCDR2和LCDR3，重链可变区的CDR(CDRH或HCDR)可以称为HCDR1、HCDR2和HCDR3。The variable region (i.e., "binding domain") allows the binding molecule to selectively recognize an epitope on an antigen and specifically bind to the epitope. That is, for example, the light chain variable region (VL) domain and the heavy chain variable region (VH) domain of the binding molecule of an antibody, or these complementary determining regions (CDR) subgroups combine to form a variable region that determines a three-dimensional antigen binding site. More specifically, the antigen binding site is determined by three CDRs on each VH and VL chain. For example, the heavy chain antibody of the present invention has an antigen binding site determined by three CDRs on the VH chain. These "complementary determining regions" or "CDRs" are short, non-continuous sequences of amino acids that are specifically positioned to form a binding domain as the antibody adopts its three-dimensional configuration in an aqueous environment. The remaining amino acids in the binding domain are called "framework (FR)" regions, which show smaller intermolecular differences. The binding domain formed by the positioned CDRs determines a surface that is complementary to the epitope on the immunoreactive antigen. This complementary surface promotes the non-covalent binding of the antibody to its complementary epitope. The amino acids that make up the CDRs and framework regions, respectively, in any given heavy or light chain variable region can be identified by conventional methods (see, "Sequences of Proteins of Immunological Interest," Kabat, E. et al., U.S. Department of Health and Human Services, (1983); and Chothia and Lesk, J. Mol. Biol., 196:901-917 (1987), which are incorporated herein by reference in their entireties). Herein, the CDRs (CDRL or LCDR) of the light chain variable region may be referred to as LCDR1, LCDR2, and LCDR3, and the CDRs (CDRH or HCDR) of the heavy chain variable region may be referred to as HCDR1, HCDR2, and HCDR3.

在本发明中，CDR的氨基酸序列均是按照Chothia定义规则所示出的(本发明的权利要求中也是按照Chothia定义规则所示出的序列)。但是，本领域人员公知，在本领域中可以通过多种方法来定义抗体的CDR，例如基于抗体的三维结构和CDR环的拓扑学的Chothia(参见例如Chothia,C.et al.,Nature,342,877-883(1989)；和Al-Lazikani,B.et al.,J.Mol.Biol.,273,927-948(1997))、基于抗体序列可变性的Kabat(参见例如Kabat,E.A.et al.(1991)Sequences of Proteins of Immunological Interest,Fifth Edition,U.S.Department of Health and Human Services,NIH Publication No.91-3242)、AbM(Martin,A.C.R.and J.Allen(2007)“Bioinformatics tools for antibody engineering,”in S.Dübel(ed.),Handbook of Therapeutic Antibodies.Weinheim:Wiley-VCH Verlag,pp.95–118)、Contact(MacCallum,R.M.et al.,(1996)J.Mol.Biol.262:732-745)、IMGT(Lefranc,M.-P.,2011(6),IMGT,the International ImMunoGeneTics Information System Cold Spring Harb Protoc.；和Lefranc,M.-P.et al.,Dev.Comp.Immunol.,27,55-77(2003))，以及基于利用大量晶体结构的近邻传播聚类(affinity propagation clustering)的North CDR定义。在本文中，对于同一个可变区可以使用多个CDR编号系统，例如Chothia、Abm、Kabat、Contact和IMGT。本领域技术人员应当理解，尽管由不同编号系统定义的CDR可能会不同，但是同一编号系统对应的CDR代表能够结合抗原表位的有效抗原结合位点。除非另有规定，否则术语给定抗体或其区(例如可变区)的“CDR”及“互补决定区”应理解为涵盖如通过本发明描述的上述已知方案中的任何一种界定的互补决定区。虽然本发明的权利要求中请求保护的范围是基于Chothia定义规则所示出的序列，但是根据其他CDR的定义规则所对应的氨基酸序列也应当落在本发明的保护范围中。In the present invention, the amino acid sequences of CDRs are all shown according to the Chothia definition rules (the sequences in the claims of the present invention are also shown according to the Chothia definition rules). However, it is well known to those skilled in the art that the CDR of an antibody can be defined in the art by a variety of methods, such as Chothia based on the three-dimensional structure of the antibody and the topology of the CDR loop (see, for example, Chothia, C. et al., Nature, 342, 877-883 (1989); and Al-Lazikani, B. et al., J. Mol. Biol., 273, 927-948 (1997)), Kabat based on antibody sequence variability (see, for example, Kabat, E.A. et al. (1991) Sequences of Proteins of Immunological Interest, Fifth Edition, U.S. Department of Health and Human Services, NIH Publication No. 91-3242), AbM (Martin, A.C.R. and J. Allen (2007) “Bioinformatics tools for antibody engineering,” in S. Dübel (ed.), Handbook of Therapeutic Antibodies. Weinheim: Wiley-VCH Verlag, pp. 95–118), Contact (MacCallum, R. M. et al., (1996) J. Mol. Biol. 262: 732–745), IMGT (Lefranc, M.-P., 2011 (6), IMGT, the International ImMunoGeneTics Information System Cold Spring Harb Protoc.; and Lefranc, M.-P. et al., Dev. Comp. Immunol., 27, 55–77 (2003)), and North CDR definitions based on affinity propagation clustering using a large number of crystal structures. In this article, multiple CDR numbering systems can be used for the same variable region, such as Chothia, Abm, Kabat, Contact and IMGT. It will be understood by those skilled in the art that although the CDRs defined by different numbering systems may be different, the CDRs corresponding to the same numbering system represent effective antigen binding sites that can bind to antigen epitopes. Unless otherwise specified, the terms "CDR" and "complementarity determining region" of a given antibody or a region thereof (such as a variable region) should be understood to cover the complementary determining region defined by any of the above-mentioned known schemes described by the present invention. Although the scope of protection claimed in the claims of the present invention is based on the sequences shown in the Chothia definition rules, the amino acid sequences corresponding to the definition rules of other CDRs should also fall within the scope of protection of the present invention.

因此，在涉及用本发明定义的具体CDR序列限定抗体时，所述抗体的范围还涵盖了这样的抗体，其可变区序列包含所述的具体CDR序列，但是由于应用了不同的方案(例如不同的指派系统规则或组合)而导致其所声称的CDR边界与本发明所定义的具体CDR边界不同。Thus, when referring to antibodies defined by specific CDR sequences defined herein, the scope of said antibodies also encompasses antibodies whose variable region sequences comprise said specific CDR sequences, but whose declared CDR boundaries differ from the specific CDR boundaries defined herein due to the application of a different scheme (e.g., a different assignment system rule or combination).

如本文所用，术语“框架区”和“构架区”可以互换使用。如本文中所使用的，术语“框架区”、“构架区”或“FR”残基是指抗体可变区中除了如上定义的CDR序列以外的那些氨基酸残基。As used herein, the terms "framework region" and "framework region" are used interchangeably. As used herein, the terms "framework region," "framework region," or "FR" residues refer to those amino acid residues in the antibody variable region excluding the CDR sequences as defined above.

本文所用的术语“二硫键”包括两个硫原子之间形成的共价键。氨基酸半胱氨酸包含可形成二硫键或桥接第二巯基基团的巯基基团。As used herein, the term "disulfide bond" includes a covalent bond formed between two sulfur atoms. The amino acid cysteine contains a sulfhydryl group that can form a disulfide bond or bridge a second sulfhydryl group.

如本文所用，术语“人源化抗体”是指非人抗体经修饰以增加与人抗体的序列同源性的抗体。人源化抗体通常保留其所源自的非人抗体的抗原结合能力并且对于人体具有较低的免疫原性。人源化抗体可以通过抗体工程化改造任何非人物种抗体或其中包含非人物种来源序列的抗体(例如嵌合抗体)来获得。非人物种例如可以包括小鼠、大鼠、兔、羊驼、鲨鱼或非人灵长类动物。由非人抗体获得人源化抗体的技术是本领域技术人员熟知的。例如，将非人抗体(例如鼠源抗体)的CDR序列移植到人抗体框架区中。在某些情况下，为了保持人源化抗体的抗原结合能力和/或稳定性，可以在人抗体框架区中保留非人抗体(例如鼠源抗体)框架序列的关键氨基酸残基，即进行“回复突变”(参见，例如Morrison et al.(1984)Proc.Natl.Acad.Sci.81(21):6851-6855；Neuberger et al.(1984)Nature 312:604-608)。As used herein, the term "humanized antibody" refers to an antibody in which a non-human antibody is modified to increase sequence homology with a human antibody. Humanized antibodies generally retain the antigen-binding ability of the non-human antibody from which they are derived and have lower immunogenicity to the human body. Humanized antibodies can be obtained by engineering any non-human species antibody or an antibody (e.g., a chimeric antibody) comprising a sequence of a non-human species. Non-human species may, for example, include mice, rats, rabbits, alpacas, sharks, or non-human primates. The technology for obtaining humanized antibodies from non-human antibodies is well known to those skilled in the art. For example, the CDR sequences of non-human antibodies (e.g., mouse antibodies) are transplanted into human antibody framework regions. In some cases, in order to maintain the antigen-binding ability and/or stability of the humanized antibody, key amino acid residues of the non-human antibody (e.g., mouse antibody) framework sequence can be retained in the human antibody framework region, i.e., "back mutation" is performed (see, for example, Morrison et al. (1984) Proc. Natl. Acad. Sci. 81(21): 6851-6855; Neuberger et al. (1984) Nature 312: 604-608).

在本发明中，术语“全人源抗体”或“全人源化抗体”通常是指将人编码抗体的基因转移至基因工程改造的抗体基因缺失动物中，使动物表达的抗体。抗体所有部分(包括抗体的可变区和恒定区)均由人类来源的基因所编码。本领域获得全人源抗体的方法可以有噬菌体展示技术、转基因小鼠技术、核糖体展示技术和RNA-多肽技术等。In the present invention, the term "fully human antibody" or "fully humanized antibody" generally refers to an antibody expressed by a genetically engineered animal by transferring a human antibody-encoding gene into an antibody gene-deficient animal. All parts of the antibody (including the variable and constant regions) are encoded by genes of human origin. Methods for obtaining fully human antibodies in the art include phage display technology, transgenic mouse technology, ribosome display technology, and RNA-peptide technology.

如本文所用，氨基酸序列的“百分比(％)序列相同性”、“序列相同性”具有本领域公认的定义，其指通过序列比对(例如通过人工检视或可公知的算法)确定的两个多肽序列之间相同的百分比。可以使用本领域技术人员已知的方法确定，例如使用可公开获得的计算机软件如BLAST、BLAST-2、Clustal Omega和FASTA软件。As used herein, "percent (%) sequence identity" or "sequence identity" of amino acid sequences has the art-recognized definition of the percentage of identity between two polypeptide sequences as determined by sequence alignment (e.g., by manual inspection or a publicly known algorithm). This can be determined using methods known to those skilled in the art, for example, using publicly available computer software such as BLAST, BLAST-2, Clustal Omega, and FASTA software.

在本文中，“源自”或“衍生自”参考氨基酸序列的氨基酸序列与参考氨基酸序列的部分或者全部相同或同源。例如，衍生自人免疫球蛋白的重链恒定区的氨基酸序列可以与其所源自的人免疫球蛋白重链恒定区的野生型序列具有至少80％、至少85％、至少90％、至少91％、至少92％、至少93％、至少94％、至少95％、至少96％、至少97％、至少98％、至少99％或100％的序列相同性。As used herein, an amino acid sequence that is "derived from" or "derived from" a reference amino acid sequence is identical or homologous to part or all of the reference amino acid sequence. For example, an amino acid sequence derived from the heavy chain constant region of a human immunoglobulin may have at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% or 100% sequence identity with the wild-type sequence of the heavy chain constant region of the human immunoglobulin from which it is derived.

可以修饰多肽中的非关键区域(例如抗体的CDR区、框架区的非关键氨基酸、恒定区的氨基酸)，例如进行一个或多个氨基酸的取代、添加和/或缺失，而不改变多肽的功能。在肽或蛋白中，合适的保守氨基酸取代是本领域技术人员已知的，并且一般可以进行而不改变所得分子的生物活性。通常，本领域技术人员认识到多肽的非必需区中的单个氨基酸取代基本上不改变生物活性(参见，例如Watson et al.,Molecular Biology of the Gene,4th Edition,1987,The Benjamin/Cummings Pub.co.,p.224)。Non-critical regions in a polypeptide (e.g., CDR regions of an antibody, non-critical amino acids in the framework regions, amino acids in the constant regions) can be modified, for example, by substitution, addition, and/or deletion of one or more amino acids without altering the function of the polypeptide. Suitable conservative amino acid substitutions in peptides or proteins are known to those skilled in the art and can generally be made without altering the biological activity of the resulting molecule. Generally, those skilled in the art recognize that single amino acid substitutions in non-essential regions of a polypeptide do not substantially alter biological activity (see, e.g., Watson et al., Molecular Biology of the Gene, 4th Edition, 1987, The Benjamin/Cummings Pub.co., p. 224).

“亲和力”或“结合亲和力”用来衡量抗体和抗原之间通过非共价作用相互结合的强度。可以用本领域已知的常规技术测定亲和力，例如生物膜干涉技术(可以采用例如Octet Fortebio检测系统)、放射免疫法、表面等离子共振法、酶联免疫测定(ELISA)或流式细胞术(FACS)等。"Affinity" or "binding affinity" is a measure of the strength of the non-covalent binding between an antibody and an antigen. Affinity can be determined using conventional techniques known in the art, such as membrane interferometry (e.g., using the Octet Fortebio assay system), radioimmunoassay, surface plasmon resonance, enzyme-linked immunosorbent assay (ELISA), or flow cytometry (FACS).

“特异性结合”一般表示结合分子，例如抗体或其片段、变体或衍生物通过其抗原结合结构域结合表位，并且该结合需要抗原结合结构域和表位之间的一些互补性。按这一定义，当结合分子通过其抗原结合结构域与表位结合比其与随机、无关的表位结合更容易时，称该结合分子“特异性结合”该表位。本文中使用术语“特异性”定性分析某一抗体与某一表位结合的相对亲和性。例如，可认为结合分子“A”比结合分子“B”对给定表位具有更高特异性，或者可以说结合分子“A”以比其对相关表位“D”的特异性更高的特异性结合表位“C”。"Specific binding" generally refers to a binding molecule, such as an antibody or a fragment, variant, or derivative thereof, that binds to an epitope through its antigen binding domain, and this binding requires some complementarity between the antigen binding domain and the epitope. By this definition, a binding molecule is said to "specifically bind" to an epitope when it is easier to bind to an epitope through its antigen binding domain than to a random, unrelated epitope. The term "specificity" is used herein to qualitatively analyze the relative affinity of an antibody for binding to an epitope. For example, it can be considered that binding molecule "A" has a higher specificity for a given epitope than binding molecule "B," or it can be said that binding molecule "A" specifically binds to epitope "C" with a higher specificity than its specificity for a related epitope "D."

如本文所用，术语“EC50”，也称为“半数有效浓度”，是指在特定的暴露时间后诱导在基线和最大值之间的50％的应答的药物、抗体或毒剂的浓度。As used herein, the term " EC50 ," also known as "median effective concentration," refers to the concentration of a drug, antibody, or toxic agent that induces a response that is 50% between baseline and maximum after a specified exposure time.

如本文所用，术语“内吞”是指通过细胞膜将待内化的物质包围，然后在细胞内萌芽形成包含摄入物质的囊泡，将外界物质带入细胞的过程，其是细胞质膜运送物质的一种方式。术语“抗体内化”是指细胞表面受体与其特异性抗体结合时，可诱导发生受体介导的内吞作用。如本文所述，DLL3抗体可以通过结合DLL3的胞外部介导细胞表面表达的DLL3蛋白的内化。可以用本领域已知的常规技术测定抗体的内吞效应，例如使用内化抗体标记。As used herein, the term "endocytosis" refers to the process by which a substance to be internalized is enclosed by the cell membrane and then buds into a vesicle containing the internalized substance, bringing the external substance into the cell. This is one way that substances are transported across the cytoplasmic membrane. The term "antibody internalization" refers to the induction of receptor-mediated endocytosis when a cell surface receptor binds to its specific antibody. As described herein, DLL3 antibodies can mediate the internalization of cell-surface-expressed DLL3 protein by binding to the extracellular portion of DLL3. The endocytic effect of antibodies can be measured using conventional techniques known in the art, such as labeling with internalizing antibodies.

如本文所用，术语“多核苷酸”和“核酸”可以互换使用，用于指脱氧核糖核苷酸的聚合物(脱氧核糖核酸，DNA)或核糖核苷酸的聚合物(核糖核酸，RNA)。“多核苷酸序列”、“核酸序列”和“核苷酸序列”可以互换使用，用来表示多核苷酸中核苷酸的排序。本领域人员应当理解，DNA编码链(有义链)与其编码的RNA可以看作具有相同的核苷酸序列，DNA编码链序列中的脱氧胸苷酸对应其编码的RNA序列中的尿苷酸。As used herein, the terms "polynucleotide" and "nucleic acid" are used interchangeably to refer to a polymer of deoxyribonucleotides (deoxyribonucleic acid, DNA) or a polymer of ribonucleotides (ribonucleic acid, RNA). "Polynucleotide sequence," "nucleic acid sequence," and "nucleotide sequence" are used interchangeably to refer to the order of nucleotides in a polynucleotide. It will be understood by those skilled in the art that a DNA coding strand (sense strand) and the RNA it encodes can be considered to have the same nucleotide sequence, with deoxythymidylic acid in the DNA coding strand sequence corresponding to uridine in the RNA sequence it encodes.

如本文所用，分离的核酸分子是从存在于核酸分子的天然来源中的其他核酸分子分离的核酸分子。诸如cDNA分子的“分离的”核酸分子可以在通过重组技术制备时基本上不含其他细胞物质或培养基，或者在化学合成时基本上不含化学前体或其他化学成分。本文所提供的示例性分离的核酸分子包括编码所提供的DLL3结合分子的分离的核酸分子。As used herein, an isolated nucleic acid molecule is one that is separated from other nucleic acid molecules present in the natural source of the nucleic acid molecule. An "isolated" nucleic acid molecule, such as a cDNA molecule, can be substantially free of other cellular material or culture medium when produced by recombinant techniques, or substantially free of chemical precursors or other chemical components when chemically synthesized. Exemplary isolated nucleic acid molecules provided herein include isolated nucleic acid molecules that encode provided DLL3 binding molecules.

如本文所用，术语“表达”包括核苷酸序列的转录和/或翻译。因此，表达可以涉及转录物和/或多肽的产生。As used herein, the term "expression" includes transcription and/or translation of a nucleotide sequence. Thus, expression can involve the production of transcripts and/or polypeptides.

如本文所用，“载体”是用于将外源多核苷酸导入宿主细胞的媒介，当载体转化入适当的宿主细胞时，外源多核苷酸得以扩增或表达。载体通常保持游离，但是可以设计为使基因或其部分整合入基因组的染色体。如本文所用，载体的定义涵盖质粒、线性化质粒、病毒载体、粘粒、噬菌体载体、噬菌粒、人工染色体(例如，酵母人工染色体和哺乳动物人工染色体)等。病毒载体包括但不限于逆转录病毒载体(包括慢病毒载体)、腺病毒载体、腺相关病毒载体、疱疹病毒载体、痘病毒载体和杆状病毒载体等。As used herein, "vector" is a medium for introducing exogenous polynucleotides into a host cell, and when the vector is transformed into an appropriate host cell, the exogenous polynucleotides are amplified or expressed. The vector usually remains free, but can be designed to integrate a gene or part thereof into a chromosome of the genome. As used herein, the definition of vector encompasses plasmids, linearized plasmids, viral vectors, cosmids, phage vectors, phagemids, artificial chromosomes (e.g., yeast artificial chromosomes and mammalian artificial chromosomes), etc. Viral vectors include, but are not limited to, retroviral vectors (including lentiviral vectors), adenoviral vectors, adeno-associated viral vectors, herpes virus vectors, pox virus vectors, and baculovirus vectors, etc.

如本文所用，“宿主细胞”是用于接受、保持、复制和扩增载体的细胞。宿主细胞还可以用来表达载体所编码的多肽。当宿主细胞分裂时，载体中所含的核酸复制，从而扩增核酸。宿主细胞可以是真核细胞或原核细胞。合适的宿主细胞包括但不限于CHO细胞、各种COS细胞、HeLa细胞、HEK细胞例如HEK 293细胞。As used herein, a "host cell" is a cell that is used to receive, maintain, replicate, and amplify a vector. A host cell can also be used to express a polypeptide encoded by the vector. When the host cell divides, the nucleic acid contained in the vector is replicated, thereby amplifying the nucleic acid. The host cell can be a eukaryotic cell or a prokaryotic cell. Suitable host cells include, but are not limited to, CHO cells, various COS cells, HeLa cells, and HEK cells such as HEK 293 cells.

术语例如“处理”或“治疗”或“以治疗”或“缓解”或“以缓解"指的是治愈、减缓、减少现存的已经诊断的病理病症或紊乱的症状，和/或截停或减缓现存的已经诊断的病理病症或紊乱的进展的治疗性措施。术语例如“预防”、“防御”、“避免”、“遏制”等指的是预防未经诊断的目标病理病症或紊乱的进展的预防性的或预防用的措施。因此，“有需要的受试者”可包括已经患有疾病的受试者；易于患有疾病的受试者；和需要预防疾病的受试者。Terms such as "treating" or "treating" or "to treat" or "alleviating" or "to alleviate" refer to therapeutic measures that cure, alleviate, reduce the symptoms of an existing, diagnosed pathological condition or disorder, and/or arrest or slow the progression of an existing, diagnosed pathological condition or disorder. Terms such as "preventing," "preventing," "avoiding," "containment," and the like refer to preventative or prophylactic measures that prevent the progression of an undiagnosed target pathological condition or disorder. Thus, a "subject in need thereof" can include a subject already suffering from a disease; a subject susceptible to a disease; and a subject in need of prevention of a disease.

如本文所用，“疗效”表示由个体的治疗所导致的效果，其改变、通常改良或改善疾病或疾病状况的症状，或者治愈疾病或疾病状况。As used herein, "therapeutic effect" refers to an effect resulting from treatment of a subject that alters, typically ameliorates or improves the symptoms of a disease or condition, or cures the disease or condition.

术语“治疗有效量”是指对“治疗”对象或哺乳动物中的疾病或病症而言有效的抗体、多肽、多核苷酸、小有机分子或其他药物的量。在癌症的情况中，治疗有效量的药物可减少癌细胞的数量；阻滞或停止癌细胞分裂、降低或阻滞肿瘤尺寸增加；抑制，例如，压制、阻滞、防止、停止、延迟或逆转癌细胞浸润至周边器官，包括，例如，癌症扩散到软组织和骨；抑制，例如，压制、阻滞、防止、收缩、停止、延迟或逆转肿瘤转移；抑制，例如，压制、阻滞、防止、停止、延迟或逆转肿瘤生长；一定程度上缓解与癌症相关的一种或多种症状，降低发病率和死亡率；改善生活质量；或这些效果的组合。就药物预防生长和/或杀伤现存的癌细胞的程度而言，其可以指代抑制细胞生长的和/或细胞毒性的。The term "therapeutically effective amount" refers to an amount of an antibody, polypeptide, polynucleotide, small organic molecule or other drug that is effective for "treating" a disease or condition in a subject or mammal. In the case of cancer, a therapeutically effective amount of a drug can reduce the number of cancer cells; block or stop cancer cell division, reduce or block the increase in tumor size; inhibit, for example, suppress, block, prevent, stop, delay or reverse cancer cell infiltration into peripheral organs, including, for example, the spread of cancer to soft tissue and bone; inhibit, for example, suppress, block, prevent, shrink, stop, delay or reverse tumor metastasis; inhibit, for example, suppress, block, prevent, stop, delay or reverse tumor growth; alleviate to some extent one or more symptoms associated with cancer, reduce morbidity and mortality; improve quality of life; or a combination of these effects. To the extent that a drug prevents growth and/or kills existing cancer cells, it can refer to being cytostatic and/or cytotoxic.

如本文所用，术语“受试者”是指哺乳动物，例如人。As used herein, the term "subject" refers to a mammal, such as a human.

本文所用的抗体编号(如PR013234、PR013235、…、PR013402、PR013406等或类似的编号)仅为区分或标识抗体或产品所用，并不意图表示这样的标识是本发明抗体或产品的特征。本领域技术人员应当理解，例如出于区分或标识的目的，其他抗体或产品同样也有可能使用这样的标识，但并非是指相同或等同的抗体或产品。类似地，实施例中所用的类似编号或标识也仅仅是用于示例方便，本发明的抗体或产品由所附权利要求中所描述的特征限定。The antibody numbers used herein (such as PR013234, PR013235, ..., PR013402, PR013406, etc. or similar numbers) are only used to distinguish or identify antibodies or products, and are not intended to indicate that such identification is a feature of the antibody or product of the present invention. It will be understood by those skilled in the art that, for example, other antibodies or products may also use such identification for the purpose of distinction or identification, but do not refer to the same or equivalent antibodies or products. Similarly, the similar numbers or identifications used in the embodiments are only for illustrative convenience, and the antibodies or products of the present invention are defined by the features described in the appended claims.

DLL3结合分子DLL3 binding molecules

本发明提供一种DLL3结合分子。一般而言，DLL3结合分子可以包括任何特异性结合DLL3的分子。The present invention provides a DLL3 binding molecule. Generally speaking, a DLL3 binding molecule can include any molecule that specifically binds to DLL3.

在一些实施方案中，DLL3结合分子是多肽或蛋白，例如抗体。In some embodiments, the DLL3 binding molecule is a polypeptide or protein, such as an antibody.

在一优选实施方案中，DLL3结合分子是针对DLL3的抗体或其抗原结合片段。In a preferred embodiment, the DLL3 binding molecule is an antibody or antigen-binding fragment thereof directed against DLL3.

DLL3结合分子包含合成抗体、重组产生的抗体、多特异性抗体(例如双特异性抗体)、人抗体、非人抗体、人源化抗体、全人源化抗体、重链抗体、纳米抗体、嵌合抗体、胞内抗体以及抗体片段，例如但不限于Fab片段、Fab'片段、F(ab’)2片段、Fv片段、二硫键连接的Fv(dsFv)、Fd片段、Fd’片段、单链Fv(scFv)、单链Fab(scFab)、双抗体、抗独特型(抗Id)抗体、或者上述任何抗体的抗原结合片段。DLL3-binding molecules include synthetic antibodies, recombinantly produced antibodies, multispecific antibodies (e.g., bispecific antibodies), human antibodies, non-human antibodies, humanized antibodies, fully humanized antibodies, heavy chain antibodies, nanobodies, chimeric antibodies, intrabodies, and antibody fragments, such as, but not limited to, Fab fragments, Fab' fragments, F(ab')2 fragments, Fv fragments, disulfide-linked Fv (dsFv), Fd fragments, Fd' fragments, single-chain Fv (scFv), single-chain Fab (scFab), diabodies, anti-idiotypic (anti-Id) antibodies, or antigen-binding fragments of any of the foregoing.

在一实施方案中，DLL3结合分子是包含重链可变区的抗体或其抗原结合片段。In one embodiment, the DLL3 binding molecule is an antibody or antigen-binding fragment thereof comprising a heavy chain variable region.

在一实施方案中，DLL3结合分子是包含重链可变区的重链抗体或其抗原结合片段。In one embodiment, the DLL3 binding molecule is a heavy chain antibody or antigen-binding fragment thereof comprising a heavy chain variable region.

在一实施方案中，DLL3结合分子是重链抗体。在另一实施方案中，DLL3结合分子是全人源化抗体。在一优选实施方案中，DLL3结合分子是重链抗体并且是全人源化抗体。In one embodiment, the DLL3 binding molecule is a heavy chain antibody. In another embodiment, the DLL3 binding molecule is a fully humanized antibody. In a preferred embodiment, the DLL3 binding molecule is a heavy chain antibody and is a fully humanized antibody.

在一实施方案中，DLL3所包含的重链可变区可以与另一分子融合；优选地，所述另一分子是免疫球蛋白的Fc结构域；最优选地，所述另一分子是免疫球蛋白G1(IgG1)的Fc结构域；最优选地，所述另一分子是人免疫球蛋白G1(IgG1)的Fc结构域。In one embodiment, the heavy chain variable region comprised by DLL3 can be fused to another molecule; preferably, the other molecule is the Fc domain of an immunoglobulin; most preferably, the other molecule is the Fc domain of immunoglobulin G1 (IgG1); most preferably, the other molecule is the Fc domain of human immunoglobulin G1 (IgG1).

在一实施方案中，DLL3结合分子是包含重链可变区与人IgG1的Fc结构域的重链抗体。In one embodiment, the DLL3 binding molecule is a heavy chain antibody comprising a heavy chain variable region and the Fc domain of human IgG1.

在一实施方案中，DLL3结合分子包含重链可变区，其中所述重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1包含SEQ ID NO:18或32所示的序列或者与SEQ ID NO:18或32所示的序列存在不超过2个氨基酸的氨基酸添加、缺失或取代的差异的氨基酸序列；和/或HCDR2包含SEQ ID NO:69、74、79、80、87或90所示的序列或者与SEQ ID NO:69、74、79、80、87或90所示的序列存在不超过2个氨基酸的氨基酸添加、缺失或取代的差异的氨基酸序列；和/或HCDR3包含SEQ ID NO:126、127、128、129、133、135、136、137、138、139、140、141、142、143、144、146、147、148、149、150、151或152所示的序列或者与SEQ ID NO:126、127、128、129、133、135、136、137、138、139、140、141、142、143、144、146、147、148、149、150、151或152所示的序列存在不超过2个氨基酸的氨基酸添加、缺失或取代的差异的氨基酸序列。In one embodiment, a DLL3 binding molecule comprises a heavy chain variable region, wherein the heavy chain variable region comprises a HCDR1, a HCDR2, and a HCDR3, and wherein HCDR1 comprises the sequence of SEQ ID NO: 18 or 32, or an amino acid sequence that differs from the sequence of SEQ ID NO: 18 or 32 by no more than 2 amino acid additions, deletions, or substitutions; and/or HCDR2 comprises the sequence of SEQ ID NO: 69, 74, 79, 80, 87, or 90, or an amino acid sequence that differs from the sequence of SEQ ID NO: 69, 74, 79, 80, 87, or 90 by no more than 2 amino acid additions, deletions, or substitutions; and/or HCDR3 comprises the sequence shown in SEQ ID NO: 126, 127, 128, 129, 133, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 146, 147, 148, 149, 150, 151 or 152, or an amino acid sequence that differs from the sequence shown in SEQ ID NO: 126, 127, 128, 129, 133, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 146, 147, 148, 149, 150, 151 or 152 by no more than 2 amino acids of amino acid addition, deletion or substitution.

在一实施方案中，HCDR1序列包含SEQ ID NO:18所示的氨基酸序列(SEQ ID NO:18；GFTFSDY)，其中第3位氨基酸被取代。在一实施方案中，SEQ ID NO:18中第3位氨基酸被取代为N(SEQ ID NO:19；GFNFSDY)。In one embodiment, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 3 is substituted. In one embodiment, the amino acid at position 3 in SEQ ID NO: 18 is substituted with N (SEQ ID NO: 19; GFNFSDY).

在一实施方案中，HCDR1序列包含SEQ ID NO:18所示的氨基酸序列(SEQ ID NO:18；GFTFSDY)，其中第4位氨基酸被取代。在一实施方案中，SEQ ID NO:18中第4位氨基酸被取代为L(SEQ ID NO:20；GFTLSDY)。In one embodiment, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 4 is substituted. In one embodiment, the amino acid at position 4 in SEQ ID NO: 18 is substituted with L (SEQ ID NO: 20; GFTLSDY).

在一实施方案中，HCDR1序列包含SEQ ID NO:18所示的氨基酸序列(SEQ ID NO:18；GFTFSDY)，其中第4位氨基酸被取代。在一实施方案中，SEQ ID NO:18中第4位氨基酸被取代为I(SEQ ID NO:21；GFTISDY)。In one embodiment, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 4 is substituted. In one embodiment, the amino acid at position 4 in SEQ ID NO: 18 is substituted with 1 (SEQ ID NO: 21; GFTISDY).

在一实施方案中，HCDR1序列包含SEQ ID NO:18所示的氨基酸序列(SEQ ID NO:18；GFTFSDY)，其中第3位氨基酸被取代。在一些实施方案中，HCDR1序列包含SEQ ID NO:18所示的氨基酸序列(SEQ ID NO:18；GFTFSDY)，其中第4位氨基酸被取代。在一些实施方案中，HCDR1序列包含SEQ ID NO:18所示的氨基酸序列(SEQ ID NO:18；GFTFSDY)，其中第3位氨基酸和第4位氨基酸被取代。在一实施方案中，SEQ ID NO:18中第3位氨基酸被取代为N，SEQ ID NO:18中第4位氨基酸被取代为I(SEQ ID NO:22；GFNISDY)。In one embodiment, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 3 is substituted. In some embodiments, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 4 is substituted. In some embodiments, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 3 and the amino acid at position 4 are substituted. In one embodiment, the amino acid at position 3 in SEQ ID NO: 18 is substituted with N, and the amino acid at position 4 in SEQ ID NO: 18 is substituted with I (SEQ ID NO: 22; GFNISDY).

在一实施方案中，HCDR1序列包含SEQ ID NO:18所示的氨基酸序列(SEQ ID NO:18；GFTFSDY)，其中第3位氨基酸被取代。在一些实施方案中，HCDR1序列包含SEQ ID NO:18所示的氨基酸序列(SEQ ID NO:18；GFTFSDY)，其中第4位氨基酸被取代。在一些实施方案中，HCDR1序列包含SEQ ID NO:18所示的氨基酸序列(SEQ ID NO:18；GFTFSDY)，其中第3位氨基酸和第4位氨基酸被取代。在一实施方案中，SEQ ID NO:18中第3位氨基酸被取代为N，SEQ ID NO:18中第4位氨基酸被取代为L(SEQ ID NO:23；GFNLSDY)。In one embodiment, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 3 is substituted. In some embodiments, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 4 is substituted. In some embodiments, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 3 and the amino acid at position 4 are substituted. In one embodiment, the amino acid at position 3 in SEQ ID NO: 18 is substituted with N, and the amino acid at position 4 in SEQ ID NO: 18 is substituted with L (SEQ ID NO: 23; GFNLSDY).

在一实施方案中，HCDR1序列包含SEQ ID NO:18所示的氨基酸序列(SEQ ID NO:18；GFTFSDY)，其中第3位氨基酸被取代。在一些实施方案中，HCDR1序列包含SEQ ID NO:18所示的氨基酸序列(SEQ ID NO:18；GFTFSDY)，其中第5位氨基酸被取代。在一些实施方案中，HCDR1序列包含SEQ ID NO:18所示的氨基酸序列(SEQ ID NO:18；GFTFSDY)，其中第3位氨基酸和第5位氨基酸被取代。在一实施方案中，SEQ ID NO:18中第3位氨基酸被取代为N，SEQ ID NO:18中第5位氨基酸被取代为N(SEQ ID NO:24；GFNFNDY)。In one embodiment, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 3 is substituted. In some embodiments, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 5 is substituted. In some embodiments, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 3 and the amino acid at position 5 are substituted. In one embodiment, the amino acid at position 3 in SEQ ID NO: 18 is substituted with N, and the amino acid at position 5 in SEQ ID NO: 18 is substituted with N (SEQ ID NO: 24; GFNFNDY).

在一实施方案中，HCDR1序列包含SEQ ID NO:18所示的氨基酸序列(SEQ ID NO:18；GFTFSDY)，其中第6位氨基酸被取代。在一实施方案中，SEQ ID NO:18中第6位氨基酸被取代为S(SEQ ID NO:25；GFTFSSY)。In one embodiment, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 6 is substituted. In one embodiment, the amino acid at position 6 in SEQ ID NO: 18 is substituted with S (SEQ ID NO: 25; GFTFSSY).

在一实施方案中，HCDR1序列包含SEQ ID NO:18所示的氨基酸序列(SEQ ID NO:18；GFTFSDY)，其中第6位氨基酸被取代。在一实施方案中，SEQ ID NO:18中第6位氨基酸被取代为N(SEQ ID NO:26；GFTFSNY)。In one embodiment, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 6 is substituted. In one embodiment, the amino acid at position 6 in SEQ ID NO: 18 is substituted with N (SEQ ID NO: 26; GFTFSNY).

在一实施方案中，HCDR1序列包含SEQ ID NO:18所示的氨基酸序列(SEQ ID NO:18；GFTFSDY)，其中第7位氨基酸被取代。在一实施方案中，SEQ ID NO:18中第7位氨基酸被取代为H(SEQ ID NO:27；GFTFSDH)。In one embodiment, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 7 is substituted. In one embodiment, the amino acid at position 7 in SEQ ID NO: 18 is substituted with H (SEQ ID NO: 27; GFTFSDH).

在一实施方案中，HCDR1序列包含SEQ ID NO:18所示的氨基酸序列(SEQ ID NO:18；GFTFSDY)，其中第6位氨基酸被取代。在一实施方案中，SEQ ID NO:18中第6位氨基酸被取代为T(SEQ ID NO:28；GFTFSTY)。In one embodiment, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 6 is substituted. In one embodiment, the amino acid at position 6 in SEQ ID NO: 18 is substituted with T (SEQ ID NO: 28; GFTFSTY).

在一实施方案中，HCDR1序列包含SEQ ID NO:18所示的氨基酸序列(SEQ ID NO:18；GFTFSDY)，其中第3位氨基酸被取代。在一实施方案中，SEQ ID NO:18中第3位氨基酸被取代为A(SEQ ID NO:29；GFAFSDY)。In one embodiment, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 3 is substituted. In one embodiment, the amino acid at position 3 in SEQ ID NO: 18 is substituted with A (SEQ ID NO: 29; GFAFSDY).

在一实施方案中，HCDR1序列包含SEQ ID NO:18所示的氨基酸序列(SEQ ID NO:18；GFTFSDY)，其中第3位氨基酸被取代。在一些实施方案中，HCDR1序列包含SEQ ID NO:18所示的氨基酸序列(SEQ ID NO:18；GFTFSDY)，其中第6位氨基酸被取代。在一些实施方案中，HCDR1序列包含SEQ ID NO:18所示的氨基酸序列(SEQ ID NO:18；GFTFSDY)，其中第3位氨基酸和第6位氨基酸被取代。在一实施方案中，SEQ ID NO:18中第3位氨基酸被取代为I，SEQ ID NO:18中第6位氨基酸被取代为N(SEQ ID NO:30；GFIFSNY)。In one embodiment, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 3 is substituted. In some embodiments, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 6 is substituted. In some embodiments, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 3 and the amino acid at position 6 are substituted. In one embodiment, the amino acid at position 3 in SEQ ID NO: 18 is substituted with I, and the amino acid at position 6 in SEQ ID NO: 18 is substituted with N (SEQ ID NO: 30; GFIFSNY).

在一实施方案中，HCDR1序列包含SEQ ID NO:18所示的氨基酸序列(SEQ ID NO:18；GFTFSDY)，其中第3位氨基酸被取代。在一些实施方案中，HCDR1序列包含SEQ ID NO:18所示的氨基酸序列(SEQ ID NO:18；GFTFSDY)，其中第6位氨基酸被取代。在一些实施方案中，HCDR1序列包含SEQ ID NO:18所示的氨基酸序列(SEQ ID NO:18；GFTFSDY)，其中第3位氨基酸和第6位氨基酸被取代。在一实施方案中，SEQ ID NO:18中第3位氨基酸被取代为I，SEQ ID NO:18中第6位氨基酸被取代为S(SEQ ID NO:31；GFIFSSY)。In one embodiment, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 3 is substituted. In some embodiments, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 6 is substituted. In some embodiments, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 3 and the amino acid at position 6 are substituted. In one embodiment, the amino acid at position 3 in SEQ ID NO: 18 is substituted with I, and the amino acid at position 6 in SEQ ID NO: 18 is substituted with S (SEQ ID NO: 31; GFIFSSY).

在一实施方案中，HCDR1序列包含SEQ ID NO:18所示的氨基酸序列(SEQ ID NO:18；GFTFSDY)，其中第4位氨基酸被取代。在一些实施方案中，HCDR1序列包含SEQ ID NO:18所示的氨基酸序列(SEQ ID NO:18；GFTFSDY)，其中第6位氨基酸被取代。在一些实施方案中，HCDR1序列包含SEQ ID NO:18所示的氨基酸序列(SEQ ID NO:18；GFTFSDY)，其中第4位氨基酸和第6位氨基酸被取代。在一实施方案中，SEQ ID NO:18中第4位氨基酸被取代为I，SEQ ID NO:18中第6位氨基酸被取代为N(SEQ ID NO:33；GFTISNY)。In one embodiment, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 4 is substituted. In some embodiments, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 6 is substituted. In some embodiments, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 4 and the amino acid at position 6 are substituted. In one embodiment, the amino acid at position 4 in SEQ ID NO: 18 is substituted with I, and the amino acid at position 6 in SEQ ID NO: 18 is substituted with N (SEQ ID NO: 33; GFTISNY).

在一实施方案中，HCDR1序列包含SEQ ID NO:18所示的氨基酸序列(SEQ ID NO:18；GFTFSDY)，其中第6位氨基酸被取代。在一实施方案中，SEQ ID NO:18中第6位氨基酸被取代为H(SEQ ID NO:34；GFTFSHY)。In one embodiment, the HCDR1 sequence comprises the amino acid sequence of SEQ ID NO: 18 (SEQ ID NO: 18; GFTFSDY), wherein the amino acid at position 6 is substituted. In one embodiment, the amino acid at position 6 in SEQ ID NO: 18 is substituted with H (SEQ ID NO: 34; GFTFSHY).

在一实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第2位氨基酸被取代。在一实施方案中，SEQ ID NO:69中第2位氨基酸被取代为G(SEQ ID NO:60；SGSGST)。In one embodiment, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO:69 (SEQ ID NO:69; SSSGST), wherein the amino acid at position 2 is substituted. In one embodiment, the amino acid at position 2 in SEQ ID NO:69 is substituted with G (SEQ ID NO:60; SGSGST).

在一实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第2位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第3位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第2位氨基酸和第3位氨基酸被取代。在一实施方案中，SEQ ID NO:69中第2位氨基酸被取代为A，SEQ ID NO:69中第3位氨基酸被取代为N(SEQ ID NO:61；SANGST)。In one embodiment, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 2 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 3 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 2 and the amino acid at position 3 are substituted. In one embodiment, the amino acid at position 2 in SEQ ID NO: 69 is substituted with A, and the amino acid at position 3 in SEQ ID NO: 69 is substituted with N (SEQ ID NO: 61; SANGST).

在一实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第1位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第2位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第1位氨基酸和第2位氨基酸被取代。在一实施方案中，SEQ ID NO:69中第1位氨基酸被取代为T，SEQ ID NO:69中第2位氨基酸被取代为G(SEQ ID NO:62；TGSGST)。In one embodiment, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 1 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 2 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 1 and the amino acid at position 2 are substituted. In one embodiment, the amino acid at position 1 in SEQ ID NO: 69 is substituted with T, and the amino acid at position 2 in SEQ ID NO: 69 is substituted with G (SEQ ID NO: 62; TGSGST).

在一实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第5位氨基酸被取代。在一实施方案中，SEQ ID NO:69中第5位氨基酸被取代为N(SEQ ID NO:63；SSSGNT)。In one embodiment, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO:69 (SEQ ID NO:69; SSSGST), wherein the amino acid at position 5 is substituted. In one embodiment, the amino acid at position 5 in SEQ ID NO:69 is substituted with N (SEQ ID NO:63; SSSGNT).

在一实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第3位氨基酸被取代。在一实施方案中，SEQ ID NO:69中第3位氨基酸被取代为N(SEQ ID NO:64；SSNGST)。In one embodiment, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO:69 (SEQ ID NO:69; SSSGST), wherein the amino acid at position 3 is substituted. In one embodiment, the amino acid at position 3 in SEQ ID NO:69 is substituted with N (SEQ ID NO:64; SSNGST).

在一实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第2位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第5位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第2位氨基酸和第5位氨基酸被取代。在一实施方案中，SEQ ID NO:69中第2位氨基酸被取代为G，SEQ ID NO:69中第5位氨基酸被取代为R(SEQ ID NO:65；SGSGRT)。In one embodiment, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 2 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 5 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 2 and the amino acid at position 5 are substituted. In one embodiment, the amino acid at position 2 in SEQ ID NO: 69 is substituted with G, and the amino acid at position 5 in SEQ ID NO: 69 is substituted with R (SEQ ID NO: 65; SGSGRT).

在一实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第2位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第3位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第2位氨基酸和第3位氨基酸被取代。在一实施方案中，SEQ ID NO:69中第2位氨基酸被取代为G，SEQ ID NO:69中第3位氨基酸被取代为N(SEQ ID NO:66；SGNGST)。In one embodiment, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 2 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 3 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 2 and the amino acid at position 3 are substituted. In one embodiment, the amino acid at position 2 in SEQ ID NO: 69 is substituted with G, and the amino acid at position 3 in SEQ ID NO: 69 is substituted with N (SEQ ID NO: 66; SGNGST).

在一实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第5位氨基酸被取代。在一实施方案中，SEQ ID NO:69中第5位氨基酸被取代为G(SEQ ID NO:67；SSSGGT)。In one embodiment, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 5 is substituted. In one embodiment, the amino acid at position 5 in SEQ ID NO: 69 is substituted with G (SEQ ID NO: 67; SSSGGT).

在一实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第2位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第6位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第2位氨基酸和第6位氨基酸被取代。在一实施方案中，SEQ ID NO:69中第2位氨基酸被取代为G，SEQ ID NO:69中第6位氨基酸被取代为I(SEQ ID NO:68；SGSGSI)。In one embodiment, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 2 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 6 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 2 and the amino acid at position 6 are substituted. In one embodiment, the amino acid at position 2 in SEQ ID NO: 69 is substituted with G, and the amino acid at position 6 in SEQ ID NO: 69 is substituted with I (SEQ ID NO: 68; SGSGSI).

在一实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第5位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第6位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第5位氨基酸和第6位氨基酸被取代。在一实施方案中，SEQ ID NO:69中第5位氨基酸被取代为D，SEQ ID NO:69中第6位氨基酸被取代为S(SEQ ID NO:70；SSSGDS)。In one embodiment, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 5 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 6 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 5 and the amino acid at position 6 are substituted. In one embodiment, the amino acid at position 5 in SEQ ID NO: 69 is substituted with D, and the amino acid at position 6 in SEQ ID NO: 69 is substituted with S (SEQ ID NO: 70; SSSGDS).

在一实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第5位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第6位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第5位氨基酸和第6位氨基酸被取代。在一实施方案中，SEQ ID NO:69中第5位氨基酸被取代为E，SEQ ID NO:69中第6位氨基酸被取代为S(SEQ ID NO:71；SSSGES)。In one embodiment, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 5 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 6 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 5 and the amino acid at position 6 are substituted. In one embodiment, the amino acid at position 5 in SEQ ID NO: 69 is substituted with E, and the amino acid at position 6 in SEQ ID NO: 69 is substituted with S (SEQ ID NO: 71; SSSGES).

在一实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第5位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第6位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第5位氨基酸和第6位氨基酸被取代。在一实施方案中，SEQ ID NO:69中第5位氨基酸被取代为N，SEQ ID NO:69中第6位氨基酸被取代为S(SEQ ID NO:73；SSSGNS)。In one embodiment, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 5 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 6 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 5 and the amino acid at position 6 are substituted. In one embodiment, the amino acid at position 5 in SEQ ID NO: 69 is substituted with N, and the amino acid at position 6 in SEQ ID NO: 69 is substituted with S (SEQ ID NO: 73; SSSGNS).

在一实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第3位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第6位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第3位氨基酸和第6位氨基酸被取代。在一实施方案中，SEQ ID NO:69中第3位氨基酸被取代为D，SEQ ID NO:69中第6位氨基酸被取代为S(SEQ ID NO:75；SSDGSS)。In one embodiment, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 3 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 6 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 3 and the amino acid at position 6 are substituted. In one embodiment, the amino acid at position 3 in SEQ ID NO: 69 is substituted with D, and the amino acid at position 6 in SEQ ID NO: 69 is substituted with S (SEQ ID NO: 75; SSDGSS).

在一实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第2位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第6位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第2位氨基酸和第6位氨基酸被取代。在一实施方案中，SEQ ID NO:69中第2位氨基酸被取代为G，SEQ ID NO:69中第6位氨基酸被取代为D(SEQ ID NO:76；SGSGSD)。In one embodiment, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 2 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 6 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 2 and the amino acid at position 6 are substituted. In one embodiment, the amino acid at position 2 in SEQ ID NO: 69 is substituted with G, and the amino acid at position 6 in SEQ ID NO: 69 is substituted with D (SEQ ID NO: 76; SGSGSD).

在一实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第3位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第6位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第3位氨基酸和第6位氨基酸被取代。在一实施方案中，SEQ ID NO:69中第3位氨基酸被取代为D，SEQ ID NO:69中第6位氨基酸被取代为D(SEQ ID NO:82；SSDGSD)。In one embodiment, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 3 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 6 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 3 and the amino acid at position 6 are substituted. In one embodiment, the amino acid at position 3 in SEQ ID NO: 69 is substituted with D, and the amino acid at position 6 in SEQ ID NO: 69 is substituted with D (SEQ ID NO: 82; SSDGSD).

在一实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第3位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第6位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第3位氨基酸和第6位氨基酸被取代。在一实施方案中，SEQ ID NO:69中第3位氨基酸被取代为D，SEQ ID NO:69中第6位氨基酸被取代为N(SEQ ID NO:83；SSDGSN)。In one embodiment, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 3 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 6 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 3 and the amino acid at position 6 are substituted. In one embodiment, the amino acid at position 3 in SEQ ID NO: 69 is substituted with D, and the amino acid at position 6 in SEQ ID NO: 69 is substituted with N (SEQ ID NO: 83; SSDGSN).

在一实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第2位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第6位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:69所示的氨基酸序列(SEQ ID NO:69；SSSGST)，其中第2位氨基酸和第6位氨基酸被取代。在一实施方案中，SEQ ID NO:69中第2位氨基酸被取代为T，SEQ ID NO:69中第6位氨基酸被取代为P(SEQ ID NO:89；STSGSP)。In one embodiment, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 2 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 6 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 69 (SEQ ID NO: 69; SSSGST), wherein the amino acid at position 2 and the amino acid at position 6 are substituted. In one embodiment, the amino acid at position 2 in SEQ ID NO: 69 is substituted with T, and the amino acid at position 6 in SEQ ID NO: 69 is substituted with P (SEQ ID NO: 89; STSGSP).

在一实施方案中，HCDR2序列包含SEQ ID NO:74所示的氨基酸序列(SEQ ID NO:74；SGSDNN)，其中第1位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:74所示的氨基酸序列(SEQ ID NO:74；SGSDNN)，其中第3位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:74所示的氨基酸序列(SEQ ID NO:74；SGSDNN)，其中第1位氨基酸和第3位氨基酸被取代。在一实施方案中，SEQ ID NO:74中第1位氨基酸被取代为N，SEQ ID NO:74中第3位氨基酸被取代为N(SEQ ID NO:72；NGNDNN)。In one embodiment, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 74 (SEQ ID NO: 74; SGSDNN), wherein the amino acid at position 1 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 74 (SEQ ID NO: 74; SGSDNN), wherein the amino acid at position 3 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 74 (SEQ ID NO: 74; SGSDNN), wherein the amino acid at position 1 and the amino acid at position 3 are substituted. In one embodiment, the amino acid at position 1 in SEQ ID NO: 74 is substituted with N, and the amino acid at position 3 in SEQ ID NO: 74 is substituted with N (SEQ ID NO: 72; NGNDNN).

在一实施方案中，HCDR2序列包含SEQ ID NO:74所示的氨基酸序列(SEQ ID NO:74；SGSDNN)，其中第5位氨基酸被取代。在一实施方案中，SEQ ID NO:74中第5位氨基酸被取代为D(SEQ ID NO:86；SGSDDN)。In one embodiment, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 74 (SEQ ID NO: 74; SGSDNN), wherein the amino acid at position 5 is substituted. In one embodiment, the amino acid at position 5 in SEQ ID NO: 74 is substituted with D (SEQ ID NO: 86; SGSDDN).

在一实施方案中，HCDR2序列包含SEQ ID NO:79所示的氨基酸序列(SEQ ID NO:79；NSDGST)，其中第2位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:79所示的氨基酸序列(SEQ ID NO:79；NSDGST)，其中第6位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:79所示的氨基酸序列(SEQ ID NO:79；NSDGST)，其中第2位氨基酸和第6位氨基酸被取代。在一实施方案中，SEQ ID NO:79中第2位氨基酸被取代为G，SEQ ID NO:79中第6位氨基酸被取代为N(SEQ ID NO:77；NGDGSN)。In one embodiment, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 79 (SEQ ID NO: 79; NSDGST), wherein the amino acid at position 2 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 79 (SEQ ID NO: 79; NSDGST), wherein the amino acid at position 6 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 79 (SEQ ID NO: 79; NSDGST), wherein the amino acid at position 2 and the amino acid at position 6 are substituted. In one embodiment, the amino acid at position 2 in SEQ ID NO: 79 is substituted with G, and the amino acid at position 6 in SEQ ID NO: 79 is substituted with N (SEQ ID NO: 77; NGDGSN).

在一实施方案中，HCDR2序列包含SEQ ID NO:79所示的氨基酸序列(SEQ ID NO:79；NSDGST)，其中第6位氨基酸被取代。在一实施方案中，SEQ ID NO:79中第6位氨基酸被取代为Y(SEQ ID NO:78；NSDGSY)。In one embodiment, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 79 (SEQ ID NO: 79; NSDGST), wherein the amino acid at position 6 is substituted. In one embodiment, the amino acid at position 6 in SEQ ID NO: 79 is substituted with Y (SEQ ID NO: 78; NSDGSY).

在一实施方案中，HCDR2序列包含SEQ ID NO:79所示的氨基酸序列(SEQ ID NO:79；NSDGST)，其中第3位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:79所示的氨基酸序列(SEQ ID NO:79；NSDGST)，其中第5位氨基酸被取代。在一些实施方案中，HCDR2序列包含SEQ ID NO:79所示的氨基酸序列(SEQ ID NO:79；NSDGST)，其中第3位氨基酸和第5位氨基酸被取代。在一实施方案中，SEQ ID NO:79中第3位氨基酸被取代为N，SEQ ID NO:79中第5位氨基酸被取代为N(SEQ ID NO:81；NSNGNT)。In one embodiment, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 79 (SEQ ID NO: 79; NSDGST), wherein the amino acid at position 3 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 79 (SEQ ID NO: 79; NSDGST), wherein the amino acid at position 5 is substituted. In some embodiments, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 79 (SEQ ID NO: 79; NSDGST), wherein the amino acid at position 3 and the amino acid at position 5 are substituted. In one embodiment, the amino acid at position 3 in SEQ ID NO: 79 is substituted with N, and the amino acid at position 5 in SEQ ID NO: 79 is substituted with N (SEQ ID NO: 81; NSNGNT).

在一实施方案中，HCDR2序列包含SEQ ID NO:79所示的氨基酸序列(SEQ ID NO:79；NSDGST)，其中第6位氨基酸被取代。在一实施方案中，SEQ ID NO:79中第6位氨基酸被取代为D(SEQ ID NO:84；NSDGSD)。In one embodiment, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 79 (SEQ ID NO: 79; NSDGST), wherein the amino acid at position 6 is substituted. In one embodiment, the amino acid at position 6 in SEQ ID NO: 79 is substituted with D (SEQ ID NO: 84; NSDGSD).

在一实施方案中，HCDR2序列包含SEQ ID NO:79所示的氨基酸序列(SEQ ID NO:79；NSDGST)，其中第6位氨基酸被取代。在一实施方案中，SEQ ID NO:79中第6位氨基酸被取代为N(SEQ ID NO:85；NSDGSN)。In one embodiment, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 79 (SEQ ID NO: 79; NSDGST), wherein the amino acid at position 6 is substituted. In one embodiment, the amino acid at position 6 in SEQ ID NO: 79 is substituted with N (SEQ ID NO: 85; NSDGSN).

在一实施方案中，HCDR2序列包含SEQ ID NO:79所示的氨基酸序列(SEQ ID NO:79；NSDGST)，其中第6位氨基酸被取代。在一实施方案中，SEQ ID NO:79中第6位氨基酸被取代为S(SEQ ID NO:88；NSDGSS)。In one embodiment, the HCDR2 sequence comprises the amino acid sequence of SEQ ID NO: 79 (SEQ ID NO: 79; NSDGST), wherein the amino acid at position 6 is substituted. In one embodiment, the amino acid at position 6 in SEQ ID NO: 79 is substituted with S (SEQ ID NO: 88; NSDGSS).

在一实施方案中，HCDR3序列包含SEQ ID NO:126所示的氨基酸序列(SEQ ID NO:126；ENGNSDY)，其中第2位氨基酸被取代。在一实施方案中，SEQ ID NO:126中第2位氨基酸被取代为S(SEQ ID NO:131；ESGNSDY)。In one embodiment, the HCDR3 sequence comprises the amino acid sequence of SEQ ID NO: 126 (SEQ ID NO: 126; ENGNSDY), wherein the amino acid at position 2 is substituted. In one embodiment, the amino acid at position 2 in SEQ ID NO: 126 is substituted with S (SEQ ID NO: 131; ESGNSDY).

在一实施方案中，HCDR3序列包含SEQ ID NO:127所示的氨基酸序列(SEQ ID NO:127；GLGEFSLGI)，其中第9位氨基酸被取代。在一实施方案中，SEQ ID NO:127中第9位氨基酸被取代为S(SEQ ID NO:130；GLGEFSLGS)。In one embodiment, the HCDR3 sequence comprises the amino acid sequence of SEQ ID NO: 127 (SEQ ID NO: 127; GLGEFSLGI), wherein the amino acid at position 9 is substituted. In one embodiment, the amino acid at position 9 in SEQ ID NO: 127 is substituted with S (SEQ ID NO: 130; GLGEFSLGS).

在一实施方案中，HCDR3序列包含SEQ ID NO:127所示的氨基酸序列(SEQ ID NO:127；GLGEFSLGI)，其中第5位氨基酸被取代。在一实施方案中，SEQ ID NO:127中第5位氨基酸被取代为L(SEQ ID NO:132；GLGELSLGI)。In one embodiment, the HCDR3 sequence comprises the amino acid sequence of SEQ ID NO: 127 (SEQ ID NO: 127; GLGEFSLGI), wherein the amino acid at position 5 is substituted. In one embodiment, the amino acid at position 5 in SEQ ID NO: 127 is substituted with L (SEQ ID NO: 132; GLGELSLGI).

在一实施方案中，HCDR3序列包含SEQ ID NO:133所示的氨基酸序列(SEQ ID NO:133；ERGSTGGWFDP)，其中第2位氨基酸被取代。在一实施方案中，SEQ ID NO:133中第2位氨基酸被取代为K(SEQ ID NO:134；EKGSTGGWFDP)。In one embodiment, the HCDR3 sequence comprises the amino acid sequence of SEQ ID NO: 133 (SEQ ID NO: 133; ERGSTGGWFDP), wherein the amino acid at position 2 is substituted. In one embodiment, the amino acid at position 2 in SEQ ID NO: 133 is substituted with K (SEQ ID NO: 134; EKGSTGGWFDP).

在一实施方案中，HCDR3序列包含SEQ ID NO:139所示的氨基酸序列(SEQ ID NO:139；GGDTVVVPEFDY)，其中第2位氨基酸被取代。在一些实施方案中，HCDR3序列包含SEQ ID NO:139所示的氨基酸序列(SEQ ID NO:139；GGDTVVVPEFDY)，其中第7位氨基酸被取代。在一些实施方案中，HCDR3序列包含SEQ ID NO:139所示的氨基酸序列(SEQ ID NO:139；GGDTVVVPEFDY)，其中第2位氨基酸和第7位氨基酸被取代。在一实施方案中，SEQ ID NO:139中第2位氨基酸被取代为D，SEQ ID NO:139中第7位氨基酸被取代为L(SEQ ID NO:145；GDDTVVLPEFDY)。In one embodiment, the HCDR3 sequence comprises the amino acid sequence of SEQ ID NO: 139 (SEQ ID NO: 139; GGDTVVVPEFDY), wherein the amino acid at position 2 is substituted. In some embodiments, the HCDR3 sequence comprises the amino acid sequence of SEQ ID NO: 139 (SEQ ID NO: 139; GGDTVVVPEFDY), wherein the amino acid at position 7 is substituted. In some embodiments, the HCDR3 sequence comprises the amino acid sequence of SEQ ID NO: 139 (SEQ ID NO: 139; GGDTVVVPEFDY), wherein the amino acid at position 2 and the amino acid at position 7 are substituted. In one embodiment, the amino acid at position 2 in SEQ ID NO: 139 is substituted with D, and the amino acid at position 7 in SEQ ID NO: 139 is substituted with L (SEQ ID NO: 145; GDDTVVLPEFDY).

在一实施方案中，HCDR1包含如SEQ ID NO:18、19、20、21、22、23、24、25、26、27、28、29、30、31、32、33或34所示的氨基酸序列。In one embodiment, HCDR1 comprises an amino acid sequence as shown in SEQ ID NO: 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33 or 34.

在一实施方案中，HCDR2包含如SEQ ID NO:60、61、62、63、64、65、66、67、68、69、70、71、72、73、74、75、76、77、78、79、80、81、82、83、84、85、86、87、88、89或90所示的氨基酸序列。In one embodiment, HCDR2 comprises an amino acid sequence as shown in SEQ ID NO: 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89 or 90.

在一实施方案中，HCDR3包含如SEQ ID NO:126、127、128、129、130、131、132、133、134、135、136、137、138、139、140、141、142、143、144、145、146、147、148、149、150、151或152所示的氨基酸序列。In one embodiment, HCDR3 comprises an amino acid sequence as shown in SEQ ID NO:126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151 or 152.

在一实施方案中，HCDR1包含如SEQ ID NO:18、19、20、21、22、23、24、25、26、27、28、29、30、31、32、33或34所示的氨基酸序列；HCDR2包含如SEQ ID NO:60、61、62、63、64、65、66、67、68、69、70、71、72、73、74、75、76、77、78、79、80、81、82、83、84、85、86、87、88、89或90所示的氨基酸序列；以及HCDR3包含如SEQ ID NO:126、127、128、129、130、131、132、133、134、135、136、137、138、139、140、141、142、143、144、145、146、147、148、149、150、151或152所示的氨基酸序列。In one embodiment, HCDR1 comprises an amino acid sequence as shown in SEQ ID NO: 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33 or 34; HCDR2 comprises an amino acid sequence as shown in SEQ ID NO: 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85 2, 83, 84, 85, 86, 87, 88, 89 or 90; and HCDR3 comprises the amino acid sequence shown in SEQ ID NO:126, 127, 128, 129, 130, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151 or 152.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、60、126所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:18, 60, 126, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:19、60、126所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:19, 60, 126, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、61、126所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:18, 61, 126, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:20、62、127所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 20, 62, 127, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、63、128所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:18, 63, 128, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:21、64、129所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 21, 64, 129, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:22、65、126所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 22, 65, and 126, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:23、65、126所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 23, 65, and 126, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、66、126所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:18, 66, 126, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、67、130所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:18, 67, 130, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、64、131所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:18, 64, 131, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:19、68、126所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:19, 68, and 126, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、68、126所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:18, 68, 126, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:20、69、132所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 20, 69, 132, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:24、66、126所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 24, 66, 126, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、70、133所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:18, 70, 133, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、70、134所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:18, 70, 134, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、71、134所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:18, 71, 134, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:25、72、135所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 25, 72, 135, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、73、134所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:18, 73, 134, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:25、74、135所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 25, 74, 135, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:26、75、136所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 26, 75, 136, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:27、76、137所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 27, 76, 137, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:25、77、138所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 25, 77, 138, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:25、78、139所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 25, 78, 139, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、79、140所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:18, 79, 140, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:26、80、141所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 26, 80, 141, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:26、81、142所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 26, 81, 142, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:25、82、143所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 25, 82, 143, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:28、83、144所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 28, 83, 144, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:29、84、143所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 29, 84, 143, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:26、85、145所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 26, 85, and 145, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:30、75、136所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 30, 75, 136, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:31、85、146所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:31, 85, and 146, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:32、86、147所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:32, 86, and 147, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、85、148所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:18, 85, 148, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:33、87、149所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 33, 87, 149, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:26、88、150所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 26, 88, and 150, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、89、151所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:18, 89, 151, respectively.

在一实施方案中，重链可变区包含HCDR1、HCDR2和HCDR3，并且其中HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:34、90、152所示的氨基酸序列。In one embodiment, the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO:34, 90, 152, respectively.

在一实施方案中，重链可变区包含1)SEQ ID NO:174、175、176、177、178、179、180、181、182、183、184、185、186、187、188、189、190、191、192、193、194、195、196、197、198、199、200、201、202、203、204、205、206、207、208、209、210、211、212、213、214、215、216或217的氨基酸序列；2)与SEQ ID NO:174、175、176、177、178、179、180、181、182、183、184、185、186、187、188、189、190、191、192、193、194、195、196、197、198、199、200、201、202、203、204、205、206、207、208、209、210、211、212、213、214、215、216或217的氨基酸序列具有至少80％、至少85％、至少90％、至少91％、至少92％、至少93％、至少94％、至少95％、至少96％、至少97％、至少98％、至少99％序列相同性的氨基酸序列；或者3)与SEQ ID NO:174、175、176、177、178、179、180、181、182、183、184、185、186、187、188、189、190、191、192、193、194、195、196、197、198、199、200、201、202、203、204、205、206、207、208、209、210、211、212、213、214、215、216或217的氨基酸序列相比具有一个或多个氨基酸的取代、添加和/或缺失的氨基酸序列，优选地，所述添加、缺失和/或取代不发生在CDR区。In one embodiment, the heavy chain variable region comprises 1) SEQ ID NO: 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210 , 211, 212, 213, 214, 215, 216 or 217 having an amino acid sequence that is at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% identical to the amino acid sequence of SEQ ID NO: 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 184, 185, 186 , 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216 or 217, and preferably, the addition, deletion and/or substitution does not occur in the CDR region.

在一实施方案中，重链可变区与另一分子融合。在一优选方案中，另一分子是免疫球蛋白的Fc结构域。在一更优选方案中，另一分子是人免疫球蛋白G1(IgG1)的Fc结构域。In one embodiment, the heavy chain variable region is fused to another molecule. In a preferred embodiment, the other molecule is the Fc domain of an immunoglobulin. In a more preferred embodiment, the other molecule is the Fc domain of human immunoglobulin G1 (IgG1).

在一实施方案中，DLL3结合分子包含1)SEQ ID NO:222、223、224、225、226、227、228、229、230、231、232、233、234、235、236、237、238、239、240、241、242、243、244、245、246、247、248、249、250、251、252、253、254、255、256、257、258、259、260、261、262、263、264或265的氨基酸序列；2)与SEQ ID NO:222、223、224、225、226、227、228、229、230、231、232、233、234、235、236、237、238、239、240、241、242、243、244、245、246、247、248、249、250、251、252、253、254、255、256、257、258、259、260、261、262、263、264或265的氨基酸序列具有至少80％、至少85％、至少90％、至少91％、至少92％、至少93％、至少94％、至少95％、至少96％、至少97％、至少98％、至少99％序列相同性的氨基酸序列；或者3)与SEQ ID NO:222、223、224、225、226、227、228、229、230、231、232、233、234、235、236、237、238、239、240、241、242、243、244、245、246、247、248、249、250、251、252、253、254、255、256、257、258、259、260、261、262、263、264或265的氨基酸序列相比具有一个或多个氨基酸的取代、添加和/或缺失的氨基酸序列，优选地，所述添加、缺失和/或取代不发生在CDR区。In one embodiment, the DLL3 binding molecule comprises 1) SEQ ID NO: 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257, 258, 259, 260 , 261, 262, 263, 264 or 265; 2) an amino acid sequence having the same amino acid sequence as SEQ ID NO: 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 232, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257, 258 8) an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% sequence identity to the amino acid sequence of SEQ ID NO: 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 232, 233, 234 4, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257, 258, 259, 260, 261, 262, 263, 264 or 265, compared to the amino acid sequence having one or more amino acid substitutions, additions and/or deletions, preferably, the additions, deletions and/or substitutions do not occur in the CDR regions.

在一实施方案中，DLL3结合分子具有下述特征中的至少一个：In one embodiment, the DLL3 binding molecule has at least one of the following characteristics:

1)具有对DLL3蛋白的亲和活性；1) Has affinity activity for DLL3 protein;

2)具有对DLL3阳性细胞的亲和活性；2) has affinity activity for DLL3-positive cells;

3)能够内化至DLL3阳性的细胞内。3) Ability to be internalized into DLL3-positive cells.

在一实施方案中，DLL3结合分子包括针对DLL3的抗体PR013234、PR013235、PR013236、PR013239、PR013240、PR013241、PR013242、PR013244、PR013246、PR013249、PR013250、PR013252、PR013253、PR013254、PR013256、PR013257、PR013258、PR013259、PR013260、PR013261、PR013262、PR013263、PR013264、PR013363、PR013364、PR013367、PR013368、PR013370、PR013371、PR013372、PR013376、PR013377、PR013378、PR013381、PR013382、PR013384、PR013387、PR013392、PR013393、PR013394、PR013397、PR013399、PR013402、PR013406，特别是DLL3抗体PR013236、PR013239、PR013240、PR013254、PR013371、PR013377、PR013393和PR013397。In one embodiment, the DLL3 binding molecules include antibodies to DLL3, such as PR013234, PR013235, PR013236, PR013239, PR013240, PR013241, PR013242, PR013244, PR013246, PR013249, PR013250, PR013252, PR013253, PR013254, PR013256, PR013257, PR013258, PR013259, PR013260, PR013261, PR013262, PR013263, PR013264, PR013363, PR013364, 13240, PR013254, PR013371, PR013377, PR013393, and PR013397.

抗体PR013234Antibody PR013234

在一方面，本发明提供一种针对DLL3的抗体PR013234或其抗原结合片段，In one aspect, the present invention provides an antibody PR013234 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:18所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:18,

HCDR2，其包含SEQ ID NO:60所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:60, and

HCDR3，其包含SEQ ID NO:126所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:126.

在一实施方案中，DLL3抗体PR013234包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013234 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:174所示氨基酸序列或与SEQ ID NO:174具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 174 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 174.

在一实施方案中，DLL3抗体PR013234包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013234包含SEQ ID NO:222所示氨基酸序列或与SEQ ID NO:222具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013234 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013234 comprises the amino acid sequence shown in SEQ ID NO:222 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:222.

抗体PR013235Antibody PR013235

在又一方面，本发明提供一种针对DLL3的抗体PR013235或其抗原结合片段，In another aspect, the present invention provides an antibody PR013235 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:19所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO: 19,

HCDR2，其包含SEQ ID NO:60所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:60, and

HCDR3，其包含SEQ ID NO:126所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:126.

在一实施方案中，DLL3抗体PR013235包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013235 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:175所示氨基酸序列或与SEQ ID NO:175具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 175 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 175.

在一实施方案中，DLL3抗体PR013235包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013235包含SEQ ID NO:223所示氨基酸序列或与SEQ ID NO:223具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013235 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013235 comprises the amino acid sequence shown in SEQ ID NO:223 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:223.

抗体PR013236Antibody PR013236

在又一方面，本发明提供一种针对DLL3的抗体PR013236或其抗原结合片段，In another aspect, the present invention provides an antibody PR013236 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:18所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:18,

HCDR2，其包含SEQ ID NO:61所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:61, and

HCDR3，其包含SEQ ID NO:126所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:126.

在一实施方案中，DLL3抗体PR013236包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013236 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:176所示氨基酸序列或与SEQ ID NO:176具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。wherein the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 176 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 176.

在一实施方案中，DLL3抗体PR013236包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013236包含SEQ ID NO:224所示氨基酸序列或与SEQ ID NO:224具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013236 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013236 comprises the amino acid sequence shown in SEQ ID NO:224 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:224.

抗体PR013239Antibody PR013239

在又一方面，本发明提供一种针对DLL3的抗体PR013239或其抗原结合片段，In another aspect, the present invention provides an antibody PR013239 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:18所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:18,

HCDR2，其包含SEQ ID NO:60所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:60, and

HCDR3，其包含SEQ ID NO:126所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:126.

在一实施方案中，DLL3抗体PR013239包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013239 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:177所示氨基酸序列或与SEQ ID NO:177具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。wherein the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 177 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 177.

在一实施方案中，DLL3抗体PR013239包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013239包含SEQ ID NO:225所示氨基酸序列或与SEQ ID NO:225具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013239 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013239 comprises the amino acid sequence shown in SEQ ID NO:225 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:225.

抗体PR013240Antibody PR013240

在又一方面，本发明提供一种针对DLL3的抗体PR013240或其抗原结合片段，In another aspect, the present invention provides an antibody PR013240 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:20所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:20,

HCDR2，其包含SEQ ID NO:62所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:62, and

HCDR3，其包含SEQ ID NO:127所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:127.

在一实施方案中，DLL3抗体PR013240包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013240 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:178所示氨基酸序列或与SEQ ID NO:178具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 178 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 178.

在一实施方案中，DLL3抗体PR013240包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013240包含SEQ ID NO:226所示氨基酸序列或与SEQ ID NO:226具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013240 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013240 comprises the amino acid sequence shown in SEQ ID NO:226 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:226.

抗体PR013241Antibody PR013241

在又一方面，本发明提供一种针对DLL3的抗体PR013241或其抗原结合片段，In another aspect, the present invention provides an antibody PR013241 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:18所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:18,

HCDR2，其包含SEQ ID NO:63所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:63, and

HCDR3，其包含SEQ ID NO:128所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:128.

在一实施方案中，DLL3抗体PR013241包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013241 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:179所示氨基酸序列或与SEQ ID NO:179具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 179 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 179.

在一实施方案中，DLL3抗体PR013241包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013241包含SEQ ID NO:227所示氨基酸序列或与SEQ ID NO:227具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013241 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013241 comprises the amino acid sequence shown in SEQ ID NO:227 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:227.

抗体PR013242Antibody PR013242

在又一方面，本发明提供一种针对DLL3的抗体PR013242或其抗原结合片段，In another aspect, the present invention provides an antibody PR013242 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:21所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:21,

HCDR2，其包含SEQ ID NO:64所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:64, and

HCDR3，其包含SEQ ID NO:129所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:129.

在一实施方案中，DLL3抗体PR013242包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013242 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:180所示氨基酸序列或与SEQ ID NO:180具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 180 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 180.

在一实施方案中，DLL3抗体PR013242包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013242包含SEQ ID NO:228所示氨基酸序列或与SEQ ID NO:228具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013242 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013242 comprises the amino acid sequence shown in SEQ ID NO:228 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:228.

抗体PR013244Antibody PR013244

在又一方面，本发明提供一种针对DLL3的抗体PR013244或其抗原结合片段，In another aspect, the present invention provides an antibody PR013244 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:22所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:22,

HCDR2，其包含SEQ ID NO:65所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:65, and

HCDR3，其包含SEQ ID NO:126所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:126.

在一实施方案中，DLL3抗体PR013244包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013244 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:181所示氨基酸序列或与SEQ ID NO:181具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 181 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 181.

在一实施方案中，DLL3抗体PR013244包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013244包含SEQ ID NO:229所示氨基酸序列或与SEQ ID NO:229具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013244 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013244 comprises the amino acid sequence shown in SEQ ID NO:229 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:229.

抗体PR013246Antibody PR013246

在又一方面，本发明提供一种针对DLL3的抗体PR013246或其抗原结合片段，In another aspect, the present invention provides an antibody PR013246 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:23所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:23,

HCDR2，其包含SEQ ID NO:65所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:65, and

HCDR3，其包含SEQ ID NO:126所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:126.

在一实施方案中，DLL3抗体PR013246包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013246 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:182所示氨基酸序列或与SEQ ID NO:182具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 182 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 182.

在一实施方案中，DLL3抗体PR013246包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013246包含SEQ ID NO:230所示氨基酸序列或与SEQ ID NO:230具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013246 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013246 comprises the amino acid sequence shown in SEQ ID NO:230 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:230.

抗体PR013249Antibody PR013249

在又一方面，本发明提供一种针对DLL3的抗体PR013249或其抗原结合片段，In another aspect, the present invention provides an antibody PR013249 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:18所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:18,

HCDR2，其包含SEQ ID NO:66所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:66, and

HCDR3，其包含SEQ ID NO:126所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:126.

在一实施方案中，DLL3抗体PR013249包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013249 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:183所示氨基酸序列或与SEQ ID NO:183具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 183 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 183.

在一实施方案中，DLL3抗体PR013249包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013249包含SEQ ID NO:231所示氨基酸序列或与SEQ ID NO:231具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013249 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013249 comprises the amino acid sequence shown in SEQ ID NO:231 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:231.

抗体PR013250Antibody PR013250

在又一方面，本发明提供一种针对DLL3的抗体PR013250或其抗原结合片段，In another aspect, the present invention provides an antibody PR013250 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:18所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:18,

HCDR2，其包含SEQ ID NO:67所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:67, and

HCDR3，其包含SEQ ID NO:130所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:130.

在一实施方案中，DLL3抗体PR013250包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013250 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:184所示氨基酸序列或与SEQ ID NO:184具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 184 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 184.

在一实施方案中，DLL3抗体PR013250包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013250包含SEQ ID NO:232所示氨基酸序列或与SEQ ID NO:232具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013250 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013250 comprises the amino acid sequence shown in SEQ ID NO:232 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:232.

抗体PR013252Antibody PR013252

在又一方面，本发明提供一种针对DLL3的抗体PR013252或其抗原结合片段，In another aspect, the present invention provides an antibody PR013252 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:18所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:18,

HCDR2，其包含SEQ ID NO:64所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:64, and

HCDR3，其包含SEQ ID NO:131所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:131.

在一实施方案中，DLL3抗体PR013252包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013252 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:185所示氨基酸序列或与SEQ ID NO:185具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 185 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 185.

在一实施方案中，DLL3抗体PR013252包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013252包含SEQ ID NO:233所示氨基酸序列或与SEQ ID NO:233具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013252 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013252 comprises the amino acid sequence shown in SEQ ID NO:233 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:233.

抗体PR013253Antibody PR013253

在又一方面，本发明提供一种针对DLL3的抗体PR013253或其抗原结合片段，In another aspect, the present invention provides an antibody PR013253 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:19所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:19,

HCDR2，其包含SEQ ID NO:68所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:68, and

HCDR3，其包含SEQ ID NO:126所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:126.

在一实施方案中，DLL3抗体PR013253包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013253 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:186所示氨基酸序列或与SEQ ID NO:186具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 186 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 186.

在一实施方案中，DLL3抗体PR013253包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013253包含SEQ ID NO:234所示氨基酸序列或与SEQ ID NO:234具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013253 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013253 comprises the amino acid sequence shown in SEQ ID NO:234 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:234.

抗体PR013254Antibody PR013254

在又一方面，本发明提供一种针对DLL3的抗体PR013254或其抗原结合片段，In another aspect, the present invention provides an antibody PR013254 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:18所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:18,

HCDR2，其包含SEQ ID NO:63所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:63, and

HCDR3，其包含SEQ ID NO:128所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:128.

在一实施方案中，DLL3抗体PR013254包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013254 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:187所示氨基酸序列或与SEQ ID NO:187具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 187 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 187.

在一实施方案中，DLL3抗体PR013254包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013254包含SEQ ID NO:235所示氨基酸序列或与SEQ ID NO:235具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013254 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013254 comprises the amino acid sequence shown in SEQ ID NO:235 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:235.

抗体PR013256Antibody PR013256

在又一方面，本发明提供一种针对DLL3的抗体PR013256或其抗原结合片段，In another aspect, the present invention provides an antibody PR013256 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:18所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:18,

HCDR2，其包含SEQ ID NO:68所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:68, and

HCDR3，其包含SEQ ID NO:126所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:126.

在一实施方案中，DLL3抗体PR013256包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013256 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:188所示氨基酸序列或与SEQ ID NO:188具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 188 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 188.

在一实施方案中，DLL3抗体PR013256包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013256包含SEQ ID NO:236所示氨基酸序列或与SEQ ID NO:236具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013256 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013256 comprises the amino acid sequence shown in SEQ ID NO:236 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:236.

抗体PR013257Antibody PR013257

在又一方面，本发明提供一种针对DLL3的抗体PR013257或其抗原结合片段，In another aspect, the present invention provides an antibody PR013257 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:20所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:20,

HCDR2，其包含SEQ ID NO:69所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:69, and

HCDR3，其包含SEQ ID NO:132所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:132.

在一实施方案中，DLL3抗体PR013257包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013257 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:189所示氨基酸序列或与SEQ ID NO:189具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 189 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 189.

在一实施方案中，DLL3抗体PR013257包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013257包含SEQ ID NO:237所示氨基酸序列或与SEQ ID NO:237具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013257 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013257 comprises the amino acid sequence shown in SEQ ID NO:237 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:237.

抗体PR013258Antibody PR013258

在又一方面，本发明提供一种针对DLL3的抗体PR013258或其抗原结合片段，In another aspect, the present invention provides an antibody PR013258 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:24所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:24,

HCDR2，其包含SEQ ID NO:66所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:66, and

HCDR3，其包含SEQ ID NO:126所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:126.

在一实施方案中，DLL3抗体PR013258包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013258 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:190所示氨基酸序列或与SEQ ID NO:190具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。wherein the heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 190 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 190.

在一实施方案中，DLL3抗体PR013258包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013258包含SEQ ID NO:238所示氨基酸序列或与SEQ ID NO:238具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013258 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013258 comprises the amino acid sequence shown in SEQ ID NO:238 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:238.

抗体PR013259Antibody PR013259

在又一方面，本发明提供一种针对DLL3的抗体PR013259或其抗原结合片段，In another aspect, the present invention provides an antibody PR013259 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:18所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:18,

HCDR2，其包含SEQ ID NO:70所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:70, and

HCDR3，其包含SEQ ID NO:133所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:133.

在一实施方案中，DLL3抗体PR013259包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013259 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:191所示氨基酸序列或与SEQ ID NO:191具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 191 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 191.

在一实施方案中，DLL3抗体PR013259包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013259包含SEQ ID NO:239所示氨基酸序列或与SEQ ID NO:239具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013259 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013259 comprises the amino acid sequence shown in SEQ ID NO:239 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:239.

抗体PR013260Antibody PR013260

在又一方面，本发明提供一种针对DLL3的抗体PR013260或其抗原结合片段，In another aspect, the present invention provides an antibody PR013260 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:18所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:18,

HCDR2，其包含SEQ ID NO:70所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:70, and

HCDR3，其包含SEQ ID NO:134所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:134.

在一实施方案中，DLL3抗体PR013260包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013260 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:192所示氨基酸序列或与SEQ ID NO:192具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 192 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 192.

在一实施方案中，DLL3抗体PR013260包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013260包含SEQ ID NO:240所示氨基酸序列或与SEQ ID NO:240具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013260 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013260 comprises the amino acid sequence shown in SEQ ID NO: 240 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 240.

抗体PR013261Antibody PR013261

在又一方面，本发明提供一种针对DLL3的抗体PR013261或其抗原结合片段，In another aspect, the present invention provides an antibody PR013261 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:18所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:18,

HCDR2，其包含SEQ ID NO:71所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:71, and

HCDR3，其包含SEQ ID NO:134所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:134.

在一实施方案中，DLL3抗体PR013261包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013261 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:193所示氨基酸序列或与SEQ ID NO:193具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 193 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 193.

在一实施方案中，DLL3抗体PR013261包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013261包含SEQ ID NO:241所示氨基酸序列或与SEQ ID NO:241具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013261 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013261 comprises the amino acid sequence shown in SEQ ID NO:241 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:241.

抗体PR013262Antibody PR013262

在又一方面，本发明提供一种针对DLL3的抗体PR013262或其抗原结合片段，In another aspect, the present invention provides an antibody PR013262 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:25所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:25,

HCDR2，其包含SEQ ID NO:72所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:72, and

HCDR3，其包含SEQ ID NO:135所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:135.

在一实施方案中，DLL3抗体PR013262包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013262 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:194所示氨基酸序列或与SEQ ID NO:194具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 194 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 194.

在一实施方案中，DLL3抗体PR013262包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013262包含SEQ ID NO:242所示氨基酸序列或与SEQ ID NO:242具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013262 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013262 comprises the amino acid sequence shown in SEQ ID NO:242 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:242.

抗体PR013263Antibody PR013263

在又一方面，本发明提供一种针对DLL3的抗体PR013263或其抗原结合片段，In another aspect, the present invention provides an antibody PR013263 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:18所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:18,

HCDR2，其包含SEQ ID NO:73所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:73, and

HCDR3，其包含SEQ ID NO:134所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:134.

在一实施方案中，DLL3抗体PR013263包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013263 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:195所示氨基酸序列或与SEQ ID NO:195具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 195 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 195.

在一实施方案中，DLL3抗体PR013263包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013263包含SEQ ID NO:243所示氨基酸序列或与SEQ ID NO:243具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013263 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013263 comprises the amino acid sequence shown in SEQ ID NO:243 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:243.

抗体PR013264Antibody PR013264

在又一方面，本发明提供一种针对DLL3的抗体PR013264或其抗原结合片段，In another aspect, the present invention provides an antibody PR013264 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:25所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:25,

HCDR2，其包含SEQ ID NO:74所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:74, and

HCDR3，其包含SEQ ID NO:135所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:135.

在一实施方案中，DLL3抗体PR013264包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013264 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:196所示氨基酸序列或与SEQ ID NO:196具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 196 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 196.

在一实施方案中，DLL3抗体PR013264包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013264包含SEQ ID NO:244所示氨基酸序列或与SEQ ID NO:244具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013264 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013264 comprises the amino acid sequence shown in SEQ ID NO:244 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:244.

抗体PR013363Antibody PR013363

在又一方面，本发明提供一种针对DLL3的抗体PR013363或其抗原结合片段，In another aspect, the present invention provides an antibody PR013363 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:26所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:26,

HCDR2，其包含SEQ ID NO:75所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:75, and

HCDR3，其包含SEQ ID NO:136所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:136.

在一实施方案中，DLL3抗体PR013363包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013363 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:197所示氨基酸序列或与SEQ ID NO:197具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 197 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 197.

在一实施方案中，DLL3抗体PR013363包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013363包含SEQ ID NO:245所示氨基酸序列或与SEQ ID NO:245具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013363 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013363 comprises the amino acid sequence shown in SEQ ID NO:245 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:245.

抗体PR013364Antibody PR013364

在又一方面，本发明提供一种针对DLL3的抗体PR013364或其抗原结合片段，In another aspect, the present invention provides an antibody PR013364 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:27所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:27,

HCDR2，其包含SEQ ID NO:76所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:76, and

HCDR3，其包含SEQ ID NO:137所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:137.

在一实施方案中，DLL3抗体PR013364包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013364 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:198所示氨基酸序列或与SEQ ID NO:198具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 198 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 198.

在一实施方案中，DLL3抗体PR013364包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013364包含SEQ ID NO:246所示氨基酸序列或与SEQ ID NO:246具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013364 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013364 comprises the amino acid sequence shown in SEQ ID NO:246 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:246.

抗体PR013367Antibody PR013367

在又一方面，本发明提供一种针对DLL3的抗体PR013367或其抗原结合片段，In another aspect, the present invention provides an antibody PR013367 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:25所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:25,

HCDR2，其包含SEQ ID NO:77所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:77, and

HCDR3，其包含SEQ ID NO:138所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:138.

在一实施方案中，DLL3抗体PR013367包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013367 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:199所示氨基酸序列或与SEQ ID NO:199具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 199 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 199.

在一实施方案中，DLL3抗体PR013367包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013367包含SEQ ID NO:247所示氨基酸序列或与SEQ ID NO:247具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013367 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013367 comprises the amino acid sequence shown in SEQ ID NO:247 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:247.

抗体PR013368Antibody PR013368

在又一方面，本发明提供一种针对DLL3的抗体PR013368或其抗原结合片段，In another aspect, the present invention provides an antibody PR013368 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:25所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:25,

HCDR2，其包含SEQ ID NO:78所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:78, and

HCDR3，其包含SEQ ID NO:139所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:139.

在一实施方案中，DLL3抗体PR013368包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013368 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:200所示氨基酸序列或与SEQ ID NO:200具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 200 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 200.

在一实施方案中，DLL3抗体PR013368包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013368包含SEQ ID NO:248所示氨基酸序列或与SEQ ID NO:248具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013368 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013368 comprises the amino acid sequence shown in SEQ ID NO:248 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:248.

抗体PR013370Antibody PR013370

在又一方面，本发明提供一种针对DLL3的抗体PR013370或其抗原结合片段，In another aspect, the present invention provides an antibody PR013370 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:18所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:18,

HCDR2，其包含SEQ ID NO:79所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:79, and

HCDR3，其包含SEQ ID NO:140所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:140.

在一实施方案中，DLL3抗体PR013370包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013370 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:201所示氨基酸序列或与SEQ ID NO:201具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 201 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 201.

在一实施方案中，DLL3抗体PR013370包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013370包含SEQ ID NO:249所示氨基酸序列或与SEQ ID NO:249具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013370 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013370 comprises the amino acid sequence shown in SEQ ID NO:249 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:249.

抗体PR013371Antibody PR013371

在又一方面，本发明提供一种针对DLL3的抗体PR013371或其抗原结合片段，In another aspect, the present invention provides an antibody PR013371 or an antigen-binding fragment thereof against DLL3.

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:26所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:26,

HCDR2，其包含SEQ ID NO:80所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:80, and

HCDR3，其包含SEQ ID NO:141所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:141.

在一实施方案中，DLL3抗体PR013371包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013371 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:202所示氨基酸序列或与SEQ ID NO:202具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 202 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 202.

在一实施方案中，DLL3抗体PR013371包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013371包含SEQ ID NO:250所示氨基酸序列或与SEQ ID NO:250具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013371 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013371 comprises the amino acid sequence shown in SEQ ID NO: 250 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 250.

抗体PR013372Antibody PR013372

在又一方面，本发明提供一种针对DLL3的抗体PR013372或其抗原结合片段，In another aspect, the present invention provides an antibody PR013372 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:26所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:26,

HCDR2，其包含SEQ ID NO:81所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:81, and

HCDR3，其包含SEQ ID NO:142所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:142.

在一实施方案中，DLL3抗体PR013372包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013372 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:203所示氨基酸序列或与SEQ ID NO:203具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 203 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 203.

在一实施方案中，DLL3抗体PR013372包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013372包含SEQ ID NO:251所示氨基酸序列或与SEQ ID NO:251具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013372 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013372 comprises the amino acid sequence shown in SEQ ID NO:251 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:251.

抗体PR013376Antibody PR013376

在又一方面，本发明提供一种针对DLL3的抗体PR013376或其抗原结合片段，In another aspect, the present invention provides an antibody PR013376 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:25所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:25,

HCDR2，其包含SEQ ID NO:82所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:82, and

HCDR3，其包含SEQ ID NO:143所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:143.

在一实施方案中，DLL3抗体PR013376包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013376 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:204所示氨基酸序列或与SEQ ID NO:204具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 204 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 204.

在一实施方案中，DLL3抗体PR013376包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013376包含SEQ ID NO:252所示氨基酸序列或与SEQ ID NO:252具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013376 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013376 comprises the amino acid sequence shown in SEQ ID NO:252 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:252.

抗体PR013377Antibody PR013377

在又一方面，本发明提供一种针对DLL3的抗体PR013377或其抗原结合片段，In another aspect, the present invention provides an antibody PR013377 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:18所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:18,

HCDR2，其包含SEQ ID NO:60所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:60, and

HCDR3，其包含SEQ ID NO:126所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:126.

在一实施方案中，DLL3抗体PR013377包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013377 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:205所示氨基酸序列或与SEQ ID NO:205具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 205 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 205.

在一实施方案中，DLL3抗体PR013377包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013377包含SEQ ID NO:253所示氨基酸序列或与SEQ ID NO:253具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013377 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013377 comprises the amino acid sequence shown in SEQ ID NO:253 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:253.

抗体PR013378Antibody PR013378

在又一方面，本发明提供一种针对DLL3的抗体PR013378或其抗原结合片段，In another aspect, the present invention provides an antibody PR013378 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:28所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:28,

HCDR2，其包含SEQ ID NO:83所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:83, and

HCDR3，其包含SEQ ID NO:144所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:144.

在一实施方案中，DLL3抗体PR013378包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013378 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:206所示氨基酸序列或与SEQ ID NO:206具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 206 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 206.

在一实施方案中，DLL3抗体PR013378包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013378包含SEQ ID NO:254所示氨基酸序列或与SEQ ID NO:254具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013378 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013378 comprises the amino acid sequence shown in SEQ ID NO:254 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:254.

抗体PR013381Antibody PR013381

在又一方面，本发明提供一种针对DLL3的抗体PR013381或其抗原结合片段，In another aspect, the present invention provides an antibody PR013381 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:29所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:29,

HCDR2，其包含SEQ ID NO:84所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:84, and

HCDR3，其包含SEQ ID NO:143所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:143.

在一实施方案中，DLL3抗体PR013381包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013381 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:207所示氨基酸序列或与SEQ ID NO:207具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 207 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 207.

在一实施方案中，DLL3抗体PR013381包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013381包含SEQ ID NO:255所示氨基酸序列或与SEQ ID NO:255具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013381 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013381 comprises the amino acid sequence shown in SEQ ID NO:255 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:255.

抗体PR013382Antibody PR013382

在又一方面，本发明提供一种针对DLL3的抗体PR013382或其抗原结合片段，In another aspect, the present invention provides an antibody PR013382 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:26所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:26,

HCDR2，其包含SEQ ID NO:85所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:85, and

HCDR3，其包含SEQ ID NO:145所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:145.

在一实施方案中，DLL3抗体PR013382包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013382 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:208所示氨基酸序列或与SEQ ID NO:208具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 208 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 208.

在一实施方案中，DLL3抗体PR013382包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013382包含SEQ ID NO:256所示氨基酸序列或与SEQ ID NO:256具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013382 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013382 comprises the amino acid sequence shown in SEQ ID NO:256 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:256.

抗体PR013384Antibody PR013384

在又一方面，本发明提供一种针对DLL3的抗体PR013384或其抗原结合片段，In another aspect, the present invention provides an antibody PR013384 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:30所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:30,

HCDR2，其包含SEQ ID NO:75所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:75, and

HCDR3，其包含SEQ ID NO:136所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:136.

在一实施方案中，DLL3抗体PR013384包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013384 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:209所示氨基酸序列或与SEQ ID NO:209具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 209 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 209.

在一实施方案中，DLL3抗体PR013384包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013384包含SEQ ID NO:257所示氨基酸序列或与SEQ ID NO:257具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013384 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013384 comprises the amino acid sequence shown in SEQ ID NO:257 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:257.

抗体PR013387Antibody PR013387

在又一方面，本发明提供一种针对DLL3的抗体PR013387或其抗原结合片段，In another aspect, the present invention provides an antibody PR013387 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:31所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:31,

HCDR2，其包含SEQ ID NO:85所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:85, and

HCDR3，其包含SEQ ID NO:146所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:146.

在一实施方案中，DLL3抗体PR013387包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013387 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:210所示氨基酸序列或与SEQ ID NO:210具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 210 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 210.

在一实施方案中，DLL3抗体PR013387包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013387包含SEQ ID NO:258所示氨基酸序列或与SEQ ID NO:258具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013387 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013387 comprises the amino acid sequence shown in SEQ ID NO:258 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:258.

抗体PR013392Antibody PR013392

在又一方面，本发明提供一种针对DLL3的抗体PR013392或其抗原结合片段，In another aspect, the present invention provides an antibody PR013392 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:18所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:18,

HCDR2，其包含SEQ ID NO:70所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:70, and

HCDR3，其包含SEQ ID NO:134所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:134.

在一实施方案中，DLL3抗体PR013392包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013392 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:211所示氨基酸序列或与SEQ ID NO:211具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 211 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 211.

在一实施方案中，DLL3抗体PR013392包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013392包含SEQ ID NO:259所示氨基酸序列或与SEQ ID NO:259具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013392 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013392 comprises the amino acid sequence shown in SEQ ID NO:259 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:259.

抗体PR013393Antibody PR013393

在又一方面，本发明提供一种针对DLL3的抗体PR013393或其抗原结合片段，In another aspect, the present invention provides an antibody PR013393 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:32所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:32,

HCDR2，其包含SEQ ID NO:86所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:86, and

HCDR3，其包含SEQ ID NO:147所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:147.

在一实施方案中，DLL3抗体PR013393包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013393 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:212所示氨基酸序列或与SEQ ID NO:212具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 212 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 212.

在一实施方案中，DLL3抗体PR013393包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013393包含SEQ ID NO:260所示氨基酸序列或与SEQ ID NO:260具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013393 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013393 comprises the amino acid sequence shown in SEQ ID NO:260 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:260.

抗体PR013394Antibody PR013394

在又一方面，本发明提供一种针对DLL3的抗体PR013394或其抗原结合片段，In another aspect, the present invention provides an antibody PR013394 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:18所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:18,

HCDR2，其包含SEQ ID NO:85所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:85, and

HCDR3，其包含SEQ ID NO:148所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:148.

在一实施方案中，DLL3抗体PR013394包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013394 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:213所示氨基酸序列或与SEQ ID NO:213具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 213 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 213.

在一实施方案中，DLL3抗体PR013394包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013394包含SEQ ID NO:261所示氨基酸序列或与SEQ ID NO:261具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013394 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013394 comprises the amino acid sequence shown in SEQ ID NO:261 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:261.

抗体PR013397Antibody PR013397

在又一方面，本发明提供一种针对DLL3的抗体PR013397或其抗原结合片段，In another aspect, the present invention provides an antibody PR013397 or an antigen-binding fragment thereof against DLL3.

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:33所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:33,

HCDR2，其包含SEQ ID NO:87所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:87, and

HCDR3，其包含SEQ ID NO:149所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:149.

在一实施方案中，DLL3抗体PR013397包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013397 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:214所示氨基酸序列或与SEQ ID NO:214具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 214 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 214.

在一实施方案中，DLL3抗体PR013397包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013397包含SEQ ID NO:262所示氨基酸序列或与SEQ ID NO:262具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013397 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013397 comprises the amino acid sequence shown in SEQ ID NO:262 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:262.

抗体PR013399Antibody PR013399

在又一方面，本发明提供一种针对DLL3的抗体PR013399或其抗原结合片段，In another aspect, the present invention provides an antibody PR013399 or an antigen-binding fragment thereof against DLL3.

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:26所示氨基酸序列，HCDR1 comprises the amino acid sequence shown in SEQ ID NO:26,

HCDR2，其包含SEQ ID NO:88所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:88, and

HCDR3，其包含SEQ ID NO:150所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:150.

在一实施方案中，DLL3抗体PR013399包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013399 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:215所示氨基酸序列或与SEQ ID NO:215具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 215 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 215.

在一实施方案中，DLL3抗体PR013399包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013399包含SEQ ID NO:263所示氨基酸序列或与SEQ ID NO:263具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013399 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013399 comprises the amino acid sequence shown in SEQ ID NO:263 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:263.

抗体PR013402Antibody PR013402

在又一方面，本发明提供一种针对DLL3的抗体PR013402或其抗原结合片段，In another aspect, the present invention provides an antibody PR013402 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:18所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:18,

HCDR2，其包含SEQ ID NO:89所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:89, and

HCDR3，其包含SEQ ID NO:151所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:151.

在一实施方案中，DLL3抗体PR013402包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013402 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:216所示氨基酸序列或与SEQ ID NO:216具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 216 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 216.

在一实施方案中，DLL3抗体PR013402包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013402包含SEQ ID NO:264所示氨基酸序列或与SEQ ID NO:264具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013402 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013402 comprises the amino acid sequence shown in SEQ ID NO:264 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:264.

抗体PR013406Antibody PR013406

在又一方面，本发明提供一种针对DLL3的抗体PR013406或其抗原结合片段，In another aspect, the present invention provides an antibody PR013406 or an antigen-binding fragment thereof against DLL3,

其中所述抗体包含重链可变区，其中wherein the antibody comprises a heavy chain variable region, wherein

所述重链可变区包含：The heavy chain variable region comprises:

HCDR1，其包含SEQ ID NO:34所示氨基酸序列，HCDR1, which comprises the amino acid sequence shown in SEQ ID NO:34,

HCDR2，其包含SEQ ID NO:90所示氨基酸序列，和HCDR2 comprising the amino acid sequence shown in SEQ ID NO:90, and

HCDR3，其包含SEQ ID NO:152所示氨基酸序列。HCDR3, which comprises the amino acid sequence shown in SEQ ID NO:152.

在一实施方案中，DLL3抗体PR013406包含重链可变区(VH)，In one embodiment, the DLL3 antibody PR013406 comprises a heavy chain variable region (VH),

其中所述重链可变区包含SEQ ID NO:217所示氨基酸序列或与SEQ ID NO:217具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。The heavy chain variable region comprises the amino acid sequence shown in SEQ ID NO: 217 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO: 217.

在一实施方案中，DLL3抗体PR013406包含重链可变区和人IgG1的Fc结构域，其中DLL3抗体PR013406包含SEQ ID NO:265所示氨基酸序列或与SEQ ID NO:265具有至少80％、至少85％、至少90％、至少95％或更高序列相同性的氨基酸序列。In one embodiment, the DLL3 antibody PR013406 comprises a heavy chain variable region and an Fc domain of human IgG1, wherein the DLL3 antibody PR013406 comprises the amino acid sequence shown in SEQ ID NO:265 or an amino acid sequence having at least 80%, at least 85%, at least 90%, at least 95% or higher sequence identity with SEQ ID NO:265.

如本文所用，术语“修饰”是指针对天然氨基酸、非天然氨基酸、天然氨基酸多肽或非天然氨基酸多肽的变化的存在。通过合成后修饰天然氨基酸、非天然氨基酸、天然氨基酸多肽或非天然氨基酸多肽；或通过共翻译或通过翻译后修饰天然氨基酸、非天然氨基酸、天然氨基酸多肽或非天然氨基酸多肽，可以获得这样的变化或修饰。形式“经修饰的或未经修饰的”意指所讨论的天然氨基酸、非天然氨基酸、天然氨基酸多肽或非天然氨基酸多肽是任选地修饰的，即，所讨论的天然氨基酸、非天然氨基酸、天然氨基酸多肽或非天然氨基酸多肽可以是经修饰或未经修饰的。As used herein, the term "modification" refers to the presence of changes to a natural amino acid, non-natural amino acid, natural amino acid polypeptide, or non-natural amino acid polypeptide. Such changes or modifications can be obtained by post-synthetic modification of a natural amino acid, non-natural amino acid, natural amino acid polypeptide, or non-natural amino acid polypeptide; or by co-translational or post-translational modification of a natural amino acid, non-natural amino acid, natural amino acid polypeptide, or non-natural amino acid polypeptide. The form "modified or unmodified" means that the natural amino acid, non-natural amino acid, natural amino acid polypeptide, or non-natural amino acid polypeptide in question is optionally modified, i.e., the natural amino acid, non-natural amino acid, natural amino acid polypeptide, or non-natural amino acid polypeptide in question can be modified or unmodified.

本发明的DLL3结合分子可以包括经修饰的或未经修饰的氨基酸，例如在抗体的生产、运输过程或储存过程中，抗体的氨基酸序列可能会发生的一些修饰，例如可能存在的，特别是CDR区的翻译后修饰(Post-translational modifications，PTMs)。在本发明中，所述翻译后修饰可以包括目前如本领域技术人员公知的修饰，包括天冬酰胺脱酰胺化(asparagine deamidation，位点NG、NS、NH等)、天冬氨酸异构化(位点DG、DP等)、N糖基化(位点NxS/T，其中x为除脯氨酸以外的任意氨基酸)及氧化等相关翻译后修饰。本发明DLL3抗体的可能的翻译后修饰如表1所示。The DLL3 binding molecules of the present invention may include modified or unmodified amino acids. For example, during the production, transportation, or storage of the antibody, the amino acid sequence of the antibody may undergo modifications, such as post-translational modifications (PTMs), particularly in the CDR regions. In the present invention, such PTMs may include modifications known to those skilled in the art, including asparagine deamidation (positions NG, NS, NH, etc.), aspartate isomerization (positions DG, DP, etc.), N-glycosylation (positions NxS/T, where x is any amino acid except proline), and oxidation, among other related PTMs. Possible PTMs of the DLL3 antibodies of the present invention are shown in Table 1.

多特异性抗原结合构建体Multispecific antigen-binding constructs

在第二方面，本发明提供一种多特异性抗原结合构建体，其包含结合DLL3的第一抗原结合部分，其中所述第一抗原结合部分包含本发明第一方面的DLL3结合分子。在一优选实施方案中，多特异性抗原结合构建体包含结合DLL3的第一抗原结合部分以及结合第二抗原的第二抗原结合部分，其中所述第一抗原结合部分包含本发明第一方面的DLL3结合分子。In a second aspect, the invention provides a multispecific antigen-binding construct comprising a first antigen-binding moiety that binds DLL3, wherein the first antigen-binding moiety comprises the DLL3-binding molecule of the first aspect of the invention. In a preferred embodiment, the multispecific antigen-binding construct comprises a first antigen-binding moiety that binds DLL3 and a second antigen-binding moiety that binds a second antigen, wherein the first antigen-binding moiety comprises the DLL3-binding molecule of the first aspect of the invention.

如本文所用，“多特异性抗原结合构建体”或“多特异性分子”是指特异性结合一个以上不同抗原或同一抗原内一个以上不同表位的构建体。多特异性抗原结合构建体可以是蛋白、蛋白复合物或抗体。多特异性抗原结合构建体包含抗体或其抗原结合片段，其可以与至少一种其它功能性分子(例如另一种肽或蛋白，诸如受体的另一种抗体或配体)缔合或连接，从而形成与至少两个不同结合位点或靶分子结合的分子。多特异性分子可能对其他相关的抗原具有交叉反应性，例如，对来自其他物种(同源)(诸如人或猴，例如食蟹猕猴或黑猩猩)的相同抗原具有交叉反应性，或者可结合两个或更多个不同抗原之间所共享的表位。示例性的多特异性分子包括多特异性抗体，例如双特异性抗体。As used herein, a "multispecific antigen-binding construct" or "multispecific molecule" refers to a construct that specifically binds to one or more different antigens or one or more different epitopes within the same antigen. A multispecific antigen-binding construct can be a protein, a protein complex, or an antibody. A multispecific antigen-binding construct comprises an antibody or antigen-binding fragment thereof that can be associated or linked to at least one other functional molecule (e.g., another peptide or protein, such as another antibody or ligand for a receptor) to form a molecule that binds to at least two different binding sites or target molecules. A multispecific molecule may have cross-reactivity to other related antigens, for example, to the same antigen from another species (homologous), such as a human or monkey, e.g., a cynomolgus macaque or a chimpanzee, or may bind to an epitope shared between two or more different antigens. Exemplary multispecific molecules include multispecific antibodies, such as bispecific antibodies.

如本文所用，术语“多特异性抗体”是指能够特异性结合两种或更多种(例如2、3、4、5或6种)不同抗原表位的抗体。多特异性抗体可以例如是双特异性、三特异性或四特异性抗体，其分别能够特异性结合2、3或4种抗原表位。如本文所用，术语“抗原表位”或“抗原决定簇”表示抗原中与抗体的抗原结合位点特异性结合的区域。抗原表位通常由抗原的化学活性表面基团(如氨基酸或糖侧链)组成且通常具有特定的三维结构性质以及特定的电荷性质。第二抗原可以为不同于DLL3的其他抗原。第二抗原也可以为DLL3，其与本发明的抗DLL3抗体或其抗原结合片段结合DLL3上不同的抗原表位。可以利用本领域常规的方法确定两种抗体结合的抗原表位是否相同，例如通过ELISA、流式细胞术或表面等离子共振等测定两种抗体对相同抗原表位的竞争结合。As used herein, the term "multispecific antibody" refers to an antibody that specifically binds to two or more (e.g., two, three, four, five, or six) different antigenic epitopes. Multispecific antibodies may be, for example, bispecific, trispecific, or tetraspecific antibodies, each capable of specifically binding to two, three, or four antigenic epitopes, respectively. As used herein, the terms "epitope" or "antigenic determinant" refer to a region of an antigen that specifically binds to the antigen-binding site of an antibody. An antigenic epitope is typically composed of chemically active surface groups (e.g., amino acids or sugar side chains) of an antigen and typically has specific three-dimensional structural properties as well as specific charge properties. The second antigen may be an antigen other than DLL3. The second antigen may also be DLL3, which binds to a different epitope on DLL3 than the anti-DLL3 antibody or antigen-binding fragment thereof of the present invention. Whether the two antibodies bind to the same epitope can be determined using conventional methods in the art, such as by measuring competitive binding of the two antibodies to the same epitope using ELISA, flow cytometry, or surface plasmon resonance.

多特异性抗体可以是多价(例如2、3、4价)抗体，即其具有多个抗原结合位点。利用感兴趣的抗体或抗原结合片段构建多特异性抗体的方法是本领域技术人员熟知的(参见，例如WO 93/08829；Suresh et al.,(1986)Methods in Enzymology,121:210；和Traunecker et al.,(1991)EMBO,10:3655-3659)。可以利用本领域已知的各种技术产生和分离多特异性抗体。例如，可以通过重组DNA技术获得编码多特异性抗体的多核苷酸，任选地将所述多核苷酸克隆入表达载体，然后用所述多核苷酸或表达载体转化宿主细胞，在合适的条件下培养转化的宿主细胞以允许所述多核苷酸或表达载体表达，最后从宿主细胞或培养基中分离和纯化多特异性抗体。还可以分别获得多特异性抗体的各个部分，例如分别获得如本文所述的第一抗原结合部分和第二抗原结合部分，然后通过酶促或化学偶联技术任选地通过接头将各个部分偶联获得特异性结合DLL3和其他抗原的多特异性抗体。Multispecific antibodies can be multivalent (e.g., 2, 3, 4-valent) antibodies, i.e., they have multiple antigen binding sites. Methods for constructing multispecific antibodies using antibodies or antigen-binding fragments of interest are well known to those skilled in the art (see, for example, WO 93/08829; Suresh et al., (1986) Methods in Enzymology, 121: 210; and Traunecker et al., (1991) EMBO, 10: 3655-3659). Multispecific antibodies can be produced and isolated using various techniques known in the art. For example, a polynucleotide encoding a multispecific antibody can be obtained by recombinant DNA technology, optionally cloned into an expression vector, and then transformed into a host cell with the polynucleotide or expression vector, cultured under appropriate conditions to allow expression of the polynucleotide or expression vector, and finally isolated and purified from the host cell or culture medium. The various portions of the multispecific antibody can also be obtained separately, for example, the first antigen-binding portion and the second antigen-binding portion as described herein can be obtained separately, and then the various portions can be coupled by enzymatic or chemical coupling techniques, optionally through a linker, to obtain a multispecific antibody that specifically binds to DLL3 and other antigens.

如本文所用，“第一抗原结合部分”和“第二抗原结合部分”表示包含抗原结合位点的、能够与抗原表位结合的氨基酸序列，其定义落入抗体或抗原结合片段的含义范围内。As used herein, "first antigen-binding portion" and "second antigen-binding portion" refer to amino acid sequences that contain an antigen-binding site and are capable of binding to an antigen epitope, and are defined within the meaning of an antibody or an antigen-binding fragment.

第一抗原结合部分可以是任何形式的抗体或抗原结合片段，包括但不限于Fv、scFv、dsFv、scdsFv、Fab、scFab、Fab'和F(ab')2。在一些实施方案中，第一抗原结合部分包含本发明的DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)。在一实施方案中，第一抗原结合部分包含VH，其包含如上所述本发明的DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)的HCDR1、HCDR2和HCDR3。The first antigen-binding moiety can be any form of antibody or antigen-binding fragment, including but not limited to Fv, scFv, dsFv, scdsFv, Fab, scFab, Fab', and F(ab') 2. In some embodiments, the first antigen-binding moiety comprises a DLL3-binding molecule of the invention (e.g., an anti-DLL3 antibody of the invention, or an antigen-binding fragment thereof). In one embodiment, the first antigen-binding moiety comprises a VH comprising the HCDR1, HCDR2, and HCDR3 of a DLL3-binding molecule of the invention (e.g., an anti-DLL3 antibody of the invention, or an antigen-binding fragment thereof), as described above.

第二抗原结合部分可以是结合任何感兴趣的抗原表位的抗体或抗原结合片段。在一实施方案中，第二抗原为不同于DLL3的其他抗原。第二抗原结合部分可以特异性结合的抗原可以包括：肿瘤抗原(例如肿瘤相关抗原和肿瘤特异性抗原)、免疫调节受体和免疫检查点分子。如本文所用，“肿瘤相关抗原”是指在肿瘤细胞中高表达，健康细胞中也存在但是表达水平较低的抗原。如本文所用，“肿瘤特异性抗原”是指肿瘤细胞中特异性表达，健康细胞中几乎不表达的抗原。肿瘤抗原的非限制性实例可以包括CD19、CD20、EGFR、GPC3、HER-2和FOLR1。免疫检查点分子的非限制性实例可以包括CTLA-4、LAG-3、PD-1、PD-L1和TIM-3。免疫调节受体可以包括例如免疫激活性受体(例如CD27、CD137、CD40、GITR和OX40)和免疫抑制性受体(例如BTLA、CTLA4、LAG-3和PD-1)。例如，第二抗原结合部分可以为免疫激活性受体的激动剂抗体或者免疫抑制性受体的拮抗剂抗体。在一实施方案中，第二抗原结合部分可以特异性结合的抗原可以包括：淋巴细胞诸如T细胞或自然杀伤(NK)细胞上的抗原。在一实施方案中，所淋巴细胞上的所述抗原为CD3、CD3ε(CD3 epsilon)、CD8、KI2L4、NKG2E、NKG2D、NKG2F、BTNL3、CD186、BTNL8、PD-1、CD195或NKG2C。The second antigen binding portion can be an antibody or antigen binding fragment that binds to any antigenic epitope of interest. In one embodiment, the second antigen is an antigen other than DLL3. Antigens that the second antigen binding portion can specifically bind to may include: tumor antigens (e.g., tumor-associated antigens and tumor-specific antigens), immunomodulatory receptors, and immune checkpoint molecules. As used herein, "tumor-associated antigen" refers to an antigen that is highly expressed in tumor cells and also present in healthy cells but at a lower expression level. As used herein, "tumor-specific antigen" refers to an antigen that is specifically expressed in tumor cells and hardly expressed in healthy cells. Non-limiting examples of tumor antigens can include CD19, CD20, EGFR, GPC3, HER-2, and FOLR1. Non-limiting examples of immune checkpoint molecules can include CTLA-4, LAG-3, PD-1, PD-L1, and TIM-3. Immunomodulatory receptors can include, for example, immune-activating receptors (e.g., CD27, CD137, CD40, GITR, and OX40) and immunoinhibitory receptors (e.g., BTLA, CTLA4, LAG-3, and PD-1). For example, the second antigen binding portion can be an agonist antibody for an immune activating receptor or an antagonist antibody for an immune inhibitory receptor. In one embodiment, the antigen to which the second antigen binding portion can specifically bind can include: an antigen on a lymphocyte such as a T cell or a natural killer (NK) cell. In one embodiment, the antigen on the lymphocyte is CD3, CD3ε (CD3 epsilon), CD8, KI2L4, NKG2E, NKG2D, NKG2F, BTNL3, CD186, BTNL8, PD-1, CD195 or NKG2C.

在一实施方案中，第二抗原结合部分特异性结合肿瘤抗原、免疫调节受体和免疫检查点分子。在一实施方案中，第二抗原结合部分特异性结合淋巴细胞诸如T细胞或自然杀伤(NK)细胞上的抗原。In one embodiment, the second antigen binding portion specifically binds to a tumor antigen, an immunomodulatory receptor, and an immune checkpoint molecule. In one embodiment, the second antigen binding portion specifically binds to an antigen on a lymphocyte such as a T cell or a natural killer (NK) cell.

在一实施方案中，第二抗原结合部分特异性结合SIRPα、PD-1、PD-L1、LAG3、TIM-3、CTLA-4、VISTA、GPC3、EGFR、HER-2、CD19、CD20、CD33、CD40、CD73、OX40、CD3、TIP-1、叶酸受体α(FOLR1)和/或其他抗原。In one embodiment, the second antigen binding portion specifically binds SIRPα, PD-1, PD-L1, LAG3, TIM-3, CTLA-4, VISTA, GPC3, EGFR, HER-2, CD19, CD20, CD33, CD40, CD73, OX40, CD3, TIP-1, folate receptor alpha (FOLR1) and/or other antigens.

第二抗原结合部分可以是任何形式的抗体或抗原结合片段，包括但不限于免疫球蛋白的单可变结构域(例如包含羊驼的VHH或鲨鱼的IgNAR可变结构域)、Fv、scFv、dsFv、scdsFv、Fab、scFab、Fab'和F(ab')2。The second antigen binding portion can be any form of antibody or antigen binding fragment, including but not limited to a single variable domain of an immunoglobulin (e.g., comprising a VHH from alpaca or an IgNAR variable domain from shark), Fv, scFv, dsFv, scdsFv, Fab, scFab, Fab', and F(ab') 2 .

第一抗原结合部分和第二抗原结合部分可以任选地通过接头连接。在一些实施方案中，第一抗原结合部分和第二抗原结合部分不通过接头连接。在另一些实施方案中，第一抗原结合部分和第二抗原结合部分通过接头连接，例如肽接头或化学键。优选地，第一抗原结合部分和第二抗原结合部分通过肽接头连接。示例性的肽接头可以包括但不限于聚甘氨酸(G)、聚丙氨酸(A)、聚丝氨酸(S)或其组合，例如GGAS、GGGS、GGGSG或者(G4S)n，其中n为1-20的整数。The first antigen binding portion and the second antigen binding portion can optionally be connected by a linker. In some embodiments, the first antigen binding portion and the second antigen binding portion are not connected by a linker. In other embodiments, the first antigen binding portion and the second antigen binding portion are connected by a linker, such as a peptide linker or a chemical bond. Preferably, the first antigen binding portion and the second antigen binding portion are connected by a peptide linker. Exemplary peptide linkers can include but are not limited to polyglycine (G), polyalanine (A), polyserine (S) or a combination thereof, such as GGAS, GGGS, GGGSG or (G 4 S) n , wherein n is an integer of 1-20.

CAR和表达CAR的免疫效应细胞CAR and CAR-expressing immune effector cells

在第三方面，本发明还提供包含本发明的DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)的嵌合抗原受体(CAR)。如本文所用，术语“嵌合抗原受体(CAR)”是指至少包含胞外结构域、跨膜结构域和胞内T细胞受体活化信号结构域的多肽，所述胞外结构域通过单特异性或多特异性抗体结合或特异性地结合单特异性或多特异性抗体上的靶标。CAR是同时提供抗原结合并激活T细胞功能的重组受体。CAR结构和工程改造描述于，例如Dotti G.et al.,(2014)Immunol Rev.257(1):107-126，其通过援引加入本文。In a third aspect, the present invention also provides a chimeric antigen receptor (CAR) comprising a DLL3 binding molecule of the present invention (e.g., an anti-DLL3 antibody of the present invention or an antigen-binding fragment thereof). As used herein, the term "chimeric antigen receptor (CAR)" refers to a polypeptide comprising at least an extracellular domain, a transmembrane domain, and an intracellular T cell receptor activation signaling domain, wherein the extracellular domain binds to or specifically binds to a target on a monospecific or multispecific antibody through a monospecific or multispecific antibody. CAR is a recombinant receptor that simultaneously provides antigen binding and activates T cell function. CAR structure and engineering are described in, for example, Dotti G. et al., (2014) Immunol Rev. 257(1):107-126, which is incorporated herein by reference.

在一示例性实施方案中，本发明的CAR从N端至C端包含：In an exemplary embodiment, the CAR of the present invention comprises, from N-terminus to C-terminus:

(a)胞外区，其包含本发明的DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)；(a) an extracellular region comprising a DLL3-binding molecule of the invention (e.g., an anti-DLL3 antibody of the invention or an antigen-binding fragment thereof);

(b)跨膜区，其连接胞外区和胞内信号区并将CAR锚定在细胞膜上；以及(b) a transmembrane region that connects the extracellular region and the intracellular signaling region and anchors the CAR to the cell membrane; and

(c)胞内信号区。(c) Intracellular signaling region.

胞外区包含本发明的DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)，使得本发明的CAR与表达DLL3的细胞(例如癌细胞)结合。The extracellular region comprises a DLL3 binding molecule of the invention (e.g., an anti-DLL3 antibody of the invention or an antigen-binding fragment thereof), such that the CAR of the invention binds to cells expressing DLL3 (e.g., cancer cells).

跨膜区连接胞外区和胞内信号区并将CAR锚定在细胞膜上。The transmembrane region connects the extracellular region and the intracellular signaling region and anchors the CAR to the cell membrane.

当CAR的胞外区与其识别的抗原结合时，胞内信号区向胞内传导T细胞受体(TCR)样信号并激活表达CAR的免疫效应细胞发挥其效应功能。When the extracellular region of CAR binds to the antigen it recognizes, the intracellular signaling region transmits T cell receptor (TCR)-like signals into the cell and activates the immune effector cells expressing CAR to exert their effector functions.

在一些实施方案中，胞内信号区还包含至少一个共刺激结构域。共刺激结构域可以促进表达CAR的免疫效应细胞在与胞外区所靶向的抗原结合后的活化。In some embodiments, the intracellular signaling region further comprises at least one co-stimulatory domain. The co-stimulatory domain can promote the activation of CAR-expressing immune effector cells after binding to the antigen targeted by the extracellular region.

在一些实施方案中，本发明的CAR还包含连接胞外区和跨膜区的间隔区。在一实施方案中，间隔区衍生自免疫球蛋白的重链恒定区。In some embodiments, the CAR of the present invention further comprises a spacer connecting the extracellular region and the transmembrane region.In one embodiment, the spacer is derived from the heavy chain constant region of an immunoglobulin.

在又一方面，本发明还涉及编码本发明的CAR的多核苷酸和包含所述多核苷酸的表达载体。In yet another aspect, the present invention also relates to a polynucleotide encoding the CAR of the present invention and an expression vector comprising the polynucleotide.

在第四方面，本发明还提供一种免疫效应细胞，其在细胞表面表达本发明的CAR。优选地，所述免疫效应细胞选自T淋巴细胞(例如细胞毒性T细胞(CTL))、巨噬细胞、自然杀伤细胞(NK)和自然杀伤T细胞(NKT)。免疫效应细胞可以靶向DLL3阳性的病变细胞(例如，DLL3阳性的癌细胞)，并被激活以启动效应功能，例如，引起DLL3阳性的癌细胞的死亡。In a fourth aspect, the present invention also provides an immune effector cell that expresses the CAR of the present invention on the cell surface. Preferably, the immune effector cell is selected from T lymphocytes (e.g., cytotoxic T cells (CTL)), macrophages, natural killer cells (NK) and natural killer T cells (NKT). Immune effector cells can target DLL3-positive diseased cells (e.g., DLL3-positive cancer cells) and be activated to start effector functions, for example, causing the death of DLL3-positive cancer cells.

抗体缀合物Antibody conjugates

在第五方面，本发明还提供了一种抗体缀合物，其包含与至少一种治疗剂缀合的本发明的DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)或者本发明的多特异性抗原结合构建体。抗体-药物缀合物(ADC)是一种典型的抗体缀合物，其中所述治疗剂可以例如为细胞毒性剂。In a fifth aspect, the present invention further provides an antibody conjugate comprising a DLL3-binding molecule of the invention (e.g., an anti-DLL3 antibody or antigen-binding fragment thereof) or a multispecific antigen-binding construct of the invention conjugated to at least one therapeutic agent. An antibody-drug conjugate (ADC) is a typical antibody conjugate, wherein the therapeutic agent can be, for example, a cytotoxic agent.

如本文所用，“缀合”是指两个或多个部分通过共价或非共价作用相互连接。在优选的实施方案中，所述缀合是共价缀合。As used herein, "conjugation" refers to the attachment of two or more moieties to each other through covalent or non-covalent interactions. In a preferred embodiment, the conjugation is covalent conjugation.

治疗剂可以选自细胞毒性剂、治疗性抗体(例如与另外的抗原特异性结合的抗体或其抗原结合片段)、放射性同位素、寡核苷酸及其类似物(例如干扰RNA)、生物活性肽、蛋白毒素(例如白喉毒素、蓖麻毒素)和酶(例如脲酶)。The therapeutic agent can be selected from cytotoxic agents, therapeutic antibodies (e.g., antibodies or antigen-binding fragments thereof that specifically bind to another antigen), radioactive isotopes, oligonucleotides and their analogs (e.g., interfering RNA), biologically active peptides, protein toxins (e.g., diphtheria toxin, ricin toxin), and enzymes (e.g., urease).

细胞毒性剂是指抑制或降低细胞的活性、功能和/或杀死细胞的物质。细胞毒性剂的实例可以包括但不限于：类美登素(maytansinoid)(例如美登素(maytansine)、奥利斯他汀类(例如MMAF、MMAE、MMAD)、多司他汀(duostatin)、念珠藻环肽(cryptophycin)、长春花生物碱类(例如长春碱、长春新碱)、秋水仙碱类、海兔毒素类、紫杉烷、紫杉醇、多西他赛、卡巴他赛、烯二炔类抗生素、细胞松弛素类、喜树碱类、蒽环类抗生素(例如道诺霉素(daunorubicin)、二羟基蒽二酮(dihydroxyanthracindione)、多柔比星(doxorubicin))、细胞毒性抗生素(例如丝裂霉素(mitomycin)、放线菌素(actinomycin)、倍癌霉素(duocarmycin)(例如CC-1065)、金霉素(auromycin)、多霉素(duomycin)、卡奇霉素(calicheamicin)、内孢霉素(endomycin)、酚霉素(phenomycin))、阿霉素、柔红霉素、卡奇霉素、顺铂(cisplatin)、溴化乙锭(ethidiumbromide)、博来霉素、丝裂霉素、光辉霉素、普拉地内酯、鬼臼毒素、依托泊苷(etoposide)、米托蒽醌、5-氟尿嘧啶、阿糖胞苷、吉西他滨、巯嘌呤、喷司他丁、氟达拉滨、克拉屈滨、奈拉滨、卡莫司汀、洛莫司汀、甲氨蝶呤、苯丙氨酸氮芥、替尼泊苷(tenoposide)、糖皮质激素等。Cytotoxic agents refer to substances that inhibit or reduce the activity, function and/or kill cells. Examples of cytotoxic agents may include, but are not limited to, maytansinoids (e.g., maytansine), auristatins (e.g., MMAF, MMAE, MMAD), duostatins, cryptophycins, vinca alkaloids (e.g., vinblastine, vincristine), colchicines, dolastatins, taxanes, paclitaxel, docetaxel, cabazitaxel, enediyne antibiotics, cytochalasins, camptothecins, anthracyclines (e.g., daunorubicin, dihydroxyanthracindione, doxorubicin), cytotoxic antibiotics (e.g., mitomycin, actinomycin), dactinomycin, dactin, dactycin, dactin ... [0013] Examples of the present invention include, but are not limited to, chloramphenicol, chlortetracycline ...

放射性同位素可以选自例如212Bi、213Bi、131I、125I、111In、177Lu、186Re、188Re、153Sm、90Y。The radioactive isotope may be selected from, for example, 212 Bi, 213 Bi, 131 I, 125 I, 111 In, 177 Lu, 186 Re, 188 Re, 153 Sm, 90 Y.

在一些实施方案中，治疗剂选自细胞毒性剂、化疗剂、放射性同位素、免疫检查点抑制剂、靶向肿瘤特异性抗原的抗体和其他抗肿瘤药物。在一优选实施方案中，治疗剂为细胞毒性剂。在又一优选实施方案中，治疗剂为放射性同位素。在又一优选实施方案中，治疗剂为免疫检查点抑制剂。In some embodiments, the therapeutic agent is selected from cytotoxic agents, chemotherapeutic agents, radioisotopes, immune checkpoint inhibitors, antibodies targeting tumor-specific antigens, and other anti-tumor drugs. In a preferred embodiment, the therapeutic agent is a cytotoxic agent. In another preferred embodiment, the therapeutic agent is a radioisotope. In another preferred embodiment, the therapeutic agent is an immune checkpoint inhibitor.

可以利用本领域已知的任何技术将治疗剂与本发明的本发明的DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)或者本发明的多特异性抗原结合构建体通过接头缀合。所述接头可以包含用于共价缀合的活性基团，例如胺、羟胺、马来酰亚胺基、羧基、苯基、硫醇、巯基或羟基。The therapeutic agent can be conjugated to the DLL3-binding molecule of the invention (e.g., an anti-DLL3 antibody or antigen-binding fragment thereof) or the multispecific antigen-binding construct of the invention via a linker using any technique known in the art. The linker can comprise a reactive group for covalent conjugation, such as an amine, hydroxylamine, maleimido, carboxyl, phenyl, thiol, sulfhydryl, or hydroxyl group.

药物组合物Pharmaceutical composition

在第六方面，本发明还提供一种药物组合物，其包含本发明的DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)、多特异性抗原结合构建体、免疫效应细胞或者抗体缀合物，以及药学上可接受的载剂。In a sixth aspect, the present invention also provides a pharmaceutical composition comprising a DLL3 binding molecule of the present invention (e.g., an anti-DLL3 antibody or antigen-binding fragment thereof of the present invention), a multispecific antigen-binding construct, an immune effector cell or an antibody conjugate, and a pharmaceutically acceptable carrier.

药学上可接受的载剂可以包括但不限于：稀释剂、粘合剂和胶粘剂、润滑剂、崩解剂、防腐剂、媒介物、分散剂、助流剂、甜味剂、包衣、赋形剂、防腐剂、抗氧化剂(如抗坏血酸、盐酸半胱氨酸、硫酸氢钠、焦亚硫酸钠、亚硫酸钠、抗坏血酸棕榈酸酯、丁羟茴醚(BHA)、丁羟甲苯(BHT)、卵磷脂、没食子酸丙酯、α-生育酚、柠檬酸、乙二胺四乙酸(EDTA)、山梨糖醇、酒石酸、磷酸等)、增溶剂、胶凝剂、软化剂、溶剂(例如，水、酒精、乙酸和糖浆)、缓冲剂(例如，磷酸盐缓冲剂、组氨酸缓冲剂和乙酸盐缓冲剂)、表面活性剂(例如非离子表面活性剂，例如聚山梨酯80、聚山梨酯20、泊洛沙姆或聚乙二醇)、抗细菌剂、抗真菌剂、等渗剂(例如海藻糖、蔗糖、甘露醇、山梨醇、乳糖、葡萄糖)、吸收延迟剂、螯合剂和乳化剂。对于包含DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)、多特异性抗原结合构建体或者抗体缀合物的药物组合物而言，合适的载剂可以选自缓冲剂(例如柠檬酸盐缓冲液、乙酸盐缓冲液、磷酸盐缓冲液、组氨酸缓冲液、组氨酸盐缓冲液)、等渗剂(例如海藻糖、蔗糖、甘露醇、山梨醇、乳糖、葡萄糖)、非离子表面活性剂(例如聚山梨酯80、聚山梨酯20、泊洛沙姆)或其组合。Pharmaceutically acceptable carriers may include, but are not limited to, diluents, binders and adhesives, lubricants, disintegrants, preservatives, vehicles, dispersants, glidants, sweeteners, coatings, excipients, preservatives, antioxidants (such as ascorbic acid, cysteine hydrochloride, sodium bisulfate, sodium metabisulfite, sodium sulfite, ascorbyl palmitate, butylated hydroxyanisole (BHA), butylated hydroxytoluene (BHT), lecithin, propyl gallate, α-tocopherol, citric acid, ethylenediaminetetraacetic acid (EDTA), sorbitan, [0014] Examples of the present invention include, but are not limited to, sucrose alcohols, tartaric acid, phosphoric acid, etc.), solubilizers, gelling agents, emollients, solvents (e.g., water, alcohol, acetic acid, and syrup), buffers (e.g., phosphate buffers, histidine buffers, and acetate buffers), surfactants (e.g., non-ionic surfactants such as polysorbate 80, polysorbate 20, poloxamer, or polyethylene glycol), antibacterial agents, antifungal agents, isotonic agents (e.g., trehalose, sucrose, mannitol, sorbitol, lactose, glucose), absorption delaying agents, chelating agents, and emulsifiers. For pharmaceutical compositions comprising a DLL3 binding molecule (e.g., an anti-DLL3 antibody or antigen-binding fragment thereof of the present invention), a multispecific antigen-binding construct, or an antibody conjugate, suitable carriers can be selected from a buffer (e.g., citrate buffer, acetate buffer, phosphate buffer, histidine buffer, histidine salt buffer), an isotonic agent (e.g., trehalose, sucrose, mannitol, sorbitol, lactose, glucose), a non-ionic surfactant (e.g., polysorbate 80, polysorbate 20, poloxamer), or a combination thereof.

本发明提供的药物组合物可以为多种剂型，包括但不限于固体、半固体、液体、粉末或冻干形式。对于包含DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)或者多特异性抗原结合构建体的药物组合物而言，优选的剂型通常可以为例如注射液和冻干粉。The pharmaceutical compositions provided herein may be in a variety of dosage forms, including but not limited to solid, semisolid, liquid, powder, or lyophilized forms. For pharmaceutical compositions comprising DLL3-binding molecules (e.g., anti-DLL3 antibodies or antigen-binding fragments thereof of the present invention) or multispecific antigen-binding constructs, preferred dosage forms include, for example, injection solutions and lyophilized powders.

可通过本领域已知的任何方法，例如通过全身或局部施用，将本文提供的药物组合物给药于受试者。给药途径包括但不限于肠胃外(例如，静脉内、腹膜内、皮内、肌肉内、皮下或腔内)、局部(例如瘤内)、硬膜外或粘膜(例如鼻内、口服、阴道、直肠、舌下或局部)。优选地，所述药物组合物适合于静脉内、肌内、皮下、肠胃外、脊柱或表皮给药(如通过注射或输注)。给药方法可以为例如注射或输注。The pharmaceutical compositions provided herein can be administered to a subject by any method known in the art, for example, by systemic or topical administration. Routes of administration include, but are not limited to, parenteral (e.g., intravenous, intraperitoneal, intradermal, intramuscular, subcutaneous, or intracavitary), topical (e.g., intratumoral), epidural, or mucosal (e.g., intranasal, oral, vaginal, rectal, sublingual, or topical). Preferably, the pharmaceutical composition is suitable for intravenous, intramuscular, subcutaneous, parenteral, spinal, or epidermal administration (e.g., by injection or infusion). Methods of administration can be, for example, injection or infusion.

本领域技术人员应当理解，确切的给药剂量将取决于各种因素，例如药物组合物的代谢动力学性质、治疗的持续时间、特定化合物的排泄速率、治疗目的、给药途径和受试者的状况，例如患者的年龄、健康状况、体重、性别、饮食、病史，以及医学领域公知的其他因素。It will be understood by those skilled in the art that the exact dosage administered will depend on various factors, such as the metabolic kinetic properties of the pharmaceutical composition, the duration of treatment, the excretion rate of the specific compound, the purpose of the treatment, the route of administration, and the condition of the subject, such as the patient's age, health status, weight, sex, diet, medical history, and other factors well known in the medical field.

作为一般性指导，本发明的DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)、多特异性抗原结合构建体或者抗体缀合物的给药剂量范围可以为约0.0001至100mg/kg，更通常为0.01至20mg/kg受试者体重。例如，给药剂量可以是0.3mg/kg体重、1mg/kg体重、3mg/kg体重、5mg/kg体重，10mg/kg体重或20mg/kg体重，或在1-20mg/kg范围内。示例性的治疗方案需要每周给药一次、每两周一次、每三周一次、每四周一次、每月一次、每3个月一次、每3-6个月一次、或起始给药间隔略短后期给药间隔加长。As a general guide, the DLL3-binding molecules of the invention (e.g., anti-DLL3 antibodies or antigen-binding fragments thereof), multispecific antigen-binding constructs, or antibody conjugates can be administered at a dosage ranging from about 0.0001 to 100 mg/kg, more typically 0.01 to 20 mg/kg of the subject's body weight. For example, the dosage can be 0.3 mg/kg, 1 mg/kg, 3 mg/kg, 5 mg/kg, 10 mg/kg, or 20 mg/kg, or within the range of 1-20 mg/kg. Exemplary treatment regimens entail dosing once a week, once every two weeks, once every three weeks, once every four weeks, once a month, once every three months, once every three to six months, or initially with a shorter dosing interval followed by a longer dosing interval.

治疗treat

在第七方面，本发明涉及本发明的DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)、多特异性抗原结合构建体、免疫效应细胞、抗体缀合物或者药物组合物在制备用于治疗受试者中疾病的药物中的用途。In a seventh aspect, the present invention relates to use of a DLL3 binding molecule (e.g., an anti-DLL3 antibody or antigen-binding fragment thereof), a multispecific antigen-binding construct, an immune effector cell, an antibody conjugate, or a pharmaceutical composition of the invention in the preparation of a medicament for treating a disease in a subject.

本发明还涉及本发明的DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)、多特异性抗原结合构建体、免疫效应细胞、抗体缀合物或者药物组合物，其用于治疗疾病。The present invention also relates to the DLL3 binding molecules of the present invention (eg, the anti-DLL3 antibodies or antigen-binding fragments thereof), multispecific antigen-binding constructs, immune effector cells, antibody conjugates or pharmaceutical compositions of the present invention for use in treating diseases.

本发明还提供一种治疗受试者中疾病的方法，所述方法包括向所述受试者给药治疗有效量的本发明的DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)、多特异性抗原结合构建体、免疫效应细胞、抗体缀合物或者药物组合物。The present invention also provides a method of treating a disease in a subject, comprising administering to the subject a therapeutically effective amount of a DLL3-binding molecule of the invention (e.g., an anti-DLL3 antibody or antigen-binding fragment thereof), a multispecific antigen-binding construct, an immune effector cell, an antibody conjugate, or a pharmaceutical composition.

本文所述“治疗有效剂量”指导致疾病症状的严重性降低，疾病无症状期的频率和持续时间增加，或者防止因疾病痛苦而引起的损伤或失能的剂量。例如，用于抗增殖作用，防止进一步肿瘤发展，减少肿瘤尺寸，减少肿瘤血管，减少癌细胞数量，抑制、延迟或减少癌症患者中肿瘤和/或恶性细胞生长和/或转移，和/或减少可以观察到与该疾病相关的一种或多种症状的治疗有效剂量。根据许多不同的因素，治疗有效剂量可以各不相同，所述因素包括给药的方式，靶位点，患者的生理状态，患者是人还是其他动物，给予的其他药物，以及治疗是预防性的还是治疗性的。在某些实施方案中，患者是人，但也可治疗非人哺乳动物，包括转基因动物。可使用本领域技术人员已知的常规方法对治疗剂量进行滴定以优化安全性和功效。As used herein, a "therapeutically effective dose" refers to a dose that results in a decrease in the severity of disease symptoms, an increase in the frequency and duration of asymptomatic periods of the disease, or prevents damage or disability caused by the suffering of the disease. For example, a therapeutically effective dose is used for anti-proliferative effects, preventing further tumor development, reducing tumor size, reducing tumor vascularity, reducing the number of cancer cells, inhibiting, delaying or reducing tumor and/or malignant cell growth and/or metastasis in cancer patients, and/or reducing one or more symptoms associated with the disease that can be observed. The therapeutically effective dose can vary depending on many different factors, including the mode of administration, the target site, the patient's physiological state, whether the patient is human or other animal, other drugs administered, and whether the treatment is prophylactic or therapeutic. In certain embodiments, the patient is human, but non-human mammals, including transgenic animals, can also be treated. The therapeutic dose can be titrated using conventional methods known to those skilled in the art to optimize safety and efficacy.

“治疗有效剂量”的本发明的DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)、多特异性抗原结合构建体、免疫效应细胞、抗体缀合物或者药物组合物优选地将细胞生长或肿瘤生长抑制至少约10％，优选至少约20％，更优选至少约30％，更优选至少约40％，更优选至少约50％，更优选至少约60％，更优选至少约70％，更优选至少约80％。抑制肿瘤生长的能力可以在预测对人肿瘤的疗效的动物模型系统中评价。或者，也可以通过检查抑制细胞生长的能力来评价，这种抑制可以通过本领域技术人员公知的实验在体外测定。有效量的本发明的DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)、多特异性抗原结合构建体、免疫效应细胞、抗体缀合物或者药物组合物能够减小肿瘤大小，或者以其他方式缓解对象的症状如预防和/或治疗转移或复发。本领域技术人员可以根据如下因素确定这种量，如受试者的大小、受试者症状的严重性和选择的特定组合物或给药途径。A "therapeutically effective dose" of a DLL3 binding molecule (e.g., an anti-DLL3 antibody or antigen-binding fragment thereof), multispecific antigen-binding construct, immune effector cell, antibody conjugate, or pharmaceutical composition of the invention preferably inhibits cell growth or tumor growth by at least about 10%, preferably at least about 20%, more preferably at least about 30%, more preferably at least about 40%, more preferably at least about 50%, more preferably at least about 60%, more preferably at least about 70%, and more preferably at least about 80%. The ability to inhibit tumor growth can be evaluated in animal model systems predictive of efficacy against human tumors. Alternatively, it can be evaluated by examining the ability to inhibit cell growth, which can be determined in vitro using assays known to those skilled in the art. An effective amount of a DLL3 binding molecule (e.g., an anti-DLL3 antibody or antigen-binding fragment thereof), multispecific antigen-binding construct, immune effector cell, antibody conjugate, or pharmaceutical composition of the invention is capable of reducing tumor size or otherwise alleviating symptoms in a subject, such as preventing and/or treating metastasis or recurrence. Such an amount can be determined by one skilled in the art based on factors such as the size of the subject, the severity of the subject's symptoms, and the specific composition or route of administration selected.

在一些实施方案中，所述疾病与DLL3异常表达相关。术语“异常表达”指与正常状态的样品(或标准样品，例如未患有与DLL3异常表达相关疾病的受试者的样品)相比，样品中蛋白表达水平过高或过低。优选地，所述疾病以DLL3高表达为特征。例如，患有或怀疑患有疾病(例如小细胞肺癌)的受试者的组织(例如肿瘤组织和癌旁组织)中高表达DLL3，而未患有所述疾病的对象的相应组织中低表达DLL3。In some embodiments, the disease is associated with abnormal DLL3 expression. The term "abnormal expression" refers to excessively high or low protein expression levels in a sample compared to a normal sample (or a standard sample, such as a sample from a subject not suffering from a disease associated with abnormal DLL3 expression). Preferably, the disease is characterized by high DLL3 expression. For example, DLL3 is highly expressed in tissues (e.g., tumor tissue and adjacent tissues) of subjects suffering from or suspected of suffering from a disease (e.g., small cell lung cancer), while DLL3 is low in the corresponding tissues of subjects not suffering from the disease.

在一实施方案中，如上所述的疾病为癌症。在一实施方案中，癌症为DLL3阳性的肿瘤。在一优选实施方案中，癌症选自小细胞肺癌(SCLC)；大细胞神经内分泌癌(LCNEC)；包括肾脏、泌尿生殖道(例如膀胱、前列腺、卵巢、宫颈和子宫内膜)、胃肠道(例如胃、结肠)、甲状腺(例如甲状腺髓样癌)、胰腺和肺的多种组织的神经内分泌肿瘤；神经胶质瘤或假性神经内分泌肿瘤(pNET)。In one embodiment, the disease as described above is cancer. In one embodiment, the cancer is a DLL3-positive tumor. In a preferred embodiment, the cancer is selected from small cell lung cancer (SCLC); large cell neuroendocrine carcinoma (LCNEC); neuroendocrine tumors of various tissues including the kidney, genitourinary tract (e.g., bladder, prostate, ovary, cervix, and endometrium), gastrointestinal tract (e.g., stomach, colon), thyroid (e.g., medullary thyroid carcinoma), pancreas, and lung; glioma or pseudoneuroendocrine tumor (pNET).

本发明的DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)、多特异性抗原结合构建体、免疫效应细胞、抗体缀合物或者药物组合物可以与至少一种或多种本文所述的治疗剂联合施用。联合施用的方式没有限制。例如，可以将上述治疗剂全部在一次施用或分开施用。The DLL3-binding molecules of the present invention (e.g., anti-DLL3 antibodies or antigen-binding fragments thereof), multispecific antigen-binding constructs, immune effector cells, antibody conjugates, or pharmaceutical compositions can be administered in combination with at least one or more therapeutic agents described herein. The manner of combined administration is not limited. For example, all of the above therapeutic agents can be administered at once or separately.

对于癌症治疗，本发明的DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)、多特异性抗原结合构建体、免疫效应细胞、抗体缀合物或者药物组合物可以与其他治疗方法联合使用，包括但不限于：手术、化学治疗、放射治疗、靶向治疗、免疫治疗、激素治疗、血管生成抑制和姑息治疗。For cancer treatment, the DLL3-binding molecules of the invention (e.g., anti-DLL3 antibodies or antigen-binding fragments thereof), multispecific antigen-binding constructs, immune effector cells, antibody conjugates, or pharmaceutical compositions of the invention can be used in combination with other treatment methods, including but not limited to surgery, chemotherapy, radiotherapy, targeted therapy, immunotherapy, hormone therapy, angiogenesis inhibition, and palliative care.

在某些实施方案中，本发明的DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)、多特异性抗原结合构建体、免疫效应细胞、抗体缀合物或者药物组合物进一步与选自以下的一种或多种治疗剂联合使用：化疗剂、放射性同位素、免疫检查点抑制剂和肿瘤抗原靶向药物。化疗剂可以包括例如：抗代谢药、烷化剂、细胞毒性剂、拓扑异构酶抑制剂、微管抑制剂。肿瘤抗原靶向药物包括但不限于靶向肿瘤相关抗原和肿瘤特异性抗原的药物。治疗剂的其他非限制性实例可以包括例如：血管生成抑制因子、脱乙酰化酶(HDAC)抑制因子、Hedgehog信号通路阻滞剂、mTOR抑制剂、p53/mdm2抑制剂、PARP抑制剂、蛋白酶体抑制剂(例如bortezomib、carfilzomib、ixazomib、Marizomib、Oprozomib)以及酪氨酸激酶抑制剂(例如BTK抑制剂)。In certain embodiments, the DLL3 binding molecules of the present invention (e.g., anti-DLL3 antibodies or antigen-binding fragments thereof), multispecific antigen-binding constructs, immune effector cells, antibody conjugates, or pharmaceutical compositions are further used in combination with one or more therapeutic agents selected from the group consisting of chemotherapeutic agents, radioisotopes, immune checkpoint inhibitors, and tumor antigen-targeted drugs. Chemotherapeutic agents may include, for example, antimetabolites, alkylating agents, cytotoxic agents, topoisomerase inhibitors, and microtubule inhibitors. Tumor antigen-targeted drugs include, but are not limited to, drugs that target tumor-associated antigens and tumor-specific antigens. Other non-limiting examples of therapeutic agents may include, for example, angiogenesis inhibitors, heparin deacetylase (HDAC) inhibitors, hedgehog signaling pathway blockers, mTOR inhibitors, p53/mdm2 inhibitors, PARP inhibitors, proteasome inhibitors (e.g., bortezomib, carfilzomib, ixazomib, marizomib, oprozomib), and tyrosine kinase inhibitors (e.g., BTK inhibitors).

在一些实施方案中，本发明的DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)、多特异性抗原结合构建体、免疫效应细胞、抗体缀合物或者药物组合物与化疗剂联合使用。在一些实施方案中，本发明的DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)、多特异性抗原结合构建体、免疫效应细胞、抗体缀合物或者药物组合物与免疫检查点抑制剂联合使用。在一些实施方案中，本发明的DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)、多特异性抗原结合构建体、免疫效应细胞、抗体缀合物或者药物组合物与放射性同位素联合使用。在一些实施方案中，本发明的DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)、多特异性抗原结合构建体、免疫效应细胞、抗体缀合物或者药物组合物与肿瘤抗原靶向药物联合使用。In some embodiments, the DLL3 binding molecules (e.g., anti-DLL3 antibodies or antigen-binding fragments thereof), multispecific antigen-binding constructs, immune effector cells, antibody conjugates, or pharmaceutical compositions of the present invention are used in combination with a chemotherapeutic agent. In some embodiments, the DLL3 binding molecules (e.g., anti-DLL3 antibodies or antigen-binding fragments thereof), multispecific antigen-binding constructs, immune effector cells, antibody conjugates, or pharmaceutical compositions of the present invention are used in combination with an immune checkpoint inhibitor. In some embodiments, the DLL3 binding molecules (e.g., anti-DLL3 antibodies or antigen-binding fragments thereof), multispecific antigen-binding constructs, immune effector cells, antibody conjugates, or pharmaceutical compositions of the present invention are used in combination with a radioisotope. In some embodiments, the DLL3 binding molecules (e.g., anti-DLL3 antibodies or antigen-binding fragments thereof), multispecific antigen-binding constructs, immune effector cells, antibody conjugates, or pharmaceutical compositions of the present invention are used in combination with a tumor antigen-targeted drug.

多核苷酸、载体和宿主细胞Polynucleotides, vectors and host cells

在第八方面，本发明提供了一种多核苷酸，其编码本发明的DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)或者多特异性抗原结合构建体。In an eighth aspect, the present invention provides a polynucleotide encoding a DLL3 binding molecule of the present invention (eg, an anti-DLL3 antibody or antigen-binding fragment thereof of the present invention) or a multispecific antigen-binding construct.

可以利用本领域已知的方法获得本发明的多核苷酸。例如，本发明的多核苷酸可以分离自噬菌体展示文库、酵母展示文库、免疫动物、永生化的细胞(例如，小鼠B细胞杂交瘤细胞、EBV介导的永生化B细胞)或者化学合成。可以从免疫动物中分离或者采用化学合成的方法制备编码结合分子(例如抗体或其抗原结合片段)或者多特异性抗原结合构建体的多核苷酸，然后利用多核苷酸构建表达载体。本发明的多核苷酸可以针对用于表达的宿主细胞进行密码子优化。The polynucleotides of the present invention can be obtained using methods known in the art. For example, the polynucleotides of the present invention can be isolated from phage display libraries, yeast display libraries, immune animals, immortalized cells (e.g., mouse B cell hybridoma cells, EBV-mediated immortalized B cells) or chemically synthesized. Polynucleotides encoding binding molecules (e.g., antibodies or their antigen-binding fragments) or multispecific antigen-binding constructs can be prepared by separation from immune animals or chemical synthesis, and then expression vectors can be constructed using the polynucleotides. The polynucleotides of the present invention can be codon-optimized for the host cells used for expression.

在第九方面，本发明还提供了包含本发明的多核苷酸的表达载体。表达载体可以进一步包含额外的多核苷酸序列，例如调控序列和抗生素抗性基因。本发明的多核苷酸可以存在于一种或多种表达载体中。在一实施方案中，本发明的多核苷酸制备为重组核酸。可使用本领域众所周知的技术制备重组核酸，例如化学合成、DNA重组技术(例如聚合酶链式反应(PCR)技术)等。In a ninth aspect, the present invention also provides an expression vector comprising a polynucleotide of the present invention. The expression vector may further comprise additional polynucleotide sequences, such as regulatory sequences and antibiotic resistance genes. The polynucleotide of the present invention may be present in one or more expression vectors. In one embodiment, the polynucleotide of the present invention is prepared as a recombinant nucleic acid. Recombinant nucleic acids can be prepared using techniques well known in the art, such as chemical synthesis, DNA recombination techniques (e.g., polymerase chain reaction (PCR) technology), etc.

在第十方面，本发明还提供了一种宿主细胞，其包含本发明的多核苷酸或表达载体。可以采用本领域已知的各种方法将本发明的多核苷酸或表达载体导入合适的宿主细胞中。这类方法包括但不限于脂质体转染、电穿孔、病毒转导和磷酸钙转染等。In a tenth aspect, the present invention further provides a host cell comprising a polynucleotide or expression vector of the present invention. The polynucleotide or expression vector of the present invention can be introduced into a suitable host cell using various methods known in the art. Such methods include, but are not limited to, liposome transfection, electroporation, viral transduction, and calcium phosphate transfection.

在优选的实施方案中，宿主细胞用于表达本发明的DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)或者多特异性抗原结合构建体。宿主细胞的实例包括但不限于原核细胞(例如细菌，例如大肠杆菌)和真核细胞(例如酵母、昆虫细胞、哺乳动物细胞)。适合于抗体表达的哺乳动物宿主细胞包括但不限于人宫颈癌细胞(HeLa细胞)、人类胚胎肾细胞(HEK细胞，例如HEK 293细胞)、中国仓鼠卵巢(CHO)细胞和其他适于表达抗体的哺乳动物细胞。In preferred embodiments, host cells are used to express the DLL3-binding molecules of the invention (e.g., anti-DLL3 antibodies or antigen-binding fragments thereof) or multispecific antigen-binding constructs of the invention. Examples of host cells include, but are not limited to, prokaryotic cells (e.g., bacteria, such as E. coli) and eukaryotic cells (e.g., yeast, insect cells, mammalian cells). Mammalian host cells suitable for antibody expression include, but are not limited to, human cervical carcinoma cells (HeLa cells), human embryonic kidney cells (HEK cells, such as HEK 293 cells), Chinese hamster ovary (CHO) cells, and other mammalian cells suitable for antibody expression.

在第十一方面，本发明还提供了一种产生本发明的DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)或者多特异性抗原结合构建体的方法，所述方法包括：In an eleventh aspect, the present invention further provides a method for producing a DLL3-binding molecule of the present invention (e.g., an anti-DLL3 antibody or antigen-binding fragment thereof of the present invention) or a multispecific antigen-binding construct of the present invention, the method comprising:

a)在合适条件下培养本发明的宿主细胞以表达本发明的DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)或者多特异性抗原结合构建体；以及a) culturing the host cells of the invention under suitable conditions to express the DLL3-binding molecules of the invention (e.g., anti-DLL3 antibodies or antigen-binding fragments thereof) or multispecific antigen-binding constructs of the invention; and

b)从宿主细胞或其培养物分离所述DLL3结合分子(例如抗DLL3抗体或其抗原结合片段)或者多特异性抗原结合构建体。b) isolating the DLL3 binding molecule (eg, an anti-DLL3 antibody or antigen-binding fragment thereof) or multispecific antigen-binding construct from the host cell or culture thereof.

试剂盒Reagent test kit

本发明还提供试剂盒，其包含本发明的DLL3结合分子(例如本发明的抗DLL3抗体或其抗原结合片段)、多特异性抗原结合构建体、免疫效应细胞、抗体缀合物或者药物组合物，以及使用说明。试剂盒还可以包含合适的容器。在某些实施方案中，试剂盒还包含给药的装置。试剂盒一般包括表明试剂盒内容物的预期用途和/或使用方法的标签。术语“标签”包括在试剂盒上或与试剂盒一起提供的或以其他方式随试剂盒提供的任何书面的或记录的材料。The present invention also provides kits comprising a DLL3-binding molecule of the invention (e.g., an anti-DLL3 antibody or antigen-binding fragment thereof), a multispecific antigen-binding construct, an immune effector cell, an antibody conjugate, or a pharmaceutical composition, and instructions for use. The kit may also comprise a suitable container. In certain embodiments, the kit further comprises a device for administration. Kits generally include a label indicating the intended use and/or instructions for use of the kit contents. The term "label" includes any written or recorded material provided on or with the kit, or otherwise accompanying the kit.

有益效果Beneficial effects

本发明的DLL3结合分子可以呈现出优异的效果，例如但不限于：1)本发明的DLL3抗体是一种全新的仅含“重链”的全人抗体，具有与人DLL3和食蟹猴DLL3结合的活性。这种重链抗体的大小只有传统IgG抗体的一半，由于不含轻链的这一特点，使得该抗体可以有利地用于双特异性抗体，并解决了轻链错配和异源二聚化的问题。(2)能够结合源自人和猴的DLL3蛋白或小鼠的DLL3蛋白。(3)能够内化至DLL3阳性的细胞内。(4)结合表位与参照抗体PR012621和PR012622相同或不同。The DLL3-binding molecules of the present invention can exhibit excellent effects, such as, but not limited to: 1) The DLL3 antibody of the present invention is a novel fully human antibody containing only a "heavy chain" and has binding activity to both human DLL3 and cynomolgus monkey DLL3. This heavy chain antibody is only half the size of a traditional IgG antibody. Due to the fact that it does not contain a light chain, it can be advantageously used in bispecific antibodies and solves the problems of light chain mispairing and heterodimerization. (2) It can bind to DLL3 proteins derived from humans and monkeys or DLL3 proteins from mice. (3) It can be internalized into DLL3-positive cells. (4) The binding epitope is the same as or different from that of the reference antibodies PR012621 and PR012622.

实施例Example

通过参考在此给出的一些具体实施例可获得对本发明的进一步的理解，这些实施例仅用于说明本发明，其无意于对本发明的范围做出任何限制。显然，可以对本发明作出多种改动和变化而不脱离本发明的实质，因此，这些改动和变化同样在本申请要求保护的范围内。本文中所用的比例包括百分比，如果没有特别指出，都按重量计。A further understanding of the present invention may be obtained by reference to some of the specific examples provided herein, which are intended to illustrate the present invention only and are not intended to limit the scope of the present invention in any way. Obviously, various modifications and variations may be made to the present invention without departing from the spirit of the present invention, and therefore, such modifications and variations are also within the scope of the present application. Ratios used herein include percentages, and unless otherwise specified, are by weight.

实施例1.全人源抗DLL3 HCAb抗体的获得Example 1. Obtaining a fully human anti-DLL3 HCAb antibody

可以利用DLL3抗原对实验动物进行免疫以获得针对DLL3特异性结合的抗体分子，该实验动物可以是小鼠、大鼠、兔、羊、骆驼等。通常，其得到的抗体分子是非人源的。在获得非人源抗体后，需要对这些分子利用抗体工程技术进行人源化改造，以降低免疫原性并提高成药性。然而，抗体的人源化过程有其技术复杂性，经过人源化改造的分子往往会降低对抗原的亲和力。另一方面，转基因技术的进步使得可以培育出基因工程化小鼠，其携带人免疫球蛋白免疫库并使其内源的鼠的免疫库缺失。这种转基因小鼠产生的抗体具有全人源的序列，因而无需再进一步做人源化改造，大大提高了治疗性抗体开发的效率。Harbour HCAb小鼠(Harbour Antibodies BV，WO2002/085945A3)是一种携带人免疫球蛋白免疫库的转基因小鼠，能够产生全新的仅“重链”抗体，该抗体的大小只有传统IgG抗体的一半。其产生的抗体仅具有人的抗体“重链”可变结构域和小鼠Fc恒定结构域。由于不含轻链的这一特点，该抗体几乎解决了轻链错配和异源二聚化的问题，使得这一技术平台能够开发出传统抗体平台难以实现的产品。Antibodies specifically binding to DLL3 can be obtained by immunizing experimental animals (e.g., mice, rats, rabbits, sheep, camels, etc.) with the DLL3 antigen. Typically, the resulting antibodies are non-human. After obtaining non-human antibodies, these molecules need to be humanized using antibody engineering techniques to reduce immunogenicity and improve drugability. However, the humanization process is technically complex, and humanized molecules often have reduced affinity for the antigen. On the other hand, advances in transgenic technology have enabled the creation of genetically engineered mice that carry a human immunoglobulin repertoire and lack the endogenous murine immune repertoire. The antibodies produced by these transgenic mice possess fully human sequences, eliminating the need for further humanization and significantly improving the efficiency of therapeutic antibody development. Harbour HCAb mice (Harbour Antibodies BV, WO2002/085945A3) are transgenic mice carrying a human immunoglobulin repertoire, capable of producing novel heavy-chain-only antibodies that are half the size of traditional IgG antibodies. The antibodies it produces only have human antibody "heavy chain" variable domains and mouse Fc constant domains. Due to the absence of light chains, the antibodies almost solve the problems of light chain mispairing and heterodimerization, making this technology platform capable of developing products that are difficult to achieve with traditional antibody platforms.

实施例1.1小鼠免疫Example 1.1 Immunization of mice

采用重组的人DLL3-his标签的蛋白(Acro，#DL3-H52H4)对6-8周龄的Harbour人源抗体转基因小鼠Harbour HCAb V2.1小鼠进行多轮免疫。在第一次免疫中，每只小鼠分别在腹腔和腋下淋巴结及腹股沟淋巴结处注射50μg免疫原蛋白和0.2mL完全弗氏佐剂(CFA，Sigma，#F5881)。为增强免疫应答，在第一次免疫接种两周后，将25μg免疫原蛋白连同200μL Ribi(Sigma佐剂系统，Sigma，#S6322)，注射到每只小鼠的腹腔内和皮下淋巴结处，随后每只老鼠每隔2周注射一次25μg免疫原和200μLRibi佐剂，加上第一次免疫共计5次。免疫时间为第0、14、28、42、56天；并且在21、35、49、61天，检测小鼠血清抗体滴度。在进行HCAb小鼠脾B细胞分离前5天，以每只小鼠25μg抗原蛋白的剂量进行最后一次增强免疫。Multiple rounds of immunization were performed on 6-8 week old Harbour HCAb V2.1 mice, transgenic mice expressing human DLL3-his tags, using recombinant human DLL3-his tagged protein (Acro, #DL3-H52H4). In the first immunization, each mouse was injected with 50 μg of immunogen protein and 0.2 mL of complete Freund's adjuvant (CFA, Sigma, #F5881) into the abdominal cavity, axillary lymph nodes, and inguinal lymph nodes, respectively. To enhance the immune response, two weeks after the first immunization, 25 μg of immunogen protein was injected with 200 μL of Ribi (Sigma Adjuvant System, Sigma, #S6322) into the abdominal cavity and subcutaneous lymph nodes of each mouse. Subsequently, each mouse was injected with 25 μg of immunogen and 200 μL of Ribi adjuvant every 2 weeks, for a total of 5 times, including the first immunization. Immunizations were performed on days 0, 14, 28, 42, and 56, and serum antibody titers were measured on days 21, 35, 49, and 61. Five days before HCAb mouse spleen B cell isolation, a final booster immunization was performed at a dose of 25 μg antigen protein per mouse.

实施例1.2血清滴度检测Example 1.2 Serum titer detection

在特定的时间点，收取小鼠血清，对血清进行10倍稀释取6个浓度(1:100、1:1000、1:10000、1:100000、1:1000000)，在包被有huDLL3-his蛋白(Acro，#DL3-H52H4)或cynoDLL3-his蛋白(Acro，#DL3-C52H9)或与DLL3不相关的蛋白的ELISA板进行ELISA检测来确定小鼠血液中抗DLL3的滴度，并经流式细胞术检测2个浓度的小鼠血清(1:100、1:1000)对DLL3高表达的CHO-K1/humanDLL3细胞和CHO-K1/cyno DLL3细胞和CHO-K1母细胞的特异反应性。空白对照组(PB)为免疫前老鼠的血清。At specific time points, mouse serum was collected and diluted 10-fold to obtain six concentrations (1:100, 1:1000, 1:10,000, 1:100,000, and 1:1,000,000). ELISA titers were determined using ELISA plates coated with huDLL3-his protein (Acro, #DL3-H52H4), cynoDLL3-his protein (Acro, #DL3-C52H9), or proteins unrelated to DLL3. Flow cytometry was used to analyze the specific reactivity of two concentrations of mouse serum (1:100 and 1:1000) against CHO-K1/humanDLL3 cells, CHO-K1/cynoDLL3 cells, and CHO-K1 blast cells expressing high DLL3 expression. A blank control (PB) consisted of serum from pre-immunized mice.

在ELISA方法中，分别用1μg/mL的huDLL3-his蛋白(Acro，#DL3-H52H4)或1μg/mL的cynoDLL3-his蛋白(Acro，#DL3-C52H9)或1μg/mL的不相关的his蛋白100μL/孔包被ELISA板(corning，#9018)，4℃孵育过夜；漂洗2次后，用含1％BSA的PBST在37℃阻断1小时；加入100μL/孔梯度稀释的血清37℃孵育1小时；漂洗3次后，加入100μL/孔1:5000稀释的anti-mouse-HRP(Bethyl，#A90-231P)，37℃孵育30分钟。漂洗3次后，加入100μL/孔TMB底物孵育约10分钟，加入50μL/孔1N HCl终止显色，读450nm的吸光度(Molecular Devices,Plus 384)。In the ELISA method, 100 μL/well of 1 μg/mL huDLL3-his protein (Acro, #DL3-H52H4) or 1 μg/mL cynoDLL3-his protein (Acro, #DL3-C52H9) or 1 μg/mL of an irrelevant his protein was coated on an ELISA plate (corning, #9018) and incubated overnight at 4°C; after rinsing twice, the plate was blocked with PBST containing 1% BSA at 37°C for 1 hour; 100 μL/well of serially diluted serum was added and incubated at 37°C for 1 hour; after rinsing three times, 100 μL/well of 1:5000 diluted anti-mouse-HRP (Bethyl, #A90-231P) was added and incubated at 37°C for 30 minutes. After rinsing three times, add 100 μL/well TMB substrate and incubate for about 10 minutes. Add 50 μL/well 1N HCl to stop color development and read the absorbance at 450 nm (Molecular Devices, Plus 384).

在FACS方法中，梯度稀释的鼠血清分别与CHO-K1/humanDLL3或CHO-K1/cynoDLL3或CHO-K1细胞4℃孵育1小时；细胞洗2次后，加入二抗Alexa FluorTM488goat anti-mouse IgG Fcγspecific(Jackson，#115-545-071)4℃孵育1小时，洗2次后，重悬细胞用流式细胞仪检测(BD CantoII)。CHO-K1细胞作为本底对照。For FACS analysis, CHO-K1/human DLL3, CHO-K1/cyno DLL3, or CHO-K1 cells were incubated with serially diluted mouse serum for 1 hour at 4°C. After washing twice, the cells were incubated with a secondary antibody, Alexa Fluor™ 488 goat anti-mouse IgG Fcγ specific (Jackson, #115-545-071), for 1 hour at 4°C. After washing twice, the cells were resuspended and analyzed by flow cytometry (BD Canto II). CHO-K1 cells served as a background control.

实施例1.3获得抗DLL3的HCAb抗体序列Example 1.3 Obtaining anti-DLL3 HCAb antibody sequences

当检测小鼠血清中DLL3特异的抗体滴度达到一定的水平后，将小鼠的脾细胞取出分离B细胞，用BD FACS AriaIICell Sorter分选CD138阳性的浆细胞和人DLL3抗原阳性的B细胞群。提取B细胞的RNA，反转录cDNA(SuperScript IV First-Strand synthesis system，Invitrogen，18091200)，然后用特异性的引物PCR扩增人VH基因。PCR引物5’-GGTGTCCAGTGTSAGGTGCAGCTG-3’(SEQ ID NO:282)，5’-AATCCCTGGGCACT GAAGAGACGGTGACC-3’(SEQ ID NO:283)。将扩增的VH基因片段构建到编码人IgG1抗体重链Fc结构域序列的哺乳动物细胞表达质粒pCAG载体中。When the titer of DLL3-specific antibodies in mouse serum reached a certain level, splenocytes were removed from the mice to isolate B cells. CD138-positive plasma cells and human DLL3 antigen-positive B cell populations were sorted using a BD FACS AriaII Cell Sorter. RNA was extracted from B cells and reverse-transcribed into cDNA (SuperScript IV First-Strand Synthesis System, Invitrogen, 18091200). The human VH gene was then amplified using PCR using specific primers: 5'-GGTGTCCAGTGTSAGGTGCAGCTG-3' (SEQ ID NO: 282) and 5'-AATCCCTGGGCACTGAAGAGACGGTGACC-3' (SEQ ID NO: 283). The amplified VH gene fragment was constructed into the mammalian cell expression plasmid pCAG vector encoding the Fc domain sequence of the human IgG1 antibody heavy chain.

构建好的质粒转染哺乳动物宿主细胞(如人胚肾细胞HEK293)进行表达获得HCAb的抗体。用流式细胞仪检测表达HCAb的上清与过表达人DLL3的CHO-K1/hu DLL3稳定细胞系或过表达食蟹猴DLL3的CHO-K1/cyno DLL3稳定细胞系的结合，同时用ELISA的方法检测表达HCAb的上清与huDLL3-his蛋白或cynoDLL3-his蛋白的结合。获得的阳性单克隆抗体利用常规的测序手段获得编码抗体分子可变结构域的核苷酸序列以及对应的氨基酸序列。去除重复序列后得到168个同时结合CHO-K1/hu DLL3和CHO-K1/cyno DLL3的具有独特序列的全人源DLL3单克隆抗体。根据人猴细胞结合能力的试验结果，选择排名靠前的44个抗体进行重组表达。44个抗体的胚系基因分析和翻译后修饰位点(PTM)分析如表1所示。The constructed plasmids were transfected into mammalian host cells (such as human embryonic kidney HEK293 cells) for expression to obtain HCAb antibodies. Flow cytometry was used to analyze the binding of HCAb-expressing supernatants to CHO-K1/hu DLL3 stable cell lines overexpressing human DLL3 or CHO-K1/cyno DLL3 stable cell lines overexpressing cynomolgus macaque DLL3. ELISA was also used to analyze the binding of HCAb-expressing supernatants to huDLL3-his or cynoDLL3-his proteins. Conventional sequencing was used to determine the nucleotide sequence encoding the variable domain of the antibody molecule and the corresponding amino acid sequence. After removing repetitive sequences, 168 fully human DLL3 monoclonal antibodies with unique sequences that bound to both CHO-K1/hu DLL3 and CHO-K1/cyno DLL3 were obtained. Based on the results of human and monkey cell binding, the top 44 antibodies were selected for recombinant expression. Germline gene analysis and post-translational modification (PTM) analysis of the 44 antibodies are shown in Table 1.

表1.HCAb抗体序列的胚系基因分析和翻译后修饰位点(PTM)分析

Table 1. Germline gene analysis and post-translational modification (PTM) analysis of HCAb antibody sequences

实施例1.4抗DLL3全人源重组抗体的表达和纯化Example 1.4 Expression and purification of fully human recombinant anti-DLL3 antibodies

将编码HCAb抗体的质粒转染哺乳动物宿主细胞(如人胚肾细胞HEK293)，利用常规的重组蛋白表达和纯化技术，可以得到纯化的抗DLL3重组重链抗体。具体地，质粒用大抽试剂盒(Macherey-Nagel，Xtra Midi)抽提质粒，以提高质粒的纯度和质量，质粒通过0.22μm过滤器(millpore)过滤。纯化后的质粒用于转染。将HEK293细胞在FreeStyleTMF17 Expression Medium培养基(Thermo，A1383504)扩培。瞬时转染开始之前，调节细胞浓度至6×105细胞/ml，于37℃8％CO2摇床中培养24小时，细胞浓度在1.2×106细胞/mL。准备30mL培养的细胞，将上述编码HCAb重链的质粒30μg质粒溶解于1.5ml Opti-MEM减血清培养基(Thermo，#31985088)，再取1.5ml Opti-MEM溶入1mg/mL PEI(Polysciences,Inc，#23966-2)120μl，静置5分钟。把PEI缓慢加入质粒中，室温孵育10分钟，边摇晃培养瓶边缓慢滴入质粒PEI混合溶液，于37℃8％CO2摇床中培养5天。5天后观测细胞活率。收集培养物，以3300G转速离心10分钟后取上清；然后将上清高速离心去除杂质。用PBS(pH7.4)平衡含有MabSelectTM(GE Healthcare Life Science，#71-5020-91AE)的重力柱(Bio-Rad，#7311550)，2-5倍柱体积冲洗。将上清样品过柱。用5-10倍柱体积的PBS冲洗柱子。再用pH3.5的0.1M甘氨酸洗脱目的蛋白，后用pH 8.0的Tris-HCl调节至中性，最后用超滤管(Millipore，UFC901024)浓缩换液至PBS缓冲液，得到纯化的抗DLL3重链抗体溶液。抗体浓度用NanoDrop检测280nm吸光度测定，抗体的纯度用SEC-HPLC和SDS-PAGE测定。经上述实验，共得到44种具有独特序列的DLL3单克隆抗体，均为IgG1亚型。The plasmid encoding the HCAb antibody is transfected into mammalian host cells (such as human embryonic kidney cells HEK293), and the purified anti-DLL3 recombinant heavy chain antibody can be obtained using conventional recombinant protein expression and purification techniques. Specifically, the plasmid was purified using a large extraction kit (Macherey-Nagel, Plasmids were extracted using Xtra Midi (Extra Midi) to improve their purity and quality, and then filtered through a 0.22 μm filter (millpore). The purified plasmids were used for transfection. HEK293 cells were cultured in FreeStyle ™ F17 Expression Medium (Thermo, A1383504). Before transient transfection, the cell concentration was adjusted to 6 × 10 5 cells/mL and cultured in a shaker at 37°C with 8% CO 2 for 24 hours, with a cell concentration of 1.2 × 10 6 cells/mL. Prepare 30 mL of cultured cells and dissolve 30 μg of the plasmid encoding the HCAb heavy chain in 1.5 mL of Opti-MEM Reduced Serum Medium (Thermo, #31985088). Then, dissolve 120 μl of 1 mg/mL PEI (Polysciences, Inc, #23966-2) in 1.5 mL of Opti-MEM and let stand for 5 minutes. Slowly add PEI to the plasmid and incubate at room temperature for 10 minutes. The plasmid-PEI mixture is then slowly added dropwise while shaking the culture flask. Culture the cells at 37°C in a shaker with 8% CO2 for 5 days. After 5 days, observe cell viability. Harvest the culture and centrifuge at 3300g for 10 minutes. The supernatant is then removed by high-speed centrifugation to remove impurities. A gravity column (Bio-Rad, #7311550) containing MabSelect ™ (GE Healthcare Life Science, #71-5020-91AE) is equilibrated with PBS (pH 7.4) and rinsed with 2-5 column volumes. The supernatant is passed through the column. The column is rinsed with 5-10 column volumes of PBS. The target protein is then eluted with 0.1 M glycine (pH 3.5), adjusted to neutrality with Tris-HCl (pH 8.0), and concentrated using an ultrafiltration tube (Millipore, UFC901024) to PBS buffer to obtain a purified anti-DLL3 heavy chain antibody solution. The antibody concentration was determined by NanoDrop assay at 280 nm absorbance, and the antibody purity was determined by SEC-HPLC and SDS-PAGE. Through the above experiments, a total of 44 DLL3 monoclonal antibodies with unique sequences were obtained, all of which were IgG1 subtype.

此外，准备了两种对比例抗体，用于后续实施例，一种是对比例1抗体PR0012621(自制)，另一种是对比例2抗体PR0012622(自制)。具体而言，对比例1抗体Tab(PR012621)是在ABBVie公司的Rovalpituzumab tesirine(Rova-T)基础上制备的对比例抗体，其重链和轻链的氨基酸序列分别如SEQ NO ID:220和SEQ NO ID:266所示。In addition, two comparative antibodies were prepared for use in subsequent examples, one being the comparative example 1 antibody PR0012621 (prepared in-house), and the other being the comparative example 2 antibody PR0012622 (prepared in-house). Specifically, the comparative example 1 antibody Tab (PR012621) was prepared based on Rovalpituzumab tesirine (Rova-T) from ABBVie, and the amino acid sequences of its heavy chain and light chain are shown in SEQ NO ID: 220 and SEQ NO ID: 266, respectively.

对比例2抗体Tab(PR012622)是在Amgen公司的AMG757基础上制备的对比例抗体，其重链和轻链的氨基酸序列分别如SEQ NO ID:221和SEQ NO ID:267所示。Comparative Example 2 Antibody Tab (PR012622) is a comparative antibody prepared based on Amgen's AMG757. The amino acid sequences of its heavy chain and light chain are shown in SEQ NO ID: 221 and SEQ NO ID: 267, respectively.

实施例1.5利用SEC-HPLC分析蛋白纯度和聚体Example 1.5 Analysis of protein purity and aggregates using SEC-HPLC

本实施例使用分析型分子尺寸排阻层析色谱法(SEC)来分析蛋白样品的纯度和聚体形式。将分析型色谱柱TSKgel G3000SWxl(Tosoh Bioscience，#08541，5μm，7.8mm×30cm)连接到高压液相色谱仪HPLC(Agilent Technologies，Agilent 1260Infinity II)，用PBS缓冲液室温下平衡至少1小时。适量蛋白样品(至少10μg)用0.22μm滤膜过滤后注射入系统，并设定HPLC程序：用PBS缓冲液将样品以1.0mL/分钟的流速流过色谱柱，最长时间为25分钟。HPLC将生成分析报告，报告出样品内不同分子尺寸组份的滞留时间。In this example, analytical size exclusion chromatography (SEC) was used to analyze the purity and aggregate form of the protein sample. An analytical column, TSKgel G3000SWxl (Tosoh Bioscience, #08541, 5 μm, 7.8 mm × 30 cm), was connected to a high-pressure liquid chromatograph (HPLC) (Agilent Technologies, Agilent 1260 Infinity II) and equilibrated with PBS buffer at room temperature for at least 1 hour. An appropriate amount of protein sample (at least 10 μg) was filtered through a 0.22 μm filter membrane and injected into the system. The HPLC program was set to flow the sample through the column at a flow rate of 1.0 mL/min with PBS buffer for a maximum of 25 minutes. The HPLC will generate an analysis report reporting the retention time of different molecular size components in the sample.

实施例1.6抗DLL3抗体序列与编号Example 1.6 Anti-DLL3 Antibody Sequences and Numbering

在本文中，CDR的氨基酸序列均是按照Chothia定义规则所示出的。但是，本领域人员公知，在本领域中可以通过多种方法来定义抗体的CDR，例如基于序列可变性的Kabat定义规则(参见，Kabat等人，免疫学的蛋白质序列，第五版，美国国立卫生研究院，贝塞斯达，马里兰州(1991))和基于结构环区域位置的Chothia定义规则(参见JMol Biol 273:927-48,1997)。在本发明的技术方案中，还可以使用包含了Kabat定义和Chothia定义的Combined定义规则来确定可变结构域序列中的氨基酸残基。其中Combined定义规则即是将Kabat定义和Chothia定义的范围相结合，基于此取了一个更大的范围，详见表2。本领域技术人员应当理解的是，除非另有规定，否则术语给定抗体或其区(例如可变区)的“CDR”及“互补决定区”应了解为涵盖如通过本发明描述的上述已知方案中的任何一种界定的互补决定区。虽然本发明的权利要求中请求保护的范围是基于Chothia定义规则所示出的序列，但是根据其他CDR的定义规则所对应的氨基酸序列也应当落在本发明的保护范围中。In this article, the amino acid sequences of CDRs are shown in accordance with the Chothia definition rules. However, it is well known to those skilled in the art that the CDRs of antibodies can be defined in the art by a variety of methods, such as the Kabat definition rules based on sequence variability (see, Kabat et al., Protein Sequences in Immunology, Fifth Edition, National Institutes of Health, Bethesda, Maryland (1991)) and the Chothia definition rules based on the position of the structural loop region (see J Mol Biol 273:927-48, 1997). In the technical solution of the present invention, the Combined definition rule comprising the Kabat definition and the Chothia definition can also be used to determine the amino acid residues in the variable domain sequence. The Combined definition rule combines the scope of the Kabat definition and the Chothia definition, and a larger scope is taken based on this, as shown in Table 2. It should be understood by those skilled in the art that, unless otherwise specified, the terms "CDR" and "complementarity determining region" of a given antibody or its region (e.g., variable region) should be understood to encompass the complementary determining region defined by any of the above-mentioned known schemes described in the present invention. Although the scope of protection requested in the claims of the present invention is based on the sequences shown in the Chothia definition rules, the amino acid sequences corresponding to the definition rules of other CDRs should also fall within the scope of protection of the present invention.

表2.Kabat定义、Chothia定义和Combined定义
Table 2. Kabat definition, Chothia definition and Combined definition

其中，Haa-Hbb可以指从抗体重链的N端开始，第aa位(Chothia编码规则)至第bb位(Chothia编码规则)的氨基酸序列。例如，H26-H35可以指从抗体重链N端开始，按照Chothia编码规则的从第26位至第35位的氨基酸序列。本领域技术人员应当知晓的是，在用Chothia编码CDR时，有些位置会有插入位点的情况(可参见http://bioinf.org.uk/abs/)。Here, Haa-Hbb may refer to the amino acid sequence starting from the N-terminus of the antibody heavy chain and starting from position aa (Chothia numbering rules) to position bb (Chothia numbering rules). For example, H26-H35 may refer to the amino acid sequence starting from the N-terminus of the antibody heavy chain and starting from position 26 to position 35 according to the Chothia numbering rules. It should be understood by those skilled in the art that when using Chothia to encode CDRs, some positions may have insertion sites (see http://bioinf.org.uk/abs/).

44个候选抗体和两个对比例抗体的氨基酸序列如表3和表4所示，采用Chothia定义CDR的方式确定CDR序列。The amino acid sequences of 44 candidate antibodies and two comparative antibodies are shown in Tables 3 and 4. The CDR sequences were determined using the Chothia definition of CDR.

表3. 44个候选抗体的氨基酸序列(SEQ ID NO:)

Table 3. Amino acid sequences of 44 candidate antibodies (SEQ ID NO:)

表4. 2个对比例抗体的氨基酸序列(SEQ ID NO:)
Table 4. Amino acid sequences of two comparative antibodies (SEQ ID NO:)

实施例2 ELISA检测抗人DLL3的HCAb单抗蛋白水平的结合能力Example 2 ELISA detection of the binding ability of anti-human DLL3 HCAb monoclonal antibody protein level

本实施例研究抗人DLL3的HCAb单抗体外结合人/猴/小鼠DLL3的活性。采用人DLL3-his tag蛋白(Acro，#DL3-H52H4)、猴DLL3-his tag蛋白(Acro，#DL3-C52H9)、小鼠DLL3-his tag蛋白(Acro，#DL3-M52H9)，进行蛋白水平上的抗体结合实验。简言之，将人/猴/小鼠DLL3分别用PBS稀释为1mg/mL，加至96孔板(Corning，#9018)中，每孔100mL，4℃下孵育过夜。弃去液体后用PBST缓冲液(pH 7.4，含0.05％tween-20)洗板3次，加入250mL 2％BSA封闭液，37℃条件下孵育1小时。弃去封闭液，并用PBST缓冲液(pH7.4，含0.05％吐温-20)洗板3次，然后将5倍浓度梯度稀释的待测抗体待加入孔中，每个孔加入100mL，37℃孵育1小时，同型抗体作为对照。PBST缓冲液(pH7.4，含0.05％吐温-20)清洗3次后，加入稀释5000倍的羊抗人HRP二抗(Jackson ImmunoResearch，#109-035-098)，37℃条件下，避光孵育1小时。PBST缓冲液(pH7.4，含0.05％tween-20)清洗3次后，添加100mL/孔TMB(Biopanda，#TMB-S-003)，室温避光放置约30分钟；每孔加入50mL/孔终止液(BBI life sciences，#E661006-0200)终止反应，置酶标仪(MDC，Spectramax Plus384)中测定450nm吸光度(OD450)值。This example investigates the in vitro binding activity of anti-human DLL3 HCAb monoclonal antibodies to human, monkey, and mouse DLL3. Protein-level antibody binding experiments were performed using human DLL3-his-tag protein (Acro, #DL3-H52H4), monkey DLL3-his-tag protein (Acro, #DL3-C52H9), and mouse DLL3-his-tag protein (Acro, #DL3-M52H9). Briefly, human, monkey, and mouse DLL3 were diluted to 1 mg/mL in PBS, added to 100 mL per well of a 96-well plate (Corning, #9018), and incubated overnight at 4°C. After discarding the liquid, the plate was washed three times with PBST buffer (pH 7.4, containing 0.05% tween-20). 250 mL of 2% BSA blocking buffer was added, and the plate was incubated at 37°C for 1 hour. The blocking buffer was discarded and the plate was washed three times with PBST buffer (pH 7.4, containing 0.05% Tween-20). Then, 100 mL of the antibody to be tested was added to each well in a 5-fold serial dilution. The plate was incubated at 37°C for 1 hour. An isotype antibody was used as a control. After washing three times with PBST buffer (pH 7.4, containing 0.05% Tween-20), a 5000-fold diluted goat anti-human HRP secondary antibody (Jackson ImmunoResearch, #109-035-098) was added and incubated at 37°C for 1 hour in the dark. After washing three times with PBST buffer (pH 7.4, containing 0.05% tween-20), add 100 mL/well TMB (Biopanda, #TMB-S-003) and place in the dark at room temperature for about 30 minutes; add 50 mL/well stop solution (BBI life sciences, #E661006-0200) to each well to terminate the reaction, and measure the absorbance at 450 nm (OD450) in a microplate reader (MDC, Spectramax Plus384).

抗体结合人DLL3、猴DLL3以及小鼠DLL3的数据汇总如表5所示。44个单克隆抗体结合细胞表面的人DLL3、猴DLL3以及小鼠DLL3的结果分别如图1A-1D、图2A-2D、3A-3D所示。The data of antibody binding to human DLL3, monkey DLL3, and mouse DLL3 are summarized in Table 5. The results of 44 monoclonal antibodies binding to human DLL3, monkey DLL3, and mouse DLL3 on the cell surface are shown in Figures 1A-1D, 2A-2D, and 3A-3D, respectively.

结果显示：The results show:

1)18个抗体如PR013234、PR013235、PR013236、PR013239、PR013240、PR013241、PR013244、PR013246、PR013254、PR013256、PR013257、PR013258、PR013259、PR013261、PR013364、PR013377、PR013392、PR013402对人DLL3、猴DLL3和小鼠DLL3有优异的结合活性，尤其是PR013364、PR013377和PR013402，具有与参照抗体PR012621相当的最大值和EC50值。1) 18 antibodies such as PR013234, PR013235, PR013236, PR013239, PR013240, PR013241, PR013244, PR013246, PR013254, PR013256, PR013257, PR013258, PR013259, PR013261, PR013364, PR013377, PR013392, and PR013402 have excellent binding activity to human DLL3, monkey DLL3, and mouse DLL3, especially PR013364, PR013377, and PR013402, with maximum values and EC50 values comparable to those of the reference antibody PR012621.

2)11个抗体具有对人DLL3、猴DLL3的结合活性，但不结合小鼠DLL3，如PR013363、PR013367、PR013368、PR013370、PR013371、PR013376、PR013378、PR013382、PR013384、PR013387、PR013399。2) 11 antibodies have binding activity to human DLL3 and monkey DLL3, but do not bind to mouse DLL3, such as PR013363, PR013367, PR013368, PR013370, PR013371, PR013376, PR013378, PR013382, PR013384, PR013387, and PR013399.

3)PR013372和PR013394仅与人DLL3有结合。3) PR013372 and PR013394 only bind to human DLL3.

表5.DLL3抗体与人/猴/小鼠DLL3蛋白的结合活性



“\”代表未检测到，低于检测下限。Table 5. Binding activity of DLL3 antibodies to human/monkey/mouse DLL3 proteins



“\” means not detected, below the detection limit.

实施例3 FACS检测抗DLL3的HCAb单抗细胞水平的结合能力Example 3 FACS detection of the binding ability of anti-DLL3 HCAb monoclonal antibody at the cellular level

本实施例是为了研究抗DLL3的HCAb单抗体外结合人/食蟹猴DLL3的活性。采用过表达人DLL3的CHO-K1细胞株(仓鼠卵巢细胞亚株)(CHO-K1/humanDLL3，在本文中也可称为CHO-K1-humanDLL3，诺纳生物)、过表达食蟹猴DLL3的CHO-K1细胞株(CHO-K1/cynoDLL3，在本文中也可称为CHO-K1-cynoDLL3，诺纳生物)、CHO-K1母细胞株(CCL-61)或高表达人DLL3的细胞系SHP-77(人小细胞肺癌细胞)(CRL-2195)，DMS79(人小细胞肺癌细胞)(CRL-2049)进行细胞水平上的抗体结合实验。简言之，分别消化收集CHO-K1/humanDLL3细胞、CHO-K1/cynoDLL3细胞、CHO-K1细胞、SHP-77细胞或DMS79细胞，并用含2％FBS的PBS重悬。将细胞密度分别调整为1×10 6细胞/mL。以100μL细胞/孔接种于96孔V底板(Corning，#3894)，随后加入100μL/孔，2倍于终浓度的5倍浓度梯度稀释的待测抗体。将细胞放置于4℃，避光孵育2小时。之后，加入100μL/孔预冷含2％FBS的PBS漂洗细胞两次，于500g、4℃下离心5分钟，弃上清。再加入100μL/孔荧光二抗(Alexa Fluor 647标记亲和纯化山羊抗人IgG二抗，与Fcγ片段特异性反应，Jackson，#109-605-098，1:1000稀释)，4℃，避光孵育1小时。用100μL/孔预冷含2％FBS的PBS洗涤细胞两次，于500g、4℃下离心5分钟，弃上清。最后，200μL/孔预冷含2％FBS的PBS重悬细胞，使用BD CantoII流式细胞仪读取荧光发光信号值。This example is to study the activity of anti-DLL3 HCAb monoclonal antibody binding to human/cynomolgus monkey DLL3 in vitro. CHO-K1 cell line (hamster ovary cell subline) overexpressing human DLL3 (CHO-K1/humanDLL3, also referred to as CHO-K1-humanDLL3, Nona Bio), CHO-K1 cell line overexpressing cynomolgus monkey DLL3 (CHO-K1/cynoDLL3, also referred to as CHO-K1-cynoDLL3, Nona Bio), CHO-K1 mother cell line ( CCL-61) or the cell line SHP-77 (human small cell lung cancer cells) that highly expresses human DLL3 ( CRL-2195), DMS79 (human small cell lung cancer cells) CRL-2049) was used for antibody binding experiments at the cellular level. Briefly, CHO-K1/humanDLL3 cells, CHO-K1/cynoDLL3 cells, CHO-K1 cells, SHP-77 cells or DMS79 cells were digested and collected, and resuspended in PBS containing 2% FBS. The cell density was adjusted to 1×10 6 cells/mL. 100 μL cells/well were seeded in a 96-well V-bottom plate (Corning, #3894), followed by the addition of 100 μL/well of a 5-fold concentration gradient dilution of the antibody to be tested at 2 times the final concentration. The cells were placed at 4°C and incubated in the dark for 2 hours. Afterwards, 100 μL/well of pre-cooled PBS containing 2% FBS was added to rinse the cells twice, centrifuged at 500g, 4°C for 5 minutes, and the supernatant was discarded. Then, add 100 μL/well of fluorescent secondary antibody (Alexa Fluor 647-conjugated affinity-purified goat anti-human IgG secondary antibody, specifically reactive with Fcγ fragments, Jackson, #109-605-098, 1:1000 dilution) and incubate at 4°C for 1 hour in the dark. Wash the cells twice with 100 μL/well of pre-chilled PBS containing 2% FBS, centrifuge at 500g at 4°C for 5 minutes, and discard the supernatant. Finally, resuspend the cells in 200 μL/well of pre-chilled PBS containing 2% FBS and read the fluorescence signal using a BD CantoII flow cytometer.

抗体结合细胞表面的人DLL3、食蟹猴DLL3、CHO-K1母细胞以及结合肿瘤细胞SHP-77、DMS79表面的DLL3的数据汇总如表6和表7所示。44个单克隆抗体结合CHO-K1/humanDLL3、CHO-K1/cynoDLL3、CHO-K1细胞和肿瘤细胞SHP-77、DMS-79的结果如图4A-4D、图5A-5D、图6A-6D、图7A-7D和图8A-8D所示。The data for antibody binding to human DLL3, cynomolgus monkey DLL3, CHO-K1 mother cells, and DLL3 on the surface of tumor cells SHP-77 and DMS79 are summarized in Tables 6 and 7. The results of 44 monoclonal antibodies binding to CHO-K1/human DLL3, CHO-K1/cyno DLL3, CHO-K1 cells, and tumor cells SHP-77 and DMS-79 are shown in Figures 4A-4D, 5A-5D, 6A-6D, 7A-7D, and 8A-8D.

结果显示：The results show:

1)9个抗体如PR013236、PR013239、PR013240、PR013254、PR013256、PR013371、PR013377、PR013393、PR013406显示出优异的结合能力，不管是与过表达DLL3的重组细胞，还是与肿瘤细胞的结合，都具有与参照抗体PR012622相当的最大值和EC50值。1) Nine antibodies, including PR013236, PR013239, PR013240, PR013254, PR013256, PR013371, PR013377, PR013393, and PR013406, showed excellent binding ability, with maximum values and EC50 values comparable to those of the reference antibody PR012622, whether binding to recombinant cells overexpressing DLL3 or to tumor cells.

2)PR013250具有结合CHO-K1母细胞的活性，显示出非特异性结合。2) PR013250 has binding activity to CHO-K1 mother cells, showing non-specific binding.

表6.抗DLL3抗体与CHO-K1/humanDLL3和CHO-K1/cynoDLL3细胞的结合活性


Table 6. Binding activity of anti-DLL3 antibodies to CHO-K1/humanDLL3 and CHO-K1/cynoDLL3 cells


表7.抗DLL3抗体与肿瘤细胞SHP-77和DMS79的结合活性


Table 7. Binding activity of anti-DLL3 antibodies to tumor cells SHP-77 and DMS79


实施例4抗DLL3抗体的理化性质检测Example 4 Physicochemical Properties of Anti-DLL3 Antibodies

实施例4.1利用HIC-HPLC分析蛋白纯度和疏水性Example 4.1 Analysis of protein purity and hydrophobicity using HIC-HPLC

使用分析型疏水相互作用层析色谱法(HIC)来分析蛋白样品的纯度和疏水性。将分析型色谱柱TSKge1 Buty1-NPR(Tosoh Bioscience，14947，4.6mm×3.5cm)连接到高压液相色谱仪HPLC(Agilent Technologies，Agilent 1260Infinity II)，用PBS缓冲液室温下平衡至少1小时。设定方法由16分钟内从100％流动相A(20mM组氨酸，1.8M硫酸铵，pH 6.0)至100％流动相B(20mM组氨酸，pH 6.0)的线性梯度，流速设定为0.7mL/分钟，蛋白样品浓度1mg/mL，进样体积20μl，检测波长280nm。采集后用ChemStation软件对色谱图进行积分并计算相关数据，生成分析报告，报告出样品内不同分子尺寸组份的滞留时间。Analytical hydrophobic interaction chromatography (HIC) was used to analyze the purity and hydrophobicity of the protein samples. The analytical column TSKge1 Buty1-NPR (Tosoh Bioscience, 14947, 4.6 mm × 3.5 cm) was connected to a high-pressure liquid chromatograph HPLC (Agilent Technologies, Agilent 1260 Infinity II) and equilibrated with PBS buffer at room temperature for at least 1 hour. The method was set to a linear gradient from 100% mobile phase A (20 mM histidine, 1.8 M ammonium sulfate, pH 6.0) to 100% mobile phase B (20 mM histidine, pH 6.0) in 16 minutes, with a flow rate of 0.7 mL/min, a protein sample concentration of 1 mg/mL, an injection volume of 20 μl, and a detection wavelength of 280 nm. After acquisition, the chromatogram was integrated and the relevant data were calculated using ChemStation software to generate an analysis report reporting the retention time of different molecular size components in the sample.

实施例4.2利用DSF测定蛋白分子的热稳定性Example 4.2 Determination of thermal stability of protein molecules using DSF

差示扫描荧光法(Differential Scanning Fluorimetry，DSF)是一种常用的高通量的测定蛋白质热稳定性的方法。它使用实时荧光定量PCR仪器通过监测与去折叠的蛋白分子结合的染料的荧光强度的变化，来反映蛋白质的变性的过程，从而反映出蛋白分子的热稳定性。本实施例利用DSF方法来测定蛋白分子热变性温度(Tm)。10μg蛋白加入96-孔PCR板(Thermo，#AB-0700/W)，接着加入2μL 100×稀释的染料SYPROTM(Invitrogen，#2008138)，然后加入缓冲液使得终体积为40μL每孔。将PCR板密封，放置于实时荧光定量PCR仪器(Bio-Rad CFX96 PCR System)，先于25℃孵育5分钟，然后以0.2℃/0.2分钟的梯度逐渐从25℃升温至95℃，在测试结束时将温度降至25℃。使用FRET扫描模式并使用Bio-Rad CFX Maestro软件进行数据分析并计算出样品的Tm。Differential Scanning Fluorimetry (DSF) is a commonly used high-throughput method for determining protein thermal stability. It uses a real-time fluorescence quantitative PCR instrument to monitor the changes in the fluorescence intensity of the dye bound to the unfolded protein molecule to reflect the process of protein denaturation, thereby reflecting the thermal stability of the protein molecule. This example uses the DSF method to determine the thermal denaturation temperature (Tm) of the protein molecule. 10μg of protein was added to a 96-well PCR plate (Thermo, #AB-0700/W), followed by the addition of 2μL of 100× diluted dye SYPROTM (Invitrogen, #2008138), and then buffer was added to make the final volume 40μL per well. The PCR plate was sealed and placed in a real-time fluorescence quantitative PCR instrument (Bio-Rad CFX96 PCR System), first incubated at 25°C for 5 minutes, then gradually heated from 25°C to 95°C with a gradient of 0.2°C/0.2 minutes, and the temperature was lowered to 25°C at the end of the test. The FRET scanning mode was used and Bio-Rad CFX Maestro software was used for data analysis and calculation of the sample Tm.

抗体的理化性质如表8所示。结果显示12个抗体具有优异的SEC-HPLC纯度。PR013236、PR013377和PR013397具有较好的亲水性，热稳定性。The physicochemical properties of the antibodies are shown in Table 8. The results showed that all 12 antibodies had excellent SEC-HPLC purity. PR013236, PR013377, and PR013397 had good hydrophilicity and thermal stability.

表8.抗体的理化性质


“数量”指100mL的纯化体系得到的抗体量。
Tm1是检测VH区域的热稳定性，Tm2是检测CH2区域的热稳定性。
“/”代表本次实验未检测到。Table 8. Physicochemical properties of antibodies


“Amount” refers to the amount of antibody obtained in 100 mL of the purification system.
Tm1 is used to measure the thermal stability of the VH region, and Tm2 is used to measure the thermal stability of the CH2 region.
“/” means that it was not detected in this experiment.

实施例5抗DLL3抗体对重组人或猴DLL3蛋白的结合亲和力和解离常数的测定Example 5 Determination of Binding Affinity and Dissociation Constant of Anti-DLL3 Antibodies to Recombinant Human or Monkey DLL3 Protein

根据制造商说明，使用Octet RED96仪器(Fortiebio)和抗人IgG Fc亲和素传感器(AHC传感器，Pall ForteBio，#18-5060)测定亲和力。具体地，用含有0.1％(w/w)BSA和0.02％(v/v)吐温20的PBS缓冲液(pH7.4)将人DLL3蛋白(Acro，#DL3-H52H4)或者猴DLL3蛋白(Acro，#DL3-C52H9)稀释至200nM，分别与AHC传感器孵育。将40nM的DLL3抗体分别与负载人DLL3蛋白或猴蛋白的AHC传感器在30℃孵育3分钟。该反应混合物在含有0.1％(v/w)BSA和0.02％(v/v)吐温20的PBS缓冲液(pH7.4)中于30℃继续孵育5分钟。Octet Red 96实时记录DLL3抗体与人DLL3蛋白或者猴DLL3蛋白的结合和分离信号。Kon、Kdis(也可称为Koff)和KD值(KD＝Kdis(Koff)/Kon)通过使用Octet软件确定。Affinity was measured using an Octet RED96 instrument (Fortiebio) and an anti-human IgG Fc avidin sensor (AHC sensor, Pall FortieBio, #18-5060) according to the manufacturer's instructions. Specifically, human DLL3 protein (Acro, #DL3-H52H4) or monkey DLL3 protein (Acro, #DL3-C52H9) was diluted to 200 nM in PBS buffer (pH 7.4) containing 0.1% (w/w) BSA and 0.02% (v/v) Tween 20 and incubated with the AHC sensor. 40 nM of DLL3 antibody was incubated with the human or monkey protein-loaded AHC sensor at 30°C for 3 minutes. The reaction mixture was further incubated in PBS buffer (pH 7.4) containing 0.1% (v/w) BSA and 0.02% (v/v) Tween 20 at 30°C for 5 minutes. The binding and dissociation signals of the DLL3 antibody to human or monkey DLL3 protein were recorded in real time using the Octet Red 96 assay. Kon, Kdis (also known as Koff), and KD values (KD = Kdis(Koff)/Kon) were determined using Octet software.

结果如表9、表10和表11所示：The results are shown in Table 9, Table 10 and Table 11:

1)测试的8个抗DLL3抗体中PR013393与人DLL3蛋白和猴DLL3蛋白的KD值均低于2个参照抗体(PR012621和PR012622)，表明其更强的人和猴DLL3蛋白的结合亲和力。1) Among the eight anti-DLL3 antibodies tested, the KD values of PR013393 for human DLL3 protein and monkey DLL3 protein were lower than those of two reference antibodies (PR012621 and PR012622), indicating its stronger binding affinity to human and monkey DLL3 proteins.

2)在与人DLL3蛋白结合亲和力方面，PR013240、PR013371的KD值与参照抗体PR012621相当。PR013397甚至于优于参照抗体PR012622。2) In terms of binding affinity to human DLL3 protein, the KD values of PR013240 and PR013371 were comparable to the reference antibody PR012621. PR013397 was even superior to the reference antibody PR012622.

3)在与食蟹猴DLL3蛋白结合亲和力方面，PR013240、PR013371、PR013377和PR013397的KD值与参照抗体PR012621相当。3) In terms of binding affinity to cynomolgus monkey DLL3 protein, the KD values of PR013240, PR013371, PR013377 and PR013397 were comparable to those of the reference antibody PR012621.

表9.DLL3抗体与人DLL3蛋白的结合亲和力测定

Table 9. Binding affinity determination of DLL3 antibodies to human DLL3 protein

表10.DLL3抗体与猴DLL3蛋白的结合与解离常数
Table 10. Binding and dissociation constants of DLL3 antibodies to monkey DLL3 protein

表11.DLL3抗体与猴DLL3蛋白的结合与解离常数

Table 11. Binding and dissociation constants of DLL3 antibodies to monkey DLL3 protein

实施例6抗DLL3抗体在靶细胞上的内吞效应Example 6 Endocytic Effect of Anti-DLL3 Antibodies on Target Cells

抗DLL3抗体可以通过结合DLL3的胞外部介导细胞表面表达的DLL3蛋白的内化。在本实施例中，测试抗DLL3抗体在24小时的内化情况。具体方法如下：CHO-K1-humanDLL3或SHP-77细胞在T-75培养瓶中用含10％血清(VivaCell，胎牛血清，#2128194)的RPMI1640(Thermo，#C11875500CP)培养和扩增。将细胞转移到15mL无菌离心管中，1000rpm室温离心5分钟，使细胞成团。吸去培养基，将细胞重新悬浮于培养基中。轻柔地吹打细胞，得到单细胞混悬液。用细胞计数板进行计数，并把细胞浓度调整至2×105个细胞/mL(CHO-K1/humanDLL3，即图9A和图9B中的CHO-K1-hDLL3)或者1×106个细胞/ml(SHP-77)，取50mL每孔细胞悬液至96孔板(Corning，#3599)内，把96孔板放在37℃，5％CO2细胞培养箱内孵育过夜。96孔板内加入50mL抗体和Zenon pHrodo(Invitrogen，#Z25611)混合液，1:1混合。37℃，5％CO2细胞培养箱内孵育24小时。然后把96孔板中的细胞消化收集至96孔V型板中(Corning，#3894)，并用PBS将细胞洗2遍，重新悬浮在150mL FACS缓冲液中(2％FBS)。用FACS ACEA Novocyte3000流式细胞仪分析细胞的荧光强度。Anti-DLL3 antibodies can mediate the internalization of cell-surface-expressed DLL3 proteins by binding to the extracellular domain of DLL3. In this example, the internalization of anti-DLL3 antibodies was tested over a 24-hour period. The specific method is as follows: CHO-K1-human DLL3 or SHP-77 cells were cultured and expanded in T-75 flasks using RPMI 1640 (Thermo, #C11875500CP) supplemented with 10% serum (VivaCell, fetal bovine serum, #2128194). Cells were transferred to a sterile 15 mL centrifuge tube and centrifuged at 1000 rpm for 5 minutes at room temperature to pellet the cells. The culture medium was aspirated and the cells were resuspended in culture medium. The cells were gently pipetted to obtain a single-cell suspension. Cells were counted using a cell counter and adjusted to a concentration of 2× 10⁵ cells/mL (CHO-K1/humanDLL3, i.e., CHO-K1-hDLL3 in Figures 9A and 9B) or 1× 10⁶ cells/mL (SHP-77). 50 mL of the cell suspension was transferred from each well of a 96-well plate (Corning, #3599) and incubated overnight in a 37°C, 5% CO₂ incubator. 50 mL of a 1:1 mixture of antibody and Zenon pHrodo (Invitrogen, #Z25611) was added to the 96-well plate. The cells were incubated in a 37°C, 5% CO₂ incubator for 24 hours. The cells were then harvested from the 96-well plate into a 96-well V-plate (Corning, #3894), washed twice with PBS, and resuspended in 150 mL of FACS buffer (2% FBS). The fluorescence intensity of the cells was analyzed using a FACS ACEA Novocyte 3000 flow cytometer.

抗DLL3抗体在CHO-K1/humanDLL3和SHP-77细胞上的内吞效应数据汇总如表12和13所示。8个单克隆抗体在细胞上的内吞结果如图9A-9B和图10A-10B所示，结果显示，PR013240，PR013371和PR013397在CHO-K1/humanDLL3和SHP-77细胞上的内吞效应和参照抗体相当。The data for the endocytosis effects of anti-DLL3 antibodies in CHO-K1/humanDLL3 and SHP-77 cells are summarized in Tables 12 and 13. The endocytosis results of the eight monoclonal antibodies are shown in Figures 9A-9B and 10A-10B. The results show that PR013240, PR013371, and PR013397 have endocytosis effects comparable to those of the reference antibody in CHO-K1/humanDLL3 and SHP-77 cells.

表12.抗DLL3抗体在CHO-K1/humanDLL3和SHP-77细胞上的内吞效应
Table 12. Endocytic effects of anti-DLL3 antibodies on CHO-K1/humanDLL3 and SHP-77 cells

表13.抗DLL3抗体在CHO-K1/humanDLL3和SHP-77细胞上的内吞效应
Table 13. Endocytic effects of anti-DLL3 antibodies on CHO-K1/humanDLL3 and SHP-77 cells

实施例7构建过表达DLL3胞外结构域及其细胞表面的子结构域的HEK293重组细胞Example 7 Construction of HEK293 recombinant cells overexpressing the DLL3 extracellular domain and its cell surface subdomains

依据全长的人DLL3基因(Uniprot:Q9NYJ7)的序列，将相应的胞外结构域及其子结构域的编码序列克隆至pcDNA3.1载体中，FLAG标签插入在信号肽和胞外结构域之间。过表达胞外结构域和子结构域的重组质粒均包含N端小鼠IgG Vk前导序列，随后是FLAG标签和相应的人DLL3结构域。为确保细胞表面人DLL3结构域的定位，所有的重组质粒后面均有Ser/Gly连接子、人EpCAM(上皮细胞粘附分子)(P16422)的跨膜结构域(aa266-288)和人EpCAM(P16422)的胞内结构域(aa 289-314)。使用FITC标记的抗FLAG抗体(克隆M2，Sigma)来验证细胞表面上的胞外结构域及其子结构域的表达。Based on the sequence of the full-length human DLL3 gene (Uniprot: Q9NYJ7), the coding sequences of the corresponding extracellular domains and their subdomains were cloned into the pcDNA3.1 vector, with a FLAG tag inserted between the signal peptide and the extracellular domain. Recombinant plasmids overexpressing the extracellular domains and subdomains all contained an N-terminal mouse IgG Vk leader sequence followed by a FLAG tag and the corresponding human DLL3 domain. To ensure cell surface localization of the human DLL3 domain, all recombinant plasmids were followed by a Ser/Gly linker, the transmembrane domain of human EpCAM (epithelial cell adhesion molecule) (P16422) (aa 266-288), and the intracellular domain of human EpCAM (P16422) (aa 289-314). FITC-conjugated anti-FLAG antibody (clone M2, Sigma) was used to verify cell surface expression of the extracellular domains and their subdomains.

DLL3的胞外结构域包括以下几个不同的子结构域：The extracellular domain of DLL3 consists of several distinct subdomains:

·人DLL3信号肽：aa 1-26Human DLL3 signal peptide: aa 1-26

·人DLL3 N-端ECD区域：aa 27-175Human DLL3 N-terminal ECD region: aa 27-175

·人DLL3 DSL：aa 176-215Human DLL3 DSL: aa 176-215

·人DLL3EGF 1：aa 216-249Human DLL3EGF 1: aa 216-249

·人DLL3EGF2：aa 274-310Human DLL3EGF2: aa 274-310

·人DLL3EGF3：aa 312-351Human DLL3EGF3: aa 312-351

·人DLL3EGF4：aa 353-389Human DLL3EGF4: aa 353-389

·人DLL3EGF5：aa 391-427Human DLL3EGF5: aa 391-427

·人DLL3EGF6：aa 429-465Human DLL3EGF6: aa 429-465

·近膜端肽段：aa 466-492Membrane-proximal peptide segment: aa 466-492

HEK293细胞上过表达各种DLL3子结构域的序列如下表所示。The sequences of various DLL3 subdomains overexpressed in HEK293 cells are shown in the table below.

表14.HEK293细胞上过表达各种DLL3子结构域的序列
Table 14. Sequences of various DLL3 subdomains overexpressed in HEK293 cells

实施例8抗DLL3抗体与DLL3蛋白的结合表位的鉴定Example 8 Identification of the Binding Epitope of Anti-DLL3 Antibodies to DLL3 Protein

过表达DLL3子结构域的重组质粒的具体信息如表14和图11A所示。这些质粒分别瞬时转染HEK239T细胞，48小时后收集细胞，用FITC标记的抗FLAG抗体检测重组细胞系的转染效率，未转染的HEK93T细胞作为转染效率检测的阴性对照。同时用1mg/mL DLL3抗体进行一抗染色。hIgG1作为同型阴性参照抗体。使用荧光抗体(Alexa Fluor 647标记亲和纯化山羊抗人IgG二抗，与Fcγ片段特异性反应，Jackson，#109-605-098，1:1000稀释)作为二抗，通过流式细胞术测量抗DLL3抗体与子结构域的瞬转细胞的结合能力。Detailed information about the recombinant plasmids overexpressing the DLL3 subdomain is shown in Table 14 and Figure 11A. These plasmids were transiently transfected into HEK239T cells. Cells were harvested 48 hours later, and the transfection efficiency of the recombinant cell lines was determined using a FITC-labeled anti-FLAG antibody. Untransfected HEK93T cells served as a negative control for transfection efficiency. Primary antibody staining was performed using 1 mg/mL DLL3 antibody. hIgG1 was used as an isotype negative reference antibody. The binding of the anti-DLL3 antibody to the transiently transfected cells containing the subdomain was measured by flow cytometry using a fluorescent secondary antibody (Alexa Fluor 647-labeled affinity-purified goat anti-human IgG, specifically reactive with Fcγ fragments, Jackson, #109-605-098, 1:1000 dilution) as a secondary antibody.

图11B示出过表达DLL3子结构域的重组质粒均瞬转至HEK293T细胞。图12A-12J示出DLL3抗体结合人DLL3的胞外端的不同区域，结果如表15所示。Figure 11B shows that recombinant plasmids overexpressing DLL3 subdomains were transiently transfected into HEK293T cells. Figures 12A-12J show that DLL3 antibodies bind to different regions of the extracellular end of human DLL3, and the results are shown in Table 15.

表15.抗DLL3抗体与DLL3蛋白的结合表位

Table 15. Binding epitopes of anti-DLL3 antibodies to DLL3 protein

序列表Sequence Listing

PR012621(抗DLL3rovalpituzumab hIgG1)

PR012621 (anti-DLL3 rovalvituzumab hIgG1)

PR012622(抗DLL3 DLL3-4_CDDI hIgG1)
PR012622 (anti-DLL3 DLL3-4_CDDI hIgG1)

PR013234(抗DLL3 N1003P003A12 HCAb hIgG1(C220S))
PR013234 (anti-DLL3 N1003P003A12 HCAb hIgG1(C220S))

PR013235(抗DLL3 N1003P003B12 HCAb hIgG1(C220S))
PR013235 (anti-DLL3 N1003P003B12 HCAb hIgG1(C220S))

PR013236(抗DLL3 N1003P003D01 HCAb hIgG1(C220S))
PR013236 (anti-DLL3 N1003P003D01 HCAb hIgG1(C220S))

PR013239(抗DLL3 N1003P005E04 HCAb hIgG1(C220S))
PR013239 (anti-DLL3 N1003P005E04 HCAb hIgG1(C220S))

PR013240(抗DLL3 N1003P005G03 HCAb hIgG1(C220S))
PR013240 (anti-DLL3 N1003P005G03 HCAb hIgG1(C220S))

PR013241(抗DLL3 N1003P007B07 HCAb hIgG1(C220S))
PR013241 (anti-DLL3 N1003P007B07 HCAb hIgG1(C220S))

PR013242(抗DLL3 N1003P008A11 HCAb hIgG1(C220S))
PR013242 (anti-DLL3 N1003P008A11 HCAb hIgG1(C220S))

PR013244(抗DLL3 N1003P008B09 HCAb hIgG1(C220S))
PR013244 (anti-DLL3 N1003P008B09 HCAb hIgG1(C220S))

PR013246(抗DLL3 N1003P008H01 HCAb hIgG1(C220S))
PR013246 (anti-DLL3 N1003P008H01 HCAb hIgG1(C220S))

PR013249(抗DLL3 N1003P013C05 HCAb hIgG1(C220S))
PR013249 (anti-DLL3 N1003P013C05 HCAb hIgG1(C220S))

PR013250(抗DLL3 N1003P015H03 HCAb hIgG1(C220S))
PR013250 (anti-DLL3 N1003P015H03 HCAb hIgG1(C220S))

PR013252(抗DLL3 N1003P017G01 HCAb hIgG1(C220S))
PR013252 (anti-DLL3 N1003P017G01 HCAb hIgG1(C220S))

PR013253(抗DLL3 N1003P018C04 HCAb hIgG1(C220S))
PR013253 (anti-DLL3 N1003P018C04 HCAb hIgG1(C220S))

PR013254(抗DLL3 N1003P018F06 HCAb hIgG1(C220S))
PR013254 (anti-DLL3 N1003P018F06 HCAb hIgG1(C220S))

PR013256(抗DLL3 N1003P019B04 HCAb hIgG1(C220S))
PR013256 (anti-DLL3 N1003P019B04 HCAb hIgG1(C220S))

PR013257(抗DLL3 N1003P020A07 HCAb hIgG1(C220S))
PR013257 (anti-DLL3 N1003P020A07 HCAb hIgG1(C220S))

PR013258(抗DLL3 N1003P020C12 HCAb hIgG1(C220S))
PR013258 (anti-DLL3 N1003P020C12 HCAb hIgG1(C220S))

PR013259(抗DLL3 N1003P022A02 HCAb hIgG1(C220S))
PR013259 (anti-DLL3 N1003P022A02 HCAb hIgG1(C220S))

PR013260(抗DLL3 N1003P024E01 HCAb hIgG1(C220S))
PR013260 (anti-DLL3 N1003P024E01 HCAb hIgG1(C220S))

PR013261(抗DLL3 N1003P024G01 HCAb hIgG1(C220S))
PR013261 (anti-DLL3 N1003P024G01 HCAb hIgG1(C220S))

PR013262(抗DLL3 N1003P027E04 HCAb hIgG1(C220S))
PR013262 (anti-DLL3 N1003P027E04 HCAb hIgG1(C220S))

PR013263(抗DLL3 N1003P028F03 HCAb hIgG1(C220S))
PR013263 (anti-DLL3 N1003P028F03 HCAb hIgG1(C220S))

PR013264(抗DLL3 N1003P030F03 HCAb hIgG1(C220S))
PR013264 (anti-DLL3 N1003P030F03 HCAb hIgG1(C220S))

PR013363(抗DLL3 N1003P041D05 HCAb hIgG1(C220S))
PR013363 (anti-DLL3 N1003P041D05 HCAb hIgG1(C220S))

PR013364(抗DLL3 N1003P041G02 HCAb hIgG1(C220S))
PR013364 (anti-DLL3 N1003P041G02 HCAb hIgG1(C220S))

PR013367(抗DLL3 N1003P044A10 HCAb hIgG1(C220S))
PR013367 (anti-DLL3 N1003P044A10 HCAb hIgG1(C220S))

PR013368(抗DLL3 N1003P044G07 HCAb hIgG1(C220S))
PR013368 (anti-DLL3 N1003P044G07 HCAb hIgG1(C220S))

PR013370(抗DLL3 N1003P046B02 HCAb hIgG1(C220S))
PR013370 (anti-DLL3 N1003P046B02 HCAb hIgG1(C220S))

PR013371(抗DLL3 N1003P046E04 HCAb hIgG1(C220S))
PR013371 (anti-DLL3 N1003P046E04 HCAb hIgG1(C220S))

PR013372(抗DLL3 N1003P046G02 HCAb hIgG1(C220S))
PR013372 (anti-DLL3 N1003P046G02 HCAb hIgG1(C220S))

PR013376(抗DLL3 N1003P047C08 HCAb hIgG1(C220S))
PR013376 (anti-DLL3 N1003P047C08 HCAb hIgG1(C220S))

PR013377(抗DLL3 N1003P047D04 HCAb hIgG1(C220S))
PR013377 (anti-DLL3 N1003P047D04 HCAb hIgG1(C220S))

PR013378(抗DLL3 N1003P047D06 HCAb hIgG1(C220S))
PR013378 (anti-DLL3 N1003P047D06 HCAb hIgG1(C220S))

PR013381(抗DLL3 N1003P048B11 HCAb hIgG1(C220S))
PR013381 (anti-DLL3 N1003P048B11 HCAb hIgG1(C220S))

PR013382(抗DLL3 N1003P049A11 HCAb hIgG1(C220S))
PR013382 (anti-DLL3 N1003P049A11 HCAb hIgG1(C220S))

PR013384(抗DLL3 N1003P051B09 HCAb hIgG1(C220S))
PR013384 (anti-DLL3 N1003P051B09 HCAb hIgG1(C220S))

PR013387(抗DLL3 N1003P052G11 HCAb hIgG1(C220S))
PR013387 (anti-DLL3 N1003P052G11 HCAb hIgG1(C220S))

PR013392(抗DLL3 N1003P054C06 HCAb hIgG1(C220S))
PR013392 (anti-DLL3 N1003P054C06 HCAb hIgG1(C220S))

PR013393(抗DLL3 N1003P054C07 HCAb hIgG1(C220S))
PR013393 (anti-DLL3 N1003P054C07 HCAb hIgG1(C220S))

PR013394(抗DLL3 N1003P054D06 HCAb hIgG1(C220S))
PR013394 (anti-DLL3 N1003P054D06 HCAb hIgG1(C220S))

PR013397(抗DLL3 N1003P054E04 HCAb hIgG1(C220S))
PR013397 (anti-DLL3 N1003P054E04 HCAb hIgG1(C220S))

PR013399(抗DLL3 N1003P054H03 HCAb hIgG1(C220S))
PR013399 (anti-DLL3 N1003P054H03 HCAb hIgG1(C220S))

PR013402(抗DLL3 N1003P055D04 HCAb hIgG1(C220S))
PR013402 (anti-DLL3 N1003P055D04 HCAb hIgG1(C220S))

PR013406(抗DLL3 N1003P058H08 HCAb hIgG1(C220S))


PR013406 (anti-DLL3 N1003P058H08 HCAb hIgG1(C220S))


Claims (20)

一种DLL3结合分子，其包含重链可变区，其中所述重链可变区包含HCDR1、HCDR2和HCDR3，并且其中A DLL3 binding molecule comprising a heavy chain variable region, wherein the heavy chain variable region comprises HCDR1, HCDR2 and HCDR3, and wherein HCDR1包含SEQ ID NO:18或32所示的序列或者与SEQ ID NO:18或32所示的序列存在不超过2个氨基酸的氨基酸添加、缺失或取代的差异的氨基酸序列；和/或HCDR1 comprises the sequence shown in SEQ ID NO: 18 or 32, or an amino acid sequence that differs from the sequence shown in SEQ ID NO: 18 or 32 by an amino acid addition, deletion or substitution of not more than 2 amino acids; and/or HCDR2包含SEQ ID NO:69、74、79、80、87或90所示的序列或者与SEQ ID NO:69、74、79、80、87或90所示的序列存在不超过2个氨基酸的氨基酸添加、缺失或取代的差异的氨基酸序列；和/或HCDR2 comprises the sequence shown in SEQ ID NO: 69, 74, 79, 80, 87 or 90, or an amino acid sequence that differs from the sequence shown in SEQ ID NO: 69, 74, 79, 80, 87 or 90 by an amino acid addition, deletion or substitution of not more than 2 amino acids; and/or HCDR3包含SEQ ID NO:126、127、128、129、133、135、136、137、138、139、140、141、142、143、144、146、147、148、149、150、151或152所示的序列或者与SEQ ID NO:126、127、128、129、133、135、136、137、138、139、140、141、142、143、144、146、147、148、149、150、151或152所示的序列存在不超过2个氨基酸的氨基酸添加、缺失或取代的差异的氨基酸序列。HCDR3 comprises the sequence shown in SEQ ID NO: 126, 127, 128, 129, 133, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 146, 147, 148, 149, 150, 151 or 152, or an amino acid sequence that differs from the sequence shown in SEQ ID NO: 126, 127, 128, 129, 133, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 146, 147, 148, 149, 150, 151 or 152 by amino acid addition, deletion or substitution of no more than 2 amino acids. 权利要求1的DLL3结合分子，其中所述DLL3结合分子是针对DLL3的抗体或其抗原结合片段。The DLL3 binding molecule of claim 1, wherein the DLL3 binding molecule is an antibody or antigen-binding fragment thereof directed against DLL3. 权利要求1或2的DLL3结合分子，其中The DLL3 binding molecule of claim 1 or 2, wherein HCDR1包含如SEQ ID NO:18、19、20、21、22、23、24、25、26、27、28、29、30、31、32、33或34所示的氨基酸序列；HCDR1 comprises the amino acid sequence set forth in SEQ ID NO: 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, 33 or 34; HCDR2包含如SEQ ID NO:60、61、62、63、64、65、66、68、69、70、71、72、73、74、75、76、77、78、79、80、81、82、83、84、85、86、87、88、89或90所示的氨基酸序列；和/或HCDR2 comprises the amino acid sequence of SEQ ID NO: 60, 61, 62, 63, 64, 65, 66, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89 or 90; and/or HCDR3包含如SEQ ID NO:126、127、128、129、131、132、133、134、135、136、137、138、139、140、141、142、143、144、145、146、147、148、149、150、151或152所示的氨基酸序列。HCDR3 comprises an amino acid sequence as shown in SEQ ID NO:126, 127, 128, 129, 131, 132, 133, 134, 135, 136, 137, 138, 139, 140, 141, 142, 143, 144, 145, 146, 147, 148, 149, 150, 151 or 152. 权利要求1-3中任一项的DLL3结合分子，其中所述重链可变区包含HCDR1、HCDR2和HCDR3，并且其中The DLL3 binding molecule of any one of claims 1 to 3, wherein the heavy chain variable region comprises HCDR1, HCDR2, and HCDR3, and wherein HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、60、126所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 18, 60 and 126, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:19、60、126所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 19, 60 and 126, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、61、126所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 18, 61 and 126, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:20、62、127所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 20, 62 and 127, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、63、128所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 18, 63 and 128, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:21、64、129所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 21, 64 and 129, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:22、65、126所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 22, 65 and 126, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:23、65、126所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 23, 65 and 126, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、66、126所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 18, 66 and 126, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、64、131所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 18, 64 and 131, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:19、68、126所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 19, 68 and 126, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、68、126所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 18, 68 and 126, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:20、69、132所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 20, 69 and 132, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:24、66、126所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 24, 66 and 126, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、70、133所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 18, 70 and 133, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、70、134所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 18, 70 and 134, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、71、134所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 18, 71 and 134, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:25、72、135所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 25, 72 and 135, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、73、134所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 18, 73 and 134, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:25、74、135所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 25, 74 and 135, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:26、75、136所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 26, 75 and 136, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:27、76、137所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 27, 76 and 137, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:25、77、138所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 25, 77 and 138, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:25、78、139所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 25, 78 and 139, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、79、140所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 18, 79 and 140, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:26、80、141所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 26, 80 and 141, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:26、81、142所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 26, 81 and 142, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:25、82、143所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 25, 82 and 143, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:28、83、144所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 28, 83 and 144, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:29、84、143所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 29, 84 and 143, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:26、85、145所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 26, 85 and 145, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:30、75、136所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 30, 75 and 136, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:31、85、146所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 31, 85 and 146, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:32、86、147所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 32, 86 and 147, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、85、148所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 18, 85 and 148, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:33、87、149所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 33, 87 and 149, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:26、88、150所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 26, 88 and 150, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:18、89、151所示的氨基酸序列；或者HCDR1, HCDR2 and HCDR3 comprise the amino acid sequences shown in SEQ ID NO: 18, 89 and 151, respectively; or HCDR1、HCDR2和HCDR3分别包含如SEQ ID NO:34、90、152所示的氨基酸序列。HCDR1, HCDR2 and HCDR3 contain the amino acid sequences shown in SEQ ID NO:34, 90 and 152, respectively. 权利要求1-4中任一项的DLL3结合分子，其中The DLL3 binding molecule of any one of claims 1 to 4, wherein 所述重链可变区包含1)SEQ ID NO:174、175、176、177、178、179、180、181、182、183、185、186、187、188、189、190、191、192、193、194、195、196、197、198、199、200、201、202、203、204、205、206、207、208、209、210、211、212、213、214、215、216或217的氨基酸序列；2)与SEQ ID NO:174、175、176、177、178、179、180、181、182、183、185、186、187、188、189、190、191、192、193、194、195、196、197、198、199、200、201、202、203、204、205、206、207、208、209、210、211、212、213、214、215、216或217的氨基酸序列具有至少80％、至少85％、至少90％、至少91％、至少92％、至少93％、至少94％、至少95％、至少96％、至少97％、至少98％、至少99％序列相同性的氨基酸序列；或者3)与SEQ ID NO:174、175、176、177、178、179、180、181、182、183、185、186、187、188、189、190、191、192、193、194、195、196、197、198、199、200、201、202、203、204、205、206、207、208、209、210、211、212、213、214、215、216或217的氨基酸序列相比具有一个或多个氨基酸的取代、添加和/或缺失的氨基酸序列，优选地，所述添加、缺失和/或取代不发生在CDR区。The heavy chain variable region comprises 1) SEQ ID NO: 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214 , 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216, or 217; 2) an amino acid sequence having the same amino acid sequence as SEQ ID NO: 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 185, 186, 187, 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 185, 186, 187, 188, 189, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216, or 217; or 3) an amino acid sequence that has at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% sequence identity to the amino acid sequence of SEQ ID NO: 174, 175, 176, 177, 178, 179, 180, 181, 182, 183, 185, 186, 187 , 188, 189, 190, 191, 192, 193, 194, 195, 196, 197, 198, 199, 200, 201, 202, 203, 204, 205, 206, 207, 208, 209, 210, 211, 212, 213, 214, 215, 216 or 217, and preferably, the addition, deletion and/or substitution does not occur in the CDR region. 权利要求1-5中任一项的DLL3结合分子，其为重链抗体和/或为全人源化抗体。The DLL3 binding molecule of any one of claims 1 to 5, which is a heavy chain antibody and/or a fully humanized antibody. 权利要求1-5中任一项的DLL3结合分子，其中所述重链可变区可以与另一分子融合；The DLL3 binding molecule of any one of claims 1 to 5, wherein the heavy chain variable region can be fused to another molecule; 优选地，所述另一分子是免疫球蛋白的Fc结构域；Preferably, the other molecule is the Fc domain of an immunoglobulin; 最优选地，所述另一分子是免疫球蛋白G1(IgG1)的Fc结构域。Most preferably, the other molecule is the Fc domain of immunoglobulin G1 (IgG1). 权利要求1-7中任一项的DLL3结合分子，其包含1)SEQ ID NO:222、223、224、225、226、227、228、229、230、231、233、234、235、236、237、238、239、240、241、242、243、244、245、246、247、248、249、250、251、252、253、254、255、256、257、258、259、260、261、262、263、264或265的氨基酸序列；2)与SEQ ID NO:222、223、224、225、226、227、228、229、230、231、233、234、235、236、237、238、239、240、241、242、243、244、245、246、247、248、249、250、251、252、253、254、255、256、257、258、259、260、261、262、263、264或265的氨基酸序列具有至少80％、至少85％、至少90％、至少91％、至少92％、至少93％、至少94％、至少95％、至少96％、至少97％、至少98％、至少99％序列相同性的氨基酸序列；或者3)与SEQ ID NO:222、223、224、225、226、227、228、229、230、231、233、234、235、236、237、238、239、240、241、242、243、244、245、246、247、248、249、250、251、252、253、254、255、256、257、258、259、260、261、262、263、264或265的氨基酸序列相比具有一个或多个氨基酸的取代、添加和/或缺失的氨基酸序列，优选地，所述添加、缺失和/或取代不发生在CDR区。The DLL3 binding molecule of any one of claims 1-7, comprising 1) SEQ ID NO: 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257, 258, 259, 260, 261, 262, 263, 264 or 265; 2) an amino acid sequence having the same amino acid sequence as SEQ ID NO: 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257, 2 58, 259, 260, 261, 262, 263, 264 or 265 has an amino acid sequence that is at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% sequence identity; or 3) an amino acid sequence that is at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99% sequence identity to the amino acid sequence of SEQ ID NO: 222, 223, 224, 225, 226, 227, 228, 229, 230, 231, 233, 234, 235, 236, 237, 238, 239, 240, 241, 242, 243, 244, 245, 246, 247, 248, 249, 250, 251, 252, 253, 254, 255, 256, 257, 258, 259, 260, 261, 262, 263, 264 or 265, compared to the amino acid sequence having one or more amino acid substitutions, additions and/or deletions, preferably, the additions, deletions and/or substitutions do not occur in the CDR regions. 权利要求1-8中任一项的DLL3结合分子，其具有以下特征中的至少一个：The DLL3 binding molecule of any one of claims 1 to 8, having at least one of the following features: 1)具有对DLL3蛋白的亲和活性；1) Has affinity activity for DLL3 protein; 2)具有对DLL3阳性细胞的亲和活性；2) has affinity activity for DLL3-positive cells; 3)能够内化至DLL3阳性的细胞内。3) Ability to be internalized into DLL3-positive cells. 一种多特异性抗原结合构建体，其包含结合DLL3的第一抗原结合部分，其中所述第一抗原结合部分包含权利要求1-9中任一项的DLL3结合分子；a multispecific antigen-binding construct comprising a first antigen-binding moiety that binds DLL3, wherein the first antigen-binding moiety comprises the DLL3-binding molecule of any one of claims 1 to 9; 优选地，所述多特异性抗原结合构建体包含结合DLL3的第一抗原结合部分以及结合第二抗原的第二抗原结合部分；Preferably, the multispecific antigen-binding construct comprises a first antigen-binding moiety that binds DLL3 and a second antigen-binding moiety that binds a second antigen; 优选地，所述第二抗原结合部分特异性结合肿瘤抗原、免疫调节受体和免疫检查点分子；Preferably, the second antigen binding moiety specifically binds to tumor antigens, immunomodulatory receptors and immune checkpoint molecules; 优选地，所述第二抗原结合部分特异性结合淋巴细胞上的抗原。Preferably, the second antigen binding moiety specifically binds an antigen on a lymphocyte. 一种嵌合抗原受体，其包含权利要求1-9中任一项的DLL3结合分子。A chimeric antigen receptor comprising the DLL3 binding molecule of any one of claims 1-9. 一种免疫效应细胞，其在表面表达权利要求11的嵌合抗原受体；优选地，所述免疫效应细胞包含细胞毒性T细胞、自然杀伤细胞、巨噬细胞或自然杀伤T细胞。An immune effector cell expressing the chimeric antigen receptor of claim 11 on its surface; preferably, the immune effector cell comprises a cytotoxic T cell, a natural killer cell, a macrophage or a natural killer T cell. 一种抗体缀合物，其包含与至少一种治疗剂缀合的权利要求1-9中任一项的DLL3结合分子或者权利要求10的多特异性抗原结合构建体。An antibody conjugate comprising the DLL3 binding molecule of any one of claims 1 to 9 or the multispecific antigen-binding construct of claim 10 conjugated to at least one therapeutic agent. 一种药物组合物，其包含权利要求1-9中任一项的DLL3结合分子、权利要求10的多特异性抗原结合构建体、权利要求12的免疫效应细胞或者权利要求13的抗体缀合物，以及药学上可接受的载剂。A pharmaceutical composition comprising the DLL3 binding molecule of any one of claims 1 to 9, the multispecific antigen-binding construct of claim 10, the immune effector cell of claim 12 or the antibody conjugate of claim 13, and a pharmaceutically acceptable carrier. 权利要求1-9中任一项的DLL3结合分子、权利要求10的多特异性抗原结合构建体、权利要求12的免疫效应细胞、权利要求13的抗体缀合物或者权利要求14的药物组合物在制备用于治疗癌症的药物中的用途；优选地，所述癌症为DLL3阳性的肿瘤；较优选地，所述癌症选自小细胞肺癌(SCLC)；大细胞神经内分泌癌(LCNEC)；包括肾脏、泌尿生殖道、胃肠道、甲状腺、胰腺和肺的多种组织的神经内分泌肿瘤；神经胶质瘤或假性神经内分泌肿瘤(pNET)。Use of the DLL3 binding molecule of any one of claims 1 to 9, the multispecific antigen-binding construct of claim 10, the immune effector cell of claim 12, the antibody conjugate of claim 13, or the pharmaceutical composition of claim 14 in the preparation of a medicament for treating cancer; preferably, the cancer is a DLL3-positive tumor; more preferably, the cancer is selected from small cell lung cancer (SCLC); large cell neuroendocrine carcinoma (LCNEC); neuroendocrine tumors of various tissues including the kidney, genitourinary tract, gastrointestinal tract, thyroid, pancreas and lung; glioma or pseudoneuroendocrine tumor (pNET). 权利要求15的用途，其中所述药物与选自以下的一种或多种治疗剂联合使用：化疗剂、放射性同位素、免疫检查点抑制剂和肿瘤抗原靶向药物。The use of claim 15, wherein the drug is used in combination with one or more therapeutic agents selected from the group consisting of chemotherapeutic agents, radioisotopes, immune checkpoint inhibitors, and tumor antigen targeting drugs. 一种多核苷酸，其编码权利要求1-9中任一项的DLL3结合分子或者权利要求10的多特异性抗原结合构建体。A polynucleotide encoding the DLL3 binding molecule of any one of claims 1 to 9 or the multispecific antigen-binding construct of claim 10. 一种表达载体，其包含权利要求17的多核苷酸。An expression vector comprising the polynucleotide of claim 17. 一种宿主细胞，其包含权利要求17的多核苷酸或权利要求18的表达载体。A host cell comprising the polynucleotide of claim 17 or the expression vector of claim 18. 一种产生权利要求1-9中任一项的DLL3结合分子或者权利要求10的多特异性抗原结合构建体的方法，所述方法包括：A method of producing the DLL3-binding molecule of any one of claims 1 to 9 or the multispecific antigen-binding construct of claim 10, the method comprising: a)在合适条件下培养权利要求19的宿主细胞以表达权利要求1-9中任一项的DLL3结合分子或者权利要求10的多特异性抗原结合构建体；以及a) culturing the host cell of claim 19 under suitable conditions to express the DLL3 binding molecule of any one of claims 1 to 9 or the multispecific antigen-binding construct of claim 10; and b)从宿主细胞或其培养物分离所述DLL3结合分子或者多特异性抗原结合构建体。b) isolating the DLL3 binding molecule or multispecific antigen-binding construct from the host cell or culture thereof.
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